Multiple Viruses Research Articles

The use of human intestinal enteroid cell cultures for detection of multiple gastroenteric viruses.

Human norovirus (HuNoV) and human astrovirus (HAstV) are viral enteric pathogens and known causative agents of acute gastroenteritis. Identifying the presence of these viruses in environmental samples such as irrigation water, or foods exposed to virus contaminated water (e.g., shellfish, agricultural crops), remains an important goal in the field of food safety. Determining if a virus species present in a sample is infectious is complicated by the recalcitrance of many enteric virus species to grow in culture, and/or the lack of a common cell culture system(s). Human intestinal enteroids (HIEs) can support the replication of HuNoV and HAstV, and thus hold promise as a platform for demonstrating the replication of multiple enteric virus species within a single sample. The objective of this study was to determine if HIEs can support co-replication of two genetically distinct human enteric viruses, HAstV3 and HuNoV GII.4[P16]. In single virus infections, HuNoV GII.4[P16] RNA levels were highest at 48-72 hpi (6.3 to 9.1 x 106 genome copy equivalents [gce]/well) and HAstV3 RNA levels were highest at 24 hpi (3.4 ×108 gce/well). HAstV3-infected cells stained positive for viral capsid protein at 24 hpi and induced the synthesis of RNA and protein expression of interferon (IFN)-beta, -lambda 1 and 2/3, peaking at 24 hpi and 48 hpi respectively. HuNoV GII.4[P16] replication was negatively impacted by HAstV3 co-infection, but HAstV3 was unaffected by HuNoV. A reduction in HuNoV GII.4[P16] RNA during co-infections was observed at 72 hpi, with partial restoration achieved using neutralizing anti-IFN-antibodies. Human intestinal enteroids can support the co-infection and replication of HuNoVGII.4[P16] and HAstV3, even at more than 100-fold excess in one virus over the other, and compounds (e.g., anti-IFN antibodies) that interfere with HIE antiviral mechanism(s) can aid in maximizing HuNoV replication during co-infection.

Detection of Rotavirus, Norovirus, and Astrovirus Causing Acute Diarrhoea in Children by Multiplex PCR in a Tertiary Care Hospital.

To evaluate the use of a multiplex real-time polymerase chain reaction (RT-PCR) test in detecting three viruses namely Rotavirus, Norovirus (genotypes 1 and 2), and Astrovirus that cause gastroenteritis in children under the age of five years. A cross-sectional study. Place and Duration of the Study: Department of Virology, Armed Forces Institute of Pathology (AFIP), Rawalpindi, Pakistan, from January to July 2023. A total of 87 children having acute diarrhoea and less than 5 years of age were included in this study from the outpatient clinic. Multiplex PCR was performed for the detection of three viruses: Rotavirus, Norovirus, and Astrovirus in stool samples of patients using a commercially available PCR kit. The data were analysed using Statistical Package for Social Sciences (SPSS) version 27:00. Out of total 87 children, aged 2.5 ± 1.5 years, 56 (64.3%) were positive for multiplex RT-PCR and 31 (35.6%) were negative. Rotavirus was identified in 24 (27.5%) children as the most common cause of acute diarrhoea, followed by Norovirus in 20 (22.9%), and Astrovirus in 7 (8%) patients, while co-infection with multiple viruses occurred in 5 (5.7%) of the cases. This study revealed viral aetiology as a significant cause of acute diarrhoea in children. Multiplex PCR in the healthcare system can make it easier to identify, treat, and control the upsurge of diarrhoea. Prompt diagnosis of viral causes can lead to the prevention of unnecessary use of antibiotics. Multiplex polymerase chain reaction, Rotavirus, Norovirus, Astroviridae, Diarrhoea.

Discovery and development of INNA-051, a TLR2/6 agonist for the prevention of complications resulting from viral respiratory infections.

