Detection, semiquantitative enumeration, and antimicrobial susceptibility of Yersinia enterocolitica in pork and chicken meats in Italy

Yersinia enterocolitica is recognized as an etiological agent of gastroenteritis, lymphadenitis, and chronic sequelae. During 2006 and 2007, 205 samples (125 pork and 80 chicken meats) were collected in Italy and tested for detection and most-probable-number (MPN) enumeration of Y. enterocolitica organisms. The microorganism was isolated from 45 samples (21.9%): 19 (15.2%) pork samples and 26 (32.5%) chicken samples. Y. enterocolitica MPN contamination levels were low, ranging from 0.30 to 1.50/g. Most (94.4%) Y. enterocolitica strains were biotype 1A (serotypes O:3; O:5; O:6,30; O:6,30-6,31; O:7,8-8-8,19; O:8; O:9; O:25,35; O:36; and O nontypeable), and 5.6% of the isolates were bioserotype 2/O:9. All isolates were tested for yadA, ail, inv, ystA, and ystB virulence sequences. The yadA gene was detected in two strains (3.7%) isolated from chicken samples: one Y. enterocolitica 2/O:9 yadA+ ail+ ystA+, and one Y. enterocolitica 1A/O:7,8-8-8,19 yadA+ inv+ ystB+. Two (3.7%) 2/O:9 strains, isolated from pork products, were ail+ ystA+. Most biotype 1A strains were ystB+ (84.3%) and inv+ (39.2%). All strains were sensitive to cefotaxime, ciprofloxacin, chloramphenicol, nalidixic acid, streptomycin, sulfonamide, tetracycline, trimethoprim, and trimethoprim-sulfamethoxazole. Resistance to gentamicin and aztreonam was observed in 1.9% of the isolates. High levels of resistance were detected toward amoxicillin-clavulanic acid (27.8%), ampicillin (75.9%), and erythromycin (100%). The authors hypothesize that Y. enterocolitica pathogenic biotypes are rather uncommon in foods when compared with their isolation rates from animal sources and that chicken meat could be contaminated as well as pig meat and its derived products.     

Prevalence of Yersinia enterocolitica in fattening pigs

The prevalence of pathogenic Yersinia enterocolitica in pig herds was monitored during six trials (at four different farrow-to-finisher farms). Samples were taken throughout the whole rearing period from birth of the piglets to the final fattening stage, and different samples were taken from these pigs during the slaughter process. Environmental samples also were evaluated to identify potential sources of on-farm infection. Y. enterocolitica was isolated using irgasan-ticarcillin-potassium chlorate broth enrichment and cefsulodin-irgasan-novobiocin agar culture. Colonies were identified using bio- and serotyping methods and by PCR assay. Pathogenic Y. enterocolitica were not isolated from fecal samples from piglets and weaners. The only fecal samples positive for Y. enterocolitica were obtained during the fattening stage. The prevalence of Y. enterocolitica in fattening pig herds ranged between 0 and 65.4%. Y. enterocolitica isolates were detected at the abattoir in 38.4% of the tonsils, in 3.8% of the ileocecal lymph nodes, on 0.3% of the carcass surfaces before chilling, and on 0% of the carcass surfaces after chilling. Almost all isolates belonged to bioserotype 4/O:3. Only one strain was identified as O:9. All isolates contained the ail gene. The yopT gene was found in 99.1% of the farm isolates but in only 76.6% of the isolates found at the abattoir from the corresponding carcasses. Although a direct link between porcine isolates and human infection has not been demonstrated, the similarity of the bioserotypes in infected pigs and humans and the presence of virulence factors in porcine isolates should encourage further studies to determine the risk of transmission of Y. enterocolitica to humans from pigs and pork products.     

