Comparative Analysis on Transcriptomics of Ivermectin Resistant and Sensitive Strains of Haemonchus Contortus

Background: Ivermectin (IVM) is one of the most important and widely used anthelmintics in veterinary medicine. However, its efficacy is increasingly compromised by widespread resistance, and the exact mechanism of IVM resistance remains unclear for most parasitic nematodes including Haemonchus contortus, a blood-sucking parasitic nematode of small ruminants.Methods: In this study, we isolated and assessed an IVM resistant strain from Zhaosu, Xinjiang, China. Subsequently, the comparative analyses on transcriptomics of IVM susceptible and resistant H. contortus adult worms were carried out using RNA sequencing and bioinformatics.Results: In total, 543 and 359 differentially expressed genes (DEGs) were identified in male and female adult worms of the resistant strain compared with the susceptible strain, respectively. The DEGs encode molecules involved in receptor activities, transport, detoxification, lipid metabolism and cuticle collagen formation. In addition, Gene Ontology (GO) analysis revealed that transcriptional changes were dominant in genes associated with ligand-gated channel activity, oxidation-reduction process, lipid metabolic process, and structural constituent of cuticle. The results support previous proposal that the IVM resistant mechanism of H. contortus involved in both neuromuscular and non-neuromuscular pathways. Finally, the quantitative RT-PCR results confirmed that the transcriptional profiles of selected DEGs (male: 8 genes, female: 10 genes) were consistent with those obtained by the RNA-Seq.Conclusions: The findings from this work provided valuable information for further studies on the IVM resistance in H. contortus.

Deltamethrin interacts with Culex quinquefasciatus odorant-binding protein: a novel potential resistance mechanism

Background Odorant-binding proteins (OBPs) play important roles in many physiological processes of mosquitoes. Previous high-throughput sequencing studies have revealed that some OBPs of Culex quinquefasciatus might be involved in the development of resistance to insecticides. Methods Based on the results of sequencing analyses, the OBP28 gene was selected for evaluation in this study. Three laboratory strains of Cx. quinquefasciatus [susceptible strain (SS), deltamethrin-resistant strain 1 (HN) and deltamethrin-resistant strain 2 (RR)] were first examined by using the Centers for Disease Control and Prevention bottle bioassay, after which the expression level of the OBP28 gene in the susceptible and deltamethrin-resistant strains was determined by real-time quantitative polymerase chain reaction. The OBP28 gene in deltamethrin-resistant strain RR was silenced using RNA interference technology. The expression level of OBP28 and the resistance level were tested in the silenced strain and control strain after microinjection of double-stranded RNA for a 48-h interference period. Four field-collected strains (henceforth ‘field strains’) of Cx. quinquefasciatus were also examined for their resistance to deltamethrin and levels of OBP28 expression. Finally, a correlation analysis between deltamethrin resistance and gene expression was carried out for all seven strains, i.e. the four field strains and the three laboratory strains. Results In the bioassay, the mortality of SS, HN and RR was 100%, 21.33% and 1.67%, respectively. The relative expression levels of OBP28 in strains HN and RR were 6.30- and 6.86-fold higher, respectively, than that of strain SS. After silencing of the OBP28 gene, the mortality of strain RR was 72.20% and that of the control strain 26.32%. The mortality of strain RR increased significantly after interference compared to that of the control strain. There was a negative correlation between OBP28 gene expression and mortality in adult mosquitoes after exposure to deltamethrin. Conclusions To our knowledge, this study shows for the first time a correlation between the expression of a gene coding for OBP and insecticide resistance in mosquitoes. The potential resistance mechanism that was elucidated provides a new target gene for the surveillance of resistance in mosquitoes. Graphical abstract

The Integrative and Conjugative Element ICE Csp POL2 Contributes to the Outbreak of Multi-Antibiotic-Resistant Bacteria for Chryseobacterium Spp. and Elizabethkingia Spp.

