Saha Biswajit, Singh Anil K, Ghosh Abhrajyoti, Bal Manjusri
Microbiology Laboratory, Department of Physiology, University College of Science, Technology and Agriculture, University of Calcutta, Kolkata, India.
Department of Chemistry, Bose Institute, Kolkata, India.
J Med Microbiol. 2008 Jan;57(Pt 1):72-79. doi: 10.1099/jmm.0.47144-0.
A pathogenic vancomycin-resistant Staphylococcus aureus (VRSA) isolate (MIC > or =64 microg ml(-1)) was obtained from a Kolkata hospital in June 2005. Species identification was confirmed by Gram staining, standard biochemical tests and PCR amplification of the nuc gene, which encodes the thermostable nuclease that is highly specific for S. aureus. The VRSA isolate was also resistant to beta-lactams (amoxicillin, ampicillin, cefepime, cefotaxime, cefuroxime, cephalexin and meticillin), chloramphenicol, streptomycin, macrolides (erythromycin and roxithromycin), clindamycin, rifampicin and trimethoprim-sulfamethoxazole. However, the isolate was susceptible to gentamicin (an aminoglycoside) and ciprofloxacin (a fluoroquinolone). The resistance to vancomycin was inducible in vitro, because the MIC of vancomycin increased from 64 microg ml(-1) initially to 1024 microg ml(-1) during culture of this VRSA strain in the presence of vancomycin. The VRSA isolate contained a large plasmid ( approximately 53.4 kb) and four small plasmids of approximately 6, 5.5, 5.1 and 1.5 kb. The large plasmid of approximately 53.4 kb harboured the vancomycin-resistance genes vanHAX, which was confirmed by PCR amplification using the same plasmid as template and, separately, primers specific for the 2.61 kb vanHAX gene cluster, vanH (969 bp), vanA (1032 bp) and vanX (609 bp). The VRSA isolate was also positive for mecA. Vancomycin resistance was successfully transferred from this VRSA donor to a vancomycin-sensitive recipient S. aureus clinical isolate by a broth mating procedure. The MIC of vancomycin for the transconjugant was 32 microg ml(-1), as against 2 microg ml(-1) for the parent strain. Nucleotide sequencing of the PCR product showed partial homology with van genes of an enterococcal transposon Tn1546-like element. This is believed to be the first Indian S. aureus isolate that has been shown to be phenotypically vancomycin-resistant, presumably due to a vanHAX analogue.
2005年6月，从加尔各答一家医院分离出一株致病性耐万古霉素金黄色葡萄球菌（VRSA，最低抑菌浓度[MIC]≥64μg/ml）。通过革兰氏染色、标准生化试验及对编码对金黄色葡萄球菌具有高度特异性的耐热核酸酶的nuc基因进行PCR扩增，确认了菌种鉴定结果。该VRSA菌株还对β-内酰胺类抗生素（阿莫西林、氨苄西林、头孢吡肟、头孢噻肟、头孢呋辛、头孢氨苄和甲氧西林）、氯霉素、链霉素、大环内酯类抗生素（红霉素和罗红霉素）、克林霉素、利福平和甲氧苄啶-磺胺甲恶唑耐药。然而，该菌株对庆大霉素（一种氨基糖苷类抗生素）和环丙沙星（一种氟喹诺酮类抗生素）敏感。该菌株对万古霉素的耐药性在体外可诱导，因为在万古霉素存在的情况下培养此VRSA菌株时，万古霉素的MIC从最初的64μg/ml增加到1024μg/ml。该VRSA菌株含有一个大约53.4kb的大质粒和四个分别约为6kb、5.5kb、5.1kb和1.5kb的小质粒。大约53.4kb的大质粒携带了万古霉素耐药基因vanHAX，以该质粒为模板进行PCR扩增，以及分别使用针对2.61kb vanHAX基因簇、vanH（969bp）、vanA（1032bp）和vanX（609bp）的引物进行PCR扩增，均证实了这一点。该VRSA菌株的mecA也呈阳性。通过肉汤交配程序，万古霉素耐药性成功地从该VRSA供体转移到了一株对万古霉素敏感的金黄色葡萄球菌临床分离株。转接合子对万古霉素的MIC为32μg/ml，而亲本菌株为2μg/ml。PCR产物的核苷酸测序显示与肠球菌转座子Tn1546样元件的van基因有部分同源性。据信这是第一株在表型上显示耐万古霉素的印度金黄色葡萄球菌分离株，可能是由于一个vanHAX类似物所致。
