Selection and Optimization of Lactic Acid Bacteria having Antifungal Properties

Abstract

Silage is the forage preservation product through anaerobic fermentation by using lactic acid bacteria (LAB). The presence of oxygen may activate the other microbe that could be contaminate and interrupt the silage processing. Fungi is a microbe that often contaminate silage processing. In Indonesia, prevention of silage contamination by fungi is necessary in order to support livestock business. By using the inoculum that able to produce antifungal compounds, the fungi contamination could be prevented. This research was dedicated to screen LAB having antifungal properties, taxonomic study and optimization of antifungal production. Screening of antifungal producing LAB was initiated by growing LAB on de Man Rogosa Sharpe Agar (MRSA) against Aspergillus fumigatus, A. flavus, and Penicillium by plate assay method (aerobic condition) and verified by deep tube technique (anerobic condition). The results showed that strain DR 1-6-2 could inhibit A. fumigatus and A. flavus in aerobic condition with percentage of inhibition was 41% and 32 %, respectively. In anaerobic condition, strain DR 1-6-2 could inhibit not only A. fumigatus, A. flavus, but also Penicillium. Identification of selected LAB was carried out using 16S rRNA gene sequencing with 27F and 1492R primer. Based on molecular analysis, strain DR 1-6-2 is belong to Lactobacillus plantarum with the degree of homology of 98%. Optimization of antifungal production by selected LAB was carried out using Response Surface Method (RSM) with Central Composite Design (CCD) on five variables (glucose, yeast extract, CH3COONa, K2HPO4, Tween 80) and five level combinations. From statistical analysis, the production of antifungal which is represented by the total lactic acid production of selected LAB is significantly influenced by glucose, combination of glucose and K2HPO4, combination of yeast extract and K2HPO4 and Tween 80. The optimum concentration of glucose is 2.91%, yeast extract 0.58%, K2HPO4 0.29% and Tween 80 0.19% with a maximum predicted lactic acid yield of 2.61%, which is increased by 59%. This result is confirmed when the optimum concentration of nutritions used. The production of lactic acid increased by 58%. Benefit of this research is utilization of indegenous LAB isolates as the silage inoculum and antifungi to prevention of contamination and to obtain a good silage quality. This LAB utilization is capable to trigger the silage production, which in turn could increase the silage utilization as animal feed, increasing the production of meat, and could support the government's efforts to achieve the national demand of meat product. In silage fermentation using L. plantarum DR 1-6-2, anaerobic condition is a requirement to be considered. Identification of other antifungal compounds produced by L. plantarum DR1-6-2 is need to be studied.