Yersinia pseudotuberculosis serotype O:1 infection in a captive Seba's short tailed-fruit bat (Carollia perspicillata) colony in Switzerland.

BACKGROUND: Between February and April 2016, a slight increase in mortality was observed in a colony consisting of 400 captive Seba's short-tailed bats (Carollia perspicillata). These animals cohabited with other nocturnal animal species in a dome of a private zoo in Switzerland. RESULTS: Gross and histological analysis of two (14.3%) out of the 13 animals submitted for necropsy within this period revealed a necrosuppurative pneumonia, hepatitis, splenitis, enterocolitis, and endometritis, with abundant intralesional colonies of Gram-negative rods. Yersinia (Y.) pseudotuberculosis serotype O:1 and biotype 1 belonging to the sequence type ST90 was isolated from the affected organs in both animals. Following this diagnosis, » of the colony (99 animals) was culled and submitted for gross and histopathological analysis, and a bacterial culture selective for Yersinia spp. of lung, liver, and spleen was performed. From these 99 animals, one gravid female was tested and found to be positive for Y. pseudotuberculosis in the absence of clinical symptoms and histopathological lesions. PCR analysis of altogether three bacterial isolates for virulence factors revealed the presence of the ail gene, and one isolate was also positive for the virF and yadA plasmid genes. CONCLUSIONS: These findings suggest that Carollia perspicillata are susceptible to lethal yersiniosis but do not represent a regular reservoir for Y. pseudotuberculosis. Culling of » of the population was sufficient to limit the spread of this infection among the colony. Moreover, no infections were detected in cohabitant nocturnal animals and caretakers, indicating that the zoonotic risk in this case was low.

A survey for selected avian viral pathogens in backyard chicken farms in Finland.

Backyard poultry are regaining popularity in Europe and increased interest in the health and management of non-commercial farms has resulted. Furthermore, commercial poultry farm owners have become concerned about the risk represented by contagious avian diseases that nearby backyard poultry could transmit. Fifty-one voluntary backyard chicken farms were visited between October 2012 and January 2013. Blood samples and individual cloacal swabs were collected from 457 chickens. In 44 farms (86%), one or more of the tested chickens had antibodies against avian encephalomyelitis and chicken infectious anaemia viruses, 24 farms (47%) had chickens seropositive for infectious bronchitis virus, 10 farms (20%) had chickens seropositive for infectious bursal disease virus, six farms (12%) had chickens seropositive for infectious laryngotracheitis virus and two farms (5.4%) had chickens seropositive for avian influenza virus. No farms had chickens seropositive for Newcastle disease virus. Of the 51 farms, five (10%) had chickens positive for coronavirus reverse transcription polymerase chain reaction. A phylogenetic analysis showed that all backyard chicken coronaviruses collected were QX type infectious bronchitis viruses. All chickens tested for avian influenza and Newcastle disease viruses using real time reverse transcription polymerase chain reaction were negative. To our knowledge, there is no evidence to date to suggest that these diseases would have been transmitted between commercial and non-commercial flocks.

Characterization of European Yersinia enterocolitica 1A strains using restriction fragment length polymorphism and multilocus sequence analysis.

UNLABELLED: Yersinia enterocolitica is currently divided into two subspecies: subsp. enterocolitica including highly pathogenic strains of biotype 1B and subsp. palearctica including nonpathogenic strains of biotype 1A and moderately pathogenic strains of biotypes 2-5. In this work, we characterized 162 Y. enterocolitica strains of biotype 1A and 50 strains of biotypes 2-4 isolated from human, animal and food samples by restriction fragment length polymorphism using the HindIII restriction enzyme. Phylogenetic relatedness of 20 representative Y. enterocolitica strains including 15 biotype 1A strains was further studied by the multilocus sequence analysis of four housekeeping genes (glnA, gyrB, recA and HSP60). In all the analyses, biotype 1A strains formed a separate genomic group, which differed from Y. enterocolitica subsp. enterocolitica and from the strains of biotypes 2-4 of Y. enterocolitica subsp. palearctica. Based on these results, biotype 1A strains considered nonpathogenic should not be included in subspecies palearctica containing pathogenic strains of biotypes 2-5. SIGNIFICANCE AND IMPACT OF THE STUDY: Yersinia enterocolitica strains are currently divided into six biotypes and two subspecies. Strains of biotype 1A, which are phenotypically and genotypically very heterogeneous, are classified as subspecies palearctica. In this study, European Y. enterocolitica 1A strains isolated from both human and nonhuman sources were characterized using restriction fragment length polymorphism and multilocus sequence analysis. The European biotype 1A strains formed a separate group, which differed from strains belonging to subspecies enterocolitica and palearctica. This may indicate that the current division between the two subspecies is not sufficient considering the strain diversity within Y. enterocolitica.

