Corrigendum: Enhanced biodegradation of phenanthrene and anthracene using a microalgal-bacterial consortium.

[This corrects the article DOI: 10.3389/fmicb.2023.1227210.].

Enhanced biodegradation of phenanthrene and anthracene using a microalgal-bacterial consortium.

Polycyclic aromatic hydrocarbons (PAHs) are chemicals that are released into the environment during activities of the petroleum industry. The bioaccumulation, carcinogenic and mutagenic potential of PAHs necessitates the bioremediation of these contaminants. However, bioremediation of PAHs has a number of limitations including the inability of a single microbe to degrade all of the PAH fraction's environmental constituents. Therefore, a different paradigm, employing microalgal-bacterial consortium (MBC), may be used to effectively remove PAHs contaminants. In this type of interaction, the microalgae and bacteria species in the consortium work together in a way that enhances the overall performance of the MBC. Bacterial species in the consortium provide essential nutrients or growth factors by degrading toxic substances and provide these to microalgae, while the microalgae species provide organic carbon for the bacterial species to grow. For the first time, the ability of Gonium pectorale (G. pectorale) microalgae to break down phenanthrene (PHE) and anthracene (ANT) was investigated. Phenanthrene was shown to be more effectively degraded by G. pectorale (98%) as compared to Bacillus licheniformis (B. licheniformis) 19%. Similarly, G. pectorale has effectively degrade anthracene (98%) as compared with B. licheniformis (45%). The consortia of G. pectorale and B. licheniformis has shown a slight increase in the degradation of PHE (96%) and ANT (99%). Our findings show that B. licheniformis did not inhibit the growth of G. pectorale and in the consortia has effectively eliminated the PAHs from the media. Therefore G. pectorale has a tremendous potential to remove PAHs from the polluted environment. Future research will be conducted to assess Gonium's capacity to eliminate PAHs that exhibit high molar masses than that of PHE and ANT.

Benzo[A]Pyrene Biodegradation by Multiple and Individual Mesophilic Bacteria under Axenic Conditions and in Soil Samples.

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.

Biogenic Synthesis and Characterization of Chitosan-CuO Nanocomposite and Evaluation of Antibacterial Activity against Gram-Positive and -Negative Bacteria.

Chitosan-copper oxide (CHT-CuO) nanocomposite was synthesized using olive leaf extract (OLE) as reducing agent and CuSO4⋅5H2O as precursor. CHT-CuO nanocomposite was prepared using an in situ method in which OLE was added to a solution of chitosan and CuSO4⋅5H2O mixture in the ratio of 1:5 (v/v) and heated at a temperature of 90 °C. The obtained CHT-CuO nanocomposite was characterized using field emission scanning electron microscopy (FE-SEM), X-ray diffraction (XRD), ultraviolet-visible (UV-Vis) spectrophotometry, energy-dispersive X-ray spectroscopy (EDAX), Fourier transform infrared spectroscopy (FTIR), and high-resolution transmission electron microscopy (TEM). TEM results indicated that CHT-CuO nanocomposite are spherical in shape with size ranging from 3.5 to 6.0 nm. Antibacterial activity of the synthesized nanocomposites was evaluated against Gram-positive (Bacillus cereus, Staphyloccous haemolytica and Micrococcus Luteus) and Gram-negative (Escherichia coli, Pseudomonas citronellolis, Pseudomonas aeruginosa, kliebisella sp., Bradyrhizobium japonicum and Ralstonia pickettii) species by cup platting or disc diffusion method. Overall, against all tested bacterial strains, the diameters of the inhibition zone of the three nanocomposites fell between 6 and 24 mm, and the order of the antimicrobial activity was as follows: CuO-1.0 > CuO-0.5 > CuO-2.0. The reference antibiotic amoxicillin and ciprofloxacin showed greater activity based on the diameter of zones of inhibition (between 15−32 mm) except for S. heamolytica and P. citronellolis bacteria strains. The nanocomposites MIC/MBC were between 0.1 and 0.01% against all tested bacteria, except S. heamolityca (>0.1%). Based on MIC/MBC values, CuO-0.5 and CuO-1.0 were more active than CuO-2.0, in line with the observations from the disc diffusion experiment. The findings indicate that these nanocomposites are efficacious against bacteria; however, Gram-positive bacteria were less susceptible. The synthesized CHT-CuO nanocomposite shows promising antimicrobial activities and could be utilized as an antibacterial agent in packaging and medical applications.

