APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Sept. 1976, p. 352-354 Copyright X) 1976 American Society for Microbiology
Vol. 32, No. 3 Printed in U.S.A.
Recovery of Yersinia enterocolitica from Streams and Lakes of California SYDNEY HARVEY, J. R. GREENWOOD, M. J. PICKETT,* AND ROBERT A. MAH Department of Bacteriology and School of Public Health, University of California, Los Angeles, California 90024 Received for publication 19 April 1976
Stream and lake water from the Mammoth Lakes region of California was sampled for Yersinia enterocolitica. From 10 of the 34 sites examined, organisms were isolated that were biochemically identified as Y. enterocolitica. Only one of the ten strains could be serologically confirmed. This strain was identified as Y. enterocolitica serotype 16. Although an outbreak of enteritis in the area prompted this study, no correlation with gastrointestinal disease could be established since the majority of the strains were untypeable. Yersinia enterocolitica is associated with a variety of clinical syndromes. These include abscesses of the spleen, colon, and neck; septicemia; peritonitis; and mesenteric lymphadenitis (2, 6, 7). Y. enterocolitica has also caused outbreaks of gastroenteritis similar to those produced by Salmonella and Shigella (1, 8, 11). Toma and Lafleur (9) (1974) reported that in some areas Y. enterocolitica exceeds Shigella and is second only to Salmonella in frequency of isolation from fecal specimens associated with acute bacterial gastroenteritis. Although the epidemiology of Y. enterocolitica infections has not been completely elucidated, it increasingly appears that water may be a significant environmental reservoir. Support for this view is found in Lassen's (1972) isolations from drinking water in Norway (4). Nine of his eleven isolates, however, were found to be new serotypes and thus had no known correlation with human or animal disease. Of his two remaining isolates, one was serotype 13, and the other was serotype 7. Although neither of these serotypes is commonly found in clinical infections, both have been isolated from humans (4, 5). In the United States a case of Y. enterocolitica septicemia was directly related to the consumption of stream water at a hunting camp in the Adirondack (New York) Mountains (2). Additionally, a mass outbreak of gastrointestinal illness was recently reported at a ski resort in Big Sky, Mont.; 41% of 1,500 guests experienced symptoms of nausea, diarrhea, abdominal pain, or vomiting. Statistical data revealed a significant correlation between amounts of well water consumed and illness. Y. enterocolitica was subsequently isolated from the unchlorinated well water which served as the resort's water supply (10). The editorial
note on this report points out, however, that the role of Y. enterocolitica as the etiological agent is unclear. The incidence of the organism in well water of the area is unknown and acute stool specimens were not examined for Y. enterocolitica. The present study was initiated as a survey for the presence of Y. enterocolitica in California lake and stream water. The study was given added impetus and a focal point when outbreaks of gastroenteritis were reported by rangers in the Mammoth Lakes region of the California Sierra Nevada mountain range. The enteritis episodes were of unknown etiology and were circumstantially linked to the consumption of lake and stream water. We took samples from 34 lakes and streams in this area duirng 1975 and processed them for Y. enterocolitica. MATERIALS AND METHODS Isolations. Samples were collected in sterile 100ml water bottles and placed in an insulated ice chest at the collection site. A commercially available prepackaged gel (Blue Ice) was used as a coolant in the ice chest. When prefrozen, this refrigerant thaws more slowly than ice and can maintain the interior of the ice chest at a temperature slightly above freezing for as long as 2 days. The samples were kept at refrigerator temperature or below by use of the ice chest and a cabin refrigerator until received at the laboratory. The elapsed time between sample collection and processing did not exceed 2 weeks. In prior tests in this laboratory, we had determined that Y. enterocolitica survives in refrigerated water for as long as 6 months. A 1-ml sample from each water bottle was inoculated into glucose broth of the following formulation: casein peptone (Difco), 1.0%; glucose, 0.5%; bromocresol purple, 0.001%; and distilled water. The inoculated broths were incubated anaerobically in a GasPak jar (BBL) at room tem352
