R K Ratho

Biomedical Informatics Centre, Chandigarh, Chandīgarh, India

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Publications (88)157.38 Total impact

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    ABSTRACT: Rubella virus outbreaks usually occur when a large numbers of susceptible individuals accumulate. The disease presents clinically with fever and maculopapular rash. The present study reports the investigation of rubella outbreak in a modern and well-planned village near Chandigarh, North India. The blood samples were collected from 39 cases with febrile rash and from 15 age and sex matched healthy controls residing in the same locality and subjected for the detection of Rubella IgM and IgG antibodies by Enzyme linked immunosorbent assay. The throat swabs, urine and blood samples from acute cases were also collected and subjected to RT-PCR using the primers targeting the E1 region. The genetic characterization of the rubella virus was carried out to identify the circulating genotypes. In the present outbreak, 13 laboratory confirmed cases were reported. Rubella IgM antibodies were detected in 12/39 (30.7%) patients. Rubella RNA could be detected in 83.3% (5/6) of urine, 22.2% (2/9) of throat swabs, and 8.3% (1/12) of blood samples. The rubella genotype responsible for the present outbreak was identified as genotype 1a. This outbreak highlights the need for the introduction of rubella vaccine in the National Immunization Programme of India to prevent outbreaks and to aim towards the eradication of this disease. This study reports the presence of genotype 1a in North India for the first time and stresses the need for further molecular work to identify the circulating strains of the virus. J. Med. Virol. © 2014 Wiley Periodicals, Inc.
    Journal of Medical Virology 08/2014; · 2.37 Impact Factor
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    ABSTRACT: Dengue is one of the most important arboviral infections caused by one of the four dengue serotypes, 1-4.
    Journal of global infectious diseases 07/2014; 6(3):109-13.
  • Journal of hepatology. Supplement / EASL 04/2014; 60(1):S299.
  • Journal of Hepatology 04/2014; 60(1):S299. · 9.86 Impact Factor
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    ABSTRACT: Human parvovirus B19 (PVB19) is linked to variety of diseases, including erythema infectiosum, transient aplastic crisis, fetal hydrops, cardiomyopathy and, recently, hepatitis and arthritis. Persistence of PVB19 in asymptomatic individuals has been reported in skin, synovium, myocardium and bone marrow. A higher level of PVB19 DNA has been observed in various tissues from cases of disease than in controls. Simultaneously, equal detection of PVB19 DNA has been shown in both cases and controls. Thus, it has become fundamental to study PVB19 DNA persistence in tissues that are unaffected by disease. This will help to better understand PVB19 DNA persistence in symptomatic and asymptomatic individuals and its possible pathogenic role in various diseases. A total of 70 adult autopsies were included and divided into seropositive (SP) and seronegative (SN) groups based on PVB19 IgG. Nested PCR for PVB19 DNA was carried out in myocardium, liver, kidney, and bone marrow. Of the 70 patients, 60 % belonged to the SP group and 40 % to the SN group. Seropositivity ranged from 50 % in the 12 to 20 year old group to 66.7 % in the 61 to 80 year old group. The viral genome was detected in 34.3 % of myocardium, 20 % of bone marrow, 10 % of kidney and 8.6 % of liver samples. There was no significant difference in the persistence rates between the SP and SN groups. The persistence of PVB19 DNA in various tissues ranged from 8.3 % to 36 % in the SP group and 10 % to 30 % in the SN group. The persistence of PVB19 DNA in all the tissues was low, and PVB19 serostatus had no influence on the persistence of PVB19 DNA.
