virF-positive Yersinia pseudotuberculosis and Yersinia enterocolitica found in migratory birds in Sweden

During spring and autumn migrations, 468 fecal samples from 57 different species of migratory birds were collected in Sweden. In total, Yersinia spp. were isolated from 12.8% of collected samples. The most commonly found species was Yersinia enterocolitica, which was isolated from 5.6% of all collected samples, followed by Y. intermedia (3.8%), Y. frederiksenii (3.0%), Y. kristensenii (0.9%), Y. pseudotuberculosis (0.6%), and Y. rohdei (0.4%). The pathogenic, virF-positive Y. pseudotuberculosis strains were recovered from three thrushes. These strains belonged to the same bioserotype, 1/O:2, but had two different profiles as determined by pulsed-field gel electrophoresis with NotI and SpeI enzymes. In addition, 10 Y. enterocolitica strains, all from barnacle geese, belonged to bioserotype 3/O:3, which is associated with human disease. Two of the strains were pathogenic, carrying the virF gene on their plasmids. All pathogenic Y. pseudotuberculosis and Y. enterocolitica strains were recovered during the spring, and as the birds were caught during active migration they likely became infected at an earlier stage of the migration, thus potentially transporting these bacterial pathogens over long geographical distances.     

Identification and characterization of pathogenic Yersinia enterocolitica isolates by PCR and pulsed-field gel electrophoresis

Approximately 550 to 600 yersiniosis patients are reported annually in Sweden. Although pigs are thought to be the main reservoir of food-borne pathogenic Yersinia enterocolitica, the role of pork meat as a vehicle for transmission to humans is still unclear. Pork meat collected from refrigerators and local shops frequented by yersiniosis patients (n=48) were examined for the presence of pathogenic Yersinia spp. A combined culture and PCR method was used for detection, and a multiplex PCR was developed and evaluated as a tool for efficient identification of pathogenic food and patient isolates. The results obtained with the multiplex PCR were compared to phenotypic test results and confirmed by pulsed-field gel electrophoresis (PFGE). In all, 118 pork products (91 raw and 27 ready-to-eat) were collected. Pathogenic Yersinia spp. were detected by PCR in 10% (9 of 91) of the raw pork samples (loin of pork, fillet of pork, pork chop, ham, and minced meat) but in none of the ready-to-eat products. Isolates of Y. enterocolitica bioserotype 4/O:3 were recovered from six of the PCR-positive raw pork samples; all harbored the virulence plasmid. All isolates were recovered from food collected in shops and, thus, none were from the patients' home. When subjected to PFGE, the six isolates displayed four different NotI profiles. The same four NotI profiles were also present among isolates recovered from the yersiniosis patients. The application of a multiplex PCR was shown to be an efficient tool for identification of pathogenic Y. enterocolitica isolates in naturally contaminated raw pork.     

Efficient subtyping of Yersinia enterocolitica bioserotype 4/O:3 with pulsed-field gel electrophoresis

The purpose of this study was to evaluate the efficiency of pulsed-field gel electrophoresis (PFGE) for differentiation of isolates of Yersinia enterocolitica bioserotype 4/O:3 recovered from pig tongues at retail level in the Helsinki area during October and November of 1996. NotI generated 15 different PFGE patterns for 128 isolates. The discriminatory index did not exceed 74% due to the presence of two predominant PFGE-types, NA1 (58/128) and NB1 (25/128). After preliminary investigations with 35 enzymes, ApaI, XbaI, XhoI and SpeI were chosen for further characterization of the isolates. The discriminatory index increased from 74% to 93% and the number of different genotypes from 15 to 30 when isolates with the same PFGE pattern with NotI were further characterized with ApaI and XhoI, indicating that PFGE can be an efficient technique for characterization of bioserotype 4/O:3.     