High prevalence of yadA-positive Yersinia enterocolitica in pig tongues and minced meat at the retail level in Finland

Polymerase chain reaction (PCR) was used to determine the prevalence of yadA-positive Yersinia enterocolitica in pig tongues and minced meat at the retail level in Finland and to confirm the yadA-positive Y. enterocolitica isolates recovered from the same samples using the conventional culture method. A total of 51 pig tongues purchased at 12 retail outlets and 255 minced meat samples purchased at 40 retail outlets in the Helsinki area were studied. The prevalence of Y. enterocolitica carrying the yadA gene was 92% in pig tongues and 25% in minced meat using PCR and 78% in tongues and 2% in minced meat with the culture method. The prevalence of yadA-positive tongues was higher (98%) when both PCR- and culture-positive results were included because Y. enterocolitica carrying the yadA gene could also be isolated in three PCR-negative tongue samples. In the minced meat samples, all PCR-negative samples were also culture-negative. With the culture method, 66 of 80 yadA-positive isolates in 38 tongues and all yadA-positive isolates (4) in four minced meat samples were recovered after selective enrichment. A total of 92 isolates of Y. enterocolitica bioserotype 4/O:3 in tongues and 5 isolates in minced meat were found, of which 13% in tongues and 20% in minced meat did not carry the yadA gene.     

致病性小肠结肠炎耶尔森氏菌分离鉴定及三重PCR检测方法的建立

为鉴定导致患病鲫鱼肛门红肿、腹部有出血点症状的病原菌,并建立一种快速检测该病原菌的方法。本研究从患病鲫鱼中分离了病原菌,采用形态学、理化特性分析及16S rDNA序列分析方法鉴定菌株。采用PCR扩增法检测该菌毒力基因,琼脂纸片扩散法检测该菌株的耐药性,致病性能验证试验检测其致病性。针对该菌ail基因、inv基因和intB基因设计了3条特异性引物,通过对反应体系和条件的优化,建立了一种检测该菌的三重PCR方法并初步应用于患病鲫鱼样品的检测中。结果显示,从患病鲫鱼心脏组织中分离了一株小肠结肠炎耶尔森氏菌（Yersinia enterocolitica）,命名为fsznc-10。检测到该菌携带ail、ystB、virF、intB毒力基因,该菌对诺氟沙星、庆大霉素等5种抗生素敏感,对鲫鱼具有一定的致病性。三重PCR方法可准确扩增出小肠结肠炎耶尔森氏菌ail、inv和intB三个目的基因,而其他菌株均未扩增出目的基因。该方法检测该菌DNA最低检出量为1.704×10?6 ng/μL,检测患病鱼心脏样品的阳性率约为86.67%,与16S rDNA序列分析方法的检测符合率为100%。本试验建立的三重PCR检测法具有特异性强、敏感性高、操作简单、成本低的优点。这为临床中小肠结肠炎耶尔森氏菌的防控和检测提供参考依据。     

Detection, isolation and enumeration of Yersinia enterocolitica from raw pork

The methods available for the isolation of Yersinia enterocolitica from foods are generally considered to be less than optimal, and methods for estimation of numbers are lacking. Such methods are needed to understand better the significance of foodborne yersiniosis and to provide data for exposure assessment. We describe a method for the detection and enumeration of Y. enterocolitica containing the pYV virulence plasmid (YeP+) in samples from pork surfaces. The method uses a multiplex PCR targeting the ail and virF genes to detect Y. enterocolitica after incubation of surface swabs in Yersinia enrichment broth according to Ossmer. Enumeration was achieved by adapting the enrichment to a most probable number (MPN) method format. A presumptive result was available within 24 h of sample receipt, and YeP+ isolates were confirmed within four days. The presence/absence and MPN methods were evaluated in a pilot survey of 34 packs of raw pork meat purchased from retail outlets in Christchurch, New Zealand. YeP+ was detected by PCR on meat from 32% of the packs, and YeP+ isolates were obtained from 18% of the samples. YeP+ were present at numbers ranging from 0.30 to 5.42 MPN/cm(2). This improved method for the detection and enumeration of YeP+ from meat samples can be used for microbiological surveys to obtain data for assessments of consumer exposure to virulent Y. enterocolitica, and in outbreak investigations.     

