Biology subjects

Duffy, C.

Publications and source records attributed to Duffy, C..

8 recordsLinked to original sources

bioRxiv2026

Restricted expression site use and extreme genome diversification drives trypanosome antigenic variation in chronic bovine infections

Trypanosoma brucei exploits an extreme form of antigenic variation to escape the mammalian immune response. This involves the progressive expression of antigenically distinct variant surface glycoproteins (VSGs) on the surface of individual parasites in the population, generating waves of parasitaemia that are successively cleared by host antibodies. Current paradigms were established using in vitro studies and acute rodent infections characterized by high parasitaemia, but natural livestock infections are characterized by low parasitaemia and chronicity. Here, we analysed the infection dynamics of isogenic parasites in mice and cattle in blood during early and chronic infections, quantitating VSG expression diversity within and between hosts, antigen type persistence in vivo and their timing of appearance. This revealed enhanced antigenic diversity in cattle but with a surprisingly reproducible temporal expression hierarchy of related VSGs between independent chronic infections. Analyses demonstrated the unexpected dominance of a single telomeric VSG expression site irrespective of host species and time of infection. Detailed prediction of mosaic VSG assembly reveals exceptional parasite genome diversification within infections involving extensive macro and micro-homology-based recombination to evolve the antigen repertoire. This diversity was restricted but not eliminated in homologous recombination mutants, which could nonetheless sustain chronic infections in mice. These data provide the first comprehensive insight into trypanosome antigenic variation in the clinically-relevant host.

microbiology↗

bioRxiv2026

Clade 2.3.4.4b H5N1 influenza virus and SARS-CoV-2 seroprevalence among owned and feral cats in Philadelphia and surrounding communities

Clade 2.3.4.4b H5NX influenza viruses have spread widely in birds since 2020. In addition to causing disease in birds, these viruses have infected a variety of mammals, including humans. Clade 2.3.4.4b H5N1 viruses are currently causing an outbreak among dairy cattle in the United States, and it is important to determine if other mammals have been exposed to H5NX viruses. Cats, specifically outdoor and feral cats, frequently predate wild birds. Recent studies have shown that cats living on dairy cattle farms can be infected with H5N1. Here, we completed serological studies to determine if owned and feral cats living in an urban environment in the United States have evidence of past H5N1 exposures. We used multianalyte bead-based assays to measure clade 2.3.4.4b hemagglutinin (HA) antibody levels in serum samples collected in July 2023 to June 2025 from 417 feral and 228 owned cats from the greater Philadelphia area. We also measured antibody levels against a panel of HAs from other human and non-human influenza viruses, and the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We completed additional H5N1 and SARS-CoV-2 neutralization assays using samples that had detectable antibodies in the multianalyte bead-based assays. One cat (0.16%) was positive for H5 antibodies and twenty cats (3.1%) were positive for SARS-CoV-2 antibodies in both binding and neutralization assays. These data suggest that cats in the Philadelphia area have not been routinely exposed to clade 2.3.4.4b H5N1 viruses but have been more commonly exposed to SARS-CoV-2. ImportanceHighly pathogenic H5NX avian influenza viruses have caused widespread disease and mortality in both wild and domestic birds, and have infected marine mammals, cattle, and humans. Recent studies have demonstrated that cats can also be infected with H5NX viruses, but it is unknown if these infections are common. In this report, we completed a seroprevalence study and found that H5NX exposure in feral and owned cats in the Philadelphia area appears to be rare. We also completed SARS-CoV-2 serological experiments and found that 3.1% of cats possessed antibodies reactive to this virus.

microbiology↗

bioRxiv2022

Characterization of Three Variants of SARS-CoV-2 in vivo Shows Host-Dependent Pathogenicity in Hamsters

