Abstract Pomegranate juice (PJ) has gained popularity because of its associated antioxidant, antimicrobial, anticancer, and anti-inflammatory properties. However, its effects against epidemiologically significant foodborne viruses have not been investigated. In the absence of culturable human noroviruses, feline calicivirus (FCV-F9), murine norovirus (MNV-1), and MS2 (ssRNA) bacteriophage were used as foodborne viral surrogates. The aim of this research was to study the effects of PJ and pomegranate polyphenols (PP) on foodborne viral infectivity. Viruses at high ( approximately 7 log(10) PFU/mL) or low ( approximately 5 log(10) PFU/mL) titers were mixed with equal volumes of PJ, 8, 16, and 32 mg/mL of PP, or water (control) and incubated for 1 h at room temperature. Viral infectivity after treatments was evaluated using standardized plaque assays. PJ decreased the titer of FCV-F9, MNV-1, and MS2 by 2.56, 1.32, and 0.32 log(10) PFU/mL, respectively, for low titers and 1.20, 0.06, and 0.63 log(10) PFU/mL, respectively, for high titers. Interestingly, FCV-F9 was undetectable after exposure to the three tested PP solutions using both low and high titers. MNV-1 at low initial titers was reduced by 1.30, 2.11, and 3.61 log(10) PFU/mL and at high initial titers by 1.56, 1.48, and 1.54 log(10) PFU/mL with 4, 8, and 16 mg/mL of PP treatment, respectively. MS2 at low initial titers was reduced by 0.41, 0.45, and 0.93 log(10) PFU/mL and at high initial titers by 0.32, 0.41, and 0.72 log(10) PFU/mL after 4, 8, and 16 mg/mL of PP treatment, respectively. PJ and PP resulted in titer reductions of foodborne virus surrogates after 1 h exposure, showing promise for use in hurdle technologies and/or for therapeutic or preventive use. To suggest the use of PJ and PP as natural remedies for foodborne viral illness prevention, their mechanism of action against viral infectivity needs to be further investigated.

In this paper we propose a specific sandwich immunoassay method for human-immunoglobulin G (HIgG). This immunoassay protocol takes advantage of sandwich binding of primary and secondary antibodies for increased specificity. Polystyrene microspheres (PS) serve as immobilizing support, site for sandwich immunoassay and then subsequently used for chemiluminescence (CL) detections. In this sandwich immunoassay, PS microspheres were modified with the primary anti-HIgG (Ab(1)) via electrostatic interaction, while CdTe nanoparticles (CdTeNPs) were modified with horseradish peroxidase labeled anti-HIgG (Ab(2)) via covalent binding. Antigen HIgG (Ag) was specifically captured by the first and secondary antibody and form sandwich immunoassay format. Combination of the remarkable sensitivity of CL method and the use of CdTe NPs as anti-HIgG-HRP carrier for the enzymatic signal amplification, provide a linear response range of HIgG from 0.01 to 300ngmL(-1) with an extremely low detection limit of 0.3pgmL(-1). This immunoassay system has many desirable merits including sensitivity, accuracy, and little required instrumentation. The assay results were compared with enzyme-linked immunosorbent assay (ELISA), and showed relatively good reliability. Significantly the new protocol may become quite promising technique for protein immune-detection as well as DNA analysis and other biological analyses.

Carbohydrate binding modules (CBMs) are specialized proteins that bind to polysaccharides and oligosaccharides. Caldanaerobius polysaccharolyticus Man5ACBM16-1/CBM16-2 bind to glucose-, mannose-, and glucose/mannose-configured substrates. The crystal structures of the two proteins represent the only examples in CBM family 16, and studies that evaluate the roles of amino acid residues in ligand binding in this family are lacking. In this study, we probed the roles of amino acids (selected based on CBM16-1/ligand co-crystal structures) on substrate binding. Two tryptophan (W20 and W125) and two glutamine (Q81 and Q93) residues are shown to be critical in ligand binding. Additionally, several polar residues that flank the critical residues also contribute to ligand binding. The CBM16-1 Q121E mutation increased affinity for all substrates tested, while the Q21G and N97R mutants exhibited decreased substrate affinity. We solved CBM/substrate co-crystal structures to elucidate the molecular basis of the increased substrate binding by CBM16-1 Q121E. The Q121, Q21, and N97 residues can be manipulated to fine tune ligand binding by the Man5A CBMs. Surprisingly, none of the eight residues investigated was absolutely conserved in CBM family 16. Thus, the critical residues in the Man5A CBMs are either not essential for substrate binding in the other members of this family, or the two CBMs are evolutionarily distinct from the members available in the current protein database. Man5A is dependent on its CBMs for robust activity, and insights from this study should serve to enhance our understanding of the interdependence of its catalytic and substrate binding modules.

