Isolation of sucrose-negativeYersinia enterocolitica biotype 3 serotype 03 strains and their pathogenicity

The sucrose-negative strains ofYersinia enterocolitica biotype 3 serotype 03 phage type 2 were isolated from cecal contents and oral cavity swabs of slaughtered pigs and from a swab of a skinner at the slaughterhouse. These organisms differed fromY. kristensenii, determined by assaying the antibiotic susceptibilities to ampicillin, carbenicillin, and cephalothin. These organisms showed positive reactions in the presence of 44 Md plasmid, a calcium dependency, autoagglutination activity, and produced diarrhea in mice and a negative reaction for pyrazinamidase activity. Plasmid digestion with restriction endonucleases isolated from this organism showed the same patterns in biotypes 3 and 4 serotype 03. Therefore, the sucrose-negative strains ofY. enterocolitica biotype 3 serotype 03 are apparently pathogenic.     

Biochemical,serological, and phage typing characteristics of 459Yersinia strains isolated from a terrestrial ecosystem

The origins of human contamination withYersinia enterocolitica are still unknown. We have investigated the major components of a terrestrial ecosystem (soil, earthworms, field voles, shrews, crops, hares, rabbits, and birds) for the presence ofYersinia. Four hundred fifty-nine strains ofYersinia were isolated. We report the first isolations of typicalY. enterocolitica belonging to classical or new biotypes and ofY. enterocolitica-like organisms (sucrose negative; rhamnose positive; melibiose and rhamnose positive) from soil samples, earthworms, crops, and birds. Sucrose-negativeY. enterocolitica strains and biotypes 1, 2, and 3, usually associated with human nonmesenteric syndromes, are predominant in soil, which can be considered as a reservoir for these biotypes.Y. enterocolitica serogroups O∶3 and O∶9, strains of which are responsible in Europe for human mesenteric syndromes, were not found in this study. The epidemiology ofY. enterocolitica infections is discussed.     

Application of Fluorescent Amplified Fragment Length Polymorphism for Comparison of Human and Animal Isolates of Yersinia enterocolitica

An amplified fragment length polymorphism (AFLP) method, developed to genotype Yersinia enterocolitica, has been used to investigate 70 representative strains isolated from humans, pigs, sheep, and cattle in the United Kingdom. AFLP primarily distinguished Y. enterocolitica strains according to their biotype, with strains dividing into two distinct clusters: cluster A, comprising largely the putatively pathogenic biotypes (BT2 to -4), and cluster B, comprising the putatively nonpathogenic biotype 1A strains and a single BT1B isolate. Within these two clusters, subclusters formed largely on the basis of serotype. However, AFLP profiles also allowed differentiation of strains within these serotype-related subclusters, indicating the high discriminatory power of the technique for Y. enterocolitica. Investigation of the relationship between strain AFLP profile and host confirmed that pigs are, and provides further proof that sheep may be, potential sources of human infection with putatively pathogenic strains. However, the results suggest that some strains causing human disease do not come from veterinary sources identifiable at this time. The distribution of some BT1A isolates within cluster A raises questions about the relationship between virulence potential and biotype.     

Yersinia strains isolated from river water: biochemical,serological, and phage typing characteristics

Twenty-fourYersinia enterocolitica-like strains were isolated from heavily contaminated river water. Twenty-three of the strains could only be isolated on deoxycholate-hydrogen sulfidelactose agar after cold-enrichment in tryptone soya broth. Biochemically, these strains exhibited the common properties ofY. enterocolitica. However, most strains were also melibiose-, rhamnose-, raffinose-, and Simmons’ citrate-positive. Two strains fermented lactose. The serological typing showed that the strains belonged to the serotypes O:1, O:14, O:38 and O:55. Four strains had a K-antigen linked to a complex antigenic structure. Two strains were autoagglutinated. One strain was agglutinated by two different serotypes. The strains belonged to the phage types X_o and X_z.     

Isolation ofYersinia enterocolitica andYersinia enterocolitica-like organisms from raw milk in Italy

Thirty samples of raw milk, originating from individual producers in the Turin area, were examined for the presence ofYersinia enterocolitica. A cold enrichment method with phosphate-buffered saline (PBS) 1/15M, pH 7.6, and sorbitol-bile-salts broth (SB) was used. After 7, 14, or 21 days at 4°–5°C, plating was performed on selective agar media directly (MacConkey agar andSalmonella-Shigella agar) after the alkali method was used. Six strains ofY. enterocolitica (biotype 1) and 32 strainsY. enterocolitica-like (threeY. fredericksenii; nineYersinia rhamnose-, melibiose+, -methyl-d-glucoside+, raffinose+, probablyYersinia intermedia biotype rhamnose-; and 20Y. intermedia) were isolated.Yersinia strains were found in 11 samples of raw milk, andY. enterocolitica in four samples.     

