Functional characterization of a conserved membrane protein, Pbs54, involved in gamete fertilization in Plasmodium berghei.

The successful completion of gamete fertilization is essential for malaria parasite transmission, and this process can be targeted by intervention strategies. In this study, we identified a conserved gene (PBANKA_0813300) in the rodent malaria parasite Plasmodium berghei, which encodes a protein of 54 kDa (designated as Pbs54). Localization studies indicated that Pbs54 is associated with the plasma membranes of gametes and ookinetes. Functional studies by gene disruption showed that the Δpbs54 parasites had no defect in asexual proliferation, gametocyte development, or gametogenesis. However, the interactions between male and female gametes were significantly decreased compared with wild-type parasites. The Δpbs54 lines did not show a further reduction in zygote and ookinete numbers during in vitro culture, indicating that the defects were probably restricted to gamete fertilization. Consistent with this finding, mosquitoes fed on Δpbs54-infected mice showed a 30.1% reduction in infection prevalence and a 74.7% reduction in oocyst intensity. Cross-fertilization assay indicated that both male and female gametes were impaired in the Δpbs54 parasites. To evaluate its transmission-blocking potential, we obtained polyclonal antibodies from mice immunized with the recombinant Pbs54 (rPbs54) protein. In vitro assays showed that anti-rPbs54 sera inhibited ookinete formation by 42.7%. Our experiments identified Pbs54 as a fertility factor required for mosquito transmission and a novel candidate for a malaria transmission-blocking vaccine.

A novel long-term intravenous combined with local treatment with human amnion-derived mesenchymal stem cells for a multidisciplinary rescued uremic calciphylaxis patient and the underlying mechanism.

Calciphylaxis is a rare disease characterized histologically by microvessel calcification and microthrombosis, with high mortality and no proven therapy. Here, we reported a severe uremic calciphylaxis patient with progressive skin ischemia, large areas of painful malodorous ulcers, and mummified legs. Because of the worsening symptoms and signs refractory to conventional therapies, treatment with human amnion-derived mesenchymal stem cells (hAMSCs) was approved. Preclinical release inspections of hAMSCs, efficacy, and safety assessment, including cytokine secretory ability, immunocompetence, tumorigenicity, and genetics analysis in vitro, were introduced. We further performed acute and long-term hAMSC toxicity evaluations in C57BL/6 mice and rats, abnormal immune response tests in C57BL/6 mice, and tumorigenicity tests in neonatal Balbc-nu nude mice. After the preclinical research, the patient was treated with hAMSCs by intravenous and local intramuscular injection and external supernatant application to the ulcers. When followed up to 15 months, the blood-based markers of bone and mineral metabolism improved, with skin soft tissue regeneration and a more favorable profile of peripheral blood mononuclear cells. Skin biopsy after 1-month treatment showed vascular regeneration with mature noncalcified vessels within the dermis, and 20 months later, the re-epithelialization restored the integrity of the damaged site. No infusion or local treatment-related adverse events occurred. Thus, this novel long-term intravenous combined with local treatment with hAMSCs warrants further investigation as a potential regenerative treatment for uremic calciphylaxis due to effects of inhibiting vascular calcification, stimulating angiogenesis and myogenesis, anti-inflammatory and immune modulation, multidifferentiation, re-epithelialization, and restoration of integrity.

Evaluation of two Plasmodium vivax sexual stage antigens as transmission-blocking vaccine candidates.

BACKGROUND: Plasmodium vivax transmission-blocking vaccines (TBVs) are receiving increasing attention. Based on excellent transmission-blocking activities of the PbPH (PBANKA_0417200) and PbSOP26 (PBANKA_1457700) antigens in Plasmodium berghei, their orthologs in P. vivax, PVX_098655 (PvPH) and PVX_101120 (PvSOP26), were selected for the evaluation of their potential as TBVs. METHODS: Fragments of PvPH (amino acids 22-304) and PvSOP26 (amino acids 30-272) were expressed in the yeast expression system. The recombinant proteins were used to immunize mice to obtain antisera. The transmission-reducing activities of these antisera were evaluated using the direct membrane feeding assay (DMFA) using Anopheles dirus mosquitoes and P. vivax clinical isolates. RESULTS: The recombinant proteins PvPH and PvSOP26 induced robust antibody responses in mice. The DMFA showed that the anti-PvSOP26 sera significantly reduced oocyst densities by 92.0 and 84.1% in two parasite isolates, respectively, whereas the anti-PvPH sera did not show evident transmission-reducing activity. The variation in the DMFA results was unlikely due to the genetic polymorphisms of the two genes since their respective sequences were identical in the clinical P. vivax isolates. CONCLUSION: PvSOP26 could be a promising TBV candidate for P. vivax, which warrants further evaluation.

Human amnion-derived mesenchymal stem cells improved the reproductive function of age-related diminished ovarian reserve in mice through Ampk/FoxO3a signaling pathway.