Comparison of the biotypes of Yersinia enterocolitica isolated from pigs, cattle and sheep at slaughter and from humans with yersiniosis in Great Britain during 1999-2000

AIMS: To investigate the relationship between livestock carriage of Yersinia enterocolitica and human disease. The biotypes/serotypes of strains recovered from the faeces of pigs, cattle and sheep at slaughter during a national survey in Great Britain in 1999-2000, were compared with those of strains isolated from human cases of yersiniosis during the same period. METHODS AND RESULTS: The faecal carriage of Y. enterocolitica by cattle, sheep and pigs at slaughter was 6.3, 10.7 and 26.1%, respectively. Yersinia enterocolitica biotype (BT) 1a was the most frequently isolated biotype from livestock (58%) and was the predominant biotype (53%) isolated from human cases over the same period. The main recognized pathogenic Y. enterocolitica biotype isolated from livestock was BT3 (O:5,27) (35% of sheep, 22% of pigs and 4% of cattle) but this biotype was not detected in any of the human isolates investigated. The major pathogenic biotypes of strains isolated from humans were BT3 (O:9) (24%) and BT4 (O:3) (19%) whereas of the veterinary isolates investigated, only pigs (11%) carried BT3 (O:9) strains. CONCLUSIONS: Because of significant overlaps in phenotypes of the veterinary and human strains it is not possible to comment on the correlation between host and pathogenicity, especially of biotype 1a. SIGNIFICANCE AND IMPACT OF THE STUDY: The data suggest that further investigations using methods with greater discriminatory power are required. However the data also suggests that pigs may be the primary reservoir for human pathogenic Y. enterocolitica infection.     

云南省新平县家鼠鼠疫疫原地小肠结肠炎耶尔森菌的调查分析

目的了解新平县家鼠鼠疫疫源地小肠结肠炎耶尔森菌的分布及病原学特征。方法采集家鼠盲肠、舌头和猪粪便、咽喉粘液以及腹泻患者粪便标本进行小肠结肠炎耶尔森菌的检测与分析。结果检测家鼠盲肠、鼠舌头、猪粪便、猪咽喉粘液物、腹泻患者粪便的标本数分别为722、722、467、237和107份,共分离到61株小肠结肠炎耶尔森菌,总检出率为2. 71%,5种标本的检出率分别为2. 63%、1. 39%、3. 85%、2.53%和7. 48%,差异有统计学意义（x^2= 16. 422,P = 0. 003）;分离株包括致病株10株、非致病株51株,有1A、2、3三种生物型和0：3、0：5、0：8等多种血清型,以及六种毒力基因型。猪、鼠、腹泻患者标本检出致病菌株数分别为9、1、0株。结论新平县家鼠鼠疫自然疫源地猪、鼠、腹泻患者是小肠结肠炎耶尔森菌的重要宿主,分离菌株具有遗传多样性,猪、鼠是小肠结肠炎耶尔森菌病的主要传染源。     

High prevalence of Yersinia enterocolitica 4:O3 on pig offal in southern Germany: a slaughtering technique problem

Prevalence and contamination routes of pathogenic Yersinia enterocolitica were studied in Southern Germany. Tonsil and faeces samples of 50 fattening pigs, 140 offal samples and 120 minced meat samples were examined. Pig and offal samples were collected from a slaughterhouse approved by the European Union, and minced meat samples from two large meat factories. Yersinia enterocolitica was isolated using direct plating, overnight enrichment and selective enrichment in MRB and ITC broth. The isolates were bio- and serotyped, and pathogenicity was studied using two plasmid-encoded virulence markers: calcium dependence and Congo red absorption. The genotypes were studied with pulsed-field gel electrophoresis using NotI enzyme. Prevalence of pathogenic Y. enterocolitica 4:O3 was 60% and 10% in tonsils and faeces of fattening pigs, respectively. Besides tonsils, prevalence of pathogenic Y. enterocolitica 4:O3 was also high in other pluck set samples, including tongues, lungs, hearts, diaphragms and livers. However, the highest isolation rate was obtained from the tonsils. Kidneys, which were not attached to the pluck set and did not hang together with tonsils on the rack, had the lowest isolation rate. Yersinia enterocolitica 4:O3 was isolated from 12% of minced meat samples. A total of 25 NotI profiles were obtained from porcine samples. The most common genotype, NBI, found in tonsils was also the most common type recovered from offal and minced meat samples. The high contamination rate of tonsils, and the indistinguishable NotI profiles obtained from tonsils and offal indicate that the tonsils contaminate offal when they are removed and hung on the rack together. When the head, with the tonsils and tongue, is not removed prior to evisceration and is not handled and inspected separately, it is difficult to control the spread of Y. enterocolitica 4:O3 from tonsils to the carcass, and subsequently, to meat.     

