Enterovirus Strains Research Articles

Detection of enterovirus RNA in pancreas and lymphoid tissues of organ donors with type 1 diabetes.

The nPOD-Virus group collaboratively applied innovative technologies to detect and sequence viral RNA in pancreas and other tissues from organ donors with type 1 diabetes. These analyses involved the largest number of pancreas samples collected to date. We analysed pancreas, spleen, pancreatic lymph nodes, and duodenum samples from the following donor groups: a) donors with type 1 diabetes (n=71), with (n=35) or without (n=36) insulin-containing islets, (b) donors with single or double islet autoantibody positivity without diabetes (n=22) and c) autoantibody-negative donors without diabetes (control donors) (n=74). Five research laboratories participated in this collaborative effort using approaches for unbiased discovery of RNA viruses (two RNA-Seq platforms), targeted detection of Enterovirus A-D species using RT-PCR, and tests for virus growth in cell-culture. Direct RNA-Seq did not detect virus signal in pancreas samples, whereas RT-PCR detected enterovirus RNA confirmed by sequencing in low amounts in pancreas samples in three of the five donor groups, namely donors with type 1 diabetes with insulin-containing islets, 16% (5/32) donors being positive, donors with single islet autoantibody positivity with 53% (8/15) donors being positive, and non-diabetic donors with 8% (4/49) being enterovirus RNA positive. Detection of enterovirus RNA was significantly more frequent in single islet autoantibody-positive donors compared to donors with type 1 diabetes with insulin-deficient islets (p-value <0.001) and control donors (p-value 0.004). In some donors, pancreatic lymph nodes were also positive. RT-PCR detected enterovirus RNA also in spleen of a small number of donors and virus enrichment in susceptible cell lines before RT-PCR resulted in much higher rate in spleen positivity, particularly in donors with type 1 diabetes. Interestingly, the enterovirus strains detected did not cause a typical lytic infection, possibly reflecting their persistence-prone nature. This was the largest coordinated effort to examine the presence of enterovirus RNA in pancreas of organ donors with type 1 diabetes, using a multitude of assays. These findings are consistent with the notion that both the subjects with type 1 diabetes and those with islet autoantibodies may carry a low-grade enterovirus infection in the pancreas and lymphoid tissues.

Receptors and Host Factors for Enterovirus Infection: Implications for Cancer Therapy.

Enteroviruses, with their diverse clinical manifestations ranging from mild or asymptomatic infections to severe diseases such as poliomyelitis and viral myocarditis, present a public health threat. However, they can also be used as oncolytic agents. This review shows the intricate relationship between enteroviruses and host cell factors. Enteroviruses utilize specific receptors and coreceptors for cell entry that are critical for infection and subsequent viral replication. These receptors, many of which are glycoproteins, facilitate virus binding, capsid destabilization, and internalization into cells, and their expression defines virus tropism towards various types of cells. Since enteroviruses can exploit different receptors, they have high oncolytic potential for personalized cancer therapy, as exemplified by the antitumor activity of certain enterovirus strains including the bioselected non-pathogenic Echovirus type 7/Rigvir, approved for melanoma treatment. Dissecting the roles of individual receptors in the entry of enteroviruses can provide valuable insights into their potential in cancer therapy. This review discusses the application of gene-targeting techniques such as CRISPR/Cas9 technology to investigate the impact of the loss of a particular receptor on the attachment of the virus and its subsequent internalization. It also summarizes the data on their expression in various types of cancer. By understanding how enteroviruses interact with specific cellular receptors, researchers can develop more effective regimens of treatment, offering hope for more targeted and efficient therapeutic strategies.

