Whole-genome sequence analysis for evaluating the safety and probiotic potential of Lactiplantibacillus pentosus 9D3, a gamma-aminobutyric acid (GABA)-producing strain isolated from Thai pickled weed

Abstract

<i>Lactiplantibacillus pentosus</i> 9D3, a prominent gamma-aminobutyric acid (GABA)-producing bacteria isolated from Thai pickled weed was characterized for its safety and probiotic properties <i>via</i> whole-genome analysis and <i>in vitro</i> testing. The whole-genome sequence of <i>L. pentosus</i> 9D3 was determined using a hybrid-sequencing approach, combining PacBio and Illumina technologies. A 3.81-Mbp genome of <i>L. pentosus</i> 9D3 consisting of one 3.65-Mbp chromosome and six plasmids (1.9-71.9 Kbp) was identified with an estimated GC content of 46.09% and 3,456 predicted genes. The strain was confirmed to be <i>Lactiplantibacillus pentosus</i> according to the high average nucleotide identity value of >95% and digital DNA-DNA hybridization scores of >70% to the <i>L. pentosus</i> type strain. Comparative genome analysis with other <i>L. pentosus</i> strains showed that the GABA-producing capability was specific to the strain 9D3. Genes related to GABA biosynthesis and transport were identified on a plasmid, pLPE-70K, indicating the acquired nature of this property. The safety of <i>L. pentosus</i> 9D3 was demonstrated through the lack of genes related to the production of toxins, biogenic amines, and antimicrobial drugs. Although the strain exhibited resistance to ampicillin and chloramphenicol, none of the antimicrobial resistance (AMR) genes were associated with mobile elements, i.e., plasmids and prophages. Therefore, the strain is considered to have low risk of transferring the AMR genes to other, potentially pathogenic bacteria. In addition, <i>L. pentosus</i> 9D3 showed good survivability in the gastrointestinal tract environment and was able to adhere to the intestinal cell <i>in vitro.</i> Therefore, <i>L. pentosus</i> 9D3 is concluded to be safe, with the potential to be used as a probiotic, exerting its health benefit through GABA production in the food system. The GABA-producing capability of the strain <i>in vivo</i> is the subject of further investigation.