Alexandre B. de Menezes

Microbiome analysis of dairy cows fed pasture or total mixed ration diets.

Understanding rumen microbial ecology is essential for the development of feed systems designed to improve livestock productivity, health and for methane mitigation strategies from cattle. Although rumen microbial communities have been studied previously, few studies have applied next-generation sequencing technologies to that ecosystem. The aim of this study was to characterize changes in microbial community structure arising from feeding dairy cows two widely used diets: pasture and total mixed ration (TMR). Bacterial, archaeal and protozoal communities were characterized by terminal restriction fragment length polymorphism of the amplified SSU rRNA gene and statistical analysis showed that bacterial and archaeal communities were significantly affected by diet, whereas no effect was observed for the protozoal community. Deep amplicon sequencing of the 16S rRNA gene revealed significant differences in the bacterial communities between the diets and between rumen solid and liquid content. At the family level, some important groups of rumen bacteria were clearly associated with specific diets, including the higher abundance of the Fibrobacteraceae in TMR solid samples and members of the propionate-producing Veillonelaceae in pasture samples. This study will be relevant to the study of rumen microbial ecology and livestock feed management.

Comparative metatranscriptomics reveals widespread community responses during phenanthrene degradation in soil

Soil microbial community response to phenanthrene was evaluated by metatranscriptomics. A marked increase in transcripts involved in aromatic compound metabolism, respiration and stress responses, and concurrent decreases in virulence, carbohydrate, DNA metabolism and phosphorus metabolism transcripts was revealed. Phenanthrene addition led to a 1.8-fold to 33-fold increase in the abundance of dioxygenase, stress response and detoxification transcripts, whereas those of general metabolism were little affected. Heavy metal P-type ATPases and thioredoxin transcripts were more abundant in the phenanthrene-amended soil, and this is the first time these proteins have been associated with polycyclic aromatic hydrocarbon (PAH) stress in microorganisms. Annotation with custom databases constructed with bacterial or fungal PAH metabolism protein sequences showed that increases in PAH-degradatory gene expression occurred for all gene groups investigated. Taxonomic determination of mRNA transcripts showed widespread changes in the bacteria, archaea and fungi, and the actinobacteria were responsible for most of the de novo expression of transcripts associated with dioxygenases, stress response and detoxification genes. This is the first report of an experimental metatranscriptomic study detailing microbial community responses to a pollutant in soil, and offers information on novel in situ effects of PAHs on soil microbes that can be explored further.

Network analysis reveals that bacteria and fungi form modules that correlate independently with soil parameters

Network and multivariate statistical analyses were performed to determine interactions between bacterial and fungal community terminal restriction length polymorphisms as well as soil properties in paired woodland and pasture sites. Canonical correspondence analysis (CCA) revealed that shifts in woodland community composition correlated with soil dissolved organic carbon, while changes in pasture community composition correlated with moisture, nitrogen and phosphorus. Weighted correlation network analysis detected two distinct microbial modules per land use. Bacterial and fungal ribotypes did not group separately, rather all modules comprised of both bacterial and fungal ribotypes. Woodland modules had a similar fungal : bacterial ribotype ratio, while in the pasture, one module was fungal dominated. There was no correspondence between pasture and woodland modules in their ribotype composition. The modules had different relationships to soil variables, and these contrasts were not detected without the use of network analysis. This study demonstrated that fungi and bacteria, components of the soil microbial communities usually treated as separate functional groups as in a CCA approach, were co-correlated and formed distinct associations in these adjacent habitats. Understanding these distinct modular associations may shed more light on their niche space in the soil environment, and allow a more realistic description of soil microbial ecology and function.

Cellulose Degradation by Micromonosporas Recovered from Freshwater Lakes and Classification of These Actinomycetes by DNA Gyrase B Gene Sequencing

ABSTRACT A number of Micromonospora strains isolated from the water column, sediment, and cellulose baits placed in freshwater lakes were shown to be able to degrade cellulose in lake water without any addition of nutrients. A selective isolation method was also developed to demonstrate that CFU arose from both spores and hyphae that inhabit the lake environment. Gyrase B gene sequencing performed on the isolates identified a number of new centers of variation within Micromonospora, but the most actively cellulolytic strains were recovered in a single cluster that equated with the type species of the genus, M. chalcea.

Molecular Biological Detection and Quantification of Novel Fibrobacter Populations in Freshwater Lakes

ABSTRACT PCR and quantitative PCR (qPCR) primers targeting the 16S rRNA gene were used to detect and quantify members of the genus Fibrobacter in lake water, sediment and colonized cotton taken from two freshwater lakes. Phylogenetic analysis identified two groups of sequences; those clustered with Fibrobacter succinogenes, the type species, and a defined cluster of clones loosely associated with several Fibrobacter sequences observed previously in clone libraries from freshwater environments. 16S rRNA gene sequences recovered in the same way from soil samples and ovine feces in the surrounding land were all F. succinogenes and did not include any from this group of the “freshwater” Fibrobacteres. In all cases, nested PCR was required to detect Fibrobacter 16S rRNA genes, and qPCR analysis of reverse transcribed bacterial community RNA confirmed their very low relative abundance on colonized cotton baits in the water column (at 0, 3, 7, 11, and 13 m) and on the sediment surface (<0.02% of total bacterial rRNA). However, in Esthwaite Water sediment itself, the relative abundance of fibrobacters was 2 orders of magnitude higher (ca. 1% of total bacterial rRNA). The presence of fibrobacters, including the cellulolytic rumen species F. succinogenes, on colonized cellulose samples and in lake sediment suggests that these organisms may contribute to the primary degradation of plant and algal biomass in freshwater lake ecosystems.

