Species-specific DNA probes for Campylobacter species isolated from pigs with proliferative enteritis

Cloned, chromosomal DNA probes from porcine isolates of Campylobacter hyointestinalis and C. mucosalis were developed for the detection and identification of these putative swine enteric pathogens. High molecular weight chromosomal DNA from each species was used to construct genomic libraries in plasmids. Recombinants were selected which hybridized strongly to the homologous organism, but not to any other species of Campylobacter. Species-specific recombinants were labeled with phosphorus-32 and tested for sensitivity by dot blot hybridization to various dilutions of DNA and bacteria from each swine species, including C. hyointestinalis, C. mucosalis, C. coli and C. jejuni. Specificity was tested by hybridizing these probes against various strains of C. hyointestinalis or C. mucosalis, and against reference strains of all other described Campylobacter species. A C. hyointestinalis-specific probe and a C. mucosalis-specific probe were identified which were capable of detecting 1 ng of DNA or 10(4) cfu by bacterial spot blotting on nylon membranes. These probes hybridized to intestinal mucosal scrapings containing C. hyointestinalis and C. mucosalis obtained from pigs with proliferative enteritis, but not to material from normal pigs. Thus, cloned, chromosomal DNA probes may be useful in the detection and identification of bacteria involved in swine proliferative enteritis.     

Use of embryonating eggs for isolation of Campylobacter species from intestines of swine with proliferative enteritis

Intestinal tissues from swine affected with proliferative enteritis were ground, filtered through a 0.65-micron pore membrane filter, diluted, and injected into 7-day-old embryonated hens' eggs via the yolk sac. At 2, 4, and 7 days later, yolk sac swab specimens taken from live embryos were cultured for Campylobacter species. Campylobacter hyointestinalis was recovered from eggs injected with tissues of swine with acute hemorrhagic proliferative enteritis at dilutions up to 10(-4). Campylobacter mucosalis was recovered from eggs injected with tissues of swine with chronic proliferative enteritis at dilutions up to 10(-6). Campylobacter coli was recovered from several specimens without lesions of proliferative enteritis and also from some specimens with lesions of proliferative enteritis. Two previously undescribed hemolytic Campylobacter species designed as hemolytic number 1 and hemolytic number 2 were recovered from normal and experimentally inoculated swine tissues. Few contaminating organisms grow in eggs and these were usually recovered at dilutions of 10(-2) or less. Recovery of Campylobacter species by use of these techniques was seldom successful in tissues stored at -70 C for more than 6 months.     

Pathology of proliferative haemorrhagic enteropathy in pigs.

Examination of the small intestine of pigs with proliferative haemorrhagic enteropathy showed changes consistent with defects in vascular permeability. Early in the disease there were many eosinophils and distension of lacteals and intercellular spaces with proteinaceous material. Later the predominant features were red blood cells and exudate in tissue spaces. This was most severe and extensive at the tips of villi which were covered by a cast of cells and fibrinous exudate. Adenomatous intestinal mucosal cells contained organisms that were free within the apical cytoplasm and were morphologically identical with those seen in the related disease, porcine intestinal adenomatosis. Also these bacteria were seen free in the subepithelial mucosal area, in blood vessels and within membrane-bound vesicles in phagocytic cells in the mucosa and its blood vessels. Mast cells were prominent in some areas as were thrombosed vessels.     

广西猪增生性肠炎的PCR诊断

猪增生性肠炎(porcine proliferative enteropathy,PPE),又称猪回肠炎,是由胞内劳森氏菌(Lawsonia intracellularis,LI)引起的接触性传染病[1].1931年首次报道,目前已经呈全球性流行,亚洲地区如台湾、日本、韩国等近年来也发现猪增生性肠炎;     

Plasmid profiles of six species of Campylobacter from human beings, swine, and sheep

