Presence and coincidence of Listeria spp., motile aeromonads and Yersinia enterocolitica in a cold-smoked salmon processing and packing plant

Environmental samples and samples of partially processed fish from a cold-smoked salmon processing and packing plant, and product samples purchased from retail outlets, were examined for the presence of Listeria monocytogenes and other Listeria spp., motile aeromonads and Yersinia enterocolitica. Listeria spp. were not isolated from raw fish but were a contaminant of processed and partially processed fish. Listeria spp. were also detected in 18.8% of surfaces in contact with fish. Unlike Listeria spp., motile aeromonads were isolated from the raw fish but not from the finished product. They were more frequently isolated from environmental sources than Listeria spp. Yersinia enterocolitica was not isolated from any of the samples tested. Little evidence was found to show a coincidence of motile aeromonads and Listeria spp., since only one subset of samples showed such a link. It is concluded that contamination by Listeria spp. was from environmental sources at the processing plant at, or beyond, the slicing stage. Reducing the number of wet areas and special cleaning and sanitation considerations for a contaminated site (the freezer seal) are suggested as ways of reducing contamination of the product.     

Experimental mixed infection of rabbits with Yersinia enterocolitica and Listeria monocytogenes

Experimental mixed infection was reproduced in rabbits after per os infection with Yersinia enterocolitica serotype 0:3 cells. Four days later some of animals were re-infected orally with Listeria monocytogenes serotype 4b cells. A third group of healthy rabbits was also infected per os with Listeria monocytogenes. The infectious process was followed dynamically from days 1-28. The experimental animals were examined for clinical, paraclinical and morphological findings. Augmentation of body temperature and alveolar macrophage number, a decreased number of peritoneal macrophages, leucopenia as well as purulent meningoencephalitis, catarrhal pneumonia, lienitis, lymphadenitis and enteritis were detected after experimental mixed infection. Both types of macrophages demonstrated a weak bactericidal activity against Yersinia enterocolitica and a highly expressed killing effect against Listeria monocytogenes. Yersinia and Listeria cells were isolated from the viscera and brain. Both species of bacteria were established intracellularly in the macrophages by electron-microscopic examination. The data received showed that mixed Yersinia enterocolitica 0:3 and Listeria monocytogenes 4b infection of rabbits runs with transitory hyperthermia as a generalized infection and is similar to the Listeria mono-infection. The immunosuppressive effect induced by oral Yersinia enterocolitica infection of rabbits promotes the expression of listerious agents.     

Occurrence of Salmonella,Campylobacter, Yersinia enterocolitica,Escherichia coli O157 and Listeria monocytogenes in Swine

The objective of this study was to investigate the occurrence of major bacterial foodborne pathogens in swine. In total, 359 samples from manure storage tanks (91) and fresh pooled faeces (268) obtained from finisher (110), sows (78) and weanlings (80) were collected and tested. Campylobacter, Salmonella, Yersinia enterocolitica, Escherichia coli O157 and Listeria monocytogenes were isolated from 36.5%, 31.5%, 5.8%, 3.3% and 3.3% of samples respectively. All E. coli O157 isolates found on 10 farms were tested but none was determined to be E. coli O157:H7. Salmonella and Campylobacter were more likely to be detected from stored manure rather than from fresh faecal samples. Yersinia enterocolitica tended to be detected more commonly from fresh samples than from manure pits. Listeria monocytogenes was not recovered from manure pits or from sow faecal samples and only infrequently found in the faeces of weanling pigs and finisher pigs. The proportion of positive samples showed a seasonal change. Salmonella was twice as likely not be recovered in winter, whereas the chance of culturing Campylobacter was higher in winter. The 113 Salmonella isolates recovered on 24 farms and the four most common serovars were Salmonella Typhimurium var. Copenhagen (31.0%), Salmonella Derby (12.4%), S. Typhimurium (10.6%) and Salmonella Agona (10.6%). Of 131 Campylobacter isolates recovered on 21 farms, 118 isolates were Campylobacter coli and 13 isolates could not be speciated. Fifteen of 21 Y. enterocolitica isolates found on 15 farms were detected in finisher pigs. The sero/biogroups of Y. enterocolitica were O3/biotype 4 (16 isolates), O6,30/biotype 1A (three isolates), O5/biotype 1A (one isolate) and O8/biotype 1B (one isolate). These findings provide baseline information on the distribution of important zoonotic pathogens in swine and indicate that pigs should be considered as a possible source of foodborne diseases in humans.     

