Biology subjects

Butt, S. L.

Publications and source records attributed to Butt, S. L..

7 recordsLinked to original sources

bioRxiv2026

Hematogenous neuroinvasion and genotype-dependent transmission of influenza A H5N1 viruses in the cat host

The spillover of highly pathogenic avian influenza (HPAI) A H5N1 virus to mammalian hosts raises major concerns due to its pandemic potential. Cats are frequently affected mammals, often succumbing to systemic and neurological disease. Here, we characterized the pathogenesis and transmissibility of two H5N1 genotypes, B3.13 and D1.1, in cats. Infected cats exhibited high-level viremia and virus shedding in nasal, oral, and fecal secretions were consistently detected. The virus replicated initially in the upper respiratory tract and lungs, followed by systemic dissemination and neuroinvasion. Notably, the virus crossed the blood-brain-barrier by infecting endothelial cells, spreading to astrocytes and neurons, causing multifocal encephalitis. D1.1-virus infection caused protracted disease with lower shedding and no transmissibility, whereas B3.13 virus caused rapid onset with efficient shedding and transmission. These findings reveal critical H5N1 neuropathogenesis mechanisms and highlight mammalian transmission potential in a species with close human contact.

microbiology↗

bioRxiv2025

The ORF6 accessory protein contributes to SARS-CoV-2 virulence and pathogenicity in the naturally susceptible feline model of infection

Here the infection dynamics, replication, and pathogenicity of a recombinant virus containing a deletion of ORF6 (rWA1{Delta}ORF6) on the backbone of the highly virulent SARS-CoV-2 WA1 virus (rWA1) were investigated and compared to the parental rWA1 virus. While both rWA1 and rWA1{Delta}ORF6 viruses replicated efficiently in cultured cells, the rWA1{Delta}ORF6 virus produced smaller plaques, suggesting reduced cell-to-cell spread. Luciferase reporter assays revealed immune suppressing effects of ORF6 on interferon and nuclear factor kappa B (NF-{kappa}B) signaling pathways. Pathogenesis assessment in cats revealed that animals inoculated with rWA1 were lethargic and presented with fever on days 2 and 4 post-infection (pi), whereas rWA1{Delta}ORF6-inoculated animals developed subclinical infection. Additionally, animals inoculated with rWA1{Delta}ORF6 presented reduced infectious virus shedding in nasal and oral secretions and broncho-alveolar lavage fluid when compared with the rWA1-inoculated cats. Similarly, the rWA1{Delta}ORF6-inoculated cats presented reduced virus replication in the respiratory tract as evidenced by lower viral loads and reduced lung inflammation on day 3 and 5 pi when compared to rWA1-inoculated animals. Host gene transcriptomic analysis revealed marked differences in differentially expressed genes (DEG) in the nasal turbinate of animals infected with rWA1 when compared to rWA1{Delta}ORF6. Importantly, type I interferon signaling was significantly upregulated in rWA1{Delta}ORF6 infected cats when compared to rWA1-inoculated animals, which is correlated to the reduced replication of rWA1{Delta}ORF6 in the upper and lower respiratory tracts of infected animals. Collectively, these results demonstrate that the SARS-CoV-2 ORF6 is an important virulence determinant of the virus contributing to the modulation of host antiviral immune responses. IMPORTANCESARS-CoV-2 encodes several proteins that inhibit host interferon responses. The accessory protein ORF6 antagonizes interferon signaling by blocking the nucleocytoplasmic trafficking of key transcription factors. In this study, we showed that ORF6 plays an important role in SARS-CoV-2 pathogenesis. While both rWA1 and rWA1{Delta}ORF6 viruses replicated efficiently in cell culture, the rWA1{Delta}ORF6 presented impaired cell-to-spread and reduced innate immune inhibition as compared to the parental rWA1. Pathogenicity study in the feline model revealed an attenuated phenotype of the rWA1{Delta}ORF6 indicating that the ORF6 is a major virulence determinant of SARS-CoV-2. These results also demonstrate that ORF6 contributed to the feline host range of SARS-CoV-2.

microbiology↗

bioRxiv2024

From birds to mammals: spillover of highly pathogenic avian influenza H5N1 virus to dairy cattle led to efficient intra- and interspecies transmission

