Microbial genomic taxonomy

A need for a genomic species definition is emerging from several independent studies worldwide. In this commentary paper, we discuss recent studies on the genomic taxonomy of diverse microbial groups and a unified species definition based on genomics. Accordingly, strains from the same microbial species share >95% Average Amino Acid Identity (AAI) and Average Nucleotide Identity (ANI), >95% identity based on multiple alignment genes, 70% in silico Genome-to-Genome Hybridization similarity (GGDH). Species of the same genus will form monophyletic groups on the basis of 16S rRNA gene sequences, Multilocus Sequence Analysis (MLSA) and supertree analysis. In addition to the established requirements for species descriptions, we propose that new taxa descriptions should also include at least a draft genome sequence of the type strain in order to obtain a clear outlook on the genomic landscape of the novel microbe. The application of the new genomic species definition put forward here will allow researchers to use genome sequences to define simultaneously coherent phenotypic and genomic groups

Marinomonas brasilensis sp. nov., isolated from the coral Mussismilia hispida, and reclassification of Marinomonas basaltis as a later heterotypic synonym of Marinomonas communis

A Gram-negative, aerobic bacterium, designated strain R-40503(T), was isolated from mucus of the reef-builder coral Mussismilia hispida, located in the Sao Sebastiao Channel, Sao Paulo, Brazil. Phylogenetic analyses revealed that strain R-40503(T) belongs to the genus Marinomonas. The 16S rRNA gene sequence similarity of R-40503(T) was above 97% with the type strains of Marinomonas vaga, M. basaltis, M. communis and M. pontica, and below 97% with type strains of the other Marinomonas species. Strain R-40503(T) showed less than 35% DNA-DNA hybridization (DDH) with the type strains of the phylogenetically closest Marinomonas species, demonstrating that it should be classified into a novel species. Amplified fragment length polymorphism (AFLP), chemotaxonomic and phenotypic analyses provided further evidence for the proposal of a novel species. Concurrently, a close genomic relationship between M. basaltis and M. communis was observed. The type strains of these two species showed 78% DDH and 63% AFLP pattern similarity. Their phenotypic features were very similar, and their DNA G+C contents were identical (46.3 mol%). Collectively, these data demonstrate unambiguously that Marinomonas basaltis is a later heterotypic synonym of Marinomonas communis. Several phenotypic features can be used to discriminate between Marinomonas species. The novel strain R-40503(T) is clearly distinguishable from its neighbours. For instance, it shows oxidase and urease activity, utilizes L-asparagine and has the fatty acid C(12:1) 3-OH but lacks C(10:0) and C(12:0). The name Marinomonas brasilensis sp. nov. is proposed, with the type strain R-40503(T) (=R-278(T) =LMG 25434(T) =CAIM 1459(T)). The DNA G+C content of strain R-40503(T) is 46.5 mol%

Photobacterium sanctipauli sp nov isolated from bleached Madracis decactis (Scleractinia) in the St Peter & St Paul Archipelago, Mid-Atlantic Ridge, Brazil

Five novel strains of Photobacterium (A-394T, A-373, A-379, A-397 and A-398) were isolated from bleached coralMadracis decactis (scleractinian) in the remote St Peter & St Archipelago (SPSPA), Mid-Atlantic Ridge, Brazil. Healthy M. decactis specimens were also surveyed, but no strains were related to them. The novel isolates formed a distinct lineage based on the 16S rRNA, recA, and rpoA gene sequences analysis. Their closest phylogenetic neighbours were Photobacterium rosenbergii, P. gaetbulicola, and P. lutimaris, sharing 96.6 to 95.8% 16S rRNA gene sequence similarity. The novel species can be differentiated from the closest neighbours by several phenotypic and chemotaxonomic markers. It grows at pH 11, produces tryptophane deaminase, presents the fatty acid C-18:0, but lacks C-16:0 iso. The whole cell protein profile, based in MALDI-TOF MS, distinguished the strains of the novel species among each other and from the closest neighbors. In addition, we are releasing the whole genome sequence of the type strain. The name Photobacterium sanctipauli sp. nov. is proposed for this taxon. The G + C content of the type strain A-394(T) (=LMG27910(T) = CAIM1892(T)) is 48.2 mol%

