Scholarly Works, Center for Emerging, Zoonotic, and Arthropod-borne Pathogens (CeZAP)

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Obesity fosters severe disease outcomes in a mouse model of coronavirus infection associated with transcriptomic abnormalities
Rai, Pallavi; Marano, Jeffrey M.; Kang, Lin; Coutermarsh-Ott, Sheryl; Daamen, Andrea R.; Lipsky, Peter E.; Weger-Lucarelli, James (Wiley, 2024-04-01)
Obesity has been identified as an independent risk factor for severe outcomes in humans with coronavirus disease 2019 (COVID-19) and other infectious diseases. Here, we established a mouse model of COVID-19 using the murine betacoronavirus, mouse hepatitis virus 1 (MHV-1). C57BL/6 and C3H/HeJ mice exposed to MHV-1 developed mild and severe disease, respectively. Obese C57BL/6 mice developed clinical manifestations similar to those of lean controls. In contrast, all obese C3H/HeJ mice succumbed by 8 days postinfection, compared to a 50% mortality rate in lean controls. Notably, both lean and obese C3H/HeJ mice exposed to MHV-1 developed lung lesions consistent with severe human COVID-19, with marked evidence of diffuse alveolar damage (DAD). To identify early predictive biomarkers of worsened disease outcomes in obese C3H/HeJ mice, we sequenced RNA from whole blood 2 days postinfection and assessed changes in gene and pathway expression. Many pathways uniquely altered in obese C3H/HeJ mice postinfection aligned with those found in humans with severe COVID-19. Furthermore, we observed altered gene expression related to the unfolded protein response and lipid metabolism in infected obese mice compared to their lean counterparts, suggesting a role in the severity of disease outcomes. This study presents a novel model for studying COVID-19 and elucidating the mechanisms underlying severe disease outcomes in obese and other hosts.
Inter-epidemic seroprevalence of Rift Valley fever virus and associated risk factors in humans in Eastern Rwanda
Nsengimana, Isidore; Kelvin, David; Uwibambe, Evodie; Rwagasore, Edson; Muvunyi, Claude M.; Eastwood, Gillian; Chengula, Augustino A.; Kasanga, Christopher J. (Public Library of Science, 2025-08-01)
Background: Rift Valley fever (RVF) is a mosquito-borne zoonosis that causes periodic and explosive epizootics/epidemics in Africa and the Arabian Peninsula. In Rwanda, RVF virus (RVFV) circulation has resulted into two major outbreaks in 2018 and 2022, both of which involving humans. Information on the magnitude of human exposure to RVFV in the country is scarce. This cross-sectional study aimed to investigate the seroprevalence of RVFV and associated risk factors in humans in the Eastern province of Rwanda, 3 years after the end of the 2018 outbreak. Methodology: A total of 552 outpatients at health facilities in five districts of the Eastern province were randomly sampled and interviewed between December 2021 and February 2022. Exposure to RVFV was examined by detection of anti-RVFV IgG/IgM antibodies in serum samples using a competitive enzyme linked immunosorbent assay (c-ELISA). Bivariate and multivariate logistic regressions were used to assess the association between risk factors and RVFV seropositivity. Results: The findings revealed an overall seroprevalence of 9.6%. The highest seropositivity, but without significant difference, was observed in Bugesera district (12.9%), followed by Kayonza, (10.8%), Kirehe (8.6%), Rwamagana (7.0%) and Ngoma (6.8%). Odds of seropositivity were significantly higher in people with a history of slaughtering animals (OR 2.26, 95% CI 1.04-4.91, p=0.03), and milking (OR 2.60, 95% CI 1.23-5.49, p=0.012). Sex and age-related differences were not significant. Conclusion: This study is a first serological survey of RVFV spillover to humans in the country and shows that rural communities in Rwanda have been exposed to RVFV. These findings suggest the Eastern province of Rwanda as a potential hotspot for RVFV circulation, and emphasize the need for a countrywide One Health-based surveillance, prevention, and control strategy to minimize the effects of RVFV in the country.