Infections with multiple antibiotic-resistant Chryseobacterium and Elizabethkingia in intensive care units have been increasing in recent years. In this study, the mobile integrative and conjugative element ICE Csp POL2, which was associated with the transmission of a carbapenem resistance gene, was identified in the genome of the multi-antibiotic-resistant strain Chryseobacterium sp.

Transcriptomic analysis of resistance and short-term induction response to pyrethroids, in Anopheles coluzzii legs

Background Insecticide-treated bed nets and indoor residual spraying comprise the major control measures against Anopheles gambiae sl, the dominant vector in sub-Saharan Africa. The primary site of contact with insecticide is through the mosquitoes’ legs, which represents the first barrier insecticides have to bypass to reach their neuronal targets. Proteomic changes and leg cuticle modifications have been associated with insecticide resistance that may reduce the rate of penetration of insecticides. Here, we performed a multiple transcriptomic analyses focusing on An. coluzzii legs. Results Firstly, leg-specific enrichment analysis identified 359 genes including the pyrethroid-binder SAP2 and 2 other chemosensory proteins, along with 4 ABCG transporters previously shown to be leg enriched. Enrichment of gene families included those involved in detecting chemical stimuli, including gustatory and ionotropic receptors and genes implicated in hydrocarbon-synthesis. Subsequently, we compared transcript expression in the legs of a highly resistant strain (VK7-HR) to both a strain with very similar genetic background which has reverted to susceptibility after several generations without insecticide pressure (VK7-LR) and a lab susceptible population (NG). Two hundred thirty-two differentially expressed genes (73 up-regulated and 159 down-regulated) were identified in the resistant strain when compared to the two susceptible counterparts, indicating an over-expression of phase I detoxification enzymes and cuticular proteins, with decrease in hormone-related metabolic processes in legs from the insecticide resistant population. Finally, we analysed the short-term effect of pyrethroid exposure on An. coluzzii legs, comparing legs of 1 h-deltamethrin-exposed An. coluzzii (VK7-IN) to those of unexposed mosquitoes (VK7-HR) and identified 348 up-regulated genes including those encoding for GPCRs, ABC transporters, odorant-binding proteins and members of the divergent salivary gland protein family. Conclusions The data on An. coluzzii leg-specific transcriptome provides valuable insights into the first line of defense in pyrethroid resistant and short-term deltamethrin-exposed mosquitoes. Our results suggest that xenobiotic detoxification is likely occurring in legs, while the enrichment of sensory proteins, ABCG transporters and cuticular genes is also evident. Constitutive resistance is primarily associated with elevated levels of detoxification and cuticular genes, while short-term insecticide-induced tolerance is linked with overexpression of transporters, GPCRs and GPCR-related genes, sensory/binding and salivary gland proteins.

Synthesis and Evaluation of Trypanocidal Activity of Chromane-Type Compounds and Acetophenones

American trypanosomiasis (Chagas disease) caused by the Trypanosoma cruzi parasite, is a severe health problem in different regions of Latin America and is currently reported to be spreading to Europe, North America, Japan, and Australia, due to the migration of populations from South and Central America. At present, there is no vaccine available and chemotherapeutic options are reduced to nifurtimox and benznidazole. Therefore, the discovery of new molecules is urgently needed to initiate the drug development process. Some acetophenones and chalcones, as well as chromane-type substances, such as chromones and flavones, are natural products that have been studied as trypanocides, but the relationships between structure and activity are not yet fully understood. In this work, 26 compounds were synthesized to determine the effect of hydroxyl and isoprenyl substituents on trypanocide activity. One of the compounds showed interesting activity against a resistant strain of T. cruzi, with a half effective concentration of 18.3 µM ± 1.1 and an index of selectivity > 10.9.

