Archaeal Populations Research Articles

Endolithic Microbial Habitats Hosted in Carbonate Nodules Currently Forming within Sediment at a High Methane Flux Site in the Sea of Japan

Concretionary carbonates in deep-sea methane seep fields are formed as a result of microbial methane degradation, called anaerobic oxidation of methane (AOM). Recently, active microorganisms, including anaerobic methanotrophic archaea, were discovered from methane seep-associated carbonate outcroppings on the seafloor. However sedimentary buried carbonate nodules are a hitherto unknown microbial habitat. In this study, we investigated the microbial community structures in two carbonate nodules collected from a high methane flux site in a gas hydrate field off the Oki islands in the Sea of Japan. The nodules were formed around sulfate-methane interfaces (SMI) corresponding to 0.7 and 2.2 m below the seafloor. Based on a geochemical analysis, light carbon isotopic values ranging from −54.91‰ to −37.32‰ were found from the nodules collected at the shallow SMI depth, which were attributed to the high contributions of AOM-induced carbonate precipitation. Signatures of methanotrophic archaeal populations within the sedimentary buried nodule were detected based on microbial community composition analyses and quantitative real-time PCR targeted 16S rRNA, and functional genes for AOM. These results suggest that the buried carbonate nodule currently develops AOM-related microbial communities, and grows depending on the continued AOM under high methane flux conditions.

Effect of substrate composition on the stability and microbial community of an anaerobic expanded granular sludge bed reactor treating printing solvent mixtures of ethanol and glycol ethers

The performance and microbial community analysis of an expanded granular sludge bed reactor (EGSB) treating wastewater polluted with mixtures of ethanol and glycol ethers –such as 1-ethoxy-2-propanol (E2P) and 1-methoxy-2-propanol (M2P)– were evaluated. The results showed good EGSB performance during start-up (100% of ethanol) in terms of global removal efficiency (RE > 95%). When glycol ethers were added, an initial adaptation period was observed of ~20 days. While the RE of M2P became complete, the RE of E2P reached only 65%. The proportion of glycol ethers was gradually increased and at the end of this study only a binary mixture of E2P and M2P was fed. In the last stage, the global RE dropped to 80% and a partial degranulation of the sludge bed occurred, revealing the importance of acetate-yielding compounds as co-substrate for maintaining the granular structure of the sludge. Microbial community analysis showed a relation between the substrate utilized in the reactor and the developed microbial populations, such as Methanosaeta, an acetate consumer, and Methanomethylovorans, a methanol consumer, which were the predominant microorganisms in the archaeal population; the predominant bacterium found was Geobacter, an ethanol consumer; this and Methanosaeta are synthropic partners.

Effects of particle size of ground alfalfa hay on caecal bacteria and archaea populations of rabbits.

This work was aimed to investigate the effects of the different particle size of ground alfalfa hay on caecal microbial and archeal communities of rabbits. One hundred-twenty New Zealand rabbits (950.3 ± 8.82 g) were allocated into four treatments, with five replicates in each treatment and six rabbits in each replicate. The particle sizes of the alfalfa meal in the four treatment diets were 2,500, 1,000, 100 and 10 µm respectively, while the other ingredients were ground through a 2.5 mm sieve. High-throughput sequencing technology was applied to examine the differences in bacteria and methanogenic archaea diversity in the caecum of the four treatment groups of rabbits. A total of 745,946 bacterial sequences (a mean of 31,081 ± 13,901 sequences per sample) and 539,227 archaeal sequences (a mean of 22,468 ± 2,443 sequences per sample) were recovered from twenty-four caecal samples, and were clustered into 9,953 and 2,246 OTUs respectively. A total of 26 bacterial phyla with 465 genera and three archaeal phyla with 10 genera were identified after taxonomic summarization. Bioinformatic analyses illustrated that Firmicutes (58.69% ∼ 68.50%) and Bacteroidetes (23.96% ∼ 36.05%) were the two most predominant bacterial phyla and Euryarchaeota (over 99.9%) was the most predominant archaeal phyla in the caecum of all rabbits. At genus level, as the particle size of alfalfa decreased from 2,500 to 10 µm, the relative abundances of Ruminococcaceae UCG-014 (P < 0.001) and Lactobacillus (P = 0.043) were increased and Ruminococcaceae UCG-005 (P = 0.012) was increased first and then decreased when the alfalfa particle size decreased, while Lachnospiraceae NK4A136 group (P = 0.016), Ruminococcaceae NK4A214 (P = 0.044), Christensenellaceae R-7 group (P = 0.019), Lachnospiraceae other (Family) (P = 0.011) and Ruminococcaceae UCG-013 (P = 0.021) were decreased. The relative abundance of Methanobrevibacter was increased from 62.48% to 90.40% (P < 0.001), whereas the relative abundance of Methanosphaera was reduced from 35.47% to 8.62% (P < 0.001). In conclusion, as the particle size of alfalfa meal decreased, both the bacterial and archaeal population in the caecum of rabbit experienced alterations, however archaea response earlier than bacteria to the decrease of alfalfa meal particle size.

