Molecular epidemiological study on Infectious Pancreatic Necrosis Virus isolates from aquafarms in Scotland over three decades.

In order to obtain an insight into genomic changes and associated evolution and adaptation of Infectious Pancreatic Necrosis Virus (IPNV), the complete coding genomes of 57 IPNV isolates collected from Scottish aquafarms from 1982 to 2014 were sequenced and analysed. Phylogenetic analysis of the sequenced IPNV strains showed separate clustering of genogroups I, II, III and V. IPNV isolates with genetic reassortment of segment A/B of genogroup III/II were determined. About 59 % of the IPNV isolates belonged to the persistent type and 32 % to the low-virulent type, and only one highly pathogenic strain (1.79 %) was identified. Codon adaptation index calculations indicated that the IPNV major capsid protein VP2 has adapted to its salmonid host. Under-representation of CpG dinucleotides in the IPNV genome to minimize detection by the innate immunity receptors, and observed positive selection in the virulence determination sites of VP2 embedded in the variable region of the main antigenic region, suggest an immune escape mechanism driving virulence evolution. The prevalence of mostly persistent genotypes, together with the assumption of adaptation and immune escape, indicates that IPNV is evolving with the host.

Monitoring Viral Genetic Variation as a Tool To Improve Molecular Diagnostics for Mumps Virus.

Here, we report how the analysis of viral genetic variation using next-generation sequencing (NGS) can be used as a tool to improve mumps virus diagnostics. Analysis of NGS data from recently circulating mumps virus isolates allowed optimization of the current mumps virus real-time reverse transcription-PCR (RT-PCR) by primer and probe modifications due to nucleotide variations. The modified assay showed a higher efficiency and sensitivity than the previously used CDC protocol for the detection of currently circulating mumps virus strains and could therefore offer better support for outbreak control. The NGS sequence data were also used to make predictions of changes in the hemagglutinin-neuraminidase protein structure that could explain possible immune escape mechanisms.

Establishment of a minigenome system for Oropouche virus reveals the S genome segment to be significantly longer than reported previously.

Oropouche virus (OROV) is a medically important orthobunyavirus, which causes frequent outbreaks of a febrile illness in the northern parts of Brazil. However, despite being the cause of an estimated half a million human infections since its first isolation in Trinidad in 1955, details of the molecular biology of this tripartite, negative-sense RNA virus remain limited. We have determined the complete nucleotide sequence of the Brazilian prototype strain of OROV, BeAn 19991, and found a number of differences compared with sequences in the database. Most notable were that the S segment contained an additional 204 nt at the 3' end and that there was a critical nucleotide mismatch at position 9 within the base-paired terminal panhandle structure of each genome segment. In addition, we obtained the complete sequence of the Trinidadian prototype strain TRVL-9760 that showed similar characteristics to the BeAn 19991 strain. By using a T7 RNA polymerase-driven minigenome system, we demonstrated that cDNA clones of the BeAn 19991 L and S segments expressed functional proteins, and also that the newly determined terminal untranslated sequences acted as functional promoters in the minigenome assay. By co-transfecting a cDNA to the viral glycoproteins, virus-like particles were generated that packaged a minigenome and were capable of infecting naive cells.

Molecular and serological findings in suspected patients with Crimean-Congo hemorrhagic fever virus in Iran.

Crimean-Congo hemorrhagic fever (CCHF) is an arthropod-borne disease of humans associated with a severe clinical picture, including hemorrhagic syndrome and a high mortality rate. CCHF virus is widely distributed throughout large areas of the world. To characterize the serological status in CCHF patients, paired clinical samples were collected from suspected CCHF patients and analyzed by microbiological and other laboratory analyses with the aim of: determining the presence of neutralizing antibodies against CCHF virus; investigating the cross-reactivity of these neutralizing antibodies against virus isolated from the same outbreak and against other available laboratory strain; and studying the relationship between the isolated virus with other virus by whole genome sequencing. Patients at Boo-Ali Hospital, Zahedan, Iran, with clinical symptoms ranging from mild to severe hemorrhagic fever were included in the study. Two serum samples were taken from each patient, the first as soon as the patient matched the criteria for CCHF notification and the second when the patient was discharged from hospital (2 weeks later). Commercial and in-house assays revealed a positive IgM signal in acute serum samples from six patients. A novel finding was that CCHF patients develop neutralizing antibodies soon after infection. Interestingly these antibodies were able to neutralize other CCHF virus strains too. The complete sequence of the Zahedan 2007 isolate, including the hitherto unknown first L-segment sequence, was identified using an original clinical sample from one patient with confirmed CCHF infection.

