Search

Desulfomicrobium sp. strain ZS1 is an obligate anaerobic, sulfate-reducing member of the Desulfobacterota from Zodletone Spring, an anoxic sulfide-rich spring in southwestern Oklahoma. Its complete genome was sequenced using a combination of Illumina and Oxford Nanopore platforms and encodes 3,364 proteins and 81 RNAs on a single chromosome.

Establishment of microbial communities in neonatal calves is vital for their growth and overall health. Feed type and associated gastrointestinal tract morphophysiological changes occurring during the pre-weaning, weaning, and post-weaning phases are known to induce shifts in microbial community diversity, structure, and function. However, while the process has received considerable attention for bacteria, our knowledge on temporal progression of anaerobic gut fungi (AGF) in calves is lacking. Here, we examined AGF communities in fecal samples from six dairy cattle collected at 24 different time points during the pre-weaning (day 1-48), weaning (day 49-60), and post-weaning (3-12 months) stages. Quantitative PCR (qPCR) indicated that AGF colonize the calves GIT within 24 hours after birth, with AGF loads slowly increasing during pre-weaning/weaning phases and drastically increasing post-weaning. Culture- independent amplicon surveys identified higher AGF alpha diversity during pre-weaning/ weaning phases, compared to post-weaning. Further, the AGF community structure underwent a drastic shift post-weaning, from a community enriched in the generaKhoyollomyces, Orpinomyces, AL3, and NY8 (some of which commonly encountered in hindgut fermenters) to one enriched in the generaCaecomyces, Piromyces, Pecoramyces, and Cyllamyces, commonly encountered in adult ruminants. Inter-calf community variability was low in the pre- weaning/weaning phases but increased post-weaning. Finally, pairwise comparison of AGF community between calves day 1 post-birth and their mothers suggest a major role for maternal transmission, with additional input from cohabitating subjects. The distinct pattern of AGF progression in calves could best be understood in-light of their narrower niche preferences, metabolic specialization, and physiological optima when compared to bacteria, hence eliciting a unique response to changes in feeding pattern and associated structural development in the GIT of calves during maturation.

The anaerobic gut fungi (AGF) represent a coherent phylogenetic clade within the Mycota. Twenty genera have been described so far. Currently, the phylogenetic and evolutionary relationships between AGF genera remain poorly understood. Here, we utilized 52 transcriptomic datasets from 14 genera to resolve AGF inter-genus relationships using phylogenomics, and to provide a quantitative estimate (amino acid identity, AAI) for intermediate rank assignments. We identify four distinct supra-genus clades, encompassing all genera producing polyflagellated zoospores, bulbous rhizoids, the broadly circumscribed genus Piromyces, and the Anaeromyces and affiliated genera. We also identify the genus Khoyollomyces as the earliest evolving AGF genus. Concordance between phylogenomic outputs and RPB1 and D1/D2 LSU, but not RPB2, MCM7, EF1α or ITS1, phylogenies was observed. We combine phylogenomic analysis and AAI outputs with informative phenotypic traits to propose accommodating 14/20 AGF genera into four families: Caecomycetaceae fam. nov. (encompassing the genera Caecomyces and Cyllamyces), Piromycetaceae fam. nov. (encompassing the genus Piromyces), emend the description of the family Neocallimastigaceae to encompass the genera Neocallimastix, Orpinomyces, Pecoramyces, Feramyces, Ghazallomyces, Aestipascuomyces and Paucimyces, as well as the family Anaeromycetaceae to include the genera Oontomyces, Liebetanzomyces and Capellomyces in addition to Anaeromyces. We refrain from proposing families for the deeply branching genus Khoyollomyces and for genera with uncertain position (Buwchfawromyces, Joblinomyces, Tahromyces, Agriosomyces and Aklioshbomyces) pending availability of additional isolates and sequence data; and these genera are designated as ‘genera incertae sedis’ in the order Neocallimastigales. Our results establish an evolutionary-grounded Linnaean taxonomic framework for the AGF, provide quantitative estimates for rank assignments, and demonstrate the utility of RPB1 as an additional informative marker in Neocallimastigomycota taxonomy.

An overwhelming majority of bacterial life remains uncharacterized. Recent efforts to assemble genomes from metagenomes have provided invaluable insights into these yet-uncultured bacterial lineages. We report on the characterization of 30 genomes belonging to three novel classes within the phylum Desulfobacterota. One class (proposed name Candidatus “Anaeroferrophillalia”) was characterized by the capacity for heterotrophic growth, either fermentatively or utilizing polysulfide, tetrathionate and thiosulfate as electron acceptors. Autotrophic growth using the Wood Ljungdahl pathway and hydrogen or Fe(II) as an electron donor could also occur in absence of organic carbon sources. The second class (proposed name Candidatus “Anaeropigmentia”) was characterized by its capacity for fermentative or aerobic growth at low oxygen thresholds using a broad range of sugars and amino acids, and the capacity to synthesize the methyl/alkyl carrier CoM, an ability that is prevalent in the archaeal but rare in the bacterial domain. Pigmentation is inferred from the capacity for carotenoids (lycopene) production, as well as the occurrence of the majority of genes involved in bacteriochlorophyll a biosynthesis. The third class (proposed name Candidatus “Zymogenia”) was characterized by the capacity for heterotrophic growth fermentatively using broad sugars and amino acids as carbon sources, and the adaptation of some of its members to hypersaline habitats. Analysis of the distribution pattern of all three classes showed their occurrence as rare community members in multiple habitats, with preferences for anaerobic terrestrial (e.g. hydrocarbon contaminated environments, wetlands, bioreactors), freshwater (e.g. ground water and gas-saturated temperate lakes), and marine (e.g. hydrothermal vents, marine sediments, and coastal sediments) environments, over oxygenated (e.g. pelagic ocean and agricultural land) settings. Special preference for some members of the class Candidatus “Zymogenia” to hypersaline environments, e.g. hypersaline microbial mats and lagoons was observed.ImportanceCulture-independent diversity surveys conducted in the last three decades have clearly demonstrated that the scope of microbial diversity is much broader than that inferred from isolation efforts. Multiple reasons have been put forth to explain the refractiveness of a wide range of the earth’s microbiome to isolation efforts. Documenting the scope of high-rank phylogenetic diversity on earth, as well as deciphering and documenting the metabolic capacities, physiological preferences, and putative ecological roles of these yet-uncultured lineages represents one of the central goals in current microbial ecology research. Recent efforts to assemble genomes from metagenomes have provided invaluable insights into these yet-uncultured lineages. This study expands our knowledge of the phylum Desulfobacterota through the characterization of 30 genomes belonging to three novel classes. The analyzed genomes were either recovered from Zodletone Spring in southwestern Oklahoma in this study, or recently binned from public metagenomes as part of the Global Earth Microbiome initiative. Our results expand the high-rank diversity within the bacterial tree of life by describing three novel classes within the phylum Desulfobacterota, document the utilization of multiple metabolic processes, e.g. iron-oxidation, aromatic hydrocarbon degradation, reduction of sulfur-cycling intermediates, and features, e.g. coenzyme M biosynthesis, and pigmentation, as salient characteristics in these novel Desulfobacterota classes.