Laboratory investigation of virulence among strains of Yersinia enterocolitica and related species isolated from pigs and pork products.

Eighty strains of Yersinia enterocolitica and related species isolated from slaughtered pigs and pork products were tested for possession of virulence-associated phenotypes by employing 12 in vivo and in vitro assays. The isolates could be broadly divided into two groups: (i) strains belonging to pathogenic bioserotypes of Y. enterocolitica that displayed virulence-associated characteristics in most or all assays and (ii) strains belonging to Y. enterocolitica biotype 1A and to related species that were largely negative in these assays. No individual test was found as a single reliable measure of virulence. All strains belonging to Y. enterocolitica serotype O:1,2,3 were pyrazinamidase positive (indicates avirulence) and autoagglutination negative but were positive in all other virulence assays. Salt aggregation was found to be a better indicator of virulence than latex particle agglutination, both of which measure surface hydrophobicity. Overall, tissue culture cell invasion provided the best selection of a subpopulation of yersiniae that are potentially virulent. However, crystal violet and Congo red binding assays among others provided good prediction of virulence at the time of testing. Our results provide further evidence that swine may constitute an important reservoir of human pathogenic strains.     

Multiple-locus variable-number tandem-repeat analysis of pathogenic Yersinia enterocolitica in China

The predominant bioserotypes of pathogenic Yersinia enterocolitica in China are 2/O: 9 and 3/O: 3; no pathogenic O: 8 strains have been found to date. Multiple-Locus Variable-Number Tandem-Repeat Analysis (MLVA) based on seven loci was able to distinguish 104 genotypes among 218 pathogenic Y. enterocolitica isolates in China and from abroad, showing a high resolution. The major pathogenic serogroups in China, O: 3 and O: 9, were divided into two clusters based on MLVA genotyping. The different distribution of Y. enterocolitica MLVA genotypes maybe due to the recent dissemination of specific clones of 2/O: 9 and 3/O: 3 strains in China. MLVA was a helpful tool for bacterial pathogen surveillance and investigation of pathogenic Y. enterocolitica outbreaks.     

The application of PCR fingerprinting to the differentiation of Yersinia enterocolitica strains isolated from humans and pigs

The distribution of different genotypes of Yersinia enterocolitica strains recovered from humans and from healthy pigs was investigated using PCR fingerprinting. The thirty six strains of Y. enterocolitica from humans, thirty five strains from pigs and Y. enterocolitica ATCC 9610 strain were included in this study. The tested strains of Y. enterocolitica belonged to O3 and O9 serogroups. The PCR fingerprinting using EAE5 primer (5' CTT AAT CTC AGT AAT GCT GGC CTT GG) made it possible to form five groups among the tested Y. enterocolitica strains. Two groups were very numerously represented by the tested strains. The thirty of Y. enterocolitica O3 strains from humans (thirty one of tested) and eighteen of Y. enterocolitica O3 strains from pigs (twenty of tested) belonged to one group. This group also included Y. enterocolitica ATCC9610 strain and four Y. enterocolitica O9 strains from pigs. All investigated Y. enterocolitica O9 strains from humans and the majority of Y. enterocolitica O9 strains isolated from pigs created a second, numerous group. The third genotype was created by two strains O9 from pigs, and the remaining two strains, isolated from pigs, belonging to O3 and O9 serogroups showed different binding patterns revealed by gel electrophoresis and created two other genotypes. The tested Y. enterocolitica strains which were isolated from humans formed only two groups but Y. enterocolitica strains isolated from pigs were found in five groups but such as the Y. enterocolitica strains from humans, the majority of strains from pigs were in first and second group. The Y. enterocolitica O3 strains regardless of their origin mostly represented the same PCR fingerprinting profile. The tested Y. enterocolitica O9 strains were more genetically diverse and represented four PCR fingerprinting profiles.     

