Environmental Letters
ISSN: 0013-9300 (Print) (Online) Journal homepage: http://www.tandfonline.com/loi/lesa17
Virus Association with Wastewater Solids D. O. Cliver To cite this article: D. O. Cliver (1975) Virus Association with Wastewater Solids, Environmental Letters, 10:3, 215-223, DOI: 10.1080/00139307509435823 To link to this article: http://dx.doi.org/10.1080/00139307509435823
Published online: 02 Sep 2009.
Submit your article to this journal
Article views: 1
View related articles
Citing articles: 8 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=2esa20 Download by: [University of Florida]
Date: 06 November 2015, At: 04:51
ENVIRONMENTAL LETTERS, 10(3),
215-223 (1975)
VIRUS ASSOCIATION WITH WASTEWATER SOLIDS
-KEY WORDS:
enteroviruses, reoviruses, sewage, sludge
Downloaded by [University of Florida] at 04:51 06 November 2015
D. 0: Cliver
Food Research Institute and Department of Bacteriology University of Wisconsin - Madison Madison, Wisconsin 53706
ABSTRACT The solids produced at an urban wastewater treatment plant, tested semiquantitatively, almost all contained human intestinal viruses.
Reoviruses and five or more types of enteroviruses were
present.
Sludge, digested anaerobically at 30'
grit contained measurable levels of viruses.
-
32' C, and
Until reliable means
of inactivating the viruses have been developed and implemented, great care should be taken in disposing of these solids. INTRODUCTION Effluent from an urban sewage treatment plant ought not contain viruses. The human intestinal viruses which occur in community wastewater should be removed during treatment.
'Removal' might
mean either inactivation (loss of the virus' ability to cause
215 Copyright 0 1976 by h!arcel Dekker, Inc. All Rights Reserved. Neither this work nor any part may be reproduced or transmitted in any form o r by any means, electronic or mechanical, including photocopying, microfilming, and recording, or by any information storage and retrieval system, without permission in writing from the publisher.
216
CLIVER
infection) or physical separation of the virus from the wastewater before the effluent is discharged. Sewage contains solids of biological and mineral origin, and other solids may be added or produced during treatment. Virus particles are too small to sediment by themselves from wastewater, but they often associate with solids which do sediment during
Downloaded by [University of Florida] at 04:51 06 November 2015
various stages of treatment. "
The factors governing association
of virus with these solids are being investigated.3 , 4 The products of sedimentation steps in sewage treatment are grit and various kinds of sludge. Each contains a good deal of water.
They must be disposed in some fashion, and they are poorly
suited to chemical disinfection.
If viruses are present, human
illnesses might result unless great care is taken in disposal of these solids. The present study included one metropolitan treatment plant and two groups of viruses. The plant serves a relatively nonindustrial community and receives 120,obO to 140,000 m3 of influent per day, depending upon the season. It produces approximately
0.8 m 3 of grit and 480 m 3 of final, anaerobically digested sludge per day.
Treatment includes screening and grit separation, primary
sedimentation, secondary treatment by the activated sludge (principally) or the trickling filter process, and terminal chlorination of the clarified effluent.
The blend of thickened primary and
secondary (principally waste activated) sludges is digested anaerobically for 12 days at 30'
-
32OC and held for eight more days at
a temperature which, depending on the season, ranges upward from 7"
- 10°C.
217
VIRUSES IN WASTEWATER SOLIDS
The viruses sought were members of the enterovirus and reovirus groups. The enteroviruses are approximately 28 nm in diameter, contain single-stranded ribonucleic acid (RNA) , and may cause a variety of human illnesses. The group includes the polioviruses, coxsackieviruses A and B, and echoviruses. All emanate from the human intestines; the polioviruses most frequently occur in sewage
Downloaded by [University of Florida] at 04:51 06 November 2015
with the feces of those who have received the oral polio vaccine. The reoviruses are 70 to 85 nm in diameter and contain doublestranded RNA.
Their role in causing human disease is uncertain.
MATERIALS AND METHODS Samples were collected in four sets over a 12-month period. Given the known seasonal and diurnal variations in the quantity and composition of sewage arriving at the plant, there is no claim that these few 'grab' samples are statistically representative of all that passes through it. Where virus has associated with solids homogeneity of virus distribution is likely to be poor. Studies of the best methods for dissociating virus from sewage solids have produced differing results.2 y 3 y 5
Optimum conditions
seem generally to include mild alkalinity, some added protein, and physical force imposed by violent mixing or by freezing and thawing. Grab samples were he3d in the laboratory to see if sedimentation occurred.