The anaerobic gut fungi (AGF) represent a coherent phylogenetic clade within the Mycota. Twenty genera have been described so far. Currently, the phylogenetic and evolutionary relationships between AGF genera remain poorly understood. Here, we utilized 53 transcriptomic datasets from 14 genera to resolve AGF inter-genus relationships using phylogenomics, and to provide a quantitative estimate (amino acid identity) for intermediate rank assignments. We identify four distinct supra-genus clades, encompassing genera producing polyflagellated zoospores, bulbous rhizoids, the broadly circumscribed genus Piromyces, and the Anaeromyces and affiliated genera. We also identify the genus Khoyollomyces as the earliest evolving AGF genus. Concordance between phylogenomic outputs and RPB1 and D/D2 LSU, but not RPB2, MCM7, or ITS1, phylogenies was observed. We combine phylogenomic analysis, and AAI outputs with informative phenotypic traits to propose accommodating 13/20 AGF genera into four families: Caecomycetaceae fam. nov. (encompassing genera Caecomyces and Cyllamyces), Piromycetaceae fam. nov. (encompassing the genus Piromyces), emend the description of fam. Neocallimastigaceae to only encompass genera Neocallimastix, Orpinomyces, Pecramyces, Feramyces, Ghazallomyces, and Aestipascuomyces, as well as the family Anaeromycetaceae to include the genera Oontomyces, Liebetanzomyces, and Capellomyces in addition to Anaeromyces. We refrain from proposing families for the deeply branching genus Khoyollomyces, and for genera with uncertain position (Buwchfawromyces, Joblinomyces, Tahromyces, Agriosomyces, Aklioshbomyces, and Paucimyces) pending availability of additional isolates and sequence data. Our results establish an evolutionary- grounded Linnaean taxonomic framework for the AGF, provide quantitative estimates for rank assignments, and demonstrate the utility of RPB1 as additional informative marker in Neocallimastigomycota taxonomy.

Members of the anaerobic gut fungi (Neocallimastigomycota) reside in the rumen and alimentary tract of larger mammalian and some reptilian, marsupial and avian herbivores. The recent decade has witnessed a significant expansion in the number of described Neocallimastigomycota genera and species. However, the difficulties associated with the isolation and maintenance of Neocallimastigomycota strains has greatly complicated comparative studies to resolve inter- and intra-genus relationships. Here, we provide an updated outline of Neocallimastigomycota taxonomy. We critically evaluate various morphological, microscopic and phylogenetic traits previously and currently utilized in Neocallimastigomycota taxonomy, and provide an updated key for quick characterization of all genera. We then synthesize data from taxa description manuscripts, prior comparative efforts and molecular sequence data to present an updated list of Neocallimastigomycota genera and species, with an emphasis on resolving relationships and identifying synonymy between recent and historic strains. We supplement data from published manuscripts with information and illustrations from strains in the authors’ collections. Twenty genera and 36 species are recognized, but the status of 10 species in the genera Caecomyces, Piromyces, Anaeromyces and Cyllamyces remains uncertain due to the unavailability of culture and conferre (cf.) strains, lack of sequence data, and/or inadequacy of available microscopic and phenotypic data. Six cases of synonymy are identified in the genera Neocallimastix and Caecomyces, and two names in the genus Piromyces are rejected based on apparent misclassification.

As tick-borne diseases continue to increase across North America, current research strives to understand how the tick microbiome may affect pathogen acquisition, maintenance, and transmission. Prior high throughput amplicon-based microbial diversity surveys of the widespread tick Dermacentor variabilis have suggested that life stage, sex, and geographic region may influence the composition of the tick microbiome. Here, adult D. variabilis ticks (n = 145) were collected from dogs and cats from 32 states with specimens originating from all four regions of the United States (West, Midwest, South, and Northeast), and the tick microbiome was examined via V4-16S rRNA gene amplification and Illumina sequencing. A total of 481,246 bacterial sequences were obtained (median 2924 per sample, range 399-11,990). Fifty genera represented the majority (80%) of the sequences detected, with the genera Allofrancisella and Francisella being the most abundant. Further, 97%, 23%, and 5.5% of the ticks contained sequences belonging to Francisella spp., Rickettsia spp., and Coxiella spp., respectively. No Ehrlichia spp. or Anaplasma spp. were identified. Co-occurrence analysis, by way of correlation coefficients, between the top 50 most abundant genera demonstrated five strong positive and no strong negative correlation relationships. Geographic region had a consistent effect on species richness with ticks from the Northeast having a significantly greater level of richness. Alpha diversity patterns were dependent on tick sex, with males exhibiting higher levels of diversity, and geographical region, with higher level of diversity observed in ticks obtained from the Northeast, but not on tick host. Community structure, or beta diversity, of tick microbiome was impacted by tick sex and geographic location, with microbiomes of ticks from the western US exhibiting a distinct community structure when compared to those from the other three regions (Northeast, South, and Midwest). In total, LEfSe (Linear discriminant analysis Effect Size) identified 18 specific genera driving these observed patterns of diversity and community structure. Collectively, these findings highlight the differences in bacterial diversity of D. variabilis across the US and supports the interpretation that tick sex and geographic region affects microbiome composition across a broad sampling distribution.

Highlights • Four bacterial strains with record resistance to Cu(II) have been isolated. • Genomic sequences are available for genome mining and studying the genetic determinants of bacterial resistance to Cu(II). • Genomic sequences provide the foundation necessary for transcriptional and functional studies of genes encoding for Cu(II) resistance mechanisms in Pseudomonas spp. • The data will be of interest for a broad community of biotechnologists and microbiologists and will aid in developing novel technologies for copper detoxification in contaminated soils and industrial wastewaters., Environmental copper pollution causes major destruction to ecological systems, which require the development of environmentally friendly biotechnological, in particular, microbial methods for copper removal. These methods rely on the availability of microorganisms resistant to high levels of copper. Here we isolated four bacterial strains with record resistance to up to 1.0 M Cu(II). The strains were isolated from ecologically diverse soil samples, and their genomes were sequenced. A 16S rRNA sequence-based phylogenetic analysis identified that all four isolates belong to the genus Pseudomonas. Particularly, strains UKR1 and UKR2 isolated from Kyiv region in Ukraine were identified as P. lactis and P. panacis, respectively, and strains UKR3 and UKR4 isolated from Svalbard Island in the Arctic Ocean and Galindez Island in Antarctica, respectively, were identified as P. veronii. Initial in-silico screening for genes encoding copper resistance mechanisms showed that all four strains encode copper resistance proteins CopA, CopB, CopD, CopA3, CopZ, as well as two-component regulatory system CusRS, all known to be associated with metal resistance in Pseudomonas genus. Further detailed studies will aim to characterize the full genomic potential of the isolates to enable their application for copper bioremediation in contaminated soils and industrial wastewaters., Graphical abstract Image, graphical abstract

We isolated and characterized 65 anaerobic gut fungal (AGF; Neocallimastigomycota) strains from fecal samples of five wild (W, axis deer, white-tailed deer, Boer goat, mouflon, and Nilgiri tahr), one zoo-housed (Z, zebra), and three domesticated (D, horse, sheep, and goat) herbivores in the US states of Texas (TX) and Oklahoma (OK), Wales (WA), and the Indian states of Kerala (KE) and Haryana (HA). Phylogenetic assessment using the D1-D2 regions of the large subunit (28S) rDNA and internal transcribed spacer 1 (ITS1) identified seven monophyletic clades that are distinct from all currently recognized AGF genera. All strains displayed monocentric thalli and produced exclusively or predominantly monoflagellate zoospores, with the exception of axis deer strains, which produced polyflagellate zoospores. Analysis of amplicon-based AGF diversity surveys indicated that zebra and horse strains are representatives of uncultured AL1 group, whereas domesticated goat and sheep strains are representatives of uncultured AL5 group, previously encountered in fecal and rumen samples of multiple herbivores. The other five lineages, all of which were isolated from wild herbivores, have not been previously encountered in such surveys. Our results significantly expand the genus-level diversity within the Neocallimastigomycota and strongly suggest that wild herbivores represent a yet-untapped reservoir of AGF diversity. We propose seven novel genera and eight novel Neocallimastigomycota species to comprise these strains, for which we propose the names

