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Guidelines Establishing Test Procedures for the
Analysis of Pollutants; Whole Effluent Toxicity
Test Methods; Final Rule
[Federal Register: November 19, 2002 (Volume 67, Number 223)]
[Rules and Regulations]
[Page 69951-69972]
From the Federal Register Online via GPO Access [wais.access.gpo.gov]
[DOCID:fr19no02-13]
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ENVIRONMENTAL PROTECTION AGENCY
40 CFR Part 136
[FRL-7408-6]
RIN 2040-AD73
Guidelines Establishing Test Procedures for the Analysis of
Pollutants; Whole Effluent Toxicity Test Methods; Final Rule
AGENCY: Environmental Protection Agency (EPA).
ACTION: Final rule.
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SUMMARY: In this final regulation, EPA ratifies approval of several
test procedures for measuring the toxicity of effluents and receiving
waters. The test procedures are commonly referred to as whole effluent
toxicity or WET test methods. EPA also withdraws two WET test methods
from the list of nationally-approved biological test procedures for the
analysis of pollutants. This action also revises some of the WET test
methods to improve performance and increase confidence in the
reliability of the results. Today's action will satisfy settlement
agreement obligations designed to resolve litigation over an earlier
rulemaking that originally approved WET test methods.
DATES: This regulation is effective December 19, 2002. For judicial
review purposes, this final rule is promulgated as of 1:00 p.m. Eastern
Standard Time on December 3, 2002 in accordance with 40 CFR 23.7. The
incorporation by reference of certain publications listed in this rule
is approved by the Director of the Federal Register as of December 19,
2002.
FOR FURTHER INFORMATION CONTACT: Marion Kelly; Engineering and Analysis
Division (4303T); Office of Science and Technology; Office of Water,
U.S. Environmental Protection Agency; Ariel Rios Building; 1200
Pennsylvania Avenue, NW; Washington, DC 20460, or call (202) 566-1045,
or E-mail at kelly.marion@epa.gov. For technical information regarding
method changes in today's rule, contact Debra L. Denton, USEPA Region
9, c/o SWRCB, 1001 I Street, Sacramento, CA 95814, or call (916) 341-
5520, or E-mail denton.debra@epa.gov.
SUPPLEMENTARY INFORMATION
I. General Information
A. Potentially Regulated Entities
B. How Can I Get Copies Of Related Information?
1. Docket
2. Electronic Access
II. Statutory Authority
III. Background
A. Regulatory History
B. Settlement Agreement
C. Proposed Rule
IV. Summary of Final Rule
A. Proposed WET Method Changes
B. Additional Revisions to WET Test Methods
C. Ratification and Withdrawal of Methods
D. Amendment to 40 CFR 136.3, Table IA
V. Changes from the Proposed Rule
A. Proposed WET Method Changes
1. Blocking by Known Parentage
2. pH Drift
3. Nominal Error Rates
4. Dilution Series
5. Dilution Waters
6. Pathogen Interference
7. EDTA in the Selenastrum capricornutum Growth Test
B. Additional Revisions to WET Test Methods
1. Variability Criteria
2. Minimum Number of Replicates
3. Test Requirements/Recommendations
4. Sample Collection and Holding Times
5. Reference Toxicant Testing
6. Sample Holding Temperature
7. Biomass
8. Total Residual Chlorine
9. Ceriodaphnia dubia Survival and Reproduction Test Termination
Criteria
10. Additional Minor Corrections
C. Ratification and Withdrawal of Methods
VI. Response to Major Comments
A. Proposed WET Method Changes
1. Cost
2. Concentration-Response Relationships
3. Confidence Intervals
B. Additional Revisions to WET Test Methods
1. Method Flexibility
2. Test Acceptability Criteria
3. Quality Assurance/Quality Control Requirements
4. Statistical Methods
C. Ratification and Withdrawal of Methods
1. Validation of Performance Characteristics
2. Interlaboratory Variability Study
3. Variability
4. Successful Test Completion Rate
5. False Positive Rate
6. Implementation
VII. Statutory and Executive Order Reviews
A. Executive Order 12866: Regulatory Planning and Review
B. Paperwork Reduction Act
C. Regulatory Flexibility Act
D. Unfunded Mandates Reform Act
E. Executive Order 13132: Federalism
F. Executive Order 13175: Consultation and Coordination with
Indian Tribal Governments
G. Executive Order 13045: Protection of Children from
Environmental Health and Safety Risks
H. Executive Order 13211: Actions Concerning Regulations that
Significantly Affect Energy Supply, Distribution, or Use
I. National Technology Transfer Advancement Act
J. Congressional Review Act
VIII. References
I. General Information
A. Potentially Regulated Entities
EPA Regions, as well as States, Territories, and Tribes authorized
to implement the National Pollutant Discharge Elimination System
(NPDES) program, issue permits that comply with the technology-based
and water quality-based requirements of the Clean Water Act. In doing
so, NPDES permitting authorities make a number of discretionary choices
associated with permit writing, including the selection of pollutants
to be measured and, in many cases, limits for those pollutants in
permits. If EPA has ``approved'' (i.e., promulgated through rulemaking)
standardized test procedures for a given pollutant, the NPDES
permitting authority must specify one of the approved testing
procedures or an EPA-approved alternate test procedure for the
measurements required under the permit. In addition, when a State,
Territory, or authorized Tribe provides certification of Federal
licenses under Clean Water Act section 401, States, Territories and
Tribes are directed to use the approved testing procedures. Categories
and entities that may be regulated include:
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Examples of potentially regulated
Category entities
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Federal, State, Territorial, Federal, State, Territorial, and Tribal
and Indian Tribal entities authorized to administer the
Governments. NPDES permitting program; Federal,
State, Territorial, and Tribal entities
providing certification under Clean
Water Act section 401.
Municipalities............... Municipal operators of NPDES facilities
required to monitor whole effluent
toxicity.
Industry..................... Private operators of NPDES facilities
required to monitor whole effluent
toxicity.
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This table is not intended to be exhaustive, but rather provides a
guide for readers regarding entities likely to be regulated by this
action. This table lists the types of entities that EPA is now aware
could potentially be regulated by
[[Page 69953]]
this action. Other types of entities not listed in the table could also
be regulated. To determine whether your facility or organization is
regulated by this action you should carefully examine 40 CFR
122.41(j)(4), 122.44(i)(1)(iv), and 122.21. If you have questions
regarding the applicability of this action to a particular entity,
consult the first person listed in the preceding FOR FURTHER
INFORMATION CONTACT section.
B. How Can I Get Copies of Related Information?
1. Docket
EPA has established an official public docket for this action under
Docket ID No. WET-X (Electronic Docket No. OW-2002-0024). The official
public docket consists of the documents specifically referenced in this
action, any public comments received, and other information related to
this action. Although a part of the official docket, the public docket
does not include Confidential Business Information (CBI) or other
information whose disclosure is restricted by statute. The official
public docket is the collection of materials that is available for
public viewing at the Office of Water (OW) Docket, in the EPA Docket
Center (EPA/DC), EPA West, Room B-102, 1301 Constitution Avenue NW.,
Washington, DC. The EPA Docket Center Public Reading Room is open from
8:30 a.m. to 4:30 p.m. EST, Monday through Friday, excluding legal
holidays. The telephone number for the Public Reading Room is (202)
566-1744, and the telephone number for the OW Docket is (202) 566-2426.
2. Electronic Access
You may access this Federal Register document electronically
through the EPA Internet under the ``Federal Register'' listings at
http://www.epa.gov/fedrgstr/.
An electronic version of the public docket is available through
EPA's electronic public docket and comment system, EPA Dockets. You may
use EPA Dockets at http://www.epa.gov/edocket/ to view public comments,
access the index listing of the contents of the official public docket,
and to access those documents in the public docket that are available
electronically. Although not all docket materials may be available
electronically, you may still access any of the publicly available
docket materials through the docket facility identified in Unit I.B.1.
Once in the system, select ``search,'' then key in the appropriate
docket identification number.
II. Statutory Authority
EPA promulgates today's rule pursuant to the authority of sections
301, 304(h), 402, and 501(a) of the Clean Water Act (``CWA'' or the
``Act''), 33 U.S.C. 1311, 1314(h), 1342, 1361(a) (the ``Act''). Section
101(a) of the Act sets forth the ``goal of restoring and maintaining
the chemical, physical, and biological integrity of the nation's
waters'' and prohibits ``the discharge of toxic pollutants in toxic
amounts.'' Section 301 of the Act prohibits the discharge of any
pollutant into navigable waters unless the discharge complies with a
National Pollutant Discharge Elimination System (NPDES) permit, issued
under section 402 of the Act. Section 304(h) of the Act requires the
Administrator of the EPA to ``promulgate guidelines establishing test
procedures for the analysis of pollutants that shall include the
factors which must be provided in any certification pursuant to section
401 of this Act or permit applications pursuant to section 402 of this
Act.'' Section 501(a) of the Act authorizes the Administrator to
``prescribe such regulations as are necessary to carry out his function
under this Act.'' EPA publishes CWA analytical method regulations at 40
CFR part 136. The Administrator also has made these test procedures
applicable to monitoring and reporting of NPDES permits (40 CFR part
122, Sec. Sec. 122.21, 122.41, 122.44, and 123.25), and implementation
of the pretreatment standards issued under section 307 of the Act (40
CFR part 403, Sec. Sec. 403.10 and 403.12).
III. Background
A. Regulatory History
On October 16, 1995, EPA amended the ``Guidelines Establishing Test
Procedures for the Analysis of Pollutants,'' 40 CFR part 136, to add a
series of standardized toxicity test methods to the list of Agency
approved methods for conducting required testing of aqueous samples
under the CWA (60 FR 53529) (WET final rule). The WET final rule
amended 40 CFR 136.3 (Tables IA and II) by adding acute toxicity
methods and short-term methods for estimating chronic toxicity. These
methods measure the toxicity of effluents and receiving waters to
freshwater, marine, and estuarine organisms. Acute methods (USEPA,
1993) generally use death of some percentage of the test organisms
during 24 to 96 hour exposure durations as the measured effect of an
effluent or receiving water. The short-term methods for estimating
chronic toxicity (USEPA, 1994a; USEPA, 1994b) use longer durations of
exposure (up to nine days) to ascertain the adverse effects of an
effluent or receiving water on survival, growth, and/or reproduction of
the organisms. The methods listed at 40 CFR part 136 for measuring
aquatic toxicity are referred to collectively as ``WET test methods,''
methods specific to measuring acute toxicity are referred to as
``acute'' test methods, and short-term methods for estimating chronic
toxicity are referred to as ``chronic'' methods.
EPA standardized the test procedures for conducting the approved
acute and chronic WET test methods in the following three method
manuals, which were incorporated by reference in the WET final rule:
Methods for Measuring the Acute Toxicity of Effluents and Receiving
Water to Freshwater and Marine Organisms, Fourth Edition, August 1993,
EPA/600/4-90/027F (acute method manual); Short-Term Methods for
Estimating the Chronic Toxicity of Effluents and Receiving Water to
Freshwater Organisms, Third Edition, July 1994, EPA/600/4-91/002
(freshwater chronic method manual); and Short-Term Methods for
Estimating the Chronic Toxicity of Effluents and Receiving Water to
Marine and Estuarine Organisms, Second Edition, July 1994, EPA/600/4-
91/003 (marine chronic method manual). EPA explains in the Technical
Support Document for Water Quality-Based Toxics Control (TSD) (USEPA,
1991) that these WET test methods, along with chemical controls and
bioassessments, are a component of EPA's integrated strategy for water
quality-based toxics control. The TSD recommends that WET tests using
the most sensitive of at least three test species from different phyla
be used for monitoring the toxicity of effluents.
Since the 1995 WET final rule, EPA has issued several rulemakings
and guidance documents in fulfillment of settlement agreements to
resolve judicial challenges to the WET final rule (see Settlement
Agreement discussion in Section III.B). On February 2, 1999, EPA
published technical corrections that incorporated into the WET final
rule an errata document to correct minor errors and omissions, provide
clarification, and establish consistency among the WET final rule and
method manuals (64 FR 4975; February 2, 1999). On July 18, 2000, EPA
announced the availability of a WET Variability Guidance Document (65
FR 44528; July 18, 2000). On July 28, 2000, EPA published the
availability of a WET Method Guidance Document (65 FR 46457; July 28,
2000). On September 28, 2001, EPA proposed specific revisions to the
WET test methods, and EPA proposed to ratify its previous
[[Page 69954]]
approval of these methods (66 FR 49794; September 28, 2001) (see
section III.C). Today, EPA takes final action on the September 2001
proposal.
B. Settlement Agreement
Following promulgation of the WET methods on October 16, 1995,
several parties challenged the rulemaking (Edison Electric Institute v.
EPA, No. 96-1062 (D.C. Cir.); Western Coalition of Arid States v. EPA,
No. 96-1124 (D.C. Cir.); and Lone Star Steel Co. v. EPA, No. 96-1157
(D.C. Cir.)). To resolve the litigation, EPA entered into settlement
agreements with the various parties and agreed to publish a technical
correction notice, publish a method guidance document and a variability
guidance document, conduct an interlaboratory variability study,
publish a peer-reviewed interlaboratory variability study report
(including a table of coefficients of variation), address pathogen
contamination, propose specific technical method changes, and propose
to ratify or withdraw WET test methods evaluated in the interlaboratory
variability study. Today's final action fulfills EPA's obligations
under the settlement agreements.
C. Proposed Rule
On September 28, 2001, EPA proposed modifications to the WET test
methods (66 FR 49794). The proposal included updates to the methods,
minor corrections and clarifications, and specific technical changes in
response to stakeholder concerns. Specifically, EPA proposed technical
changes to (1) require ``blocking'' by known parentage in the
Ceriodaphnia dubia Survival and Reproduction Test; (2) specify
procedures to control pH drift that may occur during testing; (3)
incorporate review procedures for the evaluation of concentration-
response relationships; (4) clarify recommendations regarding nominal
error rate assumptions; (5) clarify limitations in the generation of
confidence intervals; (6) add guidance on dilution series selection;
(7) clarify requirements regarding acceptable dilution waters; and (8)
add procedures for determining and minimizing the adverse impact of
pathogens in the Fathead Minnow Survival and Growth Test.
EPA also solicited comment on other modifications to improve the
performance of the methods, including the incorporation of variability
criteria and increases in the minimum number of test replicates. EPA
proposed to incorporate WET method changes into new editions of each of
the WET method manuals (USEPA, 1993; USEPA, 1994a; USEPA, 1994b) and to
update Table IA at 40 CFR part 136 to cite the new method manual
editions.
In the September 28, 2001 proposed rule, EPA also proposed to
ratify 11 of the 12 WET methods evaluated in EPA's WET Interlaboratory
Variability Study. EPA proposed to ratify the Ceriodaphnia dubia Acute
Test; Fathead Minnow Acute Test; Sheepshead Minnow Acute Test; Inland
Silverside Acute Test; Ceriodaphnia dubia Survival and Reproduction
Test; Fathead Minnow Larval Survival and Growth Test; Selenastrum
capricornutum Growth Test; Sheepshead Minnow Larval Survival and Growth
Test; Inland Silverside Larval Survival and Growth Test; Mysidopsis
bahia Survival, Growth, and Fecundity Test; and Champia parvula
Reproduction Test. To support ratification of these methods, EPA
presented the results of the WET Interlaboratory Variability Study
(USEPA, 2001a; USEPA, 2001b), a national study of 12 WET methods
involving 56 laboratories and over 700 samples. EPA proposed to
withdraw Holmesimysis costata as an acceptable substitute species for
use in the Mysidopsis bahia Acute Test method protocol. In its place,
EPA proposed a new Holmesimysis costata Acute Test protocol.
