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Sixty billion chickens are produced worldwide each year, and all are at risk from Eimeria , parasites that cause coccidiosis. Control relies widely on chemoprophylaxis, but pressure to reduce drug use in farming urges development of cost-effective vaccines. Antigens such as apical membrane antigen 1 (AMA1) offer promise as anticoccidial vaccine candidates, but experience with related apicomplexans such as Plasmodium , in which pre-existing antigenic diversity and incompatible population structure have undermined vaccine development, tempers confidence. Parasite genotyping identified enormous region-specific variation in haplotype diversity for Eimeria tenella but a contrastingly low level of polymorphism for Et AMA1. Although high levels of polyclonal Eimeria infection and hybridization indicate an ability to disseminate vaccine resistance rapidly, the low level of Et AMA1 diversity promotes vaccine development. The phylum Apicomplexa includes serious pathogens of humans and animals. Understanding the distribution and population structure of these protozoan parasites is of fundamental importance to explain disease epidemiology and develop sustainable controls. Predicting the likely efficacy and longevity of subunit vaccines in field populations relies on knowledge of relevant preexisting antigenic diversity, population structure, the likelihood of coinfection by genetically distinct strains, and the efficiency of cross-fertilization. All four of these factors have been investigated for Plasmodium species parasites, revealing both clonal and panmictic population structures with exceptional polymorphism associated with immunoprotective antigens such as apical membrane antigen 1 (AMA1). For the coccidian Toxoplasma gondii only genomic diversity and population structure have been defined in depth so far; for the closely related Eimeria species, all four variables are currently unknown. Using Eimeria tenella , a major cause of the enteric disease coccidiosis, which exerts a profound effect on chicken productivity and welfare, we determined population structure, genotype distribution, and likelihood of cross-fertilization during coinfection and also investigated the extent of naturally occurring antigenic diversity for the E . tenella AMA1 homolog. Using genome-wide Sequenom SNP-based haplotyping, targeted sequencing, and single-cell genotyping, we show that in this coccidian the functionality of Et AMA1 appears to outweigh immune evasion. This result is in direct contrast to the situation in Plasmodium and most likely is underpinned by the biology of the direct and acute coccidian life cycle in the definitive host.

Background Porcine circovirus 2 is globally noted swine pathogen with multiple genotypes associated with vast clinical and subclinical outcomes. This study aimed to isolate and characterize PCV2 genotypes circulating in southern states of India. Methods and results A total of 434 field samples comprising of serum (n = 273), tissues (n = 109) and swabs (n = 52) collected from swine during 2019 to 2021 from southern states of India were screened for PCV2 by specific polymerase chain reaction (PCR) assay. Molecular prevalence of PCV2 in southern India was found to be 12.21% (n = 53). All the 53 PCV2 positive samples were further subjected to the PCR assay with designed primers targeting full length amplification of ORF2 gene of PCV2 for molecular characterization. Randomly 32 positive samples by full length PCV2-ORF2 gene PCR were sequenced for genotyping. Signature motif and phylogenetic analysis of 32 PCV2 sequences revealed 62.5% (n = 20) prevalence of PCV2d genotype followed by 21.8% (n = 7) of PCV2h or PCV2-IM1 and 15.6% (n = 5) of PCV2b genotypes. Twenty five PCR positive field samples were subjected for virus isolation in PK15 cells and characterized. Out of 25 samples processed 5 (20%) PCV2 isolates obtained in this study were confirmed by PCR and immune fluorescence assay. Molecular characterization of PK15 adapted five PCV2 isolates confirmed circulation of PCV2d, PCV2h and PCV2b genotypes in pigs under field conditions in southern India. Conclusions Isolation and molecular epidemiological study of PCV2 in southern states of India evidences high circulation of PCV2d genotypes in field conditions in comparison to other genotypes.

Laboratory detection of rabies in most cases is based on detection of the antigen by fluorescent antibody test, however, in weak positive cases confirmative laboratory diagnosis depends on widely accepted mouse inoculation test. Cell lines like neuroblastoma have been used to isolate the virus with greater success not only to target for diagnosis, but also for molecular studies that determine the epidemiology of the circulating street rabies strains and in studies that look at the efficiency of the developed monoclonal antibodies to neutralize the different rabies strains. Due to the recent issues in obtaining ethical permission for mouse experimentation, and also the passages required in the cell lines to isolate the virus, we report herewith a co-culture protocol using the murine neuroblastoma (MNA) cells, which enable quicker isolation of street rabies virus with minimum passages. This study is not to have an alternative diagnostic assay, but an approach to produce sufficient amount of rabies virus in minimum passages by a co-culture approach in MNA cells. The MNA cells are co-cultured by topping the normal cells with infected cells every 48 h and the infectivity was followed up by performing direct fluorescent-antibody test. The co-culture approach results in 100% infectivity and hence the use of live mouse for experimentation could be avoided. Co-culture method provides an alternative for the situations with limited sample volume and for the quicker isolation of virus which warrants the wild type strains without much modification.

