Antiviral treatment with fluoxetine for rituximab-associated chronic echovirus 13 meningoencephalitis and myofasciitis.

BACKGROUND AND PURPOSE: Enterovirus infections pose a serious threat for patients with humoral deficiencies and may be lethal, whilst the efficacy of proposed treatment options such as corticosteroids, intravenous immunoglobulins and fluoxetine remains debated. METHODS: Viral clearance was investigated in a patient with rituximab-induced B-cell depletion and chronic echovirus 13 (E13) meningoencephalitis/myofasciitis in response to intravenous immunoglobulins and fluoxetine using sequential semi-quantitative E13 viral load measurements by real-time reverse transcription polymerase chain reaction. Fluoxetine concentrations in plasma and cerebrospinal fluid were determined by liquid chromatography mass spectrometry. RESULTS: Intravenous immunoglobulins appeared ineffective in this case of E13 infection, whereas virus clearance in cerebrospinal fluid was obtained after 167 days of oral fluoxetine. Since treatment with corticosteroids resulted in a flare of symptoms, rechallenge with viral load measurements was not attempted. CONCLUSION: In this report of a patient with rituximab-associated chronic echovirus 13 meningoencephalitis, viral clearance in response to single treatment options is assessed for the first time. Our observations further support the in vivo efficacy of fluoxetine against enteroviral infections. More research is needed to establish its efficacy in different enterovirus strains.

Direct next-generation sequencing diagnosis of echovirus 9 meningitis, France.

The prognosis of central nervous system infections caused by enteroviruses partially depends on the viral genotype, which is not provided by current point-of-care diagnostic methods. In this study, next-generation sequencing identified an echovirus 9 directly from the cerebrospinal fluid of a patient presenting with meningitis.

Upsurge in echovirus 30 detections in five EU/EEA countries, April to September, 2018.

An upsurge in Echovirus 30 (E30) infections, associated with meningitis/meningoencephalitis, has been observed in Denmark, Germany, the Netherlands, Norway and Sweden in the period April to September 2018, compared with 2015-2017. In total, 658 E30 infections among 4,537 enterovirus infections were detected in 15 countries between January and September 2018 and affected mainly newborns and 26-45 year-olds. National public health institutes are reminded to remain vigilant and inform clinicians of the ongoing epidemic.

Detection and genotyping of enteroviruses in cerebrospinal fluid in patients in Victoria, Australia, 2007-2013.

Genotyping by VP1 fragment polymerase chain reaction (PCR) and nucleic acid sequencing to detect enterovirus (EV) genotypes was performed directly on 729 EV PCR positive cerebrospinal fluid (CSF) samples collected between 2007 and 2012 from Victorian hospital inpatients. The overall genotype identification rate from CSF-positive material was 43%. The four most common genotypes identified were Echovirus 6 (24%), Echovirus 30 (17%), Echovirus 25 (10%), and Coxsackievirus A9 (10%), together comprising 61% of all EVs typed. The seasonal distribution of all EVs identified followed the recognized pattern of mainly summer epidemics. Three of the four predominant genotypes were present in each of the 6 years in which the study was conducted, with 20 other EV genotypes also detected, often in only a single year. Genotyping of EVs directly in CSF is faster, simpler and more sensitive than traditional virus neutralization assays performed on EV positive samples.

Outbreak of echovirus 18 meningitis in a rural Missouri community.

This is a case-control study of viral meningitis outbreak in Perry County, Missouri. All case-patients had viral meningitis per routine CSF analysis or PCR testing. Enterovirus VP1-specific RT-PCR and sequencing was performed on CSF. Univariate and multivariable logistic regression analysis was done to evaluate risk factors. The meningitis attack rate was 1/1,000 population. The main risk factor for meningitis was association with childcare. In patients with signs of meningitis, but normal routine CSF analysis, molecular testing of the CSF is helpful for conclusive diagnosis.

[Rapid enterovirus genotyping in cerebrospinal fluids: a two-year prospective study in a virology laboratory setting]. / Génotypage direct rapide des entérovirus dans le liquide céphalorachidien : étude prospective de deux ans au sein d'un laboratoire de virologie clinique.

