D19.24 - Water Microbiology

SIGNIFICANCE AND USE
5.1 Calcium Carbonate (CaCO3) buffered formalin (3 % to 5 %) can be used as a permanent preservative for zooplankton. Lugol’s iodine solution can be used to preserve zooplankton for up to one year. Thirty percent ethanol, 30 % glutaraldehyde, or 25 % vinegar (can use 3 % acetic acid solution) can be used for more temporary storage and preservation of zooplankton samples. A 25 % vinegar solution is preferred to preserve soft-bodied planktonic coelenterates.
SCOPE
1.1 This practice describes the proper procedures for preserving zooplankton samples with either formaldehyde, ethanol, glutaraldehyde, Lugol’s iodine solution, or vinegar (acetic acid).  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

ABSTRACT
This practice covers determination of the quantitative and qualitative species composition of fish in a specified area. The successful use of this technique is dependent on: (1) preventing fish from escaping the sample area and (2) retrieving all affected fish, which may take up to three days. This practice is useful in both short- and long-term studies for management and impact assessment purposes. The sample area is blocked off with a small mesh net(s) and the volume of water to be treated is calculated. The required quantity of rotenone is diluted and distributed throughout the water column in the sample area. All fish should be affected and they should be collected for processing.
SCOPE
1.1 This practice covers determination of the quantitative and qualitative species composition of fish in a specified area. The successful use of this technique is dependent on: (1) preventing fish from escaping the sample area and (2) retrieving all affected fish, which may take up to three days.  
1.2 Advantages:  
1.2.1 Easily detoxified.  
1.2.2 All native freshwater fish are susceptible, but it has low toxicity to mammals and birds.  
1.2.3 At low concentrations fish toxicity depends on species, age, and size.  
1.2.4 The suffocating action is reversible.  
1.3 Limitations:  
1.3.1 It is less effective in cold (below 20 °C) and highly alkaline water.  
1.3.2 Smaller fish and those without air bladders usually do not float.  
1.3.3 Completely random selection of sample areas is not possible.  
1.3.4 Overkill beyond sample area can sometimes occur.  
1.3.5 Food web organisms may be eliminated.  
1.4 Applications—This practice is useful in both short- and long-term studies for management and impact assessment purposes. It is adaptable to both lotic and lentic situations in littoral and limnetic areas.  
1.5 The values stated in inch-pound units are to be regarded as standard. The values given in parentheses are mathematical conversions to SI units that are provided for information only and are not considered standard.  
1.6 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazards, see Section 7.  
1.7 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 P. aeruginosa is an opportunistic pathogen and has been linked as the causative agent of numerous infections that may be transmitted through a contaminated water supply to a susceptible host.
Note 1: Fecal waste is >95 % E. coli which is found in humans and warm blooded animals.  
5.2 The membrane filtration procedure described is a rapid and reliable test method of detecting P. aeruginosa in water.
SCOPE
1.1 The test method covers the isolation and enumeration of Pseudomonas aeruginosa. Testing was performed on spiked samples using reagent grade water as the diluent from surface waters; recreational waters; ground water, water supplies; especially rural nonchlorinated sources; waste water; and saline waters. The detection limit of this test method is one microorganism per 100 mL.  
1.2 This test method was used successfully with reagent water. It is the user's responsibility to ensure the validity of this test method for surface waters, recreational waters, ground water, rural nonchlorinated sources; waste water; and saline waters.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 10.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 The advantages of collecting zooplankton with pumps are as follows:  
5.1.1 Sample size is more accurately controlled than with nets.  
5.1.2 Discrete samples can be more easily obtained both vertically and horizontally.  
5.1.3 Multiple or replicate samples can be more easily obtained.  
5.1.4 The pumps are adaptable to a variety of ecosystems less than 30 m deep.  
5.1.5 Sampling efficiency does not decrease with sample size.  
5.2 The disadvantages of collecting zooplankton with pumps are as follows:  
5.2.1 Pumps are bulky and require an electrical source.  
5.2.2 Pumps are generally more costly than nets.  
5.2.3 Pumps generally discriminate against collecting macroplankton.  
5.2.4 Pump intake tubes may be avoided by the more motile zooplankton forms.  
5.2.5 Requires a long, bulky, intake tube for deep water sampling.  
5.3 There are several special considerations that should be observed when collecting zooplankton with a pump. They are:  
5.3.1 Some pumps can fragment zooplankton and induce mortality due to their design.  
5.3.2 The pump hose must be cleared before taking the next sample.
SCOPE
1.1 This practice covers the procedures for obtaining qualitative/quantitative samples of a zooplankton community by use of pumping systems.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 This test method is useful for measuring recreational water quality and chlorinated wastewaters, although it can be used for any water suspected of contamination by fecal wastes of warm-blooded animals. The significance of finding E. coli in recreational water samples, especially samples obtained from fresh recreational waters, is that there is a risk of gastrointestinal illness, directly related to the E. coli density, associated with swimming.5  
5.2 Since small or large volumes of water or dilutions thereof can be analyzed by the MF technique, a wider range of levels of E. coli in water can be detected and enumerated than with other methods.
SCOPE
1.1 This test method describes a membrane filter (MF) procedure for the detection and enumeration of Escherichia coli, a bacterium found exclusively in the feces of humans and other warm-blooded animals. The presence of these microorganisms in water is an indication of fecal pollution and the possible presence of enteric pathogens. These bacteria are found in water and wastewater in a wide range of densities. The detection limit of this procedure is one colony forming unit (CFU) per volume filtered.  
1.2 This test method has been used successfully with temperate fresh and marine ambient waters, and wastewaters. It is the user’s responsibility to ensure the validity of this test method for waters of other types.  
1.3 The values stated in SI units are to be regarded as standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 This practice provides uniform guidance for cleaning the laboratory glassware, plasticware, and equipment used in routine microbiological analyses. However, tests that are extremely sensitive to toxic agents (such as virus assays) may require more stringent cleaning practices.2
SCOPE
1.1 In microbiology, clean glassware is crucial to ensure valid results. Previously used or new glassware must be thoroughly cleaned. Laboratory ware and equipment that are not chemically clean are responsible for considerable losses in personnel time and supplies in many laboratories. These losses may occur as down time when experiments clearly have been adversely affected and as invalid data that are often attributed to experimental error. Chemical contaminants that adversely affect experimental results are not always easily detected. This practice describes the procedures for producing chemically clean glassware.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific precautions, see Section 6, 5.7.3.1, and 8.3.1.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Sulfate-reducing bacteria are widely distributed in marine and fresh water muds which, in consequence, frequently are laden with the hydrogen sulfide produced by these organisms during dissimilatory sulfate reduction.  
5.2 It has been reported that Desulfovibrio spp. can form as much as 10 g of sulfide per litre during active multiplication. Sulfate-reducing bacteria can cause the external or internal corrosion of water or wastewater pipelines and pipelines for petroleum and natural gas. The formation of galvanic cells by massive growth of sulfate-reducing bacteria under suitable conditions makes the corrosion much worse than just the effect of the hydrogen sulfide on the metal or concrete.
SCOPE
1.1 These test methods cover the procedure for the detection and enumeration by the most probable number (MPN) technique of sulfate-reducing bacteria in water or water-formed deposits.  
1.2 Two media preparations are provided. Medium A which is prepared with reagent grade water, and Medium B which is prepared using the water to be sampled as the water source. Medium B is offered for those special conditions where sulfate-reducing bacterial strains have adapted to atypical non-fresh water environment.  
1.3 For the isolation and enumeration of thermophilic sulfate-reducing bacteria encountered in waters associated with oil and gas production, all broths, dilution blanks, and incubations must be maintained at temperatures of at least 45 °C and preferably within 5 °C at the sample temperature.  
1.4 The sensitivity of these test methods can be increased by purging the dilution blanks and tubes of media with nitrogen immediately prior to use.  
1.5 The analyst should be aware that adequate collaborative data for precision and bias statements as required by Practice D2777 are not provided. See Section 11 for details.  
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.7 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.8 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 The enterococci are indicators of the bacteriological quality for potable water, shellfish growing waters, ambient, and recreational waters. A direct relationship between swimming, associated gastroenteritis, and enterococci has been established through epidemiological studies and marine and fresh water bathing beaches. These studies have led to the development of criteria that can be used to establish bathing water standards based on established health-water quality relationships.  
5.2 Since small or large volumes of water or dilutions thereof, can be analyzed by the membrane filter technique, a wide range of levels of enterococci in water can be enumerated and detected.
SCOPE
1.1 This test method covers a membrane filter (MF) procedure for the detection and enumeration of the enterococci bacteria in water. The enterococci, which include Entero-coccus faecalis (E. faecalis), E. faecium, and their varieties are commonly found in the feces of humans and other warm-blooded animals. Although some strains are ubiquitous and not related to fecal pollution, enterococci in water are an indication of fecal pollution and the possible presence of enteric pathogens. These bacteria are found in water and wastewater in a wide range of densities. The detection limit is one colony forming unit (CFU)/volume filtered.  
1.2 This test method has been used successfully with temperate fresh and marine ambient waters, and wastewaters. It is the user’s responsibility to ensure the validity of this test method for waters of untested types.  
1.3 The values stated in SI units are to be regarded as standard. The values given in parentheses after SI units are included for information only and are not considered standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 This test provides an easy and reliable method for the detection of enterococci in water within 24 h. For recreational water (fresh and marine) testing is performed to insure areas are safe for swimming. Enterolert also can be used for testing bottled water, wastewater, ground water, and drinking water.
SCOPE
1.1 This test method covers a simple procedure for the detection of enterococci in water and wastewater. It is based on IDEXX’s patented Defined Substrate Technology (DST).2 This product, Enterolert, utilizes a nutrient indicator that fluoresces when metabolized. It can detect these bacteria at one most probable number (MPN)/100 mL within 24 h. The presence of this microorganism in water is an indication of fecal contamination and the possible presence of enteric pathogens.  
1.2 This test method can be used successfully with drinking water, source water, recreational (fresh and marine) water, wastewater, and bottled water. It is the user’s responsibility to ensure the validity of this test method for waters of untested matrices.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 The advantages of the Clarke-Bumpus plankton sampler are as follows:  
5.1.1 It will sample a discrete depth or multiple depths, depending upon the sampling design.  
5.1.2 It is a slow to medium speed sampler requiring a towing speed of three to five knots.  
5.1.3 The sample size can be easily controlled.  
5.1.4 The sampler is lightweight and can be used without auxiliary equipment.  
5.1.5 It has a relatively high filtration efficiency factor of 0.88.  
5.1.6 It is a versatile sampler and can be used in all but the shallowest waters.  
5.1.7 The flowmeter records the amount of water that passes into the net.  
5.1.8 Overspill of water at the mouth of the net due to excess speed of towing is of minimal consequence.  
5.2 The disadvantages of the Clarke-Bumpus plankton sampler are as follows:  
5.2.1 The flowmeter requires frequent maintenance including calibration and lubrication.  
5.2.2 It is not suitable for use in very small areas or shallow waters.  
5.3 There are several special considerations that shall be observed when using a Clarke-Bumpus plankton sampler. They are:  
5.3.1 The flowmeter should be calibrated and serviced frequently to ensure efficient and accurate operation.  
5.3.2 The sampler is relatively fragile, particularly the closing device and flowmeter. This necessitates careful deployment and recovery procedures.  
5.3.3 Following each collection, the net must be thoroughly washed.  
5.3.4 Special attention must be given to the strength of the cable and its attachment to avoid loss of the sampler.  
5.3.5 The sampler should not be used in beds of macrophytes, in waters containing submerged objects, or close to the bottom.  
5.3.6 The net should be inspected frequently for pin-size holes, tears, net deterioration, and other anomalies.  
5.3.7 Following use, the wet net should be suspended full length in the air in subdued light and allowed to dry.
SCOPE
1.1 This practice covers the procedures for obtaining quantitative samples of a zooplankton community by use of a Clarke-Bumpus plankton sampler.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 A rapid and routine procedure for determining biomass of the living microorganisms in cultures, waters, wastewaters, and in plankton and periphyton samples taken from surface waters is frequently of vital importance. However, classical techniques such as direct microscope counts, turbidity, organic chemical analyses, cell tagging, and plate counts are expensive, time-consuming, or tend to underestimate total numbers. In addition, some of these methods do not distinguish between living and nonliving cells.  
5.2 This test method measures the concentration of cellular-ATP present in the sample. ATP is a constituent of all living cells, including bacteria, algae, protozoa, and fungi. Consequently, the presence of cellular-ATP is an indicator of total metabolically active microbial contamination in water. ATP is not associated with matter of non-biological origin.  
5.3 The ATP (luciferin-luciferase) method is a rapid, sensitive determination of viable microbial biomass. ATP is the primary energy donor for life processes, does not exist in association with nonliving detrital material, and the amount of ATP per unit of biomass (expressed in weight) is relatively constant. (ATP per cell varies with species and physiological state of the organism.)  
5.4 This test method can be used to:  
5.4.1 Estimate viable microbial biomass in cultures and waters.  
5.4.2 Estimate the amount of total viable biomass in plankton and periphyton samples.  
5.4.3 Estimate the number of viable cells in a unispecies culture if the cATP content (or if the average amount of cATP) per cell is known.  
5.4.4 Estimate and differentiate between zooplanktonic, phytoplanktonic, bacterial, and fungal cATP through size fractionation of water samples.  
5.4.5 Measure the mortality rate of microorganisms in toxicity tests in entrainment studies, and in other situations where populations or assemblages of microorganisms are placed under stress.  
5.5 This test method is similar to Test Metho...
SCOPE
1.1 This test method covers a protocol for capturing, extracting and quantifying the cellular adenosine triphosphate (cATP) content associated with microorganisms normally found in laboratory cultures and waters in plankton and periphyton samples from waters.  
1.2 The ATP is measured using a bioluminescence enzyme assay, whereby light is generated in amounts proportional to the concentration of ATP in the samples. The light is produced and measured quantitatively as relative light units (RLU) which are converted by comparison with an ATP standard and computation to pg ATP/mL.  
1.3 This method does not remove all known chemical interferences, known to either luminesce in the 530 nm ± 20 nm range, or to quench light emitted in that range. It should not be used to determine ATP concentrations in samples with dissolved organic compounds, heavy metals or >10 000 ppm total dissolved solids. Alternative methods have been developed for determining ATP concentrations in fluids samples likely to contain such interferences (Test Methods D7687 and E2694).  
1.4 Knowledge of the concentration of ATP can be related to viable biomass or metabolic activity of microorganisms (Appendix X1).  
1.5 This test method offers a high degree of sensitivity, rapidity, accuracy, and reproducibility.  
1.6 The analyst should be aware that the precision statement pertains only to determinations in reagent water and not necessarily in the matrix being tested.  
1.7 This test method is equally suitable for use in the laboratory or field.  
1.8 The method normally detects cATP concentrations in the range of 0.1 pg cATP/mL (–1.0Log10 [pg cATP/mL]) to
4 000 000 pg cATP/mL (6.6 Log10 [pg cATP/mL]) in 50 mL water samples.  
1.9 Providing interferences can be overcome, bioluminescence is a reliable and proven method for qualifying and quantifying ATP, although the method does not differentiate between ATP from different sources, for example, from...

