Infection and Immunity Program, Monash Biomedicine Discovery Institute and Department of Microbiology, Monash Universitygrid.1002.3, Melbourne, Victoria, Australia.
Microbiol Spectr. 2022 Apr 27;10(2):e0019522. doi: 10.1128/spectrum.00195-22. Epub 2022 Apr 11.
Pasteurella multocida is a Gram-negative capsulated bacterium responsible for a range of diseases that cause severe morbidity and mortality in livestock animals. The hyaluronic acid (HA) capsule produced by P. multocida serogroup A strains is a critical virulence factor. In this study, we utilized transposon-directed insertion site sequencing (TraDIS) to identify genes essential for growth of P. multocida and combined TraDIS with discontinuous density gradients (TraDISort) to identify genes required for HA capsule production and regulation in this pathogen. Analysis of mutants with a high cell density phenotype, indicative of the loss of extracellular capsule, led to the identification of 69 genes important for capsule production. These genes included all previously characterized genes in the capsule biosynthesis locus and and , which encode known positive regulators of P. multocida capsule. Many of the other capsule-associated genes identified in this study were involved in regulation or activation of the stringent response, including and , which encode proteins that regulate the concentration of guanosine alarmones. Disruption of the autoregulatory domains in the C-terminal half of SpoT using insertional mutagenesis resulted in reduced expression of capsule biosynthesis genes and an acapsular phenotype. Overall, these findings have greatly increased the understanding of hyaluronic acid capsule production and regulation in P. multocida. The bacterial pathogen P. multocida can cause serious disease in production animals, including fowl cholera in poultry, hemorrhagic septicemia in cattle and buffalo, atrophic rhinitis in pigs, and respiratory diseases in a range of livestock. P. multocida produces a capsule that is essential for systemic disease, but the complete mechanisms underlying synthesis and regulation of capsule production are not fully elucidated. A whole-genome analysis using TraDIS was undertaken to identify genes essential for growth in rich media and to obtain a comprehensive characterization of capsule production. Many of the capsule-associated genes identified in this study were involved in the stringent response to stress, a novel finding for this important animal pathogen.
多杀巴斯德菌是一种革兰氏阴性有荚膜的细菌，可导致多种疾病，使家畜动物严重发病和死亡。多杀巴斯德菌血清 A 群菌株产生的透明质酸 (HA) 荚膜是一个关键的毒力因子。在这项研究中，我们利用转座子定向插入位点测序（TraDIS）来鉴定生长所必需的基因多杀巴斯德菌，并将 TraDIS 与不连续密度梯度（TraDISort）相结合，鉴定生产 HA 荚膜和调节的必需基因这种病原体。对具有高细胞密度表型的突变体进行分析，表明细胞外荚膜丢失，导致鉴定出 69 个对荚膜产生重要的基因。这些基因包括荚膜生物合成基因座中的所有先前表征的基因，以及编码多杀巴斯德菌荚膜的已知正调控因子的 和 。在这项研究中鉴定的许多其他与荚膜相关的基因参与了严格反应的调节或激活，包括编码调节鸟嘌呤警报素浓度的蛋白质的 和 。使用插入诱变破坏 SpoT C 端半部分的自身调节结构域导致荚膜生物合成基因表达减少和无荚膜表型。总的来说，这些发现极大地增加了对多杀巴斯德菌 HA 荚膜产生和调节的理解。细菌病原体多杀巴斯德菌可导致生产动物发生严重疾病，包括家禽的禽霍乱、牛和水牛的出血性败血症、猪的萎缩性鼻炎以及多种家畜的呼吸道疾病。多杀巴斯德菌产生荚膜，对全身疾病至关重要，但荚膜产生的合成和调节的完整机制尚未完全阐明。使用 TraDIS 进行全基因组分析，以鉴定在丰富培养基中生长所必需的基因，并对荚膜产生进行全面表征。在这项研究中鉴定的许多与荚膜相关的基因都参与了对压力的严格反应，这是对这种重要的动物病原体的一个新发现。