Susceptibility monitoring and the molecular characterization of resistance of Spodoptera frugiperda (Lepidoptera: Noctuidae) to lambda-cyhalothrin and chlorpyrifos

Spodoptera frugiperda (J. E. Smith) is a serious and widespread agricultural pest with several records of resistance to different insecticides and Bt proteins, including the neurotoxic insecticides chlorpyrifos (organophosphate) and lambda-cyhalothrin (pyrethroid). In this study, we (i) characterized and monitored the susceptibility of field populations of S. frugiperda to chlorpyrifos (194 populations) and lambda-cyhalothrin (197 populations) collected from major maize-growing regions of Brazil from 2003 to 2016, and (ii) compared gene expression levels of chlorpyrifos- and lambda-cyhalothrin-resistant strains to a susceptible reference strain (Sf-ss) of S. frugiperda. Laboratory-guided assays to monitor larval susceptibility detected average survival ranging from 29.3% to 36.0% to chlorpyrifos, and 23.1% to 68.0% to lambda-cyhalothrin at diagnostic concentration, based on LC99 of the susceptible reference strain of each insecticide. The resistance ratio of the chlorpyrifos-resistant strain (Clo-rr) was 25.4-fold and of the lambda-cyhalothrin-resistant strain (Lam-rr) was 217-fold. Differential gene expression analyses between resistant vs susceptible strains identified 1,098 differentially expressed genes (DEGs) between Clo-rr and Sf-ss, and 303 DEGs between Lam-rr and Sf-ss. Functional analyses of the DEGs revealed the up-regulation of several detoxification enzymes, mainly cytochrome P450 belonging to the CYP3 and CYP6 clans. Genes associated with regulatory processes, such as the forkhead box O (FoxO) were also up-regulated. Our data points that the resistance mechanisms of Clo-rr and Lam-rr strains of S. frugiperda to chlorpyrifos and lambda-cyhalothrin are mainly mediated by enzyme detoxification.

Malathion-resistant Tribolium castaneum has enhanced response to oxidative stress, immunity, and fitness

Many cases of insecticide resistance in insect pests give resulting no-cost strains that retain the resistance genes even in the absence of the toxic stressor. Malathion has been widely used against the red flour beetle, Tribolium castaneum Herbst. in stored products although no longer used. Malathion specific resistance in this pest has provided resistance that is long lasting and widely distributed. To understand this resistance a malathion resistant strain was challenged with a range of stressors including starvation, hyperoxia, malathion and a pathogen and the antioxidant responses and some lifecycle parameters were determined. Adult life span of malathion-specific resistant strain of T. castaneum was significantly shorter than the susceptible. Starvation and/or high oxygen reduced adult life span of both strains. Starving with and without 100% oxygen gave longer lifespan for the resistant strain, but for oxygen alone there was no difference. Under oxygen the proportional survival of the resistant strain to the adult stage was significantly higher, for both larvae and pupae, than the susceptible. The resistant strain when stressed with malathion and/or oxygen significantly increased catalase activity, but the susceptible did not. The resistant strain stressed with Paranosema whitei infection had significantly higher survival compared to the susceptible, and with almost no mortality. The malathion resistant strain of T. castaneum showed greater vigour than the susceptible in most oxidative stress situations and especially where stressors were combined. The induction of the antioxidant enzyme catalase could have helped the resistant strain to withstand oxidative stresses, including insecticidal and importantly those from pathogens. These adaptations, in the absence of insecticide, seem to support the increased immunity of host insects to pathogens seen in other insect species, such as mosquitoes. By increasing the responses to a range of stressors the resistant strain could be considered as having enhanced fitness.

Discovery of New Ginsenol-like Compounds with High Antiviral Activity

A number of framework amides with a ginsenol backbone have been synthesized using the Ritter reaction. We named the acetamide as Ginsamide. A method was developed for the synthesis of the corresponding amine and thioacetamide. The new compounds revealed a high activity against H1N1 influenza, which was confirmed using an animal model. Biological experiments were performed to determine the mechanism of action of the new agents, a ginsamide-resistant strain of influenza virus was obtained, and the pathogenicity of the resistant strain and the control strain was studied. It was shown that the emergence of resistance to Ginsamide was accompanied by a reduction in the pathogenicity of the influenza virus.