Prevalence of porcine circovirus 2 infection in pig population in Slovakia.

The prevalence of porcine circovirus 2 (PCV-2) infection in the pig population in Slovakia was investigated. Sera from pigs suspected for post-weaning multisystemic wasting syndrome (PMWS) as well as clinically healthy pigs were tested for viral DNA and specific IgM and IgG antibodies. Pigs (n = 198) were categorized to weaning, grower and fattening ones and sows. The results showed that PCV-2 antibodies were present in 53.4% of PMWS-suspects, in 50.0% of healthy pigs and in 69.0% of sows. In PMWS-suspect grower pigs, 40.7% were positive for IgM+IgG antibodies and 22.2% for viral DNA. In PMWS-suspect fattening pigs, 50.0% were positive for IgM+IgG antibodies and 25.0% for viral DNA. In healthy fattening pigs, almost 90.0% were positive for IgG antibodies and 38.5% for viral DNA. The highest proportion of PMWS-suspects was in grower pigs and specific antibodies were increasing with the age of pigs. A combination of positivities for IgG+IgM antibodies and viral DNA was a highly significant marker of PMWS. Viral DNA was detected in seropositive as well as seronegative PMWS-suspects. Overall, in all categories of pigs tested, specific antibodies and viral DNA were detected in 54.0% and 35.5%, respectively.

Multiplex PCR assay for detection of Actinobacillus pleuropneumoniae, Pasteurella multocida and Haemophilus parasuis in lungs of pigs from a slaughterhouse.

Multiplex PCR has been developed for parallel identification of Actinobacillus pleuropneumoniae, Pasteurella multocida and Haemophilus parasuis, important pathogens of swine, responsible for considerable economic losses in swine industry. Multiplex PCR and bacteriological cultivation was used to analyze lung samples from slaughterhouse pigs. From a total of 219 lung samples, 164 (74.9 %) were positive for P. multocida, 45 (20.5 %) for A. pleuropneumoniae and 4 (1.83 %) for H. parasuis. Bacteriological examination revealed that 145 samples (66.2 %) were positive for P. multocida, 31 (14.2 %) for A. pleuropneumoniae and 2 (0.91 %) for H. parasuis.

Serotypes, virulence genes, intimin types and PFGE profiles of Escherichia coli isolated from piglets with diarrhoea in Slovakia.

Two hundred and fifty Escherichia coli isolates from diarrhoeic and healthy piglets were serotyped and tested for the presence of virulence genes for fimbriae, intimin, heat-labile (LT) and heat-stable (STa and STb) enterotoxins, Stx toxins, and enteroaggregative heat-stable 1 (EAST1) enterotoxin by polymerase chain reaction (PCR). Although 220 isolates from diarrhoeic piglets belonged to 43 O serogroups and 77 O:H serotypes, 60% were of one of the 10 serogroups O2, O8, O15, O54, O84, O101, O141, O147, O149 and O157, and 60% belonged to only 10 serotypes (O8:H-, O54:H-, O84:H7, O101:H-, O141:H-, O141:H4, O147:H-, O149:H10, O163:H-, and ONT:H-). PCR showed that 79% of 220 isolates carried genes for at least one of the virulence factors tested. The gene encoding for EAST1 was the most prevalent (65%) followed by those encoding for STb (49%), LT (42%), STa (13%), and Stx2e (4%). Eighty-three (38%) of the 220 E. coli isolates carried the gene for F4 (K88), whereas genes for F18, F5 (K99), F41, F6 (P987), F17, and intimin (eae) were detected in 9%, 3%, 3%, 3%, 1%, and 3%, respectively. Seropathotype O149:H10:F4:LT/STb/EAST1 (70 isolates) was the most common, representing 32% of isolates. Pulsed-field gel electrophoresis (PFGE) analysis with XbaI of 15 O149:H10 representative isolates from diarrhoeic piglets distinguished 14 types. The 15 isolates exhibited a wide variability of distinct restriction patterns though all belonged to the same serotype (O149:H10), and all but one showed identical virulence determinants (F4, LT, STb, and EAST1). Among 30 isolates from healthy piglets only two virulence genes were detected: EAST1 (26%) and eae (17%). In total, 12 isolates were positives for the eae gene: five isolates had intimin beta1, four possessed intimin theta and three showed intimin type xiB. This is believed to be the first study describing the presence of intimin type xiB in E. coli of porcine origin.

