Journal of Clinical and Diagnostic Research, ISSN - 0973 - 709X

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Dr Bhanu K Bhakhri

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Super Speciality Paediatric Hospital and Post Graduate Teaching Institute, Noida
On Sep 2018




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Prof. Somashekhar Nimbalkar
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Chairman, Research Group, Charutar Arogya Mandal, Karamsad
National Joint Coordinator - Advanced IAP NNF NRP Program
Ex-Member, Governing Body, National Neonatology Forum, New Delhi
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On Sep 2018




Dr. Kalyani R

"Journal of Clinical and Diagnostic Research is at present a well-known Indian originated scientific journal which started with a humble beginning. I have been associated with this journal since many years. I appreciate the Editor, Dr. Hemant Jain, for his constant effort in bringing up this journal to the present status right from the scratch. The journal is multidisciplinary. It encourages in publishing the scientific articles from postgraduates and also the beginners who start their career. At the same time the journal also caters for the high quality articles from specialty and super-specialty researchers. Hence it provides a platform for the scientist and researchers to publish. The other aspect of it is, the readers get the information regarding the most recent developments in science which can be used for teaching, research, treating patients and to some extent take preventive measures against certain diseases. The journal is contributing immensely to the society at national and international level."



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Sri Devaraj Urs Academy of Higher Education and Research , Kolar, Karnataka
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Professor and Head
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Saraswati Dental College
Lucknow
On Sep 2018




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Department of Pharmacology
Calcutta National Medical College & Hospital , Kolkata




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Best regards,
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On Aug 2018




Dr. Arundhathi. S
"Journal of Clinical and Diagnostic Research (JCDR) is a reputed peer reviewed journal and is constantly involved in publishing high quality research articles related to medicine. Its been a great pleasure to be associated with this esteemed journal as a reviewer and as an author for a couple of years. The editorial board consists of many dedicated and reputed experts as its members and they are doing an appreciable work in guiding budding researchers. JCDR is doing a commendable job in scientific research by promoting excellent quality research & review articles and case reports & series. The reviewers provide appropriate suggestions that improve the quality of articles. I strongly recommend my fraternity to encourage JCDR by contributing their valuable research work in this widely accepted, user friendly journal. I hope my collaboration with JCDR will continue for a long time".



Dr. Arundhathi. S
MBBS, MD (Pathology),
Sanjay Gandhi institute of trauma and orthopedics,
Bengaluru.
On Aug 2018




Dr. Mamta Gupta,
"It gives me great pleasure to be associated with JCDR, since last 2-3 years. Since then I have authored, co-authored and reviewed about 25 articles in JCDR. I thank JCDR for giving me an opportunity to improve my own skills as an author and a reviewer.
It 's a multispecialty journal, publishing high quality articles. It gives a platform to the authors to publish their research work which can be available for everyone across the globe to read. The best thing about JCDR is that the full articles of all medical specialties are available as pdf/html for reading free of cost or without institutional subscription, which is not there for other journals. For those who have problem in writing manuscript or do statistical work, JCDR comes for their rescue.
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Reviewing articles is no less a pain staking process and requires in depth perception, knowledge about the topic for review. It requires time and concentration, yet I enjoy doing it. The JCDR website especially for the reviewers is quite user friendly. My suggestions for improving the journal is, more strict review process, so that only high quality articles are published. I find a a good number of articles in Obst. Gynae, hence, a new journal for this specialty titled JCDR-OG can be started. May be a bimonthly or quarterly publication to begin with. Only selected articles should find a place in it.
An yearly reward for the best article authored can also incentivize the authors. Though the process of finding the best article will be not be very easy. I do not know how reviewing process can be improved. If an article is being reviewed by two reviewers, then opinion of one can be communicated to the other or the final opinion of the editor can be communicated to the reviewer if requested for. This will help one’s reviewing skills.
My best wishes to Dr. Hemant Jain and all the editorial staff of JCDR for their untiring efforts to bring out this journal. I strongly recommend medical fraternity to publish their valuable research work in this esteemed journal, JCDR".



