Phylogeny-Guided Microbiome OTU-Specific Association Test (POST)

Background: The relationship between host conditions and microbiome profiles, typically characterized by operational taxonomic units (OTUs), contains important information about the microbial role in human health. Traditional association testing frameworks are challenged by the high-dimensionality and sparsity of typical microbiome profiles. Incorporating phylogenetic information is often used to address these challenges with the assumption that evolutionarily similar taxa tend to behave similarly. However, this assumption may not always be valid due to the complex effect of microbes, and phylogenetic information should be incorporated in a data-supervised fashion. Results: In this work, we propose a local collapsing test called Phylogeny-guided microbiome OTU-Specific association Test (POST). In POST, whether or not to borrow information and how much information to borrow from the neighboring OTUs in the phylogenic tree are supervised by phylogenetic distance and the outcome-OTU association. POST is constructed under the kernel machine framework to accommodate complex OTU effects and extends kernel machine microbiome tests from community-level to OTU-level. Using simulation studies, we showed that when the phylogenetic tree is informative, POST has better performance than existing OTU-level association tests. When the phylogenetic tree is not informative, POST achieves similar performance as existing methods. Finally, we show that POST can identify more outcome-associated OTUs that are of biological relevance in real data applications on bacterial vaginosis and on preterm birth. Conclusions: Using POST, we show that the power of detecting associated microbiome features can be enhanced by adaptively leveraging the phylogenetic information when testing for a target OTU. We developed an user friendly R package POSTm which is now available at CRAN (https://CRAN.R-project.org/package=POSTm) for public access.

Identification and characterization of histones in Physarum polycephalum evidence a phylogenetic vicinity of Mycetozoans to the animal kingdom

Physarum polycephalum belongs to Mycetozoans, a phylogenetic clade apart from the animal, plant and fungus kingdoms. Histones are nuclear proteins involved in genome organization and regulation and are among the most evolutionary conserved proteins within eukaryotes. Therefore, this raises the question of their conservation in Physarum and the position of this organism within the eukaryotic phylogenic tree based on histone sequences. We carried out a comprehensive study of histones in Physarum polycephalum using genomic, transcriptomic and molecular data. Our results allowed to identify the different isoforms of the core histones H2A, H2B, H3 and H4 which exhibit strong conservation of amino acid residues previously identified as subject to post-translational modifications. Furthermore, we also identified the linker histone H1, the most divergent histone, and characterized a large number of its PTMs by mass spectrometry. We also performed an in-depth investigation of histone genes and transcript structures. Histone proteins are highly conserved in Physarum and their characterization will contribute to a better understanding of the polyphyletic Mycetozoan group. Our data reinforce that P. polycephalum is evolutionary closer to animals than plants and located at the crown of the eukaryotic tree. Our study provides new insights in the evolutionary history of Physarum and eukaryote lineages.

Molecular Identification and Phylogenetic Study of a Novel Pravastatin-producing Fungal Strain

Pravastatin is a cholesterol-lowering agent of high therapeutic value which is obtained based on a twostep production process. The development of one-step fermentation process based on pravastatin-producing microfungi could be advantageous. We isolated a number of soil microfungi and screened them for pravastatin production. Among them, the isolate ESF21 was selected as an active pravastatin producer. The aim of the present study was to identify this isolate to species level and to investigate pravastatin accumulation by this microfungus. As a result, a range of standard macro- and micro-morphological characters of the ESF21 strain was documented. The isolate ESF21 was a member of the genus Penicillium. Based on the results of molecular characterization, the isolated culture was identified as Penicillium brefeldianum ESF21. To analyze the phylogenetic relationships between this microfungus and other Penicillium species, a phylogenic tree was constructed. When the time course of pravastatin production by the identified strain was investigated, the maximum level of pravastatin (196.83 mg/L) was accumulated after 10 days of fermentation. The identified strain has a scientific interest as a previously unknown wild-type fungal producer of pravastatin.

Identification of hidden N4-like viruses and their interactions with hosts in global metagenomes