Viral respiratory infection is associated with significant morbidity and mortality. The diversity of viruses implicated, coupled with their propensity for mutation, ignited an interest in host-directed antiviral therapies effective across a wide range of viral variants. Toll-like receptors (TLRs) are potential targets for the development of broad-spectrum antivirals given their central role in host immune defenses. Synthetic agonists of TLRs have been shown to boost protective innate immune responses against respiratory viruses. However, clinical success was hindered by short duration of benefit and/or induction of systemic adverse effects. INNA-051, a TLR2/6 agonist, is in development as an intranasal innate immune enhancer for prophylactic treatment in individuals at risk of complications resulting from respiratory viral infections. In vivo animal demonstrated the efficacy as prophylaxis against multiple viruses including SARS-CoV-2, influenza, and rhinovirus. Early clinical trials demonstrated an acceptable safety and tolerability profile. Intranasal delivery to the primary site of infection in humans induced a local innate host defense response characterized by innate immune cell infiltration into the nasal epithelium and activation and antiviral response genes . Taken together, the preclinical and clinical data on INNA-051 support further investigation of its use in community infection setting.

The UP1 domain is essential for the facilitation effect of HnRNP A1 on PRRSV-2 replication.

Porcine reproductive and respiratory syndrome virus (PRRSV) infection causes reproductive failure and respiratory distress and is a serious threat to the swine industry, given its continuous and rapid emergence. The knowledge of viral-host interaction could facilitate anti-PRRSV drug development. HnRNP A1 is an abundantly expressed protein which associates with RNA metabolic processes and plays multifarious roles during the reproduction cycle of multiple viruses. However, the function of porcine HnRNP A1 in PRRSV-2 replication is still unknown. Herein, HnRNP A1 was identified as a nucleocapsid (N)-binding protein for PRRSV-2. Overexpression of porcine HnRNP A1 promoted the expression of viral RNA, and viral proteins, corresponding to enhanced virus titers. While deletion of the UP1 domain abolished the HnRNP A1-mediated enhancement of PRRSV-2 replication. In addition, HnRNP A1-silencing confirmed its pro-viral effect on PRRSV-2 infectivity in porcine alveolar macrophages (PAMs). RNA pull-down and RNA immunoprecipitation verification confirmed that the UP1 domain is important for the recognition of the guanine-rich sequence (GRS) in PRRSV-2 negative RNA. Eventually, supplementation with TMPyP4, a G4 ligand, efficiently provokes the release of HnRNP A1 from GRS, thereby limiting PRRSV-2 replication. Together, these findings help to inform the mechanism by which HnRNP A1 accelerates PRRSV-2 replication, and facilitate antiviral drug design.

Screening great ape museum specimens for DNA viruses

Natural history museum collections harbour a record of wild species from the past centuries, providing a unique opportunity to study animals as well as their infectious agents. Thousands of great ape specimens are kept in these collections, and could become an important resource for studying the evolution of DNA viruses. Their genetic material is likely to be preserved in dry museum specimens, as reported previously for monkeypox virus genomes from historical orangutan specimens. Here, we screened 209 great ape museum specimens for 99 different DNA viruses, using hybridization capture coupled with short-read high-throughput sequencing. We determined the presence of multiple viruses within this dataset from historical specimens and obtained several near-complete viral genomes. In particular, we report high-coverage (> 18-fold) hepatitis B virus genomes from one gorilla and two chimpanzee individuals, which are phylogenetically placed within clades infecting the respective host species.

Delayed low-dose oral administration of 4'-fluorouridine inhibits pathogenic arenaviruses in animal models of lethal disease.

