Enterovirus Genotypes Research Articles

Epidemiology of Echovirus 30 Infections Detected in a University Hospital in Catalonia, Spain, in 1995-2020.

There is a growing interest in echovirus 30 (E30), an enterovirus responsible for neurological disease and hospitalization. There are multiple studies of outbreaks, but few that study the epidemiology over long periods of time. Our study aims to describe the clinical, epidemiological and microbiological characteristics of a series of E30 infections detected over 26 years. Data were retrospectively collected from a database of all enterovirus infections identified in our laboratory. They were detected by viral isolation or nucleic acid detection in patients presenting with respiratory or neurological infections, rash, sepsis-like syndrome, or gastroenteritis. Enterovirus genotyping was performed by amplification of the VP1 gene using RT-nested PCR, followed by sequencing and BLAST analysis. Of the 2402 enterovirus infections detected, 1619 were linked to at least one genotype and 173 were caused by E30. Clinical information was available for 158 (91.3%) patients. E30 was associated with neurological infection in 107 (67.8%) cases and it was detected almost every year. Phylogenetic analysis was performed with 67 sequences. We observed that E30 strains circulating in Catalonia from 1996 to 2016 belong to two lineages (E and F), although the majority cluster was in F. In 2018, lineage I emerged as the dominant lineage.

Intestinal microbiota has important effect on severity of hand foot and mouth disease in children

BackgroundThe incidence of hand foot and mouth disease (HFMD) has increased in recent years, making it a very common childhood illness worldwide. The relationship between different enterovirus genotypes and disease severity is not clearly understood. Given that enteroviruses are transmitted through the gastrointestinal tract, we hypothesized that variation in intestinal microorganisms of the host might play a role in the prognosis of HFMD.MethodsWe carried out a meta-transcriptomic-wide association study of fecal samples obtained from a cohort of children (254 patients, 227 tested positive for enterovirus, including 16 patients co-infectied with 2 kinds of enterovirus) with mild and severe HFMD and healthy controls.ResultsWe found there was no significant difference in the amount of each virus type between the mild and severe cases. Genes of enterovirus 71 (EV71) and coxsackievirus A (CV-A) from the severe and mild cases did not show significant clustering. Clostridium sp. L2-50 and Bacteroides stercoris ATCC 43183 were enriched in the guts of children with severe HFMD and KEGG enrichment was found between mild and severe cases.ConclusionsIntestinal microorganisms appear to interact with enterovirus to determine the progression of HFMD. Genes of Bacteroides and Clostridium may be used as predictive markers for a more efficient prognosis and intervention. The enrichment of intestinal bacteria genes with functions may facilitate the development of severe symptoms for HFMD patients.

A five-year retrospective study on the epidemiology of hand, foot and mouth disease in Sabah, Malaysia

Hand, foot, and mouth disease (HFMD) is endemic in Malaysia, with the number of cases increasing. Sabah has experienced several HFMD outbreaks, but information on the epidemiology and molecular characteristics of responsible viruses is scarce. In this study, data of 17,574 reports of HFMD cases in Sabah from 2015 to 2019 were extracted from a public health disease surveillance system and analyzed. Twenty-one swab samples from 13 children were collected from Beaufort, Sabah, during an outbreak in August 2018 for detection and serotyping of causative viruses by semi-nested reverse transcription-polymerase chain reaction (snRT-PCR) of the VP4–VP2 region and consensus degenerate hybrid oligonucleotide primer PCR of the VP1 region, respectively. Nucleotide sequencing and phylogenetic analysis were conducted by the neighbor-joining method. The average annual incidence of HFMD was 94.3 per 100,000 people, with the greatest yearly increase between 2017 and 2018. Swabs from six children were tested positive for enterovirus, of which five were positive for CVA16 and one for EV71. All CVA16 strains belonged to sub-genotype B1a, and the EV71 strain belonged to sub-genotype B5. Phylogenetic analyses indicate that enterovirus genotype shift might be responsible for the increasing trend of HFMD in Sabah, however, further study is needed.