Genomic and phenotypic attributes of novel salinivibrios from stromatolites, sediment and water from a high altitude lake

Background: Salinivibrios are moderately halophilic bacteria found in salted meats, brines and hypersaline environments. We obtained three novel conspecific Salinivibrio strains closely related to S. costicola, from Socompa Lake, a high altitude hypersaline Andean lake (approx. 3,570 meters above the sea level).Results: The three novel Salinivibrio spp. were extremely resistant to arsenic (up to 200 mM HAsO42-), NaCl (up to 15%), and UV-B radiation (19 KJ/m2, corresponding to 240 minutes of exposure) by means of phenotypic tests. Our subsequent draft genome ionsequencing and RAST-based genome annotation revealed the presence of genes related to arsenic, NaCl, and UV radiation resistance. The three novel Salinivibrio genomes also had the xanthorhodopsin gene cluster phylogenetically related to Marinobacter and Spiribacter. The genomic taxonomy analysis, including multilocus sequence analysis, average amino acid identity, and genome-to-genome distance revealed that the three novel strains belong to a new Salinivibrio species.Conclusions: Arsenic resistance genes, genes involved in DNA repair, resistance to extreme environmental conditions and the possible light-based energy production, may represent important attributes of the novel salinivibrios, allowing these microbes to thrive in the Socompa Lake. © 2014 Gorriti et al.; licensee BioMed Central Ltd.Fil: Gorriti, Marta Fabiana. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; Brasil. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; ArgentinaFil: Dias, Graciela M.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Chimetto, Luciane A.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Trindade-Silva, Amaro E.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Silva, Bruno S.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Mesquita, Milene M.A.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Gregoracci, Gustavo B.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Farias, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Planta Piloto de Procesos Industriales Microbiologicos; ArgentinaFil: Thompson, Cristiane C.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; BrasilFil: Thompson, Fabiano L.. Laboratório de Microbiologia, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brasil ; Brasi

