新疆农业科学 ›› 2023, Vol. 60 ›› Issue (12): 3113-3120.DOI: 10.6048/j.issn.1001-4330.2023.12.029
• 畜牧兽医·微生物·农业装备工程与机械化 • 上一篇
王东1(), 易海波2, 李宏博1, 秦蕾1, 徐琦琦1, 吴慧敏1, 夏利宁1()
收稿日期:
2023-04-20
出版日期:
2023-12-20
发布日期:
2024-01-03
通信作者:
夏利宁(1975-),女,湖南人,教授,博士,研究方向为兽医药理与毒理学,(E-mail)xln750530@163.com
作者简介:
王东(1994-),男,四川人,硕士研究生,研究方向为兽医药理与毒理学,(E-mail)1005621397@qq.com
基金资助:
WANG Dong1(), YI Haibo2, LI Hongbo1, QIN Lei1, XU Qiqi1, WU Huimin1, XIA Lining1()
Received:
2023-04-20
Online:
2023-12-20
Published:
2024-01-03
Correspondence author:
XIA Lining (1975-), female, Hunan, professor, PhD, research interests in veterinary pharmacology and toxicology, (E-mail)Supported by:
摘要:
【目的】分析新疆阿克苏地区猪源粪肠球菌对抗菌药物的耐药性及相关耐药基因携带情况。【方法】从阿克苏地区三个猪场采集的624份肛拭子样,通过肠球菌选择培养基结合粪肠球菌持家基因PCR检测的方法进行粪肠球菌的分离鉴定,采用琼脂稀释法对分离的粪肠球菌进行药物敏感性试验,使用PCR方法检测相关耐药基因。【结果】共分离粪肠球菌160株,分离率为25.6%(160/624)。分离的粪肠球菌对红霉素、泰乐菌素、四环素、多西环素、庆大霉素和沃尼妙林的耐药率均达到97.0%以上,值得注意的是分离的粪肠球菌对利奈唑胺的耐药率为21.3%(34/160),未发现万古霉素耐药菌株。阿克苏地区分离的猪源粪肠球菌耐药谱型在3~10耐之间,其主要谱型为6耐(28.1%)和8耐(30.6%)。【结论】三个猪场均检出ermB、ermC、tet(K)、tet(L)、tet(M)、aac(6')/aph(2")、lsa(E)基因,检出率均达到50%以上;多药耐药基因optrA、lsa(E)检出率达到25.6%(44/160)、40.0%(64/160)。
中图分类号:
王东, 易海波, 李宏博, 秦蕾, 徐琦琦, 吴慧敏, 夏利宁. 新疆阿克苏地区猪源粪肠球菌耐药性分析及相关耐药基因的检测[J]. 新疆农业科学, 2023, 60(12): 3113-3120.
WANG Dong, YI Haibo, LI Hongbo, QIN Lei, XU Qiqi, WU Huimin, XIA Lining. Drug resistance analysis of Enterococcus faecalis isolated from pigs and detection of related drug resistance genes in Aksu, Xinjiang[J]. Xinjiang Agricultural Sciences, 2023, 60(12): 3113-3120.