Regulatory variance of aspartate transcarbamoylase among strains ofYersinia enterocolitica andYersinia enterocolitica-like organisms

Aspartate transcarbamoylase (ATCase) has been isolated and characterized from 20 different strains ofYersinia enterocolitica andY. enterocolitica-like organisms. A variety of regulatory properties have emerged for the ATCases from the different strains. These regulatory properties may be used as a taxonomic tool to divideY. enterocolitica andY. enterocolitica-like organisms into separate groups. Results are in accord with the recent assignment ofY. enterocolitica andY. enterocolitica-like organisms to four DNA-relatedness groups and four correspondingYersinia species.     

Serotypes and mating types of clinical strains ofCryptococcus neoformans isolated in Taiwan

Twenty-one strains ofCryptococcus neoformans isolated from patients in Taiwan were characterized for serotypes and mating types. Slide agglutination test was performed with 8 factor-specific sera (Iatron Company, Japan) to determine the serotypes. Wheat bran agar (WBA) and malt extract agar (MEA, Wickerham) media were used for the mating tests. Twenty of the isolates were of serotype A, and one was serotype B. Except for 2 strains of serotype A, all of the serotype A strains mated withFilobasidiella neoformans var.neoformans, mating type a. The only serotype B strain mated withF. neoformans var.bacillispora mating type a in MEA medium. These data revealed the low prevalence (1/21; 4.8%) ofC. neoformans var.gattii in Taiwan, a subtropically located island.     

Characterization ofYersinia enterocolitica sensu stricto

The speciesYersinia enterocolitica is definedsensu stricto on the bases of biochemical and other phenotypic characteristics. Biochemically,Y. enterocolitica contains five major biotypes: 1 through 4 of Niléhn and of Wauters, and the trehalose-negative, metabolically inactive, socalled hare strains in biotype 5 of Niléhn and of Wauters, and biochemically atypical strains, including urease-negative, Simmons' citrate-positive, and lactose-and raffinose-positive strains.Y. enterocolitica sensu stricto was distinguishable from the newly described speciesYersinia kristensenii by sucrose and Voges-Proskauer reactions (negative inY. kristensenii). These species were previously separated by DNA relatedness.Y. enterocolitica was also separable biochemically and by DNA relatedness from the two newly proposed rhamnose-positive species,Yersinia intermedia andYersinia frederiksenii. Strain 161(=CIP 80-27=ATCC 9610) is proposed as the neotype forY. enterocolitica.     

Kinetics of infection induced byYersinia

Yersinia enterocolitica of different serotypes andY. intermedia, Y. frederiksenii, andY. kristensenii, in a total of nine strains, were inoculated intragastrically and intravenously into Swiss mice. The animals were observed daily to check for clinical alterations. Groups of five were killed intermittently at 6-h, and 3-, 6-, 10-, 15-,and 21-day periods or more after the inoculation; possible macroscopic alterations of the organs and tissues were checked. Development of infection at these periods was followed by performing viable bacterial counts on homogenates of selected tissues and the kinetics of infection was established. Clinical and pathologic alterations occurred only in the animals inoculated with the human strains ofY. enterocolitica 0:3 and 0:8, independent of the route of infection. After intragastric inoculation, theY. enterocolitica strains considered to be adapted to man were isolated from all organs and tissues, with the exception of the blood, from which only serotype 0:8 was isolated; otherYersinia strains were found only in the cecal content. After intravenous challenge, all the strains infected the organs and tissues at different times and in varied intensity, with exception of Peyer's patches and mesenteric lymph nodes, which were not infected by all theYersinia strains.     

Genotyping of Human and Porcine Yersinia enterocolitica, Yersinia intermedia, and Yersinia bercovieri Strains from Switzerland by Amplified Fragment Length Polymorphism Analysis

In this study, 231 strains of Yersinia enterocolitica, 25 strains of Y. intermedia, and 10 strains of Y. bercovieri from human and porcine sources (including reference strains) were analyzed using amplified fragment length polymorphism (AFLP), a whole-genome fingerprinting method for subtyping bacterial isolates. AFLP typing distinguished the different Yersinia species examined. Representatives of Y. enterocolitica biotypes 1A, 1B, 2, 3, and 4 belonged to biotype-related AFLP clusters and were clearly distinguished from each other. Y. enterocolitica biotypes 2, 3, and 4 appeared to be more closely related to each other (83% similarity) than to biotypes 1A (11%) and 1B (47%). Biotype 1A strains exhibited the greatest genetic heterogeneity of the biotypes studied. The biotype 1A genotypes were distributed among four major clusters, each containing strains from both human and porcine sources, confirming the zoonotic potential of this organism. The AFLP technique is a valuable genotypic method for identification and typing of Y. enterocolitica and other Yersinia spp.     