EPA invited public comment for 60 days and later extended the
comment period for an additional 45 days (66 FR 58693; November 23,
2001). EPA received 38 comment packages during the allotted comment
period.
IV. Summary of Final Rule
A. Proposed WET Method Changes
Today's action incorporates most of the method changes proposed on
September 28, 2001 (66 FR 49794) with minor modifications to address
public comments. For a summary of major changes from the proposed rule,
including proposed actions not incorporated in today's rule, see
Section V of this preamble. Method manual revisions promulgated in
today's action include:
• Minor corrections and clarifications,
• Incorporation of updated method precision data,
• Requirement for ``blocking'' by known parentage in the
Ceriodaphnia dubia Survival and Reproduction Test,
• Specification of procedures to control pH drift that may
occur during testing,
• Review procedures for the evaluation of concentration-
response relationships,
• Clarification of limitations in the generation of
confidence intervals,
• Guidance on dilution series selection,
• Clarification of requirements regarding acceptable dilution
waters,
• Procedures for determining and minimizing the adverse
impact of pathogens in the Fathead Minnow Survival and Growth Test,
• Requirement for the use of ethylenediaminetetraacetic acid
(EDTA) in the Selenastrum capricornutum Growth Test.
B. Additional Revisions to WET Test Methods
In addition to requesting comment on the specific modifications to
WET test methods mentioned above, EPA solicited comment on any
additional modifications that would improve the overall performance of
the methods. Specifically, EPA solicited comment on application of
variability criteria to test results, modification of test
acceptability criteria, and increases in test replication requirements.
In response to comments, today's final rule also incorporates the
following additional modifications to WET test methods:
• Requirement to meet specific variability criteria when
NPDES permits require sublethal WET testing endpoints expressed using
hypothesis testing,
• Increases in the required minimum number of replicates for
several tests,
• Clarification of required and recommended test conditions
for the purposes of reviewing WET test data submitted under NPDES
permits,
• Additional clarification of sample holding times,
• Clarification of requirements for reference toxicant
testing and additional guidance on evaluating reference toxicant test
results,
• Clarification of allowable sample holding temperatures,
• Clarification of biomass as the measured endpoint in
survival and growth tests,
• Clarification of requirements for measuring total residual
chlorine in WET samples,
• Modification of the test termination criteria for the
Ceriodaphnia dubia Survival and Reproduction Test to exclude the
counting of fourth brood neonates,
• Additional minor corrections identified by commenters.
C. Ratification and Withdrawal of Methods
Based on the WET Interlaboratory Variability Study, peer review
comments, and comments on the proposed rule, EPA is ratifying ten
methods evaluated in the WET
[[Page 69955]]
Interlaboratory Variability Study and withdrawing two methods. EPA is
ratifying the Ceriodaphnia dubia Acute Test; Fathead Minnow Acute Test;
Sheepshead Minnow Acute Test; Inland Silverside Acute Test;
Ceriodaphnia dubia Survival and Reproduction Test; Fathead Minnow
Larval Survival and Growth Test; Selenastrum capricornutum Growth Test;
Sheepshead Minnow Larval Survival and Growth Test; Inland Silverside
Larval Survival and Growth Test; and Mysidopsis bahia Survival, Growth,
and Fecundity Test. In accordance with EPA's Report to Congress on the
Availability, Adequacy, and Comparability of testing procedures (USEPA,
1988), EPA has confirmed that the methods ratified today are repeatable
and reproducible (i.e., exhibit adequate within-laboratory and between-
laboratory precision), available and applicable (i.e., adaptable to a
wide variety of laboratories and use widely available organisms and
supplies), and representative (i.e., predictive of receiving system
impacts). See section VI.C.1 of this preamble.
EPA's WET Interlaboratory Variability Study demonstrated that the
methods ratified today generally have a high rate of successful
completion, do not often produce false positive results, and exhibit
precision comparable to chemical methods approved at 40 CFR part 136.
Table 1 summarizes the performance characteristics for the ten WET test
methods ratified today. In ratifying these WET test methods, EPA
reaffirms the conclusion expressed in the 1995 WET final rule (60 FR 53529; October 16, 1995), that these methods, including the
modifications in today's rule, are applicable for use in NPDES permits.
Table 1.--Summary of Performance Characteristics for Ratified WET Methods
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Successful Interlaboratory
Test method test completion False positive precision (%CV)
rate (%) rate \a\ (%) \b\
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Ceriodaphnia dubia Acute Test................................ 95.2 0.00 29.0
Ceriodaphnia dubia Survival and Reproduction Test............ 82.0 3.70 35.0
Fathead Minnow Acute Test.................................... 100 0.00 20.0
Fathead Minnow Larval Survival and Growth Test............... 98.0 4.35 20.9
Selenastrum capricornutum Growth Test........................ 63.6 0.00 34.3
Mysidopsis bahia Survival, Growth, and Fecundity Test........ 97.7 0.00 41.3
Sheepshead Minnow Acute Test................................. 100 0.00 26.0
Sheepshead Minnow Larval Survival and Growth Test............ 100 0.00 10.5
Inland Silverside Acute Test................................. 94.4 0.00 38.5
Inland Silverside Larval Survival and Growth Test............ 100 0.00 43.8
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\a\ False positive rates reported for each method represent the higher of false positive rates observed for
hypothesis testing or point estimate endpoints.
\b\ Coefficients of variation (CVs) reported for each method represent the CV of LC50 values for acute test
methods and IC25 values for chronic test methods. CVs reported are based on total interlaboratory variability
(including within-laboratory and between-laboratory components of variability) and averaged across sample
types.
EPA is withdrawing the Holmesimysis costata Acute Test and the
Champia parvula Reproduction Test methods from 40 CFR part 136. EPA was
unable to obtain interlaboratory precision data for these methods in
the WET Interlaboratory Variability Study due to laboratory
unavailability. EPA was unable to contract with a minimum of six
laboratories qualified and willing to conduct these test methods within
the time frame of the Study. Due to this lack of interlaboratory
precision data generated from the Study for these methods, several
commenters recommended that these methods not be approved at 40 CFR
part 136 for national use. In response, today's action removes the
Holmesimysis costata Acute Test method (1995 version) and the Champia
parvula Reproduction Test method from the list of test methods approved
for nationwide use at 40 CFR part 136.
By withdrawing these methods from 40 CFR part 136 for nationwide
use, EPA does not reject their use on more limited bases. Today's
withdrawal simply reflects that the Agency has not validated these
methods for national use. EPA continues to support the use of these
methods for applications other than for the determination of compliance
with NPDES permit limits, as well for limited, localized, or regional
use where the methods have been validated by other entities. In
addition, EPA continues to support the use of the Holmesimysis costata
Acute Test to measure toxicity to marine organisms of the Pacific
Ocean. Because test procedures for measuring toxicity to estuarine and
marine organisms of the Pacific Ocean are not listed at 40 CFR part
136, permit writers may include (under 40 CFR 122.41(j)(4) and
122.44(i)(1)(iv)) requirements for the use of test procedures that are
not approved at part 136, such as the Holmesimysis costata Acute Test
and other West Coast WET methods (USEPA, 1995b) on a permit-by-permit
basis.
D. Amendment to 40 CFR 136.3, Table IA
Today's rule amends 40 CFR 136.3 by removing the Champia parvula
Reproduction Test method (Method 1009.0) from Table IA, modifying the
reference to acute ``mysid'' tests in Table IA to include only
Mysidopsis bahia (and not Holmesimysis costata), adding method numbers
to acute tests, revising the parameter measured in marine tests to
refer to organisms ``of the Atlantic Ocean and Gulf of Mexico,'' and
modifying footnotes and references to cite the updated versions of the
method manuals.
V. Changes From the Proposed Rule
A. Proposed WET Method Changes
On September 28, 2001, EPA proposed technical method changes to
improve the performance and clarity of WET test methods and to address
specific stakeholder concerns. These provisions were presented and
discussed in section III of the proposed rule preamble (66 FR 49794)
and detailed in the document titled, Proposed Changes to Whole Effluent
Toxicity Method Manuals (USEPA, 2001e). In today's action, EPA is
withdrawing or revising some of the proposed revisions based on
comments received on the proposed rule. These revisions are discussed
below. Other comments that EPA addressed but did not result in changes
from the proposal are discussed in section VI.
[[Page 69956]]
1. Blocking by Known Parentage
EPA proposed specific method manual modifications that would
require blocking by known parentage in the Ceriodaphnia dubia Survival
and Reproduction Test method. Today, EPA is finalizing the proposed
method changes with a minor modification to clarify that neonates from
a single known parent may be used in the initiation of more than one
test. This minor modification mitigates some commenters' concerns
regarding the increased cost of blocking by known parentage. Blocking
by known parentage requires the use of at least six neonates from each
of at least ten separate parents. If more than six neonates from a
given parent remain after allocating organisms to a test, those
remaining neonates may be discarded, used as future culture organisms,
or used in another test initiated on the same day (provided that the
neonates meet age requirements).
2. pH Drift
During the conduct of static or static-renewal WET tests, the pH in
test containers may fluctuate or drift from the initial pH value. EPA
proposed specific procedures that may be used to control this pH drift
in chronic WET tests. Today, EPA is revising the specified procedures
in response to stakeholder comments. Some commenters requested that EPA
clarify the pH that should be maintained in pH-controlled tests.
Today's action clarifies that, when the test objective is to determine
the toxicity of an effluent in the receiving water, the target pH to
maintain in a pH-controlled test is the pH of the receiving water
measured at the edge of any mixing zone authorized in a permit. When
the test objective is to determine the absolute toxicity of the
effluent, the target pH to maintain in a pH-controlled test is the pH
of the sample upon completion of collection. The revisions also clarify
that in pH-controlled tests, the pH should be maintained within +/-0.2
pH units of the target pH in freshwater chronic tests and within +/-0.3
pH units for marine/estuarine chronic tests. EPA also added guidance on
interpreting the results of parallel testing.
The revisions also remove language from the proposed method manual
changes that warned about effects from pH drift in the absence of pH-
dependent toxicants. To address the concern that the daily cycle of pH
drift and renewal caused artifactual toxicity by ``shocking'' test
organisms, EPA proposed language in the method manuals that warned of
such potential interference from pH drift even when pH-dependent
toxicants were not present. EPA specifically requested that commenters
provide ``any data that show the value of proposed pH control measures
in situations where ammonia or other pH-dependent toxicants are not
present.'' EPA did not receive such data. EPA believes that pH drift
alone is not a test interference if pH is within the organism's
tolerance range. The degree of pH drift typically observed in effluent
samples should generally only interfere with test results if the sample
contains a compound with toxicity that is pH dependent and at a
concentration that is near the toxicity threshold. Because EPA did not
receive data to suggest otherwise, EPA is removing any reference to pH
drift interference in the absence of pH-dependent toxicants.
Many commenters recommended that EPA include the proposed pH
control guidance for acute test methods as well as chronic methods
because of the insufficiency of static renewal testing to control the
pH drift and the impracticability and cost of flowthrough testing. In
today's action, EPA has not provided additional techniques that involve
modification of the sample to control pH drift in acute test methods,
because EPA believes that the current acute methods provide adequate
remedies for pH drift without modifying the sample. In acute tests, pH
drift may be remedied by more frequent test renewals or use of
flowthrough testing. While EPA agrees that flowthrough testing is more
costly than static or static renewal testing, today's action does not
impose any additional costs by requiring flowthrough testing. Today's
action simply retains the options for pH control that are currently
described in the acute method manual and does not add additional
options.
3. Nominal Error Rates
Today's action does not incorporate the proposed method manual
changes regarding nominal error rates. The method manuals maintain the
original statement recommending a nominal error rate of 0.05. EPA
proposed changes to its recommendation regarding nominal error rate
assumptions, specifically, the change from 0.05 to 0.01 under specific
circumstances. EPA proposed changes to its recommended error rate
assumptions based on the settlement agreement, which identified the
circumstances under which EPA would change its recommendations
regarding nominal error rate reductions. These specified circumstances
do not necessarily represent cases where the risk of false positive
results increase, but rather situations for which the petitioners
sought specific relief.
Commenters on the proposed rule commented that there was no
scientific justification for reducing nominal error rate assumptions in
only these circumstances and recommended reducing the nominal error
rate in all circumstances. EPA agrees with the commenters that there is
not a scientific justification for allowing reduced nominal error rates
in these specific circumstances, but disagrees that nominal error rates
should be reduced in all circumstances. Some commenters claimed that a
reduced nominal error rate is needed to improve confidence in the test
results. Reducing the nominal error rate, however, does not inherently
improve confidence in test results. Because of the relationship between
Type I and Type II statistical errors, reductions in nominal error
rates improve confidence in results that identify toxicity, but reduce
confidence in results that do not identify toxicity. This reduces the
power of the test and the chance of identifying toxic discharges,
thereby reducing environmental protection. In addition, the statistical
test designs (i.e., test replication requirements) of WET methods and
all supporting method validation data were based on a nominal error
rate of 0.05. Because there is no scientific justification for
recommending reductions in nominal error rates in the circumstances
proposed and commenters did not provide such supporting rationale or
data, EPA has not incorporated the proposed method manual
recommendations regarding nominal error rates. The method manuals
maintain the original recommendation to assume a nominal error rate of
0.05.
4. Dilution Series
EPA is finalizing the proposed guidance on the selection of
dilution series in WET testing. In addition to the proposed guidance,
EPA has made minor modifications in response to comments to further
clarify that no one particular dilution series is required. Specific
dilution series used in the WET method manuals are provided as examples
and recommendations, not requirements.
5. Dilution Waters
EPA is finalizing the proposed guidance on the selection of
dilution waters in WET testing. In addition to the proposed guidance,
EPA has made minor modifications in response to comments to further
clarify that no single dilution water type is required for all tests.
The method manuals now
[[Page 69957]]
clarify that receiving waters, synthetic waters, or synthetic waters
adjusted to approximate receiving water characteristics may be used for
dilution water, provided that the water meets the qualifications for an
acceptable dilution water. EPA clarified in the method manuals that an
acceptable dilution water is one which is appropriate for the
objectives of the test; supports adequate performance of the test
organisms with respect to survival, growth, reproduction, or other
responses that may be measured in the test (i.e., consistently meets
test acceptability criteria for control responses); is consistent in
quality; and does not contain contaminants that could produce toxicity.
EPA also provided clarification on the use of dual controls. When using
dual controls, the dilution water control should be used for
determining the acceptability of the test and for comparisons with the
tested effluent. If test acceptability criteria (e.g., minimum
survival, reproduction, or growth) are not met in the dilution water
control, the test must be repeated on a newly collected sample.
Comparisons between responses in the dilution water control and in the
culture water control can be used to determine if the dilution water,
which may be a receiving water, possesses ambient toxicity.