Loop-mediated isothermal amplification (LAMP) for detection of atypical enterovirus D68 strain VR-1197

A method that has rapidly evolved for detection of viral pathogens are loop-mediated isothermal amplification (LAMP) assays. The available LAMP assays usually target the most common viral strains, including enteroviruses, but for the atypical enterovirus D68 strain VR-1197 this method has not yet been developed. Enterovirus D68 are known for severe respiratory distress in children, and atypical strains are less likely to be detected by traditional methods. This study targets the atypical EVD68 strain VR-1197 and have developed a rapid detection method saving time when differentiating enterovirus strains. This study present method development and review the sensitivity and specificity compared to traditional RT-qPCR, and wet lab cross reactivity with other airway pathogens. The EVD68 VR-1197 assay can be a rapid POC (Point of care) test for atypical EVD68 VR-1197 and have the potential as reliable detection method with minimal technological requirements.

Changing Epidemiology of Hand, Foot, and Mouth Disease Causative Agents and Contributing Factors.

Hand, foot, and mouth disease (HFMD) is a common viral infection primarily affecting children. It causes vesicles on the skin and inside the mouth. Although most cases get better on their own, severe cases can lead to complications such as brain stem encephalitis, meningoencephalitis, acute flaccid paralysis, and pulmonary edema. Hand, foot, and mouth disease is caused by various enteroviruses, with enterovirus A71 (EV-A71) and coxsackievirus A16 being the most common. However, recent studies have shown a shift in the molecular epidemiology of HFMD-causing pathogens, with coxsackievirus A6 and coxsackievirus A10 causing more infections. In addition, extensive recombination events have been identified among enterovirus strains, which may have a role in faster evolution and extinction of dominant enterovirus serotypes. Other strains of enterovirus can also cause severe complications, and there has been an increase in mortality associated with brain stem encephalitis in children under 3 years of age and teenagers. Currently, there are no effective antiviral therapies available to treat enterovirus infections. Vaccines against EV-A71 have been approved and are now used in mainland China. Studying the changing epidemiology of HFMD pathogens and the evolution patterns of its causative agents is crucial in developing effective prevention and control strategies. Increased interest in the molecular epidemiology of HFMD causative agents has led to a better understanding of the critical drivers of HFMD outbreaks, which can inform efforts to prevent and control the disease.

Detection and type identification of non-polio enteroviruses in children against the background of acute intestinal infections of various etiologies: 2018–2023

Introduction. Enteroviruses (EV) are characterized by: species and type diversity, polymorphism of clinical manifestations, a tendency to epidemic spread, and are often the cause of disease outbreaks, which determines the relevance of monitoring EV strains in various clinical forms of infection, including in conditions of anti-epidemic measures. The aim of the study: to characterize the prevalence and diversity of non-polio enteroviruses (NPEV) types in children with acute intestinal infection (AII) in the period 2018–2023, including the COVID-19 pandemic. Materials and methods. The RT-PCR method was used to study 7302 samples of feces from children hospitalized with a diagnosis of AII in the infectious diseases hospital of Nizhny Novgorod. Genotyping of EV strains was carried out using fragment Sanger sequencing of the genome region encoding capsid protein 1 (VP1) and the online resource BLAST. Results. EVs were found in 5.0 ± 0.3% (1.7–7.8%), both in mono- and mixed infections with other enteric viruses. The long-term dynamics of the frequency of EV detection and the incidence of EV infection in children in the Nizhny Novgorod region was characterized by a sharp decrease in indicators in 2020 against the backdrop of the introduction of anti-epidemic measures. When genotyping 299 strains, 41 types of NPEV of 4 species were identified. The spectrum included the main pathogens of exanthema and neuroinfections and rare types found in “minor” or intestinal forms of infection. During the study period, a redistribution of NPEV species was established. Before the pandemic, the ratio of Enterovirus A : Enterovirus B : Enterovirus C species was as follows — 41.0 : 46.7 : 12.3%; during the 2020 pandemic season the ratio was 0.0 : 37.5 : 62.5%; after the lifting of restrictive measures — 47 : 29 : 23%, which may be due to the different effectiveness of the restrictive measures on the mechanisms of transmission of EVs of different types. Conclusion. The genetic diversity of NPEVs detected in children with AII complements information on the typical composition of the territorial enterovirus population. In children with AII, when the airborne transmission of SARS-CoV-2 was blocked, there was a decrease in the frequency of detection of viruses of the Enterovirus B type, the absence of detection of Enterovirus A and the constant presence of Enterovirus C.

