Determination of antibiotic resistance profiles and biofilm production of Staphylococcus spp. isolated from Anatolian water buffalo milk with subclinical mastitis.

Mastitis is one of the most crucial diseases of dairy animals. Especially subclinical mastitis (SCM) has negative impacts on of dairy economy in term of reducing milk quality and quantity also premature culling and cost of therapy. Staphylococci are important etiological agents in SCM. The aim of the study was to investigate the biofilm production and antibiotic resistance profiles of Staphylococcus spp. other than S. aureus isolated from milks of Anatolian water buffalo with subclinical mastitis. Twenty-two coagulase negative staphylococci (CNS) identified phenotypically were also identified with PCR as Staphylococcus spp. other than S. aureus. Biofilm productions were investigated both by Congo Red Agar Method and PCR. The antibiotic resistance profiles of the isolates were determined by Disc Diffusion Method and they were antibiotyped. Only three (13.6%) isolates were biofilm positive both phenotypically and genotypically. All isolates except for two were resistant against at least two antibiotics. Multidrug-resistance among the isolates was low (13.6%). Antibiotyping results showed that the similarities among the strains were between 30-100%. Genotyping of the strains revealed that a genetic heterogeneity was found among CNS isolates and their similarities were between 43% and 93%. In conclusion, CNS isolates identified as subclinical mastitis agents in buffaloes showed a high antibiotic resistance profile especially against oxacillin and vancomycin. Further studies should be conducted to investigate new mechanisms and/or genes responsible for antibiotic resistance in buffaloes.

Molecular analysis and genotyping of methicillin-resistant Staphylococcus aureus strains isolated from different clinical sources.

BACKGROUND: Investigating genetic relatedness between methicillin-resistant Staphylococcus aureus (MRSA) strains from humans and different animal species may clarify the epidemiological characteristic of MRSA infections together. AIM: The aim of the study was to perform genotypic characterization and type strains of MRSA isolated from different clinical sources, by molecular techniques. MATERIALS AND METHODS: The molecular characterization of the strains was performed by polymerase chain reaction (PCR), using several specific oligonucleotides. These were as follows: S. aureus species-specific sau gene, mecA gene coding PBP2a responsible for methicillin resistance, femA gene coding for a protein, which influences the level of methicillin resistance of S. aureus, and is universally present in all MRSA strains; spa gene coding for protein A; coa gene coding for coagulase, and blaZ gene coding for the production of beta-lactamase. To determine the genetic diversity of these strains, random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) was performed. RESULTS: Among the 415 S. aureus strains, 61 were phenotypically identified as MRSA, and confirmed as S. aureus by amplification of sau gene. However, 90.16% of the strains were mecA positive, while all were negative for femA gene. The presence and polymorphism of coa and spa genes were investigated and 83.60% and 18.03% strains were positive for coa and spa, respectively. While these strains were grouped into six coa-types by PCR, no polymorphism was found for spa gene among strains having only single 190 bp of the band. bla genes were found in 75.40% of strains. These strains were divided into 12 RAPD types. CONCLUSIONS: The results showed the relatively high heterogeneity and variation of coa gene among MRSA strains, while further studies on sequencing of these strains may identify which sequence type is predominant in this region.

Development and validation of glycoprotein-based native-subunit vaccine for fish against Aeromonas hydrophila.

Aeromonas hydrophila is known to be causative agent of an infection named as Bacterial haemorrhagic septicaemia or red pest in freshwater fish. The aim of this study was to develop and validate the glycoprotein-based fish vaccine against Aeromonas hydrophila. For this aim, after identification and characterization of A. hydrophila isolates from fish farms, one A. hydrophila isolate was selected as vaccine strain. Antigenic glycoproteins of this vaccine strain were determined by Western blotting and glycan detection kit. The connection types of these glycoproteins were examined by glycoprotein differentiation kit. Two glycoproteins, molecular weights of 19 and 38 kDa, with SNA connection type were selected for use in vaccination trials. After their purification by SNA-specific lectin and size-exclusion chromatography, protection studies with purified proteins were performed. For challenge trials, four experimental fish groups were designated: Group I (with montanide), Group II (with montanide and ginseng), Group III [with Al(OH)3 ] and Group IV [with Al(OH)3 and ginseng]. The survival ratings of fish were determined, and protection was calculated as 21.56%, 29.41%, 69.83% and 78.88% in groups I, II, III and IV, respectively. In conclusion, A. hydrophila glycoproteins with Al(OH)3 and ginseng could be used as a safe and effective vaccine for fish.

Characterization of Enterococcus faecalis isolates originating from different sources for their virulence factors and genes, antibiotic resistance patterns, genotypes and biofilm production.

In this study, 72 Enterococcus faecalis isolates originating from humans (n=39), dogs (n=26) and cats (n=7) were investigated for some virulence factors, some virulence genes, antibiotic resistance phenotypes, randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) patterns and biofilm production. Of the isolates, 31 (43.1%) were positive for gelatinase, 11 (15.3%) for aggregation substance and cytolysine, 38 (52.8%) for gelE and 34 (47.2%) for asa1 genes. All isolates were found to be negative for hyl, esp and cylA genes. All isolates were found to be resistant to nalidixic acid and kanamycin. On the other hand, all isolates were cited for susceptible to amoxicillin. Vancomycin resistance genes (vanA, vanB, vanC1/C2 or vanD) have not been detected in any of the phenotypically vancomycin resistant isolates. Isolates from humans, dogs and cats were grouped into 8, 2 and 4 antibiotypes depending upon susceptibilities to 12 different antibiotics. In all human, dog and cat isolates, 9, 12 and 2 genotypes were determined by RAPD-PCR, respectively. Nine (34.6%) of the dog isolates were found to be positive for biofilm production. This study showed that multiple antibiotic resistance among human isolates is more frequent than in dog and cat isolates.