6. Pathogen Interference
In today's action, EPA finalizes the proposed guidance on
controlling pathogen interference in the Fathead Minnow Larval Survival
and Growth Test with several modifications to address commenter
concerns. Some commenters were concerned that the proposed guidance
allowed the use of pathogen control techniques such as UV,
chlorination, filtration, and antibiotics only after the recommended
modified test design (fewer fish per cup) failed to control pathogen
interference. Today's revisions clarify that EPA recommends pathogen
control techniques that do not modify the sample, such as the modified
test design technique, over ones that do. Upon approval by the
regulatory authority, however, analysts also may use various sample
sterilization techniques that modify the sample to control pathogen
interference, provided that parallel testing of altered and unaltered
samples further confirms the presence of pathogen interference and
demonstrates successful pathogen control.
The manuals also now provide further explanation regarding the
purpose for and required extent of pathogen source determination.
Commenters were concerned that EPA was requiring permittees to generate
data that was irrelevant to correcting for pathogen test interference.
This is not the case. Determining whether tests are adversely affected
by pathogens in the effluent or pathogens in the receiving water used
for test dilution is an important first step in selecting an
appropriate pathogen control technique. If the source of interfering
pathogens in the test is the receiving water used as the dilution
water, then pathogen interference may be controlled by simply using an
alternative dilution water. If the source of interfering pathogens in
the test is the effluent, then pathogen control techniques are
appropriate to control the interference. To further address the
comments, EPA removed mention of pathogen source identification beyond
determining whether the pathogen source was the effluent or dilution
water. EPA also made several minor modifications in response to
comments, including an acknowledgment that pathogen control techniques
may not eliminate pathogens, but should minimize the adverse influence
of pathogens so that test results are not confounded by mortality due
to pathogens.
7. EDTA in the Selenastrum capricornutum Growth Test
In the WET Interlaboratory Variability Study, EPA found that
performance of the Selenastrum capricornutum Growth Test was much
higher (lower interlaboratory variability and lower false positive
rate) when the test was conducted with EDTA (ethylenediaminetetraacetic
acid). Based on this finding, EPA proposed to recommend the use of EDTA
in the Selenastrum capricornutum Growth Test. Several commenters
expressed concern that EPA only recommended, rather than required, the
use of EDTA. Commenters stated that this recommendation was not
sufficient to ensure the acceptable performance of the method and
encouraged EPA to require the use of EDTA. To address these comments,
the Selenastrum capricornutum Growth Test now requires the addition of
EDTA to nutrient stock solutions when conducting the Selenastrum
capricornutum Growth Test and submitting data under NPDES permits. To
address concerns that EDTA may interfere with (i.e., mask) the toxicity
of metals, the method continues to caution that the addition of EDTA
may cause the Selenastrum capricornutum Growth Test to underestimate
the toxicity of metals. EPA cautions regulatory authorities to consider
this possibility when selecting test methods for monitoring effluents
that are suspected to contain metals. As recommended in EPA's Technical
Support Document for Water Quality-Based Toxics Control (TSD) (USEPA,
1991), the most sensitive of at least three test species from different
phyla should be used for monitoring the toxicity of effluents.
B. Additional Revisions to WET Test Methods
1. Variability Criteria
Today's action incorporates mandatory variability criteria for five
chronic test methods. EPA recommends the use of point estimation
techniques over hypothesis testing approaches for calculating endpoints
for effluent toxicity tests under the NPDES Permitting Program.
However, to reduce the within-test variability and to increase
statistical sensitivity when test endpoints are expressed using
hypothesis testing rather than the preferred point estimation
techniques, variability criteria must be applied as a test review step
when NPDES permits require sublethal hypothesis testing endpoints
(i.e., no observed effect concentration (NOEC) or lowest observed
effect concentration (LOEC)) and the effluent has been determined to
have no toxicity at the permitted receiving water concentration These
variability criteria must be applied for the following methods: Fathead
Minnow Larval Survival and Growth Test; Ceriodaphnia dubia Survival and
Reproduction Test; Selenastrum capricornutum Growth Test; Mysidopsis
bahia Survival, Growth, and Fecundity Test; and Inland Silverside
Larval Survival and Growth Test. Within-test variability, measured as
the percent minimum significant difference (PMSD), must be calculated
and compared to upper bounds established for test PMSDs. Under this new
requirement, tests conducted under NPDES permits that fail to meet the
variability criteria (i.e., PMSD upper bound) and show ``no toxicity''
at the permitted receiving water concentration (i.e., no significant
difference from the control at the receiving water concentration or
above) are considered invalid and must be repeated on a newly collected
sample. Lower bounds on the PMSD are also applied, such that test
concentrations shall not be considered toxic (i.e., significantly
different from the control) if the relative difference from the control
is less than the lower PMSD bound.
In the proposed rule, EPA solicited comment on the required use of
upper and lower PMSD bounds in the calculation of NOEC and LOEC values.
[[Page 69958]]
According to the proposed approach, any test treatment with a
percentage difference from the control (i.e., [mean control response--
mean treatment response]/ mean control response * 100) that is greater
than the upper PMSD bound would be considered as significantly
different; and any test treatment with a percentage difference from the
control that is less than the lower PMSD bound would not be considered
as significantly different.
EPA received comments on this proposed approach that expressed
concern that variability criteria were used only to adjust NOEC and
LOEC values and not to invalidate tests. Commenters argued that the
proposed approach does not control variability unless tests failing to
meet the variability criteria are invalidated. In response to these
comments, EPA has modified the application of variability criteria in
today's action. Rather than implementing variability criteria as a
component of endpoint calculation, today's method modifications
implement variability criteria (upper and lower PMSD bounds) as a test
review step that is required when NPDES permits require sublethal WET
testing endpoints expressed using hypothesis testing for the five test
methods previously listed. Reviewed tests that fail to meet the
variability criteria and do not detect toxicity at the receiving water
concentration are invalid and must be repeated on a newly collected
sample.
EPA received comments both for and against implementation of
variability criteria as test acceptability criteria. To balance these
comments, the final rule implements the variability criteria as a
required test review step when NPDES permits require sublethal WET
testing endpoints expressed using hypothesis testing for the five test
methods previously listed. As such, the variability criteria have the
potential to invalidate highly variable tests. Invalidation, however,
is contingent upon other data evaluation steps. For instance, tests
that exceed the variability criteria are only invalidated when the test
also fails to detect toxicity at the permitted receiving water
concentration. The method manuals continue to restrict use of the term
``test acceptability criteria'' to biological measurements in test
controls (i.e., control survival, reproduction, and growth) that
independently assess test acceptability. Unlike the variability
criteria instituted today, the use of ``test acceptability criteria''
to invalidate tests are not contingent on any other data evaluation
steps. For this reason, the term ``test acceptability criteria'' is not
applicable to the variability criteria established in today's action.
EPA received comments that recommended alternative measures for
controlling within test variability, such as limits on the coefficient
of variation (CV) for the control treatment. In developing variability
criteria, EPA considered other measures of test precision, including
the standard deviation and coefficients of variation for treatments and
control, minimum significant difference (MSD), and the mean square for
error from the analysis of variance of treatment effects. EPA considers
the PMSD to be the measure that is most easily understood and that is
most directly applied to determination of NOEC and LOEC values. The
PMSD quantifies the smallest percentage difference between the control
and a treatment (effluent dilution) that could be declared as
statistically significant. It thus includes exactly that variability
affecting determination of the NOEC and LOEC. The CV for the control or
any one treatment, or selected treatments, represents only a portion of
the variability affecting the NOEC and LOEC. Some State or Regional WET
programs have requirements on the CV for the control and the treatment
representing the receiving water concentration (RWC). Such requirements
can provide finer control over the variability influencing a single
comparison between the control and the RWC treatment. The PMSD upper
bound provides control over the total within-test variability and is
intended specifically for multi-concentration tests in which the NOEC
or LOEC are determined by using hypothesis testing. Regulatory
authorities may continue to use variability control strategies adopted
within their jurisdiction, but when NPDES permits require sublethal WET
testing endpoints expressed using hypothesis testing, the variability
criteria required by today's action must be implemented as well.
Requiring such variability criteria provides national consistency and
control of WET test precision when hypothesis testing approaches are
chosen. In today's action, EPA reiterates the recommendation of the
method manuals and the TSD (USEPA, 1991) by stating that for the NPDES
Permit Program, point estimation techniques are preferred over
hypothesis testing approaches for calculating endpoints for effluent
toxicity tests.
EPA received comments that the upper and lower bounds established
for PMSD variability criteria were arbitrary or unrepresentative. EPA
established the proposed variability criteria as performance-based
standards set at the 10th and 90th percentiles of PMSD values from
EPA's evaluation of national reference toxicant test data (USEPA,
2000c). In today's action, EPA has revised the variability criteria to
reflect the 10th and 90th percentiles of PMSD values based on EPA's
Interlaboratory Variability Study. The use of data from this study
reflects not only tests performed on reference toxicants, but tests
performed on effluents, receiving waters, and non-toxic ``blank''
samples as well. Data from this study also is representative of
qualified laboratories that routinely conduct WET testing for
permittees (see Section VI.C.2 of this preamble). In method
development, EPA routinely uses such data from interlaboratory
validation studies to set performance-based criteria.
In September 2001, EPA proposed variability criteria for four
methods. Some commenters recommended that EPA expand the variability
criteria to other test methods and other test endpoints. EPA did not
propose variability criteria for the Selenastrum capricornutum Growth
Test and the Sheepshead Minnow Larval Survival and Growth Test because
these methods showed lower within-test variability in EPA's evaluation
of national reference toxicant test data (USEPA, 2000c). EPA's WET
Interlaboratory Variability Study confirmed that the Sheepshead Minnow
Larval Survival and Growth Test was less variable than the methods for
which EPA proposed variability criteria, however, the Selenastrum
capricornutum Growth Test showed comparable within-test variability to
methods for which EPA proposed variability criteria. For this reason,
EPA is today requiring variability criteria for the Selenastrum
capricornutum Growth Test in addition to the four methods for which
variability criteria were proposed.
As previously stated in the method manuals (USEPA, 1993; USEPA,
1994a; USEPA, 1994b) and EPA's Technical Support Document (USEPA,
1991), EPA recommends the use of point estimation techniques over
hypothesis testing approaches for calculating endpoints for effluent
toxicity tests under the NPDES Permitting Program. EPA is instituting
variability criteria to reduce within-test variability and to increase
statistical sensitivity when test endpoints are expressed using
hypothesis testing rather than the preferred point estimation
techniques. For the five methods for which EPA is instituting
variability criteria when test results are analyzed by hypothesis test
methods, less than 90% of tests are able to detect
[[Page 69959]]
a 25% reduction in growth or reproduction (from the control treatment)
as statistically significant using the hypothesis test. A 25% reduction
in growth or reproduction is equivalent to the effect level measured
using the preferred point estimation endpoint for chronic methods
(i.e., the IC25). Instituting variability criteria for these five
chronic methods will improve the overall statistical sensitivity when
using hypothesis testing and allow hypothesis testing approaches to
achieve a level of statistical sensitivity that is more comparable to
the preferred point estimation endpoint (IC25).
EPA is not requiring variability criteria for the Sheepshead Minnow
Larval Survival and Growth Test, because the WET Interlaboratory
Variability Study confirmed that this method is less variable than the
five methods for which EPA is requiring variability criteria. In EPA's
WET Interlaboratory Variability Study, all Sheepshead Minnow Larval
Survival and Growth Tests were able to detect effects of 25% or less as
statistically significant in hypothesis testing without instituting
variability criteria. The 90th percentile PMSD for the Sheepshead
Minnow Larval Survival and Growth Test was 17%, compared to 29%, 47%,
30%, 37%, and 28% for the five methods for which EPA is requiring
variability criteria. For the chronic methods that were not evaluated
in the WET Interlaboratory Variability Study, EPA does not have
sufficient data to support the implementation of mandatory variability
criteria at this time.
EPA is not requiring variability criteria for survival endpoints of
acute methods because, in general, these methods are less variable than
sublethal chronic test methods, and hypothesis testing approaches are
able to achieve a level of statistical sensitivity similar to the
preferred point estimation endpoint for acute methods and survival
endpoints (i.e., the LC50). The preferred point estimation endpoint for
the analysis of survival in acute methods is the LC50, which represents
an effect level of 50% mortality. Over 90% of acute tests in the WET
Interlaboratory Variability Study were able to detect effects of 50%
mortality or less as statistically significant in hypothesis testing
without instituting variability criteria. The 90th percentile of PMSD
values in the WET Interlaboratory Variability Study was 39% for the
Fathead Minnow Acute Test, 25% for the Ceriodaphnia dubia Acute Test,
17% for the Sheepshead Minnow Acute Test, and 31% for the Inland
Silverside Acute Test. Based on these measured PMSD values, well over
90% of acute tests should be able to detect effects at the LC50 as
statistically significant without instituting variability criteria.
By requiring application of variability criteria today in five
methods, EPA does not intend to discourage permitting authorities from
applying variability criteria for other endpoints or methods, or from
applying more stringent variability criteria for the five chronic
methods subject to today's action. While EPA continues to recommend
that permitting authorities apply variability criteria to additional
methods as recommended in EPA guidance (USEPA, 2000c), today's rule
does not require such variability criteria for additional methods or
endpoints.
2. Minimum Number of Replicates
EPA solicited comment on increasing the minimum number of
replicates in certain WET tests from three to four. Commenters were
supportive of this proposed change and stated that this change was
needed to support the use of non-parametric hypothesis tests as
outlined in the method manuals. In today's action, EPA is increasing
the minimum number of replicates as proposed.
3. Test Requirements/Recommendations
Several commenters on the proposed rule expressed concern that WET
methods do not adequately differentiate between mandatory test
conditions (i.e., those required using the words ``must'' or ``shall'')
and discretionary test conditions (i.e., those recommended using the
word ``should''). Commenters claimed that this situation causes
difficulty in reviewing, validating, and certifying test results
submitted under NPDES permits. To address this concern, EPA modified
the WET methods to clearly distinguish between required and recommended
test conditions for the purposes of reviewing WET test data submitted
under NPDES permits. In today's action, EPA has modified the tables of
test conditions and test acceptability criteria presented in the method
manuals for each method, such that each test condition is identified as
required or recommended. In addition, EPA has added to each method
manual a section on test review. This section provides guidance on the
review of sampling and handling procedures, test acceptability
criteria, test conditions, statistical methods, concentration-response
relationships, reference toxicant testing, and test variability. This
section also establishes two new requirements for WET test review:
mandatory review of concentration-response relationships and, for some
methods, the mandatory variability criteria described earlier.
4. Sample Collection and Holding Times
In today's action, EPA has further clarified the requirements for
sample collection and sample holding times. EPA made these
modifications in response to comments requesting additional
clarification and additional flexibility. In today's action, EPA has
not modified the default maximum 36 hour sample holding time (up to 72
hours with regulatory authority approval), which must be met for first
use of the sample, but EPA has provided additional clarification and
additional flexibility for the use of samples for test renewals when
the samples meet the initial sample holding times for first use. Sample
holding times apply to ``first use of the sample,'' and samples may be
used for renewal at 24, 48, and/or 72 hours after first use.
The method manuals also now provide additional flexibility when
shipment of renewal samples is delayed during an ongoing test. If
shipping problems (e.g., unsuccessful Saturday delivery) are
encountered with renewal samples after a test has been initiated, the
permitting authority may allow the continued use of the most recently
used sample for test renewal. EPA also clarified that sample collection
on days one, three, and five is the recommended (not required) sample
collection scheme. A minimum of three samples are required for seven-
day chronic tests, but variations in the sampling scheme (i.e., the
days on which new samples are collected) also are allowed.