SIGNIFICANCE AND USE
5.1 This plate format is useful for the routine monitoring of culturable, waterborne bacteria in potable and non-potable waters. The significance of finding these bacteria can help with identifying water quality or water system problems or evaluate compliance with maintenance protocols. This test method uses small volumes of water, or dilutions thereof, and provides an easy and reliable method that eliminates media preparation and reduces laboratory waste.
SCOPE
1.1 This test method describes a simple procedure for the quantification of culturable, waterborne bacteria in potable water (drinking water, bottled water, and dental water, for example) and non-potable waters (cooling towers, for example).  
1.1.1 The EasyDisc2, 3 plate format is designed to test 1 mL of a water sample on a 47 mm gridded plate containing a growth reagent embedded to the plate’s inner surface.  
1.1.2 Detection is based on colorimetric technology in which viable, aerobic, heterotrophic, waterborne bacteria grow when present in the water sample, displaying a color reaction which allows for a simplified visualization of colony growth.  
1.2 Each plate can accurately detect up to 300 colony forming units per 1 mL (CFU/1 mL) of sample. To increase the quantification range, a sample dilution can be used. Adjust the CFU/mL result to reflect dilutions.  
1.3 This test method can be used for potable (for example, drinking, bottled, and dental) waters and non-potable waters such as cooling tower waters. It is the user’s responsibility to adhere to all requirements by local regulations and ensure the validity of this test method for waters other than those tested as part of the Interlaboratory Study (ILS).  
1.4 The values stated in SI units are to be regarded as the standard. No other units of measurement are included in this standard.  
1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.6 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Microbiological water testing procedures using membrane filtration are based on the premise that all bacteria within a specific size range will be retained by the membrane filter used. If the membrane filter does not retain these bacteria, false negative results or lowered density estimates may occur that could have serious repercussions due to the presence of unrecognized potential health hazards in the water being tested, especially in drinking water.  
5.2 This procedure as devised will enable the user to test each membrane filter lot number for its ability to retain all bacteria equal to, or larger than, the stated membrane pore size.
SCOPE
1.1 This test method covers a procedure to test membrane filters for their ability to retain bacteria whose diameter is equal to or slightly larger than membrane filters with pore size rated at 0.40 to 0.45 μm.  
1.2 The procedures described are for the use of user laboratories as differentiated from manufacturers’ laboratories.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 This test provides an easy and reliable method for the detection of L. pneumophila in potable and non-potable waters in 7 days.  
5.2 Routine monitoring for L. pneumophila determines whether implemented control measures are effective, such as those outlined in a water safety program (2).  
5.2.1 Water system management is necessary to maintain L. pneumophila concentrations below hazardous levels. Through routine measurement of L. pneumophila levels, a monitoring program can ensure that control measures are effective and implemented when necessary in response to increasing levels. Water samples may be examined for L. pneumophila during epidemiological investigations as part of local authority, industrial, or hospital programs, or in order to validate treatment control methods. Routine sampling could also be carried out based on risk assessments or on local, state, or federal requirements or guidelines.
SCOPE
1.1 This test method describes a simple procedure for the detection of Legionella pneumophila (L. pneumophila) in potable water and non-potable waters (cooling towers, for example). This procedure describes a liquid culture method based on a bacterial enzyme technology. The detection of L. pneumophila is signaled through the utilization of a substrate present in the Legiolert reagent. L. pneumophila cells grow rapidly and reproduce using the rich supply of amino acids, vitamins and other nutrients present in the Legiolert reagent. Actively growing strains of L. pneumophila use the added substrate to produce a brown color indicator or produce turbid growth with or without brown coloration. Legiolert can detect this bacterial species at the following minimum concentrations based on the protocol employed:  
1.1.1 Potable Water:  
1.1.1.1 ≥1 organism / 100 mL at 7 days for 100 mL potable protocol.
1.1.1.2 ≥1 organism / 10 mL at 7 days for 10 mL potable protocol.  
1.1.2 Non-potable Water:  
1.1.2.1 ≥1 organism / 1.0 mL at 7 days for 1.0 mL non-potable protocol.
1.1.2.2 ≥1 organism / 0.1 mL at 7 days for 0.1 mL non-potable protocol.  
1.1.3 This test method can be used for potable (drinking) waters and non-potable waters such as cooling tower waters (1).3 It is the user’s responsibility to ensure the validity of this test method for waters of untested matrices.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 Fish sampling includes a number of lethal and non-lethal practices.  
4.2 This guide provides an overview of commonly used fish sampling practices.  
4.3 This summary serves as a brief accounting of options available to personnel responsible for determining the fish sampling practice or practices that best serve the sampling objectives.
SCOPE
1.1 This guide covers the use of lethal and non-lethal collection practices for fish.  
1.1.1 Lethal practices include the use of rotenone and antimycin which are used to collect or eradicate fish; numerous chemicals have been used but presently only rotenone and antimycin are U.S. Environmental Protection Agency (EPA)-approved for this use.  
1.2 Non-lethal collection practices typically do not cause mortality to fish.  
1.2.1 Non-lethal practices include surface or bank observation, underwater observation, gill netting, beach seines, hoop nets, fyke nets, trap nets, electroshocking, minnow traps, enclosure (pop drop and throw) traps, angler surveys, commercial surveys.  
1.3 The focus of this guide is to provide sampling practices for fish collection. This standard does not cover the identification of species or any statistical methods for the sampling data.  
1.4 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Refer to the MSDSs for all chemicals used in this procedure.  
1.6 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 Numerous ASTM test methods and practices (for example: Test Methods D5259 and D5392, and Practices D6974 and E2563) report colony counts as their measured parameter.  
4.2 These practices provide a uniform set of counting, calculating, and reporting procedures for ASTM test methods in microbiology.    
Section  
A—Counting Colonies on Membrane Filters  
6  
B—Counting Colonies on Pour Plates  
7  
C—Counting Colonies on Spread Plates  
8  
4.3 The counting rules provide a best attainable estimate of microorganisms in the sample, since the samples cannot be held and reanalyzed at a later date.
SCOPE
1.1 These practices cover recommended procedures for counting colonies and reporting colony-forming units (CFU) on membrane filters (MF) and standard pour and spread plates.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Data on the chlorophyll content of the algae have the following applications:  
5.1.1 To provide estimates of algal biomass and productivity.  
5.1.2 To provide general information on the taxonomic composition (major groups) of the algae, based on the relative amounts of chlorophyll a, b, and c, and the physiological condition of algal communities, which is related to the relative abundance of pheopigments.  
5.1.3 To determine long-term trends in water quality.  
5.1.4 To determine the trophic status of surface waters.  
5.1.5 To detect adverse effects of pollutants on plankton and periphyton in receiving waters.  
5.1.6 To determine maximum growth rates and yields in algal growth potential tests.
SCOPE
1.1 These practices include the extraction and the measurement of chlorophyll a, b, and c, and pheophytin a in freshwater and marine plankton and periphyton. Three practices are provided as follows:  
1.1.1 Spectrophotometric, trichromatic practice for measuring chlorophyll a, b, and c.  
1.1.2 Spectrophotometric, monochromatic practice for measuring chlorophyll a corrected for pheophytin a; and for measuring pheophytin a.  
1.1.3 Fluorometric practice for measuring chlorophyll a corrected for pheophytin a; and for measuring pheophytin a.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Specific precautionary statements are given in Section 8.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Filamentous iron bacteria is a general classification for microorganisms that utilize ferrous iron as a source of energy and are characterized by the deposition of ferric hydroxide in their mucilaginous sheaths. The process is continuous with these growths, and over a period of time large accumulations of slimy brown deposits can occur. Iron bacteria may clog water lines, reduce heat transfer, and cause staining; objectionable odors may arise following death of the bacteria. The organic matter in the water is consequently increased, and this in turn favors the multiplication of other bacteria.
SCOPE
1.1 This practice covers the determination of filamentous iron bacteria (FIB) by microscopic examination. This practice provides for the identification of the following genera of bacteria found in water and water-formed deposits: Siderocapsa, Gallionella (Dioymohelix), Sphaerotilus, Crenothrix, Leptothrix, and Clonothrix.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 This test method is useful for rapid determination of viable active biomass concentrations in cooling tower waters. The efficiency of cooling towers is directly affected by the concentration of biomass in the cooling tower waters. As biomass concentrations increase, biofilm formation occurs resulting in a decrease in the efficiency of heat exchange in the tower. Current tests for monitoring the biomass concentration in cooling towers require at least 36 h for growth of the microorganisms on a solid agar surface for counting. Replication of microorganisms over the 36-h period before results are available creates an aqueous environment which is no longer represented by the data generated. Timely test results can assist in minimizing biocide addition to control biomass concentrations. Kool Kount provides data within hours to allow for more precise control of active biomass concentrations in the waters.
SCOPE
1.1 This test method covers the determination of viable active biomass in cooling tower water in the range from 102 to 108 cfu/mL. It is a semiquantitative test method.  
1.2 This test method was used successfully with reagent water, physiologic saline, and cooling tower waters. It is the user’s responsibility to ensure the validity of this test method for waters of untested matrices.  
