Piperlongumine (PL) is a bioactive alkaloid and medicinal compound of piperamide isolated from the long pepper (Piper longum Linn). It has demonstrated bactericidal action against Mycobacterium tuberculosis (MTB), the cause of pulmonary tuberculosis; nevertheless, immunomodulatory activity had not been identified for it in MTB-triggered granulomatous inflammation. This study investigated if piperlongumine could inhibit such inflammation. Mycobacterium tuberculosis strain H37Rv was subjected to a broth microdilution assay. Piperlongumine at 5, 15, and 25 μg/mL, 0.2% dimethyl sulphoxide as control or 4 μM of dexamethasone were tested in vitro on MH-S murine alveolar macrophages. BALB/c mice were orally administered PL at 50, 100 and 150 mg/kg b.w. after trehalose-6,6-dimycolate (TDM) stimulation. Chemokine and cytokine concentrations were determined in lung supernatants. Flow cytometry and Western blot analysis were performed to determine phosphorylated spleen tyrosine kinase (Syk), c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) pathways. Piperlongumine inhibited inflammatory mediators and adherence of lymphocyte function-associated antigen 1 to MH-S cells following TDM activation. It also improved macrophage clearance of MTB. In TDM-stimulated MH-S cells, PL significantly influenced the macrophage inducible Ca²⁺-dependent lectin receptor (Mincle)-Syk-ERK signalling pathway. Oral dosing of PL effectively suppressed the development of pulmonary granulomas and inflammatory reactions in the TDM-elicited mouse granuloma model. PL as an inhibitor of MTB-triggered granulomatous inflammation may be an effective complementary treatment for mycobacterial infection.

OBJECTIVE To develop a noninvasive biomarker-based detection system specific for Mycobacterium bovis for monitoring infection in wild animals. SAMPLE Serum samples from 8 experimentally infected yearling white-tailed deer (Odocoileus virginianus) and 3 age-matched control deer and from 393 Minnesota Department of Natural Resources hunter-harvested white-tailed deer in northwest Minnesota. PROCEDURES 8 yearling deer were inoculated with 2 × 10(8) CFUs of virulent M bovis strain 1315 (day 0), and sera were obtained on days 0, 19, 48, and 60; sera were obtained from 3 uninoculated control deer on those same days. Sera from these deer and 9 M bovis-positive hunter-harvested deer were tested for 3 Mycobacterium-specific biomarkers (MB1895c, MB2515c, and polyketide synthase 5) by use of an indirect ELISA. That same ELISA was used to test sera obtained from 384 exposed noninfected deer in northwest Minnesota from 2007 through 2010, concurrent with an outbreak of tuberculosis involving cattle and deer in that region. RESULTS ELISA results revealed that tuberculosis infection could be detected as early as 48 days after inoculation in experimentally infected deer. Results for 384 deer sera revealed that prevalence of tuberculosis decreased over the 4-year period. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that the prevalence of tuberculosis in Minnesota deer decreased after 2009 but tuberculosis may have persisted (as subclinical disease) at extremely low levels, as indicated by the presence of low concentrations of circulating biomarkers. Biomarker-based diagnostic tests may offer a specific approach for early identification of M bovis infection.

Objective: To assess the prevalence of Mycobacterium bovis infection in cattle and wild ruminants (WRs) in a wildlife-livestock interface area (WLIA) of the Mexican highland plateau. Animals: 24,400 cattle from 793 herds (including 17,351 commercially slaughtered cattle) and 142 WRs (110 white-tailed deer [Odocoileus virginianus], 20 red deer [Cervus elaphus], and 12 North American elk [Cervus canadensis]) harvested via controlled hunting. Procedures: Cattle were serially tested for M bovis infection via caudal fold tuberculin and comparative cervical tuberculin tests during field surveillance. Carcasses of cattle and WRs were inspected for gross lesions; samples suggestive of tuberculosis were analyzed via histologic evaluation and mycobacterial culture (HMC). A PCR assay to detect Mycobacterium tuberculosis complex organisms was performed to confirm positive results of HMC. Results: WRs had inflammatory lesions in lungs and lymph nodes, although HMC results did not indicate M bovis infection. Eight cattle had positive results for both tuberculin tests, and 31 had positive results for HMC of grossly detected lesions; all were from 7 herds, and ≥ 1 cow in each herd had positive PCR assay results. These 7 herds were depopulated; adjacent herds and herds related via commerce were quarantined. Calculated true prevalence of M bovis infection was 0.86% (95% confidence interval, 0.24% to 1.49%) in cattle; M bovis was not detected in any WRs. Conclusions and Clinical Relevance: M bovis infection was present in cattle. Although transmission to WRs in this WLIA was not detected, diagnosis and prevention activities should be implemented and consolidated to prevent potential M bovis transmission between cattle and WRs.