An epidemiological survey on bovine and ovine babesiosis in Kurdistan Province, western Iran

The aim of this study was to determine the prevalence of the Babesia infection in domestic animals in Kurdistan Province of Iran for the first time. In this survey, 9,111 domestic livestock, including cattle and sheep, were randomly sampled and examined from 500 flocks in Kurdistan Province from July 2007 to September 2009. Thin peripheral blood smears were taken and then stained by Giemsa staining method. From a total of 9,111 collected samples, 2,642 were sheep and 6,469 were cattle. Babesia spp. is detected in 1,359 (51.4%) out of sheep samples and 136 (2.1%) out of cattle samples by direct examination of blood smear. Altogether, the prevalence rate of Babesia infection was 16.4% (n = 1,495) in both animal groups. Babesia ovis and Babesia bigemina were the most prevalent species found in sheep and cattle, respectively. The relatively high prevalence of Babesia infection in livestock indicates the epizootic stability status of babesiosis in the western part of Iran.     

Multiplex PCR method for differentiating highly pathogenic Yersinia enterocolitica and low pathogenic Yersinia enterocolitica,and Yersinia pseudotuberculosis

A multiplex PCR method for rapid and sensitive diagnosis, differentiating three pathogenic Yersinia groups such as the highly pathogenic Y. enterocolitica, including serotype O8, low pathogenic Y. enterocolitica, and Y. pseudotuberculosis, was developed. Four primer pairs were chosen to detect the genes fyuA, ail, inv, and virF, responsible for the virulence in pathogenic Yersinia species. Under the multiplex PCR conditions, the unique band patterns for the highly pathogenic Y. enterocolitica, low pathogenic Y. enterocolitica, and Y. pseudotuberculosis were generated from Yersinia strains. The detection limit of this method was 10¹–10³ CFU per reaction tube. This multiplex PCR method could detect highly pathogenic Y. enterocolitica O8 from the wild rodent fecal samples that were culture-positive. Therefore, the new multiplex PCR method developed in this study is a useful tool for rapid and sensitive diagnosis, distinguishing three pathogenic Yersinia groups.     

Serological cross-reactions between different Brucella species and Yersinia enterocolitica. Immunodiffusion and immunoelectrophoresis

By immunodiffusion and immunoelectrophoresis tests in agarose serological cross-reactions were demonstrated between Yersinia enterocolitica type IX and Brucella strains from four species (Brucella abortus, Brucella melitensis, Brucella suis and Brucella neotomae). No qualitative differences between these strains in their tendencies to cross-react with Yersinia enterocolitica type IX were observed. Brucella canis and Brucella ovis, which have nonsmooth colonial morphology, gave no demonstrable cross-reaction with Yersinia enterocolitica type IX.The results of absorption tests and qualitative staining reaction of the obtained precipitation lines suggest that the antigenic determinants common to Brucella and Yersinia enterocolitica type IX seemed to be associated with the outer layer and in the lipopolysaccharide complex of the respective bacteria. By immunodiffusion and immunoelectrophoresis it was possible to identify in hyperimmune sera those antibodies that derive from Brucella and Yersinia enterocolitica type IX.Keyword: serological cross-reaction, Brucella species, Yersinia enterocolitica serotype IX, immunodiffusion, immunoelectrophoresis     

Assessment of superficial contamination of bovine and ovine carcasses from the municipal slaughterhouse of Constantine in Algeria

The neck, shoulder, flank, and thigh of 36 bovine and 30 ovine carcasses were swabbed for bacteriological analyses. The greatest microbial load was found on the neck. The site averages for the flora analyzed indicated that the levels of contamination were greater than those in similar studies in France, Morocco, and Tunisia.     

