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首页> 外文期刊>Infection and immunity >Inactivation of Pasteurella(Mannheimia) haemolytica Leukotoxin Causes Partial Attenuation of Virulence in a Calf Challenge Model
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Inactivation of Pasteurella(Mannheimia) haemolytica Leukotoxin Causes Partial Attenuation of Virulence in a Calf Challenge Model

机译：小牛挑战模型中溶血巴斯德氏菌（曼海姆）失活白细胞毒素导致部分毒力减弱。

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The leukotoxin of Pasteurella (Mannheimia)haemolytica is believed to play a significant role in pathogenesis, causing cell lysis and apoptosis that lead to the lung pathology characteristic of bovine shipping fever. Using a system for Cre-lox recombination, a nonpolar mutation within thelktC transacylase gene of the leukotoxin operon was created. The lktC locus was insertionally inactivated using a loxP-aph3-loxP cassette, and then the aph3marker was excised from the chromosome by Cre recombinase expressed from a P. haemolytica plasmid. The resultinglktC strain (SH2099) secretes inactive leukotoxin and carries no known antibiotic resistance genes. Strain SH2099 was tested for virulence in a calf challenge model. We inoculated 3 × 108 or 3 × 109 CFU of wild-type or mutant bacteria into the lungs of healthy, colostrum-deprived calves via transthoracic injection. Animals were observed for clinical signs and for nasal colonization for 4 days, after which they were euthanized and necropsied. The lower inoculum (3 × 108 CFU) caused significantly fewer deaths and allowed lung pathology to be scored and compared, while the 3 × 109 CFU dose of either the wild-type or mutant was lethal to ≥50% of the calves. The estimated 50% lethal dose of SH2099 was four times higher than that of the wild-type strain. Lung lesion scores were reduced twofold in animals inoculated with the mutant, while clinical scores were nearly equivalent for both strains. The wild-type and mutant strains were equally capable of colonizing the upper respiratory tracts of the calves. In this study, the P. haemolytica lktC mutant was shown to be less virulent than the parent strain.
机译：溶血巴斯德氏杆菌（曼氏假单胞菌）的白细胞毒素被认为在发病机理中起重要作用，引起细胞裂解和细胞凋亡，从而导致牛运输热的肺部病理特征。使用Cre-lox重组系统，在白细胞毒素操纵子的thelktC转酰基酶基因内创建了一个非极性突变。使用loxP-aph3-loxP盒插入灭活lktC基因座，然后通过溶血毕赤酵母质粒表达的Cre重组酶从染色体上切除aph3marker。所得的lktC菌株（SH2099）分泌失活的白细胞毒素且不携带已知的抗生素抗性基因。在小牛激发模型中测试菌株SH2099的毒力。我们通过经胸腔注射将3×108或3×109 CFU的野生型或突变细菌接种到健康的，初乳剥夺的小牛的肺中。观察动物的临床体征和鼻部定植4天，然后对其进行安乐死并进行尸检。较低的接种量（3×108 CFU）造成的死亡显着减少，并且可以对肺病理进行评分和比较，而野生型或突变体的3×109 CFU剂量对小牛的≥50％致死。估计SH2099的50％致死剂量是野生型菌株的四倍。接种该突变体的动物的肺部病变评分降低了两倍，而两种菌株的临床评分几乎相等。野生型和突变型菌株同样能够在小牛的上呼吸道定居。在这项研究中，溶血性疟原虫lktC突变体显示出比亲本菌株低的毒性。

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