网箱养殖青石斑鱼的溃疡病病原

The pathogeny of ulcer disease in Epinephlus awoara

2002年夏季厦门同安湾一带气候炎热,降雨量少,海水盐度偏高,同安湾刘五店的网箱养殖石斑鱼大面积暴发溃疡病。本研究调查了石斑鱼溃疡病暴发的特征和症状,主要表现为肌肉溃疡坏死、眼球脱落、鱼骨暴露等。从病鱼体表及内脏分离出优势菌群命名为TS 628,经回归感染证实TS 628就是引发本次石斑鱼溃疡病的病原菌。对病原菌进行鉴定发现,该菌革兰氏染色呈阴性,电镜下观察菌体呈短杆状,极端单鞭毛,综合研究该菌在形态、生理生化、16SrDNA同源性及药物敏感性等方面的特性,基本确认分离到的病原菌为哈维氏弧菌(Vibrioharveyi),该菌对氯霉素、壮观霉素等多种抗生素敏感,对万古霉素、青霉素G等抗生素不敏感。哈维氏弧菌是水产养殖病害常见病原菌,但作为养殖石斑鱼的病原菌在国内属首次报道。

The summer of 2002 was hot and dry in Xiamen, and the seawater salinity was higher than usual, sometimes reaching 38 in Tongan Bay. From the end of May, ulcer disease occured on a large scale in Tongan Bay. In the middle of June, a heavy rain brought about high mortality among the cultured groupers, which caused serious loss. The characteristics of ulcer disease were observed. Infected groupers displayed sluggish swimming and refused feeding, several days later, the groupers' eyes swelled and became ulcerated, the tail turned red, the scales became detached and the back muscle gradually ulcerated, then the eyes even dropped out and the muscle became so necrotic that the spine was exposed, finally the diseased fishes died. Anatomized the dead fish and found that the livers and gills were pale and the gallbladder was distended. The time course from appearance of disease signs to death lasted about a week. Dominant bacteria, which were Gram-negative and seen short rod with single polar flagellum under electron microscope, were isolated and designated TS-628. In artifical infection test all fish of the experimental group died on the third day after injected with bacterial suspension, while all the fish in control group showed no signs till the end of another week observation. The dead grouper had pale livers and gills and small ulcerations on the caudal fin and anal fins. These were the same signs as the natural infected fishes. The re-isolate also had the same characteristics as TS-628, which proved the isolate TS-628 was the pathogenic bacteria that triggered this ulcer disease. Different methods were used to identify the pathogenic bacteria in this study. The identification result through VITEK-AMS system GNI card was that TS-628 was Burkholderia mallei, with 99% reliability. While traditional biochemical identification revealed that TS-628 exhibited relVibrio harveyi through comparison with Bergey's Manual Determinative Bacteriology. In order to confirm absolutely different results above, further researches were carried out to identify TS-628 again. So 471bp sequence of TS-62816S rRNA gene was amplify and compared with all DNA sequences (1192858 in total) in genebank (GenBank+EMBL+DDBJ+PDB), homology analyses showed that 16 sequences were picked out to have the highest similarity, with 98% identity. These 16 sequences all originated fromV. harveyV. carchiariae which also belongs to harveyi because their similarity in physiological and biological characteristics and DNA homology. According to morphological features, physiological and biochemical characteristics and 16S rRNA gene homology comparison of the bacteria, the patV. harveyi. Drug sensitivity test showed that the pathogenic bacteria were highly sensitive to chloramphenicol, actinospectcinV. harveyi is a kind of pathogenic bacterium commonly found iV. harveyi is reported as the pathogenic bacteria of cage-cultured grouper in China. And V. harveyi should be regarded as an opportunistic pathogen which has close relation to temperature and salinity and easily causes vibriosis under conditions of high temperatures and drought. Therefore, it is necessary to guard agV. harveyi vibriosis in such summer days.