Multiple-locus variable-number tandem-repeat analysis in genotyping Yersinia enterocolitica strains from human and porcine origins.

Sporadic and epidemiologically linked Yersinia enterocolitica strains (n = 379) isolated from fecal samples from human patients, tonsil or fecal samples from pigs collected at slaughterhouses, and pork samples collected at meat stores were genotyped using multiple-locus variable-number tandem-repeat analysis (MLVA) with six loci, i.e., V2A, V4, V5, V6, V7, and V9. In total, 312 different MLVA types were found. Similar types were detected (i) in fecal samples collected from human patients over 2 to 3 consecutive years, (ii) in samples from humans and pigs, and (iii) in samples from pigs that originated from the same farms. Among porcine strains, we found farm-specific MLVA profiles. Variations in the numbers of tandem repeats from one to four for variable-number tandem-repeat (VNTR) loci V2A, V5, V6, and V7 were observed within a farm. MLVA was applicable for serotypes O:3, O:5,27, and O:9 and appeared to be a highly discriminating tool for distinguishing sporadic and outbreak-related strains. With long-term use, interpretation of the results became more challenging due to variations in more-discriminating loci, as was observed for strains originating from pig farms. Additionally, we encountered unexpectedly short V2A VNTR fragments and sequenced them. According to the sequencing results, updated guidelines for interpreting V2A VNTR results were prepared.

Characteristics of Yersinia enterocolitica biotype 1A strains isolated from patients and asymptomatic carriers.

Yersinia enterocolitica biotype 1A strains are frequently isolated from the environment, foods, and animals, and also from humans with yersiniosis. There are controversial reports on the pathogenicity of biotype 1A strains. In this study, 811 fecal samples from asymptomatic humans from Switzerland were studied for the presence of Y. enterocolitica. Nine (1.1%) of the 811 samples were positive for Y. enterocolitica 1A. These strains were compared with 12 Y. enterocolitica 1A strains from Swiss patients with diarrhea isolated in the same year. Almost all (20/21) Y. enterocolitica 1A strains carried the ystB gene, seven strains carried the hreP gene, and none carried the ail, ystA, myfA, yadA, or virF genes. Most (17/21) Y. enterocolitica 1A strains belonged to two major clusters, A and B, by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Strains of cluster B were only isolated from humans with diarrhea; however, ystB and hreP genes were detected in strains from both clinical and non-clinical samples and from strains of clusters A and B. Using ribotyping, six restriction patterns among biotype 1A strains were obtained with HindIII enzyme. The most common ribotype (RT I) was found in strains isolated from humans with and without diarrhea. All biotype 1A strains had a unique NotI profile by pulsed-field gel electrophoresis (PFGE), showing a very high genetic diversity. In this study, Y. enterocolitica 1A strains from clinical and non-clinical samples could not be clearly differentiated from each other. More research is needed in order to prove that biotype 1A strains are a primary cause for human yersiniosis and not only a secondary finding.

Pathogenic Yersinia enterocolitica O:3 isolated from a hunted wild alpine ibex.