Molecular Genotyping of Circulating Enterovirus in the Lazio Region from 2012 to 2023.

Enteroviruses (EVs) are ubiquitous viruses that circulate worldwide, causing sporadic or epidemic infections, typically during the summer and fall. They cause a broad spectrum of illnesses, ranging from an unspecified febrile clinical presentation to a severe illness. EVs are recognized to be the most frequent etiological agents of aseptic meningitis in children. However, as the infection is usually mild and self-limiting, it remains underestimated, and the epidemiology of EVs is poorly understood. To date, no vaccine or effective therapy for all types of enteroviruses is available, and EVs constitute a public health concern. Here, we investigated the molecular epidemiology of EV strains circulating in the Lazio region over a 10-year time span (2012-2023) by using a sequence-typing approach and phylogenetic analysis. The epidemiological trend of EV infection has undergone changes during the SARS-CoV-2 pandemic (2020-2021), which resulted in a modification in terms of the number of diagnosed cases and seasonality. From 2022, the circulation of EVs showed a behavior typical of the pre-pandemic period, although changes in predominantly circulating strains have been noted. Both epidemic and sporadic circulation events have been characterized in the Lazio region. Further analyses are needed to better characterize any strain with higher potential pathogenic power and to identify possible recombinant strains.

Enterovirus 71 Activates Plasmacytoid Dendritic Cell-Dependent PSGL-1 Binding Independent of Productive Infection.

Enterovirus 71 (EV71) is a significant causative agent of hand, foot, and mouth disease, with potential serious neurologic complications or fatal outcomes. The lack of effective treatments for EV71 infection is attributed to its elusive pathogenicity. Our study reveals that human plasmacytoid dendritic cells (pDCs), the main type I IFN-producing cells, selectively express scavenger receptor class B, member 2 (SCARB2) and P-selectin glycoprotein ligand 1 (PSGL-1), crucial cellular receptors for EV71. Some strains of EV71 can replicate within pDCs and stimulate IFN-α production. The activation of pDCs by EV71 is hindered by Abs to PSGL-1 and soluble PSGL-1, whereas Abs to SCARB2 and soluble SCARB2 have a less pronounced effect. Our data suggest that only strains binding to PSGL-1, more commonly found in severe cases, can replicate in pDCs and induce IFN-α secretion, highlighting the importance of PSGL-1 in these processes. Furthermore, IFN-α secretion by pDCs can be triggered by EV71 or UV-inactivated EV71 virions, indicating that productive infection is not necessary for pDC activation. These findings provide new insights into the interaction between EV71 and pDCs, suggesting that pDC activation could potentially mitigate the severity of EV71-related diseases.

Diversity of enteroviruses in cerebrospinal fluid specimens collected from hospitalised patients in the private and public sector in South Africa.

Enteroviruses cause a wide range of neurological illnesses such as encephalitis, meningitis, and acute flaccid paralysis. Two types of enteroviruses, echovirus E4 and E9, have recently been detected in South Africa and are known to be associated with meningitis and encephalitis. The objective of this study was to characterize enterovirus strains detected in cerebrospinal fluid specimens of hospitalized patients in the private and public sector to identify genotypes associated with meningitis and encephalitis. From January 2019 to June 2021 enterovirus positive nucleic acid samples were obtained from a private (n = 116) and a public sector (n = 101) laboratory. These enteroviruses were typed using a nested set of primers targeting the VP1 region of the enterovirus genome, followed by Sanger sequencing and BLASTn analysis. Forty-two percent (91/217) of the strains could be genotyped. Enterovirus B species was the major species detected in 95% (86/91) of the specimens, followed by species C in 3% (3/91) and species A in 2% (2/91) of the specimens. Echovirus E4 and E9 were the two major types identified in this study and were detected in 70% (64/91) and in 10% (9/91) of specimens, respectively. Echovirus E11 has previously been identified in sewage samples from South Africa, but this study is the first to report Echovirus E11 in cerebrospinal fluid specimens from South African patients. The genotypes identified during this study are known to be associated with encephalitis and meningitis. The predominant detection of echovirus E4 followed by E9 corresponds with other studies conducted in South Africa.