Genotyping of non-polio enteroviruses associated with acute flaccid paralysis in Thailand in 2013 and 2014

BackgroundAcute flaccid paralysis (AFP) surveillance was conducted as part of the World Health Organization’s strategy for completely eradicating poliomyelitis and leaving non-polio enteroviruses NPEVs as one of the main potential causes of AFP. We aimed to detect NPEV in association with AFP.MethodsWe used 459 isolates reported to be Negative Polio and some NPEVs by the World Health Organization Polio Regional Reference Laboratory (Thailand), which had been obtained during polio surveillance programmes conducted in Thailand in 2013–2014. Of 459 isolates, 35 belonged to the genus Enterovirus by RT-PCR and genotyping by DNA sequencing.ResultsThis study found 17 NPEV genotypes, with 3, 13 and 1 belonging to enterovirus (EV) species A (EV-A), EV-B, and EV-C, respectively. The EV-A types identified included coxsackievirus A2 (CA2), CA4, and EV71, typically associated with hand, foot and mouth diseases. EV-B is the most prevalent cause of AFP in Thailand, while CA21 was the only type of EV-C detected. The EV-B species (13/35; 76.5%) constituted the largest proportion of isolates, followed by EV-A (3/35; 17.6%) and EV-C (1/35; 5.9%). For the EV-B species, Echovirus (E) 30 and CVB were the most frequent isolates. E30, CVB, E14, and E6 were considered endemic strains.ConclusionNPEVs, e.g. CA4, are reported for the first time in Thailand. Despite some limitations to this study, this is the first report on the circulation patterns of NPEVs associated with AFP in Thailand. AFP surveillance has unearthed many unknown NPEVs and, the cases of death due to AFP occur annually. Therefore, it is important to study NPEVs in the wake of the eradication of poliovirus in the context of the continued incidence of paralysis.

Analysis of enterovirus genotypes in the cerebrospinal fluid of children associated with aseptic meningitis in Liaocheng, China, from 2018 to 2019

BackgroundAseptic meningitis is most often caused by enteroviruses (EVs), but EVs associated with aseptic meningitis have not yet been reported in Liaocheng. The aim of this study was to determine the prevalence and genetic characteristics of EVs causing aseptic meningitis in children in Liaocheng.MethodsWe reviewed the epidemiological and clinical characteristics of 504 paediatric cases of aseptic meningitis in Liaocheng from 2018 to 2019 and analysed the phylogeny of the predominant EV types causing this disease.ResultsA total of 107 children were positive for EV in cerebrospinal fluid samples by nested PCR. Most of the positive patients were children 13 years old or younger and had symptoms such as fever, headache and vomiting (P < 0.05). The seasons with the highest prevalence of EV-positive cases were summer and autumn. The 107 EV sequences belonged to 8 serotypes, and echovirus types 18, 6 and 11 were the three dominant serotypes in Liaocheng during the 2-year study period. Phylogenetic analyses demonstrated that the E18 and E6 isolates belonged to subgenotype C2, while the E11 isolates belonged to subgenotype D5. VP1 analysis suggested that only one lineage of these three types was cocirculating in the Liaocheng region.ConclusionsThis study demonstrated the diverse EV genotypes contributing to a large outbreak of aseptic meningitis in Liaocheng. Therefore, large-scale surveillance is required to assess the epidemiology of EVs associated with aseptic meningitis and is important for the diagnosis and treatment of aseptic meningitis in Liaocheng.

Molecular Epidemiology of Enterovirus in Children with Central Nervous System Infections.

Limited recent molecular epidemiology data are available for pediatric Central Nervous System (CNS) infections in Europe. The aim of this study was to investigate the molecular epidemiology of enterovirus (EV) involved in CNS infections in children. Cerebrospinal fluid (CSF) from children (0–16 years) with suspected meningitis–encephalitis (ME) who were hospitalized in the largest pediatric hospital of Greece from October 2017 to September 2020 was initially tested for 14 common pathogens using the multiplex PCR FilmArray® ME Panel (FA-ME). CSF samples positive for EV, as well as pharyngeal swabs and stools of the same children, were further genotyped employing Sanger sequencing. Of the 330 children tested with FA-ME, 75 (22.7%) were positive for EV and 50 different CSF samples were available for genotyping. The median age of children with EV CNS infection was 2 months (IQR: 1–60) and 44/75 (58.7%) of them were male. There was a seasonal distribution of EV CNS infections, with most cases detected between June and September (38/75, 50.7%). EV genotyping was successfully processed in 84/104 samples: CSF (n = 45/50), pharyngeal swabs (n = 15/29) and stools (n = 24/25). Predominant EV genotypes were CV-B5 (16/45, 35.6%), E30 (10/45, 22.2%), E16 (6/45, 13.3%) and E11 (5/45, 11.1%). However, significant phylogenetic differences from previous described isolates were detected. No unusual neurologic manifestations were observed, and all children recovered without obvious acute sequelae. Specific EV circulating genotypes are causing a significant number of pediatric CNS infections. Phylogenetic analysis of these predominant genotypes found genetic differences from already described EV isolates.

