Echovirus 30 detection in an outbreak of acute myalgia and rhabdomyolysis, Brazil 2016-2017.

OBJECTIVES: To describe an outbreak of acute myalgia accompanied by elevated levels of muscle enzymes that occurred in the northeast region of Brazil from December 2016 through to May 2017. METHODS: Clinical data were analysed and laboratory tests were performed in 86 specimens obtained from 52 individuals with suspected acute myalgia. A broader reactive enterovirus real-time RT-PCR followed by a semi-nested PCR amplification of partial VP1 gene were performed to identify the causative agent. RESULTS: Eighty-six clinical samples were received in our laboratory during the myalgia outbreak. Median age of individuals was 39 years. Sudden acute myalgia and dark urine were the most common symptoms. Creatine phosphokinase levels were elevated with mean value ∼16 893 U/L. Human enterovirus was detected in 67% (58/86) of the patient's specimens (urine, serum, faeces and rectal swab). The enterovirus positivity per patient was 82.7% (43/52). Echovirus 30 (E-30) (82% of the typed specimens, 18/22; 76.4% (13/17) of the typed specimens per patient) was the main enterovirus identified. In addition to E-30, CV-A16 (1/22) and E-6 (3/22) were detected in 4% and 14% of the typed specimens, respectively. No deaths occurred. CONCLUSION: The 2016-2017 outbreak of acute myalgia that occurred in the northeast region of Brazil can be associated with E-30. Despite the clinical manifestations, a favourable outcome was observed for all patients.

Rubella virus genotype 1G and echovirus 9 as etiologic agents of exanthematous diseases in Brazil: insights from phylogenetic analysis.

The aim of the present study was to identify the rubella virus (RV) and enterovirus (EV) genotypes detected during the Epidemiological Surveillance on Exanthematic Febrile Diseases (VIGIFEX) study and to perform phylogenetic analysis. Ten RV- and four EV-positive oropharyngeal samples isolated from cell culture were subjected to RT-PCR and sequencing. Genotype 1G and echovirus 9 (E-9) was identified in RV- and EV-positive samples, respectively. The RV 1G genotype has been persisting in Brazil since 2000-2001. No evidence of E-9 being involved in exanthematic illness in Brazil has been reported previously. Differential laboratory diagnosis is essential for management of rash and fever disease.

Molecular diagnosis of echovirus 30 as the etiological agent in an outbreak of aseptic meningitis in Panama: May-June 2008.

INTRODUCTION: Aseptic meningitis outbreaks are commonly caused by viral pathogens with enterovirus a common etiological agent. Between May and June of 2008, an outbreak of 173 cases of aseptic meningitis occurred in the Chiriqui Province of Panama. Molecular techniques were used to identify the etiological agent. METHODOLOGY: Cerebrospinal fluid (CSF) samples from 75 patients were received at the Gorgas Memorial Institute for Health Studies.  RNA extraction and one-step RT-PCR were performed on each sample to determine the presence of enterovirus.  Thirty-four samples which were positive for enterovirus were subject to group-specific PCR, sequencing, and phylogenetic analysis to identify the etiological agent of the outbreak. RESULTS: The CSF of 58 subjects was found positive for the enterovirus family using RT-PCR. Thirty-four samples were found to belong to the enterovirus B group. Phylogenetic analysis of four successfully sequenced samples revealed echovirus 30 as the etiological agent. CONCLUSION: Echovirus 30 is reported as the likely cause of an outbreak of aseptic meningitis in Panama, the first since the 1980s.

Echovirus 30 associated with cases of aseptic meningitis in state of Pará, Northern Brazil.

Investigation of the aetiology of viral meningitis in Brazil is most often restricted to cases that occur in the Southern and Southeastern Regions; therefore, the purpose of this study is to describe the viral meningitis cases that occurred in state of Pará, Northern Brazil, from January 2005-December 2006. The detection of enterovirus (EV) in cerebrospinal fluid was performed using cell culture techniques, RT-PCR, nested PCR and nucleotide sequencing. The ages of the 91 patients ranged from < one year old to > 60 years old (median age 15.90 years). Fever (87.1%), headache (77.0%), vomiting (61.5%) and stiffness (61.5%) were the most frequent symptoms. Of 91 samples analyzed, 18 (19.8%) were positive for EV. Twelve were detected only by RT- PCR followed by nested PCR, whereas six were found by both cell culture and RT-PCR. From the last group, five were sequenced and classified as echovirus 30 (Echo 30). Phylogenetic analyses revealed that Echo 30 detected in Northern Brazil clustered within a unique group with a bootstrap value of 100% and could constitute a new subgroup (4c) according to the phylogenetic tree described by Oberste et al. (1999). This study described the first molecular characterization of Echo 30 in Brazil and this will certainly contribute to future molecular analyses involving strains detected in other regions of Brazil.

