tdh+与tdh−副溶血弧菌耐药表型与基因型差异分析

肖文静; 陈美玲; 赵文轩; 李臻鹏; 张天姿; 卢盼; 李杰; 阚飙; 逄波

doi:10.3784/jbjc.202103080104

tdh⁺与tdh⁻副溶血弧菌耐药表型与基因型差异分析

Comparison of antibiotic resistance phenotype and genotype between tdh⁺ and tdh⁻ strains of Vibrio parahaemolyticus

摘要

摘要:
目的描述tdh⁺与tdh⁻副溶血弧菌表型与基因型耐药差异，分析耐药基因型与耐药表型之间的关系。
方法采用K-B纸片法检测208株tdh⁺与73株tdh⁻副溶血弧菌对临床及水产养殖中常用的7类16种抗生素的敏感性，使用新一代测序技术获得实验菌株的全基因序列，通过序列比对分析基因组中耐药基因及耐药相关基因突变情况。分析耐药表型与基因型之间的关系。
结果 tdh⁺菌株和tdh⁻菌株对亚胺培南均100%敏感。 tdh⁺菌株对头孢吡肟、阿奇霉素耐药率（0.48%、8.65%）高于tdh⁻菌株（0.00%、4.11%）；tdh⁻菌株对氨苄西林、头孢西汀、头孢曲松、环丙沙星、萘啶酸、链霉素、卡那霉素、庆大霉素、磺胺甲恶唑、复方新诺明、四环素、多西环素、氯霉素耐药率均高于tdh⁺菌株。 tdh⁻菌株的多重耐药率（34.25%）高于tdh⁺菌株（19.23%）。耐药基因共检出5类7种，其中β-内酰胺类bla_CARB基因在所有的tdh⁺菌株和tdh⁻菌株均检出；四环素类tet（34）基因在tdh⁺与tdh⁻菌株基因携带率分别为99.52%、98.63%；喹诺酮类耐药基因qnrS5 仅在tdh⁺菌株中检出；tdh⁻菌株中叶酸代谢抑制类（sul2）、四环素类［tet（35）、tet（59）］、苯丙醇类（floR）耐药基因检出率高于tdh⁺菌株。共在14株（tdh⁺9株，tdh⁻5株）细菌中检索出6种耐药基因点突变方式，均为编码16S rRNA的rrs基因突变。
结论 tdh⁻副溶血弧菌耐药现象较tdh⁺副溶血弧菌更为严重； 16S rRNA的rrs基因突变是除aph（3''）-Ib、aac（3）-Ⅱ、aac（6’） -I、aph（3''）-Id等耐药基因外副溶血弧菌对氨基糖胺类药物耐药的主要机制之一；副溶血弧菌中，可用bla_CARB耐药基因存在情况预测副溶血弧菌氨苄西林耐药表型；除此之外其余抗生素耐药表型与其对应耐药基因之间未见明确的对应的关系。

Abstract:
Objective To compare the phenotypic and genotypic antibiotic resistance difference between tdh⁺ and tdh⁻ strains of Vibrio parahaemolyticus, and analyze the relationship between antibiotic resistance genotype and phenotype.
Methods The resistance patterns of 208 tdh⁺ and 73 tdh⁻ strains of V. parahaemolyticus to 16 antibiotics (7 kinds) commonly used in clinical treatment and aquaculture were determined using K-B test. The whole gene sequences of the strains were obtained by new generation sequencing technology. The mutations of antibiotic resistance genes and related genes in the genome were analyzed by sequence alignment. The relationship between phenotype and genotype was analyzed.
Results Both tdh⁺ and tdh⁻ strains were 100% sensitive to imipenem. The drug-resistance rates of tdh⁺ strains (0.48%, 8.65%) to cefepime and azithromycin were higher than that of tdh⁻ strains (0.00%, 4.11%); the drug-resistance rates of tdh⁻ strains to ampicillin, cefoxitin, ceftriaxone, ciprofloxacin, nalidixic acid, streptomycin, kanamycin, gentamicin, sulfamethoxazole, sulfamethoxazole, tetracycline, doxycycline and chloramphenicol were higher than those of tdh⁺ strains. The multidrug resistance rate of tdh⁻ strains (34.25%) was higher than that of tdh⁺ strains (19.23%). There were 5 types and 7 kinds of drug resistance genes, among which β - lactam gene (bla_CARB) was detected in all the tdh⁺ strains and tdh⁻ strains; the tetracycline resistance tet (34) gene carriage rates in tdh⁺and tdh⁻ strains were 99.52% and 98.63%, the quinolone resistance gene qnrS5 was detected only in tdh⁺ strain. The detection rates of folate metabolism inhibitors (sul2), tetracycline ［tet（35）、tet（59）］ and phenylpropanol (floR) resistance genes in tdh⁻ strains were higher than those in tdh⁺ strains. A total of 14 strains of V. parahaemolyticus (9 tdh⁺ strains, 5 tdh⁻ strains) were screened for six point mutations of drug-resistance genes, all of which were rrs gene mutations encoding 16S rRNA.
Conclusion The drug resistance of tdh⁻ V. parahaemolyticus strains was more serious than that of tdh⁺ V. parahaemolyticus strains; the rrs gene mutation of 16S rRNA was one of the main resistance mechanisms of V. parahaemolyticus to aminoglycosamines except for resistance genes: aph(3'')-Ib, aac(3)-Ⅱ, aac(6’) –I and aph(3'')-Id. bla_CARB gene can be used to predict the ampicillin resistance phenotype of V. parahaemolyticus, bur there was no clear relationship between other antibiotic resistance phenotypes and the corresponding resistance genes

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tdh+与tdh−副溶血弧菌耐药表型与基因型差异分析

Comparison of antibiotic resistance phenotype and genotype between tdh+ and tdh− strains of Vibrio parahaemolyticus

tdh⁺与tdh⁻副溶血弧菌耐药表型与基因型差异分析

Comparison of antibiotic resistance phenotype and genotype between tdh⁺ and tdh⁻ strains of Vibrio parahaemolyticus