1.3 The values stated in SI units are to be regarded as standard. The values given in parentheses after SI units are provided for information only and are not considered standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Microbiological water testing procedures using membrane filtration are based on the premise that all bacteria within a specific size range will be retained by the membrane filter used. If the membrane filter does not retain these bacteria, false negative results or lowered density estimates may occur that could have serious repercussions due to the presence of unrecognized potential health hazards in the water being tested, especially in drinking water.  
5.1.1 This procedure as devised will enable the user to test each membrane filter lot number for its ability to retain all bacterial equal to, or larger than, the stated membrane pore size.  
5.2 Since this membrane is often used to sterilize nonautoclavable liquids, it is essential that the retention characteristics of this membrane are stable.
SCOPE
1.1 This test method covers a procedure to test membrane filters for their ability to retain bacteria whose diameter is equal to or slightly larger than the 0.2-µm pore size of the membrane filter.  
1.2 The procedures described are for the use of user laboratories as differentiated from manufacturers’ laboratories.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
3.1 The advantages of using conical tow nets are as follows:  
3.1.1 They are relatively inexpensive and highly versatile in a variety of inland, estuarine, coastal, and marine waters.  
3.1.2 They can be used from a small or large powered boat with a minimum of auxiliary equipment.  
3.1.3 They can be used to collect qualitative samples and semiquantitative samples when fitted with a flowmeter and even better samples when fitted with a companion meter on the outside of the hoop to monitor filtering efficiency.  
3.2 The disadvantages of conical tow nets are as follows:  
3.2.1 When equipped with a flowmeter they require frequent maintenance including calibration and, in some types, lubrication.  
3.2.2 They are effective only where drawn through a stream of water having considerable thickness. They are not suitable for collecting samples from a small or restricted region.  
3.2.3 They are not suitable for collecting in very shallow water.  
3.2.4 They are clogged by grass beds, coelenterates, and filamentous algae.  
3.2.5 When used with a flowmeter, they collect only qualitative samples, or semiquantitative samples.  
3.2.6 When sampling discrete depths using a horizontal tow, the sample can be contaminated from other depths during the deployment and retrieval of the samples if opening and closing devices are not used.  
3.3 There are several special considerations that shall be observed when using conical tow nets. They are:  
3.3.1 Conical tow net samplers are designed to be towed at speeds less than three knots; however, greater speeds have been used for the larger nets with a concomitant increase in capture.2  
3.3.2 A conical tow net 0.5 m in diameter or larger shall be used to reduce avoidance by organisms.2  
3.3.3 The nets shall be washed frequently and inspected for pin-size holes, tears, net deterioration, and other anomalies.  
3.3.4 Nets should be allowed to dry while suspended full length in air and in subdued light prior to storage.  
...
SCOPE
1.1 This practice covers the procedure for obtaining qualitative samples of a zooplankton community by use of conical tow nets. Nets will collect most zooplankton, but some forms will avoid nets.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 Although many laboratories are presently isolating viruses from sludge, a valid comparison of data generated has not been possible because of the lack of a standard test method(s).
SCOPE
1.1 This practice is used for the recovery of viruses from wastewater sludges and favors the enteroviruses.  
1.2 Both procedures are applicable to raw, digested, and dewatered sludges.    
Sections  
Procedure A—Adsorption  
7 to 10  
Procedure B—Sonication  
11 to 15  
1.3 This practice was tested on standardized sludges as described in 10.1. It is the user's responsibility to ensure the validity of this practice for untested matrices.  
1.4 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.6 Only adequately trained personnel should be allowed to perform these procedures and should use safety precautions recommended by the U.S. Public Health Service, Center for Disease Control,2  for work with potentially hazardous biological organisms.  
1.7 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Sulfate reducing archaea and bacteria are known to contribute to microbiologically influenced corrosion.  
5.2 Sulfate-reducing bacteria are widely distributed in marine and fresh water muds which, in consequence, frequently are laden with the hydrogen sulfide produced by these organisms during dissimilatory sulfate reduction.  
5.3 Traditional, culture-dependent methods such as those described in Test Methods D4412, prescribe incubation periods of as long as 21 days before assigning a below detection limit (BDL) score to a specimen. Moreover, it is well known that not all SRP will proliferate in the nutrient media specified in Test Methods D4412.  
5.4 This test method uses ELISA technology to provide semi-quantitative, culture-independent, SRP bioburden test results in less than 30 min.  
5.4.1 Because all the reagents and supplies used are non-hazardous and prepackaged for single test use, this test method does not require any apparatus other than a laboratory timer. Consequently, it can be performed at or near the point of sample collection.  
5.4.2 The opportunity to minimize the delay between sample collection, testing, and results availability translates into timely use of the data to drive preventive and corrective SRB control measures.
SCOPE
1.1 This test method provides a protocol for using enzyme-linked immunosorbent assay (ELISA) technology to test water samples for the enzyme adenosine 5’-phosphosulfate reductase (APSr) concentration.  
1.1.1 APSr is present in all known sulfate reducing protists (SRP – sulfate reducing bacteria – SRB – and sulfate reducing archaea – SRA).  
1.1.2 As reported in U.S. Patent 4,999,286, APS reductase concentration can be used as a surrogate parameter for estimating SRA bioburdens (Appendix X1 compares results from Test Methods D8243, D4412, and quantitative polymerase chain reaction – qPCR – testing).  
1.2 This test method has been validated in tap water, oilfield produced water (salinities ranging from 100 g L-1 to 600 g L-1), and fuel-associated water (commonly referred to as bottoms-water).  
1.3 This test method detects APS reductase semi-quantitatively in the range of 0.001M to 0.1M – correlating to 102 SRP/mL to 106 SRP/mL.  
1.3.1 As described in Appendix X2 test method sensitivity can be increased 10-fold to 100-fold. However, the precision statistics provided in X apply only to 10-mL specimens.  
1.4 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Some specific hazards statements are given in Section 9 on Hazards.  
1.6 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 A rapid and routine procedure for determining biomass of the living microorganisms in cultures, waters, wastewaters, and in plankton and periphyton samples taken from surface waters is frequently of vital importance. However, classical techniques such as direct microscope counts, turbidity, organic chemical analyses, cell tagging, and plate counts are expensive, time-consuming, or tend to underestimate total numbers. In addition, some of these methods do not distinguish between living and nonliving cells.  
5.2 This test method measures the concentration of cellular-ATP present in the sample. ATP is a constituent of all living cells, including bacteria, algae, protozoa, and fungi. Consequently, the presence of cellular-ATP is an indicator of total metabolically active microbial contamination in water. ATP is not associated with matter of non-biological origin.  
5.3 The ATP (luciferin-luciferase) method is a rapid, sensitive determination of viable microbial biomass. ATP is the primary energy donor for life processes, does not exist in association with nonliving detrital material, and the amount of ATP per unit of biomass (expressed in weight) is relatively constant. (ATP per cell varies with species and physiological state of the organism.)  
5.4 This test method can be used to:  
5.4.1 Estimate viable microbial biomass in cultures and waters.  
5.4.2 Estimate the amount of total viable biomass in plankton and periphyton samples.  
5.4.3 Estimate the number of viable cells in a unispecies culture if the cATP content (or if the average amount of cATP) per cell is known.  
5.4.4 Estimate and differentiate between zooplanktonic, phytoplanktonic, bacterial, and fungal cATP through size fractionation of water samples.  
5.4.5 Measure the mortality rate of microorganisms in toxicity tests in entrainment studies, and in other situations where populations or assemblages of microorganisms are placed under stress.  
5.5 This test method is similar to Test Metho...
SCOPE
1.1 This test method covers a protocol for capturing, extracting and quantifying the cellular adenosine triphosphate (cATP) content associated with microorganisms normally found in laboratory cultures and waters in plankton and periphyton samples from waters.  
1.2 The ATP is measured using a bioluminescence enzyme assay, whereby light is generated in amounts proportional to the concentration of ATP in the samples. The light is produced and measured quantitatively as relative light units (RLU) which are converted by comparison with an ATP standard and computation to pg ATP/mL.  
1.3 This method does not remove all known chemical interferences, known to either luminesce in the 530 nm ± 20 nm range, or to quench light emitted in that range. It should not be used to determine ATP concentrations in samples with dissolved organic compounds, heavy metals or >10 000 ppm total dissolved solids. Alternative methods have been developed for determining ATP concentrations in fluids samples likely to contain such interferences (Test Methods D7687 and E2694).  
1.4 Knowledge of the concentration of ATP can be related to viable biomass or metabolic activity of microorganisms (Appendix X1).  
1.5 This test method offers a high degree of sensitivity, rapidity, accuracy, and reproducibility.  
1.6 The analyst should be aware that the precision statement pertains only to determinations in reagent water and not necessarily in the matrix being tested.  
1.7 This test method is equally suitable for use in the laboratory or field.  
1.8 The method normally detects cATP concentrations in the range of 0.1 pg cATP/mL (–1.0Log10 [pg cATP/mL]) to
4 000 000 pg cATP/mL (6.6 Log10 [pg cATP/mL]) in 50 mL water samples.  
1.9 Providing interferences can be overcome, bioluminescence is a reliable and proven method for qualifying and quantifying ATP, although the method does not differentiate between ATP from different sources, for example, from...

