IgM-positive Sera Research Articles

Evidence for the presence of autochthonous (locally acquired) cases of acute hepatitis E virus infections in Italy since the 80s

BackgroundAutochthonous (locally acquired) cases of acute hepatitis E virus have been recently reported in several developed countries. AimTo evidence cases, if any, and characteristics of acute hepatitis E virus infections in North-East of Italy several years ago. MethodsIn 2014, stored sera of 165 nonA–nonB acute hepatitis referred to the hospital of Padua during the period 1978–1991 were tested for hepatitis C virus antibodies by EIA III and for anti-hepatitis E virus IgM by Wantai HEV IgM ELISA. Anti-hepatitis E virus IgM positive sera were tested by Real Star HEV RT-PCR kit (Altona Diagnostics, Hamburg, Germany). ResultsNinety-six (58.1%) sera resulted anti-HCV positive, and thus classified as acute C hepatitis. None of these subjects was anti-HEV IgM positive. Out of the 69 anti-HCV negative cases, 4 (5.8%) resulted anti-HEV IgM positive (one case hepatitis E virus-RNA positive), with an increasing trend from 2.8% during the years 1978–1984 to 9.1% during the years 1985–1991. All cases occurred in Italian patients with no travel abroad history. ConclusionsThere is evidence for the presence of autochthonous cases of acute hepatitis E virus infections in Italy since 80s.

Determination of anti-Mycobacterium tuberculosis IgM and IgG antibodies in individuals in contact with patients with active tuberculosis.

Determination of anti-Mycobacterium tuberculosis IgM and IgG antibodies in individuals in contact with patients with active tuberculosis.

Bilateral herpetic keratouveitis in an immunocompetent patient

Case reportWe report the case of an immunocompetent male who presented with a limbal-adjacent scleritis and interstitial keratitis in the left eye. A few days later a new dendritiform ulcer in his right eye and bilateral progressive worsening with granulomatous uveitis in both eyes were observed. A thorough review of systems revealed positive serum IgM titles for herpes simplex virus. DiscussionIn the context of a bilateral keratouveitis refractory to conventional treatment it is mandatory to rule out the herpetic origin based on the different forms of clinical presentation of this virus.

Queratouveítis herpética bilateral en paciente inmunocompetente

Case reportWe report the case of an immunocompetent male who presented with a limbal-adjacent scleritis and interstitial keratitis in the left eye. A few days later a new dendritiform ulcer in his right eye and bilateral progressive worsening with granulomatous uveitis in both eyes were observed. A thorough review of systems revealed positive serum IgM titles for herpes simplex virus. DiscussionIn the context of a bilateral keratouveitis refractory to conventional treatment it is mandatory to rule out the herpetic origin based on the different forms of clinical presentation of this virus.

Evaluation of bio-conjugation methods for obtaining of synthetic positive control for IgM-capture ELISA

The enzyme-linked immunosorbent assay (ELISA) is the most informative and versatile method of serological diagnostics. The possibility of detecting by ELISA specific antibodies of different classes allow to differentiate primary infectious process and its remission, exacerbation and chronic disease (differential diagnosis). This approach is implemented in the methodology for evaluation of patients for the presence of humoral immune response to TORCH-infections pathogens (toxoplasmosis, rubella, cytomegalovirus, herpes simplex viruses’ infections, and some others). Therefore, testing for presence of specific IgG and IgM antibodies against TORCH-infections pathogens in blood serum is an important element of motherhood and childhood protection. The essential problem in the production of IgM-capture ELISA diagnostic kits is obtaining of positive control. The classic version of positive control is human blood serum (plasma) containing specific antibodies. But specific IgM-positive sera are insignificant raw materials. This fact can seriously restrict the production of diagnostic kits, especially in the event of large-scale production. We have suggested the methodological approach to using of synthetic positive controls in IgM-capture ELISA kits based on conjugate of normal human IgM and monoclonal antibodies against horseradish peroxidase. It is found that this task can be fulfilled by means of NHS ester-maleimide-mediated conjugation (by sulfosuccinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate), reductive amination-mediated conjugation (by sodium periodate) and glutaraldehyde-mediated conjugation. It was found that conjugates of normal human IgM and monoclonal antibodies against horseradish peroxidase obtained using NHS ester-mediated maleimide conjugation and periodate method were similar by molecular weight, whereas conjugate synthesized by glutaraldehyde method comprised at least three types of biopolymers with close molecular weight. It was found that synthetic positive control obtained by different methods was characterized by higher titer compared to IgM-positive high-titer serum. However, positive control obtained by NHS ester-mediated maleimide conjugation had the best titration profile characteristics.