PCR assay for detection and differentiation of K88ab(1), K88ab(2), K88ac, and K88ad fimbrial adhesins in E. coli strains isolated from diarrheic piglets.

Primers were designed and prepared and conditions were determined for PCR detection and differentiation of enterotoxigenic E. coli bacterial strains isolated from diarrheic pigs. Primers K88/1 and K88/2 are 25 bp oligomers that correspond to a region of genes encoding one of serological variants of the K88 antigen (K88ab(1), K88ab(2), K88ac or K88ad). A positive result of PCR is an amplificate of 792 bp in size for K88ab and K88ad variant or 786 bp for K88ac variant. The individual serological variants of genes of the K88 antigen could be differentiated by cutting the obtained PCR amplificates by restriction endonucleases. The PCR analysis of 674 E. coli strains isolated from diarrheic pigs showed that 184 strains were K88 positive. By using restriction endonucleases the K88-positive strains were in 4 cases classified as K88ab variant, 180 as K88ac variant and none contained gene for the K88ad variant. Ninety-five % coincidence with serological examination using K88ab, K88ac and K88ad specific antibodies was shown.

Distribution of virulence genes in Escherichia coli strains isolated from diarrhoeic piglets in the Slovak Republic.

Ninety-two Escherichia coli isolates from 14 to 28-day-old piglets that died because of diarrhoea were examined for genes for fimbriae (F4, F5, F6, F18 and F41), enterotoxins (STa, STb and LT), verotoxin (VT2e or Stx2e) and enteroaggregative heat-stable enterotoxin 1 (EAST1) by polymerase chain reaction. Twenty-two strains (24%) carried a gene for F4, whereas genes for F18, F6 and F5 + F41 were detected in 10.8, 3.3 and 1.1% of strains respectively. Genes for STb, LT, STa and Stx2e were detected in 40.2, 26.1, 14.1 and 1.1% of strains respectively. The astA gene was detected in 49 (53.3%) isolates, 35 of which also carried genes for enterotoxins and/or fimbriae. The major genotypes reached at (in decreasing order of prevalence) were F4/STb/LT/EAST1, F18/STa/STb/EAST1, STb/EAST1, F6/STa/STb/EAST1 and F18/STb/EAST1.

Genotyping of fimbrial adhesins in Escherichia coli strains isolated from Slovak piglets suffering from diarrhea.

One-hundred sixty Escherichia coli isolates obtained from piglets with diarrhea from different parts of Slovakia were examined for the presence of genes coding for F4, F5, F6 and F41 fimbrial adhesins, and hemolytic activity. According to polymerase chain reaction tests 74 (46%) E. coli isolates were positive for primers that detected genes coding for fimbrial adhesins. Of these 74 isolates, 64 were positive for genes encoding for F4+, four for F5+, five for F6+, and one for both F41+ and F5+ adhesins.

[Occurrence and characteristics of staphylococcal enterotoxin antibodies in gynecologic patients]. / Výskyt a charakterizácia protilátok proti stafylokokovým enterotoxínom u gynekologických pacientiek.

OBJECTIVE: To control the occurrence of the antibodies against Staphylococcal type A and type B enterotoxin in gynaecological patients and in selected patients to determine the thermodynamic parameters of antibodies against Staphylococcal enterotoxins in their blood samples. DESIGN: Retrospective clinical study. SETTING: 2nd Department of Obstetrics and Gynaecology, P. J. Safárik University Kosice, Slovak Republic; Research Institute of Veterinary Medicine, Kosice, Slovak Republic; Department of Microbiology and Immunology, University of Veterinary Medicine, Kosice, Slovak Republic; Department of Food Microbiology and Toxicology, Food Research Institute, University of Wisconsin, Madison, USA. METHODS: The occurrence of antibodies against Staphylococcal type A and type B enterotoxin was determined in 68 patients hospitalized in Department of Obstetrics and Gynecology in Kosice. RESULTS: The occurrence of antibodies against Staphylococcal enterotoxins was determined by radioimmunoassay (RIA) in blood samples of 45 (66%) patients. The antibodies against Staphylococcal type A enterotoxin were determined in 36 (53%) patients and the antibodies against Staphylococcal type B enterotoxin were determined in 9 (13%) patients. The antibodies against both type A and type B enterotoxins were determined simultaneously in blood samples of 10% of all patients. The thermodynamic parameters of the antibodies were determined in 5 patients with positive serum findings. CONCLUSION: With regard to the existence of heterogeneous clinical findings in large amount of patients with antibodies against Staphylococcal enterotoxins, the next study of Staphylococcal enterotoxins role in pathogenesis of wide spectrum of diseases is necessary.