Twenty-four isolates representing 6 species of Campylobacter were screened for plasmids. A large plasmid with an approximate molecular weight of 38 Mdal was detected in 5 C jejuni isolates originally recovered from diarrheic human beings, in one isolate of C coli recovered from diarrheic pigs, and in 1 isolate of C sputorum ssp mucosalis and 2 isolates of C hyointestinalis recovered from pigs with proliferative enteritis. One isolate of C coli and 1 isolate of C hyointestinalis contained an additional smaller plasmid with an approximate molecular weight of 1.6 Mdal; this plasmid was partially mapped by restriction endonuclease digestion. Fifteen Campylobacter isolates contained no detectable plasmids: 2 C coli, 2 C sputorum ssp mucosalis, 2 C fecalis, 1 C fetus ssp fetus, and 8 C hyointestinalis isolates. In summary, 37.5% of the Campylobacter isolates contained a 38-Mdal plasmid, with 8% having both 38 Mdal and 1.6-Mdal plasmids; 62.5% contained no detectable plasmids.     

Antimicrobial susceptibility in thermophilic Campylobacter species isolated from pigs and chickens in South Africa

Campylobacter jejuni is one of the leading causes of sporadic food-borne bacterial disease in humans. In intensive poultry and pig rearing systems the use of oral antibiotics is essential to maintain health. Consequently, there is a high risk for the thermophilic Campylobacter jejuni and C. coli resident in the intestinal tract of food animals to develop resistance to commonly used antibiotics. Contamination of meat or eggs with pathogenic strains of resistant Campylobacter could, therefore, result in a form of campylobacteriosis in humans that is difficult to treat. The aim of this investigation was to determine the antimicrobial susceptibility of thermophilic Campylobacter spp. isolated from pigs and poultry by the broth microdilution minimum inhibitory concentration (MIC) test. A total of 482 samples from the Western Cape and Gauteng provinces was collected and analysed. Thirty-eight Campylobacter isolates were obtained. Analysis of data revealed that C. jejuni strains mainly of poultry origin were more resistant to the fluoroquinolones, macrolides and tetracyclines and the C. coli strains were more resistant to the macrolides and lincosamides. Multi-resistance was also detected in 4 Campylobacter strains from the Western Cape. With the exception of tetracyclines, strains from high health Gauteng broiler farms were susceptible to antibiotics used to treat Campylobacter infections.     

Biological properties and genetic analysis of the ompA locus in chlamydiae isolated from swine.

Eight strains of Chlamydia psittaci isolated from swine with pneumonia, pleuritis, pericarditis, and enteritis were characterized through analysis of the major outer membrane protein gene ompA by a two-step polymerase chain reaction, by their interactions with cells in culture, and by the morphologic features and ultrastructure of intracellular inclusions. Amplified chlamydial ompA DNA fragments were differentiated by restriction endonuclease digestion. Chlamydial isolates were separated into 2 types on the basis of ompA restriction fragment length polymorphism. Strains of type L71 had finely granular inclusions, whereas those of type 1710S contained pleomorphic reticulate bodies (RB) in the inclusions, which are characteristic of aberrant chlamydial developmental forms. Chlamydial types L71 and 1710S required centrifuge-assisted inoculation for efficient infection of cell cultures. Cultivation in cell culture medium containing cycloheximide increased the numbers of chlamydial inclusions about 1.5-fold. These strains formed few elementary bodies in yolk sac cells of chicken embryos. Ultrastructurally, unique doublet RB were observed, particularly in strains of the ompA type L71. These doublets consisted of 2 RB, bounded by a cytoplasmic membrane, contained within a common cell wall and an extended periplasmic space. Ultrastructural examination of strains of the ompA type 1710S confirmed the aberrant chlamydial developmental forms, but evidence of viral infection of the RB as a cause of these aberrant forms was not found. The strain S45 isolated from intestinal sites of swine was a trachoma restriction fragment length polymorphism type. With the mouse biotype, it represented the second isolate from animals of Chlamydia trachomatis.     