Zahedan rhabdovirus, a novel virus detected in ticks from Iran.

BACKGROUND: Rhabdoviridae infect a wide range of vertebrates, invertebrates and plants. Their transmission can occur via various arthropod vectors. In recent years, a number of novel rhabdoviruses have been identified from various animal species, but so far only few tick-transmitted rhabdoviruses have been described. METHODS: We isolated a novel rhabdovirus, provisionally named Zahedan rhabdovirus (ZARV), from Hyalomma anatolicum anatolicum ticks collected in Iran. The full-length genome was determined using 454 next-generation sequencing and the phylogenetic relationship to other rhabdoviruses was analyzed. Inoculation experiments in mammalian Vero cells and mice were conducted and a specific PCR assay was developed. RESULTS: The complete genome of ZARV has a size of 11,230 nucleotides (nt) with the typical genomic organization of Rhabdoviridae. Phylogenetic analysis confirms that ZARV is closely related to Moussa virus (MOUV) from West Africa and Long Island tick rhabdovirus (LITRV) from the U.S., all forming a new monophyletic clade, provisionally designated Zamolirhabdovirus, within the Dimarhabdovirus supergroup. The glycoprotein (G) contains 12 conserved cysteins which are specific for animal rhabdoviruses infecting fish and mammals. In addition, ZARV is able to infect mammalian Vero cells and is lethal for mice when inoculated intracerebrally or subcutaneously. The developed PCR assay can be used to specifically detect ZARV. CONCLUSION: The novel tick-transmitted rhabdovirus ZARV is closely related to MOUV and LITRV. All three viruses seem to form a new monophyletic clade. ZARV might be pathogenic for mammals, since it can infect Vero cells, is lethal for mice and its glycoprotein contains 12 conserved cysteins only found in animal rhabdoviruses. The mammalian host of ZARV remains to be identified.

Genetic and biological characterization of selected Changuinola viruses (Reoviridae, Orbivirus) from Brazil.

The genus Orbivirus of the family Reoviridae comprises 22 virus species including the Changuinola virus (CGLV) serogroup. The complete genome sequences of 13 CGLV serotypes isolated between 1961 and 1988 from distinct geographical areas of the Brazilian Amazon region were obtained. All viral sequences were obtained from single-passaged CGLV strains grown in Vero cells. CGLVs are the only orbiviruses known to be transmitted by phlebotomine sandflies. Ultrastructure and molecular analysis by electron microscopy and gel electrophoresis, respectively, revealed viral particles with typical orbivirus size and morphology, as well as the presence of a segmented genome with 10 segments. Full-length nucleotide sequencing of each of the ten RNA segments of the 13 CGLV serotypes provided basic information regarding the genome organization, encoded proteins and genetic traits. Segment 2 (encoding VP2) of the CGLV is uncommonly larger in comparison to those found in other orbiviruses and shows varying sizes even among different CGLV serotypes. Phylogenetic analysis support previous serological findings, which indicate that CGLV constitutes a separate serogroup within the genus Orbivirus. In addition, six out of 13 analysed CGLV serotypes showed reassortment of their genome segments.

Impact of the COVID-19 related border restrictions on influenza and other common respiratory viral infections in New Zealand.

