| Haemophilus parasuis is gram negative bacterium, which can cause swine polyserositis, meningitis,arthritis and other diseases in pigs. H. parasuis is conditional pathogenic bacteria and often causesecondary infections in some viral diseases such as porcine pseudorabies virus, swine flu and classicalswine fever.In addition, H. parasuis could have mixed infection with immune suppression diseases,such as porcine reproductive and respiratory syndrome syndrome and porcine circovirus disease andresulted in major economic loses in swine farm, At present, the prevention and treatment of the H.parasuis infection mainly depends on antibiotic use. However, Due to the long-term and irrational useof antibiotics in veterinary clinical, antibiotic resistance has emerged, which brings great challenges forprevention and treatment of the infections caused by drug-resistant strains. It is very importmant tocarry out long-term monitoring on the drug sensitivity for the control and prevention of H.parasuisinfection and study on mechanisms of antibiotic resistance could provide the theoretical support forcontrolling the spread of resistant strains.The aim of this study was to investigate antibiotic resistance in H.parasuis isolates in Shanghai andthereby to provide data support for strategy of antibiotic use in clinical and prevention of H. parasuisinfection. 62 H. parasuis were isolated from 484 samples. Minimal inhibitory concentrations（MICs） of21 antibiotics were determined by agar dilution method. Susceptibility test showed that all isolates weresusceptible to azithromycin, tilmicosin, tiamulin, amikacin, ceftiofur and levofloxacin. At least 90% ofthem were susceptible to erythromycin（60, 96.8%）, florfenicol（59, 95.3%）, chloramphenicol（60, 96.8%）,gentamicin（60, 96.8%）, spectinomycin（61, 98.4%）, penicillin（57, 91.9%）, ampicillin（57, 91.9%） andciprofloxacin（61, 98.4%）. Resistance to tetracycline and enrofloxacin were found in 75.8% and 30.6%of the isolates, respectively. Resistance to at least 2 antibiotics was observed in 37.1% of the isolates,9.7% strains are resistant to more than three kinds of antibiotics. The most high resistant strain canresistant to 5 kinds of drugs, but the proportion is low（1,1.5%）.In this study, we firstly identified 3 H. parasuis strains that showed florfenicol resistance. PCRdetection and subsequent sequencing showed that the three H. parasuis isolates carried the efflux pumpgene floR. It is 1215 bp, consistented to other the size of floR gene in the genbank database. It showed97% identify with amino acid sequence of the floR gene found in Escherichia coli（GenBank AccessionNo:NC012692）. Plasmid extraction, electrophoresis and southern blot showed that floR gene is locatedon a 6 kb plasmid in all the three isolates, but not on the genome. The three plasmids were completelysequenced by Primer Walking. Sequnece anlysis showed that the three floR-carrying plasmids wereexact identical. This plasmid was named pHPSF1. Bioinformatics analysis showed that the plasmidpHPSF1 contains 5 open reading frames, including three genes involved in plasmidmobilization andreplication（mob A/L, mobC and Rep）, florfenicol resistance gene floR and transcriptional regulator lysR.Database search and sequence comparison showed that the fragments of plasmid pHPSF1 exhibited highhomology with several resistant plasmid identified in animal pathogens, suggesting that, under theantibiotic selection pressure, several recombination events take place between these small plasmids andfinally results in new resistance plasmid. The results of this study showed that the situation of antibioticresistance is not serious in H. Parasuis isolates from Shanghai. However, some strains that wereresistant to the antibiotics used in clinical have emerged. It is necessary to carry out long-term monitorthe resistant strains further study about the transmission of resistant strains, in order to more effectivelyprevent and control the epidemic of drug-resistant strains to guarantee the economic benefits oflivestock breeding. |
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