Metatranscriptomics revealed the molecular characterization of circulating enterovirus strains causing aseptic meningitis in children in Wuxi, China

Enteroviruses are major etiological agents of aseptic meningitis globally, however information on circulating enterovirus types associated with this disease in Wuxi, China is limited. In this study, cerebrospinal fluid samples were collected from 20 pediatric aseptic meningitis cases in a Wuxi hospital in 2020 and subjected to metagenomic analysis to detect pathogens. Enterovirus B was detected in 9 cases, including 7 echovirus 18 (E18) and 2 echovirus 11 (E11) strains. The E18 strains exhibited 87.5–98.2% nucleotide identity and phylogenetically clustered with other China E18 strains, while the E11 strains showed 97.59% identity and clustered within the D5 subgroup along with other China E11 strains. One E18 strain was identified as a novel recombinants with a distinct recombination breakpoint within 3D gene. These findings expand knowledge on enteroviruses associated with pediatric aseptic meningitis in Wuxi, and highlight the circulation of genetically diverse E18 and E11 strains, including novel E18 recombinants. Characterization of enterovirus diversity by metagenomic analysis is important for molecular diagnosis and epidemiological tracking of aseptic meningitis cases. Continued surveillance of circulating enterovirus strains in Wuxi that may cause future outbreaks is warranted.

Genetic diversity and recombination of bovine enterovirus strains in China.

Bovine enterovirus (BEV) infection is an emerging disease in China that is characterized by digestive, respiratory, and reproductive disorders. In this study, we first reported two novel EV-E subtypes detected in cattle herds in China, unveiled the coinfection of two enterovirus species (EV-E/EV-F) and different subtypes (EV-E2/EV-E7, EV-E1/EV-E7, and EV-E3/EV-E6) in the same cattle herds, and revealed the enterovirus genetic exchange in intraspecies and interspecies recombination. These results provide an important update of enterovirus prevalence and epidemiological aspects and contribute to a better understanding of enterovirus genetic diversity, evolution, and pathogenesis.

Attenuating RNA Viruses with Expanded Genetic Codes to Evoke Adjustable Immune Response in PylRS-tRNACUAPyl Transgenic Mice.

Ribonucleic acid (RNA) viruses pose heavy burdens on public-health systems. Synthetic biology holds great potential for artificially controlling their replication, a strategy that could be used to attenuate infectious viruses but is still in the exploratory stage. Herein, we used the genetic-code expansion technique to convert Enterovirus 71 (EV71), a prototypical RNA virus, into a controllable EV71 strain carrying the unnatural amino acid (UAA) Nε-2-azidoethyloxycarbonyl-L-lysine (NAEK), which we termed an EV71-NAEK virus. After NAEK supplementation, EV71-NAEK could recapitulate an authentic NAEK time- and dose-dependent infection in vitro, which could serve as a novel method to manipulate virulent viruses in conventional laboratories. We further validated the prophylactic effect of EV71-NAEK in two mouse models. In susceptible parent mice, vaccination with EV71-NAEK elicited a strong immune response and protected their neonatal offspring from lethal challenges similar to that of commercial vaccines. Meanwhile, in transgenic mice harboring a PylRS-tRNACUAPyl pair, substantial elements of genetic-code expansion technology, EV71-NAEK evoked an adjustable neutralizing-antibody response in a strictly external NAEK dose-dependent manner. These findings suggested that EV71-NAEK could be the basis of a feasible immunization program for populations with different levels of immunity. Moreover, we expanded the strategy to generate controllable coxsackieviruses for conceptual verification. In combination, these results could underlie a competent strategy for attenuating viruses and priming the immune system via artificial control, which might be a promising direction for the development of amenable vaccine candidates and be broadly applied to other RNA viruses.