353
WATER ISOLATION OF Y. ENTEROCOLITICA
VOL. 32, 1976
TABLE 1. Biochemical reactions of Yersinia enterocolitica water isolatesa T
Yenterocoli-
KIA
Oxi U
LIA
MoI tiMity tlt
MIOb ONPG VP Gel Cit ADH Glu Man Inos Sor Rha Suc Mel Amy Ara
tica
strain
A_
3BP7 K/or/-
-
+ K/Al- +-+
+
A 4Sc3 K/or/-
-
+ K/A/- +++
+
K/A/-
+
-
_ -
-
-
+
-
+
-
+
-
+
+
+
+
-
+
+
-
-
+
+
-
+
-
-
-
+
+
-
-
+ +
+ +
-
-
-
-
+ +
- - -
+ +
+ +
-
-
+ +
+ +
+ +
+ +
+ +
+ +
-
+ +
+ +
-
+
-
+
+
-
+
+
+
-
+
+
-
-
+ K/A/- +-+
+
-
-
K/A/- K/A/K/A/- K/A/- _
+ K/A/- +-+ + K/A/- +-+
+ +
-
_ -
+ K/A/- +++ + K/A/- +++
+ +
-
-
-
-
-
-
A CF-1 K/or/- -
+ K/A/- +++
+
-
-
7E 6-1 4-1 14-2 16-3
+
+
-
A K/or/-
-
+ K/A/- +++
+
-
-
-
-
+
+
-
+
+
+
-
+
+
-
A 2SI-1 K/or/-
-
+ K/A/- +++
+
-
-
+
-
+
+
-
+
+
+
+
+
+
-
H-2
a Abbreviations: KIA, Kligler iron agar; Oxi, oxidase; U, urease; LIA, lysine iron agar; MIO, motility-indole-ornithine; ONPG, o-nitrophenyl-,&/-galactopyranoside; VP, Voges-Proskauer; Gel, gelatin; Cit, citrate; ADH, arginine dihydrolase; Glu, glucose; Man, mannitol; Inos, inositol; Sor, sorbitol; Rha, rhamnose; Suc, sucrose; Mel, melibiose; Amy, amygdalin; Ara, arabinose; K, alkaline; A, acid; ®, acid + gas. b MIO at room temperature; other tests at 35°C.
perature and checked at 24-h intervals, up to a maximum of 96 h, for visual evidence of acid production. When acid production was detected, three loopfuls of broth were subcultured to a MacConkeyTween-80 plate and streaked for isolation. MacConkey-Tween-80 medium was developed by W. H. Lee (Abstr. Annu. Meet. Am. Soc. Microbiol. 1975, P 16, p. 202) for detection of Y. enterocolitica in food. This medium was prepared by adding 0.02% CaCl2 and 1.0% (vol/vol) Tween-80 (Difco) to MacConkey agar (Difco) before autoclaving. These plates were incubated for 48 h anaerobically at room temperature and then examined for colonies indicative of Y. enterocolitica. Isolation media were incubated anaerobically to prevent overgrowth by the strict aerobes found in natural waters. All plates were stored at room temperature aerobically for an additional 24 h to detect any subsequent Tween hydroly-
was rubbed into the paper with a platinum loop. Motility, ornithine decarboxylase production, and indole production were determined by using dehydrated motility-indole-ornithine (Difco) medium prepared and dispensed according to the manufacturer's directions. The medium was stabbed to the bottom and incubated for 18 to 24 h at room temperature. Motility at 35°C was determined in motility agar prepared by adding 1% tryptose (Difco) and 0.8% infusion agar (Difco) to distilled water. The medium was dispensed in 5-ml amounts into tubes (13 by 100 mm) and autoclaved for 15 min at 121°C. Motility agar was inoculated by stabbing 5 mm into the center of the medium and incubated at 35°C for 18 to 24 h. Other biochemical results were determined with the API Enteric 20 system (Analytab Products, Inc.) incubated for 18 to 24 h at 35°C.
S1S.
RESULTS Ten strains of Y. enterocolitica were isolated from 10 of the sites examined; the cultures were identified biochemically. Table 1 shows the results of these tests. These differ partly from the majority of Y. enterocolitica isolates in that strains 2SI-1, 14-2, 16-3, CF-1, and H-2 produce acid from rhamnose, strains 2SI-1 and CF-1 are citrate positive, strains 3BP7, 7E, 6-1, and 4-1 failed to produce acid from sucrose and only two
Biochemical studies. Potential Y. enterocolitica colonies were inoculated into tubes of Kligler iron agar (KIA) medium (Difco) which were read after 24 h of incubation at 35°C. All KIA slants giving Y. enterocolitica-like reactions (slant, alkaline; butt, acid with or without small amounts of gas; no H2S) were further biochemically characterized. Oxidase production was tested by using filter paper strips saturated (then drained and dried at 35°C) with 1 g% tetramethylparaphenylene diamine dihydrochloride in 0.2 g% ascorbic acid. For the test a bit of cell paste
354
HARVEY ET AL.