Leading Enterovirus Genotypes Causing Hand, Foot, and Mouth Disease in Guangzhou, China: Relationship with Climate and Vaccination against EV71.

(1) Background: Assignment of pathogens to the correct genus, species, and type is vital for controlling infectious epidemics. However, the role of different enteroviruses during hand, foot, and mouth disease (HFMD) epidemics and the major contributing factors remain unknown. (2) Methods: HFMD cases from 2016 to 2018 in Guangzhou, China were collected. The relationship between HFMD cases and genotype frequency, as well as the association between genotype frequency and climate factors, were studied using general linear models. We transformed the genotype frequency to the isometric log-ratio (ILR) components included in the model. Additionally, vaccination rates were adjusted in the climate-driven models. (3) Results: We observed seasonal trends in HFMD cases, genotype frequency, and climate factors. The model regressing case numbers on genotype frequency revealed negative associations with both the ILRs of CAV16 (RR = 0.725, p < 0.001) and EV71 (RR = 0.421, p < 0.001). The model regressing genotype frequency on driven factors showed that the trends for EV71 proportions were inversely related to vaccination rate (%, β = −0.152, p = 0.098) and temperature (°C, β = −0.065, p = 0.004). Additionally, the trends for CVA16 proportions were inversely related to vaccination rate (%, β = −0.461, p = 0.004) and temperature (°C, β = −0.068, p = 0.031). The overall trends for genotype frequency showed that EV71 decreased significantly, while the trends for CVA16 increased annually. (4) Conclusions: Our findings suggest a potential pathway for climate factors, genotype frequency, and HFMD cases. Our study is practical and useful for targeted prevention and control, and provides environmental-based evidence.

Distribution of enterovirus genotypes detected in clinical samples in Hungary, 2010-2018.

This report provides the findings of a retrospective surveillance study on the emergence and circulation of enteroviruses with their associated clinical symptoms over a nine-year period detected at the National Enterovirus Reference Laboratory in Hungary between 2010-2018.Enterovirus (EV) detection and genotyping were performed directly from clinical samples. From 4,080 clinical specimens 25 EV types were identified with a median age of patients of 5 years and 68% of all cases affected children aged 10 years or younger, although infections occurred in all age-groups. In 130 cases neurological symptoms were recorded, in 123 cases the infection presented in skin related signs including hand, foot, and mouth disease (HFMD), herpangina and rash. In 2010 EV-A71 was found to cause the majority of diagnosed EV infections while in 2011 and from 2014-2018, Coxsackievirus (CV)-A6 was identified most often. Echovirus E6 accounted for the most cases in 2012 and Echovirus 30 dominated in 2013. EV-D68 was identified only in 2010 and 2013.Widespread circulation of several EV-A and EV-B viruses with occasional occurrence of EV-C and EV-D was detected. The ability of EVs to cause severe infections in sporadic cases and regular outbreaks highlight the importance of continued monitoring of circulating EV types.

Enteroviral infection in Ural Federal District and West Siberia: results of epidemiologic observation involving molecular-genetic methods