PURPOSE: It has been reported that priming irradiation or conditioning irradiation with a low dose of X-rays in the range of 0.02-0.1 Gy induces a p53-dependent adaptive response in mammalian cells. The aim of the present study was to clarify the effect of space radiations on the adaptive response. METHODS AND MATERIALS: Two human lymphoblastoid cell lines were used; one cell line bears a wild-type p53 (wtp53) gene, and another cell line bears a mutated p53 (mp53) gene. The cells were frozen during transportation on the space shuttle and while in orbit in the International Space Station freezer for 133 days between November 15, 2008 and March 29, 2009. After the frozen samples were returned to Earth, the cells were cultured for 6 h and then exposed to a challenging X-ray-irradiation (2 Gy). Cellular sensitivity, apoptosis, and chromosome aberrations were scored using dye-exclusion assays, Hoechst33342 staining assays, and chromosomal banding techniques, respectively. RESULTS: In cells exposed to space radiations, adaptive responses such as the induction of radioresistance and the depression of radiation-induced apoptosis and chromosome aberrations were observed in wtp53 cells but not in mp53 cells. CONCLUSION: These results have confirmed the hypothesis that p53-dependent adaptive responses are apparently induced by space radiations within a specific range of low doses. The cells exhibited this effect owing to space radiations exposure, even though the doses in space were very low.

Mutations in PKD1, which encodes polycystin-1 (PC1), contribute to >85% of cases of autosomal dominant polycystic kidney disease (ADPKD). The planar cell polarity (PCP) pathway is necessary for the oriented cell division and convergent extension that establishes and maintains the structure of kidney tubules, but the role of this pathway in the pathophysiology of ADPKD is incompletely understood. Here, we show that inactivation of Pkd1 in postnatal developing mouse kidneys leads to a defect in oriented cell division in precystic kidney tubules. We also observed this defect in precystic Pkd1-inactivated mature kidneys subjected to ischemia-reperfusion injury as a "third hit." Cystic kidneys exhibited striking upregulation and activation of Frizzled 3 (Fz3), a regulator of PCP, and its downstream effector, CDC42. Precystic kidneys demonstrated upregulation of CDC42, but the localization of the polarity proteins Par3 and Par6 was similar to control. Fz3 was expressed on the cilia of cystic kidneys but barely detected on the cilia of normal kidneys. In vitro, PC1 and Fz3 antagonized each other to control CDC42 expression and the rate of cell migration in HEK293T cells. Taken together, our data suggest that PC1 controls oriented cell division and that aberrant PCP signaling contributes to cystogenesis.

We have developed a unique metal nanoparticle (mNPs)-based assay to detect sequence-specific interactions between transcription factor and its corresponding DNA-binding elements. This assay exploits the interparticle-distance dependent optical properties of noble mNPs as sensing element and utilizes specific protein-DNA interactions to control the dispersion status of the mNPs. The assay involves two sets of double-stranded (ds)DNA modified-mNPs, each carrying a half site segment of a functional DNA sequence for the protein of interest. Each of these half sites is designed to contain a short complementary sticky end that introduces base-pairing forces to facilitate particle aggregation and to form a transient full dsDNA sequence. The detection of specific protein-DNA binding is founded on the premise that the mixture of these two sets of dsDNA-mNPs experiences a remarkable particle aggregation under certain salt conditions; whereas the aggregation can be retarded in the presence of a specific protein that binds and stabilizes the transient full dsDNA structure and therefore introduces steric protection forces between particles. We have demonstrated the concept using estrogen receptor alpha and its response elements, with gold and silver NPs as the sensing platform. UV-vis spectroscopy, transmission electron spectroscopy, and dynamic light scattering measurements were conducted to provide full characterization of the particle aggregation/dispersion mechanism. Differing from most of the mNP-based colorimetric sensors that are designed based on the analyte-induced aggregation mechanism, current protein binding-stabilization sensing strategy reduces the false signals caused by unrelated particle destabilizing effects. It is expected that this assay principle can be directed toward other transcription factors by simply changing the recognition sequence to form different segmented dsDNA-mNP constructs.