Dr. Mamta Gupta
Consultant
(Ex HOD Obs &Gynae, Hindu Rao Hospital and associated NDMC Medical College, Delhi)
Aug 2018




Dr. Rajendra Kumar Ghritlaharey

"I wish to thank Dr. Hemant Jain, Editor-in-Chief Journal of Clinical and Diagnostic Research (JCDR), for asking me to write up few words.
Writing is the representation of language in a textual medium i e; into the words and sentences on paper. Quality medical manuscript writing in particular, demands not only a high-quality research, but also requires accurate and concise communication of findings and conclusions, with adherence to particular journal guidelines. In medical field whether working in teaching, private, or in corporate institution, everyone wants to excel in his / her own field and get recognised by making manuscripts publication.


Authors are the souls of any journal, and deserve much respect. To publish a journal manuscripts are needed from authors. Authors have a great responsibility for producing facts of their work in terms of number and results truthfully and an individual honesty is expected from authors in this regards. Both ways its true "No authors-No manuscripts-No journals" and "No journals–No manuscripts–No authors". Reviewing a manuscript is also a very responsible and important task of any peer-reviewed journal and to be taken seriously. It needs knowledge on the subject, sincerity, honesty and determination. Although the process of reviewing a manuscript is a time consuming task butit is expected to give one's best remarks within the time frame of the journal.
Salient features of the JCDR: It is a biomedical, multidisciplinary (including all medical and dental specialities), e-journal, with wide scope and extensive author support. At the same time, a free text of manuscript is available in HTML and PDF format. There is fast growing authorship and readership with JCDR as this can be judged by the number of articles published in it i e; in Feb 2007 of its first issue, it contained 5 articles only, and now in its recent volume published in April 2011, it contained 67 manuscripts. This e-journal is fulfilling the commitments and objectives sincerely, (as stated by Editor-in-chief in his preface to first edition) i e; to encourage physicians through the internet, especially from the developing countries who witness a spectrum of disease and acquire a wealth of knowledge to publish their experiences to benefit the medical community in patients care. I also feel that many of us have work of substance, newer ideas, adequate clinical materials but poor in medical writing and hesitation to submit the work and need help. JCDR provides authors help in this regards.
Timely publication of journal: Publication of manuscripts and bringing out the issue in time is one of the positive aspects of JCDR and is possible with strong support team in terms of peer reviewers, proof reading, language check, computer operators, etc. This is one of the great reasons for authors to submit their work with JCDR. Another best part of JCDR is "Online first Publications" facilities available for the authors. This facility not only provides the prompt publications of the manuscripts but at the same time also early availability of the manuscripts for the readers.
Indexation and online availability: Indexation transforms the journal in some sense from its local ownership to the worldwide professional community and to the public.JCDR is indexed with Embase & EMbiology, Google Scholar, Index Copernicus, Chemical Abstracts Service, Journal seek Database, Indian Science Abstracts, to name few of them. Manuscriptspublished in JCDR are available on major search engines ie; google, yahoo, msn.
In the era of fast growing newer technologies, and in computer and internet friendly environment the manuscripts preparation, submission, review, revision, etc and all can be done and checked with a click from all corer of the world, at any time. Of course there is always a scope for improvement in every field and none is perfect. To progress, one needs to identify the areas of one's weakness and to strengthen them.
It is well said that "happy beginning is half done" and it fits perfectly with JCDR. It has grown considerably and I feel it has already grown up from its infancy to adolescence, achieving the status of standard online e-journal form Indian continent since its inception in Feb 2007. This had been made possible due to the efforts and the hard work put in it. The way the JCDR is improving with every new volume, with good quality original manuscripts, makes it a quality journal for readers. I must thank and congratulate Dr Hemant Jain, Editor-in-Chief JCDR and his team for their sincere efforts, dedication, and determination for making JCDR a fast growing journal.
Every one of us: authors, reviewers, editors, and publisher are responsible for enhancing the stature of the journal. I wish for a great success for JCDR."



Thanking you
With sincere regards
Dr. Rajendra Kumar Ghritlaharey, M.S., M. Ch., FAIS
Associate Professor,
Department of Paediatric Surgery, Gandhi Medical College & Associated
Kamla Nehru & Hamidia Hospitals Bhopal, Madhya Pradesh 462 001 (India)
E-mail: drrajendrak1@rediffmail.com
On May 11,2011




Dr. Shankar P.R.