Heat resistance in liquids of Enterococcus spp., Listeria spp., Escherichia coli, Yersinia enterocolitica, Salmonella spp. and Campylobacter spp

The aim of the work was to collect, evaluate, summarize and compare heat resistance data reported for Campylobacter, Enterococcus, Escherichia, Listeria, Salmonella and Yersinia spp. The work was limited to resistance in liquids with pH values 6-8. Results obtained under similar experimental conditions were sought. Thermal destruction lines for the various bacterial groups studied were constructed using log10 D values and treatment temperatures. There was a good linear relationship between log10 D and temperature with Escherichia coli, listerias and salmonellas. For campylobacters, enterococci and yersinias the relationships were weaker but, nevertheless, present. Using the slopes of the lines and their 95% confidence limits, z values and their 95% confidence limits were calculated. z values were compared with z values obtained from reports. The equations for the lines were also used for calculation of predicted means of D values at various treatment temperatures. 95% confidence limits on predicted means of D values and on predicted individual D values were also calculated. Lines and values are shown in figures and tables. Differences in heat resistance noted between and within the bacterial groups studied are discussed.     

Antagonistic effect of chosen lactic acid bacteria strains on Yersinia enterocolitica species in model set-ups,meat and fermented sausages

The present study was aimed at determining the influence of 15 strains of lactic acid bacteria on the growth of 8 Yersinia enterocolitica strains in model set-ups, and in meat and ageing fermented sausages. The investigations were performed within the framework of three alternate stages which differed in respect to the products studied, the number of Lactobacillus sp. strains and, partly, methodological approach. The ratio between lactic acid bacteria and Yersinia enterocolitica strains studied was, depending on the variant of experiment, 1:1, 1:2 and 2:1, respectively. The study also considered water activity (aw) and pH of the products investigated. The results suggest that all the lactic acid bacteria strains used within the framework of the model set-ups had antagonistic effect on all the Salmonella sp. strains. However, this ability was not observed with respect to of tested lactic acid bacteria strains in meat and fermented sausage. This ability was possessed by one of the strains investigated--Lactobacillus helveticus T 78. The temperature and time of the incubation of sausages, but not aw and pH, were found to have a distinct influence on the antagonistic interaction between the bacteria tested.     

Competitive exclusion of Yersinia enterocolitica biotype 4, serotype O:3 by Yersinia enterocolitica biotype 1A,serotype O:6,30 in tissue culture and in pigs

AIMS: To study the adhesion properties of a biotype 4, serotype O:3 (human pathogenic) strain of Yersinia enterocolitica and to determine if adhesion in vitro and colonisation in vivo can be prevented by competition with a biotype 1A, serotype O:6,30 (non-pathogenic) strain. To study interaction between Y. enterocolitica biotype 4, serotype O:3 and cultured epithelial cells using the synthetic tripeptide arginine-glycine-aspartic acid (RGD).

METHODS: The human intestinal epithelial (HEp-2) cell line was used for in vitro studies. Inocula of Y. enterocolitica biotype 4, serotype O:3 radiolabelled using tritium were incubated with HEp-2 cells and RGD tripeptide, or with Y. enterocolitica biotype 1A, serotype O:6,30 sequentially or concurrently, then washed and lysed, and radioactivity measured to determine the effect of RGD on adhesion, and competitive exclusion of pathogenic by non-pathogenic bacteria. For in vivo studies, two groups of 5-week-old piglets (n=5/group) were sequentially inoculated orally with 5x10⁹ colony forming units (cfu) of either a non-pathogenic biotype 1A, serotype O:6,30 strain of Y. enterocolitica followed by a pathogenic biotype 4, serotype O:3 strain, or vice versa. Pigs were monitored for carriage of strains using bacterial culture and a multiplex polymerase chain reaction (PCR).

RESULTS: The RGD tripeptide significantly inhibited adherence of the pathogenic Y. enterocolitica strain to cultured epithelial cells, suggesting that adhesion involved the RGD tripeptide sequence. The non-pathogenic biotype 1A, serotype O:6,30 strain of Y. enterocolitica prevented adhesion of the pathogenic strain to cells in vitro when allowed to adhere first. Pathogenic Y. enterocolitica was consistently isolated from rectal swabs from 80-100% of pigs on all sampling occasions but not from oral swabs after 14 days in pigs first inoculated with the non- pathogenic strain or at 26 days in pigs first inoculated with the pathogenic strain.

