Evaluation of hydroxylamine oxidoreductase as a functional and phylogenetic marker to differentiate Nitrosomonas spp.
Evaluation of hydroxylamine oxidoreductase as a functional and phylogenetic marker to differentiate Nitrosomonas spp.
复制标题
DOI：
10.1002/jobm.201200378
复制
发表时间：
2014-04
影响因子：
3.1
通讯作者：
Radhika P. Keluskar;A. Desai
中科院分区：
文献类型：
--
作者：
Radhika P. Keluskar;A. Desai
亚硝化单胞菌属属于变形菌门的β -亚纲，包括密切相关的物种。本研究利用AOB特有的氨单加氧酶（amoA）和羟胺氧化还原酶（hao）基因片段，尝试单链确认多态性（SSCP）等序列无关技术来解析AOB。研究中观察到的分离株羟胺氧化还原酶（HAO）酶谱的变化也作为另一种与序列无关的方法来证实观察结果。通过amoA基因片段的SSCP分析和hao基因片段的SSCP分析，将欧洲亚硝化单胞菌（标准菌株）和环境样品富集得到的12株分离菌株分别分为6个和4个类群，而通过hao酶活性染色，将它们区分为6个不同的类群。在丰富度和均匀度方面，amoA基因片段优于hao基因片段，Simpson多样性指数为- 0.85。然而，综合使用这些分子方法（amoA和hao基因片段的SSCP）和hao酶谱在AOB指纹图谱中提供了更好的分辨率和均匀性，对AOB多样性研究有重要贡献。hao基因SSCP分析对AOB分离物的分类与基于16S rRNA基因的序列分析基本一致，因此适合作为系统发育标记。
Nitrosomonas genus belongs to beta‐subclass of Proteobacteria and encompasses closely related species. Sequence independent techniques like single strand confirmation polymorphism (SSCP) was attempted in the present study to resolve AOB using ammonia monooxygenase (amoA) and hydroxylamine oxidoreductase (hao) gene fragments, unique to AOB. Variation in hydroxylamine oxidoreductase (HAO) enzyme zymogram of isolates observed in the study was also explored as an additional sequence independent method to substantiate the observations. Nitrosomonas europaea (standard strain) and 12 isolates, obtained by enriching environmental samples, were differentiated into six and four groups by SSCP analyses of amoA and hao gene fragments, respectively, whereas they could be resolved into six distinct groups through activity staining of HAO enzyme. amoA gene fragment was therefore found to be better than hao gene fragment in resolving the studied AOB based on richness and evenness with Simpson's index of diversity – 0.85. However, the ensembled use of these molecular methods (SSCP of amoA and hao gene fragments) and HAO enzyme zymogram in fingerprinting AOB provide better resolution and evenness, contributing significantly in AOB diversity studies. Grouping of AOB isolates by hao gene SSCP analysis followed almost the same pattern as that by 16S rRNA gene based sequence analysis, hence it is suitable as a phylogenetic marker.