Taxonomy of prokaryontes isolated from cnidarians of São Sebastião, SP, Brasil

Orientadores: Fabiano Lopes Thompson, Marcelo BrocchiTese (doutorado) - Universidade Estadual de Campinas, Instituto de BiologiaResumo: Microrganismos têm um papel fundamental na saúde de cnidários, estabelecendo relações ecológicas diversas, de parasitismo até mutualismo. O conhecimento acerca da diversidade de procariontes associados com cnidários e corais no Brasil ainda é escasso. Este é o primeiro estudo enfocando a caracterização taxonômica de bactérias heterotróficas incluindo fixadoras de nitrogênio (N2) e especialmente vibrios isolados do muco do coral endêmico Mussismilia hispida e de zoantídeos simpátricos Palythoa caribaeorum, Palythoa variabilis e Zoanthus solanderi da região de São Sebastião-SP, Brasil. Ao todo, foram obtidos 488 isolados bacterianos destes cnidários, empregando-se os meios de cultivo Ágar Marinho (AM), Ágar Fixação de Nitrogênio (NFb) e Ágar Tiosulfato-Citrato-Bile-Sacarose (TCBS). Os cnidários foram coletados em três praias (Grande, Portinho e Preta) na região de São Sebastião em 2005 e 2006. A maioria dos isolados obtidos pertenciam à classe Gammaproteobacteria. Mais da metade foram identificados como vibrios com base em sequências parciais do gene 16S rRNA. Vibrios do grupo core corresponderam a 80 % das bactérias potencialmente fixadoras de N2 e revelaram alta similaridade de sequência no gene pyrH com V. harveyi e V. alginolyticus. Dezenove isolados representativos, pertencentes às espécies V. harveyi, V. alginolyticus, V. campbellii e V. parahaemolyticus foram capazes de crescer 6 vezes sucessivas em meio NFb e alguns deles mostraram forte atividade da nitrogenase pelo teste de redução de acetileno (ARA). Cerca de 150 vibrios isolados em TCBS foram caracterizados por meio de sequências de 16S rRNA, recA e pyrH. Os taxa mais abundantes foram V. harveyi, V. rotiferianus, V. campbellii, V. alginolyticus, V. mediterranei (=V. shillonii), V. chagasii, V. tubiashii e uma espécie nova de Vibrio. Com exceção de V. chagasii que foi encontrado apenas no muco de M. hispida, as demais espécies foram observadas nos diferentes hospedeiros, não havendo fortes evidências da presença de espécies hospedeiro-específicas. A variabilidade intra-específica dos vibrios foi avaliada por meio de padrões de banda de elementos palindrômicos extragênicos repetitivos (rep-PCR). Esta análise revelou alta diversidade genômica entre os isolados. Cada isolado apresentava um padrão de rep-PCR distinto, indicando uma grande diversidade de populações co-existindo no mesmo habitat. As identificações geradas pelo rep-PCR concordaram com as identificações obtidas pelo MLSA (Multilocus Sequence Analysis). Diversos isolados de diferentes grupos microbianos apresentaram menos de 97 % de similaridade em sequências do gene 16S rRNA com espécies conhecidas, indicando que elas seriam possíveis espécies novas. Ao todo, sete espécies novas foram descritas por meio da taxonomia polifásica: Marinobacterium coralli sp. nov., Marinomonas brasilensis sp. nov., Photobacterium swingsii sp. nov., P. jeanii sp. nov., Vibrio communis sp. nov., V. variabilis sp. nov. e V. marinum sp. nov. Os dados fenotípicos, quimiotaxonômicos, genotípicos e filogenéticos obtidos a partir da descrição das sete espécies novas demonstraram que há uma ampla variabilidade fenotípica intra-específica. Este estudo determinou a identidade dos principais grupos de bactérias cultivadas do muco de corais, sobretudo, os vibrios. Uma das espécies de vibrios mais abundantes no muco dos corais é V. communis sp. nov. A fixação de N2 representa um fenótipo comum entre as espécies de vibrios estudadas, sugerindo uma função positiva no holobionte.Abstract: Microorganisms have a key role in the health of cnidarians establishing ecologic relationships, of parasitism to mutualism. The knowledge on the diversity of prokaryotes associated with corals in Brazil is very limited. This is the first study on the taxonomic characterization of heterotrophic bacteria, including putative N2-fixing and specially vibrios isolated from mucus of endemic coral Mussismilia hispida and the sympatric zoanthids Palythoa caribaeorum, Palythoa variabilis and Zoanthus solanderi. Cnidarian specimens were isolated in the beaches Grande, Portinho and Preta from São Sebastião region, SP, Brazil, in 2005 and 2006. A total of 488 isolates were obtained using different culture media, i.e. Marine Agar (MA), Nitrogen Fixation (NFb) and Thiosulphate-Citrate- Bile-Sucrose Agar (TCBS). The majority of the isolates fell within the class Gammaproteobacteria and more than half of them were vibrios based on 16S rRNA gene sequences. Based on pyrH gene sequences 80 % of the putative N2-fixing bacteria clustered with the Vibrio core species group (i.e. V. harveyi and V. alginolyticus). Nineteen representative isolates of V. harveyi, V. alginolyticus, V. campbellii and V. parahaemolyticus) were capable of growing six successive times in nitrogen-free medium and some of them showed strong nitrogenase activity by means of the acetylene reduction assay (ARA). About 150 vibrios isolated on TCBS were characterized by means of 16S rRNA, recA and pyrH gene sequences. The most abundant taxa were V. harveyi, V. rotiferianus, V. campbellii, V. alginolyticus, V. mediterranei (=V. shillonii), V. chagasii, V. tubiashii and a new Vibrio species. With the exception of V. chagasii which was found only in the mucus of M. hispida, the other species appeared in different hosts with no strong evidence for the presence of host-specific species. A high genomic diversity was observed using rep-PCR. Each vibrio isolate represented a different co-occuring population, suggesting huge intra-especific diversity. There was a complete agreement between the grouping based on rep-PCR and identification based on MLSA (Multilocus Sequence Analysis). Several isolates had less than 97 % similarity towards know species based on 16S rRNA gene sequence, indicating that they are possibly new species. In total, seven new species were described based on polyphasic taxonomy: Marinobacterium coralli sp. nov., Marinomonas brasilensis sp. nov. Photobacterium swingsii sp. nov., P. jeanii sp. nov., Vibrio communis sp. nov., V. variabilis sp. nov. and V. marinum sp. nov. The new species were characterized by polyphasic taxonomy. Based on phenotypic, chimiotaxonomic, genotypic and phylogenetic data obtained from description of new species, it was demonstrated a huge intra-specific phenotypic variability. This study determined the taxonomic identity of heterotrophic bacteria, especially Vibrio, isolates from cnidarians. V. communis sp. nov. is one of the most frequently found species in the corals. Nitrogen fixation is a common phenotypic trait among vibrios, suggesting that these bacteria may have a positive function in the holobiont.DoutoradoGenetica de MicroorganismosDoutor em Genetica e Biologia Molecula

