Echovirus Research Articles

The combination of pleconaril, rupintrivir, and remdesivir efficiently inhibits enterovirus infections in vitro, delaying the development of drug-resistant virus variants

Enteroviruses are a significant global health concern, causing a spectrum of diseases from the common cold to more severe conditions like hand-foot-and-mouth disease, meningitis, myocarditis, pancreatitis, and poliomyelitis. Current treatment options for these infections are limited, underscoring the urgent need for effective therapeutic strategies. To find better treatment option we analyzed toxicity and efficacy of 12 known broad-spectrum anti-enterovirals both individually and in combinations against different enteroviruses in vitro. We identified several novel, synergistic two-drug and three-drug combinations that demonstrated significant inhibition of enterovirus infections in vitro. Specifically, the triple-drug combination of pleconaril, rupintrivir, and remdesivir exhibited remarkable efficacy against echovirus (EV) 1, EV6, EV11, and coxsackievirus (CV) B5, in human lung epithelial A549 cells. This combination surpassed the effectiveness of single-agent or dual-drug treatments, as evidenced by its ability to protect A549 cells from EV1-induced cytotoxicity across seven passages. Additionally, this triple-drug cocktail showed potent antiviral activity against EV-A71 in human intestinal organoids. Thus, our findings highlight the therapeutic potential of the pleconaril-rupintrivir-remdesivir combination as a broad-spectrum treatment option against a range of enterovirus infections. The study also paves the way towards development of strategic antiviral drug combinations with virus family coverage and high-resistance barriers.

Analysis of the distribution characteristics of enterovirus types based on environmental surveillance from 2013 to 2021 in Fujian Province, China

Environmental surveillance (ES) is a useful approach for monitoring circulating viruses, including polioviruses (PVs) and non-polio enteroviruses (NPEVs). In this study, the results of nine years of ES from 2013 to 2021 at six sampling sites in three cities in Fujian Province, China, were summarized. It showed that the sewage samples contained abundant viruses, but the positive rate was affected by different sampling sites. From the 520 samples, 431 PVs, 1,713 NPEVs, and 281 human adenoviruses (HAdVs) were isolated. PV isolates had been markedly affected following the adjustment of the immunization strategy. All but one PV isolate were Sabin-like strains without wild PVs. One isolate was vaccine-derived PV type 3 with 10 variation points in the VP1 region. After May 2016, PV type 2 was no longer detected, and PV type 3 became a superior serotype. Of 1,713 NPEVs, 24 serotypes were identified, including echovirus11 (E11), E6, coxsackievirus B3 (CVB3), CVB5, E7, and E3 were the predominant serotypes (37.65%, 20.96%, 11.50%, 8.87%, 8.23%, and 7.06%, respectively). The temporal dynamic of the six common serotypes was inconsistent. E3 was frequently isolated, but the number of isolates was low, with no obvious peaks. E6, E7, and CVB3 exhibited periodic changes with a high peak every three to four years, and E11 only had one high peak lasting four years. Summer-fall peaks of the echoviruses and spring-winter peaks of CVB were observed in the monthly distribution of virus isolation. The infectious isolates of various serotypes of different species identified from the sewage samples showed that ES is an essential part of pathogen surveillance.

Increased detection of Echovirus 6-associated meningitis in patients hospitalized during the COVID-19 pandemic, Israel 2021-2022.

Outbreaks of enteroviral meningitis occur periodically and may lead to hospitalization and severe disease. To analyze and describe the meningitis outbreak in patients hospitalized in Israel in 2021-2022, during the COVID-19 pandemic. In December 2021, before the emergence of the SARS-CoV-2 omicron variant, an off-season increase in enterovirus (EV) infections was observed among patients hospitalized with meningitis. In January 2022, enterovirus cases decreased by 66% in parallel with the peak of the Omicron wave, and then increased rapidly by 78% in March (compared with February) after a decline in Omicron cases. Sequencing of the enterovirus-positive samples showed a dominance of echovirus 6 (E-6) (29%) before and after the Omicron wave. Phylogenetic analysis found that all 29 samples were very similar and all clustered in the E-6 C1 subtype. The main E-6 symptoms observed were fever and headache, along with vomiting and neck stiffness. The median patient age was 25 years, with a broad range (0-60 years). An upsurge in enterovirus cases was observed after the decline of the SARS-CoV-2 omicron wave. The dominant subtype was E-6, which was present prior to the emergence of the omicron variant, but increased rapidly only after the omicron wave decline. We hypothesize that the omicron wave delayed the rise in E-6-associated meningitis.