Background: N4-like viruses, with specific genomic features and propagation signatures, comprise a unique viral clade within the Podoviridae family. N4-like viruses are commonly characterized by the N4-like major capsid protein (MCP) and a giant virion-encapsulated RNA polymerase (N4-like RNAP) with a size of approximately 3,500-aa, which is the largest viral protein so far described. To date, our understanding of N4-like viruses is largely derived from 80 viral isolates that infect bacteria. Thus, it is necessary to expand the diversity of N4-like viruses in culturing-independent methods.Methods: A Hidden-Markov-Module based method was designed based on two characterized N4-specific marker genes, major capsid protein and N4-like virion-encapsulated RNA polymerase. Viral sub-clades were classified based on the monophyly presented in phylogenic tree and the results of pangenome analysis. Further analysis assessed different distribution patterns, genomic properties, hosts’ metabolism reprogramming potentialities, significance of viral tRNA and horizontal gene transfer landscape.Results: We identified 1,000 N4-like virus sequences from genomes and metagenomes representing diverse habitats from around the world. N4-like viruses have been classified into 27 sub-clades and detected in almost all habitats from pole to pole, including some novel habitats, such as oral mucosa and Antarctica. Virulent factors might be crucial for some human-associated N4-like viruses to reprogram the metabolism of host cells and mediate their pathogenic ability through horizontal gene transfer. From the pangenome analysis, the protein diversity was expended over 7-fold and 17 conserved house-keeping genes were identified. Transcriptional compensation of tRNA indicates that producing progeny virion might be the main significance of viral tRNAs. From the horizontal gene transfer network, some N4-like viral sub-clades were observed that potentially infect some important human pathogens, such as Campylobacteria and Veillonella , which have not been considered as potential hosts of N4-like virus or even any virus.Conclusion: This study expands the knowledge of N4-like viruses via global metagenomic datasets, reveals the novel ecological and genomic signatures of these viruses and will provide the backbone for further N4-like virus studies.

Phylogenetic molecular evolution and recombination analysis of complete genome of human parechovirus in Thailand

Human parechovirus (HPeV), which is a member of the Picornavirus group of viruses, is a pathogen that is reported to be associated with manifestations that include respiratory tract involvement, gastroenteritis, sepsis-like symptom, and central nervous system complication. Until now, nineteen genotypes have been identified. The lack of proofreading property of viral RNA-dependent RNA polymerase (RdRp) together with recombination among the intra- and inter-genotypes of the virus results in high diversity. However, data specific to the molecular evolutionary perspective of the complete genome of HPeV remains limited. This study aimed to analyze the phylogenetic, molecular evolution, and recombination characteristics of the complete genome of HPeV strains isolated in Thailand during 2009–2012. Fifty-eight samples that were previously confirmed to be HPeV positive and then evaluated for genotyping were subjected to complete genome amplification to generate ten overlapping PCR fragments using a set of in-house designed primers. The same position of the viral genome was read in triplicate using direct Sanger sequencing. All samples were classified into the same previously defined genotypes in both whole-genome and VP1 phylogenic tree. However, sample B1091/HPeV14/2011 exhibited discordant grouping between whole-genome and VP1 on the phylogenetic tree. Bootscan analysis revealed that B1091/HPeV14/2011 inherited from two genotypic viruses, including VP1 from HPeV14, and the rest of the genome from HPeV1B. The results of this study provide important insights into the molecular evolution of and recombination in the viral genome of HPeV that will improve and accelerate our ability to develop treatment and prophylactic strategies in the future.

Clinical Origin and Species Distribution of Fusarium spp. Isolates Identified by Molecular Sequencing and Mass Spectrometry: A European Multicenter Hospital Prospective Study

Fusarium spp. are widespread environmental fungi as well as pathogens that can affect plants, animals and humans. Yet the epidemiology of human fusariosis is still cloudy due to the rapidly evolving taxonomy. The Mass Spectrometry Identification database (MSI) has been developed since 2017 in order to allow a fast, accurate and free-access identification of fungi by matrix-assisted laser desorption ionization—time of flight (MALDI-TOF) mass spectrometry. Taking advantage of the MSI database user network, we aim to study the species distribution of Fusarium spp. isolates in an international multicenter prospective study. This study also allowed the assessment of the abilities of miscellaneous techniques to identify Fusarium isolates at the species level. The identification was performed by PCR-sequencing and phylogenic-tree approach. Both methods are used as gold standard for the evaluation of mass spectrometry. Identification at the species complex was satisfactory for all the tested methods. However, identification at the species level was more challenging and only 32% of the isolates were correctly identified with the National Center for Biotechnology Information (NCBI) DNA database, 20% with the Bruker MS database and 43% with the two MSI databases. Improvement of the mass spectrometry database is still needed to enable precise identification at the species level of any Fusarium isolates encountered either in human pathology or in the environment.

