| Klebsiella pneumoniae is a common opportunistic pathogen,which can cause pneumonia,enteritis and septicemia in animals under certain conditions.In recent years,the isolation rate of multi-drug resistant K.pneumoniae is increasing,which increases the difficulty of clinical treatment of bacterial infections.Maxingshigan granules,a traditional Chinese medicine formula for treating respiratory tract infections,which has many kinds of pharmacological action including but not limited to anti-inflammatory,antiviral effect,and antibacterial effect,and it is a refined preparation of Maxingshigan decoction.In this study,the prevalence and drug resistance of K.pneumoniae from fresh feces of chicks and laying hens were investigated and analyzed.The antibacterial effect of Maxingshigan granules on multidrug-resistant K.pneumoniae isolates in vitro was also explored,in order to provide a theoretical basis for clinical drug use,animal health breeding and clinical animal drug resistance monitoring.The results are as following:（1）Isolation and identification of K.pneumoniae:In this study,60 fresh feces of healthy chicks and laying hens were collected from a large-scale layer farm in Ningxia.A total of 48K.pneumoniae strains were identified by PCR and VITEK2 Compact test,including 36 strains（36/36）from chicks and 12 strains（12/24）from laying hens.The isolation rate of K.pneumoniae in chick feces was significantly higher than that in laying hens.（2）Antibiotic resistance analysis and virulence gene detection of K.pneumoniae isolates:The sensitivity of 48 K.pneumoniae strains to 13 antibacterial drugs was tested by standard broth microdilution method.Results show that K.pneumoniae isolates had high resistance to spectinomycin（79.17%）,tetracycline（81.25%）,florfenicol（50.00%）,sulfisoxazole（72.91%）and trimethoprim-sulfamethoxazole（72.91%）.Isolates showed low resistance to amoxicillin/clavulanic acid,cephalosporins and quinolones,with resistance rates ranging from14.58%to 27.08%.All isolates were sensitive to colistin and meropenem.Multi-drug resistant strains accounted for 72.92%of the isolates,and 10.42%strains were resistant to seven kinds of antibacterial drugs.The 4 multi-drug resistance were prevalent（22.92%）.The resistance rate of chick fecal isolates was higher than that of laying hens.The carrying distribution of drug resistance and virulence genes of isolates was detected by PCR.A total of 6 drug resistance genes and 4 virulence genes were detected,including aad A1,tet A,oqx A,oqx B,bla TEM and qnr B genes,uge,wab G,kfu BC and ent B.（3）Antibacterial activity of Maxingshigan granules against multidrug-resistant K.pneumoniae:The effect of Maxingshigan granules on the growth of multidrug-resistant K.pneumoniae was determined by microbroth dilution method and time-growth curve.The antibacterial effect of Maxingshigan granules combined with antibacterial drugs and the effect on the expression level of drug-resistant genes were performed by checkerboard method and RT-PCR.Maxingshigan granules can inhibit the growth of multidrug-resistant K.pneumoniae isolates and delay the entry of K.pneumoniae from the lag phase into the logarithmic growth phase,and the MIC value was 125 mg/m L.The combination of Maxingshigan granules and enrofloxacin showed synergistic effect or additive effect on K.pneumoniae isolates.The relative expressions of bla TEM,aad A1 and tet A m RNA of K.pneumoniae were significantly down-regulated after the treatment of Maxingshigan granules（P<0.01）,while the transcription levels of oqx B and oqx A were not significantly affected（P>0.05）.In conclusion,the drug resistance of K.pneumoniae from laying hens is relatively serious,mainly in chicks.Maxingshigan granules have bacteriostatic activity against multidrug-resistant K.pneumoniae,and can significantly inhibit the expression of partial drug resistance genes of K.pneumoniae. |