BACKGROUND: New Zealand's (NZ) complete absence of community transmission of influenza and respiratory syncytial virus (RSV) after May 2020, likely due to COVID-19 elimination measures, provided a rare opportunity to assess the impact of border restrictions on common respiratory viral infections over the ensuing 2 years. METHODS: We collected the data from multiple surveillance systems, including hospital-based severe acute respiratory infection surveillance, SHIVERS-II, -III and -IV community cohorts for acute respiratory infection (ARI) surveillance, HealthStat sentinel general practice (GP) based influenza-like illness surveillance and SHIVERS-V sentinel GP-based ARI surveillance, SHIVERS-V traveller ARI surveillance and laboratory-based surveillance. We described the data on influenza, RSV and other respiratory viral infections in NZ before, during and after various stages of the COVID related border restrictions. RESULTS: We observed that border closure to most people, and mandatory government-managed isolation and quarantine on arrival for those allowed to enter, appeared to be effective in keeping influenza and RSV infections out of the NZ community. Border restrictions did not affect community transmission of other respiratory viruses such as rhinovirus and parainfluenza virus type-1. Partial border relaxations through quarantine-free travel with Australia and other countries were quickly followed by importation of RSV in 2021 and influenza in 2022. CONCLUSION: Our findings inform future pandemic preparedness and strategies to model and manage the impact of influenza and other respiratory viral threats.

Full-length genome sequence of Ntaya virus.

Presentation of pyrosequencing data and phylogenetic analysis for the full genome of Ntaya virus, type virus of the Ntaya virus group of the Flaviviridae isolated in Cameroon in 1966.

Genetic characterization of Yug Bogdanovac virus.

We present pyrosequencing data and phylogenetic analysis for the full genome of Yug Bogdanovac virus (YBV), a member of the Vesicular stomatitis virus serogroup of the Rhabdoviridae isolated from a pool of Phlebotomus perfiliewi sandflies collected in Serbia in 1976. YBV shows very low nucleotide identities to other members of the Vesicular stomatitis virus serogroup and does not contain a reading frame for C'/C proteins.

Clarifying Bunyamwera virus riddles of the past.

Pyrosequencing data and phylogenetic analysis for the full genome of Ilesha virus, Ngari virus and Calovo virus are described clarifying their much discussed relationship within the species Bunyamwera virus of the genus Orthobunyavirus of the Bunyaviridae.

Development of real-time RT-PCR assays to detect measles virus on the Hologic Panther Fusion® System.

BACKGROUND: In 2019, Aotearoa New Zealand (NZ) experienced its worst measles outbreak since 1997. Due to declining childhood vaccination rates since the beginning of the SARS-CoV-2 pandemic, NZ is at serious risk of another major measles outbreak. Our laboratory provides diagnostic services to NZ's Southern region. In 2019 the Southern region experienced the greatest number of cases outside of Auckland and Northland, however we did not have a validated measles PCR assay in our laboratory. OBJECTIVES: We sought to develop reverse transcription real-time polymerase chain reaction (RT-PCR) assays for measles on the Hologic Panther Fusion® System by utilising its open access function. STUDY DESIGN: Previously published real-time RT-PCR assays were modified and optimised to detect wild-type measles virus (LDT-Mea), and the vaccine strain of measles virus (LDT-MeaVacA), on the Hologic Panther Fusion® System. The assays were clinically validated. RESULTS: The LDT-Mea assay has a limit of detection (LoD) of 0.1 CCID50, while the LDT-MeaVacA assay is less sensitive with a LoD of 1 CCID50. Using 27 samples, the clinical sensitivity and specificity was 100% for both assays. Other common respiratory viruses were found not to cross-react with either the LDT-Mea or LDT-MeaVacA assays. CONCLUSION: We have successfully adapted and validated for diagnostic use on the Hologic Panther Fusion® System previously published assays to detect wild-type and vaccine strains of the measles virus. The implementation of measles testing on this system will greatly improve the turn-around time for measles testing, and better support the measles public health response, for our region.

Circulation of influenza and other respiratory viruses during the COVID-19 pandemic in Australia and New Zealand, 2020-2021.