Cost-effective complete genome sequencing using the MinION platform for identification of recombinant enteroviruses.

Enteroviruses (EVs) are a group of viruses that cause various human illnesses. While the CODEHOP (COnsensus-DEgenerate Hybrid Oligonucleotide Primer) method can generate VP1 gene fragments for enterovirus genotyping, it is unable to detect recombinant strains. Recent advances in viral genome sequencing using next-generation sequencing technologies have enabled comprehensive analyses. However, the high cost poses a challenge for widespread adoption. To address this issue, this study proposes a cost-effective approach for generating complete enterovirus genome sequences using the Oxford Nanopore MinION sequencer. This protocol not only facilitates the generation of accurate genome sequences for various enterovirus strains but also allows for the differentiation of co-infections from viral isolates. In addition, the method can generate polyprotein sequences as well as peptide sequences of the upstream ORF (uORF) whose expression can impact virus infection. Through the analysis of complete enterovirus genomes, this study successfully identified seven enterovirus A71 isolates obtained during the 2018 enterovirus outbreak in Malaysia and Taiwan as recombinants between enterovirus A71 and coxsackievirus A2. Furthermore, our study has made a significant discovery by establishing a strong correlation between uORF trees and the epidemics of EVA71. This finding highlights the potential of uORF sequences as valuable indicators for monitoring and understanding the spread of EVA71 infections. We also identified notable amino acid changes in the transmembrane domain of the uORF protein within a newly identified lineage. These findings provide crucial insights into the molecular characteristics and evolutionary dynamics of EVA71, offering valuable information for future research and intervention strategies. IMPORTANCE By employing a cost-effective approach for complete genome sequencing, the study has enabled the identification of novel enterovirus strains and shed light on the genetic exchange events during outbreaks. The success rate of genome sequencing and the scalability of the protocol demonstrate its practical utility for routine enterovirus surveillance. Moreover, the study's findings of recombinant strains of EVA71 and CVA2 contributing to epidemics in Malaysia and Taiwan emphasize the need for accurate detection and characterization of enteroviruses. The investigation of the whole genome and upstream ORF sequences has provided insights into the evolution and spread of enterovirus subgenogroups. These findings have important implications for the prevention, control, and surveillance of enteroviruses, ultimately contributing to the understanding and management of enterovirus-related illnesses.

Molecular characterization and pathogenicity evaluation of enterovirus G isolated from diarrheic piglets.

Four enterovirus G (EV-G) strains were isolated from diarrheic piglets that were negative for common swine enteric viruses. The spherical enterovirus particles of roughly 30-nm diameter were observed under transmission electron microscopy by using plaque-purified enterovirus. The complete genome sequence analysis revealed that each of four enteroviruses contained a papain-like cysteine protease (PLCP) gene between the 2C and 3A junction regions of the viral genome. This insertion encoded a predicted protease similar to the PLCP of porcine torovirus. The phylogenetic analysis based on complete genome with and without PLCP gene revealed that the four isolated EV-G strains were grouped together with global enterovirus G1-PLCP strains, and more closely related to EV-G/PLCP strains previously detected in China, Japan, and Korea (90.3%-92.2% similarities based on nucleotides). The cell susceptibility test demonstrated that the isolated EV-G could infect and replicate in cell lines from various host species. Furthermore, pathogenicity evaluation showed that the isolated EV-Gs induced mild diarrhea, pyrexia, and reduced body weight in infected piglets. The epidemiological investigation revealed a high prevalence of EV-G in swine herds. Together, our findings demonstrate that the isolated EV-G is pathogenic in piglets and may be advantageous in providing more trustworthy data on the evolution and pathological properties of EV-G. IMPORTANCE Enterovirus G is a species of positive-sense single-stranded RNA viruses associated with several mammalian diseases. The porcine enterovirus strains isolated here were chimeric viruses with the PLCP gene of porcine torovirus, which grouped together with global EV-G1 strains. The isolated EV-G strain could infect various cell types from different species, suggesting its potential cross-species infection risk. Animal experiment showed the pathogenic ability of the isolated EV-G to piglets. Additionally, the EV-Gs were widely distributed in the swine herds. Our findings suggest that EV-G may have evolved a novel mechanism for broad tropism, which has important implications for disease control and prevention.