5. Reference Toxicant Testing
Today's action clarifies the purpose and requirements of reference
toxicant testing and the appropriate use of reference toxicant test
results. Several commenters identified inconsistencies in the
requirements for reference toxicant testing and recommended that EPA
clarify the purpose of generating reference toxicant test data. In
today's action, EPA clarifies that reference toxicant testing is used
to (1) initially demonstrate acceptable laboratory performance, (2)
assess the sensitivity and health of test organisms, and (3) document
ongoing laboratory performance. EPA has made method manual
modifications consistent with this stated purpose. Regardless of the
source of test organisms (in-house cultures or purchased from external
suppliers), the testing laboratory must perform at least one acceptable
reference toxicant test per month for each type of toxicity test method
conducted in that month. If a test
[[Page 69960]]
method is conducted only monthly, or less frequently, a reference
toxicant test must be performed concurrently with each effluent
toxicity test. This requirement will document ongoing laboratory
performance and assess organism sensitivity and consistency when
organisms are cultured in-house. When organisms are obtained from
external suppliers, concurrent reference toxicant tests must be
performed with each effluent sample, unless the test organism supplier
provides control chart data from at least the last five months of
reference toxicant testing. This requirement assesses organism
sensitivity and health when organisms are obtained from external
vendors. To initially demonstrate acceptable laboratory performance,
the method manuals require a laboratory to obtain consistent, precise
results with reference toxicants before it performs toxicity tests with
effluents under NPDES permits.
In today's action, EPA also clarifies the appropriate use of
reference toxicant test results. Commenters recommended that EPA
provide additional guidance on evaluating reference toxicant test
results and using these results to validate toxicity tests on test
samples of unknown toxicity. In response, EPA clarifies that reference
toxicant test results should not be used as a de facto criterion for
rejection of individual effluent or receiving water tests. Reference
toxicant testing is used for evaluating the sensitivity and consistency
of organisms over time and for documenting initial and ongoing
laboratory performance. EPA clarified the steps to take when more than
1 in 20 reference toxicant tests falls outside of control chart limits,
or when a reference toxicant test result falls ``well'' outside of
control limits. Under these circumstances, the laboratory should
investigate sources of variability, take corrective actions to reduce
identified sources of variability, and perform an additional reference
toxicant test during the same month.
In response to comments that reference toxicant testing only
compares variability within a laboratory, EPA added guidance for
evaluating test precision among laboratories and for limiting excessive
variability in reference toxicant testing. EPA has recommended that
laboratories compare the calculated coefficient of variation, also
referred to as the CV (i.e., standard deviation/mean), of the IC25 or
LC50 for the 20 most recent data points to the distribution of
laboratory CVs reported nationally for reference toxicant testing
(USEPA, 2000c). If the calculated CV exceeds the 75th percentile of CVs
reported nationally for LC50s or IC25s, the laboratory should use the
75th and 90th percentiles to calculate warning and control limits,
respectively, and the laboratory should investigate options for
reducing variability.
Several commenters recommended standardizing reference toxicants
and acceptance ranges for reference toxicant test results. Other
comments opposed mandatory reference toxicants and required acceptance
ranges claiming that insufficient guidance and data are available for
instituting such requirements and that such requirements would impose
additional costs on laboratories. In today's action, EPA is not
requiring the use of specific reference toxicants or setting required
acceptance ranges for reference toxicant testing. EPA agrees that
requiring specific reference toxicants and acceptance ranges would
increase laboratory costs. Many laboratories would be forced to develop
initial and ongoing documentation of laboratory performance (e.g.,
reference toxicant control charts) using a new reference toxicant. For
these laboratories, years of historic performance information using the
original reference toxicant would be rendered useless. In addition, EPA
believes that certain advantages gained by requiring reference toxicant
acceptance ranges are already provided by method modifications
instituted in today's action. For instance, today's action institutes
variability criteria when NPDES permits require sublethal WET testing
endpoints expressed using hypothesis testing. This method modification
limits WET test variability, which would be one of the primary purposes
of any standardized reference toxicant acceptance ranges.
6. Sample Holding Temperature
Today's action clarifies the allowable sample holding temperatures
for WET samples as 0[deg]-6[deg]C. EPA received comments that the
Agency should establish acceptable ranges for the current sampling
holding temperature of 4[deg]C. EPA has defined the acceptable range as
0[deg]-6[deg]C based on current NELAC (National Environmental
Laboratory Accreditation Conference) standards which state that, ``for
samples with a specified storage temperature of 4[deg]C, storage at a
temperature above the freezing point of water to 6[deg]C shall be
acceptable'' (NELAC, 2001). EPA also clarifies that hand-delivered
samples used on the day of collection do not need to be cooled to
0[deg]-6[deg]C prior to test initiation.
7. Biomass
Today's action clarifies that the sublethal endpoint used in
survival and growth tests is based on the number of initial organisms
exposed. Comments expressed concern that by calculating the chronic
endpoint based on the number of initial organisms (rather than
surviving organisms), the growth endpoint was in error and biased. EPA
disagrees. In the 1995 WET final rule, EPA changed the test endpoint
from a growth endpoint that was based on the number of surviving
organisms, to a combined growth and survival endpoint that is based on
the number of initial organisms. This does not represent an error in
the endpoint calculation, but rather a change in the endpoint itself.
EPA made this change: (1) to provide consistency with other methods
(e.g., Ceriodaphnia dubia Survival and Reproduction Test) that
incorporate survival along with sublethal effects, and (2) because the
survival and growth endpoint is a more sensitive measure than the
growth endpoint alone. While the 1995 WET final rule changed the test
endpoint to a combined survival and growth endpoint, the method manuals
continued to refer to the endpoint as a ``growth'' endpoint. Today's
action clarifies that the endpoint is, in fact, a combined survival and
growth endpoint that is more accurately termed biomass.
8. Total Residual Chlorine
Today's action clarifies the requirements for measuring total
residual chlorine in WET test samples. Several commenters stated that
certain requirements for measuring total residual chlorine were
unnecessary when the absence of the chemical has already been
determined. In response to these comments, EPA has clarified that if
total residual chlorine is not detected in effluent or dilution water
at test initiation, it is unnecessary to measure total residual
chlorine at test solution renewal or at test termination. If total
residual chlorine is detected at test initiation, then measurement of
total residual chlorine at test solution renewal and test termination
would continue to be required. EPA also has clarified that the
measurement of total residual chlorine is unnecessary in laboratory
prepared synthetic dilution water.
Commenters also recommended that EPA remove the requirement for the
analysis of total residual chlorine immediately following sample
collection. EPA has maintained this requirement in today's action,
because information on chlorine at the site and
[[Page 69961]]
time of collection is important for evaluating the effectiveness of
chlorination/dechlorination processes and comparing the results of WET
testing with instream effects.
9. Ceriodaphnia dubia Survival and Reproduction Test Termination
Criteria
Commenters recommended various modifications to the test
termination criteria in the Ceriodaphnia dubia Survival and
Reproduction Test. Some commenters recommended a strict seven-day test,
and others recommended that the test last no longer than seven days.
Other commenters recommended that the test be terminated when 80% of
control females produce three broods, rather than the current criteria
of 60%. Still other commenters recommended that fourth brood neonates
not be counted. To evaluate the recommended approaches to terminating
Ceriodaphnia dubia Survival and Reproduction Tests, EPA analyzed test
data from the WET Interlaboratory Variability Study using each of the
recommended test termination criteria. EPA compared the recommended
criteria to the current criteria by calculating within-test variability
and successful test completion rates under each of the test termination
scenarios. While some of the recommended test termination criteria
(such as termination when 80% of control females produce three broods
or a maximum of seven days) slightly improved the within-test
variability of the method (from a median PMSD of 23.2% to 19.9%), these
criteria caused significant reductions in successful test completion
(from 83% successful completion to 66%). Only the recommendation to
exclude fourth brood neonates resulted in a decrease in within-test
variability without an offsetting decrease in the rate of successful
test completion. Based on these results, EPA is modifying the
Ceriodaphnia dubia Survival and Reproduction Test to specify that
neonates from fourth broods are excluded from the number of neonates
counted in the test. With the exception of excluding fourth brood
neonates, EPA is maintaining the current test termination criteria.
These criteria state that the test is terminated when 60% or more of
the surviving control females have produced their third brood, or at
the end or eight days, whichever occurs first. These criteria may be
met at six, seven, or eight days.
10. Additional Minor Corrections
Some commenters identified additional errors in the WET method
manuals or the proposed changes that EPA was not aware of at the time
of proposal. In today's action, EPA has made these additional
corrections and minor clarifications.
C. Ratification and Withdrawal of Methods
In the September 28, 2001 proposal, EPA proposed to ratify the
following eleven test methods evaluated in the WET Interlaboratory
Variability Study: Ceriodaphnia dubia Acute Test; Fathead Minnow Acute
Test; Sheepshead Minnow Acute Test; Inland Silverside Acute Test;
Ceriodaphnia dubia Survival and Reproduction Test; Fathead Minnow
Larval Survival and Growth Test; Selenastrum capricornutum Growth Test;
Sheepshead Minnow Larval Survival and Growth Test; Inland Silverside
Larval Survival and Growth Test; Mysidopsis bahia Survival, Growth, and
Fecundity Test; and Champia parvula Reproduction Test. EPA proposed to
withdraw the Holmesimysis costata Acute Test and, in its place,
proposed a revised version of the method. As explained previously, EPA
is ratifying ten of these methods today based on the results of EPA's
WET Interlaboratory Variability Study that demonstrate the adequacy,
availability, and comparability of the methods (see Section IV.C). For
these ten methods, EPA generated sufficient interlaboratory validation
data, and those data justify ratification. EPA's WET Interlaboratory
Study evaluated interlaboratory precision, successful test completion
rates, and false positive rates of the WET methods from the testing of
over 700 samples in 56 laboratories. For each method ratified in
today's action, EPA obtained interlaboratory data on four sample
matrices from at least seven laboratories to as many as 35
laboratories.
Several commenters expressed concern that EPA did not properly
validate WET test methods, specifically, the Champia parvula
Reproduction Test and the Holmesimysis costata Acute Test. EPA was
unable to obtain interlaboratory precision data for these methods in
the WET Interlaboratory Variability Study. Because these WET methods
are not used widely in NPDES permits, EPA was unable to contract with a
minimum of six laboratories qualified and willing to conduct these test
methods within the time frame of the Study. In the proposed rule, EPA
supported these methods with intralaboratory precision data and limited
interlaboratory precision data (two trials of the Holmesimysis costata
Acute Test in two laboratories), but commenters questioned the
sufficiency of such data for validating methods for nationwide use, as
well as the necessity to approve such methods for nationwide use.
EPA has reviewed its proposal to ratify the Champia parvula
Reproduction Test in light of comments received and has decided to
withdraw the method from the list of nationally-approved test methods
at 40 CFR part 136. At the current time, an insufficient number of
laboratories nationwide have the capabilities to perform the method. As
noted, EPA was thus unable to obtain a rigorous multi-laboratory
performance data set to comprehensively evaluate this method. EPA had
predicted that as the requirements for use of this organism in the
NPDES permit program increased, the resulting increase in market demand
would result in an increase in the number of laboratories capable of
performing the test. However, the number of permits requiring the
Champia parvula chronic test has remained low (DeGraeve et al., 1998),
so few laboratories have invested in developing Champia parvula
cultures or standard operating procedures for the method. While today's
action removes the Champia parvula chronic test method from the 40 CFR
part 136 listing, EPA retains the standardized method in the marine
chronic method manual with an explanation that the method is not listed
at 40 CFR part 136 for nationwide use. Accordingly, retention of the
method in the method manual continues to enable standardization of the
method for developmental and other non-regulatory purposes and may
foster laboratories to maintain or even develop expertise in performing
the method.
EPA also has reviewed its proposal of the Holmesimysis costata
Acute Test in light of comments received. As proposed, EPA now
withdraws Holmesimysis costata as an acceptable species for use in the
Mysidopsis bahia Acute Test method. EPA does not, however, promulgate
the proposed Holmesimysis costata Acute Test method as a nationally-
approved method at 40 CFR part 136 at this time. Because the
Holmesimysis costata Acute Test is used in only a small number of
permits on the West Coast, EPA was unable to obtain sufficient
interlaboratory data on this method during the time that the WET
Interlaboratory Variability Study was conducted to support today's
rulemaking. While today's action removes the Homesimysis costata Acute
Test from the 40 CFR part 136 listing, EPA includes the proposed method
in the method manual with an explanation
[[Page 69962]]
that the method has not yet been approved at 40 CFR part 136 for
nationwide use.
Three commenters, including the California State Water Resources
Control Board, supported ratification of the Holmesimysis costata Acute
Test method. The California State Water Resources Control Board added
that ratification of this method was ``particularly important, as it is
the only method employing a marine species that is indigenous to the
Pacific coast.'' The California State Water Resources Control Board has
been proactive in developing, testing, validating, and implementing WET
test methods specific to West Coast species (USEPA, 1995b), and EPA
does not intend to frustrate that effort by today's action. For this
reason, EPA is specifying in Table IA of 40 CFR part 136 that the
marine acute and marine chronic test methods ratified in today's
rulemaking measure toxicity to estuarine and marine organisms ``of the
Atlantic Ocean and Gulf of Mexico.'' By defining the parameter measured
by promulgated marine methods as toxicity to organisms ``of the
Atlantic Ocean and Gulf of Mexico,'' today's action does not displace
West Coast methods that have been approved for use in States such as
California. Because test procedures for measuring toxicity to estuarine
and marine organisms of the Pacific Ocean are not listed at 40 CFR part
136, permit writers may include (under 40 CFR 122.41(j)(4) and
122.44(i)(1)(iv)) requirements for the use of test procedures that are
not approved at part 136, such as West Coast WET methods (USEPA, 1995b)
on a permit-by-permit basis. Furthermore, this rule does not preclude
permit writers addressing marine or estuarine waters of the Pacific
Ocean from requiring, on a permit-by-permit basis, any method
designated as approved for ``estuarine and marine organisms of the
Atlantic Ocean and Gulf of Mexico,'' where such method is suitable for
the specific application.
VI. Response to Major Comments
EPA encouraged public participation in this rulemaking and
requested comments on the proposed revision and ratification of WET
methods. EPA also requested data supporting comments, if available.
Thirty-eight stakeholders provided comments on the proposal.
Stakeholders included eight laboratories, eight regulatory authorities,
11 industries/industry groups, nine publicly-owned treatment works
(POTWs), and two environmental consulting companies.
This section summarizes major comments received on the proposed
rule that were not previously addressed in Section V and provides a
summary of EPA's responses. The complete comment summary and response
document can be found in the public record for this final rule.
A. Proposed WET Method Changes
EPA received comments on each of the proposed method changes, and
those comments that prompted modifications to the proposed method
changes are discussed in pection V of this preamble. Other substantial
comments on proposed method changes follow.