This article provides an outline of the classification of the kingdom Fungi (including fossil fungi. i.e. dispersed spores, mycelia, sporophores, mycorrhizas). We treat 19 phyla of fungi. These are Aphelidiomycota, Ascomycota, Basidiobolomycota, Basidiomycota, Blastocladiomycota, Calcarisporiellomycota, Caulochytriomycota, Chytridiomycota, Entomophthoromycota, Entorrhizomycota, Glomeromycota, Kickxellomycota, Monoblepharomycota, Mortierellomycota, Mucoromycota, Neocallimastigomycota, Olpidiomycota, Rozellomycota and Zoopagomycota. The placement of all fungal genera is provided at the class-, order- and family-level. The described number of species per genus is also given. Notes are provided of taxa for which recent changes or disagreements have been presented. Fungus-like taxa that were traditionally treated as fungi are also incorporated in this outline (i.e. Eumycetozoa, Dictyosteliomycetes, Ceratiomyxomycetes and Myxomycetes). Four new taxa are introduced: Amblyosporida ord. nov. Neopereziida ord. nov. and Ovavesiculida ord. nov. in Rozellomycota, and Protosporangiaceae fam. nov. in Dictyosteliomycetes. Two different classifications (in outline section and in discussion) are provided for Glomeromycota and Leotiomycetes based on recent studies. The phylogenetic reconstruction of a four-gene dataset (18S and 28S rRNA, RPB1, RPB2) of 433 taxa is presented, including all currently described orders of fungi., Nalin N. Wijayawardene thanks Mushroom Research Foundation and National Science Foundation of China (No. NSFC 31950410558) for financially supporting this project. Kevin D. Hyde acknowledges the Foreign Experts Bureau of Yunnan Province, Foreign Talents Program (2018; grant no. YNZ2018002), Thailand Research grants entitled Biodiversity, phylogeny and role of fungal endophytes on above parts of Rhizophora apiculata and Nypa fruticans (grant no: RSA5980068), the future of specialist fungi in a changing climate: baseline data for generalist and specialist fungi associated with ants, Rhododendron species and Dracaena species (grant no: DBG6080013), Impact of climate change on fungal diversity and biogeography in the Greater Mekong Subregion (grant no: RDG6130001). H.T. Lumbsch thanks support by the Grainger Bioinformatics Center. E. Malosso is grateful to CAPES for financial support (grant no. 88881.062172/2014-01). B.T. Goto, G.A. Silva and K. Jobim, L.C. Maia acknowledges CNPq (Brazilian Scientific Council, grants no. 465.420/2014-1, 307.129/2015-2 and 408011/2016-5) and CAPES for support. The study was partially supported by the National Science Centre, Poland, under Grants No. 2015/17/D/NZ8/00778 and 2017/25/B/NZ8/00473 to Julia Pawłowska. The research of Martin Kukwa received support from the National Science Centre (NCN) in Poland (project no 2015/17/B/NZ8/02441). Alan J.L. Phillips acknowledges the support from UID/MULTI/04046/2019 Research Unit grant from FCT, Portugal to BioISI. H. Zhang is financially supported by the National Natural Science Foundation of China (Project ID: NSF 31500017). S. Boonmee would like to thank the Thailand Research Fund (Project No. TRG6180001). Dong-Qin Dai and Li-Zhou Tang would like to thank the National Natural Science Foundation of China (No. NSFC 31760013, NSFC 31260087, NSFC 31460561), the Scientific Research Foundation of Yunnan Provincial Department of Education (2017ZZX186) and the Thousand Talents Plan, Youth Project of Yunnan Provinces for support. R. Phookamsak, M. Doilom, D. N. Wanasinghe, S.C. Karunarathna and J.C. Xu express sincere appreciations to Key Research Program of Frontier Sciences of the Chinese Academy of Sciences (grant no. QYZDY-SSW-SMC014) for research financial support. R. Phookamsak thanks the Yunnan Provincial Department of Human Resources and Social Security (grant no. Y836181261), Chiang Mai University and National Science Foundation of China (NSFC) project code 31850410489 for research financial support. S.C. Kaunarathna thanks CAS President’s International Fellowship Initiative (PIFI) for funding his postdoctoral research (No. 2018PC0006) and the National Science Foundation of China (NSFC) for funding this work under the project code 31851110759. S. Tibpromma would like to thank the International Postdoctoral Exchange Fellowship Program (number Y9180822S1), CAS President’s International Fellowship Initiative (PIFI) (number 2020PC0009), China Postdoctoral Science Foundation and the Yunnan Human Resources, and Social Security Department Foundation for funding her postdoctoral research. Yuri S. Tokarev, Elena S. Nassonova and Irma V. Issi are indebtful to Yuliya Y. Sokolova (Institute of Cytology RAS, St. Petersbug, Russia) and Anastasia V. Simakova (Tomsk State University, Tomsk, Russia) for kind permission of reproduction of electron microscopy images of Metchnikovella incurvata and Crepidulospora beklemishevi, respectively. Yuri S. Tokarev and Irma V. Issi thank Russian Foundation of Basic Research, grant number 17-04-00871 (taxonomy of Rozellomycota). Elena S. Nassonova thank Russian Foundation of Basic Research, grant number 18-04-01359 (early evolution of Microsporidia, phylogeny of Metchnikovellida). Adam Flakus and Pamela Rodriguez-Flakus are greatly indebted to all staff of the Herbario Nacional de Bolivia, Instituto de Ecología, Universidad Mayor de San Andrés, La Paz and the SERNAP (http://sernap.gob.bo), for their generous cooperation providing permits, assistance and facilities support for scientific studies. The research of AF and PRF were financially supported by the National Science Centre (NCN) in Poland (DEC-2013/11/D/NZ8/03274). Adam Flakus and Pamela Rodriguez-Flakus received additional support under statutory funds from the W. Szafer Institute of Botany, Polish Academy of Sciences, Krakow, Poland. The authors would like to thank Yunnan Innovation Platform for Development and Utilization of Symbiotic Fungi Resources for finance support. Li-Fang Zhang would like to thank grant-in-aid from Science and Technology Department of Yunnan Province (2018FD080) for finance support. Chun-Ying Deng thanks the Biodiversity Survey and Assessment Project of the Ministry of Ecology and Environment, China (2019HJ2096001006). Yingqian Kang would like to thank Guizhou Scientific Plan Project [(2019) 2873]; Excellent Youth Talent Training Project of Guizhou Province [(2017) 5639]; Guiyang Science and Technology Project [(2017) No. 5-19]; Talent Base Project of Guizhou Province, China [FCJD2018-22]; Research Fund of Education Bureau of Guizhou Province, China [(2018) 481]. D. N. Wanasinghe would like to thank the CAS President’s International Fellowship Initiative (PIFI) for funding his postdoctoral research (number 2019PC0008), the National Science Foundation of China and the Chinese Academy of Sciences for financial support under the following grants: 41761144055, 41771063 and Y4ZK111B01. Yuri K. Novozhilov and Oleg N. Shchepin acknowledge support from the Russian Foundation of Basic Research, project 18-04-01232 А. Ivana Kušan, Neven Matočec, Armin Mešić and Zdenko Tkalčec are grateful to Croatian Science Foundation for their financial support under the project grant HRZZ-IP-2018-01-1736 (ForFungiDNA). K. Tanaka would like to thank the Japan Society for the Promotion of Science (JSPS 19K06802)

