Baijayantimala Mishra

An update on crimean congo hemorrhagic Fever.

Crimean Congo hemorrhagic fever (CCHF) is one of the deadly hemorrhagic fevers that are endemic in Africa, Asia, Eastern Europe, and the Middle East. It is a tick-borne zoonotic viral disease caused by CCHF virus of genus Nairovirus (family Bunyaviridae). CCHF not only forms an important public health threat but has a significant effect on the healthcare personnel, especially in resource-poor countries. India was always a potentially endemic area until an outbreak hit parts of Gujarat, taking four lives including the treating medical team. The current review is an attempt to summarize the updated knowledge on the disease particularly in modern era, with special emphasis on nosocomial infections. The knowledge about the disease may help answer certain questions regarding entry of virus in India and future threat to community.

Usefulness of RT-PCR for the diagnosis of Japanese encephalitis in clinical samples

The present study was carried out between July 2003 and December 2005 in PGIMER, Chandigarh, India and aimed to compare IgM capture ELISA and nested RT-PCR for the diagnosis of Japanese encephalitis (JE). The samples collected were cerebrospinal fluid and blood from 40 febrile patients with encephalitis (n=40, group I) and blood samples from febrile patients without encephalitis residing in JE endemic areas (n=45, group II). Overall, in CSF samples JE specific RNA was detected in 9/40 (22.5%), while 7/28 (25%) patients showed the presence of specific IgM antibodies. Only 28 CSF samples could be subjected to both RT-PCR and IgM and, among these, 13 cases were found to be confirmed JE based on IgM and/or RT-PCR positivity. Among the confirmed cases, 6 (6/13, 46.5%) could be detected by RT-PCR alone, 4 (4/13, 30.7%) by IgM capture ELISA and 3 (3/13, 23.1%) patients were positive by both the methods. All the RT-PCR positive cases had presented within 5 d of onset of illness. The serum samples of only 16 patients in group I could be tested for IgM antibodies and 5 (31.25%) were found to be positive, while in group II, 11.1% (5/45) positivity was observed. JE specific RNA could not be detected in serum samples of either group of patients. This study highlights the need for carrying out RT-PCR in CSF samples, compared to IgM antibody detection, for the early detection of JEV.

NS1 antigen as an early diagnostic marker in dengue: report from India

Detection of specific IgM antibodies by ELISA forms the mainstay for diagnosis of dengue infection. However, IgM antibodies develop after 4 to 5 days of infection. The methods for early diagnosis include virus isolation and reverse transcriptase polymerase chain reaction (RT-PCR) which need a sophisticated laboratory. Another alternative that has recently come up is NS1 antigen detection. The present study compared IgM antibody detection with NS1 antigen for the diagnosis of acute dengue in 87 samples. NS1 antigen could be detected with good sensitivity (71-100%) till day 3 of fever, whereas IgM had a sensitivity of 0% to 50% at this time. On day 4 of illness, both the tests had comparative sensitivity. Beyond day 4, IgM antibody detection was superior to NS1. Both these diagnostic modalities were also compared with RT-PCR in 40 acute samples. NS1 detected additional 15 samples, which were missed by PCR. NS1 antigen is an early diagnostic marker that is feasible in a routine diagnostic laboratory.

Pathology and virology findings in cases of fatal influenza A H1N1 virus infection in 2009-2010.

Bal A, Suri V, Mishra B, Bhalla A, Agarwal R, Abrol A, Ratho R K & Joshi K 
(2012) Histopathology 60, 326–335 
Pathology and virology findings in cases of fatal influenza A H1N1 virus infection in 2009–2010

TNF-α promoter polymorphism: a factor contributing to the different immunological and clinical phenotypes in Japanese encephalitis

BackgroundMore than three billion populations are living under the threat of Japanese encephalitis in South East Asian (SEA) countries including India. The pathogenesis of this disease is not clearly understood and is probably attributed to genomic variations in viral strains as well as the host genetic makeup. The present study is to determine the role of polymorphism of TNF-alpha promoter regions at positions -238G/A, -308G/A, -857C/T and -863C/A in the severity of Japanese encephalitis patients.MethodsTotal of 142 patients including 66 encephalitis case (IgM/RT-PCR positive), 16 fever cases (IgM positive) without encephalitis and 60 apparently healthy individuals (IgG positive) were included in the study. Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) using site specific restriction enzymes were implemented for polymorphism study of TNF alpha promoter.ResultsFollowing the analysis of the digestion patterns of four polymorphic sites of the TNF- alpha promoter region, a significant association was observed between the allele -308A and -863C with the patients of Japanese encephalitis.ConclusionsTNF- alpha 308 G/A has been shown to be associated with elevated TNF- alpha transcriptional activity. On the other hand, polymorphism at position -863C/A in the promoter region has been reported to be associated with reduced TNF- alpha promoter activity and lower plasma TNF levels. As per the literature search, this is the first study to identify the role of TNF- alpha promoter in JE infection. Our results show that subjects with - 308A and -863C alleles are more vulnerable to the severe form of JE infection.

