环境有机碳源抑制食弧菌嗜盐噬菌弧菌(Halobacteriovorax vibrionivorans)的捕食生长

Environmental organic carbon sources inhibit the predatory life of Halobacteriovorax vibrionivorans

摘要:

【背景】蛭弧菌类群(Bdellovibrio-And-Like Organisms，BALOs)的生长所需碳源主要来源于宿主菌，而环境中各类碳源对其生长的影响还有待探究。【目的】探究不同环境有机碳源对食弧菌嗜盐噬菌弧菌(Halobacteriovorax vibrionivorans)捕食生长的影响，为其后续捕食机制和微生物菌剂的应用研究提供理论基础。【方法】以溶藻弧菌(Vibrio alginolyticus)为宿主，采用96孔板测定细胞吸光度法和双层平板法测定不同糖类化合物、酵母提取物和胰蛋白胨对食弧菌嗜盐噬菌弧菌Y22捕食生长的影响，设置热致死宿主和活宿主、人工海水和Tris-HCl (25 g/L NaCl)培养体系的对比试验，结合菌株基因组信息分析，探索其可能的生长影响机制。【结果】食弧菌嗜盐噬菌弧菌Y22不具备转运外界糖类的相关基因，不能利用环境中的糖类物质作为碳源；宿主溶藻弧菌可以利用蔗糖、麦芽糖、甘露醇生长且产酸，降低人工海水混合培养体系的pH值，从而抑制菌株Y22的捕食；葡萄糖不仅可以改变人工海水混合培养体系的pH值，而且可影响宿主的细胞特性，抑制菌株Y22的捕食识别过程；宿主无法利用淀粉、α-乳糖生长，该类碳源不影响菌株Y22捕食生长。菌株Y22具有蛋白和多肽膜转运蛋白基因，可以通过分解并摄取宿主细胞蛋白质类物质以获取碳源和氮源。外源添加质量浓度为1?5 g/L酵母提取物和胰蛋白胨都会抑制菌株Y22捕食生长，抑制效果随浓度增加而加强，酵母提取物浓度超过4 g/L、胰蛋白胨浓度超过5 g/L，菌株Y22捕食现象几乎不可见，抑制效果最强。【结论】环境中糖类可通过影响宿主菌代谢产酸或细胞特性，从而影响食弧菌嗜盐噬菌弧菌的捕食生长。食弧菌嗜盐噬菌弧菌可以吸收利用环境中的蛋白或多肽，并因此抑制其捕食生长。该研究结果将为嗜盐噬菌弧菌捕食机制的进一步探究和嗜盐噬菌弧菌微生物菌剂的开发应用提供重要的理论基础。

Abstract:

[Background] The carbon sources required for the growth of Bdellovibrio-and-like organisms are mainly derived from the host, and the effects of various carbon sources in the environment remain to be explored. [Objective] The effects of different extracellular organic carbon sources on the predatory life of Halobacteriovorax vibrionivorans was investigated to provide a theoretical basis for subsequent research on the predation mechanism and microbiological control agent against pathogenic bacteria. [Methods] Vibrio alginolyticus was used as host strain, an absorbance-based assay with 96-well plates for cell lysis and double-layer plate method were applied to determine the effects of different carbohydrates, yeast extract and tryptone on the predatory growth of H. vibrionivorans Y22. Comparative experiments between nonviable heat-deactivated and viable hosts, artificial seawater and Tris-HCl (25 g/L NaCl) culture system, were adopted to explore possible growth mechanism of strain Y22, combined with genomic analysis of strain Y22 and its host strain. [Results] Strain Y22 had no phosphotransferase system and ABC transporters of sugars, and could not use extracellular carbohydrates as carbon sources; The host V. alginolyticus could utilize sucrose, maltose, mannitol for growth and produce acid, which lead to the decreased pH value of the artificial seawater co-culture system, thereby block the predation of strain Y22. Glucose could not only decrease the pH value of artificial seawater co-culture system, but also changed the cell characteristics of the host, thereby block the predation recognition process of strain Y22. The host could not grow with starch and α-lactose, and these carbon sources did not affect the predation of strain Y22. Strain Y22 contained protein, polypeptide and amino acid membrane transporter genes, which allowed it to utilize the host protein as carbon sources and nitrogen sources. Around 1?5 g/L external yeast extract and tryptone could inhibit the predation of strain Y22, and the predation-inhibiting effect was found to be dose dependent. When the concentration of yeast extract was over 4 g/L, and the concentration of tryptone was over 5 g/L, the predation phenomenon of strain Y22 was strongly inhibited and almost invisible. [Conclusion] Environmental carbohydrates will affect the metabolic activity of host bacteria, acidify the co-culture system, and change the cell characteristics of the host, which inhibit the predatory activity of H. vibrionivorans. H. vibrionivorans can absorb and utilize the environmental protein or peptide, by which its predatory growth will be inhibited. The results will provide further insights into the predation mechanism and antibacterial application of Halobacteriovorax.