Infections with the highly pathogenic avian influenza (HPAI) H5N1 clade 2.3.4.4b virus have resulted in the death of millions of domestic birds and thousands of wild birds in the U.S. since January, 20221-4 Throughout this outbreak, spillovers of the virus to mammals have been frequently documented5-12. Here, we report the detection of HPAI H5N1 virus in dairy cattle herds across several states in the U.S. The affected cows displayed clinical signs encompassing decreased feed intake, altered fecal consistency, respiratory distress, and decreased milk production with abnormal milk. Infectious virus and RNA were consistently detected in milk collected from affected cows. Viral staining in tissues revealed a distinct tropism of the virus for the epithelial cells lining the alveoli of the mammary gland in cows. Analysis of whole genome sequences obtained from dairy cows, birds, domestic cats, and a racoon from affected farms indicated multidirectional interspecies transmissions. Epidemiologic and genomic data revealed efficient cow-to-cow transmission after healthy cows from an affected farm were transported to a premise in a different state. These results demonstrate the transmission of HPAI H5N1 clade 2.3.4.4b virus at a non-traditional interface and to a new and highly relevant livestock species, underscoring the ability of the virus to cross species barriers.

microbiology↗

bioRxiv2022

White-tailed deer (Odocoileus virginianus) may serve as a wildlife reservoir for nearly extinct SARS-CoV-2 variants of concern

The spillover of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from humans into white-tailed deer (WTD) and its ability to transmit from deer-to-deer raised concerns about the role of WTD in the epidemiology and ecology of the virus. In the present study, we conducted a comprehensive investigation to assess the prevalence, genetic diversity, and evolution of SARS-CoV-2 in WTD in the State of New York (NY). A total of 5,462 retropharyngeal lymph node (RPLN) samples collected from free-ranging hunter-harvested WTD during the hunting seasons of 2020 (Season 1, September-December 2020, n=2,700) and 2021 (Season 2, September-December 2021, n=2,762) were tested by SARS-CoV-2 real-time RT-PCR. SARS-CoV-2 RNA was detected in 17 samples (0.6%) from Season 1 and in 583 (21.1%) samples from Season 2. Hotspots of infection were identified in multiple confined geographic areas of NY. Sequence analysis of SARS-CoV-2 genomes from 164 samples demonstrated the presence multipls SARS-CoV-2 lineages as well as the co-circulation of three major variants of concern (VOCs) (Alpha, Gamma, and Delta) in WTD. Our analysis suggests the occurrence of multiple spillover events (human-to-deer) of the Alpha and Delta lineages with subsequent deer-to-deer transmission of the viruses. Detection of Alpha and Gamma variants in WTD long after their broad circulation in humans in NY suggests that WTD may serve as a wildlife reservoir for VOCs no longer circulating in humans. Thus, implementation of continuous surveillance programs to monitor SARS-CoV-2 dynamics in WTD are warranted, and measures to minimize virus transmission between humans and animals are urgently needed. SIGNIFICANCEWhite-tailed deer (WTD) are highly susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and are known to efficiently transmit the virus to other susceptible animals. Evidence of natural exposure or infection of wild WTD in North America raised significant concerns about their role on the ecology of the virus and its impact on the control of the coronavirus disease 2019 (COVID-19) pandemic. This comprehensive study demonstrates widespread infection of SARS-CoV-2 in the WTD populations across the State of New York. Additionally, we showed co-circulation of three major SARS-CoV-2 variants of concern (VOCs) in this wildlife population, long after their broad circulation in humans. These findings indicate that WTD - the most abundant large mammal in North America - may serve as a reservoir for variant SARS-CoV-2 strains that no longer circulate in the human population.

microbiology↗

bioRxiv2021

Putative novel avian paramyxoviruses identified from wild bird surveillance (United States, 2016?2018) and retrospective analysis of APMV-2s and APMV-6s in GenBank