Members of the anaerobic gut fungi (Neocallimastigomycota) reside in the rumen and alimentary tract of larger mammalian and some reptilian, marsupial, and avian herbivores. The recent decade has witnessed a significant expansion in the number of described Neocallimastigomycota genera and species. However, the lack of a centralized culture collection, coupled with difficulties associated with their isolation and maintenance has greatly complicated comparative studies to resolve inter- and intra-genus relationships. Here, we provide an updated outline of Neocallimastigomycota taxonomy. We critically evaluate various morphological, microscopic, and phylogenetic traits previously and currently utilized in Neocallimastigomycota taxonomy, and provide an updated key for quick characterization of all genera. We then synthesize data from taxa description manuscripts, prior comparative efforts, and molecular sequence data to present an updated list of Neocallimastigomycota genera and species, with emphasis on resolving relationships and identifying synonymy between recent and historic strains. We supplement data from published manuscripts with information and illustrations from strains in the authors’ collections. Twenty genera and thirty-six species are recognized, but the status of ten species in the genera Caecomyces, Piromyces, Anaeromyces, and Cyllamyces remains uncertain due to the unavailability of culture and conferre (cf.) strains, lack of sequence data, and/or inadequacy of available microscopic and phenotypic data. Six cases of synonymy are identified in the genera Neocallimastix and Caecomyces, and two names in the genus Piromyces are rejected based on apparent misclassification. &nbsp

Cultured Myxococcota are predominantly aerobic soil inhabitants, characterized by their highly coordinated predation and cellular differentiation capacities. Little is currently known regarding yet-uncultured Myxococcota from anaerobic, nonsoil habitats. We analyzed genomes representing one novel order (o__JAFGXQ01) and one novel family (f__JAFGIB01) in the Myxococcota from an anoxic freshwater spring (Zodletone Spring) in Oklahoma, USA. Compared to their soil counterparts, anaerobic Myxococcota possess smaller genomes and a smaller number of genes encoding biosynthetic gene clusters (BGCs), peptidases, one- and two-component signal transduction systems, and transcriptional regulators. Detailed analysis of 13 distinct pathways/processes crucial to predation and cellular differentiation revealed severely curtailed machineries, with the notable absence of homologs for key transcription factors (e.g., FruA and MrpC), outer membrane exchange receptor (TraA), and the majority of sporulation-specific and A-motility-specific genes. Further, machine learning approaches based on a set of 634 genes informative of social lifestyle predicted a nonsocial behavior for Zodletone Myxococcota. Metabolically, Zodletone Myxococcota genomes lacked aerobic respiratory capacities but carried genes suggestive of fermentation, dissimilatory nitrite reduction, and dissimilatory sulfate-reduction (in f_JAFGIB01) for energy acquisition. We propose that predation and cellular differentiation represent a niche adaptation strategy that evolved circa 500 million years ago (Mya) in response to the rise of soil as a distinct habitat on Earth. IMPORTANCE The phylum Myxococcota is a phylogenetically coherent bacterial lineage that exhibits unique social traits. Cultured Myxococcota are predominantly aerobic soil-dwelling microorganisms that are capable of predation and fruiting body formation. However, multiple yet-uncultured lineages within the Myxococcota have been encountered in a wide range of nonsoil, predominantly anaerobic habitats, and the metabolic capabilities, physiological preferences, and capacity of social behavior of such lineages remain unclear. Here, we analyzed genomes recovered from a metagenomic analysis of an anoxic freshwater spring in Oklahoma, USA, that represent novel, yet-uncultured, orders and families in the Myxococcota. The genomes appear to lack the characteristic hallmarks for social behavior encountered in Myxococcota genomes and displayed a significantly smaller genome size and a smaller number of genes encoding biosynthetic gene clusters, peptidases, signal transduction systems, and transcriptional regulators. Such perceived lack of social capacity was confirmed through detailed comparative genomic analysis of 13 pathways associated with Myxococcota social behavior, as well as the implementation of machine learning approaches to predict social behavior based on genome composition. Metabolically, these novel Myxococcota are predicted to be strict anaerobes, utilizing fermentation, nitrate reduction, and dissimilarity sulfate reduction for energy acquisition. Our results highlight the broad patterns of metabolic diversity within the yet-uncultured Myxococcota and suggest that the evolution of predation and fruiting body formation in the Myxococcota has occurred in response to soil formation as a distinct habitat on Earth.

Cultured Myxococcota are predominantly aerobic soil inhabitants, characterized by their highly coordinated predation and cellular differentiation capacities. Little is currently known regarding yet-uncultured Myxococcota from anaerobic, nonsoil habitats. We analyzed genomes representing one novel order (o__JAFGXQ01) and one novel family (f__JAFGIB01) in the Myxococcota from an anoxic freshwater spring (Zodletone Spring) in Oklahoma, USA. Compared to their soil counterparts, anaerobic Myxococcota possess smaller genomes and a smaller number of genes encoding biosynthetic gene clusters (BGCs), peptidases, one- and two-component signal transduction systems, and transcriptional regulators. Detailed analysis of 13 distinct pathways/processes crucial to predation and cellular differentiation revealed severely curtailed machineries, with the notable absence of homologs for key transcription factors (e.g., FruA and MrpC), outer membrane exchange receptor (TraA), and the majority of sporulation-specific and A-motility-specific genes. Further, machine learning approaches based on a set of 634 genes informative of social lifestyle predicted a nonsocial behavior for Zodletone Myxococcota. Metabolically, Zodletone Myxococcota genomes lacked aerobic respiratory capacities but carried genes suggestive of fermentation, dissimilatory nitrite reduction, and dissimilatory sulfate-reduction (in f_JAFGIB01) for energy acquisition. We propose that predation and cellular differentiation represent a niche adaptation strategy that evolved circa 500 million years ago (Mya) in response to the rise of soil as a distinct habitat on Earth. IMPORTANCE The phylum Myxococcota is a phylogenetically coherent bacterial lineage that exhibits unique social traits. Cultured Myxococcota are predominantly aerobic soil-dwelling microorganisms that are capable of predation and fruiting body formation. However, multiple yet-uncultured lineages within the Myxococcota have been encountered in a wide range of nonsoil, predominantly anaerobic habitats, and the metabolic capabilities, physiological preferences, and capacity of social behavior of such lineages remain unclear. Here, we analyzed genomes recovered from a metagenomic analysis of an anoxic freshwater spring in Oklahoma, USA, that represent novel, yet-uncultured, orders and families in the Myxococcota. The genomes appear to lack the characteristic hallmarks for social behavior encountered in Myxococcota genomes and displayed a significantly smaller genome size and a smaller number of genes encoding biosynthetic gene clusters, peptidases, signal transduction systems, and transcriptional regulators. Such perceived lack of social capacity was confirmed through detailed comparative genomic analysis of 13 pathways associated with Myxococcota social behavior, as well as the implementation of machine learning approaches to predict social behavior based on genome composition. Metabolically, these novel Myxococcota are predicted to be strict anaerobes, utilizing fermentation, nitrate reduction, and dissimilarity sulfate reduction for energy acquisition. Our results highlight the broad patterns of metabolic diversity within the yet-uncultured Myxococcota and suggest that the evolution of predation and fruiting body formation in the Myxococcota has occurred in response to soil formation as a distinct habitat on Earth.

