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This study was performed to evaluate the change in seasonal abundance and distribution of individual mosquito vectors (Culex, Anopheles, and Aedes) in relation to the climatic factors in Eastern Province, Saudi Arabia, for the study period of 2014. The association between mosquito abundance and environmental parameters was investigated using bivariate and multivariate analysis. The study showed the range of temperature and relative humidity required for individual mosquito larvae abundance varies for Culex, Anopheles, and Aedes. However, no variation was observed in the range of temperature and relative humidity required for the abundance of adult Culex and Anopheles. The results revealed a negative relationship between mosquito larval/adult abundance and temperature (Total number of larva/adult is 671/11 in July, While it is 2462/221 in January). There is a link between relative humidity and rainfall, as the three climatic factors together were responsible for 33.1% (R2 = 0.331), 54.6% (R2 = 0.546), and 86.6% (R2 = 0.866) of the variance on Culex, Anopheles, and Aedes larvae, respectively. The effects of the three climatic parameters of temperature, relative humidity, and rainfall on mosquito larval and adult abundance were discussed. In addition, influences of other environmental factors on larval/adult mosquito distribution and abundance were also explained.

Thus far, only a handful of bacterial strains that can independently degrade and utilize benzo[a]pyrene (BaP) as the sole carbon source have been isolated and characterized. Here, three new bacterial strains, JBZ1A, JBZ2B, and JBZ5E, were isolated from contaminated soil and, using 16S rRNA sequencing, were identified as Bradyrhizobium japonicum, Micrococcus luteus, and Bacillus cereus, respectively. The growth ability of each individual strain and a consortium of all strains in the presence of BaP (4–400 µmol·l−1, pH 7, 37°C) was identified by the doubling time (dt). The results illustrated that dt decreased with increasing BaP concentrations for individual strain and the consortium. The optimum growth conditions of the consortium were 37°C, 0.5% NaCl (w/v), and pH 7. Under these conditions, the degradation rate was 1.06 µmol·l−1·day−1, whereas that of individual strains ranged 0.9–0.38 µmol·l−1·day−1. B. cereus had the strongest contribution to the consortium’s activity, with a degradation rate of 0.9 µmol·l−1·day−1. The consortium could also remove BaP spiked with soil, but at a lower rate (0.01 µmol.l−1.day−1). High-performance liquid chromatography–high-resolution tandem mass spectrometry permitted the detection of the metabolites of these strains, and a biodegradation pathway has been proposed.

To date, only a handful of bacterial strains that can independently degrade and utilize benzo[a]pyrene (BaP) as the sole carbon source has been isolated and characterized. Here, three new bacterial strains—JBZ1A, JBZ2B, and JBZ5E—were isolated from contaminated soil and, using 16S rRNA sequencing, were identified as Brad rhizobium japonicum, Micrococcus luteus, and Bacillus cereus, respectively. The growth ability of each individual strain and a consortium of all strains in the presence of BaP (4–400 µmol·L−1, pH 7, 37 °C) was identified by the doubling time (dt). The results illustrate that dt decreased with increasing BaP concentrations for individual strains and the consortium. The optimum growth conditions of the consortium were 37 °C, 0.5% NaCl (w/v), and pH 7. Under these conditions, the degradation rate was 1.06 µmol·L−1·day−1, whereas that of individual strains ranged from 0.9 to 0.38 µmol·L−1·day−1. B. cereus had the strongest contribution to the consortium’s activity, with a degradation rate of 0.9 µmol·L−1·day−1. The consortium could also remove BaP spiked with soil but at a lower rate (0.01 µmol L−1.day−1). High-performance liquid chromatography–high-resolution tandem mass spectrometry permitted the detection of the metabolites of these strains, and a biodegradation pathway is proposed.

Polycyclic aromatic hydrocarbons (PAHs), which consist of low-molecular-weight PAHs (LMW-PAHs) and high-molecular-weight PAHs (HMW-PAHs), form an important class of pollutants. Pyrene and benzo[a]pyrene (BaP) are the main pollutants belonging to HMW-PAHs, and their degradation by microorganisms remains an important strategy for their removal from the environments. Extensive studies have been carried out on the isolation and characterisation of microorganisms that actively degrade LMW-PAHs, and to a certain extent, the HMW-PAH pyrene. However, so far, limited work has been carried out on BaP biodegradation. BaP consists of five fused aromatic rings, which confers this compound a high stability, rendering it less amenable to biodegradation. The current review summarizes the emerging reports on BaP biodegradation. More specifically, work carried out on BaP bacterial degradation and current knowledge gaps that limit our understanding of BaP degradation are highlighted. Moreover, new avenues of research on BaP degradation are proposed, specifically in the context of the development of “omics” approaches