Echovirus 30 associated with cases of aseptic meningitis in state of Pará, Northern Brazil

Investigation of the aetiology of viral meningitis in Brazil is most often restricted to cases that occur in the Southern and Southeastern Regions; therefore, the purpose of this study is to describe the viral meningitis cases that occurred in state of Pará, Northern Brazil, from January 2005-December 2006. The detection of enterovirus (EV) in cerebrospinal fluid was performed using cell culture techniques, RT-PCR, nested PCR and nucleotide sequencing. The ages of the 91 patients ranged from < one year old to > 60 years old (median age 15.90 years). Fever (87.1 percent), headache (77.0 percent), vomiting (61.5 percent) and stiffness (61.5 percent) were the most frequent symptoms. Of 91 samples analyzed, 18 (19.8 percent) were positive for EV. Twelve were detected only by RT- PCR followed by nested PCR, whereas six were found by both cell culture and RT-PCR. From the last group, five were sequenced and classified as echovirus 30 (Echo 30). Phylogenetic analyses revealed that Echo 30 detected in Northern Brazil clustered within a unique group with a bootstrap value of 100 percent and could constitute a new subgroup (4c) according to the phylogenetic tree described by Oberste et al. (1999). This study described the first molecular characterization of Echo 30 in Brazil and this will certainly contribute to future molecular analyses involving strains detected in other regions of Brazil.

[Outbreak of viral meningitis caused by echovirus type 4 in Misiones province]. / Brote epidémico de meningitis viral causado por echovirus tipo 4 en la provincia de Misiones.

A descriptive retrospective study was carried out to describe an epidemic outbreak of enteroviral meningitis in Misiones. We reviewed records of 143 children from 1 month to 14 years of age who were hospitalized with aseptic meningitis in the Pediatric Hospital of Posadas from August to December 2005. Increased number of cases was observed between weeks 33 to 50 which reached a maximum peak in weeks 47 and 48, confirming an outbreak. The median of age was 8 years old, 55.2% were males. Eighty percent of cases were in 5 to 14 years old children. The average length of time spent in the hospital was 4.5+/-1.7 days, no deaths were reported. We performed cell counts, chemical and bacterial studies of CSF, and culture or RT-Nested/PCR for enteroviruses. Isolates were serotyped by RT-PCR amplification and genetic sequencing. Cell counts were from 6 to 5040 cells/mm3. Ninety two percent had less than 500 cells/mm3 and 43.5% had lymphocyte predominance. Glucose levels were normal with slightly elevated protein counts in 56% of cases. Of the cultured samples, 28% (17/60) showed cytopathic effect compatible with enterovirus. RT-n-PCR detected enterovirus in 73% (43/59) of the analyzed CSF. Echovirus type 4 was identified in 6 of them. The positive indicator obtained by combining both techniques was 83% (58/70).

An outbreak of concurrent echovirus 30 and coxsackievirus A1 infections associated with sea swimming among a group of travelers to Mexico.

BACKGROUND: Enteroviruses are shed in human stool and can cause a wide spectrum of illness. They are the leading cause of aseptic meningitis. METHODS: In 2004, the Connecticut Department of Public Health investigated a meningitis cluster among persons returning from a school-organized trip to Mexico. RESULTS: Among 29 travelers (25 teenagers and 4 adult chaperones), 21 became acutely ill. Viral culture and nucleic acid amplification testing of stool (n=27) and cerebrospinal fluid (n=4) specimens identified enteroviral infection in 20 of 28 travelers from whom any specimen was obtained; 4 had echovirus 30 only, 11 had coxsackievirus (CV) A1 only, 4 had both echovirus 30 and CVA1, and 1 had CVA5 only. Illness onset dates were tightly clustered 4 days after a prolonged swim in the Gulf of Mexico. Time spent swimming was significantly associated with the odds of enteroviral infection (univariate odds ratio for each additional hour swimming, 14.3; 95% confidence interval, 1.3-154.3). Headache, fever, vomiting, and nausea occurred more frequently among the echovirus 30-infected travelers than among the uninfected control subjects (P< .05). The most frequent symptoms among travelers infected with only CVA1 identified were nausea and diarrhea (36% each), but neither was significantly associated with CVA1 infection; 5 patients with CVA1 infection were asymptomatic. CONCLUSIONS: We identified multiple enteroviruses among the travelers. Clustered illness onsets suggest point-source exposure, which likely was a sea swim in sewage-contaminated seawater. Novel molecular amplification and sequencing methodologies were required to recognize the rarely identified CVA1, but it is ambiguous whether CVA1 infection caused illness. Travelers should be aware of risks associated with swimming in natural waters when visiting areas where there is limited sewage treatment.