UNLABELLED: Enterovirus (EV - 68 serotypes) infections comprise a wide spectrum of clinical presentations including infections of the central nervous system. In severe clinical presentation or epidemics, the precise identification of the involved serotype is necessary. OBJECTIVES: To perform enterovirus genotyping directly in cerebrospinal fluid (CSF) samples, and to assess its feasibility in a laboratory setting. METHODS: Enterovirus genotyping was carried out directly with CSF specimens tested for the diagnostic procedure by amplifying the complete 1D gene encoding the VP1 protein of the HEV-B serotypes (the most frequent) - providing results in two days. Secondly, sequences 1A/1B encoding the VP4/VP2 capsid proteins, respectively, were analysed (results in five days). RESULTS: Direct enterovirus genotyping allowed the identification of enterovirus involved in 77 out of 81 (95%) meningitis cases between January 2006 and December 2007. In combination with the indirect genotyping of enterovirus isolates, identification of the type was achieved in 94 out of 97 (96.9%) patients included in the study. The most frequent serotypes were echovirus 6 (E6) and 13 in 2006, coxsackievirus B2 and E30 in 2007. Four children presented an EV71 associated meningitis. CONCLUSION: When prospectively applied in a laboratory setting, direct enterovirus genotyping in CSF samples allows the identification of the involved enterovirus in two to five days. This time frame is relevant for an optimal patient management, the rapid identification of a new enterovirus variant or in the context of an epidemic alert.

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.

Mosaic genome structure of echovirus type 30 that circulated in Taiwan in 2001.

An echovirus type 30 (E30) outbreak occurred in Taiwan in 2001. In this study, one 1998 and nineteen 2001 enterovirus isolates from cerebrospinal fluid (CSF) of children with meningitis were genetically analyzed. Although negative results were obtained using the E30-specific monoclocal antibody in an immunofluorescent assay (IFA) test of all 20 isolates, molecular typing by partial VP1 sequences and subsequent neutralization test identified them as E30. Among those, seven of them were misidentified as echovirus type 4 (E4) when E4-specific monoclonal antibody was used. Complete genome sequences of one E30 isolate (TW-2513) that were IFA-positive to E4 and another (TW-3182) that was IFA-negative to both E30 and E4 were determined and analyzed. The overall percentage nucleotide identity in the structural coding region (P1) between these two isolates is 98.4, while those in the nonstructural regions P2 and P3 are only 83.2 and 84.4, respectively, indicating that the two 2001 Taiwanese E30 strains were probably recombinant. Recombination analysis of these two E30 genomes revealed that their genome structures are mosaic, which might have been formed gradually and frequently over time.

Echovirus 15 and autumn meningitis outbreak among children, Patras, Greece, 2005.

BACKGROUND: Enteroviruses are the most common cause of aseptic meningitis, presenting in epidemic or endemic form. OBJECTIVES: To determine the causative agent of an aseptic meningitis outbreak in autumn, 2005 in Patras, Greece. STUDY DESIGN: Cerebrospinal fluid (CSF) samples taken during May 2005-February 2006 from children admitted to the Children Hospital of Patras with signs of aseptic meningitis were tested for the presence of enteroviral RNA. Typing was performed by nucleotide analysis. RESULTS: Enteroviruses were detected in 11 (57.9%) of 19 tested CSF samples. In a 12-day period (27 October-7 November 2005) five aseptic meningitis cases were observed. Echovirus 15 was detected in all five cases, and differed from the prototype strain by 27.6%. Enteroviruses before and after this cluster of cases were of different serotypes (Echovirus 9, Echovirus 6). All patients with Echovirus 15 infection were male with a mean age of 7.7 years (2 months-13 years), all recovered successfully. CONCLUSIONS: This is the first report of a cluster of aseptic meningitis cases caused by Echovirus 15. The causative agent was a new variant of Echovirus 15.

[Etiology of aseptic meningitis prevalent in Xuzhou].

BACKGROUND: To find the pathogenic agents of aseptic meningitis prevalent in Xuzhou of Jiangsu province in 2001. METHODS: The enterovirus (EV) was cultured from CSF of the patients and identified with anti-serum by neutralization test. Neutralization titer of antibody in paired sera from meningitis children was determined. EV RNA was detected by RT-PCR. RESULTS: Four strains of Coxsackievirus B5, 2 strains of Coxsackievirus B3 and 1 strain of Echovirus 7 were isolated from 22 CSF specimens. The isolation rate of virus was 31.8% (7/22), 21 CSF were tested by RT-PCR, the positive rate of EV RNA was 52.4% (11/21); 57.9% (11/19) of patients paired-sera had over 4 folds antibody rise or became seroconverted. CONCLUSION: Enterovirus was the pathogenic agent of aseptic meningitis prevalent in Xuzhou of Jiangsu province, the main serotype of the virus was Coxsackievirus B5.

