•Detecting M. bovis antibodies in cattle sera after a tuberculin skin test (TST) may improve bovine tuberculosis control.•Used as an anamnestic test, the ELISA reported herein had high levels of ...specificity and positive predictive value.•The ELISA fulfilled the World Organization for Animal Health criteria.•This serological ELISA used 15–17 days after a negative TST could be used on dairy farms to improve livestock health.
Programs for the eradication of bovine tuberculosis (bTB) focus on the tuberculin skin test (TST) and slaughter of reactor cattle. However, the disease remains an animal health concern in several countries and improving the efficiency of the TST has become a critical issue. The detection of Mycobacterium bovis antibodies in serum, within weeks after the TST, may be a rapid and inexpensive way to improve bTB control. This study reports the validation of an enzyme-linked immunosorbent assay (ELISA) to detect bovine tuberculosis as an ancillary test to TST in dairy farms in Argentina.
The estimated validation parameters were within the established requirements of the World Organization for Animal Health (OIE). The test demonstrated high repeatability, with coefficients of variation <25%. High test reproducibility through interlaboratory testing was also found, with an estimated Pearson coefficient of 0.9648 (95% confidence intervals 0.9315–0.9820). The ELISA detected tuberculous cattle unidentified by the TST. Of 43 animals sent to slaughterhouses that were ELISA positive 15–17 days after a negative TST, 36 were confirmed as infected with M. bovis by histopathology and IS6110 PCR. According to ROC curve analysis of results of 145 cattle from M. bovis-free herds and the 36 M. bovis-infected cattle, at a corrected optical density cut-off point of 0.3853, specificity was 95.95% and the positive predictive value at this cut-off was 83.72%. The ELISA detection test validated in this study could be readily applied in dairy farms, to complement a prior TST and improve livestock health.
In a previous study, we evaluated the degree of virulence of Mycobacterium avium subsp. paratuberculosis (Map) strains isolated from cattle in Argentina in a murine model. This assay allowed us to ...differentiate between high-virulent MapARG1347 and low-virulent MapARG1543 strains. To corroborate whether the differences in virulence could be attributed to genetic differences between the strains, we performed Whole Genome Sequencing and compared the genomes and gene content between them and determined the differences related to the reference strain MapK10. We found 233 SNPs/INDELS in one or both strains relative to Map K10. The two strains share most of the variations, but we found 15 mutations present in only one of the strains. Considering NS-SNP/INDELS that produced a severe effect in the coding sequence, we focus the analysis on four predicted proteins, putatively related to virulence. Survival of MapARG1347 strain in bMDM was higher than MapARG1543 and was more resistant to acidic pH and H2O2 stresses than MapK10. The genomic differences between the two strains found in genes MAP1203 (a putative peptidoglycan hydrolase), MAP0403 (a putative serine protease) MAP1003c (a member of the PE-PPE family) and MAP4152 (a putative mycofactocin binding protein) could contribute to explain the contrasting phenotype previously observed in mice models.
•WGS of Argentinian isolates with contrasting pathogenic phenotypes.•Phylogenetic analysis with whole genome sequences available from GenBank.•Identification of NS-SNP and INDELS by comparative genomic analysis.
DNA damage induces apoptosis and many apoptotic genes are regulated via alternative splicing (AS), but little is known about the control mechanisms. Here we show that ultraviolet irradiation (UV) ...affects cotranscriptional AS in a p53-independent way, through the hyperphosphorylation of RNA polymerase II carboxy-terminal domain (CTD) and a subsequent inhibition of transcriptional elongation, estimated in vivo and in real time. Phosphomimetic CTD mutants not only display lower elongation but also duplicate the UV effect on AS. Consistently, nonphosphorylatable mutants prevent the UV effect. Apoptosis promoted by UV in cells lacking p53 is prevented when the change in AS of the apoptotic gene
bcl-x is reverted, confirming the relevance of this mechanism. Splicing-sensitive microarrays revealed a significant overlap of the subsets of genes that have changed AS with UV and those that have reduced expression, suggesting that transcriptional coupling to AS is a key feature of the DNA-damage response.
