Molecular subtyping of Staphylococcus aureus isolates from cases of bovine mastitis in Brazil (original) (raw)
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Brazilian Journal of Microbiology, 2001
Contamination of fresh milk with Staphylococcus aureus was assessed comparatively through routine phenotypic (coagulase tube test and coagulase slide test) and genotypic (PCR) screening of 128 S. aureus strains isolated from 555 milk samples. These samples were collected from 362 cows with subclinical mastitis, hosted in different dairy herds at various locations of the Northern and Northeastern rural areas of the State of Rio de Janeiro, 39.7% of which were CMT-positive. All S. aureus isolates tested positive for the presence of the coagulase gene by PCR and the isolates could be grouped into four distinct classes according to the size of the PCR product. The strains also yielded variable results when assayed with coagulase test. Taken together, these data indicate the existence of extensive polymorphism at the coagulase gene locus in the genus Staphylococcus and exemplifies the extent of molecular and phenotypic heterogeneity associated with the strains circulating in rural herds.
Acta Scientiarum. Biological Sciences, 2010
Evaluation of a simplified key for the identification of coagulase-positive Staphylococcus isolated from bovine mastitis. Three hundred fourty four strains of coagulasepositive Staphylococcus (CPS), isolated from mastitis cases, underwent phenotypic and genotypic tests to evaluate the efficiency of a simplified key, based on phenotypic tests for the discrimination of these microorganisms. The tests consisted of amplification of the femA gene and hemolysis in blood agar, production of acetoin and fermentation of maltose, mannitol and trehalose. Strains that showed negative results in the amplification test of the femA gene or that were not identified as Staphylococcus aureus (S. aureus) by phenotypic tests were tested with the APISTAPH kit (Biomériux-France), for precise identification of species. Phenotypic tests revealed 338 strains (98.25%) as S. aureus, three strains (0.86%) as Staphylococcus hyicus, and three microorganisms (0.86%) as Staphylococcus intermedius. PCR demonstrated that 338 (98.25%) strains belonged to the S. aureus species, confirming the results for 336 strains from 338 identified, through a simplified phenotypic key. A high rate of correlation (98.83%) was verifeid between the results of genotypic and phenotypic tests for the identification of S. aureus, demonstrating the applicability of the proposed key, for the discrimination of this microorganism in CPS isolated from bovine mastitis.
2006
Mastitis is a disease of dairy cattle worldwide that causes decreased milk quality/yield, resulting in major economic losses. Mastitis is caused by a wide variety of organisms, including Staphylococcus aureus as a major pathogen. The purpose of this research was to characterize 20 Staphylococcus spp. isolates from the mammary gland of mastitic, lactating, backyard cattle, using both the Random Amplified Polymorphic DNA (RAPD) analysis, and sensitivity tests to 13 antibiotics and 1 detergent. Sensitivity tests to 13 antibiotics showed that all isolates were resistant to penicillin but sensitive to cephalothin, erythromycin, and trimethoprim. All isolates were susceptible to a 90% similarity in the RAPD band pattern, while isolates SA35 & SA36 were identical to each other. Nevertheless, all other isolates showed different amplification profiles. Isolates SA7 & SA8, SA30 & SA32, SA35 & SA36 had similar antibiotic resistance/RAPD patterns, suggesting re-isolation. Staphylococcus isolate...
The Veterinary Journal, 2007
The genetic relatedness of coagulase (coa) positive Staphylococcus aureus isolated from cows with subclinical mastitis in Turkey was investigated by polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) analysis. Among 700 milk samples positive in the California Mastitis Test (CMT), species specific PCR identified 200 (28.6%) isolates as S. aureus and 161 (80.5%) of these isolates were positive for the 3 0 end of the coa gene by PCR. Most isolates (n = 135, 83.9%) produced a single band on coa PCR, with molecular sizes ranging from 500 to 1400 bp, whereas a small number of isolates (n = 26, 16.1%) yielded two amplification products. Coa RFLP analysis using AluI and Hin6I revealed 23 and 22 band patterns, respectively. The detection of double bands by coa PCR, previously reported in human isolates, suggests that milking personnel can play a role in the transmission of S. aureus.
Turkish Journal of Veterinary & Animal Sciences
The objective of this work was to study the protein patterns, plasmid profiles, and antibiotic susceptibility of Staphylococcus intermedius and Staphylococcus aureus isolates originating from mastitic mammary glands of dairy cattle in different parts of Konya province. A total of 114 Staphylococcus species were isolated and identified by conventional bacteriological methods from bovine mastitis. Of the total isolates 77 were identified as S. aureus and 37 as S. intermedius. Intra-and inter-species diversities in the coagulase-positive staphylococci were investigated by analysis of whole-cell protein profiles using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Plasmid profiling also demonstrated that 75 S. aureus isolates and 36 S. intermedius isolates contained plasmid. In addition, 88.3% of S. aureus and 59.4% of S. intermedius isolates were resistant to penicillin. Sixty-six of the 77 S. aureus isolates were also resistant to amoxicillin+clavulanic acid (85.7%). The corresponding number for S. intermedius was 17 (45.9%). Only 1 S. aureus isolate was resistant to danofloxacin. One of each of the Staphylococcus isolates was resistant to methicillin. Results from the study showed that the susceptibility of S. intermedius isolates to antibiotics used widely in mastitis therapy is a matter of concern.