APPL. ENVIRON. MICROBIOL.
The fact that the majority of isolations were made from sites only infrequently visited by campers argues against human pollution as the source of the organisms. It has been suggested in the literature that animals, both wild and domestic, may also constitute an important reservoir (3). We have begun sampling animal droppings and other environmental specimens from the Mammoth Lakes region in an effort to determine the distribution of Y. enterocolitica DISCUSSION in this area. Y. enterocolitica may be even more Of the 10 strains isolated, five (3BP7, 4Sc3, ubiquitous than our findings have indicated, 7E, 6-1, 4-1) came from different sampling sites but no resolution of its distribution can be made along the same creek. These sites were at ap- until selective media are available for quantifyproximately 5-mile (ca. 8 km) intervals; three of ing its presence in various samples. these (3BP7, 4Sc3, 4-1) were below and two (7E, LITERATURE CITED 6-1) were above established areas of transient J., M. Laverdiere, B. Martineau, and L. and permanent human habitations. The other 1. Delorme, Lafleur. 1974. Yersiniosis in children. Can. Med. five isolations were from five different lakes. Assoc. J. 110:281-284. One of these lakes (2SI) is the source of a main 2. Keet, E. E. 1974. Yersinia enterocolitica septicemia. N.Y. State J. Med. 74:2226-2230. branch of the creek from which the other isolaE. V. 1972. Yersinia enterocolitica isolated tions were made. Two of the remaining four 3. Langford, from animals in the Fraser Valley of British Columlakes (14-2, 16-3) are in close proximity to one bia. Can. Vet. J. 13:109-113. another and drain into this same main branch. 4. Lassen, J. 1972. Yersinia enterocolitica in drinking water. Scand. J. Infect. Dis. 4:125-127. The other two lakes (CF-1, H-2) have no conB. 1973. Host range, temperature characterisnection with the drainage group discussed 5. Nilehn, tics, and serologic relationships among Yersinia above and are a considerable distance from it phages. Contrib. Microbiol. Immunol. 2:59-67. and from each other. One of these lakes (CF-1) 6. Rabson, A. R., A. F. Hallett, and H. J. Koornhof. 1975. Generalized Yersinia enterocolitica infection. J. is relatively inaccessible and at an elevation of 11,300 feet (ca. 4,200 m). All of the lakes are 7. Infect. Dis. 131:447-451. Sonnenwirth, A. C., and R. E. Weaver. 1970. Yersinia above 9,000 feet (ca. 2,280 m) in elevation. The enterocolitica. N. Engl. J. Med. 283:1468. sampling reported here was done during the 8. Toivanen, P., A. Toivanen, L. Olkkonen, and S. Aantaa. 1973. Hospital outbreak of Yersinia enteromonths of June, July, and August since most of colitica infection. Lancet 1:801-803. this area is impenetrable during the other 9. Toma, S., and L. Lafleur. 1974. Survey on the months of the year. incidence of Yersinia enterocolitica infection in The degree of pathogenicity for man of the Canada. Appl. Microbiol. 28:469-473. present isolates is unknown. The prototype of 10. U.S. Department of Health, Education, and Welfare. 1975. Morbid. Mortal. Week. Rep. 24:141-142. serotype 16 was isolated from man, but since H., and T. Maruyama. 1972. The first sucthe remaining nine isolates are presently clas- 11. Zen-Yoji, cessful isolations and identification of Yersinia sified as untypeable, no correlation with gasenterocolitica from human cases in Japan. Jpn. J. trointestinal disease has been established. Microbiol. 16:493-500.
strains, 14-2 and 16-3, produced acid from inositol. Five of the strains, 3BP7, 4Sc3, 2SI-1, CF-1, and H-2, produced varying amounts of gas in kligler iron agar medium. Only one of the ten strains proved typeable by the system presently used at the Microbial Diseases Laboratory of the California Department of Health. This was culture 4Sc3 which they identified as Y. enterocolitica serotype 16.