Spread of SARS-CoV-2 Genomes on Genomic Index Maps of Hierarchy - Compared with B.1.1.7 Lineage on BLAST

COVID-19 patients worldwide are conveniently described by position information to collect samples, and modern GIS maps are useful to show influenced flows and numbers of patients on various regions of a pendamic. From an analysis viewpoint, it is more interesting to organize genomic information into a phylogenic tree with multiple branches and leaves in representations. Clusters of genomes are organized as phylogenic trees to represent intrinsic information of genomes. However, there are structural difficulties in projecting phylogenetic information into 2D distributions as GIS maps naturally.Considering advanced generating schemes of phylogenetic trees, information entropy provides ultra optimal properties in the minimum computational complexity, superior flexibility, better stability, improved reliability and higher quality on global constructions.In this paper, a novel projection is proposed to arrange SARS-CoV-2 genomes by genomic indexes to make a structural organization as 2D GIS maps. For any genome, there is a unique invariant under certain conditions to provide an absolute position on a specific region. In this hierarchical framework, it is possible to use a visual tool to represent any selected region for clustering genomes on refined effects. Applied diversity measure to a given set of genomes, equivalent clusters and complementary visual effects are provided between genomic index maps and phylogenetic trees. Sample genomes of three UK new lineages are aligned by BLAST as a basis on both RNA-dependent RNA polymerase RDRP segments and whole genomes. Selected regions and various projections show spread effects of five thousand SARS-CoV-2 genomes in 72 countries on both RDRP and whole genomes, and six special countries/regions are selected on genomic index maps.Based on genomic index maps, one SNV of two genomes on B.1.1.7 lineage can be identified from a unit of 10^4 probability measure to a unit of 10^6 difference for genomic indexes on a special ‘G’ projection to extract the finest variation.Further exploration on optimal classification and phylogenetic analysis of genomic index maps and phylogenetic trees on SARS-CoV-2 genomes worldwide are discussed.

Impaired tumor immune response in metastatic tumors is a selective pressure for neutral evolution in CRC cases

A Darwinian evolutionary shift occurs early in the neutral evolution of advanced colorectal carcinoma (CRC), and copy number aberrations (CNA) are essential in the transition from adenoma to carcinoma. In light of this primary evolution, we investigated the evolutionary principles of the genome that foster postoperative recurrence of CRC. CNA and neoantigens (NAG) were compared between early primary tumors with recurrence (CRCR) and early primary tumors without recurrence (precancerous and early; PCRC). We compared CNA, single nucleotide variance (SNV), RNA sequences, and T-cell receptor (TCR) repertoire between 9 primary and 10 metastatic sites from 10 CRCR cases. We found that NAG in primary sites were fewer in CRCR than in PCRC, while the arm level CNA were significantly higher in primary sites in CRCR than in PCRC. Further, a comparison of genomic aberrations of primary and metastatic conditions revealed no significant differences in CNA. The driver mutations in recurrence were the trunk of the evolutionary phylogenic tree from primary sites to recurrence sites. Notably, PD-1 and TIM3, T cell exhaustion-related molecules of the tumor immune response, were abundantly expressed in metastatic sites compared to primary sites along with the increased number of CD8 expressing cells. The postoperative recurrence-free survival period was only significantly associated with the NAG levels and TCR repertoire diversity in metastatic sites. Therefore, CNA with diminished NAG and diverse TCR repertoire in pre-metastatic sites may determine postoperative recurrence of CRC.

Genomic insights into methicillin-resistant Staphylococcus aureus spa type t899 isolates belonging to different sequence types

Methicillin-resistant Staphylococcus aureus (MRSA) presenting the spa type t899 is commonly associated with sequence type (ST) 9, but also increasingly linked to ST398. This study provides a genomic insight into the diversity of t899 isolates using cgMLST, SNP-based phylogeny and the description of selected antimicrobial resistance and virulence markers. The SNP-based phylogenic tree showed that isolates sharing the same spa type (t899) but different STs highly diverged in their core and accessory genomes, revealing discriminant antimicrobial-resistant (AMR) and virulence markers. Our results highlighted that, in a surveillance context where only spa typing is used, an additional multiplex PCR for the detection of the tet(M), sak and seg genes would valuably allow to distinguish ST9 from ST398 isolates on a routine basis. Importance This study showed the genetic diversity and population structure of S. aureus presenting the same spa type t899 but belonging to different STs. Our findings revealed that these isolates vary deeply in their core and accessory genomes, contrary to what is regularly inferred from studies using spa typing only. Given that identical spa types can be associated with different STs, and knowing that spa typing only is not appropriate for S. aureus isolates that underwent major recombination events which include the passage of the spa gene (such as in t899-positive MRSA), the combination of both MLST and spa typing methods is recommended. However, spa typing only is still largely used in surveillance studies and basic characterization. Our data suggest that additional markers, such as tet(M), sak and seg genes, could be implemented in an easy and cheap manner in order to identify S. aureus lineages with a higher accuracy.