Comparison of two serological diagnosis tests for bovine paratuberculosis.

Mycobacterium avium subsp. paratuberculosis (MAP) is the causal agent of paratuberculosis (PTBC), a chronic infectious granulomatous enteritis of ruminants. The PTBC diagnosis with commercial ELISA has limitations in sensitivity and specificity, and its results depend on the state of progress of the disease. This research aimed to evaluate two different ELISAs: (a) an "in-house" ELISA with a sonicated antigen obtained from a MAP I47 strain, and (b) a commercial ELISA. In total, the evaluated sample consisted of 394 bovine serum samples from 12 farms in Argentina with high (5-9%) and low (≤ 0.05%) prevalence of PTBC. The evaluation of the new antigen (2.5 µg/mL) was against a 1:50 dilution of the M. phlei faced sera. The cut-off point, sensitivity, and specificity determinations of both techniques were by ROC curve analysis. The area under the curve for the I47 ELISA was 0.9 (CI 95%, 0.93-0.97). With a cut-off point of 8.8%, the sensitivity was 84.3% and the specificity 96.6%. The agreement between both techniques was 0.7 (CI 95%, 0.6-0.8). These results indicate a high discriminative capacity to differentiate positive and negative bovine sera of MAP infection with the I47 ELISA. This result would represent an advantage to dispense with the imported kit.

Enzyme-linked immunosorbent assay as complement of intradermal skin test for the detection of mycobacterium bovis infection in cattle.

Diagnostic tests based on cell-mediated immunity are used in programs for the control and eradication of bovine tuberculosis (bTB), which is mainly caused by Mycobacterium bovis. Additional serological assays could be performed as an ancillary method to detect an infected animal that fails to produce an immune response against the intradermal reaction (IDR), the official bTB test. In this study, we evaluated the effectiveness of an enzyme-linked immunosorbent assay (ELISA) that uses bovine PPD as a capture antigen as a complement to the IDR in herds with confirmed cases of bTB. The study was conducted in two stages. First, a panel of 200 serum samples was analyzed by ELISA. The sensitivity and specificity obtained were 60% and 99%, respectively. The subsequent stage consisted of evaluating 7,494 bovines from 14 selected dairy farms. The number of animals yielding a IDR negative/ELISA positive result were 200. A necropsy analysis of 33 of these IDR negative/ELISA positive animals revealed that 30 (91%) presented granulomatous lesions and positive M. bovis isolation. This finding confirmed bTB in most cases. Altogether, the results obtained in the present study suggest that the combined use of IDR and ELISA is an effective strategy to improve the control of bTB in endemic herds.

Identification of novel vaccine candidates against cryptosporidiosis of neonatal bovines by reverse vaccinology.

The apicomplexan protozoan Cryptosporidium parvum is an important causative agent of diarrhea of neonatal bovines. Vaccination has been proposed as an advantageous strategy against cryptosporidiosis of calves since besides protection against disease it has also the potential to prevent dissemination of infective oocysts into the environment. Antigens anchored to the parasite surface via glycosylphosphatidylinositol (GPI) are implicated in host cell attachment and invasion and represent promising vaccine candidates. A reverse vaccinology approach was employed to (i) identify the GPI-anchored proteome of C. parvum using available web-based bioinformatic tools and (ii) characterize previously unrecognized novel vaccine antigens. Altogether, 14 putative GPI-anchored proteins could be determined of which CpH1 and CpSUB2 as well as GP60 were further characterized. Sequencing and comparison of GP60, CpH1, and CpSUB1 alleles amplified from different geographic isolates showed a high degree of conservation. All three antigens were recombinant expressed and immunoblotted using sera of 12 Cryptosporidium-infected calves sampled at age periods 1-11 and 12-28 days after birth. Specific antibody reactions against the studied antigens were detected in all analyzed calves, demonstrating their immunreactivity and expression, and recognition in vivo at an early stage of host infection. Besides the acknowledged GP60 vaccinogen, the presented reverse vaccinology approach reveals the additional vaccine candidates CpH1 and CpSUB1 for inclusion into a subunit vaccine formulation.

Interethnic variation of CYP2C19 alleles, 'predicted' phenotypes and 'measured' metabolic phenotypes across world populations.

