Abstract: Lactobacilli are employed in probiotic food preparations and as feed additives in poultry and livestock, due to health benefits associated with their consumption. The objective of this study was to evaluate and compare the probiotic potential of ten lactobacilli strains isolated from commercial dairy food products and animal rumen contents in New Zealand. Genetic identification of the isolates revealed that all belonged to the genus Lactobacillus, specifically the species L.reuteri, L. rhamnosus and L.plantarum. All isolates did not show any haemolytic behaviour. Isolates of dairy origin showed better tolerance to low pH stress. On the other hand, rumen isolates exhibited a higher tolerance to presence of bile salts. All isolates exhibited resistance to aminoglycoside antibiotics, however most were sensitive to ampicillin. Isolates of rumen origin demonstrated a higher inhibitory effect on Listeria monocytogenes, Enterobacter aerogenes and Salmonella menston. Bacterial adherence of all isolates increased with a decrease in pH. This screening study on lactobacilli isolates has assessed and identified potential probiotic candidates for further evaluation.
Abstract: Methyloversatilisuniversalis FAM5 utilizes single carbon compounds such as methanol or methylamine as a sole source of carbon and energy. Expression profiling reveals distinct sets of genes altered during growth on methylamine vs methanol. As expected, all genes for the N-methylglutamate pathway were induced during growth on methylamine. Among other functions responding to the aminated source of C1-carbon, are a heme-containing amine dehydrogenase (Qhp), a distant homologue of formaldehyde activating enzyme (Fae3), molybdenum-containing formate dehydrogenase, ferredoxin reductase, a set of homologues to urea/ammonium transporters and amino-acid permeases. Mutants lacking one of the functional subunits of the amine dehydrogenase (ΔqhpA) or Δfae3 showed no growth defect on C1-compounds. M.universalis FAM5 strains with a lesion in the H4-folate pathway were not able to use any C1-compound, methanol or methylamine. Genes essential for C1-assimilation (the serine cycle and glyoxylate shunt) and H4MTP-pathway for formaldehyde oxidation showed similar levels of expression on both C1-carbon sources. M. universalis FAM5 possesses three homologs of the formaldehyde activating enzyme, a key enzyme of the H4MTP-pathway. Strains lacking the canonical Fae (fae1)lost the ability to grow on both C1-compounds. However, upon incubation on methylamine the fae1-mutant produced revertants (Δfae1R), which regained the ability to grow on methylamine. Double and triple mutants (Δfae1RΔfae3,or Δfae1RΔfae2 or Δfae1RΔfae2Δfae3) constructed in the revertantstrainbackground showed growth similar to the Δfae1R phenotype. The metabolic pathways for utilization of methanol and methylamine in Methyloversatilisuniversalis FAM5 are reconstructed based on these gene expression and phenotypic data.
Abstract: Bioengineering holds great promise to provide fast and efficient biocatalysts for methanol-based biotechnology, but necessitates proven methods to optimize physiology in engineered strains. Here, we highlight experimental evolution as an effective means for optimizing an engineered Methylobacterium extorquens AM1. Replacement of the native formaldehyde oxidation pathway with a functional analog substantially decreased growth in an engineered Methylobacterium, but growth rapidly recovered after six hundred generations of evolution on methanol. We used whole-genome sequencing to identify the basis of adaptation in eight replicate evolved strains, and examined genomic changes in light of other growth and physiological data. We observed great variety in the numbers and types of mutations that occurred, including instances of parallel mutations at targets that may have been “rationalized” by the bioengineer, plus other “illogical” mutations that demonstrate the ability of evolution to expose unforeseen optimization solutions. Notably, we investigated mutations to RNA polymerase, which provided a massive growth benefit but are linked to highly aberrant transcriptional profiles. Overall, we highlight the power of experimental evolution to present genetic and physiological solutions for strain optimization, particularly in systems where the challenges of engineering are too many or too difficult to overcome via traditional engineering methods.
Abstract: Methylotrophs, which can utilize methane and/or methanol as sole carbon and energy sources, are key players in the carbon cycle between methane and CO2, the two most important greenhouse gases. This review describes the relationships between methylotrophs and plants, and between methanotrophs (methane-utilizers, a subset of methylotrophs) and heterotrophic bacteria. Some plants emit methane and methanol from their leaves, and provide methylotrophs with habitats. Methanol-utilizing methylotrophs in the genus Methylobacterium are abundant in the phyllosphere and have the ability to promote the growth of some plants.Methanotrophs also inhabit the phyllosphere, and methanotrophs with high methane oxidation activities have been found on aquatic plants. Both plant and environmental factors are involved in shaping the methylotroph community on plants. Methanotrophic activity can be enhanced by heterotrophic bacteria that provide growth factors (e.g., cobalamin). Information regarding the biological interaction of methylotrophs with other organisms will facilitate a better understanding of the carbon cycle that is driven by methylotrophs.
Abstract: Northern temperate forest soils and Sphagnum-dominated peatlands are a major source and sink of methane. In these ecosystems, methane is mainly oxidized by aerobic methanotrophic bacteria, which are typically found in aerated forest soils, surface peat, and Sphagnum moss. We contrasted methanotrophic bacterial diversity and abundances from the (i) organic horizon of forest soil; (ii) surface peat; and (iii) submerged Sphagnum moss from Cranesville Swamp Preserve, West Virginia, using multiplex sequencing of bacterial 16S rRNA (V3 region) gene amplicons. From ~1 million reads, >50,000 unique OTUs (Operational Taxonomic Units), 29 and 34 unique sequences were detected in the Methylococcaceae and Methylocystaceae, respectively, and 24 potential methanotrophs in the Beijerinckiaceae were also identified. Methylacidiphilum-like methanotrophs were not detected. Proteobacterial methanotrophic bacteria constitute <2% of microbiota in these environments, with the Methylocystaceae one to two orders of magnitude more abundant than the Methylococcaceae in all environments sampled. The Methylococcaceae are also less diverse in forest soil compared to the other two habitats. Nonmetric multidimensional scaling analyses indicated that the majority of methanotrophs from the Methylococcaceae and Methylocystaceae tend to occur in one habitat only (peat or Sphagnum moss) or co-occurred in both Sphagnum moss and peat. This study provides insights into the structure of methanotrophic communities in relationship to habitat type, and suggests that peat and Sphagnum moss can influence methanotroph community structure and biogeography.
Abstract: Gram-positive methylotrophic bacteria have been known for a long period of time, some serving as model organisms for characterizing the specific details of methylotrophy pathways/enzymes within this group. However, genome-based knowledge of methylotrophy within this group has been so far limited to a single species, Bacillus methanolicus (Firmicutes). The paucity of whole-genome data for Gram-positive methylotrophs limits our global understanding of methylotrophy within this group, including their roles in specific biogeochemical cycles, as well as their biotechnological potential. Here, we describe the isolation of seven novel strains of Gram-positive methylotrophs that include two strains of Bacillus and five representatives of Actinobacteria classified within two genera, Arthrobacter and Mycobacterium. We report whole-genome sequences for these isolates and present comparative analysis of the methylotrophy functional modules within these genomes. The genomic sequences of these seven novel organisms, all capable of growth on methylated amines, present an important reference dataset for understanding the genomic basis of methylotrophy in Gram-positive methylotrophic bacteria. This study is a major contribution to the field of methylotrophy, aimed at closing the gap in the genomic knowledge of methylotrophy within this diverse group of bacteria.