We report a new assay system for the detection of miR-21 in cancer cells. The new assay works at room temperature and it does not involve enzymatic amplification. It consists a hairpin smart probe, designed to specifically recognize miR-21 target sequence. We tested the performance and sequence recognition capability of the smart probe to confirm desired specifications. We used the smart probe for the sequence-specific recognition of synthetic miR-21 oligonucleotides as well as mismatch sequences and we found that the probe recognizes the target sequence-specifically, while discriminating against mismatched sequences. We determined the limit of detection and limit of quantitation for the miR-21 oligonucleotides to be 1.72 nM and 5.78 nM, respectively, while the sensitivity is 6.90 × 10

Rumex vesicarius (RVE) mediated chitosan–AgNPs composite was produced in situ by using an aqueous extract of Rumex vesicarius leaves as the reducing agent to reduce Ag+ to Ag0. The synthesized composite was evaluated as a sweet (CO2) corrosion inhibitor (CI) for C1018 carbon steel (CS) in 3.5 wt% NaCl solution under static and hydrodynamic conditions. The corrosion inhibitive performance was evaluated using electrochemical impedance spectroscopy (EIS), linear polarization resistance (LPR), and potentiodynamic polarization (PDP) techniques, as well as scanning electron microscopy (SEM)/energy dispersive X-ray spectroscopy (EDAX), and atomic force microscopy (AFM) on corroded C1018 CS without and with additives. The effect of concentration, immersion time, temperature, and rotation speed on the CI performance of the composite was also investigated. The corrosion inhibitive effect increased with increasing composite dosage, with the highest inhibition efficiency (IE) acquired at the maximum composite dosage of 0.3%. Beyond this concentration, the IE decline with increasing concentration. Furthermore, IE was found to increase with immersion time and decline with a temperature rise from 25 to 40 °C, with the optimum temperature of 60 °C found to accelerate corrosion without and with RVE-mediated Chi–AgNPs composite. Under high shear stress, the Chi–AgNPs composite exhibits moderate corrosion inhibition under hydrodynamic conditions. The surface analysis results validate the formation of a protective covering due to composite adsorption on the CS surface. The RVE-mediated chitosan–AgNPs composite could be recommended as a CI for C1018 CS in sweet (CO2) corrosion environments at ambient temperature.

MalariaGEN is a data-sharing network that enables groups around the world to work together on the genomic epidemiology of malaria. Here we describe a new release of curated genome variation data on 7,000 Plasmodium falciparum samples from MalariaGEN partner studies in 28 malaria-endemic countries. High-quality genotype calls on 3 million single nucleotide polymorphisms (SNPs) and short indels were produced using a standardised analysis pipeline. Copy number variants associated with drug resistance and structural variants that cause failure of rapid diagnostic tests were also analysed. Almost all samples showed genetic evidence of resistance to at least one antimalarial drug, and some samples from Southeast Asia carried markers of resistance to six commonly-used drugs. Genes expressed during the mosquito stage of the parasite life-cycle are prominent among loci that show strong geographic differentiation. By continuing to enlarge this open data resource we aim to facilitate research into the evolutionary processes affecting malaria control and to accelerate development of the surveillance toolkit required for malaria elimination.

MalariaGEN is a data-sharing network that enables groups around the world to work together on the genomic epidemiology of malaria. Here we describe a new release of curated genome variation data on 7,000 Plasmodium falciparum samples from MalariaGEN partner studies in 28 malaria-endemic countries. High-quality genotype calls on 3 million single nucleotide polymorphisms (SNPs) and short indels were produced using a standardised analysis pipeline. Copy number variants associated with drug resistance and structural variants that cause failure of rapid diagnostic tests were also analysed. Almost all samples showed genetic evidence of resistance to at least one antimalarial drug, and some samples from Southeast Asia carried markers of resistance to six commonly-used drugs. Genes expressed during the mosquito stage of the parasite life-cycle are prominent among loci that show strong geographic differentiation. By continuing to enlarge this open data resource we aim to facilitate research into the evolutionary processes affecting malaria control and to accelerate development of the surveillance toolkit required for malaria elimination.

Three strains of novel bacteria were isolated from oil-contaminated sediment from the Arabian Gulf (Brevibacillus brevis T2C2008, Proteus mirabilis T2A12001, and Rhodococcus quinshengi TA13008). The isolated strains were tested for their degrading efficacy of low and high molecular hydrocarbon (naphthalene and pyrene). The efficacy of the two-hydrocarbon degradation by the isolates bacterial was determined at a temperature of 25 °C and 37 °C and pH of 5.0 and 9.0. In inoculated media at 37 °C, Rhodococcus qinshengi fully metabolized naphthalene and degrade 56% of pyrene. Brevibacillus brevis break down over 80% of naphthalene at room temperatures (25 °C). However, it was found that P. mirabilis and R. qinshengi biodegraded nearly 94% of naphthalene in the incubated media. The capacity for pyrene and naphthalene degradation in varying pH and temperature conditions was shown to be significant in Rhodococcus qinshengi because of its mineralization exceeding 50% across the tested pH and temperature. This implies that the isolated strains are ideal for biodegradation of contaminated sediment with naphthalene and pyrene.