Quantitative Detection of Active Vibrios Associated with White Plague Disease in Mussismilia braziliensis Corals

Over recent decades several coral diseases have been reported as a significant threat to coral reef ecosystems causing the decline of corals cover and diversity around the world. The development of techniques that improve the ability to detect and quantify microbial agents involved in coral disease will aid in the elucidation of disease cause, facilitating coral disease detection and diagnosis, identification and pathogen monitoring, pathogen sources, vectors, and reservoirs. The genus Vibrio is known to harbor pathogenic strains to marine organisms. One of the best-characterized coral pathogens is Vibrio coralliilyticus, an aetilogic agent of White Plague Disease (WPD). We used Mussismilia coral tissue (healthy and diseased specimens) to develop a rapid reproducible detection system for vibrios based on RT-QPCR and SYBR chemistry. We were able to detect total vibrios in expressed RNA targeting the 16S rRNA gene at 5.23 × 10 6 copies/μg RNA and V. coralliilyticus targeting the pyrH gene at 5.10 × 10 3 copies/μg RNA in coral tissue. Detection of V. coralliilyticus in diseased and in healthy samples suggests that WPD in the Abrolhos Bank may be caused by a consortium of microorganism and not only a single pathogen. We developed a more practical and economic system compared with probe uses for the real-time detection and quantif ication of vibrios from coral tissues by using the 16S rRNA and pyrH gene. This qPCR assay is a reliable tool for the monitoring of coral pathogens, and can be useful to prevent, control, or reduce impacts in this ecosystem

Marinobacterium coralli sp. nov., isolated from mucus of coral (Mussismilia hispida)