Yeast-like microorganisms in eye infections.

The proportion of yeast species involved in eye infections in 11 patients was examined. The presence of yeast organisms as causative agents of endophthalmitis was found in corneal smears (n = 4), conjunctival swabs (4), and vitreous fluid (3). Altogether 5 strains of Candida albicans, 2 strains of C. krusei and one strain each of C. guilliermondii, C. parapsilosis, C. tropicalis and Cryptococcus neoformans were isolated from the clinical material. The hematogenic origin of endophthalmitis was proved in 7 cases on the basis of positive blood samples and in 2 cases by the isolation of yeasts from the tip of an intravenous catheter. Endophthalmitis-supporting risk factors such as indwelling intravenous catheters, prolonged use of broad-spectrum antibiotics and chemotherapy, surgical intervention, diabetes mellitus, and malignancy were observed in the patients.

Incidence of Candida species isolated from human skin and nails: a survey.

Distribution of Candida species was investigated by examining 245 samples from skin lesions and nails. The isolates were identified using standard laboratory methods including germ tube test, micromorphology of colonies on rice agar, the commercial kit, saccharide assimilation and fermentation tests. Eight species of Candida were identified: C. albicans accounted for 56.4% of the isolates, C. parapsilosis 29.1, C. tropicalis 7.8, C. pulcherrima 2.9, C. guilliermondii 1.5, C. krusei and C. zeylanoides for 0.9% each, and C. robusta for 0.5%. The factors significantly associated with colonization were prolonged antibiotic therapy, parenteral nutrition, low birth body mass of infants, intubation, duration of stay in hospital, indwelling intravenous catheter, malignancies, diabetes, surgery, and obesity.

Monoclonal antibodies against the common polysaccharide of Streptococcus agalactiae.

A monoclonal antibody giving a dominant reaction with the group-specific polysaccharide of streptococcus group B in an ELISA test has been developed. The purified polysaccharide exhibited a high positivity with reference anti B streptococcal antiserum in the ELISA test. Cross-tests of antibodies with other groups of streptococci provided a minimum cross-reaction only in the case of G streptococci. Monoclonal antibodies were prepared using Streptococcus agalactiae S 589 MT strain isolated from a case of bovine mastitis which does not express Ia, Ib, II, III, IV and V type antigens, nor C, R and X protein antigens.

The production of K88 antigen by Escherichia coli and Salmonella typhimurium strains with recombinant DNA.

Expression of K88ab antigen by strains with recombinant DNA, differing in molecular weight or promoter, was subjected to investigation. Strains with recombinant DNA produced greater amounts of antigen than did field isolates. Maximal production was recorded for the E. coli C600 strain with 10.85 kb recombinant DNA with promoter P1 of the pBR322 vector. The substitution of promoter Ptac for promoter P1 did not result in an increased expression of K88ab antigen. The production of K88ab antigen by Salmonella typhimurium TM333 with recombinant DNA was on a level comparable to that of E. coli C600 or E. coli HB101 strains with the same recombinant DNA.

Construction of recombinant K88 DNA with Ptac promoter.

The gene encoding K88ab was localized on the 11.6 kb HindIII-HindIII fragment of 74 kb plasmid DNA of E. coli 7301. The smallest recombinant DNA producing the K88ab antigen was obtained by excision of the 5.15 kb EcoRI-EcoRI fragment from recombinant DNA composed of the 11.6 kb K88ab fragment in the pBR322 vector. The size of the smallest fragment was 6.5 kb. Expression of the K88ab antigen was controlled by the P1 promoter of the pBR322 vector. Substitution of promoter Ptac for promoter P1 made it possible to achieve expression of the K88ab antigen by E. coli MT. Substitution of promoter PL for promoter P1 failed to achieve expression of the K88ab antigen in the recipient strains used.