Features of the etiological structure of non-polio enterovirus infection in children in the pre-pandemic and post-pandemic periods

BACKGROUND: Regional features of the etiological structure of nonpolio enterovirus infection (NPEVI) in children in the pre- and post-pandemic periods must be registered.&#x0D; AIM: To determine the features of the etiological structure of NPEVI in children in the pre- and post-pandemic periods in the KhantyMansiysk Autonomous Okrug.&#x0D; MATERIALS AND METHODS: Statistical observation data for 20132022 and results of molecular genetic studies of the structure of NPEVI in children in the pre- and post-pandemic periods were analyzed.&#x0D; RESULTS: The tendency of the incidence of NPEVI in the KhantyMansiysk Autonomous Okrug to increase in the pre-COVID-19 pandemic and a predominance of children aged 6 years has been established. In the study period up to 2019 (pre-pandemic), enteroviruses Coxsackie A6 in combination with E A71 were dominant, for which the calculated trends of the short-term forecast were confirmed in 2022. In the same year, these enteroviruses were the most common causes of focal diseases with exanthemic forms of NPEVI. In 2022, the circulation of eight more enteroviruses genovariants was recorded, among which Coxsackie A10 EV prevailed (20.5% of cases), which caused the majority of enterovirus vesicular pharyngitis cases. Enterovirus meningitis was registered in 5 (11.4%) children with the release of ECHO 6, ECHO 9, and Coxsackie A9, which were not noted in other clinical forms and circulated irregularly and in smaller numbers during the pre-pandemic period than in post-pandemic period (2022).&#x0D; CONCLUSIONS: The analysis of the monitoring data of the NPEVI in KhantyMansiysk Autonomous Okrug revealed a trend of a gradual increase in incidence in children, which must be considered when organizing epidemiological surveillance of NPEVI. The completeness of the registration of NPEVI cases with mandatory genotyping of the detected enterovirus strains will allow to more accurately determine the spectrum of strains endemic to this territory, assess the probability of their spread, forecast about the true level of morbidity, and conduct appropriate scientifically based antiepidemic and preventive measures among children with NPEVI in KhantyMansiysk Autonomous Okrug.

Potential of a bivalent vaccine for broad protection against enterovirus 71 and coxsackie virus 16 infections causing hand, foot, and mouth disease

Hand, foot, and mouth disease (HFMD) is a highly contagious viral infection that is mainly caused by enterovirus 71 (EV71) and coxsackievirus 16 (CVA16). As there are no specific therapeutics for HFMD, the development of a bivalent vaccine is required to cover a broad range of infections. In this study, the effectiveness of novel monovalent and bivalent vaccines targeting EV71 C4a and CVA16 was investigated for their ability to prevent viral infections in neonatal human scavenger receptor class B member 2 (hSCARB2) transgenic mice. As hSCARB2 serves as a key viral receptor for EV71, these transgenic mice are susceptible to EV71 strains and facilitate viral binding, internalization, and uncoating processes. Antisera prepared by vaccine immunization were transferred to 2-day-old hSCARB2 transgenic mice, which were then infected with EV71 C4a or CVA16 virus. The antisera generated by each monovalent or bivalent vaccine effectively protected against EV71 C4a and CVA16 infections. The examination of tissue damage and viral contents in various organs indicated that both monovalent and bivalent antisera reduced EV71 C4a viral load in the brainstem, and no significant tissue damage was observed. During CVA16 infection, the monovalent and bivalent antisera significantly reduced viral contents in both the brainstem and muscles. These results suggest that passive immunity by monovalent and bivalent antisera can effectively protect against EV71 C4a and CVA16 infections. Thus, the development of a bivalent vaccine that can provide broad protection against both CV and EV infections may be a promising strategy in preventing HFMD.