Brote epidémico de meningitis viral causado por echovirus tipo 4 en la provincia de Misiones / Outbreak of viral meningitis caused by echovirus type 4 in Misiones province

Se realizó un estudio retrospectivo a fin de describir un brote epidémico de meningitis causado por enterovirus, que comprometió a 143 niños de 1 mes a 14 años internados en el Hospital Pediátrico de Posadas (Misiones) con diagnóstico de meningitis aséptica, entre agosto y diciembre de 2005. Se observó un aumento de casos entre las semanas 33 a 50, con un pico máximo entre las semanas 47 y 48, lo que confirmó el brote. La mediana de edad de los niños afectados fue de 8 años y el 55,2% fueron varones. El 80% de los casos se observó entre escolares (5 a 14 años). El promedio del tiempo de internación fue de 4,5 ± 1,7 días, y no se registraron fallecidos. Los LCR se estudiaron mediante examen citoquímico y estudios bacteriológicos y virológicos (aislamiento viral, RT- PCR anidada e identificación molecular mediante secuenciación génica). Los recuentos de células en LCR variaron entre 6 y 5040 células /mm3, el 92% fueron inferiores a 500 células/mm3 y el 43,5% mostró predominio linfocitario. El 56% presentó concentraciones de glucosa normal, con proteínas ligeramente elevadas. El 28% de las muestras estudiadas por cultivo (17/60) mostró efecto citopático, compatible con enterovirus. La RT-PCR anidada permitió detectar enterovirus en un 73% de las muestras (43/59), con 6 casos que se tipificaron como echovirus tipo 4. El índice de positividad al combinar ambas técnicas alcanzó el 83%.

A descriptive retrospective study was carried out to describe an epidemic outbreak of enteroviral meningitis in Misiones. We reviewed records of 143 children from 1 month to 14 years of age who were hospitalized with aseptic meningitis in the Pediatric Hospital of Posadas from August to December 2005. Increased number of cases was observed between weeks 33 to 50 which reached a maximum peak in weeks 47 and 48, confirming an outbreak. The median of age was 8 years old, 55.2% were males. Eighty percent of cases were in 5 to 14 years old children. The average length of time spent in the hospital was 4.5±1.7 days, no deaths were reported. We performed cell counts, chemical and bacterial studies of CSF, and culture or RT-Nested/PCR for enteroviruses. Isolates were serotyped by RT-PCR amplification and genetic sequencing. Cell counts were from 6 to 5040 cells/mm3. Ninety two percent had less than 500 cells/mm3 and 43.5% had lymphocyte predominance. Glucose levels were normal with slightly elevated protein counts in 56% of cases. Of the cultured samples, 28% (17/60) showed cytopathic effect compatible with enterovirus. RT-n-PCR detected enterovirus in 73% (43/59) of the analyzed CSF. Echovirus type 4 was identified in 6 of them. The positive indicator obtained by combining both techniques was 83% (58/70).

RT-PCR for confirmation of echovirus 30 isolated in Belém, Brazil.

Echovirus (Echo) 30 or human enterovirus B is the most frequent enterovirus associated with meningitis cases. Epidemics and outbreaks of this disease caused by Echo 30 have occurred in several countries. In Brazil, Echo 30 has been isolated from sporadic cases and outbreaks that occurred mainly in the south and southeast regions. We used RT-PCR to examine Echo 30 isolates from meningitis cases detected from March 2002 to December 2003 in Belém, state of Pará, in northern Brazil. The patients were attended in a Basic Health Unit (State Health Secretary of Pará), where cerebrospinal fluid (CSF) was collected and stored in liquid nitrogen. Weekly visits were made by technicians from Evandro Chagas Institute to the health unit and samples were stored at -70 degrees C in the laboratory until use. HEp-2 and RD cell lines were used for viral isolation and neutralization with specific antisera for viral identification. RNA extraction was made using Trizol reagent. The RT-PCR was made in one step, and the total mixture (50 microL) was composed of: RNA, reaction buffer, dNTP, primers, Rnase inhibitor, reverse transcriptase, Taq polymerase and water. The products were visualized in agarose gel stained with ethidium bromide, visualized under UV light. Among the 279 CSF samples examined, 30 (10.7%) were EV positive, 29 being Echo 30 and one was Cox B. Nineteen Echo 30 were examined with RT-PCR; 18 tested positive (762 and 494 base pairs). The use of this technique permitted viral identification in less time than usual, which benefits the patient and is of importance for public-health interventions.

