Characterization of lactic acid bacteria from fermented fish product (Bekasam)

Abstract

Bekasam is an Indonesian fermented fish product that has sour taste and mostly contain lactic acid bacteria (LAB). This study aimed to obtain and characterize LAB isolates from bekasam and to study their potency in inhibiting the growth of pathogenic bacteria, i.e. Escherichia coli, Salmonella typhimurium ATCC 14028, Bacillus cereus, Staphylococcus aureus, and Listeria monocytogenes. LAB were isolated from bekasam using MRSA media supplemented with CaCO3 0.5%. Incubation was done at 37°C for 48 hours. The pure cultures were verified as LAB based on morphological and biochemical characteristics. LAB were obtained, then they were selected for their antimicrobial activity and further the determination of their antimicrobial compounds. Identification for the selected isolates was based on 16S rDNA sequences, followed by production of organic acids. From eight bekasam samples, total of LAB was 1.4 x108-9.0x108 CFU/g. Seventy four isolates were successfully isolated. It was found that 62 isolates (84%) belonged to LAB. Twenty three isolates could inhibit the growth of the five pathogenic bacteria in vitro. The highest inhibition zone was on S. aureus. However, neutralized supernatant of the LAB culture did not inhibit the growth of the pathogenic bacteria. While, cell free supernatant at pH 5 and 6 from 11 isolates did inhibit the growth of the pathogenic bacteria. BI(3), BP(3), BP(20) and SK(5) isolates growed in MRSB medium in vitro, They produced H2O2 concentrations are much smaller than the production of organic acids. The highest of antimicrobial activity was SK(5) isolates. Pellet of protein precipated from fourth isolates showed inhibitory zone against pathogenic bacteria, this inhibition is thought to have come from the bacteriocin, but will need more detailed testing. BI (3), BP (3) and BP (20) isolates showed antimicrobial activity of precipitated protein against E. coli, L. monocytogenes, and S. typhimurium, respectively, with concentration of ammonium sulfate at 40%, 10% and 70-80%, respectively. While SK (5) isolates showed antimicrobial activity only against S. typhimurium with concentration of ammonium sulfat at 40%, 60% and 70%. Molecular identification based on 16S rDNA sequence revealed that BI(3), BP(3), and BP(20) isolates were Pediococcus pentosaceusi IE 3 with similarity of 98%, 97%, and 98%, respectively. While SK(5) isolates showed 93% similarity to Lactobacillus plantarum subsp. plantarum NC 8. Productivity of organic acids from BI(3) isolates was the best than the other three isolates. The dominant organic acid content of BP (3) and SK (5) isolates were lactic acid while the BI (3) and BP (20) isolates were acetic acid. Thus BI (3), BP (3), BP (20) and SK (5) showed antimicrobial activity which could be useful in food preservation.