virF-Positive Yersinia pseudotuberculosis and Yersinia enterocolitica Found in Migratory Birds in Sweden

During spring and autumn migrations, 468 fecal samples from 57 different species of migratory birds were collected in Sweden. In total, Yersinia spp. were isolated from 12.8% of collected samples. The most commonly found species was Yersinia enterocolitica, which was isolated from 5.6% of all collected samples, followed by Y. intermedia (3.8%), Y. frederiksenii (3.0%), Y. kristensenii (0.9%), Y. pseudotuberculosis (0.6%), and Y. rohdei (0.4%). The pathogenic, virF-positive Y. pseudotuberculosis strains were recovered from three thrushes. These strains belonged to the same bioserotype, 1/O:2, but had two different profiles as determined by pulsed-field gel electrophoresis with NotI and SpeI enzymes. In addition, 10 Y. enterocolitica strains, all from barnacle geese, belonged to bioserotype 3/O:3, which is associated with human disease. Two of the strains were pathogenic, carrying the virF gene on their plasmids. All pathogenic Y. pseudotuberculosis and Y. enterocolitica strains were recovered during the spring, and as the birds were caught during active migration they likely became infected at an earlier stage of the migration, thus potentially transporting these bacterial pathogens over long geographical distances.     

Identification and Characterization of Pathogenic Yersinia enterocolitica Isolates by PCR and Pulsed-Field Gel Electrophoresis

Approximately 550 to 600 yersiniosis patients are reported annually in Sweden. Although pigs are thought to be the main reservoir of food-borne pathogenic Yersinia enterocolitica, the role of pork meat as a vehicle for transmission to humans is still unclear. Pork meat collected from refrigerators and local shops frequented by yersiniosis patients (n = 48) were examined for the presence of pathogenic Yersinia spp. A combined culture and PCR method was used for detection, and a multiplex PCR was developed and evaluated as a tool for efficient identification of pathogenic food and patient isolates. The results obtained with the multiplex PCR were compared to phenotypic test results and confirmed by pulsed-field gel electrophoresis (PFGE). In all, 118 pork products (91 raw and 27 ready-to-eat) were collected. Pathogenic Yersinia spp. were detected by PCR in 10% (9 of 91) of the raw pork samples (loin of pork, fillet of pork, pork chop, ham, and minced meat) but in none of the ready-to-eat products. Isolates of Y. enterocolitica bioserotype 4/O:3 were recovered from six of the PCR-positive raw pork samples; all harbored the virulence plasmid. All isolates were recovered from food collected in shops and, thus, none were from the patients' home. When subjected to PFGE, the six isolates displayed four different NotI profiles. The same four NotI profiles were also present among isolates recovered from the yersiniosis patients. The application of a multiplex PCR was shown to be an efficient tool for identification of pathogenic Y. enterocolitica isolates in naturally contaminated raw pork.     

