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.

Evaluating the carnivorous efficacy of Utricularia aurea (Lamiales: Lentibulariaceae) on the larval stages of Anopheles stephensi, Culex quinquefasciatus, and Aedes aegypti (Diptera: Culicidae).

The emergence of insecticide resistance in mosquitoes necessitates the exploration and validation of sustainable biological strategies for controlling mosquitoes in their natural habitats. We assessed the predatory effect of Utricularia aurea Lour (Lamiales: Lentibulariaceae), an aquatic carnivorous plant found in the Indian subcontinent, Japan, and Australia, on 4 instars of Anopheles stephensi Liston, Culex quinquefasciatus Say, and Aedes aegypti Linn (Diptera: Culicidae), in the laboratory and field settings. In the laboratory setting, predation of larvae by U. aurea was highest during the first hour when it predated 45%, 61%, and 58% of first instars of An. stephensi, Cx. quinquefasciatus, and, Ae. aegypti, respectively, and, within 12 h, U. aurea preyed upon ~95% of the first, second, and third instars of the 3 mosquito species, ~80% of the fourth instars of An. stephensi and Ae. aegypti, and ~60% of fourth instars of Cx. quinquefasciatus. The predatory effect of U. aurea varied with mosquito species and instar. Broadly, predation risk declined with the increase of the instar size. In the field setting, at the end of 16 days, U. aurea predated 76% and 71% of the immature An. stephensi and Ae. aegypti, respectively. Our findings suggest U. aurea can be utilized as a potential biocontrol agent for controlling mosquito larvae in natural habitats; however, the current claim warrants additional investigations in a variety of natural habitats.

An assessment of fluorescence- and absorbance-based assays to study metal-oxide nanoparticle ROS production and effects on bacterial membranes.

The production and inevitable release of engineered nanoparticles requires rapid approaches to screen for their potential effects in environmental organisms, including bacteria. In bacteria, engineered nanoparticle effects can initiate at the cell membrane, for example by structurally damaging membranes or inhibiting energy transduction. Commercially available fluorescence- and absorbance-based assays could allow for rapidly assaying engineered nanoparticle effects on bacterial membranes, but there are limitations, including that: 1) assays are not currently configured to operate as part of a comprehensive high-throughput screening system, since assay conditions vary widely and formats are mostly high-volume and thus low-throughput, and; 2) engineered nanoparticles can interfere with assay reagents or function, yielding false-negative or -positive outcomes. Here, key assays to study reactive oxygen species (total ROS, and superoxide) production, and impacts on bacterial membrane integrity, membrane potential, and electron transport chain activity, are assessed for their potential use as a comprehensive system to test for nanoparticle effects in bacteria. To address (1), assays are adapted for simultaneous use in 96-well microplates under harmonized conditions. To address (2), a general scheme to test for engineered nanoparticle interferences with assay reagents and function is conceived, and used to study assay interferences by three nanoscale metal-oxides: nano-TiO2 , nano-CeO2 , and nano-ZnO. The results show that the selected assays can be used as a suite, and that nanoparticle interferences, when they occur, can be systematically investigated and often accounted for.

Nano-Nd2O3 reduced soil bacterial community function by altering the relative abundance of rare and sensitive taxa.

Nanoparticulate-Nd2O3 (nano-Nd2O3) has been excessively utilized in agriculture, industry, and medicine. Hence, nano-Nd2O3 can have environmental implications. However, the impact of nano-Nd2O3 on alpha diversity, composition, and function of soil bacterial communities has not been thoroughly evaluated. We amended soil to achieve different concentrations of nano-Nd2O3 (0, 10, 50, and 100 mg kg-1 soil) and incubated the mesocosms for 60 days. On days 7 and 60 of the experiment, we measured the effect of nano-Nd2O3 on alpha diversity and composition of soil bacterial community. Further, the effect of nano-Nd2O3 on the function of soil bacterial community was assessed based on changes in the activities of the six potential enzymes that mediate the cycling of nutrients in the soil. Nano-Nd2O3 did not alter the alpha diversity and composition of the soil bacterial community; however, it negatively affected community function in a dose-dependent manner. Specifically, the activities of ß-1,4-glucosidase and ß-1,4-n-acetylglucosaminidase that mediate soil carbon and nitrogen cycling, respectively, were significantly affected on days 7 and 60 of the exposure. The effect of nano-Nd2O3 on the soil enzymes correlated with changes in relative abundances of the rare and sensitive taxa, viz., Isosphaerales, Isosphaeraceae, Ktedonobacteraceae, and Streptomyces. Overall, we provide information for the safe implementation of technological applications that use nano-Nd2O3.

Effect of modeled reduced gravity conditions on bacterial morphology and physiology.