Occurrence of Yersinia spp. in wild ruminants was studied and the strains were characterized to get more information on the epidemiology of enteropathogenic Yersinia in the wildlife. In total, faecal samples of 77 red deer, 60 chamois, 55 roe deer and 27 alpine ibex were collected during 3 months of the hunting season in 2011. The most frequently identified species was Y. enterocolitica found in 13%, 10%, 4% and 2% of roe deer, red deer, alpine ibex and chamois, respectively. Interestingly, one Y. enterocolitica O:3 strain, isolated from an alpine ibex, carried the important virulence genes located on the virulence plasmid (yadA and virF) and in the chromosome (ail, hreP, myfA and ystA). Most of the Y. enterocolitica strains belonged to biotype 1A of which 14 were ystB positive. Further studies are needed to clarify the importance of alpine ibex as a reservoir of pathogenic Y. enterocolitica.

Detection and characterization of Shiga toxin-producing Escherichia coli in faeces and lymphatic tissue of free-ranging deer.

Shiga toxin-producing Escherichia coli (STEC) have led to outbreaks worldwide and are considered emerging pathogens. Infections by STEC in humans have been reported after consumption of mainly beef, but also deer. This study investigated the occurrence of STEC in deer in Germany. The virulence genes eae, e-hlyA and saa, the stx subtypes, pulsed-field gel electrophoresis (PFGE) patterns and serovars were studied. In total, 120 samples of 60 animals were screened by real-time polymerase chain reaction (PCR). The PCR results showed a high detection rate of stx genes (83%). Mainly faecal samples, but also some lymphatic tissue samples, tested stx-positive. All isolates carried stx2, were eae-negative and carried e-hlyA in 38% and saa in 9% of samples. Serovars (O88:[H8], O174:[H8], O146:H28) associated with human diseases were also identified. In some animals, isolates from lymphatic tissue and faecal samples showed undistinguishable PFGE patterns. The examined deer were shown to be relevant reservoirs of STEC with subtype stx2b predominating.

Yersinia enterocolitica strains associated with human infections in Switzerland 2001-2010.

Yersinia enterocolitica infections are common in humans. However, very scarce data are available on the different biotypes and virulence factors of human strains, which has proved to be problematic to assess the clinical significance of the isolated strains. In this study, the presence of the ail gene and distribution of different bio- and serotypes among human Y. enterocolitica strains and their possible relation to the genotype and antimicrobial resistance were studied. In total, 128 Y. enterocolitica strains isolated from human clinical samples in Switzerland during 2001-2010 were characterised. Most (75 out of 128) of the Y. enterocolitica strains belonged to biotypes 2, 3 or 4 and carried the ail gene. One of the 51 strains that belonged to biotype 1A was also ail positive. Most of the ail-positive strains belonged to bioserotype 4/O:3 (47 out of 76) followed by 2/O:9 (22 out of 76). Strains of bioserotype 4/O:3 were dominant among patients between 20 and 40 years old and strains of biotype 1A dominate in patients over 40 years. Strains belonging to biotypes 2, 3 and 4, which all carried the ail gene, exhibited a high homogeneity with PFGE typing. Y. enterocolitica 2/O:5,27 and 2/O:9 strains showed resistance to amoxicillin/clavulanic acid and cefoxitin, but Y. enterocolitica 4/O:3 strains did not.

Evaluation of isolation methods for pathogenic Yersinia enterocolitica from pig intestinal content.

AIMS: The aim of this study was to evaluate the efficiency of four isolation methods for the detection of pathogenic Yersinia enterocolitica from pig intestinal content. METHODS AND RESULTS: The four methods comprised of 15 isolation steps using selective enrichments (irgasan-ticarcillin-potassium chlorate and modified Rappaport broth) and mildly selective enrichments at 4 or 25 degrees C. Salmonella-Shigella-desoxycholate-calcium chloride agar, cefsulodin-irgasan-novobiocin agar were used as plating media. The most sensitive method detected 78% (53/68) of the positive samples. Individual isolation steps using cold enrichment as the only enrichment or as a pre-enrichment step with further selective enrichment showed the highest sensitivities (55-66%). All isolation methods resulted in high numbers of suspected colonies not confirmed as pathogenic Y. enterocolitica. CONCLUSIONS: Cold enrichment should be used in the detection of pathogenic Y. enterocolitica from pig intestinal contents. In addition, more than one parallel isolation step is needed. SIGNIFICANCE AND IMPACT OF THE STUDY: The study shows that depending on the isolation method used for Y. enterocolitica, the detected prevalence of Y. enterocolitica in pig intestinal contents varies greatly. More selective and sensitive isolation methods need to be developed for pathogenic Y. enterocolitica.

