Department of Crop Sciences, Tshwane University of Technology, Pretoria, South Africa.
Department of Chemistry, Tshwane University of Technology, Arcadia Campus, Pretoria, South Africa
Appl Environ Microbiol. 2018 Jan 2;84(2). doi: 10.1128/AEM.01712-17. Print 2018 Jan 15.
Cowpea derives most of its N nutrition from biological nitrogen fixation (BNF) via symbiotic bacteroids in root nodules. In Sub-Saharan Africa, the diversity and biogeographic distribution of bacterial microsymbionts nodulating cowpea and other indigenous legumes are not well understood, though needed for increased legume production. The aim of this study was to describe the distribution and phylogenies of rhizobia at different agroecological regions of Mozambique using PCR of the BOX element (BOX-PCR), restriction fragment length polymorphism of the internal transcribed spacer (ITS-RFLP), and sequence analysis of ribosomal, symbiotic, and housekeeping genes. A total of 122 microsymbionts isolated from two cowpea varieties (IT-1263 and IT-18) grouped into 17 clades within the BOX-PCR dendrogram. The PCR-ITS analysis yielded 17 ITS types for the bacterial isolates, while ITS-RFLP analysis placed all test isolates in six distinct clusters (I to VI). BLAST sequence analysis of 16S rRNA and four housekeeping genes (, , , and ) showed their alignment with and species. The results revealed a group of highly diverse and adapted cowpea-nodulating microsymbionts which included , , , and a novel sp., as well as , , and in Mozambican soils. Discordances observed in single-gene phylogenies could be attributed to horizontal gene transfer and/or subsequent recombinations of the genes. Natural deletion of 60 bp of the region was observed in isolate TUTVU7; however, this deletion effect on DNA gyrase function still needs to be confirmed. The inconsistency of with core gene phylogenies suggested differences in the evolutionary history of both chromosomal and symbiotic genes. A diverse group of both and species responsible for cowpea nodulation in Mozambique was found in this study. Future studies could prove useful in evaluating these bacterial isolates for symbiotic efficiency and strain competitiveness in Mozambican soils.
豇豆主要通过根瘤中的共生菌固氮（BNF）从生物固氮中获取氮营养。在撒哈拉以南非洲，豆科植物和其他本地豆科植物的结瘤细菌微生物共生体的多样性和生物地理分布尚不清楚，但对于增加豆科植物的产量是必要的。本研究的目的是使用 BOX 元件（BOX-PCR）聚合酶链反应、内部转录间隔区（ITS-RFLP）的限制性片段长度多态性以及核糖体、共生和管家基因的序列分析，描述莫桑比克不同农业生态区的根瘤菌分布和系统发育。从两个豇豆品种（IT-1263 和 IT-18）中分离出的 122 个微生物共生体在 BOX-PCR 系统发育树中分为 17 个分支。PCR-ITS 分析为细菌分离物产生了 17 种 ITS 类型，而 ITS-RFLP 分析将所有测试分离物置于六个不同的聚类（I 至 VI）中。16S rRNA 和四个管家基因（、、、和）的 BLAST 序列分析表明，它们与和物种的序列一致。结果表明，存在一群高度多样化和适应豇豆结瘤的微生物共生体，其中包括、、和一种新的、以及、和在莫桑比克土壤中。单基因系统发育中观察到的不匹配可能归因于基因的水平转移和/或随后的重组。在分离株 TUTVU7 中观察到 区域的 60bp 自然缺失；然而，这种缺失对 DNA 拓扑异构酶功能的影响仍需确认。与核心基因系统发育不一致表明，染色体和共生基因的进化历史存在差异。本研究发现，莫桑比克豇豆结瘤的微生物共生体包括和物种的一个多样化群体。未来的研究可以证明评估这些细菌分离物在莫桑比克土壤中的共生效率和菌株竞争力是有用的。
