Biology subjects

Agustinho, D. P.

Publications and source records attributed to Agustinho, D. P..

5 recordsLinked to original sources

bioRxiv2026

Rapid phylogenomic analysis for viral surveillance and metagenomic profiling with Omni2Tree

Phylogenomic surveillance is limited not by sequencing throughput, but by the difficulty of converting heterogeneous raw data into reliable evolutionary inference, particularly for low-titer and contaminated viral field samples. Here we present Omni2Tree, an assembly-free framework that reconstructs viral phylogenies directly from raw sequencing reads and generates easily shareable interactive reports and genome-wide entropy profiles to identify diversification. In H5N1 benchmark analyses, Omni2Tree maintained accurate placement and topological stability even under low coverage, unlike assembly or reference based methods. Omni2Tree generated an annotated phylogeny for 64-sample H5N1 field surveillance dataset from the eastern USA in under 3 hours. Omni2Tree recovered known phylogenetic structure and key variability insights across 1,328 hepatitis C virus and 707 human cytomegalovirus datasets, and resolved co-infecting respiratory viruses in clinical metagenomic samples. By enabling direct analysis from raw reads, Omni2Tree supports faster, more portable, and more decentralized phylogenomic surveillance across outbreak, clinical, and resource-limited settings.

bioinformatics↗

bioRxiv2026

Two closely related β-1,2-xylosyltransferases differentially impact fungal glycan synthesis

Cryptococcus neoformans is an opportunistic fungal pathogen that causes pulmonary infection in immunocompromised patients, which in severe cases leads to fatal meningoencephalitis. Cryptococcus exhibits unique glycobiology that plays important roles in pathogenesis. Unlike model yeast and other common fungal pathogens, Cryptococcus incorporates xylose, a five-carbon monosaccharide, into its glycans. One trimer motif, which consists of xylose in {beta}-1,2 linkage to the reducing mannose of an -1,3-mannose dimer, occurs in key cryptococcal glycoconjugates that include protein N- and O-linked glycans, glycosylinositol phosphorylceramides (GIPCs), and the capsule polysaccharides glucuronoxylomannan (GXM) and glucuronoxylomannogalactan (GXMGal). We previously identified cryptococcal {beta}-1,2-xylosyltransferase 1 (Cxt1), which catalyzes formation of this motif in GIPCs, GXM, and GXMGal. Here, we report the discovery of a second enzyme, cryptococcal {beta}-1,2-xylosyltransferase 2 (Cxt2). Through characterization of cells that lack one or both corresponding genes (CXT1 and CXT2), we have dissected the biological roles of these enzymes, which are overlapping but not identical. Notably, Cxt1 and Cxt2 colocalize in the Golgi, influence capsule in a strain-dependent manner, and together are responsible for all xylose addition to O-glycans. Overall, our work highlights unique roles of these two enzymes and fills a gap in understanding of cryptococcal glycan synthesis. IMPORTANCECryptococcus neoformans is an opportunistic fungal pathogen that causes almost 150,000 deaths each year worldwide. Cryptococcus synthesizes unique glycan structures that play important roles in its biology and pathogenesis. One abundant component of these structures is xylose, a five-carbon monosaccharide. Because xylose is not used by many fungal organisms, including model yeast, we have limited information about how cells add it to their glycans. Here we report a xylosyltransferase enzyme that performs this function, and we characterize specific biological roles of this protein and a closely related one we discovered earlier. We find that these proteins together perform all detectable xylose addition to an important class of protein-linked glycans (O-glycans). They also both participate in other synthetic processes, although this varies with the specific enzyme and strain background. These results contribute to our understanding of cryptococcal glycan synthesis and underscore the importance of testing multiple background strains.

microbiology↗

bioRxiv2025

Evolutionary dynamics of Respiratory Syncytial Virus in pre-pandemic, pandemic, and post-pandemic periods in Houston, Texas, USA