The immunopathogenesis of infectious bronchitis virus (IBV) infection in the chicken is reviewed. While infectious bronchitis (IB) is considered primarily a disease of the respiratory system, different IBV strains may show variable tissue tropisms and also affect the oviduct and the kidneys, with serious consequences. Some strains replicate in the intestine but apparently without pathological changes. Pectoral myopathy has been associated with an important recent variant. Several factors can influence the course of infection with IBV, including the age, breed and nutrition of the chicken, the environment and intercurrent infection with other infectious agents. Immunogenic components of the virus include the S (spike) proteins and the N nucleoprotein. The humoral, local and cellular responses of the chicken to IBV are reviewed, together with genetic resistance of the chicken. In long-term persistence of IBV, the caecal tonsil or kidney have been proposed as the sites of persistence. Antigenic variation among IBV strains is related to relatively small differences in amino acid sequences in the S1 spike protein. However, antigenic studies alone do not adequately define immunological relationships between strains and cross-immunisation studies have been used to classify IBV isolates into 'protectotypes'. It has been speculated that changes in the S1 protein may be related to differences in tissue tropisms shown by different strains. Perhaps in the future, new strains of IBV may arise which affect organs or systems not normally associated with IB.

The present study examined 434 field samples including serum (n = 273), swabs from natural orifices (n = 52) and postmortem tissue samples (n = 109) from both suspected and asymptomatic swine from Andhra Pradesh, Karnataka, Kerala, Pondicherry, Tamil Nadu, and Telangana states in southern India. All the samples were processed for molecular screening of PCV3 by specific PCR assay. Overall molecular positivity rate of PCV3 was found to be 0.7% in southern India with one sample positive from each state of Tamil Nadu, Kerala and Telangana. All the three PCR positive PCV3 samples are detected from reproductive failures and were processed and propagated in PK15 cell line for virus isolation. Out of 3 samples processed, one (INDKL9PK76) PCV3 isolate could be obtained in this study and it was confirmed by specific PCR at third and fifth passage levels. Sequencing of PCV3 positive PCR amplicon (INDKL9PK76) revealed 1004 nucleotides and BLAST analysis confirmed partial sequence of the PCV3 genome. The aligned contig sequence was submitted to GenBank under the accession number of MW627201. PCV3 sequence in this study revealed 99% homology with PCV3 isolates from Europe and China. Phylogentic analysis of the PCV3 isolate-INDKL9PK76 sequence along with established PCV3 genotypes revealed clustering within PCV3 genotypes. Characterization of PCV3 (INDKL9PK76) isolate based on deduced amino acid composition of PCV3-capsid protein revealed “A” (alanine) and “R” (arginine) at 24th and 27th residues respectively confirming the incidence of PCV3a genotype. This study evidences PCV3 associated reproductive failure in domestic pigs for the first time in southern India.

Cell mediated immune response to immunoaffinity chromatography purified midgut antigen of Rhipicephalus haemaphysaloides ticks in rabbits was studied by using lymphocyte transformation test. This test was carried out by using 5-bromo-2′-deoxy-uridine kit method. The blastogenic response of peripheral blood lymphocytes of normal rabbit to different concentrations of antigen and mitogen (Con A) showed that 2 μg of antigen and 2 μg of mitogen gave maximum stimulation index. The antigen specific responsiveness of immunized rabbits with affinity purified 35 kDa midgut antigen was highly significant ( P  ≤ 0.01) compared to mitogen. The maximum lymphocyte stimulation index (LSI) of 2.47 was observed on 49 day post immunization in immunized group. The lymphocytes separated from control animal cultured in RPMI1640 medium with 2 μg of antigen and 2 μg of mitogen (Con A) were never stimulated and their LSI values were below 2.0.

