Integrated Genome-Based and In Vivo Safety Evaluation of Lactiplantibacillus plantarum M2, an Isolate Obtained from the Amazonian Fruit Humiria balsamifera

Abstract

Lactiplantibacillus plantarum M2, previously isolated from the Amazonian fruit Humiria balsamifera , has shown a promising probiotic profile. This study performed an integrated safety evaluation combining genome-informed analyses with a 28-day repeated-dose oral toxicity assessment in a murine model. In silico genome screening revealed no detectable signatures associated with pathogenicity, virulence, or acquired antimicrobial resistance. Functional marker profiling and comparative analyses across reference strains further indicated that M2 harbors a conserved repertoire of genomic features associated with probiotic-relevant functions, including potential traits related to gastrointestinal survival, adhesion, oxidative stress response, and biofilm formation, with specific enrichment in substrate utilization pathways associated with niche adaptation. In vivo, repeated oral administration of L. plantarum M2 to Swiss mice did not result in mortality, adverse clinical signs, or treatment-related alterations in body weight trajectories in either sex, despite the expected variation over time (females: time effect p = 0.0018; males: time effect p < 0.0001). Mean daily food intake was higher in probiotic-treated animals in both sexes (females: mean difference = 3.54 g/day, p = 0.0016; males: mean difference = 2.84 g/day, p = 0.0008). A reduction in alanine aminotransferase levels was observed in females (49.40 ± 6.18U/L in controls vs. 42.33 ± 3.67 U/L in treated animals, p = 0.0428), without associated histopathological alterations in the liver. Taken together, these findings indicate that L. plantarum M2 does not elicit detectable adverse effects under the experimental conditions employed and provide a preclinical safety framework supporting its further investigation for oral applications.

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