Development of broad-spectrum antiviral therapies is critical for outbreak and pandemic preparedness against emerging and reemerging viruses. Viruses inducing hemorrhagic fevers cause high morbidity and mortality in humans and are associated with several recent international outbreaks, but approved therapies for treating most of these pathogens are lacking. Here, we show that 4'-fluorouridine (4'-FlU; EIDD-2749), an orally available ribonucleoside analog, has antiviral activity against multiple hemorrhagic fever viruses in cell culture, including Nipah virus, Crimean-Congo hemorrhagic fever virus, orthohantaviruses, and arenaviruses. We performed preclinical in vivo evaluation of oral 4'-FlU against two arenaviruses, Old World Lassa virus (LASV) and New World Junín virus (JUNV), in guinea pig models of lethal disease. 4'-FlU demonstrated both advantageous pharmacokinetic characteristics and high efficacy in both of these lethal disease guinea pig models. Additional experiments supported protection of the infected animals even when 4'-FlU delivery was reduced to a low dose of 0.5 milligram per kilogram. To demonstrate clinical utility, 4'-FlU treatment was evaluated when initiated late in the course of infection (12 or 9 days after infection for LASV and JUNV, respectively). Delayed treatment resulted in rapid resolution of clinical signs, demonstrating an extended window for therapeutic intervention. These data support the use of 4'-FlU as a potent and efficacious treatment against highly pathogenic arenaviruses of public health concern with a virus inhibition profile suggesting broad-spectrum utility as an orally available antiviral drug against a wide variety of viral pathogens.

Viral susceptibility and innate immune competency of Carollia perspicillata bat cells produced for virological studies.

Multiple viruses that are highly pathogenic in humans are known to have evolved in bats. How bats tolerate infection with these viruses, however, is poorly understood. As viruses engage in a wide range of interactions with their hosts, it is essential to study bat viruses in a system that resembles their natural environment like bat-derived in vitro cellular models. However, stable and accessible bat cell lines are not widely available for the broader scientific community. Here, we generated in vitro reagents for the Seba's short-tailed bat (Carollia perspicillata), tested multiple methods of immortalization, and characterized their susceptibility to virus infection and response to immune stimulation. Using a pseudotyped virus library and authentic virus infections, we show that these C. perspicillata cell lines derived from a diverse array of tissues are susceptible to viruses bearing the glycoprotein of numerous orthohantaviruses, including Andes and Hantaan virus and are also susceptible to live hantavirus infection. Furthermore, stimulation with synthetic double-stranded RNA prior to infection with VSV and MERS-CoV induced a protective antiviral response, demonstrating the suitability of our cell lines to study the bat antiviral immune response. Taken together, the approaches outlined here will inform future efforts to develop in vitro tools for virology from non-model organisms and these C. perspicillata cell lines will enable studies on virus-host interactions in bats.

Monitoring Influenza A (H1N1, H3N2), RSV, and SARS-CoV-2 Using Wastewater-Based Epidemiology: A 2-Year Longitudinal Study in an Indian Megacity Covering Omicron and Post-Omicron Phases.

The bourgeoning field of wastewater-based epidemiology (WBE) for the surveillance of several respiratory viruses which includes Influenza A, H1N1pdm09, H3N2, respiratory syncytial viruses (RSV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is of interest for public health concerns. However, there are few long-term monitoring studies globally. In this study, respiratory viruses were detected and quantified from 11 sewer sheds by utilizing reverse transcription-quantitative polymerase chain reaction analysis in Pune city, India, from Jan 2022 to Dec 2023. The RNA fragments of respiratory viruses were detected in sewage samples before clinical cases were reported, underscoring the potential of WBE for early detection and monitoring within the population. The Spearman correlation of wastewater viral copies was positively and significantly correlated with the clinically positive case of H1N1pdm09 (ρ = 0.55, p = 1.4 × 10-9), H3N2 (ρ = 0.25, p = 9.9 × 10-3), and SARS-CoV-2 (ρ = 0.43, p = 4.1 × 10-6). The impact of public health interventions on the circulation of infectious respiratory diseases showed a significant difference in the viral load during the period when many preventing measures were carried out against the COVID-19 pandemic (restriction phase), compared to the period when no such preventive measures are followed (no-restriction phase) for Influenza A, H1N1pdm09, H3N2, and RSV with p-value < 0.05, which indicates the influence of health policy implementation in controlling disease spread. The present study provides an effective approach to detecting multiple respiratory viruses from wastewater and provides insights into the epidemiology of respiratory illnesses. The WBE aids in providing information on the spread of pathogens (viruses) in the community, offering a proactive strategy for public health management, allowing for timely interventions and implementing targeted measures to mitigate the spread of these viruses under one health approach.