Objective-To determine whether cats exposed at a residence were infected with Mycobacterium bovis, whether the tuberculin skin test can identify cats infected with M bovis, and whether an ELISA could identify tuberculosis-infected cats. Animals-20 domestic cats exposed to a cat with laboratory-confirmed disseminated M bovis infection. Procedure-Cats were administered a tuberculin skin test and monitored for 72 hours. Blood and fecal samples were collected. Cats were then euthanatized, and postmortem examinations were performed. Tissues were examined grossly and histologically for signs of mycobacteriosis. Pooled tissue samples and fecal samples were submitted for mycobacterial culture. Blood samples were examined for evidence of tuberculosis by use of a comparative ELISA. Results-4 cats had positive responses for the ELISA, and 2 cats had suspicious responses. All tuberculin skin tests yielded negative results. No gross or histologic lesions of tuberculosis were detected in any tissues, and mycobacteria were not isolated from tissues or feces obtained from the 20 cats. Conclusions and Clinical Relevance-All cats that had positive or suspicious responses for the ELISA were offspring of the cat with tuberculosis. Evidence of tuberculosis was not seen in other cats at the residence, the owner, or the attending veterinarian. The most likely source of tuberculosis for the infected cat was through the consumption of M bovis-infected wildlife carcasses or offal. Because M bovis is endemic in wildlife in northeastern Michigan, there is a risk of exposure to tuberculosis in companion animals, their owners, and attending veterinarians.

Postmortem examinations were done on 51 wood bison (Bison bison athabascae) killed as part of a multidisciplinary research project in the Mackenzie Bison Sanctuary, Northwest Territories, Canada, between 1986 and 1988. There was no gross, histological or bacteriological evidence of brucellosis or tuberculosis in these bison. Traumatic lesions were seen in one calf that had been attacked by wolves and a second calf that had been gored. Antibody titers to Brucella abortus were not found in sera from these 51 animals or an additional 112 wood bison that were chemically-immobilized or killed in the Sanctuary between 1986 and 1990. The combined prevalence of the diseases in the population could not have exceeded 5.95% for the necropsy survey to have missed finding at least one infected animal, and the prevalence of brucellosis in the population would have had to be less than 1.95% for the broader serological survey to have failed to find at least one reactor animal on the battery of tests. These results, and the cumulative epidemiological information on brucellosis and tuberculosis in bison, indicate that bovine brucellosis and tuberculosis are not enzootic in the wood bison population in and around the Mackenzie Bison Sanctuary, and suggest that the population is free of these diseases. However, this expanding population is at risk of contracting both diseases from the infected bison population in and around nearby Wood Buffalo National Park.

