Biomedical subjects
Publications and source records attributed to C Natarajan.
This is a case report of a 71-year-old man who presented with an incidental finding of a mass in a solitary right testis. A scrotal ultrasound scan showed the typical features of a testicular epidermoid cyst. Tumour markers were not elevated and the patient was managed non-operatively. On reviewing the literature, we found no previous report on the non-operative management of testicular epidermoid cysts
The epidemiological role of small ruminants in foot-and-mouth disease (FMD) outbreaks has been generally neglected. Although, the disease in these species is sub-clinical in nature, their role as virus carriers represents a reservoir for further infection and spread of disease. Data on the usefulness of polyvalent FMD vaccine (FMDV) in goats is scant. Thus, the present study was undertaken to evaluate the benefits of a highly potent polyvalent FMDV in goats. In the present investigations, FMDV quadrivalent double oil emulsion (Montanide ISA 206) vaccines were tested in goats at reduced doses of 2 ml per animal (antigen payload 3.5 microg per serotype per dose). The oil adjuvant elicited superior immune response at any given period than aluminium hydroxide gel (AGS) vaccine and the rapidity of development of response was quicker. The duration of immunity also appeared to be maintained for long period. The differences in immune response between two adjuvant groups were statistically significant (P<0.05). The differences were apparent even in kinetics of immune response. Unlike cattle, goats were found to be late responders for oil-adjuvanted vaccine. Our results indicate possible universal usage of double oil emulsion vaccines for disease control programs irrespective of species of animals.
Foot-and-mouth disease (FMD) is an economically important disease of cloven-hoofed animals. The multiplicity of FMDV serotypes in animals poses a central problem in the policy of vaccination and is of much concern to health authorities. Hence it is the practice of vaccination with polyvalent vaccine for prophylactic measure. In the present report, we analysed the early antibody responses elicited by FMDV quadrivalent (FMDV O, A, C and Asia 1 serotypes) double emulsion (Montanide ISA 206) vaccines in cattle. We observed variations between various viral serotypes in eliciting early antibody response although neutralizing antibody response against all the four serotypes were detected as early as fourth day following vaccination. The duration of immunity also appeared to maintain for long period. The neutralizing antibody titres were maintained well above 2log(10) even after 6 months of vaccination irrespective of serotypes. Thus, allows the possibilities of two vaccinations per year for the maintenance of herd immunity.
Vaccine against foot-and-mouth disease (FMD) is prepared after inactivating the virus produced in cell culture. Inactivation of the FMD virus (FMDV) was earlier done by formaline. However, several vaccine outbreaks, which occurred in Europe revealed that the formaline treatment is not highly effective for virus inactivation. Subsequently, binary ethyleneimine (BEI) was identified as an effective inactivation reagent for FMDV. However, these chemical reagents are likely to have effect on whole virus particle whose integrity is essential for vaccine potency. Therefore, a need is felt to develop non-chemical methods. We have studied induction of endonucleolytic activity as an alternative method for virus inactivation. This method of inactivation was compared with the chemical methods, and found to be highly effective for virus inactivation. The effects of endonucleolytic activity on the integrity of virus capsid was studied using antibodies raised against recombinant proteins, which elicited antibodies against major epitopes present on the surface of the virus. Further, the effect of the agents on the integrity of the virus capsid was studied by using antigen capture PCR (Ag-RT/PCR) which detects the whole virus. The studies showed that inactivation of the virus by induction of endonucleolytic activity is more effective besides maintaining virus integrity. The effect of various inactivating agents on four serotypes of FMDV has also been studied and found to have varying effects, depending on serotype.
Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a nuclear receptor transcription factor that regulates adipocyte differentiation and glucose homeostasis. PPARgamma agonists are potent therapeutic agents for the treatment of type 2 diabetes and obesity. PPARgamma agonists also prevent inflammation in animal models, suggesting their use for the treatment of human inflammatory diseases. Experimental allergic encephalomyelitis (EAE) is a Th1 cell-mediated inflammatory demyelinating disease model of multiple sclerosis (MS) and IL-12 plays a crucial role in the pathogenesis of EAE and MS. In this study we have examined the effect of PPARgamma agonists on the pathogenesis of EAE. In vivo treatment of SJL/J mice with PPARgamma agonists, 15-deoxydelta(12,14) prostaglandin J2 or Ciglitazone, decreased the duration and clinical severity of active immunization and adoptive transfer models of EAE. PPARgamma agonists inhibited EAE in association with a decrease in IL-12 production and differentiation of neural antigen-specific Th1 cells. In vitro treatment of activated T cells with PPARgamma agonists inhibited IL-12-induced activation of JAK-STAT signaling pathway and Th1 differentiation. These findings highlight the fact that PPARgamma agonists regulate central nervous system inflammation and demyelination by inhibiting IL-12 production, IL-12 signaling and Th1 differentiation in EAE.