A comparative study on environmental surveillance of enterovirus: Using a two-phase separation method and a filtration method with a mixed cellulose ester (MCE) membrane

This study aimed to compare the sensitivity of two-phase separation and the filtration method using a mixed cellulose ester (MCE) membrane to detect enteroviruses in sewage samples. From December 2015 to July 2016, four domestic sewage samples (1 L/sample) were collected monthly from the Guangzhou Liede Sewage Treatment Plant, and each sewage sample was divided into two aliquots (500 mL). The sewage sample was concentrated using the two-phase separation and the filtration method using an MCE membrane, and the treated solutions were inoculated into cells for enterovirus isolation. Polymerase chain reaction amplification, VP1 sequencing, and enterovirus molecular typing were performed on the positive isolates. The detection rates of poliovirus (PV) and non-polio enterovirus (NPEV) obtained using the filtration method using an MCE membrane were higher than those using the two-phase separation method. McNemar's test showed that the detection rates of PV, NPEV, type 1 Sabin-like (SL1), type 2 Sabin-like (SL2), and type 3 Sabin-like (SL3) strain were not statistically significant (P > 0.05). In Guangdong Province, China, the detection rates for PV and NPEV were 53.13% and 62.50% (20/32), respectively. Twenty-seven PVs were isolated, three highly variable strains of the type 1 vaccine, with seven nucleotide substitutions in the VP1 region, compared with the type 1 Sabin strains. Eighty-seven strains of NPEV were isolated and nine serotypes were identified, among which coxsackievirus B3 (CVB3), echovirus 6(E6), and echovirus 11(E11) were the dominant strains. The filtration method using an MCE membrane is more sensitive than two-phase separation and can be used as a robust, sensitive, and cost-effective method to isolate enteroviruses from sewage.

Tomato flu-a tell-tale sign of another pandemic or just a benign manifestation of hand foot mouth disease

The world is still dealing with the COVID-19 pandemic, but of a gentle variant. In the midst of the global crisis, in God's own land (Kerala), media highlighted cases of febrile illness similar to flu with red round skin lesions that bear a resemblance to tomatoes that have emerged in children younger than 5 years, known as tomato flu or tomato fever. It could be an aftereffect of chikungunya or dengue, rather than a new viral illness. Numerous reports claim that the tomato flu is truly a viral HFMD variant. Hand, foot, and mouth disease (HFMD) is a common febrile illness characterised by a rash caused by enteroviruses, including Coxsackie A16 (CA16), EV A71, Coxsackie A6, Coxsackie B, and Echo viruses.

Bacterial matrix metalloproteases and serine proteases contribute to the extra-host inactivation of enteroviruses in lake water

Enteroviruses are ubiquitous contaminants of surface waters, yet their fate in presence of microbial congeners is poorly understood. In this work, we investigated the inactivation of Echovirus-11 (E11) and Coxsackievirus-A9 (CVA9) by bacteria isolated from Lake Geneva. Incubation of E11 or CVA9 in biologically active lake water caused inactivation of 2- and 4-log10, respectively, within 48 h. To evaluate the antiviral action of individual bacterial species, we isolated 136 bacterial strains belonging to 31 genera from Lake Geneva. The majority of isolates (92) induced decay of at least 1.5-log10 of CVA9, whereas only 13 isolates induced a comparable inactivation on E11. The most extensive viral decay was induced by bacterial isolates producing matrix metalloproteases (MMPs). Correspondingly, the addition of a specific MMP inhibitor to lake water reduced the extent of inactivation for both viruses. A lesser, though significant protective effect was also observed with inhibitors of chymotrypsin-like or trypsin-like proteases, suggesting involvement of serine proteases in enterovirus inactivation in natural systems. Overall, we demonstrate the direct effect of bacterial proteases on the inactivation of enteroviruses and identify MMPs as effective controls on enteroviruses’ environmental persistence.