SIGNIFICANCE AND USE
5.1 P. aeruginosa is an opportunistic pathogen and has been linked as the causative agent of numerous infections that may be transmitted through a contaminated water supply to a susceptible host.
Note 1: Fecal waste is >95 % E. coli which is found in humans and warm bloodied animals.  
5.2 The membrane filtration procedure described is a rapid and reliable test method of detecting P. aeruginosa in water.
SCOPE
1.1 The test method covers the isolation and enumeration of Pseudomonas aeruginosa. Testing was performed on spiked samples using reagent grade water as the diluent from surface waters; recreational waters; ground water, water supplies; especially rural nonchlorinated sources; waste water; and saline waters. The detection limit of this test method is one microorganism per 100 mL.  
1.2 This test method was used successfully with reagent water. It is the user's responsibility to ensure the validity of this test method for surface waters, recreational waters, ground water, rural nonchlorinated sources; waste water; and saline waters.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 10.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 Sulfate-reducing bacteria are widely distributed in marine and fresh water muds which, in consequence, frequently are laden with the hydrogen sulfide produced by these organisms during dissimilatory sulfate reduction.  
5.2 It has been reported that Desulfovibrio spp. can form as much as 10 g of sulfide per litre during active multiplication. Sulfate-reducing bacteria can cause the external or internal corrosion of water or wastewater pipelines and pipelines for petroleum and natural gas. The formation of galvanic cells by massive growth of sulfate-reducing bacteria under suitable conditions makes the corrosion much worse than just the effect of the hydrogen sulfide on the metal or concrete.
SCOPE
1.1 These test methods cover the procedure for the detection and enumeration by the most probable number (MPN) technique of sulfate-reducing bacteria in water or water-formed deposits.  
1.2 Two media preparations are provided. Medium A which is prepared with reagent grade water, and Medium B which is prepared using the water to be sampled as the water source. Medium B is offered for those special conditions where sulfate-reducing bacterial strains have adapted to atypical non-fresh water environment.  
1.3 For the isolation and enumeration of thermophilic sulfate-reducing bacteria encountered in waters associated with oil and gas production, all broths, dilution blanks, and incubations must be maintained at temperatures of at least 45°C and preferably within 5°C at the sample temperature.  
1.4 The sensitivity of these test methods can be increased by purging the dilution blanks and tubes of media with nitrogen immediately prior to use.  
1.5 The analyst should be aware that adequate collaborative data for precision and bias statements as required by Practice D2777 are not provided. See Section 11 for details.  
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.7 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.8 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 The enterococci are indicators of the bacteriological quality for potable water, shellfish growing waters, ambient, and recreational waters. A direct relationship between swimming, associated gastroenteritis, and enterococci has been established through epidemiological studies and marine and fresh water bathing beaches. These studies have led to the development of criteria that can be used to establish bathing water standards based on established health-water quality relationships.  
5.2 Since small or large volumes of water or dilutions thereof, can be analyzed by the membrane filter technique, a wide range of levels of enterococci in water can be enumerated and detected.
SCOPE
1.1 This test method covers a membrane filter (MF) procedure for the detection and enumeration of the enterococci bacteria in water. The enterococci, which include Entero-coccus faecalis (E. faecalis), E. faecium, and their varieties are commonly found in the feces of humans and other warm-blooded animals. Although some strains are ubiquitous and not related to fecal pollution, enterococci in water are an indication of fecal pollution and the possible presence of enteric pathogens. These bacteria are found in water and wastewater in a wide range of densities. The detection limit is one colony forming unit (CFU)/volume filtered.  
1.2 This test method has been used successfully with temperate fresh and marine ambient waters, and wastewaters. It is the user’s responsibility to ensure the validity of this test method for waters of untested types.  
1.3 The values stated in SI units are to be regarded as standard. The values given in parentheses after SI units are included for information only and are not considered standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 This practice provides uniform guidance for cleaning the laboratory glassware, plasticware, and equipment used in routine microbiological analyses. However, tests that are extremely sensitive to toxic agents (such as virus assays) may require more stringent cleaning practices.2
SCOPE
1.1 In microbiology, clean glassware is crucial to ensure valid results. Previously used or new glassware must be thoroughly cleaned. Laboratory ware and equipment that are not chemically clean are responsible for considerable losses in personnel time and supplies in many laboratories. These losses may occur as down time when experiments clearly have been adversely affected and as invalid data that are often attributed to experimental error. Chemical contaminants that adversely affect experimental results are not always easily detected. This practice describes the procedures for producing chemically clean glassware.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific precautions, see Section 6, 5.7.3.1, and 8.3.1.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 The advantages of the Clarke-Bumpus plankton sampler are as follows:  
4.1.1 It will sample a discrete depth or multiple depths, depending upon the sampling design.  
4.1.2 It is a slow to medium speed sampler requiring a towing speed of three to five knots.  
4.1.3 The sample size can be easily controlled.  
4.1.4 The sampler is lightweight and can be used without auxiliary equipment.  
4.1.5 It has a relatively high filtration efficiency factor of 0.88.  
4.1.6 It is a versatile sampler and can be used in all but the shallowest waters.  
4.1.7 The flowmeter records the amount of water that passes into the net.  
4.1.8 Overspill of water at the mouth of the net due to excess speed of towing is of minimal consequence.  
4.2 The disadvantages of the Clarke-Bumpus plankton sampler are as follows:  
4.2.1 The flowmeter requires frequent maintenance including calibration and lubrication.  
4.2.2 It is not suitable for use in very small areas or shallow waters.  
4.3 There are several special considerations that shall be observed when using a Clarke-Bumpus plankton sampler. They are:  
4.3.1 The flowmeter should be calibrated and serviced frequently to ensure efficient and accurate operation.  
4.3.2 The sampler is relatively fragile, particularly the closing device and flowmeter. This necessitates careful deployment and recovery procedures.  
4.3.3 Following each collection, the net must be thoroughly washed.  
4.3.4 Special attention must be given to the strength of the cable and its attachment to avoid loss of the sampler.  
4.3.5 The sampler should not be used in beds of macrophytes, in waters containing submerged objects, or close to the bottom.  
4.3.6 The net should be inspected frequently for pin-size holes, tears, net deterioration, and other anomalies.  
4.3.7 Following use, the wet net should be suspended full length in the air in subdued light and allowed to dry.
SCOPE
1.1 This practice covers the procedures for obtaining quantitative samples of a zooplankton community by use of a Clarke-Bumpus plankton sampler.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 This test provides an easy and reliable method for the detection of enterococci in water within 24 h. For recreational water (fresh and marine) testing is performed to insure areas are safe for swimming. Enterolert also can be used for testing bottled water, wastewater, ground water, and drinking water.
SCOPE
1.1 This test method covers a simple procedure for the detection of enterococci in water and wastewater. It is based on IDEXX’s patented Defined Substrate Technology (DST).2 This product, Enterolert, utilizes a nutrient indicator that fluoresces when metabolized. It can detect these bacteria at one most probable number (MPN)/100 mL within 24 h. The presence of this microorganism in water is an indication of fecal contamination and the possible presence of enteric pathogens.  
1.2 This test method can be used successfully with drinking water, source water, recreational (fresh and marine) water, wastewater, and bottled water. It is the user’s responsibility to ensure the validity of this test method for waters of untested matrices.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 Calcium Carbonate (CaCO3) buffered formalin (3 to 5 %) can be used as a permanent preservative for zooplankton. Lugol’s iodine solution can be used to preserve zooplankton for up to one year. Thirty percent ethanol, 30 % glutaraldehyde, or 25 % vinegar (can use 3 % acetic acid solution) can be used for more temporary storage and preservation of zooplankton samples. A 25 % vinegar solution is preferred to preserve soft-bodied planktonic coelenterates.
SCOPE
1.1 This practice describes the proper procedures for preserving zooplankton samples with either formaldehyde, ethanol, glutaraldehyde, Lugol’s iodine solution, or vinegar (acetic acid).  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 The advantages of collecting zooplankton with pumps are as follows:  
4.1.1 Sample size is more accurately controlled than with nets.  
4.1.2 Discrete samples can be more easily obtained both vertically and horizontally.  
4.1.3 Multiple or replicate samples can be more easily obtained.  
4.1.4 The pumps are adaptable to a variety of ecosystems less than 30-m deep.  
4.1.5 Sampling efficiency does not decrease with sample size.  
4.2 The disadvantages of collecting zooplankton with pumps are as follows:  
4.2.1 Pumps are bulky and require an electrical source.  
4.2.2 Pumps are generally more costly than nets.  
4.2.3 Pumps generally discriminate against collecting macroplankton.  
4.2.4 Pump intake tubes may be avoided by the more motile zooplankton forms.  
4.2.5 Requires a long, bulky, intake tube for deep water sampling.  
4.3 There are several special considerations that should be observed when collecting zooplankton with a pump. They are:  
4.3.1 Some pumps can fragment zooplankton and induce mortality due to their design.  
4.3.2 The pump hose must be cleared before taking the next sample.
SCOPE
1.1 This practice covers the procedures for obtaining qualitative/quantitative samples of a zooplankton community by use of pumping systems.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.4 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
5.1 This test method is useful for measuring recreational water quality and chlorinated wastewaters, although it can be used for any water suspected of contamination by fecal wastes of warm-blooded animals. The significance of finding E. coli in recreational water samples, especially samples obtained from fresh recreational waters, is that there is a risk of gastrointestinal illness, directly related to the E. coli density, associated with swimming.5  
5.2 Since small or large volumes of water or dilutions thereof can be analyzed by the MF technique, a wider range of levels of E. coli in water can be detected and enumerated than with other methods.
SCOPE
1.1 This test method describes a membrane filter (MF) procedure for the detection and enumeration of Escherichia coli, a bacterium found exclusively in the feces of humans and other warm-blooded animals. The presence of these microorganisms in water is an indication of fecal pollution and the possible presence of enteric pathogens. These bacteria are found in water and wastewater in a wide range of densities. The detection limit of this procedure is one colony forming unit (CFU) per volume filtered.  
1.2 This test method has been used successfully with temperate fresh and marine ambient waters, and wastewaters. It is the user’s responsibility to ensure the validity of this test method for waters of other types.  
1.3 The values stated in SI units are to be regarded as standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