    Archives of Virology 04/2014; · 2.28 Impact Factor
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    ABSTRACT: The conventional method of transfection of suspension cells by chemical has proven to be very difficult. We present a new transfection protocol, wherein, low-speed centrifugation of cell culture plates immediately after adding the lipid: DNA complex significantly enhances the transfection efficiency. Peripheral blood mononuclear cells (PBMCs) were transfected with BLOCK-iT™ Fluorescent Oligo (scrambled siRNA) and lipofectamine complex using conventional and low-speed centrifugation modified transfection protocols. The efficiency of transfection was determined using flowcytometer and cell viability was checked using MTT assay. Incorporation of low-speed centrifugation significantly enhances the transfection efficiency of BLOCK-iT™ in the suspension culture of PBMCs as compared to conventional transfection method (99.8% vs 28.3%; P < 0.0001), even at a low concentration of 40 picomoles without affecting the cell viability. Centrifugation enhanced transfection (CET) technique is simple, time-saving and novel application without compromising the cell viability in the context of recently popular RNA interference in suspension cultures of PBMCs. This undemanding modification might be applicable to a wide variety of cell lines and solve crucial problem of researchers working with RNA interference in suspension cultures.
    Indian journal of medical microbiology 04/2014; 32(2):164-8.
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    ABSTRACT: Background and aimsEvery year globally WHO reports 20 million Hepatitis E virus (HEV) infections. The disease occurs as sporadic cases or focused outbreaks and has potentials to cause massive epidemics. The reservoir of HEV during inter-epidemic period is not well characterized. The sporadic cases usually lack history of contact with clinically overt HEV patients. In the present context we evaluated the occurrence of subclinical HEV as a possible reservoir in endemic region.Methods Blood samples were collected from 67 apparently-healthy individuals and 10 acute viral hepatitis (AVH) patients during two HEV outbreaks in North India. The serum samples were tested for anti-HEV IgM, IgG, HEV-IgG avidity index, HEV viral-load and conventional-PCR followed by sequencing and phylogenetic analysis.ResultsA total of 14 (20.89%) apparently-healthy individuals showed the presence of anti-HEV IgM and IgG. Out of 14 based on HEV-IgG avidity index 9 (64.28%) had secondary-exposure, 4 (28.57%) had primary-exposure, while one patient had intermediate avidity. Subclinical subjects with primary-exposure had significantly higher anti-HEV IgM index as compared to secondary-exposure (p= 0.0028). Viral load in clinically jaundiced patients was significantly higher as compared to subclinical subjects (p<0.0001). Phylogenetic analysis showed HEV sequences retrieved from subclinical individuals clustered along with AVH patients, suggesting matched source. The significantly low viral-load in subclinical subjects hints towards the dose dependency for progression of clinical manifestation.Conclusion We document subclinical HEV with low level viremia occurs during outbreak settings and goes un-noticed, which helps maintaining the virus in nature possibly leading to its endemicity.This article is protected by copyright. All rights reserved.
    Liver international: official journal of the International Association for the Study of the Liver 04/2014; · 3.87 Impact Factor
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    ABSTRACT: Hepatitis E virus (HEV) is the causative agent of hepatitis E. It can be asymptomatic, associated with acute self-limiting hepatitis or acute liver failure. The conventional diagnosis of HEV infection relies on anti-HEV IgM serology. The collection of blood samples by venepunture for laboratory confirmation is often difficult during an outbreak. Thus, testing the specimens of dried blood spots (DBS) on filter papers can prove to be a feasible alternative. The present study aimed to evaluate the applicability of anti-HEV IgM detection from DBS samples and the stability of anti-HEV IgM detection at varied time interval, at various storage temperatures. Paired blood and DBS sample were collected from 44 jaundiced patients and eight healthy controls during HEV outbreaks. The DBS were tested for anti-HEV IgM by available ELISA kit with in-house modifications. Three cut offs were determined, that is, the CO1: kit cut-off, CO2: mean of negative controls above 3SD and CO3: area under Receiver operating Curve. The sensitivity of anti-HEV IgM detection ranged from 86-91%. The maximum sensitivity (91%) and specificity (100%) was obtained using CO3. Maximum stability of anti-HEV IgM antibodies (100%) was observed till 65 days at 4°C. Storage at 37°C significantly reduced anti-HEV IgM positivity, wherein 42.85% sample became negative by 45 days. DBS showed good sensitivity and specificity for detecting anti-HEV IgM and can be considered an alternate to serum sample. Moreover, anti-HEV IgM was stable at 4°C, which makes DBS a preferred method for storage and transportation of the sample to reference laboratory. J. Med. Virol. © 2013 Wiley Periodicals, Inc.