Microfluidic qPCR for detection of 21 common respiratory viruses in children with influenza-like illness

Multiple respiratory viruses lead to high morbidity and mortality, yet global surveillance platforms focus primarily on seasonal influenza viruses. The COVID-19 pandemic and new RSV vaccines highlight the importance of a broader approach. Upper respiratory tract swabs from children aged 24–59 months presenting with influenza-like illness in The Gambia were collected during follow-up of a live-attenuated influenza vaccine randomised controlled trial in 2017–18. A microfluidic quantitative polymerase chain reaction (qPCR) assay was established and used to detect 21 respiratory viruses. 76.6% of samples had one or more viruses detected (n = 121/158). The viruses detected most frequently were rhinovirus (n = 37/158, 23.4%) and adenovirus (n = 34/158, 21.5%), followed by parainfluenza virus 3, influenza B and human metapneumovirus B. A third of positive samples had multiple viruses detected (two n = 31/121, 25.6%; three n = 9/121, 7.4%). Our data demonstrates how microfluidic qPCR is a useful tool for high-throughput, comprehensive detection of multiple respiratory viruses in surveillance platforms. Rapidly changing epidemiology exemplifies the need for new, broader approaches to virus surveillance in low-resource settings to respond to future epidemics and to guide the need for and use of new prevention and therapeutic measures.

Chronic Viral Reactivation and Associated Host Immune Response and Clinical Outcomes in Acute COVID-19 and Post-Acute Sequelae of COVID-19.

Chronic viral infections are ubiquitous in humans, with individuals harboring multiple latent viruses that can reactivate during acute illnesses. Recent studies have suggested that SARS-CoV-2 infection can lead to reactivation of latent viruses such as Epstein-Barr Virus (EBV) and cytomegalovirus (CMV), yet, the extent and impact of viral reactivation in COVID-19 and its effect on the host immune system remain incompletely understood. Here we present a comprehensive multi-omic analysis of viral reactivation of all known chronically infecting viruses in 1,154 hospitalized COVID-19 patients, from the Immunophenotyping Assessment in a COVID-19 Cohort (IMPACC) study, who were followed prospectively for twelve months. We reveal significant reactivation of Herpesviridae, Enteroviridae, and Anelloviridae families during acute stage of COVID-19 (0-40 days post-hospitalization), each exhibiting distinct temporal dynamics. We also show that viral reactivation correlated with COVID-19 severity, demographic characteristics, and clinical outcomes, including mortality. Integration of cytokine profiling, cellular immunophenotyping, metabolomics, transcriptomics, and proteomics demonstrated virus-specific host responses, including elevated pro-inflammatory cytokines (e.g. IL-6, CXCL10, and TNF), increased activated CD4+ and CD8+ T-cells, and upregulation of cellular replication genes, independent of COVID-19 severity and SARS-CoV-2 viral load. Notably, persistent Anelloviridae reactivation during convalescence (≥3 months post-hospitalization) was associated with Post-Acute Sequelae of COVID-19 (PASC) symptoms, particularly physical function and fatigue. Our findings highlight a remarkable prevalence and potential impact of chronic viral reactivation on host responses and clinical outcomes during acute COVID-19 and long term PASC sequelae. Our data provide novel immune, transcriptomic, and metabolomic biomarkers of viral reactivation that may inform novel approaches to prognosticate, prevent, or treat acute COVID-19 and PASC.

