Marine Science Institute, College of Science, University of the Philippines Diliman, 1101, Quezon City, Philippines.
National Institute of Molecular Biology and Biotechnology, College of Science, University of the Philippines Diliman, 1101, Quezon City, Philippines.
Mar Biotechnol (NY). 2018 Dec;20(6):685-705. doi: 10.1007/s10126-018-9840-6. Epub 2018 Jun 23.
Carbohydrate-active enzymes (CAZymes) have significant biotechnological potential as agents for degradation or modification of polysaccharides/glycans. As marine macroalgae are known to be rich in various types of polysaccharides, seaweed-associated bacteria are likely to be a good source of these CAZymes. A genomics approach can be used to explore CAZyme abundance and diversity, but it can also provide deep insights into the biology of CAZyme producers and, in particular, into molecular mechanisms that mediate their interaction with their hosts. In this study, a Gram-negative, aerobic, rod-shaped, carrageenolytic, and culturable marine bacterium designated as AOL6 was isolated from a diseased thallus of a carrageenan-producing farmed rhodophyte, Kappaphycus alvarezii (Gigartinales, Rhodophyta). The whole genome of this bacterium was sequenced and characterized. Sequence reads were assembled producing a high-quality genome assembly. The estimated genome size of the bacterium is 4.4 Mb and a G+C content of 52%. Molecular phylogenetic analysis based on a complete sequence of 16S rRNA, rpoB, and a set of 38 single-copy genes suggests that the bacterium is an unknown species and represents a novel genus in the family Cellvibrionaceae that is most closely related to the genera Teredinibacter and Saccharophagus. Genome comparison with T. turnerae T7901 and S. degradans 2-40 reveals several features shared by the three species, including a large number of CAZymes that comprised > 5% of the total number of protein-coding genes. The high proportion of CAZymes found in the AOL6 genome exceeds that of other known carbohydrate degraders, suggesting a significant capacity to degrade a range of polysaccharides including κ-carrageenan; 34% of these CAZymes have signal peptide sequences for secretion. Three putative κ-carrageenase-encoding genes were identified from the genome of the bacterium via in silico analysis, consistent with the results of the zymography assay (with κ-carrageenan as substrate). Genome analysis also indicated that AOL6 relies exclusively on type 2 secretion system (T2SS) for secreting proteins (possibly including glycoside hydrolases). In relation to T2SS, the product of the pilZ gene was predicted to be highly expressed, suggesting specialization for cell adhesion and secretion of virulence factors. The assignment of proteins to clusters of orthologous groups (COGs) revealed a pattern characteristic of r-strategists. Majority of two-component system proteins identified in the AOL6 genome were also predicted to be involved in chemotaxis and surface colonization. These genomic features suggest that AOL6 is an opportunistic pathogen, adapted to colonizing polysaccharide-rich hosts, including carrageenophytes.
碳水化合物活性酶（CAZymes）在降解或修饰多糖/聚糖方面具有重要的生物技术潜力。由于海洋大型藻类富含各种类型的多糖，因此海藻相关细菌可能是这些 CAZymes 的良好来源。基因组学方法可用于探索 CAZyme 的丰度和多样性，但也可以深入了解 CAZyme 产生者的生物学特性，特别是介导它们与宿主相互作用的分子机制。在这项研究中，从一种卡拉胶生产养殖红藻（Gigartinales，Rhodophyta）卡拉胶病叶中分离到一株革兰氏阴性、需氧、杆状、卡拉胶降解和可培养的海洋细菌，命名为 AOL6。对该细菌的全基因组进行了测序和表征。序列读取组装产生了高质量的基因组组装。该细菌的估计基因组大小为 4.4 Mb，G+C 含量为 52%。基于 16S rRNA、rpoB 和一组 38 个单拷贝基因的完整序列的分子系统发育分析表明，该细菌是一种未知物种，代表细胞动弯杆菌科中的一个新属，与 Teredinibacter 和 Saccharophagus 属最为密切相关。与 T. turnerae T7901 和 S. degradans 2-40 的基因组比较揭示了这三个物种的几个共同特征，包括大量 CAZymes，占编码基因总数的>5%。在 AOL6 基因组中发现的 CAZymes 比例高于其他已知的碳水化合物降解菌，表明其具有降解包括 κ-卡拉胶在内的多种多糖的显著能力；其中 34%的 CAZymes 具有信号肽序列用于分泌。通过计算机分析从细菌基因组中鉴定了三个推定的 κ-卡拉胶酶编码基因，与胶凝试验结果（以 κ-卡拉胶为底物）一致。基因组分析还表明，AOL6 仅依赖于 II 型分泌系统（T2SS）来分泌蛋白（可能包括糖苷水解酶）。与 T2SS 相关，预测 pilZ 基因的产物高度表达，表明其对细胞粘附和毒力因子分泌的特异性。将蛋白质分配到直系同源群（COG）簇中揭示了与 r-策略者相关的模式。在 AOL6 基因组中鉴定的大多数双组分系统蛋白也被预测参与趋化作用和表面定植。这些基因组特征表明，AOL6 是一种机会性病原体，适应于定殖富含多糖的宿主，包括卡拉胶植物。