ABSTRACT
This practice covers determination of the quantitative and qualitative species composition of fish in a specified area. The successful use of this technique is dependent on: (1) preventing fish from escaping the sample area and (2) retrieving all affected fish, which may take up to three days. This practice is useful in both short- and long-term studies for management and impact assessment purposes. The sample area is blocked off with a small mesh net(s) and the volume of water to be treated is calculated. The required quantity of rotenone is diluted and distributed throughout the water column in the sample area. All fish should be affected and they should be collected for processing.
SCOPE
1.1 This practice covers determination of the quantitative and qualitative species composition of fish in a specified area. The successful use of this technique is dependent on: (1) preventing fish from escaping the sample area and (2) retrieving all affected fish, which may take up to three days.  
1.2 Advantages:  
1.2.1 Easily detoxified.  
1.2.2 All native freshwater fish are susceptible, but it has low toxicity to mammals and birds.  
1.2.3 At low concentrations fish toxicity depends on species, age, and size.  
1.2.4 The suffocating action is reversible.  
1.3 Limitations:  
1.3.1 It is less effective in cold (below 20°C) and highly alkaline water.  
1.3.2 Smaller fish and those without air bladders usually do not float.  
1.3.3 Completely random selection of sample areas is not possible.  
1.3.4 Overkill beyond sample area can sometimes occur.  
1.3.5 Food web organisms may be eliminated.  
1.4 Applications—This practice is useful in both short- and long-term studies for management and impact assessment purposes. It is adaptable to both lotic and lentic situations in littoral and limnetic areas.  
1.5 The values stated in inch-pound units are to be regarded as standard. The values given in parentheses are mathematical conversions to SI units that are provided for information only and are not considered standard.  
1.6 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. For specific hazards, see Section 7.  
1.7 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

