Phenotypic and molecular identification and antibiotic susceptibility of endophytic bacteria associated with in vitro plantain (Musa spp.) plantlets

Abstract

The proliferation of endophytic bacteria represents a critical challenge for the viability of Musa spp. micropropagation, compromising the health and development of explants during their in vitro establishment. This study aimed to molecularly identify endophytic bacteria associated with plantain vitroplants and to evaluate the efficacy of five antibiotics for their control. The research was conducted between April 2022 and September 2023 at the National Center for Tissue Culture of INTA-CNIA and the Agricultural Microbiology Laboratory of the National Agrarian University, Managua, Nicaragua. A total of 690 symptomatic explants of the Cuerno Enano, Cuerno Gigante, and CEMSA ¾ varieties, obtained from seven subculture in multiplication micropropagation stages, were analyzed. Bacterial isolates were obtained from internal tissues of leaves and stems through surface disinfection and incubation in nutrient agar; subsequently, morphological and biochemical characterization was performed, followed by amplification and sequencing of the 16S rRNA gene in three bacterial isolates to establish their taxonomic identity. The susceptibility of nine bacterial isolates to five broad-spectrum antibiotics was determined using the disk diffusion method under a completely randomized experimental design. Of the 690 explants evaluated, 485 (70.29%) showed symptoms of endophytic contamination, with variable incidence among subcultures (52%–100%). From 24 bacterial isolates, the genera Pseudomonas (10 isolates), Staphylococcus, Micrococcus, and Stenotrophomonas were preliminarily identified. Sequencing confirmed the presence of Stenotrophomonas maltophilia (isolates 19 and 25) and Bacillus cereus (isolate 57). In the susceptibility evaluation, isolates T58, T59, and T38 were the most sensitive, with inhibition values exceeding 80%, particularly under ampicillin and chloramphenicol, which surpassed 85% effectiveness, whereas T19, T62, and T68 exhibited marked resistance with values close to 0%; isolate T19 was the most resistant in the population. These findings underscore that the effectiveness of antimicrobial treatments intrinsically depends on the sensitivity profile of the prevalent endophytic microbiota, highlighting the need to implement sanitation protocols tailored to the local bacterial composition.