The objective of this work was to reveal the incidence patterns of enteroviral infection (EVI) in the constituent territories of Ural Federal District (UFD) and West Siberia in modern times. Materials and methods . We used the data from forms of federal statistical EVI monitoring for the period from 2017 to 2019 as well as those from the epidemiological investigation reports obtained from the disease clusters. The genotype of the enteroviruses was determined in specimens of biomaterial from 1,213 patients and their contacts using the molecular genetic method. Results and discussion . In most of the regions presented, an unfavourable epidemic situation with a trend towards further increase in the incidence still remains. Enteroviral infection has typical estivo-autumnal seasonality and age structure. Its most common clinical form is exanthematous lesion of skin and mucous membranes. The intensity and dynamics of the disease clusters formation closely correlates with the level and seasonal dynamics of the EVI incidence in the given area. A total of 36 enterovirus genotypes were detected. Most of the typed strains belonged to species A (39.8 %) and В (55.1 %), and their ratio did not change considerably from 2017 to 2019. However, the intraspecific type structure was subject to major variation. Conclusion . In most regions of UFD and West Siberia, an unfavourable epidemic situation with EVI with a trend towards further increase in the incidence still remains. Genotyping of the detected strains with subsequent phylogenetic analysis permitted more precise identification of the spectrum of the imported enterovirus strains and of those endemic for a specific area as well as evaluating the rate of their spread and their epidemic potential. Studying the variability of the enteroviral population demonstrated high intraspecific dynamics of the genotypes that maintains the intensity of the epidemic. Importation of new genovariants of Coxsackievirus B5 and echoviruses-30 and-11 with a high epidemic potential into the territory of UFD and West Siberia deserves particular attention.

Multiple genotypes of enterovirus G carrying a papain-like cysteine protease (PL-CP) sequence circulating on two pig farms in Japan: first identification of enterovirus G10 carrying a PL-CP sequence.

Two and three genotypes of enterovirus G (EV-G) carrying a papain-like cysteine protease (PL-CP) sequence were detected on two pig farms and classified into genotypes G1 and G10, and G1, G8, and G17, respectively, based on VP1 sequences. A G10 EV-G virus bearing a PL-CP sequence was detected for the first time. Phylogenetic analysis of the P2 and P3 regions grouped the viruses by farm with high sequence similarity. Furthermore, clear recombination break points were detected in the 2A region, suggesting that PL-CP EV-G-containing strains gained sequence diversity through recombination events among the multiple circulating EV-G genotypes on the farms.

Enterovirus infections in pediatric patients hospitalized with acute gastroenteritis in Chiang Mai, Thailand, 2015-2018.

BackgroundInfection with viruses especially rotavirus, norovirus, astrovirus, and adenovirus has been known to be a major cause of acute gastroenteritis in children under 5 years of age globally, particularly in developing countries. Also, some genotypes of enteroviruses (EVs) have been reported to be associated with gastroenteritis. This study is aimed to investigate the prevalence and genotype diversity of EV in children admitted to hospitals with acute gastroenteritis.MethodsA total of 1,736 fecal specimens were collected from children hospitalized with diarrhea in Chiang Mai, Thailand from 2015 to 2018. All specimens were tested for the presence of EV by RT-PCR of the 5′ untranslated region. The genotypes of EV were further identified by nucleotide sequencing and phylogenetic analysis of the viral protein 1 (VP1) gene.ResultsEV was detected in 154 out of 1,736 specimens (8.9%) throughout the study period. The prevalence of EV detected in 2015, 2016, 2017, and 2018 was 7.2%, 9.0%, 11.2%, and 8.6%, respectively. EV was detected all year round with a high prevalence during rainy season in Thailand. Overall, 37 genotypes of EV were identified in this study. Among these, coxsackievirus (CV)-A24 and CV-B5 (7.5% each), and EV-C96 (6.8%) were the common genotypes detected.ConclusionThis study demonstrates the prevalence, seasonal distribution, and genotype diversity of EV circulating in children hospitalized with acute gastroenteritis in Chiang Mai, Thailand during the period 2015 to 2018.

Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing.

Enteroviruses are small RNA viruses that affect millions of people each year by causing an important burden of disease with a broad spectrum of symptoms. In routine diagnostic laboratories, enteroviruses are identified by PCR-based methods, often combined with partial sequencing for genotyping. In this proof-of-principle study, we assessed direct RNA sequencing (DRS) using nanopore sequencing technology for fast whole-genome sequencing of viruses directly from clinical samples. The approach was complemented by sequencing the corresponding viral cDNA via Illumina MiSeq sequencing. DRS of total RNA extracted from three different enterovirus-positive stool samples produced long RNA fragments, covering between 59% and 99.6% of the most similar reference genome sequences. The identification of the enterovirus sequences in the samples was confirmed by short-read cDNA sequencing. Sequence identity between DRS and Illumina MiSeq enterovirus consensus sequences ranged between 94% and 97%. Here, we show that nanopore DRS can be used to correctly identify enterovirus genotypes from patient stool samples with high viral load and that the approach also provides rich metatranscriptomic information on sample composition for all life domains.