BACKGROUND: Bacterial phenotypes result from responses to environmental conditions under which these organisms grow; reduced gravity has been demonstrated in many studies as an environmental condition that profoundly influences microorganisms. In this study, we focused on low-shear stress, modeled reduced gravity (MRG) conditions and examined, for Escherichia coli and Staphlyococcus aureus, a suite of bacterial responses (including total protein concentrations, biovolume, membrane potential and membrane integrity) in rich and dilute media and at exponential and stationary phases for growth. The parameters selected have not been studied in E. coli and S. aureus under MRG conditions and provide critical information about bacterial viability and potential for population growth. RESULTS: With the exception of S. aureus in dilute Luria Bertani (LB) broth, specific growth rates (based on optical density) of the bacteria were not significantly different between normal gravity (NG) and MRG conditions. However, significantly higher bacterial yields were observed for both bacteria under MRG than NG, irrespective of the medium with the exception of E. coli grown in LB. Also, enumeration of cells after staining with 4',6-diamidino-2-phenylindole showed that significantly higher numbers were achieved under MRG conditions during stationary phase for E. coli and S. aureus grown in M9 and dilute LB, respectively. In addition, with the exception of smaller S. aureus volume under MRG conditions at exponential phase in dilute LB, biovolume and protein concentrations per cell did not significantly differ between MRG and NG treatments. Both E. coli and S. aureus had higher average membrane potential and integrity under MRG than NG conditions; however, these responses varied with growth medium and growth phase. CONCLUSIONS: Overall, our data provides novel information about E. coli and S. aureus membrane potential and integrity and suggest that bacteria are physiologically more active and a larger percentage are viable under MRG as compared to NG conditions. In addition, these results demonstrate that bacterial physiological responses to MRG conditions vary with growth medium and growth phase demonstrating that nutrient resources are a modulator of response.

Rare-earth metal oxide nanoparticles decouple the linkage between soil bacterial community structure and function by selectively influencing potential keystone taxa.

Excessive production and application of rare-earth metal oxide nanoparticles warrants assessment of their environmental risks. Little is known about the impact of these nanoparticles on soil bacterial communities. We quantified the effects of nano-Gd2O3 and nano-La2O3, at the different concentrations and exposure regimes, on soil bacterial community structure and function as well as the structure-function relationship. Further, we constructed and analyzed a co-occurrence network to identify and characterize potential keystone taxa that were related to the enzyme activities and responded to the increasing concentrations of nanoparticles. Both nano-Gd2O3 and nano-La2O3 significantly altered the bacterial community structure and function in a concentration-dependent manner; however, these negative effects were observed on day 1 or day 7 but not on day 60, indicating that these effects were transient and the bacterial communities can mitigate the effect of these nanoparticles over time. Interestingly, the nanoparticle exposure decoupled the relationship between the structure and function of the soil bacterial communities. The decoupling was due to changes in the composition and relative abundances of potential keystone taxa related to bacterial community functions. Altogether, we provide insights into the interactions between the rare-earth metal oxide nanoparticles and soil bacterial communities. Our results facilitate the environmental risk assessment and safe usage of rare-earth metal oxide nanoparticles.

Enrichment of potential degrading bacteria accelerates removal of tetracyclines and their epimers from cow manure biochar amended soil.

The excessive usage of tetracyclines in animal husbandry and aquaculture invariably leads to deterioration of the microbial quality of nearby soils. We previously reported the accelerated removal of tetracyclines and their intermediates from the cow manure biochar amended soil (CMB). However, little is known about the underlying changes in the microbial community that mediate the accelerated removal of tetracyclines from the CMB. Here, we compared the concentration of parent tetracyclines along with their intermediates, microbial biomass, and microbial (fungal and bacterial) community in CMB and the control soil (CK) on the day of 1, 5, 10, 20, 30, 45, and 60. The biochar amendment accelerated the removal of tetracyclines and their epimers from the soil. Bacterial community composition varied between the CMB and CK. The relative abundance and richness of the bacteria that correlated with the degradation of tetracyclines and their epimers was significantly higher in the CMB as compared to the CK. Specifically, the CMB had a more intricate network of the degrading bacteria with the three keystone genera viz. Acidothermus sp., Sphingomonas sp., and Blastococcus sp., whereas, the CK had a simple network with Sphingomonas sp. as the keystone genus. Overall, the biochar amendment accelerated the removal of tetracyclines and their epimers through the enrichment of potential tetracycline degrading bacteria in the soil; thus, it can be applied for the in situ remediation of soils contaminated with tetracyclines.

Thiacloprid exposure perturbs the gut microbiota and reduces the survival status in honeybees.

Honeybees (Apis mellifera) offer ecosystem services such as pollination, conservation of biodiversity, and provision of food. However, in recent years, the number of honeybee colonies is diminishing rapidly, which is probably linked to the wide use of neonicotinoid insecticides. Middle-aged honeybees were fed with 50% (w/v) sucrose solution containing 0, 0.2, 0.6, and 2.0 mg/L thiacloprid (a neonicotinoid insecticide) for up to 13 days, and on each day of exposure experiment, percentage survival, sucrose consumption, and bodyweight of honeybees were measured. Further, changes in honeybee gut microbial community were examined using next-generation 16S rDNA amplicon sequencing on day 1, 7, and 13 of the exposure. When compared to control-treatment, continuous exposure to high (0.6 mg/L) and very high (2.0 mg/L) concentrations of thiacloprid significantly reduced percentage survival of honeybees (p < 0.001) and led to dysbiosis of their gut microbial community on day 7 of the exposure. However, during subsequent developmental stages of middle-aged honeybees (i.e. on day 13), their gut microbiome recovered from dysbiosis that occurred previously due to thiacloprid exposure. Taken together, improper application of thiacloprid can cause loss of honeybee colonies, while the microbial gut community of honeybee is an independent variable in this process.