Non-enzymatic detection of miR-21 in cancer cells using a homogeneous mix-and-read smart probe assay.

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 × 1011 c.p.sM-1. More importantly, we showed that the smart probe-based method is also sensitive and selective for miR-21 when applied to crude extractions from MCF-7 cancer cell line at room temperature, with the results showing high fluorescence signals for the MCF-7 samples while showing much less signals for samples that did not contain miR-21. Thus, this new smart probe system constitutes a homogeneous, mix-and-read detection technique that can provide reliable diagnostics of miR-21 cancer biomarker at room temperature.

Degradation of benzo[a]pyrene by halophilic bacterial strain Staphylococcus haemoliticus strain 10SBZ1A.

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.

Biodegradation of selected hydrocarbons by novel bacterial strains isolated from contaminated Arabian Gulf sediment.

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.

Preparation of Silver/Chitosan Nanofluids Using Selected Plant Extracts: Characterization and Antimicrobial Studies Against Gram-Positive and Gram-Negative Bacteria.

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-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 (8.0-36 nm) than those produced with DPLE (10-43 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.

Current Status of and Future Perspectives in Bacterial Degradation of Benzo[a]pyrene.

Benzo[a]pyrene (BaP) is one the main pollutants belonging to the high-molecular-weight PAHs (HMW-PAHs) class and its degradation by microorganisms remains an important strategy for its removal from the environment. Extensive studies have been carried out on the isolation and characterisation of microorganisms that can actively degrade low-molecular-weight PAHs (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 chemical 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.

Characterisation and microbial community analysis of lipid utilising microorganisms for biogas formation.

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 <4 ml.L-1.day-1 at 40 g.L-1. Similar results were observed with the use of LCFAs. However, GLYC concentrations up to 100g.L-1 did not affect the rate of biogas formation. Acidic pH, temperature > = 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.

Current Status of the Degradation of Aliphatic and Aromatic Petroleum Hydrocarbons by Thermophilic Microbes and Future Perspectives.

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 medium, 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 was carried out. This work has identified the characteristics of thermophiles, and unraveled specific catabolic pathways of petroleum products that are only found with 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 were proposed.

Biodegradation of high-molecular-weight polycyclic aromatic hydrocarbons under anaerobic conditions: Overview of studies, proposed pathways and future perspectives.

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.

Amodiaquine resistance in Plasmodium berghei is associated with PbCRT His95Pro mutation, loss of chloroquine, artemisinin and primaquine sensitivity, and high transcript levels of key transporters.

Background: The human malaria parasite Plasmodium falciparum has evolved complex drug evasion mechanisms to all available antimalarials. To date, 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 serial technique to select amodiaquine resistance by submitting the parasites to continuous amodiaquine pressure. We then employed the 4-Day Suppressive Test to monitor emergence of resistance 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 P. berghei ANKA to amodiaquine pressure yielded resistant parasite within thirty-six passages. The effective dosage that reduced 90% of parasitaemia (ED 90) of sensitive line and resistant line were 4.29mg/kg and 19.13mg/kg, respectively. After freezing at -80ºC for one month, the resistant parasite remained stable with an ED 90 of 18.22mg/kg. Amodiaquine resistant parasites are also resistant to chloroquine (6fold), artemether (10fold), primaquine (5fold), piperaquine (2fold) and lumefantrine (3fold). Sequence analysis of Plasmodium berghei chloroquine resistant transporter revealed His95Pro mutation. No variation was identified in Plasmodium berghei multidrug resistance gene-1 (Pbmdr1), Plasmodium berghei deubiquitinating enzyme-1 or Plasmodium berghei Kelch13 domain nucleotide sequences. Amodiaquine resistance is also accompanied by high mRNA transcripts of key transporters; Pbmdr1, V-type/H+ pumping pyrophosphatase-2 and sodium hydrogen ion exchanger-1 and Ca 2+/H + antiporter. Conclusions: Selection of amodiaquine resistance yielded stable "multidrug-resistant'' parasites and thus may be used to study common resistance mechanisms associated with other antimalarial drugs. Genome wide studies may elucidate other functionally important genes controlling AQ resistance in P. berghei.