Approaches for recovering and analyzing genomes belonging to novel, hitherto unexplored bacterial lineages have provided invaluable insights into the metabolic capabilities and ecological roles of yet-uncultured taxa. The phylum Acidobacteria is one of the most prevalent and ecologically successful lineages on earth yet, currently, multiple lineages within this phylum remain unexplored. Here, we utilize genomes recovered from Zodletone spring, an anaerobic sulfide and sulfur-rich spring in southwestern Oklahoma, as well as from multiple disparate soil and non-soil habitats, to examine the metabolic capabilities and ecological role of members of the family UBA6911 (group18) Acidobacteria. The analyzed genomes clustered into five distinct genera, with genera Gp18_AA60 and QHZH01 recovered from soils, genus Ga0209509 from anaerobic digestors, and genera Ga0212092 and UBA6911 from freshwater habitats. All genomes analyzed suggested that members of Acidobacteria group 18 are metabolically versatile heterotrophs capable of utilizing a wide range of proteins, amino acids, and sugars as carbon sources, possess respiratory and fermentative capacities, and display few auxotrophies. Soil-dwelling genera were characterized by larger genome sizes, higher number of CRISPR loci, an expanded carbohydrate active enzyme (CAZyme) machinery enabling de-branching of specific sugars from polymers, possession of a C1 (methanol and methylamine) degradation machinery, and a sole dependence on aerobic respiration. In contrast, non-soil genomes encoded a more versatile respiratory capacity for oxygen, nitrite, sulfate, trimethylamine N-oxide (TMAO) respiration, as well as the potential for utilizing the Wood Ljungdahl (WL) pathway as an electron sink during heterotrophic growth. Our results not only expand our knowledge of the metabolism of a yet-uncultured bacterial lineage, but also provide interesting clues on how terrestrialization and niche adaptation drives metabolic specialization within the Acidobacteria.ImportanceMembers of the Acidobacteria are important players in global biogeochemical cycles, especially in soils. A wide range of Acidobacterial lineages remain currently unexplored. We present a detailed genomic characterization of genomes belonging to the family UBA6911 (also known as group 18) within the phylum Acidobacteria. The genomes belong to different genera and were obtained from soil (genera Gp18_AA60 and QHZH01), freshwater habitats (genera Ga0212092 and UBA6911), and anaerobic digestor (Genus Ga0209509). While all members of the family shared common metabolic features, e.g. heterotrophic respiratory abilities, broad substrate utilization capacities, and few auxotrophies; distinct differences between soil and non-soil genera were observed. Soil genera were characterized by expanded genomes, higher numbers of CRISPR loci, larger carbohydrate active enzyme (CAZyme) repertoire enabling monomer extractions from polymer side chains, and methylotrophic (methanol and methylamine) degradation capacities. In contrast, non-soil genera encoded more versatile respiratory capacities for utilizing nitrite, sulfate, TMAO, and the WL pathway, in addition to oxygen as electron acceptors. Our results not only broaden our understanding of the metabolic capacities within the Acidobacteria, but also, provide interesting clues on how terrestrialization shaped Acidobacteria evolution and niche adaptation.

Serratia marcescens is a Gram-negative bacterium with both environmental and host-associated strains. Pigmentation is reportedly inversely correlated with infection frequency, and prodigiosin is one of Serratia pigments that has medical and industrial applications. Here, we report the draft genome sequence of prodigiosin-hyperproducing Serratia marcescens strain N2, isolated from Cairo, Egypt. The sequence is assembled into 142 contigs, with a combined size of 5,570,793 bp. The assembled genome carries typical S. marcescens genes, with potential prodigiosin-biosynthesizing genes detected.

Life emerged and diversified in the absence of molecular oxygen. The prevailing anoxia and unique sulfur chemistry in the Paleo-, Meso- and Neoarchean, and early Proterozoic eons may have supported microbial communities that are drastically different than those currently thriving on the earth’s surface. Zodletone spring in southwestern Oklahoma represents a unique habitat where spatial sampling could substitute for geological eons: from the anoxic, surficial light exposed sediments simulating a preoxygenated earth, to overlaid water column where air exposure simulates the relentless oxygen intrusion during the Neo Proterozoic. We document a remarkably diverse microbial community in the anoxic spring sediments, with 340/516 (65.89%) of genomes recovered in a metagenomic survey belonging to 200 bacterial and archaeal families that were either previously undescribed or that exhibit an extremely rare distribution on the current earth. Such diversity is underpinned by the widespread occurrence of sulfite-, thiosulfate, tetrathionate-, and sulfur-reduction, and paucity of sulfate-reduction machineries in these taxa; hence greatly expanding lineages mediating reductive sulfur cycling processes in the tree of life. Analysis of the overlaying water community demonstrated that oxygen intrusion lead to the development of a significantly less diverse community dominated by well-characterized lineages and a prevalence of oxidative sulfur cycling processes. Such transition from ancient novelty to modern commonality underscores the profound impact of the great oxygenation event on the earth’s surficial anoxic community. It also suggests that novel and rare lineages encountered in current anaerobic habitats could represent taxa once thriving in an anoxic earth, but have failed to adapt to earth’s progressive oxygenation.

We analyzed five metagenome-assembled genomes (MAGs) belonging to the rare, yet-uncultured phylum CSSED10-310 recovered from the anoxic sediments of Zodletone Spring (Oklahoma, USA). Our analysis suggests their potential involvement in sulfite respiration.

The accumulation of genomes of uncultured organisms has highlighted the need for devising a taxonomic and nomenclature scheme to validate names and prevent redundancies. We here report on the recovery and analysis of four phylogenetically related genomes recovered from an anoxic sulfide and sulfur-rich spring (Zodletone spring) in southwestern Oklahoma. Phylogenetic analysis based on 120 single copy markers attested to their position as a novel distinct bacterial phylum. Genomic analysis suggests Gram-negative flagellated organisms that possess type IV pili. The organisms are predicted to be rod-shaped, slow-growers, with an anoxic, heterotrophic, and fermentative lifestyle. Predicted substrate utilization pattern includes multiple amino acids, dipeptides, tripeptides, and oligpopeptides; as well as few sugars. Predicted auxotrophies include proline, vitamin B6, lipoic acid, biotin, and vitamin B12. Assessment of the putative global distribution pattern of this novel lineage suggests its preference to anoxic marine, terrestrial, hydrocarbon-impacted, and freshwater habitats. We propose the candidatus name Krumholzibacterium zodletonense gen. nov, sp. nov. for Zgenome0171T, with the genome serving as the type material for the novel family Krumholzibacteriaceae fam. nov., order Krumholzibacteriales ord. nov., class Krumholzibacteria class nov., and phylum Krumholzibacteriota phyl. nov. The type material genome assembly is deposited in GenBank under accession number QTKG01000000.

The recent leveraging of genome-resolved metagenomics has generated an enormous number of genomes from novel uncultured microbial lineages yet left many clades undescribed. Here, we present a global analysis of genomes belonging to Binatota (UBP10), a globally distributed, yet-uncharacterized bacterial phylum. All orders in Binatota encoded the capacity for aerobic methylotrophy using methanol, methylamine, sulfomethanes, and chloromethanes as the substrates. Methylotrophy in Binatota was characterized by order-specific substrate degradation preferences, as well as extensive metabolic versatility, i.e., the utilization of diverse sets of genes, pathways, and combinations to achieve a specific metabolic goal. The genomes also encoded multiple alkane hydroxylases and monooxygenases, potentially enabling growth on a wide range of alkanes and fatty acids. Pigmentation is inferred from a complete pathway for carotenoids (lycopene, β- and γ-carotenes, xanthins, chlorobactenes, and spheroidenes) production. Further, the majority of genes involved in bacteriochlorophyll a, c, and d biosynthesis were identified, although absence of key genes and failure to identify a photosynthetic reaction center preclude proposing phototrophic capacities. Analysis of 16S rRNA databases showed the preferences of Binatota to terrestrial and freshwater ecosystems, hydrocarbon-rich habitats, and sponges, supporting their potential role in mitigating methanol and methane emissions, breakdown of alkanes, and their association with sponges. Our results expand the lists of methylotrophic, aerobic alkane-degrading, and pigment-producing lineages. We also highlight the consistent encountering of incomplete biosynthetic pathways in microbial genomes, a phenomenon necessitating careful assessment when assigning putative functions based on a set-threshold of pathway completion.IMPORTANCE A wide range of microbial lineages remain uncultured, yet little is known regarding their metabolic capacities, physiological preferences, and ecological roles in various ecosystems. We conducted a thorough comparative genomic analysis of 108 genomes belonging to the Binatota (UBP10), a globally distributed, yet-uncharacterized bacterial phylum. We present evidence that members of the order Binatota specialize in methylotrophy and identify an extensive repertoire of genes and pathways mediating the oxidation of multiple one-carbon (C1) compounds in Binatota genomes. The occurrence of multiple alkane hydroxylases and monooxygenases in these genomes was also identified, potentially enabling growth on a wide range of alkanes and fatty acids. Pigmentation is inferred from a complete pathway for carotenoids production. We also report on the presence of incomplete chlorophyll biosynthetic pathways in all genomes and propose several evolutionary-grounded scenarios that could explain such a pattern. Assessment of the ecological distribution patterns of the Binatota indicates preference of its members to terrestrial and freshwater ecosystems characterized by high methane and methanol emissions, as well as multiple hydrocarbon-rich habitats and marine sponges.

