Luine Rosele Vidal

Acute gastroenteritis and enteric viruses in hospitalised children in southern Brazil: aetiology, seasonality and clinical outcomes.

Viral acute gastroenteritis (AG) is a significant cause of hospitalisation in children younger than five years. Group A rotavirus (RVA) is responsible for 30% of these cases. Following the introduction of RVA immunisation in Brazil in 2006, a decreased circulation of this virus has been observed. However, AG remains an important cause of hospitalisation of paediatric patients and only limited data are available regarding the role of other enteric viruses in these cases. We conducted a prospective study of paediatric patients hospitalised for AG. Stool samples were collected to investigate human adenovirus (HAdV), RVA, norovirus (NoV) and astrovirus (AstV). NoV typing was performed by nucleotide sequencing and phylogenetic analysis. From the 225 samples tested, 60 (26%) were positive for at least one viral agent. HAdV, NoV, RVA and AstV were detected in 16%, 8%, 6% and 0% of the samples, respectively. Mixed infections were found in nine patients: HAdV/RVA (5), HAdV/NoV (3) and HAdV/NoV/RVA (1). The frequency of fever and lymphocytosis was significantly higher in virus-infected patients. Phylogenetic analysis of NoV indicated that all of these viruses belonged to genotype GII.4. The significant frequency of these pathogens in patients with AG highlights the need to routinely implement laboratory investigations.

Laboratory Diagnosis, Epidemiology, and Clinical Outcomes of Pandemic Influenza A and Community Respiratory Viral Infections in Southern Brazil

ABSTRACT Community respiratory viruses (CRVs) are commonly associated with seasonal infections. They have been associated with higher morbidity and mortality among children, elderly individuals, and immunosuppressed patients. In April 2009, the circulation of a new influenza A virus (FLUA H1N1v) was responsible for the first influenza pandemic of this century. We report the clinical and epidemiological profiles of inpatients infected with CRVs or with FLUA H1N1v at a tertiary care hospital in southern Brazil. In addition, we used these profiles to evaluate survivor and nonsurvivor patients infected with FLUA H1N1v. Multiplex reverse transcription-PCR (RT-PCR) and real time RT-PCR were used to detect viruses in inpatients with respiratory infections. Record data from all patients were reviewed. A total of 171 patients were examined over a period of 16 weeks. Of these, 39% were positive for FLUA H1N1v, 36% were positive for CRVs, and 25% were negative. For the FLUA H1N1v- and CRV-infected patients, epidemiological data regarding median age (30 and 1.5 years), myalgia (44% and 13%), need for mechanical ventilation (44% and 9%), and mortality (35% and 9%) were statistically different. In a multivariate analysis comparing survivor and nonsurvivor patients infected with influenza A virus H1N1, median age and creatine phosphokinase levels were significantly associated with a severe outcome. Seasonal respiratory infections are a continuing concern. Our results highlight the importance of studies on the prevalence and severity of these infections and that investments in programs of clinical and laboratory monitoring are essential to detect the appearance of new infective agents.

Quantitation of cerebrospinal fluid lactic acid in infectious and non-infectious neurological diseases

Abstract Background: The differential diagnosis between bacterial and viral meningitis is not easy in some cases. Cerebrospinal fluid (CSF) analysis is essential for establishing this diagnosis. The objectives were to quantitate lactic acid (LA) concentrations in bacterial and viral meningitis, and other central nervous system (CNS) diseases in order to evaluate the diagnostic utility of CSF LA for discriminating bacterial from viral meningitis. Methods: CSF LA was measured in 139 CSF samples from seven groups: viral meningitis with classic CSF; suspicion of viral meningitis with neutrophils in CSF; bacterial meningitis; non-infectious neurological diseases; chronic meningitis; traumatic lumbar puncture (LP) and normal CSF. Results: CSF LA was higher in bacterial meningitis 8.7+5.4 mmol/L compared with viral meningitis (1.9+0.6) and the other groups (p<0.0001). CSF LA in the groups with viral meningitis was not different compared to groups with non-infectious CNS diseases and chronic meningitis. The ability of CSF LA to discriminate bacterial from viral meningitis showed a sensitivity of 80% and specificity of 97%, positive predictive value (PPV) of 94%, and negative predictive value (NPV) of 89%. Conclusions: CSF LA is a powerful test to discriminate bacterial from viral meningitis with high sensitivity, specificity and predictive values. CSF LA can help in the cases with diagnostic uncertainty. Clin Chem Lab Med 2009;47:755–61.

Adenovirus respiratory infection: significant increase in diagnosis using PCR comparing with antigen detection and culture methods.