Synthetic peptides for efficient discrimination of anti-enterovirus antibodies at the serotype level

Enteroviruses are important human pathogens, causing a broad spectrum of diseases from minor common colds to fatal myocarditis. However, certain disease syndromes are caused by one or few serotypes. Serotype identification is difficult due to the laborious neutralization tests that lack of sensitivity, while in commercial ELISAs homotypic antibodies’ activities are largely masked by the recognition of genera-specific epitopes by heterotypic antibodies. In the present study homotypic assays were developed with the ability to discriminate different enterovirus serotypes. Seventy-three children sera, positive for IgM antibodies against enterovirus genus and 49 healthy children were examined for the presence of antibodies against 14 synthetic peptides derived from a non-conserved region of the VP1 protein of coxsackieviruses B2, B3, B4, B5, A9, A16, A24, echoviruses 6, 7, 9, 11, 30, enterovirus 71 and parechovirus 1. 50% of the anti-enterovirus IgM positive sera (>150BU) reacted with the peptides with the majority of them to preferentially recognize one of them, supporting the homotypic nature of our assay. Inhibition studies yielded homologous inhibition rates 67–95% suggesting that specific peptide recognition actually occurred. The diagnostic value of our assay was tested in blood samples drawn over a 1.5-year period from a 5-year old patient. The anti-enterovirus reactivity was clearly attributed to echovirus serotype 11. The IgM/IgG antibody ratio was reversed 4 months later and subsequently IgM antibodies dropped below the cutoff point. In this paper we demonstrate that our assay can be used to discriminate between antibodies targeting different enterovirus serotypes.

Potential Impact of Different Cytomegalovirus (CMV) IgM Assays on an Algorithm Requiring IgM Reactivity as a Criterion for Measuring CMV IgG Avidity

The measurement of cytomegalovirus (CMV) IgG avidity is a powerful tool for identifying individuals with recent CMV infection. Because such patients are expected to be positive for CMV IgM, several investigators have suggested that CMV IgG-positive sera first be screened for CMV IgM and then only the IgM-reactive sera be tested for avidity. We investigated the impact of different CMV IgM assays on such a reflexing algorithm using a panel of 369 consecutive IgG-positive serum samples submitted for avidity testing. A bead-based immunofluorescent assay (BIFA) identified 105 IgM-positive serum samples, whereas an IgM-capture enzyme immunoassay (EIA) identified 48 IgM-positive serum samples; this marked difference led us to evaluate additional CMV IgM assays. An enzyme-linked immunofluorescent assay (ELFA) and a chemiluminescent immunoassay (CIA) were used to test all sera with discordant BIFA/EIA results, all sera with concordant positive results, and selected sera with concordant negative results. The findings indicated that the ELFA would identify 74 CMV IgM-positive samples and the CIA would identify 64. Of the 23 low-avidity serum samples, 2 were IgM negative by BIFA, 3 by ELFA and CIA, and 4 by EIA; of the 23 intermediate-avidity serum samples, 6 were IgM negative by BIFA, 10 by ELFA, and 15 by EIA and CIA. In both these avidity groups, BIFA IgM-negative sera were also negative by the other 3 assays. These findings demonstrate that an algorithm requiring CMV IgM reactivity as a criterion for CMV IgG avidity testing does not identify all low-avidity sera and thus misses some cases of acute CMV infection.

Hypoproteinemia in childhood: What should you look for?

