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Macrophages, as sentinels of robust host immunity, are key regulators of innate immune responses against invading mycobacteria; however, pathogenic mycobacteria survive in the infected host by subverting host innate immunity. Infection dependent expression of early secreted antigenic target protein 6 (ESAT-6) by Mycobacterium tuberculosis is strongly correlated with subversion of innate immune responses against invading mycobacteria. As a part of multifaceted immunity to mycobacterial infection, induced expression of cyclooxygenase-2 (COX-2) may act as an important influencing factor towards effective host immunity. In the current investigation, we demonstrate that ESAT-6 triggers COX-2 expression both in vitro and in vivo in a TLR2 dependent manner. Signaling perturbation data suggest that signaling dynamics of PI3K and p38 and JNK1/2 MAPK assume critical importance in ESAT-6 triggered expression of COX-2 in macrophages. Interestingly, ESAT-6 triggered PI3K-MAPK signaling axis holds the capacity to regulate coordinated activation of NF-kappa B and AP-1. Overall, current investigation provides mechanistic insights into ESAT-6 induced COX-2 expression and unravels TLR2 mediated interplay of PI3K and MAPK signaling axis as a rate-determining step during intricate host immune responses. These findings would serve as a paradigm to understand pathogenesis of mycobacterial infection and clearly pave a way towards development of novel therapeutics. (C) 2011 Elsevier Ltd. All rights reserved.

Background: Mycobacterium tuberculosis, a causative agent of chronic tuberculosis disease, is widespread among some animal species too. There is paucity of information on the distribution, prevalence and true disease status of tuberculosis in Asian elephants (Elephas maximus). The aim of this study was to estimate the sensitivity and specificity of serological tests to diagnose M. tuberculosis infection in captive elephants in southern India while simultaneously estimating sero-prevalence. Methodology/Principal Findings: Health assessment of 600 elephants was carried out and their sera screened with a commercially available rapid serum test. Trunk wash culture of select rapid serum test positive animals yielded no animal positive for M. tuberculosis isolation. Under Indian field conditions where the true disease status is unknown, we used a latent class model to estimate the diagnostic characteristics of an existing (rapid serum test) and new (four in-house ELISA) tests. One hundred and seventy nine sera were randomly selected for screening in the five tests. Diagnostic sensitivities of the four ELISAs were 91.3-97.6% (95% Credible Interval (CI): 74.8-99.9) and diagnostic specificity were 89.6-98.5% (95% CI: 79.4-99.9) based on the model we assumed. We estimate that 53.6% (95% CI: 44.6-62.8) of the samples tested were free from infection with M. tuberculosis and 15.9% (97.5% CI: 9.8 - to 24.0) tested positive on all five tests. Conclusions/Significance: Our results provide evidence for high prevalence of asymptomatic M. tuberculosis infection in Asian elephants in a captive Indian setting. Further validation of these tests would be important in formulating area-specific effective surveillance and control measures.

Acting primarily through its specific G protein-coupled receptor termed FPr, prostaglandin (PG) F2αinduces regression of the corpus luteum (CL) at the end of a non-fertile oestrous cycle. This study was aimed at cloning a full-length cDNA for FPr and determining its expression and protein concentrations during different stages of CL development in the water buffalo. Serum progesterone and StAR expression were determined to establish temporal relationships between indices of steroidogenesis and changes in FPr expression at different stages of CL development. In contrast to the dairy cow, the stage IV CL (day 20 of the oestrous cycle) did not appear to be functionally regressed in the buffalo. Molecular cloning of a cDNA encoding the buffalo FPr yielded a full length 2193 bp FPr cDNA containing a single open reading frame encoding a 362 amino acid protein with seven putative membrane-spanning domains. The deduced buffalo FPr amino acid sequence possesses a high degree of identity with the other mammalian homologues. Steady state concentration of buffalo FPr transcript increased (P> 0.05) from stage I to stage II/III, and declined at 18 h post PGF2αinjection. The FPr concentration expressed as fmol/μg of plasma membrane protein showed an increase (P> 0.05) from stage I (1.98 ± 0.10), through stage II/III (2.42 ± 0.48) to stage IV (2.77 ± 0.18). High affinity FPr was observed in stage I (Kd4.86 nmol) and stage II/III (Kd6.28 nmol) while low affinity FPr (Kd19.44 nmol) was observed in stage IV. In conclusion, we have cloned a full length FPr cDNA from buffalo cow CL and observed that FPr mRNA expression, receptor number and affinity did not vary significantly (P> 0.05) within the luteal phase of the oestrous cycle.

During reproductive life, only a selected few ovarian follicles mature and ovulate, while the vast majority of follicles undergo a degenerative process called atresia. Recent studies have indicated that follicular atresia is mediated through apoptosis of follicular granulosa cells. The objectives of the present study were to determine the time of onset of apoptosis in granulosa cells of preovulatory follicles and to evaluate the consequences of gonadotropin withdrawal on mitogen-activated protein (MAP) kinase activities. Bonnet monkeys (Macaca radiata) undergoing controlled ovarian stimulation cycles were utilized for stimulation of multiple follicles, and granulosa cells were retrieved from preovulatory follicles at 24, 48, 72, and 96 h after stopping gonadotropin treatment. Serum and follicular fluid estradiol concentrations were highest at 24 h but declined precipitously (P < 0.05) to reach the lowest concentrations at 96 h; however, progesterone concentrations during this period did not increase, indicating the absence of luteinization. Quantitative analysis of genomic DNA by 3'-end labeling revealed the presence of low-molecular-weight fragments from 48 h onward, but by agarose gel electrophoresis, DNA laddering could be visualized only after 72 h. Messenger RNA expression for Bax, caspase-2, and caspase-3 increased with the onset of apoptosis. Immunoblot analysis of MAP kinases in lysates of granulosa cells (48-72 h) indicated increased (P < 0.05) levels of phosphorylated extracellular response kinase-1 and -2, Jun N-terminal kinase (JNK)-1 and -2, and p38. However, in vitro kinase assay data indicated that only phospho-JNK and -p38 activities were higher at 72 h compared to 24 h. These results demonstrate that granulosa cells of preovulatory follicles undergo apoptosis and that increased activities of phospho-JNK and -p38 are correlated with apoptosis in the primate.