RT-PCR for confirmation of echovirus 30 isolated in Belém, Brazil

Echovirus (Echo) 30 or human enterovirus B is the most frequent enterovirus associated with meningitis cases. Epidemics and outbreaks of this disease caused by Echo 30 have occurred in several countries. In Brazil, Echo 30 has been isolated from sporadic cases and outbreaks that occurred mainly in the south and southeast regions. We used RT-PCR to examine Echo 30 isolates from meningitis cases detected from March 2002 to December 2003 in Belém, state of Pará, in northern Brazil. The patients were attended in a Basic Health Unit (State Health Secretary of Pará), where cerebrospinal fluid (CSF) was collected and stored in liquid nitrogen. Weekly visits were made by technicians from Evandro Chagas Institute to the health unit and samples were stored at -70°C in the laboratory until use. HEp-2 and RD cell lines were used for viral isolation and neutralization with specific antisera for viral identification. RNA extraction was made using Trizol reagent. The RT-PCR was made in one step, and the total mixture (50 æL) was composed of: RNA, reaction buffer, dNTP, primers, Rnase inhibitor, reverse transcriptase, Taq polymerase and water. The products were visualized in agarose gel stained with ethidium bromide, visualized under UV light. Among the 279 CSF samples examined, 30 (10.7 percent) were EV positive, 29 being Echo 30 and one was Cox B. Nineteen Echo 30 were examined with RT-PCR; 18 tested positive (762 and 494 base pairs). The use of this technique permitted viral identification in less time than usual, which benefits the patient and is of importance for public-health interventions.

[Intrathecal synthesis pattern of immunoglobulins in meningoencephalitis epidemic due to echovirus 9]. / Patrones de síntesis intratecal de inmunoglobulinas en epidemia de meningoencefalitis por echovirus 9.

INTRODUCTION: Epidemics of meningoencephalitis due to echovirus 9 were commonly occurred when a children population become susceptible for the first time in front the virus. OBJECTIVE: To present the intrathecal synthesis pattern of immunoglobulins of the epidemic that affected Cuba in 1999 and to probe the usefulness of reibergram and antibody index in the diagnostic and characterization of the outbreak. PATIENTS AND METHODS: 23 pediatric patients suffering from viral meningoencephalitis due to echovirus 9 were studied in the income moment. Serum and cerebrospinal fluid IgA, IgM, IgG, albumin and glucose were quantified. Cerebrospinal fluid total protein content and lactate were quantified. Titles of antibodies against echo 9 and Coxsackie A9 and differential cell count were performed. RESULTS: A mean of 555 cells/10 6 L mainly lymphocytes were obtained. Glucose in cerebrospinal fluid was over 50%, serum glucose and lactate levels below 2.1 mmol/L. In the reibergram an absence of intrathecal synthesis was predominant (15/23), IgM synthesis (6/23) and IgM+IgA (2/23). Blood cerebrospinal fluid dysfunction was observed in 15 patients. The mean antibody index was 1,8 for echo 9 and 0,9 for Coxsackie A9. CONCLUSIONS: The intrathecal synthesis pattern of immunoglobulins was different from other enterovirus and from echovirus 9 in non epidemic situations before this epidemic, probably with alteration of viral genome.

[Neuroimmune epidemiological value of reibergram in the first echovirus 16 meningoencephalitis outbreak in Cuba]. / Valor neuroinmunoepidemiologico del reibergrama en la primera epidemia de meningoencefalitis por echovirus 16 en Cuba.

INTRODUCTION: During the third quarter of 2000, an outbreak of echovirus 16 meningoencephalitis was firstly occurred in Cuba and produced vomiting, headache, fever and exanthem that differentiate it from other enterovirus epidemies. OBJECTIVE: To describe the intrathecal immunoglobulin synthesis pattern of the epidemie from the reibergram. PATIENTS AND METHODS: Diagnostic serum and cerebrospinal fluid from 18 children during the acute phase by nephelometrie assay, besides cytochemical and virological study. RESULTS: A predominant absence of intrathecal immunoglobulin synthesis and two patients with IgA and IgM synthesis was produced. 66% of cerebrospinal blood barrier dysfunction, 6.26 10 3 mean albumin ratio, lactate below 2.1 mmol/L and glucose concentration in cerebrospinal fluid was 50% over glucose blood content and 168 cells 10 6 L mainly lymphocytes were observed. CONCLUSIONS: The intratecal immunoglobulin synthesis pattern differs from other enterovirus outbreaks that have affected this population and it seems the one found for the diagnostic period in adults. This finding alerts the possibility of genetic changes in echovirus 16 strain, interesting from the neuroimmunoepidemiological point of view.