Introduction: Menetrier’s disease (MD) is a fundus and body gastritis frequently associated with hypoalbuminemia secondary to protein loss through the gastricmucosa. Clinical findings include vomiting, diarrhoea, abdominal pain, anorexia and peripheral oedema inmost cases. In one third of cases Cytomegalovirus (CMV) it is demonstrated having a central role in the pathogenesis and in childhood MD is usually a benign and self-limiting condition, lasting for a few weeks. Patients and methods: We present a 2 yrs old girl with important protein-loss admitted to our Unit; we describe diagnosis and management discussing the causative role of CMV in MD. Results: Two yrs-old girl with a few days history of diarrhoea, cough and fever especially during the night, important peripheral and periorbital oedemas presented at our Unit. Serological tests revealed anemia, iron deficiency, WBC elevated, hyponatremia, hypocalcemia, low level of thyroid hormones, hypoproteinemia, hypoalbuminemia and she was treated by human albumin. Antigliadin, antiendomysial and anti-tissue transglutaminase antibodies were negative. Urinary protein loss was excluded and her liver synthetic function was normal. Abdominal ultrasonography was negative for abdominal ascitic fluid. Nephrological cause was excluded and gastrointestinal causes were studied. Upper gastrointestinal endoscopy was performed and it showed mild multinodular duodenitis and a widely bleeding mucosa with large areas of erosions in the gastric body and fundus. Histologic examination confirmed a profound chronic active hypertrophic gastritis with intense regenerative epithelial hyperplasia. Tissue culture for Helicobacter Pylori and intranuclear CMV inclusion bodies were negative. Duodenal biopsy revealed normal mucosa and villous that excluded coeliac syndrome. An infectious cause is hypothised and serological tests for virus, bacteria and parasites revealed positive serum IgM antibodies against CMV; CMV DNAwas detected in blood through polymerase chain reaction (PCR). During her stay in hospital, shewas treatedwith PPI and another human albuminwas not necessary because of the improvement of her clinical condition. Next upper endoscopy after 20 days showed healing of duodenitis and an erythematous mucosa in the gastric body and fundus without erosions. She continued therapy with PPI and at clinical follow-up 4weeks later shewas free of symptomswith satisfactory weight gain and her haemoglobin, serum electrolytes and serum protein levels were normal, including IgA levels that exclude any Ig deficit. Conclusions: One of the major causes of MD in children is the CMV infection that should be suspected in every childwhopresents with protein-losing gastropathy. Upper endoscopy with histological examination and PCR level should be performed. Usually only supportive care is required, according to literature that describes MD in children as a transient and self-limiting condition. PO26

Anti-hepatitis E virus antibody prevalence in French expatriate workers

The hepatitis E virus (HEV) is a leading cause of acute hepatitis in developing countries and an emerging pathogen in Europe. HEV seroprevalence has rarely been assessed in cohorts of travelers, and previous studies have reported a very low rate of exposure. We assessed HEV seroprevalence in French expatriate workers. The prevalence of HEV IgG and IgM was assessed among 43 French expatriate workers using two commercial microplate enzyme immunoassays (Adaltis and Wantai). Additionally HEV IgG-positive sera were tested with an immunoblot assay (recomLine), while IgM-positive sera were tested with a rapid immunochromatographic assay (Assure). The prevalence of anti-HEV IgG was 3.7 times higher in French expatriates than in comparable blood donors from the same area. A discrepancy was evidenced between the HEV IgG results obtained by the Wantai and Adaltis assays (48.8% vs. 30.2%). Expatriation from France, including to areas not recognized as hyperendemic for HEV, may expose individuals to HEV infection. This issue warrants further study; in particular, serology should be compared before and after travel. The most sensitive Wantai serological assay should be used for epidemiological studies to obtain better insight into the epidemiology of HEV.

Asian Musk Shrew as a Reservoir of Rat Hepatitis E Virus, China

Asian Musk Shrew as a Reservoir of Rat Hepatitis E Virus, China

High prevalence of rat hepatitis E virus in wild rats in China

Serum samples from a total of 713 wild rats captured in Zhanjiang city in China from December 2011 to September 2012 were investigated for the prevalence of rat hepatitis E virus (HEV) by exploring rat HEV-specific antibodies and RNA. By an ELISA based on recombinant rat HEV-like particles (HEV-LPs), 23.3% (166/713) of the rats were positive for anti-HEV IgG, and 8.3% (59/713) were positive for anti-HEV IgM. The IgG-positive rates in Rattus norvegicus, Bandicota indica, Rattus flavipectus, Rattus rattoides losea, and Rattus rattus hainanus, were 27.8% (64/230), 23.0% (40/174), 19.9% (34/171), 21.5% (26/121), and 11.8% (2/17), while the IgM-positive rates were 8.3% (19/230), 6.9% (12/174), 8.2% (14/171), 10.7% (13/121), and 5.9% (1/17), respectively. The IgG-positive rate of the rats captured in rural areas, 24.1% (84/348), was higher than that in the central area of Zhanjiang city, 15.1% (32/212). The highest IgG-positive rates, as high as 45.3% (39/86), were detected in wild rats trapped in the garbage dump. Twelve of the 59 IgM-positive serum samples were positive for HEV RNA, which was detected in all of the wild rat species except R. rattus hainanus. A phylogenetic analysis of the partial genome of rat HEV ORF1 indicated that all of the 12 HEV strains belong to rat HEV, and no other genotype HEV were detected. The rat HEV from Zhangjiang city could be classified into three separated clusters, suggesting that the infection due to rat HEV with a variety of genome entities occurs extensively among wild rats in China.