Pseudomonas aeruginosa infects patients with cystic fibrosis, burns, wounds and implants. Previously, our group showed that elevated Ca2+ positively regulates the production of several virulence factors in P. aeruginosa , such as biofilm formation, production of pyocyanin and secreted proteases. We have identified a Ca2+-regulated β-propeller putative phytase, CarP, which is required for Ca2+ tolerance, regulation of the intracellular Ca2+ levels, and plays a role in Ca2+ regulation of P. aeruginosa virulence. Here, we studied the conservation of carP sequence and its occurrence in diverse phylogenetic groups of bacteria. In silico analysis revealed that carP and its two paralogues PA2017 and PA0319 are primarily present in P. aeruginosa and belong to the core genome of the species. We identified 155 single nucleotide alterations within carP, 42 of which lead to missense mutations with only three that affected the predicted 3D structure of the protein. PCR analyses with carP-specific primers detected P. aeruginosa specifically in 70 clinical and environmental samples. Sequence comparison demonstrated that carP is overall highly conserved in P. aeruginosa isolated from diverse environments. Such evolutionary preservation of carP illustrates its importance for P. aeruginosa adaptations to diverse environments and demonstrates its potential as a biomarker.

We report on the isolation of the previously uncultured Neocallimastigomycota SK4 lineage by two independent research groups from a wild aoudad sheep rumen sample (Texas, USA) and an alpaca fecal sample (Baden-Württemberg, Germany). Isolates from both locations showed near identical morphological and microscopic features, forming medium-sized white filamentous colonies with a white center of sporangia on agar roll tubes and a heavy biofilm in liquid media. Microscopic analysis revealed monocentric thalli, and spherical polyflagellated zoospores with 7–20 flagella. Zoospore release occurred through an apical pore as well as by sporangial wall rupturing, a duality that is unique amongst described AGF strains. Isolates were capable of growing on a wide range of mono-, oligo-, and polysaccharides substrates. Phylogenetic assessment based on the D1-D2 large rRNA subunit (D1-D2 LSU) and internal transcribed spacer-1 (ITS-1) regions demonstrated high sequence identity (minimum identity of 99.07% and 96.96%, respectively) between all isolates; but low sequence identity (92.4% and 86.7%, respectively) to their closest cultured relatives. D1-D2 LSU phylogenetic trees grouped the isolates as a new monophyletic clade within the Orpinomyces-Neocallimastix-Pecoramyces-Feramyces-Ghazallamyces supragenus group. D1-D2 LSU and ITS-1 sequences from the obtained isolates were either identical, or displayed extremely high sequence similarity to sequences recovered from the same Aoudad sheep sample on which isolation was conducted, as well as several sequences recovered from domestic sheep and few other herbivores. Interestingly, members of the SK4 clade seem to be encountered in animals grazing on summer pasture. We hence propose accommodating these novel isolates in a new genus, Aestipascuomyces (derived from the Latin word for “summer pasture”), and a new species, A. dupliciliberatus. The type strain is Aestipascuomyces dupliciliberatus strain R4.

The recent leveraging of genome-resolved metagenomics has opened a treasure trove of genomes from novel uncultured microbial lineages, yet left many clades undescribed. We here present a global analysis of genomes belonging to the Binatota (UBP10), a globally distributed, yet-uncharacterized bacterial phylum. All orders in the Binatota encoded the capacity for aerobic methylotrophy using methanol, methylamine, sulfomethanes, chloromethanes, and potentially methane as substrates. Methylotrophy in the Binatota was characterized by order-specific substrate degradation preferences, as well as extensive metabolic versatility, i.e. the utilization of diverse sets of genes, pathways and combinations to achieve a specific metabolic goal. The genomes also encoded an arsenal of alkane hydroxylases and monooxygenases, potentially enabling growth on a wide range of alkanes and fatty acids. Pigmentation is inferred from a complete pathway for carotenoids (lycopene, β and γ carotenes, xanthins, chlorobactenes, and spheroidenes) production. Further, the majority of genes involved in bacteriochlorophyll a, c, and d biosynthesis were identified; although absence of key genes and failure to identify a photosynthetic reaction center precludes proposing phototrophic capacities. Analysis of 16S rRNA databases showed Binatota’s preferences to terrestrial and freshwater ecosystems, hydrocarbon-rich habitats, and sponges supporting their suggested potential role in mitigating methanol and methane emissions, alkanes degradation, and nutritional symbiosis with sponges. Our results expand the lists of methylotrophic, aerobic alkane degrading, and pigment-producing lineages. We also highlight the consistent encountering of incomplete biosynthetic pathways and gene shrapnel in microbial genomes, a phenomenon necessitating careful assessment when assigning putative functions based on a set-threshold of pathway completion.

The anaerobic gut fungi (AGF, Neocallimastigomycota) reside in the alimentary tracts of herbivores where they play a central role in the breakdown of plant material. Here, we report on the development of the hypervariable domains D1/D2 of the large ribosomal subunit (D1/D2 LSU) as a barcoding marker for the AGF. We generated a reference D1/D2 LSU database for all cultured AGF genera, as well as the majority of candidate genera encountered in prior internal transcribed spacer 1 (ITS1)-based surveys. Subsequently, a D1/D2 LSU-based diversity survey using long read PacBio SMRT sequencing was conducted on faecal samples from 21 wild and domesticated herbivores. Twenty-eight genera and candidate genera were identified, including multiple novel lineages that were predominantly, but not exclusively, identified in wild herbivores. Association between certain AGF genera and animal lifestyles, or animal host family was observed. Finally, to address the current paucity of AGF isolates, concurrent isolation efforts utilizing multiple approaches to maximize recovery yielded 216 isolates belonging to 12 different genera, several of which have no prior cultured-representatives. Our results establish the utility of D1/D2 LSU and PacBio sequencing for AGF diversity surveys, the culturability of multiple AGF taxa, and demonstrate that wild herbivores represent a yet-untapped reservoir of AGF diversity.

The anaerobic gut fungi (Neocallimastigomycota), first described almost 50 years ago, hold extraordinary potential for biotechnology. Anaerobic fungi could contribute to bioenergy and bio-based chemical production via their ability to degrade lignocellulose, may enhance animal health and production, and are now being revealed to have interesting biosynthetic potential for development. In this chapter, we briefly describe the biology of these species, discuss recent efforts to identify and exploit anaerobic fungal enzymes, and survey challenges and opportunities for their adoption in a variety of biotechnology applications.

