Phát hiện gen mã hóa carbapenemase trên chủng acinetobacter baumannii bằng kỹ thuật lamp

I H C QU C GIA TP HHCM I H C BÁCH KHOA NG PHÁT HI N GEN MÃ HÓA CARBAPENEMASE TRÊN CH NG ACINETOBACTER BAUMANNII B NG K THU T LAMP Chuyên ngành: Công Ngh Sinh H c Mã s : 60420201 LU TP H CHÍ MINH 2021 c hoàn thành t i: Cán b i h c Bách Khoa ng d n khoa h c: TS Nguy -HCM n PGS.TS Nguy Cán b ch m nh n xét 1: TS Nguy Cán b ch m nh n xét 2: TS Hoàng Anh Hoàng Lu cb ov t ih (Online Google Meet) Thành ph n H m: PGS.TS.Nguy n Ti n Th ng Ch t ch TS Hu nh Ng c Oanh Th ký TS Nguy Ph n bi n TS Hoàng Anh Hoàng Ph n bi n PGS.TS.Nguy y viên Xác nh n c a Ch t ch H ng Khoa qu n lý chuyên ngành sau lu CH T CH H c s a ch a (n u có) NG PGS.TS Nguy n Ti n Th ng NG KHOA I H C QU C GIA TP.HCM I H C BÁCH KHOA C NG HÒA XÃ H I CH c l p - T - H nh phúc T NAM NHI M V LU H tên h ng MSHV: 1770181 Chuyên ngành: Công Ngh Sinh H c Mã s : 60420201 TÀI: Phát hi n gen mã hóa carbapenemase ch ng Acinetobacter baumannii b ng k thu t LAMP NHI M V VÀ N I DUNG: n gen mã hóa carbapenemase ch ng Acinetobacter baumannii b ng k thu Áp d c ng s i hai m c tiêu c công b b i Norihisa Yamamoto phát hi n t l gen mã hóa carbapenemase bla OXA-23 khu n l c Acinetobacter baumannii phân l p t m u b nh ph m lâm sàng nh c hi u c a k thu t LAMP vi c phát hi n gen mã hóa carbapenemase bla OXA-23 Acinetobacter baumannii m u b nh ph m lâm sàng NGÀY GIAO NHI M V : 22/02/2021 NGÀY HOÀN THÀNH NHI M V : 13/06/2021 CÁN B NG D N: TS Nguy n ; PGS.TS Nguy Tp HCM, ngày CÁN B TS Nguy CH NHI M B PGS.TS Lê Th Th tháng NG D N n PGS.TS Nguy NG KHOA 21 L IC Tôi xin trân tr ng c t u ki ng h , tơi hồn thành lu Tơi xin trân tr ng c ng TS Nguy vi n Th ng Nh n u ki ng khoa Vi Sinh , B nh ng d n th c nghi m nghiên c u t i B nh vi n Tôi xin trân tr ng c Th y Tiên Công Ngh Sinh H c Th y Cô công tác t i b Ch nhi m b môn tr , gi ng d y giúp tơi hồn thành lu Tơi xin trân tr ng c c b nh vi n Gia An 115, Ban Giám c b nh vi n Th ng Nh cung c tr , u ki n th c nghi Cu i cùng, xin g i l i c i b n nh cho ti p t c ph giúp tơi hồn thành t tài n nh ng nghi ng n bó tơi, ngu ng l c u h c t p cơng tác 2021 H c viên ng TĨM T T LU tv ch ng nghiên c u xu t phát t th c ti n v hi n tr ng nhi m khu n b nh vi tài áp d ng l i quy trình k thu t c a Yamamoto, nh m phát hi n gen bla OXA-23 mã hóa carbapenemase A baumannii Các s li u công b c h c vi n nhóm nghiên c u th c hi n tr c ti p m u b nh ph m c i b nh t i B nh vi n Gia An 115 B nh vi n Th ng Nh li u có tính th c t bám sát vào th c tr ng nhi m khu n t i b nh vi kh xác cao v t c hi u 100% gi i h n phát hi n 10 CFU/mL cho vi c phát hi n gen bla OXA-23 K t qu c m kháng Carbapemem A baumannii cao, v y c n có m t gi i pháp nhanh chóng nh m k p th i phát hi nhóm vi sinh v t t l n a vi c th c hi n lu ABSTRACT The method of questioning and choosing a research direction comes from the reality of hospital infections, the subject is applying Yamamoto's technical process to detect the gene bla OXA-23 encoding carbapenemase on A baumannii The data published in the topic, performed by the institute and the research team directly on patient samples at Gia An 115 Hospital and Thong Nhat Dong Nai Hospital, these are real data, and closely follow the infection situation at the hospital The study confirmed that the LAMP method is a molecular diagnostic method with high accuracy with a specificity of 100% and a detection limit of 10 CFU/mL for the detection of the bla OXA-23 gene The results of the study show that the level of resistance to Carbapemem on A baumannii is 77.41%, this is a rather high rate, so a quick solution is needed to promptly detect and prevent groups of microorganisms drug resistance This is also the aim and meaning of the thesis L cơng trình nghiên c u c a h c viên nhóm nghiên c u, khơng chép c a t nghiên c th c hi n N i dung lý thuy t lu tham kh o cơng trình nghiên c u c, d ch tài li u, t ng h p c viên có s d ng m t s tài li u c công b li u, n m m nh ng k t qu lu hi c th c c l p b i h c viên nhóm nghiên