Anaerobic digestion of mixed urban biowaste: The microbial community shift towards stability

Anaerobic digestion is applied worldwide to treat food waste (FW) with the aim of obtaining renewable bioenergy by exploiting the methane gas produced. However, there are several problems in practical applications, primarily due to system instability. Although exhaustive knowledge regarding anaerobic microbial community composition has been established, few studies have investigated long-term correlations between microbial consortia, operative conditions and feedstock characteristics. Here, microbial community shifts as a response to feedstock variations were investigated in long-term semi-continuous systems, which were evaluated by an in situ cell detection method and 16S rRNA gene amplicon sequencing. FW digestion showed progressive system instability caused by the inhibition of methanogens, which resulted in volatile fatty acid accumulation and process failure at the low organic loading rate (OLR). Conversely, by co-digesting FW with waste-activated sludge (WAS), a stable process with methane yields of up to 0.27 Nm3 kg−1VSfed for OLR = 1.7 gVS L−1d−1 was achieved. This stabilizing effect was not related to the buffering capacity of WAS, but to its capacity to avoid volatile fatty acid accumulation and falls in pH by overcoming methanogenic activity inhibition. WAS addition promoted the establishment of a stable and active archaeal population in anaerobic co-digestion (AcoD) reactors. The continuous supply of trace elements together with the seeding of microbial functional groups were the main drivers that positively affected process stability.

Microbial diversity and long-term geochemical trends in the euxinic zone of a marine, meromictic lake

Hypoxic and anoxic niches of meromictic lakes are important sites for studying the microbial ecology of conditions resembling ancient Earth. The expansion and increasing global distribution of such environments also means that information about them serves to understand future phenomena. In this study, a long-term chemical dataset (1996–2015) was explored together with seasonal (in 2015) information on the diversity and abundance of bacterial and archaeal communities residing in the chemocline, monimolimnion and surface sediment of the marine meromictic Rogoznica Lake. The results of quantitative PCR assays, and high-throughput sequencing, targeting 16S rRNA genes and transcripts, revealed a clear vertical structure of the microbial community with Gammaproteobacteria (Halochromatium) and cyanobacteria (Synechococcus spp.) dominating the chemocline, Deltaproteobacteria and Bacteroidetes dominating the monimolimnion, and significantly more abundant archaeal populations in the surface sediment, most of which affiliated to Nanoarchaeota. Seasonal changes in the community structure and abundance were not pronounced. Diversity in Rogoznica Lake was found to be high, presumably as a consequence of stable environmental conditions accompanied by high dissolved carbon and nutrient concentrations. Long-term data indicated that Rogoznica Lake exhibited climate changes that could alter its physico-chemical features and, consequently, induce structural and physiological changes within its microbial community.

A genomic view of the reef-building coral Porites lutea and its microbial symbionts.