Characterization of Yersinia enterocolitica 4/O:3 isolates from tonsils of Bavarian slaughter pigs.

AIMS: Yersinia enterocolitica 4/O:3 isolates of slaughter pigs originating from different farms were characterized to study the distribution of different genotypes at farm. A correlation between the genotypes and the resistance patterns was also examined. METHODS AND RESULTS: Hundred and eighty-seven ail-positive Y. enterocolitica 4/O:3 isolates recovered from pigs originating from 31 Bavarian farms in 2000, 2003 and 2004 were characterized. PFGE using NotI, ApaI and XhoI enzymes revealed 31 genotypes. The most common genotype was found in 13% of the pigs. From most farms (71%), only one genotype was found. Some genotypes were found during different years. Low resistance was noted to streptomycin (9%), sulphamethoxazole (9%), amoxicillin/clavulanic acid (5%) and tetracycline (1%) by agar disc diffusion method. CONCLUSIONS: Several genotypes were found. Some genotypes were widely distributed and persisted for years. Farm-specific genotypes may exist. No clear relation between the genotypes and antimicrobial patterns was found. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides data on the genetic diversity of Bavarian pig strains and antimicrobial resistance. It may be of interest for other countries where Y. enterocolitica strains are genotyped to get more information about the strain distribution of this pathogen.

Whole genome sequence analysis of antimicrobial resistance genes, multilocus sequence types and plasmid sequences in ESBL/AmpC Escherichia coli isolated from broiler caecum and meat.

Plasmid-encoded extended-spectrum ß-lactamase and AmpC gene-carrying Escherichia coli (ESBL/AmpC E. coli) is an increasing cause of human infections worldwide. Increasing carbapenem and colistin resistance further complicate treatment of these infections. The aim of this study was to assess the occurrence of ESBL/AmpC E. coli in different broiler flocks and farms, as well as in broiler meat, in a country with no antimicrobial usage in broiler production. An additional goal was to assess the genetic characteristics of ESBL/AmpC E. coli isolates by using whole genome sequencing (WGS). Altogether 520 caecal swabs and 85 vacuum-packed broiler meat samples were investigated at the slaughterhouse level. WGS of the bacterial isolates revealed acquired antimicrobial resistance (AMR) genes, multilocus sequence types (MLST) and plasmid sequences. ESBL/AmpC E. coli was identified in 92 (18%) of the caecum and 27 (32%) of the meat samples. ESBL/AmpC E. coli-carrying birds derived from six (33%) out of 18 farms. Of the two blaESBL/AmpC genes detected by PCR, blaCMY-2 (96%) was predominant over blaCTX-M-1 (4%). Furthermore, WGS revealed an additional AMR gene sul2. Carbapenemase, colistin, and other AMR genes were not detected from the isolates of either the caecal or meat samples. Altogether seven MLSTs (ST101, ST117, ST212, ST351, ST373, ST1594 and an unknown ST) and a variety of different plasmid sequences (IncB/O/K/Z, IncI1, IncFII, IncII, IncFIB, IncFIC, IncX1 and an additional set of Col-plasmids) were detected. This is the first study on genomic epidemiology of ESBL/AmpC E. coli on broiler farms and flocks with no antimicrobial usage, by using WGS analysis. Results show that ESBL/AmpC E. coli occurrence is common both in the caecum and in the packaged meat. However, compared to other European countries, the occurrence is low and the presence of AMR genes other than blaCMY-2 and blaCTX-M-1 is rare. More studies are needed to understand the ESBL/AmpC E. coli occurrence in broiler production to prevent the meat from contamination during slaughter and processing, thereby also preventing zoonotic transmission of ESBL/AmpC E. coli. Additionally, more studies are needed to understand the ecology and fitness cost of Enterobacteriaceae plasmids in animal production in order to prevent their acquisition of plasmid-encoded antimicrobial resistance genes such as carbapenem and colistin resistance genes, as this would pose a great hazard to food safety.