BackgroundRespiratory syncytial virus (RSV) is a leading cause of severe lower respiratory tract infections in infants and poses significant risks to immuno-compromised individuals and the elderly. The COVID-19 pandemic disrupted typical RSV seasonality, leading to unusual patterns of viral circulation and resurgence. However, the impact of these disruptions on RSV evolutionary dynamics remains incompletely understood. This study aimed to investigate the evolutionary dynamics of RSV in Houston, Texas, across pre-pandemic (November 2015 - March 2020), pandemic (April 2021 - June 2023), and post-pandemic periods (July 2023 - December 2024), focusing on genetic diversity, lineage dynamics, and selective pressures. MethodsIn one of the largest comprehensive RSV genomic evolutionary analyses, we analyzed 1,344 RSV-positive respiratory samples collected from children seeking outpatient care or hospitalized with acute respiratory infections between November 2015 to February 2024, which is one of the largest comprehensive RSV genomic evolutionary analyses. We successfully sequenced the whole genomes for 87.5% of the isolates (606 RSV/A and 570 RSV/B). To assess genetic diversity, lineage dynamics, and selective pressures, we conducted phylogenetic analysis, variant calling, and dN/dS ratio calculations. We clustered variant prevalence patterns using k-means and used Shannon entropy to quantify genetic variability. We performed statistical regression analyses to evaluate the accumulation of variants and their impact on gene expression. ResultsRSV circulation in Houston was markedly disrupted during the pandemic, with an absence of cases in 2020-2021 followed by off-season surges from 2021 to 2023. Phylogenetic analysis revealed distinct lineage dynamics, with RSV/A showing remarkable persistence of specific lineages such as A.D.1 and emergence of new lineages including A.D.3. In contrast, RSV/B underwent a dramatic restructuring, dominated by the B.D.E.1 lineage in the pandemic and post-pandemic period. Bayesian molecular clock analysis estimated nucleotide substitution rates of 9.37 x 10-4 (95% HPD: 7.99 x 10-4 - 1.07 x 10-3) substitutions/site/year for RSV/A and 9.79 x 10-4 (95% HPD: 8.96 x 10-4 - 1.06 x 10-3) for RSV/B. The time to the most recent common ancestor (tMRCA) of the dominant RSV/A clade, A.D.1, is dated to approximately 2010 (95% HPD: 2008-2011), while for RSV/B the dominant clade- B.D.E.1 traces back to 2002 (95% HPD: 1997-2007). Despite reduced lineage diversity, RSV/B accumulated non-synonymous variants at nearly twice the rate of RSV/A, notably in the M2-2 gene, reflecting high within-lineage variability. The significant increase in non-synonymous mutations in M2-2 in RSV/B during the pandemic and post-pandemic time phases strongly correlated with increased transcriptional activity. Additionally, antigenic site mutations in the F protein, particularly in RSV/B, were also observed, with implications for immune evasion and effectiveness of the current monoclonal antibody immunoprophylaxis intervention. ConclusionsThe COVID-19 pandemic significantly impacted RSV evolution, leading to reduced genetic diversity during the pandemic and the emergence of novel lineages post-pandemic. RSV/B exhibited more dynamic evolutionary changes, particularly in the M2-2 gene, suggesting potential adaptive advantages. Ongoing genomic surveillance coupled with functional studies is crucial for monitoring variants and assessing the clinical impact of these mutations on transmission, disease severity, fitness, and long-term effectiveness of RSV prevention and treatment strategies.

evolutionary biology↗

bioRxiv2023

Unbiased discovery of natural sequence variants that influence fungal virulence

Isolates of Cryptococcus neoformans, a fungal pathogen that kills over 120,000 people each year, differ from a 19-megabase reference genome at a few thousand up to almost a million DNA sequence positions. We used bulked segregant analysis and association analysis, genetic methods that require no prior knowledge of sequence function, to address the key question of which naturally occurring sequence variants influence fungal virulence. We identified a region containing such variants, prioritized them, and engineered strains to test our findings in a mouse model of infection. At one locus we identified a 4-nt variant in the PDE2 gene, which severely truncates its phosphodiesterase product and significantly alters virulence. Our studies demonstrate a powerful and unbiased strategy for identifying key genomic regions in the absence of prior information, suggest revisions to current assumptions about cAMP levels and about common laboratory strains, and provide significant sequence and strain resources to the community.

microbiology↗

bioRxiv2020

The transcription factor Pdr802 regulates Cryptococcus neoformans Titan cell production and pathogenicity

Cryptococcus neoformans is a ubiquitous, opportunistic fungal pathogen that kills almost 200,000 people worldwide each year. It is acquired when mammalian hosts inhale the infectious propagules; these are deposited in the lung and, in the context of immunocompromise, may disseminate to the brain and cause lethal meningoencephalitis. Once inside the host, C. neoformans undergoes a variety of adaptive processes, including secretion of virulence factors, expansion of a polysaccharide capsule that impedes phagocytosis, and the production of giant (Titan) cells. The transcription factor Pdr802 is one regulator of these responses to the host environment. Expression of the corresponding gene is highly induced under host-like conditions in vitro and is critical for C. neoformans dissemination and virulence in a mouse model of infection. Direct targets of Pdr802 include the quorum sensing proteins Pqp1, Opt1 and Liv3; the transcription factors Stb4, Zfc3 and Bzp4, which regulate cryptococcal brain infectivity and capsule thickness; the calcineurin targets Had1 and Crz1, important for cell wall remodeling and C. neoformans virulence; and additional genes related to resistance to host temperature and oxidative stress, and to urease activity. Notably, cryptococci engineered to lack Pdr802 showed a dramatic increase in Titan cells, which are not phagocytosed and have diminished ability to directly cross biological barriers. This explains the limited dissemination of pdr802 mutant cells to the central nervous system and the consequently reduced virulence of this strain. The role of Pdr802 as a negative regulator of Titan cell formation is thus critical for cryptococcal pathogenicity. IMPORTANCEThe pathogenic yeast Cryptococcus neoformans presents a worldwide threat to human health, especially in the context of immunocompromise, and current antifungal therapy is hindered by cost, limited availability, and inadequate efficacy. After the infectious particle is inhaled, C. neoformans initiates a complex transcriptional program that integrates cellular responses and enables adaptation to the host lung environment. Here we describe the role of the transcription factor Pdr802 in the response to host conditions and its impact on C. neoformans virulence. We identified direct targets of Pdr802 and also discovered that it regulates cellular features that influence movement of this pathogen from the lung to the brain, where it causes fatal disease. These findings advance our understanding of a serious disease.

microbiology↗