BIODIVERSITAS ISSN: 1412-033X Volume 20, Number 4, April 2019 E-ISSN: 2085-4722 Pages: 1063-1068 DOI: 10.13057/biodiv/d200418 Isolation and molecular identification of antimicrobial-producing Lactic Acid Bacteria from chao, South Sulawesi (Indonesia) fermented fish product RISKY NURHIKMAYANI 1 , BUDI SETIADI DARYONO 2 , ENDAH RETNANINGRUM 1, 1 Microbiology Laboratory, Faculty of Biology, Universitas Gadjah Mada. Jl. Teknika Selatan Sekip Utara, Sleman 55281, Yogyakarta, Indonesia. Tel.: +62-274-580839, Fax.: +62-274-6492355,  email: endahr@ugm.ac.id 2 Genetic and Breeding Laboratory, Faculty of Biology, Universitas Gadjah Mada. Jl. Teknika Selatan Sekip Utara, Sleman 55281, Yogyakarta, Indonesia Manuscript received: 23 May 2018. Revision accepted: 23 March 2019. Abstract. Nurhikmayani R, Daryono BS, Retnaningrum E. 2019. Isolation and molecular identification of antimicrobial-producing Lactic Acid Bacteria from chao, South Sulawesi (Indonesia) fermented fish product. Biodiversitas 20: 1063-1068. Chao is traditional fermented fish made of anchovy (Stolephorus sp.), rice, salt, and fermented naturally on controlled environment. The product comes from Pangkep Regency, South Sulawesi. Chao has the potential for the source of high-grade lactic acid bacteria isolates, especially for antimicrobial compound. Antimicrobial compound produced by lactic acid bacteria is a biopreservative compound with many advantages. The purposes of this investigation were to isolate antimicrobial-producing lactic acid bacteria from chao and to identify their isolates based on the molecular properties. Lactic acid bacteria were isolated from chao using Man Ragosa Sharpe Agar (MRSA) medium added with 0.1 % CaCO3, followed by the ability test to produce antimicrobial compound against Staphylococcus aureus FNCC 0047 as an indicator. The spectrum of crude antimicrobial as cell-free supernatant (CFS) was determined by antagonism against S. aureus FNCC 0047 and Escherichia coli FNCC 0049 on agar well diffusion test. Selected isolates of antimicrobial-producing lactic acid bacteria molecularly identified from 16S rRNA marker using primer 27F and 1492R. We obtained four isolates of lactic acid bacteria that showed the antimicrobial activity. This crude antimicrobial compound could inhibit both gram-positive Staphylococcus aureus FNCC0047 and gram-negative bacteria Escherichia coli FNCC0049. The highest crude antimicrobial activity of isolates was observed on IB1C strain. The results of molecular characterization suggested that the IB1C and IB3B strain were identified as Lactobacillus plantarum, whereas IB3E and IB3F were identified as Pediococcus pentosaceus. Keywords: Antimicrobial, chao, lactic acid bacteria, Lactobacillus plantarum, Pediococcus pentosaceus INTRODUCTION Chao is a local fermented fish product of coastal communities from Pangkep Regency, South Sulawesi, Indonesia. It is produced by soaking the raw materials, including anchovy (Stolephorus sp.), rice and salt, then fermented naturally on controlled environment (Karim et al. 2013). The salt content of chao fermentation ranges from 15-20% that inhibit the growth of the non-tolerant microbes. Lactic acid bacteria are tolerant and adaptable to saline environment with concentrations of 10-18% (Sanches 2008; Agussalim 2017). This makes lactic acid bacteria becomes the dominant microorganism in chao fermentation products. Chao is also known to have a sour taste that indicates the main microorganism for fermentation activity is the lactic acid bacteria (LAB) group (Lakong 2003). The LAB was divided into 12 genera, i.e. Lactococcus, Leuconostoc, Pediococcus, Streptococcus, Lactobacillus, Enterococcus, Aerococcus, Vagococcus, Tetragenococcus, Carnobacterium, Weissella, and Oenococcus. These bacterial species in the LAB group are designated as LAB according to their ability to metabolize lactic acid (Ray 2005). Lactic acid bacteria are gram-positive bacteria, endospores-negatives, catalase-negative, usually non- motile and producing antimicrobial components (Axelsson 2004). Antimicrobial substances of LAB such as lactic acid, hydrogen peroxide, diacetyl, and bacteriocin can inhibit pathogenic bacteria and spoilage bacteria. The ability of LAB to produce antimicrobial components can be used as a natural preservative (Adams and Moss 2008). The LAB are known as GRAS (Generally Regarded as Safe) because they are not toxic and do not produce a toxin, so they are not harmful to health, even these microorganism are useful for health (USFDA 2015). The use of LAB as a natural preservative agent can reduce the use of harmful synthetic preservatives. The ability of LAB as natural preservative has been widely reported, especially from fermented food. Indonesia has many fermented food but only few studies about Indonesian’s fermented fish are available in literature such as jambal roti, peda (Indriati et al. 2006), bekasam (Desniar et al. 2015; Budiarto et al. 2016), and rusip (Koesoemawardani et al. 2018). As far as we know, there has been no research on antimicrobial- producing lactic acid bacteria from chao. This research aimed to isolate and identify antimicrobial-producing LAB from chao, South Sulawesi Fermented Fish Product.