If not, a small portion of the sample was processed for
testing in tissue culture. Where sediment did collect at the bottom of a vessel, its volume was recorded, and it was resuspended in a small volume-of supernatant fluid for further processing. Phosphate-buffered saline (pH 7.4) plus 10% fetal calf serum was
218
CLIVER
added at a rate which depended on the turbidity of the sample; the volume of diluent did not exceed that of the sample fraction being processed. The suspension was homogenized for 30 sec in a Servall OmniMixer (80~). After three or more freeze-thaw cycles, the sediment was discarded, and the supernatant fluid was treated with 5% chloro-
Downloaded by [University of Florida] at 04:51 06 November 2015
form or passed through a glass fiber prefilter and 0.2 pm porosity membrane filter (Gelman GA-8) to remove bacteria.
The resulting
extract was tested in primary rhesus (Placaca mulatta) monkey kidney cells produced by methods described elsewhere.6
Effects
seen within 24 hr were considered toxic, rather than viral. Virus was detected by microscopically visible cytopathic effects (CPE) in cultures maintained with liquid medium or by areas of cell degeneration (plaques) produced in cultures maintained with either of two semisolid media.
Enteroviruses and reoviruses
could be differentiated presumptively on the basis of differences i n their patterns of CPE.
Further, reoviruses were essentially
unable to produce plaques unless the culture medium contained pancreatin,' and they were retained absolutely by a 50 nm porosity Nuclepore filter. Serologic identification was attempted only for enteroviruses. Antisera to the three types of polioviruses and the s i x types of coxsackieviruses B were used in an agar-diffusion neutralization test adapted from that of Kalter.8
Virus
concentrations are expressed in plaque-forming units (PFU) per milliliter of the original grab sample.
219
VIRUSES I N WASTEWATER SOLIDS RESULTS p r e l i m i n a r y tests showed t h a t v i r u s w a s r e a d i l y d e t e c t a b l e i n t h e i n f l u e n t r a w sewage and w a s p r e f e r e n t i a l l y a s s o c i a t e d with t h e solids..
The v i r u s r e c o v e r e d from t h e s o l i d s may n o t
have b e e n a l l t h a t was p r e s e n t , s o any c o n c e n t r a t i o n r e p o r t e d should b e r e g a r d e d as a minimum.
Added l a b o r a t o r y p o l i o v i r u s
Downloaded by [University of Florida] at 04:51 06 November 2015
d i d n o t seem t o a s s o c i a t e s i m i l a r l y w i t h t h e s o l i d s , s o a l l o b s e r v a t i o n s had t o b e based upon n a t u r a l l y - o c c u r r i n g v i r u s . T a b l e 1 summarizes t h e r e s u l t s o b t a i n e d w i t h raw sewage sampled on f o u r o c c a s i o n s .
The numbers p r e s e n t e d are n o t s o
p r e c i s e as t o show s e a s o n a l t r e n d s , though t h e i n c i d e n c e of e n t e r o v i r u s e s o t h e r t h a n t h e v a c c i n e p o l i o v i r u s e s would be e x p e c t e d t o peak i n t h e donor p o p u l a t i o n d u r i n g l a t e summer.
Any of
t h e s e l e v e l s of v i r u s could have o c c u r r e d w i t h l e s s t h a n 1%of t h e donor p o p u l a t i o n i n f e c t e d .
P o l i o v i r u s e s 1 and 3 from t h e 8/26/74
sample were t h e o n l y v i r u s e s s e r o t y p e d s u c c e s s f u l l y from t h e r a w sewage samples. TABLE I V i r u s e s Found i n Raw Sewage Samples Sampling Date
Enteroviruses (PFU/ml)
Reoviruses (PFU/ml>
12/21/73
0.013
n o t determined
61121 74
0.17
0.17
8/26/74
0.06
0.06
11/1/74
0.09
0.06
220
CLIVER Raw sewage was sampled by dipping from the top of the grit
tank.