Vol. 32, No. 3 Printed in U.S.A.
Recovery of Yersinia enterocolitica from Streams and Lakes of California SYDNEY HARVEY, J. R. GREENWOOD, M. J. PICKETT,* AND ROBERT A. MAH Department of Bacteriology and School of Public Health, University of California, Los Angeles, California 90024 Received for publication 19 April 1976
Stream and lake water from the Mammoth Lakes region of California was sampled for Yersinia enterocolitica. From 10 of the 34 sites examined, organisms were isolated that were biochemically identified as Y. enterocolitica. Only one of the ten strains could be serologically confirmed. This strain was identified as Y. enterocolitica serotype 16. Although an outbreak of enteritis in the area prompted this study, no correlation with gastrointestinal disease could be established since the majority of the strains were untypeable. Yersinia enterocolitica is associated with a variety of clinical syndromes. These include abscesses of the spleen, colon, and neck; septicemia; peritonitis; and mesenteric lymphadenitis (2, 6, 7). Y. enterocolitica has also caused outbreaks of gastroenteritis similar to those produced by Salmonella and Shigella (1, 8, 11). Toma and Lafleur (9) (1974) reported that in some areas Y. enterocolitica exceeds Shigella and is second only to Salmonella in frequency of isolation from fecal specimens associated with acute bacterial gastroenteritis. Although the epidemiology of Y. enterocolitica infections has not been completely elucidated, it increasingly appears that water may be a significant environmental reservoir. Support for this view is found in Lassen's (1972) isolations from drinking water in Norway (4). Nine of his eleven isolates, however, were found to be new serotypes and thus had no known correlation with human or animal disease. Of his two remaining isolates, one was serotype 13, and the other was serotype 7. Although neither of these serotypes is commonly found in clinical infections, both have been isolated from humans (4, 5). In the United States a case of Y. enterocolitica septicemia was directly related to the consumption of stream water at a hunting camp in the Adirondack (New York) Mountains (2). Additionally, a mass outbreak of gastrointestinal illness was recently reported at a ski resort in Big Sky, Mont.; 41% of 1,500 guests experienced symptoms of nausea, diarrhea, abdominal pain, or vomiting. Statistical data revealed a significant correlation between amounts of well water consumed and illness. Y. enterocolitica was subsequently isolated from the unchlorinated well water which served as the resort's water supply (10). The editorial
note on this report points out, however, that the role of Y. enterocolitica as the etiological agent is unclear. The incidence of the organism in well water of the area is unknown and acute stool specimens were not examined for Y. enterocolitica. The present study was initiated as a survey for the presence of Y. enterocolitica in California lake and stream water. The study was given added impetus and a focal point when outbreaks of gastroenteritis were reported by rangers in the Mammoth Lakes region of the California Sierra Nevada mountain range. The enteritis episodes were of unknown etiology and were circumstantially linked to the consumption of lake and stream water. We took samples from 34 lakes and streams in this area duirng 1975 and processed them for Y. enterocolitica. MATERIALS AND METHODS Isolations. Samples were collected in sterile 100ml water bottles and placed in an insulated ice chest at the collection site. A commercially available prepackaged gel (Blue Ice) was used as a coolant in the ice chest. When prefrozen, this refrigerant thaws more slowly than ice and can maintain the interior of the ice chest at a temperature slightly above freezing for as long as 2 days. The samples were kept at refrigerator temperature or below by use of the ice chest and a cabin refrigerator until received at the laboratory. The elapsed time between sample collection and processing did not exceed 2 weeks. In prior tests in this laboratory, we had determined that Y. enterocolitica survives in refrigerated water for as long as 6 months. A 1-ml sample from each water bottle was inoculated into glucose broth of the following formulation: casein peptone (Difco), 1.0%; glucose, 0.5%; bromocresol purple, 0.001%; and distilled water. The inoculated broths were incubated anaerobically in a GasPak jar (BBL) at room tem352