Yersinia enterocolitica Isolates from Wild Boars Hunted in Lower Saxony,Germany

Yersiniosis is strongly associated with the consumption of pork contaminated with enteropathogenic Yersinia enterocolitica, which is harbored by domestic pigs without showing clinical signs of disease. In contrast to data on Y. enterocolitica isolated from conventionally reared swine, investigations into the occurrence of Y. enterocolitica in wild boars in Germany are rare. The objectives of the study were to get knowledge about these bacteria and their occurrence in wild boars hunted in northern Germany by isolation of the bacteria from the tonsils, identification of the bioserotypes, determination of selected virulence factors, macrorestriction analysis, multilocus sequence typing (MLST), and testing of antimicrobial susceptibility. Altogether, tonsils from 17.1% of 111 tested wild boars were positive for Y. enterocolitica by culture methods. All but two isolates belonged to biotype (BT) 1A, with the majority of isolates bearing a ystB nucleotide sequence which was revealed to have 85% identity to internal regions of Y. enterocolitica heat-stable enterotoxin type B genes. The remaining Y. enterocolitica isolates were identified to be BT 1B and did not carry the virulence plasmid. However, two BT 1A isolates carried the ail gene. Macrorestriction analysis and results from MLST showed a high degree of genetic diversity of the isolates, although the region where the samples were taken was restricted to Lower Saxony, Germany, and wild boars were shot during one hunting season. In conclusion, most Y. enterocolitica isolates from wild boars investigated in this study belonged to biotype 1A. Enteropathogenic Y. enterocolitica bioserotypes 4/O:3 and 2/O:9, usually harbored by commercially raised pigs in Europe, could not be identified.     

The influence ofYersinia enterocolitica on the recovery of cyclophosphamide-damaged bone marrow in mice

Mice were treated with cyclophosphamide (CY) to induce reversible bone marrow damage and peripheral leukopenia and were infected intraperitoneally withYersinia enterocolitica to study comparatively the effects of both virulent and avirulent isogenic strains on marrow recovery. As a general rule, virulentY. enterocolitica increased the CY-induced damage, whereas the avirulent isogenic derivative improved the marrow recovery. In some instances, infection with virulentY. enterocolitica of CY-pretreated mice resulted in the death of animals.     

Addition of new serogroups and improvement of the antigenic designs ofYersinia pseudotuberculosis

Eight strains ofYersinia pseudotuberculosis, untypable for any established serogroups, were studied. They were isolated from humans (three), dogs (two), cat (one), and rats (two), and the general characteristics of the strains were identical with those ofY. pseudotuberculosis. On the basis of results of O-agglutinin absorption tests, these strains were classified into three new serogroups. The new serogroups proposed were serogroups IIC, VII, and VIII, and a new O-antigenic scheme ofY. pseudotuberculosis was designed.     

Spontaneous binding ofYersinia enterocolitica to murine leukocytes

Twelve strains ofYersinia enterocolitica were examined for their ability to bind spontaneously to murine leukocytes. Each of eight HeLa cell invasive strains exhibited nonselective binding to peritoneal leukocytes, lymph node leukocytes, and thymocytes, whereas four noninvasive strains lacked binding properties. Like the HeLa cell invasion, the binding ofY. enterocolitica to leukocytes was much less efficient for bacteria grown at 37°C than for bacteria grown at 22°C. The binding properties were not influenced by the virulence plasmid that codes for Vwa⁺ phenotype. This leukocyte binding test is proposed as a simple assay for invasive properties ofY. enterocolitica.     

Rôle of pigeons in the spread of salmonellosis: Incidence of different types ofSalmonella typhi-murium var.Copenhagen in pigeons,man, and other animals

The strains ofSalmonella typhimurium var.Copenhagen isolated from pigeons were characterized by biochemical tests and by phage typing. They were rarely found to cause disease in man. These strains were also rarely found in animals through which they might infect man. It can be concluded that the rôle of pigeons in the spread of salmonellosis in man is small.All strains of phage type 85, and the few strains of phage type 47, biotype 6 were var.Copenhagen. These strains were often found in man. If found in animals, they were practically always isolated from pigs.     