A descriptive study of the frequency and characteristics of proliferative enteropathy in swine in Ontario by analyzing routine animal health surveillance data

Routine surveillance data, collected on pathology submissions at the Animal Health Laboratory in Guelph between 1992 and 1997, were analyzed to determine demographic, clinical, and pathologic characteristics of cases of proliferative enteropathy and the frequency of this condition relative to other infectious enteric diseases in swine in Ontario. The most commonly reported disease was Escherichia coli enteritis (average cases/year = 70.0). Among infectious enteropathies that occur typically in neonatal pigs, coccidiosis (28.4 cases/year) and rotaviral enteritis (5.6 cases/year) were reported. Among infectious enteropathies generally associated with diarrhea in weaner and grower/finisher pigs, the most frequently reported was proliferative enteropathy (27.6 cases/year), followed by swine dysentery (23.3 cases/year), transmissible gastroenteritis (19.6 cases/year), and salmonellosis (8.4 cases/year). Diarrhea and bloody diarrhea were reported in 29% and 31%, respectively, of herds diagnosed with proliferative enteropathy. Important gross intestinal lesions included mucosal hypertrophy (62% of cases), hemorrhage (47%), and mucosal necrosis (34%). Histologic intestinal lesions included epithelial hyperplasia (90% of cases), mucosal necrosis (59%), and inflammation (49%). Our results suggest that proliferative enteropathy is a major infectious enteric disease in grower/finisher pigs in Ontario.     

Molecular detection of Campylobacter species and Cytolethal distending toxin isolated from chicken livers in Tabriz

Introduction and purpose: Campylobacter jejuni and coli are zoonotic bacteria commonly associated With human diarrhea and usually transmit through consumption of meat and poultry contaminated products such as heart and liver. Cytolethal distending toxin (cdt) in Campylobacter spp. is among the significant virulence factors of these bacteria in the intestine. The purpose of this study was to determine the prevalence of Campylobacter spp. and presence of cdt genes among isolated bacteria. Materials and methods: In this cross sectional study, 100 chicken livers were examined. Detection, bacterial enumeration and isolation of Campylobacter spp. was done using Campylobacter selective agar media containing Campylobacter growth supplement, gram staining, catalase and oxidase production, hippurate hydrolysis and PCR molecular technique. Also the presence of cdt genes were detected using PCR assay. Results: From 100 studied liver samples, 43 were contaminated with Campylobacter spp. Among them 31(72 %) samples had Campylobacter jejuni and 12 (28 %) had Campylobacter coli. All Campylobacter jejuni isolates contained cdtA ،cdtB and cdtC genes. However, all of these genes detected in 9 (75 %) of isolated coli. Conclusion: The results of this study showed that great percentages of chicken livers in Tabriz were contaminated with Campylobacter.     

Comparison of intestinal mucosa homogenate and pure culture of the homologous Lawsonia intracellularis isolate in reproducing proliferative enteropathy in swine

Little information is available on reproduction of proliferative enteropathy (PE) using a virulent pure culture of Lawsonia intracellularis. Reproduction of the disease using PE-diseased mucosa homogenates, however, is well-characterized. The aims of this study were to evaluate and compare clinical signs, growth performance and the severity of lesions in pigs inoculated with intestinal mucosa homogenate or pure culture of the homologous L. intracellularis isolate. Five-week-old pigs were inoculated with pure culture of L. intracellularis (isolate PHE/MN1-00; n=10), PE-diseased mucosa (n=10), or control media (n=4). The L. intracellularis isolate PHE/MN1-00 used in the pure culture inoculum was extracted from a fragment of the same intestine used to prepare the mucosa homogenate. Clinical signs and growth performance were evaluated throughout the study. Fecal shedding was evaluated in all animals weekly during the experiment. All animals were euthanized 22 days post-inoculation, the intestines were examined grossly and histologically. Results showed that both the infection procedures reproduced clinical disease, macroscopic and histologic lesions typical of PE. Fecal shedding was detected in animals in both challenge groups. In conclusion, the L. intracellularis isolate PHE/MN1-00 reproduces typical clinical signs and lesions of PE similar to the homologous infection with an intestinal mucosa homogenate.     

