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.

Adiponectin/leptin ratio and insulin resistance in pregnancy.

OBJECTIVE: Pregnancy is characterized by progressive insulin resistance. The present study evaluated whether the adiponectin/leptin ratio is associated with insulin resistance in pregnancy, since this ratio has been shown to be associated with insulin resistance in obesity, type 2 diabetes mellitus, metabolic syndrome and polycystic ovary syndrome. METHODS: In this cross-sectional study, adiponectin and leptin concentrations were measured in pregnant women using enzyme-linked immunosorbent assays. Insulin resistance was assessed using the homeostasis model assessment of insulin resistance (HOMA-IR). RESULTS: Mean ± SD age of the participants (n = 74) was 30.76 ± 4.27 years, mean ± SD gestational age was 26.81 ± 3.52 weeks and median body mass index (BMI) before pregnancy was 22.68 kg/m(2) (interquartile range 20.75-26.79 kg/m(2)). There was a significant correlation between the HOMA-IR and leptin concentration, but not between the HOMA-IR and adiponectin concentration. There was a significant inverse correlation between the HOMA-IR and adiponectin/leptin ratio. The adiponectin/leptin ratio was inversely correlated with BMI before pregnancy. CONCLUSION: The adiponection/leptin ratio inversely correlates with HOMA-IR in pregnancy.

Adipocytokines and insulin resistance across various degrees of glucose tolerance in pregnancy.

OBJECTIVE: Gestational diabetes mellitus is characterized by progressive insulin resistance. Adipocytokines are thought to be associated with insulin resistance. This cross-sectional study evaluated the associations between serum concentrations of several adipocytokines and insulin resistance at different stages of glucose tolerance in pregnancy, using the homeostasis model assessment of insulin resistance (HOMA-IR) as a reference. METHODS: According to oral glucose tolerance test results, 74 pregnant women were divided into three groups: normal glucose tolerance (n = 25); intermediate glucose tolerance (n = 19); gestational diabetes mellitus (n = 30). Adiponectin, leptin, resistin, visfatin and retinol-binding protein 4 (RBP4) concentrations were measured using enzyme-linked immuno sorbent assays. RESULTS: Groups were comparable regarding age, week of gestation and body mass index before gestation. There were statistically significant between-group differences in HOMA-IR, but no significant differences regarding serum adipocytokine concentrations. CONCLUSIONS: Adipo nectin, leptin, resistin, visfatin and RBP4 were not associated with the degree of glucose tolerance in pregnancy. Concentrations of these adipocytokines are not sufficiently sensitive to replace HOMA- IR in pregnancy.

Comparison of alterations in insulin signalling pathway in adipocytes from Type II diabetic pregnant women and women with gestational diabetes mellitus.

AIMS/HYPOTHESIS: The cellular mechanisms for the insulin resistance in pregnancy and gestational diabetes mellitus are not known. The membrane protein plasma cell glycoprotein PC-1 has been identified as an inhibitor of insulin receptor tyrosine kinase activity and could have a role in insulin resistance. This study aimed to examine the effects of insulin on glucose transport and changes in insulin receptor tyrosine phosphorylation, IRS-1 and PC-1. METHODS: Adipocytes were obtained either during elective cesarean section from three groups of subjects (Type II diabetic pregnant women ( n=6) women with gestational diabetes mellitus ( n=10) and pregnant women with normal glucose tolerance ( n=6) as pregnant control subjects) or during elective gynaecological surgery from non-pregnant ( n=6) control subjects. RESULTS: Insulin stimulated glucose transport was reduced by 50% in women with gestational diabetes mellitus and 70% in pregnant women with Type II diabetes, compared to the non-pregnant control subjects. After maximal insulin stimulation of adipocytes, IRTK phosphorylation was reduced by 29.5% in women with gestational diabetes mellitus and 44.5% in women with Type II diabetes, compared to the non-pregnant control subjects. We also found that IRS-1 phosphorylation was reduced by 32% and 48%, respectively. On the other hand, PC-1 content in adipocytes in women with gestational diabetes mellitus increased by 320% and 668% in Type II diabetic women, compared to the non-pregnant control subjects. CONCLUSIONS/INTERPRETATION: Our results indicate that women with gestational diabetes mellitus and Type II diabetes have increased PC-1 content and suggest that this could contribute to lower phosphorylation levels of IRTK and IRS-1. Furthermore, these postreceptor defects in insulin signalling pathway are greater in both groups compared to the women with normal pregnancy. However, results from women with Type II diabetes show that pre-existing insulin resistance lead to an even greater deterioration of the signalling pathway.

Romano-Ward syndrome: case report, family study and signal averaged electrocardiogram.

A family with the Romano-Ward syndrome is described. A 28-year-old woman had episodes of syncope due to self-terminating ventricular tachycardia torsades de pointes. During lidocaine treatment she developed a sustained ventricular tachycardia and cardiopulmonary resuscitation was necessary. Propranolol reduced ventricular ectopic activity, but the QTc interval remained prolonged. A "threshold" in QT duration for the generation of complex ventricular arrhythmias was observed in the patient. There were no late ventricular potentials noted in the signal averaged ECG in the patient and in members of her family.