One-Step Multiplex RT-PCR Method for Detection of Melon Viruses.

This study presents a one-step multiplex reverse transcription polymerase chain reaction (RT-PCR) method for the simultaneous detection of multiple viruses affecting melon crops. Viruses such as Watermelon mosaic virus (WMV), Cucumber mosaic virus (CMV), Zucchini yellow mosaic virus (ZYMV), Squash mosaic virus (SqMV), Tobacco mosaic virus (TMV), Papaya ring spot virus (PRSV), and Melon yellow spot virus (MYSV) pose a great threat to melons. The mixed infection of these viruses is the most common observation in the melon-growing fields. In this study, we surveyed northern Xingjiang (Altay, Changji, Wujiaqu, Urumqi, Turpan, and Hami) and southern Xingjiang (Aksu, Bayingolin, Kashgar, and Hotan) locations in Xinjiang province and developed a one-step multiplex RT-PCR to detect these melon viruses. The detection limits of this multiplex PCR were 103 copies/μL for ZYMV and MYSV and 102 copies/μL for WMV, SqMV, PRSV, CMV, and TMV. The detection results in the field showed 242 samples were infected by one or more viruses. The multiplex RT-PCR protocol demonstrated rapid, simultaneous, and relatively effective detection of viruses such as WMV, CMV, ZYMV, SqMV, TMV, PRSV, and MYSV. The technique is designed to identify these melon viruses in a single reaction, enhancing diagnostic efficiency and reducing costs, thus serving as a reference for muskmelon anti-virus breeding in Xinjiang.

Dynamin independent endocytosis is an alternative cell entry mechanism for multiple animal viruses.

Mammalian receptor-mediated endocytosis (RME) often involves at least one of three isoforms of the large GTPase dynamin (Dyn). Dyn pinches-off vesicles at the plasma membrane and mediates uptake of many viruses, although some viruses directly penetrate the plasma membrane. RME is classically interrogated by genetic and pharmacological interference, but this has been hampered by undesired effects. Here we studied virus entry in conditional genetic knock-out (KO) mouse embryonic fibroblasts lacking expression of all three dynamin isoforms (Dyn-KO-MEFs). The small canine parvovirus known to use a single receptor, transferrin receptor, strictly depended on dynamin. Larger viruses or viruses known to use multiple receptors, including alphaviruses, influenza, vesicular stomatitis, bunya, adeno, vaccinia, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and rhinoviruses infected Dyn-KO-MEFs, albeit at higher dosage than wild-type MEFs. In absence of the transmembrane protease serine subtype 2 (TMPRSS2), which normally activates the SARS-CoV-2 spike protein for plasma membrane fusion, SARS-CoV-2 infected angiotensin-converting enzyme 2 (ACE2)-expressing MEFs predominantly through dynamin- and actin-dependent endocytosis. In presence of TMPRSS2 the ancestral Wuhan-strain bypassed both dynamin-dependent and -independent endocytosis, and was less sensitive to endosome maturation inhibitors than the Omicron B1 and XBB variants, supporting the notion that the Omicron variants do not efficiently use TMPRSS2. Collectively, our study suggests that dynamin function at endocytic pits can be essential for infection with single-receptor viruses, while it is not essential but increases uptake and infection efficiency of multi-receptor viruses that otherwise rely on a functional actin network for infection.

TwinDemic detection: A non-enzymatic signal amplification system for on-site detection of multiple respiratory viruses

The simultaneous occurrence of multiple respiratory viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 [CoV]) and influenza A virus (IAV), presents a challenge for accurate diagnosis and treatment. To address this, we developed the TwinDemic Detection (TDD) system—an integrated point-of-care diagnostic platform that can simultaneously detect CoV and IAV using a multi-respiratory virus identification approach. The TDD system contains a transparent poly(methyl methacrylate) microfluidic chip with a hydrogel-based gene detection sensor and a handheld fluorescence reader. This system uses a fuel-stimulated DNA probe to detect viral RNA and amplify the fluorescence signal without the need for enzymatic or thermal controls. The system’s performance was tested using 45 human nasopharyngeal samples; results revealed positive and negative predictive values of 93.3 % and 96.7 % for CoV, and 100 % and 96.7 % for IAV, respectively. This system enables the accurate and simultaneous detection of multiple respiratory viruses for public health surveillance and shows potential for use as a point-of-care molecular test to monitor a wider range of viruses in the future.

