Lanthanides have been considered unimportant for biological processes. In methylotrophic bacteria, however, a methanol dehydrogenase (MDH) encoded by xoxF was recently found to be lanthanide dependent, while the classic-type mxaFI is calcium dependent. XoxF-type MDHs are more widespread in diverse bacterial genera, suggesting their importance for methylotrophy. Methylobacterium species, representative methylotrophic and predominating alphaproteobacteria in the phyllosphere, contain both types and regulate their expression depending on the availability of lanthanides. RNA-seq analysis showed that the regulation takes place not only for MDH genes but also the subsequent formaldehyde oxidation pathways and respiratory chain, which might be due to the direct oxidation of methanol to formate by XoxF. In addition, a considerable number of genes of unknown function, including AT-rich genes, were found to be upregulated in the presence of lanthanides. This study provides first insights into the specific reaction of methylotrophic bacteria to the presence of lanthanides, emphasizing the biological relevance of this trace metal.

Metabolomic analysis revealed that Methylobacterium cells accumulate a large amount of ergothioneine (EGT), which is a sulfur-containing, non-proteinogenic, antioxidative amino acid derived from histidine. EGT biosynthesis and its role in methylotrophy and physiology for plant surface-symbiotic Methylobacterium species were investigated in this study. Almost all Methylobacterium type strains can synthesize EGT. We selected one of the most productive strains (M. aquaticum strain 22A isolated from a moss), and investigated the feasibility of fermentative EGT production through optimization of the culture condition. Methanol as a carbon source served as the best substrate for production. The productivity reached up to 1000 μg/100 ml culture (1200 μg/g wet weight cells, 6.3 mg/g dry weight) in 38 days. Next, we identified the genes (egtBD) responsible for EGT synthesis, and generated a deletion mutant defective in EGT production. Compared to the wild type, the mutant showed better growth on methanol and on the plant surface as well as severe susceptibility to heat treatment and irradiation of ultraviolet (UV) and sunlight. These results suggested that EGT is not involved in methylotrophy, but is involved in their phyllospheric lifestyle fitness of the genus in natural conditions.

Bradyrhizobium sp. S23321 is an oligotrophic bacterium isolated from paddy field soil. Although S23321 is phylogenetically close to Bradyrhizobium japonicum USDA110, a legume symbiont, it is unable to induce root nodules in siratro, a legume often used for testing Nod factor-dependent nodulation. The genome of S23321 is a single circular chromosome, 7,231,841 bp in length, with an average GC content of 64.3%. The genome contains 6,898 potential protein-encoding genes, one set of rRNA genes, and 45 tRNA genes. Comparison of the genome structure between S23321 and USDA110 showed strong colinearity; however, the symbiosis islands present in USDA110 were absent in S23321, whose genome lacked a chaperonin gene cluster (groELS3) for symbiosis regulation found in USDA110. A comparison of sequences around the tRNA-Val gene strongly suggested that S23321 contains an ancestral-type genome that precedes the acquisition of a symbiosis island by horizontal gene transfer. Although S23321 contains a nif (nitrogen fixation) gene cluster, the organization, homology, and phylogeny of the genes in this cluster were more similar to those of photosynthetic bradyrhizobia ORS278 and BTAi1 than to those on the symbiosis island of USDA110. In addition, we found genes encoding a complete photosynthetic system, many ABC transporters for amino acids and oligopeptides, two types (polar and lateral) of flagella, multiple respiratory chains, and a system for lignin monomer catabolism in the S23321 genome. These features suggest that S23321 is able to adapt to a wide range of environments, probably including low-nutrient conditions, with multiple survival strategies in soil and rhizosphere.