    Journal of Medical Virology 12/2013; · 2.37 Impact Factor
  • Journal of global infectious diseases 07/2013; 5(3):121-2.
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    ABSTRACT: Mumps is a vaccine-preventable disease that usually occurs as a self-limiting parotitis, but it can also lead to several life-threatening complications, including pancreatitis, meningitis, and encephalitis. The molecular epidemiology of the virus is poorly understood. The present study describes an outbreak of mumps virus infection in Punjab, India. The etiology was confirmed by serology and RNA detection to be mumps virus in 72 % of the cases and 50 % of contacts. This study, for the first time, revealed the mumps virus genotypes circulating in the Indian subcontinent as subtype G2 of genotype G.
    Archives of Virology 05/2013; · 2.28 Impact Factor
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    ABSTRACT: Hepatitis E virus (HEV) is implicated in many outbreaks of viral hepatitis in the Indian subcontinent. The conventional diagnosis of such outbreaks rests on the detection of anti-HEV IgM antibodies. However, IgM antibodies develop after 4-5 days of infection. An early-diagnostic marker is imperative for timely diagnosis of the outbreak and also initiation of control measures. This study aimed to determine the use of hepatitis E virus antigen detection as an early diagnostic marker in an outbreak in comparison to anti-HEV IgM and RT-PCR analyses. Forty samples were collected during a suspected outbreak of viral hepatitis due to HEV. A total of 36 samples were positive for one or more HEV markers. The positivity for anti-HEV IgM, HEV antigen, and RT-PCR was 91.6%, 69.4%, and 47.2% respectively. RT-PCR and HEV antigen detection gave the highest positive results (100%) in the first 3 days of illness. Positive HEV PCR declined to 54% by Days 4-7, whereas HEV antigen and IgM detection were 88% and 100%, respectively. Sequencing of representative HEV samples indicated that the strains responsible for this outbreak belonged to genotype I, subtype 1a. HEV antigen was found to be an early diagnostic marker of acute infection. HEV antigen was detected in three additional cases in the early phase (1-3 days), and they had no detectable anti-HEV IgM antibodies. These three samples were also positive for HEV RNA. After Day 7, anti-HEV IgM was the main diagnostic indicator of infection. J. Med. Virol. © 2013 Wiley Periodicals, Inc.
    Journal of Medical Virology 02/2013; · 2.37 Impact Factor
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    ABSTRACT: Hepatitis E virus (HEV) is an important cause of hepatitis in developing nations. Disease spans from asymptomatic infection to acute viral hepatitis (AVH) and acute liver failure (ALF). Cell-mediated immunity (CMI) is less studied. Studies document CMI in HEV patients using [ 3 H]-thymidine incorporation (radioactive in nature). The aim of this study was to evaluate the antigenicity of recombinant HEV ORF 2 peptide (452-617 a.a) (pORF2) by non-radioactive MTT assay and detecting the proliferation indices of primary PBMC culture. A total of 27 laboratory confirmed HEV patients (16 AVH and 11 ALF) and 20 apparently healthy individuals (HC) were included. PBMCs were isolated, plated and stimulated with pORF2. After an incubation of 4 days, cells were looked for blastogenic transformation and subjected to MTT assay. PI of AVH, ALF and healthy controls were found to be 3.249 ± 0.219, 1.748 ± 0.076 and 0.226 ± 0.017, respectively. PI of AVH Vs HC, ALF Vs HC and AVH Vs ALF were found to be significantly higher ( P < 0.0001). This study demonstrates MTT to be an adaptable technique to evaluate CMI in HEV patients. Recombinant pORF2 was found to be antigenic in nature and PBMCs from AVH patients were immunologically more reactive than ALF patients.