Despite representing the majority of the world's foot-and-mouth disease (FMD)-susceptible livestock, sheep and goats have generally been neglected with regard to their epidemiological role in the spread of FMD. In the present investigations, FMD virus quadrivalent double emulsion (Montanide ISA 206) vaccines were tested in sheep. The oil adjuvant elicited a better immune response at any time than did aluminum hydroxide gel vaccine, and the response developed quicker. The animals maintained their neutralizing antibody titers at >3 log(10) for the duration of the trial (90 days). Sheep were found to be late responders to serotypes A, C, and Asia-1; a clear upward shift in titer was observed at 60 days postvaccination. However, development of the immune response to serotype O in sheep was superior to that in cattle and goats.
An immunobiosensor using a piezo electric (PZ) crystal was developed and standardized for foot and mouth disease (FMD) diagnosis and virus typing. A 6MHz quartz crystal was used as the frequency determining element. Foot and mouth disease virus (FMDV) type specific antibody raised in rabbits/monoclonal antibody was coated on the crystal surface and the resonance measured. One microlitre of the 10% aqueous suspension of the clinical sample (tongue or foot epithelium) was applied on both surfaces of the crystal and the resonance recorded. A difference in resonance of more than -2.5Hz was obtained in positive samples (homologous antigen and antibody). The test was standardized initially using various dilutions of FMD tissue culture antigen. Repeatability and sensitivity were also tested and it was found that the crystals could be washed and reused eight times. The test could be used for FMDV type specifically and no cross-reaction between FMDV types was observed. The shelf-life of the antibody-coated crystal stored at room temperature was 18 weeks. Application of the biosensor test to the FMDV clinical samples confirmed virus typing results when compared with enzyme-linked immunoabsorbent assay (ELISA) and it could also detect virus in ELISA negative samples and mixed virus infections.
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In a health care environment dominated by the growth of managed care organizations, generic competition, therapeutic substitution and drug utilization review, drug development is an extremely risky proposition. Consequently, it is imperative to incorporate a mechanistic approach to drug development that combines a thorough understanding of a drug at the molecular/cellular level with a rigorous preclinical, and clinical pharmacology program. This should enable the sponsor to evaluate multiple hypotheses during the early "learning" phases of clinical development (Phases I and IIA) and to eliminate nonpromising candidates early on while drug development costs are low. Clinical research done properly in the early stages of drug development will also set the stage for designing and conducting optimal "confirming" registrational Phase IIB/III studies for promising drug candidates. Pharmacokinetics (PK) and pharmacodynamics (PD) modeling and simulation are crucial components of a mechanistic approach to optimal drug development and their application has significant impact in both early and late development efforts. This communication describes several applications of pharmacokinetics and pharmacodynamics modeling and simulation that were important in guiding, optimizing and ensuring the success of development efforts for drug candidates in the therapeutic areas of cardiology and oncology. These examples are used to illustrate and discuss the use of the current state-of-the-art in pharmacokinetics and pharmacodynamics modeling and simulation at numerous stages in the development cycle and to postulate on future directions in this area.
Truncated proteins corresponding to the C-terminal half of VP1 of four vaccine strains and two field variants of foot-and-mouth disease virus (FMDV) were expressed in E. coli. The expressed proteins were affinity purified and their type specific reactivity was confirmed by immunoprecipitation with anti-virus antibodies. Antibodies were raised against the purified proteins in guinea pigs and the type specificity of the anti peptide antibodies was confirmed by antigen capture reverse transcription polymerase chain reaction (Ag-RT/PCR) where the sera against a particular type captured the homologous virus. Antibodies were purified by immuno-affinity chromatography and tested for specificity by various serological tests. Using the purified proteins and the antibodies raised against them, tests like ELISA, Ag-RT/PCR, and latex agglutination test (LAT) were standardized. Application of the reagents in various tests was studied by screening a few field samples and by nucleotide sequencing. Specific reactivity of antibodies raised against expressed protein was seen with both vaccine virus and field samples. Thus E. coli expressed proteins and antibodies to them may form an alternative and cheap source of diagnostic reagents. The studies showed that antibodies against peptides were mono-specific and therefore may be used in LAT for rapid typing of FMDV and Ag-RT/PCR for typing ELISA negative field samples.
The technique of capturing of foot-and-mouth disease virus (FMDV) from clinical material in microcentrifuge tubes coated with type-specific antibodies and amplifying the viral sequences by RT/PCR in the same tube, promoted the detection and serotyping of FMDV with high sensitivity and specificity. The efficiency of antigen capturing and shelf life of the coated tubes was improved by glutaraldehyde fixation of antibodies to the tubes. Virus in infected tissues, even after storage for 25-30 years at 70 degrees C, could be successfully typed by this method. Conserved sequences flanking the variable region of immunoreactive VP1 gene of FMDV were used as primers in the assay and hence the nucleotide sequence analysis of the product could reveal the strain variation. The test has been found to be at least 125-fold more sensitive than type specific ELISA and of comparable sensitivity as other protocols for detection of FMDV by RT/PCR.