Noninvasive Analysis of Peptidoglycan from Living Animals
Ocius, Karl L.; Kolli, Sree H.; Ahmad, Saadman S.; Dressler, Jules M.; Chordia, Mahendra D.; Jutras, Brandon L.; Rutkowski, Melanie R.; Pires, Marcos M. (American Chemical Society, 2024-04-09)
The role of the intestinal microbiota in host health is increasingly revealed in its contributions to disease states. The host-microbiome interaction is multifactorial and dynamic. One of the factors that has recently been strongly associated with host physiological responses is peptidoglycan from bacterial cell walls. Peptidoglycan from gut commensal bacteria activates peptidoglycan sensors in human cells, including the nucleotide-binding oligomerization domain-containing protein 2. When present in the gastrointestinal tract, both the polymeric form (sacculi) and depolymerized fragments can modulate host physiology, including checkpoint anticancer therapy efficacy, body temperature and appetite, and postnatal growth. To utilize this growing area of biology toward therapeutic prescriptions, it will be critical to directly analyze a key feature of the host-microbiome interaction from living hosts in a reproducible and noninvasive way. Here we show that metabolically labeled peptidoglycan/sacculi can be readily isolated from fecal samples collected from both mice and humans. Analysis of fecal samples provided a noninvasive route to probe the gut commensal community including the metabolic synchronicity with the host circadian clock. Together, these results pave the way for noninvasive diagnostic tools to interrogate the causal nature of peptidoglycan in host health and disease.
Once bitten, twice shy: A modeling framework for incorporating heterogeneous mosquito biting into transmission models
Dahlin, Kyle J. M.; Robert, Michael A.; Childs, Lauren M. (Springer, 2025-11-01)
The risk and intensity of mosquito-borne disease outbreaks are tightly linked to the frequency at which mosquitoes feed on blood, also known as the biting rate. However, standard models of mosquito-borne disease transmission inherently assume that mosquitoes bite only once per reproductive cycle—an assumption commonly violated in nature. Drivers of multiple biting, such as host defensive behaviors or climate factors, also affect the mosquito gonotrophic cycle duration (GCD), a quantity customarily used to estimate the biting rate. Here, we present a novel framework for incorporating more complex mosquito biting behaviors into transmission models. This framework can account for heterogeneity in and linkages between mosquito biting rates and multiple biting. We provide general formulas for the basic offspring number, N0, and basic reproduction number, R0, threshold measures for mosquito population and pathogen transmission persistence, respectively. To exhibit its flexibility, we expand on specific models derived from the framework that arise from empirical, phenomenological, or mechanistic modeling perspectives. Using the gonotrophic cycle duration as a standard quantity to make comparisons among the models, we show that assumptions about the biting process strongly affect the relationship between GCD and R0. While under the standard assumption of one bite per reproductive cycle, R0 is an increasing linear function of the inverse of the GCD, alternative models of the biting process can exhibit saturating or concave relationships. Critically, from a mechanistic perspective, decreases in the GCD can lead to substantial decreases in R0. Through sensitivity analysis of the mechanistic model, we determine that parameters related to probing and ingesting success are the most important targets for disease control. This work highlights the importance of incorporating the behavioral dynamics of mosquitoes into transmission models and provides a method for evaluating how individual-level interventions against mosquito biting scale up to determine population-level mosquito-borne disease risk.
Regioselective Annulation of 6-Carboxy-Substituted Pyrones as a Two-Carbon Unit in Formal [4+2] Cycloaddition Reactions
Kohanov, Zachary A.; Shuvo, Suzzudul Islam; Lowell, Andrew N. (American Chemical Society, 2024-06-13)
Heterocycles serve as a critical motif in chemistry, but despite being present in more than 85% of pharmaceuticals, there are limited methods for their construction. Here, we describe the incorporation of intact pyrone (2H-pyran-2-one) into larger ring systems via annulation. In a formal [4 + 2] cycloaddition, the pyrone regioselectively accepts a benzylic anion as a nucleophile in a conjugate addition fashion, with the subsequent pyrone-derived enolate attaching to a pendant ester on the initial nucleophile. Subsequent base-driven enolate formation and elimination establish aromaticity of the newly formed ring. After optimization of this process using an NMR-based assessment to overcome solubility and separation challenges, the reaction was successfully applied to a library of 6-ester and -amide-substituted pyrones and using a phenyl ester and other substituted sulfoxides. This technology enables the incorporation of intact pyrone rings into more complex systems, such as for the total synthesis of the natural product thermorubin.
