Malaysian Journal of Microbiology, Vol 9(1) 2013, pp. 13-23 http://dx.doi.org/10.21161/mjm.44012 Malaysian Journal of Microbiology Published by Malaysian Society for Microbiology (In since 2011) 13 Identification of Elizabethkingia meningoseptica from American bullfrog (Rana catesbeiana) farmed in Sabah, Malaysia using PCR method and future management of outbreak Julian Ransangan 1 *, Noramini Zainuri 1 , Tamrin Mohammad Lal 1 , Boniface Jintoni 2 and Vui Sheong Chung 2 1 Microbiology and Fish Disease Laboratory, Borneo Marine Research Institute, Universiti Malaysia Sabah, 88400, Jalan UMS, Kota Kinabalu, Sabah, Malaysia. 2 Fisheries Department Sabah, Wisma Pertanian, Jalan Tasik, 88628 Kota Kinabalu, Sabah, Malaysia. E-mail: liandra@ums.edu.my Received 28 April 2012; Received in revised form 17 July 2012; Accepted 19 July 2012 ABSTRACT 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. Keywords: American bullfrog, Elizabethkingia meningoseptica, cataract, red-leg syndrome, Malaysia INTRODUCTION American bullfrog, Rana catesbeiana, is not native to Malaysia but it was introduced in the country for aquaculture purposes. The farming of this anuran species has gained popularity because of its excellent attributes of adaptability to various tropical environmental conditions and relatively large size, with rich muscle mass. In Malaysia, the American bullfrog is farmed commercially to satisfy both local and international markets. Initially, there were only 12 bullfrog farms operating in Malaysia with the annual production of 80 tons of anuran meat (Kechick, 1995). After 14 years of introduction, the frog meat industry in Malaysia has undergone much development whereby the current meat production is estimated at 100 tons per month (Lee et al., 2009). However, in the recent years the production of bullfrog in some farms in Malaysia is limited by the high mortality due to diseases. The most frequently occurring diseases are the ‘red-leg’ syndrome and cataract. These diseases affect mainly the adult frogs. The cataract is characterized by opaque eye lens, lethargic behaviour and loss of appetite. The frog with red- leg syndrome seemed to have limited hopping ability with no appetite for food. These two diseases can sometimes concurrently occur in the same individual frog. The diseases caused mortality within few days to weeks after the onset. Previous study on various tissues of bullfrog with redleg syndrome showed presence of variety bacterial pathogens which include Aeromonas hydrophila, Elizabethkingia (Chryseobacterium) meningoseptica, *Corresponding author