Adenovirus (AdV) respiratory infections are usually described as being associated with high mortality rates. Laboratory diagnosis is essential for the establishment of the appropriate therapy, and for guiding the implementation of preventive measures in order to prevent the spread of the infection. Aiming to analyze the sensitivity and specificity of the laboratorial diagnosis methods available, we compared antigen detection by indirect immunofluorescence assay (IF), and a specific nested polymerase chain reaction (PCR), to detect AdV in respiratory samples collected from patients admitted to hospital with acute respiratory disease. Positive samples were inoculated into a cell culture to confirm the results. We analyzed 381 samples from the nasopharyngeal aspirates collected during the year 2008; of these, 2.6% tested were positive for adenovirus through IF and 10% through PCR; positive isolation was obtained in 40% and 26% of these cases, respectively. Most infected patients were children under six months of age, and despite of the fact that a significant number of patients required intensive care, the mortality rate was low (5%). In conclusion, molecular methods were found to be useful for rapid diagnosis of adenovirus infections with higher sensitivity than antigen detection; their introduction permitted a significant increase in diagnoses of adenovirus infections.

Impact of human metapneumovirus infection on in and outpatients for the years 2006-2008 in Southern Brazil

The human metapneumovirus (hMPV), member of the Paramyxoviridae family, has been reported as an important agent involved with acute respiratory infections (ARIs). The aim of this study is to identify hMPV as the etiological agent of ARIs on in and outpatients in the city of Curitiba, Southern Brazil, and describe clinical data of hMPV subtyping. A retrospective study was performed in 1,572 respiratory samples over a period of three years. hMPV was detected by reverse transcription-polymerase chain reaction and subtyping was performed by nucleotide sequencing. hMPV was present in 61 (3.9%) samples and subtypes A1, A2a, B1 and B2 were detected. The incidence of hMPV was higher in outpatients (5.9%), whose mean age was 19.7 years (range 6 months-75 years old), than in inpatients (3%), whose mean age was 7.6 months (range 1 month-26 years old). The outpatients had upper respiratory tract infections with flu-like symptoms and all hospitalized children had lower respiratory tract infections. A pediatric patient died from complications associated with hMPV A2a infection. hMPV has been reported as a respiratory pathogen in all age groups. No correlation was observed between viral subtype and disease severity in the samples of this study.

Molecular epidemiology of HIV-1 clades in Southern Brazil

Human immunodeficiency virus (HIV) clades B and C account for more than 60% of the HIV-1 infections worldwide. In this paper, we describe the profiles of patients infected with subtypes of HIV-1 from the state of Paraná, Southern Brazil, and correlate them with demographic and epidemiological findings. A retrospective analysis of HIV cases reported from 1999-2007 was also performed. Data from 293 patients were reviewed and 245 were older than 13 (58% female). The distribution of clades was as follows: B 140 (57%), C 67 (23%), F 24 (10%) and mosaic or unique recombinant forms (URFs) 24 (10%). Of the 48 patients younger than 13 years of age (62.5% male), vertical transmission occurred in 46 and the distribution of clades was as follows: B 14 (29%), C 24 (50%), F 7 (15%) and URFs 6 (13%). There was no significant difference in mortality between HIV-1 subtypes. In both groups, patients infected with clade C tended to have higher rates of injection drug use exposure risk.

The epidemiology and antigenic characterization of influenza viruses isolated in Curitiba, South Brazil

Several studies conducted all over the world have reported that the influenza virus is associated with great morbidity and mortality rates. In this study, we analyzed the incidence of the influenza virus between 2000 and 2003 in Curitiba. We studied 1621 samples obtained from outpatients and hospitalized patients of both sexes and all ages. The study was conducted at the local primary care health units (outpatients) and at the tertiary care unit (hospitalized) of the General Hospital of the Federal University in the state of Paraná, Brazil. Nasopharyngeal aspirates and, eventually, bronchoalveolar lavage were assayed for the presence of viral antigens, either by indirect immunofluorescence or cell culture. Of the samples studied, 135 (8.3%) were positive for influenza virus, and of those, 103 (76.3%) were positive for type A and 32 (23.7%) for type B. Additionally, positive samples were analyzed by reverse transcription followed by polymerase chain reaction and subtypes H1 and H3 were identified from this group. A high incidence of positive samples was observed mainly in the months with lower temperatures. Furthermore, outpatients showed a higher incidence of influenza viruses than hospitalized patients.

