Molecular Characterization of Lactic Acid Organisms Isolated from Spontaneous Fermentation of Cassava- fufu and gari

Abstract

Aim: The present work was aimed at ensuring the accurate taxonomic identification of the fermentation organisms previously isolated from spontaneously fermented cassava for the purpose of safety assessment and quality assurance. Study Design: Purposive sampling method was used as all the five isolated organisms (from the previous study); 2 bacterial and 3 fungal organisms were characterized using molecular methods. Place and Duration of Study:  The genotypically characterized organisms are from a previous study carried out in May 2016 where lactic acid bacterial and fungal organisms were isolated from spontaneously fermented cassava.  Methodology: Genomic DNA was extracted from the lactic acid microorganisms and this was used as a template in a polymerase chain reaction (PCR) reaction. The 16s rRNA genes were amplified for the bacterial isolates while nuclear ribosomal internal transcribed spacer (ITS) genes were targeted for the fungal isolates. The amplicons were electrophoresed on 2% agarose gel prepared with Tris Borate Ethylenediamintetraacetate (TBE) buffers stained with ethidium bromide. The sizes of the corresponding amplicon captured on gel images were compared with that of the ladder to determine their sizes. Furthermore, sequences of the PCR products were analyzed and the chromatograms subjected to BLAST (Basic Local Alignment Search Tool) analyses to identify the lactic acid organisms. Results: The 3 fungal isolates were identified as Schizophyllum commune (MK 431022), Aspergillus oryzae (MK434151), Aspergillus sydowii (MK434152), while the bacterial isolates were identified as Bacillus spp (MK450345) and Bacillus spp (MK 449018). A high level of discrepancies was observed when the results of identification of the same isolates by biochemical methods in an earlier report were compared with the results from the present study using molecular methods.  Conclusion: The identification and characterization of the isolates and the discrepancies between the outcomes of the biochemical and molecular methods underscore the limitations of phenotypic (biochemical) methods in characterizing organisms that may eventually be used as starter organisms in food fermentation. The present result makes quality assurance and safety assessment of foods prepared using these organisms as starter organisms possible.