Genotyping of enteroviruses and human parechoviruses highlights their diversity in Northwest Ethiopia.

Enteroviruses (EVs) and human parechoviruses (HPeVs) infections are associated with various forms of disease, including gastroenteritis. As information on the molecular epidemiology of these viruses is limited in Ethiopia, the genetic diversity of EV and HPeV was investigated in the Northwestern part of the country. Of the total 450 stool samples obtained from infants and young children with diarrhea, 157 (34.9%) were positive for EV and 49 (10.9%) for HPeV RNA when tested by real-time reverse transcription polymerase chain reaction. Genotyping was performed by sequencing of the EV VP1 gene and the HPeV VP3/VP1 gene, respectively. Genotyping of EV was successful in 118 samples. Thereof, 82 (69.5%) belonged to non-polio EVs as a broad range of genotypes within species C, B, and A. Sabin polioviruses were found in 36 cases. HPeV sequences were also heterogeneous with a relative dominance of genotype 3. In conclusion, diverse EV and HPeV genotypes were found cocirculating in Northwest Ethiopia. The findings highlight the importance of continuous surveillance of these viruses in Ethiopia.

Virome diversity analysis reveals novel enteroviruses and a human picobirnavirus in stool samples from African green monkeys with diarrhea

It is important to identify viruses in animals because most infectious diseases in humans are caused by viruses of zoonotic origin. African green monkey is a widely used non-human primate model in biomedical investigations. In this study, total RNAs were extracted from stool samples of 10 African green monkeys with diarrhea. High-throughput sequencing was used to characterize viromes. PCR and Sanger sequencing were used to determine the full genome sequences. Great viral diversity was observed. The dominant viruses were enteroviruses and picobirnaviruses. Six enterovirus genomes and a picobirnavirus RNA-dependent RNA polymerase sequence were characterized. Five enteroviruses belonged to two putative new genotypes of species Enterovirus J. One enterovirus belonged to EV-A92. The picobirnavirus RNA-dependent RNA polymerase sequence had the highest nucleotide similarity (93.48%) with human picobirnavirus isolate GPBV6C2. The present study helped to identify the potential zoonotic viruses in African green monkeys. Further investigations are required to elucidate their pathogenic roles in animals and humans.

Tracking echovirus eleven outbreaks in Guangdong, China: a metatranscriptomic, phylogenetic, and epidemiological study.

In April 2019, a suspect cluster of enterovirus cases was reported in a neonatology department in Guangdong, China, resulting in five deaths. We aimed to investigate the pathogen profiles in fatal cases, the circulation and transmission pattern of the viruses by combining metatranscriptomic, phylogenetic, and epidemiological analyses. Metatranscriptomic sequencing was used to characterize the enteroviruses. Clinical and environmental surveillance in the local population was performed to understand the prevalence and genetic diversity of the viruses in the local population. The possible source(s), evolution, transmission, and recombination of the viruses were investigated by incorporating genomes from the current outbreak, from local retrospective surveillance, and from public databases. Metatranscriptomic analysis identified Echovirus 11 (E11) in three fatal cases. Seroprevalence of neutralization antibody to E11 was 35 to 44 per cent in 3–15 age groups of general population, and the viruses were associated with various clinical symptoms. From the viral phylogeny, nosocomial transmissions were identified and all E11 2019 outbreak strains were closely related with E11 strains circulating in local population 2017–19. Frequent recombination occurred among the 2019 Guangdong E11 outbreak strains and various genotypes in enterovirus B species. This study provides an example of combining advanced genetic technology and epidemiological surveillance in pathogen diagnosis, source(s), and transmission tracing during an infectious disease outbreak. The result highlights the hidden E11 circulation and the risk of viral transmission and infection in the young age population in China. Frequent recombination between Guangdong-like strains and other enterovirus genotypes also implies the prevalence of these emerging E11 strains.