Competitive Binding of Viral Nuclear Localization Signal Peptide and Inhibitor Ligands to Importin-α Nuclear Transport Protein
Delfing, Bryan M.; Laracuente, Xavier E.; Jeffries, William; Luo, Xingyu; Olson, Audrey; Foreman, Kenneth W.; Petruncio, Greg; Lee, Kyung Hyeon; Paige, Mikell; Kehn-Hall, Kylene; Lockhart, Christopher; Klimov, Dmitri K. (American Chemical Society, 2024-06-13)
Venezuelan equine encephalitis virus (VEEV) is a highly virulent pathogen whose nuclear localization signal (NLS) sequence from capsid protein binds to the host importin-alpha transport protein and blocks nuclear import. We studied the molecular mechanisms by which two small ligands, termed I1 and I2, interfere with the binding of VEEV's NLS peptide to importin-alpha protein. To this end, we performed all-atom replica exchange molecular dynamics simulations probing the competitive binding of the VEEV coreNLS peptide and I1 or I2 ligand to the importin-alpha major NLS binding site. As a reference, we used our previous simulations, which examined noncompetitive binding of the coreNLS peptide or the inhibitors to importin-alpha. We found that both inhibitors completely abrogate the native binding of the coreNLS peptide, forcing it to adopt a manifold of nonnative loosely bound poses within the importin-alpha major NLS binding site. Both inhibitors primarily destabilize the native coreNLS binding by masking its amino acids rather than competing with it for binding to importin-alpha. Because I2, in contrast to I1, binds off-site localizing on the edge of the major NLS binding site, it inhibits fewer coreNLS native binding interactions than I1. Structural analysis is supported by computations of the free energies of the coreNLS peptide binding to importin-alpha with or without competition from the inhibitors. Specifically, both inhibitors reduce the free energy gain from coreNLS binding, with I1 causing significantly larger loss than I2. To test our simulations, we performed AlphaScreen experiments measuring IC50 values for both inhibitors. Consistent with in silico results, the IC50 value for I1 was found to be lower than that for I2. We hypothesize that the inhibitory action of I1 and I2 ligands might be specific to the NLS from VEEV's capsid protein.
The PTP4A3 inhibitor KVX-053 reduces Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) virulence, inflammation, and development of acute lung injury in K18-hACE2 mice
Colunga-Biancatelli, Ruben M. L.; Solopov, Pavel A.; Woodson, Caitlin M.; Allen, Irving C.; Akhrymuk, Ivan; Akhrymuk, Maryna; Heath, Brittany N.; Ivester, Hannah M.; Day, Tierney; Austin, Dan E.; Kehn-Hall, Kylene; Lazo, John S.; Sharlow, Elizabeth R.; Catravas, John D. (2025-10-28)
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has caused a global health crisis, marked by high transmissibility and virulence. Despite widespread vaccination efforts, effective treatments for COVID-19, particularly for severe cases leading to Acute Respiratory Distress Syndrome (ARDS), remain limited. This study investigates the efficacy of KVX-053, a protein tyrosine phosphatase type IVA (PTP4A3) small molecule inhibitor, in modulating SARS-CoV-2-induced inflammation and lung injury using in vitro cell models and in vivo K18-hACE2 transgenic mice. KVX-053 reduced in vitro pro-inflammatory cytokine expression, including TNFα, CXCL10, and CXCL11, without impacting viral replication or cell viability. K18-hACE2 mice treated with KVX-053 demonstrated marked improvement in clinical scores and reduced histological evidence of lung injury compared to untreated SARS-CoV-2-infected controls. KVX-053 mitigated the activation of key inflammatory mediators in the lung, including NLRP3 inflammasomes, IL-6, and phosphorylated STAT3, effectively curbing the “cytokine storm” associated with severe COVID-19. Importantly, treatment preserved lung parenchymal integrity, reduced edema, and minimized macrophage infiltration. Our findings highlight PTP4A3 as a potential critical regulator of the inflammatory response during SARS-CoV-2 infection. KVX-053, a potent and selective PTP4A3 inhibitor, emerges as a promising host-directed therapeutic strategy for mitigating ARDS and inflammation-driven lung injury in SARS-CoV-2 and potentially other respiratory viral infections. Future studies are required to optimize dosing strategies, elucidate precise molecular mechanisms, and validate these findings in clinical settings.