引物名称 Primer | 引物序列5'-3' Sequences | 片段大小 Products(bp) | 退火温度 Annealing Temperature(℃) |
---|---|---|---|
E.faecalis-F | TCAAGTACAGTTAGTCTTTATTAG | 941 | 55 |
E.faecalis-R | ACGATTCAAAGCTAACTGAATCAGT | ||
16S-F | AGAGTTTGATCCTGGCTCAG | 1466 | 54 |
16S-R | ACGGCTACCTTGTTACGACTT |
表1 肠球菌鉴定引物序列
Tab.1 Primer sequence for identification of Enterococcus faecalis
引物名称 Primer | 引物序列5'-3' Sequences | 片段大小 Products(bp) | 退火温度 Annealing Temperature(℃) |
---|---|---|---|
E.faecalis-F | TCAAGTACAGTTAGTCTTTATTAG | 941 | 55 |
E.faecalis-R | ACGATTCAAAGCTAACTGAATCAGT | ||
16S-F | AGAGTTTGATCCTGGCTCAG | 1466 | 54 |
16S-R | ACGGCTACCTTGTTACGACTT |
引物名称 Primer | 引物序列5'-3' Sequences | 片段大小 Products(bp) | 退火温度 Annealing Temperature(℃) |
---|---|---|---|
ermA-F | TCTAAAAAGCATGTAAAAGAA | 645 | 53 |
ermA-R | CTTCGATAGTTTATTAATATTAGT | ||
ermB-F | GAAAAAGTACTCAACCAAATA | 639 | 50 |
ermB-R | AGTAACGGTACTTAAATTGTTTAC | ||
ermC-F | TCGAAACGTAATATAGATAAA | 642 | 47.5 |
ermC-R | GCTAATATTGTTTAAATCGTCAAT | ||
tet(L)-F | GTTGCGCGCTATATTCCAAA | 788 | 51.7 |
tet(L)-R | TTAAGCAAACTCATTCCAGC | ||
tet(K)-F | TTAGGTGAAGGGTTAGGTCC | 718 | 49.9 |
tet(K)-R | GCAAACTCATTCCAGAAGCA | ||
tet(M)-F | ACAGAAAGCTTATTATATAAC | 171 | 48.5 |
tet(M)-R | TGGCGTGTCTATGATGTTCAC | ||
fexA-F | GTACTTGTAGGTGCAATTACGGCTGA | 1272 | 57 |
fexA-R | CGCATCTGAGTAGGACATAGCGTC | ||
fexB-F | TTCCCACTATTGGTGAAAGGAT | 750 | 51.9 |
fexB-R | GCAATTCCCTTTTATGGACGTT | ||
cfr-F | TGAAGTATAAAGCAGGTTGGGAGT | 746 | 53.2 |
cfr-R | ACCATATAATTGACCACAAGCAGC | ||
optrA-F | AGGTGGTCAGCGAACTAA | 1395 | 49.9 |
optrA-R | ATCAACTGTTCCCATTCA | ||
lsa(E)-F | TTGTACGGAATGTATGG | 675 | 49.3 |
lsa(E)-R | TTCGCTTCTATTAAGCACTCTT | ||
aac(6')/aph(2")-F | CCAAGAGCAATAAGGGATACC | 672 | 58 |
aac(6')/aph(2")-R | ACCCTCAAAAACTGTTGTTGC |
表2 相关耐药基因引物序列
Tab.2 Primer sequence of related drug resistance genes
引物名称 Primer | 引物序列5'-3' Sequences | 片段大小 Products(bp) | 退火温度 Annealing Temperature(℃) |
---|---|---|---|
ermA-F | TCTAAAAAGCATGTAAAAGAA | 645 | 53 |
ermA-R | CTTCGATAGTTTATTAATATTAGT | ||
ermB-F | GAAAAAGTACTCAACCAAATA | 639 | 50 |
ermB-R | AGTAACGGTACTTAAATTGTTTAC | ||
ermC-F | TCGAAACGTAATATAGATAAA | 642 | 47.5 |
ermC-R | GCTAATATTGTTTAAATCGTCAAT | ||
tet(L)-F | GTTGCGCGCTATATTCCAAA | 788 | 51.7 |
tet(L)-R | TTAAGCAAACTCATTCCAGC | ||
tet(K)-F | TTAGGTGAAGGGTTAGGTCC | 718 | 49.9 |
tet(K)-R | GCAAACTCATTCCAGAAGCA | ||
