Standardization of a multiplex PCR for the identification of coagulase-positive Staphylococcus

Gandra, Eliézer Ávila; Fernandez, Maria Aparecida; Silva, Jorge Adolfo; Silva, Wladimir Padilha da

doi:10.1590/s0101-20612011000400019

Cited by 9 publications

(3 citation statements)

References 6 publications

(11 reference statements)

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“…Coagulase-positive Staphylococci ( S. aureus , Staphylococcus intermedius , and Staphylococcus hyicus ) were identified based on morphology, pigmentation, coagulase test, Voges Proskauer, and the ONPG test ( Capurro et al, 1999 ). Specific polymerase chain reaction (PCR) tests were used to confirm S. aureus strains using primers NUC1 and NUC2 described by ( Gandra et al, 2011 ) ( Supplementary Figure S1A ).…”

Section: Methodsmentioning

confidence: 99%

Antibacterial Effect of Copper on Microorganisms Isolated from Bovine Mastitis

et al. 2016

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The antimicrobial properties of copper have been recognized for several years; applying these properties to the prevention of diseases such as bovine mastitis is a new area of research. The aim of the present study was to evaluate in vitro the antimicrobial activity of copper on bacteria isolated from subclinical and clinical mastitis milk samples from two regions in Chile. A total of 327 microorganisms were recovered between March and September 2013, with different prevalence by sample origin (25 and 75% from the central and southern regions of Chile, respectively). In the central region, Escherichia coli and coagulase negative Staphylococci (CNS) were the most frequently detected in clinical mastitis cases (33%), while in the southern region S. uberis, S. aureus, and CNS were detected with frequencies of 22, 21, and 18%, respectively. Antibiotic susceptibility studies revealed that 34% of isolates were resistant to one or more antibiotics and the resistance profile was different between bacterial species and origins of isolation of the bacteria. The minimum inhibitory concentration of copper (MIC-Cu) was evaluated in all the isolates; results revealed that a concentration as low as 250 ppm copper was able to inhibit the great majority of microorganisms analyzed (65% of isolates). The remaining isolates showed a MIC-Cu between 375 and 700 ppm copper, and no growth was observed at 1000 ppm. A linear relationship was found between the logarithm of viable bacteria number and time of contact with copper. With the application of the same concentration of copper (250 ppm), CNS showed the highest tolerance to copper, followed by S. uberis and S. aureus; the least resistant was E. coli. Based on these in vitro results, copper preparations could represent a good alternative to dipping solutions, aimed at preventing the presence and multiplication of potentially pathogenic microorganisms involved in bovine mastitis disease.

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Section: Methodsmentioning

confidence: 99%

Antibacterial Effect of Copper on Microorganisms Isolated from Bovine Mastitis

et al. 2016

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“…Specific primers were designed for amplifying a segment of the nuc gene (a gene encoding the thermostable endonuclease of coagulase-positive Staphylococcus ) by real-time PCR sybr green I. The gene sequence for each of the three Staphylococcus species considered in this study was aligned using the GenBank/NCBI ( 26 ) software for identification of sequence homologies shared among the nuc gene ( 27 ). We used the forward and reverse primers described by Sasaki et al ( 28 ) for the in silico search.…”

Section: Methodsmentioning

confidence: 99%

Molecular Detection and Characterization of the mecA and nuc Genes From Staphylococcus Species (S. aureus, S. pseudintermedius, and S. schleiferi) Isolated From Dogs Suffering Superficial Pyoderma and Their Antimicrobial Resistance Profiles

et al. 2020

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Canine superficial pyoderma (CSP) is a bacterial infection secondary to several skin diseases of the dog. Staphylococcus pseudintermedius, which is a commensal bacterium of the dog's skin, is the leading agent found in dogs affected by CSP, which can progress to deep pyoderma. It is also of clinical significance because S. pseudintermedius strains carry antimicrobial resistance genes, mainly the mecA gene. In this descriptive longitudinal study, molecular characterization of bacterial isolates from dogs affected by CSP was performed in addition to phenotyping, antimicrobial profiling, and assessment of resistance carriage status. Fifty dogs (24 females and 26 males) attending the CES University Veterinary Teaching Hospital were included in the study. CSP was confirmed according to clinical signs and cytological examination. Swabs were taken from active skin lesions for bacterial culture, and phenotyping and antimicrobial resistance profiles were assessed using API-Staph phenotyping and the Kirby-Bauer method, respectively. We also performed molecular detection and characterization of the mecA and nuc encoding gene of coagulase-positive Staphylococci. The mecA gene frequency was established by qPCR amplification of a 131bp gene fragment. Data were evaluated by descriptive statistics. Erythema, peeling, pruritus, and alopecia were the predominant symptoms (72, 56, and 46%, respectively). We isolated bacteria compatible with Staphylococcus species from all samples tested. González-Domínguez et al. mecA/nuc Genes From Staphylococcus Species API phenotyping showed 83.1 to 97.8% compatibility with S. pseudintermedius. PCR-genotyping resulted in 15, 3, and 1 isolates positive for S. pseudintermedius, S. aureus, and S. schleiferi, respectively. Isolated strains showed high susceptibility to Imipenem, Ampicillin/Sulbactam, and Rifampicin (100, 94, and 92%, respectively). The highest resistance was against Vancomycin and Trimethoprim/Sulfamethoxazole (98 and 74%, respectively). S. pseudintermedius, S. aureus, and S. schleiferi isolates were cloned and shared 96% sequence homology. Finally, we found 62% carriage status of the mecA gene in isolates of CSP patients, although only 36% of the isolates were methicillin-resistant. Identification of three Staphylococcus species causing CSP, high-level resistance against conventional antimicrobials, and carriage of the mecA gene highlight the importance of performing molecular characterization of bacteria causing dermatological conditions in dogs.

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“…The development of PCR-based methods for the detection of pathogens or virulence factors (Shahi et al, 2013;Branquinho & Ferreira, 2012;Gandra et al, 2011) can be used for the initial screening of the presence of microorganisms in samples of milk and dairy products. Furthermore, the present study, through direct isolation of bacterial DNA in milk samples, aimed to develop the multiplex PCR to assess the prevalence of E. coli and STEC in samples of fresh milk.…”

Section: Molecular Characterization Of Stec In Milk Samplesmentioning

confidence: 99%