Low Occurrence of Extended-Spectrum ß-lactamase-Producing Escherichia coli in Finnish Food-Producing Animals.

ESBL/AmpC-producing Escherichia coli is increasingly isolated from humans and animals worldwide. The occurrence of ESBL/AmpC-producing E. coli was studied in food-producing animals in Finland, a country with a low and controlled use of antimicrobials in meat production chain. A total of 648 cattle, 531 pig, 495 broiler and 35 turkey faecal samples were collected from four Finnish slaughterhouses to determine the presence of extended-spectrum ß-lactamase (ESBL/AmpC)-producing E. coli. In addition, 260 broiler and 15 turkey samples were screened for carbapenemase-producing E. coli. Susceptibility to different class of cephalosporins and meropenem was determined with disc diffusion tests according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Determination of ESBL/AmpC production was performed with a combination disc diffusion test according to the recommendations of the European Food Safety Authority (EFSA). Plasmidic blaESBL/AmpC  genes were characterized by polymerase chain reaction and sequencing. A collection of isolates producing AmpC enzyme but not carrying plasmidic blaAmpC  was analysed by PCR and sequencing for possible chromosomal ampC promoter area mutations. Altogether ESBL/AmpC-producing E. coli was recovered from five cattle (0.8%), eight pig (1.5%) and 40 broiler samples (8.1%). No ESBL/AmpC-producing E. coli was found in turkey samples. Carbapenem resistance was not detected. Altogether ESBL/AmpC-producing E. coli was found on 4 (2.0%), 3 (4.5%) and 14 (25%) cattle, pig and broiler farms, respectively. From cattle samples 3 (27%) blaCTX-M-1  and from broiler samples 13 (33%) blaCTX-M-1  and 22 (55%) blaCMY-2 gene-carrying isolates were detected. In pigs, no plasmidic blaESBL/AmpC gene-carrying isolates were found. In all analysed isolates, the same mutations in the promoter region of chromosomal ampC were detected. The results showed low occurrence of ESBL/AmpC-producing E. coli in Finnish food-producing animals. In pigs, plasmidic blaESBL/AmpC -carrying E. coli was not detected at all.

Bacterial quality and safety of packaged fresh leafy vegetables at the retail level in Finland.

Consumption of packaged fresh leafy vegetables, which are convenient ready-to-eat products, has increased during the last decade. The number of foodborne outbreaks associated with these products has concurrently increased. In our study, (1) label information, (2) O2/CO2 composition, (3) bacterial quality and (4) safety of 100 fresh leafy vegetables at the retail level were studied in Finland during 2013. Bacterial quality was studied using aerobic bacteria (AB) and coliform bacteria (CB) counts, and searching for the presence of Escherichia coli, Listeria and Yersinia. The safety was studied by the presence of Salmonella, ail-positive Yersinia, stx-positive E. coli (STEC) and Listeria monocytogenes using PCR and culturing. Important label information was unavailable on several packages originating from different companies. The packaging date was missing on all packages and the date of durability on 83% of the packages. Storage temperature was declared on 62% of the packages and 73% of the packages contained information about prewashing. The batch/lot number was missing on 29% of the packages. Very low oxygen (O2) (<1%) and elevated carbon dioxide (CO2) (2-22%) concentrations were measured in all packages labelled to contain a protective atmosphere. O2 and CO2 concentrations varied widely in the rest of the packages. AB and CB counts were high in the leafy vegetable samples varying between 6.2 and 10.6 and 4.2-8.3logcfu/g, respectively. In most of the samples, the AB and CB counts exceeded 10(8) and 10(6)cfu/g, respectively. A positive correlation was observed between the AB and CB counts. E. coli was isolated from 15% of the samples and Yersinia from 33%. L. monocytogenes was isolated from two samples and ail-positive Y. enterocolitica in one. Using PCR, STEC was detected in seven samples, and Salmonella and ail-positive Y. enterocolitica in two samples each. The AB and CB mean values of products originating from different companies varied widely. High AB and CB counts and pathogenic bacteria were detected in ready-to-eat products not needing washing before use. Our study shows that the bacterial quality and safety of packaged fresh leafy vegetables is poor and label information on the packages is inadequate. More studies are needed concerning the impact of a protective atmosphere on bacterial growth, and the impact of washing for removing bacteria.