1. Cost
Several commenters expressed concern that proposed method
modifications will increase test costs. Of the WET method modifications
instituted in today's action, only four are additional mandatory
changes that have the potential to increase test costs. These four
modifications include: (1) The requirement for blocking by known
parentage in the Ceriodaphnia dubia Survival and Reproduction Test; (2)
the requirement to review test results for concentration-response
relationships; (3) the incorporation of mandatory variability criteria
for certain test methods when NPDES permits require sublethal WET
testing endpoints expressed using hypothesis testing; and (4) the
increase in the minimum number of replicates for the Fathead Minnow
Larval Survival and Growth Test, Selenastrum capricornutum Growth Test,
Sheepshead Minnow Larval Survival and Growth Test, Inland Silverside
Larval Survival and Growth Test, and Sea Urchin Fertilization Test. EPA
believes that the overall cost increases due to these changes will be
minor and that the potential benefits of these modifications outweigh
the incremental costs. EPA has estimated that the total cost of these
modifications for all permittees will be less than five million dollars
per year nationwide for all tests (Table 2 and USEPA, 2002). EPA
believes that these costs also would be alleviated by a potential
reduction in costs for retesting and additional investigations (e.g.,
toxicity identification evaluations). The modifications should result
in improved test performance and increased confidence in the
reliability of testing results.
Table 2.--Estimated Total Cost Resulting From WET Method Modifications
Required by Today's Action (From USEPA, 2002)
------------------------------------------------------------------------
Modification Cost ($/yr)
------------------------------------------------------------------------
Blocking-by-parentage...................................... $352,592
Concentration-response relationship........................ 98,069
Increased replicates....................................... 886,634
Variability criteria....................................... 2,595,873
------------
Total.................................................. 3,933,168
------------------------------------------------------------------------
2. Concentration-Response Relationships
Today, EPA is finalizing proposed method modifications to require
the review of concentration-response relationships for all multi-
concentration tests. Under this requirement, the concentration-response
relationship generated for each multi-concentration test must be
reviewed to ensure that calculated test results are interpreted
appropriately. In conjunction with this requirement, EPA has provided
recommended guidance for concentration-response relationship review
(USEPA, 2000a).
Several commenters expressed concern that the proposed method
modifications require that the concentration-response relationship be
reviewed but does not require that a concentration-response
relationship be established before determining that toxicity is
present. Commenters recommended that EPA require the establishment of a
``valid'' concentration-response relationship prior to determining
toxicity. Though within the scope of the proposed rule, EPA does not
consider such a requirement appropriate for several reasons. First, WET
methods and the WET testing program rely on the measurement of specific
test endpoints (NOECs, LC50s, IC25s) for determining toxicity, not on
achievement of specified concentration-response patterns. Second, the
concentration-response guidance is a component of test review that
ensures that test endpoints, which are used to determine toxicity, are
calculated and interpreted appropriately. Second, concentration-
response relationships are empirical; and a single definition for a
``valid'' concentration-response relationship is not appropriate. A
range of toxicants may produce an infinite range of different shaped
responses. In addition, a single response pattern may be due to several
different reasons, some indicating toxicity, and some not. For example,
the presence of pathogens,
[[Page 69963]]
considered an adverse effect confounding WET tests, may produce the
same concentration-response pattern as a true toxicant. For this
reason, EPA designed the guidance as a step-by-step review process that
investigates the causes for non-ideal concentration-response patterns
and provides for proper interpretation of test endpoints. Third, WET
testing has inherent characteristics that may limit the ability to
achieve ideal concentration-response relationships. For instance, WET
testing is constrained to 100% effluent sample as the highest test
concentration. This sometimes inhibits the ability to establish an
ideal concentration-response relationship that extends gradually from
no effect at one concentration to complete effect at some higher
concentration. Traditional toxicology on pure substances, from which
the concentration-response relationship concept is borrowed, is not
similarly constrained. Test concentrations can be increased or lowered
until an ideal response is generated. The typical WET test design of
five concentrations and a control also may limit the ability to
generate ideal concentration-response relationships. The location or
spacing of these five concentrations may miss the gradual transition
from no effect to complete effects. In traditional toxicology using
pure substances, tests can be rerun with altered or additional test
concentrations of the same compound, but in WET testing each individual
sample and test is unique and cannot be exactly duplicated due to the
complex and dynamic nature of the test samples over time. Non-ideal
concentration-response relationships will occasionally be encountered
in WET testing, and the goal of concentration-response relationship
review is to properly interpret these non-ideal patterns.
Fourth, the concentration-response relationship guidance has been
shown to be very effective at reducing false positives. For instance,
in the WET Interlaboratory Variability Study, the use of the
concentration-response relationship guidance reduced false positive
incidences from above 14% to below 5% for some methods (USEPA, 2001a).
3. Confidence Intervals
EPA is finalizing the proposed method modifications that provide
guidance when confidence intervals are not generated. This guidance
clarifies that confidence intervals may not be generated by EPA
software when test data do not meet specific assumptions required by
the statistical methods, when point estimates are outside of the test
concentration range, or when specific limitations imposed by the
software are encountered. EPA also provides guidance for proceeding
under each circumstance. Some commenters stressed the importance of
obtaining confidence intervals in all circumstances and recommended
that EPA use confidence intervals in assessing the reliability of
results and determining compliance. EPA believes that the failure to
generate confidence intervals should not adversely affect WET test
result reporting because confidence intervals surrounding point
estimates are not currently reported in the Permit Compliance System
(the national database tracking compliance with NPDES permits) or used
in compliance determinations. Compliance with permit requirements is
based on the point estimate itself and not confidence intervals
surrounding the estimate. This approach is no different in WET testing
than in chemical testing, where compliance is also based on the
analytical result itself. EPA demonstrated in the WET Interlaboratory
Variability Study that the WET methods provide adequate precision and
adequate protection from false positives. Therefore, EPA is not
altering the compliance determination approach to include the use of
confidence intervals.
B. Additional Revisions to WET Test Methods
In addition to receiving comment on proposed method modifications,
EPA received comments recommending additional method modifications.
Those recommendations that EPA incorporated in today's action and those
comments that prompted additional modifications are discussed in
section V of this preamble. Other substantial comments on additional
method changes are discussed below.
1. Method Flexibility
EPA received comments that requested additional requirements be
added to WET test methods, as well as comments that WET test methods
are overly restrictive and would benefit from additional flexibility.
As with all promulgated methods, EPA has attempted to balance these two
opposing objectives. EPA has prescribed certain method elements when
necessary to ensure the reliability of results, and allowed flexibility
in other method elements so that the performance of analytical methods
can be optimized. As noted in section V.B.3, EPA reevaluated the use of
mandatory and discretionary terms in the WET test methods to ensure
that the terms are included in the manuals as intended.
EPA received comments that WET test methods do not adequately
distinguish between required and recommended procedures. In response,
EPA modified the tables of test conditions and test acceptability
criteria presented in the method manuals for each method, such that
each item is identified as required or recommended. In addition, EPA
added to each method manual a section on test review. This section
provides direction on the review of sampling and handling procedures,
test acceptability criteria, test conditions, statistical methods,
concentration-response relationships, reference toxicant testing, and
test variability.
EPA believes that these method modifications clarify the
requirements for acceptable WET test results submitted under NPDES
permits. However, EPA acknowledges that these method modifications will
not solve all commenters concerns regarding inconsistencies in WET test
review and acceptance. In the WET test methods, EPA established the
minimum requirements for acceptable WET tests. In some cases, NPDES
permits incorporate recommendations from the WET test method manuals as
requirements in the permit (on a permit-by-permit basis). Authorized
States retain the authority to establish more stringent requirements or
to require additional procedures, test conditions, or QC elements.
Thus, WET requirements ultimately reflected as NPDES permit
requirements may continue to differ among States.
2. Test Acceptability Criteria
In the proposed rule, EPA solicited comments on increasing the test
acceptability criteria for mean control reproduction in the
Ceriodaphnia dubia Survival and Reproduction Test and mean control
weight in the Fathead Minnow Larval Survival and Growth Test. EPA also
requested that commenters submit supporting data. EPA received comments
both in favor of and opposed to increasing test acceptability criteria
for these methods, but these comments were not accompanied by
supporting data. Because EPA does not currently possess and did not
receive data indicating that such changes would improve the performance
of the methods, EPA is not modifying the survival, growth and
reproduction test acceptability criteria for these methods in today's
action.
EPA also received comments recommending the Agency establish
requirements for additional test acceptability criteria, such as limits
on
[[Page 69964]]
control variability. Today's action does establish mandatory
variability criteria when NPDES permits require sublethal WET testing
endpoints expressed using hypothesis testing. EPA has incorporated
these variability criteria as a required test review step for five
methods rather than as test acceptability criteria, meaning that,
depending on the reviewed result, retesting may be necessary. EPA
continues to use the term ``test acceptability criteria'' only to refer
to the evaluation of biological measurements in test controls (i.e.,
control survival, reproduction, and growth).
3. Quality Assurance/Quality Control Requirements
Some commenters expressed concern that WET test methods do not
contain adequate quality assurance/quality control (QA/QC)
requirements. Each of the toxicity test method manuals contains
separate, detailed, QA/QC guidelines, and each analytical method within
these manuals discusses all aspects of the tests which are related to
QA/QC. Section 4 of each method manual provides QA/QC requirements and
guidance for facilities, equipment, and test chambers; test organisms;
culturing and test dilution water; effluent and receiving water
sampling and handling; test conditions; food quality; test
acceptability criteria; calibration and standardization; replication
and test sensitivity; demonstrating acceptable laboratory performance;
documenting ongoing laboratory performance; and record keeping. The
primary QA/QC requirements of WET test methods, as contained in section
4 of the method manuals, remain the requirements for acceptable
biological performance (survival, reproduction, and growth) in test
controls and the requirement for the routine analysis of reference
toxicants. In today's action, however, EPA added additional QA/QC
requirements including the required review of concentration-response
relationships and mandatory variability criteria when NPDES permits
require sublethal WET testing endpoints expressed using hypothesis
testing. EPA believes that the QA/QC requirements of WET tests will
adequately ensure that results are reliable and of known and acceptable
quality.
4. Statistical Methods
Several commenters recommended that EPA approve and use alternative
statistical methods (such as percent effect approaches and Generalized
Linear Models). EPA has not included such alternative statistical
methods in today's modifications to WET test methods. EPA believes that
the statistical methods currently recommended in the WET methods are
appropriate, and acknowledges that these recommended statistical
methods are not the only appropriate techniques. The method manuals
state that, ``the statistical methods recommended in this manual are
not the only possible methods of statistical analysis.'' The
recommended statistical methods described in the method manuals were
selected because they are ``(1) applicable to most of the different
toxicity test data sets for which they are recommended, (2) powerful
statistical tests, (3) hopefully ``easily'' understood by
nonstatisticians, and (4) amenable to use without a computer, if
necessary'' (see subsection 9.4.1.2 of USEPA, 1994a).
Several commenters also expressed concern over bias introduced by
the smoothing technique that is used in the recommended Inhibition
Concentration Procedure (ICp). EPA has acknowledged in the method
manuals and in method guidance (USEPA, 2000a) that the smoothing
process may result in an upward adjustment in the control mean. EPA has
provided guidance on concentration-response relationship review that
corrects anomalous results that may arise from this smoothing procedure
(USEPA, 2000a). This guidance warns that results from point estimation
techniques should be interpreted carefully when the response pattern
includes stimulation at low concentrations and no significant effect at
higher concentrations. Under these conditions, the smoothing process
could result in anomalous results, so EPA guidance recommends
evaluating the ICp calculation without smoothing in these cases. If the
percent effect at the receiving water concentration (RWC) is less than
25% when calculated without smoothing, and the response at the RWC is
not statistically significantly different from the control response,
then a calculated IC25 of less than the RWC should be noted as
anomalous and the effluent determined to be non-toxic at the RWC.
C. Ratification and Withdrawal of Methods
1. Validation of Performance Characteristics
Several commenters stated that EPA did not properly validate WET
test methods because it did not evaluate essential performance
characteristics. Commenters referenced EPA's Report to Congress on the
Availability, Adequacy, and Comparability of Testing Procedures (USEPA,
1988) and stated that EPA failed to validate the following performance
characteristics required by this report: accuracy, precision, dynamic
range, detection limits, interferences, ruggedness (applicability),
reporting, and representativeness/method comparability. EPA disagrees
with this assertion and maintains that the WET test methods ratified in
today's action were adequately validated according to all of the
applicable criteria identified in the 1988 Report to Congress.
The list of performance characteristics cited by the commenters is
provided in the 1988 Report to Congress within the context of chemical
methods, and several of these characteristics are not applicable to
biological test methods such as the WET methods that EPA is ratifying
today. The 1988 Report to Congress specifically notes that not all such
criteria apply to biological testing. The Report explains that the
generation of scientifically accurate and valid biological measurements
for environmental pollutants requires approximately the same criteria
for assessing the adequacy of a method as previously described for
chemical analyses, however, there are several differences which are
important. Detection limits and dynamic range are specifically listed
as characteristics that ``are not usually appropriate concepts for all
biological measurements unless instrumentation is required.'' Because
some performance characteristics listed in the 1988 Report to Congress
for chemical methods are not applicable to biological test methods, EPA
did not (and, in fact, could not) evaluate those inapplicable
performance characteristics for WET test method validation.
In ratifying the previously approved WET test methods, EPA applied
the availability, adequacy, and comparability criteria identified in
the Report as relevant to biological measurements. The WET test methods
ratified today are ``available'' because EPA has identified a
sufficient number of laboratories that can conduct the test and culture
the test organisms. The ratified WET test methods are ``adequate''
because the multi-laboratory tests (as well as aggregation of single
laboratory tests) demonstrate high degrees of precision; the tests are
reproducible. In addition, the manuals identify interferences and ways
to control interference. Finally, the test acceptability criteria for
control performance and requirements for reference toxicant testing
provide sufficient standards to ensure data integrity, absent the
``calibration'' procedures available with non-living analytical
instrumentation.
[[Page 69965]]
The Report specifically identified detection limits and dynamic
range as performance characteristics that are usually not applicable to
biological measurements, and the 1988 conclusions remain true today. In
addition, accuracy is a performance characteristic that is not
completely applicable to WET testing. Accuracy as a performance
characteristic of a measurement system describes the closeness of
measured results to a known result. Chemical methods generally measure
some surrogate property (e.g., absorption of light at a particular
wavelength) of an analyte (e.g., copper) to determine the concentration
of that analyte. To confirm that the surrogate measure accurately
represents the true concentration of the analyte, the pure analyte can
be weighed, diluted to a known concentration, and measured using the
analytical procedure under study. This procedure cannot be conducted
for whole effluent toxicity. Toxicity cannot be purified, weighed, or
diluted to a known concentration of ``toxicity.'' Toxicity is only
defined by its effects on organisms, and it is these effects that are
directly measured in the toxicity test. Because toxicity is inherently
defined by the measurement system (a ``method-defined analyte''), and
toxicity cannot be independently measured apart from a toxicity test,
accuracy as a performance characteristic is not completely applicable.
The inapplicability of the accuracy performance characteristic does not
mean that WET tests are not accurate or that permittees are incapable
of certifying the accuracy of WET test results reported on discharge
monitoring reports. It means simply that the procedures commonly used
in analytical testing to measure the performance characteristic that is
termed ``accuracy'' cannot be applied to WET test methods.