CONCLUSIONS: A non-pathogenic strain of Y. enterocolitica reduced adhesion of a human pathogenic strain in vitro but not in vivo.     

Competitive exclusion of Yersinia enterocolitica biotype 4, serotype O:3 by Yersinia enterocolitica biotype 1A, serotype O:6,30 in tissue culture and in pigs

AIMS: To study the adhesion properties of a biotype 4, serotype O:3 (human pathogenic) strain of Yersinia enterocolitica and to determine if adhesion in vitro and colonisation in vivo can be prevented by competition with a biotype 1A, serotype O:6,30 (non-pathogenic) strain. To study interaction between Y. enterocolitica biotype 4, serotype O:3 and cultured epithelial cells using the synthetic tripeptide arginine-glycine-aspartic acid (RGD). METHODS: The human intestinal epithelial (HEp-2) cell line was used for in vitro studies. Inocula of Y. enterocolitica biotype 4, serotype O:3 radiolabelled using tritium were incubated with HEp-2 cells and RGD tripeptide, or with Y. enterocolitica biotype 1A, serotype O:6,30 sequentially or concurrently, then washed and lysed, and radioactivity measured to determine the effect of RGD on adhesion, and competitive exclusion of pathogenic by non-pathogenic bacteria. For in vivo studies, two groups of 5-week-old piglets (n=5/group) were sequentially inoculated orally with 5 x 10(9) colony forming units (cfu) of either a non-pathogenic biotype 1A, serotype O:6,30 strain of Y. enterocolitica followed by a pathogenic biotype 4, serotype O:3 strain, or vice versa. Pigs were monitored for carriage of strains using bacterial culture and a multiplex polymerase chain reaction (PCR). RESULTS: The RGD tripeptide significantly inhibited adherence of the pathogenic Y. enterocolitica strain to cultured epithelial cells, suggesting that adhesion involved the RGD tripeptide sequence. The non-pathogenic biotype 1A, serotype O:6,30 strain of Y. enterocolitica prevented adhesion of the pathogenic strain to cells in vitro when allowed to adhere first. Pathogenic Y. enterocolitica was consistently isolated from rectal swabs from 80-100% of pigs on all sampling occasions but not from oral swabs after 14 days in pigs first inoculated with the non-pathogenic strain or at 26 days in pigs first inoculated with the pathogenic strain. CONCLUSIONS: A non-pathogenic strain of Y. enterocolitica reduced adhesion of a human pathogenic strain in vitro but not in vivo.     

Yersinia enterocolitica infection in goat. A serological and bacteriological investigation

The distribution of agglutinating antibodies to Yersinia enterocolitica type 2 in sera from goats in an infected herd, and in 190 other animals from different parts of the country was studied. The faecal excretion of Yersinia enterocolitica from the same animals was also examined. Experimental inoculations of two goats were carried out. The serological results indicate that subclinical cases had occurred in the infected herd during the enzooty and during the following months. Faecal excretion of the organism was observed during the first month after the acute phase of the disease. It was found to be difficult to induce the disease experimentally, but clinical signs were observed in 1 goat after injection of live Yersinia enterocolitica intraperitoneally. The Widal-titre rose to 1/1250 during the first 2 weeks after the inoculation and then fell to 1/80 during the next 2 months. The serological results indicate infections with Yersinia enterocolitica, if a Widal-titre of at least 1/80–1/160 in the first month after the acute phase of a disease and a fall to a low level during the following months are found. Among animals from different partst of the country 16.3 % had a Widal-titre which indicated infection with Yersinia enterocolitica during the previous 3–6 months.     

Serological cross-reactions between different brucella species and Yersinia enterocolitica. Agglutinating activity of 19 S and 7 S antibodies aginst somatic antigen of Brucella abortus and Yersinia enterocolitica type IX

