Functional studies of Plasmodium falciparum putative SURF1 in Saccharomyces cerevisiae.

BACKGROUND AND OBJECTIVES: The mitochondrial electron transport chain (mtETC) of Plasmodium falciparum is an important drug target. Identification and functional validation of putative mitochondrial proteins of the mtETC is critical for drug development. Many of the regulatory subunits and assembly factors of cytochrome c oxidase readily identifiable in humans and yeast are missing in P. falciparum. Here, we describe our efforts to identify and validate the function of putative Pfsurf1, a key assembly factor of complex IV of the mtETC. METHODS: Multiple sequence alignment of SURF 1/Shy 1 was carried out in Clustal X 2.1. Phylogenetic tree was constructed using "Draw tree" option in Clustal X, and was analyzed using interactive Tree of Life software. To identify the conserved sequences, domain search was done using Jalview version 2.8.2 (BLOSUM 62 scoring). The haploid Saccharomyces cerevisiae strain (BY4741) containing the null allele shy1 (Orf: YGR112w) (shy1::Kan) was complemented with putative Pfsurf1 to study its ability to rescue the growth defect. RESULTS: Similarity searches of PfSURF1-like protein in the Pfam shows statistically significant E = 4.7e-10 match to SURF1 family. Sequence alignment of PfSURF1 with other SURF1-like proteins reveals the conservation of transmembrane domains, α-helices and ß-pleated sheets. Phylogenetic analysis clusters putative PfSURF1 with apicomplexan SURF1-like proteins. Yeast complementation studies show that Pfsurf1 can partially rescue the yeast shy1 mutant, YGR112w. INTERPRETATION & CONCLUSION: Bioinformatics and complementation studies in yeast show that P. falciparum's SURF1 is the functional ortholog of human SURF1 and yeast Shy1.

Hydroxychloroquine as prophylaxis or treatment for COVID-19: What does the evidence say?

Hydroxychloroquine (HCQ), an antimalarial has been proposed as possible treatment for coronavirus disease-2019 (COVID-19). India has approved the use of HCQ for prophylaxis of asymptomatic health workers treating suspected or confirmed COVID-19 cases, and asymptomatic household contacts of confirmed patients. The U.S. Food and Drug Administration has issued Emergency Use Authorization for the use of HCQ to treat COVID-19 in adolescents and adults. In this review, we go over the available evidence for and against HCQ's use as prophylaxis or treatment for COVID-19, especially in the Indian context.

Susceptibility of wild and colonized Anopheles stephensi to Plasmodium vivax infection.

BACKGROUND: As much as 80% of global Plasmodium vivax infections occur in South Asia and there is a shortage of direct studies on infectivity of P. vivax in Anopheles stephensi, the most common urban mosquito carrying human malaria. In this quest, the possible effects of laboratory colonization of mosquitoes on infectivity and development of P. vivax is of interest given that colonized mosquitoes can be genetically less divergent than the field population from which they originated. METHODS: Patient-derived P. vivax infected blood was fed to age-matched wild and colonized An. stephensi. Such a comparison requires coordinated availability of same-age wild and colonized mosquito populations. Here, P. vivax infection are studied in colonized An. stephensi in their 66th-86th generation and fresh field-caught An. stephensi. Wild mosquitoes were caught as larvae and pupae and allowed to develop into adult mosquitoes in the insectary. Parasite development to oocyst and sporozoite stages were assessed on days 7/8 and 12/13, respectively. RESULTS: While there were batch to batch variations in infectivity of individual patient-derived P. vivax samples, both wild and colonized An. stephensi were roughly equally susceptible to oocyst stage Plasmodium infection. At the level of sporozoite development, significantly more mosquitoes with sporozoite load of 4+ were seen in wild than in colonized populations.

ATP synthase complex of Plasmodium falciparum: dimeric assembly in mitochondrial membranes and resistance to genetic disruption.

The rotary nanomotor ATP synthase is a central player in the bioenergetics of most organisms. Yet the role of ATP synthase in malaria parasites has remained unclear, as blood stages of Plasmodium falciparum appear to derive ATP largely through glycolysis. Also, genes for essential subunits of the F(O) sector of the complex could not be detected in the parasite genomes. Here, we have used molecular genetic and immunological tools to investigate the localization, complex formation, and functional significance of predicted ATP synthase subunits in P. falciparum. We generated transgenic P. falciparum lines expressing seven epitope-tagged canonical ATP synthase subunits, revealing localization of all but one of the subunits to the mitochondrion. Blue native gel electrophoresis of P. falciparum mitochondrial membranes suggested the molecular mass of the ATP synthase complex to be greater than 1 million daltons. This size is consistent with the complex being assembled as a dimer in a manner similar to the complexes observed in other eukaryotic organisms. This observation also suggests the presence of previously unknown subunits in addition to the canonical subunits in P. falciparum ATP synthase complex. Our attempts to disrupt genes encoding ß and γ subunits were unsuccessful, suggesting an essential role played by the ATP synthase complex in blood stages of P. falciparum. These studies suggest that, despite some unconventional features and its minimal contribution to ATP synthesis, P. falciparum ATP synthase is localized to the parasite mitochondrion, assembled as a large dimeric complex, and is likely essential for parasite survival.

Futuristic Non-antibiotic Therapies to Combat Antibiotic Resistance: A Review.

The looming problem of resistance to antibiotics in microorganisms is a global health concern. The drug-resistant microorganisms originating from anthropogenic sources and commercial livestock farming have posed serious environmental and health challenges. Antibiotic-resistant genes constituting the environmental "resistome" get transferred to human and veterinary pathogens. Hence, deciphering the origin, mechanism and extreme of transfer of these genetic factors into pathogens is extremely important to develop not only the therapeutic interventions to curtail the infections, but also the strategies to avert the menace of microbial drug-resistance. Clinicians, researchers and policymakers should jointly come up to develop the strategies to prevent superfluous exposure of pathogens to antibiotics in non-clinical settings. This article highlights the present scenario of increasing antimicrobial-resistance in pathogenic bacteria and the clinical importance of unconventional or non-antibiotic therapies to thwart the infectious pathogenic microorganisms.

Functional studies of Plasmodium falciparum's prohibitin1 and prohibitin 2 in yeast.

Prohibitins (PHBs) are evolutionarily conserved mitochondrial integral membrane proteins, shown to regulate mitochondrial structure and function, and can be classified into PHB1 and PHB2. PHB1 and PHB2 have been shown to interact with each other, and form heterodimers in mitochondrial inner membrane. Plasmodium falciparum has orthologues of PHB1 and PHB2 in its genome, and their role is unclear. Here, by homology modelling and yeast two-hybrid analysis, we show that putative Plasmodium PHBs (Pf PHB1 and Pf PHB2) interact with each other, which suggests that they could form supercomplexes of heterodimers in Plasmodium, the functional form required for optimum mitochondrial function.