Investigation of virulence genes in clinical isolates of Yersinia enterocolitica

In this study, we aimed to investigate the distribution of virulence genes in clinical isolates of pathogenic Yersinia enterocolitica. Two thousand six hundred stool samples were collected from 2600 patients with diarrhea, and were tested using the culture method and real-time PCR. Then, all isolates of pathogenic Y. enterocolitica cultured from the culture method were examined for virulence genes (inv, ail, ystA, ystB, ystC, yadA, virF) by PCR and for the presence of plasmid by four phenotypic tests. As a result, 160 pathogenic strains were successfully detected by the culture method, including bio/serotype 1A/unknown (4), 1B/unknown (8), 2/O:9 (39), 2/unknown (7), 3/O:3 (22), 3/unknown (6), 4/O:3 (55), 4/unknown (10) and 5/unknown (9). The positive rate of virulence genes tested in 160 isolates was inv (100%), ail (94%), ystA (93%), ystB (7.5%), ystC (5%), yadA (89%) and virF (82%) while the phenotypic test included autoagglutination (87%), binding of crystal violet (89%), calcium-dependent growth (74%) and Congo red absorption (78%), respectively. Finally, we found that not all pathogenic Y. enterocolitica necessarily carry all traditional virulence genes in both chromosomes and plasmids to cause illness. Perhaps, some of them, lacking some traditional virulence genes, contain other unknown virulence markers that interact with each other and play an important role in the diverse pathogenesis of pathogenic Y. enterocolitica.     

Digoxigenin-labelled inv- and ail-probes for the detection and identification of pathogenic Yersinia enterocolitica in clinical specimens and naturally contaminated pig samples

A non-radioactive colony hybridization method was developed for the rapid detection of Yersinia enterocolitica in primary isolates and for differentiation between pathogenic and non-pathogenic strains. The method is based on, respectively, the presence of the inv -locus in all Yersinia spp. and the presence of the ail -gene in pathogenic Y. enterocolitica only. Hybridization results with ail -probes of 132 strains of Y. enterocolitica were in good agreement with pathogenicity phenotypes as indicated by a tissue culture invasion (TCI) assay and by serotyping. All TCI⁺ strains and only two TCI^- strains were positive by hybridization with ail. Hybridization results with inv - or ail -probes of 150 primary isolates of human, animal or slaughterhouse origin were compared with those of conventional methods to detect and identify Y. enterocolitica. All samples that were positive for Yersinia spp. by cultivation (four of 66) or were positive for pathogenic Y. enterocolitica by cultivation and serotyping (six of 84) were also positive by hybridization with, respectively, the inv - or ail -probe. In three slaughterhouse swab samples, in which Yersinia spp. were not detected by cultivation (2%), strong positive hybridization signals were obtained with the inv - and/or ail -probe. Four other swab samples which were negative by cultivation produced weak positive signals by hybridization with inv - and/or ail - probes. These results indicate that the method can be used for (1) the identification of pathogenic Y. enterocolitica isolates and (2) the detection of Yersinia spp. in primary isolates of naturally contaminated samples.     

Transmission of Yersinia enterocolitica 4/O:3 to pets via contaminated pork

AIMS: This study was conducted to investigate sources of Yersinia enterocolitica 4/O:3 infections in dogs and cats. METHODS AND RESULTS: Transmission of Y. enterocolitica 4/O:3 to pets via contaminated pork was studied using PFGE with NotI, ApaI and XhoI enzymes. A total of 132 isolates, of which 16 were from cat and dog faeces and 116 from raw pork samples, were recovered in Finland during 1998-99. Cat 1, whose diet consisted mostly of raw pig hearts and kidneys, excreted Y. enterocolitica 4/O:3 of genotype G4. This predominant genotype was also found in isolates recovered from the pig heart, liver, kidney, tongue and ear, and minced pork samples. Dog 2, which was fed raw minced pork, excreted Y. enterocolitica of genotype G13. This genotype was also identified in isolates recovered from the pig heart, kidney and tongue, and minced pork samples. CONCLUSION: These results show that raw pork can be an important source of Yersinia enterocolitica 4/O:3 infections in dogs and cats. Significance and Impact of the Study: Raw pork should not be given to pets.     