O:8 serotype Yersinia enterocolitica strains in China

Serotypes O:3, O:8 and O:9 Yersinia enterocolitica strains carrying virulence determinants are common pathogens causing human infections. In many years of surveillance in China for Y. enterocolitica, no pathogenic O:8 strains have been found where the isolated O:8 serotypes lacked the major virulence genes and in contrast to O:3 and O:9 strains, none of the O:8 isolates were from humans. These O:8 isolates lack ail, ystA, yadA and virF genes but possess the ystB gene and all belong to Biotype 1A. These O:8 strains did not kill mice and could protect immunized mice against challenge with a pathogenic O:8 strain. Compared to the Chinese pathogenic O:3 and O:9 strains which have similar pulsed-field gel electrophoresis patterns, the 39 Chinese O:8 animal and food isolates were different from the pathogenic O:8 reference strains. This suggests the O:8 strains lacking virulence determinants may not disseminate rapidly in humans and are maintained in animal reservoirs; and therefore exhibit higher variance and divergence from the virulent type.     

Contamination of carcasses, offals, and the environment with yadA-positive Yersinia enterocolitica in a pig slaughterhouse

This study was carried out in order to evaluate the contamination of the pig-slaughtering line with pathogenic Yersinia enterocolitica carrying the yadA gene. A total of 292 samples were collected from the slaughterhouse; 131 swab samples from pig carcasses, ears, livers, kidneys, and hearts; 89 swab samples from the environment; and 72 sedimentation samples from the air. All surface samples were studied with both the polymerase chain reaction (PCR) and culture methods. The contamination rate of edible pig offals was high with both methods. Using PCR, the detection rates of yadA-positive Y. enterocolitica for livers, kidneys, and hearts were 38, 86, and 63%, respectively, and using the culture method, the detection rates were 31, 69, and 50%, respectively. Pathogenic Y. enterocolitica was also detected from different environmental sites in the slaughterhouse. Using PCR, 13% of the surface samples from the environment were contaminated with yadA-positive Y. enterocolitica. PCR-positive samples were found on the brisket saw, the hook from which the pluck set (heart, lungs, esophagus, trachea, diaphragm, liver, kidneys, and tongue with tonsils) hang, the knife used for evisceration, the floors in the eviscerating area and the weighing area, the meat-cutting table, the aprons used by trimming workers, the computer used in the meat-inspection area, and the coffeemaker used by slaughterhouse workers. The respective detection rate (6%) was considerably lower when we used the culture method. Pathogenic Y. enterocolitica was isolated from the air in the bleeding area. Bioserotype 4/O:3 was the only pathogenic bioserotype isolated in this study. A total of 113 isolates of type 4/O:3 were characterized with pulsed-field gel electrophoresis using NotI and XbaI digests. By combining these profiles, nine different pulsotypes were obtained, the most common of which (1a) was found in 19 (61%) of 31 samples from different sites. This is the same type that has dominated in pig tonsils, which suggests that tonsils may be the source of Y. enterocolitica contamination in the slaughterhouse. The four pulsotypes (1a, 4g, 6g, and 19q) found on edible offals were the same as those found in tonsils, which supports our hypothesis that tonsils are the contamination source for the liver, heart, and kidneys.     

Prevalence of pathogenic Yersinia enterocolitica in pigs slaughtered at a Swiss abattoir

Human yersiniosis is the third most common enteric disease after campylobacteriosis and salmonellosis in many European countries. However, epidemiological data on the prevalence of pathogenic Yersinia enterocolitica in animals and humans is insufficient. Pigs are assumed to be the main reservoir of pathogenic Y. enterocolitica because pig is so far the only animal species from which pathogenic strains have frequently been isolated. This work was conducted to study the frequency of ail-positive Y. enterocolitica in pigs slaughtered at a Swiss abattoir. In total, 212 pig tonsils were screened by real-time PCR and culture methods. The prevalence rate of ail-positive Y. enterocolitica in pigs at slaughter was 88% and 34% with PCR and culture methods, respectively. The 148 ail-positive isolates from the 72 culture-positive tonsils were bio-and serotyped. The most common bioserotype was 4/O:3 found in 96% (69/72) of the culture-positive samples. However, pig was also shown to be a reservoir for ail-positive Y. enterocolitica belonging to bioserotypes 2/O:5,27 and 2/O:9, which were detected in 8% (6/72) and 1% (1/72) of the culture-positive samples, respectively. Using PFGE with NotI, only a limited number of different patterns was found. In all, 6 genotypes were obtained when 86 isolates of bioserotype 4/O:3 from 69 samples were characterised and two genotypes (N1 and N4) dominated. The biotype 4 differs clearly from biotype 2 with PFGE. Antimicrobial resistance testing of 77 ail-positive Y. enterocolitica isolates from 72 samples studied with disc-diffusion revealed that all strains were sensitive to cefotaxime, chloramphenicol, ciprofloxacin and tetracycline, which are antimicrobial agents used for treatment of human disease. The isolates of bioserotype 2/O:5,27 differed from the isolates of bioserotypes 2/O:9 and 4/O:3 in resistance to ampicillin and amoxicillin/clavulanic acid.     