Notwithstanding the previous explanation, one component of accuracy
can be described for WET tests. The American Society for Testing and
Materials (ASTM) defines accuracy as ``a measure of the degree of
conformity of a single test result generated by a specific procedure to
the assumed or accepted true value and includes both precision and
bias'' (ASTM, 1998; emphasis added). Bias is defined as ``the
persistent positive or negative deviation of the average value of a
test method from the assumed or accepted true value'' (ASTM, 1998).
Precision is defined as ``the degree of agreement of repeated
measurements of the same property, expressed in terms of dispersion of
test results about the arithmetical mean result obtained by repetitive
testing of a homogeneous sample under specified conditions' (ASTM,
1998). Like ASTM, the 1988 Report to Congress (USEPA, 1988) also
explains that accuracy includes both bias and precision. As explained
previously, EPA conducted an Interlaboratory Variability Study of the
ratified methods in order to, among other things, generate a quantified
estimate of the precision for each method studied. WET tests are
therefore amenable to the precision portion of accuracy. It is the bias
portion of accuracy that is not applicable to WET test methods and
cannot be described for WET as it is described for chemical analytes.
The additional performance characteristics listed in the 1988
Report to Congress, namely precision, interferences, ruggedness
(applicability), reporting, and representativeness, are applicable to
biological test methods, and EPA evaluated and considered these
characteristics in ratifying the WET test methods. To establish the
precision of the methods, EPA conducted an Interlaboratory Variability
Study for each of the WET methods ratified today. From the Study, EPA
established single-laboratory and multi-laboratory precision estimates
for multiple sample matrices for each method (USEPA, 2001a; USEPA,
2001b). EPA also conducted a study of within laboratory precision
measured when testing reference toxicants (USEPA, 2000c). In today's
action, EPA is modifying the WET method manuals to include this new and
updated single-laboratory and multi-laboratory precision data for each
method. Precision data from the WET Interlaboratory Variability Study
confirmed that the WET test methods provided adequate precision (CVs
ranged from 10.5 to 43.8%). The measured precision ranges for the
ratified toxicity tests demonstrate the tests are comparable to (no
more variable than) chemical analytical methods approved at 40 CFR part
136. Finally, the precision had improved since the time the methods
were promulgated in 1995, thus confirming EPA's conclusions that
precision would improve with time, i.e., as analysts developed more
expertise the methods would be ``validated by use.''
In addition to precision, EPA evaluated and considered the
performance characteristic of interferences. Each WET test method
contains a section describing possible test interferences. In today's
action, EPA has expanded that section to address two additional
interference concerns that were raised by stakeholders by including
guidance for controlling test interference that could be due to pH
drift in the test and interference caused by pathogens.
EPA also evaluated and considered the performance characteristic of
ruggedness or applicability. The methods ratified today use materials
that are widely available and organisms that can be easily cultured in
the laboratory. By conducting a national interlaboratory study of these
methods, EPA also confirmed that the methods are adaptable to a wide
variety of laboratories and that the methods generate reproducible
results in those laboratories. In the WET Interlaboratory Variability
Study, EPA documented successful test completion rates of 63.6% to 100%
for WET methods. EPA anticipates that method modifications instituted
today will improve the successful test completion rate for methods at
the bottom of this range, such as the Selenastrum capricornutum Growth
Test. Today, EPA is requiring the use of EDTA in this test. As
laboratories gain experience in performing the test with EDTA, EPA
anticipates that successful test completion rates will improve. See
section VI.C.4 of this preamble.
EPA also considered the aspect of result reporting in its
development and validation of WET test methods. Each method manual
contains a section devoted to test review and reporting. In today's
action, EPA has supplemented this section by providing guidance on the
review of sampling and handling, test acceptability criteria, test
conditions, statistical methods, concentration-response relationships,
reference toxicant testing, and test variability. In addition, EPA
clarified the required and recommended test conditions when submitting
data under NPDES permits.
EPA documented and considered the representativeness or
comparability of WET methods. Prior to approving the WET test methods
in the 1995 WET final rule, EPA conducted several studies that
demonstrated the ability of WET tests to predict impacts of effluents
on the biological integrity of receiving waters (USEPA, 1991). In a
1995 workshop of nationally recognized WET experts (the Society of
Environmental Toxicology and Chemistry's Pellston Workshop), including
those from academia, government, and the regulated community (e.g.,
POTWs and industry), the experts concluded that ``WET testing is an
effective tool for predicting receiving system impacts when appropriate
considerations of
[[Page 69966]]
exposure are considered'' (Waller et al., 1996). The workgroup also
agreed that ``further laboratory-to-field validation is not essential
for the continued use of WET testing'' (Waller et al., 1996).
2. Interlaboratory Variability Study
Several commenters expressed concern that EPA used data from the
Interlaboratory Variability Study that was of poor quality and would
have been discarded in a regulatory context. In conducting the WET
Interlaboratory Variability Study, EPA's objective was to validate the
WET methods as promulgated. EPA was not attempting to validate the
diversity of testing requirements that may be implemented in various
States. State regulatory authorities retain the discretion to enhance
the requirements of a method for implementation in their State as well
as to require procedures that EPA otherwise recommends. In the WET
Interlaboratory Variability Study, EPA appropriately evaluated data
according to the promulgated methods and ASTM guidance for measuring
interlaboratory method precision. EPA accurately invalidated tests
according to test acceptability criteria specified in each method. EPA
acknowledges that the promulgated methods allow flexibility in the
review of test conditions. The method manuals state that departures in
specified test condition ranges do not necessarily invalidate test
results. In today's action EPA modified the methods to better clarify
this allowable flexibility. For the purposes of reviewing data
submitted under NPDES permits, the manuals now clearly distinguish
between requirements of the method and recommended test condition
ranges.
Several commenters expressed concern that EPA did not use the
results of reference toxicant tests from the WET Interlaboratory
Variability Study to qualify or disqualify data. EPA agrees. EPA used
reference toxicant tests in the manner in which they are described in
the method manuals. Failure of reference toxicant tests do not
necessarily invalidate a test. In today's action, EPA has incorporated
method modifications to clarify reference toxicant testing requirements
and the appropriate use of reference toxicant test data. EPA has
clarified that reference toxicant test results should not be used as a
de facto criterion for rejection of individual effluent or receiving
water tests, but rather, reference toxicant testing is used for
evaluating the health and sensitivity of organisms over time and for
documenting initial and ongoing laboratory performance.
Several commenters expressed concern that too few data points were
used to estimate method performance in the WET Interlaboratory
Variability Study. In accordance with ASTM guidance on determining
interlaboratory method precision, EPA set a data quality objective of a
minimum of six complete and useable data sets for each WET test method
evaluated in the Study. To meet this data quality objective, EPA
endeavored to sponsor a minimum of nine laboratories per method. For
all of the methods that EPA is ratifying today, seven or more
laboratories participated in interlaboratory testing. For several
individual sample matrices and test method combinations that were
tested (blank sample analyzed using the Selenastrum capricornutum
Growth Test, receiving water sample analyzed using the Selenastrum
capricornutum Growth Test without EDTA, and the receiving water sample
analyzed using the Inland Silverside Acute Test), fewer than six
useable data sets were obtained. EPA did not, however, establish
precision criteria in today's rule based on results from a single
sample matrix. EPA tested four sample matrices (blank, reference
toxicant, effluent, and receiving water) with each test method, and
precision estimates were based on the combined results of reference
toxicant, effluent and receiving water testing. Because multiple sample
matrices were used to generate precision estimates, more than six
useable data sets were used for each method. In fact, at least 17 data
sets were used to establish precision estimates for each method.
Several commenters also expressed concern that the selection of
laboratories for the WET Interlaboratory Variability Study was biased.
EPA disagrees. EPA believes that the laboratories that participated in
the WET Interlaboratory Variability Study were representative of the
laboratory community that commonly conducts WET testing for permittees.
From the outset, EPA and the regulated community wanted to ensure that
participants in the Study were representative. Industry trade groups,
such as AMSA (Association of Metropolitan Sewerage Agencies), surveyed
their member permittees to identify the laboratories that provide their
routine WET testing services. AMSA requested that members sponsor those
laboratories' participation in the Study. Of the 55 participant
laboratories involved in the Study, 44 (or 80%) were specifically
recommended by AMSA with commitments from AMSA members to sponsor such
laboratories' participation in the Study. Thirty-seven of these
laboratories were ultimately sponsored by AMSA members to analyze
samples using one or more methods. The remaining seven laboratories had
commitments of sponsorship from AMSA members, but were ultimately
sponsored by EPA in the Study because their bids were among the nine
lowest. The high percentage (80%) of laboratories in the Study that
were sponsored by permittees for participation demonstrates that the
laboratories involved in the Study are representative of those that
commonly conduct WET testing for permittees.
Several commenters expressed concern that a majority of
laboratories did not detect toxicity in the reference toxicant sample
type distributed for the Ceriodaphnia dubia Survival and Reproduction
Test method. Prior to interlaboratory testing in the WET
Interlaboratory Variability Study, referee laboratories conducted
preliminary testing to determine the appropriate composition of samples
to prepare for the Study. This preliminary testing was important for
ensuring that test samples prepared for the Study produced results
within the test concentration range. Despite these preliminary testing
efforts, the spiking level selected for the reference toxicant sample
type in the Ceriodaphnia dubia Survival and Reproduction Test method
was insufficient to produce the targeted level of effect. The spiking
concentration of KCl for this sample was selected to achieve an IC25 of
approximately 50% sample based on preliminary testing, but the spiked
sample missed this targeted effect level. The prepared sample was only
slightly toxic and could not be detected as toxic in 67% of tests.
Depending on the sensitivity of test organisms at individual
laboratories, some laboratories identified the sample as toxic, while
other laboratories did not. Similarly, marginally toxic effluents may
exhibit intermittent toxicity in routine monitoring. In such cases,
permittees and regulatory authorities should consult EPA guidance that
addresses marginal and intermittent toxicity (USEPA, 1991; USEPA,
2000c; USEPA, 2001f).
The reference toxicant sample used in the Study also was prepared
as an ampule that was reconstituted at each participant laboratory.
This reconstitution process also likely produced minor variations (from
laboratory to laboratory) in the final sample composition that
influenced whether toxicity was detected. While the concentration of
potassium ions was not measured in each final reconstituted
[[Page 69967]]
sample, conductivity was measured and can be used as an approximate
surrogate measure. In samples that showed toxicity, the average
conductivity was 873 [mu]mhos, and in samples that did not show
toxicity, the average conductivity was 797 [mu]mhos. The differences in
conductivity between tests that indicated toxicity and tests that did
not were statistically significantly different (at the alpha = 0.05
level). This finding indicates that those samples which were less
diluted in the reconstitution process, were also more likely to be
toxic.
Several commenters also expressed concern over the way EPA handled
outlier data points in the WET Interlaboratory Variability Study. EPA
believes that outliers were treated according to standard practice and
according to ASTM standards for measuring method precision. EPA
identified outliers using ASTM's h and k statistics, and discarded
outliers only when a probable cause for the outlier was identified. In
all, only eight tests in the entire study of 698 tests were excluded
based on outlier analysis.
3. Variability
Several commenters stated that the variability of the WET methods
(measured in terms of CV) is too high for use in NPDES permits.
Commenters also recommended that specific steps be taken to account for
variability in the permit limit derivation and compliance determination
process. EPA believes that the WET Interlaboratory Variability Study
accurately estimated the precision of WET test methods, and that this
precision is adequate for regulatory use of the WET methods. The
precision measured for the WET test methods is comparable to that of
chemical methods. While EPA agrees with commenters that WET test
methods cannot be compared in all aspects to chemical methods, the
comparison of interlaboratory precision values does demonstrate that
WET test methods are no more variable than other methods approved at 40
CFR part 136 and used for regulatory compliance purposes.
In a recent peer-reviewed guidance document (USEPA, 2000c), EPA
thoroughly evaluated the issue of WET test method variability and
accounting for such variability in NPDES applications. The document
concluded that ``comparisons of WET method precision with method
precision for analytes commonly limited in NPDES permits clearly
demonstrate that the variability of the promulgated WET methods is
within the range of variability experienced in other types of [required
regulatory]
analyses.'' The analytical variability of WET test methods
is accounted for appropriately in the development of permit limits
derived according to EPA's Technical Support Document (TSD) (USEPA,
1991). The TSD approach accounts for both effluent variability and
method variability. The TSD statistical approach to determination of
reasonable potential and permit limit derivation considers combined
effluent and analytical variability through the CV of measured effluent
values. Because the determination of effluent variability is based on
empirical measurements, the variability estimated for effluent
measurements includes the variability of pollutant levels, sampling
variability, and a smaller component owed to method variability.
EPA does not recommend additional approaches or factors to account
for variability, because the TSD approach appropriately accounts for
method variability in the permit derivation process. In the guidance
document, EPA evaluated additional approaches to account for
variability in the permit derivation process and concluded that such
approaches would not ensure adequate protection of water quality. The
TSD approach was designed to provide a reasonable degree of protection
for water quality as well as from effluent and analytical variability.
Alternative approaches would undermine these objectives.
Some commenters expressed specific concern that the Selenastrum
capricornutum Growth Test method was too variable. EPA believes that
the variability of the Selenastrum capricornutum Growth Test method, as
measured in the WET Interlaboratory Variability Study (USEPA, 2001a)
and variability guidance document (USEPA, 2000c), is acceptable for the
intended regulatory use of the methods. EPA observed in the WET
Interlaboratory Variability Study that the variability of the
Selenastrum capricornutum Growth Test method was lower when the method
was conducted with the addition of EDTA. In today's action, EPA is
removing the option to conduct the test without the addition of EDTA
when data is submitted under NPDES permits. EPA believes that this
modification will improve the overall performance of the test method.
False positive rates decreased from 33.3% to 0.00% and interlaboratory
variability decreased from 58.5% to 34.3% when EDTA was added. EPA
cautions, however, that the required addition of EDTA may make the
Selenastrum capricornutum Growth Test less sensitive, thus less useful,
for measuring the toxicity of some test samples, specifically, samples
that contain toxic levels of metals.
4. Successful Test Completion Rate
Some commenters stated that EPA incorrectly calculated successful
test completion rates in the WET Interlaboratory Variability Study by
failing to invalidate tests that did not meet specific test condition
ranges. As previously discussed (see section VI.C.2 of this preamble),
EPA accurately invalidated tests according to the test acceptability
criteria specific to each method, and successful test completion rates
were based on meeting these criteria. EPA acknowledges that the
promulgated methods allow flexibility in the review of test conditions.
The method manuals state that departures in specified test condition
ranges do not necessarily invalidate test results. In today's action
EPA has modified the methods to better clarify this allowable
flexibility. For the purposes of reviewing data submitted under NPDES
permits, the manuals now clearly distinguish between requirements of
the method and recommended test condition ranges.
Several commenters stated that the successful test completion rate
measured for the Ceriodaphnia dubia Survival and Reproduction Test
method was unacceptable and indicates a lack of ruggedness. EPA
believes that the successful test completion rate observed for the
Ceriodaphnia dubia Survival and Reproduction Test method in the WET
Interlaboratory Variability Study was artificially suppressed by very
poor performance in a small subset of laboratories. Only ten of the 34
participant laboratories performed invalid tests, but eight of these
laboratories performed invalid tests on 50% or more of the samples
tested. The low rate of successful test completion in these eight
laboratories may have been influenced by the Study's strict testing
schedule, which required that each test be conducted on a given day and
that all tests be conducted within a 15-day time period. When invalid
tests conducted in a given laboratory were likely due to marginal or
poor health of the test organism cultures, then it was logical that the
laboratory would fail a high percentage of tests during the Study
because culture health was unlikely to fully recover within 15 days.