Antisera from rabbits immunized with acetone-killed whole cells and with lipopolysaccharides from Brucella abortus (B.a.) and Yersinia enterocolitica type IX (Y.e.) were gel-filtered on Sephadex G-200 columns.All the antisera had serologically cross-reacting agglutinins against B.a. and Y.e. both in the 19 S and in the 7 S antibody fractions.The homologous and the heterologous agglutinating activity of 19 S and of 7 S antibodies was tested before and after treatment with dithiothreitol (DTT) and subsequent alkylation. Unlike 19 S antibodies, 7 S antibodies were resistant to the DTT reduction.The results from heterologous absorptions of the respective 19 S and 7 S antibody fractions indicated that 19 S antibodies both to B.a. and to Y.e. had a greater tendency towards being absorbed to the cross-reacting antigenic determinants than had the corresponding 7 S antibodies. The agglutination titres for 19 as well as 7 S antibody fractions derived from the immunizations with B.a. (both whole cells and LPS) fell more markedly after heterologous absorptions than did the analogous titres for corresponding Y.e. antibody fractions. Possible explanations of these differences are discussed.     

Serological crossreactivity between Brucella abortus and Yersinia enterocolitica 0:9 III. Specificity of the in vitro antigen-specific gamma interferon test for bovine brucellosis diagnosis in experimentally Yersinia enterocolitica 0:9-infected cattle

The course of immunological reaction in 10 Yersinia enterocolitica 0:9 experimentally-infected heifers was followed using the conventional brucellosis tests complement fixation test (CFT), serum agglutination test (SAT) and brucella card test (BCT), and a recently developed Brucella antigen-specific gamma interferon (IFN-γ) test. Initially, the animals were exposed orally to 10¹⁰ colony-forming units (CFU) of Y. enterocolitica 0:9. Four weeks later, they were inoculated intravenously with 10⁸ CFU of Y. enterocolitica 0:9 cells. After oral inoculation, the response in the conventional brucellosis tests was minimal. Only after intravenous inoculation were CFT and SAT titres and BCT reactions comparable to natural, false positive brucellosis reactors. After oral exposure the Brucellergen-stimulated release of IFN-γ peaked at values above the cut-off stimulation index of 2.5 in 80% of the heifers. After intravenous inoculation, stimulation indices above 2.5 were present in only 10% of the animals. Two B. abortus infected control cattle showed stimulation indices of 3.1 and 3.4, and a negative control animal exhibited a stimulation index of 1.0. These findings show, in contrast to a previous study, that the Brucellergen-specific IFN-γ assay cannot be used as a specific and discriminatory test for B. abortus infections.     

Demonstration of Listeria monocytogenes by immunohistochemistry in formalin-fixed brain tissues from natural cases of ovine and bovine encephalitis

In the present work, evidence of Listeria monocytogenes antigens based on the avidin-biotin complex (ABC) immunoperoxidase technique was performed on formalin-fixed central nervous system tissues (CNS) from a total of 23 natural cases of encephalitis (four ovine and 19 bovine). Listeria monocytogenes serotype 4 was isolated from 10 of 17 cultured specimens. Meningoencephalitis characterized by focal necrosis, microabscesses, perivascular cuffing, and gliosis with presence of macrophages and/or neutrophils was observed at histological examination. Positive L. monocytogenes antigens were successfully identified by immunohistochemistry (IHC) in the CNS of all 23 cases. Paraffin-embedded tissues assayed were stored up for 17 years. Morbidity of the outbreaks was between 0.3-3% and 0.1-1% for ovine and bovine cases, respectively. In all the ovine cases, flocks involved were under extensive grazing conditions. In nine of the 19 bovine cases (47.3%), supplementation with corn silage was used. The ABC test can help as a practical tool for the diagnosis of natural cases of L. monocytogenes encephalitis on formalin-fixed specimens from ovine and bovine.     

感染小肠结肠炎耶尔森菌的黄颡鱼脾脏和肾脏组织病理学观察

为了解小肠结肠炎耶尔森菌感染黄颡鱼后对其脾脏和肾脏免疫器官结构的影响,通过人工感染小肠结肠炎耶尔森菌致病后,观察黄颡鱼脾脏和肾脏显微和超微结构的病理变化。结果显示,发病黄颡鱼腹部肿大,剖检可见脾脏、肾脏肿大;肾小球萎缩,肾小管上皮细胞水肿、变性;脾组织内充满大量红细胞;脾脏、肾脏细胞中线粒体均出现不同程度的肿胀,嵴断裂,囊泡化。这表明该菌对黄颡鱼具有明显的致病性,可导致其脾脏和肾脏发生严重的病变。