"On looking back through my Gmail archives after being requested by the journal to write a short editorial about my experiences of publishing with the Journal of Clinical and Diagnostic Research (JCDR), I came across an e-mail from Dr. Hemant Jain, Editor, in March 2007, which introduced the new electronic journal. The main features of the journal which were outlined in the e-mail were extensive author support, cash rewards, the peer review process, and other salient features of the journal.
Over a span of over four years, we (I and my colleagues) have published around 25 articles in the journal. In this editorial, I plan to briefly discuss my experiences of publishing with JCDR and the strengths of the journal and to finally address the areas for improvement.
My experiences of publishing with JCDR: Overall, my experiences of publishing withJCDR have been positive. The best point about the journal is that it responds to queries from the author. This may seem to be simple and not too much to ask for, but unfortunately, many journals in the subcontinent and from many developing countries do not respond or they respond with a long delay to the queries from the authors 1. The reasons could be many, including lack of optimal secretarial and other support. Another problem with many journals is the slowness of the review process. Editorial processing and peer review can take anywhere between a year to two years with some journals. Also, some journals do not keep the contributors informed about the progress of the review process. Due to the long review process, the articles can lose their relevance and topicality. A major benefit with JCDR is the timeliness and promptness of its response. In Dr Jain's e-mail which was sent to me in 2007, before the introduction of the Pre-publishing system, he had stated that he had received my submission and that he would get back to me within seven days and he did!
Most of the manuscripts are published within 3 to 4 months of their submission if they are found to be suitable after the review process. JCDR is published bimonthly and the accepted articles were usually published in the next issue. Recently, due to the increased volume of the submissions, the review process has become slower and it ?? Section can take from 4 to 6 months for the articles to be reviewed. The journal has an extensive author support system and it has recently introduced a paid expedited review process. The journal also mentions the average time for processing the manuscript under different submission systems - regular submission and expedited review.
Strengths of the journal: The journal has an online first facility in which the accepted manuscripts may be published on the website before being included in a regular issue of the journal. This cuts down the time between their acceptance and the publication. The journal is indexed in many databases, though not in PubMed. The editorial board should now take steps to index the journal in PubMed. The journal has a system of notifying readers through e-mail when a new issue is released. Also, the articles are available in both the HTML and the PDF formats. I especially like the new and colorful page format of the journal. Also, the access statistics of the articles are available. The prepublication and the manuscript tracking system are also helpful for the authors.
Areas for improvement: In certain cases, I felt that the peer review process of the manuscripts was not up to international standards and that it should be strengthened. Also, the number of manuscripts in an issue is high and it may be difficult for readers to go through all of them. The journal can consider tightening of the peer review process and increasing the quality standards for the acceptance of the manuscripts. I faced occasional problems with the online manuscript submission (Pre-publishing) system, which have to be addressed.
Overall, the publishing process with JCDR has been smooth, quick and relatively hassle free and I can recommend other authors to consider the journal as an outlet for their work."



Dr. P. Ravi Shankar
KIST Medical College, P.O. Box 14142, Kathmandu, Nepal.
E-mail: ravi.dr.shankar@gmail.com
On April 2011
Anuradha

Dear team JCDR, I would like to thank you for the very professional and polite service provided by everyone at JCDR. While i have been in the field of writing and editing for sometime, this has been my first attempt in publishing a scientific paper.Thank you for hand-holding me through the process.


Dr. Anuradha
E-mail: anuradha2nittur@gmail.com
On Jan 2020

Important Notice

Original article / research
Year : 2012 | Month : April | Volume : 6 | Issue : 2 | Page : 188 - 191

Rotaviral Diarrhoea in Children: A Comparison of PAGE with ELISA

Venkatesh V.N., Prashanth H.V., K.G. Bhat, Subha D.S., Sudhindra K.S., Farheen Fathima

1. Associate Professor, Dept of Microbiology, Basaweshwara Medical College, Chitradurga, Karnataka, India. 2. Associate Professor, Dept of Microbiology, Sri Siddhartha Medical college, Agalakote, Tumkur, Karnataka, India. 3. Professor and Head of Department, Maratha Mandal Dental College, Belgaum, Karnataka, India. 4. Assistant Professor, Dept of Microbiology, Basaweshwara Medical College, Chitradurga, Karnataka, India. 5. Assistant Professor, Dept of Microbiology, Basaweshwara Medical College, Chitradurga, Karnataka, India. 6. Mrs Farheen Fathima, Assistant Professor, Dept of Microbiology, Basaweshwara Medical College, Chitradurga, Karnataka, India.