    Indian Journal of Medical Microbiology 01/2013; 31(1):64-8. · 0.91 Impact Factor
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    ABSTRACT: This paper introduces a new optimization technique known as Swine Influenza Model based Optimization (SIMBO). It is mimicked from Susceptible–Infectious–Recovered (SIR) models of swine flu. The development of SIMBO follows through treatment (SIMBO-T), vaccination (SIMBO-V) and quarantine (SIMBO-Q) based on probability. The SIMBO variants can be used to optimize complex multimodal functions with improved convergence and accuracy. Firstly, swine flue test based on the dynamic threshold identifies a confirmed case of swine flue. After a confirmed case of swine flue in the community, the susceptible are advised to go for the swine flue vaccination to acquire immunity. The confirmed case of swine flue is quarantined from the population. The suspected cases are treated with antiviral. The amount of antiviral drugs given to individual is dependent on patients with or without complications as well as current health of individual. In SIMBO-V and SIMBO-Q, state of the individual is updated directly through vaccination/quarantine and indirectly through treatment. The nonlinear momentum factors restrict the individuals’ treatment and state inside the defined limits without checking the health every day. SIMBO variants can easily be implemented on parallel computer architecture without having over burden or modifications. The SIMBO-T, SIMBO-V and SIMBO-Q are tested with thirteen standard benchmark functions and results are compared with other optimization techniques. The results validate that, the SIMBO variants perform comparably better. The performance of SIMBO variants are evaluated in terms of quality of optima, number of times heating stopping criteria, convergence, Fitness Evaluations (FEs), t-test, statistical parameters and analysis of variance test (ANOVA). A real time application in video motion estimation is also considered by authors to test the efficiency of the SIMBO variants. The results of motion estimation using proposed variants seems to be faster than the published methods by maintaining similar peak signal to noise ratio.
    Applied Soft Computing 01/2013; 13(1):628–653. · 2.68 Impact Factor
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    ABSTRACT: 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 RNA extraction for constitutive gene expression and pathogen detection. Sixteen archival formalin-fixed paraffin-embedded (FFPE) myocardial tissues were subjected to RNA extraction by Trizol, SDS, and RNeasy FFPE kit followed by RT-PCR and Taqman Real-Time PCR to study the expression of housekeeping genes. RNA was extracted from all 16 myocardial tissues (100%) by RNeasy FFPE kit, as compared to 14/16 by Trizol and 8/10 by SDS methods. The expression of Glyceraldehyde-3-phosphate dehydrogenase (GAPDH)was observed in RNA extracted by RNeasy FFPE kit and Trizol. High yield of RNA was obtained by RNeasy FFPE kit than Trizol (P = 0.002) and SDS(P = 0.012). Of the three methods, RNeasy FFPE kit was evaluated for Enterovirus RNA detection in 16 other histopathologically confirmed FFPE tissues of dilated cardiomyopathy (DCM) cases and Enterovirus genome was detected in 4/16 (25%) FFPE tissues of DCM cases. The enteroviral sequences of the viral isolates revealed 99% homology with Human coxsackievirus B5. The Qiagen RNeasy FFPE kit resulted in significantly high reproducibility of RNA from FFPE myocardial tissues, which are suitable for amplification by Taq-Man Real-Time and RT-PCR. Thus, the results show that these FFPE tissues can be used for gene expression, pathogen detection, and epidemiological studies.