Wastewater-based epidemiology revealed in advance the increase of enterovirus circulation during the Covid-19 pandemic

Wastewater-based epidemiology (WBE) was conducted to track Enteroviruses (EVs) circulation in the Milan metropolitan area (Northern Italy) during Covid-19 pandemic (March 2020–December 2022).202 composite 24-hour wastewater samples (WWSs) were collected weekly from March 24, 2020, to December 29, 2022 at the inlet of two wastewater treatment plants (WWTP) in Milan (1.5 million inhabitants). EV-RNA was quantified and molecular characterization of non-polio EVs (NPEV) was performed by Sanger sequence analysis. Data from WWS were matched with virological data collected in the framework of Influenza-Like Illness (ILI) surveillance in the same place and time.EV-RNA was identified in 88.2 % of WWSs. The peak in EVs circulation was observed in late August 2020 (upon conclusion of the first national lockdown), in late August 2021, and in mid-April 2022.EV-RNA concentration in WWS (normalized as copies/d/1000 people) at peak of circulation presented a yearly increase (2020: 2.47 × 1010; 2021: 6.81 × 1010; 2022: 2.14 × 1011). This trend overlapped with trend in EV-positivity rate in ILI cases, expanded from 21.7 % in 2021 to 55.6 % in 2022. EV trends in WWS preceded clinical sample detections in 2021 and 2022 by eight and five weeks, respectively, acting as an early warning of outbreak.Although sequencing of EV-positive WWSs revealed the presence of multiple EV strains, typing remained inconclusive. Molecular characterization of EVs in clinical samples revealed the co-circulation of several genotypes: EV-A accounted for 60 % of EVs, EV-B for 16.7 %, EV-D68 for 23.3 %.EVs were circulating in Milan metropolitan area between March 2020 and December 2022. The epidemiological trends unfolded the progressive accumulation of EV transmission in the population after removal of Covid-19 restrictions. The increased circulation of EVs in 2021–2022 was identified at least 35 days in advance compared to the analysis of clinical data. The inconclusive results of Sanger sequencing lookout for improvement and innovative molecular approaches to deepen track EVs.

Phylogeographic dynamics and molecular characteristics of Enterovirus 71 in China.

Enterovirus 71 (EV71) and coxsackievirus (CV-A16) are the major etiological agents of hand, foot and mouth disease (HFMD). This report reviewed the full-length genomic sequences of EV71 identified in different provinces of China between 1998 and 2019 (a total of 312) in addition to eight worldwide reference genomes to address the genomic evolution and genetic events. The main prevalent EV71 strians in China are C4 genotypes, co-circulating with a few A, B5, C1, and C2 subgenotypes. A new emerging subgenotype in China was identified and classified as B6 genotype. Phylogeographic analysis revealed multiple branches, where a Jiangsu strain 2006-52-9 (GenBank ID: KP266579.1) was linked to different subgenotypes through multiple long mutant branches, including the CV-A16 viruses through the A genotype. Furthermore, identification of 28 natural recombination events suggests that the emergence of new genotypes are associated with intratypic recombination involving EV71 strains and intertypic recombination between EV71 and CV-A16 strains. Compared with the structural proteins, the non-structural proteins of EV71 seem to be highly variable with the highest variable regions of peptidase C3 (3C protein), P2A, and the N-terminus of RNA-dependent RNA polymerase. This study updates the phylogenetic and phylogeographic information of EV71 and provides clues to the emergence of new genotypes of EV71 based on genetics.

Identification of a Conserved, Linear Epitope on VP3 of Enterovirus A Species Recognized by a Broad-Spectrum Monoclonal Antibody.