Elevated circulating miR-494 in plasma of children with enterovirus 71 induced hand, foot, and mouth disease and its potential diagnostic value.

Hand, foot, and mouth disease (HFMD) is a common pediatric disease, whose outcome depends of the enterovirus genotype infecting the patient. The present study is focused on the potential diagnostic value and the role of circulating microRNA-494 (miR-494) in enterovirus 71-induced more severe form of HFMD. We included 102 children with enterovirus 71 (EV71)-induced HFMD, 42 coxsackievirus A16 (CA16)-induced HFMD and 102 healthy controls. The plasma and serum samples were collected. The expression level of circulating miR-494 was determined by RT-PCR method. Moreover, ROC curve has been drawn to evaluate the sensitivity and specificity of circulating miR-494 for the diagnosis of EV71-induced HFMD. Furthermore, the correlation between the circulating miR-494 and the levels of interleukin 6 (IL-6), interleukin 4 (IL-4) and interferon γ (IFN-γ) in the serum of patients were analyzed. Circulating miR-494 was significantly increased in plasma of children with EV71-induced HFMD compared with the healthy children or CA16-induced HFMD, and level of miR-494 in the EV71 severe group was significantly higher than the EV71 mild group. Moreover, results of ROC analysis suggested that miR-494 is a sensitive biomarker to distinguish EV71 patients from healthy controls and CA16 patients. Furthermore, IL-6 and IFN-γ were elevated in serum of patients with EV71-induced HFMD and the level of circulating miR-494 in patients with EV71-induced HFMD was positively correlated with the serum levels of both IL-6 and IFN-γ, respectively. Circulating miR-494 was abnormally up-regulated in plasma of the children with EV71-induced HFMD, and miR-494 may serve as potential biomarker for the diagnosis and treatment of the disease. Keywords: miR-494; HFMD; biomarker; enterovirus 71; inflammation.

National Epidemiology and Evolutionary History of Four Hand, Foot and Mouth Disease-Related Enteroviruses in China from 2008 to 2016.

Hand, foot and mouth disease (HFMD) is a major public health concern in China. The most predominant enteroviruses that cause HFMD have traditionally been attributed to enterovirus A71 (EVA71) and coxsackievirus A16 (CVA16). Since its first large outbreak in 2008, the dominant HFMD pathogens are constantly changing. In 2013 and 2015, CVA6 exceeded both EVA71 and CVA16 to become the leading cause of HFMD in some provinces. However, there still lacks a comprehensive overview on the molecular epidemiology and evolution of HFMD-related enteroviruses at the national level. In this study, we performed systematic epidemiological analyses of HFMD-related enteroviruses using the data of 64 published papers that met the inclusion criteria, and conducted phylogenetic analyses based on 12,080 partial VP1 sequences identified in China before 31st June 2018. We found that EVA71 prevalence has decreased sharply but other enteroviruses have increased rapidly from 2008 to 2016 and that one subtype of each enterovirus is represented during the epidemic. In addition, four genotypes EVA71_C4, CVA16_B1, CVA6_D and CVA10_C are the most predominant enterovirus strains and collectively they cause over 90% of all HFMD cases in China according to the phylogenetic trees using representative partial VP1 sequences. These four major enterovirus genotypes have different geographical distributions, and they may co-circulate with other genotypes and serotypes. These results suggest that more molecular epidemiological studies should be performed on several enteroviruses simultaneously, and such information should have implications for virological surveillance, disease management, vaccine development and policy-making on the prevention and control of HFMD.Electronic supplementary materialThe online version of this article (10.1007/s12250-019-00169-2) contains supplementary material, which is available to authorized users.

Global epidemiology of nonpolio enteroviruses causing severe neurological complications: A systematic review and meta-analysis.