353
WATER ISOLATION OF Y. ENTEROCOLITICA
VOL. 32, 1976
TABLE 1. Biochemical reactions of Yersinia enterocolitica water isolatesa T
Yenterocoli-
KIA
Oxi U
LIA
MoI tiMity tlt
MIOb ONPG VP Gel Cit ADH Glu Man Inos Sor Rha Suc Mel Amy Ara
tica
strain
A_
3BP7 K/or/-
-
+ K/Al- +-+
+
A 4Sc3 K/or/-
-
+ K/A/- +++
+
K/A/-
+
-
_ -
-
-
+
-
+
-
+
-
+
+
+
+
-
+
+
-
-
+
+
-
+
-
-
-
+
+
-
-
+ +
+ +
-
-
-
-
+ +
- - -
+ +
+ +
-
-
+ +
+ +
+ +
+ +
+ +
+ +
-
+ +
+ +
-
+
-
+
+
-
+
+
+
-
+
+
-
-
+ K/A/- +-+
+
-
-
K/A/- K/A/K/A/- K/A/- _
+ K/A/- +-+ + K/A/- +-+
+ +
-
_ -
+ K/A/- +++ + K/A/- +++
+ +
-
-
-
-
-
-
A CF-1 K/or/- -
+ K/A/- +++
+
-
-
7E 6-1 4-1 14-2 16-3
+
+
-
A K/or/-
-
+ K/A/- +++
+
-
-
-
-
+
+
-
+
+
+
-
+
+
-
A 2SI-1 K/or/-
-
+ K/A/- +++
+
-
-
+
-
+
+
-
+
+
+
+
+
+
-
H-2
a Abbreviations: KIA, Kligler iron agar; Oxi, oxidase; U, urease; LIA, lysine iron agar; MIO, motility-indole-ornithine; ONPG, o-nitrophenyl-,&/-galactopyranoside; VP, Voges-Proskauer; Gel, gelatin; Cit, citrate; ADH, arginine dihydrolase; Glu, glucose; Man, mannitol; Inos, inositol; Sor, sorbitol; Rha, rhamnose; Suc, sucrose; Mel, melibiose; Amy, amygdalin; Ara, arabinose; K, alkaline; A, acid; ®, acid + gas. b MIO at room temperature; other tests at 35°C.
perature and checked at 24-h intervals, up to a maximum of 96 h, for visual evidence of acid production. When acid production was detected, three loopfuls of broth were subcultured to a MacConkeyTween-80 plate and streaked for isolation. MacConkey-Tween-80 medium was developed by W. H. Lee (Abstr. Annu. Meet. Am. Soc. Microbiol. 1975, P 16, p. 202) for detection of Y. enterocolitica in food. This medium was prepared by adding 0.02% CaCl2 and 1.0% (vol/vol) Tween-80 (Difco) to MacConkey agar (Difco) before autoclaving. These plates were incubated for 48 h anaerobically at room temperature and then examined for colonies indicative of Y. enterocolitica. Isolation media were incubated anaerobically to prevent overgrowth by the strict aerobes found in natural waters. All plates were stored at room temperature aerobically for an additional 24 h to detect any subsequent Tween hydroly-
was rubbed into the paper with a platinum loop. Motility, ornithine decarboxylase production, and indole production were determined by using dehydrated motility-indole-ornithine (Difco) medium prepared and dispensed according to the manufacturer's directions. The medium was stabbed to the bottom and incubated for 18 to 24 h at room temperature. Motility at 35°C was determined in motility agar prepared by adding 1% tryptose (Difco) and 0.8% infusion agar (Difco) to distilled water. The medium was dispensed in 5-ml amounts into tubes (13 by 100 mm) and autoclaved for 15 min at 121°C. Motility agar was inoculated by stabbing 5 mm into the center of the medium and incubated at 35°C for 18 to 24 h. Other biochemical results were determined with the API Enteric 20 system (Analytab Products, Inc.) incubated for 18 to 24 h at 35°C.
S1S.
RESULTS Ten strains of Y. enterocolitica were isolated from 10 of the sites examined; the cultures were identified biochemically. Table 1 shows the results of these tests. These differ partly from the majority of Y. enterocolitica isolates in that strains 2SI-1, 14-2, 16-3, CF-1, and H-2 produce acid from rhamnose, strains 2SI-1 and CF-1 are citrate positive, strains 3BP7, 7E, 6-1, and 4-1 failed to produce acid from sucrose and only two
Biochemical studies. Potential Y. enterocolitica colonies were inoculated into tubes of Kligler iron agar (KIA) medium (Difco) which were read after 24 h of incubation at 35°C. All KIA slants giving Y. enterocolitica-like reactions (slant, alkaline; butt, acid with or without small amounts of gas; no H2S) were further biochemically characterized. Oxidase production was tested by using filter paper strips saturated (then drained and dried at 35°C) with 1 g% tetramethylparaphenylene diamine dihydrochloride in 0.2 g% ascorbic acid. For the test a bit of cell paste
354
HARVEY ET AL.