Yersinia enterocolitica O9 as a Possible Barrier Against Y. pestis in Natural Plague Foci in Ningxia, China

A survey of Yersinia spp, as related to plague control, was made in Haiyuan of Ganning loess plateau plague focus, Yanchi of Inner Mongolia plateau plague focus, and Yinchuan city, as a control area, in Ningxia, China. In Haiyuan, where the main plague reservoir was Mongolian ground squirrel (Citellus alaschanicus) living in the prairie, Y. enterocolitica O9 was frequently isolated from pigs, dogs, rodents living in and around houses, but only rarely from hare and Mongolian ground squirrel. In Yanchi, where the main plague reservoir was Mongolian gerbil (Meriones unguiculatus) living in the prairie and Y. pestis, which was isolated from rodents up to 1991, Y. enterocolitica O9 was sometimes isolated from pigs and rodents. In all areas, some strains of Y. enterocolitica O3 and Y. pseudotuberculosis serotypes 3 and 4b were also isolated from pigs, dogs, and from rodents. We propose that an epidemiological link exists between the prevalence of Y. pestis and Y. enterocolitica O9 in domestic and rodents living in these areas in China. The residential area in Haiyuan may be protected against Y. pestis by the domestic animals and rodents which acquired cross-protection against Y. pestis by infection with Y. enterocolitica O9, but this is not the case in the Yanchi district. Received: 14 February 2000 / Accepted: 17 July 2000     

Enterotoxin production at refrigeration temperature byYersinia enterocolitica andYersinia enterocolitica-like bacteria

Enterotoxin production after cultivation for 7 days in a refrigerator (3–6°C) was indicated for 4 of 20 strains ofYersinia enterocolitica andY. enterocolitica-like bacteria, by use of the infant mouse assay. These four strains were isolated from wild-living small mammals and water. Three of these isolates (Y. kristensenii, serogroups 11 and 28) were enterotoxigenic at 22 and 37°C as well as at refrigeration temperature. The remaining strain (Y. enterocolitica sensu stricto, serogroup 6) produced enterotoxin only at refrigeration temperature and at 22°C. The results indicate thatY. enterocolitica andY. enterocolitica-like bacteria may be capable of causing food intoxication after food storage at refrigeration temperature. A potential clinical significance of theY. enterocolitica enterotoxin in cold-blooded animals such as fish is suggested.     

Enterotoxigenic porcine and human isolates ofYersinia enterocolitica in Sweden

The production of heat-stable and heat-labile enterotoxins byYersinia enterocolitica was studied in 69 strains from healthy swine and in 24 strains from humans with acute diarrhea. All of the human strains were of serotype O3, and 20 (83%) of them produced heat-stable enterotoxin detectable in the infant mouse assay. All were negative in the Chinese Hamster Ovary (CHO) cell test for detection of heat-labile enterotoxin. Of the 69 porcine strains, which were of twelve serotypes plus 9 nontypable strains, 26 (38%) gave a positive infant mouse test. Of the porcine isolates of serotype O3, 42% were enterotoxigenic. A high incidence of enterotoxigenicity was also apparent among six other serotypes (53%). All porcine strains were negative in the CHO cell test. However, of seven culture supernatants from these porcine strains, three gave positive reactions in rabbit skin permeability tests, two of which were also positive in rabbit loop tests. Heat treatment of the supernatants abolished the reactivity in both tests. It is concluded that production of a heatstable enterotoxin is fairly common in porcine and human strains ofY. enterocolitica of serotype O3 in Sweden.     

Survey on the Incidence of Yersinia enterocolitica Infection in Canada

Data pertaining to 278 cultures of Yersinia enterocolitica isolated in Canada are summarized in this paper. Of this amount, 256 were isolated from humans, whereas 22 were of nonhuman sources. The typing of these cultures is presented together with their geographical location. Y. enterocolitica serotype O:3 biotype 4, phage type 9b, was practically the only serotype present in the Province of Quebec. This serotype O:3 was also predominant in Ontario, followed by serotypes O:5,27 and O:6,30; other serotypes were seldom isolated. In the central and western areas of Canada, Y. enterocolitica was occasionally isolated; the strains were indole-positive, serotypes O:5,27, O:8, and O:4,32.     

Application of fluorescent amplified fragment length polymorphism for comparison of human and animal isolates of Yersinia enterocolitica

An amplified fragment length polymorphism (AFLP) method, developed to genotype Yersinia enterocolitica, has been used to investigate 70 representative strains isolated from humans, pigs, sheep, and cattle in the United Kingdom. AFLP primarily distinguished Y. enterocolitica strains according to their biotype, with strains dividing into two distinct clusters: cluster A, comprising largely the putatively pathogenic biotypes (BT2 to -4), and cluster B, comprising the putatively nonpathogenic biotype 1A strains and a single BT1B isolate. Within these two clusters, subclusters formed largely on the basis of serotype. However, AFLP profiles also allowed differentiation of strains within these serotype-related subclusters, indicating the high discriminatory power of the technique for Y. enterocolitica. Investigation of the relationship between strain AFLP profile and host confirmed that pigs are, and provides further proof that sheep may be, potential sources of human infection with putatively pathogenic strains. However, the results suggest that some strains causing human disease do not come from veterinary sources identifiable at this time. The distribution of some BT1A isolates within cluster A raises questions about the relationship between virulence potential and biotype.