tet(M)-F | ACAGAAAGCTTATTATATAAC | 171 | 48.5 |
tet(M)-R | TGGCGTGTCTATGATGTTCAC | ||
fexA-F | GTACTTGTAGGTGCAATTACGGCTGA | 1272 | 57 |
fexA-R | CGCATCTGAGTAGGACATAGCGTC | ||
fexB-F | TTCCCACTATTGGTGAAAGGAT | 750 | 51.9 |
fexB-R | GCAATTCCCTTTTATGGACGTT | ||
cfr-F | TGAAGTATAAAGCAGGTTGGGAGT | 746 | 53.2 |
cfr-R | ACCATATAATTGACCACAAGCAGC | ||
optrA-F | AGGTGGTCAGCGAACTAA | 1395 | 49.9 |
optrA-R | ATCAACTGTTCCCATTCA | ||
lsa(E)-F | TTGTACGGAATGTATGG | 675 | 49.3 |
lsa(E)-R | TTCGCTTCTATTAAGCACTCTT | ||
aac(6')/aph(2")-F | CCAAGAGCAATAAGGGATACC | 672 | 58 |
aac(6')/aph(2")-R | ACCCTCAAAAACTGTTGTTGC |
样品数(份) Sample source (Items) | 粪肠球 菌数(株) No.of Enterococcus faecalis (Strain) | 分离率 Isolation rate(%) | |
---|---|---|---|
A猪场 A farm | 269 | 44 | 16.4 |
B猪场 B farm | 234 | 81 | 34.6 |
C猪场 C farm | 121 | 35 | 28.9 |
合计 Total | 624 | 160 | 25.6 |
表3 阿克苏地区三个猪场猪源粪肠球菌分离
Table 3 Situation of isolation of Enterococcus faecalis from pigs in three pig farms in Aksu Area
样品数(份) Sample source (Items) | 粪肠球 菌数(株) No.of Enterococcus faecalis (Strain) | 分离率 Isolation rate(%) | |
---|---|---|---|
A猪场 A farm | 269 | 44 | 16.4 |
B猪场 B farm | 234 | 81 | 34.6 |
C猪场 C farm | 121 | 35 | 28.9 |
合计 Total | 624 | 160 | 25.6 |
药物名称 Drug names | 粪肠球菌的耐药率/%(耐药数) Drug resistance rate of Enterococcus faecalis/%(No.of drug resistance) | |||
---|---|---|---|---|
A养殖场 (A farm)n=44 | B养殖场 (B farm)n=81 | C养殖场 (C farm)n=35 | 总计 Total n=160 | |
氨苄西林Ampicillin | 0(0)a | 0(0)a | 0(0)a | 0(0) |
红霉素Erythromycin | 97.7(43)a | 100.0(81/81)a | 94.3(33)a | 98.1(157) |
泰乐菌素Tylosin | 97.7(43)ab | 100.0(81/81)b | 91.4(32)a | 97.5(156) |
利奈唑胺Linezolid | 11.4(5)a | 35.8(29)b | 0.0(0)a | 21.3(34) |
四环素Tetracycline | 100.0(44)a | 100.0(81)a | 97.1(34)a | 99.4(159) |
多西环素Doxycycline | 100.0(44)a | 100.0(81)a | 100.0(35)a | 100.0(160) |
万古霉素Vancomycin | 0.0(0)a | 0.0(0)a | 0.0(0)a | 0(0) |
氟苯尼考Florfenicol | 29.6(13)a | 54.3(42)b | 0.0(0)c | 30.6(57) |
左氧氟沙星Levofloxacin | 11.4(5)a | 96.3(78)b | 8.6(3)b | 53.8(86) |
恩诺沙星Enrofloxacin | 27.3(12)a | 96.3(78/81)b | 5.7(2)c | 57.5(92) |
庆大霉素Gentamicin | 100.0(44/)a | 100.0(81)a | 100.0(35)a | 100.0(160) |
沃尼妙林Valnemulin | 100.0(44)a | 100.0(81)a | 100.0(35)a | 100.0(160) |
表4 阿克苏地区三个猪场猪源肠球菌的耐药变化
Table 4 Situation of resistant Enterococcus faecalis isolated from three pig farms in Aksu Area