During the growing seasons of 2006-2008, feeding tests and field studies were conducted in Beijing, China, to investigate the effects of transgenic Bacillus thuringiensis (Bt) poplar (BGA-5) expressing the Cry3A protein (0.0264-0.0326% of the total soluble protein) on target and non-target pests and the arthropod community. The effects of BGA-5 on the target pest Plagiodera versicolora (Coleoptera, Chrysomelidae) and a non-target pest Clostera anachoreta (Lepidoptera, Notodontidae), were assessed under laboratory conditions. Total mortality of P. versicolora larvae fed with BGA-5 leaves was significantly higher than that of the control (P < 0.05). The exuviation index of P. versicolora larvae fed with BGA-5 tended to be higher than that of CK, but it was not significantly different. The pupation rate and eclosion rate of the survived larvae fed with BGA-5 were lower than that of CK, but it was also not significantly different. Additionally, no significant differences were detected in the mortality, exuviations index, pupation rate, or eclosion rate of C. anachoreta fed with leaves of transgenic and non-transgenic poplars. Furthermore, the arthropod communities in the Bt poplar and CK field stands were similar, as indicated by four diversity indices (Berge-Parker index, Shannon-Wiener indices, evenness index, and Simpson's inverted index) and the Bray-Curtis index. Therefore, the Bt-Cry3A poplar decreased damage by the target pest (P. versicolora), had no effects on a non-target pest (C. anachoreta), and generally did not have any significant negative effect on the poplar arthropod community.

Abstract Cytokine-induced killer (CIK) cells have shown antitumor activity against several tumor cells both in vitro and in vivo. This study reports on the large-scale expansion of CIK cells and also present preliminary results from a pilot clinical trial. Sixteen (16) patients with renal cell carcinoma (RCC), all of whom had metastases after radical nephrectomy and adjuvant therapy using interferon-alpha (IFN-alpha) and/or interleukin-2 (IL-2), were treated with CIK cells. CIK cells were generated from peripheral blood mononuclear cells (PBMCs) and incubated in the presence of IFN-gamma followed by OKT3 and IL-2. Treatment schedule consisted of two to three cycles of CIK cell infusions at an interval of 3 weeks. A total of 46 infusions were administered to 16 metastatic RCC (mRCC) patients. The median number of transferred cells per treatment was 6.7 x 10(9) (range, 2.5-12.3). At a 60:1 effector-target cell ratio, CIK cells killed 51.4% and 32.1% of two human kidney tumor cell lines (293 and SK-RC-42), respectively. After CIK cell infusion, the percentage of CD3(+), CD8(+), CD3(+)CD56(+), and NKG2D(+) cells and the intracellular products of two type 1 cytokines (IFN-gamma and tumor necrosis factor alpha) significantly increased in the patients' PBMCs. Toxicity was minimal, and there were no immediate adverse reactions to the infusions. Three (3) patients had complete response, 1 patient had partial response, and 6 patients had stable disease. These results showed that adoptive CIK cell immunotherapy is a safe and effective treatment, which may have essential benefits for the improvement of the immunologic function in mRCC patients and play an important role in the treatment of mRCC.

A sensitive QDs-based RLS assay method for the detection of Newcastle disease virus (NDV) antibody has been developed. CdTe quantum dots (QDs) were conjugated with Newcastle disease virus and used as RLS-based probes to detect NDV antibody. The electrostatic interaction between CdTe QDs and NDV resulted in enhanced resonance light scattering (RLS) signal characterized at 555nm. Upon the addition of NDV antibody, QDs-NDV formed dispersive immunocomplex that can decrease the RLS signal. The decreased RLS intensity at 555nm (DeltaI(RLS)) was linearly proportional to the concentration of NDV antibody (C(anti-NDV)) in the range of 0.5-50ng/mL, with correlation coefficient of 0.974 and detection limit of 0.1ng/mL under the optimization conditions. The proposed method was applied to the determination of NDV antibody in spiked samples with satisfactory results.

