食品动物源耐甲氧西林金黄色葡萄球菌MLSB类 抗生素耐药性调查

【目的】调查中国6个省食品动物源耐甲氧西林金黄色葡萄球菌（livestock-associated methicillin- resistant staphylococcus aureus,LA-MRSA）对大环内酯类-林可酰胺类-链阳菌素B类（macrolides, lincosamides and type B streptogramin, MLS_B）抗生素的耐药情况及MLS_B耐药基因的流行情况。【方法】从广东、河南、河北、福建、山东以及湖南等6省采集的食品动物源（猪、鸡以及鸭）样品约6 500份,分离金黄色葡萄球菌,通过苯唑西林药物敏感性测定和mecA,mecC基因检测鉴定MRSA菌株。采用二倍琼脂稀释法和微量肉汤稀释法测定LA-MRSA菌株对MLS_B以及其他常见的10种抗菌药物的敏感性,采用PCR方法检测MLS_B耐药基因（ermA-C和ereA-B）以及其他16种常见耐药基因。【结果】6 500份动物源样品中分离出480株金黄色葡萄球菌,其中101株为LA-MRSA,LA-MRSA分离率为1.54%（101/6500）,金黄色葡萄萄球菌中LA-MRSA检出率达21.04%（101/480）。LA-MRSA菌株对MLS_B类抗生素呈现高度耐药的情况,耐药率高达99.00%以上,对其他常见的抗菌药物氟苯尼考、四环素、头孢噻肟以及庆大霉素等抗菌药物也均呈现高度耐药,耐药率为94.00%—99.00%。MLS_B耐药基因检测结果显示,ermC的检出率最高,为100.00%;其次是ereB,为79.20%,接着是ermA和ereA,检出率分别为45.54%和40.59%,而ermB的检出率最低,为11.88%。其他耐药基因的检测结果显示fexA、tetL、aadA1、aph(4’)-Ia的检出率较高,分别是92.10%、97.02%、97.29%和83.17%;其次是tet(M),检出率为71.29%;而aac(6’)-Ib、aac(3’)-Ic、 aph(3’)-II、aph(3’)-IV、optrA、tet(A)、tet(C)的检出率相对较低,分别是49.50%、46.53%、39.60%、37.62%、33.67%、29.70%和20.79%;接着检出率更低的是cfr、tet(K)、lnuA以及lnuF,分别为17.82%、14.85%、5.94%和3.96%。进一步分析LA-MRSA菌株中MLS_B耐药基因检出个数与对受试抗菌药物的耐药数目之间的关系,发现随着动物源MRSA菌株对受试抗菌药物的耐药数目增加时,菌株中MLS_B耐药基因检出个数也在增加。【结论】动物源MRSA菌株对MLS_B高度耐药,其与菌株中ermC基因的高检出率一致,并且LA-MRSA菌株多重耐药情况严重,这与MLS_B耐药基因检出个数以及其他耐药基因的高检出率密切相关。因此,养殖场中MLS_B类药物使用应规范使用,以减少动物源MRSA菌株的多重耐药性。

Investigation on the Antibiotic Resistance of Staphylococcus Methicillin-Resistant MLSB from Food Animals in Six Provinces of China

【Objective】 The aim of this study was to investigate the prevalence of genes encoding resistance to macrolides, lincosamides and streptogramins (MLS_B) among methicillin-resistant Staphylococcus aureus from food animals in China. 【Method】Staphylococcus aureus were isolated from about 6 500 samples of pig, chicken and duck origins among six different provinces, including Guangdong, Henan, Hebei, Fujian and so on, in China. LA-MRSA was confirmed from staphylococcus aureus through detecting the susceptibility of oxacillin and the presence of mecA and mecC genes. Susceptibility test of LA-MRSA resistant to MLS_B and other common antibiotics were determined by agar dilution method and microdilution broth method. And the MLS_B resistance genes (ermA-C and ereA-B) and other common resistant genes were detected by PCR. 【Result】Four hundred and eighty Staphylococcus aureus isolates and 101 LA-MRSA isolates were found among about 6 500 samples, and the isolation rate of Staphylococcus aureus and the detection rate of LA-MRSA among Staphylococcus aureus isolates were 1.54% (101/6500) and 21.04% (101/480), respectively. Susceptibility test showed high resistance rate of LA-MRSA to MLS_B was found with over 99.00% among LA-MRSA isolates. High resistance rate to other common antibiotics was also found among LA-MRSA, including florfenicol, tetracycline, cefotaxime and gentamycin with the resistance rates of 99.01%, 96.39%, 96.39%, 96.39% and 94.05%, respectively. MLS_B resistance genes detection showed that ermC was found in all LA-MRSA isolates, followed by ereB (79.20%), ermA (45.54%) and ereA (40.59%), while, ermB was present with a low detection rate (11.88%). Other drug-resistant genes test results showed that fexA, tetL, aadA1, aph (4') - Ia detection rate was higher, which was 92.10%, 97.02%, 97.29% and 83.17%, respectively; secondly, tet (M), the detection rate was 71.29%; the detection rate of aac (6') - Ib, aac (3') - Ic, aph (3') -II, aph (3') -IV, optrA, tet (A) and tet (C) was relatively low, which was 49.50%, 46.53%, 39.60%, 37.62%, 33.67%, 29.70% and 20.79%, respectively; the detection rate of cfr, tet (K), lnuA and lnuF was the lowest, which was 17.82%, 14.85%, 5.94% and 3.96%, respectively. The relationship of the number of MLS_B resistance genes and the number of resistance to antibiotics was further analyzed, which indicated the increased number of resistance to antibiotics was corresponding with the increased number of MLS_B resistance genes and high detection rate of other resistant genes among the LA-MRSA isolates. 【Conclusion】In conclusion, in this study, a high resistance to MLS_B was found and it was consistent to the high detection rate of ermC gene among LA-MRSA isolates. The multi-drug resistance of LA-MRSA isolates was serious and this was closely related to the number of MLS_B resistance genes. Therefore, it is urgently needed to take action in the rational use of antibiotics, as to reducing the emergence of multi-drug resistance LA-MRSA.