Identifying to Manage: Characterization of Bacterial Contaminants in Sugarcane Meristem Tissue Culture and Antibiotic Sensitivity of Bacillus spp.

Sugarcane is Louisiana’s most economically important agricultural crop, contributing $1.6 billion to the state’s economy in 2024. Its semi-perennial cultivation system relies heavily on vegetative propagation, increasing the risk of systemic pathogen dissemination via infected propagative material, particularly viral and bacterial agents such as sugarcane and sorghum mosaic viruses (SCMV and SrMV), Leifsonia xyli subsp. xyli and Xanthomonas albilineans. Apical meristem tissue culture (MTC) has emerged as a reliable method for generating disease-free propagative material, commonly referred to as seed cane, but potential widespread losses due to microbial contamination in the MTC process can pose a critical challenge to production. In this study, we aimed to (1) identify the main bacterial contaminants in MTC procedures, (2) evaluate their susceptibility to different antibiotics, and (3) propose improved contamination mitigation strategies. Bacterial contaminant isolates were obtained from five commercial sugarcane cultivars during the MTC sterilization process. Sixty isolates were identified by 16S rRNA sequencing, with 83% belonging to the genus Bacillus, notably B. safensis, B. aerius, and B. altitudinis, all known sugarcane endophytes. Antibiotic sensitivity assays using a modified disk diffusion method revealed significant differences in efficacy among compounds and concentrations. Cephalothin, cephalexin, cloxacillin, and timentin were the most effective antibiotics across Bacillus spp., while cefotaxime, erythromycin, and chloramphenicol exhibited minimal inhibition. This study is the first to characterize bacterial contaminants in Louisiana’s sugarcane MTC labs and provides actionable recommendations to mitigate contamination using targeted antibiotic treatments, improving the reliability of MTC for “clean” seed cane production.