BACKGROUND:: The mechanisms of ventilator-induced lung injury, an iatrogenic inflammatory condition induced by mechanical ventilation, are not completely understood. Toll-like receptor 4 (TLR4) signaling via the adaptor protein myeloid differentiation factor 88 (MyD88) is proinflammatory and plays a critical role in host immune response to invading pathogen and noninfectious tissue injury. The role of TLR4-MyD88 signaling in ventilator-induced lung injury remains incompletely understood. METHODS:: Mice were ventilated with low or high tidal volume (HTV), 7 or 20 ml/kg, after tracheotomy for 4 h. Control mice were tracheotomized without ventilation. Lung injury was assessed by: alveolar capillary permeability to Evans blue albumin, wet/dry ratio, bronchoalveolar lavage analysis for cell counts, total proteins and cytokines, results of histopathological examination of the lung, and plasma cytokine levels. RESULTS:: Wild-type mice subjected to HTV had increased pulmonary permeability, inflammatory cell infiltration/lung edema, and interleukin-6/macrophage-inflammatory protein-2 in the lavage compared with control mice. In HTV, levels of inhibitor of kappaB alpha decreased, whereas phosphorylated extracellular signal-regulated kinases increased. TLR4 mutant and MyD88 mice showed markedly attenuated response to HTV, including less lung inflammation, pulmonary edema, cell number, protein content, and the cytokines in the lavage. Furthermore, compared with wild-type mice, both TLR4 mutant and MyD88 mice had significantly higher levels of inhibitor of kappaB alpha and reduced extracellular signal-regulated kinase phosphorylation after HTV. CONCLUSIONS:: TLR4-MyD88 signaling plays an important role in the development of ventilator-induced lung injury in mice, possibly through mechanisms involving nuclear factor-kappaB and mitogen-activated protein kinase pathways.

A novel method was developed for the analysis of four beta-blockers, namely sotalol, carteolol, bisoprolol, and propranolol, in human urine by coupling carrier-mediated liquid phase microextraction (CM-LPME) to high performance liquid chromatography (HPLC). By adding an appropriate carrier in organic phase, simultaneous extraction and enrichment of hydrophilic (sotalol, carteolol, and bisoprolol) and hydrophobic (propranolol) drugs were achieved. High enrichment factors were obtained by optimizing the compositions of the organic phase, the acceptor solution, the donor solution, the stirring rate, and the extraction time. The linear ranges were from 0.05 to 10.0mgL(-1) for sotalol and carteolol, and from 0.05 to 8.0mgL(-1) for bisoprolol and propranolol. The limits of detection (S/N=3) were 0.01mgL(-1) for sotalol, carteolol, and bisoprolol, and 0.005mgL(-1) for propranolol. The relative standard deviations were lower than 6%. The developed method exhibited high analyte preconcentration and excellent sample clean-up effects with little solvent consumption and was found to be sensitive and suitable for simultaneous determination of the above four drugs spiked in human urine. Furthermore, the successful analysis of propranolol in real urine specimens revealed that the determination of beta-blockers in human urine is feasible using the present method.

Abstract Microfluidic methods for cellular studies can significantly reduce costs due to reduced reagent and biological specimen requirements compared with many traditional culture techniques. However, current types of readouts are limited and this lack of suitable readouts for microfluidic cultures has significantly hindered the application of microfluidics for cell-based assays. The In-Cell Western (ICW) technique uses quantitative immunocytochemistry and a laser scanner to provide an in situ measure of protein quantities in cells grown in microfluidic channels of arbitrary geometries. The use of ICWs in microfluidic channels was validated by a detailed comparison with current macroscale methods and shown to have excellent correlation. Transforming growth factor-beta-induced epithelial-to-mesenchymal transition of an epithelial cell line was used as an example for further validation of the technique as a readout for soluble-factor-based assays performed in high-throughput microfluidic channels. The use of passive pumping for sample delivery and laser scanning for analysis opens the door to high-throughput quantitative microfluidic cell-based assays that integrate seamlessly with existing high-throughput infrastructure.

In 2005 a pollution accident occurred in the Songhua River, which is geographically located next to groundwater supply plants. This caused public concern about the transport and fate of nitrobenzene (NB) in the groundwater. This paper discusses the mechanisms and effects of the transport and fate of NB in groundwater based on pilot scale experiments conducted in the laboratory, including a simulation experiment, bench-scale batch tests and a one-dimensional numerical model. Parallel batch tests showed that the adsorption of NB to the clay and sand followed the Langmuir-type isotherm, and clay had a greater NB adsorption capacity than sand. NB biodegradation in different conditions was well fitted by the Monod equation and the q(max) values varied from 0.018 to 0.046 h(-1). Results indicated that NB's biodegradation was not affected by the initial NB concentration. Numerical modeling results indicated a good match between computed and observed data, and in the prediction model NB entered the groundwater after the pollution accident. However, the highest concentration of NB was much lower than the allowable limit set by the national standard (0.017 mg/L).