Avian paramyxoviruses (APMVs) (subfamily Avulavirinae) have been isolated from over 200 species of wild and domestic birds from around the world. The International Committee on Taxonomy of Viruses (ICTV) currently defines 22 different APMV species, with Avian orthoavulavirus 1 (whose viruses are designated as APMV-1) being the most frequently studied due to its economic burden to the poultry industry. Less is known about other APMV species, including limited knowledge on the genetic diversity in wild birds and there is a paucity of public whole genome sequences for APMV-2 to -22. The goal of this study was to use MinION sequencing to genetically characterize APMVs isolated from wild bird swab samples collected during 2016-2018 in the United States. Multiplexed MinION libraries were prepared using a random strand-switching approach using 37 egg-cultured, influenza-negative, hemagglutination-positive samples. Thirty-five APMV isolates that had complete polymerase coding sequences were speciated using ICTVs current Paramyxoviridae phylogenetic methodology. Viruses from APMV-1, -4, -6, -8 were classified, one putative novel species (Avian orthoavulavirus 23) was identified from viruses isolated in this study, two putative new APMV species (Avian metaavulavirus 24 and 27) were identified from viruses isolated in this study and from retrospective GenBank sequences, and two putative new APMV species (Avian metaavulavirus 25 and 26) were identified solely from retrospective GenBank sequences. Furthermore, co-infections of APMVs were identified in a subset of the samples. The potential limitations of the branch length being the only speciation criterion and the potential benefit of a group pairwise distance analysis are discussed. ImportanceMost species of APMVs are understudied and/or underreported and many species were incidentally identified from asymptomatic wild birds; however, the disease significance of APMVs in wild birds is not fully determined. The rapid rise in high-throughput sequencing coupled with avian influenza surveillance programs have identified 12 different APMV species in the last decade and have challenged the resolution of classical serological methods to identify new viral species. Currently, ICTVs only criterion for Paramyxoviridae species classification is the requirement of a branch length >0.03 using a phylogenetic tree constructed from polymerase (L) amino acid sequences. The results from this study identify one new APMV species, propose four additional new APMV species, and highlight that the criterion may have insufficient resolution for APMV species demarcation and that refinement or expansion of this criterion may need to be established for Paramyxoviridae speciation.

microbiology↗

bioRxiv2019

Real-time, MinION-based, amplicon sequencing for lineage typing of infectious bronchitis virus from upper respiratory samples

Infectious bronchitis (IB) causes significant economic losses in the global poultry industry. Control of infectious bronchitis is hindered by the genetic diversity of the causative agent, infectious bronchitis virus (IBV), which has led to the emergence of several serotypes that lack complete serologic cross-protection. While serotyping by definition requires immunologic characterization, genotyping is an efficient means to identify IBVs detected in samples. Sanger sequencing of the S1 subunit of the spike gene is currently used to genotype IBV; however, the universal S1 PCR was created to work from cultured IBV and it is inefficient at detecting mixed isolates. This paper describes a MinION-based AmpSeq method that genetically typed IBV from clinical samples, including samples with multiple isolates. Total RNA was extracted from fifteen tracheal scrapings and choanal cleft swab samples, randomly reverse transcribed, and PCR amplified using modified S1-targeted primers. Amplicons were barcoded to allow for pooling of samples, processed per manufacturers instructions into a 1D MinION sequencing library, and sequenced on the MinION. The AmpSeq method detected IBV in 13 of 14 IBV-positive samples. AmpSeq accurately detected and genotyped both IBV lineages in three of five samples containing two IBV lineages. Additionally, one sample contained three IBV lineages, and AmpSeq accurately detected two of the three. Strain identification, including detection of different strains from the same lineage, was also possible with this AmpSeq method. The results demonstrate the feasibility of using MinION-based AmpSeq for rapid and accurate identification and lineage typing of IBV from oral swab samples.

microbiology↗

bioRxiv2018

Rapid and sensitive virulence prediction and identification of Newcastle disease virus genotypes using third-generation sequencing

Newcastle disease (ND) outbreaks are global challenges to the poultry industry. Effective management requires rapid identification and virulence prediction of the circulating Newcastle disease viruses (NDV), the causative agent of ND. However, these diagnostics are hindered by the genetic diversity and rapid evolution of NDVs. A highly sensitive amplicon sequencing (AmpSeq) workflow for virulence and genotype prediction of NDV samples using a third-generation, real-time DNA sequencing platform is described using both egg-propagated virus and clinical samples. 1D MinION sequencing of barcoded NDV amplicons was performed on 33 egg-grown isolates, (23 unique lineages, including 15 different NDV genotypes), and from 15 clinical swab samples from field outbreaks. Assembly-based data analysis was performed in a customized, Galaxy-based AmpSeq workflow. For all egg-grown samples, NDV was detected and virulence and genotype were predicted. For clinical samples, NDV was detected in ten of eleven NDV samples. Six of the clinical samples contained two mixed genotypes, of which the MinION method detected both genotypes in four of those samples. Additionally, testing a dilution series of one NDV sample resulted in detection of NDV with a 50% egg infectious dose (EID50) as low as 101 EID50/ml. This was accomplished in as little as 7 minutes of sequencing time, with a 98.37% sequence identity compared to the expected consensus. The high sensitivity, fast sequencing capabilities, accuracy of the consensus sequences, and the low cost of multiplexing allowed for identification of NDV of different genotypes circulating worldwide. This general method will likely be applicable to other infections agents.

molecular biology↗