MULTIPLEX-PCR FOR DETECTION OF β-LACTAM RESISTANCE IN Staphylococcus spp.

Nakadomari, Giovana Hashimoto; Charalo, Amanda Carmen; Pavan, Ana Claudia Lemes; Vignoto, Vanessa Kelly Capóia; Sfaciotte, Ricardo Antonio Pilegi; Wosiacki, Sheila Rezler

doi:10.4025/revcivet.v6i2.45050

Cited by 3 publications

(3 citation statements)

References 16 publications

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“…To detect the beta-lactam resistance genes (mecA, mecC and blaZ) present in MRS isolates, Nakadomari et al (2019) multiplex PCR technique was performed. Staphylococcus aureus ATCC 43330 reference strain was used as a positive control to all genes.…”

Section: Methodsmentioning

confidence: 99%

Detection of the main multiresistant microorganisms in the environment of a teaching veterinary hospital in Brazil

et al. 2021

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Contamination of the veterinary hospital environment with multiresistant pathogens endangers not only hospitalized animals, but also the workplace safety of veterinarians and nurses, animal guardians and, when in case of a teaching hospital, veterinary students. The objective of this study was to map the main points of bacterial contamination of a veterinary teaching hospital in Brazil to identify multiresistant microorganisms and their antimicrobial resistance genes. Samples were collected from 39 different locations of a veterinary school hospital which comprised a pool according to each hospital environment. In certain environments, more than one pool has been collected. All samples were collected in quadruplicates for the selective isolation of the main multiresistant microorganisms: methicillin-resistant Staphylococcus (MRS), vancomycin resistant Enterococcus (VRE), cephalosporinases and/or extended-spectrum beta-lactamase-producing Gram-negative bacteria (ESBL) and Carbapenemase-producing (CP). After isolation and identification of isolates, multiplex-PCR reactions were performed to detect the main genes for each microorganism and antimicrobial susceptibility tests with the main antibiotics used for each bacterial group according to CLSI. Of the 39 veterinary teaching hospital sites collected, all (100%) had at least one of the microorganisms surveyed, and 17.95% (n=7) of the sites were able to isolate the four pathogens. From the 94 pools collected, it was possible to isolate MRS in 81.91% (n=77), VRE in 12.77% (n=12), cephalosporinases and/or ESBL in 62.77% (n=59) and CP in 24.47%. (n=23). Regarding MRS, the mecA gene was detected in all isolates. All isolated VREs were identified as Enterococcus faecalis and presented the vanA gene. Regarding cephalosporinases and/or ESBL, 89.83% (n=53) of the isolates presented the blaTEM gene, 57.63% (n=34) the blaOXA-1 gene, 37.29% (n=22) blaCTX-M gene from some group (1, 2, 9 ou 8/25) and 20.34% (n=12) the blaSHV gene. It was possible to identify the main microorganisms responsible for causing nosocomial infections in humans (VRE, MRS, ESBL and CP) in the veterinary hospital environment, suggesting a source of infection for professionals and students of veterinary medicine, placing a high risk for public health.

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Section: Methodsmentioning

confidence: 99%

Detection of the main multiresistant microorganisms in the environment of a teaching veterinary hospital in Brazil

et al. 2021

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“…A amplificação foi realizada em termociclador nas seguintes condições de tempo e temperatura: primeira etapa de desnaturação à 94°C por 7 minutos seguidos de 40 ciclos de 94°C por 1 minuto, 54°C por 1 minuto e 72°C por 1 minuto, terminando por uma etapa de extensão final à 72°C por 7 minutos. Os produtos amplificados foram submetidos à eletroforese em gel de agarose a 1,5% corado com SYBR® safe DNA e gel stain e visualizado sob luz ultravioleta (NAKADOMARI et al, 2019). A contagem de colônias características de Staphylococcus spp.…”

Section: Methodsunclassified

EFEITOS DE REGULADORES VEGETAIS NA PRODUTIVIDADE BIOLÓGICA DE PLANTAS DE SOJA (Glycine max (L.) Merrill)

Campos¹,

Ono²

2019

A Face Multidisciplinar Das Ciências Agrárias 3

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Todo o conteúdo deste livro está licenciado sob uma Licença de Atribuição Creative Commons. Atribuição 4.0 Internacional (CC BY 4.0).O conteúdo dos artigos e seus dados em sua forma, correção e confiabilidade são de responsabilidade exclusiva dos autores. Permitido o download da obra e o compartilhamento desde que sejam atribuídos créditos aos autores, mas sem a possibilidade de alterá-la de nenhuma forma ou utilizá-la para fins comerciais.

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“…As a result, this assay is more accurate, especially for highly diverse and rapidly evolving pathogens, and time saving with a lower operational cost. The primers and amplification conditions used in this assay must be carefully designed and properly evaluated by electrophoresis so that all primers work at the same annealing temperature, and amplification products must be of different molecular sizes [41][42][43]. Therefore, the objectives of this research were to establish a sensitive, rapid, and specific multiplex PCR assay for accurate detection of R. solani AG-3 and to apply multiplex PCR technology for the detection of R. solani AG-3 in soil and potato tubers.…”

Section: Introductionmentioning

confidence: 99%

Establishment and Application of a Multiplex PCR Assay for the Rapid Detection of Rhizoctonia solani Anastomosis Group (AG)-3PT, the Pathogen Causing Potato Black Scurf and Stem Canker

et al. 2022

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Rhizoctonia solani anastomosis group 3 (AG-3) is the main causative agent of the soil-borne disease known as potato black scurf, which poses a huge threat to potato production. Rapid and accurate identification of R. solani AG-3 isolates in soil and potato seed tubers prior to planting is essential for good production. In this study, a multiplex PCR assay was established for the detection of R. solani AG-3. Two pairs of target-specific primers were designed from sequences for endopolygalacturonase and pyridoxine biosynthesis genes downloaded from GenBank. The main factors influencing PCR amplification, such as annealing temperature and primer concentration, were optimized. Results show that the proposed multiplex PCR assay is highly sensitive and specific for the target genes in the pathogen even when the DNA concentration is reduced to 20 fg/μL. The resulting calibration plot shows a linear relationship between electrophoretic band peaks and genomic DNA concentration (R2 = 0.98). The primer specificity was confirmed by applying them to other R. solani AG groups and plant pathogen species on which no amplicons were produced. Using the primers, we successfully detected small amounts of R. solani AG-3 present in soil and potato tuber samples. Taken together, the detection assay developed in this study has high sensitivity, strong specificity, and accuracy and can be used to detect and identify soil and potato seed tubers infected with Rhizoctonia solani AG-3.

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MULTIPLEX-PCR FOR DETECTION OF β-LACTAM RESISTANCE IN Staphylococcus spp.

Cited by 3 publications

References 16 publications

Detection of the main multiresistant microorganisms in the environment of a teaching veterinary hospital in Brazil

Detection of the main multiresistant microorganisms in the environment of a teaching veterinary hospital in Brazil

EFEITOS DE REGULADORES VEGETAIS NA PRODUTIVIDADE BIOLÓGICA DE PLANTAS DE SOJA (Glycine max (L.) Merrill)

Establishment and Application of a Multiplex PCR Assay for the Rapid Detection of Rhizoctonia solani Anastomosis Group (AG)-3PT, the Pathogen Causing Potato Black Scurf and Stem Canker

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