Assessment of potentially probiotic properties of Lactobacillus strains isolated from chickens.

This study was performed in order to isolate lactobacilli from chicken droppings and to select strains with the most promising probiotic properties. Lactobacillus strains were isolated from a flock of healthy laying hens. The first selection criterion was the ability to inhibit the growth of Salmonella Enteritidis. Then the tolerance to low pH and bile salt, the ability to coaggregate with pathogenic bacteria and hydrogen peroxide production were evaluated. Four isolates showing the best antagonistic activity against Salmonella Enetritidis were selected for further research. All isolates tested tolerated low pH and bile salt, likewise all produced hydrogen peroxide. They efficiently coaggregated with C. perfringens and relatively less with E. coli. Isolate 03'04 displayed above-average results in all criteria, thus it is considered as a potential probiotic for chickens, and will be further evaluated for health promoting effect in animals. The results presented in this study confirm the strain specific probiotic properties and prove the probiotic potential of isolate 03'04. Strong antagonistic properties against C. perfringens exhibited by certain Lactobacillus strains indicate the possibility to use them as a component of probiotic supplement in necrotic enteritis of poultry.

Genetic characterization of coagulase-positive staphylococci isolated from healthy pigeons.

Coagulase-positive staphylococci (CoPS) are opportunistic veterinary pathogens, of which Staphylococcus aureus, S. delphini and S. intermedius can be isolated from pigeons. The biochemical identification of S. delphini and S. intermedius isolates may be incorrect, because of their phenotypic similarity. The purpose of the present study was to isolate and identify CoPS from domestic and feral pigeons and to determine their genetic relatedness by PFGE. A total number of 31 isolates of CoPS were obtained, 15 were identified as S. delphini group B, six as S. aureus, four as S. delphini group A, three as S. intermedius and three as S. schleiferi subsp. coagulans. The results indicate that S. delphini group B is the predominant CoPS species among pigeons studied. PFGE restriction patterns of S. delphini group A and S. delphini group B form separate clusters, demonstrating their genetic heterogeneity. Indistinguishable or very similar PFGE patterns observed among S. delphini group B isolates from domestic and feral pigeons confirm the possibility of CoPS transmission between these birds.

PFGE and AFLP genotyping of Staphylococcus aureus subsp. anaerobius isolated from goats with Morel's disease.

Staphylococcus aureus subsp. anaerobius is the etiological agent of the Morel's disease in sheep and goats. The disease presents with subcutaneous abscesses, located mainly in the superficial lymph nodes. Forty-one isolates of S. aureus subsp. anaerobius were collected from two outbreaks of the Morel's disease in Poland in years 2006-2008. Analysis of DNA SmaI digests by PFGE showed that 35 of 41 isolates belonged to the same PFGE type, identical to the type strain of S. aureus subsp. anaerobius ATCC 35844, confirming high level of clonality of the species. The DNA patterns of the remaining identical 6 isolates, different from the reference strain only by two bands, were found closely related. Genotyping performed with AFLP technique revealed two clonal groups including 16 and 25 isolates, respectively. The study indicated that AFLP technique might be a better discriminatory tool for genetic analysis of S. aureus subsp. anaerobius isolates, when compared to PFGE.

Antibiotic resistance of canine Staphylococcus intermedius group (SIG)--practical implications.

A total of 221 SIG strains were isolated from clinical samples of canine origin submitted to the Diagnostic Laboratory of the Division of Bacteriology and Molecular Biology at the Warsaw University of Life Sciences in Warsaw during the period 2006-2010. The aim of the study was to investigate the frequency of prevalence of methicillin-resistant SIG strains and to determine the MIC values of cephalotin, amoxicillin/clavulanic acid, ciprofloxacin, clindamycin, gentamicin, chloramphenicol, mupirocin for a collection of randomly selected 79 strains belonging to Staphylococcus intermedius group (SIG), including 23 mecA-positive and 56 mecA-negative strains. All isolates were identified as belonging to SIG based on their phenotypic properties and PCR amplification of S. intermedius-specific fragment of the 16S rRNA gene. The mecA gene was detected in 26 (12%) of 221 SIG strains. All tested mecA-negative SIG strains were susceptible to amoxicillin/clavulanic acid and cephalotin. One of the 56 mecA-negative SIG strains was resistant to ciprofloxacin, six (11%) to gentamicin. It was found that sixteen (29%) of 56 mecA-negative SIG strains were resistant to clindamycin. Most of the mecA-positive SIG strains were resistant to ciprofloxacin (96%), clindamycin (96%), and gentamicin (96%). Only one MRSIG strain was resistant to chloramphenicol. All examined mecA-positive SIG strains were found to be susceptible to mupirocin. Our results imply that staphylococcal multidrug resistance has become more prevalent, which could lead to difficulties in effective treatment. With some resistant strains the only therapeutic possibility are antimicrobial agents important in human medicine. New regulations for veterinary medicine concerning appropriate therapy of infections caused by multidrug-resistat staphylococci are needed.

The first description of gastric Helicobacter in free-ranging wild boar (Sus scrofa) from Poland.