Molecular characterization of Yersinia enterocolitica by pulsed-field gel electrophoresis and hybridization of DNA fragments to ail and pYV probes.

Sixty strains of Yersinia enterocolitica from five serogroups (O:3; O:9; O:8; O:5; and O:5,27) and eight non-Y. enterocolitica strains, recovered from diverse sources (humans, animals, food, and the environment) in Europe, Argentina, and the United States, were examined by the pulsed-field gel electrophoresis (PFGE) technique of contour clamped homogeneous electric field electrophoresis (CHEF) by using NotI and XbaI as restriction enzymes. NotI and XbaI generated 36 and 33 restriction endonuclease digestion profiles (REDP), respectively. By combining the results of both enzymes, 42 unique genomic groups were differentiated. DNA fragments were transferred to nylon membranes and hybridized with digoxigenin-labelled oligonucleotide probes to the ail gene and virulence plasmid to determine hybridization patterns and the potential virulence of the strains. The strains were tested for the presence of the plasmid by PFGE-CHEF and phenotypic characteristics encoded for by the virulence plasmid. Thirty of the 60 Y. enterocolitica strains tested harbored the virulence plasmid. The specificity of the ail and pYV probes was 100% when tested with 68 Yersinia strains and 19 different non-Yersinia strains. Sixteen selected Y. enterocolitica strains were tested for their virulence by lethality in iron- and desferrioxamine-sensitized mice. No correlation between REDP and the virulence of the strains was observed. The observed REDP and the hybridization patterns were very homogeneous within a serogroup and independent of the source of isolation. In addition, PFGE-CHEF was shown to be valuable in identifying and confirming serogroups. Principal component analysis of Dice similarity indices from REDP was an excellent tool for determining genetic relatedness among strains.     

Isolation of Yersinia enterocolitica and related species from foods in France

The occurrence of Yersinia enterocolitica and related species (Y. intermedia, Y. frederiksenii, Y. kristensenii) in foods from France was investigated by using different enrichment procedures. Initially, seven procedures were evaluated with pork products. These methods included a cold preenrichment in yeast extract-rose bengal broth or in phosphate-sorbitol-bile medium, followed by selective enrichment either in Pastone-sucrose-Tris-azide broth, in modified Rappaport broth, or in bile-oxalate-sorbose broth, and then isolation onto Hektoen or cefsulodin-irgasan-novobiocin agar with or without KOH pretreatment. The best enrichment procedure in terms of percentage of positive samples obtained within the shortest time was the combination of phosphate-sorbitol-bile and bile-oxalate-sorbose with alkali treatment before isolation onto cefsulodin-irgasan-novobiocin agar. This system was then used to analyze foods other than pork. An average contamination rate of 33.5% was observed for 666 samples analyzed; pork products were by far the most contaminated, especially the so-called tartinette (96.8% of positive samples) which contained up to five different strains of Yersinia spp. Environmental serogroups of Y. enterocolitica O:5, O:39,41, O:6, and O:7,8 were predominant, but no isolate of either human pathogenic type (O:3 or O:9) was obtained.     

New enrichment method for isolation of pathogenic Yersinia enterocolitica serogroup O:3 from pork

A new enrichment medium for the recovery of pathogenic Yersinia enterocolitica serogroup O:3 from naturally infected meat products based on three selective agents, Irgasan, ticarcillin, and potassium chlorate (ITC), was compared with several other one- or two-step enrichments. Y. enterocolitica serogroup O:3 was recovered from 96.5% of 29 pork tongues, 24% of 50 ground pork samples, 16% of 25 masseter muscle samples, and 61% of tonsils. ITC was by far the most sensitive method for the recovery of Y. enterocolitica O:3, especially from ground meat and masseter muscles, while cold and two-step enrichments yielded better results for nonpathogenic strains. Plating of ITC enrichments onto SS-deoxycholate-calcium agar gave overall better results than plating onto cefsulodin-Irgasan-novobiocin agar for serogroup O:3.     