Animal models are used in preclinical trials to test vaccines, antivirals, monoclonal antibodies, and immunomodulatory drug therapies against SARS-CoV-2. However, these drugs often do not produce equivalent results in human clinical trials. Here, we show how different animal models infected with some of the most clinically relevant SARS-CoV-2 variants, WA1/2020, B.1.617.2/Delta, B.1.1.529/Omicron and BA5.2/Omicron, have independent outcomes. We show that in mice, B.1.617.2 is more pathogenic, followed by WA1, while B.1.1.529 showed an absence of clinical signs. Only B.1.1.529 was able to infect C57BL/6J mice, which lack the human ACE2 receptor. B.1.1.529-infected ACE2 mice had different T cell profiles compared to infected K18-hACE2 mice, while viral shedding profiles and viral titers in lungs were similar between the ACE2 and the C57BL/6J mice. These data suggest B.1.1.529 virus adaptation to a new host and shows that asymptomatic carriers can accumulate and shed virus. Next, we show how B.1.617.2, WA1 and BA5.2/Omicron have similar viral replication kinetics, pathogenicity, and viral shedding profiles in hamsters, demonstrating that the increased pathogenicity of B.1.617.2 observed in mice is host-dependent. Overall, these findings suggest that small animal models are useful to parallel human clinical data, but the experimental design places an important role in interpreting the data. IMPORTANCEThere is a need to investigate SARS-CoV-2 variants phenotypes in different animal models due to the lack of reproducible outcomes when translating experiments to the human population. Our findings highlight the correlation of clinically relevant SARS-CoV-2 variants in animal models with human infections. Experimental design and understanding of correct animal models are essential to interpreting data to develop antivirals, vaccines, and other therapeutic compounds against COVID-19.

microbiology↗

bioRxiv2022

Genome-wide CRISPR screens of T cell exhaustion identify chromatin remodeling factors that limit T cell persistence

T cell exhaustion limits anti-tumor immunity, but the molecular determinants of this process remain poorly understood. Using a chronic antigen stimulation assay, we performed genome-wide CRISPR/Cas9 screens to systematically discover genetic regulators of T cell exhaustion, which identified an enrichment of epigenetic factors. In vivo CRISPR screens in murine and human tumor models demonstrated that perturbation of several epigenetic regulators, including members of the INO80 and BAF chromatin remodeling complexes, improved T cell persistence in tumors. In vivo paired CRISPR perturbation and single-cell RNA sequencing revealed distinct transcriptional roles of each complex and that depletion of canonical BAF complex members, including Arid1a, resulted in the maintenance of an effector program and downregulation of terminal exhaustion-related genes in tumor-infiltrating T cells. Finally, Arid1a-depletion limited the global acquisition of chromatin accessibility associated with T cell exhaustion and led to improved anti-tumor immunity after adoptive cell therapy. In summary, we provide a comprehensive atlas of the genetic regulators of T cell exhaustion and demonstrate that modulation of the epigenetic state of T cell exhaustion can improve T cell responses in cancer immunotherapy.

immunology↗

bioRxiv2021

An invariant Trypanosoma vivax vaccine antigen inducing protective immunity

Trypanosomes are protozoan parasites that cause infectious diseases including human African trypanosomiasis (sleeping sickness), and nagana in economically-important livestock animals1,2. An effective vaccine against trypanosomes would be an important control tool, but the parasite has evolved sophisticated immunoprotective mechanisms including antigenic variation3 that present an apparently insurmountable barrier to vaccination. Here we show using a systematic genome-led vaccinology approach4 and a murine model of Trypanosoma vivax infection5 that protective invariant subunit vaccine antigens can be identified. Vaccination with a single recombinant protein comprising the extracellular region of a conserved cell surface protein localised to the flagellum membrane termed "invariant flagellum antigen from T. vivax" (IFX) induced long-lasting protection. Immunity was passively transferred with immune serum, and recombinant monoclonal antibodies to IFX could induce sterile protection and revealed multiple mechanisms of antibody-mediated immunity, including a major role for complement. Our discovery identifies a vaccine candidate for an important parasitic disease that has constrained the socioeconomic development of sub-Saharan African countries6 and challenges long-held views that vaccinating against trypanosome infections cannot be achieved.

microbiology↗

bioRxiv2020

Subjugating Translational Inhibition in Response to Ribosomal Protein Insufficiency by a Herpesvirus Ribosome-Associated Protein