Molecular characterization of echovirus 9 strains isolated from hand-foot-and-mouth disease in Kunming, Yunnan Province, China

Echovirus 9 (E9) belongs to the species Enterovirus B. So far, 12 whole genome sequences of E9 are available in GenBank. In this study, we determined the whole genomic sequences of five E9 strains isolated from the stools of patients with hand-foot-and-mouth disease in Kunming, Yunnan Province, China, in 2019. Their nucleotide and amino acid sequences shared 80.8–80.9% and 96.4–96.8% identity with the prototype Hill strain, respectively, and shared 99.3–99.9% and 99.1–99.8% mutual identity, respectively. Recombination analyses revealed that intertype recombination had occurred in the 2C and 3D regions of the five Yunnan E9 strains with coxsackieviruses B5 and B4, respectively. This study augmented the whole genome sequences of E9 in the GenBank database and extended the molecular characterization of this virus in China.

Clinico-Pathological Correlations of Poststreptococcal Glomerulonephritis

Abstract Postinfectious glomerulonephritis is associated with bacterial, viral, fungal, and parasitic infectious agents and histologically appears most often as acute diffuse endocapillary or proliferative glomerulonephritis secondary infection with: group A streptococcus, streptococcus viridans, staphilococus aureus, diploccocus pneumoniae, Brucella melitensis, Salmonella typhi, Yershinia enterocolitica, Plasmodium falciparum, meningococcus, Mycoplasma, Klebsiella, varicella, variola, mumps. Less commonly, it appears as diffuse crescentic glomerulonephritis and a lot of infectious causes are incriminated like: streptococcus, legionella, varicella, Treponema pallidum or as focal crescentic glomerulonephritis: streptococcus A. It rarely appears as mesangiocapillary glomerulonephritis secondary infection with: streptococcus viridans, hepatitis C virus; diffuse or focal mesangial proliferative glomerulonephritis: hepatitis B virus, salmonella, adenovirus, influenza virus, salmonella; focal segmental, necrotizing and sclerosing glomerulonephritis: bacterial endocarditis; membranous glomerulonephritis: hepatitis B virus, syphilis, filarial, Mycobacterium, plasmodium falciparum; focal proliferative: Mycoplasma; mesangiolytic glomerulonephritis :Echo virus. Poststreptococcal glomerulonephritis (PSGN) is caused by prior infection with specific nephritogenic strains of group A beta-hemolytic streptococcus. The clinical presentation of PSGN varies from asymptomatic, microscopic hematuria to the full-blown acute nephritic syndrome, characterized by red to brown urine, proteinuria (which can reach the nephrotic range), edema, hypertension, and acute kidney injury. The prognosis is generally favorable, especially in children, but in some cases, the long-term prognosis is not benign. Managing a case of PSGN requires cooperation between internists, nephrologists, infectious disease consultants, pharmacists, and nursing staff, functioning as an interprofessional team, to provide excellent care for their patients.

Meningitis

The term aseptic meningitis encompasses all types of inflammations of the brain meninges other than that caused by pus producing organisms. It is usually a mild illness. Etiology of aseptic meningitis is very wide and includes many infections - both viral and non viral, drugs, malignancy and systemic illness. The most common cause is viral infection and enteroviruses - Coxsackie and ECHO viruses account for more than half of all cases. Clinical manifestations include headache, fever, malaise, photophobia and meningeal signs. Convulsions, neurological deficits and severe obtundation are rare except with certain non viral infectious meningitis. Diagnostic work up includes blood and cerebrospinal fluid (CSF) examination and serology for infectious meningitis. The polymerase chain reaction is a rapid and accurate method for detection of microbial DNA in CSF. Treatment is mainly supportive, except for the nonviral infectious etiology.