Corals and the reef ecosystems that they support are in global decline due to increasing anthropogenic pressures such as climate change1. However, effective reef conservation strategies are hampered by a limited mechanistic understanding of coral biology and the functional roles of the diverse microbial communities that underpin coral health2,3. Here, we present an integrated genomic characterization of the coral species Porites lutea and its microbial partners. High-quality genomes were recovered from P. lutea, as well as a metagenome-assembled Cladocopium C15 (the dinoflagellate symbiont) and 52 bacterial and archaeal populations. Comparative genomic analysis revealed that many of the bacterial and archaeal genomes encode motifs that may be involved in maintaining association with the coral host and in supplying fixed carbon, B-vitamins and amino acids to their eukaryotic partners. Furthermore, mechanisms for ammonia, urea, nitrate, dimethylsulfoniopropionate and taurine transformation were identified that interlink members of the holobiont and may be important for nutrient acquisition and retention in oligotrophic waters. Our findings demonstrate the critical and diverse roles that microorganisms play within the coral holobiont and underscore the need to consider all of the components of the holobiont if we are to effectively inform reef conservation strategies.

Implication of Viral Infections for Greenhouse Gas Dynamics in Freshwater Wetlands: Challenges and Perspectives.

Viruses are non-living, acellular entities, and the most abundant biological agents on earth. They are widely acknowledged as having the capacity to influence global biogeochemical cycles by infecting the bacterial and archaeal populations that regulate carbon and nutrient turnover. Evidence suggests that the majority of viruses in wetlands are bacteriophages, but despite their importance, studies on how viruses control the prokaryotic community and the concomitant impacts on ecosystem function (such as carbon cycling and greenhouse gas flux) in wetlands are rare. Here we investigate virus-prokaryote interactions in freshwater wetland ecosystems in the context of their potential influence on biogeochemical cycling. Specifically, we (1) synthesize existing literature to establish current understanding of virus-prokaryote interactions, focusing on the implications for wetland greenhouse gas dynamics and (2) identify future research priorities. Viral dynamics in freshwater wetlands have received much less attention compared to those in marine ecosystems. However, based on our literature review, within the last 10 years, viral ecology studies on freshwater wetlands have increased twofold. Despite this increase in literature, the potential implication of viral infections on greenhouse gas emission dynamics is still a knowledge gap. We hypothesize that the rate of greenhouse gas emissions and the pool of sequestered carbon could be strongly linked to the type and rate of viral infection. Viral replication mechanism choice will consequently influence the microbial efficiency of organic matter assimilation and thus the ultimate fate of carbon as a greenhouse gas or stored in soils.

Biological pretreatment with Trametes versicolor to enhance methane production from lignocellulosic biomass: A metagenomic approach

The presence of poorly biodegradable components in lignocellulosic biomass limits the methane recovery in anaerobic digesters. The main reason to go for aerobic pretreatment before anaerobic digestion (AD) is to enable enzymatic cleavage of the aromatic rings in lignin by oxygen since it cannot be efficiently degraded under anaerobic conditions. In this study, the advantage of highly-cellulolytic white-rot fungi Trametes versicolor was taken by aerobic pretreatment prior to anaerobic co-digestion of cow manure and selected cereal crop materials (i.e. wheat, rye, barley, triticale) harvested at different stages. Fungal pretreatment improved the methane yield by 10%–18% and cellulose degradation up to 80%. Furthermore, higher volatile fatty acid (VFA) speciation was found in the anaerobic digesters upon fungal pretreatment. 16S rRNA gene amplicon sequencing revealed a more diverse microbial community in the fungal-pretreated anaerobic digesters. Generally, typically-detected bacterial species dominated the digesters; except that Synergistetes was only enriched in the fungal-pretreated digesters. Although Methanosarcianease was the predominant methanogenic archaea, a more diverse methanogenic population was identified in the fungal-pretreated digesters in which Methanobacteriaceaa and Methanomibrobiaceae also took role during biomethanation. Comparatively more unique microbiome of biogas reactors upon fungal pretreatment synergistically affected VFA production, cellulose degradation and eventually methane yield in an affirmative way. Considering the functional importance of bacterial and methanogenic archaeal populations, elevated knowledge of the microbial structures is essential for minimizing process failures and for creating strategies for process optimization of lignocellulose based-AD.