Variations in the amino acid sequence of Foot-and-Mouth Disease Virus (FMDV) structural proteins are the basis for the antigenic diversity of the virus. Majority of antigenic sites for the virus neutralization are present on VP1, the major immunogenic protein. However, a few conformational epitopes are present on the structural proteins VP2 and VP3. The nucleotide sequence encoding all the four structural proteins (P1 region) of FMDV type Asia 1 Ind 63/72 was determined. The nucleotide and the deduced amino acid sequence of P1 of Asia 1 of Indian strain was compared with that of Asia 1 Israel strain. Differences were observed at 284 (14%) nucleotide positions resulting in 69 (10%) amino acid changes. The variation in the derived amino acid sequence is the highest in VP1 (14.4%) followed by VP2 (10%), VP3 (6.4%) and VP4 (3%). Deletion of two amino acids, which was observed in the case of Indian strain as well as in Israel strain indicated that these deletions are specific for type Asia 1. The P1 sequence was also compared with the corresponding region of the other serotypes O1K, A12, Cl and SAT-1. The sequence has been submitted to EMBL data bank, under accession number Y09949.
Two outbreaks of foot-and-mouth disease (FMD) in vaccinated cattle were investigated wherein a mixed infection due to FMD virus (FMDV) types O and Asia 1 was detected by sandwich enzyme-linked immunosorbent assay (ELISA) and confirmed by antigen capture polymerase chain reaction (PCR). The clinical picture and the epidemiological data on these outbreaks are presented. The isolated virus strains were compared to the respective vaccine strains by means of monoclonal antibody (MAb) profiling and nucleotide sequence analysis. The probable cause of the mixed FMDV infection and its significance in disease control are discussed.
Vaccination is the only pragmatic approach to control foot and mouth disease in India. Strict quality control measures are essential to supply potent vaccine to the field application, in addition to monitoring the performance of the vaccine in the field. During the process of monitoring, an outbreak of FMD in vaccinated animals caused by type "O" virus in Tanjavur district of Tamil Nadu and a type "O" virus from unvaccinated herd of Karnataka were studied. Field isolates and vaccine virus were sequenced and analyzed. Data indicated that the virus from the outbreak in vaccinated cattle was a variant which could escape neutralization by antibodies against vaccine virus.
Five fusion experiments were conducted with spleen cells from Balb/c mice immunized with purified 146S antigen of foot and mouth disease virus type 'C' (vaccine strain). Monoclones (31) thus developed were isotyped as IgM (3), IgG1 (6), IgG2a (5), IgG2b (3) and IgG3 (14). Eleven clones isotyped as IgM, IgG2a and IgG2b showed neutralizing activity in virus neutralization and plaque reduction tests. Six of the neutralizing clones precipitated 146S virus in Ouchterlony reaction. On the basis of location of MAb reactive epitopes in relation to intact virus (146S), 12S particles and VP1 in ELISA test, the clones were classified as Class II (6), Class III (11) and Class IV (14). These clones may be useful for purposes of antigen detection from field isolates and for estimation of antibody titres in vaccinated animals.
The Indian cattle is known to be more tolerant to tropical infections than the European cattle. In order to verify the genetic basis of this variation, the DR B exon-2 of the major histocompatibility locus, known for coding the antigen recognition site, from the Hallikar breed of Indian cattle was amplified by PCR, cloned and sequenced. Comparison of this sequence with the information available on taurus cattle brought out six unique nucleotide changes and three amino acid changes. The amino acid positions were at 17, 72 and 87. A major variable region was observed at amino acid position 85 to 87 from all the alleles so far reported for the bovine locus.
The antigenic variation in Foot and Mouth Disease Virus (FMDV) is very high. The effective strategy to control the Foot and Mouth Disease (FMD) in India which is a habitat of four serotypes O, A, C and Asia 1, is by regular vaccination, using the vaccine strain most suitable for the local situation. India is an endemic country with the disease being widely distributed. Selection of vaccine strain should therefore need the information on the circulating viruses. Asia 1 causes the second largest number of disease outbreaks in India. As there is no information available with respect to the extent of antigenic variation in FMDV type Asia 1, we have studied FMDV isolates from vaccinated and unvaccinated animals from different parts of the country and compared their relationship with Asia 1 vaccine virus. The immunogenic, hypervariable region of viral protein 1 (VP1) gene was amplified by RT-PCR and sequenced. Analysis of sequence data showed that the viruses from two field outbreaks of Southern India were closely related to each other when compared to the isolate from the North and all the three isolates are away from the vaccine virus.
A pair of oligomers of 20 and 23 bp were designed for amplifying a 381 bp sequence from glycoprotein IV gene of bovine herpesvirus 1. The primer pairs were used for amplifying genomic DNA of BHV-1 directly from cell culture fluids under different experimental conditions such as, untreated cell culture fluid, thermal denaturation and proteinase K treatment in presence of detergent. The results reveal that direct thermal denaturation of cell culture fluid is sufficient to detect the virus by polymerase chain reaction.