Rotavirus infection in a tertiary hospital: laboratory diagnosis and impact of immunization on pediatric hospitalization

BACKGROUND AND OBJECTIVES Rotavirus (RV) is the main etiological agent of diarrhea in childhood; its laboratory diagnosis is crucial to guide the clinical management and prevention of its spread. RV immunization was introduced in Brazilian 6-month-old children in 2006. The present study was aimed to evaluate three methodologies used for human RV detection in stool samples obtained from patients hospitalized due to gastroenteritis in a teaching hospital and report the impact of RV immunization in hospitalization by diarrhea. METHODS 293 stool samples collected in the 2001-2008 period were analyzed by enzyme immunoassay (EIA), latex agglutination (LA) and polyacrylamide gel electrophoresis (PAGE). RESULTS Rotavirus was detected in 34.8% of samples by LA assay, 28.3% of samples by EIA assay and in 25.6% of samples by PAGE assay. Considering the PAGE method as gold standard, the sensitivity, specificity and accuracy of EIA were 94.6%, 94.4% and 94.5%, and to LA were 82.6%, 81.6% and 81.9%, respectively. CONCLUSION These results indicate that antigen detection by EIA is a rapid, sensitive and specific method, and could be used in large-scale applications for screening stool samples suspected of RV infection. This study showed decreased incidence of RV infection in hospitalized children prior to the implementation of the national immunization program against RV.

Quantification of cerebrospinal fluid lactic acid in the differential diagnosis between HIV chronic meningitis and opportunistic meningitis.

Abstract Background: Approximately 40% of HIV infected patients have chronic meningitis at various stages during the infection, 59% are asymptomatic. This is a diagnosis of exclusion and a confounding factor in cerebrospinal fluid (CSF) analysis, any other causes of chronic meningitis by opportunistic or co-infection must be ruled out. The aim of this study was to analyze CSF lactic acid (LA) as an adjuvant biomarker in chronic meningitis due to HIV. Methods: CSF LA was quantified in 223 CSF samples by the Dimension AR (Dade Behring, Deerfield, IL, USA), distributed into nine groups: 1) HIV positive with an increase in CSF WBCs (n=17); 2) HIV positive with normal CSF (n=20); 3) enterovirus meningitis (n=33); 4) Herpesviridae meningoencephalitis (n=30); 5) fungal meningitis (n=25); 6) tuberculosis (TB) meningitis (n=17); 7) toxoplasmosis (n=18); 8) neurosyphilis (n=6); 9) control group (n=57). Results: CSF LA (median; IQR) was higher in samples with TB meningitis (5.5; 2.9–7.5 mmol/L) and Cryptococcus neoformans meningitis (3.9; 2.7–5.8 mmol/L) compated with samples with HIV chronic meningitis (1.7; 1.4–1.9 mmol/L) and other groups (p≤0.0001). For the diagnosis of HIV chronic meningitis, using a cut-off of 3.5 mmol/L, CSF LA showed high sensitivity and negative predictive value, although low specificity. Conclusions: CSF LA helps to discriminate between C. neoformans or TB meningitis and HIV chronic meningitis: CSF LA can be included with the methods currently used to identify these specific pathogens, though it does not replace them. It is rapid, inexpensive and easy to perform, and can be used in developing countries.

Human cytomegalovirus detection by real-time PCR and pp65-antigen test in hematopoietic stem cell transplant recipients: a challenge in low and middle-income countries

Abstract Aim: Cytomegalovirus (CMV) infection is one of the most common complications in patients submitted to hematopoietic stem cell transplant (HSCT). Pre-emptive therapy has been indicated in patients with laboratory evidence of CMV replication. The aims of this study were to compare real-time PCR or pp65 antigen assay methodologies to detect CMV replication in HSCT patients, define a viral load threshold for initiation of pre-emptive therapy, and assess the feasibility of its implementation in hospitals of countries with low and middle income. Material and methods: Human CMV detection by real-time PCR and pp65 antigen assay was carried out in blood and plasma samples of HSCT patients collected weekly during 3 months. Pre-emptive therapy was based on CMV antigenemia results. Results: Twenty-one patients were monitored with a total of 227 samples collected; 13 (62%) patients were children. A poor correlation was observed between qualitative results, though quantitative results showed statistically significant difference, with higher viral loads detected in patients with positive antigenemia. Compared to a positive antigenemia, a cutoff value of 1067·5 copies/ml, 3·03 log10/ml, for viral load was obtained with 100% sensitivity and 71% specificity. Conclusion: CMV real-time PCR in whole blood was suitable for monitoring HSCT patients. However, its high cost is a limiting factor, and it could be used to monitor selected patients, those with prolonged leukopenia and underweight children, and subsequently switched to pp65 antigen test. Further studies involving larger numbers of patients should be performed to confirm this statement.