Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523, USA.
J Bacteriol. 2011 Jul;193(13):3265-75. doi: 10.1128/JB.00093-11. Epub 2011 Apr 29.
Genotyping of Francisella tularensis (A1a, A1b, A2, and type B) and Francisella novicida has identified multiple differences between species and among F. tularensis subspecies and subpopulations. Variations in virulence, geographic distribution, and ecology are also known to exist among this group of bacteria, despite the >95% nucleotide identity in their genomes. This study expands the description of phenotypic differences by evaluating the ability of F. tularensis and F. novicida to degrade chitin analogs and produce active chitinases. Endochitinase activities were observed to vary among F. tularensis and F. novicida strains. The activity observed for F. tularensis strains was predominantly associated with whole-cell lysates, while the chitinase activity of F. novicida localized to the culture supernatant. In addition, the overall level of chitinase activity differed among the subpopulations of F. tularensis and between the species. Bioinformatic analyses identified two new putative chitinase genes (chiC and chiD), as well as the previously described chiA and chiB. However, the presence of these four open reading frames as intact genes or pseudogenes was found to differ between Francisella species and F. tularensis subspecies and subpopulations. Recombinant production of the putative chitinases and enzymatic evaluations revealed ChiA, ChiB, ChiC, and ChiD possessed dissimilar chitinase activities. These biochemical studies coupled with bioinformatic analyses and the evaluation of chiA and chiC knockouts in F. tularensis A1 and A2 strains, respectively, provided a molecular basis to explain the differential chitinase activities observed among the species and subpopulations of Francisella.
对土拉弗朗西斯菌（A1a、A1b、A2 和 B 型）和新疏螺旋体的基因分型鉴定表明，在物种之间以及在土拉弗朗西斯菌亚种和亚群之间存在多种差异。尽管它们的基因组中有>95%的核苷酸同一性，但已知在毒力、地理分布和生态方面也存在差异。本研究通过评估土拉弗朗西斯菌和新疏螺旋体降解壳聚糖类似物和产生活性壳聚糖酶的能力，扩展了表型差异的描述。观察到土拉弗朗西斯菌和新疏螺旋体菌株之间的内切几丁质酶活性存在差异。观察到的土拉弗朗西斯菌菌株的活性主要与全细胞裂解物相关，而新疏螺旋体的壳聚糖酶活性定位于培养上清液中。此外，土拉弗朗西斯菌亚群和物种之间的壳聚糖酶活性总体水平存在差异。生物信息学分析鉴定了两个新的推定的几丁质酶基因（chiC 和 chiD），以及先前描述的 chiA 和 chiB。然而，这些四个开放阅读框作为完整基因或假基因的存在，在弗朗西斯菌属物种和土拉弗朗西斯菌亚种和亚群之间存在差异。推定的几丁质酶的重组生产和酶学评估表明 ChiA、ChiB、ChiC 和 ChiD 具有不同的几丁质酶活性。这些生化研究结合生物信息学分析和土拉弗朗西斯菌 A1 和 A2 菌株中 chiA 和 chiC 敲除的评估，为解释在物种和弗朗西斯菌亚群之间观察到的差异几丁质酶活性提供了分子基础。