SIGNIFICANCE AND USE
4.1 Numerous ASTM test methods and practices (for example: Test Methods D5259 and D5392, and Practices D6974 and E2563) report colony counts as their measured parameter.  
4.2 These practices provide a uniform set of counting, calculating, and reporting procedures for ASTM test methods in microbiology.    
Section  
A—Counting Colonies on Membrane Filters  
6  
B—Counting Colonies on Pour Plates  
7  
C—Counting Colonies on Spread Plates  
8  
4.3 The counting rules provide a best attainable estimate of microorganisms in the sample, since the samples cannot be held and reanalyzed at a later date.
SCOPE
1.1 These practices cover recommended procedures for counting colonies and reporting colony-forming units (CFU) on membrane filters (MF) and standard pour and spread plates.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

SIGNIFICANCE AND USE
5.1 Sulfate-reducing bacteria are widely distributed in marine and fresh water muds which, in consequence, frequently are laden with the hydrogen sulfide produced by these organisms during dissimilatory sulfate reduction.  
5.2 It has been reported that Desulfovibrio spp. can form as much as 10 g of sulfide per litre during active multiplication. Sulfate-reducing bacteria can cause the external or internal corrosion of water or wastewater pipelines and pipelines for petroleum and natural gas. The formation of galvanic cells by massive growth of sulfate-reducing bacteria under suitable conditions makes the corrosion much worse than just the effect of the hydrogen sulfide on the metal or concrete.
SCOPE
1.1 These test methods cover the procedure for the detection and enumeration by the most probable number (MPN) technique of sulfate-reducing bacteria in water or water-formed deposits.  
1.2 Two media preparations are provided. Medium A which is prepared with reagent grade water, and Medium B which is prepared using the water to be sampled as the water source. Medium B is offered for those special conditions where sulfate-reducing bacterial strains have adapted to atypical non-fresh water environment.  
1.3 For the isolation and enumeration of thermophilic sulfate-reducing bacteria encountered in waters associated with oil and gas production, all broths, dilution blanks, and incubations must be maintained at temperatures of at least 45°C and preferably within 5°C at the sample temperature.  
1.4 The sensitivity of these test methods can be increased by purging the dilution blanks and tubes of media with nitrogen immediately prior to use.  
1.5 The analyst should be aware that adequate collaborative data for precision and bias statements as required by Practice D2777 are not provided. See Section 11 for details.  
1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.7 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

