Harper Marina, John Marietta, Turni Conny, Edmunds Mark, St Michael Frank, Adler Ben, Blackall P J, Cox Andrew D, Boyce John D
Department of Microbiology, Monash University, Melbourne, Australia Australian Research Council Centre of Excellence in Structural and Functional Microbial Genomics, Monash University, Melbourne, Australia.
Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, Brisbane, Australia.
J Clin Microbiol. 2015 Feb;53(2):477-85. doi: 10.1128/JCM.02824-14. Epub 2014 Nov 26.
Pasteurella multocida is a Gram-negative bacterial pathogen that is the causative agent of a wide range of diseases in many animal species, including humans. A widely used method for differentiation of P. multocida strains involves the Heddleston serotyping scheme. This scheme was developed in the early 1970s and classifies P. multocida strains into 16 somatic or lipopolysaccharide (LPS) serovars using an agar gel diffusion precipitin test. However, this gel diffusion assay is problematic, with difficulties reported in accuracy, reproducibility, and the sourcing of quality serovar-specific antisera. Using our knowledge of the genetics of LPS biosynthesis in P. multocida, we have developed a multiplex PCR (mPCR) that is able to differentiate strains based on the genetic organization of the LPS outer core biosynthesis loci. The accuracy of the LPS-mPCR was compared with classical Heddleston serotyping using LPS compositional data as the "gold standard." The LPS-mPCR correctly typed 57 of 58 isolates; Heddleston serotyping was able to correctly and unambiguously type only 20 of the 58 isolates. We conclude that our LPS-mPCR is a highly accurate LPS genotyping method that should replace the Heddleston serotyping scheme for the classification of P. multocida strains.
多杀性巴氏杆菌是一种革兰氏阴性细菌病原体，是包括人类在内的许多动物物种中多种疾病的病原体。一种广泛用于区分多杀性巴氏杆菌菌株的方法涉及赫德尔斯顿血清分型方案。该方案于20世纪70年代初开发，使用琼脂凝胶扩散沉淀试验将多杀性巴氏杆菌菌株分为16种菌体或脂多糖（LPS）血清型。然而，这种凝胶扩散试验存在问题，在准确性、可重复性以及优质血清型特异性抗血清的来源方面都存在困难。利用我们对多杀性巴氏杆菌LPS生物合成遗传学的了解，我们开发了一种多重PCR（mPCR），能够根据LPS外核心生物合成位点的基因组织来区分菌株。将LPS-mPCR的准确性与使用LPS组成数据作为“金标准”的经典赫德尔斯顿血清分型进行了比较。LPS-mPCR对58株分离株中的57株进行了正确分型；赫德尔斯顿血清分型仅能对58株分离株中的20株进行正确且明确的分型。我们得出结论，我们的LPS-mPCR是一种高度准确的LPS基因分型方法，应取代赫德尔斯顿血清分型方案用于多杀性巴氏杆菌菌株的分类。
