Cloning and expression of Rift Valley fever virus nucleocapsid (N) protein and evaluation of a N-protein based indirect ELISA for the detection of specific IgG and IgM antibodies in domestic ruminants

Serodiagnosis of Rift Valley fever (RVF) currently relies on the use of live or inactivated whole virus as antigens. The recombinant nucleocapsid (N) protein of RVF virus was tested for diagnostic applicability in an indirect enzyme-linked immunosorbent assay (I-ELISA), using sera from experimentally infected sheep (n=128), vaccinated sheep (n=240), and field-collected sera from sheep (n=251), goats (n=362) and cattle (n=100). The N-protein based I-ELISA performed at least as good as VN and HI tests. In goat the diagnostic sensitivity (D-Sn) and specificity (D-Sp) of the I-ELISA was 100% when using the anti-species IgG conjugate. Using protein G as a detection system, the D-Sn and D-Sp in goats were 99.4% and 99.5%, in sheep field sera both 100%, in cattle 100% and 98.3%, respectively. The I-ELISA based on recombinant N-protein has the potential to complement the traditional assays for serodiagnosis of RVF. Advantages of the N-protein are its safety, stability and cost-effectiveness in use and production.     

Metabolomic fingerprint of cabbage resistance to Mamestra brassicae L. (Lepidoptera: Noctuidae)

BACKGROUND

Plants defend themselves from insect feeding by activating specific metabolic pathways. We performed a metabolomic analysis to compare the metabolome reorganization that occurs in the leaves of two genotypes of cabbage (one partially resistant and one susceptible) when attacked by Mamestra brassicae caterpillars.

RESULTS

The comparison of the metabolomic reorganization of both genotypes allowed us to identify 43 metabolites that are specifically associated with the insect feeding response in the resistant genotype. Of these, 19% are lipids or lipid-related compounds, most of which are modified fatty acids. These include glycosylated, glycerol-binding and oxidized fatty acids, the majority being associated with the oxylipin pathway. Some of the identified lipids are unlikely to be produced by plants and may be the result of biochemical reactions in the caterpillar oral secretions. A further 16% are phenylpropanoids. Interestingly, some phenylpropanoids were not present in the susceptible genotype, making them possible candidates for specific resistance-related compounds.

CONCLUSION

Our results suggest that glucosinolates do not have a clear role in the resistance to M. brassicae feeding on cabbage. Using an untargeted metabolomics approach, we associated the regulation of metabolic pathways related to lipid signalling and phenylpropanoid compounds with the resistance to this pest. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Effects of intravenous administration of neurokinin receptor subtype-selective agonists on gonadotropin-releasing hormone pulse generator activity and luteinizing hormone secretion in goats

Recent evidence suggests that neurokinin B (NKB), a member of the neurokinin (tachykinin) peptide family, plays a pivotal role in gonadotropin-releasing hormone (GnRH) pulse generation. Three types of neurokinin receptors (NKRs), NK1R, NK2R and NK3R, are found in the brain. Although NKB preferentially binds to NK3R, other NKRs are possibly also involved in NKB action. The present study examined the effects of intravenous administration of the NKR subtype-selective agonists GR73632 (NK1R), GR64349 (NK2R), and senktide (NK3R) on GnRH pulse generator activity and luteinizing hormone (LH) secretion. Multiple-unit activity (MUA) was monitored in ovariectomized goats (n = 5) implanted with recording electrodes. Characteristic increases in MUA (MUA volleys) were considered GnRH pulse generator activity. Although three NKR agonists dose-dependently induced an MUA volley and an accompanying increase in LH secretion, the efficacy in inducing the volley markedly differed. As little as 10 nmol of senktide induced an MUA volley in all goats, whereas a dose of 1000 nmol was only effective for the NK1R and NK2R agonists in two and four goats, respectively. When the treatment failed to evoke an MUA volley, no apparent change was observed in the MUA or LH secretion. Similar effects of the NK2R and NK3R agonists were observed in the presence of estradiol. The results demonstrated that NK3R plays a predominant role in GnRH pulse generation and suggested that the contributions of NK1R and NK2R to this mechanism may be few, if any, in goats.     

Marinopyrrole Derivatives as Potential Antibiotic Agents against Methicillin-Resistant Staphylococcus aureus (III)

The marine natural product, marinopyrrole A (1), was previously shown to have significant antibiotic activity against Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA). Although compound (1) exhibits a significant reduction in MRSA activity in the presence of human serum, we have identified key modifications that partially restore activity. We previously reported our discovery of a chloro-derivative of marinopyrrole A (1a) featuring a 2–4 fold improved minimum inhibitory concentration (MIC) against MRSA, significantly less susceptibility to serum inhibition and rapid and concentration-dependent killing of MRSA. Here, we report a novel fluoro-derivative of marinopyrrole A (1e) showing an improved profile of potency, less susceptibility to serum inhibition, as well as rapid and concentration-dependent killing of MRSA.     

Changes in Phenolic Compounds in Garlic (<Emphasis Type="Italic">Allium sativum</Emphasis> L.) Owing to the Cultivar and Location of Growth

The contents of total phenolic compounds, flavonoids, and phenolic acids were determined in selected garlic cultivars grown at four locations. The total phenolic content varied from 3.4 mg gallic acid equivalents (GAE)/g of dry matter (dm) to 10.8 mg GAE/g of dm with a mean value of 6.5 mg GAE/g of dm. The myricetin, quercetin, kaempferol, and apigenin flavonoids were not detected in any of the samples. Caffeic acid and ferulic acid were the major phenolic acids found with mean values of 2.9 mg/kg of dm and 2.6 mg/kg of dm, respectively. The mean contents of vanillic, p-hydroxybenzoic, and p-coumaric acids were comparable (0.4–0.8 mg/kg of dm), and the level of sinapic acid was negligible (< 0.1 mg/kg of dm). There was a significant effect of location but an insignificant effect of genotype on contents of caffeic, vanillic, p-hydroxybenzoic, and p-coumaric acids. However, genotype but not location affected the contents of total phenolics and ferulic acid. On average, the white garlic cultivars and Chinese garlic cultivars contained higher contents of total phenolics and ferulic acid than the purple garlic cultivars. However, the differences in the total phenolic content between the purple and white garlic cultivars were not significant.     

Design of new α-conotoxins: from computer modeling to synthesis of potent cholinergic compounds

A series of 14 new analogs of α-conotoxin PnIA Conus pennaceus was synthesized and tested for binding to the human α7 nicotinic acetylcholine receptor (nAChR) and acetylcholine-binding proteins (AChBP) Lymnaea stagnalis and Aplysia californica. Based on computer modeling and the X-ray structure of the A. californica AChBP complex with the PnIA[A10L, D14K] analog, single and multiple amino acid substitutions were introduced in α-conotoxin PnIA aimed at compounds of higher affinity and selectivity. Three analogs, PnIA[L5H], PnIA[A10L, D14K] and PnIA[L5R, A10L, D14R], have high affinities for AChBPs or α7 nAChR, as found in competition with radioiodinated α-bungarotoxin. That is why we prepared radioiodinated derivatives of these α-conotoxins, demonstrated their specific binding and found that among the tested synthetic analogs, most had almost 10-fold higher affinity in competition with radioactive α-conotoxins as compared to competition with radioactive α-bungarotoxin. Thus, radioiodinated α-conotoxins are a more sensitive tool for checking the activity of novel α-conotoxins and other compounds quickly dissociating from the receptor complexes.