It is not surprising that virus was also found in the grit
which had settled to the bottom of the tank. Fluid drained from the grit through a Miracloth (Chicopee Mills) filter contained unidentified enterovirus at a level of approximately 1 PFU/ml. The distinction between grit and primary sludge is fairly
Downloaded by [University of Florida] at 04:51 06 November 2015
arbitrary in this instance. Both are collections of solids sedimented spontaneously from the raw influent wastewater, but the grit settles faster and has a lower water content. The virus in the raw wastewater samples seemed to be associated with solids,
so
it was to be expected that a good deal of virus occurred
in primary sludge. Recorded levels of enteroviruses ranged from 2.4 to 15 PFU/ml in various samples, and reovirus was present at
>1 PFU/ml of primary sludge. Identification of five isolated enterovirus strains was attempted: two were poliovirus 3 , one was coxsackievirus B 5 , and two were not identifiable. Several solids associated with secondary wastewater treatment were collected on various,occasions and tested. These included waste and return activated sludge, trickling filter stones and solids sloughed from them, and blended sludge on its way to digestion. Only the sloughed trickling filter solids failed to yield virus, and this may have been due to the method of testing them. All other samples contained enteroviruses (including polioviruses2 and 3 plus some unidentified types); as high as 5 PFLJ/ml were present in a sample of return activated sludge. Reoviruses ranged from below detectable levels in some of these samples to near 5 PFU/ml in a sample of waste activated sludge.
221
VIRUSES IN WASTEWATER SOLIDS Knowing that viruses are present in the sewage plant is no
direct cause for concern, since only the grit leaves the plant in approximately the form that was tested.
We were more interested
in knowing whether virus in these solids could withstand the digestion process, as Lund3 had reported. The results shown in Table 2 indicate that viruses were consistently present after digestion. The samples
Downloaded by [University of Florida] at 04:51 06 November 2015
from which no enteroviruses were isolated had been treated with chloroform, which ought not have affected the viruses; however, enteroviruses were detected when membrane filtration was substituted for the chloroform treatment. None of these enteroviruses was identified by our methods. DISCUSSION This limited study seems to have been the first in the United States in which the emphasis has been upon viruses associated with the solid products of wastewater treatment.
It is being reported
more because of the questions i t raises than of those it answers. Viruses were expected to occur in _raw sewage sludge, but perhaps not in digested sludge. Antiviral factors in sludge TABLE I1 Viruses Found in Digested Sludge Samples Sampling Date
Enteroviruses (PFU/d)
Reoviruses (PFUIml)
12/21/73
0
8
6/12/74
0
3
8126174
0.8
4,4
11/1/74
2
2.4
222
CLIVER
d i g e s t i o n might include temperature, pH, and microbial a c t i v i t y . Temperatures of 32°C o r below do nQt d e s t r o y v i r u s e s r a p i d l y .
The
sewage treatment process monitored does n o t i n c l u d e liming, so our samples seldom reached pH 8.
Enteroviruses a r e biodegradable?
and v i r u s e s i n sludge are degraded under some conditions,"
9
but
t h e f i n a l product i n t h e p r e s e n t study s t i l l had v i r u s i n i t .
Downloaded by [University of Florida] at 04:51 06 November 2015
The volume r a t i o of raw sewage i n f l u e n t t o d i g e s t e d sludge i s approximately 300 a t t h i s p l a n t , s o t h e v i r u s t i t e r of t h e
d i g e s t e d sludge might have been as much a s 300 times t h a t of t h e i n f l u e n t & The numerical t i t e r s reported should n o t b e taken too s e r i o u s l y , b u t i t does appear t h a t a good d e a l of v i r u s w a s ina c t i v a t e d ? r a t h e r than merely separated i n t o t h e s o l i d phase, during sewage treatment. On t h e o t h e r hand, sludge d i g e s t e d i n t h i s way is a v i r u s r i c h substance which dither wants f u r t h e r treatment (perhaps a t a higher temperature) p r v e r y c a u t i o u s d i s p o s a l t o avoid hazards t o human h e a l t h .
Grit, too, should be handled w i t h care.
Given
Lund's r e p o r t 3 t h a t v i r u s sometimes withstood 50 days a t 5OoC i n sludge, a v e r y s u b s t a n t i a l i n c r e a s e i n temperature might be required. ACKNOWLEDGMENTS
T h i s r e s e a r c h w a s supported by t h e College of A g r i c u l t u r a l and L i f e Sciences, University of Wisconsin, Madison, and by the S t a t e of Wisconsin Department of Natural Resources.
