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

Phosphorus-mediated succession of microbial nitrogen, carbon, and sulfur functions in rice-driven saline-alkali soil remediation

Although rice cultivation holds potential for restoring unproductive saline-alkali soils and increasing food production, the mechanisms underlying the relationship between microbial functions and soil element turnover remain unclear. To clarify this relationship, this study investigated the soil physicochemical properties and microbial functions during remediation in saline-alkali soil by rice cultivation over 2, 4, 6, 8, 11, 12, 20, and 23 years. The results indicated rice cultivation markedly improved soil nutrients, soil nutrient stoichiometry, and soil aggregate stability. Additionally, rice cultivation significantly increased the microbial functions involved in nutrient cycling, such as nitrogen fixation, carbon fixation, methanogenesis, dissimilatory sulfate reduction, and thiosulfate oxidation. However, these nitrogen (N), carbon (C), and sulfur (S) cycle-related functions exhibited a similar “increase-peak-decrease” successional pattern with the years of remediation, reaching optimal levels when rice was continuously grown for 11–16 years. Furthermore, correlation analysis demonstrated that the succession of soil microbial N, C, and S functions during saline-alkali soil restoration closely related to changes in soil properties, particularly the availability of phosphorus (P). Therefore, we propose to prioritize the management of P during saline-alkali soil remediation. In conclusion, this study provides a comprehensive understanding of the microbial N, C, and S functions and soil P in the remediation of saline-alkali soils mediated by rice crop

Enhancing phosphate-solubilising microbial communities through artificial selection

Microbial communities, acting as key drivers of ecosystem processes, harbour immense potential for sustainable agriculture practices. Phosphate-solubilising microorganisms, for example, can partially replace conventional phosphate fertilisers, which rely on finite resources. However, understanding the mechanisms and engineering efficient communities poses a significant challenge. In this study, we employ two artificial selection methods, environmental perturbation, and propagation, to construct phosphate-solubilising microbial communities. To assess trait transferability, we investigate the community performance in different media and a hydroponic system with Chrysanthemum indicum. Our findings reveal a distinct subset of phosphate-solubilising bacteria primarily dominated by Klebsiella and Enterobacterales. The propagated communities consistently demonstrate elevated levels of phosphate solubilisation, surpassing the starting soil community by 24.2% in activity. The increased activity of propagated communities remains consistent upon introduction into the hydroponic system. This study shows the efficacy of community-level artificial selection, particularly through propagation, as a tool for successfully modifying microbial communities to enhance phosphate solubilisation

Stable isotope probing reveals compositional and functional shifts in active denitrifying communities along the soil profile in an intensive agricultural area

Denitrifying microbial communities in the vadose zone play an essential role in eliminating the nitrate leached from agricultural practices. This nitrate could otherwise contaminate groundwater and threaten public health. Here, we utilized stable isotope probing combined with amplicon sequencing and functional gene quantification to inspect the composition and function of heterotrophic denitrifying microorganisms along a 9-m soil profile in an intensive agricultural area. Dramatic differences in the composition of the active denitrifiers were uncovered between the surface soil and deep layers of the vadose zone. The main denitrifying bacterial taxa identified from 13C-DNA fractions were Pseudomonadaceae (Pseudomonas), Rhodocyclaceae (Azoarcus), and Burkholderiaceae in the surface soil (0–0.2 m), and were Pseudomonadaceae (Pseudomonas), Burkholderiaceae, Bacillaceae (Bacillus), and Paenibacillaceae (Ammoniphilus) in the deep layers (0.5–9.0 m). Analysis of the functional genes (nirS, nirK, and nosZ) in isotope-labeled DNA revealed an upward nos/nir ratio with increasing soil depth, which may account for the higher nitrous oxide emission potential in the surface soil, as compared to the deeper sand-rich, low organic carbon layers. This study improves our understanding of active denitrifying microbes in the vadose zone and helps in developing techniques to reduce nitrate pollution in groundwater

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

Arbuscular mycorrhizal fungi originated from soils with a fertility gradient highlight a strong intraspecies functional variability

Characterization and selection of arbuscular mycorrhizal fungal (AMF) taxa to design inocula tailored to meet a spectrum of needs is a crucial first step to achieve specific beneficial agronomic functions. Commonly, commercial microbial inocula are based on generalist single AM fungal taxa, having low genetic variability and not offering efficiency and stability when applied in agroecosystems. In this study, we investigated the AMF functional variability at inter- and intra-species levels by characterizing colonization traits, host growth, and mineral uptake of single-spore AM fungi isolated from soils with a fertility gradient. Nineteen single-spore cultures, showing high spore density and AMF colonization, were phylogenetically assigned to different isolates of 3 AMF species (i.e. Entrophospora claroidea, Funneliformis mosseae and Archaeospora trappei). A higher functional variability in infectivity and effectiveness was detected among isolates within AMF species (25 % of total variance) than among AMF species. Most of AMF isolates of F. mosseae have a better outcome in terms of plant growth, although with a performance gradient, while the isolates of E. claroidea showed a variable functional pattern, and those of A. trappei a less variable pattern. Overall, isolates originating from the soil of the conventional arable field with higher pH and phosphorous availability promoted the uptake of plant nutrients, while those originating from soils with higher SOM and plant diversity promoted plant growth. On the contrary, the infectivity traits of the AM fungi were more conserved, as they were not affected by the environmental parameters of the soils of origin. Finally, we highlighted that soil pH played an important role in shaping the pattern of AMF functionality. Boosting the isolation and cultivation of AMF taxa, originating from agricultural and natural soils, is shown to be a key step in exploiting AMF diversity and designing the new generation of microbial inoculants

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

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

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