Chitosan/silver nanofluids were prepared using Phoenix dactylifera (DPLE) or Rumex vesicarius (HEL) extracts as the reducing agent, characterized using Fourier-transform infrared spectroscopy (FTIR), ultraviolet&ndash, visible (UV-vis), X-ray diffraction (XRD), and transmission electron microscope (TEM). The antimicrobial effect of the nanofluids against Gram positive, Bacillus licheniformis, Staphylococcus haemolyticus, Bacillus cereus, and Micrococcus luteus, and Gram-negative Pseudomonas aeruginosa, Pseudomonas citronellolis, and Escherichia coli bacteria has been studied. The nanoparticles were polydispersed in the chitosan matrix and are highly stable. The zeta potential of the silver nanoparticles in DPLE- and HEL-mediated composites is +46 mV and +56 mV, respectively. The FTIR results reveal that the free carboxylate groups in the plant biomaterial took part in stabilization process. HEL is a stronger reducing agent than DPLE and nanoparticles generated with HEL are smaller (8.0&ndash, 36 nm) than those produced with DPLE (10&ndash, 43 nm). DPLE- and HEL-mediated composites effectively inhibit the growth of the studied bacteria but HEL-mediated composite exhibited higher effect. The higher antimicrobial activity of HEL-mediated composite is linked to the smaller nanoparticles. The foregoing results indicate that HEL extract can be used in the green production of potential antimicrobial chitosan/silver nanofluids for biomedical and packaging applications.

The ability of the human malaria parasite, Plasmodium falciparum to develop resistance against mainstay drugs remains a public health problem. Currently, the antimalarial drugs, lumefantrine (LM), and piperaquine (PQ) are essential components of the mainstay artemisinin-based therapies used for the treatment of malaria globally. Here, we used a model parasite Plasmodium berghei, to investigate the mechanisms of LM and PQ resistance. We employed known resistance reversing agents (RA): probenecid, verapamil, or cyproheptadine to study the mechanisms of LM and PQ resistance in the standard 4-day suppressive test. We then employed reverse genetics to assess the impact of deleting or over-expressing plausible genes associated with the metabolism and transport of drugs. We show that only, cyproheptadine at 5mgkg-1 restored LM activity by above 65% against LM-resistant parasites (LMr) but failed to reinstate PQ activity against PQ-resistant parasites (PQr). Whereas the PQr had lost significant susceptibility to LM, the three RA, cyproheptadine verapamil, and probenecid restored LM potency by above 70%, 60%, and 55% respectively against the PQr. We thus focused on the mechanisms of LM resistance in PQr. Here we show the partial deletion of the cysteine desulfurase (SUFS) and overexpression of the Ferredoxin NADP+ reductase (FNR) genes in the PQr parasite achieved two results; i) abolished the impact of RA on LM activity; ii) restored the susceptibility of PQr to LM alone. Our findings associated SUFS and FNR protein with the action of LM and RA action in P. berghei. We demonstrate that the incorporation of any of the RA into an antimalarial combination that comprises LM would augment LM activity and concomitantly antagonize the emergence of LM resistance derived from PQ pressure. The impact of RA, deletion of SUFS, and overexpression of FNR on LM activity need to be tested in Plasmodium falciparum.nnAuthor summaryLumefantrine (LM) and piperaquine (PQ) are essential drugs for the treatment of malaria globally. Here, we used Plasmodium berghei, a model parasite that infects rodents to study how parasites escape killing by PQ and LM. We first used a second drug: probenecid, verapamil, or cyproheptadine to enhance the activity of LM or PQ. We show that cyproheptadine restores LM activity against LM-resistant parasites (LMr) but failed to reestablish PQ activity against PQ-resistant parasites (PQr). Since PQr is resistant to LM, combining LM with either cyproheptadine, verapamil, or probenecid reinstates LM activity against PQr. We then focused mainly on LM resistance in PQr. After genetically manipulating the PQr, we reveal that cysteine desulfurase (SUFS) and ferredoxin NADP+reductase (FNR) regulate LM capacity to kill parasites. Decreasing the level of SUFS or increasing FNR levels in the PQr makes the parasites susceptible to LM but abolishes the impact of probenecid, verapamil, and cyproheptadine on LM activity. Overall, we provide clues on the link between SUFS and FNR in the action of LM and RA in P. berghei. This study provides a basis for an in-depth analysis of how LM mediates parasites kill and how the parasite escapes LM action in Plasmodium falciparum.