I thank Folaranmi V.
Alofe, Kenneth H. Green, David J. Hurley, Kenneth D. Kostenbader,
Jr., and Beverly M. Thompson f o r t h e i r p a r t i c i p a t i o n .
VIRUSES IN WASTEWATER SOLIDS
223
REFERENCES
1. Lund, E. Water Research 7:873-879, 1973. 2. Moore, B. E. D., L . Funderburg, B. P. Sagik, and J. F. Malina, Jr. In J. F. Malina, Jr., and B. P. Sagik (eds.), Virus Survival in Water and Wastewater Systems. Center for Research in Water --Resources, University of Texas at Austin, 1974. pp. 3-15. 3. Lund, E. In S. H. Jenkins (ed.), Advances in Water Pollution Research, Vol. 1. Pergamon Press, Oxford, 1971. pp. 2411 - 2415. 4 . Schaub, S. A., C. A. Sorber, and G. W. Taylor. In J. F. Malina, Jr., and B. P. Sagik (eds.), Virus Survival in Water and Waste-. water Systems. Center for Research in Water Resources, University of Texas at Austin, 1974. pp. 71-83. 5. Korchak, G. I. Laboratornoye Dyelo 3:170-172, 1970. 6. Cliver, D. O., and R. M. Herrmann. Health Lab. Sci. 6:5-17, 1969. 7. Wallis, C., J. L. Melnick, and F. Rapp. J. Bacteriol. 92:155-160, 1966. 8. Kalter, S. S. Procedures for Routine Laboratory Diagnosis of Virus and Rickettsia1 Diseases. Burgess, Minneapolis, 1963. pp. 49-51. 9. Cliver, D. O., and J. E. Herrmann. Water Research 6:797-805, 1972. 10. Malina, J. F., Jr., K. R. Ranganathan, B. E. D. Moore, and B. P. Sagik. In J. F. Malina, Jr., and B. P. Sagik (eds.), Virus Survival --in Water and Wastewater Systems. Center for Research in Water Resources, University of Texas at Austin, 1974. pp. 95-114.
Downloaded by [University of Florida] at 04:51 06 November 2015
-
Received May 27, 1975 Accepted June 5 , 1975
ISSN: 0013-9300 (Print) (Online) Journal homepage: http://www.tandfonline.com/loi/lesa17
Virus Association with Wastewater Solids D. O. Cliver To cite this article: D. O. Cliver (1975) Virus Association with Wastewater Solids, Environmental Letters, 10:3, 215-223, DOI: 10.1080/00139307509435823 To link to this article: http://dx.doi.org/10.1080/00139307509435823
Published online: 02 Sep 2009.
Submit your article to this journal
Article views: 1
View related articles
Citing articles: 8 View citing articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=2esa20 Download by: [University of Florida]
Date: 06 November 2015, At: 04:51
ENVIRONMENTAL LETTERS, 10(3),
215-223 (1975)
VIRUS ASSOCIATION WITH WASTEWATER SOLIDS
-KEY WORDS:
enteroviruses, reoviruses, sewage, sludge
Downloaded by [University of Florida] at 04:51 06 November 2015
D. 0: Cliver
Food Research Institute and Department of Bacteriology University of Wisconsin - Madison Madison, Wisconsin 53706
ABSTRACT The solids produced at an urban wastewater treatment plant, tested semiquantitatively, almost all contained human intestinal viruses.
Reoviruses and five or more types of enteroviruses were
present.
Sludge, digested anaerobically at 30'
grit contained measurable levels of viruses.
-
32' C, and
Until reliable means
of inactivating the viruses have been developed and implemented, great care should be taken in disposing of these solids. INTRODUCTION Effluent from an urban sewage treatment plant ought not contain viruses. The human intestinal viruses which occur in community wastewater should be removed during treatment.
'Removal' might
mean either inactivation (loss of the virus' ability to cause
215 Copyright 0 1976 by h!arcel Dekker, Inc. All Rights Reserved. Neither this work nor any part may be reproduced or transmitted in any form o r by any means, electronic or mechanical, including photocopying, microfilming, and recording, or by any information storage and retrieval system, without permission in writing from the publisher.