Isolation of Yersinia enterocolitica and related species from foods in France.

The occurrence of Yersinia enterocolitica and related species (Y. intermedia, Y. frederiksenii, Y. kristensenii) in foods from France was investigated by using different enrichment procedures. Initially, seven procedures were evaluated with pork products. These methods included a cold preenrichment in yeast extract-rose bengal broth or in phosphate-sorbitol-bile medium, followed by selective enrichment either in Pastone-sucrose-Tris-azide broth, in modified Rappaport broth, or in bile-oxalate-sorbose broth, and then isolation onto Hektoen or cefsulodin-irgasan-novobiocin agar with or without KOH pretreatment. The best enrichment procedure in terms of percentage of positive samples obtained within the shortest time was the combination of phosphate-sorbitol-bile and bile-oxalate-sorbose with alkali treatment before isolation onto cefsulodin-irgasan-novobiocin agar. This system was then used to analyze foods other than pork. An average contamination rate of 33.5% was observed for 666 samples analyzed; pork products were by far the most contaminated, especially the so-called tartinette (96.8% of positive samples) which contained up to five different strains of Yersinia spp. Environmental serogroups of Y. enterocolitica O:5, O:39,41, O:6, and O:7,8 were predominant, but no isolate of either human pathogenic type (O:3 or O:9) was obtained.     

New enrichment method for isolation of pathogenic Yersinia enterocolitica serogroup O:3 from pork.

A new enrichment medium for the recovery of pathogenic Yersinia enterocolitica serogroup O:3 from naturally infected meat products based on three selective agents, Irgasan, ticarcillin, and potassium chlorate (ITC), was compared with several other one- or two-step enrichments. Y. enterocolitica serogroup O:3 was recovered from 96.5% of 29 pork tongues, 24% of 50 ground pork samples, 16% of 25 masseter muscle samples, and 61% of tonsils. ITC was by far the most sensitive method for the recovery of Y. enterocolitica O:3, especially from ground meat and masseter muscles, while cold and two-step enrichments yielded better results for nonpathogenic strains. Plating of ITC enrichments onto SS-deoxycholate-calcium agar gave overall better results than plating onto cefsulodin-Irgasan-novobiocin agar for serogroup O:3.     

A synthetic oligonucleotide probe and a cloned polynucleotide probe based on the yopA gene for detection and enumeration of virulent Yersinia enterocolitica

We compared a synthetically produced 19-mer oligonucleotide probe with a polynucleotide probe consisting of a cloned fragment of the virulence gene yopA for their relative efficiencies in identification and enumeration of virulent Yersinia enterocolitica. The probes were used in DNA-DNA colony hybridization assays to differentiate 70 Yersinia strains with known plasmid profiles. All 19 strains harboring the 40- to 50-megadalton virulence plasmid were positive in the hybridization assay, whereas their isogenic derivatives lacking this plasmid were negative. Both probes correctly identified plasmid-bearing variants of Y. enterocolitica serogroups O:3, O:5,27, O:8, O:9, O:13, and O:21 from three continents. In contrast, none of the probes hybridized with DNA from 32 environmental yersiniae belonging to 26 serogroups not associated with disease. Colony hybridization was used to detect and enumerate virulent Y. enterocolitica in three artificially contaminated food samples. Despite a large background of indigenous bacteria (3 x 10(4) CFU), the efficiency of enumeration ranged from 33 to 82%. The use of nylon filters did not impair the growth of virulent yersiniae. Both probes showed a perfect concordance in their specific differentiation and enumeration of virulent Y. enterocolitica. DNA colony hybridization with these two probes permitted rapid and reliable identification of all common pathogenic serogroups without the need for enrichment or esoteric identification protocols.