Acute Flaccid Paralysis surveillance in Gaza Strip, Palestine

Acute Flaccid Paralysis (AFP) is a clinical syndrome characterized by rapid onset of weakness, in the respiratory muscles, swallowing and limbs; and death because of respiratory muscles failure. AFP surveillance is crucial, the clinicians are required to notify AFP cases among children below 15 years old to the epidemiology department with completion of AFP surveillance form. In this study, the researcher analyzed AFP surveillance investigation forms to determine the clinical picture, viral etiology and the incidence of AFP among children under 15 years. Material and method (Study design): The research study conducted through cross sectional study, based on AFP investigation forms in the epidemiology department. Results: The final diagnosis of the cases were as follow, 71.1% “27 cases” were GBS, 10.5 % were Encephalitis, 7.9% were Neuropathy, 2.6% “ one case only” for each of the following Aseptic meningitis, Transverse myelitis, Tuphus fever and Subspinal muscular atrophy. The incidence rate among children less than 15 years old which range from 0.58 / 100.000 in 2006 to 1.2 / 100.000 in 2012; Most of the cases 89.5 % had fully vaccinated and 10.5% had not fully vaccinated due to their ages less than one year, 65.8 “25 cases” had residual weakness, all of the cases classified as discarded according to the expert committee decision; no one case dead, all of the samples sent to the Tal-Hashomir laboratory in the occupied land in 1948 for viral isolation; most of the results 81.6% revealed that no virus isolated, 11.8% isolated ECHO virus, 2.6% isolated Coxsachie virus and 4 % isolated NPEV. Conclusion: No wild polio virus in Palestine which is polio free since 1984 and certified in 2006, our success being attributed to: start of 2 doses of IPV and 5 doses of OPV since 1978, high immunization coverage more than 95% at district level and effective AFP surveillance system in Palestine. The study recommended to encourage polio immunization and all clinicians are required to report all AFP cases and submitting at least two stool samples to the epidemiology department (at least 24 hours apart within 14 days of onset of paralysis).

Molecular epidemiological characteristics of echovirus 6 in mainland China: extensive circulation of genotype F from 2007 to 2018

Echovirus 6 (E6) is associated with various clinical diseases and is frequently detected in environmental sewage. Despite its high prevalence in humans and the environment, little is known about its molecular phylogeography in mainland China. In this study, 114 of 21,539 (0.53%) clinical specimens from hand, foot, and mouth disease (HFMD) cases collected between 2007 and 2018 were positive for E6. The complete VP1 sequences of 87 representative E6 strains, including 24 strains from this study, were used to investigate the evolutionary genetic characteristics and geographical spread of E6 strains. Phylogenetic analysis based on VP1 nucleotide sequence divergence showed that, globally, E6 strains can be grouped into six genotypes, designated A to F. Chinese E6 strains collected between 1988 and 2018 were found to belong to genotypes C, E, and F, with genotype F being predominant from 2007 to 2018. There was no significant difference in the geographical distribution of each genotype. The evolutionary rate of E6 was estimated to be 3.631 × 10-3 substitutions site-1 year-1 (95% highest posterior density [HPD]: 3.2406 × 10-3-4.031 × 10-3 substitutions site-1 year-1) by Bayesian MCMC analysis. The most recent common ancestor of the E6 genotypes was traced back to 1863, whereas their common ancestor in China was traced back to around 1962. A small genetic shift was detected in the Chinese E6 population size in 2009 according to Bayesian skyline analysis, which indicated that there might have been an epidemic around that year.

Up-to-date approaches to forecast of epidemiological situation with incidence of enteroviral meningitis

This work evaluates the results of long-term dynamic monitoring of non-polio enteroviruses (NPEV) circulation in the indicator subpopulation in Yekaterinburg (children aged 3 to 6 years old), isolation of this group of viruses from specimens of sewage and the spectrum of causative agents detected in the liquor of patients with enteroviral meningitis (EVM). We established a high comparability rate of Coxsackie B and ECHO virus serotypes etiologically significant in EVM and isolated from healthy carriers. NPEV detected in sewage waters over the observation period were presented mainly by Coxsackie B viruses (84.0 %), while ECHO viruses (75.6 %) dominated among the causative agents of EVM. A conclusion was made about low informative value of the sewage study results for evaluation and forecast of the EVM epidemic situation as well as about the rationale for including the screening studies of NPEV circulation in the indicator subpopulation into the system of virological and molecular genetic monitoring.

Molecular epidemiology of enteroviruses associated with severe hand, foot and mouth disease in Shenzhen, China, 2014-2018

