Investigation of Immune Biomarkers Using Subcutaneous Model of M. tuberculosis Infection in BALB/c Mice: A Preliminary Report

Evaluation and screening of vaccines against tuberculosis depends on development of proper cost effective disease models along with identification of different immune markers that can be used as surrogate endpoints of protection in preclinical and clinical studies. The objective of the present study was therefore evaluation of subcutaneous model of M.tuberculosis infection along with investigation of different immune biomarkers of tuberculosis infection in BALB/c mice. Groups of mice were infected subcutaneously with two different doses : high (2×106 CFU) and low doses (2×102 CFU) of M.tuberculosis and immune markers including humoral and cellular markers were evaluated 30 days post M.tuberculosis infections. Based on results, we found that high dose of subcutaneous infection produced chronic disease with significant (p<0.001) production of immune markers of infection like IFNγ, heat shock antigens (65, 71) and antibody titres against panel of M.tuberculosis antigens (ESAT-6, CFP-10, Ag85B, 45kDa, GroES, Hsp-16) all of which correlated with high bacterial burden in lungs and spleen. To conclude high dose of subcutaneous infection produces chronic TB infection in mice and can be used as convenient alternative to aerosol models in resource limited settings. Moreover assessment of immune markers namely mycobacterial antigens and antibodies can provide us valuable insights on modulation of immune response post infection. However further investigations along with optimization of study protocols are needed to justify the outcome of present study and establish such markers as surrogate endpoints of vaccine protection in preclinical and clinical studies in futur

PCR diagnosis of tick-borne pathogens in Maharashtra state, India indicates fitness cost associated with carrier infections is greater for crossbreed than native cattle breeds

Tick-borne pathogens (TBP) are responsible for significant economic losses to cattle production, globally. This is particularly true in countries like India where TBP constrain rearing of high yielding Bos taurus, as they show susceptibility to acute tick borne disease (TBD), most notably tropical theileriosis caused by Theileria annulata. This has led to a programme of cross breeding Bos taurus (Holstein-Friesian or Jersey) with native Bos indicus (numerous) breeds to generate cattle that are more resistant to disease. However, the cost to fitness of subclinical carrier infection in crossbreeds relative to native breeds is unknown, but could represent a significant hidden economic cost. In this study, a total of 1052 bovine blood samples, together with associated data on host type, sex and body score, were collected from apparently healthy animals in four different agro-climatic zones of Maharashtra state. Samples were screened by PCR for detection of five major TBPs: T. annulata, T. orientalis, B. bigemina, B. bovis and Anaplasma spp.. The results demonstrated that single and co-infection with TBP are common, and although differences in pathogen spp. prevalence across the climatic zones were detected, simplistic regression models predicted that host type, sex and location are all likely to impact on prevalence of TBP. In order to remove issues with autocorrelation between variables, a subset of the dataset was modelled to assess any impact of TBP infection on body score of crossbreed versus native breed cattle (breed type). The model showed significant association between infection with TBP (particularly apicomplexan parasites) and poorer body condition for crossbreed animals. These findings indicate potential cost of TBP carrier infection on crossbreed productivity. Thus, there is a case for development of strategies for targeted breeding to combine productivity traits with disease resistance, or to prevent transmission of TBP in India for economic benefit

Identification and characterization of a novel infectious bursal disease virus from outbreaks in Maharashtra Province of India

Aim: The study was undertaken to isolate infectious bursal disease virus (IBDV) from clinical cases in broiler and cockerel flocks of Maharashtra state, India, and its molecular epidemiological investigation. Materials and Methods: The morbid bursal tissues were collected from flocks suspected for IBD. The samples were subjected for virus adaptation in primary chicken embryo fibroblast (CEF) cells followed by confirmation by reverse transcription polymerase chain reaction (RT-PCR) for partial VP2 sequence and phylogenetic analysis. Results: The isolation of IBDV from field samples took seven blind passages for adaptation in CEF. The cytopathic effects included rounding, aggregation, vacuolation, and detachment of the cells. The RT-PCR showed amplification of 627 bp amplicon specific to the primers for VP2 gene fragment which confirmed successful adaptation and isolation of IBDV using CEF. The nucleotide and deduced amino acids based on phylogeny clustered the current isolate in a distinct clade with classical virulent and antigenic variants. It showed divergence from very virulent (vv) and vaccine strains of Indian origin. The isolate showed unique amino acid substitution at A329V as compared to all other IBDVs. The variation in key amino acids was reported at A222, I242, Q249, Q253, A256, T270, N279, T284, I286, L294, N299, and V329. It shared conserved amino acids at position A222, I242, and Q253 as reported in vvIBDV isolates. However, the amino acids reported at position T270, N279, T284, L294, and N299 are conserved in classic, antigenic variant and attenuated strains of IBDV. The amino acids at positions N279 and T284 indicated that the isolate has key amino acids for cell culture replication. Conclusion: The IBDV field isolate does not reveal the full nucleotide sequence signature of vvIBDV as well as vaccine strains. Hence, we can conclude that it might not belong to vvIBDVs of Indian origin and the vaccine strain used in the region. This may be suggestive of the evolution of the IBDV in the field due to the coexistence of circulating field strains and live attenuated hot strains, resulting into morbidity and mortality, warranting the need for safer protective vaccines, and implementation of stringent biosecurity measures to minimize loss to farmers