Effects of orange juice pH on survival, urease activity and DNA profiles of Yersinia enterocolitica and Yersinia pseudotuberculosis stored at 4 degree C

The objective of this study was to determine the survival, growth rate and possible cellular adaptation mechanisms of Y. pseudotuberculosis and Y. enterocolitica in orange juice under different pH conditions. Yersinia was inoculated in orange juice with adjusted pH levels of 3.9, 4.0, and 7.0 and stored at 4 C for 3, 24, 72 and 168 hours (h). The inter-and intra-species variation is significant to the pH and time of incubation variables (p<0.05). At 3.9 pH the CFU (colony forming units) count decreased significantly.At pH 3.9 and 4.0, Y. enterocolitica and Y. pseudotuberculosis survived for at least 30 days and 15 days, respectively. Yersinia that survived under low pH in orange juice revealed enhanced urease activity within 12 h of incubation. The attachment gene (ail) could not be detected by PCR in Y. enterocolitica from undiluted sample incubated for 24 h or longer. Moreover, the FesI-restriction profile was altered when Y. pseudotuberculosis was stored at pH 4.0 orange juice for 7 days. These results indicate that Yersinia could survive and grow at low pH and the survival mechanisms could also enable the bacteria to survive the stomach pH barrier to cause enteric infection.     

Yersinia pseudotuberculosis with limited genetic diversity is a common finding in tonsils of fattening pigs

A total of 425 pig tonsils, including 210 tonsils from fattening pigs and 215 from sows, from seven different abattoirs in Finland were studied for the occurrence of Yersinia pseudotuberculosis from 1999 to 2000. The mean prevalence of Y. pseudotuberculosis in fattening pig tonsils was 4%, varying from 0 to 10% between slaughterhouses. Y. pseudotuberculosis was not recovered from sow tonsils. All 30 Y. pseudotuberculosis isolates from eight pig tonsils were recovered after cold enrichment. Seventeen isolates from seven tonsils were found after cold enrichment for 14 days, followed by alkali treatment. Y. pseudotuberculosis was not isolated after direct plating, overnight enrichment, or selective enrichment. All 30 isolates belonged to bioserotype 2/0:3 and carried the virF gene in the virulence plasmid. The isolates exhibited calcium dependence and Congo red absorption. The pyrazinamidase test gave variable results. All isolates were characterized with pulsed-field gel electrophoresis (PFGE). Using SpeI, NotI, and XbaI enzymes, seven, five, and two different PFGE patterns were obtained, respectively. A total of 11 genotypes, gI to gXI, identified by a combination of the various SpeI, NotI, and XbaI profiles, were detected. Three pigs were found to carry more than one genotype. Overall, variations between PFGE patterns were small, indicating genetic homogeneity among pig strains of bioserotype 2/0:3.     

Evaluation of a Yersinia adhesion gene (yadA) specific PCR for the identification of enteropathogenic Yersinia enterocolitica

A total of 101 Yersinia enterocolitica strains was investigated with a PCR assay [Blais and Phillipe, Food Control, 6 (1995) 211-214] targeting the Yersinia adhesin gene (yadA) responsible for autoagglutination. Compared to the autoagglutination test the PCR assay has a specificity of 100% but a sensitivity of only 70%. This failure might be caused by the sequence heterogeneity of yadA.     

