Antibiotic resistance and multi-locus sequence typing of non-O157 Shiga toxin-producing Escherichia coli from livestock in Dongtai, Jiangsu
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摘要:
目的 了解江苏省东台地区家畜来源非O157产志贺毒素大肠埃希菌(STEC)抗生素耐药表型、耐药基因,以及多位点序列分型(MLST)情况。 方法 于2019年5月,江苏省东台市采集家畜粪便样本301份(羊粪便231份,牛粪便70份),分离非O157 STEC菌株。 采用改良微量肉汤法测定菌株对21种抗生素最小抑菌浓度(MIC);通过全基因组测序技术对O∶H血清型、耐药基因和多位点序列型别进行预测。 结果 在68株非O157 STEC中,有32株对至少1种药物耐药。 其中,STEC对四环素耐药率最高(42.6%),其次分别对阿奇霉素(36.8%)、复方新诺明(35.3%)、链霉素(32.3%)、氯霉素(30.9%)、环丙沙星(29.4%)等9种抗生素耐药;共识别出25种耐药基因,其中,氨基糖苷类耐药基因和叶酸途径抑制剂类耐药基因携带率最高(44.1%),其次为四环素类耐药基因tet(A)(42.6%);MLST将分离株分为13种ST型,其中ST43(19.1%)和ST155(16.2%)比例较高。 最小生成树显示,具有相同血清型的STEC菌株多聚集在一起。 STEC中4种ST型别(ST25、ST40、ST43、ST675)分别与引起肠溶血性尿毒综合征相关的肠出血性大肠埃希菌(HUSEC)聚类成簇。 结论 江苏东台地区家畜中非O157 STEC耐药形势复杂,且存在多重耐药现象。 此外,该地区STEC菌株对人群存在潜在威胁,家畜作为非O157 STEC宿主应引起重视。 -
关键词:
- 产志贺毒素大肠埃希菌 /
- 耐药性 /
- 耐药基因 /
- 多位点序列分型
Abstract:Objective To investigate and characterize the antibiotic resistance, antibiotic resistance genes, and multi-locus sequence typing (MLST) of non-O157 Shiga toxin-producing Escherichia coli (STEC) isolated from livestock in Jiangsu. Methods In May 2019, a total of 301 stool samples, including 231 sheep stool samples and 70 cattle stool samples, were collected in Dongtai of Jiangsu province for the isolation of non-O157 STEC. The modified micro broth method was used to determine the minimum inhibitory concentration (MIC) of 21 antibiotics to the isolates. The serotypes, antibiotic resistance genes and MLST of the isolates were analyzed by whole-genome sequencing (WGS). Results The drug sensitivity test showed that of the 68 non 157-STEC strains, 32 were at least resistant to one antibiotic. It was found that STEC showed the highest resistance rate to tetracycline (42.6%), followed by azithromycin (36.8%), trimethoprim-sulfamethoxazole (TMP/SMX) (35.3%), streptomycin (32.3%), chloramphenicol (30.9%) and ciprofloxacin (29.4%). A total of 25 AMR genes were identified. The majority of STEC isolates carried the antibiotic resistance genes associated with aminoglycoside (44.1%), TMP/SMX (44.1%), and tetracycline (42.6%). MLST showed that the isolates were divided into 13 sequence types (STs), among which ST43 (19.1%) and ST155 (16.2%) had higher proportions. The minimum spanning tree (MST) indicated that most STEC strains with the same serotypes were clustered together. Four STs of STEC strains, including ST25, ST40, ST43 and ST675, were clustered with HUSEC strains. Conclusion The antibiotic resistance of non-O157 STEC in livestock is complex, and there are multi-drug resistant strains. In addition, STEC strains in this area pose a potential threat to the population, and close attention needs to be paid to livestock, as an important host of non-O157 STEC. -
