Drosophila melanogaster Y Chromosome Genes Affect Male Sensitivity to Microbial Infections

The genders of Drosophila melanogaster vary in their sensitivities to microbial pathogens. While many of the immunity-related genes are located on the X chromosome, the polymorphisms within the Y chromosome were also shown to affect the immunity of flies. In this study, we investigated the necessity of individual genes on the Y chromosome (Y-genes) for male sensitivity to microbes. We identified several Y-genes whose genetic inactivation either increases or decreases the sensitivity of males to gastrointestinal infections with fungal Saccharomyces cerevisiae and bacterial Serratia liquefaciens. Specifically, the loss of function mutations in fly kl-5 and Ppr-Y Y-genes lead to increased and decreased sensitivity of males to fungal challenge, respectively, compared to female sensitivity. In contrast, mutations in Drosophila Pp1-Y1, kl-5, kl-3, Ppr-Y, CCY, and FDY Y-genes lead to increased sensitivity of males to bacterial infection, compared to females. Moreover, while these Y-genes are necessary, the Y chromosome is not sufficient for the sensitivity of males to microbes, since the sensitivity of XXY females to fungal and bacterial challenges was not different from the sensitivity of wild-type female flies, compared to males. This study assigns a new immunity-related function to numerous Y-genes in D.melanogaster

Identification of clinically approved small molecules that inhibit growth and affect transcript levels of developmentally regulated genes in the African trypanosome.

Trypanosoma brucei are unicellular parasites endemic to Sub-Saharan Africa that cause fatal disease in humans and animals. Infection with these parasites is caused by the bite of the tsetse fly vector, and parasites living extracellularly in the blood of infected animals evade the host immune system through antigenic variation. Existing drugs for Human and Animal African Trypanosomiasis are difficult to administer and can have serious side effects. Resistance to some drugs is also increasing, creating an urgent need for alternative trypanosomiasis therapeutics. We screened a library of 1,585 U.S. or foreign-approved drugs and identified 154 compounds that inhibit trypanosome growth. As all of these compounds have already undergone testing for human toxicity, they represent good candidates for repurposing as trypanosome therapeutics. In addition to identifying drugs that inhibit trypanosome growth, we wished to identify small molecules that can induce bloodstream form parasites to differentiate into forms adapted for the insect vector. These insect stage parasites lack the immune evasion mechanisms prevalent in bloodstream forms, making them vulnerable to the host immune system. To identify drugs that increase transcript levels of an invariant, insect-stage specific surface protein called procyclin, we engineered bloodstream reporter parasites that express Green Fluorescent Protein (GFP) following induction or stabilization of the procyclin transcript. Using these bloodstream reporter strains in combination with automated flow cytometry, we identified eflornithine, spironolactone, and phenothiazine as small molecules that increase abundance of procyclin transcript. Both eflornithine and spironolactone also affect transcript levels for a subset of differentiation associated genes. While we failed to identify compounds that increase levels of procyclin protein on the cell surface, this study is proof of principle that these fluorescent reporter parasites represent a useful tool for future small molecule or genetic screens aimed at identifying molecules or processes that initiate remodeling of the parasite surface during life cycle stage transitions

Antifungal drug repurposing

Control of fungal pathogens is increasingly problematic due to the limited number of effective drugs available for antifungal therapy. Conventional antifungal drugs could also trigger human cytotoxicity associated with the kidneys and liver, including the generation of reactive oxygen species. Moreover, increased incidences of fungal resistance to the classes of azoles, such as fluconazole, itraconazole, voriconazole, or posaconazole, or echinocandins, including caspofungin, anidulafungin, or micafungin, have been documented. Of note, certain azole fungicides such as propiconazole or tebuconazole that are applied to agricultural fields have the same mechanism of antifungal action as clinical azole drugs. Such long-term application of azole fungicides to crop fields provides environmental selection pressure for the emergence of pan-azole-resistant fungal strains such as Aspergillus fumigatus having TR34/L98H mutations, specifically, a 34 bp insertion into the cytochrome P450 51A (CYP51A) gene promoter region and a leucine-to-histidine substitution at codon 98 of CYP51A. Altogether, the emerging resistance of pathogens to currently available antifungal drugs and insufficiency in the discovery of new therapeutics engender the urgent need for the development of new antifungals and/or alternative therapies for effective control of fungal pathogens. We discuss the current needs for the discovery of new clinical antifungal drugs and the recent drug repurposing endeavors as alternative methods for fungal pathogen control

Antifungal drug repurposing

Control of fungal pathogens is increasingly problematic due to the limited number of effective drugs available for antifungal therapy. Conventional antifungal drugs could also trigger human cytotoxicity associated with the kidneys and liver, including the generation of reactive oxygen species. Moreover, increased incidences of fungal resistance to the classes of azoles, such as fluconazole, itraconazole, voriconazole, or posaconazole, or echinocandins, including caspofungin, anidulafungin, or micafungin, have been documented. Of note, certain azole fungicides such as propiconazole or tebuconazole that are applied to agricultural fields have the same mechanism of antifungal action as clinical azole drugs. Such long-term application of azole fungicides to crop fields provides environmental selection pressure for the emergence of pan-azole-resistant fungal strains such as Aspergillus fumigatus having TR34/L98H mutations, specifically, a 34 bp insertion into the cytochrome P450 51A (CYP51A) gene promoter region and a leucine-to-histidine substitution at codon 98 of CYP51A. Altogether, the emerging resistance of pathogens to currently available antifungal drugs and insufficiency in the discovery of new therapeutics engender the urgent need for the development of new antifungals and/or alternative therapies for effective control of fungal pathogens. We discuss the current needs for the discovery of new clinical antifungal drugs and the recent drug repurposing endeavors as alternative methods for fungal pathogen control

Anthrax toxin component, Protective Antigen, protects insects from bacterial infections.

Anthrax is a major zoonotic disease of wildlife, and in places like West Africa, it can be caused by Bacillus anthracis in arid nonsylvatic savannahs, and by B. cereus biovar anthracis (Bcbva) in sylvatic rainforests. Bcbva-caused anthrax has been implicated in as much as 38% of mortality in rainforest ecosystems, where insects can enhance the transmission of anthrax-causing bacteria. While anthrax is well-characterized in mammals, its transmission by insects points to an unidentified anthrax-resistance mechanism in its vectors. In mammals, a secreted anthrax toxin component, 83 kDa Protective Antigen (PA83), binds to cell-surface receptors and is cleaved by furin into an evolutionary-conserved PA20 and a pore-forming PA63 subunits. We show that PA20 increases the resistance of Drosophila flies and Culex mosquitoes to bacterial challenges, without directly affecting the bacterial growth. We further show that the PA83 loop known to be cleaved by furin to release PA20 from PA63 is, in part, responsible for the PA20-mediated protection. We found that PA20 binds directly to the Toll activating peptidoglycan-recognition protein-SA (PGRP-SA) and that the Toll/NF-κB pathway is necessary for the PA20-mediated protection of infected flies. This effect of PA20 on innate immunity may also exist in mammals: we show that PA20 binds to human PGRP-SA ortholog. Moreover, the constitutive activity of Imd/NF-κB pathway in MAPKK Dsor1 mutant flies is sufficient to confer the protection from bacterial infections in a manner that is independent of PA20 treatment. Lastly, Clostridium septicum alpha toxin protects flies from anthrax-causing bacteria, showing that other pathogens may help insects resist anthrax. The mechanism of anthrax resistance in insects has direct implications on insect-mediated anthrax transmission for wildlife management, and with potential for applications, such as reducing the sensitivity of pollinating insects to bacterial pathogens