The N-fixing legume Periandra mediterranea constrains the invasion of an exotic grass (Melinis minutiflora P. Beauv) by altering soil N cycling

Melinis minutiflora is an invasive species that threatens the biodiversity of the endemic vegetation of the campo rupestre biome in Brazil, displacing the native vegetation and favouring fire spread. As M. minutiflora invasion has been associated with a high nitrogen (N) demand, we assessed changes in N cycle under four treatments: two treatments with contrasting invasion levels (above and below 50%) and two un-invaded control treatments with native vegetation, in the presence or absence of the leguminous species Periandra mediterranea. This latter species was considered to be the main N source in this site due to its ability to fix N2 in association with Bradyrhizobia species. Soil proteolytic activity was high in treatments with P. mediterranea and in those severely invaded, but not in the first steps of invasion. While ammonium was the N-chemical species dominant in plots with native species, including P.mediterranea, soil nitrate prevailed only in fully invaded plots due to the stimulation of the nitrifying bacterial (AOB) and archaeal (AOA) populations carrying the amoA gene. However, in the presence of P. mediterranea, either in the beginning of the invasion or in uninvaded plots, we observed an inhibition of the nitrifying microbial populations and nitrate formation, suggesting that this is a biotic resistance strategy elicited by P. mediterranea to compete with M. minutiflora. Therefore, the inhibition of proteolytic activity and the nitrification process were the strategies elicited by P.mediterranea to constrain M.munitiflora invasion.

Determination of microbial numbers in anaerobically digested biofertilisers

ABSTRACT This study aimed to quantity total numbers of bacteria, fungi and archaea in different types of commercial liquid anaerobic digestates, and to identify common patterns in their microbial numbers post-digestion and possible implications of their use as biofertiliser. Relationships between microbial numbers and physical–chemical traits of the digestates were also investigated. Quantification was performed using culturable and molecular (quantitative PCR) approaches. Bacterial and fungal CFUs ranged up to five orders of magnitude (105–1010; 0–105 g−1 DW, respectively) between different types of anaerobic digestates. Bacterial, archaeal and fungal gene copy numbers (GCN) varied by two orders of magnitude (108–1010; 107–109; 104–106 g−1 DW, respectively) between digestates. All microbial variables analysed showed significant differences between the different types of anaerobic digestate investigated (p < 0.05). Culturable microbial numbers for fungi (6.43 × 104 CFU g−1 DW) were much lower than for bacteria (2.23 × 109 CFU g−1 DW). Gene copy numbers were highest for bacteria (16S) (1.09 × 1010 g−1 DW), followed by archaea (16S) (5.87 × 108 g−1 DW), and fungi (18S) (1.77 × 106 g−1 DW). Liquid anaerobic digestates were predominantly dominated by bacteria, followed by archaeal and fungal populations. At 50% similarity level, the microbial profiles of the eleven anaerobic digestates tested separated into just two groups, indicating a broad relative degree of similarity in terms of microbial numbers. Higher bacterial (16S) GCN was associated with low OM and C/N ratio in digestates.

Effect of different co-treatments of waste activated sludge on biogas production and shaping microbial community in subsequent anaerobic digestion

Different pretreatment approaches enhance hydrolysis and biogas production of waste activated sludge (WAS). However, the best WAS pretreatment method may vary for different purposes. The aim of this study was to compare different combined treatments using potassium ferrate, alkali, and ultrasonication treatment in terms of hydrolysis, biogas production, and microbial community structure during subsequent anaerobic digestion. The soluble chemical oxygen demand (SCOD) increased sharply during sludge pretreatment. Co-treatment with potassium ferrate and ultrasonication (PF + ULT) provided the highest concentration of SCOD (10,206 mg/L) and volatile fatty acids (VFAs) (9160 mg/L). Maximum cumulative methane production, 752.6 mL during 21 days of subsequent anaerobic digestion, was achieved for alkali and ultrasonication (ALK + ULT) co-treated sludge, a 39.3% improvement compared with raw sludge. Our results indicate that PF + ULT and ALK + ULT co-treatments are suitable for methanogenesis in subsequent anaerobic digestion. Co-treatment typically improves biogas production and sludge hydrolysis relative to a single pretreatment. Illumina sequencing of 16S rRNA genes showed that sludge subjected to ALK pretreatment and ALK + ULT co-treatment contained the same predominant populations, and the community structure of PF was similar to PF + ULT. Principal coordinate analysis (PCoA) based on weighted UniFrac distance indicated differences in the microbial communities of the pretreated and raw sludges. The predominant archaeal populations were Methanomassiliicoccus and Methanobacterium in the ALK + ULT and ALK pretreated sludges, and the majority were Methanomassiliicoccus (76.82%) in PF + ULT co-treated sludge, indicating that different pretreatment methods could substantially shape archaeal communities of pretreated sludge. These results reveal that sludge pretreatment not only impacts sludge hydrolysis and biogas production, but also significantly influences the microbial community in subsequent anaerobic digestion.