Zoonotic Public Health Hazards in Backyard Chickens.

Backyard poultry has become increasingly popular in industrialized countries. In addition to keeping chickens for eggs and meat, owners often treat the birds as pets. However, several pathogenic enteric bacteria have the potential for zoonotic transmission from poultry to humans but very little is known about the occurrence of zoonotic pathogens in backyard flocks. The occurrence and the antimicrobial resistance of Salmonella enterica, Campylobacter spp., Listeria monocytogenes and enteropathogenic Yersinia spp. was studied in 51 voluntary backyard chicken farms in Finland during October 2012 and January 2013. Campylobacter isolates were further characterized by pulsed-field gel electrophoresis (PFGE), and the occurrence of ESBL/AmpC-producing E. coli was investigated. The findings from this study indicate that backyard chickens are a reservoir of Campylobacter jejuni strains and a potential source of C. jejuni infection for humans. Backyard chickens can also carry L. monocytogenes, although their role as a primary reservoir is questionable. Campylobacter coli, Yersinia pseudotuberculosis and Salmonella enterica were only found sporadically in the faecal and environmental samples of backyard poultry in Finland. No Yersinia enterocolitica carrying the virulence plasmid was isolated. All pathogens were highly susceptible to most of the antimicrobials studied. Only a few AmpC- and no ESBL-producing E. coli were found.

Meat Juice Serology and Improved Food Chain Information as Control Tools for Pork-Related Public Health Hazards.

The seroprevalence of Salmonella spp., pathogenic Yersinia spp., Toxoplasma gondii and Trichinella spp. was studied in 1353 finishing pigs from 259 farms that were allocated according to farm types: large fattening farms (≥ 1000 pig places), small fattening farms (< 1000 pig places) and farrow-to-finish farms. The antibodies were analysed with commercial ELISA kits in meat juice samples that were collected at Finnish slaughterhouses. Salmonella antibodies were rare (3% of pigs, 14% of farms) when the cut-off optical density (OD) value 0.2 was used. Antibodies to pathogenic Yersinia spp. and T. gondii were detected in 57% of pigs and 85% of farms (OD ≥ 0.3) and in 3% of pigs and 9% of farms (OD ≥ 0.15), respectively. No antibodies to Trichinella spp. were detected (OD ≥ 0.3). The European Food Safety Authority (EFSA) considers Salmonella spp., Yersinia enterocolitica, T. gondii and Trichinella spp. as the most relevant biological hazards in the context of meat inspection of pigs. The seroprevalence of these important zoonotic pathogens was low in Finland, except that of Yersinia. The seroprevalence of Toxoplasma was significantly higher in pigs originating from small-scale fattening farms (P < 0.05). Strong positive correlation was observed at the animal level between Salmonella and Yersinia seropositivity and between Salmonella and Toxoplasma seropositivity (P < 0.05). We suggest that these results reflect the level and importance of biosecurity measures applied on the farms. Meat juice serology at slaughter is a useful tool for targeting measures to control these pathogens. The information obtained from analyses should be used as part of the food chain information (FCI).

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.

Different Yersinia enterocolitica 4:O3 genotypes found in pig tonsils in Southern Germany and Finland.

The distribution of different genotypes of Y. enterocolitica 4:O3 strains recovered from pig tonsils in Southern Germany and Finland in 1999-2000 was investigated. A total of 96 and 207 Y. enterococolitica 4:O3 isolates recovered from 47 and 66 tonsils of finishing pigs in Germany and Finland, respectively, were characterised with PFGE using NotI enzyme. In all, 39 different NotI profiles were obtained, only one of which, NB1, was found in both Germany and Finland. All strains were further characterised with ApaI and XhoI enzymes. When the 54 German and 74 Finnish strains were characterised with all three enzymes, 51 genotypes were obtained. The 23 genotypes found in German strains differed from the 28 found in Finnish strains. These results indicate that Y. enterocolitica 4:O3 genotypes have a differential geographical distribution and thus can be used in epidemiological studies.