In the anaerobic process, fat-oil-grease (FOG) is hydrolysed to long-chain fatty acids (LCFAs) and glycerol (GLYC), which are then used as substrates to produce biogas. The increase in FOG and LCFAs inhibits methanogenesis, and so far, most work investigating this inhibition has been carried out when FOG or LCFAs were used as co-substrates. In the current work, the inhibition of methanogenesis by FOG, LCFAs and GLYC was investigated when used as sole substrates. To gain more insight on the dynamics of this process, the change of microbial community was analysed using 16S rRNA gene amplicon sequencing. The results indicate that, as the concentrations of cooking olive oil (CO, which represents FOG) and LCFAs increase, methanogenesis is inhibited. For instance, at 0.01 g. L-1 of FOG, the rate of biogas formation was around 8 ml.L-1.day-1, and this decreased to = 45°C and NaCl > 3% led to a significant decrease in the rate of biogas formation. Microbial community analyses were carried out from samples from 3 different bioreactors (CO, OLEI and GLYC), on day 1, 5 and 15. In each bioreactor, microbial communities were dominated by Proteobacteria, Firmicutes and Bacteroidetes phyla. The most important families were Enterobacteriaceae, Pseudomonadaceae and Shewanellaceae (Proteobacteria phylum), Clostridiacea and Ruminococcaceae (Firmicutes) and Porphyromonadaceae and Bacteroidaceae (Bacteroidetes). In CO bioreactor, Proteobacteria bacteria decreased over time, while those of OLEI and GLYC bioreactors increased. A more pronounced increase in Bacteroidetes and Firmicutes were observed in CO bioreactor. The methanogenic archaea Methanobacteriaceae and Methanocorpusculaceae were identified. This analysis has shown that a set of microbial population is selected as a function of the substrate.

Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants from incomplete combustion and petroleum products. As the molecular weight increases, PAHs become more recalcitrant to biodegradation. A bacterial strain capable of metabolizing the four fused aromatic ring PAH pyrene was isolated and characterized. The analysis of 16S rRNA gene revealed that it belongs to Achromobacter xylosoxidans species. A. xylosoxidans PY4 can utilize pyrene as the sole source of carbon. PY4 has a doubling time (dt) of less than 1 day when it grows in the presence of 1–5 mg l−1 pyrene, a dt range similar to that of the most efficient pyrene biodegrading bacteria described so far. The optimal pyrene degradation conditions are at pH 7–9, 37–40 °C, and 0–2.5% NaCl. PY4 also utilizes salicylic acid, catechol, naphthalene, anthracene and phenanthrene. PY4 degrades more than 50% of 100 mg l−1 of pyrene, within the first 15 days, at a rate of 0.069 day−1, R2 = 0.99. The metabolites include monohydroxypyrene, 1-methoxyl-2-H-benzo[h]chromene-2-carboxylic acid, 9,10-phenanthrenequinone, 1-methoxyl-trans-2′-carboxybenzalpyruvate, and dibutyl-phthalate. Up-expressed proteins in response to pyrene are involved in cell homeostasis, genetic information synthesis and storage, and chemical stress. Among these proteins are 4-hydroxyphenylpyruvate dioxygenase and homogentisate 1,2-dioxygenase, involved in the lower pyrene degradation pathway.

Silver nanoparticles (AgNPs) have been synthesized in the presence of Strawberry fruit extract (SBFE) at room temperature. The synthesized AgNPs was characterized by UV-vis spectroscopy, SEM, EDS, XRD, TEM and FTIR. The UV-vis spectra of the AgNPs show SPR band at 450 nm. TEM results indicate that AgNPs are spherical in shape and size range between 7–65 nm. Antibacterial activity of the synthesized AgNPs has been assessed against Pseudomonas aeruginosa and Bacillus licheniformis. The results show that AgNPs exhibit inhibitory effect and effect is a function of AgNPs concentration. The antibacterial activity of the prepared AgNPs has been compared with two antibiotics, amoxicillin and ciprofloxacin. It is found that the antibiotics perform better than AgNPs.

Contamination of the environment by petroleum products is a growing concern worldwide, and strategies to remove these contaminants have been evaluated. One of these strategies is biodegradation, which consists of the use of microorganisms. Biodegradation is significantly improved by increasing the temperature of the milieu, thus, the use of thermophiles, microbes that thrive in high-temperature environments, will render this process more efficient. For instance, various thermophilic enzymes have been used in industrial biotechnology because of their unique catalytic properties. Biodegradation has been extensively studied in the context of mesophilic microbes, and the mechanisms of biodegradation of aliphatic and aromatic petroleum hydrocarbons have been elucidated. However, in comparison, little work has been carried out on the biodegradation of petroleum hydrocarbons by thermophiles. In this paper, a detailed review of the degradation of petroleum hydrocarbons (both aliphatic and aromatic) by thermophiles has been carried out. This work has identified the characteristics of thermophiles, and unravelled specific catabolic pathways of petroleum products that are only found in thermophiles. Gaps that limit our understanding of the activity of these microbes have also been highlighted, and finally, different strategies that can be used to improve the efficiency of degradation of petroleum hydrocarbons by thermophiles have been proposed.