Several glitches have been observed in young, isolated radio pulsars, while a clear detection in accretion-powered X-ray pulsars is still lacking. We use the Pizzochero snowplow model for pulsar ...glitches as well as starquake models to determine for the first time the expected properties of glitches in accreting pulsars and their observability. Since some accreting pulsars show accretion-induced long-term spin-up, we also investigate the possibility that anti-glitches occur in these stars. We find that glitches caused by quakes in a slow accreting neutron star are very rare and their detection extremely unlikely. On the contrary, glitches and anti-glitches caused by a transfer of angular momentum between the superfluid neutron vortices and the non-superfluid component may take place in accreting pulsars more often. We calculate the maximum jump in angular velocity of an anti-glitch and we find that it is expected to be ΔΩa − gl ≈ 10-5 − 10-4 rad s-1. We also note that since accreting pulsars usually have rotational angular velocities lower than those of isolated glitching pulsars, both glitches and anti-glitches are expected to have long rise and recovery timescales compared to isolated glitching pulsars, with glitches and anti-glitches appearing as a simple step in angular velocity. Among accreting pulsars, we find that GX 1+4 is the best candidate for the detection of glitches with currently operating X-ray instruments and future missions such as the proposed Large Observatory for X-ray Timing (LOFT).
Mycobacterium avium sp. avium (MAA), M. avium sp. hominissuis (MAH), and M. avium sp. paratuberculosis (MAP) are the main members of the M. avium complex (MAC) causing diseases in several hosts. The ...aim of this study was to describe the genetic diversity of MAC isolated from different hosts. Twenty-six MAH and 61 MAP isolates were recovered from humans and cattle, respectively. GenoType CM® and IS1311-PCR were used to identify Mycobacterium species. The IS901-PCR was used to differentiate between MAH and MAA, while IS900-PCR was used to identify MAP. Genotyping was performed using a mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) scheme (loci: 292, X3, 25, 47, 3, 7, 10, 32) and patterns (INMV) were assigned according to the MAC-INMV database (http://mac-inmv.tours.inra.fr/). Twenty-two (22/26, 84·6%) MAH isolates were genotyped and 16 were grouped into the following, INMV 92, INMV 121, INMV 97, INMV 103, INMV 50, and INMV 40. The loci X3 and 25 showed the largest diversity (D: 0·5844), and the global discriminatory index (Hunter and Gaston discriminatory index, HGDI) was 0·9300. MAP (100%) isolates were grouped into INMV 1, INMV 2, INMV 11, INMV 8, and INMV 5. The HGDI was 0·6984 and loci 292 and 7 had the largest D (0·6980 and 0·5050). MAH presented a higher D when compared with MAP. The MIRU-VNTR was a useful tool to describe the genetic diversity of both MAH and MAP as well as to identify six new MAH patterns that were conveniently reported to the MAC-INMV database. It was also demonstrated that, in the geographical region studied, human MAC cases were produced by MAH as there was no MAA found among the human clinical samples.
The lungs of patients with cystic fibrosis (CF) are characterized by an exaggerated inflammation driven by secretion of IL-8 from bronchial epithelial cells and worsened by Pseudomonas aeruginosa ...infection. To identify novel antiinflammatory molecular targets, we previously performed a genetic study of 135 genes of the immune response, which identified the c.2534C>T (p.S845L) variant of phospholipase C-β3 (PLCB3) as being significantly associated with mild progression of pulmonary disease. Silencing PLCB3 revealed that it potentiates the Toll-like receptor's inflammatory signaling cascade originating from CF bronchial epithelial cells. In the present study, we investigated the role of the PLCB3-S845L variant together with two synthetic mutants paradigmatic of impaired catalytic activity or lacking functional activation in CF bronchial epithelial cells. In experiments in which cells were exposed to P. aeruginosa, the supernatant of mucopurulent material from the airways of patients with CF or different agonists revealed that PLCB3-S845L has defects of 1) agonist-induced Ca
release from endoplasmic reticulum and rise of Ca
concentration, 2) activation of conventional protein kinase C isoform β, and 3) induction of IL-8 release. These results, besides identifying S845L as a loss-of-function variant, strengthen the importance of targeting PLCB3 to mitigate the CF inflammatory response in bronchial epithelial cells without blunting the immune response.