Mini review: Update on bioaugmentation in anaerobic processes for biogas production.

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.

Characterization of aerobic oil and grease-degrading bacteria in wastewater.

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.

Bioaugmentation: An Emerging Strategy of Industrial Wastewater Treatment for Reuse and Discharge.

A promising long-term and sustainable solution to the growing scarcity of water worldwide is to recycle and reuse wastewater. In wastewater treatment plants, the biodegradation of contaminants or pollutants by harnessing microorganisms present in activated sludge is one of the most important strategies to remove organic contaminants from wastewater. However, this approach has limitations because many pollutants are not efficiently eliminated. To counterbalance the limitations, bioaugmentation has been developed and consists of adding specific and efficient pollutant-biodegrading microorganisms into a microbial community in an effort to enhance the ability of this microbial community to biodegrade contaminants. This approach has been tested for wastewater cleaning with encouraging results, but failure has also been reported, especially during scale-up. In this review, work on the bioaugmentation in the context of removal of important pollutants from industrial wastewater is summarized, with an emphasis on recalcitrant compounds, and strategies that can be used to improve the efficiency of bioaugmentation are also discussed. This review also initiates a discussion regarding new research areas, such as nanotechnology and quorum sensing, that should be investigated to improve the efficiency of wastewater bioaugmentation.

Malaria in the Era of Food Fortification With Folic Acid.

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.

The MSPDBL2 codon 591 polymorphism is associated with lumefantrine in vitro drug responses in Plasmodium falciparum isolates from Kilifi, Kenya.

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).

Piperaquine and Lumefantrine resistance in Plasmodium berghei ANKA associated with increased expression of Ca2+/H+ antiporter and glutathione associated enzymes.

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), Ca(2+)/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.

Polymorphisms in Plasmodium falciparum chloroquine resistance transporter and multidrug resistance 1 genes: parasite risk factors that affect treatment outcomes for P. falciparum malaria after artemether-lumefantrine and artesunate-amodiaquine.

Adequate clinical and parasitologic cure by artemisinin combination therapies relies on the artemisinin component and the partner drug. Polymorphisms in the Plasmodium falciparum chloroquine resistance transporter (pfcrt) and P. falciparum multidrug resistance 1 (pfmdr1) genes are associated with decreased sensitivity to amodiaquine and lumefantrine, but effects of these polymorphisms on therapeutic responses to artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL) have not been clearly defined. Individual patient data from 31 clinical trials were harmonized and pooled by using standardized methods from the WorldWide Antimalarial Resistance Network. Data for more than 7,000 patients were analyzed to assess relationships between parasite polymorphisms in pfcrt and pfmdr1 and clinically relevant outcomes after treatment with AL or ASAQ. Presence of the pfmdr1 gene N86 (adjusted hazards ratio = 4.74, 95% confidence interval = 2.29 - 9.78, P < 0.001) and increased pfmdr1 copy number (adjusted hazards ratio = 6.52, 95% confidence interval = 2.36-17.97, P < 0.001 : were significant independent risk factors for recrudescence in patients treated with AL. AL and ASAQ exerted opposing selective effects on single-nucleotide polymorphisms in pfcrt and pfmdr1. Monitoring selection and responding to emerging signs of drug resistance are critical tools for preserving efficacy of artemisinin combination therapies; determination of the prevalence of at least pfcrt K76T and pfmdr1 N86Y should now be routine.