216
CLIVER
infection) or physical separation of the virus from the wastewater before the effluent is discharged. Sewage contains solids of biological and mineral origin, and other solids may be added or produced during treatment. Virus particles are too small to sediment by themselves from wastewater, but they often associate with solids which do sediment during
Downloaded by [University of Florida] at 04:51 06 November 2015
various stages of treatment. "
The factors governing association
of virus with these solids are being investigated.3 , 4 The products of sedimentation steps in sewage treatment are grit and various kinds of sludge. Each contains a good deal of water.
They must be disposed in some fashion, and they are poorly
suited to chemical disinfection.
If viruses are present, human
illnesses might result unless great care is taken in disposal of these solids. The present study included one metropolitan treatment plant and two groups of viruses. The plant serves a relatively nonindustrial community and receives 120,obO to 140,000 m3 of influent per day, depending upon the season. It produces approximately
0.8 m 3 of grit and 480 m 3 of final, anaerobically digested sludge per day.
Treatment includes screening and grit separation, primary
sedimentation, secondary treatment by the activated sludge (principally) or the trickling filter process, and terminal chlorination of the clarified effluent.
The blend of thickened primary and
secondary (principally waste activated) sludges is digested anaerobically for 12 days at 30'
-
32OC and held for eight more days at
a temperature which, depending on the season, ranges upward from 7"
- 10°C.
217
VIRUSES IN WASTEWATER SOLIDS
The viruses sought were members of the enterovirus and reovirus groups. The enteroviruses are approximately 28 nm in diameter, contain single-stranded ribonucleic acid (RNA) , and may cause a variety of human illnesses. The group includes the polioviruses, coxsackieviruses A and B, and echoviruses. All emanate from the human intestines; the polioviruses most frequently occur in sewage
Downloaded by [University of Florida] at 04:51 06 November 2015
with the feces of those who have received the oral polio vaccine. The reoviruses are 70 to 85 nm in diameter and contain doublestranded RNA.
Their role in causing human disease is uncertain.
MATERIALS AND METHODS Samples were collected in four sets over a 12-month period. Given the known seasonal and diurnal variations in the quantity and composition of sewage arriving at the plant, there is no claim that these few 'grab' samples are statistically representative of all that passes through it. Where virus has associated with solids homogeneity of virus distribution is likely to be poor. Studies of the best methods for dissociating virus from sewage solids have produced differing results.2 y 3 y 5
Optimum conditions
seem generally to include mild alkalinity, some added protein, and physical force imposed by violent mixing or by freezing and thawing. Grab samples were he3d in the laboratory to see if sedimentation occurred.
If not, a small portion of the sample was processed for
testing in tissue culture. Where sediment did collect at the bottom of a vessel, its volume was recorded, and it was resuspended in a small volume-of supernatant fluid for further processing. Phosphate-buffered saline (pH 7.4) plus 10% fetal calf serum was
218
CLIVER
added at a rate which depended on the turbidity of the sample; the volume of diluent did not exceed that of the sample fraction being processed. The suspension was homogenized for 30 sec in a Servall OmniMixer (80~). After three or more freeze-thaw cycles, the sediment was discarded, and the supernatant fluid was treated with 5% chloro-
Downloaded by [University of Florida] at 04:51 06 November 2015
form or passed through a glass fiber prefilter and 0.2 pm porosity membrane filter (Gelman GA-8) to remove bacteria.
The resulting
extract was tested in primary rhesus (Placaca mulatta) monkey kidney cells produced by methods described elsewhere.6
Effects
seen within 24 hr were considered toxic, rather than viral. Virus was detected by microscopically visible cytopathic effects (CPE) in cultures maintained with liquid medium or by areas of cell degeneration (plaques) produced in cultures maintained with either of two semisolid media.
Enteroviruses and reoviruses
could be differentiated presumptively on the basis of differences i n their patterns of CPE.
Further, reoviruses were essentially
unable to produce plaques unless the culture medium contained pancreatin,' and they were retained absolutely by a 50 nm porosity Nuclepore filter. Serologic identification was attempted only for enteroviruses. Antisera to the three types of polioviruses and the s i x types of coxsackieviruses B were used in an agar-diffusion neutralization test adapted from that of Kalter.8
Virus
concentrations are expressed in plaque-forming units (PFU) per milliliter of the original grab sample.
219
VIRUSES I N WASTEWATER SOLIDS RESULTS p r e l i m i n a r y tests showed t h a t v i r u s w a s r e a d i l y d e t e c t a b l e i n t h e i n f l u e n t r a w sewage and w a s p r e f e r e n t i a l l y a s s o c i a t e d with t h e solids..