A Gram-negative, aerobic bacterium, designated R-40509(T), was isolated from mucus of the reef builder coral (Mussismilia hispida) located in the Sao Sebastiao Channel, Sao Paulo, Brazil. The strain was oxidase-positive and catalase-negative, and required Na(+) for growth. Its phylogenetic position was in the genus Marinobacterium and the closest related species were Marinobacterium sediminicola, Marinobacterium maritimum and Marinobacterium stanieri; the isolate exhibited 16S rRNA gene sequence similarities of 97.5-98.0% with the type strains of these species. 16S rRNA gene sequence similarities with other type strains of the genus Marinobacterium were below 96%. DNA-DNA hybridizations between strain R-40509T and the type strains of the phylogenetically closest species of the genus Marinobacterium revealed less than 70% DNA-DNA relatedness, supporting the novel species status of the strain. Phenotypic characterization revealed that the strain was able to grow at 15-42 degrees C and in medium containing up to 9% NaCl. The isolate could be differentiated from phenotypically related species by several features, including its ability to utilize c-alanine, L-alanine, bromosuccinic acid, beta-hydroxybutyric acid and a-ketovaleric acid, but not acetate or L-arabinose. It produced acetoin (Voges-Proskauer), but did not have esterase lipase (08) or catalase activities. It possessed C(18:1)omega 7(C) (35%), summed feature 3 (iso-C(15:0) 2-OH and/or C16:1 omega 7(C); 25%) and C(16:0) (22%) as major cellular fatty acids. The DNA G+C content was 58.5 mol%. The name Marinobacterium coralli sp. nov. is proposed to accommodate this novel isolate; the type strain is R-40509(T) (=LMG 25435(T) =CAIM 1449(T))

Vibrios dominate as culturable nitrogen-fixing bacteria of the Brazilian coral mussismilia hispida

Taxonomic characterization was performed on the putative N2-fixing microbiota associated with the coral species Mussismilia hispida, and with its sympatric species Palythoa caribaeorum, P. variabilis, and Zoanthus solanderi, off the coast of São Sebastião (São Paulo State, Brazil). The 95 isolates belonged to the Gammaproteobacteria according to the 16S rDNA gene sequences. In order to identify the isolates unambiguously, pyrH gene sequencing was carried out. The majority of the isolates (n=76) fell within the Vibrio core group, with the highest gene sequence similarity being towards Vibrio harveyi and Vibrio alginolyticus. Nineteen representative isolates belonging to V. harveyi (n=7), V. alginolyticus (n=8), V. campbellii (n=3), and V. parahaemolyticus (n=1) were capable of growing six successive times in nitrogen-free medium and some of them showed strong nitrogenase activity by means of the acetylene reduction assay (ARA). It was concluded that nitrogen fixation is a common phenotypic trait among Vibrio species of the core group. The fact that different Vibrio species can fix N2 might explain why they are so abundant in the mucus of different coral species314312319CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPNão temNão temThe authors acknowledge grants from FAPESP, CNPq, FUJB and IFS. L.A. Chimetto acknowledges a PhD scholarship provided by CNPq. We would like to thank Mariza Gondo, Patrícia L. Ramos (ICB-USP), and Álvaro Migotto (CEBIMAR-USP) for technical assistance and valuable comment

Vibrios dominate as culturable nitrogen-fixing bacteria of the Brazilian coral Mussismilia hispida

Taxonomic characterization was performed on the putative N-2-fixing microbiota associated with the coral species Mussismilia hispida, and with its sympatric species Palythoa caribaeorum, P. variabilis, and Zoanthus solanderi, off the coast of Sao Sebastiao (Sao Paulo State, Brazil). The 95 isolates belonged to the Gammaproteobacteria according to the 16S rDNA gene sequences. In order to identify the isolates unambiguously, pyrH gene sequencing was carried out. The majority of the isolates (n = 76) fell within the Vibrio core group, with the highest gene sequence similarity being towards Vibrio harveyi and Vibrio alginolyticus. Nineteen representative isolates belonging to V. harveyi (n = 7), V. alginolyticus (n = 8), V. campbellii (n = 3), and V parahaemolyticus (n = 1) were capable of growing six successive times in nitrogen-free medium and some of them showed strong nitrogenase activity by means of the acetylene reduction assay (ARA). It was concluded that nitrogen fixation is a common phenotypic trait among Vibrio species of the core group. The fact that different Vibrio species can fix N, might explain why they are so abundant in the mucus of different coral species. (C) 2008 Published by Elsevier GmbH.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESPCNPqConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FUJBFUJBIFSIF