The present study evaluates the worldwide frequency distribution of CYP2C19 alleles and CYP2C19 metabolic phenotypes ('predicted' from genotypes and 'measured' with a probe drug) among healthy volunteers from different ethnic groups and geographic regions, as well as the relationship between the 'predicted' and 'measured' CYP2C19 metabolic phenotypes. A total of 52 181 healthy volunteers were studied within 138 selected original research papers. CYP2C19*17 was 42- and 24-fold more frequent in Mediterranean-South Europeans and Middle Easterns than in East Asians (P<0.001, in both cases). Contrarily, CYP2C19*2 and CYP2C19*3 alleles were more frequent in East Asians (30.26% and 6.89%, respectively), and even a twofold higher frequency of these alleles was found in Native populations from Oceania (61.30% and 14.42%, respectively; P<0.001, in all cases), which may be a consequence of genetic drift process in the Pacific Islands. Regarding CYP2C19 metabolic phenotype, poor metabolizers (PMs) were more frequent among Asians than in Europeans, contrarily to the phenomenon reported for CYP2D6. A correlation has been found between the frequencies of CYP2C19 poor metabolism 'predicted' from CYP2C19 genotypes (gPMs) and the poor metabolic phenotype 'measured' with a probe drug (mPMs) when subjects are either classified by ethnicity (r=0.94, P<0.001) or geographic region (r=0.99, P=0.002). Nevertheless, further research is needed in African and Asian populations, which are under-represented, and additional CYP2C19 variants and the 'measured' phenotype should be studied.

Subclinical ketosis in dairy cows: prevalence and risk factors in grazing production system.

The purpose of this study was to estimate the prevalence of Subclinical ketosis (SCK) between 4 and 19 days in milk (DIM) in a grazing production system and investigate the importance of potential risk factors for SCK. This cross-sectional study was conducted in dairy cows (n = 107), which had more of two parities. The concentration of ß-hydroxybutyric (BHB) in blood was quantified through a hand-held meter. Potential risk factors evaluated were calving interval (CI), milk yield in previous lactation, metritis, dystocia, calf sex (male), parity (≤3 vs. ≥4) and pre-partum body condition score (BCS ≤ 3.5 vs. ≥3.75). Prevalence of SCK was 10.3% (95% CI 4.7-15) between 4 and 19 DIM. Risk factors identified were the occurrence of both metritis and pre-partum BCS ≥ 3.75. Cows with metritis had 4.9 (95% CI 1.17-20.98) times more risk of developing SCK than cows without metritis. And the cows with pre-partum BCS ≥ 3.75 had 5.25 (95% CI 1.32-21.11) times more risk of developing SCK than cows with pre-partum BCS ≤ 3.5. Metritis could induce a lower feed intake and promote the development of SCK. High pre-partum BCS could induce a greater mobilization of body reserves altering liver function and aggravating post-partum NEB. The results are indicative of the expected prevalence of SCK in grazing production system. Factors associated could help to identify cattle at risk of SCK and improve the management of strategies to limit the effects.

Recurrent lacunar infarction following a previous lacunar stroke: a clinical study of 122 patients.

OBJECTIVE: To determine clinical variables related to recurrent lacunar infarction following a previous lacunar stroke. METHODS: A total of 122 out of 733 consecutive patients with lacunar infarction collected from a hospital based registry between 1986 and 2004 were readmitted because of a recurrent lacunar infarction. In a subset of 59 patients, cognition was assessed using the Mini-Mental State Examination (MMSE). Predictors of lacunar infarction recurrence were assessed by logistic regression analysis. RESULTS: First lacunar infarction recurrence occurred in 101 patients (83%) and multiple recurrences in 21. The mean time between first ever lacunar infarction and recurrent lacunes was 58.3 months (range 2-240). In the subset of 59 patients in whom cognition was studied, cognitive impairment, defined as an MMSE score <24, was detected in 16% (8/49) of patients with first lacunar infarction recurrence and in 40% (4/10) of those with multiple lacunar infarction recurrences. In the multivariate analysis, hypertension (odds ratio 2.01, 95% CI 1.23 to 3.30) and diabetes (odds ratio 1.62, 95% CI 1.07 to 2.46) were significant predictors of lacunar stroke recurrence, whereas hyperlipidaemia was inversely associated (odds ratio 0.52, 95% CI 0.30 to 0.90). CONCLUSIONS: Hypertension and diabetes were significant factors related to recurrent lacunar infarction. Hyperlipidaemia appeared to have a protective role. Cognitive impairment was a frequent finding in patients with multiple lacunar infarction recurrences.