    Journal of Clinical Laboratory Analysis 07/2012; 26(4):279-85. · 1.36 Impact Factor
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    ABSTRACT: Traditionally, rabies diagnosis is made by demonstration of rabies viral antigen by direct immunofluorescence (DIF) and mouse inoculation test (MIT). The present study was carried out to evaluate the role of reverse transcriptase-polymerase chain reaction (RT-PCR) in comparison with these conventional techniques for the diagnosis of rabies. Skin biopsies, corneal impression smears and saliva sample were collected ante-mortem and brain tissue and CSF were collected post-mortem from ten clinically suspected rabies patients. DIF, Seller staining, MIT and RT-PCR were performed on the patients' samples for the diagnosis of rabies. The ability of RT-PCR to detect rabies virus earlier as compared to other assays was tested both for reference virus as well as clinical isolates. All samples taken ante-mortem were negative for DIF test. Six of 10 post-mortem brain tissues of the clinically suspected patients were positive both by RT-PCR and MIT, of these six, five were positive by DIF test and four were positive by Seller stain. RT-PCR could detect the rabies virus earlier as compared to DIF, both from clinical isolates and fixed rabies virus. The present results showed 100 per cent sensitivity and specificity of RT-PCR as compared to 83.3 per cent of DIF and 66.7 per cent of Sellers stain for diagnosis of rabies. RT-PCR also detected rabies viral infection earlier as compared to conventional tests and can also be used on ante-mortem samples. Thus, the present study shows the usefulness of RT-PCR as an alternative to MIT for the confirmation of rabies diagnosis.
    The Indian Journal of Medical Research 06/2012; 135(6):837-42. · 2.06 Impact Factor
  • International journal of infectious diseases: IJID: official publication of the International Society for Infectious Diseases 06/2012; 16S:e48. · 2.17 Impact Factor
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    ABSTRACT: 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. To study the viral markers in patients with HPS. 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). 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.
    Indian Journal of Pathology and Microbiology 04/2012; 55(2):215-7. · 0.68 Impact Factor
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    ABSTRACT: More 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. Total 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. Following 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. TNF-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.
    BMC Infectious Diseases 01/2012; 12:23. · 3.03 Impact Factor
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    ABSTRACT: Myocarditis and dilated cardiomyopathy (DCM) are common causes of morbidity and mortality in children and adults. Recently, the human coxsackievirus and adenovirus receptor (CAR), a common receptor for coxsackieviruses and adenoviruses, was discovered and its increased expression has been reported in patients with DCM and myocarditis. To measure the expression of CAR in myocardial tissues of patients with DCM and its cellular localization in DCM cases. Formalin-fixed myocardial tissues collected during autopsy from 26 cases of DCM, and 20 cases each of noncardiac disease and cardiac disease other than DCM were included as the test group, and control groups A and B, respectively. Expression of CAR was studied using immunohistochemical staining of myocardial tissue with a CAR-specific rabbit polyclonal antibody. CAR messenger RNA was semiquantified by reverse transcription polymerase chain reaction followed by agarose gel analysis and measurement of band intensity. CAR positivity in DCM cases was found to be 96% (25 of 26) compared with 30% in control group A and 40% in control group B. CAR was found to be expressed in myocytes, endothelial and interstitial cells; however, positivity in myocytes was significantly higher than in other cells in all groups. The site of CAR expression was predominantly the sarcolemma along with cytoplasm in cardiomyocytes. The present study highlighted the increased expression of CAR in DCM cases, with localization in myocytes and endothelial cells.
    Experimental and clinical cardiology 01/2012; 17(4):183-186. · 1.10 Impact Factor
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    Indian journal of public health 01/2012; 56(2):168-9.

Publication Stats

294 Citations
157.38 Total Impact Points

Institutions

  • 2002–2014
    • Biomedical Informatics Centre
      Chandigarh, Chandīgarh, India
  • 1997–2014
    • Postgraduate Institute of Medical Education and Research
      • • Department of Virology
      • • Department of Internal Medicine
      • • Department of Parasitology
      Chandigarh, Chandīgarh, India
  • 2011
    • AIIMS Bhopal All India Institute of Medical Sciences
      Bhopal, Madhya Pradesh, India
    • Loyola University Maryland
      Baltimore, Maryland, United States