Depth-related differences in archaeal populations impact the isoprenoid tetraether lipid composition of the Mediterranean Sea water column

Thaumarchaeota are one of the most abundant groups of Archaea in the marine water column. Their membrane consists of isoprenoid glycerol dibiphytanyl glycerol tetraethers (GDGTs) which are applied in the widely used TEX86 proxy to reconstruct past sea surface temperatures (SSTs). However, in some specific marine systems, such as the Mediterranean Sea, core-top TEX86-derived temperatures do not seem to reflect annual mean SSTs. This has been attributed to contributions of deep-water dwelling Thaumarchaeota. Here, we investigate the potential causes of this bias by studying both the archaeal diversity as well as the intact polar lipid (IPL) GDGT composition in the Mediterranean water column by a combined 16S rRNA gene amplicon sequencing and a lipidomic approach on suspended particulate matter (SPM) at different water depths. The archaeal distribution showed a dominance of archaea other than Thaumarchaeota, i.e. Marine Euryarchaeota group II and III in the upper epipelagic waters (0–100 m deep), while Thaumarchaeota (Marine group I; MGI) dominated the subsurface and the deeper waters. This shift in the archaeal community composition coincided with a decrease in IPL GDGT-0 and increase of IPL crenarchaeol. The ratio of GDGT-2/GDGT-3 increased with water depth, but values were lower than observed in deep marine waters of some other regions. The increase of the GDGT-2/GDGT-3 ratio coincided with the high relative abundance of deep-water MGI, which may be linked to the high temperature and salinity found in specific water masses of the Mediterranean Sea. We conclude that these particularities of the Mediterranean Sea are responsible for the overestimated SST based on TEX86.

Functional relationship between ammonia-oxidizing bacteria and ammonia-oxidizing archaea populations in the secondary treatment system of a full-scale municipal wastewater treatment plant

The abundance of ammonia-oxidizing bacteria and archaea and their amoA genes from the aerobic activated sludge tanks, recycled sludge and anaerobic digesters of a full-scale wastewater treatment plant (WWTP) was determined. Polymerase chain reaction and denaturing gradient gel electrophoresis were used to generate diversity profiles, which showed that each population had a consistent profile although the abundance of individual members varied. In the aerobic tanks, the ammonia-oxidizing bacterial (AOB) population was more than 350 times more abundant than the ammonia-oxidizing archaeal (AOA) population, however in the digesters, the AOA population was more than 10 times more abundant. Measuring the activity of the amoA gene expression of the two populations using RT-PCR also showed that the AOA amoA gene was more active in the digesters than in the activated sludge tanks. Using batch reactors and ddPCR, amoA activity could be measured and it was found that when the AOB amoA activity was inhibited in the anoxic reactors, the expression of the AOA amoA gene increased fourfold. This suggests that these two populations may have a cooperative relationship for the oxidation of ammonia.

Microbial population dynamics in temperature‐phased anaerobic digestion of municipal wastewater sludge

