Isolation of Novel Members of the Family Verrucomicrobiaceae from Methane-Oxidizing Enrichment Cultures and Description of Prosthecobacter flavus sp. nov.

Abstract The Prosthecobacter is one of the first genera described within the bacterial phylum Verrucomicrobiota. This genus accommodates Gram-negative, aerobic, chemoheterotrophic, prosthecate bacteria with a fusiform cell morphology, which inhabit freshwater ecosystems and wastewater treatment plants. The number of described Prosthecobacter species as well as knowledge of their functional roles in the environment remain limited. This study revealed that members of this genus develop in methane-oxidizing enrichment cultures retrieved from freshwater sediments. Two yellow-pigmented isolates of non-motile prosthecate bacteria, strains Pr1 T and Pr3, were obtained from Methylocystis - and Methylomonas -dominated enrichment cultures and were identified as members of the genus Prosthecobacter. These strictly aerobic bacteria utilized a wide range of sugars, some organic acids and grew well on polysaccharides of microbial origin, such as gellan gum and xanthan gum. Methane and methanol were not utilized. Apparently, development of these verrucomicrobia in methane-oxidizing enrichment cultures occurred due to the utilization of exopolysaccharides produced by methanotrophic bacteria. Growth was observed at temperatures between 10 and 35°C (with an optimum at 25°C) and at pH values between 5.5 and 7.5 (with an optimum at pH 7.0). The closest taxonomically characterized phylogenetic relative was Prosthecobacter fluviatilis HAQ-1 T , with 16S rRNA gene sequence similarity of 98.31% to strains Pr1 T and Pr3. The genome of strain Pr1 T was 7.19 Mb in size and contained ~5800 putative protein-coding sequences, one rRNA operon, and 57 tRNA genes. The DNA G + C content was 59.8%. The average nucleotide identity between the genome sequences of strain Pr1 T and Prosthecobacter fluviatilis HAQ-1 T was 83.43%. Based on the genomic and phenotypic difference between strains Pr1 T and Pr3 and other described species of the genus Prosthecobacter , the two novel isolates were classified as representing a novel species of this genus, Prosthecobacter flavus sp. nov. Strain Pr1 T (=VKM B-4037 T = BIM B-2223 T ) is the type strain of the new species.