SIGNIFICANCE AND USE
5.1 A rapid and routine procedure for determining biomass of the living microorganisms in cultures, waters, wastewaters, and in plankton and periphyton samples taken from surface waters is frequently of vital importance. However, classical techniques such as direct microscope counts, turbidity, organic chemical analyses, cell tagging, and plate counts are expensive, time-consuming, or tend to underestimate total numbers. In addition, some of these methods do not distinguish between living and nonliving cells.  
5.2 This test method measures the concentration of cellular-ATP present in the sample. ATP is a constituent of all living cells, including bacteria, algae, protozoa, and fungi. Consequently, the presence of cellular-ATP is an indicator of total metabolically active microbial contamination in water. ATP is not associated with matter of non-biological origin.  
5.3 The ATP (luciferin-luciferase) method is a rapid, sensitive determination of viable microbial biomass. ATP is the primary energy donor for life processes, does not exist in association with nonliving detrital material, and the amount of ATP per unit of biomass (expressed in weight) is relatively constant. (ATP per cell varies with species and physiological state of the organism.)  
5.4 This test method can be used to:  
5.4.1 Estimate viable microbial biomass in cultures, waters, and wastewaters.  
5.4.2 Estimate the amount of total viable biomass in plankton and periphyton samples.  
5.4.3 Estimate the number of viable cells in a unispecies culture if the cATP content (or if the average amount of cATP) per cell is known.  
5.4.4 Estimate and differentiate between zooplanktonic, phytoplanktonic, bacterial, and fungal cATP through size fractionation of water, and wastewater samples.  
5.4.5 Measure the mortality rate of microorganisms in toxicity tests in entrainment studies, and in other situations where populations or assemblages of microorganisms are placed under stress.  
5.5 This test m...
SCOPE
1.1 This test method covers a protocol for capturing, extracting and quantifying the cellular adenosine triphosphate (cATP) content associated with microorganisms normally found in laboratory cultures, waters, wastewaters, and in plankton and periphyton samples from waters.  
1.2 The ATP is measured using a bioluminescence enzyme assay, whereby light is generated in amounts proportional to the concentration of ATP in the samples. The light is produced and measured quantitatively as relative light units (RLU) which are converted by comparison with an ATP standard and computation to pg ATP/mL.  
1.3 This method does not remove all known chemical interferences, known to either luminesce in the 530 ± 20 nm range, or to quench light emitted in that range. It should not be used to determine ATP concentrations in samples with dissolved organic compounds, heavy metals or >10 000 ppm total dissolved solids. Alternative methods have been developed for determining ATP concentrations in fluids samples likely to contain such interferences (Test Methods D7687 and E2694).  
1.4 Knowledge of the concentration of ATP can be related to viable biomass or metabolic activity of microorganisms (Appendix X1).  
1.5 This test method offers a high degree of sensitivity, rapidity, accuracy, and reproducibility.  
1.6 The analyst should be aware that the precision statement pertains only to determinations in reagent water and not necessarily in the matrix being tested.  
1.7 This test method is equally suitable for use in the laboratory or field.  
1.8 The method normally detects cATP concentrations in the range of 0.1 pg cATP/mL (–1.0Log10 [pg cATP/mL]) to 4 000 000 pg cATP/mL (6.6 Log10 [pg cATP/mL]) in 50 mL water samples.  
1.9 Providing interferences can be overcome, bioluminescence is a reliable and proven method for qualifying and quantifying ATP, although the method does not differentiate between ATP from different sources, for exa...

SIGNIFICANCE AND USE
5.1 Pseudomonas aeruginosa is an opportunistic pathogen, and has been linked as the causative agent of numerous infections that may be transmitted through a contaminated water supply to a susceptible host.
Note 1: Fecal waste is >95 % E. coli which is found in humans and warm bloodied animals.  
5.2 The membrane filtration procedure described is a rapid and reliable test method of detecting P. aeruginosa in water.
SCOPE
1.1 The test method covers the isolation and enumeration of Pseudomonas aeruginosa. Testing was performed on spiked samples using reagent grade water as the diluent from surface waters; recreational waters; ground water, water supplies; especially rural nonchlorinated sources; waste water; and saline waters. The detection limit of this test method is one microorganism per 100 mL.  
1.2 This test method was used successfully with reagent water. It is the user's responsibility to ensure the validity of this test method for surface waters, recreational waters, ground water, rural nonchlorinated sources; waste water; and saline waters.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 10.

SIGNIFICANCE AND USE
5.1 Filamentous iron bacteria is a general classification for microorganisms that utilize ferrous iron as a source of energy and are characterized by the deposition of ferric hydroxide in their mucilaginous sheaths. The process is continuous with these growths, and over a period of time large accumulations of slimy brown deposits can occur. Iron bacteria may clog water lines, reduce heat transfer, and cause staining; objectionable odors may arise following death of the bacteria. The organic matter in the water is consequently increased, and this in turn favors the multiplication of other bacteria.
SCOPE
1.1 This practice covers the determination of filamentous iron bacteria (FIB) by examination under the microscope. The practice provides for the identification of the following genera of bacteria found in water and water-formed deposits: Siderocapsa, Gallionella (Dioymohelix), Sphaerotilus, Crenothrix,  Leptothrix, and Clonothrix.  
1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.3 This standard does not purport to address the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

SIGNIFICANCE AND USE
5.1 This test method is useful for measuring recreational water quality and chlorinated wastewaters, although it can be used for any water suspected of contamination by fecal wastes of warm-blooded animals. The significance of finding E. coli  in recreational water samples, especially samples obtained from fresh recreational waters, is that there is a risk of gastrointestinal illness, directly related to the E. coli  density, associated with swimming.5  
5.2 Since small or large volumes of water or dilutions thereof can be analyzed by the MF technique, a wider range of levels of E. coli in water can be detected and enumerated than with other methods.
SCOPE
1.1 This test method describes a membrane filter (MF) procedure for the detection and enumeration of Escherichia coli, a bacterium found exclusively in the feces of humans and other warm-blooded animals. The presence of these microorganisms in water is an indication of fecal pollution and the possible presence of enteric pathogens. These bacteria are found in water and wastewater in a wide range of densities. The detection limit of this procedure is one colony forming unit (CFU) per volume filtered.  
1.2 This test method has been used successfully with temperate fresh and marine ambient waters, and wastewaters. It is the user's responsibility to ensure the validity of this test method for waters of other types.  
1.3 The values stated in SI units are to be regarded as standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.

SIGNIFICANCE AND USE
5.1 This test provides an easy and reliable method for the detection of enterococci in water within 24 h. For recreational water (fresh and marine) testing is performed to insure areas are safe for swimming. Enterolert also can be used for testing bottled water, wastewater, and drinking water.
SCOPE
1.1 This test method covers a simple procedure for the detection of enterococci in water and wastewater. It is based on IDEXX’s patented Defined Substrate Technology (DST).2 This product, Enterolert, utilizes a nutrient indicator that fluoresces when metabolized. It can detect these bacteria at one colony forming unit (CFU)/100 mL within 24 h. The presence of this microorganism in water is an indication of fecal contamination and the possible presence of enteric pathogens.  
1.2 This test method can be used successfully with drinking water, source water, recreational (fresh and marine) water, wastewater, and bottled water. It is the user’s responsibility to ensure the validity of this test method for waters of untested matrices.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

SIGNIFICANCE AND USE
5.1 The enterococci are indicators of the bacteriological quality for potable water, shellfish growing waters, ambient, and recreational waters. A direct relationship between swimming, associated gastroenteritis, and enterococci has been established through epidemiological studies and marine and fresh water bathing beaches. These studies have led to the development of criteria that can be used to establish bathing water standards based on established health-water quality relationships.  
5.2 Since small or large volumes of water or dilutions thereof, can be analyzed by the membrane filter technique, a wide range of levels of enterococci in water can be enumerated and detected.
SCOPE
1.1 This test method covers a membrane filter (MF) procedure for the detection and enumeration of the enterococci bacteria in water. The enterococci, which include Entero-coccus faecalis (E. faecalis), E. faecium, and their varieties are commonly found in the feces of humans and other warm-blooded animals. Although some strains are ubiquitous and not related to fecal pollution, enterococci in water are an indication of fecal pollution and the possible presence of enteric pathogens. These bacteria are found in water and wastewater in a wide range of densities. The detection limit is one colony forming unit (CFU)/volume filtered.  
1.2 This test method has been used successfully with temperate fresh and marine ambient waters, and wastewaters. It is the user's responsibility to ensure the validity of this test method for waters of untested types.  
1.3 The values stated in SI units are to be regarded as the standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.

SIGNIFICANCE AND USE
4.1 Although many laboratories are presently isolating viruses from sludge, a valid comparison of data generated has not been possible because of the lack of a standard test method(s).
SCOPE
1.1 This practice is used for the recovery of viruses from wastewater sludges and favors the enteroviruses.  
1.2 Both procedures are applicable to raw, digested, and dewatered sludges.    
Sections  
Procedure A—Adsorption  
6 to 10  
Procedure B—Sonication  
11 to 15
1.3 This practice was tested on standardized sludges as described in 10.1. It is the user's responsibility to ensure the validity of this practice for untested matrices.  
1.4 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.5 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.  
1.6 Only adequately trained personnel should be allowed to perform these procedures and should use safety precautions recommended by the U.S. Public Health Service, Center for Disease Control,2 for work with potentially hazardous biological organisms.

