Interspecies transmission of porcine-originated G4P[6] rotavirus A between pigs and humans: a synchronized spatiotemporal approach

As a leading viral cause of acute gastroenteritis in both humans and pigs, rotavirus A (RVA) poses a potential public health concern. Although zoonotic spillover of porcine RVA strains to humans is sporadic, it has been detected worldwide. The origin of chimeric human–animal strains of RVA is closely linked to the crucial role of mixed genotypes in driving reassortment and homologous recombination, which play a major role in shaping the genetic diversity of RVA. To better understand how genetically intertwined porcine and zoonotic human-derived G4P[6] RVA strains are, the present study employed a spatiotemporal approach to whole-genome characterization of RVA strains collected during three consecutive RVA seasons in Croatia (2018–2021). Notably, sampled children under 2 years of age and weanling piglets with diarrhea were included in the study. In addition to samples tested by real-time RT-PCR, genotyping of VP7 and VP4 gene segments was conducted. The unusual genotype combinations detected in the initial screening, including three human and three porcine G4P[6] strains, were subjected to next-generation sequencing, followed by phylogenetic analysis of all gene segments, and intragenic recombination analysis. Results showed a porcine or porcine-like origin for each of the eleven gene segments in all six RVA strains. The G4P[6] RVA strains detected in children most likely resulted from porcine-to-human interspecies transmission. Furthermore, the genetic diversity of Croatian porcine and porcine-like human G4P[6] strains was propelled by reassortment events between porcine and porcine-like human G4P[6] RVA strains, along with homologous intragenotype and intergenotype recombinations in VP4, NSP1, and NSP3 segments. Described concurrent spatiotemporal approach in investigating autochthonous human and animal RVA strains is essential in drawing relevant conclusions about their phylogeographical relationship. Therefore, continuous surveillance of RVA, following the One Health principles, may provide relevant data for assessing the impact on the protectiveness of currently available vaccines

Molekularna in epidemiološka opredelitev novih različic sesalčjega ortoreovirusa

Recently, the mammalian orthoreovirus (MRV) isolate SI-MRV01 with high similarity to newly described bat MRVs was first observed in a child with severe diarrhoea. Furthermore, a closely related MRVs were described in a dog with haemorrhagic enteritis and pigs with diarrhoea. The aim of this doctoral dissertation was to determine the prevalence of MRVs in bats, dogs and hospitalised children with acute gastroenteritis to reveal the occurrence and genetic variability of these viruses. In addition, MRV seroprevalence in dogs and children was investigated. The particular focus was on the occurrence of SI-MRV01 variants in target populations. To determine MRV prevalence, archive bat guano samples, stool samples of hospitalised children with diarrhoea and prospective dog stool samples were analysed with a broad-spectrum RT-PCR and a specific real time RT-PCR, targeting a novel bat MRV. Subsequently, MRV isolates were propagated in cell line and whole-genomes were obtained with NGS sequencing. For MRV seroprevalence study, dog sera and sera samples from children were analysed with serum-neutralisation test on MRV serotype reference strains (Reo1TL, Reo2TJ, Reo3TD) and bat isolate SI-MRV01. A novel bat MRV variant with 99% whole genome identity to isolate SI-MRV01 is present in Slovenian bats. Overall, SI-MRV01 prevalence was at 9.1%. Five independent strains were successfully isolated in cell culture following whole genome sequence determination from the archived bat guano samples. In addition to SI-MRV01-like variant four further genetically distant bat MRV variants were identified. Acute MRV infection was detected in one hospitalised child with diarrhoea (0.2%) and in zero dogs (0%). Neutralising antibodies against SI-MRV01 were detected in 7.5% children‘s sera and in 81.3% dogs‘ sera. MRVs are frequently present in bats, indicating bats as possible reservoir of circulating MRVs. Serology analysis of dog sera indicates possible route of infection via pets.Nedavno smo pri otroku s hudo drisko odkrili sesalčji ortoreovirus (MRV), poimenovan SI-MRV01, ki je izjemno soroden netopirskim MRV, na novo odkritim pri netopirjih v Italiji in Nemčiji. Poleg tega so bili sorodni MRV opisani tudi pri psu s hemoragičnim enteritisom in prašiču z drisko. Cilj te doktorske naloge je bil določiti pogostnost pojavljanja MRV pri netopirjih, psih in hospitaliziranih otrocih z akutnim gastroenteritisom. Želeli smo odkriti pojavljanje in genetsko raznolikost teh virusov. Poleg tega smo raziskali seroprevalenco MRV pri psih in otrocih. Posebno pozornost smo posvetili pojavljanju različic SI-MRV01. Za določanje pogostnosti pojavljanja MRV smo arhivske netopirske vzorce gvana, vzorce blata pri hospitaliziranih otrocih in prospektivno zbrane pasje iztrebke analizirali s široko-spektralnim RT-PCR in SI-MRV01 specifičnim RT-PCR v realnem času. Izolate MRV smo namnožili na celični liniji in analizirali njihove celotne genome s sekvenciranjem naslednje generacije. Za seroprevalenčno študijo MRV smo pasje in otroške serume analizirali s serum-nevtralizacijskim testom, pri čemer smo uporabili referenčne seve MRV (Reo1TL, Reo2TJ, Reo3TD) in netopirski izolat iz otroka SI-MRV01. Pri slovenskih netopirjih smo dokazali nove netopirske različice MRV z 99 % podobnostjo na nivoju celotnega genoma z izolatom SI-MRV01. Celokupna pogostnost pojavljanja SI-MRV01 je 9,1 %. Iz arhivskih vzorcev netopirskega gvana smo na celični kulturi uspešno osamili 5 neodvisnih izolatov in jim določili nukleotidno zaporedje celotnega genoma. Poleg različic SI-MRV01 smo določili 4 genetsko bolj oddaljene netopirske različke MRV. Akutno okužbo z MRV smo določili pri enem hospitaliziranem otroku z drisko (0,2 %) in pri nobenem psu (0 %). Nevtralizirajoča protitelesa proti izolatu SI-MRV01 smo določili pri 7,5 % otroških serumih in pri 81,3 % pasjih serumih. MRV so pogosto prisotni pri netopirjih, ki predstavljajo možen vir krožečih MRV. Serološka analiza pasjih serumov nakazuje možno okužbo z MRV preko domačih ljubljenčkov