Manganese superoxide dismutase (Mn-SOD; SOD(2)) is an important antioxidant defense enzyme. In this study, a full-length Mn-SOD cDNA was cloned from the cDNA library of Phascolosoma esculenta. The cDNA is 1385 bp in length, including an open reading frame (ORF) of 681 bp encoding 226 amino acids. The predicted protein has a calculated molecular weight of 25.2 kDa and a theoretical isoelectric point of 5.96. BLAST analysis revealed the predicted protein shared 70% identity with homologues in Caenorhabditis elegans and Gallus gallus. The SOD gene was inserted into E. coli expression plasmid pET-28a (+) to produce pET-SOD. The recombinant Mn-SOD of P. esculenta was expressed following IPTG induction, and verified by Western blot analysis using antiserum from immunized mice. Furthermore, fluorescent real-time PCR analysis revealed varying degrees of induction of SOD(2) mRNA expression in the blood of P. esculenta exposed to heavy metals (Cd(2+), Cu(2+) and Zn(2+)) and thermal stresses (4 degrees C and 37 degrees C).

A new detection format for multiplexed analysis based on fluorescent encoded magnetic composite nanoparticles is presented. Two kinds of virus were analyzed by this new method: equine influenza virus (EIV) and equine infectious anemia virus (EIAV). Firstly, EIV antigen and EIAV antigen were conjugated to two kinds of fluorescent encoded magnetic composite nanoparticles, while the green-emitting CdTe quantum dots (QDs) were attached to the antibody of EIV and EIAV. Then both green-emitting CdTe QD-labeled antibodies and antigens labeled with fluorescent encoded magnetic composite nanoparticles were used to form an immunoassay system for the detection of EIV and EIAV antigens. The method is time-saving and has higher sensitivity (1.3 ng mL(-1) for EIV antigens and 1.2 ng mL(-1) for EIAV antigens) than the conventional methods. A competitive immunoassay method based on this analysis system was used to detect EIV and EIAV antigens in spiked serum samples with satisfactory results.

Gaseous microemboli (GME) remain a challenge for cardiopulmonary bypass procedures in adult as well as pediatric cardiac surgery patients. The present study tested the effectiveness of a new adult membrane oxygenator in models both with and without an integrated arterial filter to evaluate GME trapping capability and determine membrane pressure drops at various flow rates and temperatures. The experimental circuit included a RotaFlow centrifugal blood pump, Quadrox-i (n = 8) or Quadrox (n = 8) adult microporous membrane oxygenator, and Sorin adult tubing package. A Sorin Cardiovascular VVR 4000i venous reservoir served as pseudo-patient. The circuit was primed with 900 mL heparinized human red blood cells and 300 mL Lactated Ringer's solution. The final hematocrit was 36%. Tests were performed at different flow rates (4 L/min, 5 L/min, and 6 L/min) and temperatures (35 degrees and 30 degrees). Five mL of bolus air was injected into the venous line over 15 seconds using a syringe connected to a 3/8 x 1/2 luer connector. The Quadrox-i adult microporous membrane oxygenator with integrated arterial filter had a similar pressure drop at 4 L/min and 35 degrees C compared with Quadrox membrane oxygenator whereas it had higher pressure drops at 5 L/min and 6 L/min (p < .001). Quadrox-i adult microporous membrane oxygenator reduced the total emboli count and total emboli volume delivered to the pseudo-patient at all flow rates (p < .001).The emboli handling of Quadrox-i adult microporous membrane oxygenator was not affected by flow rate and temperature. Compared with the traditional Quadrox oxygenator, Quadrox-i adult microporous membrane oxygenator with integrated arterial filter and Softline coating has improved GME handling capacity.

