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Biomedical subjects

B Pattnaik

Publications and source records attributed to B Pattnaik.

At least 19 recordsLinked to original sources

The safety and efficacy of the oral rabies vaccine SAG2 in Indian stray dogs.

India is one of the countries with the highest prevalence of human rabies throughout the world. Dogs are primarily responsible for rabies transmission. Among them, stray dogs play a major role in that country. Parenteral vaccination programmes are insufficient to eliminate rabies partly due to difficulties in establishing satisfactory immunisation coverage in the dog population in view of the high proportion of stray dogs. Oral vaccination may be a useful adjunct to parenteral vaccination by increasing dog vaccination coverage. Safety, immunogenicity and efficacy of Rabidog SAG2 bait were evaluated in Indian stray dogs in captivity. Safety of SAG2 was demonstrated by the absence of adverse clinical sign, salivary excretion and absence of replication of the vaccine strain in brain and salivary glands of 21 vaccinated dogs, even when immunodepressed. Efficacy was shown 109 days post-vaccination after challenge with a highly virulent street rabies virus which killed all five controls whereas all nine vaccinated dogs survived, despite the fact that only five out of nine had seroconverted before challenge.

Administration, Oral↗

Genetic typing of bovine viral diarrhoea virus isolates from India.

Thirteen BVDV isolates collected in four geographic regions of India between 2000 and 2002 were typed in 5'-UTR. To confirm results of genetic typing, selected viruses were also analysed in the N(pro) region. Phylogenetic analysis revealed that all Indian BVDV isolates belong to BVDV-1b (Osloss-like group). Despite a long distance between the farms from which the viruses were isolated there was no correlation between the origin of viral isolates and their position in a phylogenetic tree. Higher genetic similarity of Indian BVDV isolates was observed most probably due to the uncontrolled movement of cattle as well as the uncontrolled use of semen from bulls for breeding of local and farm cattle in different states of India.

Animals↗

Serotype C foot-and-mouth disease virus isolates from India belong to a separate so far not described lineage.

Complete 1D gene sequences of 13 Indian foot-and-mouth disease virus (FMDV) type C field isolates and a vaccine strain (C-Bombay/64) were determined. All the field isolates showed a greater genetic homogeneity (95-100%) among themselves and were 19.7-21.2% divergent from the vaccine strain. In the phylogenetic analysis, the Indian field isolates formed a separate lineage (lineage VII) different from the previously identified six lineages (lineage I-VI) in type C FMDV [J. Virol. 66 (1992) 3557]. The vaccine strain was grouped with European lineage (lineage II). Comparison of the deduced amino acid sequences of antigenic sites A and C of field isolates showed no significant variation from the vaccine strain. One-way serological relationship determined in ELISA showed antigenic closeness of the field isolates with C-Bombay/64.

Amino Acid Sequence↗

Sequence analysis of the RNA polymerase gene of foot-and-mouth disease virus serotype Asia1.

The complete nucleotide (nt.) sequence of the RNA polymerase (3D) gene and 81 nt. in the 3'-untranslated region of foot-and-mouth disease virus (FMDV) serotype Asial (IND63/72) was determined and compared with the sequence of other FMDV serotypes. The 3D genomic region was 1410 nt. long encoding 470 amino acids with an inframe stop codon (TAA) at nt. position 1411-1413. The deduced amino acid sequence of the protein showed 8 conserved motifs as reported in other picornaviruses, 2 of which are 100% identical across the serotypes. Antigenic regions in the polymerase protein were predicted and found to be located at the N-terminus of the protein. The phylogenetic analysis showed that the FMD viruses were segregated into different clusters based on geographical origin; the Asia1 virus did not cluster tightly with any of the geographical groups.

Amino Acid Sequence↗

Genetic and antigenic analysis of type A foot-and-mouth disease viruses isolated in India during 1987-1996.