Utilization of Fc receptors as a mucosal vaccine strategy against an intracellular bacterium, Francisella tularensis

Numerous studies have demonstrated that targeting Ag to Fc receptors (FcR) on APCs can enhance humoral and cellular immunity. However, studies are lacking that examine both the use of FcR-targeting in generating immune protection against infectious agents and the use of FcRs in the induction of mucosal immunity. Francisella tularensis is a category A intracellular mucosal pathogen. Thus, intense efforts are underway to develop a vaccine against this organism. We hypothesized that protection against mucosal infection with F. tularensis would be significantly enhanced by targeting inactivated F. tularensis live vaccine strain (iFt) to FcRs at mucosal sites, via intranasal immunization with mAb-iFt complexes. These studies demonstrate for the first time that: 1) FcR-targeted immunogen enhances immunogen-specific IgA production and protection against subsequent infection in an IgA-dependent manner, 2) FcgammaR and neonatal FcR are crucial to this protection, and 3) inactivated F. tularensis, when targeted to FcRs, enhances protection against the highly virulent SchuS4 strain of F. tularensis, a category A biothreat agent. In summary, these studies show for the first time the use of FcRs as a highly effective vaccination strategy against a highly virulent mucosal intracellular pathogen

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Not AvailableWe present here the draft genome sequence of Listeria monocytogenes CIIMS-NV-3, a serovar 4b strain isolated from the vaginal swab of a female patient from central India. The availability of this genome may provide useful information on virulence characteristics for comparative genomic analysis.Not Availabl

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Not AvailableBrucellosis is a highly contagious zoonotic infection affecting livestock and human beings. The disease has been reported worldwide except in few countries where it has been eradicated. The prevalence of brucellosis among cattle from 11 farms having a history of abortions was studied. A total of 481 samples comprising of blood, milk, vaginal swabs, vaginal discharges, placental tissues and fetal tissues were collected from 296 animals. Clinical samples were processed for the isolation of Brucella. Serum samples (n = 296) were tested by Rose Bengal Plate Test (RBPT) and indirect ELISA. A total of 90 (30.40%) and 123 (41.55%) samples were positive by RBPT and indirect ELISA, respectively. Also 27.02% samples were positive by both the tests. Brucella isolates (n = 8) were recovered from clinical samples using Brucella selective media. All the isolates demonstrated PCR amplification for the bcsp31 and IS711 genes. Amplification of B. abortus specific primer was demonstrated by all the isolates in AMOS PCR indicating isolates to be of either B. abortus biotype 1, 2 or 4. Risk factors for transmission of brucellosis among cattle population were studied by field surveys. It was observed that lack of awareness about brucellosis (OR = 8.739, P = 0.138) and inadequate floor space (OR = 0.278, P = 0.128) were crucial risk factors for transmission of bovine brucellosis.Not Availabl

Biofilm-Forming Abilities of <i>Listeria monocytogenes</i> Serotypes Isolated from Different Sources

<div><p>A total of 98 previously characterized and serotyped <i>L</i>. <i>monocytogenes</i> strains, comprising 32 of 1/2a; 20 of 1/2b and 46 of 4b serotype, from clinical and food sources were studied for their capability to form a biofilm. The microtiter plate assay revealed 62 (63.26%) strains as weak, 27 (27.55%) strains as moderate, and 9 (9.18%) strains as strong biofilm formers. Among the strong biofilm formers, 6 strains were of serotype 1/2a and 3 strains were of serotype 1/2b. None of the strain from 4b serotype exhibited strong biofilm formation. No firm correlation (p = 0.015) was noticed between any serotype and respective biofilm formation ability. Electron microscopic studies showed that strong biofilm forming isolates could synthesize a biofilm within 24 h on surfaces important in food industries such as stainless steel, ceramic tiles, high-density polyethylene plastics, polyvinyl chloride pipes, and glass. Cell enumeration of strong, moderate, and weak biofilm was performed to determine if the number of cells correlated with the biofilm-forming capabilities of the isolates. Strong, moderate, and weak biofilm showed 570±127× 10³ cells/cm², 33±26× 10³ cells/cm², 5±3× 10³ cells/cm², respectively, indicating that the number of cells was directly proportional to the strength of the biofilm. The hydrophobicity index (HI) analysis revealed higher hydrophobicity with an increased biofilm formation. Fatty acid methyl esterase analysis revealed the amount of certain fatty acids such as iso-C15:0, anteiso-C15:0, and anteiso-C17:0 fatty acids correlated with the biofilm-forming capability of <i>L</i>. <i>monocytogenes</i>. This study showed that different strains of <i>L</i>. <i>monocytogenes</i> form biofilm of different intensities which did not completely correlate with their serotype; however, it correlated with the number of cells, hydrophobicity, and amount of certain fatty acids.</p></div

Strong, moderate, and weak biofilm-forming capabilities of <i>L</i>. <i>monocytogenes</i> isolates from different sources analyzed by microtiter plate assay.

<p>Strong, moderate, and weak biofilm-forming capabilities of <i>L</i>. <i>monocytogenes</i> isolates from different sources analyzed by microtiter plate assay.</p