To investigate the prevalence and resistance against antimicrobials of
Antimicrobials, which play a crucial part in animal and human health care, are commonly used to treat bacterial infections and to prevent further spread of the infections within animal groups [
There are many kinds of molecular mechanisms underlying antimicrobial resistance in bacteria. The mechanisms mediated by integrons are an example of the most studied ones. Integrons, one kind of genetic element, are able to capture exogenous genes and disseminate them through plasmids or transposons in both interspecific and intraspecific ways [
Genotyping methods of bacteria have been important in surveillance and analysis, and they enable determination of sources of outbreaks. MLST is based on DNA sequencing data, which makes it specific, repeatable, comparable, and suitable for detecting the evolution of the strains and genetic analysis of microbial populations [
Most previous studies were focused on the prevalence of antimicrobial resistance of
From February to May 2016, 60 faecal samples (20 samples per farm) were randomly collected from diarrhea rabbits on farms in Ningyang, Xintai, and Dongping regions in Tai’an, China. The samples were independently collected from individual animals and the farms were chosen based on their scale (the breeding stock was >1500 heads). According to current statistics, there are about 15 relatively large rabbit farms in Tai’an, and each farm presumably produces >4000 heads annually. Because the sampling process did not harm the animals, ethical approval was not required for the study.
Isolation and identification of
Antimicrobial susceptibility was tested by the Minimal Inhibition Concentration (MIC) method. Bacteria were cultured at 37°C in the LB broth medium for 6 h. The concentration of
The
Primers for PCR in this study.
Primer name | Sequence (5′→ 3′) | Reference |
---|---|---|
| F: ATTCTTGAAGACGAAAGGGC | [ |
R: ACGCTCAGTGGAACGAAAAC | ||
| F: CACTCAAGGATGTATTGTG | [ |
R: TTAGCGTTGCCAGTGCTCG | ||
| F: ACACAATACATATCAACTTCGC | [ |
R: AGTGTGTTTAGAATGGTGATC | ||
| F: TTT GGT TCCGCG CTA TCT G | [ |
R: TAC TCC GAG CAC CAA ATC CG | ||
| F: CGCTTTGCGATGTGCAG | [ |
R: ACCGCGATATCGTTGGT | ||
| F: ATTTCTCACGCCAGGATTTG | [ |
R: TGCCAGGCACAGATCTTGAC | ||
| F: CGACCTKAGCGGCACTGAAT | [ |
R: GAGCAACGAYGCCTGGTAGYTG | ||
| F: ATGGAAACCTACAATCATAC | [ |
R: AAAAACACCTCGACTTAAGT | ||
| F: TTGCGATGCTCTATGAGTGGCTA | [ |
R: CTCGAATGCCTGGCGTGTTT | ||
| F: GCAGGTCCAGCAGCGGGTAG | [ |
R: CTTCCTGCCCGAGTATCGTG | ||
| F: ACCTACTCCCAACATCAGCC | [ |
R: ATATAGATCTCACTACGCGC | ||
| F: ACTGTGATGGGATACGCGTC | [ |
R: CTCCGTCAGCGTTTCAGCTA | ||
| F: CACAAGAACGTGGTCCGCTA | [ |
R: AACAGGTAAGCATCCGCATC | ||
| F: CTTCAGGATGGCAAGTTGGT | [ |
R: TCATCTCGTTCTCCGCTCAT | ||
| F: ATGTTACGCAGCAGGGCAGTCG | [ |
R: CGTCAGATCAATATCATCGTGC | ||
| F: TTGCGATGCTCTATGAGTGGCTA | [ |
R: CTCGAATGCCTGGCGTGTTT | ||
| F: GCGCCTTTCCTTTGGGTTCT | [ |
R: CCACCCGTTCCACGTTGTTA | ||
| F: CATTAATAGGCGCATCGCTG | [ |
R: TGAAGGTCATCGATAGCAGG | ||
| F: TGGTGACGGTGTTCGGCATTC | [ |
R: GCGAGGGTTTCCGAGAAGGTG | ||
| F: CGGCATCGTCAACATAACC | [ |
R: GTGTGCGGATGAAGTCAG | ||
| F: CATTCTAGAAAACAGTCGTAGTTCG | [ |
R: CATCTGCAGCTAACCTAGGGCTTTGGA | ||
| F: TGTCATTTACGGCATACTCG | [ |
R: ATCAGGCATCCCATTCCCAT | ||
| F: CTGAGGGTGTCGTCATCTAC | [ |
R: GCTCCGACAATGCTGACTAT |
The universal primers for the amplification of Class I integron gene cassette genes were detected by PCR. The primers of cassette FP and cassette RP were designed according to reference: 5′-TCATGGCTTGTTATGACTGT-3′ and 5′-GTAGGGCTTATTATGCACGC-3′. The amplification consisted of an initial denaturation at 94°C for 5 min, 30 cycles of denaturation at 94°C for 60 s, annealing at 56°C for 55 s, and extension at 68°C for 6 min. A final extension for 10 min at 72°C was also applied. PCR products were purified and sequenced for the further analysis [
According to
A total of 55
From Table
Susceptibility of
Antibiotics | Disk diffusion [ | ||
---|---|---|---|
S | I | R | |
AML | 53 (96.4%) | 0 | 2 (3.6%) |
AMK | 55 (100%) | 0 | 0 |
AMP | 15 (27.3%) | 4 (7.3%) | 36 (65.5%) |
C | 41 (74.5%) | 3 (5.5%) | 11 (20.0%) |
CAZ | 0 | 0 | 0 |