In this study, we investigated the epidemiology and molecular characteristics of enteroviruses associated with severe hand, foot and mouth disease (HFMD) in Shenzhen, China, during 2014-2018. A total of 137 fecal specimens from patients with severe HFMD were collected. Enterovirus (EV) types were determined using real-time reverse transcription polymerase chain reaction (RT-PCR), RT nested PCR, and sequencing. Sequences were analyzed using bioinformatics programs. Of 137 specimens tested, 97 (70.8%), 12 (8.8%), and 10 (7.3%) were positive for EV-A71, coxsackievirus A6 (CVA6), and CVA16, respectively. Other pathogens detected included CVA2 (2.9%, 4/137), CVA10 (2.9%, 4/137), CVA5 (0.7%, 1/137), echovirus 6 (E6) (0.7%, 1/137) and E18 (0.7%, 1/137). The most frequent complication in patients with proven EV infections was myoclonic jerk, followed by aseptic encephalitis, tachypnea, and vomiting. The frequencies of vomiting and abnormal eye movements were higher in EV-A71-infected patients than that in CVA6-infected or CVA16-infected patients. Molecular phylogeny based on the complete VP1 gene revealed no association between the subgenotype of the virus and disease severity. Nevertheless, 12 significant mutations that were likely to be associated with virulence or the clinical phenotype were observed in the 5’UTR, 2Apro, 2C, 3A, 3Dpol and 3’UTR of CVA6. Eight significant mutations were observed in the 5’UTR, 2B, 3A, 3Dpol and 3’UTR of CVA16, and 10 significant mutations were observed in the 5’UTR, VP1, 3A and 3Cpro of CVA10. In conclusion, EV-A71 is still the main pathogen causing severe HFMD, although other EV types can also cause severe complications. Potential virulence or phenotype-associated sites were identified in the genomes of CVA6, CVA16, and CVA10.Electronic supplementary materialThe online version of this article (10.1007/s00705-020-04734-z) contains supplementary material, which is available to authorized users.

Etiology and molecular-epidemiological characteristics of viral meningo-encephalitis in Zhejiang province, 2002-2018

Objective: To understand the pathogens and molecular-epidemiologic characteristics of viral meningo-encephalitis in Zhejiang province during 2002 to 2018. Methods: All the samples were collected from suspected patients admitted to the hospitals under the monitoring program. Of the total samples, 2 173 were cerebrospinal fluids while the other 455 were stool specimens. Cerebrospinal fluid (CSF) samples were subject to real-time qPCR for the detection of Human enterovirus (HEV), Mumps virus (MuV), Herpes simplex virus (HSV), Cytomegalovirus (CMV) and Japanese encephalitis virus (JEV). Stool sample were subject to real-time qPCR for HEV. ELISA was used to detect the IgM antibodies in CSF, in the 5 kinds of virus mentioned above. VP1 genes from all RNA-positive specimen were amplified, sequenced, for typing and for evolution analysis. Results: 871 (40.1%) of the 2 173 samples were detected as HEV nucleic acid positive during 2002 to 2018. 654 (38.1%) of the 1 718 CSF sample were HEV nucleic acid positive while 217 (47.7%) of the 455 stool sample were HEV nucleic acid positive. Among the total positive nucleic acid sample, 670 of them were VP1 sequence positive, including 5 HEV-A and 665 HEV-B. There were 23 HEV serotypes, including Coxsackievirus (CV) CVA4, CVA6, CVA9, CVA10, CVB1-5, Echovirus (EchoV; E) E3, E4, E6,E7, E9, E11, E14, E16, E18, E21, E25, E30, E33 and EV-71. The top three serotypes went to E30, E6 and CVB5. These three serotypes presented enhanced viral activity in every several years. 795 CSF samples were detected as virus nucleic acid positive, including 374 HEV, 6 MuV, 5 HSV and 5 CMV, from 2012 to 2015 and in 2018. 5 kinds of IgM antibodies were detected simultaneously in 368 CSF samples, including 2 HEV positive, 6 JEV positive and 1 MuV positive for 5 viruses, respectively. Except for EV-71, there were 517 EchoV and 152 CV viruses presented, with the ratio of 3.4∶1. These two kinds of viruses alternately changed for each predominant epidemic strains in every 3-5 years. Based on VP1, results from the phylogenetic tree showed that HEV from Zhejiang province clustered into HEV-A and HEV-B clades respectively. E30 developed both h and i sub-genotypes. Conclusions: HEV-B seemed the main pathogen for viral meningo-encephalitis in Zhejiang province. Ratio of positive detection on EchoV was significantly higher than that on CV. These two kinds of virus alternately presented changing tendency in every several years. Predominant epidemic strains E30, CVB5 and E6 were presenting enhanced viral activity, also in every several years. High correlation was found in both HEV viral activity from the surveillance sites and in time line of the viral meningo-encephalitis outbreaks.