Neuropsychiatric symptoms are common in immunocompetent adult patients with Toxoplasma gondii acute lymphadenitis

Chronic toxoplasmosis has been shown to be strongly associated with a range of neuropsychiatric effects including schizophrenia and suicide. However there have not been any prospective, community-based studies of the neuropsychiatric effects of acute toxoplasmosis in adult immunocompetent patients. Adult patients with a positive serum IgM anti-Toxoplasma gondii test result, in the context of an acute illness with lymphadenopathy, were invited to complete a questionnaire seeking information relating to the nature, severity, and duration of symptoms in the months following the diagnosis of acute toxoplasmosis. Laboratory testing identified a total of 187 adults who had a positive serum IgM anti-T. gondii test result between 1 January and 30 November 2011. Consent to contact 108/187 (58%) patients was provided by their family doctor; 37 (34%) of these 108 patients completed and returned the questionnaire. Questionnaires from the 31/108 (29%) patients who reported swollen lymph nodes during their illness were included in the study. Fatigue (90%), headache (74%), difficulty concentrating (52%), and muscle aches (52%) were the most commonly reported symptoms. These symptoms commonly persisted for at least 4 weeks. Twenty-seven of 31 (87%) subjects reported a moderate or severe reduction in their overall physical and mental health during the first 2 months of illness. Acute toxoplasmosis in immunocompetent adults commonly causes moderately severe neuropsychiatric symptoms that might result from replication of the organism in the central nervous system with consequent effects on brain function. Patients should be advised that such symptoms are common and reassured that they usually resolve completely within a few months.

Analysis of genotype IV distribution of hepatitis E virus infections in Wuhan by sequencing the open reading frame 3 gene

To determine the distribution of genotype IV among hepatitis E virus (HEV) infections in Wuhan by sequencing the open reading frame (ORF) 3 gene of HEV clinical isolates. Serum samples were collected from 103 individuals who tested positive for the anti-HEV IgM antibody, and total HEV RNA was extracted for targeted gene sequencing analysis. Reverse transcription-nested polymerase chain reaction (PCR) was used to amplify two fragments of the ORF3 gene (5020 to 5392 nt and 5347 to 5956 nt, EF570133). The two PCR products were sequenced and the sequences were stitched with the ContigExpress program and used to determine the HEV genotype. Both ORF3 gene fragments were amplified in 18 out of the 103 anti-HEV IgM-positive serum samples. These 18 HEV isolates shared 92.5% to 99.4% identity with each other at the nucleotide level. Nucleotide sequence homology analysis of the HEV genotypes I, II, III, and IV indicated the highest homology was with genotype IV; specifically, homology with genotype I was 83.5% to 86.7%, with genotype II was 83.2% to 85.2%, with genotype III was 84.6% to 87.2%, and with genotype IV was 92.0% to 96.5%. Targeted sequencing of the HEV ORF3 gene facilitated genotyping of clinical isolates. Using this method, it was determined that nearly 20% of HEV clinical isolates from Wuhan belong to genotype IV.

Production and Evaluation of Toxoplasma gondii Recombinant GRA7 for Serodiagnosis of Human Infections

The precise diagnosis of the acute toxoplasmosis in pregnant women and immunocompromsied patients has critical importance. Most of the commercially available assays use the whole Toxoplasma soluble extract as the antigen. However, the assays currently available for the detection of specific anti-Toxoplasma antibodies may vary in their abilities to detect serum immunoglobulins, due to the lack of a purified standardized antigen. The aim of this study was production and evaluation of the usefulness of the recombinant Toxoplasma gondii GRA7 antigen for the serodiagnosis of Toxoplasma gondii IgM and IgG by ELISA. A total of 70 T. gondii IgM positive sera, 74 T. gondii IgG positive sera, and 60 sera from subjects who were not infected with T. gondii were examined. These sera were shown different absorbance values in ELISA test. To control the specificity of the rGRA7 other parasitic diseases, for example, echinococcosis, malaria, leishmaniasis, fascioliasis, and strongyloidiasis were tested of which none showed positive results. Sensitivity and specificity of the generated recombinant IgG ELISA in comparison with commercial ELISA (com ELISA) were 89% and 90%, and the sensitivity and specificity of the generated recombinant IgM ELISA were 96% and 90%, respectively. The results obtained here show that this antigen is useful for diagnostic purposes.