We examined the feasibility of stimulating methanogenic crude oil biodegradation through the combined supply of CO2 and protein-rich matter in anaerobic microcosms, without the external supply of H2, and using formation waters and crude oil collected from the Stillwater and Cushing oil fields of Oklahoma, USA. We found that, depending on the chemical and microbiological composition of the formation water, protein-rich matter can stimulate methanogenic crude oil biodegradation if supplied along with CO2. For the low salinity (116, 714 mg/L) Stillwater oil field formation water, during the early stages of the stimulation process, CH4 was produced by the microbiological reduction of the supplied CO2 with H2 produced from the syntrophic/fermentative biodegradation of the supplied protein-rich matter. As indicated by 16S rRNA gene community analysis, this resulted in a syntrophic enrichment of a methanogenic crude oil-degrading microbial community and led to >42% crude oil biodegradation with the concurrent production of CH4 after 100 days of incubation at 50 °C. For the high salinity (176,665 mg/L) Cushing oil field formation water, although H2 was produced, enrichment of a methanogenic crude oil-degrading microbial community—and thus CH4 production—was not possible. We tested two types of protein-rich matter (yeast extract and isolated soy protein) and found that isolated soy protein can be used as a substitute for yeast extract in field-scale applications. These findings have large implications toward the coupling of geological CO2 storage and microbial enhanced oil recovery in depleted oil reservoirs.

Anaerobic gut fungi are common inhabitants of the alimentary tracts of herbivorous animals. Nine different Neocallimastigomycota genera have been described so far. However, culture-independent diversity surveys suggest the presence of numerous, yet-uncultured Neocallimastigomycota genera. Here, we report on the isolation and characterization of the first cultured representatives of Neocallimastigomycota clade AL6, originally identified in culture-independent surveys of fecal samples from captive wild animals. Six strains were isolated from rumen and fecal samples of a wild Barbary sheep (Ammotragus lervia) and a wild fallow deer (Dama dama) in Texas, USA. The isolates displayed medium-sized (3-7 mm) circular beige colonies with filamentous edges and a dark center on agar roll tubes. Microscopic analysis revealed monocentric thalli with both endogenous and exogenous sporangial development patterns. Zoospores were spherical, with a diameter of 9.6 ±1.9 μm, and polyflagellated, with 7-16 flagella. Phylogenetic analysis based on nuc rDNA ITS1 region and D1-D2 domains of nuc 28S rDNA revealed that the isolated strains formed a single monophyletic and bootstrap-supported clade distinct from all currently described Neocallimastigomycota genera. Substrate utilization experiments using the type strain (F3a) demonstrated robust and fast growth on sugars and plant biomass, as well as the capability to metabolize a wide range of mono-, oligo-, and polysaccharides including galactose, arabinose, alginate, and pectin. On the basis of the morphological, physiological, and phylogenetic analyses, we propose to accommodate these isolates in a new genus Feramyces (derived from the Latin word for “wild” to reflect their isolation and apparent distribution in undomesticated herbivores), and a new species, F. austinii. The type species is Feramyces austinii strain F3a.

The anaerobic gut fungi (AGF) inhabit the rumen and alimentary tracts of multiple ruminant and nonruminant herbivores, belong to a distinct phylum-level lineage (Neocallimastigomycota), and play an important role in plant biomass degradation in many herbivores. As part of a wider effort to obtain AGF with high lignocellulolytic capacities, we isolated and characterized four different AGF strains from the feces of cattle and sheep. Microscopically, isolates produced monocentric thalli and monoflagellated zoospores. Phylogenetic analysis revealed that all isolates formed a monophyletic cluster with strong bootstrap support as a sister clade to the genus Orpinomyces and close to Neocallimastix, an unexpected result because these two genera of AGF form polyflagellated zoospores. Isolates displayed a smooth biofilm-like growth in liquid medium and formed small (0.5-1 mm) pinpoint circular colonies on agar roll tubes. Both endogenous and exogenous sporangia were observed with variable shapes and sizes. Zoospores were mainly spherical, with diameters ranging between 3.8 and 12.5 µm, and mostly a single flagellum. All strains exhibited similar substrate utilization patterns and comparable cellulolytic and xylanolytic activities. Similar ITS1 sequences falling within the same distinctive clade were found on GenBank, with all environmental samples obtained from diverse ruminant and pseudoruminant hosts from three continents, but not from any hindgut-fermenting hosts. Given the high level of sequence divergence between our strains and closest cultured representatives and their distinct microscopic/macroscopic features, we propose a new genus, Pecoramyces, from the name of the taxonomic infraorder Pecora ("horned ruminants" or "higher ruminants"; derived from the Latin word for horned livestock), and a new species, P. ruminantium (since occurrence seems to be specific to ruminant/pseudoruminant foregut, but not hindgut-fermenting mammals).

The genus Pantoea is a predominant member of host-associated microbiome. We here report on the genomic analysis of Pantoea eucrina strain Russ that was isolated from a trashcan at Oklahoma State University, Stillwater, OK. The draft genome of Pantoea eucrina strain Russ consists of 3,939,877bp of DNA with 3704 protein-coding genes and 134 RNA genes. This is the first report of a genome sequence of a member of Pantoea eucrina. Genomic analysis revealed metabolic versatility with genes involved in the metabolism and transport of all amino acids as well as glucose, fructose, mannose, xylose, arabinose and galactose, suggesting the organism is a versatile heterotroph. The genome also encodes an extensive secretory machinery including types I, II, III, IV, and Vb secretion systems, and several genes for pili production including the new usher/chaperone system (pfam 05,229). The implications of these systems for opportunistic pathogenesis are discussed.

Microbacterium oleivorans is a predominant member of hydrocarbon-contaminated environments. We here report on the genomic analysis of M. oleivorans strain Wellendorf that was isolated from an indoor door handle. The partial genome of M. oleivorans strain Wellendorf consists of 2,916,870bp of DNA with 2831 protein-coding genes and 49 RNA genes. The organism appears to be a versatile mesophilic heterotroph potentially capable of hydrolysis a suite of carbohydrates and amino acids. Genomic analysis revealed metabolic versatility with genes involved in the metabolism and transport of glucose, fructose, rhamnose, galactose, xylose, arabinose, alanine, aspartate, asparagine, glutamate, serine, glycine, threonine and cysteine. This is the first detailed analysis of a Microbacterium oleivorans genome.

We report on the characterization of a novel genomic assembly (ARYD3) recovered from formation water (17.6% salinity) and crude oil enrichment amended by isolated soy proteins (0.2%), and incubated for 100 days under anaerobic conditions at 50°C. Phylogenetic and phylogenomic analysis demonstrated that the ARYD3 is unaffiliated with all currently described bacterial phyla and candidate phyla, as evident by the low AAI (34.7%), shared gene content (19.4%), and 78.9% 16S rRNA gene sequence similarity to Halothiobacillus neapolitanus, its closest cultured relative. Genomic characterization predicts a slow-growing, non-spore forming, and non-motile Gram-negative rod. Adaptation to high salinity is potentially mediated by the production of the compatible solutes cyclic 2,3-diphosphoglycerate (cDPG), α-glucosylglycerate, as well as the uptake of glycine betaine. Metabolically, the genome encodes primarily aminolytic capabilities for a wide range of amino acids and peptides. Interestingly, evidence of propionate degradation to succinate via methyl-malonyl CoA was identified, suggesting possible capability for syntrophic propionate degradation. Analysis of ARYD3 global distribution patterns identified its occurrence in a very small fraction of Earth Microbiome Project datasets examined (318/27,068), where it consistently represented an extremely rare fraction (maximum 0.28%, average 0.004%) of the overall community. We propose the Candidatus name Mcinerneyibacterium aminivorans gen. nov, sp. nov. for ARYD3T, with the genome serving as the type material for the novel family Mcinerneyibacteriaceae fam. nov., order Mcinerneyibacteriales ord. nov., class Mcinerneyibacteria class nov., and phylum Mcinerneyibacteriota phyl. nov. The type material genome assembly is deposited in GenBank under accession number VSIX00000000.