Importance of Micromonospora spp. as Colonizers of Cellulose in Freshwater Lakes as Demonstrated by Quantitative Reverse Transcriptase PCR of 16S rRNA

ABSTRACT The relative abundance of micromonosporas in the bacterial communities inhabiting cellulose baits, water columns, and sediments of two freshwater lakes was determined by quantitative PCR (qPCR) of reverse-transcribed 16S rRNA. Micromonospora spp. were shown to be significant members of the active bacterial population colonizing cellulosic substrates in the lake sediment, and their increased prevalence with greater depth was confirmed by enumeration of CFU.

C/N ratio drives soil actinobacterial cellobiohydrolase gene diversity.

ABSTRACT Cellulose accounts for approximately half of photosynthesis-fixed carbon; however, the ecology of its degradation in soil is still relatively poorly understood. The role of actinobacteria in cellulose degradation has not been extensively investigated despite their abundance in soil and known cellulose degradation capability. Here, the diversity and abundance of the actinobacterial glycoside hydrolase family 48 (cellobiohydrolase) gene in soils from three paired pasture-woodland sites were determined by using terminal restriction fragment length polymorphism (T-RFLP) analysis and clone libraries with gene-specific primers. For comparison, the diversity and abundance of general bacteria and fungi were also assessed. Phylogenetic analysis of the nucleotide sequences of 80 clones revealed significant new diversity of actinobacterial GH48 genes, and analysis of translated protein sequences showed that these enzymes are likely to represent functional cellobiohydrolases. The soil C/N ratio was the primary environmental driver of GH48 community compositions across sites and land uses, demonstrating the importance of substrate quality in their ecology. Furthermore, mid-infrared (MIR) spectrometry-predicted humic organic carbon was distinctly more important to GH48 diversity than to total bacterial and fungal diversity. This suggests a link between the actinobacterial GH48 community and soil organic carbon dynamics and highlights the potential importance of actinobacteria in the terrestrial carbon cycle.

Fifty important research questions in microbial ecology

Abstract Microbial ecology provides insights into the ecological and evolutionary dynamics of microbial communities underpinning every ecosystem on Earth. Microbial communities can now be investigated in unprecedented detail, although there is still a wealth of open questions to be tackled. Here we identify 50 research questions of fundamental importance to the science or application of microbial ecology, with the intention of summarising the field and bringing focus to new research avenues. Questions are categorised into seven themes: host‐microbiome interactions; health and infectious diseases; human health and food security; microbial ecology in a changing world; environmental processes; functional diversity; and evolutionary processes. Many questions recognise that microbes provide an extraordinary array of functional diversity that can be harnessed to solve real‐world problems. Our limited knowledge of spatial and temporal variation in microbial diversity and function is also reflected, as is the need to integrate micro‐ and macro‐ecological concepts, and knowledge derived from studies with humans and other diverse organisms. Although not exhaustive, the questions presented are intended to stimulate discussion and provide focus for researchers, funders and policy makers, informing the future research agenda in microbial ecology.

Linking fungal–bacterial co-occurrences to soil ecosystem function

Fungi and bacteria are major players in soil biogeochemical cycles, however, most studies linking soil processes to microbial function ignore the potential role of interactions between these groups. A small number of studies have used correlation network analyses to investigate fungal-bacterial co-occurrences in soil, and revealed differences, as well as overlaps, in the ecosystem roles of these groups. These results contradict the view that fungi and bacteria are two distinct functional groups which can be studied in isolation. A more comprehensive understanding of the interplay between soil properties, biogeochemical cycles and the interactions between fungi and bacteria will be an important step towards improving the prediction and management of soil ecosystem services.

The soil microbiome at the Gi-FACE experiment responds to a moisture gradient but not to CO2 enrichment.

The soil bacterial community at the Giessen free-air CO2 enrichment (Gi-FACE) experiment was analysed by tag sequencing of the 16S rRNA gene. No substantial effects of CO2 levels on bacterial community composition were detected. However, the soil moisture gradient at Gi-FACE had a significant effect on bacterial community composition. Different groups within the Acidobacteria and Verrucomicrobia phyla were affected differently by soil moisture content. These results suggest that modest increases in atmospheric CO2 may cause only minor changes in soil bacterial community composition and indicate that the functional responses of the soil community to CO2 enrichment previously reported at Gi-FACE are due to factors other than changes in bacterial community composition. The effects of the moisture gradient revealed new information about the relationships between poorly known Acidobacteria and Verrucomicrobia and soil moisture content. This study contrasts with the relatively small number of other temperate grassland free-air CO2 enrichment microbiome studies in the use of moderate CO2 enrichment and the resulting minor changes in the soil microbiome. Thus, it will facilitate the development of further climate change mitigation studies. In addition, the moisture gradient found at Gi-FACE contributes new knowledge in soil microbial ecology, particularly regarding the abundance and moisture relationships of the soil Verrucomicrobia.