Immune Evasion by the NSs Protein of Rift Valley Fever Virus: A Viral Houdini Act
Petraccione, Kaylee; Omichinski, James G.; Kehn-Hall, Kylene (MDPI, 2025-10-21)
Rift Valley fever virus (RVFV) is a negative-sense arbovirus that causes several severe diseases, including hemorrhagic fever in ruminants and humans. There are currently no FDA-approved vaccines or therapeutics for RVFV. The viral nonstructural protein NSs acts like a molecular Harry Houdini, the world-famous escape artist, to help the virus evade the host’s innate immune response and serves as the main virulence factor of RVFV. In this review, we discuss the molecular mechanisms by which NSs interacts with multiple factors to modulate host processes, evade the host immune response, and facilitate viral replication. The impact of NSs mutations that cause viral attenuation is also discussed. Understanding the molecular mechanisms by which NSs evades the host innate immune response is crucial for developing novel therapeutics and vaccines targeting RVFV.
Obesity's Unexpected Influence: Reduced Alphavirus Transmission and Altered Immune Activation in the Vector
Rai, Pallavi; Webb, Emily M.; Paulson, Sally L.; Kang, Lin; Weger-Lucarelli, James (Wiley, 2024-11-01)
Chikungunya virus (CHIKV) and Mayaro virus (MAYV) are emerging/re-emerging alphaviruses transmitted by Aedes spp. mosquitoes and responsible for recent disease outbreaks in the Americas. The capacity of these viruses to cause epidemics is frequently associated with increased mosquito transmission, which in turn is governed by virus-host-vector interactions. Although many studies have explored virus-vector interactions, significant gaps remain in understanding how vertebrate host factors influence alphavirus transmission by mosquitoes. We previously showed that obesity, a ubiquitous vertebrate host biological factor, reduces alphavirus transmission potential in mosquitoes. We hypothesized that alphavirus-infected obese bloodmeals altered immune genes and/or pathways in mosquitoes, thereby inhibiting virus transmission. To test this, we conducted RNA sequencing (RNA-seq) and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) on midgut RNA from mosquitoes fed on alphavirus-infected lean and obese mice. This approach aimed to identify potential antiviral or proviral genes and pathways altered in mosquitoes after consuming infected obese bloodmeals. We found upregulation of the Toll pathway and downregulation of several metabolic and other genes in mosquitoes fed on alphavirus-infected obese bloodmeals. Through gene knockdown studies, we demonstrated the antiviral role of Toll pathway and proviral roles of AAEL009965 and fatty acid synthase (FASN) in the transmission of alphaviruses by mosquitoes. Therefore, this study utilized obesity to identify factors influencing alphavirus transmission by mosquitoes and this research approach may pave the way for designing broadly effective antiviral measures to combat mosquito-borne viruses, such as releasing transgenic mosquitoes deficient in the identified genes.
Highly Active Oligoethylene Glycol Pleuromutilins via Systematic Linker Synthesis/One-Pot Attachment and a Microscale Solubility Method
Breiner, Logan M.; Slowinski, Roman P.; Lowell, Andrew N. (American Chemical Society, 2024-12-18)
The semisynthetic derivatization of natural products is crucial for their continued development as antibiotics. While commercial pleuromutilin derivatives depend on amines for solubility, we demonstrate the high activity and solubility of oligoethylene glycol-substituted pleuromutilins achieved via a one-pot deprotection/attachment approach using thiolates protected as thioesters. The bifunctional linker synthesis is versatile and can be broadly applied to other chemistries. Antibacterial assays revealed this simple glycolate modification enhanced inhibition 4-8-fold relative to that of pleuromutilin. A new microscale solubility method is also introduced.
