Analysis Of Biological Characteristics Of Klebsiella Pneumoniae In A Zoo Of Eastern Area Of China

Present study is designed to identify the cause of acute death in female monkey Nomascus leucogenys at Fuzhou zoo.The lesions of disease were observed by necropsy examination and by recording the pathological changes in the tissue of internal organs.Different internal organ tissue were collected and observed under microscope by H&E staining method.We also isolated the causative bacterium and characterized it by VITEK-2 automatic identification analyzer.Later we also performed biochemical methods like use of carbon and nitrogen sources,dynamic experiment,and oxidase test to confirm the bacterial specie.Bacterial colonies were observed for physiological characteristics on a TSA plate under aerobic,anaerobic and microaerophillic conditions.The observation of colony characteristics is taken on several identified and selective medium plates.Minimum inhibitory concentrations(MICs)of different antibiotics were determined by GN-13 susceptibility test card.Genetic Identification:PCR amplification was conducted for partial gene DNA of bacterial 16S rDNA and the sequencing result was blasted to find the closest Parental Origins in NCBI.Determination of Pathogenesis:To checkpathogenes several mice were injected with bacteria via three different methods,intraperitoneal injection,subcutaneous injection and intramuscular injection.The overall changes of appearance,mental state,body weight and internal changes appearance,as well as the pathological changes of different organs were abserved.Conclusion:(1)The body of dead Nomascus leucogenys was dissected and observed from the angle of substantial organs lesions and histopathologic changes.The heart surface was smooth;mural thrombus appeared in right ventricle;hyperemia and edema occurred in myocardial matrix;local lesion of intima was preminent;myocardial fibrosis happened beneath the intima,accompanied with mural thrombosis.The lung surface was grey white;the section was red brown;thromboembolism was seen in local bronchi artery;expansion and hyperemia of capillaries was observed on alveolar wall;fusion of pulmonary alveoli was found.Liver capsule was complete;the hepatic lobule structure was normal;expansion and congestion was seen in central veins and hepatic sinusoid.Other substantial organs presented no obvious clinical and pathological lesions.Moreover,congestion occurred in respiratory system only.Acute respiratory failure caused by pulmonary thromboembolism is assumed to be the direct reason of death.(2)Klebsiella pneumonia separated from the heart blood was biochemically identified as pneumonia subspecies by VITEK-2 COMPACT system,with the probability of 95%.The supplementary biochemical identification results met the biochemical characteristics of Klebsiella pneumonia subspecies.Streak inoculation was conducted for this bacterium in 2%bovine serum TSA.The bacterium grew vigorously under micro aerobic and aerobic conditions,while it grew slowly under anaerobic condition.Characteristic bacterial colony could be observed on differential medium and enteric bacteria selective medium.The experimental result of combining VITEK-2 COMPACT system and AST-GN13 drug allergy was ESBLs(-).In other words,this bacterium will not generate Extended-Spectrum ?-Lactamases,and it can resist ampicillin.(3)Isolated culture was carried out to purify Klebsiella pneumonia;PCR reaction conditions and programs were designed.16SrDNA sequence alignment result of PCR purified product presented high homology with Klebsiella pneumonia and reported in GeneBank,reaching 99.3%.(4)The death rate of rats with abdominal infection of high-concentration bacterium liquid(about 2.0×1010 CFU/mL)was 100%.No death happened in intramuscular injection group and subcutaneous injection group.According to the dissection and examination result of dead rats,swelling of varying degrees was observed in their splenic organs,thymus glands and lungs;hemorrhagic spot existed on the surface;the liver color was deepened and hemorrhagic spot also appeared.Rats can tolerate Klebsiella pneumonia at the concentration lower than 3.0×108CFU/mL.Therefore,after the tolerance period,the weight of rats increased,and the increase rate of weight gain was similar to that of normal saline group.