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This is a Quantitative Study study (also classified as: Experimental Study).

September 30, 2025Open Access

Characterization and 16S rRNA-Based Identification of Antimicrobial Peptide-Producing Lactic Acid Bacteria from Traditional Nigerian Non-Dairy Fermentations

OPOluwabukola Atinuke PopoolaNational Science and Technology Development Agency OAOlubunmi Olaitan AgarryUniversity of Abuja AOA. A. OrukotanLagos State University

Key Points

Significant antimicrobial activity against pathogens was observed, with 60.4% of isolates inhibiting at least one target.
Thirteen isolates were confirmed to produce antimicrobial peptides, with inhibition zones ranging from 10.33 to 15.00 mm.
Isolates were identified via 16S rRNA sequencing, revealing three distinct LAB species, some with potential novelty.
Traditional Nigerian non-dairy fermented foods present valuable LAB sources for natural food biopreservatives in combating antimicrobial resistance.

Abstract

Abstract Background Lactic acid bacteria (LAB) produce antimicrobial peptides (AMPs) with activity against foodborne and clinically relevant pathogens, offering clean-label alternatives to chemical preservatives and potential tools to combat antimicrobial resistance. These bioactives have applications in food safety, medicine, and agriculture, contributing to global sustainability goals. Traditional Nigerian non-dairy fermented foods remain an underexplored source of such bacteria. This study aimed to isolate, identify, and evaluate AMP-producing LAB from soy milk, tigernut milk, and sorghum gruel for their antimicrobial activity, and to assess their potential for novel strain discovery. Methods Spontaneously fermented soy milk, tiger nut milk, and sorghum gruel were prepared under laboratory conditions, yielding 174 LAB isolates. These were characterized by Gram reaction, catalase, spore formation, motility, and carbohydrate fermentation tests. Antimicrobial activity was initially assessed using agar spot tests against Staphylococcus aureus ATCC 25923 and Escherichia coli ATCC 25922. Active isolates underwent agar well diffusion assays against S. aureus, E. coli, Pseudomonas aeruginosa ATCC 27853, and Salmonella Typhi ATCC 6539. Neutralization with catalase and pH adjustment distinguished peptide-mediated inhibition. Six representative AMP-producing isolates were identified by 16S rRNA gene sequencing and phylogenetic analysis, with sequences deposited in GenBank (PV983358–PV983363). Results Of the 96 presumptive LAB screened, 58 (60.4%) inhibited at least one target pathogen. Agar well diffusion assays confirmed AMP production in 13 isolates, with inhibition zones of 10.33–15.00 mm. Peptide-mediated inhibition was retained in the 13 isolates following catalase and pH neutralization. Sequencing identified isolates as Lactiplantibacillus plantarum, Lactobacillus fermentum, and Limosilactobacillus fermentum. One strain, L. fermentum Atinukensis-TNA14, shared only 93.54% sequence identity with its closest type strain, indicating potential novelty and warranting whole genome sequencing. Conclusion Traditional Nigerian non-dairy fermented foods harbour diverse LAB with significant AMP-producing capacity, including a putative novel species, underscoring their potential as sources of natural food biopreservatives. Future studies will focus on whole genome sequencing and functional characterisation of the novel strain, as well as evaluating the stability and efficacy of its AMPs in model food systems.

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