Different genotypes of Yersinia enterocolitica 4/O:3 strains widely distributed in butcher shops in the Munich area.

The distribution of Yersinia spp. in butcher shops in the Munich area was studied. The isolates recovered were then characterised with pulsed-field gel electrophoresis (PFGE) to identify possible contamination routes. A total of 298 samples were collected from eight small butcher shops between June and August in 2001. Of these, 113 were surface samples from carcasses, offal and raw pork products, and 185 were environmental surface samples from tools, equipment and processing areas. The samples were studied with direct plating, overnight enrichment in nonselective broth and selective enrichment in two different enrichment broths. The Yersinia isolates recovered were characterised with PFGE using NotI, ApaI and XhoI DNA restriction enzymes. Yersinia was recovered from all eight butcher shops, and pathogenic Yersinia enterocolitica 4/O:3 was present in six shops. The occurrence of this pathogen on raw pork products varied from 8% to 25%. Pathogenic Y. enterocolitica 4/O:3 was isolated from two environmental samples: a worktable and a chain glove. Most (18/24) of the Yersinia-positive samples were found already after direct plating. Forty-nine Yersinia isolates from 24 samples were studied with PFGE. Twelve genotypes (I-XII) were obtained among Y. enterocolitica 4/O:3 when 33 isolates from 16 samples were characterised with NotI, ApaI and XhoI enzymes. The genotypes of Y. enterocolitica 4/O:3 strains differed among butcher shops. In most (5/6) shops, more than one genotype was found, indicating different contamination sources. In conclusion, raw pork products from butcher shops are frequently contaminated with different genotypes of pathogenic Y. enterocolitica 4/O:3, thus serving as an important transmission vehicle from butcher shops to humans.

Characterisation of sucrose-negative Yersinia enterocolitica 4/O:3 isolates recovered from pig tonsils.

Sucrose-negative Yersinia enterocolitica isolates of bioserotype 4/O:3 have been recovered for the first time. They were found in 2% of the tonsils of clinically healthy fattening pigs. These sucrose-negative Y. enterocolitica isolates could not be differentiated from Y. kristensenii isolates using API 20E; thus, they were identified using PCR and sequencing. Using pulsed-field gel electrophoresis (PFGE). NotI profiles of sucrose-negative Y. enterocolitica 4/O:3 isolates showed a high similarity to sucrose-positive Y. enterocolitica 4/O:3 isolates. This study demonstrated that sucrose-negative Y. enterocolitica 4/O:3 isolates of porcine origin can harbour virulence genes; plasmid-encoded virulence markers were found in 8 out of 11 isolates and all isolates contained chromosomal-encoded virulence markers. Thus, the pathogenicity of sucrose-negative Yersinia isolates should always be assessed.

Microbiological quality and shelf-life of vacuum-packaged 'gravad' rainbow trout stored at 3 and 8 degrees C.

Microbiological and sensory changes of vacuum-packaged 'gravad' rainbow trout slices were studied during storage at 3 and 8 degrees C. At the time of spoilage, after 27 and 20 days of storage at 3 and 8 degrees C, respectively, both mesophilic viable counts (MVC) and psychrotrophic viable counts (PVC) reached 10(6)-10(7) cfu/g at 3 degrees C and 10(7)-10(5) cfu/g at 8 degrees C. H2S-producing bacteria constituted a high proportion of the PVCs and lactic acid bacteria (LAB) counts were lower than the other determined bacterial counts. Sensory scores decreased with increasing MVC and PVC. The judges considered samples unfit for human consumption at MVC and PVC levels exceeding 10(6) and 10(7) cfu/g for samples stored at 3 and 8 degrees C, respectively. At respective levels of 10(7) and 10(8) cfu/g, most of the samples were deemed unfit. The main reasons for sensory rejection at both storage temperatures were the lack of the typical product odour or an ammonia off-odour and colour change to dark violet. The shelf-lives of the rainbow trout slices based on microbiological and sensory analyses were 20 days and 18 days at 3 and 8 degrees C, respectively.