Molecular characterization of hepatitis E virus in patients with acute hepatitis in Venezuela

Hepatitis E virus (HEV) causes a common infection in developing countries. HEV infection occurs as outbreaks, as sporadic clinical cases and as large epidemics in endemic areas. The objective of this study was to determine the presence of HEV infection in patients with clinical suspicion of hepatitis A virus (HAV) infection, referred to the Instituto Nacional de Higiene "Rafael Rangel" in Venezuela. Seventy-four sera were tested for anti-HAV and anti-HEV IgM antibodies. HEV-RNA was amplified from anti-HEV IgM positive sera using nested reverse transcription polymerase chain reaction for ORF1 (RNA dependent RNA polymerase region) and the amplicons sequenced for phylogenetic analysis. The frequency of anti-HEV IgM was 22/74 (30%) in the samples tested. Dual infection with HAV and HEV was found in 31% (12/39) of anti-HAV IgM positive patients. Viremia was detected in 3/22 (14%) of sera positive for anti-HEV IgM. Two HEV strains were classified as genotype 1 and one as genotype 3, which were closely related to Yam 67 (north of India) and US1 isolates from the USA, respectively. These findings suggest that HEV is an important cause of acute viral hepatitis in Venezuela as a single infection or co-infection with HAV, with high morbidity in children and young adults suggesting that this infection is endemic in Venezuela.

Application and expression of Toxoplasma gondii surface antigen 2 (SAG2) and rhoptry protein 2 (ROP2) from recombinant Escherichia coli strain

The gene encoding surface antigen 2 (SAG2) or rhoptry protein 2 (ROP2) of Toxoplasma gondii was cloned into the plasmid pGEX-4T-1 and subsequently expressed in Escherichia coli as a glutathione-s-transferase (GST) fusion protein. The characteristics of purified GST-SAG2 or GST-ROP2 were analysed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analysis. The specific IgG of a panel of serum samples provided by the National Institute for the Control of Pharmaceutical and Biological Products were tested with commercial ELISA and the lateral flow immunoassay (LFIA) based on GST-SAG2, GST-ROP2 or GST-SAG2+ROP2. A total of 1096 sera and saliva samples from pregnant women were tested by GST-SAG2+ROP2-LFIA. In total, 20 T. gondii IgM positive sera (1.82%), 81 T. gondii IgG positive sera (7.4%) and 23 T. gondii IgA positive saliva (2.1%) were finally confirmed. The SAG2+ROP2 specific IgG and IFN-γ producing CD8+ T cells were induced in mice immunised with GST-SAG2+ROP2. The results indicate that GST-SAG2+ROP2 protein can be used as an antigen for diagnosing T. gondii infection and provide a strategy for development of subunit vaccines for protection against T. gondii infection.

The coastal areas of Yantai human and swine hepatitis E virus genotyping analysis

Survey of the coastal city of Yantai, from human and swine hepatitis E virus (HEV) genotype correlation. Application of reverse transcription nested polymerase chain reaction (RT-nPCR) method for local acute sporadic hepatitis E patients,normal population of HEV-IgM positive and local pig farm pigs were HEV RNA detection. And HEV RNA positive samples for cloning sequencing and sequence analysis. In 16 patients with acute sporadic hepatitis E in 7 cases of RNA positive stool specimens of HEV; 51 IgM positive sera of normal people in specimens with 1 HEV RNA positive; 34 pig bile specimens with 1 HEV RNA positive. Sequence analysis revealed the region HEV strains and swine strains in the ORF2 region of nucleotide sequence homology is 87%-98.1%. 7 strains of hepatitis E virus genotype in patients and 1 strains of swine hepatitis E virus genotypes are type IV, gene sequence homology between the 87%-98.1%; there were 6 patients and porcine gene sequence homology in 93.9%-98.1% between,for type a subtype; 1 patients and porcine gene sequence homology in 87%, for the type D subtype. Normal population of 1 cases of hepatitis E virus genotype for I type D subtype. Human and porcine HEV ORF2 gene fragment and HEV part I-IV representative strains were compared, and the nucleotide sequence homology were 82.5%-100%, 81.7%-92.9%, 81.4%-93.9%, 84.9%-100%. The area population prevalence of HEV in the presence of 2 genotype 3 subtype genes, mainly to IV A, in pigs with popular HEV gene with a high homology; HEV type I in the crowd disperses in the presence of.