Forty, newly hatched, unsexed broiler chicks were fed diets containing 10 ppm cyclopiazonic acid (CPA) and 1 ppm T-2 toxin (T2) either individually or in combination for 28 days to study the immunopathological effects. Lymphoid organs revealed lymphocytolysis and lymphoid depletion in all toxin fed birds. Thymic and splenic CD+4 and CD+8 lymphocytes decreased significantly (p<0.01) in toxin fed birds when compared to the control. Thymic CD+8 lymphocytes of T2 and CPA-T2 showed significant (p<0.01) decrease from that of CPA and control groups. Splenic CD+4 and CD+8 lymphocytes showed significant (p<0.01) decrease in CPA and CPA-T2 fed groups when compared to the control. The T2 group did not differ significantly from that of control. The stimulation index (SI) of splenocytes to concavalin A revealed significant (p<0.01) decrease in all toxin fed birds. Significant (p<0.01) decrease were observed for the haemagglutination inhibition (HI) titres to Newcastle disease virus vaccine F strain (NDV) of birds fed CPA, T2 and in combination. Significant (p<0.01) interaction was found for lymphocyte subsets, SI and HI titres to NDV. The study indicated the immunosuppressive effect of these toxins either alone or in combination in broiler chicks.

A comparative study of cytokine and toll-like receptor (TLR) mRNA expression in 3 weeks old indigenous and commercial chickens infected with a very virulent strain of Infectious bursal disease virus (IBDV) was performed using a custom-made microarray chip. In uninfected indigenous chickens, the basal levels of interleukin (IL) 15 were lower and IL 16 was higher than their commercial counterparts. In the IBDV infected indigenous chickens, only IL16 gene expression was down regulated, while TLR3 expression was up regulated significantly. In the IBDV infected commercial chickens IL15, IL16 and TLR3 were down regulated. But, IL1-β, IL2, IL8, IL12, IL17, interferon (IFN)-α and β were significantly increased compared with the control. In IBDV infected indigenous chickens, IL15, IFN-γ, beta-defensin and TLR3 were up regulated compared to virus-infected commercial chickens. The results suggested that up regulation of TLR3, a ligand for double-stranded (ds) RNA probably could account for the possible clinical resistance in these birds. There was a 5.2 fold difference by quantitative real-time RT-PCR between indigenous and commercial chickens in TLR3 mRNA expression. Therefore, TLR3, a receptor for dsRNA could be a putative molecule that could play a role in differential innate and adaptive immune responses to IBDV in commercial and indigenous chickens.

Parvoviruses are ubiquitous pathogens that cause fatal disease in cats. Feline panleukopenia virus (FPV) is a primitive virus reported first and canine parvovirus (CPV) evolved from FPV and was reported later. Both induce disease in cats and dogs with correlative signs. FPV in domestic cats is genetically diverse and some strains may differ from those used for vaccination. In this study, a virus of FPV strain, ABT/MVC/2022/FPV/001, was identified from a fecal sample of the suspected cat with severe haemorrhagic gastroenteritis. The phylogenetic analysis and complete genome sequence of the strain share 99.75% nucleotide identity with FPV variant MH559110 belonging to Tamil Nadu, India. The results also reveal similarities to strains isolated from Italy, Belgium, and China. The deduced amino acid sequence of isolated strain revealed specific amino acid substitution (Pro5Ala, Phe6Val, His7Gln, Asn9Asp, Lys16Arg, Lys19Arg, Asn52Lys, Gly58Trp, Thr66Ser, Lys67Arg, Leu70His, Asn373Asp and Ala390Thr) which differed from MH559110 and other strains. The complete genomic analysis revealed that the FPV strain circulating in India is evolving rapidly with unique antigenic variations between field FPV, CPV and vaccine strains which may be the major cause for vaccine failure in vaccinated cats.

This study used 56 aborted and stillborn fetuses from organized swine farms in Tamil Nadu and Kerala, southern states of India. All samples were screened by using a PCR assay that targets the NS1 gene for PPV. Furthermore, the PCR positive samples were subjected to amplification of the VP2 gene of PPV1 with designed primers and sequenced for further study. The PCR screening of 56 samples found that 14.3% (n = 8) were positive for PPV genome. According to VP2 gene–based PCR for PPV1, 897 bp specific amplicons were detected in all eight of the samples. Two of the eight positive samples (L17 and T5) were sequenced and annotated randomly. The BLAST analysis of contig sequence INDTNCHN-T5 revealed 100% sequence homology with Chinese PPV1genome, whereas sequence from INDTNCHN-L17 revealed 99.43% sequence homology with Spain, Chinese, and German. PPV1 sequences and both the sequences INDTNCHN-T5 and INDTNCHN-L17 were submitted to the GenBank under the accession numbers MW822566 and MW822567 respectively. A phylogenetic analysis of the sequences in this study revealed specific grouping along with PPV1 strains in cluster E. Amino acid analysis of both isolated sequences in addition to the reference sequence from PPV1 showed variations in position 215 (I to T) in both the isolates, variation at position 228 (Q to E) in T5 isolate and variations at position 59 (L to M) and 314 (K to E) in L17 isolate. This study represents the first report of PPV1 cluster E in Tamil Nadu, southern India.