Outbreaks of hand, foot and mouth disease (HFMD) have occurred frequently in the Asian-Pacific region over the last two decades, caused mainly by the serotypes in Enterovirus A species. High-quality monoclonal antibodies (mAbs) are needed to improve the accuracy and efficiency of the diagnosis of enteroviruses associated HFMD. In this study, a mAb 1A11 was generated using full particles of CV-A5 as an immunogen. In indirect immunofluorescence and Western blotting assays, 1A11 bound to the viral proteins of CV-A2, CV-A4, CV-A5, CV-A6, CV-A10, CV-A16, and EV-A71 of the Enterovirus A and targeted VP3. It has no cross-reactivity to strains of Enterovirus B and C. By mapping with over-lapped and truncated peptides, a minimal and linear epitope 23PILPGF28 was identified, located at the N-terminus of the VP3. A BLAST sequence search of the epitope in the NCBI genus Enterovirus (taxid: 12059) protein database indicates that the epitope sequence is highly conserved among the Enterovirus A species, but not among the other enterovirus species, first reported by us. By mutagenesis analysis, critical residues for 1A11 binding were identified for most serotypes of Enterovirus A. It may be useful for the development of a cost-effective and pan-Enterovirus A antigen detection for surveillance, early diagnosis and differentiation of infections caused by the Enterovirus A species.

Coxsackievirus A11 is an immunostimulatory oncolytic virus that induces complete tumor regression in a human non-small cell lung cancer

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality worldwide. Innovative treatment is required to improve overall survival rates for advanced NSCLC. Oncolytic virotherapy using enteroviruses has emerged as a promising anticancer strategy. To identify a novel, potent virotherapy with an improved safety profile, we assessed the oncolytic activity of 28 enteroviral strains and focused on coxsackievirus A11 (CVA11). CVA11 infection caused extensive oncolytic activity in all three of the examined human NSCLC cell lines, with high intercellular adhesion molecule-1 (ICAM-1) expression associated with greater CVA11-induced cytotoxicity. In vitro inhibition analysis using a pan-caspase inhibitor and western blot detection of cleaved poly (ADP-ribose) polymerase (PARP) indicated that apoptosis partly contributed to CVA11-driven cytotoxicity. CVA11 infection-induced immunogenic cell death in vitro was strongly suggested by substantial calreticulin expression and release of high mobility group box-1 protein (HMGB1). Moreover, in vivo treatment of human NSCLC xenografts with intratumoral CVA11 injection caused complete tumor regression in all treated mice, without significant weight loss. Our findings indicate that novel oncolytic virotherapy utilizing CVA11 may be less toxic and more effective than current treatments for human NSCLC, thus warranting further investigation in clinical trial settings, especially in combination with immunotherapy.

Molecular epidemiological and clinical aspects of enterovirus infection in the south of Russia

Objective: the study of molecular epidemiological and clinical aspects of EVI in the Rostov region for the period 2006-2020. to improve the disease surveillance system. Research methods: epidemiological, virological, molecular biological (PCR, sequencing, phylogenetic analysis), microbiological, statistical. Materials and methods: the analysis of medical records of 139 patients with EVI was carried out. Samples of biomaterial (feces, throat swab) from 17293 people, samples from environmental objects (2710 samples), strains of enteroviruses (EV) in the amount of 142 specimens were studied. Results: in the Rostov region until 2013. sporadic incidence of EVI prevailed with registration from 1 to 38 cases per year. June 2013 there was a sharp increase in the incidence of EVI with the formation of local foci in organized groups, associated with the circulation of a new genotype — EV 71 type of subgenotype C4 of “Chinese” origin (622 patients with EVI were registered, one death). The clinical features of the disease were determined: acute onset with manifestations of intoxication, foot and mouth disease-like syndromes, followed by the development of CNS pathology in 37.4% of patients. According to the results of sequencing of biomaterial samples from patients and virus carriers for the period 2006-2020. EVs of 22 types were detected. Conclusions: EVs are subject to intense genetic variability, due to which new genovariants pathogenic for humans may appear. The change in EV genotypes, which dominated the circulation among the population of the Rostov region, determined the rise in the incidence of EVI in 2013. A significant diversity of non-polio EV genotypes was revealed, while the structure of EV genovariants changed in different years.