Clinical and diagnostic findings of an echovirus meningitis outbreak in the north west of England.

INTRODUCTION: An outbreak of echovirus meningitis occurred in the north west of England in 2001. This paper reviewed the clinical features and the role of different diagnostic methods. METHODS: This was a prospective study of adults admitted to a regional infectious disease unit with a probable diagnosis of meningitis, March to August 2001. RESULTS: Half the 40 cases were male; median age was 28 (range 16-51) years. Fifteen of 38 (39.5%) were smokers, and 20 of 24 (83.3%) had close contact with children. Median (range) duration of symptoms was 1.1 (0.25-7) days. Symptoms included headache (100%), photophobia (87.5%), and nausea (67.5%), and severity ranged from minimal signs to those consistent with a meningoencephalitis. The diagnosis was confirmed virologically in 29 of 40 (72%); echovirus 30 was isolated from six. Cerebrospinal fluid (CSF) enterovirus polymerase chain reaction (PCR) was positive in 26 of 32 (81%), and CSF virus culture in 3 of 16 (19%). Thirty one per cent of CSF samples had a neutrophil predominance, and 3 of 29 (10%) virologically confirmed cases had normal CSF microscopy and biochemistry. CONCLUSION: CSF microscopy may be normal or suggest bacterial meningitis in a substantial minority of cases of echovirus meningitis. CSF PCR for enterovirus seems to be more sensitive than virus culture of CSF, although PCR does not yield information on circulating virus type. Early and accurate diagnosis could reduce both use of parenteral antibiotics and length of hospital stay with both morbidity and cost implications. Close contact with children may be a risk factor, particularly if good hygiene measures are not practised.

Echovirus 30, Jiangsu Province, China.

An outbreak of aseptic meningitis occurred in the northern area of Jiangsu Province in China from January to July in 2003. A total of 1,681 cases were involved in this outbreak, and 99% of patients were <15 years of age. To identify the etiologic agent, 66 cerebrospinal fluid specimens were tested by cell culture. Eighteen showed an enteroviruslike cytopathic effect on MRC-5 human fetal diploid lung cells. An enterovirus primer-mediated reverse transcriptase-polymerase chain reaction, a standard neutralization assay, and sequencing of the complete capsid-encoding (VP1) gene identified the 18 isolates (FDJS03) as echovirus 30. At least a 10% difference was seen in nucleotide sequences of VP1 between FDJS03 isolates and other global strains of echovirus 30. Phylogenetic analysis based on complete sequences of VP1 was performed to further characterize the FDJS03 isolates. This report is the first to identify a distinct lineage of echovirus 30 as a probable cause of this outbreak.

Cytokine profile in cerebrospinal fluid of children with echovirus type 4 meningitis.

Cytokines play a role in meningeal inflammation and leukocyte recruitment. Research has demonstrated that levels of different cytokines are elevated in aseptic and viral meningitis. Unfortunately, previous data were confounded by the inclusion of multiple viral agents as a study group. The aims of the study were to determine the cerebrospinal fluid concentrations of various cytokines in an outbreak of a single viral agent and to correlate between cytokine levels and leukocytes. Cerebrospinal fluid samples, collected during an outbreak of echovirus type 4 meningitis in infants and children in Israel, were tested for routine characteristics. In addition, cytokine levels were measured in 71 meningitis patients and compared with those of 11 nonmeningitis patients. Concentrations of interleukin-6 (2417 +/- 2713 vs 28 +/- 20 pg/mL; P < 0.01) and interferon gamma (36 +/- 38 vs 4.8 +/- 0.9 pg/mL; P < 0.01) were significantly higher in patients with meningitis than in the control group, whereas soluble intercellular adhesion molecule-1 (1.12 +/- 2.6 vs 0.06 +/- 0.1 ng/mL) levels did not differ significantly. In addition, only interleukin-6 levels correlated with leukocyte counts in viral meningitis patients. Interleukin-6 was the most sensitive and specific characteristic in predicting meningitis in this homogeneous group of patients. Furthermore, only interleukin-6 correlated with leukocyte counts in the cerebrospinal fluid.

Outbreak of aseptic meningitis among adults in southern Taiwan.