AbstractBACKGROUNDThe present study compared the performance and capability of thermophilic–mesophilic, temperature‐phased anaerobic digestion (TPAD) of wastewater sludge at 45 and 55 °C in terms of rate of hydrolysis and methanogenesis at two point interval times – December and January – with operational conditions kept constant (total solid content 6.5%, solids retention time 12.5 days).RESULTSThe reduction of volatile solids (VS) was 77% at 45 °C TPAD‐1 and 72% at 55 °C TPAD‐II. The accumulation of ammonia and volatile fatty acids (VFAs; propionic acid) were observed to be below the inhibitory range (&gt;3000 mg L−1), whereas the methane (CH4) production (45 °C, 3.55 ± 0.47 L CH4 L−1 day−1; 35 °C, 1.44 ± 0.12 L CH4 L−1 day−1) remained considerably higher in TPAD‐I than TPAD‐II. Furthermore, the TPAD‐II system suffered from certain degree of instability due to high level of total VFAs (6087 ± 1578 mg L−1), low buffering capacity, increased level of total NH3 (2982 ± 219 mg L−1) and free NH3 (246 ± 25 mg L−1), and relatively reduced CH4 production (1.69 ± 0.1 L L−1 day). The bacterial and archaeal population of the TPADs investigated by 454 pyrosequencing and Illumina sequencing showed, in both TPAD systems, a bacterial community dominated by Firmicutes, followed by Bacteriodetes, Proteobacteria, Synergistetes and Actiniobacteria. The archaeal community was dominated by Methanimicrobia (74–84% Methanosarcina) and Methanobacteria (15–27% Methanobacterium). A progression from genus Clostridium to Coprothermobacter and Tepidanaerobacter, and Methanocarcina to Methanothermobacter and Methanobacterium was observed in TPAD‐II.CONCLUSIONSThis study proved the processes driving the dynamics of key microbial population and its correlation with hydrolytic functionality of TPAD systems. © 2019 Society of Chemical Industry

Simultaneous biomethanisation of endogenous and imported CO2 in organically loaded anaerobic digesters

In-situ biomethanisation reduces the CO2 in biogas to CH4 via direct H2 injection into an anaerobic digester, but volumetric methane production (VMP) is limited by organic loading. Ex-situ biomethanisation, where gaseous substrates are fed to pure or mixed cultures of hydrogenotrophic methanogens, offers higher VMP but requires an additional reactor and supply of essential nutrients. This work combined the two approaches in a novel hybrid application achieving simultaneous in-situ and ex-situ biomethanisation within an organically-loaded anaerobic digester receiving supplementary biogas. Conventional stirred-tank digesters were first acclimated to H2 addition, increasing biogas methane content from 50% to 95% and VMP from 0.86 to 1.51 L L−1 day−1 at a moderate loading rate of 3 g organic chemical oxygen demand per L per day (g CODorg L−1 day−1). Externally-produced biogas was then added to demonstrate simultaneous biomethanisation of endogenous and imported CO2. This further increased VMP to 2.76 L L−1 day−1 without affecting organic substrate degradation. In-situ CO2 reduction can alter digester pH by reducing bicarbonate buffering: the combined process operated stably at around pH 8.0 with 3–5% CO2 in the headspace. Microbial community analysis indicated the process was mediated by bacterial syntrophic acetate oxidation and highly enriched hydrogenotrophic methanogenic archaea (up to 97% of the archaeal population). This approach presents the opportunity to retrofit a single digester for H2 injection to convert and upgrade biogas from several others, minimising capital and operating costs by utilising both existing infrastructure and waste-derived feedstock nutrients for simultaneous biogas upgrading and power-to-methane.

Semicontinuous anaerobic digestion of protease pretreated Chlorella biomass for volatile fatty acids production

AbstractBACKGROUNDAnaerobic digestion (AD) could be designed as a source of volatile fatty acids (VFAs). However, acidogenesis optimization for novel substrates such as Chlorella vulgaris biomass needs to be investigated considering parameters such as temperature (T), organic loading rate (OLR), hydraulic retention time (HRT) and the adaptation of the sludge to temperature and substrate.RESULTSThe best organic matter conversion into VFAs (CODVFA/CODin; COD, chemical oxygen demand) was achieved with HRT = 8 d, adapted anaerobic sludge (AAS) and OLR = 1.5 gCOD L−1 d−1 (CODVFA/CODin = 39.8 ± 1.0% and productivity VFA = 0.5 ± 0.1 g L−1 d−1). Acetic and butyric acids represented 50% of the total VFAs. The microbiota related to acidogenesis and acetogenesis (Firmicutes 55% of the operational taxonomic units in R5) and the low archaeal population resulted in VFA accumulation at 25 °C.CONCLUSIONSThe use of low HRT and temperatures promoted VFAs production especially when AAS was employed. Microbial communities were strikingly different to the ones often found in AD targeted at biogas production. The relevance of the Firmicutes phylum (≤55% in R3 and R5) and euryarchaeota absence at 25 °C contributed to VFA accumulation. The use of AAS reported an increase in Actinobacteria species. © 2019 Society of Chemical Industry

CRISPR Spacers Indicate Preferential Matching of Specific Virioplankton Genes.

