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| 海带高、低畸形率幼孢子体的附生菌群分析 |
| Analysis of Epiphytic Bacterial Communities on Saccharina japonica Sporelings with Low and High Malformation Disease Severity |
| 投稿时间：2025-04-09 修订日期：2025-05-07 |
| DOI： |
| 中文关键词: 海带 幼孢子畸形 附生菌群 功能途径 |
| 英文关键词: Saccharina japonica Sporeling malformation disease Epiphytic bacterial community Functional pathway |
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| 中文摘要: |
| 幼孢子体畸形病(Sporeling malformation disease)是制约海带育苗产业发展的重要病害之一,其发生机制尚未明确。本研究以威海某育苗基地不同畸形率(低发病率组Low vs.高发病率组High)海带幼孢子体为对象,利用高通量测序分析了其附生菌群的多样性、结构及功能途径,揭示了附生菌群结构与功能变化与病害发生的关联。结果显示,除Shannon指数外,High组菌群Alpha多样性指数均显著降低(P<0.05),表明菌群丰富度显著降低。基于Bray-Curtis距离的主坐标分析及组间距离比较分析表明,两组幼孢子体附生菌群结构差异显著(单因素方差分析, P<0.05)。两组最主要的细菌类群包括α-变形菌纲(Alphaproteobacteria),γ-变形菌纲(Gammaproteobacteria),δ-变形菌纲(Deltaproteobacteria)和拟杆菌门(Bacteroidetes),但Low组富集数量更多的差异菌群。其中,生丝单胞菌(Hyphomonas)、无形杆菌(Amorphus)、短小杆菌属(Curtobacterium)及拟杆菌门细菌等与生长发育相关的细菌在Low组显著富集,而High组显著性富集海棒杆菌(Pelagibacterium)、食酸菌(Acidovorax)及红球菌(Rohodococcus)等潜在病原菌。功能预测结果显示,Low组显著富集嘧啶代谢、维生素和辅酶A合成、免疫防御等与生长发育和保护性相关的通路,而High组显著富集苯甲酸、类固醇和烯类降解以及多糖合成等与侵染相关的通路。本研究为阐明海带畸形病的微生物驱动机制提供了新视角,对病害防控具有重要的指导意义。 |
| 英文摘要: |
| Sporeling malformation disease, characterized by abnormal cell proliferation and tissue disintegration, causes catastrophic losses in the seedling production of kelp Saccharina japonica. Previous efforts have implicated diverse factors responsible for disease occurrence, including environmental stressors (eg, inadequate light exposure), maturity of parental kelp (eg, unmatured or overmatured), alginate-decomposing bacteria, and so on. Traditional culture-dependent approaches, focusing on sulfate-reducing bacteria and alginate-decomposing bacteria, determined potential relationships with disease occurrence, indicating the role played by the epiphytic bacterial members. However, these studies failed to explain the complex pathogenesis, and the precise microbial etiology remains elusive. Recent advances in holobiont theory suggest that the dysbiosis of epiphytic microbiota, rather than individual pathogens, could drive disease progression. Our previous study has also shown that dysbiosis of the epiphytic bacterial community correlates with the severity of sporeling malformation disease, and this disruption, in turn, might modify host–bacteria interactions to enhance disease severity. In the present study, by analyzing the diversity, structure, and functional profiles of the epiphytic bacteria on sporelings with different malformation rates, we seek to obtain more data related to the relationships between the epiphytic bacterial communities and the incidence of sporeling malformation disease using in situ sporeling samples. Through microscopic observation in 2018, two groups of biological samples (i.e., the Low and High groups) were collected from a workshop in a typical kelp seedling hatchery in Weihai, China. The malformation rate in the Low group was estimated to be approximately 2~6%, while that in the High group reached approximately 10~12%. Epiphytic bacterial DNA was extracted from the samples and the hypervariable regions V5-V7 of the 16S rRNA gene were amplified and sequenced using the Illumina NovaSeq platform. The sequences were processed using USEARCH and QIIME for quality control, chimera removal, denoise, and taxonomic assignment. Alpha and Beta diversity analyses were performed to compare the bacterial community diversity and structure between the two groups. LEfSe analysis was used to identify differentially abundant bacterial taxa, and PICRUSt2 was employed to predict the functional profiles of the epiphytic bacterial communities. The results showed that the Alpha diversity indices of the bacterial communities in the High group were significantly lower than those in the Low group, except for the Shannon index, indicating a reduced richness of bacterial communities in the High group. Principal coordinates analysis (PCoA) based on Bray-Curtis distances revealed a distinct separation of the bacterial community structures between the Low and High groups with 32.74% variance explained by PCo1, although not significant (Adonis R2=0.28, P=0.10). Whereas, inter- and intra-group comparisons of Bray-Curtis distances did reveal significant differences (one-way analysis of variance [ANOVA] with Tukey’s test, P = 0.029 < 0.05), indicating significant community structure associated with sporelings with different malformation rates. The bacterial phyla in both groups were dominated by Alphaproteobacteria (51.9%±0.1%), Gammaproteobacteria (19.8%±1.3%), Deltaproteobacteria (14.4%±3.3%), and Bacteroidetes (9.7%±1.2%), and the predominated bacterial genera included Halobacteriovorax, Thalassospira, Methylotenera, Nautella, and Marinobacter. Their relative abundances were different between the Low and High groups, which further indicated the community transition from low to high disease severity. By exploring differentially abundant taxa, it is determined that the Low group tended to be enriched in mutualists, such as Hyphomonas and Amorphus (morphogenesis inducer), Flexivirga and Curtobacterium (plant growth promoter), and bacterial members in Bacteroidetes (common colonizer and morphogenesis inducer). In comparison, putative pathogenic taxa were determined to be more abundant in the High group. For example, Pelagibacterium (FicT toxin carrier to kill the host), Acidovorax (phenolic compound degrader to facilitate invasion), and Rhodococcus (toxin carrier and phytohormone disruptor). From a functional perspective, the Low group had a higher abundance of pathways related to growth, development, and defense, such as pyrimidine metabolism, vitamin and coenzyme A synthesis, and immune defense. In comparison, functional prediction analysis indicated that the High group upregulated xenobiotic degradation (benzoate and steroids), limonene and pinene degradation, and virulence-associated polysaccharide biosynthesis (arabinoglyctan and lipoarabinomannan biosynthesis). The present study provides new insights into the microbial mechanisms underlying sporeling malformation disease. The significant differences in the structure and function of the epiphytic bacterial communities between the Low and High groups suggest that the disruption of the bacterial community may contribute to the development of the disease, which is consistent with our previous study. The findings highlight the importance of maintaining a healthy microbial community for sporeling growth and development and suggest potential targets for disease prevention and control. Future research should focus on the changes in whole community functions using different omics methods and exploring the interactions between the host, environment, and certain isolated bacterial strains in the context of disease development. This study not only enriches our understanding of the microbial ecology of kelp diseases but also has important practical implications for the kelp farming industry. By identifying key microbial taxa and functional pathways associated with the disease, the findings may guide the development of microbial-based strategies for disease management, thereby contributing to the sustainable development of kelp cultivation. |
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