ASTM D5244-92(1998)
(Practice)Standard Practice for Recovery of Enteroviruses from Waters
Standard Practice for Recovery of Enteroviruses from Waters
SCOPE
1.1 This practice covers a uniform procedure for the concentration of viruses from collected samples.
1.2 This practice describes a virus adsorption-elution cartridge filter procedure for recovering viruses from drinking water. Volumes of 400 L or more are processed for samples of drinking water quality.
1.3 The principles of this practice are also applicable to sewages, effluents, and surface waters without technical modifications.
1.4 Although specifically designed for recovery of human enteroviruses, this practice also may be applied to some other human enteric viruses, that have to be determined by specific testing.
1.5 The consistency of this practice was determined from method evaluation studies with poliovirus-seeded drinking water samples.
1.6 The values stated in SI units are to be regarded as the standard.
1.7 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. 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 for work with potentially hazardous biological organisms.
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An American National Standard
Designation: D 5244 – 92 (Reapproved 1998)
Standard Practice for
Recovery of Enteroviruses from Waters
This standard is issued under the fixed designation D 5244; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (e) indicates an editorial change since the last revision or reapproval.
1. Scope 3. Terminology
1.1 This practice covers a uniform procedure for the con- 3.1 Definitions:
centration of viruses from collected samples. 3.1.1 For definitions of terms used in this practice, refer to
1.2 This practice describes a virus adsorption-elution car- Terminology D 1129.
tridge filter procedure for recovering viruses from drinking 3.2 Definitions of Terms Specific to This Standard:
water. Volumes of 400 L or more are processed for samples of 3.2.1 cell monolayer—a single layer of cells grown on a
drinking water quality. glass or plastic surface to which they are securely attached.
1.3 The principles of this practice are also applicable to 3.2.2 enterovirus—a genus of the family Picornaviridae.
sewages, effluents, and surface waters without technical modi- Members of this genus are 22 to 30 nm in diameter, contain a
fications. positive single-stranded RNA, are stable under acid conditions
1.4 Although specifically designed for recovery of human and are resistant to ether. Included in this genus are poliovirus,
enteroviruses, this practice also may be applied to some other coxsackievirus, and echovirus.
human enteric viruses, that have to be determined by specific 3.2.3 enteric virus—a general term denoting a virus that
testing. normally enters by the oral route, is capable of multiplying in
1.5 The consistency of this practice was determined from cells of the alimentary canal and is found in stool specimens. In
method evaluation studies with poliovirus-seeded drinking addition to the enterovirus, included under this general term are
water samples. such agents as adenovirus, rotavirus, Norwalk virus, astrovirus,
1.6 The values stated in SI units are to be regarded as the and calicivirus.
standard. 3.2.4 plaque—an area of clearing caused by the cytopathic
1.7 This standard does not purport to address all of the effects of virus on a susceptible cell monolayer.
safety problems, if any, associated with its use. It is the
4. Summary of Practice
responsibility of the user of this standard to establish appro-
priate safety and health practices and determine the applica- 4.1 A commercially available negatively charged cartridge-
type filter is used to recover low levels of virus from water. The
bility of regulatory limitations prior to use. Only adequately
trained personnel should be allowed to perform these proce- viruses adsorbed to this filter matrix are released by passage of
beef extract-glycine reagent (pH 9.0) through the filter. The
dures and should use safety precautions recommended by the
U.S. Public Health Service Center for Disease Control for work eluted viruses are further concentrated by organic flocculation.
This consists of lowering the pH of the beef extract to 3.5,
with potentially hazardous biological organisms.
separating the resulting floc, and solubilizing the floc in a
2. Referenced Documents
relatively small volume of phosphate solution to release the
2.1 ASTM Standards: bound viruses.
D 1129 Terminology Relating to Water
3 5. Significance and Use
D 1193 Specification for Reagent Water
5.1 Enteric viruses of public health significance are present
in the aquatic environment.
5.2 Enteric viruses have been detected in treated water
This practice is under the jurisdiction of ASTM Committee D-19 on Water and
supplies.
is the direct responsibility of Subcommittee D19.24 on Water Microbiology.
5.3 Enteric viruses are responsible for a wide range of
Current edition approved May 15, 1992. Published September 1992.
Biological Safety in Microbiological and Biomedical Laboratories, Richard-
illnesses, ranging from hepatitis to gastroenteritis.
son, J. H., and Barkley, W. E., Eds., U.S. Dept. of Health and Human Services,
5.4 This practice is applicable to the recovery of many
Public Health Service, Centers for Disease Control and National Institutes of Health,
plaque-forming enteric viruses from waters when used in
HHS Publication No. (NIH) 88-8395, 2nd Ed, May 1988.
Annual Book of ASTM Standards, Vol 11.01. conjunction with cell culture assay systems.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.
D 5244 – 92 (1998)
5.5 The principles of this practice are applicable without 8. Procedure
technical modifications for monitoring for viruses based on the
8.1 Conditioning of Sample:
use of gene probe technology.
8.1.1 Dechlorinate water, if necessary, with 0.8 mL of
sodium thiosulfate solution for each litre of test sample to be
6. Apparatus
collected.
6.1 Holder, for 10-in. (25.4 cm) cartridge filter. 8.1.2 Acidify test sample to pH 3.5 with HCl (1 + 9).
Rapidly mix acid into sample to prevent pH levels from
6.2 Filterite Cartridge Filter, negatively charged pleated
fiberglass, 10-in. (25.4 cm), 0.45-μm pore size. becoming sufficiently low in parts to inactivate viruses.
8.1.3 Condition each litre of acidified test sample with 1.2
6.3 pH Meter, measuring to an accuracy of at least 0.1 pH
unit, equipped with a combination-type electrode. mL of aluminum chloride solution. Final concentration of
AlCl is 0.0005 M.
6.4 Magnetic Stirrer, with stir bars.
6.5 Positive Pressure Source, equipped with a pressure 8.2 Adsorption of Viruses:
8.2.1 Force the pH adjusted test sample from 8.1.3 through
gage. Deliver to filte
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