Viral infection exerts selection pressure on marine microbes, as virus-induced cell lysis causes 20 to 50% of cell mortality, resulting in fluxes of biomass into oceanic dissolved organic matter. Archaeal and bacterial populations can defend against viral infection using the clustered regularly interspaced short palindromic repeat (CRISPR)-associated (Cas) system, which relies on specific matching between a spacer sequence and a viral gene. If a CRISPR spacer match to any gene within a viral genome is equally effective in preventing lysis, no viral genes should be preferentially matched by CRISPR spacers. However, if there are differences in effectiveness, certain viral genes may demonstrate a greater frequency of CRISPR spacer matches. Indeed, homology search analyses of bacterioplankton CRISPR spacer sequences against virioplankton sequences revealed preferential matching of replication proteins, nucleic acid binding proteins, and viral structural proteins. Positive selection pressure for effective viral defense is one parsimonious explanation for these observations. CRISPR spacers from virioplankton metagenomes preferentially matched methyltransferase and phage integrase genes within virioplankton sequences. These virioplankton CRISPR spacers may assist infected host cells in defending against competing phage. Analyses also revealed that half of the spacer-matched viral genes were unknown, some genes matched several spacers, and some spacers matched multiple genes, a many-to-many relationship. Thus, CRISPR spacer matching may be an evolutionary algorithm, agnostically identifying those genes under stringent selection pressure for sustaining viral infection and lysis. Investigating this subset of viral genes could reveal those genetic mechanisms essential to virus-host interactions and provide new technologies for optimizing CRISPR defense in beneficial microbes.IMPORTANCE The CRISPR-Cas system is one means by which bacterial and archaeal populations defend against viral infection which causes 20 to 50% of cell mortality in the ocean. We tested the hypothesis that certain viral genes are preferentially targeted for the initial attack of the CRISPR-Cas system on a viral genome. Using CASC, a pipeline for CRISPR spacer discovery, and metagenome data from oceanic microbes and viruses, we found a clear subset of viral genes with high match frequencies to CRISPR spacers. Moreover, we observed a many-to-many relationship of spacers and viral genes. These high-match viral genes were involved in nucleotide metabolism, DNA methylation, and viral structure. It is possible that CRISPR spacer matching is an evolutionary algorithm pointing to those viral genes most important to sustaining infection and lysis. Studying these genes may advance the understanding of virus-host interactions in nature and provide new technologies for leveraging CRISPR-Cas systems in beneficial microbes.

Co-occurring genomic capacity for anaerobic methane and dissimilatory sulfur metabolisms discovered in the Korarchaeota.

Phylogenetic and geological evidence supports the hypothesis that life on Earth originated in thermal environments and conserved energy through methanogenesis or sulfur reduction. Here we describe two populations of the deeply rooted archaeal phylum Korarchaeota, which were retrieved from the metagenome of a circumneutral, suboxic hot spring that contains high levels of sulfate, sulfide, methane, hydrogen and carbon dioxide. One population is closely related to 'Candidatus Korarchaeum cryptofilum OPF8', while the more abundant korarchaeote, 'Candidatus Methanodesulfokores washburnensis', contains genes that are necessary for anaerobic methane and dissimilatory sulfur metabolisms. Phylogenetic and ancestral reconstruction analyses suggest that methane metabolism originated in the Korarchaeota, whereas genes for dissimilatory sulfite reduction were horizontally transferred to the Korarchaeota from the Firmicutes. Interactions among enzymes involved in both metabolisms could facilitate exergonic, sulfite-dependent, anaerobic oxidation of methane to methanol; alternatively, 'Ca. M. washburnensis' could conduct methanogenesis and sulfur reduction independently. Metabolic reconstruction suggests that 'Ca. M. washburnensis' is a mixotroph, capable of amino acid uptake, assimilation of methane-derived carbon and/or CO2 fixation by archaeal type III-b RuBisCO for scavenging ribose carbon. Our findings link anaerobic methane metabolism and dissimilatory sulfur reduction within a single deeply rooted archaeal population and have implications for the evolution of these traits throughout the Archaea.