药物名称 Drug names | 粪肠球菌的耐药率/%(耐药数) Drug resistance rate of Enterococcus faecalis/%(No.of drug resistance) | |||
---|---|---|---|---|
A养殖场 (A farm)n=44 | B养殖场 (B farm)n=81 | C养殖场 (C farm)n=35 | 总计 Total n=160 | |
氨苄西林Ampicillin | 0(0)a | 0(0)a | 0(0)a | 0(0) |
红霉素Erythromycin | 97.7(43)a | 100.0(81/81)a | 94.3(33)a | 98.1(157) |
泰乐菌素Tylosin | 97.7(43)ab | 100.0(81/81)b | 91.4(32)a | 97.5(156) |
利奈唑胺Linezolid | 11.4(5)a | 35.8(29)b | 0.0(0)a | 21.3(34) |
四环素Tetracycline | 100.0(44)a | 100.0(81)a | 97.1(34)a | 99.4(159) |
多西环素Doxycycline | 100.0(44)a | 100.0(81)a | 100.0(35)a | 100.0(160) |
万古霉素Vancomycin | 0.0(0)a | 0.0(0)a | 0.0(0)a | 0(0) |
氟苯尼考Florfenicol | 29.6(13)a | 54.3(42)b | 0.0(0)c | 30.6(57) |
左氧氟沙星Levofloxacin | 11.4(5)a | 96.3(78)b | 8.6(3)b | 53.8(86) |
恩诺沙星Enrofloxacin | 27.3(12)a | 96.3(78/81)b | 5.7(2)c | 57.5(92) |
庆大霉素Gentamicin | 100.0(44/)a | 100.0(81)a | 100.0(35)a | 100.0(160) |
沃尼妙林Valnemulin | 100.0(44)a | 100.0(81)a | 100.0(35)a | 100.0(160) |
多药耐药数 No.of multidrug resistance | 粪肠球菌多药耐药率/%(多药耐药数) Multidrug resistance rate of Enterococcus faecalis /% (No.of multidrug resistant) | |||
---|---|---|---|---|
A养殖场 A farm n=44 | B养殖场 B farm n=81 | C养殖场 C farm n=35 | 合计 Total n=160 | |
3 | 0.0(0)a | 0.0(0)a | 2.9(1)a | 0.6(1) |
4 | 2.3(1)ab | 0.0(0)b | 5.7(2)a | 1.9(3) |
5 | 4.5(2)a | 0.0(0)a | 0.0(0)a | 1.3(2) |
6 | 38.6(17)a | 0.0(0)b | 80.0(28)c | 28.1(45) |
7 | 31.8(14)a | 1.2(1)b | 8.6(3)b | 11.3(18) |
8 | 20.5(9)a | 48.1(39)b | 2.9(1)a | 30.6(49) |
9 | 2.3(1)a | 17.3(14)b | 0.0(0)a | 9.4(15) |
10 | 0.0(0)a | 33.3(27)b | 0.0(0)a | 16.9(27) |
表5 阿克苏地区3个猪场猪源粪肠球菌的多药耐药变化
Table 5 Results of multidrug resistance of Enterococcus faecalis from pigs in three pig farms in Aksu Area
多药耐药数 No.of multidrug resistance | 粪肠球菌多药耐药率/%(多药耐药数) Multidrug resistance rate of Enterococcus faecalis /% (No.of multidrug resistant) | |||
---|---|---|---|---|
A养殖场 A farm n=44 | B养殖场 B farm n=81 | C养殖场 C farm n=35 | 合计 Total n=160 | |
3 | 0.0(0)a | 0.0(0)a | 2.9(1)a | 0.6(1) |
4 | 2.3(1)ab | 0.0(0)b | 5.7(2)a | 1.9(3) |
5 | 4.5(2)a | 0.0(0)a | 0.0(0)a | 1.3(2) |
6 | 38.6(17)a | 0.0(0)b | 80.0(28)c | 28.1(45) |
7 | 31.8(14)a | 1.2(1)b | 8.6(3)b | 11.3(18) |
8 | 20.5(9)a | 48.1(39)b | 2.9(1)a | 30.6(49) |
9 | 2.3(1)a | 17.3(14)b | 0.0(0)a | 9.4(15) |
10 | 0.0(0)a | 33.3(27)b | 0.0(0)a | 16.9(27) |
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