表 1 68株非O157 STEC菌株血清型、毒力基因及ST型别分布
Table 1. Distribution of serotypes, virulence genes and sequence types (STs) in 68 non-O157 STEC strains
血清型 菌株数(株) 菌株来源 毒力基因 ST型别 O6∶H10 13 羊 stx1 43 O155∶H21 10 羊 stx2 683 O21∶H25 7 羊 stx1 155 O22∶H8 7 羊 stx1 11997 O66∶H45 7 羊 stx2 971 O38∶H26 6 羊 stx1 10 OX18∶H21 6 羊 stx1 40 O8∶H21 2 羊 stx1 155 O43∶H2 2 羊 stx2 937 O66∶H21 2 羊 stx1 155 O76∶H19 2 羊 stx1 675 O5∶H19 1 羊 stx1+stx2 447 O128∶H2 1 羊 stx2 25 O150∶H8 1 羊 stx1 906 O182∶H25 1 牛 stx1+eae 300 表 2 68株非O157 STEC药敏试验结果
Table 2. Antibiotic susceptibility test results of 68 non-O157 STEC strains
抗菌药物 菌株耐药分布 敏感(S) 中介(I) 耐药(R) 四环素 39(57.4) 0(0.0) 29(42.6) 阿奇霉素 43(63.2) 0(0.0) 25(36.8) 复方新诺明 44(64.7) 0(0.0) 24(35.3) 链霉素 1(1.5) 45(66.2) 22(32.3) 氯霉素 47(69.1) 0(0.0) 21(30.9) 环丙沙星 43(63.2) 5(7.4) 20(29.4) 氨苄西林 56(82.4) 0(0.0) 12(17.6) 萘啶酸 58(85.3) 0(0.0) 10(14.7) 氨曲南 58(85.3) 1(1.5) 9(13.2) 头孢噻肟 59(86.8) 0(0.0) 9(13.2) 多黏菌素E 0(0.0) 68(100) 0(0.0) 头孢他啶 65(95.6) 3(4.4) 0(0.0) 氨苄西林–舒巴坦 66(97.1) 2(2.9) 0(0.0) 厄他培南 68(100.0) 0(0.0) 0(0.0) 美罗培南 68(100.0) 0(0.0) 0(0.0) 亚胺培南 68(100.0) 0(0.0) 0(0.0) 头孢他啶–阿维巴坦 68(100.0) 0(0.0) 0(0.0) 头孢西丁 68(100.0) 0(0.0) 0(0.0) 替加环素 68(100.0) 0(0.0) 0(0.0) 阿米卡星 68(100.0) 0(0.0) 0(0.0) 呋喃妥因 68(100.0) 0(0.0) 0(0.0) 注:括号外数据为菌株数,括号内数据为百分比(%) 表 3 非O157 STEC菌株多重耐药组合
Table 3. Combination of MDR in non-O157 STEC strains
抗菌药物组合 耐药菌株数(株) CHL+SXT+CTX+TET+CIP+AZM+STR+AMP+ATM 8 CHL+TET+STR 4 SXT+TET+CIP+NAL+AZM+STR 4 CHL+SXT+TET+AZM 3 SXT+TET+CIP+NAL+AZM 3 CHL+SXT+TET+CIP+AZM+STR+AMP 2 CHL+SXT+CTX+TET+CIP+AZM+STR+AMP 1 CHL+SXT+TET+AZM+STR 1 CHL+SXT+TET+CIP+AZM 1 CHL+SXT+TET+CIP+AZM+STR+AMP+ATM 1 TET+NAL+STR 1 注:CHL. 氯霉素;SXT. 复方新诺明;CTX. 头孢噻肟;TET. 四环素;AZM. 阿奇霉素;STR. 链霉素;AMP. 氨苄西林;ATM. 氨曲南;CIP. 环丙沙星;NAL. 萘啶酸 表 4 非O157 STEC携带耐药基因和表型耐药关系
Table 4. Correlation between antibiotic resistance genes and antibiotic resistance phenotypes in non-O157 STEC strains
药物类别 耐药基因组合(菌株数) 携带耐药基因菌株总数(株) 表型耐药菌株总数(株) 耐药基因携带率(%) 氨基糖苷类 aac(3)-IId+aph(3')-Ia+aph(3'')-Ib+aph(6)-Id(9)
aadA1(7)
aac(3)-IId+aac(6')-Ib-cr+aph(3'')-Ib+aph(6)-Id(6)
aph(3'')-Ib+aph(6)-Id(5)
aac(3)-IId+aac(6')-Ib-cr+aph(3'')-Ib+aph(6)-Id+aadA16(1)
aac(3)-IId+aph(3'')-Ia+aph(6)-Id(1)
aac(3)-IId(1)30 22 44.1 叶酸途径抑制剂 sul2+sul3+dfrA14(10)
sul1+dfrA1(7)
sul2(5)
sul1+dfrA27(4)
sul1+sul2+dfrA27(3)
sul1(1)30 24 44.1 四环素类 tet(A)(29) 29 29 42.6 喹诺酮类 gyrA+qnrS1(9)
qnrS1(8)
aac(6')-Ib-cr+qnrB6(7)
gyrA(3)
qnrB6(1)28 23 41.2 大环内酯类 mph(A)(25) 25 25 36.8 氯霉素类 floR(21) 21 21 30.9 β-内酰胺类 blaCTX-M-55(10)
blaTEM-1B(2)12 12 17.6 注:字体加粗基因为耐药基因型和耐药表型不一致者 -
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