The exploitation of petroleum oil generates a considerable amount of “produced water or petroleum waste effluent (PWE)” that is contaminated with polycyclic aromatic hydrocarbons (PAHs), including Benzo[a]pyrene (BaP). PWE is characterised by its high salinity, which can be as high as 30% NaCl, thus the exploitation of biodegradation to remove PAHs necessitates the use of active halophilic microbes. The strain 10SBZ1A was isolated from oil contaminated soils, by enrichment experiment in medium containing 10% NaCl (w/v). Homology analyses of 16S rRNA sequences identified 10SBZ1A as a Staphylococcus haemoliticus species, based on 99.99% homology (NCBI, accession number GI: MN388897). The strain could grow in the presence of 4–200 μmol l-1 of BaP as the sole source of carbon, with a doubling time of 17–42 h. This strain optimum conditions for growth were 37 oC, 10% NaCl (w/v) and pH 7, and under these conditions, it degraded BaP at a rate of 0.8 μmol l-1 per day. The strain 10SBZ1A actively degraded PAHs of lower molecular weights than that of BaP, including pyrene, phenanthrene, anthracene. This strain was also capable of removing 80% of BaP in the context of soil spiked with BaP (10 μmol l-1 in 100 g of soil) within 30 days. Finally, a metabolic pathway of BaP was proposed, based on the identified metabolites using liquid chromatography-high resolution tandem mass spectrometry. To the best of our knowledge, this is the first report of a halophilic BaP degrading bacterial strain at salinity > 5% NaCl.

Bioremediation is based on microorganisms able to use pollutants either as a source of carbon or in co-metabolism, and is a promising strategy in cleaning the environment. Using soil contaminated with petroleum products from an industrial area in Saudi Arabia (Jubail), and after enrichment with the polycyclic aromatic hydrocarbon (PAH) naphthalene, a Methylobacterium radiotolerans strain (N7A0) was isolated that can grow in the presence of naphthalene as the sole source of carbon. M. radiotolerans is known to be resistant to gamma radiation, and this is the first documented report of a strain of this bacterium using a PAH as the sole source of carbon. The commonly reported Pseudomonas aeruginosa (strain N7B1) that biodegrades naphthalene was also identified, and gas chromatography analyses have shown that the biodegradation of naphthalene by M. radiotolerans and P. aeruginosa did follow both the salicylate and phthalate pathways.

A bacterial consortium that degrades cooking oil (CO) has been isolated in wastewater (WW) samples, by enrichment in olive CO. This consortium could degrade 90% of CO within 7-9 days (from an initial 1% [w/v]), and it is more active at alkaline conditions. The 16S ribonucleic acid (RNA) gene analysis showed that it contains five bacterium species: Stenotrophomonas rhizophila, Sphingobacterium sp., Pseudomonas libanensis, Pseudomonas poae and Pseudomonas aeruginosa. This consortium can degrade the free fatty acids (FFA): palmitic, stearic, oleic, linoleic and linolenic acids; glycerol, glucose and amylose; and albumin, but could not efficiently degrade carboxymethyl-cellulose. Each strain could also degrade CO and FFAs. The level of bacterial crude-activity of extracellular lipases was found to be between 0.2 and 4U/ml. Using synthetic WW, the consortium could reduce 80% of the chemical oxygen demand [from 10550 ± 2828 mg/l], 80% of nitrogen (from 410 ± 78 mgl/l) and 57% of phosphorus (from 93 ± 25 mg/l). Thus, this consortium can be utilized in the removal of CO from WW.

The biodegradation of low- and high-molecular-weight polycyclic aromatic hydrocarbons (PAHs) (LWM-PAHs and HMW-PAHs, respectively) has been studied extensively under aerobic conditions. Molecular O2 plays 2 critical roles in this biodegradation process. O2 activates the aromatic rings through hydroxylation prior to ring opening and serves as a terminal electron acceptor (TEA). However, several microorganisms have devised ways of activating aromatic rings, leading to ring opening (and thus biodegradation) when TEAs other than O2 are used (under anoxic conditions). These microorganisms belong to the sulfate-, nitrate-, and metal-ion-reducing bacteria and the methanogens. Although the anaerobic biodegradation of monocyclic aromatic hydrocarbons and LWM-PAH naphthalene have been studied, little information is available about the biodegradation of HMW-PAHs. This manuscript reviews studies of the anaerobic biodegradation of HMW-PAHs and identifies gaps that limit both our understanding and the efficiency of this biodegradation process. Strategies that can be employed to overcome these limitations are also discussed.

We investigated the mechanisms of resistance of two antimalarial drugs piperaquine (PQ) and lumefantrine (LM) using the rodent parasite Plasmodium berghei as a surrogate of the human parasite, Plasmodium falciparum. We analyzed the whole coding sequence of Plasmodium berghei chloroquine resistance transporter (Pbcrt) and Plasmodium berghei multidrug resistance gene 1(Pbmdr-1) for polymorphisms. These genes are associated with quinoline resistance in Plasmodium falciparum. No polymorphic changes were detected in the coding sequences of Pbcrt and Pbmdr1 or in the mRNA transcript levels of Pbmdr1. However, our data demonstrated that PQ and LM resistance is achieved by multiple mechanisms that include elevated mRNA transcript levels of V-type H+ pumping pyrophosphatase (vp2), Ca2+/H+ antiporter (vcx1), gamma glutamylcysteine synthetase (ggcs) and glutathione-S-transferase (gst) genes, mechanisms also known to contribute to chloroquine resistance in P. falciparum and rodent malaria parasites. The increase in ggcs and gst transcript levels was accompanied by high glutathione (GSH) levels and elevated activity of glutathione-S-transferase (GST) enzyme. Taken together, these results demonstrate that Pbcrt and Pbmdr1 are not associated with PQ and LM resistance in P. berghei ANKA, while vp2, vcx1, ggcs and gst may mediate resistance directly or modulate functional mutations in other unknown genes.