Энтеровирусная инфекция у детей в Краснодарском крае, клинико- эпидемиологическая характеристика

Objective. To analyze age-specific etiology and clinical manifestations of enteroviral infections (EVIs) in children residing in Krasnodar region. Patients and methods. This retrospective study included 3,213 patients with confirmed EVIs registered between 2006 and 2018 and admitted to the Specialized Clinical Children's Hospital for Infectious Diseases. Polymerase chain reaction was used to confirm the diagnosis. Results. We found that the most common enteroviruses circulating in Krasnodar region for many years were Coxsackie B virus and ECHO viruses 6, 11, and 30. The majority of patients (75.1%) had isolated forms of infection. Almost one-third of children (27.8%) demonstrated central nervous system lesions (primarily meningitis) that were more common in children aged 3 to 12 years. Other clinical forms were observed in young children. Almost all children hospitalized (91%) attended kindergartens. Conclusion. Coxsackie B virus and ECHO viruses 6, 11, and 30 have been circulating in Krasnodar region for the last 12 years; enterovirus type 71 has recently been found in this area. The highest proportion of EVI cases requiring inpatient treatment was observed among young children, in whom EVIs primarily manifested with meningitis. Primary school children usually had herpangina and exanthema, while older children developed epidemic myalgia. Our findings indicate the need for a set of measures aimed at reducing the incidence of EVIs among children of Krasnodar region. Key words: children, enteroviral infection, Krasnodar region, epidemiology, clinical manifestations, diagnosis

RESEARCH OF BIOLOGICAL PROPERTIES OF ENTEROVIRUS STRAINS ASSOCIATED WITH ISCHEMIC STROKE

Introduction: The research of biological properties of enteroviruses associated with ischemic stroke (IS) allows us to identify their intratypic differences. The aim: to identify genetic markers of strains of enteroviruses associated with IS. Materials and methods: 11 strains of enteroviruses isolated from the serum of patients with IS were identified in the virus neutralization test. Genetic markers of isolated strains (Abent, marker S, marker rct40) were determined. Results: Eleven strains of enteroviruses were isolated from the serum of patients with IS. Eight viruses: Coxsackie B viruses (serotypes 2, 3, 4) and ECHO viruses (serotypes 6, 9, 27 (two strains), 29) were identified in these strains. Other three strains of enteroviruses were unidentified. Different combinations of genetic markers were found. Seven strains of enteroviruses (Coxsackie B2, B3, ECHO 6, ECHO 9, ECHO 27 (two strains) and one unidentified virus) had virulence markers: Abent-, rct40+ and S-. Three strains (Coxsackie B4, ECHO 29, one unidentified virus) had markers: Abent-, rct40+, S+. Another one unidentified virus had markers: Abent+, rct40+, S -. Conclusions: All 11 isolates of enteroviruses associated with IS had rct40+ marker, 10 of the 11 isolates had marker Abent- and 8 of 11 isolates had marker S-. The research of genetic markers allows to perform typic and intratypic differentiation of strains of enteroviruses associated with the IS.

Molecular epydemiology of enterovirus infection in the Republic of Belarus in 2016–2017

Enteroviruses are widespread human pathogens characterized by a high level of a genetic diversity. They cause different clinical forms of infection. The aim of the present study was to analyze the molecular epidemiology of enterovirus infection in the application to the structure of its clinical forms in 2016–2017.ECHO viruses predominated among patients with aseptic meningitis and were prevailing group of enteroviruses in 2016 (all ECHO viruses – 58%, including ECHO 9 – 26%, ECHO 6 –14%, ECHO 16 – 10%). In 2017, Coxsackieviruses prevailed (68%), that were including Coxsackievirus B5 (31%), Coxsackievirus B1, Coxsackievirus B4 and Coxsackievirus A6 (9% of each serotype). Coxsackieviruses were detected more frequently in patients with vesicular pharyngitis and unspecified enterovirus infection. The results of the molecular epidemiological analysis indicated that the prevalence of ECHO viruses in 2016 and Coxsackieviruses B in 2017 was due to the emergence of numerous new genovariants of these viruses.

Molecular Characterization and Phylogenetic Analysis of Enteroviruses and Hepatitis A Viruses in Sewage Samples, Northern Italy, 2016.

