Laboratory Animals (1978) 12, 149-150.
149
Water-borne Bacillus licheniformis infection in mice D. J. M. WRIGHT,'" D. J. FROST'" & P. EATONt .••Department of Microbiology and London, W68RF
t Animal
Unit, Charing Cross Hospital Medical School, Fulham Palace Road,
Summary A water-borne Bacillus licheniformis infection was associated with depressed haemoglobin content, white cell and platelet count. The epidemic was resolved by changing from tanked to mains water supply.
In the course of studies of experimental infection of CBA/Ca mice with Borrelia duttoni, an increase in the severity of infection was observed (as mortality) when normally the mice survived. Concurrently the productivity of the breeding colony, which was kept in a separate part of the animal house, decreased from 1· 2 to O· 5 pups per female per week. Analysis of the haemoglobin content, white-cell count and platelet count of uninfected stock mice showed a marked depression from normal values (Table I). Closer analysis revealed that the extreme haematological changes were not confined to this particular inbred strain, but affected in some degree all other strains of mice kept in the animal house, with the exception of animals kept in a germ-free environment. By contrast, mice purchased from a commercial source maintained normal haematological values and were not prone to fatal Borrelia infection provided that they were supplied with distilled water, or animal house tap water which had been autoclaved. The drinking water bottles were also autoclaved. When these mice were given untreated animal-house water depression of white-cell and platelet counts occurred, and reached their lowest level within a week. Similarly, mice bred in the animal house, but given sterilized drinking water, showed a return of white-cell and platelet counts to normal within 2 weeks and the haemoglobin content and reticulocyte count increased simultaneously.
Table 1. Haematological changes in the CHA/Ca mouse colony Means of 10 samples ± 1 standard deviation Normal
mouse values White-cell count (109/1) Platelet count (10911) Haemoglobin concentration (g/dl)
9·6 485 16·8
± 0·5 ± 60 ± 0·8
UninJected
stock mouse 4·6 171 9·8
± ± ±
0·4 48 1·1
Chemical analysis of the water revealed a concentration of copper of 0·02 mg/I, zinc O· 2 mg/I, with iron, lead, manganese, nickel and chromium VI+ all less than 0·01 mg/I. These were all within the normal values for chemical composition of water in the area. The presumptive coliform count was less than 1 per 100 ml of water. The total bacterial count was greater than 1800 per 100 ml using McCready's probability tables (Cruickshank, Duguid, Marmion & Swain, 1975). 100 ml tap water was filtered through a 0·22.um cellulose filter (Millipore (UK) Ltd, Abbey Road, London, NW 10 7SP) and culture of the filter yielded 7 different Bacillus colonies, 5 of which were B. licheniformis (Cowan, 1975). A week after providing the mice with a continuous supply of tap water, numerous colonies of B. licheniformis were isolated on bloodagar plates from faeces and caecal contents. This finding persisted for 2 weeks after returning to a sterile water supply. None of the mice supplied with tap water remained as carriers of B. licheniformis. Faecal carriage of the organism was not detected when 30 freshly-introduced mice were examined. The changes in white-cell and platelet counts could be reproduced experimentally within 2 hours by intraperitoneal inoculation of a O· I ml suspension of 108 B. licheniformis. These changes persisted for 3 days, but then returned to normal values. Faeces and blood cultures were persistently negative following experimental intraperitoneal infection. Decreasing dilutions of an overnight nutrient broth culture were prepared and given orally to 30 mice, the mice heing supplied with sterile water. The smallest concentration of organisms which depressed the whitecell and platelet counts was 100 bacteria. This occurred a week after ingestion. Controls treated with sterile broth showed no change in their haematological values. Organisms were isolated from faeces for up to 10 days after oral administration. Heart blood cultures were persistently sterile. There was again no evidence of permanent faecal carriage. Similar haematological changes were observed following intraperitoneal inoculation of a stock culture of B. licheniformis (NTC 10341) received from the National Collection of Type Cultures. Filtrates of a broth culture of B. licheniformis through a 0·22 ,urn cellulose filter failed to produce these haematological changes after both oral or intraperitoneal administration.
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Wright, Frost & Eaton
150 Cultures of Bacillus species other than B. anthracis have been shown to be patho'genic for mice, especially B. cereus (Bonventre & Johnson, 1970). This is the 1st report of an epidemic of a B. licheniformis waterborne infection. It was resolved by changing the drinking water supply from a roof storage cistern to mains water.
References Bonventre, P. F. & Johnson, C. E. (1970). Bacterial protein toxins. In Microbial toxins (ed. T. C. Montie, S. Kadis & S. J. Ajl), vol. 3, pp. 415-435. New York: Academic Press. Cowan, S. T. (1975). Cowan and Steel's manual for the
Acknowledgements
We wish to thank Dr P. C. Turnbull of the Food Hygiene Laboratory, Public Health Service, Colindale for verifying the strain of Bacillus species, and Mr P. A. H. Sperring of the Thames Water Authority for the chemical analysis of the water.
identification of medical bacteria. 2nd cd., p. 71. Cambridge: Cambridge University Press. Cruikshank, R., Duguid, J. P., Marmion, B. P. & Swain, R. H. A. (1975). Medical microbiology, 12th cd., vol. 2, pp. 273-276. Edinburgh: Churchill-Livingstone.