Twenty-three foot-and-mouth disease virus (FMDV) type A field isolates, recovered from different outbreaks during 1987-1996 in India, were subjected to antigenic and genetic analysis. The isolates showed a close antigenic relationship to the current vaccine strain (IND 17/77) in micro-neutralization test conducted using a vaccine strain (IND 17/77) antiserum and a peptide (aa 136-151 of VP1 protein of the A22/Azerbaijan/65 strain) antiserum. However, the isolates revealed minor antigenic differences in their reactivity with three neutralizing monoclonal antibodies (MAbs) recognizing trypsin-sensitive conformation-independent epitopes of the vaccine virus strains. Phylogenetic relationship between the isolates was carried out employing a part of the 1D gene (168 nucleotides at the 3'-end). Additional seven type A Indian field isolates reported earlier were included in the analysis. The percent similarity among the Indian isolates varied from 82.7% to 99.4% at nucleotide level, and from 83.9% to 100% at amino acid level. These observations clearly demonstrate genetic heterogeneity of the field isolates. The current vaccine strain IND 17/77 showed divergence of 9.7% at nucleotide level and 5.6% at amino acid level from the A22 Iraq 24/64 isolate. The field strains were divergent from the vaccine strain IND 17/77 by 5.6%-14.6% and 3.7% 13.7% at nucleotide and amino acid level, respectively. In the phylogenetic tree, the isolates were distributed into 21 genetic groups. The clustering pattern of the isolates in the phylogenetic tree revealed no specific distribution pattern of the foot-and-mouth disease (FMD) outbreaks in relation to their geographical locations, caused by unrestricted animal movement and endemic nature of the disease.

Amino Acid Sequence↗

GABAC receptors are localized with microtubule-associated protein 1B in mammalian cone photoreceptors.

Protein MAP1B was recently reported to link GABA(C) receptors to the cytoskeleton at neuronal synapses. This interaction was demonstrated in the mammalian retina, where GABA(C) receptors were thought to be exclusively expressed in bipolar cells. Our previous studies on cultured photoreceptors suggested however the presence of GABA(C) receptors in cones. To further investigate GABA(C) receptor expression in cones, we measured GABA responses in mammalian photoreceptors in situ, and we examined the distribution of the receptor and that of protein MAP1B in the mammalian outer retina. Photoreceptors were recorded from flat-mounted retinas of retinal degeneration mice at an age when the retina becomes cone-dominated after rod cell death. GABA(A) and GABA(C)-gated currents were produced only in cones but not rods. Recording freshly dissociated retinal cells from wild-type C57 mice confirmed the presence of GABA(A) and GABA(C) receptors in cones. Immunohistochemical labeling of mouse and rat retinal sections localized GABA(C) receptors to cone terminals that were identified by peanut agglutinin lectin staining. As expected from previous studies on bipolar cells, the punctate immunostaining was not restricted to cone terminals in the outer plexiform layer. MAP1B immunolabeling was obtained in rat and pig retinas and was similarly found in cone terminals identified by the peanut agglutinin lectin staining. These results provide physiological and histological evidence that cones receive a GABA feedback in the mammalian retina and are consistent with the notion that protein MAP1B links GABA(C) receptors to the cytoskeleton at postsynaptic sites.

Animals↗

Monoclonal antibodies to an Indian strain of type A foot-and-mouth disease virus.

A set of five neutralizing monoclonal antibodies (MAbs) to an Indian strain (IND17/77) of type A (subtype A22) foot-and-mouth disease (FMD) virus (FMDV) was used in the study. Four of the MAbs (27S, 37S, 85S, and 143S) identified a trypsin-sensitive (TS) epitope(s) and were specific for VP1, while the remaining MAb (145S) reacted with a trypsin-resistant (TR) epitope and was specific for VP3 in Western blot analysis. Both the epitopes (TS and TR) were conformation-independent in nature. Results obtained in MAb-competition enzyme-linked immunosorbent assay (ELISA), and profiling of the (MAb) neutralization-escape mutants in ELISA and cross-neutralization test revealed two overlapping TS epitopes (27S/37S and 85S/143S) on the virus. Variation at both these epitopes was observed in some field isolates of serotype A. Comparison of deduced amino acid sequence in the VP1 region (aa 140-213) between the parent virus and the mutants identified Gly148 and Arg153 as critical for the formation of both the TS epitopes. Substitution of R153 by Gly or Ser was observed in mutants with no reactivity for the MAbs 85S/143S. However, these mutants maintained partial reactivity with MAbs 27S/37S, and substitution of Gly148 by Glu eliminated both the epitopes. No amino acid substitution was observed in the VP1 region of aa 200-213. Efficient neutralization of the MAb neutralization escape mutants (MAb-resistant (MAR) mutants) by bovine vaccinate serum (BVS) indicated involvement of other epitopes on the virion surface in eliciting neutralizing antibodies following vaccination.

Amino Acid Substitution↗

GABAA and GABAC receptors in adult porcine cones: evidence from a photoreceptor-glia co-culture model.