Larval environment reshapes mosquito disease risk via phenotypic and molecular plasticity
Chandrasegaran, Karthikeyan; Walker, Melody; Marano, Jeffrey M.; Rami, Spruha; Bisese, Adaline; Weger-Lucarelli, James; Lahondère, Chloé; Robert, Michael A.; Childs, Lauren M.; Vinauger, Clément (2025-06-21)
Early-life environmental conditions can exert profound, lasting effects on adult phenotypes, with major consequences for fitness and disease transmission, especially in holometabolous insects like mosquitoes, which are a key vector species. Yet, the molecular mechanisms through which juvenile environments shape adult physiology and behavior via transstadial effects remain largely unresolved. Here, we demonstrate that larval competition, a key ecological stressor, profoundly alters adult body size, survival, reproductive output, host-seeking behavior, olfactory neurophysiology, and vector competence in the mosquito Aedes aegypti. Crucially, using transcriptomic profiling and integrative network analyses, we identify seven regulatory hub genes whose expression is strongly associated with size-dependent variation in olfactory behavior, reproductive investment, and Zika virus transmission potential. These hub genes belong to gene modules enriched for functions in chemosensory processing, metabolic regulation, and signal transduction, revealing a molecular framework mediating environmentally induced plasticity across metamorphosis. Integrating these empirical findings into a transmission model, we show that incomplete larval control can inadvertently increase outbreak risk by producing larger, longer-lived, and more competent vectors. Our results uncover molecular mechanisms underpinning phenotypic plasticity in disease vectors and highlight the critical need to account for transstadial effects in models of vector-borne disease transmission.
Guinea Pigs Are Not a Suitable Model to Study Neurological Impacts of Ancestral SARS-CoV-2 Intranasal Infection
Joyce, Jonathan D.; Moore, Greyson A.; Thompson, Christopher K.; Bertke, Andrea S. (MDPI, 2025-05-15)
Neurological symptoms involving the central nervous system (CNS) and peripheral nervous system (PNS) are common complications of acute COVID-19 as well as post-COVID conditions. Most research into these neurological sequalae focuses on the CNS, disregarding the PNS. Guinea pigs were previously shown to be useful models of disease during the SARS-CoV-1 epidemic. However, their suitability for studying SARS-CoV-2 has not been experimentally demonstrated. To assess the suitability of guinea pigs as models for SARS-CoV-2 infection and the impact of SARS-CoV-2 infection on the PNS, and to determine routes of CNS invasion through the PNS, we intranasally infected wild-type Dunkin-Hartley guinea pigs with ancestral SARS-CoV-2 USA-WA1/2020. We assessed PNS sensory neurons (trigeminal ganglia, dorsal root ganglia), autonomic neurons (superior cervical ganglia), brain regions (olfactory bulb, brainstem, cerebellum, cortex, hippocampus), lungs, and blood for viral RNA (RT-qPCR), protein (immunostaining), and infectious virus (plaque assay) at three- and six-days post infection. We show that guinea pigs, which have previously been used as a model of SARS-CoV-1 pulmonary disease, are not susceptible to intranasal infection with ancestral SARS-CoV-2, and are not useful models in assessing neurological impacts of infection with SARS-CoV-2 isolates from the early pandemic.
Neurological manifestations of encephalitic alphaviruses, traumatic brain injuries, and organophosphorus nerve agent exposure
VanderGiessen, Morgen; de Jager, Caroline; Leighton, Julia; Xie, Hehuang; Theus, Michelle H.; Johnson, Erik; Kehn-Hall, Kylene (Frontiers, 2024-12-13)
Encephalitic alphaviruses (EEVs), Traumatic Brain Injuries (TBI), and organophosphorus nerve agents (NAs) are three diverse biological, physical, and chemical injuries that can lead to long-term neurological deficits in humans. EEVs include Venezuelan, eastern, and western equine encephalitis viruses. This review describes the current understanding of neurological pathology during these three conditions, provides a comparative review of case studies vs. animal models, and summarizes current therapeutics. While epidemiological data on clinical and pathological manifestations of these conditions are known in humans, much of our current mechanistic understanding relies upon animal models. Here we review the animal models findings for EEVs, TBIs, and NAs and compare these with what is known from human case studies. Additionally, research on NAs and EEVs is limited due to their classification as high-risk pathogens (BSL-3) and/or select agents; therefore, we leverage commonalities with TBI to develop a further understanding of the mechanisms of neurological damage. Furthermore, we discuss overlapping neurological damage mechanisms between TBI, NAs, and EEVs that highlight novel medical countermeasure opportunities. We describe current treatment methods for reducing neurological damage induced by individual conditions and general neuroprotective treatment options. Finally, we discuss perspectives on the future of neuroprotective drug development against long-term neurological sequelae of EEVs, TBIs, and NAs.