The v i r u s r e c o v e r e d from t h e s o l i d s may n o t
have b e e n a l l t h a t was p r e s e n t , s o any c o n c e n t r a t i o n r e p o r t e d should b e r e g a r d e d as a minimum.
Added l a b o r a t o r y p o l i o v i r u s
Downloaded by [University of Florida] at 04:51 06 November 2015
d i d n o t seem t o a s s o c i a t e s i m i l a r l y w i t h t h e s o l i d s , s o a l l o b s e r v a t i o n s had t o b e based upon n a t u r a l l y - o c c u r r i n g v i r u s . T a b l e 1 summarizes t h e r e s u l t s o b t a i n e d w i t h raw sewage sampled on f o u r o c c a s i o n s .
The numbers p r e s e n t e d are n o t s o
p r e c i s e as t o show s e a s o n a l t r e n d s , though t h e i n c i d e n c e of e n t e r o v i r u s e s o t h e r t h a n t h e v a c c i n e p o l i o v i r u s e s would be e x p e c t e d t o peak i n t h e donor p o p u l a t i o n d u r i n g l a t e summer.
Any of
t h e s e l e v e l s of v i r u s could have o c c u r r e d w i t h l e s s t h a n 1%of t h e donor p o p u l a t i o n i n f e c t e d .
P o l i o v i r u s e s 1 and 3 from t h e 8/26/74
sample were t h e o n l y v i r u s e s s e r o t y p e d s u c c e s s f u l l y from t h e r a w sewage samples. TABLE I V i r u s e s Found i n Raw Sewage Samples Sampling Date
Enteroviruses (PFU/ml)
Reoviruses (PFU/ml>
12/21/73
0.013
n o t determined
61121 74
0.17
0.17
8/26/74
0.06
0.06
11/1/74
0.09
0.06
220
CLIVER Raw sewage was sampled by dipping from the top of the grit
tank.
It is not surprising that virus was also found in the grit
which had settled to the bottom of the tank. Fluid drained from the grit through a Miracloth (Chicopee Mills) filter contained unidentified enterovirus at a level of approximately 1 PFU/ml. The distinction between grit and primary sludge is fairly
Downloaded by [University of Florida] at 04:51 06 November 2015
arbitrary in this instance. Both are collections of solids sedimented spontaneously from the raw influent wastewater, but the grit settles faster and has a lower water content. The virus in the raw wastewater samples seemed to be associated with solids,
so
it was to be expected that a good deal of virus occurred
in primary sludge. Recorded levels of enteroviruses ranged from 2.4 to 15 PFU/ml in various samples, and reovirus was present at
>1 PFU/ml of primary sludge. Identification of five isolated enterovirus strains was attempted: two were poliovirus 3 , one was coxsackievirus B 5 , and two were not identifiable. Several solids associated with secondary wastewater treatment were collected on various,occasions and tested. These included waste and return activated sludge, trickling filter stones and solids sloughed from them, and blended sludge on its way to digestion. Only the sloughed trickling filter solids failed to yield virus, and this may have been due to the method of testing them. All other samples contained enteroviruses (including polioviruses2 and 3 plus some unidentified types); as high as 5 PFLJ/ml were present in a sample of return activated sludge. Reoviruses ranged from below detectable levels in some of these samples to near 5 PFU/ml in a sample of waste activated sludge.
221
VIRUSES IN WASTEWATER SOLIDS Knowing that viruses are present in the sewage plant is no
direct cause for concern, since only the grit leaves the plant in approximately the form that was tested.
We were more interested
in knowing whether virus in these solids could withstand the digestion process, as Lund3 had reported. The results shown in Table 2 indicate that viruses were consistently present after digestion. The samples
Downloaded by [University of Florida] at 04:51 06 November 2015
from which no enteroviruses were isolated had been treated with chloroform, which ought not have affected the viruses; however, enteroviruses were detected when membrane filtration was substituted for the chloroform treatment. None of these enteroviruses was identified by our methods. DISCUSSION This limited study seems to have been the first in the United States in which the emphasis has been upon viruses associated with the solid products of wastewater treatment.