1. Edge contrast enhancement is an integrated visual function based on the complex centre-surround organization of the cone photoreceptor light response. While centre responses result from direct light activation, surround responses are thought to result from lateral inhibition mediated by horizontal cells. This feedback signal has been attributed to GABAA receptors which have been found in lower vertebrate cones. 2. In order to study the GABA response of adult mammalian photoreceptors, we designed a culture system consisting of isolated photoreceptors seeded on a layer of retinal glial cells. Mature rods and cones required the presence of Muller glial cells to survive and develop neurites; they degenerated in the absence of glial cells. 3. Cone photoreceptors generated large GABA responses whereas rod photoreceptors did not respond to GABA applications. 4. Cone GABA responses consisted of two distinct components, one suppressed by the GABAA receptor blockers bicuculline and SR95531, and the second by the GABAC receptor antagonists TPMPA and imidazole-4-acetic acid (I4AA). Pentobarbital greatly increased the GABAA receptor component whereas it did not affect, or even reduced, the GABAC receptor component. During long GABA applications, GABAA receptor currents desensitized by 78%, contrasting with the sustained GABAC response. 5. Expression of GABAC receptors in cone photoreceptors was confirmed by anti-rho-subunit immunolabelling of porcine retinal sections. 6. These results indicate that both GABAA and GABAC receptors may participate in the feedback synapse from horizontal cells to cone photoreceptors in the mammalian retina.

Animals↗

Genetic heterogeneity of Indian field isolates of foot-and-mouth disease virus serotype O as revealed by partial sequencing of 1D gene.

The sequence of 165 nucleotides at the 3' end of the 1D gene, determined from RT PCR amplified cDNA fragments, of 25 type O strains isolated from different parts/regions of India during 1987 1995 and the vaccine strain (R2/75) currently in use in India were subjected to phylogenetic analysis. One isolate from the neighbouring country Nepal was also included in the study. The virus/ field strains showed high degree of genetic heterogeneity among themselves with % divergence in nucleotide sequence ranging from 1.2 to 19.4%. The Indian strains were much away (13.3 20.6%) from the exotic type O strains of O1BFS, O1K, and O1Campos. The type O strains analyzed were classified into three genotypes basing on level of divergence observed in nucleotide sequence. The type O vaccine virus (R2/75) was > 71% divergent (7.3-15.2%) from the field strains which revealed significant ( > 5%) genetic heterogeneity between the two. The phylogenetic analysis identified three distinct lineages, viz., (i) lineage 1 represented by the exotic strains, (ii) lineage 2 represented by 25 of the field strains which clustered into seven subgroups/sublines (2a-2g), and (iii) lineage 3 represented by a unique field isolate which shared the branching/origin with the vaccine strain. The lineage 2 which comprised of 25 of the 26 type O field strains analyzed, was placed almost at equidistance from the lineages 1 and 3 in the phylogenetic tree. The vaccine strain was closer to the viruses in lineage 2. Though there was no specific distribution pattern of sequences in different geographical regions of India, the viruses/ sequences in subgroup 2f appeared to be restricted to the southern states. Comparison of deduced amino acid sequence in the immunodominant regions 133-160 and 200-208 of the 1D gene product (VP1) showed that the two viruses in lineage 3 had unique amino acid residues at the positions 138 (D), 139 (G), 144 (I), and 158 (A) compared to rest of the strains including the exotic ones. Comparison of amino acid residues at critical positions 144, 148, 149, 151, 153, 154, and 208 revealed similarity between the type O strains analyzed. The virus strains showed variation (V/L/I) at position 144. One field strain showed replacement from Q149-->E and another from P208-->L. Thus, the study revealed that the type O FMD virus populations circulating in India and causing disease outbreaks are genetically much heterogeneous but related at the immunodominant region of VP1 polypeptide, and there are more than one genetically distinct virus populations in almost every region of the country which is possible due to unrestricted animal movement in the country. The involvement of vaccine virus in disease outbreaks was ruled out as the field strains (excluding the one in lineage 3) were phylogenetically distinct from it.

Amino Acid Sequence↗

Antigenic features of foot-and-mouth disease virus serotype Asia1 as revealed by monoclonal antibodies and neutralization-escape mutants.