APPL. ENVIRON. MICROBIOL.
The fact that the majority of isolations were made from sites only infrequently visited by campers argues against human pollution as the source of the organisms. It has been suggested in the literature that animals, both wild and domestic, may also constitute an important reservoir (3). We have begun sampling animal droppings and other environmental specimens from the Mammoth Lakes region in an effort to determine the distribution of Y. enterocolitica DISCUSSION in this area. Y. enterocolitica may be even more Of the 10 strains isolated, five (3BP7, 4Sc3, ubiquitous than our findings have indicated, 7E, 6-1, 4-1) came from different sampling sites but no resolution of its distribution can be made along the same creek. These sites were at ap- until selective media are available for quantifyproximately 5-mile (ca. 8 km) intervals; three of ing its presence in various samples. these (3BP7, 4Sc3, 4-1) were below and two (7E, LITERATURE CITED 6-1) were above established areas of transient J., M. Laverdiere, B. Martineau, and L. and permanent human habitations. The other 1. Delorme, Lafleur. 1974. Yersiniosis in children. Can. Med. five isolations were from five different lakes. Assoc. J. 110:281-284. One of these lakes (2SI) is the source of a main 2. Keet, E. E. 1974. Yersinia enterocolitica septicemia. N.Y. State J. Med. 74:2226-2230. branch of the creek from which the other isolaE. V. 1972. Yersinia enterocolitica isolated tions were made. Two of the remaining four 3. Langford, from animals in the Fraser Valley of British Columlakes (14-2, 16-3) are in close proximity to one bia. Can. Vet. J. 13:109-113. another and drain into this same main branch. 4. Lassen, J. 1972. Yersinia enterocolitica in drinking water. Scand. J. Infect. Dis. 4:125-127. The other two lakes (CF-1, H-2) have no conB. 1973. Host range, temperature characterisnection with the drainage group discussed 5. Nilehn, tics, and serologic relationships among Yersinia above and are a considerable distance from it phages. Contrib. Microbiol. Immunol. 2:59-67. and from each other. One of these lakes (CF-1) 6. Rabson, A. R., A. F. Hallett, and H. J. Koornhof. 1975. Generalized Yersinia enterocolitica infection. J. is relatively inaccessible and at an elevation of 11,300 feet (ca. 4,200 m). All of the lakes are 7. Infect. Dis. 131:447-451. Sonnenwirth, A. C., and R. E. Weaver. 1970. Yersinia above 9,000 feet (ca. 2,280 m) in elevation. The enterocolitica. N. Engl. J. Med. 283:1468. sampling reported here was done during the 8. Toivanen, P., A. Toivanen, L. Olkkonen, and S. Aantaa. 1973. Hospital outbreak of Yersinia enteromonths of June, July, and August since most of colitica infection. Lancet 1:801-803. this area is impenetrable during the other 9. Toma, S., and L. Lafleur. 1974. Survey on the months of the year. incidence of Yersinia enterocolitica infection in The degree of pathogenicity for man of the Canada. Appl. Microbiol. 28:469-473. present isolates is unknown. The prototype of 10. U.S. Department of Health, Education, and Welfare. 1975. Morbid. Mortal. Week. Rep. 24:141-142. serotype 16 was isolated from man, but since H., and T. Maruyama. 1972. The first sucthe remaining nine isolates are presently clas- 11. Zen-Yoji, cessful isolations and identification of Yersinia sified as untypeable, no correlation with gasenterocolitica from human cases in Japan. Jpn. J. trointestinal disease has been established. Microbiol. 16:493-500.
strains, 14-2 and 16-3, produced acid from inositol. Five of the strains, 3BP7, 4Sc3, 2SI-1, CF-1, and H-2, produced varying amounts of gas in kligler iron agar medium. Only one of the ten strains proved typeable by the system presently used at the Microbial Diseases Laboratory of the California Department of Health. This was culture 4Sc3 which they identified as Y. enterocolitica serotype 16.
