Cophenetic correlation analysis as a strategy to select phylogenetically informative proteins: an example from the fungal kingdom

<p>Abstract</p> <p>Background</p> <p>The construction of robust and well resolved phylogenetic trees is important for our understanding of many, if not all biological processes, including speciation and origin of higher taxa, genome evolution, metabolic diversification, multicellularity, origin of life styles, pathogenicity and so on. Many older phylogenies were not well supported due to insufficient phylogenetic signal present in the single or few genes used in phylogenetic reconstructions. Importantly, single gene phylogenies were not always found to be congruent. The phylogenetic signal may, therefore, be increased by enlarging the number of genes included in phylogenetic studies. Unfortunately, concatenation of many genes does not take into consideration the evolutionary history of each individual gene. Here, we describe an approach to select informative phylogenetic proteins to be used in the Tree of Life (TOL) and barcoding projects by comparing the cophenetic correlation coefficients (CCC) among individual protein distance matrices of proteins, using the fungi as an example. The method demonstrated that the quality and number of concatenated proteins is important for a reliable estimation of TOL. Approximately 40–45 concatenated proteins seem needed to resolve fungal TOL.</p> <p>Results</p> <p>In total 4852 orthologous proteins (KOGs) were assigned among 33 fungal genomes from the Asco- and Basidiomycota and 70 of these represented single copy proteins. The individual protein distance matrices based on 531 concatenated proteins that has been used for phylogeny reconstruction before <abbrgrp><abbr bid="B14">14</abbr></abbrgrp> were compared one with another in order to select those with the highest CCC, which then was used as a reference. This reference distance matrix was compared with those of the 70 single copy proteins selected and their CCC values were calculated. Sixty four KOGs showed a CCC above 0.50 and these were further considered for their phylogenetic potential. Proteins belonging to the cellular processes and signaling KOG category seem more informative than those belonging to the other three categories: information storage and processing; metabolism; and the poorly characterized category. After concatenation of 40 proteins the topology of the phylogenetic tree remained stable, but after concatenation of 60 or more proteins the bootstrap support values of some branches decreased, most likely due to the inclusion of proteins with lowers CCC values. The selection of protein sequences to be used in various TOL projects remains a critical and important process. The method described in this paper will contribute to a more objective selection of phylogenetically informative protein sequences.</p> <p>Conclusion</p> <p>This study provides candidate protein sequences to be considered as phylogenetic markers in different branches of fungal TOL. The selection procedure described here will be useful to select informative protein sequences to resolve branches of TOL that contain few or no species with completely sequenced genomes. The robust phylogenetic trees resulting from this method may contribute to our understanding of organismal diversification processes. The method proposed can be extended easily to other branches of TOL.</p

The reach of the genome signature in prokaryotes

BACKGROUND: With the increased availability of sequenced genomes there have been several initiatives to infer evolutionary relationships by whole genome characteristics. One of these studies suggested good congruence between genome synteny, shared gene content, 16S ribosomal DNA identity, codon usage and the genome signature in prokaryotes. Here we rigorously test the phylogenetic signal of the genome signature, which consists of the genome-specific relative frequencies of dinucleotides, on 334 sequenced prokaryotic genome sequences. RESULTS: Intrageneric comparisons show that in general the genomic dissimilarity scores are higher than in intraspecific comparisons, in accordance with the suggested phylogenetic signal of the genome signature. Exceptions to this trend, (Bartonella spp., Bordetella spp., Salmonella spp. and Yersinia spp.), which have low average intrageneric genomic dissimilarity scores, suggest that members of these genera might be considered the same species. On the other hand, high genomic dissimilarity values for intraspecific analyses suggest that in some cases (e.g.Prochlorococcus marinus, Pseudomonas fluorescens, Buchnera aphidicola and Rhodopseudomonas palustris) different strains from the same species may actually represent different species. Comparing 16S rDNA identity with genomic dissimilarity values corroborates the previously suggested trend in phylogenetic signal, albeit that the dissimilarity values only provide low resolution. CONCLUSION: The genome signature has a distinct phylogenetic signal, independent of individual genetic marker genes. A reliable phylogenetic clustering cannot be based on dissimilarity values alone, as bootstrapping is not possible for this parameter. It can however be used to support or refute a given phylogeny and resulting taxonomy

PhyloFunDB: A Pipeline to Create and Update Functional Gene Taxonomic Databases

The increase in sequencing capacity has amplified the number of taxonomically unclassified sequences in most databases. The classification of such sequences demands phylogenetic tree construction and comparison to currently classified sequences, a process that demands the processing of large amounts of data and use of several different software. Here, we present PhyloFunDB, a pipeline for extracting, processing, and inferring phylogenetic trees from specific functional genes. The goal of our work is to decrease processing time and facilitate the grouping of sequences that can be used for improved taxonomic classification of functional gene datasets