It is being reported
more because of the questions i t raises than of those it answers. Viruses were expected to occur in _raw sewage sludge, but perhaps not in digested sludge. Antiviral factors in sludge TABLE I1 Viruses Found in Digested Sludge Samples Sampling Date
Enteroviruses (PFU/d)
Reoviruses (PFUIml)
12/21/73
0
8
6/12/74
0
3
8126174
0.8
4,4
11/1/74
2
2.4
222
CLIVER
d i g e s t i o n might include temperature, pH, and microbial a c t i v i t y . Temperatures of 32°C o r below do nQt d e s t r o y v i r u s e s r a p i d l y .
The
sewage treatment process monitored does n o t i n c l u d e liming, so our samples seldom reached pH 8.
Enteroviruses a r e biodegradable?
and v i r u s e s i n sludge are degraded under some conditions,"
9
but
t h e f i n a l product i n t h e p r e s e n t study s t i l l had v i r u s i n i t .
Downloaded by [University of Florida] at 04:51 06 November 2015
The volume r a t i o of raw sewage i n f l u e n t t o d i g e s t e d sludge i s approximately 300 a t t h i s p l a n t , s o t h e v i r u s t i t e r of t h e
d i g e s t e d sludge might have been as much a s 300 times t h a t of t h e i n f l u e n t & The numerical t i t e r s reported should n o t b e taken too s e r i o u s l y , b u t i t does appear t h a t a good d e a l of v i r u s w a s ina c t i v a t e d ? r a t h e r than merely separated i n t o t h e s o l i d phase, during sewage treatment. On t h e o t h e r hand, sludge d i g e s t e d i n t h i s way is a v i r u s r i c h substance which dither wants f u r t h e r treatment (perhaps a t a higher temperature) p r v e r y c a u t i o u s d i s p o s a l t o avoid hazards t o human h e a l t h .
Grit, too, should be handled w i t h care.
Given
Lund's r e p o r t 3 t h a t v i r u s sometimes withstood 50 days a t 5OoC i n sludge, a v e r y s u b s t a n t i a l i n c r e a s e i n temperature might be required. ACKNOWLEDGMENTS
T h i s r e s e a r c h w a s supported by t h e College of A g r i c u l t u r a l and L i f e Sciences, University of Wisconsin, Madison, and by the S t a t e of Wisconsin Department of Natural Resources.
I thank Folaranmi V.
Alofe, Kenneth H. Green, David J. Hurley, Kenneth D. Kostenbader,
Jr., and Beverly M. Thompson f o r t h e i r p a r t i c i p a t i o n .
VIRUSES IN WASTEWATER SOLIDS
223
REFERENCES
1. Lund, E. Water Research 7:873-879, 1973. 2. Moore, B. E. D., L . Funderburg, B. P. Sagik, and J. F. Malina, Jr. In J. F. Malina, Jr., and B. P. Sagik (eds.), Virus Survival in Water and Wastewater Systems. Center for Research in Water --Resources, University of Texas at Austin, 1974. pp. 3-15. 3. Lund, E. In S. H. Jenkins (ed.), Advances in Water Pollution Research, Vol. 1. Pergamon Press, Oxford, 1971. pp. 2411 - 2415. 4 . Schaub, S. A., C. A. Sorber, and G. W. Taylor. In J. F. Malina, Jr., and B. P. Sagik (eds.), Virus Survival in Water and Waste-. water Systems. Center for Research in Water Resources, University of Texas at Austin, 1974. pp. 71-83. 5. Korchak, G. I. Laboratornoye Dyelo 3:170-172, 1970. 6. Cliver, D. O., and R. M. Herrmann. Health Lab. Sci. 6:5-17, 1969. 7. Wallis, C., J. L. Melnick, and F. Rapp. J. Bacteriol. 92:155-160, 1966. 8. Kalter, S. S. Procedures for Routine Laboratory Diagnosis of Virus and Rickettsia1 Diseases. Burgess, Minneapolis, 1963. pp. 49-51. 9. Cliver, D. O., and J. E. Herrmann. Water Research 6:797-805, 1972. 10. Malina, J. F., Jr., K. R. Ranganathan, B. E. D. Moore, and B. P. Sagik. In J. F. Malina, Jr., and B. P. Sagik (eds.), Virus Survival --in Water and Wastewater Systems. Center for Research in Water Resources, University of Texas at Austin, 1974. pp. 95-114.
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Received May 27, 1975 Accepted June 5 , 1975