Hantavirus in rodents in the United States: Temporal and spatial trends and report of new hosts
Astorga, Francisca; Alkishe, Abdelghafar; Paansri, Paanwaris; Mantilla, Gabriel; Escobar, Luis E. (Wiley, 2025-03-16)
In North America, the rodent-borne hantavirus pulmonary syndrome ispredominantly caused by the Sin Nombre virus, typically associated with the deer mouse Peromyscus maniculatus. Utilizing data from the National Ecological Observatory Network (NEON) hantavirus program, we assessed factors that may influence the spatial and temporal distribution of hantavirus inrodent populations across the United States. Between 2014 and 2019, the NEON hantavirus program conducted 104,379 small mammal captures and collected 14,004 blood samples from 49 species at 45 field sites. Our study identified 296 seropositive samples across 15 rodent species, including 8 Peromyscusspecies. We describe six new species with hantavirus seropositive samples not previously reported as hantavirus hosts. The highest number of seropositivesamples was obtained from Pe. maniculatus (n = 116; 2.9% seroprevalence),followed by Peromyscus leucopus (n = 96; 2.8%) and Microtus pennsylvanicus(n = 33; 4.2%). Hantavirus seroprevalence showed an uneven spatial distribution, with the highest seroprevalence found in Virginia (7.8%, 99 seropositivesamples), Colorado (5.7%, n = 37), and Texas (4.8%, n = 19). Hantavirus sero-positive samples were obtained from 32 sites, 10 of which presented seropositive samples in species other than Pe. maniculatus or Pe. leucopus. Seroprevalence was inconsistent across years but showed intra-annual bimodal trends, and in Pe. maniculatus and Pe. leucopus, the number of captures correlated with sero-prevalence in the following months. Seroprevalence was higher in adult males, with only one seropositive sample obtained from a juvenile Peromyscus truei. Higher body mass, presence of scrotal testes, and nonpregnant status were associated with higher seropositivity. The NEON dataset, derived from a multiyear and structured surveillance system, revealed the extensive distribution of hantavirus across broad taxonomic and environmental ranges. Future research should consider winter season surveillance and continued analyses of stored samples for a comprehensive spatiotemporal study of hantavirus circulation in wildlife. Global changes are expected to affect the dynamics of rodent populations by affecting their availability of resources and demography and, consequently, may modify transmission rates of rodent-borne zoonotic pathogens such as hantavi-rus. This study can be considered a baseline to assess hantavirus patterns across host taxa, geographies, and seasons in the United States.
Evolution at Spike protein position 519 in SARS-CoV-2 facilitated adaptation to humans
Cereghino, C.; Michalak, K.; DiGiuseppe, S.; Yu, D.; Faraji, A.; Sharp, A.K.; Brown, Anne M.; Kang, L.; Weger-Lucarelli, James; Michalak, P. (Springer Nature, 2024)
As the COVID-19 pandemic enters its fourth year, the pursuit of identifying a progenitor virus to SARSCoV- 2 and understanding the mechanism of its emergence persists, albeit against the backdrop of intensified efforts to monitor the ongoing evolution of the virus and the influx of new mutations. Surprisingly, few residues hypothesized to be essential forSARS-CoV-2 emergence and adaptation to humans have been validated experimentally, despite the importance that these mutations could contribute to the development of effective antivirals. To remedy this,we searched for genomic regions in the SARS-CoV-2 genome that show evidence of past selection around residues unique to SARSCoV- 2 compared with closely related coronaviruses. In doing so, we identified a residue at position 519 in Spike within the receptor binding domain that holds a static histidine in human-derived SARSCoV- 2 sequences but an asparagine in SARS-related coronaviruses from bats and pangolins. In experimental validation, the SARS-CoV-2 Spike protein mutant carrying the putatively ancestral H519N substitution showed reduced replication in human lung cells, suggesting that the histidine residue contributes to viral fitness in the human host. Structural analyses revealed a potential role of Spike residue 519 in mediating conformational transitions necessary for Spike prior to binding with ACE2. Pseudotyped viruses bearing the putatively ancestral N519 also demonstrated significantly reduced infectivity in cells expressing the human ACE2 receptor compared to H519. ELISA data corroborated that H519 enhances Spike binding affinity to the human ACE2 receptor compared to the putatively ancestral N519. Collectively, these findings suggest that the evolutionary transition at position 519 of the Spike protein played a critical role inSARS-CoV-2 emergence and adaptation to the human host. Additionally, this residue presents as a potential drug target for designing small molecule inhibitors tailored to this site.