EPA believes that measuring an individual laboratory's rate of
successful test completion over a 15-day period may not be
representative of that laboratory's overall successful test completion
rate. For instance, several laboratories had successful test completion
rates of 0% during the WET
[[Page 69968]]
Interlaboratory Variability Study. Obviously, this result is not
indicative of the laboratory's overall successful test completion rate.
If so, the laboratory would not be in business or would not have been
able to prequalify for participation in the Study. EPA believes that
successful test completion rates for this method are higher in routine
use because testing laboratories are allowed flexibility in the timing
of sample collection and can avoid initiating tests during periods of
marginal to poor culture health.
Some commenters expressed concern that the successful test
completion rate for the Selenastrum capricornutum Growth Test method
was too low. In today's action, EPA is removing the option to conduct
the test without the addition of EDTA. EPA believes that this
modification will improve successful test completion rates for the
method as laboratories consistently culture and test with EDTA. The
successful test completion rate of 63.6% (when conducted with EDTA) was
in part due to laboratory inexperience in using both the with and
without-EDTA techniques. For example, two laboratories that cultured
organisms without EDTA and generally conducted tests without EDTA
showed poor successful test completion rates (failing eight of eight
tests) when EDTA was used. These laboratories failed all eight tests
conducted with EDTA and passed all but one test (seven of eight)
without EDTA. Commenters point out that laboratories were prequalified
for participation in the WET Interlaboratory Variability Study, but
this prequalification required only experience with the method, not
experience with both the with and without-EDTA procedures of the
method. Some laboratories cultured organisms and typically conducted
tests with EDTA, and other laboratories cultured organisms and
typically conducted tests without EDTA.
5. False Positive Rate
Several comments stated that EPA underestimated the false positive
rates measured in the WET Interlaboratory Variability Study and that
the measured rates are unacceptably high for regulatory use. In the
context of WET methods, the false positive rate is the rate at which
tests conducted on non-toxic dilution waters indicate the presence of
toxicity (i.e., NOEC, LC50, or IC25 test endpoints are <100% effluent).
EPA disagrees with comments that stated that false positive rates for
WET test methods are unacceptably high. EPA's WET Interlaboratory
Variability Study conclusively showed that measured false positive
rates were below the theoretical rate of 5% estimated for the methods.
Measured false positive rates were 3.7% for the Ceriodaphnia dubia
Survival and Reproduction Test method, 4.35% for the Fathead Minnow
Larval Survival and Growth Test method, and 0% for all other methods
evaluated in the WET Interlaboratory Variability Study (with the
exception of the Selenastrum capricornutum Growth Test conducted
without EDTA, which EPA is removing as an option in today's action). A
total of 150 valid WET tests were conducted on blank samples in the
Study. Of these, only two tests (1.3%) resulted in a false positive
result.
The WET Interlaboratory Variability Study conclusively demonstrated
that the false positive rate of WET methods is at or below the level
expected for the methods. While this rate is low (below 5%), false
positives do occur. EPA accounts for this possibility in the compliance
and enforcement guidance. EPA policy states that ``EPA does not
recommend that the initial response to a single exceedance of a WET
limit, causing no known harm, be a formal enforcement action with a
civil penalty'' (USEPA, 1995a). EPA policy suggests additional testing
is an appropriate initial response to a single WET limit exceedance.
Several commenters expressed concern that WET tests do not have
method detection limits as contained in chemical methods to protect
from reporting false positive results. As previously discussed (see
section VI.C.1 of this preamble), method detection limit concepts are
not applicable to WET test methods and have not been applied
historically to toxicity testing methods developed by EPA or by
voluntary consensus standards bodies.
EPA established the method detection limit (MDL) concept
specifically for chemical methods, where results generally consist of a
single measurement of the pollutant of interest by an analytical
instrument. The MDL concept uses information about the variability of
the measurement system to determine a response level at which the
measurement can be reliably distinguished from background ``noise,''
thus providing protection from false positive results. In WET testing,
the final result is not based on a single measurement, but is the
product of a series of replicated measurements on a range of effluent
concentrations. The additional measurements, controls, replication, and
statistical approaches included in the WET test method ``measurement
system'' ensure that measured responses can be reliably distinguished
from background noise.
While results from chemical methods may rely on a single instrument
measurement, each WET test is designed as an experiment. WET tests
contain at least six treatments, each replicated from four to ten
times. Measurements are made on each replicate of each treatment, so
that results reflect average responses and the variability of those
responses can be estimated. Each test also includes a control
treatment, which is also replicated. This control treatment provides a
measure of the background response and the ``noise'' or variability
associated with that response.
The control response is then compared to the response in effluent
treatments using statistical methods to test the hypothesis that
treatments containing effluent are not significantly different from the
control treatment. If this hypothesis is rejected (considering the
measured background or control responses, the treatment responses, and
the variability associated with those responses), then the effluent is
considered toxic. Hypothesis testing techniques provide protection from
false positive results by specifically setting the Type I error rate
allowed in rejecting the null hypothesis. Point estimation techniques
use regression analysis to determine the effluent concentration that
produces a specified level of response (e.g., the IC25 endpoint
specifies a 25% difference between control and effluent treatment
response in order for the effluent to be determined as toxic). In this
case, false positive protection is inherently provided by the level of
response required for generation of the selected endpoint. EPA believes
that the test design employed in WET testing (including controls,
replication, and hypothesis testing or point estimation) provides
adequate protection from false positives.
6. Implementation
Some commenters commented on issues specifically related to the
implementation of WET permits, such as reasonable potential
determinations, independent applicability of WET limits, discharge
monitoring report certifications, and use of WET methods in NPDES
permits. Many such comments are beyond the scope of this rulemaking. In
the proposed rulemaking, EPA invited comments ``only on the conduct of
WET test methods and not on the implementation of WET control
strategies through NPDES permits.'' EPA recognizes that NPDES
permittees have continuing
[[Page 69969]]
concerns about implementation of WET requirements in NPDES permits. In
a `WHEREAS' clause to the Settlement Agreement described previously,
EPA acknowledged that the provisions of the Settlement Agreement, which
focused primarily on test methodology and, to a lesser extent,
interpretation of test results, did not address all of the litigants'
concerns regarding applicability of WET testing requirements to
particular waterbodies (with specific reference to intermittent or
effluent dependent waterbodies located in the Arid West) and did not
address many of the litigants' concerns regarding regulatory
implementation of WET control programs (e.g., toxicity identification
evaluation requirements, toxicity reduction evaluation requirements,
compliance determinations, and trigger thresholds). In addition, the
Settlement Agreement also acknowledged that the 1995 rule, which
incorporated the WET test methods in dispute, did not specify means to
adjust for the frequency, duration, or magnitude of instream exposure
conditions, and that such decisions are to be made by the regulatory
authority in the context of water quality standard setting and/or NPDES
permitting decisions. EPA continues to acknowledge these continuing
concerns and will continue to address implementation concerns as they
arise in concrete circumstances or through guidance, as appropriate.
VII. Statutory and Executive Order Reviews
A. Executive Order 12866: Regulatory Planning and Review
Under Executive Order 12866 (58 FR 51735; October 4, 1993), the
Agency must determine whether the regulatory action is ``significant''
and therefore subject to Office of Management and Budget (OMB) review
and the requirements of the Executive Order. The Executive Order
defines ``significant regulatory action'' as one that is likely to
result in a rule that may:
(1) Have an annual effect on the economy of $100 million or more,
or adversely affect in a material way the economy, a sector of the
economy, productivity, competition, jobs, the environment, public
health or safety, or State, local, or Tribal governments or
communities;
(2) Create a serious inconsistency or otherwise interfere with an
action taken or planned by another agency;
(3) Materially alter the budgetary impact of entitlements, grants,
user fees, or loan programs or the rights and obligations of recipients
thereof; or
(4) Raise novel legal or policy issues arising out of legal
mandates, the President's priorities, or the principles set forth in
the Executive Order.
It has been determined that this rule is not a ``significant
regulatory action'' under the terms of Executive Order 12866 and is
therefore not subject to OMB review.
B. Paperwork Reduction Act
This action does not impose an information collection burden under
the provisions of the Paperwork Reduction Act, 44 U.S.C. 3501 et seq.
This rule revises and ratifies test methods that are currently approved
for use in NPDES permits and does not impose any additional information
collection requirements.
Burden means the total time, effort, or financial resources
expended by persons to generate, maintain, retain, or disclose or
provide information to or for a Federal agency. This includes the time
needed to review instructions; develop, acquire, install, and utilize
technology and systems for the purposes of collecting, validating, and
verifying information, processing and maintaining information, and
disclosing and providing information; adjust the existing ways to
comply with any previously applicable instructions and requirements;
train personnel to be able to respond to a collection of information;
search data sources; complete and review the collection of information;
and transmit or otherwise disclose the information.
An Agency may not conduct or sponsor, and a person is not required
to respond to a collection of information unless it displays a
currently valid OMB control number. The OMB control numbers for EPA's
regulations are listed in 40 CFR part 9 and 48 CFR chapter 15.
C. Regulatory Flexibility Act
The Regulatory Flexibility Act (RFA) generally requires an agency
to prepare a regulatory flexibility analysis of any rule subject to
notice and comment rulemaking requirements under the Administrative
Procedure Act or any other statute unless the agency certifies that the
rule will not have a significant economic impact on a substantial
number of small entities. Small entities include small businesses,
small organizations, and small governmental jurisdictions.
For purposes of assessing the impacts of today's rule on small
entities, small entity is defined as: (1) A small business as defined
by the U.S. Small Business Administration definitions at 13 CFR
121.201; (2) a small governmental jurisdiction that is a government of
a city, county, town, school district or special district with a
population of less that 50,000; and (3) a small organization that is
any not-for-profit enterprise which is independently owned and operated
and is not dominant in its field.
After considering the economic impacts of today's final rule on
small entities, I certify that this action will not have a significant
economic impact on a substantial number of small entities. Today's rule
revises and ratifies EPA WET test methods currently approved for use at
40 CFR part 136. Overall, the costs of these revisions are minimal.
While some of the revisions may increase costs (e.g., quality control
requirements), EPA believes that these costs will be alleviated by a
potential reduction in retesting and additional investigations (e.g.,
accelerated testing, toxicity identification evaluations, or toxicity
reduction evaluations) by the permittee that may result from improved
test performance and increased confidence in the reliability of testing
results. Many of the laboratories that conduct WET testing are already
implementing the additional requirements, further minimizing any
potential cost increases. EPA estimates that the average incremental
cost per permit per year for today's method revisions is $276. Because
monitoring frequency is typically less frequent for small entities than
large entities, EPA expects the average incremental cost per permit per
year to be even less than $276 for small entities. Using a cost of $276
and average revenue information for small governmental jurisdictions
and businesses, EPA estimates that the incremental costs for these
method revisions are less than 0.1 percent of revenue for small
entities.
D. Unfunded Mandates Reform Act
Title II of the Unfunded Mandates Reform Act of 1995 (UMRA), Public
Law 104-4, establishes requirements for Federal agencies to assess the
effects of their regulatory actions on State, Tribal, and local
governments and the private sector. Under section 202 of the UMRA, EPA
generally must prepare a written statement, including a cost-benefit
analysis, for proposed and final rules with ``Federal mandates'' that
may result in expenditures to State, Tribal, and local governments, in
the aggregate, or to the private sector, of $100 million or more in any
one year. Before promulgating an EPA rule for which a written statement
is needed, section 205 of the UMRA generally requires EPA to identify
and consider a reasonable number of regulatory alternatives and adopt
the least costly, most cost-
[[Page 69970]]
effective or least burdensome alternative that achieves the objectives
of the rule. The provisions of section 205 do not apply when they are
inconsistent with applicable law. Moreover, section 205 allows EPA to
adopt an alternative other than the least costly, most cost-effective
or least burdensome alternative if the Administrator publishes with the
final rule an explanation of why that alternative was not adopted.
Before EPA establishes any regulatory requirements that may
significantly or uniquely affect small governments, including Tribal
governments, it must have developed under section 203 of the UMRA a
small government agency plan. The plan must provide for the
notification of potentially affected small governments, enabling
officials of affected small governments to have meaningful and timely
input in the development of EPA regulatory proposals with significant
Federal intergovernmental mandates, and informing, educating, and
advising small governments on compliance with the regulatory
requirements.
EPA has determined that today's rule does not contain a Federal
mandate that may result in expenditures of $100 million or more for
State, Tribal, and local governments, in the aggregate, or the private
sector in any one year. This rule promulgates revisions to WET test
methods that are currently approved for use in NPDES permits and
certification of Federal licenses under the CWA. The revisions are
minor and the cost to implement them is minimal. Thus, today's rule is
not subject to sections 202 and 205 of the UMRA. For the same reasons,
EPA has also determined that this rule contains no regulatory
requirements that might significantly or uniquely affect small
governments. Thus, today's rule also is not subject to the requirements
of section 203 of the UMRA.
E. Executive Order 13132: Federalism
Executive Order 13132, entitled ``Federalism'' (64 FR 43255; August
10, 1999), requires EPA to develop an accountable process to ensure
``meaningful and timely input by State and local officials in the
development of regulatory policies that have federalism implications.''
``Policies that have federalism implications'' is defined in the
Executive Order to include regulations that have ``substantial direct
effects on the States, on the relationship between the national
government and the States, or on the distribution of power and
responsibilities among the various levels of government.''
This final rule does not have federalism implications. It will not
have substantial direct effects on the States, on the relationship
between the national government and the States, or on the distribution
of power and responsibilities among the various levels of government,
as specified in Executive Order 13132. Today's rule promulgates
revisions to WET test methods that are currently approved for use in
NPDES permits and certification of Federal licenses under the CWA. The
revisions are minor and the cost to implement them is minimal. Thus,
Executive Order 13132 does not apply to this rule.
F. Executive Order 13175: Consultation and Coordination with Indian
Tribal Governments
Executive Order 13175, entitled ``Consultation and Coordination
with Indian Tribal Governments'' (59 FR 22951; November 9, 2000),
requires EPA to develop an accountable process to ensure ``meaningful
and timely input by Tribal officials in the development of regulatory
policies that have Tribal implications.'' ``Policies that have Tribal
implications'' is defined in the Executive Order to include regulations
that have ``substantial direct effects on one or more Indian Tribes, on
the relationship between the Federal government and the Indian Tribes
or on the distribution of power and responsibilities between the
Federal government and Indian Tribes.''
This final rule does not have Tribal implications. It will not have
substantial direct effects on Tribal governments, on the relationship
between the Federal government and Indian Tribes, or on the
distribution of power and responsibilities between the Federal
government and Indian Tribes, as specified in Executive Order 13175.
Today's rule promulgates revisions to WET test methods that are
currently approved for use in NPDES permits and certification of
Federal licenses under the CWA. The revisions are minor and the cost to
implement them is minimal. Thus, Executive Order 13175 does not apply
to this rule.