Comparison of culture, multiplex, and 5' nuclease polymerase chain reaction assays for the rapid detection of Yersinia enterocolitica in swine and pork products

Bacteriological culture was compared with multiplex and fluorogenic (TaqMan) polymerase chain reaction (PCR) assays for the detection of attachment invasion locus (ail)-bearing Yersinia enterocolitica in market weight swine, chitterlings, and ground pork. The TaqMan assay detected 1 pg of purified Y. enterocolitica DNA, whereas conventional gel-based PCR detected I ng of the same. The presence of ail-bearing Y. enterocolitica was tested in pork and feces artificially inoculated with Y. enterocolitica strain NADC 5561. The sensitivity limits of culture, multiplex, and TaqMan PCR assays were 4 x 10(3), 4 x 10(2), and 0.4 CFU/g, respectively, for the artificially inoculated pork. The sensitivity limits were 4 x 10(2), 4 x 10(2), and 0.4 CFU/g, respectively, for feces after a 48-h enrichment in a Yersinia selective broth. By the culture method, Y. enterocolitica was not detected in any of the swine specimens (n = 2,403) examined. By contrast, it was detected in 48 (2%) of the swine samples screened using the multiplex PCR and in 656 (27.2%) of these samples using the TaqMan assay. Using the culture method, Y. enterocolitica was detected in 8% of chitterling samples (n = 350) and in none of the ground pork samples (n = 350). It was identified in 27% of the chitterling samples using multiplex PCR and in 79% of these samples using the TaqMan assay. Ten percent of the ground pork samples contained Y. enterocolitica, as determined by the multiplex PCR, and 38% based on the TaqMan assay. The results suggest that pork products harbor more ail-bearing Y. enterocolitica than selected organs of freshly slaughtered hogs and that the TaqMan assay is more sensitive than either the multiplex PCR or traditional culture methods.     

Prevalence, antibiotic susceptibility, and virulence factors of Yersinia enterocolitica and related species from ready-to-eat vegetables available in Korea

A total of 673 ready-to-eat vegetable samples were collected in Korea from 2001 to 2002 and analyzed for the presence of Yersinia spp. We analyzed biotypes, serotypes, and susceptibility to 12 antibiotics and tested for virulence genes of pathogenic Yersinia enterocolitica isolates by PCR assay. Among the samples, 27 (4.0%) were found to be contaminated with Yersinia spp. Among the 27 strains of Yersinia spp. isolates, 18 strains (66.7%) of Y. enterocolitica, 5 strains (18.5%) of Y. frederiksenii, 3 strains (11.1%) of Y. intermedia, and 1 strain (3.7%) of Y. kristensenii were identified. According to the serotypes of Y. enterocolitica isolates, O:3 (11.1%) and O:5 (11.1%) were the most predominant, followed by O:8 (5.6%) and others (72.2%). For biotypes of Y. enterocolitica isolates, 1A (77.8%) was the most predominant, followed by 3B (11.1%), 3 (5.6%), and 5A (5.6%). Also, an antibiotic susceptibility test showed that Y. enterocolitica isolates were very susceptible to the antibiotics tested but highly resistant to ampicillin (94%), cephalothin (100%), and carbenicillin (83%). PCR assays with specific primers derived from yst and ail genes of Y. enterocolitica were applied to confirm the presence of pathogenic Y. enterocolitica. Among the 18 strains of Y. enterocolitica isolates, only 3 strains (O:3/1A, UT/3B, and UT/1A isolated from Chinese cabbage, onion, and spinach, respectively) were shown to have a virulence gene.     

Detection of chromosomal and plasmid--encoded virulence determinants in Yersinia enterocolitica and other Yersinia spp. isolated from food animals in Greece