Current Challenges and Pitfalls in Soil Metagenomics

Soil microbial communities are essential components of agroecological ecosystems that influence soil fertility, nutrient turnover, and plant productivity. Metagenomics data are increasingly easy to obtain, but studies of soil metagenomics face three key challenges: (1) accounting for soil physicochemical properties; (2) incorporating untreated controls; and (3) sharing data. Accounting for soil physicochemical properties is crucial for better understanding the changes in soil microbial community composition, mechanisms, and abundance. Untreated controls provide a good baseline to measure changes in soil microbial communities and separate treatment effects from random effects. Sharing data increases reproducibility and enables meta-analyses, which are important for investigating overall effects. To overcome these challenges, we suggest establishing standard guidelines for the design of experiments for studying soil metagenomics. Addressing these challenges will promote a better understanding of soil microbial community composition and function, which we can exploit to enhance soil quality, health, and fertility

Responses of Acidobacteria Granulicella sp. WH15 to High Carbon Revealed by Integrated Omics Analyses

The phylum Acidobacteria is widely distributed in soils, but few representatives have been cultured. In general, Acidobacteria are oligotrophs and exhibit slow growth under laboratory conditions. We sequenced the genome of Granulicella sp. WH15, a strain obtained from decaying wood, and determined the bacterial transcriptome and proteome under growth in poor medium with a low or high concentration of sugar. We detected the presence of 217 carbohydrate-associated enzymes in the genome of strain WH15. Integrated analysis of the transcriptomic and proteomic profiles showed that high sugar triggered a stress response. As part of this response, transcripts related to cell wall stress, such as sigma factor σW and toxin–antitoxin (TA) systems, were upregulated, as were several proteins involved in detoxification and repair, including MdtA and OprM. KEGG metabolic pathway analysis indicated the repression of carbon metabolism (especially the pentose phosphate pathway) and the reduction of protein synthesis, carbohydrate metabolism, and cell division, suggesting the arrest of cell activity and growth. In summary, the stress response of Granulicella sp. WH15 induced by the presence of a high sugar concentration in the medium resulted in the intensification of secretion functions to eliminate toxic compounds and the reallocation of resources to cell maintenance instead of growth

Impact of Different Trace Elements on the Growth and Proteome of Two Strains of Granulicella, Class “Acidobacteriia”

Acidobacteria represents one of the most dominant bacterial groups across diverse ecosystems. However, insight into their ecology and physiology has been hampered by difficulties in cultivating members of this phylum. Previous cultivation efforts have suggested an important role of trace elements for the proliferation of Acidobacteria, however, the impact of these metals on their growth and metabolism is not known. In order to gain insight into this relationship, we evaluated the effect of trace element solution SL10 on the growth of two strains (5B5 and WH15) of Acidobacteria belonging to the genus Granulicella and studied the proteomic responses to manganese (Mn). Granulicella species had highest growth with the addition of Mn, as well as higher tolerance to this metal compared to seven other metal salts. Variations in tolerance to metal salt concentrations suggests that Granulicella sp. strains possess different mechanisms to deal with metal ion homeostasis and stress. Furthermore, Granulicella sp. 5B5 might be more adapted to survive in an environment with higher concentration of several metal ions when compared to Granulicella sp. WH15. The proteomic profiles of both strains indicated that Mn was more important in enhancing enzymatic activity than to protein expression regulation. In the genomic analyses, we did not find the most common transcriptional regulation of Mn homeostasis, but we found candidate transporters that could be potentially involved in Mn homeostasis for Granulicella species. The presence of such transporters might be involved in tolerance to higher Mn concentrations, improving the adaptability of bacteria to metal enriched environments, such as the decaying wood-rich Mn environment from which these two Granulicella strains were isolated

Perfil de virulência de dez isolados de Paracoccidioides brasiliensis: associação com morfologia e padrão genético

Ten isolates of Paracoccidioides brasiliensis were examined for differences in virulence in outbred mice intravenously inoculated with the fungus, associated with mycelial morphology, and genetic patterns measured by random amplified polymorphic DNA (RAPD). Virulence was evaluated by viable yeast cell recovery from lungs and demonstration of histopathologic lesions in different organs. The results showed that the isolates presented four virulence degrees: high virulence, intermediate, low and non-virulence. RAPD clustered the isolates studied in two main groups with 56% of genetic similarity. Strains with low virulence, Pb265 or the non-virulent, Pb192, showed glabrous/cerebriform morphology and high genetic similarity (98.7%) when compared to the other isolates studied. The same was observed with Bt79 and Bt83 that shared 96% genetic similarity, cottony colonies and high virulence. The RAPD technique could only discriminate P. brasiliensis isolates according to glabrous/cerebriform or cottony colonies, and also high from low virulence strains. Isolates with intermediate virulence such as Pb18, Pb18B6, Bt32 and Bt56 showed variability in their similarity coefficient suggesting that RAPD was able to detect genetic variability in this fungal specie. Virulence profile of P. brasiliensis demonstrated that both mycelial morphologic extreme phenotypes may be associated with fungal virulence and their in vitro subculture time. Thus, RAPD technique analysis employed in association with virulence, morphologic and immunologic aspects might prove adequate to detect differences between P. brasiliensis isolates.Dez isolados de P. brasiliensis foram avaliados em relação à patogenicidade por inoculação intravenosa em camundongos e associação com morfologia miceliana e padrão genético por amplificação genônica do DNA polimórfico (RAPD). A patogenicidade, avaliada por recuperação de fungos viáveis a partir de tecido pulmonar e por lesões histopatológicas em diferentes órgãos, mostrou que os isolados apresentaram quatro graus de virulência: alta virulência, virulência intermediária, baixa virulência e não virulência. A técnica de RAPD agrupou os isolados em dois grupos com 56% de similaridade genética. Amostras com baixa virulência Pb265 ou não virulência Pb192 apresentaram morfologia glabra/cerebriforme e alta similaridade genética (98,7%) quando comparadas com os outros isolados estudados. O mesmo foi observado com os isolados Bt79 e Bt83, que compartilharam 96% de semelhança genética, colônias cotonosas e alta virulência. Essa técnica pode discriminar apenas isolados com morfologia glabra da cotonosa e com alta e baixa virulência. Isolados com virulência intermediária como Pb18, Pb18B6, Bt32 e Bt54 mostraram variabilidade no coeficiente de similaridade, sugerindo que a técnica de RAPD permite mostrar variabilidade genética nessa espécie fúngica. O estudo do perfil de virulência das amostras de P. brasiliensis demonstrou que os dois fenótipos extremos de morfologia miceliana podem ser associados com a virulência do fungo e com o tempo de subcultivo in vitro. Assim, a análise de RAPD, utilizada em conjunto com aspectos de virulência, morfológicos e imunológicos pode ser considerada adequada para detectar diferenças entre isolados de P. brasiliensis