The effect of T-2 toxin, a radiomimetic immunosuppressive agent, on resistance to the facultative intracellular bacterial pathogens Listeria monocytogenes (strain EGD), Mycobacterium bovis (BCG Copenhagen 1331), and Salmonella typhimurium was determined. Female Swiss ICR mice were given a single dose of T-2 toxin (4 mg/kg of body weight) by gastric gavage. On the seventh day after toxin administration, the mice were infected by intraperitoneal inoculation with L monocytogenes, S typhimurium, or M bovis. Mice given the toxin also were exposed to respirable droplet nuclei containing L monocytogenes or M bovis. The effect of the toxin on the course of infection was monitored by observing mortality or by enumeration of bacteria in te spleen or lungs of infected mice. The toxin increased resistance to infection with L monocytogenes initiated by intraperitoneal inoculation, but reduced resistance to M bovis infection initiated by intraperitoneal inoculation. The toxin had no appreciable effect on the course of salmonellosis or on resistance to infection initiated by inhalation of L monocytogenes or M bovis aerosols. Therefore, it was concluded that T-2 toxin does not necessarily reduce resistance to infection in mice. The toxin's effect on the course of in vivo bacterial infections depends on the nature of the infective agent and the route of inoculation.

Lymphocyte stimulation tests (LST), performed using 6 antigen preparations, were compared individually and in pairs. The tests were performed on 433 blood samples collected from elk in Mycobacterium bovis-infected herds. These elk were killed as part of Agriculture and Agri-Food Canada's bovine tuberculosis eradication policy, and mycobacterial culture results were obtained from tissues of each animal The LST, which had the highest total sum of sensitivity and specificity, was a comparative test that used M bovis purified protein derivative (PPD) and M paratuberculosis (johnin) PPD. This test had a sensitivity of 76%, with confidence limits (CL) of 63 to 85% for this estimate, and specificity of 77% (CL, 72 to 81%). The LST, using only M bovis PPD antigen, had a sensitivity of 70% (CL, 57 to 80%) and specificity of 74% (CL, 69 to 79%); when it was compared with culture results, using the kappa statistic, agreement was only 32%. This indicated that the LST identified different elk than did M bovis isolation tests.

A study to determine and compare the sensitivity of the caudal fold tuberculin test (CFT) and a commercial gamma-interferon (gamma-IFN) assay for diagnosis of bovine tuberculosis was conducted. A dairy herd with approximately a third of the cattle infected with Mycobacterium bovis was chosen for this study. All cattle from this herd were slaughtered, and tissue specimens for bacteriologic culturing and histologic examination were collected. Results of the CFT and gamma-IFN assay were compared with results of bacteriologic culturing and histologic examination to determine test sensitivity. Results were analyzed, using each of the following 4 standards to classify cattle as infected: positive test result by bacteriologic culturing only; histologic examination only; bacteriologic culturing and histologic examination; and bacteriologic culturing or histologic examination. Sensitivity of the CFT ranged from 80.4 to 84.4%, depending on the standard of comparison. Sensitivity of the gamma-IFN assay ranged from 55.4 to 97.1%, depending on the standard of comparison and on the method of interpretation. The CFT was significantly (P < 0.001) more sensitive than the gamma-IFN assay for diagnosis of bovine tuberculosis when the gamma-IFN assay was conducted and interpreted as instructed by the manufacturer. Maximum overall sensitivity was achieved when results of the CFT and gamma-IFN assay were interpreted in parallel.

Systemic administration of dexamethasone led to a significant reduction in the size of the tuberculin reaction in response to intradermal injection of bovine purified protein derivative in 18 cattle experimentally sensitized to Mycobacterium bovis (P < 0.01) and 8 cattle naturally infected with M bovis (P < 0.001). The reaction in 6 of the 7 M bovis-infected cattle that received dexamethasone was classified as negative for the standard interpretation of the single intradermal comparative tuberculin test. Significantly fewer BoCD2+ (P < 0.05) and BoCD4+ T cells (P < 0.001) were present at the reaction site and in blood of dexamethasone-treated cattle, compared with untreated control cattle. Significantly fewer cells expressing the interleukin-2 receptor and WC1+ gamma delta T cells (P < 0.001), and a significantly greater number of cells expressing the ACT2 antigen (P < 0.05) were found at the reaction site in dexamethasone-treated cattle than in controls. The number of BoCD8+ T cells at the reaction site and in blood was not significantly affected by administration of dexamethasone. In vitro production of interferon-gamma by lymphocytes incubated with bovine purified protein derivative also was significantly lower (P < 0.01) in the dexamethasone-treated cattle.