Sonal Saxena

A sindbis virus replicon-based DNA vaccine encoding the rabies virus glycoprotein elicits immune responses and complete protection in mice from lethal challenge

A sindbis virus replicon-based DNA vaccine encoding rabies virus glycoprotein (G) was developed by subcloning rabies G gene into a sindbis virus replicon-based vaccine vector (pAlpha). The self-amplification of RNA transcripts and translation efficiency of rabies G was analyzed in pAlpha-Rab-G-transfected mammalian cells using RT-PCR, SDS-PAGE and Western blot analysis. The transfected cells also showed induction of apoptosis which is an important event in the enhancement of immune responses. Further, immune responses induced with replicon-based rabies DNA vaccine (pAlpha-Rab-G) was compared with conventional rabies DNA vaccine and commercial cell culture vaccine (Rabipur) in intramuscularly injected mice. The mice immunized with replicon-based rabies DNA vaccine induced humoral and cell mediated immune responses better than conventional rabies DNA vaccine however, comparable to Rabipur vaccine. On challenge with rabies virus CVS strain, replicon-based rabies DNA vaccine conferred complete protection similar to Rabipur. These results demonstrate that replicon-based rabies DNA vaccine is effective in inducing both humoral and cellular immune responses and can be considered as effective vaccine against rabies.

Induction of immune responses and protection in mice against rabies using a self-replicating RNA vaccine encoding rabies virus glycoprotein

A self-replicating RNA vaccine encoding rabies virus glycoprotein gene was developed utilizing sindbis virus RNA replicon. The in vitro transcribed RNA (Sin-Rab-G RNA) was transfected in mammalian cells and analysed for self-replication and expression of rabies glycoprotein. To generate immune responses against rabies, mice were immunized with 10microg of Sin-Rab-G RNA and immune responses developed were compared with mice immunized with rabies DNA vaccine and commercial cell culture vaccine (Rabipur). The self-replicating rabies RNA vaccine generated cellular and humoral IgG responses similar to rabies DNA vaccine. On challenge with rabies virus CVS strain, rabies RNA vaccine conferred protection similar to rabies DNA vaccine. These results demonstrated that replicon-based self-replicating rabies RNA vaccine with 10microg dose was effective in inducing immune responses and protection similar to rabies DNA vaccine.

Risk Factor Analysis and Microbial Etiology of Surgical Site Infections following Lower Segment Caesarean Section

Background. Lower segment caesarean section (LSCS) is a common mode of delivery now and surgical site infection is the second most common infectious complication in these patients. This study was planned with this background to have a comprehensive approach to SSI following LSCS. Methods. 500 consecutive patients undergoing LSCS, irrespective of indication, were studied. A questionnaire was developed to assess the risk factors associated with development of SSI. All patients were followed up from day one of surgery till discharge and then up till the postoperative day 30 after discharge. Results. SSI was identified in 121 (24.2%) out of 500 patients. In all age groups, Gram-negative bacilli were the commonest finding. The commonest isolate was Acinetobacter species (32.03%) followed by Staphylococcus aureus and coagulase negative Staphylococcus (21.09%). 23.8% of Staphylococcus aureus strains were MRSA. By multivariate logistic regression premature rupture of membrane (PROM), antibiotics given earlier than 2 hours and increased duration of stay in the hospital were found to be significant. Conclusions. A proper assessment of risk factors that predispose to SSI and their modification may help in reduction of SSI rates. Also, frequent antimicrobial audit and qualitative research could give an insight into the current antibiotic prescription practices and the factors affecting these practices.

Isolation of Coxiella burnetii from bovines with history of reproductive disorders in India and phylogenetic inference based on the partial sequencing of IS1111 element

In the present study, a total of 158 blood samples from 148 bovines and 10 dogs having a history of reproductive disorders were screened for Coxiella burnetii by trans-PCR method. In case of bovines, 6.08% (9/148) blood samples comprised of 4.54% (4/88) cattle and 8.33% (5/60) buffaloes turned out to be positive for C. burnetii DNA while all the samples from dogs (10) were found negative. Of the 9 PCR-positive bovine blood samples, the organism could be isolated only from 3 cases of buffaloes by chick embryo inoculation method. Further, to predict the homology and genetic diversity, the recovered C. burnetii isolates designated as Y1, Y3 and Y7 were partially sequenced for IS1111 gene. On phylogenetic analysis, Y3 and Y7 isolates clustered to a common node away from Y1 isolate. This study may enlighten the nature of circulating C. burnetii isolates in different parts of the world. To the best of our knowledge, this appears to be the first report describing phylogenic analysis of C. burnetii isolates based on IS1111 gene sequence.