Identification of novel reassortant mammalian orthoreoviruses from bats in Slovenia

Background: Recently, mammalian orthoreoviruses (MRVs) were detected for the first time in European bats, and the closely related strain SI-MRV01 was isolated from a child with severe diarrhoea in Slovenia. Genetically similar strains have also been reported from other mammals, which reveals their wide host distribution. The aim of this study was to retrospectively investigate the occurrence and genetic diversity of MRVs in bats in Slovenia, from samples obtained throughout the country in 2008 to 2010, and in 2012 and to investigate the occurrence of the novel SI-MRV01 MRV variant in Slovenian bats. Results: The detection of MRVs in bat guano was based on broad-range RT-PCR and specific bat MRV real-time RT-PCR. Subsequently, MRV isolates were obtained from cell culture propagation, with detailed molecular characterisation through whole-genome sequencing. Overall, bat MRVs were detected in 1.9% to 3.8% of bats in 2008, 2009 and 2012. However, in 2010 the prevalence was 33.0%, which defined an outbreak of the single SI-MRV01 strain. Here, we report on the identification of five MRV isolates of different serotypes that are designated as SI-MRV02, SI-MRV03, SI-MRV04, SI-MRV05 and SI-MRV06. There is high genetic variability between these characterised isolates, with evident genome reassortment seen across their genome segments. Conclusions: In conclusion, we have confirmed the presence of the SI-MRV01 strain in a Slovenian bat population. Moreover, according to genetic characterisation of S1 genome segment, all three MRV serotypes were present in the bat population. In this study, five independent MRV isolates were obtained and detailed whole genome analysis revealed high diversity between them. This study generates new information about the epidemiology and molecular characteristics of emerging bat MRV variants, and provides important molecular data for further studies of their pathogenesis and evolution

The impact and complete genome characterisation of viruses involved in outbreaks of gastroenteritis in a farrow-to-finish holding

Abstract Viral enteric pathogens continuously burden intensive pig farming, causing gastrointestinal diseases of epidemic and endemic nature. The present study investigated two diarrhoea outbreaks on a large farrow-to-finish holding and subsequent circulation of outbreak-related enteric viruses. These viruses were characterised by whole genome sequencing, and statistical evaluation of the impact on specific production metrics was performed. The results provided evidence that the Porcine epidemic diarrhoea virus–swine enteric coronavirus (PEDV–SeCoV) S gene recombinant strain was responsible for the first outbreak, whilst Rotavirus A (RVA) in a mixed infection with Rotavirus B (RVB) and porcine kobuvirus (PKV) probably caused the second diarrhoea outbreak. Whole genome characterisation revealed a porcine origin of all viruses involved and significant heterogeneity of RVB strain, proposing four novel genotypes and changes in RVB VP1 genotype classification. The statistical evaluation confirmed only a minor disturbance in the number of weaned pigs per sow, with statistical forecasting showing positive trends. A follow-up study corroborated the endemicity of RVA and PKV, in contrast to PEDV–SeCoV. Punctual, comprehensive and timely investigation of diarrhoea outbreaks is a prerequisite for applying adequate pig health and biosecurity management. Calculating such outbreaks' impact on production metrics can potentially shape future decisions on management improvements

4L12 zaključna razstava likovnih del študentov 4. letnika LP

Razstava 4. letnika LP PeF v Ljubljani - 2011/201

4L12 zaključna razstava likovnih del študentov 4. letnika LP

Razstava 4. letnika LP PeF v Ljubljani - 2011/201