Correspondence Address :
Venkatesh V.N.
Associate Professor,
Dept of Microbiology, Basaweshwara medical college,
Chitradurga, Karnataka, India.
Phone : 07353242510
E-mail: vadnalvenk@rediffmail.com

Abstract

Introduction: Tuberculosis (TB) is a global health problem. An early diagnosis and an effective treatment are essential to prevent the spread of infection and to reduce the disease burden. Though the causative bacterium for this disease was discovered in the eighteenth century, its diagnosis in the twenty-first century is still a dogma.

Objectives: To compare of the rapid slide culture (RSC) method with the growth on Lowenstein Jensen (LJ) media and to know the sensitivity and specificity of the rapid slide culture method and the acid fast smear examination in comparison with the growth on the L J media.

Materials and Methods: One early morning sputum sample was collected from 220 clinically suspected pulmonary tuberculosis cases. All the samples were subjected to three tests: 1. Zeihl Neelsen Staining 2. Rapid slide culture and 3. Culture on Lowenstein Jensen media and the results were compared.

Results: Of the total 220 sputum samples which were tested by all the three methods, 51 samples (23.18 %) were found to be smear positive, 75 (34.09%) were found to be positive by the RSC method, 81 (36.81%) were found to be positive by the LJ culture method and 38(17.27%) samples were found to be positive by all the three methods and 93 (42.27%) samples were found to be positive by any one method. 127 (57.72%) samples were negative by all the three tests The sensitivity of RSC in comparison with the LJ culture was 88.88% with a specificity of 97.8%, whereas the smear showed a sensitivity of 49.4 % and a specificity of 92.1%.

Discussion: The RSC method is rapid, sensitive and more specific than microscopy. Hence, this method can be adopted by any simple laboratory for the diagnosis of tuberculosis.

Keywords

Rotavirus, Polyacrylamide gel electrophoresis (PAGE) technique, Enzyme - linked immunosorbent assay (ELISA), Diahorrea

Introduction
Rotavirus infects almost all children by the age of five, both in the developing and developed countries (1). Rotavirus is composed by 11 double-stranded RNA segments surrounded by three concentric protein layers. The outer capsid consists of VP7 (a glycoprotein) and VP4 (a protease-sensitive protein) which carry independent neutralization and protective antigens (2). In temperate climates, rotavirus is most often detected in the winter and rarely in the summer, whereas in the tropics it is found all year round, with less-defined seasonal variation (3). Of the approximately 600,000 annual deaths due to rotavirus (RV) worldwide, more than 150,000 occur in India. Also, 20 to 30 percent hospitalized cases of diarrhea are due to rotaviruses (4).

Clinically rotavirus gastroenteritis is characterized by profuse diarrhea, mild fever and vomiting leading to mild to severe dehydration. The clinical manifestations of rotavirus diarrhea alone are not sufficiently distinctive to permit diagnosis (1). The laboratory diagnosis of rotavirus infection is done mainly by ELISA, which require expensive commercial kits and reagents as also expensive instruments. Hence, not many laboratories are able to diagnose rotavirus infection. In view of this we undertook to evaluate the reliability of the Polyacrylamide gel electrophoresis (PAGE) technique as developed by Herring et al (5).