G. Executive Order 13045: Protection of Children From Environmental
Health and Safety Risks
Executive Order 13045 (62 FR 19885; April 23, 1997) applies to any
rule that: (1) is determined to be ``economically significant'' as
defined under Executive Order 12866, and (2) concerns an environmental
health or safety risk that EPA has reason to believe may have a
disproportionate effect on children. If the regulatory action meets
both criteria, the Agency must evaluate the environmental health or
safety effects of the planned rule on children, and explain why the
planned regulation is preferable to other potentially effective and
reasonably feasible alternatives considered by the Agency. This rule is
not subject to the Executive Order because it is neither ``economically
significant'' as defined in Executive Order 12866, nor does it concern
an environmental health or safety risk that EPA has reason to believe
may have a disproportionate effect on children.
H. Executive Order 13211: Actions Concerning Regulations That
Significantly Affect Energy Supply, Distribution, or Use
This rule is not subject to Executive Order 13211, ``Actions
Concerning Regulations That Significantly Affect Energy Supply,
Distribution, or Use'' (66 FR 28355; May 22, 2001) because it is not a
significant regulatory action under Executive Order 12866.
I. National Technology Transfer and Advancement Act
Section 12(d) of the National Technology Transfer and Advancement
Act of 1995, (``NTTAA''), Public Law 104-113, section 12(d) (15 U.S.C.
272 note), directs EPA to use voluntary consensus standards in its
regulatory activities unless to do so would be inconsistent with
applicable law or otherwise impractical. Voluntary consensus standards
are technical standards (e.g., material specifications, test methods,
sampling procedures, business practices) that are developed or adopted
by voluntary consensus standards bodies (VCSBs). The NTTAA directs EPA
to provide Congress, through the Office of Management and Budget (OMB),
explanations when the Agency decides not to use available and
applicable voluntary consensus standards.
This rulemaking would revise existing EPA WET test methods. For the
methods that EPA is revising, the Agency did not conduct a search to
identify potentially applicable voluntary consensus standards, because
the revisions EPA is promulgating today would merely incorporate more
specificity and detail into currently approved EPA test methods. EPA
did, however, consult available voluntary consensus standards, such as
ASTM standards, for guidance in conducting the Interlaboratory
Variability Study and in defining certain performance characteristics
of the methods.
J. Congressional Review Act
The Congressional Review Act, 5 U.S.C. 801 et seq., as added by the
Small Business Regulatory Enforcement
[[Page 69971]]
Fairness Act of 1996 (SBREFA), generally provides that before a rule
may take effect, the agency promulgating the rule must submit a rule
report, which includes a copy of the rule, to each House of the
Congress and to the Comptroller General of the United States. EPA will
submit a report containing this rule and other required information to
the U.S. Senate, the U.S. House of Representatives, and the Comptroller
General of the United States prior to publication of the rule in the
Federal Register. A major rule cannot take effect until 60 days after
it is published in the Federal Register. This action is not a ``major
rule'' as defined by 5 U.S.C. 804(2). This rule will be effective on
December 19, 2002.
VIII. References
American Society for Testing and Materials. 1998. Standard
Practice for Determination of Precision and Bias of Applicable Test
Methods of Committee D-19 on Water. D 2777-98. American Society for
Testing and Materials, Philadelphia, PA.
DeGraeve, G.M., G.J. Smith, W.H. Clement, D.O. McIntyre, and T.
Forgette. 1998. WET Testing Program: Evaluation of Practices and
Implementation. Project 94-HHE-1. Water Environment Research
Foundation, Alexandria, VA.
Edison Electric Institute et al. v. EPA, Settlement Agreement,
July 24, 1998. U.S. Court of Appeals, D.C. Circuit, No. 96-1062.
Lone Star Steel v. EPA, Settlement Agreement, March 4, 1998.
U.S. Court of Appeals, D.C. Circuit, No. 96-1157.
National Environmental Laboratory Accreditation Conference,
2001. Quality Systems. National Environmental Laboratory
Accreditation Conference.
U.S. Environmental Protection Agency. 1988. Availability,
Adequacy, and Comparability of Testing Procedures for the Analysis
of Pollutants Established Under Section 304(h) of the Federal Water
Pollution Control Act; Report to Congress. EPA/600/9-87/030. U.S.
Environmental Protection Agency, Environmental Monitoring Systems
Laboratory, Cincinnati, OH.
U.S. Environmental Protection Agency. 1991. Technical Support
Document for Water Quality-Based Toxics Control. EPA/505/2-90/001.
U.S. Environmental Protection Agency, Office of Water Enforcement
and Permits, and Office of Water Regulations and Standards,
Washington, DC.
U.S. Environmental Protection Agency. 1993. Methods for
Measuring the Acute Toxicity of Effluents and Receiving Waters to
Freshwater and Marine Organisms, 4th ed. EPA/600/4-90/027F. U.S.
Environmental Protection Agency, Environmental Monitoring Systems
Laboratory, Cincinnati, OH.
U.S. Environmental Protection Agency. 1994a. Short-term Methods
for Estimating the Chronic Toxicity of Effluents and Receiving
Waters to Freshwater Organisms, 3rd ed. EPA/600/4-91/002. U.S.
Environmental Protection Agency, Environmental Monitoring Systems
Laboratory, Cincinnati, OH.
U.S. Environmental Protection Agency. 1994b. Short-term Methods
for Estimating the Chronic Toxicity of Effluents and Receiving
Waters to Marine and Estuarine Organisms, 2nd ed. EPA/600/4-91/003.
U.S. Environmental Protection Agency, Environmental Monitoring
Systems Laboratory, Cincinnati, OH.
U.S. Environmental Protection Agency. 1995a. National Policy
Regarding Whole Effluent Toxicity Enforcement, July 6, 1995,
memorandum from Brian Maas, U.S. Environmental Protection Agency,
Office of Enforcement and Compliance Assurance, Washington DC.
U.S. Environmental Protection Agency. 1995b. Short-term Methods
for Estimating the Chronic Toxicity of Effluents and Receiving
Waters to West Coast Marine and Estuarine Organisms, 1st ed. EPA/
600/R-95/136. U.S. Environmental Protection Agency, National
Exposure Research Laboratory, Cincinnati, OH.
U.S. Environmental Protection Agency. 1995c. Whole effluent
toxicity: guidelines establishing test procedures for the analysis
of pollutants, final rule. 60 FR 53529-53563.
U.S. Environmental Protection Agency. 1999. Whole effluent
toxicity: guidelines establishing test procedures for the analysis
of pollutants, whole effluent toxicity tests; final rule, technical
correction. 64 FR 4975-4991.
U.S. Environmental Protection Agency. 2000a. Method Guidance and
Recommendations for Whole Effluent Toxicity (WET) Testing (40 CFR
Part 136). EPA/821/B-00/004. U.S. Environmental Protection Agency,
Office of Water, Washington, DC.
U.S. Environmental Protection Agency. 2000b. Notice of
availability, ``Understanding and accounting for method variability
in WET applications under the NPDES program.'' 65 FR 44528-44529.
U.S. Environmental Protection Agency. 2000c. Understanding and
Accounting for Method Variability in Whole Effluent Toxicity
Applications Under the National Pollutant Discharge Elimination
System Program. EPA/833/R-00/003. U.S. Environmental Protection
Agency, Office of Wastewater Management, Washington, DC.
U.S. Environmental Protection Agency. 2000d. Whole effluent
toxicity test methods guidance; notice of availability. 65 FR 46457-
46458.
U.S. Environmental Protection Agency. 2001a. Final Report:
Interlaboratory Variability Study of EPA Short-term Chronic and
Acute Whole Effluent Toxicity Test Methods, Vol. 1. EPA /821/B-01/
004. U.S. Environmental Protection Agency, Office of Water,
Washington, DC.
U.S. Environmental Protection Agency. 2001b. Final Report:
Interlaboratory Variability Study of EPA Short-term Chronic and
Acute Whole Effluent Toxicity Test Methods, Vol. 2: Appendix. EPA/
821/B-01-005. U.S. Environmental Protection Agency, Office of Water,
Washington, DC.
U.S. Environmental Protection Agency. 2001c. Guidelines
establishing test procedures for the analysis of pollutants; whole
effluent toxicity test methods; proposed rule. 66 FR 49794-49816.
U.S. Environmental Protection Agency. 2001d. Guidelines
establishing test procedures for the analysis of pollutants; whole
effluent toxicity test methods; extension of comment period. 66 FR
58693-58694.
U.S. Environmental Protection Agency. 2001e. Proposed Changes to
Whole Effluent Toxicity Method Manuals. EPA/821/B-01/002. U.S.
Environmental Protection Agency, Office of Water, Washington, DC.
U.S. Environmental Protection Agency. 2001f. Clarifications
Regarding Toxicity Reduction and Identification Evaluations in the
National Pollutant Discharge Elimination System Program. U.S.
Environmental Protection Agency, Office of Water, Washington, DC.
U.S. Environmental Protection Agency. 2002. Cost Estimate for
Required Method Modifications in EPA's WET Final Rule, October 16,
2002, memorandum from Marion Kelly, U.S. Environmental Protection
Agency, Office of Water, Office of Science and Technology,
Washington, DC.
Waller, T.W., L.P. Ammann, W.J. Birge, K.L. Dickson, P.B. Dorn,
N.E. LeBlanc, D.I. Mount, B.R. Parkhurst, H.R. Preston, S.C.
Schimmel, A. Spacie, and G.B. Thursby. 1996. Discussion synopsis,
predicting instream effects from WET tests. Chapter 9 In: Whole
Effluent Toxicity Testing: An Evaluation of Methods and Prediction
of Receiving System Impacts. D.R. Grothe, K.L. Dickson, and D.K.
Reed-Judkins, eds., SETAC Press, Pensacola, FL, pp. 271-286.
List of Subjects at 40 CFR Part 136
Environmental protection, Incorporation by reference, Reporting and
recordkeeping requirements, Water pollution control.
Dated: November 8, 2002.
Christine Todd Whitman,
Administrator.
For the reasons set out in the preamble, title 40, chapter I of the
Code of Federal Regulations, is amended as follows:
PART 136--GUIDELINES ESTABLISHING TEST PROCEDURES FOR THE ANALYSIS
OF POLLUTANTS
1. The authority citation for part 136 continues to read as
follows:
Authority: Secs. 301, 304(h), 307, and 501(a), Pub. L. 95-217,
91 Stat. 1566, et seq. (33 U.S.C. 1251, et seq.) (The Federal Water
Pollution Control Act Amendments of 1972 as amended by the Clean
Water Act of 1977).
2. Section 136.3 is amended:
a. In Table IA of paragraph (a) by revising entries 6 to 9.
b. In paragraph (b) by revising references (34), (38), and (39).
c. In paragraph (b) by removing and reserving reference (42).
The revisions read as follows:
[[Page 69972]]
Sec. 136.3 Identification of test procedures.
(a) * * *
Table IA.--List of Approved Biological Methods
--------------------------------------------------------------------------------------------------------------------------------------------------------
Standard methods
Parameter and units Method \1\ EPA 18th, 19th, 20th Ed. ASTM USGS
--------------------------------------------------------------------------------------------------------------------------------------------------------
* * * * * * *
Aquatic Toxicity:
6. Toxicity, acute, fresh Ceriodaphnia dubia acute..................... \7\
water organisms, LC50, Daphnia pulex and Daphnia magna acute........ 2002.0
percent effluent. Fathead minnow, Pimephales promelas, and \7\
Bannerfin shiner, Cyprinella leedsi, acute. 2021.0
\7\
2001.0
Rainbow trout, Oncorhynchus mykiss, and brook \7\
trout, Salvelinus fontinalis, acute. 2019.0
7. Toxicity, acute, estuarine Mysid, Mysidopsis, bahia, acute.............. \7\
and marine organisms of the Sheepshead minnow, Cyprinodon variegatus, 2007.0
Atlantic Ocean and Gulf of acute. \7\
Mexico, LC50, percent Silverside, Menidia beryllina, Menidia 2004.0
effluent. menidia, and Menidia peninsulae, acute. \7\
2006.0
8. Toxicity, chronic, fresh Fathead minnow, Pimephales promelas, larval \8\
water organisms, NOEC or survival and growth. 1000.0
IC25, percent effluent. Fathead minnow, Pimephales promelas, embryo- .........
larval survival and teratogenicity. \8\
1001.0
Daphnia, Ceriodaphnia dubia, survival and \8\
reproduction. 1002.0
Green alga, Selenastrum capricornutum, growth \8\
1003.0
9. Toxicity, chronic, Sheepshead minnow, Cyprinodon variegatus, \9\
estuarine and marine larval survival and growth. 1004.0
organisms of the Atlantic Sheepshead minnow, Cyprinodon variegatus, .........
Ocean and Gulf of Mexico, embryo-larval survival and teratogenicty. \9\
NOEC or IC25, percent Inland silverside, Menidia beryllina, larval 1005.0
effluent. survival and growth. .........
Mysid, Mysidopsis bahia, survival, growth, \9\
and fecundity. 1006.0
.........
\9\
1007.0
Sea urchin, Arbacia punctulata, fertilization \9\
1008.0
--------------------------------------------------------------------------------------------------------------------------------------------------------
\1\ The method must be specified when results are reported.
* * * * * * *
\7\ USEPA. October 2002. Methods for Measuring the Acute Toxicity of Effluents and Receiving Waters to Freshwater and Marine Organisms. Fifth Edition.
U.S. Environmental Protection Agency, Office of Water, Washington, D.C. EPA 821-R-02-012.
\8\ USEPA. October 2002. Short-Term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Freshwater Organisms. Fourth
Edition. U.S. Environmental Protection Agency, Office of Water, Washington, D.C. EPA 821-R-02-013.
\9\ USEPA. October 2002. Short-Term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Marine and Estuarine Organisms.
Third Edition. U.S. Environmental Protection Agency, Office of Water, Washington, D.C. EPA 821-R-02-014.
* * * * * * *
* * * * *
(b) * * *
References, Sources, Costs, and Table Citations
* * * * *
(34) USEPA. October 2002. Methods for Measuring the Acute Toxicity
of Effluents and Receiving Waters to Freshwater and Marine Organisms.
Fifth Edition. U.S. Environmental Protection Agency, Office of Water,
Washington, D.C. EPA 821-R-02-012. Available from: National Technical
Information Service, 5285 Port Royal Road, Springfield, Virginia 22161,
Publ. No. PB2002-108488. Table IA, Note 7.
* * * * *
(38) USEPA. October 2002. Short-Term Methods for Estimating the
Chronic Toxicity of Effluents and Receiving Waters to Freshwater
Organisms. Fourth Edition. U.S. Environmental Protection Agency, Office
of Water, Washington, D.C. EPA 821-R-02-013. Available from: National
Technical Information Service, 5285 Port Royal Road, Springfield,
Virginia 22161, Publ. No. PB2002-108489. Table IA, Note 8.
(39) USEPA. October 2002. Short-Term Methods for Estimating the
Chronic Toxicity of Effluents and Receiving Waters to Marine and
Estuarine Organisms. Third Edition. U.S. Environmental Protection
Agency, Office of Water, Washington, D.C. EPA 821-R-02-014. Available
from: National Technical Information Service, 5285 Port Royal Road,
Springfield, Virginia 22161, Publ. No. PB2002-108490. Table IA, Note 9.
* * * * *
(42) [RESERVED]
* * * * *
[FR Doc. 02-29072 Filed 11-18-02; 8:45 am]
BILLING CODE 6560-50-P
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