Local and Noninvasive Glyco-Virus Checkpoint Nanoblockades Restrict Sialylation for Prolonged Broad-Spectrum Epidemic Virus Therapy.

The coronavirus disease 2019 (COVID-19) pandemic has driven major advances in virus research. The role of glycans in viral infection has been revealed, with research demonstrating that terminal sialic acids are key receptors during viral attachment and infection into host cells. However, there is an urgent demand for universal tools to study the mechanism of sialic acids in viral infections, as well as to develop therapeutic agents against epidemic viruses through the downregulation of terminal sialic acid residues on glycans acting as a glyco-virus checkpoint to accelerate virus clearance. In this study, we developed a robust sialic acids blockade tool termed local and noninvasive glyco-virus checkpoint nanoblockades (LONG NBs), which blocked cell surface sialic acids by endogenously and continuously inhibiting the de novo sialic acids biosynthesis pathway. Furthermore, LONG NBs could accurately characterize the sialic acid-dependent profiles of multiple virus variants and protected the host against partial SARS-CoV-2, rotavirus, and influenza A virus infections after local and noninvasive administration. Our results suggest that LONG NBs represent a promising tool to facilitate in-depth research on the mechanism of viral infection, and serve as a broad-spectrum protectant against existing and emerging viral variants via glyco-virus checkpoint blockade.

Dengue Outbreak Caused by Multiple Virus Serotypes and Lineages, Colombia, 2023-2024.

Dengue cases rose to record levels during 2023-2024. We investigated dengue in Valle del Cauca, Colombia, to determine if specific virus serotypes or lineages caused its large outbreak. We detected all 4 serotypes and multiple lineages, suggesting that factors such as climatic conditions were likely responsible for increased dengue in Colombia.

ZC3HAV1 facilitates STING activation and enhances inflammation

Stimulator of interferon genes (STING) is vital in the cytosolic DNA-sensing process and critical for initiating the innate immune response, which has important functions in host defense and contributes to the pathogenesis of inflammatory diseases. Zinc finger CCCH-type antiviral protein 1 (ZC3HAV1) specifically binds the CpG dinucleotides in the viral RNAs of multiple viruses and promotes their degradation. ZAPS (ZC3HAV1 short isoform) is a potent stimulator of retinoid acid-inducible gene I (RIG-I) signaling during the antiviral response. However, how ZC3HAV1 controls STING signaling is unclear. Here, we show that ZC3HAV1 specifically potentiates STING activation by associating with STING to promote its oligomerization and translocation from the endoplasmic reticulum (ER) to the Golgi, which facilitates activation of IRF3 and NF-κB pathway. Accordingly, Zc3hav1 deficiency protects mice against herpes simplex virus-1 (HSV-1) infection- or 5,6-dimethylxanthenone-4-acetic acid (DMXAA)-induced inflammation in a STING-dependent manner. These results indicate that ZC3HAV1 is a key regulator of STING signaling, which suggests its possible use as a therapeutic target for STING-dependent inflammation.