Archaea dominate the microbial community in an ecosystem with low-to-moderate temperature and extreme acidity

BackgroundThe current view suggests that in low-temperature acidic environments, archaea are significantly less abundant than bacteria. Thus, this study of the microbiome of Parys Mountain (Anglesey, UK) sheds light on the generality of this current assumption. Parys Mountain is a historically important copper mine and its acid mine drainage (AMD) water streams are characterised by constant moderate temperatures (8–18 °C), extremely low pH (1.7) and high concentrations of soluble iron and other metal cations.ResultsMetagenomic and SSU rRNA amplicon sequencing of DNA from Parys Mountain revealed a significant proportion of archaea affiliated with Euryarchaeota, which accounted for ca. 67% of the community. Within this phylum, potentially new clades of Thermoplasmata were overrepresented (58%), with the most predominant group being “E-plasma”, alongside low-abundant Cuniculiplasmataceae, ‘Ca. Micrarchaeota’ and ‘Terrestrial Miscellaneous Euryarchaeal Group’ (TMEG) archaea, which were phylogenetically close to Methanomassilicoccales and clustered with counterparts from acidic/moderately acidic settings. In the sediment, archaea and Thermoplasmata contributed the highest numbers in V3-V4 amplicon reads, in contrast with the water body community, where Proteobacteria, Nitrospirae, Acidobacteria and Actinobacteria outnumbered archaea. Cultivation efforts revealed the abundance of archaeal sequences closely related to Cuniculiplasma divulgatum in an enrichment culture established from the filterable fraction of the water sample. Enrichment cultures with unfiltered samples showed the presence of Ferrimicrobium acidiphilum, C. divulgatum, ‘Ca. Mancarchaeum acidiphilum Mia14’, ‘Ca. Micrarchaeota’-related and diverse minor (< 2%) bacterial metagenomic reads.ConclusionContrary to expectation, our study showed a high abundance of archaea in this extremely acidic mine-impacted environment. Further, archaeal populations were dominated by one particular group, suggesting that they are functionally important. The prevalence of archaea over bacteria in these microbiomes and their spatial distribution patterns represents a novel and important advance in our understanding of acidophile ecology. We also demonstrated a procedure for the specific enrichment of cell wall-deficient members of the archaeal component of this community, although the large fraction of archaeal taxa remained unculturable. Lastly, we identified a separate clustering of globally occurring acidophilic members of TMEG that collectively belong to a distinct order within Thermoplasmata with yet unclear functional roles in the ecosystem.

Influence of different biological control agents and compost on total and nitrification-driven microbial communities at rhizosphere and soil level in a lettuce - Fusarium oxysporum f. sp. lactucae pathosystem.

The response of rhizosphere and bulk soil indigenous microbial communities focusing on nitrifiers was evaluated after the application of different biological control agents (BCAs; Bacillus, Trichoderma, Pseudomonas) and compost in controlling lettuce Fusarium wilt. Experiments were conducted 'insitu' over two lettuce cropping seasons. Total fungal, bacterial and archaeal populations and the nitrifiers were analysed using quantitative polymerase chain reaction method. The pathogen, Fusarium oxysporum forma specialis lactucae (FOL), Bacillus, Trichoderma and Pseudomonas and three antifungal genes (chiA, 2,4-diacetylphloroglucinol - phlD and HCN synthase - hcnAB genes) were also assessed. Quantitative data were corroborated with disease severity (DS), potential nitrification activity and soil chemical parameters. The application of BCAs and compost resulted in the disease reduction by as much as 69%, confirmed by significant negative correlations between Bacillus subtilis, Trichoderma and Pseudomonas sp. abundances and DS. The FOL presence in the untreated control resulted in the nitrifiers niche differentiation. The used treatments were efficient against Fusarium wilt and did not influence negatively the nontarget microbial communities. The use of BCAs and compost appears as an effective and safe strategy to implement sustainable agricultural practices.