Enteroviruses (EVs) and Hepatitis A Viruses (HAVs) are human pathogens with a wide spectrum of clinical manifestations. The monitoring of sewage samples enables to monitor the EVs and HAVs in circulation among the general population and recognize possible outbreaks. This study focused on the molecular characterization and phylogenetic analysis of the EVs and HAVs identified in 33 sewage samples collected every 15days at the influent of a wastewater treatment plant located in Northern Italy from March to October 2016. According to the results of the molecular characterization, the most frequently identified viruses were Echovirus 6 (E-6), E-11 and HAV-IA. The phylogenetic analyses indicated the rapid genetic evolution of E-6 and E-1; noteworthy, most E-11 strains clustered with a strain isolated from a clinical sample collected in the same geographical area over the same period by our laboratory. Most of the HAV strains detected clustered with epidemic HAV-IA strains identified during the European hepatitis A outbreak that occurred in 2016-2017 affecting men who have sex with men (MSM). The detection of environmental HAV strains before and at the beginning of its spread amongst humans demonstrated that this outbreak could have been predicted by monitoring sewage samples. Moreover, conducting a genetic comparison between the HAV and EV strains identified in sewage and clinical samples may improve knowledge of viral epidemiology. EV and HAV molecular environmental surveillance may prove useful for identifying viral circulation and for issuing early warning alerts on possible outbreaks among the human population.

Identification and Phylogenetic Characterization of Human Enteroviruses Isolated from Cases of Aseptic Meningitis in Brazil, 2013-2017.

Aseptic meningitis is a common viral infection associated with human enteroviruses. The aim of the present study was to identify and characterize the enteroviruses associated with outbreaks and sporadic cases of aseptic meningitis that occurred in different regions of Brazil between 2013 and 2017. Cerebrospinal fluids obtained from patients admitted to public health facilities were analyzed. A total of 303 patients were positive for Human Enteroviruses (EV) by cell culture isolation with a median isolation rate throughout the year of 12%. We were able to identify enterovirus serotypes in 295 clinical specimens. Nineteen different serotypes were identified; the large majority corresponded to HEV-B species. Echovirus 30 (E-30) and Echovirus 6 (E-6) were the most prevalent genotypes (66.8%). Sequence analysis suggested that circulating E-30 was closely related to E-30 from other American countries; while E-6 was derived from Europe. Most of the patients consisted of children ≤ 15 years old. The temporal distribution of all aseptic meningitis and EV-positive cases showed an obvious seasonal pattern during autumn. Our results have provided valuable information about the enteroviral etiology of the aseptic meningitis cases in Brazil pointing to the importance of enterovirus surveillance in neurological diseases.

Comparison of the detection performance of two different one-step-combined test strips with fluorescent microspheres or colored microspheres as tracers for influenza A and B viruses

BackgroundInfluenza A and B viruses mainly cause respiratory infectious disease. Till now, few tests are able to simultaneously detect both, especially in primary medical establishments.MethodsThis study was designed to compare the performance of two different one-step-combined test strips for the detection of influenza A and B: one strip with fluorescent microspheres for tracers (FMT); and the other strip with colored microspheres for tracers (CMT). To test the strips, cultures of influenza A, B, and other pathogenic viruses were used, in addition to 1085 clinical specimens from symptomatic patients with respiratory infections. Real-time RT-PCR was also considered as a reference method used to detect the different results of FMT and CTM.ResultsDetection thresholds for influenza A and B cultures using serial dilutions revealed that the sensitivity of FMT was higher than that of CMT (both P < 0.05). With the culture mixtures of Coxsackie virus (A16), enteric cytopathic human orphan virus (ECHO type30), enterovirus (EV71), rotavirus (LLR strain), and enteric adenovirus (AdV 41), specificity assessment demonstrated that there was no cross reaction during the usage of the two test strips as shown by the results which were negative. In the detection of influenza A in 1085 clinical specimens, the total coincidence rate was 96.7%, the positive coincidence rate was 97.1%, and the negative coincidence rate was 96.7%. In the case of influenza B detection, the total coincidence rate was 99.1%, the positive coincidence rate was 92.6%, and the negative coincidence rate was 98.5%. In addition, with influenza A or B real-time RT-PCR detection method, the results showed that, for influenza A, 26 of the 33 specimens that negative with CMT but positive with FMT, showed positive results, and none of the 3 specimens that positive with CMT but negative with FMT showed a positive result; For influenza B, 12 of the 15 specimens that negative with CMT but positive with FMT, showed positive results, and none of the 5 specimens that positive with CMT but negative with FMT showed a positive result.ConclusionsFMT performed better than CMT in the combined detection of influenza A and B viruses.