Material and Methods

The study included samples from 200 children belonging to both sexes who attended the pediatric clinics in civil hospital with a complaint of diarrhea. Inclusion Criteria • Children not older than 5 years of age group. • Children with profuse watery diarrhea • Diarrhoea less than 10 days duration. Exclusion Criteria • Children above 5 years. • Diarrhoea more than 10 days. Stool samples were investigated for Rota virus by PAGE (Polyacrylamide gel electrophoresis) and ELISA (Enzyme linked immunosorbant assay). ELISA was done on a 10% suspension in PBS by using ‘polyclonal’ ELISA for the group A rotavirus antigen detection. (Developed at the national institute of virology {NIV} Pune) (6). PAGE and silver staining technique were performed as per the method of herring et al (5) and Merill et al (7). Briefly a 0.5ml of 0.1 M sodium acetate solution containing 1 percent sodium dodecyl sulphate and 0.5ml phenol chloroform mixture was added to 100 mg of fecal sample. This was vortexed and centrifuged at 7000rpm for 2 minutes. The aqueous upper layer containing the double stranded RNA was removed for electrophoresis and run on gel of size 14×16cm and 0.75mm thickness with 7 wells. Ten percent polyacrylemide gels with 3 percent stacking gel were used. Each well was loaded with 40μl of RNA extract to which 10 μl of sample buffer containing 0.5 M Tris base, 1 percent bromophenol blue and 20 percent glycerol were added. The running buffer consisted of Tris glycine PH 8.8. Discontinuous electrophoresis was carried out as described by Laemlli at 30 mA for 3 hrs at room temperature (8). Finally, the double stranded RNA was visualized by silver staining. The gel was gently lifted of the glass and the stacking gel was cutoff and bottom gel was placed in washing solution consisting of 200 ml ethonol (95 %) and acetic acid (5%) and continuously rocked for 25 to 30 minutes. Next washing solution was drained of and 0.011 silver nitrate added for 50 minutes and then drained off. The gel was then briefly rinsed twice with distilled water. Developing solution (NaoH 15 gram, 3.8ml formaldehyde dissolved• Diarrhoea less than 10 days duration. Exclusion Criteria • Children above 5 years. • Diarrhoea more than 10 days. Stool samples were investigated for Rota virus by PAGE (Polyacrylamide gel electrophoresis) and ELISA (Enzyme linked immunosorbant assay). ELISA was done on a 10% suspension in PBS by using ‘polyclonal’ ELISA for the group A rotavirus antigen detection. (Developed at the national institute of virology {NIV} Pune) (6). PAGE and silver staining technique were performed as per the method of herring et al (5) and Merill et al (7). Briefly a 0.5ml of 0.1 M sodium acetate solution containing 1 percent sodium dodecyl sulphate and 0.5ml phenol chloroform mixture was added to 100 mg of fecal sample. This was vortexed and centrifuged at 7000rpm for 2 minutes. The aqueous upper layer containing the double stranded RNA was removed for electrophoresis and run on gel of size 14×16cm and 0.75mm thickness with 7 wells. Ten percent polyacrylemide gels with 3 percent stacking gel were used. Each well was loaded with 40μl of RNA extract to which 10 μl of sample buffer containing 0.5 M Tris base, 1 percent bromophenol blue and 20 percent glycerol were added. The running buffer consisted of Tris glycine PH 8.8. Discontinuous electrophoresis was carried out as described by Laemlli at 30 mA for 3 hrs at room temperature (8). Finally, the double stranded RNA was visualized by silver staining. The gel was gently lifted of the glass and the stacking gel was cutoff and bottom gel was placed in washing solution consisting of 200 ml ethonol (95 %) and acetic acid (5%) and continuously rocked for 25 to 30 minutes. Next washing solution was drained of and 0.011 silver nitrate added for 50 minutes and then drained off. The gel was then briefly rinsed twice with distilled water. Developing solution (NaoH 15 gram, 3.8ml formaldehyde dissolved Microbiology Section Venkatesh V.N., Prashanth H.V., K.G. Bhat, Subha D.S., Sudhindra K.S., Farheen Fathima www.jcdr.net Venkatesh V.N. et al., Rotaviral Diarrhoea in Children Journal of Clinical and Diagnostic Research. 2012 April, Vol-6(2): 188-191 189 in 500ml distilled water) was added for 5 to 10 minutes. This was replaced with stopping solution namely 5% acetic acid for 5 min and examined for the eleven bands. Total time for PAGE and silver staining was approximately 5 hours which included 15 min for RNA extraction, 3h for run and 2h for staining. In each run a control strain i.e., SA-11 (Simian rotavirus strain) was run which was obtained from NIV Pune. Culture of stool samples were done to know the association of common enteric pathogen with rotavirus positive cases by using standard culture techniques (9).

