Human louse Pediculus humanus is a cosmopolitan obligatory blood-feeding ectoparasite causing pediculosis and transmitting many bacterial pathogens. Control of infestation is difficult due to the developed resistance to insecticides that mainly target GABA (γ-aminobutyric acid) receptors. Previous work showed that Pediculus humanus humanus (Phh) GABA receptor subunit resistance to dieldrin (RDL) is the target of lotilaner, a synthetic molecule of the isoxazoline chemical class. To enhance our understanding of how insecticides act on GABA receptors, two other GABA receptor subunits were cloned and characterized: three variants of Phh-grd (glycine-like receptor of Drosophila) and one variant of Phh-lcch3 (ligand-gated chloride channel homolog 3). Relative mRNA expression levels of Phh-rdl, Phh-grd, and Phh-lcch3 revealed that they were expressed throughout the developmental stages (eggs, larvae, adults) and in the different parts of adult lice (head, thorax, and abdomen). When expressed individually in the Xenopus oocyte heterologous expression system, Phh-GRD1, Phh-GRD2, Phh-GRD3, and Phh-LCCH3 were unable to reconstitute functional channels, whereas the subunit combinations Phh-GRD1/Phh-LCCH3, Phh-GRD1/Phh-RDL, and Phh-LCCH3/Phh-RDL responded to GABA in a concentration-dependent manner. The three heteromeric receptors were similarly sensitive to the antagonistic effect of picrotoxin and fipronil, whereas Phh-GRD1/Phh-RDL and Phh-LCCH3/Phh-RDL were respectively about 2.5-fold and 5-fold more sensitive to ivermectin than Phh-GRD1/Phh-LCCH3. Moreover, the heteropentameric receptor constituted by Phh-GRD1/Phh-LCCH3 was found to be permeable and highly sensitive to the extracellular sodium concentration. These findings provided valuable additions to our knowledge of the complex nature of GABA receptors in human louse that could help in understanding the resistance pattern to commonly used pediculicides. SIGNIFICANCE STATEMENT Human louse is an ectoparasite that causes pediculosis and transmits several bacterial pathogens. Emerging strains developed resistance to the commonly used insecticides, especially those targeting GABA receptors. To understand the molecular mechanisms underlying this resistance, two subunits of GABA receptors were cloned and described: Phh-grd and Phh-lcch3. The heteromeric receptor reconstituted with the two subunits was functional in Xenopus oocytes and sensitive to commercially available insecticides. Moreover, both subunits were transcribed throughout the parasite lifecycle.

Introduction. Cryptococcosis is a ubiquitous opportunistic fungal disease caused by Cryptococcus neoformans var. grubii. It has high global morbidity and mortality among HIV patients and non-HIV carriers with 99% and 95%, respectively. Furthermore, the increasing prevalence of undesired toxicity profile of antifungal, multidrug-resistant organisms and the scarcity of FDA-authorized vaccines were the hallmark in the present days. This study was undertaken to design a reliable epitope-based peptide vaccine through targeting highly conserved immunodominant heat shock 70 kDa protein of Cryptococcus neoformans var. grubii that covers a considerable digit of the world population through implementing a computational vaccinology approach. Materials and Methods. A total of 38 sequences of Cryptococcus neoformans var. grubii’s heat shock 70 kDa protein were retrieved from the NCBI protein database. Different prediction tools were used to analyze the aforementioned protein at the Immune Epitope Database (IEDB) to discriminate the most promising T-cell and B-cell epitopes. The proposed T-cell epitopes were subjected to the population coverage analysis tool to compute the global population’s coverage. Finally, the T-cell projected epitopes were ranked based on their binding scores and modes using AutoDock Vina software. Results and Discussion. The epitopes (ANYVQASEK, QSEKPKNVNPVI, SEKPKNVNPVI, and EKPKNVNPVI) had shown very strong binding affinity and immunogenic properties to B-cell. (FTQLVAAYL, YVYDTRGKL) and (FFGGKVLNF, FINAQLVDV, and FDYALVQHF) exhibited a very strong binding affinity to MHC-I and MHC-II, respectively, with high population coverage for each, while FYRQGAFEL has shown promising results in terms of its binding profile to MHC-II and MHC-I alleles and good strength of binding when docked with HLA-C12:03. In addition, there is massive global population coverage in the three coverage modes. Accordingly, our in silico vaccine is expected to be the future epitope-based peptide vaccine against Cryptococcus neoformans var. grubii that covers a significant figure of the entire world citizens.

The human louse (Pediculus humanus) is an obligatory blood feeding ectoparasite with two ecotypes: the human body louse (Pediculus humanus humanus), a competent vector of several bacterial pathogens, and the human head louse (Pediculus humanus capitis), responsible for pediculosis and affecting millions of people around the globe. GABA (γ-aminobutyric acid) receptors, members of the cys-loop ligand gated ion channel superfamily, are among the main pharmacological targets for insecticides. In insects, there are four subunits of GABA receptors: resistant-to-dieldrin (RDL), glycin-like receptor of drosophila (GRD), ligand-gated chloride channel homologue3 (LCCH3), and 8916 are well described and form distinct phylogenetic clades revealing orthologous relationships. Our previous studies in the human body louse confirmed that subunits Phh-RDL, Phh-GRD, and Phh-LCCH3 are well clustered in their corresponding clades. In the present work, we cloned and characterized a putative new GABA receptor subunit in the human body louse that we named HoCas, for Homologous to Cys-loop α like subunit. Extending our analysis to arthropods, HoCas was found to be conserved and clustered in a new (fifth) phylogenetic clade. Interestingly, the gene encoding this subunit is ancestral and has been lost in some insect orders. Compared to the other studied GABA receptor subunits, HoCas exhibited a relatively higher expression level in all development stages and in different tissues of human body louse. These findings improved our understanding of the complex nature of GABA receptors in Pediculus humanus and more generally in arthropods.

Human louse (Pediculus humanus) is an obligatory blood feeding ectoparasite with two ecotypes: human body louse (Pediculus humanus humanus), a competent vector of several bacterial pathogens and the human head louse (Pediculus humanus capitis) responsible for pediculosis and affecting millions of people around the globe. GABA (γ-aminobutyic acid) receptors, members of the cys-loop ligand gated ion channels superfamily, are among the main pharmacological targets for insecticides. In insects, four subunits of GABA receptors: resistant to dieldrin (RDL), glycin like receptor of drosophila (GRD), ligand gated chloride channel homologue3 (LCCH3), and 8916 are well described and form distinct phylogenetic clades revealing orthologous relationships. Our previous studies in human body louse confirmed that subunits Phh-RDL, Phh-GRD and Phh-LCCH3 are well clustered in their corresponding clades. In the present work, we cloned and characterized a putative new GABA receptor subunit in human body louse that we named HoCas for Homologous to Cys-loop alpha like subunit. Extending our analysis to arthropods, HoCas was found to be conserved and clustered in a new (fifth) phylogenetic clade. Interestingly, the gene encoding this subunit is ancestral and has been lost in some insect orders. Compared to the other studied GABA receptor subunits, HoCas exhibited a relatively higher expression level in all development stages and in different tissues of human body louse. These findings improved our understanding of the complex nature of GABA receptors in Pediculus humanus and more generally in arthropods.