Một xét nghiệm Polymerase Spiral Reaction mới để phát hiện nhanh các gen Staphylococcus aureus kháng penicillin

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Tác giả: Hữu Nghĩa Đặng, Hồng Phúc Nguyễn, Văn Thạch Phan, Hồng Diễm Trần, Thị Quỳnh Như Trần, Quang Hiếu Vũ

Ngôn ngữ: vie

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Thông tin xuất bản: Tạp chí Khoa học và Công nghệ - Đại học Nguyễn Tất Thành, 2023

Mô tả vật lý: tr.20

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ID: 340486

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Bài báo đã phát triển phương pháp trực quan phản ứng phân tử nhằm phát hiện chủng Staphylococcus aureus kháng penicillin dựa trên phản ứng xoắn ốc polymerase (PSR). Phương pháp này thực hiện phản ứng trong điều kiện đẳng nhiệt bằng cách sử dụng enzyme Bst DNA polymerase và hai cặp mồi được thiết kế nhắm mục tiêu vào gen blaZ mã hóa tính kháng penicillin ở S. aureus. Khi khảo sát điều kiện phản ứng cho thấy, ở nhiệt độ 65 °C và nồng độ cặp mồi F2/R2 = 0,1 µM và F1-Nr/R1-N = 1,6 µM, thì phản ứng PSR cho kết quả tối ưu chỉ trong vòng 50 phút. Giới hạn phát hiện của phương pháp PSR với DNA bộ gen và tế bào vi khuẩn lần lượt là 10 pg/μL và 5 CFU/mL trên mỗi phản ứng, nhạy hơn so với phương pháp PCR. Ngoài ra, phương pháp này có tính đặc hiệu cao, do không xảy phản ứng chéo với các chủng vi khuẩn gây bệnh khác. Kết quả cho thấy phương pháp xét nghiệm PSR dễ thực hiện, có độ tin cậy cao
nên được ứng dụng để kiểm tra và khảo sát thực trạng kháng penicillin của S. aureus trong bệnh viện và cộng đồng.In this study, a visual method was developed to detect and identify penicillin-resistant Staphylococcus aureus (PRSA) based on the polymerase spiral reaction (PSR). This method can be performed under isothermal conditions using Bst DNA polymerase and two-pair primers specifically designed to target the blaZ gene, which encodes penicillin-resistance in S. aureus. Investigation of reaction conditions showed that at the temperature of 65 °C and the concentration of primer pairs F2/R2 = 0.1 µM and F1-Nr/R1-N = 1.6 µM, the reaction gave optimal PSR result within only 50 minutes. The detection limit of PSR in genomic DNA and bacterial cells were 10 pg/μL and 5 CFU/mL, respectively, which were more sensitive than with standard PCR. In addition, this method was shown to be highly specific, as non-PRSA bacteria negligibly interfered with PRSA detection. These results indicated that a novel, effortless and reliable PSR assay developed in this study has potential application in the screening for penicillin-resistant of S. aureus in hospitals and communities.

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