Acute neuromuscular weakness associated with dengue infection

Background:Dengue infections may present with neurological complications. Whether these are due to neuromuscular disease or electrolyte imbalance is unclear.Materials and Methods:Eighty-eight patients of dengue fever required hospitalization during epidemic in year 2010. Twelve of them presented with acute neuromuscular weakness. We enrolled them for study. Diagnosis of dengue infection based on clinical profile of patients, positive serum IgM ELISA, NS1 antigen, and sero-typing. Complete hemogram, kidney and liver functions, serum electrolytes, and creatine phosphokinase (CPK) were tested. In addition, two patients underwent nerve conduction velocity (NCV) test and electromyography.Results:Twelve patients were included in the present study. Their age was between 18 and 34 years. Fever, myalgia, and motor weakness of limbs were most common presenting symptoms. Motor weakness developed on 2nd to 4th day of illness in 11 of 12 patients. In one patient, it developed on 10th day of illness. Ten of 12 showed hypokalemia. One was of Guillain-Barré syndrome and other suffered from myositis; they underwent NCV and electromyography. Serum CPK and SGOT raised in 8 out of 12 patients. CPK of patient of myositis was 5098 IU. All of 12 patients had thrombocytopenia. WBC was in normal range. Dengue virus was isolated in three patients, and it was of serotype 1. CSF was normal in all. Within 24 hours, those with hypokalemia recovered by potassium correction.Conclusions:It was concluded that the dengue virus infection led to acute neuromuscular weakness because of hypokalemia, myositis, and Guillain-Barré syndrome. It was suggested to look for presence of hypokalemia in such patients.

National Prevalence Estimates for Cytomegalovirus IgM and IgG Avidity and Association between High IgM Antibody Titer and Low IgG Avidity

Primary cytomegalovirus (CMV) infection of the mother during pregnancy presents risk of CMV infection of the fetus with resulting permanent disability. CMV IgM antibody is generated following primary CMV infection but also can appear during nonprimary CMV infection and is thus of limited diagnostic use by itself. In contrast, the presence of low CMV IgG avidity has been shown to be a unique and reliable serologic indicator of primary CMV infection. We measured CMV IgG and IgM antibody levels and IgG avidity in sera from a population sample of 6,067 U.S. women aged 12 to 49 years from NHANES (National Health and Nutrition Examination Survey). The CMV IgG prevalence was 58% overall and increased strongly with age. The CMV IgM prevalence was 3.0% overall and remained relatively flat across age groups. The prevalence of low IgG avidity was 2.0% overall, decreased sharply with age, and was seen mainly among IgM-positive sera. Fourteen to 18% of the CMV IgM-positive sera were low IgG avidity, presumably representing primary CMV infection. High CMV IgM antibody titer was a strong predictor of low IgG avidity. The ability to reliably identify primary CMV infection during pregnancy is important for management of the pregnancy, including possible treatment options for the fetus. Both IgM and IgG avidity measurements provide useful clinical information for evaluating primary CMV infection, although commercial tests for CMV IgG avidity are not yet widely available in the United States.

Acute West Nile virus neuroinvasive infections: Cross‐reactivity with dengue virus and tick‐borne encephalitis virus

Cross-reactions in serology are common among flaviviruses. During the outbreak of West Nile virus (WNV) infections in Greece in 2010, WNV IgM-positive serum and cerebrospinal fluid samples were tested for the presence of IgM and IgG antibodies against Dengue virus (DENV) and tick-borne encephalitis virus. Higher cross-reactivity was observed in IgM antibodies between WNV and DENV; however, the index of the WNV antibodies was in all cases higher than that of the DENV antibodies. There is a need for caution when evaluating serologic results of flaviviral infections, while efforts have to be focused on the development of diagnostic assays with increased specificity.