First epidemic of echovirus 16 meningitis in Cuba.

From April to September 2000, an epidemic of aseptic meningitis spread throughout Cuba, with 16,943 reported cases. Virologic studies identified echovirus 16 as the cause of this epidemic. This is the first reported isolate of echovirus 16 from patients with viral meningitis in Cuba.

[Viral meningoencephalitis caused by enterovirus in Cuba from 1990-1995]. / Meningoencefalitis virales por enterovirus en Cuba en el período 1990-1995.

The results of a study on enterovirus as causal agents of viral meningoencephalitis from 1990 to 1995 are described. From the 1458 samples of patients studied, 586 were from feces, 108 from cerebrospinal fluid and 1095 were paired sera. The samples for viral isolation were inoculated in two different cellular systems. A total of 225 samples (32.42%) were found to be positive; the highest number of isolations (n = 217) was found in feces and in diploid human lung fibroblast cells (PHuE-1). Antibody determination were carried out using neutralization by micromethod test with 10 enterovirus antigens (Echovirus 4, 6, 9, 11 and Coxsackievirus B1-6). The isolated virus was also used as antigen during epidemic periods. There were 3 epidemic outbreaks: Coxsackievirus A9 (1990-1991), Echovirus 30 (1994), and Coxsackievirus B5 (1995). Coxsackievirus A9 and Echovirus 30 have been related to viral meningoencephalitis epidemics in Cuba since 1970. In paired sera 66.84% were positive, Echovirus 6 and 11 exhibited higher positivity. When positive results by isolation and serology were considered together, more than 80% of the studied cases were associated to enterovirus infection. These results show the importance of enteroviruses as cause of viral meningoencephalitis in our country.

Echoviroses / Echovirus infections

Trata das echoviroses sob aspectos da etiologia, epidemiologia, patogenia e patologia, manifestaçöes clínicas e tratamento. (MC)

Brote epidémico de meningitis aséptica: experiencias en un hospital infantil privado / Epidemic of aseptic meningitis experience of private hospital

Se informa acerca de las experiencias obtenidas en 193 niños con meningitis aséptica, estudiados durante un brote epidémico registrado entre febrero y julio de 1992. Los cultivos de líquido cefalorraquídeo hechos en el Instituto de Referencia Epidemiológica permitieron identificar al virus Echo 30 como un agente a la meningitis en 48 niños

[Epidemic outbreak of viral meningitis caused by type 30 ECHO virus]. / Brote epidémico de meningitis viral causado por virus ECHO tipo 30.

The clinical records of 15 children admitted to Hospital Infantile de México Federico Gómez with diagnosis of viral meningitis were reviewed. They were part of 19 patients admitted with this diagnosis during a 5 week period (March 22 to April 30, 1992) and represent a significant increase of aseptic meningitis compared with the same periods of previous years at Hospital Infantile de Mexico and in Mexico City where there is an ongoing epidemic outbreak of this entity. All the patients studied had spinal fluid findings consistent with viral meningitis and in 4 of them on ECHO virus type 30 was isolated at the Enterovirus Section of the CDC, Atlanta Georgia USA.

Aseptic meningitis due to echovirus 4 in Panama City, Republic of Panama.

Between September 23 and November 30, 1981, 1,032 children from Panamá City, Republic of Panamá were hospitalized with aseptic meningitis. Forty-four per cent of the cases were in the age group 5-9 years old; the disease was mild and self-limiting with an average hospital stay of five days. Echovirus 4 isolates were obtained from 48 of 160 patients. To identify risk factors associated with the epidemic, the authors randomly selected 10 per cent of hospitalized cases and conducted a family-based seroepidemiologic study. The closest neighboring house with at least one child younger than 15 years was similarly studied as a control. In total, 182 households and 1,083 of 1,177 residents were included. No risk factors ascertained by the study were associated with aseptic meningitis; however, several factors were related to recent echovirus 4 infection. Overall, 56 per cent of cases had echovirus 4 antibody, as did 29 per cent of their family members and 19 per cent of control family members. Children who attended kindergarten or primary school were more likely to have antibody than other household members and, within case families, individuals involved in child-care had an excess risk for infection.