The distribution of Yersinia strains in animal reservoirs was examined in 835 food animals (pigs, chickens, sheep, cows) from different Greek departments (Attica, Fthiotida, Viotia and Evia) over a one year period. The isolated strains were characterized with respect to the presence of chromosomal (yst) and plasmid-encoded virulence determinants (virF, yadA) and their antimicrobial susceptibility was tested.In total, Yersiniaspp. were obtained from 9.94% of the 835 food animals at slaughter that were sampled in this study. There was no statistically significant seasonal distribution, nor was any significant departmental distribution observed. From the 83 isolated Yersinia strains, 76 (91,57%) belonged to Y. enterocolitica (58 were of serotype O:3/biotype 4 and 18 strains were non O:3, non O:9), 3 belonged to Y. pseudotuberculosis, 2 to Y. kristensenii and 2 to Y. intermedia. Y. enterocolitica O:3/4 was mainly isolated from the pigs, while Y. enterocolitica non O:3, non O:9 was from the chickens. The strains were grouped into 5 genotypes, with respect to the presence or absence of the virulence genes. A significant predominance of genotype V, the one carrying all the three virulence genes, was observed in the strains isolated from the pigs. Complete susceptibility to most of the 3rd and to the 4th generation cephalosporins and to ciprofloxacin, was observed among the isolates. Remarkable was the association between the presence of each virulence gene separately and resistance to some antimicrobials, a matter of further investigation.     

多重PCR快速检测三种食品病原菌

建立一种快速、经济、实用的可以同步检测食品中金黄色葡萄球菌、沙门氏菌、小肠结肠炎耶尔森氏菌的多重聚合酶链式反应(polymerase chain reaction,PCR)的方法。根据金黄色葡萄球菌的nuc基因,沙门氏菌的invA基因,小肠结肠炎耶尔森氏菌的ail基因,分别设计了三对引物,对单个基因PCR和单管多重PCR扩增进行特异性、灵敏性实验以及优化反应体系。三对引物能特异性扩增出236、475、127bp的目的条带。建立的多重PCR同时对3种食源性致病菌进行检测具有较高的灵敏度,灵敏度金黄色葡萄球菌为102CFU/mL、沙门氏菌为102CFU/mL、小肠结肠炎耶尔森氏菌为102CFU/mL。初步建立能同步、简便、快速、灵敏地检测食品中金黄色葡萄球菌、沙门氏菌和小肠结肠炎耶尔森氏菌的三重PCR方法。     

Occurrence of Yersinia enterocolitica in milk and dairy products in Morocco.

A total of 227 samples of milk and dairy products were examined for the presence of Yersinia enterocolitica. Yersinia spp. were recovered from 11 of 30 raw milks (36.6%), one of 20 pasteurized milks (5%), 15 of 63 traditional fermented milks (23.8%), seven of 94 cheeses and one of 20 cream samples (5%). The overall incidence of Y. enterocolitica in milk and dairy products was 6.6%. The other Yersinia species were Y. intermedia, Y. kristensenii, Y. frederiksenii and Y. pseudotuberculosis. Y. enterocolitica was detected only in raw milk (30% of the samples), in traditional fermented milks (6.3%) and in raw milk-made cheese (4%). The majority of the Y. enterocolitica isolates were of biotype 1 (environmental strains). The Celfulodin-Irgasan-Novobiocin (CIN) Agar was found to be more efficient than the Mac Conkey Agar in the isolation of Yersinia organisms.     

Isolation and characterization of Yersinia enterocolitica from swine feces recovered during the National Animal Health Monitoring System Swine 2000 study

A national study was conducted for the isolation of pathogenic Yersinia enterocolitica in pig feces in the United States as part of the National Animal Health Monitoring System's Swine 2000 study. Fecal samples collected from swine operations from September 2000 to March 2001 from 77 production sites in 15 of the top 17 swine-producing states were tested for the presence of pathogenic Y. enterocolitica. After enrichment of swine fecal samples in irgasan-ticarcillin-potassium chlorate broth, the enriched cultures were plated on cefsulodin-irgasan-novobiocin agar for isolation of presumptive Y. enterocolitica. The isolates were confirmed as pathogenic Y. enterocolitica by the fluorogenic 5' nuclease PCR assay targeting the chromosomal attachment invasion ail gene. Of 2793 fecal samples tested, 106 (3.80%) ail-positive strains of Y. enterocolitica were isolated. These 106 ail-positive isolates originated from 7 of the 15 participating states. The predominant serotype O:3 (n = 79 of 106) was distributed in five states (n = 5 of 7). Serotype O:5 (n = 27 of 106) was also found in five states (n = 5 of 7). All isolates contained the virulence plasmid and expressed virulence-associated phenotypic characteristics. These results indicate that swine in the United Stares harbor Y. enterocolitica that can potentially cause human illness.     