c u Tác gi lu ng M CL C T VI T T T DANH M C HÌNH VÀ B NG TV NG QUAN 1.1 Vi khu n Acinetobacter baumannii 1.1.1 m sinh lý 1.1.2 c Protein màng (Porin) Y u t phong t a t bào (LPS, Capsule) Enzym S n xu t màng sinh h c 1.1.3 kháng kháng sinh Aminoglycoside Carbapenem Fluoroquinolone 10 Cephalosporin 11 Sulbactam 11 Rifampicin 11 Tetracycline 11 Polymyxin 12 Gen kháng thu c 12 1.1.4 Nhi m khu ng lâm sàng c a A baumannii 13 Nhi m khu ng hô h p 13 Nhi m khu n máu 13 Nhi m khu ng ti t ni u 14 Viêm màng não 14 1.2 Kháng sinh Carbapenem 15 C u t o hóa h c 15 tác d ng 16 Ph tác d ng 16 1.2.1 S kháng Carbapenem Acinetobacter baumannii 16 phi enzym 17 enzym 17 1.2.2 Các gen mã hóa carbapenemase 19 Insertion sequences 19 Intergon 19 Resistance island 19 Plasmid 20 1.2.3 n gen mã hóa carbapenemase 20 u hình 20 21 c phân t 21 1.3 K thu t LAMP 23 1.3.1 Khái ni m 23 1.3.2 c a LAMP 23 1.3.3 Phát hi n s n ph m c a LAMP 27 1.3.4 m c a LAMP 28 1.4 Các nghiên c u liên quan 29 U 31 2.1 ng nghiên c u 31 2.2 tóm t c th c hi n m c tiêu nghiên c u 32 2.2.1 tóm t t m c tiêu 32 2.2.2 tóm t t m c tiêu 33 2.3 Th m nghiên c u 34 2.4 V t li u nghiên c u 34 2.5 u 35 2.5.1 M u b nh ph m lâm sàng 35 2.5.2 35 y b nh ph m th ch 35 35 2.5.3 lý m u 36 2.5.4 t DNA 37 2.5.5 n di gel 37 2.5.6 Thi t l p ph n ng phát hi n s n ph m c a LAMP 38 2.5.7 Thi t l p ph n ng phát hi n s n ph m PCR 39 T QU NGHIÊN C U 41 3.1 K t qu kh o sát m u b nh ph m lâm sàng 41 3.2 K t qu 42 3.3 K t qu phát hi n gen mã hóa blaOXA-23 c a LAMP khu n l c 44 3.4 nh gi i h n phát hi n c a ph n ng LAMP 47 3.5 K t qu LAMP phát hi n gen blaOXA-23 m u b nh ph m lâm sàng 49 3.6 u qu c a LAMP 50 3.7 Bàn lu n 52 T LU N, KI N NGH 56 4.1 K t lu n 56 4.2 Ki n ngh ng phát tri n c tài 57 TÀI LI U THAM KH O 59 T VI T T T A.Baumannii Acinetobacter baumannii ABC Acinetobacter baumannii - calcoaceticus BIP Backward inner primer CHDL Carbapenem- CRAB Carbapenem resistance Acintobacter baumannii CRAB Carbapenem-resistant Acinetobacter baumannii FIP Forward inner primer ICU Intensive Care Unit LAMP Loop mediated isothermal amplification LB Loop primer backward LF Loop primer forward PBP Penicillin-binding proteins PCR Polymerase chain reaction VAP ventilator associated pneumonia WHO World Health Organization -lactamases 58 cho vi c m r ng k thu t LAMP phát hi n gen mã hóa carbapenemase nhóm vi sinh v t khác c th c hi n th i gian d ch b nh Covid-19 x y ra, nên s b nh nhân n i trú t i b nh vi n có th gi vi c thu th p m u b nh ph m u b nh ph v y có th s ng khơng cho tài, chúng tơi ch thu th m A baumannii m sốt h baumannii K t qu ch phát hi cs baumannii c 31 ng m u khiêm t n, ng gen mã hóa carbapenemase A c gen bla OXA-23, bla OXA-51, gen bla OXA-58 n th y s hi n di n nhóm m u N u có th m r ng s nghiên c u, có th t ng ng m u c h t t l gen mã hóa carbapenemase A 59 TÀI LI U THAM KH O [1] N R Naylor et al Antimicrob Resist Infect Control, vol 7, p 58, 2018, doi: 10.1186/s13756-018-0336-y [2] evolut Pathog Dis, vol 71, no 3, pp 292 301, Aug 2014, doi: 10.1111/2049-632X.12125 [3] Curr Pharm Des, vol 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C QU C GIA TP.HCM C NG HÒA XÃ H I CH T NAM I H C BÁCH KHOA c l p T H nh phúc Phái: Nam Sinh ngày: 28/10/1987 Tôn giáo: Không tân, Tp.HCM Email: lamtrungtruong@gmail.com -HCM: 26/03/2014 Ngày 72 Quá Ngày Ngày 2005 2010 Tp.HCM 2008 2013 Tp.HCM 2013 2017 2018 2021 - Tp.HCM - Tp.HCM Gia An 115 2021 Lâm ... Sinh H c Mã s : 60420201 TÀI: Phát hi n gen mã hóa carbapenemase ch ng Acinetobacter baumannii b ng k thu t LAMP NHI M V VÀ N I DUNG: n gen mã hóa carbapenemase ch ng Acinetobacter baumannii. .. gen bla OXA-40 n gen repAci2 gen bla OXA-23 liên quan gen repAci1 [90] 1.2.3 n gen mã hóa carbapenemase S liên quan lâm sàng vi c phát hi n nhanh chóng gen mã hóa carbapenemase gi a ch ng A baumannii. .. Norihisa Yamamoto phát hi n t l gen mã hóa carbapenemase bla OXA-23 khu n l c Acinetobacter baumannii phân l p t m u b nh ph m lâm sàng nh c hi u c a k thu t LAMP vi c phát hi n gen mã hóa carbapenemase