RUI: Hypercapnic Hypoxia Impacts Shrimp Immune Defenses Against Bacterial Pathogens

与陆地生物一样，大多数海洋生物需要氧气才能生存。 近年来，世界沿海沿着沃茨中的溶解氧水平一直在下降。 海水含氧量的下降（缺氧）归因于沿海地区集约化农业和城市化带来的许多因素。 在水生环境中，缺氧通常伴随着水质其他方面的变化，包括溶解的二氧化碳和酸度或pH值。实验室和实地调查支持这样一种观点，即这套水质变化，称为高碳酸血症缺氧，可以增加常驻生物体在暴露于致病微生物或病原体后感染和疾病的风险。 沿海地区是具有重要商业价值的鳍类和贝类的重要繁殖地。 对虾，包括白色，棕色，太平洋和虎虾，是一个重要的甲壳类动物群体，生活和繁殖在沿海河口。 初步研究表明，高碳酸血症性缺氧会增加太平洋对虾（Litopenaeus vannamei）在接触细菌病原体副溶血性弧菌后的感染率和死亡率。 高碳酸血症缺氧可能直接增强细菌本身的分布、存活或生长，或抑制虾宿主的免疫防御。 本研究的目的是探讨寄主L.凡纳滨对虾及其致病菌副溶血性弧菌可能对高碳酸血症性低氧敏感。 血细胞，这些细胞主要负责甲壳类动物的免疫防御，可能是敏感的溶解氧和二氧化碳的水平，或pH值。为了研究这种可能性，本研究的第一个目标是确定是否组织水平的变化，溶解的气体和酸度，伴随亚致死性高碳酸血症缺氧可以抑制孤立的血细胞杀死或抑制细菌生长的能力。 一些证据也表明，高碳酸血症缺氧的生理反应，如增加通气率，血淋巴循环和细胞-细胞聚集，可能会改变血细胞在虾的正常分布，并降低清除感染性病原体的效率。 为了解决这种可能性，本研究的第二个目的是检查是否高碳酸血症缺氧改变血细胞运动到鳃或其他主要组织隔室的正常模式后，注射活细菌。这些运动将使用分子和免疫学探针进行监测，这些探针对参与免疫防御的血细胞相关分子具有特异性。 第三组实验将研究氧气、二氧化碳和pH值的变化是否会改变虾清除细菌感染的效率。 在这些研究中，L.将给凡纳滨对虾注射一种能产生绿色荧光蛋白的转基因副溶血性弧菌。 基因改造将使研究人员能够跟踪细菌的去向，在哪里被杀死，以及最终如何从虾中消除。 这些研究的结果预计将提供基本信息，可用于保护一个关键的自然资源，Penaeid虾，同时增加我们对呼吸，离子调节和甲壳类动物抗病性功能之间的相互作用的理解。

Like their terrestrial counterparts, most marine organisms require oxygen to live. In recent years the levels of dissolved oxygen in waters along the world's coastlines has been decreasing. This decline in water oxygen (hypoxia) has been attributed to many factors stemming from intensive agriculture and urbanization in the coastal zone. In aquatic environments, hypoxia is often accompanied by changes in other aspects of water quality, including dissolved carbon dioxide and acidity or pH. Laboratory and field investigations support the idea that this suite of changes in water quality, referred to as hypercapnic hypoxia, can increase the risk of resident organisms to infection and disease following exposure to a disease-causing microbe, or pathogen. Coastal zones are critical breeding grounds for major groups of commercially important fin and shellfish. Penaeids, including the white, brown, Pacific and tiger shrimps, are such an important group of crustaceans that live and breed in coastal estuaries. Preliminary studies have shown that hypercapnic hypoxia increases the rate of infection and death in the Pacific shrimp, Litopenaeus vannamei, following exposure to the bacterial pathogen, Vibrio parahaemolyticus. Hypercapnic hypoxia may directly enhance the distribution, survival, or growth of the bacterium itself, or suppress immune defenses of the shrimp host. The objectives of the proposed studies are to examine three factors of the interaction between the host L. vannamei and its pathogen, V. parahaemolyticus, that might be sensitive to hypercapnic hypoxia. Hemocytes, those cells principally responsible for immune defense in crustaceans, may be sensitive to levels of dissolved oxygen and carbon dioxide, or to pH. To examine this possibility, the first objective of this study is determine whether tissue-level changes in dissolved gasses and acidity that accompany sublethal hypercapnic hypoxia can suppress the ability of isolated hemocytes to kill or inhibit the growth of bacteria. Several lines of evidence also suggest that physiological responses to hypercapnic hypoxia, such as increases in ventilation rate, hemolymph circulation and cell-cell aggregation, might alter the normal distribution of hemocytes in the shrimp and reduce the efficiency of clearing an infectious pathogen. To address this possibility, the second objective of this study is to examine whether hypercapnic hypoxia alters the normal pattern of hemocyte movement to the gill or to other major tissue compartments following an injection of live bacteria. These movements will be monitored using both molecular and immunological probes specific for hemocyte-associated molecules that are involved in immune defense. A third set of experiments will examine whether changes in oxygen, carbon dioxide and pH can alter the efficiency with which the shrimp clears a bacterial infection. In these studies, L. vannamei will be injected with a genetically-modified V. parahaemolyticus that produces green fluorescent protein. The genetic modification will allow investigators to track where the bacteria goes, where it is killed, and how it is ultimately eliminated from the shrimp. The results of these studies are expected to provide basic information that can be used toward protecting a key natural resource, the Penaeid shrimp, while increasing our understanding of interactions between the functions of respiration, ion regulation and disease resistance in crustaceans.