Ferromagnetic carbon-coated Fe nanoparticles (core size of 15 nm, saturated magnetization of M(s) = 218 emu g(-1) and coercivity of H(c) = 62 Oe), fabricated at a mild temperature, demonstrate a strong ability to effectively remove more than 95 wt% of Cr(vi) in waste water via carbon shell physical adsorption, which is much higher than the commercially available Fe NPs.

OBJECTIVES: To explore the effect of transient continuous subcutaneous insulin infusion (CSII) on beta cell function, insulin resistance and vascular endothelial injury in newly diagnosed type 2 diabetic patients and its potential mechanism. METHODS: Ten patients with newly diagnosed type 2 diabetes mellitus (T2DM) accepted CSII for two weeks. Intravenous glucose tolerance test (IVGTT) and hyperinsulinemia euglycemia clamp test were performed before and after CSII. Serum soluble E-selectin (sE-selectin) was used to evaluate the injury of vascular endothelial cell, while serum high sensitivity C reactive protein (hsCRP) and soluble CD(14) (sCD(14)) were both used to assess inflammatory condition. RESULTS: (1) Compared with those before treatment, the blood glucose levels of IVGTT, the area under the curve of the blood glucose, glycosylated hemoglobin, TC and LDL-C in the patients were decreased after CSII (P < 0.05 or 0.01). (2) Compared with those before treatment, the insulin levels of IVGTT (except the fasting insulin), the area under the curve of insulin and acute insulin response were all increased after CSII (P < 0.05 or 0.01). (3) Compared with that before treatment, the glucose infusion ratio in the clamp test [(3.46 +/- 1.66) mgxkg(-1)xmin(-1) increased to (7.14 +/- 2.37) mgxkg(-1)xmin(-1)] and HOMA-beta elevated, while HOMA-IR declined (P < 0.05 or 0.01 in all). (4) Compared with those before treatment, the levels of serum sE-selectin, sCD(14) and hsCRP were decreased (P < 0.01, except for hsCRP). CONCLUSION: Transient intensive insulin therapy in patients with newly diagnosed T2DM is useful to restore beta cell function, attenuate insulin resistance, repair vascular endothelial injury and improve the disorder of blood sugar and lipid. The mechanism may be related with the inhibition of inflammation in patients.

Sperm collection methods and the effect of osmolality, ions, sugar, temperature, pH and dilution ratio on sperm motility were investigated in guppies Poecilia reticulata. The present study revealed that the sperm was motile in a wide range of osmolalities (200-470 mOsm kg(-1)) either in Hanks balanced-salt solution (HBSS) or in non-electrolyte solutions such as glucose or sucrose. Sperm collected from crushing testes yielded lower motility and shorter motility duration than samples collected without crushing but gentle disruption. Dilution ratios within the range of 1:50 to 1:500 of sperm to HBSS had minimal effect on sperm motility during extended refrigerated storage. Examination of storage temperature showed that refrigerated storage at 4 degrees C was superior to room temperature (25 degrees C). Sperm was found to tolerate a wide range of pH from 5.6 to 7.8, but motility was affected negatively by pH values >7.8.

Quinine resistance (QNR) in Plasmodium falciparum has been detected in many malaria-endemic regions of the world. Genetic polymorphisms in at least four genes are implicated in QN susceptibility, and their significance often depends on the genetic background of the parasites. In this study, we have culture-adapted 60 P. falciparum clinical isolates from the China-Myanmar border and assessed their in vitro responses to QN. Our results showed that >50%parasite isolates have displayed reduced sensitivity to QN with the half maximal inhibitory concentration (IC50) above 500 nM. Genotyping of pfcrt found that an overwhelming proportion of the parasite population had the chloroquine-resistant genotype, whereas pfmdr1 mutation genotypes and gene amplification were rare. Genotyping of P. falciparum Na(+)/H(+) exchanger (pfnhe1) gene at the minisatellite ms4760 locus identified 10 hyplotypes. Haplotype 7, which harbors three copies of the DNNND repeat, was the most predominant, accounting for nearly half of the parasite isolates. Correlation studies did not reveal significant association of the polymorphisms in pfcrt and pfmdr1 genes with QN response. However, the ms4760 haplotypes were highly associated with in vitro QN responses. In particular, parasite isolates with increased DNNND copy number tended to have significantly reduced QN susceptibility, whereas parasite isolates with higher NHNDNHNNDDD copy number had increased QN susceptibility. This study provided further support for the importance of pfnhe1 polymorphisms in influencing QNR in P. falciparum.