Downloaded from lan.sagepub.com at University of Texas Libraries on June 5, 2015
149
Water-borne Bacillus licheniformis infection in mice D. J. M. WRIGHT,'" D. J. FROST'" & P. EATONt .••Department of Microbiology and London, W68RF
t Animal
Unit, Charing Cross Hospital Medical School, Fulham Palace Road,
Summary A water-borne Bacillus licheniformis infection was associated with depressed haemoglobin content, white cell and platelet count. The epidemic was resolved by changing from tanked to mains water supply.
In the course of studies of experimental infection of CBA/Ca mice with Borrelia duttoni, an increase in the severity of infection was observed (as mortality) when normally the mice survived. Concurrently the productivity of the breeding colony, which was kept in a separate part of the animal house, decreased from 1· 2 to O· 5 pups per female per week. Analysis of the haemoglobin content, white-cell count and platelet count of uninfected stock mice showed a marked depression from normal values (Table I). Closer analysis revealed that the extreme haematological changes were not confined to this particular inbred strain, but affected in some degree all other strains of mice kept in the animal house, with the exception of animals kept in a germ-free environment. By contrast, mice purchased from a commercial source maintained normal haematological values and were not prone to fatal Borrelia infection provided that they were supplied with distilled water, or animal house tap water which had been autoclaved. The drinking water bottles were also autoclaved. When these mice were given untreated animal-house water depression of white-cell and platelet counts occurred, and reached their lowest level within a week. Similarly, mice bred in the animal house, but given sterilized drinking water, showed a return of white-cell and platelet counts to normal within 2 weeks and the haemoglobin content and reticulocyte count increased simultaneously.
Table 1. Haematological changes in the CHA/Ca mouse colony Means of 10 samples ± 1 standard deviation Normal
mouse values White-cell count (109/1) Platelet count (10911) Haemoglobin concentration (g/dl)
9·6 485 16·8
± 0·5 ± 60 ± 0·8
UninJected
stock mouse 4·6 171 9·8
± ± ±
0·4 48 1·1
Chemical analysis of the water revealed a concentration of copper of 0·02 mg/I, zinc O· 2 mg/I, with iron, lead, manganese, nickel and chromium VI+ all less than 0·01 mg/I. These were all within the normal values for chemical composition of water in the area. The presumptive coliform count was less than 1 per 100 ml of water. The total bacterial count was greater than 1800 per 100 ml using McCready's probability tables (Cruickshank, Duguid, Marmion & Swain, 1975). 100 ml tap water was filtered through a 0·22.um cellulose filter (Millipore (UK) Ltd, Abbey Road, London, NW 10 7SP) and culture of the filter yielded 7 different Bacillus colonies, 5 of which were B. licheniformis (Cowan, 1975). A week after providing the mice with a continuous supply of tap water, numerous colonies of B. licheniformis were isolated on bloodagar plates from faeces and caecal contents. This finding persisted for 2 weeks after returning to a sterile water supply. None of the mice supplied with tap water remained as carriers of B. licheniformis. Faecal carriage of the organism was not detected when 30 freshly-introduced mice were examined. The changes in white-cell and platelet counts could be reproduced experimentally within 2 hours by intraperitoneal inoculation of a O· I ml suspension of 108 B. licheniformis. These changes persisted for 3 days, but then returned to normal values. Faeces and blood cultures were persistently negative following experimental intraperitoneal infection. Decreasing dilutions of an overnight nutrient broth culture were prepared and given orally to 30 mice, the mice heing supplied with sterile water. The smallest concentration of organisms which depressed the whitecell and platelet counts was 100 bacteria. This occurred a week after ingestion. Controls treated with sterile broth showed no change in their haematological values. Organisms were isolated from faeces for up to 10 days after oral administration. Heart blood cultures were persistently sterile. There was again no evidence of permanent faecal carriage. Similar haematological changes were observed following intraperitoneal inoculation of a stock culture of B. licheniformis (NTC 10341) received from the National Collection of Type Cultures. Filtrates of a broth culture of B. licheniformis through a 0·22 ,urn cellulose filter failed to produce these haematological changes after both oral or intraperitoneal administration.
Downloaded from lan.sagepub.com at University of Texas Libraries on June 5, 2015
Wright, Frost & Eaton
150 Cultures of Bacillus species other than B. anthracis have been shown to be patho'genic for mice, especially B. cereus (Bonventre & Johnson, 1970). This is the 1st report of an epidemic of a B. licheniformis waterborne infection. It was resolved by changing the drinking water supply from a roof storage cistern to mains water.
References Bonventre, P. F. & Johnson, C. E. (1970). Bacterial protein toxins. In Microbial toxins (ed. T. C. Montie, S. Kadis & S. J. Ajl), vol. 3, pp. 415-435. New York: Academic Press. Cowan, S. T. (1975). Cowan and Steel's manual for the
Acknowledgements
We wish to thank Dr P. C. Turnbull of the Food Hygiene Laboratory, Public Health Service, Colindale for verifying the strain of Bacillus species, and Mr P. A. H. Sperring of the Thames Water Authority for the chemical analysis of the water.
identification of medical bacteria. 2nd cd., p. 71. Cambridge: Cambridge University Press. Cruikshank, R., Duguid, J. P., Marmion, B. P. & Swain, R. H. A. (1975). Medical microbiology, 12th cd., vol. 2, pp. 273-276. Edinburgh: Churchill-Livingstone.
Downloaded from lan.sagepub.com at University of Texas Libraries on June 5, 2015