Results

During the study 51 out of 200 samples (25.5%) were positive for rotavirus infection by either PAGE or ELISA methods. Children belonging to the study group were in relation to their ages in months as <6 months, 6-12 m, 13- 24 m, 25-36 m, 37-48 m, and ≥49 m (Table/Fig 1). Maximum incidence of rotavirus infection was seen in age group of 6 m-24 m (32.3%), whereas age groups <6 and >24 months showed an incidence of 9.8%. The study shows a statistically significant difference (Z = 4.27, P = 0.001) in the incidence of rotavirus infection between the age groups 6-24 months and < 6 months and >24 months. The youngest patient found to be positive for rotavirus infection in this was 4 months old and the oldest was 60 months (5 years).

Out of a total of 51 children showing evidence of rotavirus infection, 34 were male and 17 were female giving a ratio of 2:1 with male predominance. Rotavirus positive samples were found throughout the study period from November to July, except in the month of July where no cases were detected. Maximum incidence of rotavirus positive samples was noted in January (28%) and February (28%). The incidence showed a declining trend between March to June i.e. from 12 % to 2 % (Table/Fig 2).

All the 200 samples were separately subjected to ELISA and PAGE. A total of 51 (25.5%) of them were found to be positive for rotavirus by either methods. 46(23%) samples were shown positive by ELISA method alone. Where as, in PAGE 50 (25%) samples were positive. Among 45 samples which were positive by both ELISA and PAGE methods, 31 showed long and 14 short electrophoretypes.

Excellent correlation of ELISA and PAGE results was found in 194 of 200 (97%) specimens. 45 (22.5%) were positive and 149(74.5%) were negative for rotavirus as shown by both the methods. Remaining 6(3%) samples showed conflicting results between ELISA and PAGE. Among these 5(2.5%) ELISA negative sampleswere clearly shown to be rotavirus positive by a single PAGE test. This demonstrates sensitivity of PAGE over the ELISA method. Only 1(0.5%) sample with positive ELISA result was shown to be PAGE negative. There was a perceptible though statistically non significant (p= 0.07) difference between the proportion of ELISA +, PAGE – samples (1/200) and ELISA–, PAGE+ (5/200). 51 samples were found positive by at least one method. 46 ELISA positive and 50 PAGE positive. Hence, relative sensitivity of PAGE and ELISA were 98% and 90 % respectively. This suggests that PAGE is more sensitive than ELISA.

A nalysis of RNA pattern: A total of 50-rotavirus positive sample by PAGE were studied for their RNA migration pattern on poly acrylamide gel. The migration patterns were classified as long and short; the ‘long’ RNA pattern recognized by faster migration of gene segments 10 and 11 and ‘short’ pattern in which there was slower migration of gene segments 10 and 11. The migration pattern of SA11 virus (Simian virus) was documented as a control strain. Control strain SA11 migrates down with 9 distinct bands. Gene segments 3, 4, 8 and 9 co migrate (10). There were 31 long electrophoretypes and 14 short electrophoretype observed in our study. Analysis of RNA pattern in 5 of the ELISA negative, but PAGE positive samples showed 4 long electrophoretypes and 1 Short electrophoretype.

A ssociated enteric pathogens: All 51 of the rotavirus positive cases did not show simultaneous infection with bacterial pathogens. Whereas in the remaining (n=149) in whom rotavirus couldn’t be demonstrated by both ELISA and PAGE method, pathogenic bacteria were isolated in 37 samples out of 149 samples with an isolation rate of 24.83%. Among the bacterial pathogens isolated, E. coli were isolated in 22 samples (59.46%), Vibrio cholera in 13 (35. 14%) and Klebsiella in 2 of the samples (5.40%). No shigella or salmonella were detected.

Discussion

During the current study 51 out of 200 samples (25.5%) were positive for rotavirus infection by either PAGE or ELISA methods. The available data highlights the importance of rotavirus as a cause of diarrhea in children, which is severe enough to deserve specialized care. The observed proportion of 25.5% of all diarrhea cases being associated with rotavirus falls within the range of values reported by workers from India. The reported positivity varies from 10.5% to 70.7% [4,11,12]. The positivity rates also vary between various settings, i.e. hospitalizations, symptomatic and asymptomatic infections and nosocomial infections (13). In this study majority of children who showed evidence of rotavirus infection belonged to the age group of 6 months to 24 months(32.3%), whereas other children <6 and >24 months accounted for only in 9.8%. Many investigators from different parts of India expressed their similar views about more prevalence of rotavirus infection occurring in the age group of 6-24 months [4,14,15,16]. It appeared that infants below 4 months of age were initially protected to some extent by maternal antibodies against severe diarrhoea due to rotavirus (4). The greater risks of infants and young children in the interim period of 6 to 12 months with declined levels of maternal antibodies to rotavirus infection have been documented (4).