A novel mutation (S227T) in domain II of the envelope gene of Japanese encephalitis virus circulating in North India

Japanese encephalitis (JE) is an important arboviral infection of public health concern. There is a significant variation in mortality (10-30%) in JE viral infection. Epidemics of JE have become regular features in the northern states of India. The recent resurgence of the A226V mutation leading to a widespread Chikungunya epidemic motivated the investigators to search for any such mutational occurrence with Japanese encephalitis virus (JEV) isolated from this region. This study looked for mutation of clinical strains at amino-acid positions 176, 177, 227, 244, 264 and 279. A novel mutation S227T was detected corresponding to the loop region of domain II, E gene of JEV in comparison to Indian and other isolates from different parts of the world. Genotype III was found to be circulating in this geographical area. Further studies are required to ascertain its role in JE pathogenesis and vector competency.

Utility of multiplex reverse transcriptase-polymerase chain reaction for diagnosis and serotypic characterization of dengue and chikungunya viruses in clinical samples ☆

The reemergence of chikungunya virus (CHIKV) has compounded the already existing dengue problem because of clinical similarities and common vector, demanding the need for a rapid and specific diagnosis. Thus, dengue chikungunya multiplex reverse transcriptase-polymerase chain reaction (DCmRT-PCR) was developed and validated for simultaneous detection of dengue and chikungunya viral infections and its utility in virus serotyping. Blood samples from 97 suspected dengue and chikungunya cases and 10 healthy controls were subjected to dengue and chikungunya conventional RT-PCR and DCmRT-PCR. Thirty-one of 97 samples were positive for dengue or chikungunya viral RNA by RT-PCR and DCmRT-PCR with 100% concordance. DCmRT-PCR products were cycle sequenced. Seven dengue virus strains were clustered within genotype III of DENV-3 and 4 within genotype III of DENV-1, whereas chikungunya sequences were clustered within the Central/East African genotype. DCmRT-PCR was found to be a potential rapid test for simultaneous detection of dengue and CHIKV in clinical samples along with dengue serotyping.

Congenital rubella and cytomegalovirus infections in and around Chandigarh

AIMS This study has analyzed the role of rubella and cytomegalovirus (CMV) in infections of children and pregnant women. SETTINGS AND DESIGN The study was carried out in a tertiary care hospital. Data from blood samples from pregnant women (asymptomatic and also women with obstetric problems) and children (suspected of intrauterine infections) that were received in the laboratory over a period of 8 years were analysed. MATERIALS AND METHODS The samples were tested for rubella- and CMV-specific IgM antibodies by capture enzyme linked immunosorbent assay. RESULTS In children, the overall positivity for rubella- and CMV-specific IgM antibodies was 2.8% and 12.5%, respectively. In asymptomatic pregnant females, rubella positivity was 0.7% while in women with obstetric complications it was 3.4%. IgM antibody positivity in cases of CMV was 7.8% in both asymptomatic pregnant women and also in women with obstetric complications. CONCLUSIONS The study indicated that infection with CMV is more common than the rubella virus. The incidence of rubella has reduced over the past few years. Hence, screening for rubella infection may be reserved for women with obstetric complications only. The routine screening for CMV among all antenatal cases is a debatable issue.

Viral markers in patients with hemophagocytosis: a prospective study in a tertiary care hospital.

BACKGROUND Hemophagocytic syndrome (HPS) is a rare clinicopathological condition characterized by the activation of macrophages with prominent hemophagocytosis in bone marrow and other reticulo-endothelial systems. HPS can be familial or secondary to infections including viruses. AIM To study the viral markers in patients with HPS. MATERIALS AND METHODS Serum samples of patients with HPS and control group were screened for anti EBV VCA IgM, and IgG, anti-Parvo B19 IgM, and anti-CMV IgM antibodies using commercially available ELISA kits and CMV and ParvoB19 DNA by polymerase chain reaction (PCR). RESULTS AND DISCUSSION The present prospective study reports the profile of viral markers in HPS cases from north India. Among the 14 HPS cases 43% (6/14) were positive for at least one viral marker tested, of which EBV was found to be the most prevalent (3/6: 50%) followed by parvovirus B19(2/6: 33%) and cytomegalovirus (1/6: 17%). Mortality was noted in 33% of virus associated HPS patients. Our study highlights the higher association of Epstein-Barr virus (EBV) with HPS as compared to other viruses along with higher rate of mortality in both parvovirus B 19 and EBV associated HPS.

Ribonucleic acid extraction from archival formalin fixed paraffin embedded myocardial tissues for gene expression and pathogen detection.

Archival tissue samples preserved in formalin are a great source of treasure for biomedical research and diagnostics. Formalin, though is a good preservative, causes the modification of nucleic acid limiting the application of fixed tissues. The present study evaluated three methods of RNAextraction for constitutive gene expression and pathogen detection.