Age distribution of patients with multiple High-Risk Human Papilloma Virus (HR-HPV) genotypes and HPV vaccine recommendations by age

Introduction: The aim of this study to identify the age distribution of patients with multiple high risk Human Papilloma Virus (HR-HPV) genotypes and to evaluate HPV vaccination programs through age distribution. Methods: Patients between the ages of 20-70, who had cervical screening (cervical smear) and HPV genotype tests between January 2017 and December 2021 in Gynecology and Obstetrics Department in Tınaztepe University Hospital, were included in this study. HR-HPV genotype tests and age information which were performed evaluated retrospectively. Results: The study consisted of 66 patients analysis with multiple HR-HPV genotypes. The median age of the patients with multiple HR- HPV was found to be 32.6 years old. The most frequent combination of multiple HR-HPV was found two sub-types of HR-HPV (66.7%), median age was found as 36.6 years old. In this study the frequency for three sub-types of HR-HPV was found 24.2% and median age for this group was found to be 34.7 years old. We found frequency in patients with four or more sub-types of HR-HPV as 9.1%, and the median age for this group as 33.1 years old was found. Conclusion: The most common cause of cervical cancer is HPV infection. Multiple HR-HPV infection progression may promote high risk cervical lesions and neoplasia. HPV genotype testing results considering combination of subtypes may direct individual treatment, follow-up protocols for patients. HPV vaccination research may comprise multiple HR-HPV subtypes relations and age distribution for optimal immunization.

The Genetic Characterization of Lake Sinai Virus in Colony Losses Apiaries in Türkiye

Objective: Honeybees (Apis mellifera) have a unique role in natural pollination and maintaining biodiversity in the ecosystem. The alarming increase in unexpected colony losses, mysterious bee deaths, and the tragic extinction of entire colonies (Colony collapsed disorder- CCD) have sounded a global alarm, demanding immediate attention and collaborative action to address these critical challenges in bee breeding. Diseases, parasites, and pathogens significantly threaten colony health. Türkiye is a significant honey producer, providing an ideal environment for beekeeping due to its unique eco-geographical features. Unexpected colony losses and bee deaths are also questions of concern for beekeepers in Türkiye. Material Method: In this study investigated honey bee viruses in apiaries experiencing sudden bee death losses and CCD-like symptoms between 2021 and 2023 in Türkiye, involving genetic analysis of the LSV RdRp gene region. The honeybee and varroa samples were obtained from 52 colonies in 26 apiaries complaining of unexpected bee deaths and CCD-like symptoms between May 2021 and September 2023. Result: The results showed a high DWV, BQCV, and LSV prevalence, respectively. The sampled apiaries were infested mild-moderate- high grade with Varroa mites. Following PCR results, DWV, BQCV, LSV, IAPV, CBPV, and SBV positivity was detected at 69.2% (n=18), 50% (n=13), 38.46% (n=10), 26.9% (n=7), 19.2% (n=5) and 3.8% (n=1), respectively. High rates of multiple virus coexisting and high varroa infestation were noted in colonies with heavy losses and CCD-like complaints. The RdRp gene from two LSV samples (TrLSV-6474, TrLSV-6517) was sequenced. Turkish LSV samples (TrLSVs) showed a 72.88% homology of each other and clustered LSV4 branches in the phylogenetic tree. Turkish LSV sequences showed a closer similarity rate than reference sequences in GenBank with Asian Korean, Chinese, and Japanese LSV sequences. Conclusion: Further investigation is needed to comprehend the implications of elevated LSV populations on colony losses. The execution of genetic research with a more extensive sample size can significantly enhance the demonstration of species diversity and provide valuable insights into the influence of LSV variants on honeybee health and the management of diseases.

