浒苔暴发前后青岛沿岸水体和藻体的细菌群落结构与生态环境因子相关性分析

Correlation analysis of bacterial community structure and eco-environmental factors in water and algae along Qingdao Coast before and after the outbreak of Ulva prolifera

关键词:  浒苔  绿潮  生态环境因子  可培养细菌  弧菌  毒力基因  耐药基因

Key words:  U. prolifera  Green tide  Ecological environmental factors  Culturable bacteria  Vibrio  Virulence genes  Resistance genes

摘要:

自2008年起，浒苔(Ulva prolifera)暴发给海洋环境生态、水产养殖业、旅游业和公共卫生带来严重影响。为进一步探究浒苔暴发对近岸环境中细菌群落结构与生态因子的影响，本研究对浒苔暴发前后青岛沿岸主要滨海区域水体、浒苔藻体中可培养细菌群落的组成与丰度及环境理化指标进行了比较分析。结果显示，浒苔暴发后，青岛沿岸水体的化学需氧量(COD)、氨氮、无机氮、砷离子和活性磷酸盐含量均显著升高(P<0.05)，溶解氧、Cd、Pb、Cr、Cu等金属离子的含量显著降低(P<0.05)；聚集地浒苔藻体中有机碳含量低于非聚集地(P<0.05)，大肠菌群、粪大肠菌群、硫化物、Hg、Cd、Zn和Cu离子含量高于非聚集地(P<0.05)。可培养细菌分析结果显示，浒苔暴发后，青岛市沿岸水体与藻体中可培养细菌的数量增长超1个数量级，弧菌(Vibrio)占比增加了38%；基于16S rDNA的细菌鉴定结果显示，浒苔暴发前，青岛市沿岸海水中的主要优势菌为长巨牡蛎弧菌(Vibrio gigantis)、塔斯玛尼亚弧菌(Vibrio tasmaniensis)和大西洋弧菌(Vibrio atlanticus)；浒苔暴发后，青岛市沿岸海水中的主要优势菌为强壮弧菌(Vibrio fortis)、溶藻弧菌(Vibrio alginolyticus)和欧文斯氏弧菌(Vibrio owensii)。优势弧菌的毒力基因与耐药基因携带风险检测结果表明，浒苔暴发后，弧菌携带毒力基因(VAC、vhh、chxA、tlh和flaA)和耐药基因(strA、strB、QnrVC和sul2)的类别和检出率均增高。研究结果初步揭示了浒苔暴发对青岛市沿岸环境中细菌群落结构及环境生态的影响，查明了浒苔暴发产生的有害微生物种类及其携带的毒力基因、耐药基因风险，为黄渤海近岸海域浒苔绿潮防控、海洋生态保护、水产健康养殖和公共卫生治理提供了理论依据。

Abstract:

Green tide is a harmful ecological phenomenon caused by the explosive proliferation or high aggregation of certain large green algae (such as Ulva prolifera) in seawater under specific environmental conditions, leading to discoloration of the water. The green tide formed by a large number of floating algae can not be effectively disposed of in a short period of time, which will lead to the accumulation and decay of green algae, thereby affecting the coastal ecological landscape. The outbreak of U. prolifera has been one of the main causes of green tide occurrences in the Yellow Sea since 2008. U. prolifera is an algal plant belonging to the family Ulvaceae, which has the characteristics of high nutrient absorption efficiency, fast growth rate and strong reproductive ability, and is more competitive than other species. The explosive proliferation of a large number of U. prolifera during a short period of time has a serious impact on marine environmental ecology, aquaculture, tourism and public health and safety. The study mainly investigated the environmental physicochemical parameters, cultivable microorganisms, dominant culture bacteria, and virulence and resistance genes of dominant bacteria in water and algae samples collected before and after the outbreak of U. prolifera in the main coastal areas of Qingdao were investigated. Environmental physicochemical factors, including the coliform and fecal coliform contents, chemical oxygen demand (COD), biochemical oxygen demand (BOD5), ammonia nitrogen (NH3-N) levels, nitrite, active phosphate, inorganic nitrogen, cyanide, sulfide, arsenic, mercury, copper, zinc, selenium, total chromium, nickel, cadmium, and lead ion levels, in the seawater were assessed. At the same time, the levels of organic carbon, sulfides, coliforms, fecal coliforms, and mercury, arsenic, cadmium, lead, zinc, copper, and chromium ions in the algae were measured. Research on cultivable microorganisms included an assessment of the abundance and composition of cultivable bacteria and Vibrio species in seawater and algae. The distribution of common virulence genes (VAC, vhh, chxA, tlh, and flaA) and resistance genes (strA, strB, QnrVC, and sul2) in the dominant Vibrio bacteria was examined to evaluate the potential virulence risk of the dominant Vibrio bacteria. Comparative analysis results showed that after the outbreak of U. prolifera, the COD, NH3-N, inorganic nitrogen, arsenic ion, and active phosphate levels in the coastal waters of Qingdao increased significantly (P<0.05), and the levels of dissolved oxygen and cadmium, lead, chromium, copper, and other metal ions decreased significantly (P<0.05). The organic carbon content in the algal samples from the aggregated areas of U. prolifera was lower than that from non-aggregated areas. In contrast, the content of coliforms, fecal coliforms, sulfide, mercury, cadmium, zinc, and copper ions significantly increased compared to that in open areas (P<0.05). Moreover, the proportion of Vibrio bacteria increased from 7% to 45%, representing an increase of 38%. The bacterial identification results based on 16S rDNA showed that before the outbreak of U. prolifera, there were 79 dominant bacterial and 62 Vibrio strains in the coastal waters and algae bodies of Qingdao, belonging to 22 species. The main Vibrio species were V. gigantis, V. tasmaniensis, and V. atlanticus. After the outbreak of U. prolifera, 79 dominant bacterial and 65 Vibrio strains were isolated, belonging to 24 species, with the main Vibrio species being V. fortis, V. alginolyticus, and V. owensii. The distribution of virulence genes (VAC, vhh, chxA, tlh, and flaA) and resistance genes (strA, strB, QnrVC, and sul2) in dominant Vibrio bacteria isolated from water bodies and algae before and after the outbreak of U. prolifera was tested. The results showed that only two virulence genes, tlh and flaA, were detected before the outbreak of U. prolifera, and no resistance genes were detected. Five virulence genes and four resistance genes were identified after the outbreak of U. prolifera. Correlation analysis between the number of cultivable bacteria and the environmental physicochemical factors in the water and algae of Qingdao coastal waters before and after the outbreak of U. prolifera showed that the abundance of cultivable bacteria and Vibrio species in the water and algae was correlated with environmental factors such as temperature, pH, NH3-N, nitrite, inorganic nitrogen, active phosphate, total chromium, and nickel. The results revealed the impact of the outbreak of U. prolifera on the bacterial community structure and environmental ecology in the coastal environment of Qingdao. At the same time, the harmful microbial species caused by the outbreak of U. prolifera and the risk of virulence genes and drug resistance genes carried by the outbreak of U. prolifera were evaluated. The relevant research results provide a theoretical basis for the prevention and control of green tide, marine ecological protection, healthy aquaculture in the coastal waters of the Yellow Sea and Bohai Sea.