Background: The human malaria parasite Plasmodium falciparum has evolved drug evasion mechanisms to all available antimalarials. The combination of amodiaquine-artesunate is among the drug of choice for treatment of uncomplicated malaria. In this combination, a short-acting, artesunate is partnered with long-acting, amodiaquine for which resistance may emerge rapidly especially in high transmission settings. Here, we used a rodent malaria parasite Plasmodium berghei ANKA as a surrogate of P. falciparum to investigate the mechanisms of amodiaquine resistance. Methods: We used the ramp up approach to select amodiaquine resistance. We then employed the 4-Day Suppressive Test to measure the resistance level and determine the cross-resistance profiles. Finally, we genotyped the resistant parasite by PCR amplification, sequencing and relative quantitation of mRNA transcript of targeted genes. Results: Submission of the parasite to amodiaquine pressure yielded resistant line within thirty-six passages. The effective doses that reduced 90% of parasitaemia (ED 90) of the sensitive and resistant lines were 4.29mg/kg and 19.13mg/kg respectively. The selected parasite retained resistance after ten passage cycles in the absence of the drug and freezing at -80ºC for one month with ED 90 of 20.34mg/kg and 18.22mg/kg. The parasite lost susceptibility to chloroquine by (6-fold), artemether (10-fold), primaquine (5-fold), piperaquine (2-fold) and lumefantrine (3-fold). Sequence analysis of Plasmodium berghei chloroquine-resistant transporter revealed His95Pro mutation. We found no variation in the nucleotide sequences of Plasmodium berghei multidrug resistance gene-1 (Pbmdr1), Plasmodium berghei deubiquitinating enzyme-1 or Plasmodium berghei Kelch13 domain. However, high mRNA transcripts of essential transporters; Pbmdr1, V-type/H+ pumping pyrophosphatase-2 and sodium hydrogen ion exchanger-1 and Ca 2+/H + antiporter accompanies amodiaquine resistance. Conclusions: The selection of amodiaquine resistance yielded stable “multidrug-resistant’’ parasites and thus may be used to study shared resistance mechanisms associated with other antimalarial drugs. Genome-wide analysis of the parasite may elucidate other functionally relevant genes controlling AQ resistance in P. berghei.

Anaerobic digestion (AD) is increasingly being used and exploited as a strategy to generate biomethane, which can be used as a renewable and clean energy. AD rests on the biodegradation of organic compounds in anaerobic condition, and these organic compounds are generally agricultural-, industrial- and domestic-wastes. However, problems of AD decrease efficiency, as the result of bioreactor stress, are generally encountered. The primarily cause of this stress is the presence of high concentrations of inhibitory substances such as nitrate, sulfate, heavy metals and oxygen among others. Another cause of AD decrease efficiency is the use of organic compounds that are less amenable to biodegradation such as lignocellulosic compounds. One of the strategies to overcome these limitations is the addition in bioreactors of “stress resistant”- or “efficient biomethane generating”- microorganisms to improve AD process. This strategy, known as bioaugmentation, has been used for the last 15 years to increase biomethane production. In this review, work carried out on this bioaugmentation process has been summarised, and new strategies that could be used or exploited to improve the success of this approach have also been discussed.

Background The spread of resistance to chloroquine (CQ) led to its withdrawal from use in most countries in sub-Saharan Africa in the 1990s. In Malawi, this withdrawal was followed by a rapid reduction in the frequency of resistance to the point where the drug is now considered to be effective once again, just nine years after its withdrawal. In this report, the polymorphisms of markers associated with CQ-resistance against Plasmodium falciparum isolates from coastal Kenya (Kilifi) were investigated, from 1993, prior to the withdrawal of CQ, to 2006, seven years after its withdrawal. Changes to those that occurred in the dihydrofolate reductase gene (dhfr) that confers resistance to the replacement drug, pyrimethamine/sulphadoxine were also compared. Methods Mutations associated with CQ resistance, at codons 76 of pfcrt, at 86 of pfmdr1, and at codons 51, 59 and 164 of dhfr were analysed using PCR-restriction enzyme methods. In total, 406, 240 and 323 isolates were genotyped for pfcrt-76, pfmdr1-86 and dhfr, respectively. Results From 1993 to 2006, the frequency of the pfcrt-76 mutant significantly decreased from around 95% to 60%, while the frequency of pfmdr1-86 did not decline, remaining around 75%. Though the frequency of dhfr mutants was already high (around 80%) at the start of the study, this frequency increased to above 95% during the study period. Mutation at codon 164 of dhf r was analysed in 2006 samples, and none of them had this mutation. Conclusion In accord with the study in Malawi, a reduction in resistance to CQ following official withdrawal in 1999 was found, but unlike Malawi, the decline of resistance to CQ in Kilifi was much slower. It is estimated that, at current rates of decline, it will take 13 more years for the clinical efficacy of CQ to be restored in Kilifi. In addition, CQ resistance was declining before the drug's official withdrawal, suggesting that, prior to the official ban, the use of CQ had decreased, probably due to its poor clinical effectiveness.