Specimens of gastric mucosa of 17 free-ranging wild boars (Sus scrofa) shot in the Central Poland during 2007/2008 hunting season were investigated for the presence of Helicobacter species. Histopathology, Helicobacter genus-specific 16S rRNA PCR, and DNA sequence analysis were employed. In PCR analysis the presence of Helicobacter's DNA was detected in one stomach. Obtained sequence analysis showed its relatedness to Helicobacter heilmannii type 2. In histopathology of the PCR-positive sample the presence of tightly coiled spiral bacteria was detected on the surface of the antral mucosa, in gastric pits and lumen of the upper parts of antral glands. Potential pathologic significance of the presence of Helicobacter in the stomach of free-ranging wild boars was obscured by the parasitic invasion-caused gastritis, and remains unknown.

Protective effect of potentially probiotic Lactobacillus strain on infection with pathogenic bacteria in chickens.

The probiotic potential of a Lactobacillus salivarius 3d strain isolated from chicken faeces was assessed in one day old chickens. Lactobacillus salivarius 3d was administered per os at a concentration of 10(8) cfu in 100 microl of PBS. The chickens were then challenged with pathogenic bacteria: Salmonella Enteritidis, Campylobacter jejuni and Clostridium perfringens. Samples of caecal contents and livers were collected after 1, 2, 3, 7 and 14 days after infection. Lactobacilli and pathogenic bacterial cell counts were determined in the samples. This study showed that L. salivarius 3d reduced the number of Salmonella Enteritidis and Clostridium perfringens in the group of chickens treated with Lactobacillus. Therefore it may be concluded that L. salivarius 3d may be used as a potential probiotic for chickens.

Antibiotic resistance patterns and occurrence of mecA gene in Staphylococcus intermedius strains of canine origin.

We have evaluated 102 Staphylococcus intermedius isolates of canine origin for susceptibility to antimicrobial primary agents, i.e. penicillin, amoxicillin, amoxicillin with clavulanic acid, cefuroxime, trimethoprim/sulfonamides, neomycin, streptomycin, gentamicin, norfloxacin, tetracycline, vancomycin, erythromycin and secondary agents, i.e., chloramphenicol, ciprofloxacin, lincomycin, teicoplanin, rifampicin, imipenem, mupirocin. Antimicrobial sensitivity was examined using the disk diffusion method and performed according to NCCLS quidelines. Methicillin resistance was detected using the disk diffusion method with oxacillin, and the occurrence of mecA gene was detected by PCR. Resistance to streptomycin, penicillin, amoxicillin, neomycin, followed by tetracycline was predominant. From 14 mecA-positive strains, 12 were multidrug-resitant, and the remaining two showed atypical susceptibility. One strain resistant to oxacillin in the disc diffusion method was mecA-negative, suggesting a different mechanism of resistance. Our results indicate that the emergence of S. intermedius resistance to methicillin may be underestimated. In case of clinical multidrug-resitant S. intermedius isolates, resistance to methicillin should be considered.

Effect of Helicobacter sp. infection on the number of antral gastric endocrine cells in swine.

The aim of the present study was to evaluate the effect of Candidatus Helicobacter suis (CHS) and other Helicobacter sp. different from Candidatus Helicobacter suis (non-Candidatus Helicobacter suis, non-CHS) infection on the number of endocrine G and D cells and G/D cells ratio in antral gastric mucosa in swine. Twenty nine stomachs were obtained from clinical healthy pigs about 6 months old and weighing approximately 100-120 kg after slaughter at abattoir located in central Poland. From each stomach samples of the antral gastric mucosa were taken for histopathology, and PCR examination for presence of Helicobacter genus and Candidatus Helicobacter suis. Samples for histopathology and immunohistochemistry were fixed in 10% buffered formalin. To reveal the expression of gastrin- and somatostatin-producing cells specific antibodies were used. Selected endocrine cells were counted in the midzone of pyloric glands, the results were expressed as a mean of the number of immunoreactive cells in one microscopic field, and as the ratio of gastrin to somatostatin cells (G/D). It can be concluded that some species of swine Helicobacter can alter the number of endocrine cells in gastric antral mucosa. Some of these alterations, for example increase the number of G cells, decrease of the D cells and especially increase of ratio G to D cells can be responsible for development of gastroesophageal ulcers in swine.

Molecular characterization of Staphylococcus pseudintermedius strains isolated from clinical samples of animal origin.

The aim of this study was to determine the species distribution among 44 randomly selected clinical isolates (30 mecA-positive and 14 mecA-negative) of animal origin previously identified as Staphylococcus intermedius by phenotypic tests and species-specific PCR amplification of the 16S rRNA gene. For this purpose, we used a multiplex PCR for the detection of the nuc gene and restriction fragment length polymorphism analysis of pta gene amplified by PCR. Both methods allow discrimination of Staphylococcus pseudintermedius from the other closely related members of the S. intermedius group and other coagulase-positive staphylococci isolated from animals. Genetic diversity of S. pseudintermedius strains was analyzed by staphylococcal protein A-encoding gene (spa) typing. Multiplex PCR method was used to identify staphylococcal cassette chromosome mec (SCCmec) type in mecA-positive strains. All isolates previously identified as S. intermedius were shown to belong to S. pseudintermedius. According to PCR-based SCCmec typing, SCCmecIII was the most prevalent type (n = 23), and solely seven isolates were designated as non-typeable. Furthermore, the assessment of spa-typing results revealed that the majority of all strains (n = 27) harbored spa type t02, and 17 strains were classified as non-typeable.