Microbiology

Abstract Members of the genus Candidatus Accumulibacter are important in many wastewater treatment systems performing enhanced biological phosphorus removal (EBPR). The Accumulibacter lineage can be subdivided phylogenetically into multiple clades, and previous work showed that these clades are ecologically distinct. The complete genome of Candidatus Accumulibacter phosphatis strain UW-1, a member of Clade IIA, was previously sequenced. Here, we report a draft genome sequence of Candidatus Accumulibacter spp. strain UW-2, a member of Clade IA, assembled following shotgun metagenomic sequencing of laboratory-scale bioreactor sludge. We estimate the genome to be 80–90% complete. Although the two clades share 16S rRNA sequence identity of >98.0%, we observed a remarkable lack of synteny between the two genomes. We identified 2317 genes shared between the two genomes, with an average nucleotide identity (ANI) of 78.3%, and accounting for 49% of genes in the UW-1 genome. Unlike UW-1, the UW-2 genome seemed to lack genes for nitrogen fixation and carbon fixation. Despite these differences, metabolic genes essential for denitrification and EBPR, including carbon storage polymer and polyphosphate metabolism, were conserved in both genomes. The ANI from genes associated with EBPR was statistically higher than that from genes not associated with EBPR, indicating a high selective pressure in EBPR systems. Further, we identified genomic islands of foreign origins including a near-complete lysogenic phage in the Clade IA genome. Interestingly, Clade IA appeared to be more phage susceptible based on it containing only a single Clustered Regularly Interspaced Short Palindromic Repeats locus as compared with the two found in Clade IIA. Overall, the comparative analysis provided a genetic basis to understand physiological differences and ecological niches of Accumulibacter populations, and highlights the importance of diversity in maintaining system functional resilience.

AbstractBackgroundHuanglongbing (HLB) is a worldwide devastating disease of citrus. There are no effective control measures for this newly emerging but century-old disease. Previously, we reported a combination of Penicillin G and Streptomycin was effective in eliminating or suppressing the associated bacterium, ‘CandidatusLiberibacter asiaticus’ (Las).ResultsHere we report the bacterial composition and community structure in HLB-affected citrus plants during a growing season and while being treated with antibiotic combinations PS (Penicillin G and Streptomycin) and KO (Kasugamycin and Oxytetracycline) using the Phylochip™ G3 array. Both antibiotic treatments resulted in significantly lower Las bacterial titers (Pr<0.05) and hybridization scores. Of the 50,000+ available operational taxonomic units (OTUs) on PhyloChip™ G3, 7,028 known OTUs were present in citrus leaf midribs. These OTUs were from 58 phyla, of which five contained 100 or more OTUs,Proteobacteria(44.1%), Firmicutes(23.5%),Actinobacteria(12.4%),Bacteroidetes(6.6%) andCyanobacteria(3.2%). In the antibiotic treated samples, the number of OTUs decreased to a total of 5,599. The over-all bacterial diversity decreased with the antibiotic treatments, as did the abundance of 11 OTUs withinProteobacteria,Firmicutes,BacteroidetesandPlanctomycetes. Within theProteobacteria, ten OTUs representing the classγ-proteobacteria increased in abundance after four months of treatment, when the Las bacterium was at its lowest level in the HLB-affected citrus field plants.ConclusionsOur data revealed thatProteobacteriawas constantly the dominant bacterial phylum recovered from citrus leaf midribs, with the α-proteobacterial and the γ-proteobacterial classes vying for prevalence. In addition, the level of bacterial diversity found in the leaf midribs of field citrus was greater than previously described. Bacterial cells in close proximity may be able to modify their microenvironment, making the composition of the microbial community an important factor in the ability of Las to cause HLB progression. A low Las level was seen as an annual fluctuation, part of the bacterial population dynamics, and as a response to the antibiotic treatments.

Three strictly anaerobic, methanogenic strains JH1T, JH4 and JH8 were isolated from rumen of the Korean native cattle (HanWoo; Bos taurus coreanae) in South Korea. The colonies were circular, opaque, and slightly yellowish. Phylogenetic analyses of 16S rRNA gene and mcrA (encoding α subunit of methyl-coenzyme M reductase) sequences confirmed the affiliation of the novel strains with the Methanobacteriales , and Methanobrevibacter wolinii SHT was the most closely related species. The 16S rRNA gene and mcrA sequence similarities between strains JH1T, JH4 and JH8 and M. wolinii SHT were 96.2 and 89.0 % respectively, and DNA–DNA hybridization of the isolates and M. wolinii DSM 11976T showed a 20 % reassociation. Strain JH1T exhibited 92 % DNA–DNA relatedness with strains JH4 and JH8, and their 16S rRNA gene and mcrA sequences were identical. Cells stained Gram-positive and were non-motile rods, 1.5–1.8 µm long and 0.6 µm wide. The strains were able to use H2/CO2 and formate. The optimum temperature and pH ranges for growth were 37–40 °C and pH 6.5–7.0. The DNA G+C content of strain JH1T was 28 mol%. Based on data from this study using a polyphasic approach, the three strains represent a novel species of genus Methanobrevibacter , for which the name Methanobrevibacter boviskoreani sp. nov. is proposed. The type strain is JH1T ( = KCTC 4102T = JCM 18376T).

A novel halotolerant actinomycete, designated strain BNT52T, was isolated from soil collected from Cihanbeyli Salt Mine in the central Anatolia region of Turkey, and examined using a polyphasic taxonomic approach. The isolate was found to have chemical and morphological properties typical of the genus Amycolatopsis and formed a distinct phyletic line in the 16S rRNA gene tree. Strain BNT52T was most closely related to Amycolatopsis nigrescens CSC17Ta-90T (96.7 %), Amycolatopsis magusensis KT2025T (96.6 %), Amycolatopsis sulphurea DSM 46092T (96.6 %), Amycolatopsis dongchuanensis YIM 75904T (96.5 %), Amycolatopsis ultiminotia RP-AC36T (96.4 %) and Amycolatopsis sacchari DSM 44468T (96.4 %). Sequence similarities with other strains of species of the genus Amycolatopsis were lower than 96.2 %. The isolate grew at 20–37 °C, pH 6–12 and in the presence of 0–10 % (w/v) NaCl. The cell wall of the novel strain contained meso-diaminopimelic acid and arabinose and galactose as the diagnostic sugars. Major fatty acids were iso-C16 : 0 2-OH and iso-C16 : 0. The predominant menaquinone was MK-9(H4). The polar lipids detected were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol and phosphatidylmethylethanolamine. The genomic DNA G+C content was 68.8 mol%. On the basis of the data from this polyphasic taxonomic study, strain BNT52T represents a novel species within the genus Amycolatopsis for which the name Amycolatopsis cihanbeyliensis sp. nov. is proposed (type strain BNT52T = KCTC 29065T = NRRL B-24886T = DSM 45679T).