Ortiz-Román Luisa, Riquelme-Neira Roberto, Oñate Angel
Laboratory of Molecular Immunology, Department of Microbiology, Faculty of Biological Sciences, Universidad de Concepción, Concepción, Chile; Biogenesis Research Group, Faculty of Agricultural Sciences, Universidad de Antioquia, Medellín, Colombia.
Laboratory of Molecular Immunology, Department of Microbiology, Faculty of Biological Sciences, Universidad de Concepción, Concepción, Chile.
Vet Microbiol. 2014 Aug 6;172(1-2):279-84. doi: 10.1016/j.vetmic.2014.05.005. Epub 2014 May 14.
One of the properties of bacteria is their capacity to acquire large fragments of genomic DNA from other bacteria or to loose important parts of their own genome. Such fragments include genomic islands (GIs); nine GIs are present in Brucella, including genomic island 3 (GI-3), present in B. abortus, B. melitensis and B. ovis. The GI-3 have 29 open reading frames (ORFs) most of them with unknown function. Within the GI-3, the ORFs BAB1_0267 encodes a hypothetical protein sharing a SH3 domain and BAB1_270 a zinc-dependent metallopeptidase. We have obtained deletion mutants for BAB1_0267 and BAB1_0270 ORFs present within GI-3, which have been named the Δ0267 and Δ0270, respectively; in both cases the mutation did not affect the growth of bacteria. Both mutants were evaluated with respect to their growth rates, their ability to invade and replicate in the non-professional and professional phagocytes, HeLa and J774.A1 cells, respectively. Their persistence in the spleens of mice was also evaluated. The mutants efficiently invaded HeLa and J774.A1 cells but both mutants showed a decreased intracellular survival in macrophages and HeLa cells 72 and 96 h post-infection, respectively, and were non-detected in J774.A1 cells 120 h post infection. With respect to in vivo persistence Δ0267 was detected through the fourth week while Δ0270 decreased at 7 days disappearing the second week. Our results indicated that deletion of BAB1_0267 and BAB1_270 are necessary to establish an optimal infectious process in B. abortus 2308, having more effect the deletion of ORF BAB1_0270. Therefore these ORFs, principally BAB1_0270 are important virulent of B. abortus.
细菌的特性之一是它们能够从其他细菌获取大片段基因组DNA，或者丢失自身基因组的重要部分。这些片段包括基因组岛（GI）；布鲁氏菌中有九个基因组岛，其中基因组岛3（GI-3）存在于流产布鲁氏菌、羊种布鲁氏菌和绵羊布鲁氏菌中。GI-3有29个开放阅读框（ORF），其中大多数功能未知。在GI-3内，ORF BAB1_0267编码一种具有SH3结构域的假定蛋白，BAB1_270编码一种锌依赖性金属肽酶。我们获得了GI-3内存在的BAB1_0267和BAB1_0270 ORF的缺失突变体，分别命名为Δ0267和Δ0270；在这两种情况下，突变均不影响细菌的生长。对这两种突变体的生长速率、在非专职和专职吞噬细胞（分别为HeLa和J774.A1细胞）中侵袭和复制的能力进行了评估。还评估了它们在小鼠脾脏中的持久性。突变体能够有效侵袭HeLa和J774.A1细胞，但在感染后72小时和96小时，两种突变体在巨噬细胞和HeLa细胞中的细胞内存活率均下降，且在感染后120小时在J774.A1细胞中未检测到。关于体内持久性，Δ0267在第四周仍可检测到，而Δ0270在第7天减少，在第二周消失。我们的结果表明，缺失BAB1_0267和BAB1_270对于流产布鲁氏菌2308建立最佳感染过程是必要的，其中ORF BAB1_0270的缺失影响更大。因此，这些ORF，主要是BAB1_0270，是流产布鲁氏菌的重要毒力因子。
