Aims: Vibrio harveyi causes vibriosis to Asian seabass (Lates calcarifer). The disease spreads rapidly among fish stocked in the same cage. It causes high mortality especially in weak and small sized fish stocked at high density in poorly managed net cage. Study to determine the virulence levels of the bacterial pathogen in various aquaculture animals is a key to prevent vibriosis in marine aquaculture. Methodology and Result: Isolation of bacteria from diseased Asian seabass was done using tryptic soy agar (TSA) and thiosulphate citrate bile sucrose agar (TCBS) plates. Virulence of two strains of Vibrio harveyi (VHJR4 and VHJR7) was tested against clinically healthy aquaculture animals. The analysis revealed that the two bacterial strains differ in pathogenicity. The V. harveyi strain VHJR7 was virulent to Asian seabass at 1.40 x 104 c.f.u. g-1, humpback grouper (Cromileptis altivelis) at LD50 8.33 x 103 c.f.u. g-1 and black tiger shrimp (Penaeus monodon) at LD50 3.26 x 104 c.f.u. g-1 , respectively. The V. harveyi strain VHJR4 was not virulent to Asian seabass and humpback grouper but it caused mortality to black tiger shrimp at LD50 1.32 x 106 c.f.u. g-1. Phenotypically, the two strains shared most of the biochemical features except that the V. harveyi strain VHJR7 was a urease positive and grew at 8.5 % NaCl, and at 10 °C. The percentage similarity of nucleotide sequences of 16S rDNA in V. harveyi VHJR4 and V. harveyi VHJR7 was higher (99%) but reduced at 95 % in hemolysin gene. Conclusion, significance and impact of study: Pathogenic strain of V. harveyi causes mortality and affects aquaculture production of Asian seabass. Hence, vaccine development against the bacterial pathogen is urgently needed for sustainability of Asian seabass aquaculture in Malaysia.

Aims: High demand for frog meat in Malaysia especially the American bullfrog (Rana catesbeiana) has promoted intensive farming of the animal. However, the farming of American bullfrog is restricted by the occurrence of diseases. This study reports the first isolation of Elizabethkingia meningoseptica from specimens of American bullfrog that suffer from cataract and ‘red-leg’ syndrome. Methodology and Result: The pathogen was isolated from eyes and internal organs (liver, kidney and spleen) of the diseased bullfrog specimens. All the bacterial isolates were subjected to phenotypic characterization and antibiotic susceptibility assay, and further identified by using the 16S rDNA sequencing analysis. We designed two pair of specific PCR primers (22-25 mers) which are complimentary to the β-lactamase gene in the reference strain of E. meningoseptica ATCC49470. The result showed all the bacterial isolates shared similar phenotypic characters and antibiotic susceptibility. BLAST analysis of the 16S rDNA sequences indicated that the bacterial isolates had very high sequence homology (100%) with E. meningospetica ATCC49470 and E. meningoseptica isolates from mosquito. The two PCR primers were very specific to E. meningoseptica isolates of this study. Conclusion, significance and impact of study: This is the first isolation and characterization of bacterial pathogen, E. meningoseptica in cultured American bullfrog (Rana catesbeina) that suffered from eye cataract and ‘red-leg’ syndrome in Sabah, Malaysia. It is suspected that one of the possible transmission routes of the bacterial pathogen could be via mosquito bites. The findings suggest that there is urgent requirement for standard guideline of good farming practice to be adopted in frog farms throughout the country. Such a guideline can help in minimizing economic losses, preventing transmission of the zoonotic bacterial pathogen to farm workers, and sustaining the industry in Malaysia and upgrading frog meat quality for international market.

Aims: Vibrio harveyi is a serious pathogen for marine organisms particularly in hatcheries and grow-out ponds that attack their immune system. The rapid detection of V. harveyi is urgently needed to prevent bacterial spread. Here we described a rapid and specific visual detection method based on the Loop-Mediated Isothermal Amplification in combination with Lateral Flow Dipstick (LAMP-LFD). Methodology and results: A set of six novel primers were designed to target the toxR gene. These include the biotin-labelled inner primer that complements specifically to the target sequences. The resulting biotinylated LAMP amplicons were hybridised to the FAM-labelled probe resulting in lateral flow detection on the dipstick. The addition of loop primers improved the reaction time of LAMP by more than half and rapid detection was observed within 10-15 min. In comparison, the sensitivity of PCR-UV analysis was only at 104 copies while LAMP-LFD was able to detect lower amounts at 103 copies. The LFD provided higher specificity and selectivity since hybridization with specific probes to the LAMP amplicons was employed. In addition, detection of V. harveyi infected grouper was successful using the LAMP-LFD method described here. Conclusion, significance and impact of study LAMP-LFD is specific to V. harveyi. Our method provides a useful tool to rapidly detect and monitor the outbreaks of the pathogen.