Neutralizable antigenic sites/epitopes of serotype Asial foot-and-mouth disease virus (strain IND63/72) were identified using monoclonal antibodies (mabs) and their neutralization-escape mutants. Relative affinity/reactivity of the mabs for viral (both native and trypsin-cleaved) and subviral antigens in enzyme-linked immunosorbent assay (ELISA) showed dominance of trypsin-sensitive and conformation-dependent neutralizable antigenic sites. Characterization of neutralization escape mutants identified at least four independent trypsin-sensitive neutralizable antigenic sites on Asial FMD virus. One site was identified by mabs B3, 1A, 24, 2A, 40 and 63, second site by mabs 34 and 81, third site by mab 72 and fourth site by mab 89. The reaction profile of the mabs with selected field isolates in ELISA identified four different neutralization epitopes within the site B3/1A/24/2A/40/63.

Animals↗

Escape mutants of foot-and-mouth disease virus selected by monoclonal antibodies directed to a trypsin-sensitive neutralization epitope.

Five monoclonal antibodies (MoAbs) against Indian reference/vaccine strain of foot-and-mouth disease (FMD) virus subtype A22 (IND17/77) and a guinea pig antibody against a synthetic peptide representing amino acids (aa) 136-151 of VP1 polypeptide of A22 virus were used in the study. All the antibodies either failed to react or showed a reduced reactivity with trypsin-treated (TT)-146 S virus particles in enzyme-linked immunosorbent assay (ELISA), and could neutralize the infectivity of the reference virus. The antibodies were hence identified as specific to a trypsin-sensitive neutralizable antigenic site of the virus. Using the antibodies we isolated mutants which showed either no or reduced reactivity with the homologous as well as heterologous antibodies in ELISA. The mutants could not be neutralized with the respective antibodies but were efficiently neutralized with the serum from vaccinated cattle (BVS). These results indicated that the antibodies elicited in cattle following vaccination protected them adequately against the mutants selected and that the trypsin-sensitive neutralizable antigenic site of FMD A22 virus as identified by the MoAbs may not be dominant in eliciting a neutralizing antibody response in vaccinated cattle.

Animals↗

One-tube and one-buffer system of RT-PCR amplification of 1D gene of foot-and-mouth disease virus field isolates.

A method of reverse transcription (RT) and polymerase chain reaction (PCR) amplification of 1D (VP1) gene of foot-and-mouth disease (FMD) virus using one reaction mixture containing both avian myeloblastosis virus (AMV) reverse transcriptase (RTase) and Tfl DNA polymerase is described. The procedure was time saving, made use of a single buffer for both RT and subsequent amplification and performed better than the two-step procedure usually conducted with Moloney murine leukemia virus (MMLV) RTase and Taq DNA polymerase for amplification of the VP1 gene of field isolates of FMD virus serotypes O,-A, C and Asia 1. The failure to amplify the VP1 gene of many type O and Asia 1 viruses using MMLV RTase-Taq polymerase enzyme system could be overcome by performing RT of the viral genome at a higher temperature (48 degrees C) with AMV RTase which is not possible with MMLV RTase.

Animals↗

Evaluation of primers for PCR amplification of RNA polymerase gene sequences of foot-and-mouth disease virus.

Eight oligonucleotide primers in 7 different combinations were used to amplify 3D gene sequences of foot-and-mouth disease virus (FMDV) by reverse transcription-polymerase chain reaction (RT-PCR). Six of the primers were designed at this laboratory. All the primer combinations could specifically amplify 3D gene sequences of FMDV serotypes O, A, and C. The largest fragment amplified was of 1,393 bp and the smallest was of 208 bp in size. The 1,393 bp fragment included sequences from the preceeding P18 region of FMDV genome. The second largest fragment of 734 bp included sequences from the 3'-extracistronic region of viral genome. The remaining fragments were amplified either from the 3'- or 5'-half of the 3D gene. Specific amplification of the entire 3D gene in fragments of different size showed sequence conservation in the 3D genomic region of FMDV and usefulness of the primers reported in detecting inapparent or persistent FMDV infection in susceptible animals by RT-PCR.

Animals↗

Experimental studies on Marek's disease in Japanese quail (Coturnix coturnix japonica).

Day-old quails experimentally infected with Marek's disease (MD) virus of quail origin developed lymphoid tumors. The severity of the disease increased considerably with serial passage. Tumor transplants could be made with cells derived from gross tumors in skeletal muscles, spleen cells, and blood from MD-affected quails. After five to six serial transplants, the tumor could not be transplanted further. Marek's disease tumor-associated surface antigen (MATSA) was demonstrated in lymphoid cells of spleen and peripheral blood lymphocytes of MD-affected quails. The MATSA of quail differed from the MATSA of chicken. Chickens were susceptible to MD virus isolated and propagated in quails.

Animals↗