Detection of Salmonella spp., Yersinia enterocolitica and verocytotoxin-producing Escherichia coli O157 in pigs at slaughter in Italy

From December 1999 to December 2000, 150 pigs were randomly selected in two large abattoirs of northern Italy. Caecal material and carcass swabs were collected and examined for Salmonella, Yersinia enterocolitica, and Escherichia coli O157. Tonsils were examined for Salmonella and Y. enterocolitica. Salmonella was isolated from the intestinal content of 55 (36.7%) specimens, from 8 (5.3%) tonsils, and from 9 (6.0%) carcasses. Ten different serotypes were detected; the more common were Salmonella derby (37.8%), Salmonella bredeney (21.6%), and Salmonella typhimurium (14.8%). S. typhimurium isolates that belonged to phage-types DT104 and DT208 were 45% and 27.3%, respectively; 18.2% belonged to U302 and 9.1% were non-typeable. Y. enterocolitica was detected in the intestinal matter of 6 (4.0%) slaughtered pigs and in 22 (14.7%) tonsils; however, this pathogen was not found on carcasses. The majority of Y. enterocolitica isolates (82.1%) belonged to serotype O:3 biotype 4, one (3.6%) belonged to serotype O:9, and 13% did not belong to any known biotype. Verocytotoxin-producing E. coli (VTEC) O157 was isolated from the intestinal content of one (0.7%) slaughtered pig and from one (0.7%) carcass; four (2.7%) faecal samples contained E. coli O157 strains negative for the presence of both eae and VT genes.     

双正交优化多重PCR检测食源性致病菌的研究

为了建立快速检测食源性致病菌的多重PCR检测方法,根据沙门氏菌fimY基因、单增李斯特氏菌hlyA基因和小肠结肠炎耶尔森氏菌ail基因设计3对特异性引物,利用双正交法分别对多重PCR反应体系中的3对引物比进行L9(33)正交优化和Taq酶、dNTPs、镁离子、混合引物的添加量进行L16(44)正交优化,并对Buffer的反应浓度进行优化。结果扩增出了3条特异性目的条带,建立的多重PCR方法具有很好的特异性。人工污染食品,9h富集培养增菌后的检出限可达100cfu/g。因此,该检测方法具有较强的实际应用价值,为检测多种食源性致病菌提供了有效的方法参考。     

Prevalence of Yersinia enterocolitica in market weight hogs in the United States

Pigs are the major animal reservoir for Yersinia enterocolitica strains, which are potentially pathogenic for humans. The goals of this study were (i) to estimate the individual animal and on-farm prevalences of Y. enterocolitica in hogs based on tonsil samples collected during National Animal Health Monitoring System Swine 2002 study and (ii) to use these data with data previously published for fecal samples to determine on-farm risk factors for Y. enterocolitica. Tonsil swabs (1,218) and fecal samples (2,847) were collected on 124 farms located in the top 17 pork-producing states. Ten percent of tonsils (122 of 1,218 samples) were positive in irgasan-tiracillin-chlorate (ITC) enrichment broth by real-time PCR, but only 5.6% of samples (68 of 1,218) were positive after subculture on the more selective cefsulodin-irgasan-novobiocin (CIN) agar. For tonsils, the on-farm prevalence based on real-time PCR detection of the ail gene in ITC enrichment broth cultures was 32% (32 of 100 premises sampled); the prevalence based on subculture in CIN agar was 19.6% (20 of 102 premises). Results of bacteriological isolation and real-time PCR analysis of tonsils and feces were combined to estimate prevalence (individual animal and farm), which was subsequently correlated with 40 farm management practices. Four factors and their accompanying odds ratios (ORs) were identified in the final regression model: location in a central state (OR = 0.3), vaccination for Escherichia coli (OR = 3.0), percentage of deaths due to scours (OR = 3.5), and presence of meat or bone meal in grower-finisher diet (OR = 4.1).