Genotypic analysis of Staphylococcus aureus from milk of dairy cows with mastitis in Argentina
Epidemiology and Infection, 2001
Staphylococcus aureus is the most prevalent pathogen causing mastitis of dairy ruminants. This study was developed to ascertain the genotypes and genealogical relationship among strains isolated from milk of bovines with mastitis in Argentina. Molecular epidemiological analysis of S. aureus was performed on 112 isolates from 21 districts. Clonality was assessed by SmaI pulsed-field gel electrophoresis (PFGE) typing, automated EcoRI ribotyping and restriction enzyme analysis of plasmid (REAP) DNA profiles. A total of 22 band patterns distributed in four clusters were found by SmaI PFGE analysis. The similarity of clusters 2, 3 and 4 with cluster 1 was 0n73, 0n69 and 0n33, respectively, and 101 of 112 isolates belonged in cluster 1. PFGE band patterns from 42 isolates within cluster 1 were indistinguishable from each other (type A). The second largest group of isolates with indistinguishable PFGE band patterns was subtype A11, which was composed of 19 isolates. Automated ribotyping assigned the 112 isolates into 13 ribotypes. Among these, the most prevalent ribotypes I and VI were composed of 49 and 35 isolates respectively. Although there was certain correspondence between PFGE genotypes and ribotypes, further discrimination was achieved by combining both methods. REAP DNA profile analysis was useful to provide even further discrimination between isolates with identical PFGE genotype and ribotype. The most prevalent S. aureus strains A\I and A11\VI were widely distributed in the country and were not restricted to individual nearby locations. Prevalence of these two strains varied consecutively within a period of 8 years. Whether the shift in type prevalence was due to selection of a phenotypic trait remains undisclosed.
Comparative Clinical Pathology, 2010
The aim of this study was the genotypical characterization of 58 Staphylococcus aureus isolates from nine dairy herds in the Tabriz and Urmia regions of east and west Azerbaijan provinces, Iran. In this study, 58 S. aureus isolates from 370 milk samples from cows with clinical and subclinical mastitis were analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the aroA gene. Amplification of the aroA gene resulted in a single amplicon with a size of approximately 1,153 bp from all 58 isolates of S. aureus. To obtain the RFLP patterns of the isolates, the PCR products were digested with TaqI restriction enzyme and the fragments separated by gel electrophoresis. Four distinct RFLP patterns were observed among the studied isolates. Three out of four detected genotypes had the same RFLP patterns (A, B, and N) as reported by previous studies. The fourth newly detected genotype in this study was named H. Genotypes A and B were the most frequent, being observed in 24 (41.38%) and 29 (50%) isolates, respectively. Genotypes N and H comprised 1.7% and 6.9% of all isolates, respectively. With the exception of the RFLP pattern N, which was observed only in the Tabriz region, all other patterns were found in both Tabriz and Urmia regions. The results demonstrate that strain variations of S. aureus could occur within and between herds and also between different regions, although a few genotypes of S. aureus were predominant in bovine mastitis. This study also indicated that PCR amplification of the aroA gene is specific for S. aureus identification.
Indian Journal of Comparative Microbiology, Immunology and Infectious Diseases, 2016
To understand the prevalence and characteristics of Staphylococci causing goat mastitis in Algeria, mastitis milk samples were collected and cultured on both blood agar and mannitol salt 2 agar medium. Staphylococci isolates were identified by API-Staph gallery and tested for antibiotic susceptibility by disk diffusion method. DNA microarrays analysis was performed on S. aureus strains in order to detect virulence factors, including toxins, and to assign the isolates to theirs MLST clonal complexes. Overall, 86/266 (32.3%) goats suffered from mastitis, whose 32.5% (28/86) infected with Staphylococci. Most of strains were susceptible to many antibiotics, except for penicillin, tetracycline and fusidic acid, and the resistance incidence of which were 21.4%, 25%, and 10.7% respectively. Only twelve (42.8%) isolates were resistant to utmost two antibiotics and bi-drug resistance was associated to four Coagulase negative Staphylococci (CNS) strains. Nine different species were identified in CNS isolates while S. aureus strains were clustered within CC130-agr3-[lukF-P83, lukM+] (four isolates; 50%), CC6-agr1 (three isolates; 37.5%) and one agr4 non-typables cluster. Enterotoxins genes were found in 62.5% of the S. aureus strains. Of which sec, sel were the most prevalent (62.5%: n=5) followed by sea (12.5%: n=1). CC130 showed edinB, tst and tetK (2/4) genes and CC6 were positive to cna and fosB genes. The current investigation provide the data for prevalence of Staphylococci in goats in Tizi-Ouzou region and S. aureus characterization which will help in tracking evolution of epidemic strains and their control methods.