Plasmodium falciparum parasites resistant to the combination sulfadoxine-pyrimethamine are spreading in Africa, particularly in East Africa. This is a matter of concern because there are no other affordable drugs available. This article provides the evidence indicating that sulfadoxine-pyrimethamine resistance can be reversed in vitro and discusses how this information might be exploited to extend the therapeutic lifetime of sulfadoxine-pyrimethamine in vivo.

Background The emergence of artemisinin-resistant P. falciparum malaria in South-East Asia highlights the need for continued global surveillance of the efficacy of artemisinin-based combination therapies. Methods On the Kenyan coast we studied the treatment responses in 474 children 6–59 months old with uncomplicated P. falciparum malaria in a randomized controlled trial of dihydroartemisinin-piperaquine vs. artemether-lumefantrine from 2005 to 2008. (ISRCTN88705995) Results The proportion of patients with residual parasitemia on day 1 rose from 55% in 2005–2006 to 87% in 2007–2008 (odds ratio, 5.4, 95%CI, 2.7–11.1; P37.5°C, 2.8, 1.9–4.1; P

The folate pathway represents a powerful target for combating rapidly dividing systems such as cancer cells, bacteria and malaria parasites. Whereas folate metabolism in mammalian cells and bacteria has been studied extensively, it is understood less well in malaria parasites. In two articles, we attempt to reconstitute the malaria folate pathway based on available information from mammalian and microbial systems, in addition to Plasmodium-genome-sequencing projects. In part I, we focused on folate enzymes that are already used clinically as anticancer drug targets or that are under development in drug-discovery programs. In this article, we discuss mammalian folate enzymes that have not yet been exploited as potential drug targets, and enzymes that function in the de novo folate-synthesis pathway of the parasite – a particularly attractive area of attack because of its absence from the mammalian host.

Resistance to the antifolate sulfadoxine-pyrimethamine (SP), the current mass-treatment antimalarial drug, is associated with selection of point mutations in dihydrofolate reductase and dihydropteroate synthase. Among these mutations, the leucine 164 dihydrofolate reductase mutation (Leu-164) is associated with higher levels of SP resistance; this mutation is also associated with a decrease in the efficacy of chlorproguanil/dapsone, a newly developed antifolate antimalarial drug. Leu-164 has been detected in Southeast Asia and South America, regions where SP is no longer effective. Surprisingly, this mutation has not yet been detected in Africa, using the standard protocol based on PCR-RFLP, despite high SP resistance. In this paper, we discuss briefly the reasons why Leu-164 has not yet been selected in Africa and we propose a means that may slow down the selection of this mutation.

Resistance to the antimalarial drug sulfadoxine-pyrimethamine (SP) emerged in Plasmodium falciparum from Asia in the 1960s and subsequently spread to Africa. It is not known whether alleles that confer SP resistance also arose independently in Africa. We defined the coding region and microsatellite haplotypes of dhfr alleles in P. falciparum collected in Kilifi, Kenya, during 1987--2006, which spans the period when SP was first introduced. Isolates that carried a double-mutant or triple-mutant dhfr allele were detected at a low frequency, even during 1987--1988. Each of 2 double mutants carried a unique haplotype, and both were related to wild-type haplotypes from the same population. The number of isolates that carried a triple-mutant dhfr allele increased rapidly after introduction of SP and shared the haplotype of the triple mutant derived form Asia. We observed no triple-mutant alleles with haplotypes related to those of the Africa-derived wild-type and double-mutant alleles.

Background Previous investigations indicate that methotrexate, an old anticancer drug, could be used at low doses to treat malaria. A phase I evaluation was conducted to assess the safety and pharmacokinetic profile of this drug in healthy adult male Kenyan volunteers. Methods Twenty five healthy adult volunteers were recruited and admitted to receive a 5 mg dose of methotrexate/day/5 days. Pharmacokinetics blood sampling was carried out at 2, 4, 6, 12 and 24 hours following each dose. Nausea, vomiting, oral ulcers and other adverse events were solicited during follow up of 42 days. Results The mean age of participants was 23.9 ± 3.3 years. Adherence to protocol was 100%. No grade 3 solicited adverse events were observed. However, one case of transiently elevated liver enzymes, and one serious adverse event (not related to the product) were reported. The maximum concentration (Cmax) was 160-200 nM and after 6 hours, the effective concentration (Ceff) was Conclusion Low-dose methotraxate had an acceptable safety profile. However, methotrexate blood levels did not reach the desirable Ceff of 250-400-nM required to clear malaria infection in vivo. Further dose finding and safety studies are necessary to confirm suitability of this drug as an anti-malarial agent.