A total of 38 adult cases of aseptic meningitis were diagnosed based on clinical manifestations as well as examination of cerebrospinal fluid at Kaohsiung Veterans General Hospital in 2001. The majority (31 cases, 82%) of cases occurred between June and August, and most (33 cases, 87%) of them aged from 18 to 35 years (median age, 25 years). The male-to-female ratio was 2.16:1. Common clinical presentations included headache, fever, nausea or vomiting, and symptoms of common cold. On initial cerebrospinal fluid examination, white cell counts were above 500 cells/mm3 in 7 (18%) cases, and neutrophils predominated in 10 (28%) of 36 cases. The cerebrospinal fluid protein concentration was below 100 mg/dL in 29 (77%) of 38 cases, and hypoglycorrhachia (cerebrospinal fluid/serum glucose ratio <0.5) was seen in 9 (24%) cases. Enterovirus as a definite etiology was confirmed by isolation from cerebrospinal fluid in 17 (45%) of 38 cases and was the presumptive etiology by isolation from the stool or throat swab in 2 (5%) cases. Of the 19 cases with positive viral isolation, echovirus serotype 30 accounted for the majority (15 cases, 79%), followed by echovirus serotype 6 (3 cases, 16%), with one (5%) case undetermined. The viral isolation rate from cerebrospinal fluid correlated to the day of cerebrospinal fluid sampling after disease onset (chi2 = 12.05, p = 0.007). All patients were discharged uneventfully without receiving antibiotic therapy.

First epidemic of aseptic meningitis due to echovirus type 13 among Spanish children.

Echoviruses are the commonest cause of aseptic meningitis (AM). Echovirus type 13 (EV-13) was the second enterovirus serotype associated with different local outbreaks of AM in Spain between February and October 2000. It was the first time that an epidemic AM caused by this virus was recognized in Spain. The index case appeared in the Canary Islands (Canarias). The EV-13 virus was isolated from 135 patients, predominantly from cerebrospinal fluid (CSF). All isolates were from children under 13 years. The age specific peak incidence was in infants under 1 year. Most patients had fever, headache and other meningeal signs. This enterovirus serotype, not previously detected in Spain, caused severe illness with a high attack rate.

[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.

A hospital outbreak of aseptic meningitis due to echovirus type 30 in Antalya, Turkey.

We analyzed clinical and laboratory findings of 23 hospitalized patients with aseptic meningitis in the Department of Pediatrics, Akdeniz University Hospital. The patients presented with the classic symptoms and signs of aseptic meningitis. Protein levels of the cerebrospinal fluid (CSF) samples ranged from 18 to 99 mg/dl, with a mean of 36.5 +/- 4.9 mg/dl. The mean ratio of CSF glucose compared to blood samples was 0.73. Echovirus type 30 was identified in CSF and/or stool samples of 19 patients. Four patients had negative virus culture. The outcome was favorable in all patients. We thought that this outbreak of aseptic meningitis in our department might denote a summer outbreak in the city. However, this remained unproven since field investigations could not be completed. Advances in virus culture or polymerase chain reaction techniques and satisfactory medical records may help patient care by promoting early diagnosis and by eliminating unnecessary antibiotic therapy, allowing epidemiological studies.

[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.

Development and evaluation of a nucleic acid sequence based amplification (NASBA) protocol for the detection of enterovirus RNA in cerebrospinal fluid samples.

A nucleic acid sequence based (NASBA) assay for the generic detection of enterovirus RNA in cerebrospinal fluid (CSF) samples was developed and compared with an established reverse transcription/nested polymerase chain reaction (PCR) protocol. The sensitivity of NASBA followed by detection of amplicons with a biotinylated oligonucleotide probe was < or = ten copies of enterovirus RNA, indicating that enterovirus NASBA achieves a similar sensitivity as nested PCR. Moreover, NASBA detected a panel of 22 different serotypes of the species poliovirus, human enterovirus A, human enterovirus B and human enterovirus C completely. For evaluating NASBA as a diagnostic tool, 61 CSF samples of patients suffering from aseptic meningitis were tested in parallel with NASBA and nested PCR. NASBA detected enterovirus RNA in four CSF samples, two of these were also positive by nested PCR and two other CSF samples were positive only by nested PCR (in total six positive samples). All other 55 of 61 CSF samples were concordantly enterovirus negative by both methods. In conclusion, the more simple to handle 'one step' NASBA is as sensitive as nested PCR and may be used as an alternative method for the detection of enterovirus RNA in CSF samples. Enterovirus NASBA is a 'one step' RNA amplification procedure that is less prone to cross-contamination compared to a three step nested PCR.