Bacterial community composition and diversity of two different forms of an organic residue of bioenergy crop

The use of residue of sugarcane ethanol industry named vinasse in fertirrigation is an established and widespread practice in Brazil. Both non-concentrated vinasse (NCV) and concentrated vinasse (CV) are used in fertirrigation, particularly to replace the potassium fertilizer. Although studies on the chemical and organic composition of vinasse and their impact on nitrous oxide emissions when applied in soil have been carried out, no studies have evaluated the microbial community composition and diversity in different forms of vinasse. We assessed the bacterial community composition of NCV and CV by non-culturable and culturable approaches. The non-culturable bacterial community was assessed by next generation sequencing of the 16S rRNA gene and culturable community by isolation of bacterial strains and molecular and biochemical characterization. Additionally, we assessed in the bacterial strains the presence of genes of nitrogen cycle nitrification and denitrification pathways. The microbial community based on 16S rRNA sequences of NCV was overrepresented by Bacilli and Negativicutes while CV was mainly represented by Bacilli class. The isolated strains from the two types of vinasse belong to class Bacilli, similar to Lysinibacillus, encode for nirK gene related to denitrification pathway. This study highlights the bacterial microbial composition particularly in CV what residue is currently recycled and recommended as a sustainable practice in sugarcane cultivation in the tropics

Structural and functional variation in soil fungal communities associated with litter bags containing maize leaf

Soil fungi are key players in the degradation of recalcitrant organic matter in terrestrial ecosystems. To examine the organisms and genes responsible for complex organic matter degradation in soil, we tracked changes in fungal community composition and expressed genes in soil adjacent to mesh bags containing maize leaves undergoing decomposition. Using high-throughput sequencing approaches, changes in fungal community composition were determined by targeting 18S rRNA gene sequences, whereas community gene expression was examined via a metatranscriptomic approach. The majority of the 93 000 partial 18S rRNA gene sequences generated, were affiliated with the Ascomycota and Basidiomycota. Fungal diversity was at least 224 operational taxonomic units at the 97% similarity cutoff level. During litter degradation, the relative proportion of Basidiomycota increased, with a decrease in Ascomycota : Basidiomycota ratios over time. The most commonly detected decomposition-associated fungi included Agaricomycetes and Tremellales as well as unclassified Mucoromycotina. The majority of protein families found in the metatranscriptomic data were affiliated to fungal groups described to degrade plant-derived cellulose, such as Mucoraceae, Chaetomiaceae, Sordariaceae, Sebacinaceae, Tremellaceae, Psathyrellaceae and Schizophyllaceae. The combination of high-throughput rRNA gene-based and metatranscriptomic approaches provided perspectives into the organisms and genes involved in complex organic matter in soi

Microbial Extracellular Polymeric Substances: Ecological Function and Impact on Soil Aggregation

A wide range of microorganisms produce extracellular polymeric substances (EPS), highly hydrated polymers that are mainly composed of polysaccharides, proteins, and DNA. EPS are fundamental for microbial life and provide an ideal environment for chemical reactions, nutrient entrapment, and protection against environmental stresses such as salinity and drought. Microbial EPS can enhance the aggregation of soil particles and benefit plants by maintaining the moisture of the environment and trapping nutrients. In addition, EPS have unique characteristics, such as biocompatibility, gelling, and thickening capabilities, with industrial applications. However, despite decades of research on the industrial potential of EPS, only a few polymers are widely used in different areas, especially in agriculture. This review provides an overview of current knowledge on the ecological functions of microbial EPSs and their application in agricultural soils to improve soil particle aggregation, an important factor for soil structure, health, and fertility