Antigenic characterization of an H3N2 swine influenza virus isolated from pigs with proliferative and necrotizing pneumonia in Quebec.

A new strain of swine influenza A virus, designated A/Swine/Saint-Hyacinthe/150/90 has been isolated from pigs with severe proliferative and necrotizing pneumonia in Quebec. The antigenic characterization of the hemagglutinin was performed by hemagglutination inhibition test, immunoblot and indirect immunoprecipitation using polyclonal antisera. Only the last test was able to detect an antigenic relationship between the hemagglutinin of this isolate and an H3 subtype influenza virus. The immunoprecipitation test was a useful alternative for determining the hemagglutinin of influenza A virus subtypes. The neuraminidase inhibition test demonstrated a reactivity between the A/Swine/Saint-Hyacinthe/150/90 and antiserum against a N2 subtype influenza virus. Our results indicate that this new strain isolated for the first time in the porcine population of Canada is related to A/Sw/Hong Kong/76 H3N2 swine influenza virus.     

Prevalence of Campylobacter spp isolated from the intestinal tract of pigs raised in an integrated swine production system

OBJECTIVE: To enumerate the prevalence of Campylobacter isolates in the intestinal tract of market-weight swine raised in an integrated swine operation in Texas. SAMPLE POPULATION: Samples of cecal contents were collected from 595 pigs (mean body weight, 110 kg [242 lb]) at time of slaughter. Pigs were off-spring of Yorkshire-Landrace sows and Duroc or Hampshire boars. Pigs originated from 4 farrow-to-finish farms. PROCEDURE: During a 9-month period, visits were made to a slaughter plant to remove cecal contents from market-weight hogs. Samples were obtained from 50 pigs/visit from designated farms so that samples were obtained 3 times from pigs of each of 4 farms. Isolation of Campylobacter spp was accomplished by use of enrichment broth and restrictive media, using microaerophilic conditions. RESULTS: Campylobacter spp were isolated from 70 to 100% of the pigs, depending on the farm and the date the samples were collected. Campylobacter coli was isolated from 20 to 100% (mean, 60%) of samples, and C jejuni was isolated from 0 to 76% (mean, 31%) of samples. Campylobacter lari was isolated from 2 pigs. Concentrations of C coli or C jejuni ranged from 10(3) to 10(7) colony-forming units/g of cecal content. CONCLUSIONS AND CLINICAL RELEVANCE: Campylobacter coli generally is accepted as a common inhabitant of the intestinal tract of swine. However, analysis of results of this study suggests that a relatively high prevalence of C jejuni may be found in pigs raised on specific farms.     

Biochemical and serological properties of Actinobacillus pleuropneumoniae biotype 2 strains isolated from swine

The biochemical and serological properties of 21 strains of Actinobacillus pleuropneumoniae biotype 2 isolated from haemorrhagic necrotic pleuropneumonia of swine were examined. For serologic typing, the indirect haemagglutination (IHA) and the double gel-diffusion tests were used. On the basis of their soluble surface antigens, our A. pleuropneumoniae biotype 2 isolates could be assigned to two proposed serotypes. Serotype 1 comprised 11 strains and serotype 2 comprised 10 strains. All strains contained two surface antigen components. In the strains belonging to serotype 1, one of the antigens was identical with the serotype-specific antigen of Pasteurella haemolytica T4. Both antigens of serotype 2 strains proved to be type-specific. Four strains received from Switzerland, including the holotype strain of A. pleuropneumoniae biotype 2, and three strains isolated from swine in the G.D.R. belonged to serotype 2. Both the double gel diffusion and the IHA tests detected a 2-way cross-reaction between biotype 1, serotype 2 and biotype 2, serotype 2 strains of A. pleuropneumoniae, which could be eliminated using cross-absorbed sera.