Diseases caused by bacteria, fungi, and viruses pose a great threat to aquaculture. As DNA microarrays can be used to detect multiple pathogens, here we reported an array with the potential to simultaneously detect 13 bacterial and 11 viral pathogens of aquatic animals. The array included 853 oligonucleotide probes (20-40 mer) complementary to various virus-specific sequences and four chromosomal loci (16S rRNA, gyrB, dnaJ, and recA) of bacteria. Multiplex PCR, phi29 DNA polymerase, and a Klenow fragment-based method were evaluated for amplifying and labeling the nucleic acid of pathogens. While array hybridization signals were most intense using pathogen sequences amplified by multiplex PCR, the phi29 DNA polymerase method was more convenient and ideal since it did not require sequence-specific primers that could bias against detecting novel pathogens. The feasibility of the phi29 DNA polymerase-based microarray strategy was also demonstrated by detecting multiple unknown pathogens from four samples of diseased fish and shrimps.

中文翻译：

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一种通用的随机DNA扩增和标记策略，用于微阵列检测水生动物的多种病原体。

由细菌，真菌和病毒引起的疾病对水产养殖构成了巨大威胁。由于DNA微阵列可用于检测多种病原体，因此我们在此报告了一种具有同时检测水生动物的13种细菌和11种病毒病原体潜力的阵列。该阵列包括与各种病毒特异性序列互补的853个寡核苷酸探针（20-40聚体）和细菌的四个染色体基因座（16S rRNA，gyrB，dnaJ和recA）。评价了多重PCR，phi29 DNA聚合酶和基于Klenow片段的方法，用于扩增和标记病原体的核酸。尽管使用多重PCR扩增的病原体序列产生的阵列杂交信号最为强烈，phi29 DNA聚合酶方法更方便，更理想，因为它不需要可能偏向于检测新病原体的序列特异性引物。通过从四个患病鱼类和虾样品中检测出多种未知病原体，也证明了基于phi29 DNA聚合酶的微阵列策略的可行性。