Objective: Circulation patterns of influenza and other respiratory viruses have been globally disrupted since the emergence of coronavirus disease (COVID-19) and the introduction of public health and social measures (PHSMs) aimed at reducing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission. Methods: We reviewed respiratory virus laboratory data, Google mobility data and PHSMs in five geographically diverse regions in Australia and New Zealand. We also described respiratory virus activity from January 2017 to August 2021. Results: We observed a change in the prevalence of circulating respiratory viruses following the emergence of SARS-CoV-2 in early 2020. Influenza activity levels were very low in all regions, lower than those recorded in 2017-2019, with less than 1% of laboratory samples testing positive for influenza virus. In contrast, rates of human rhinovirus infection were increased. Respiratory syncytial virus (RSV) activity was delayed; however, once it returned, most regions experienced activity levels well above those seen in 2017-2019. The timing of the resurgence in the circulation of both rhinovirus and RSV differed within and between the two countries. Discussion: The findings of this study suggest that as domestic and international borders are opened up and other COVID-19 PHSMs are lifted, clinicians and public health professionals should be prepared for resurgences in influenza and other respiratory viruses. Recent patterns in RSV activity suggest that these resurgences in non-COVID-19 viruses have the potential to occur out of season and with increased impact.

Confusions in orbivirus protein classification.

An extensive comparative analysis of orbivirus genomes revealed four cases of unclear numeration and protein designation, due to confused reference to protein size or segment size by which they are encoded. A concise nomenclature based on type species, sequence homology and functional characteristics independent of segment or protein size is suggested.

Genetic characterization of Bhanja virus and Palma virus, two tick-borne phleboviruses.

The genomes of Bhanja virus (BHAV) and Palma virus (PALV) two tick-borne viruses hitherto grouped into the Bhanja virus antigenic complex of the Bunyaviridae were determined by pyrosequencing. Phylogenetic analysis groups all three segments of BHAV and PALV into a distinct clade of tick-borne phleboviruses together with the newly described severe fever with thrombocytopenia syndrome virus and Uukuniemi virus. The terminal signature sequences which are signatures for taxonomic grouping and important for virus replication and RNA transcription show marked differences in the L- and S-segments.

Genetic characterization of Erve virus, a European Nairovirus distantly related to Crimean-Congo hemorrhagic fever virus.

Erve virus (ERVEV) is a European Nairovirus that is suspected to cause severe headache (thunderclap headache) and intracerebral hemorrhage. The mode of transmission to humans (ticks or mosquitoes) is still unknown. Currently, no standardized testing method for ERVEV exists and only a small partial sequence of the polymerase gene is available. Here, we present the first complete genome sequence of ERVEV S, M, and L segments. Phylogenetic comparison of the amino acid sequence of the L-protein (RNA-dependent RNA polymerase) revealed only 48 % homology to available L-protein sequences of other Nairoviruses like Crimean-Congo hemorrhagic fever virus, Nairobi sheep disease virus, Hazara virus, Kupe virus, and Dugbe virus. Among themselves, these Nairoviruses show 62-89 % homology in the L-protein sequences. Therefore, ERVEV seems to be only distantly related to other Nairoviruses. The new sequence data can be used for the development of diagnostic methods and the identification of the natural vector.

Clinical, Laboratory, and Molecular Epidemiology of an Outbreak of Aseptic Meningitis Due to a Triple-Recombinant Echovirus in Ashburton, New Zealand.

Here, we describe a small enterovirus outbreak including nine cases of aseptic meningitis in a New Zealand hospital in 2017. Most patients had a lymphocytic predominance in the CSF, their length of stay was short, and there were no paediatric cases or ICU admissions. VP1 genotyping revealed that the outbreak was caused by an echovirus E30 strain closely related to strains reported from the US, UK, Brazil, and Denmark. They all form a separate cluster within lineage "h", which leads to the proposal of establishing a new lineage tentatively named "j" for this group of echovirus E30 strains. However, whole genome sequencing and reference mapping to echovirus E30 sequences showed very poor mapping of reads to the 3' half of the genome. Further bioinformatic analysis indicated that the causative agent of this outbreak might be a mosaic triple-recombinant enterovirus composed of echovirus E6, echovirus E11, and echovirus E30 genome segments.