42 Clinical features and severity of COVID-19 with respiratory virus coinfections versus SARS-CoV-2 monoinfection in hospitalized children: A Canadian national surveillance study

Abstract Background The co-circulation of SARS-CoV-2 alongside other respiratory viruses has led to the risk of coinfections, potentially intensifying the severity of cases, especially in children. Objectives This study examined the epidemiology, clinical characteristics, and outcomes of SARS-CoV-2 respiratory virus coinfections in comparison to SARS-CoV-2 monoinfections in hospitalized children. Design/Methods A nationwide surveillance study was conducted to assess paediatric COVID-19-related hospitalizations across Canada from April 2020 to May 2022. Data were captured through two datasets covering ~90% of all Canadian tertiary-care paediatric beds: The Canadian Paediatric Surveillance Program and the Canadian Immunization Monitoring Program, ACTive. Coinfections were defined as the simultaneous detection of SARS-CoV-2 and ≥1 other respiratory virus. Severe infection was defined as intensive care, ventilatory, or hemodynamic support needs, organ systems complications, or death. Variables and outcomes were summarized and compared, and risk ratios were computed using Poisson regression, adjusting for age, gender, comorbid conditions, SARS-CoV-2 lineage, and vaccination status. Results Out of 1501 COVID-19-related hospitalizations, 163 (10.9%) had documented coinfections with 44/163 [27%] involving SARS-COV-2 plus ≥2 other respiratory viruses. The majority of SARS-CoV-2 monoinfections (1176/1501, 87.9%) and coinfections (139/163, 85.3%) belonged to the Omicron era (Figure A). RSV (66/163, 40.5%) and Enterovirus/rhinovirus (61/163, 37.4%) coinfections were the most common. Coinfection cases were significantly younger than monoinfection cases (median age 1.2 [IQR:0.3-3.3] vs 1.6 [IQR:0.3-7.0] years, p=0.04). Overall, severe disease was more common among cases with any coinfection (38.0%) than SARS-CoV-2 monoinfection (25.7%; adjusted risk ratio 1.45, 95% confidence interval 1.17-1.80) (Figure B). In particular, we observed that severe outcomes were significantly higher in SARS-CoV-2 coinfections compared to monoinfections during the Omicron era (Figure C-D). Overall, 26/61 (42.6%) Enterovirus/Rhinovirus and 20/66 (30.3%) RSV coinfections were associated with severe outcomes. Conclusion Children with documented coinfections had more severe respiratory disease compared to SARS-COV-2 monoinfections. However, children with severe COVID-19 may have been more likely tested for multiple viruses, leading to a risk of underestimation of coinfections among children with milder disease. Further work is needed to assess how different virus coinfections may affect children and which, if any, may be a predominant driver of disease severity. 1Severe disease was defined as intensive care, ventilatory, or hemodynamic support requirements, organ system complications, or death. 2Lineage was first assigned based on available genetic sequencing data. If missing, lineage was imputed based on the dominant circulating lineage at the provincial level according to the GISAID database.

Metagenomic Detection of Multiple Viruses in Monk Parakeet (Myiopsitta monachus) in Australia.

Birds are known to harbour many pathogens, including circovirus, herpesviruses, adenoviruses and Chlamydia psittaci. Some of these pose zoonotic risks, while others, such as beak and feather disease virus (BFDV), have a significant impact on the conservation of endangered bird species. This study was aimed to determine the faecal virome of a group of apparently healthy Monk parakeet using high-throughput sequencing. Fresh faecal samples were collected from four Monk parakeets at a pet shop in Melbourne, Australia. Virus enrichment and nucleic acid extraction were performed on the faecal samples, followed by high-throughput sequencing at the Australian Genome Research Facility (AGRF). Utilising an established pipeline for high-throughput sequencing data analysis, this study revealed the presence of three viruses of the families Circoviridae, Parvoviridae and Adenoviridae. Subsequent sequence comparison and phylogenetic analyses further confirmed that the detected viruses belong to the genera Chaphamaparvovirus (unassigned species), Circovirus (species Circovirus parrot) and Siadenovirus (species Siadenovirus viridis). Despite non-pathogenicity, the existence of multiple viruses within a bird species underscores the risk of these viruses spreading into the pet trade. Detection and a better understanding of avian viruses are crucial for the establishment of appropriate management and biosecurity measures in the domestic and international bird trade, which ultimately supports the conservation of vulnerable bird species.