Cytomegalovirus (CMV) disease represents a major cause of post-transplantation morbidity and mortality. To estimate the risk of infection and monitor response to antiviral therapy, current guidelines ...suggest combination of viral load monitoring with direct assessment of CMV-specific immune response. We used enzyme-linked immunospot (ELISpot) for the evaluation of CMV-specific T-cell response in kidney transplant recipients with CMV viremia and investigated how information gained could help manage CMV infection.
Seventeen patients on pre-emptive antiviral therapy and CMV quantitative polymerase chain reaction (qPCR) ≥500 copies/mL (first episode after transplantation) were assessed using ELISpot and divided into Weak (9 patients with baseline ELISpot <25 spot-forming colonies SFCs/200,000 peripheral blood mononuclear cells PBMCs) and Strong Responders (8 patients with baseline ELISpot ≥25 SFCs/200,000 PBMCs). CMV-specific T-cell response, infection severity, viral load, and antiviral therapy were prospectively recorded and compared between groups at 1, 2, and 24 months of follow-up.
Demographic and transplant characteristics of Weak and Strong Responders were similar. No episodes of CMV disease were observed. Weak Responders were more likely to experience CMV syndrome (56% vs 36.5%) and late virus reactivation (56% vs 25%) than Strong Responders. Weak Responders showed higher baseline median viral loads (19,700 vs 9265 copies/mL) and needed antiviral therapy for longer (179 vs 59.5 days). T-cell response showed 2 main patterns: early and delayed.
ELISpot provides prognostic information about infection severity, risk of late reactivation, and response to therapy. Randomized trials, evaluating the need for antiviral therapy in kidney transplant recipients with asymptomatic infection and effective virus-specific T-cell immune response, are warranted.
•ELISpot shows CMV-specific T-cell response in kidney transplant recipients.•CMV-specific T-cell response influences clinical course of the disease.•Strong virus-specific immune response can mitigate CMV infection.•Antiviral therapy could be tailored according to ELISpot results.
Temporal coherence (i.e., the degree of synchronicity of a given variable among ecological units within a predefined space) has been shown for several limnological features among temperate lakes, ...allowing predictions about the structure and function of ecosystems. However, there is little evidence of temporal coherence among tropical aquatic systems, where the climatic variability among seasons is less pronounced. Here, we used data from long-term monitoring of physical, chemical and biological variables to test the degree of temporal coherence among 18 tropical coastal lagoons. The water temperature and chlorophyll-a concentration had the highest and lowest temporal coherence among the lagoons, respectively, whereas the salinity and water colour had intermediate temporal coherence. The regional climactic factors were the main factors responsible for the coherence patterns in the water temperature and water colour, whereas the landscape position and morphometric characteristics explained much of the variation of the salinity and water colour among the lagoons. These results indicate that both local (lagoon morphometry) and regional (precipitation, air temperature) factors regulate the physical and chemical conditions of coastal lagoons by adjusting the terrestrial and marine subsidies at a landscape-scale. On the other hand, the chlorophyll-a concentration appears to be primarily regulated by specific local conditions resulting in a weak temporal coherence among the ecosystems. We concluded that temporal coherence in tropical ecosystems is possible, at least for some environmental features, and should be evaluated for other tropical ecosystems. Our results also reinforce that aquatic ecosystems should be studied more broadly to accomplish a full understanding of their structure and function.
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