Neuropathogenesis of Encephalitic Alphaviruses in Non-Human Primate and Mouse Models of Infection
Woodson, Caitlin M.; Carney, Shannon K.; Kehn-Hall, Kylene (MDPI, 2025-02-14)
Encephalitic alphaviruses, including eastern, Venezuelan, and western equine encephalitis virus (EEEV, VEEV, and WEEV, respectively) are New World alphaviruses primarily transmitted by mosquitos that cause debilitating and lethal central nervous system (CNS) disease in both humans and horses. Despite over one hundred years of research on these viruses, the underpinnings of the molecular mechanisms driving virally induced damage to the CNS remain unresolved. Moreover, virally induced encephalitis following exposure to these viruses causes catastrophic damage to the CNS, and survivors of infection often suffer from permanent neurological sequelae as a result of sustained neuroinflammation and neurological insults encountered. Animal models are undoubtedly invaluable tools in biomedical research, where physiologically relevant models are required to study pathogenesis and host–pathogen interactions. Here, we review the literature to examine nonhuman primate (NHP) and mouse models of infection for EEEV, VEEV, and WEEV. We provide a brief overview of relevant background information for each virus, including geography, epidemiology, and clinical disease. The primary focus of this review is to describe neuropathological features associated with CNS disease in NHP and mouse models of infection and compare CNS invasion and neuropathogenesis for aerosol, intranasal, and subcutaneous routes of exposure to EEEV, VEEV, and WEEV.
Increased risks of mosquito-borne disease emergence in temperate regions of South America
Estallo, Elizabet L.; Lopez, Maria Soledad; Luduena-Almeida, Francisco; Madelon, Magali I.; Layun, Federico; Robert, Michael A. (Elsevier, 2024-11-14)
Identification of Potential Vectors and Detection of Rift Valley Fever Virus in Mosquitoes Collected Before and During the 2022 Outbreak in Rwanda
Nsengimana, Isidore; Hakizimana, Emmanuel; Mupfasoni, Jackie; Hakizimana, Jean Nepomuscene; Chengula, Augustino A.; Kasanga, Christopher J.; Eastwood, Gillian (MDPI, 2025-01-08)
Rift Valley fever virus (RVFV) is an emerging mosquito-borne arbovirus of One Health importance that caused two large outbreaks in Rwanda in 2018 and 2022. Information on vector species with a role in RVFV eco-epidemiology in Rwanda is scarce. Here we sought to identify potential mosquito vectors of RVFV in Rwanda, their distribution and abundance, as well as their infection status. Since an outbreak of RVF occurred during the study period, data were obtained both during an interepidemic period and during the 2022 Rwanda RVF outbreak. Five districts of the eastern province of Rwanda were prospected using a combination of unbaited light traps and Biogents (BG Sentinel and Pro) traps baited with an artificial human scent during three periods, namely mid-August to mid-September 2021, December 2021, and April to May 2022. Trapped mosquitoes were morphologically identified and tested for viral evidence using both RT-PCR and virus isolation methods on a Vero cell line. A total of 14,815 adult mosquitoes belonging to five genera and at least 17 species were collected and tested as 765 monospecific pools. Culex quinquefasciatus was the most predominant species representing 72.7% of total counts. Of 527 mosquito pools collected before the 2022 outbreak, a single pool of Cx. quinquefasciatus showed evidence of RVFV RNA. Of 238 pools collected during the outbreak, RVFV was detected molecularly from five pools (two pools of Cx. quinquefasciatus, two pools of Anopheles ziemanni, and one pool of Anopheles gambiae sensu lato), and RVFV was isolated from the two pools of Cx. quinquefasciatus, from Kayonza and Rwamagana districts, respectively. Minimum infection rates (per 1000 mosquitoes) of 0.4 before the outbreak and 0.6–7 during the outbreak were noted. Maximum-likelihood phylogenetic analysis indicates that RVFV detected in these mosquitoes is closely related to viral strains that circulated in livestock in Rwanda and in Burundi during the same RVF outbreak in 2022. The findings reveal initial evidence for the incrimination of several mosquito species in the transmission of RVFV in Rwanda and highlight the need for more studies to understand the role of each species in supporting the spread and persistence of RVFV in the country.