Co-infection of Peruvian horse sickness virus and West Nile virus associated with neurological diseases in horses from Brazil.

In 2018, during the surveillance for West Nile virus (WNV) in horses with neurological clinical signs in the state of Espírito Santo (Brazil), 19 animals were investigated, and 52 biological samples were collected for WNV diagnostic. One brain sample was positive for WNV by RT-qPCR and the virus was isolated in C6/36 cell culture and sequenced. We obtained a nearly complete genome of WNV co-infected with Peruvian horse sickness virus (PHSV) in the cell culture. After confirmation of PHSV by next-generation sequencing, a new PHSV RT-qPCR protocol was developed, which was used to detect another horse positive only for PHSV. This assay provides a simple and direct method for easy identification of PHSV from biological samples from horses and may become a useful tool in the epidemiological surveillance of this virus. It is the first case of PHSV in Brazil, and only the third country overall to report, 23 years after the first confirmed notification in Peru. Moreover, it is the first reported co-infection of PHSV and WNV in a horse with neurological signs, confirmed by RT-qPCR.

Rhinothermy delivered by nasal high flow therapy in the treatment of the common cold: a randomised controlled trial.

BACKGROUND: The common cold is the most common infectious disease affecting humans and has a substantial economic impact on society. Human rhinoviruses, which cause almost two-thirds of colds, have demonstrated temperature-dependent replication which is optimal between 33°C and 35°C. METHODS: This randomised, single-blind, parallel-group trial completed at a single-centre in New Zealand, recruited 170 participants aged 18-75 years (mean age 27.5 years) who were within 48 hours of common cold symptom onset and had a symptom score (the Modified Jackson Score (MJS)) ≥7 and a negative point-of-care test for influenza. Participants were blinded to the intervention and randomised (1:1) to 5 days of either nasal high flow rhinothermy (rNHF) (100% humidified air delivered at 35 L/min and 41°C for 2 hours daily) (n=85) or 'sham' rhinothermy (100% humidified air delivered at 10 L/min and 31°C for 10 min daily) (n=85) and completed daily symptom diaries, which included the MJS, for 14 days, to investigate whether rNHF reduced common cold symptom severity and duration compared with 'sham' rhinothermy. RESULTS: An intention-to-treat superiority analysis included all randomised participants and showed no difference between treatment groups for the primary outcome, the day 4 MJS analysed by analysis of covariance: mean (SD) 6.33 (3.97) for rNHF vs 5.8 (3.15) for 'sham'; estimated difference (95% CI) 0.37 (-0.69 to 1.42), p=0.49. There was no difference in time until resolution of symptoms: mean (SD) 5.96 (4.47) days for rNHF vs 6.42 (4.09) days for 'sham'; estimated difference (95% CI) 1.02 (0.75 to 1.38), p=0.91. There were no serious adverse events related to the study treatments. CONCLUSIONS: This well-powered, single-blind randomised controlled trial does not provide evidence that 5 days of rNHF (100% humidified air heated to 41°C delivered at 35 L/min for 2 hours daily) reduces common cold symptom severity or duration. However, investigation of rNHF in the treatment of influenza is warranted. TRIAL REGISTRATION NUMBER: ACTRN12617001340325.

Impact of the COVID-19 nonpharmaceutical interventions on influenza and other respiratory viral infections in New Zealand.

Stringent nonpharmaceutical interventions (NPIs) such as lockdowns and border closures are not currently recommended for pandemic influenza control. New Zealand used these NPIs to eliminate coronavirus disease 2019 during its first wave. Using multiple surveillance systems, we observed a parallel and unprecedented reduction of influenza and other respiratory viral infections in 2020. This finding supports the use of these NPIs for controlling pandemic influenza and other severe respiratory viral threats.