Risk Factor Analysis in Clinical Isolates of ESBL and MBL (Including NDM-1) Producing Escherichia coli and Klebsiella Species in a Tertiary Care Hospital

BACKGROUND Extended-spectrum β-lactamase (ESBL) and metallo-β-lactamase (MBL) producing Gram negative organisms are emerging as a worldwide public health concern. AIM To elucidate risk factors for infection with ESBL and MBL (also NDM-1) producing E. coli and Klebsiella spp. MATERIALS AND METHODS A prospective observational study was conducted from November 2010 to March 2012. ESBL production was detected using ESBL E-test, MBL by MBL E-test and NDM-1 by polymerase chain reaction (PCR). Risk factors analysed includes age, sex, clinical specimen, type of infection, duration of hospital stay prior to collection of sample, admitting ward, antimicrobial susceptibility, previous antibiotics used, co-morbid illnesses like diabetes mellitus, immunodeficiency, low birth weight, respiratory/neurological/cardiac/haematological/liver diseases, malignancy, urinary or central venous catheter, ventilatory support, surgical procedures and dialysis. STATISTICAL ANALYSIS z-test or Fishers exact test. RESULTS E. coli - ESBL producing isolates E. coli revealed female preponderance, equal incidence of hospital and community acquired infections, mostly from surgical wards, isolated from urine, age group among females >20-30 years and among males >28 days-1 year. They showed high resistance to cephalosporins, monobactam, penicillin but low resistance to carbapenems and aminoglycosides. Co-morbid conditions observed were surgery, urinary catheterisation, haematological disease, ventilatory support, diabetes mellitus and neurological disease. MBL producing strains were mainly from females, surgical wards, (including both NDM-1 isolates), hospital acquired infections, isolated from body fluids (NDM-1 positive), female genital tract specimen and urine (one NDM-1 positive). NDM-1 positive isolates belonged to age groups >5-10 year and >0-28 days and underwent surgery and urinary catheterisation. Klebsiella spp.- ESBL producing isolates showed female preponderance, hospital acquired infections, from surgical wards, high resistance levels to cephalosporins, fluoroquinolones, monobactam, but low levels to carbapenems, among males isolated from pus in age group >0-28 days and >28 days -1 year and among females from urine in >20-30 years, no significant difference when correlated with risk factors. MBL (NDM-1) producing isolates were mainly from females with age range 0 days to 70 years, mainly admitted to ICU/postoperative wards with urinary catheter in-situ, ventilatory support, surgery, diabetes mellitus, haematological and neurological disease. CONCLUSION Risk factors for infections due to ESBL and MBL producing Gram Negative Bacteria (GNB) should be clearly identified to reduce their spread and to optimise antibiotic use.

Multiplexed Autoantibody Signature for Serological Detection of Canine Mammary Tumours

Spontaneously occurring canine mammary tumours (CMTs) are the most common neoplasms of female unspayed dogs and are of potential importance as models for human breast cancer as well. Mortality rates are thrice higher in dogs as compared to humans with breast cancer, which can partly be attributed to lack of diagnostic techniques for their early detection. Human breast cancer studies reveal role of autoantibodies in early cancer diagnosis and also the usefulness of autoantibody panels in increasing the sensitivity, as well as, specificity of diagnostic assays. Therefore, in this study, we took advantage of high-throughput Luminex technique for developing a multiplex assay to detect autoantibody signatures against 5 canine mammary tumour-associated autoantigens (TAAs). These TAAs were expressed separately as fusion proteins with halo tag at the N-terminus, which allows easy and specific covalent coupling with magnetic microspheres. The multiplex assay, comprising a panel of candidate autoantigens (TPI, PGAM1, MNSOD, CMYC & MUC1) was used for screening circulating autoantibodies in 125 dog sera samples, including 75 mammary tumour sera and 50 healthy dog sera. The area under curve (AUC) of the combined panel of biomarkers is 0.931 (p < 0.0001), which validates the discriminative potential of the panel in differentiating tumour patients from healthy controls. The assay could be conducted in 3hrs using only 1ul of serum sample and could detect clinical cases of canine mammary tumour with sensitivity and specificity of 78.6% and 90%, respectively. In this study, we report for the first time a multiplexed assay for detection of autoantibodies in canine tumours, utilizing luminex technology and halo-tag coupling strategy. Further to the best of our knowledge, autoantibodies to CMYC and MUC1 have been reported for the first time in canines in this study.

HYPERSPECTRAL IMAGING (HSI): APPLICATIONS IN ANIMAL AND DAIRY SECTOR

Hyperspectral imaging (HSI), also known as imaging spectroscopy or 3D spectroscopy, combines imaging and spectroscopy into a single system. With a high resolution measurement of spectral signatures, HSI is able to provide critical information of the target. Thus it is useful for various scientific and industrial applications, including food safety and disease diagnosis. Due to constantly increasing demands for safe animal products, there is pressure on the processing sector for applications of advanced, high throughput methods for non-destructive quality analysis of animal products. In this context, HSI finds its applications for grading, classification, quality & composition analysis of animal products including meat, egg, milk etc. Further, the technique is also a useful tool in poultry sector for assessment of wholesomeness and quality control of chicken carcasses, as well as, chicken meat products. In fish industry also, the technique has established its potential for determining freshness and quality attributes of sea-foods. Apart from quality control of animal products, HSI has also demonstrated its usefulness for disease diagnosis in animal models and for detection of mammary cancers in dogs. Thus, the future of HSI technology in animal industry is promising and associated with multivariate analysis, HSI technique will further dominate in animal products authentication and analysis in the future also.