A Phage-Based Approach to Identify Antivirulence Inhibitors of Bacterial Type IV Pili
Shimozono, Tori M.; Vogelaar, Nancy J.; O'Hara, Megan T.; Yang, Zhaomin (Wiley, 2025-01-17)
The increasing threat of antibiotic resistance underscores the urgent need for innovative strategies to combat infectious diseases, including the development of antivirulants. Microbial pathogens rely on their virulence factors to initiate and sustain infections. Antivirulants are small molecules designed to target virulence factors, thereby attenuating the virulence of infectious microbes. The bacterial type IV pilus (T4P), an extracellular protein filament that depends on the T4P machinery (T4PM) for its biogenesis, dynamics and function, is a key virulence factor in many significant bacterial pathogens. While the T4PM presents a promising antivirulence target, the systematic identification of inhibitors for its multiple protein constituents remains a considerable challenge. Here we report a novel high-throughput screening (HTS) approach for discovering T4P inhibitors. It uses Pseudomonas aeruginosa, a high-priority pathogen, in combination with its T4P-targeting phage, φKMV. Screening of a library of 2168 compounds using an optimised protocol led to the identification of tuspetinib, based on its deterrence of the lysis of P. aeruginosa by φKMV. Our findings show that tuspetinib also inhibits two additional T4P-targeting phages, while having no effect on a phage that recognises lipopolysaccharides as its receptor. Additionally, tuspetinib impedes T4P-mediated motility in P. aeruginosa and Acinetobacter species without impacting growth or flagellar motility. This bacterium-phage pairing approach is applicable to a broad range of virulence factors that are required for phage infection, paving ways for the development of advanced chemotherapeutics against antibiotic-resistant infections.
Identification of small molecule inhibitors of the Chloracidobacterium thermophilum type IV pilus protein PilB by ensemble virtual screening
McDonald-Ramos, Jay S.; Hicklin, Ian K.; Yang, Zhaomin; Brown, Anne M. (Elsevier, 2024-08-16)
Antivirulence strategy has been explored as an alternative to traditional antibiotic development. The bacterial type IV pilus is a virulence factor involved in host invasion and colonization in many antibiotic resistant pathogens. The PilB ATPase hydrolyzes ATP to drive the assembly of the pilus filament from pilin subunits. We evaluated Chloracidobacterium thermophilum PilB (CtPilB) as a model for structure-based virtual screening by molecular docking and molecular dynamics (MD) simulations. A hexameric structure of CtPilB was generated through homology modeling based on an existing crystal structure of a PilB from Geobacter metallireducens. Four representative structures were obtained from molecular dynamics simulations to examine the conformational plasticity of PilB and improve docking analyses by ensemble docking. Structural analyses after 1 μs of simulation revealed conformational changes in individual PilB subunits are dependent on ligand presence. Further, ensemble virtual screening of a library of 4234 compounds retrieved from the ZINC15 database identified five promising PilB inhibitors. Molecular docking and binding analyses using the four representative structures from MD simulations revealed that top-ranked compounds interact with multiple Walker A residues, one Asp-box residue, and one arginine finger, indicating these are key residues in inhibitor binding within the ATP binding pocket. The use of multiple conformations in molecular screening can provide greater insight into compound flexibility within receptor sites and better inform future drug development for therapeutics targeting the type IV pilus assembly ATPase.

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