Survival and growth of the anaerobic gut fungi (AGF; Neocallimastigomycota) in the herbivorous gut necessitate the possession of multiple abilities absent in other fungal lineages. We hypothesized that horizontal gene transfer (HGT) was instrumental in forging the evolution of AGF into a phylogenetically distinct gut-dwelling fungal lineage. The patterns of HGT were evaluated in the transcriptomes of 27 AGF strains, 22 of which were isolated and sequenced in this study, and 4 AGF genomes broadly covering the breadth of AGF diversity. We identified 277 distinct incidents of HGT in AGF transcriptomes, with subsequent gene duplication resulting in an HGT frequency of 2 to 3.5% in AGF genomes. The majority of HGT events were AGF specific (91.7%) and wide (70.8%), indicating their occurrence at early stages of AGF evolution. The acquired genes allowed AGF to expand their substrate utilization range, provided new venues for electron disposal, augmented their biosynthetic capabilities, and facilitated their adaptation to anaerobiosis. The majority of donors were anaerobic fermentative bacteria prevalent in the herbivorous gut. This study strongly indicates that HGT indispensably forged the evolution of AGF as a distinct fungal phylum and provides a unique example of the role of HGT in shaping the evolution of a high-rank taxonomic eukaryotic lineage. IMPORTANCE The anaerobic gut fungi (AGF) represent a distinct basal phylum lineage (Neocallimastigomycota) commonly encountered in the rumen and alimentary tracts of herbivores. Survival and growth of anaerobic gut fungi in these anaerobic, eutrophic, and prokaryote-dominated habitats necessitates the acquisition of several traits absent in other fungal lineages. We assess here the role of horizontal gene transfer as a relatively fast mechanism for trait acquisition by the Neocallimastigomycota postsequestration in the herbivorous gut. Analysis of 27 transcriptomes that represent the broad diversity of Neocallimastigomycota identified 277 distinct HGT events, with subsequent gene duplication resulting in an HGT frequency of 2 to 3.5% in AGF genomes. These HGT events have allowed AGF to survive in the herbivorous gut by expanding their substrate utilization range, augmenting their biosynthetic pathway, providing new routes for electron disposal by expanding fermentative capacities, and facilitating their adaptation to anaerobiosis. HGT in the AGF is also shown to be mainly a cross-kingdom affair, with the majority of donors belonging to the bacteria. This study represents a unique example of the role of HGT in shaping the evolution of a high-rank taxonomic eukaryotic lineage.

Arbuscular Mycorrhizal fungi (AMF, Glomeromycota) form obligate symbiotic associations with the roots of most terrestrial plants. Our understanding of the molecular mechanisms enabling AMF propagation and AMF-host interaction is currently incomplete. Analysis of AMF proteomes could yield important insights and generate hypotheses on the nature and mechanism of AMF-plant symbiosis. Here, we examined the extraradical mycelium proteomic profile of the arbuscular mycorrhizal fungus Rhizophagus irregularis grown on Ri T-DNA transformed Chicory roots in a root organ culture setting. Our analysis detected 529 different peptides that mapped to 474 translated proteins in the R. irregularis genome. R. irregularis proteome was characterized by a high proportion of proteins (9.9 % of total, 21.4 % of proteins with functional prediction) mediating a wide range of signal transduction processes, e.g. Rho1 and Bmh2, Ca-signaling (calmodulin, and Ca channel protein), mTOR signaling (MAP3K7, and MAPKAP1), and phosphatidate signaling (phospholipase D1/2) proteins, as well as members of the Ras signaling pathway. In addition, the proteome contained an unusually large proportion (53.6 %) of hypothetical proteins, the majority of which (85.8 %) were Glomeromycota-specific. Forty-eight proteins were predicted to be surface/membrane associated, including multiple hypothetical proteins of yet-unrecognized functions. However, no evidence for the overproduction of specific proteins, previously implicated in promoting soil health and aggregation was obtained. Finally, the comparison of R. irregularis proteome to previously published AMF proteomes identified a core set of pathways and processes involved in AMF growth. We conclude that R. irregularis growth on chicory roots requires the activation of a wide range of signal transduction pathways, the secretion of multiple novel hitherto unrecognized Glomeromycota-specific proteins, and the expression of a wide array of surface-membrane associated proteins for cross kingdom cell-to-cell communications.

The Wood-Ljungdahl (WL) pathway is an important component of the metabolic machinery in multiple anaerobic prokaryotes, including numerous yet-uncultured bacterial phyla. The pathway can operate in the reductive and oxidative directions, enabling a wide range of metabolic processes. Here, we present a detailed analysis of 14 newly acquired, previously analysed, and publicly available genomic assemblies belonging to the candidate phylum Bipolaricaulota (candidate division OP1, and candidatus Acetothermia), where the occurrence of WL pathway appears to be universal. In silico analysis of predicted metabolic capabilities indicates that the pathway enables homoacetogenic fermentation of sugars and amino acids in all three Bipolaricaulota orders (RBG-16-55-9, UBA7950 and Bipolaricaulales). In addition, members of RBG-16-55-9 appear to possess the additional capacity for syntrophic acetate oxidation using the WL pathway; as well as for respiratory growth using oxygen or nitrate. Anabolically, all UBA7950, and the majority of the Bipolaricaulales genomes possess the capacity for autotrophic growth using the WL pathway. Our results highlight the WL-enabled metabolic versatility in the Bipolaricaulota, emphasize the need for examining the WL pathway in context of the overall metabolic circuitry in uncultured taxa, and demonstrate the value of comparative genomic analysis for providing a detailed overview of metabolic potential in a target microbial lineage and its potential functional niche in an ecosystem.

116 pages, 1 figure, 4 tables, 3 appendices, This revision of the classification of eukaryotes follows that of Adl et al., 2012 [J. Euk. Microbiol. 59(5)] and retains an emphasis on protists. Changes since have improved the resolution of many nodes in phylogenetic analyses. For some clades even families are being clearly resolved. As we had predicted, environmental sampling in the intervening years has massively increased the genetic information at hand. Consequently, we have discovered novel clades, exciting new genera and uncovered a massive species level diversity beyond the morphological species descriptions. Several clades known from environmental samples only have now found their home. Sampling soils, deeper marine waters and the deep sea will continue to fill us with surprises. The main changes in this revision are the confirmation that eukaryotes form at least two domains, the loss of monophyly in the Excavata, robust support for the Haptista and Cryptista. We provide suggested primer sets for DNA sequences from environmental samples that are effective for each clade. We have provided a guide to trophic functional guilds in an appendix, to facilitate the interpretation of environmental samples, and a standardized taxonomic guide for East Asian users, Research support was provided as follows: SMA by NSERC 249889‐2007; DB by NERC NE/H009426/1 and NE/H000887/1; MWB by NSF 1456054; FB by a Fellowship from Science for Life Laboratory and VR/2017‐04563; PC by EU‐Horizon 2020 research and innovation program through the SponGES project 679849 [...]; IC by CSF 18‐18699S; BE by RCN TaxMArc 268286/GMR; LG by ANR HAPAR (ANR‐14‐CE02‐0007); VH MK JL by ERDF; MEYS with ERC 771592 CZ 1.05/1.1.00/02.0109 BIOCEV; SK by RSF 16‐14‐10302; MK by CSF GA18‐28103S; CEL by NSF 1541510 and NIH‐AI124092; EL by CAM: 2017‐T1/AMB‐5210; and by grant 2017‐T1/AMB‐5210 from the program >Atracción de talentos> from the Consejería de Educación, Juventud y Deporte, Comunidad de Madrid; JL by ERC CZ LL1601 and OPVVV 16_019/0000759; MP by NSF DEB‐1455611; DJR by the Beatriu de Pinós postdoctoral programme of the Government of Catalonia's Secretariat for Universities and Research of the Ministry of Economy and Knowledge; CLS by the intramural research program of the National Library of Medicine, National Institutes of Health; AS by RSF 17‐14‐01391 and RFBR 16‐04‐01454 NY by NSF DEB 1557102; VZ by RFBR 16‐34‐60102 mol‐a‐dk; UniEuk and EukRef by the Gordon and Betty Moore Foundation