Early Detection of Endobacteria in Rhizopus spp.

Abstract

Rhizopus spp. are commonly used in bio-industrial processes such as manufacturing traditional fermented foods. However, one species, R. microsporus has been reported to contain toxin-producing endobacteria. Therefore, it is essential to evaluate the Rhizopus strains' safety before application as tempe inoculum. The study aimed to detect the presence of endobacteria from seven strains of Rhizopus spp. isolated from Tempe and four strains from other substrates using molecular and staining techniques. Molecular analyses were conducted using a PCR approach of 16S rDNA with primer sets 10F and 1541R. The presence of endobacteria was confirmed with the LIVE /DEAD® bacterial viability kit. The bacteria were identified molecularly by a phylogenetic approach using the Neighbour-Joining method and p-distance model in MEGA 6 with species of endo hyphal bacteria of Mortierella elongata as an outgroup. Molecularly, only R. microsporus IPBCC 13.1131 isolated from tempe in Cilacap, Central Java, was found to contain endobacteria. The presence of endobacteria was confirmed by the appearance of the red and green luminescence in hyphae, not in the spores. These endobacteria are identified as Curtobacterium sp. The study is the first report of described endo hyphal Curtobacterium from R. microsporus tempe starter.