Sex distribution of rotavirus positive children in our study showed a Male: female ratio of 2:1. Similar male predominance in the percentage incidence of rotavirus infection was reported by some of the authors [2,17]. Analysis of seasonal variation pertaining to rotavirus revealed that cooler months had increased rate of rotavirus associated diarrhea than the hotter months. Similar observations were made by some reports from India and other countries [4,18,19,20]. It has been observed that temperature influences the stability of human and animal rotavirus that contributes to the efficient transmission of the human rota virus (4). Moreover the influence of low relative humidity in the home has been suggested as a facilitating factor for the survival of rotaviruses on surface. This is suggestive of the indirect but important influence of meteorological factors on the complex epidemiology of human rotavirus infection (4).

In our study we did not find simultaneous infection with bacterial pathogens in rotavirus positive cases. Some of the authors [14,21] showed an association of bacterial pathogens with rotavirus positive cases. Various enteropathogens isolated in their study were E coli, Salmonella, Shigella and V. cholera and the isolation of these bacterial pathogens was higher in rota virus negative cases. This finding co relates with our study. E coli, V cholera and Klebsiella sps were the bacterial isolates in our rota virus negative cases i.e., in 37 of 149 cases.

The earliest technique used to diagnose Rotavirus infection was direct electron microscopy. The identification of virus is done based on morphology. Hence, it is 100% specific. It suffers from low sensitivity being able to detect only about 100,000,000 particles /ml (22). Immune electron microscopy (IEM) increased the sensitivity of electron microscopy by a factor of 100, detecting about 1,000,000 particles/ml. The principle disadvantages are the need for electron microscope, and very careful titration to determine the optimum ratio of antigen and antibody and prozone phenomenon (23). Isolation of rota virus has a sensitivity of about 500 infectious particles/ml (24). This level of sensitivity is reached by ELISA with much less labour.

In our study a complete concordance of ELISA and PAGE results were observed in 194 (97%) of the 200 tested specimens. This finding closely correlates with the findings of other authors who found a 96.7% to 97.14% [10,25,26] concordance results between ELISA and PAGE methods. The remaining 6 (3%) samples showed conflicting results. In a lone sample in which the O.D value of ELISA test was 0.195, this value was almost at the cutoff level, the possibility of this sample being positive by ELISA test is doubtful. Negative result of the same sample in PAGE method is difficult to explain, the possibility of presence of lot of empty virus particles or due to low concentration of viral RNA in the fecal specimen and insufficient extraction of viral RNA could be possible.

On the other hand, 5 of the samples which gave positive results by PAGE method were negative by ELISA test. These 5 samples had a typical 4-2-3-2 RNA pattern. The reason for their being ELISA negative thus remains unexplained, however blocking factors (27) or the presence of inhibitory substance (28) in stools might have been responsible. The samples containing predominantly complete particles can also give false negative results (29). Since, the group antigen is not exposed. Earlier studies [30,31] have also reported PAGE to be the most sensitive technique although some are of view that it is laborious procedure. How ever, the PAGE system used in this study was very simple to perform and the results were available on the same day. The main requirement was of trained personnel and proper standardization of the technique. Most reports states that the greatest advantage of PAGE and silver stain method are its lack of ambiguity and the fact that it provides information about viral electropherotypes. More over it generated epidemiological data regarding the circulation of strains in the community.

Conclusion

The modified PAGE system was thus found to be reliable, rapid, no expensive reagents were required and simple enough to establish in small laboratories, in which facilities and budgets are limited. Locally available reagents from HI media were used. A locally produced slab gel electrophoresis system with power pack was the only equipment required. This method could be used for the routine diagnosis of rotavirus infection in the laboratory.

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DOI and Others

DOI: JCDR/3907:2004

Financial OR OTHER COMPETING INTERESTS:
None.

Date Of Submission: Dec 05, 2011
Date Of Peer Review: Feb 07, 2012
Date Of Acceptance: Feb 20, 2012
Date Of Publishing: Apr 15, 2012

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