ABSTRACT
This practice covers determination of the quantitative and qualitative species composition of fish in a specified area. The successful use of this technique is dependent on: (1) preventing fish from escaping the sample area and (2) retrieving all affected fish, which may take up to three days. This practice is useful in both short- and long-term studies for management and impact assessment purposes. The sample area is blocked off with a small mesh net(s) and the volume of water to be treated is calculated. The required quantity of rotenone is diluted and distributed throughout the water column in the sample area. All fish should be affected and they should be collected for processing.
SCOPE
1.1 This practice covers determination of the quantitative and qualitative species composition of fish in a specified area. The successful use of this technique is dependent on: (1) preventing fish from escaping the sample area and (2) retrieving all affected fish, which may take up to three days.  
1.2 Advantages:  
1.2.1 Easily detoxified.  
1.2.2 All native freshwater fish are susceptible, but it has low toxicity to mammals and birds.  
1.2.3 At low concentrations fish toxicity depends on species, age, and size.  
1.2.4 The suffocating action is reversible.  
1.3 Limitations:  
1.3.1 It is less effective in cold (below 20°C) and highly alkaline water.  
1.3.2 Smaller fish and those without air bladders usually do not float.  
1.3.3 Completely random selection of sample areas is not possible.  
1.3.4 Overkill beyond sample area can sometimes occur.  
1.3.5 Food web organisms may be eliminated.  
1.4 Applications—this practice is useful in both short- and long-term studies for management and impact assessment purposes. It is adaptable to both lotic and lentic situations in littoral and limnetic areas.  
1.5 The values stated in inch-pound units are to be regarded as the standard. The values given in parentheses are for information only.  
1.6 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. For specific hazards, see Section 6.

SIGNIFICANCE AND USE
5.1 This test method is useful for rapid determination of viable active biomass concentrations in cooling tower waters. The efficiency of cooling towers is directly affected by the concentration of biomass in the cooling tower waters. As biomass concentrations increase, biofilm formation occurs resulting in a decrease in the efficiency of heat exchange in the tower. Current tests for monitoring the biomass concentration in cooling towers require at least 36 h for growth of the microorganisms on a solid agar surface for counting. Replication of microorganisms over the 36-h period before results are available creates an aqueous environment which is no longer represented by the data generated. Timely test results can assist in minimizing biocide addition to control biomass concentrations. Kool Kount provides data within hours to allow for more precise control of active biomass concentrations in the waters.
SCOPE
1.1 This test method covers the determination of viable active biomass in cooling tower water in the range from 102 to 108 cfu/mL (1). It is a semiquantitative test method.  
1.2 This test method was used successfully with reagent water, physiologic saline, and cooling tower waters. It is the user's responsibility to ensure the validity of this test method for waters of untested matrices.  
1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. For specific hazard statements, see Section 9.

SIGNIFICANCE AND USE
5.1 Pseudomonas aeruginosa is an opportunistic pathogen, and has been linked as the causative agent of numerous infections that may be transmitted through a contaminated water supply to a susceptible host.Note 1—Fecal waste is >95 % E. coli which is found in humans and warm bloodied animals.  
5.2 The membrane filtration procedure described is a rapid and reliable test method of detecting P. aeruginosa in water.
SCOPE
1.1 The test method covers the isolation and enumeration of Pseudomonas aeruginosa. Testing was performed on spiked reagent grade water samples.  
1.2 It is the user’s responsibility to ensure the validity of this method for surface waters, ground waters, recreational waters fresh and marine), wastewaters.  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 10.

SIGNIFICANCE AND USE
3.1 The  advantages  of using conical tow nets are as follows:  
3.1.1 They are relatively inexpensive and highly versatile in a variety of inland, estuarine, coastal, and marine waters.  
3.1.2 They can be used from a small or large powered boat with a minimum of auxiliary equipment.  
3.1.3 They can be used to collect qualitative samples and semiquantitative samples when fitted with a flowmeter and even better samples when fitted with a companion meter on the outside of the hoop to monitor filtering efficiency.  
3.2 The disadvantages  of conical tow nets are as follows:  
3.2.1 When equipped with a flowmeter they require frequent maintenance including calibration and, in some types, lubrication.  
3.2.2 They are effective only where drawn through a stream of water having considerable thickness. They are not suitable for collecting samples from a small or restricted region.  
3.2.3 They are not suitable for collecting in very shallow water.  
3.2.4 They are clogged by grass beds, coelenterates, and filamentous algae.  
3.2.5 When used with a flowmeter, they collect only qualitative samples, or semiquantitative samples.  
3.2.6 When sampling discrete depths using a horizontal tow, the sample can be contaminated from other depths during the deployment and retrieval of the samples if opening and closing devices are not used.  
3.3 There are several  special considerations  that shall be observed when using conical tow nets. They are:  
3.3.1 Conical tow net samplers are designed to be towed at speeds less than three knots; however, greater speeds have been used for the larger nets with a concomitant increase in capture.2  
3.3.2 A conical tow net 0.5 m in diameter or larger shall be used to reduce avoidance by organisms.2  
3.3.3 The nets shall be washed frequently and inspected for pin-size holes, tears, net deterioration, and other anomalies.  
3.3.4 Nets should be allowed to dry while suspended full length in air and in subdued light prior to stor...
SCOPE
1.1 This practice covers the procedure for obtaining qualitative samples of a zooplankton community by use of conical tow nets. Nets will collect most zooplankton, but some forms will avoid nets.  
1.2 This standard does not purport to address all of the safety problems, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

SIGNIFICANCE AND USE
4.1 The advantages of collecting zooplankton with pumps are as follows:  
4.1.1 Sample size is more accurately controlled than with nets.  
4.1.2 Discrete samples can be more easily obtained both vertically and horizontally.  
4.1.3 Multiple or replicate samples can be more easily obtained.  
4.1.4 The pumps are adaptable to a variety of ecosystems less than 30-m deep.  
4.1.5 Sampling efficiency does not decrease with sample size.  
4.2 The disadvantages of collecting zooplankton with pumps are as follows:  
4.2.1 Pumps are bulky and require an electrical source.  
4.2.2 Pumps are generally more costly than nets.  
4.2.3 Pumps generally discriminate against collecting macroplankton.  
4.2.4 Pump intake tubes may be avoided by the more motile zooplankton forms.  
4.2.5 Requires a long, bulky, intake tube for deep water sampling.  
4.3 There are several special considerations that should be observed when collecting zooplankton with a pump. They are:  
4.3.1 Some pumps can fragment zooplankton and induce mortality due to their design.  
4.3.2 The pump hose must be cleared before taking the next sample.
SCOPE
1.1 This practice covers the procedures for obtaining qualitative/quantitative samples of a zooplankton community by use of pumping systems.  
1.2 This standard does not purport to address all of the safety problems, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

SIGNIFICANCE AND USE
3.1 The significance of using chemical fish toxicants is that more complete population analyses or total eradication, or both, can be accomplished. Target species can be selectively eradicated by varying concentrations. This provides a very effective tool in fisheries investigations and management programs. Water conditions (that is, pH, temperature, alkalinity, and so forth) and morphology can be limiting factors.  
3.2 Rotenone—Rotenone used as a fish toxicant is highly versatile and can be used effectively to collect fish samples; to eradicate fish; and to selectively remove certain fish species.  
3.2.1 Its effectiveness is reduced in cold  
3.3 Antimycin—Antimycin is versatile in the selective removal of scalefish or even more selectively against certain centrarchids (sunfish) and minnows.  
3.3.1 Its effectiveness is reduced in water with pH above 8.5.
SCOPE
1.1 This classification covers rotenone and antimycin which are used to collect or eradicate fish; numerous chemicals have been used but presently only rotenone and antimycin are EPA approved for this use.

SIGNIFICANCE AND USE
4.1 Calcium Carbonate (CaCO3) buffered formalin (3 to 5 %) can be used as a permanent preservative for zooplankton. Lugol's iodine solution can be used to preserve zooplankton for up to one year. Thirty percent ethanol, 30 % glutaraldehyde, or 25 % vinegar (can use 3 % acidic acid solution) can be used for more temporary storage and preservation of zooplankton samples. A 25 % vinegar solution is preferred to preserve soft-bodied planktonic coelenterates.
SCOPE
1.1 This practice describes the proper procedures for preserving zooplankton samples with either formaldehyde, ethanol, glutaraldehyde, Lugol's iodine solution, or vinegar (acetic acid).  
1.2 This standard does not purport to address all of the safety problems, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.