Enteroviruses are RNA viruses found as commensals in the human gut and respiratory system, which may cause a wide spectrum of disease. Enteroviruses may cause severe neurologic complications including acute flaccid paralysis (AFP) and encephalitis and are the most commonly diagnosed agents of viral meningitis. Outbreaks of more severe disease are often associated with particular genotypes, such as enterovirus-A71 causing rhombencephalitis and AFP. There are more than 300 described genotypes of human enterovirus, with overlaps in clinical phenotypes between genotypes, and uncertainty about which genotypes are more prevalent in neurological manifestations. A systematic review of observational studies was conducted to evaluate the most prevalent enterovirus genotypes causing AFP, encephalitis, and meningitis. The genotyping methods and sampling sites were compiled as secondary outcomes. Sources included MEDLINE, Embase (publications until January 2019), and references selected from included studies. Meta-analyses using a random effects model were performed to calculate the pooled proportion of enterovirus genotypes in each disease. Ninety-six publications met the eligibility criteria, comprising 3779 AFP cases, 1140 encephalitis cases, and 32 810 meningitis cases. Enterovirus-A71 was most frequently associated with AFP (pooled proportion 0.12, 95% CI, 0.05-0.20) and encephalitis (0.77, 95% CI, 0.61-0.91). Echovirus 30 (0.35, 95% CI, 0.27-0.42) was the most predominant genotype in meningitis cases. Genotypes were most commonly determined using VP1 RT- reverse transcription-polymerase chain reaction, and most samples assessed were cerebrospinal fluid. With the emergence of enteroviruses as an increasing cause of neurological diseases, surveillance and testing need to increase to identify the aetiology of the most common and most severe disorders.

Rapid and Cost-Efficient Enterovirus Genotyping from Clinical Samples Using Flongle Flow Cells.

Enteroviruses affect millions of people worldwide and are of significant clinical importance. The standard method for enterovirus identification and genotyping still relies on Sanger sequencing of short diagnostic amplicons. In this study, we assessed the feasibility of nanopore sequencing using the new flow cell “Flongle” for fast, cost-effective, and accurate genotyping of human enteroviruses from clinical samples. PCR amplification of partial VP1 gene was performed from multiple patient samples, which were multiplexed together after barcoding PCR and sequenced multiple times on Flongle flow cells. The nanopore consensus sequences obtained from mapping reads to a reference database were compared to their Sanger sequence counterparts. Using clinical specimens sampled over different years, we were able to correctly identify enterovirus species and genotypes for all tested samples, even when doubling the number of barcoded samples on one flow cell. Average sequence identity across sequencing runs was >99.7%. Phylogenetic analysis showed that the consensus sequences achieved with Flongle delivered accurate genotyping. We conclude that the new Flongle-based assay with its fast turnover time, low cost investment, and low cost per sample represents an accurate, reproducible, and cost-effective platform for enterovirus identification and genotyping.

EPIDEMIOLOGIC FEATURES OF CLUSTERED MORBIDITY CAUSED BY ENTEROVIRAL INFECTIONS IN THE URAL FEDERAL DISTRICT AND WESTERN SIBERIA IN 2017

Aim. Delineation of epidemiologic features of clustered morbidity caused by enteroviral infections (EVI) in the Ural Federal District (UFD) and West Siberia in 2017. Materials and methods. Retrospective epidemiologic analysis of clustered EVI morbidity was performed with the data obtained from the standardized Federal statistical observation reports and the data provided by the regional Departments of the Federal Service for Supervision of Consumer Rights Protection and Human Well-Being (Rospotrebnadzor). Detection and identification of enteroviruses was carried out by molecular genetic methods. Results. Totally 51 focus of clustered EVI was registered. The incidence density and time course of outbreaks closely correlated with the general EVI incidence level (r = 0.65; p &lt; 0.05) and seasonal variations (r = 0.80; p &lt; 0.05) at the given territory. Droplet nuclei transmission was considered the main route of infection in 68.2% of cases, in 22.0% - as an accessory route, and in 14.6% - as an obligate route. Contact transmission via fomites was reported as an obligate route in 9.8% of outbreaks. In the etiological structure, the proportion of Enterovirus A species was 51.4% (Coxsackievirus A6 (CV-A6) - 43.2%), Enterovirus B species - 70.3% (Echovirus 30 (E30) - 48.6%). Conclusion. In 2017 the clustered morbidity caused by EVI in the UFD and Western Siberia involved mainly children under 6 years of age from preschool institutions, where outbreaks of low incidence density with short duration were observed. Two genotypes of enteroviruses (E30 and CV-A6) were prevalent in the etiological structure of the outbreaks, therefore determining predominant clinical forms of EVI: aseptic meningitis or hand, foot, and mouth disease.