In addition to being required for protein synthesis, ribosomes and ribosomal proteins (RPs) also regulate mRNA translation in uninfected and virus-infected cells. By individually depleting 85 RPs using RNAi, we found overall protein synthesis in uninfected primary fibroblasts was more sensitive to RP-depletion than those infected with herpes simplex virus-1 (HSV-1). Although representative RP-depletion (uL3, uS4, uL5) inhibited protein synthesis in cells infected with other DNA viruses, HSV-1-infected cell protein synthesis unexpectedly endured and required a single virus-encoded gene product, VP22. During individual RP-insufficiency, VP22-expressing HSV-1 replicated better than a VP22-deficient variant. Furthermore, VP22 cosedimented with ribosomes and polyribosomes in infected cells. This identifies VP22 as a virus-encoded, polyribosome-associated protein that compensates for RP-insufficiency to support viral protein synthesis and replication. Moreover, it reveals an unanticipated class of virus-encoded, ribosome-associated effectors that reduce the dependence of protein synthesis upon RPs and broadly support translation during physiological stress such as infection.

microbiology↗

bioRxiv2020

The ΔF508 CFTR defect: molecular mechanism of suppressor mutation V510D and the contribution of transmembrane helix unraveling

Cystic fibrosis (CF) results from mutations within the gene encoding the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), a transmembrane chloride channel found on the apical surface of epithelial cells. The most common CF-causing mutation results in a deletion of phenylalanine 508 ({Delta}F508-CFTR), a residue normally found within the NBD1 domain. Loss of F508 causes NBD1 to be less thermodynamically stable and prevents proper tertiary folding of CFTR. As a result, CFTR is not properly trafficked to the cell surface. Recently, progress has been made towards the development of small molecule "correctors" that can restore CFTR tertiary structure and stabilize the channel to overcome the instability inherent in {Delta}F508-CFTR. However, the resultant improvement in channel activity has been modest, and the need for potent correctors remains. To fully inform such efforts, a better understanding of the molecular pathology associated with {Delta}F508-CFTR is required. Here we present a comprehensive study of the impact of F508 deletion on both purified NBD1 and full-length CFTR. Through the use of homology modeling, molecular dynamics simulations, mutational analysis, biochemical, biophysical and functional characterization studies, we obtained insight into how the {Delta}F508 mutation may lead to helical unraveling of transmembrane domains 10 and 11 (TM10, TM11), and how the known suppressor mutations V510D and R1070W, as well as novel second site suppressor mutations (SSSMs) identified in this work, may act to rescue {Delta}F508-CFTR maturation and trafficking.

biochemistry↗

bioRxiv2019

Variant antigen diversity in Trypanosoma vivax is not driven by recombination

African trypanosomes are vector-borne haemoparasites that cause African trypanosomiasis in humans and animals. Parasite survival in the bloodstream depends on immune evasion, achieved by antigenic variation of the Variant Surface Glycoprotein (VSG) coating the trypanosome cell surface. Recombination, or rather directed gene conversion, is fundamental in Trypanosoma brucei, as both a mechanism of VSG gene switching and of generating antigenic diversity during infections. Trypanosoma vivax is a related, livestock pathogen also displaying antigenic variation, but whose VSG lack key structures necessary for gene conversion in T. brucei. Thus, this study tests a long-standing prediction that T. vivax has a more restricted antigenic repertoire. Here we show that global VSG repertoire is broadly conserved across diverse T. vivax clinical strains. We use sequence mapping, coalescent approaches and experimental infections to show that recombination plays little, if any, role in diversifying T. vivax VSG sequences. These results explain interspecific differences in disease, such as propensity for self-cure, and indicate that either T. vivax has an alternate mechanism for immune evasion or else a distinct transmission strategy that reduces its reliance on long-term persistence. The lack of recombination driving antigenic diversity in T. vivax has immediate consequences for both the current mechanistic model of antigenic variation in African trypanosomes and species differences in virulence and transmission strategy, requiring us to reconsider the wider epidemiology of animal African trypanosomiasis.

microbiology↗