The present research is focused on cultivation of microalgae strain Chlorella vulgaris for bio-fixation of CO2 coupled with biomass production. In this regard, a single semi-batch vertical tubular photobioreactor and four similar photobioreactors in series have been employed. The concentration of CO2 in the feed stream was varied from 2 to 12 % (v/v) by adjusting CO2 to air ratio. The amount of CO2 capture and algae growth were monitored by measuring decrease of CO2 concentration in the gas phase, microalgal cell density, and algal biomass production rate. The results show that 4 % CO2 gives maximum amount of biomass (0.9 g L(-1)) and productivity (0.118 g L(-1) day(-1)) of C. vulgaris in a single reactor. In series reactors, average productivity per reactor found to be 0.078 g L(-1) day(-1). The maximum CO2 uptake for single reactor also found with 4 % CO2, and it is around 0.2 g L(-1) day(-1). In series reactors, average CO2 uptake is 0.13 g L(-1) day(-1) per reactor. TOC analysis shows that the carbon content of the produced biomass is around 40.67 % of total weight. The thermochemical characteristics of the cultivated C. vulgaris samples were analyzed in the presence of air. All samples burn above 200 °C and the combustion rate become faster at around 600 °C. Almost 98 wt% of the produced biomass is combustible in this range.

Food fortified with folic acid has been available for consumption in North America for over a decade. This strategy has led to an increase in folate levels in the general population and, more importantly, a significant decrease in the incidence of neural tube defects. However, this increase in folate intake has been associated with a greater risk of cancer disease. Many African countries are now embracing this concept; however, because folate promotes malaria parasite division, as it does in cancer cells, there is a possibility of malaria exacerbation if folate intake is increased. A precedent for such a concern is the now compelling evidence showing that an increase in iron intake can lead to a higher malaria risk; as a result, mass administration of iron in malaria-endemic areas is not recommended. In this article, we review work on the effect of folate on malaria parasites. Although this topic has received little research attention, the available data suggest that the increase in folate concentration could be associated with an increase in malaria infection. Thus, the introduction of food fortification with folic acid in malaria-endemic areas should be attended by precautionary programs to monitor the risk of malaria.

Background: The human malaria parasite Plasmodium falciparum has evolved drug evasion mechanisms to all available antimalarials. The combination of amodiaquine-artesunate is among the drug of choice for treatment of uncomplicated malaria. In this combination, a short-acting, artesunate is partnered with long-acting, amodiaquine for which resistance may emerge rapidly especially in high transmission settings. Here, we used a rodent malaria parasite Plasmodium berghei ANKA as a surrogate of P. falciparum to investigate the mechanisms of amodiaquine resistance. Methods: We used the ramp up approach to select amodiaquine resistance. We then employed the 4-Day Suppressive Test to measure the resistance level and determine the cross-resistance profiles. Finally, we genotyped the resistant parasite by PCR amplification, sequencing and relative quantitation of mRNA transcript of targeted genes. Results: Submission of the parasite to amodiaquine pressure yielded resistant line within thirty-six passages. The effective doses that reduced 90% of parasitaemia (ED90) of the sensitive and resistant lines were 4.29mg/kg and 19.13mg/kg respectively. The selected parasite retained resistance after ten passage cycles in the absence of the drug and freezing at -80ºC for one month with ED90 of 20.34mg/kg and 18.22mg/kg. The parasite lost susceptibility to chloroquine by (6-fold), artemether (10-fold), primaquine (5-fold), piperaquine (2-fold) and lumefantrine (3-fold). Sequence analysis of Plasmodium berghei chloroquine-resistant transporter revealed His95Pro mutation. We found no variation in the nucleotide sequences of Plasmodium berghei multidrug resistance gene-1 (Pbmdr1), Plasmodium berghei deubiquitinating enzyme-1 or Plasmodium berghei Kelch13 domain. However, high mRNA transcripts of essential transporters; Pbmdr1, V-type/H+ pumping pyrophosphatase-2 and sodium hydrogen ion exchanger-1 and Ca2+/H+ antiporter accompanies amodiaquine resistance. Conclusions: The selection of amodiaquine resistance yielded stable “multidrug-resistant’’ parasites and thus may be used to study shared resistance mechanisms associated with other antimalarial drugs. Genome-wide analysis of the parasite may elucidate other functionally relevant genes controlling AQ resistance in P. berghei.

The mechanisms of drug resistance development in the Plasmodium falciparum parasite to lumefantrine (LUM), commonly used in combination with artemisinin, are still unclear. We assessed the polymorphisms of Pfmspdbl2 for associations with LUM activity in a Kenyan population. MSPDBL2 codon 591S was associated with reduced susceptibility to LUM ( P = 0.04). The high frequency of Pfmspdbl2 codon 591S in Kenya may be driven by the widespread use of lumefantrine in artemisinin combination therapy (Coartem).