CIP | 43 (78.2%) | 2 (3.6%) | 10 (18.2%) |
CRO | 0 | 0 | 0 |
FOX | 54 (98.2%) | 0 | 1 (1.8%) |
GEN | 46 (83.6%) | 3 (5.5%) | 6 (10.9%) |
IMP | 0 | 0 | 0 |
NA | 10 (18.2%) | 15 (27.3%) | 30 (54.5%) |
SXT | 35 (63.6%) | 2 (3.6%) | 18 (32.7%) |
TB | 46 (83.6%) | 8 (14.5%) | 1 (1.8%) |
TET | 8 (14.5%) | 4 (7.3%) | 43 (78.2%) |
Resistance phenotype, ST, incidence of Class I integrons, and resistance gens in
Number | Location | ST | Resistance phenotype | Integrons/resistance |
---|---|---|---|---|
(1) | Ningyang | ST302 | AMP-NA-TET | |
(2) | Ningyang | ST302 | AMP-TET | |
(3) | Ningyang | ST302 | AMP-NA-TET | Class 1 |
(4) | Ningyang | ST302 | AMP-TET | |
(5) | Ningyang | ST468 | AMP-C-NA-SXT-TET | Class 1 |
(6) | Ningyang | ST302 | AMP-CIP-NA-SXT-TET | |
(7) | Ningyang | ST468 | AMP-C-NA-SXT-TET | Class 1 |
(8) | Ningyang | ST302 | AMP-TET | |
(9) | Ningyang | ST302 | AMP-TET | |
(10) | Ningyang | ST468 | AMP-CIP-NA-SXT-TET | |
(11) | Ningyang | ST302 | AMP-TET | Class 1 (dfrA1 + aadA1), |
(12) | Ningyang | ST370 | | |
(13) | Ningyang | ST370 | | |
(14) | Ningyang | ST302 | AMP-TET | |
(15) | Ningyang | ST370 | | |
(16) | Ningyang | ST302 | AMP-NA-TET | |
(17) | Ningyang | ST302 | AMP-TET | |
(18) | Ningyang | ST370 | | |
(19) | Xintai | ST370 | SXT | |
(20) | Xintai | ST87 | GEN-SXT | Class 1 |
(21) | Xintai | ST302 | AMP-GEN-SXT-TET | |
(22) | Xintai | ST302 | AMP-NA-TET | |
(23) | Xintai | ST370 | | |
(24) | Xintai | ST302 | | |
(25) | Xintai | ST314 | NA-TET | Class 1 |
(26) | Xintai | ST302 | AMP-C-NA-SXT-TET | |
(27) | Xintai | ST302 | AMP-FOX-NA-TET | |
(28) | Xintai | ST302 | TET | |
(29) | Xintai | ST302 | AMP-NA-SXT | Class 1 |
(30) | Xintai | ST636 | | |
(31) | Xintai | ST370 | AMP-NA-TET | |
(32) | Xintai | ST468 | AMP-NA-TET | |
(33) | Xintai | ST468 | AMP-C-NA-SXT | Class 1 |
(34) | Xintai | ST302 | AMP-NA-TET | |
(35) | Xintai | ST302 | AMP-NA-TET | |
(36) | Xintai | ST302 | AMP-NA-TET | |
(37) | Xintai | ST302 | AMP -TET | Class 1 |
(38) | Xintai | ST370 | NA-TET | Class 1 |
(39) | Dongping | ST461 | AMP-NA-SXT | |
(40) | Dongping | ST731 | TET | |
(41) | Dongping | ST739 | AMP-NA-TET | |
(42) | Dongping | ST370 | TET | |
(43) | Dongping | ST739 | AMP-CIP- NA-SXT-TET | |
(44) | Dongping | ST370 | TET | |
(45) | Dongping | ST88 | AML-AMP-C-CIP-GEN- NA-SXT-TET | Class 1 |
(46) | Dongping | ST370 | TET | |
(47) | Dongping | ST739 | AMP-CIP-NA-SXT-TET | |
(48) | Dongping | ST88 | AMP-C-CIP-GEN-NA-SXT-TET | |
(49) | Dongping | ST2 | AMP-C-CIP-NA-SXT-TET | |
(50) | Dongping | ST88 | AMP-C-CIP-GEN-NA-SXT-TET | Class 1 |
(51) | Dongping | ST353 | C-TET | Class 1 |
(52) | Dongping | ST370 | NA-TET | Class 1 |
(53) | Dongping | ST88 | C-CIP-NA-TET | Class 1 |
(54) | Dongping | ST24 | AML-AMP-TET | Class 1 |
(55) | Dongping | ST88 | AMP-C-CIP-GEN-NA-SXT-TB-TET | Class 1 |
Two kinds of
Class I integrons were detected in 17
13 different kinds of STs were identified among all the 55 strains, including 3 STs (ST302, ST468, and ST370) from samples derived from Ningyang, 6 STs (ST87, ST302, ST314, ST370, ST468, and ST636) from samples derived from Xintai, and 8 STs (ST2, ST24, ST88, ST353, ST370, ST461, ST731, and ST739) from samples derived from Dongping. It was shown that ST302 (22/55, 40.0%) was the most prevalent type, followed by ST370 (12/55, 21.8%) (Table
Colibacillosis of rabbits, a common disease in rabbits breeding, had become one of the major infectious diseases that endanger the rabbit breeding industry [
In this study, Class I integrons were detected in 17
In this study, the highest isolation rate of 40% (22/55) was found for ST302 which had not yet been reported related to infections. It was different from that of previous studies which found that ST40 is the most common ST from rabbits in Italy [
Our findings exhibit the molecular characterization of antimicrobial resistance in
The authors declare that they have no conflicts of interest.
Xiaonan Zhao and Jie Yang have contributed equally to this work.
This work was supported by the National Key R&D Project of China (2017YFD0500105 and 2016YFD0501608), Taishan Scholars Program (201511023), and Funds of Shandong “Double Tops” Program.