BACKGROUND: Age-related diminished ovarian reserve (AR-DOR) reduced the quality of oocytes, resulting in decreased female fertility. Aging is tightly related to abnormal distribution and function of mitochondria, while mitophagy is a major process to maintain normal quality and quantity of mitochondria in cells, especially in oocytes which containing a large number of mitochondria to meet the demand of energy production during oocyte maturation and subsequent embryonic development. Ampk/FoxO3a signaling is crucial in the regulation of mitophagy. It is reported mesenchymal stem cells (MSCs) can improve ovarian function. Here we aim to explore if human amnion-derived mesenchymal stem cells (hAMSCs) are effective in improving ovarian function in AR-DOR mice and whether Ampk/FoxO3a signaling is involved. METHODS: The AR-DOR model mice were established by 32-week-old mice with 3-8 litters, significantly low serum sex hormone levels and follicle counts. The old mice were divided into 5 treatment groups: normal saline (NS, control), 1% human serum albumin (HSA, resolver), low dose (LD, 5.0 × 106cells/kg), middle dose (MD, 7.5 × 106cells/kg), and high dose (HD, 10.0 × 106cells/kg). The prepared hAMSCs were injected through tail vein. Serum sex hormone level, follicle counts, fertilization rate, gestation rate, little size, apoptosis of granulosa and stromal cells, expression level of Sod2, Ampk, and ratio of phosphorylated FoxO3a to total FoxO3a in ovaries were examined. RESULTS: Our results show that after hAMSC transplantation, the ovarian function in AR-DOR mice was significantly improved, meanwhile the apoptosis of granulosa and stromal cells in the ovaries was significantly repressed, the expression level of Ampk and the ratio of phosphorylated FoxO3a to total FoxO3a both were significantly increased, meanwhile increased Sod2 expression was also observed. CONCLUSION: Our results demonstrate hAMSC transplantation via tail-injection can improve ovarian function of AR-DOR mice through Ampk/FoxO3a signaling pathway.

Prenatal diagnosis of an aberrant ductus venosus draining into the coronary sinus using two- and three-dimensional echocardiography: a case report.

BACKGROUND: Ductus venosus (DV) abnormalities may be associated with intracardiac or extracardiac deformities, chromosomal anomalies, and/or congestive heart failure. Aberrant DV connecting with the coronary sinus (CS) is rare and the prenatal diagnosis presents challenges for most examiners. CASE PRESENTATION: A 35-year-old pregnant woman, gravida 2, para 1, was referred to our center at 27 gestational weeks for a full evaluation of fetal cardiac anomalies. Transverse scans indicated normal cardiac anatomy except for a dilated CS; we then scanned sagittal planes to clarify the reasons for the CS dilatation. High-definition flow imaging (HDFI) together with radiant flow (R-flow) imaging was used to delineate the aberrant DV returning to the CS, enabling the diagnosis. Three-dimensional (3D) technology was also used to obtain color-rendered images showing the spatial relationships of the vessels involved, thus confirming the two-dimensional (2D) diagnosis. Chromosomal analysis revealed a normal karyotype. The neonate appeared healthy and the echocardiogram showed a normal cardiac anatomy except for a dilated CS with the DV closed and imperceptible. CONCLUSIONS: The aberrant course of the DV returning to the CS was clearly demonstrable by traditional 2D echocardiography using HDFI and the R-flow technique. We deem it helpful to trace the inflow of the dilated CS to make the differential diagnosis. The 3D modality might also provide additional spatial information on the associated vessels and thereby assist in prenatal diagnosis.

Prenatal diagnosis of coarctation of the aorta with a long and angled isthmus by two- and three-dimensional echocardiography: a case report.

BACKGROUND: Prenatal diagnosis of coarctation of the aorta (CoA) is challenging for most examiners. The malformation often occurs at the aortic isthmus, which is a short segment between the origin of the left subclavian artery and the insertion of the ductus. We report herein a rare case of CoA with a long, angled, and hypoplastic isthmus. The echocardiographic characteristics and postmortem findings are presented to approach the skill of fetal diagnosis. CASE PRESENTATION: A pregnant women undergone fetal echocardiography at 26 + 3 gestational weeks in our center. Conventional two-dimensional echocardiography (2DE) showed that ascending aorta went straight upward branching three brachiocephalic arteries without the appearance of the arch, suggesting the possibility of an interrupted aortic arch. Three-dimensional echocardiography (3DE) using spatiotemporal image correlation (STIC) and high-definition flow imaging technique was performed to obtain the 3D rendered images, which clearly showed the arch and its angled junction with the slim isthmus in space. Intra-uterine fetal death occurred and an autopsy was performed. The gross findings showed the angled hypoplastic aortic isthmus in detail and thus confirmed the prenatal diagnosis. CONCLUSIONS: Traditional 2DE may be limited in showing the angled hypoplastic aortic isthmus, while the 3DE STIC technique can provide additional spatial information to show great arteries in detail, help to find tiny vessels, and thus benefit the examiners to make an accurate diagnosis.

Role of Plasmodium falciparum Kelch 13 Protein Mutations in P. falciparum Populations from Northeastern Myanmar in Mediating Artemisinin Resistance.

Mutations in the Plasmodium falciparum Kelch 13 (PfK13) protein are associated with artemisinin resistance. PfK13 is essential for asexual erythrocytic development, but its function is not known. We tagged the PfK13 protein with green fluorescent protein in P. falciparum to study its expression and localization in asexual and sexual stages. We used a new antibody against PfK13 to show that the PfK13 protein is expressed ubiquitously in both asexual erythrocytic stages and gametocytes and is localized in punctate structures, partially overlapping an endoplasmic reticulum marker. We introduced into the 3D7 strain four PfK13 mutations (F446I, N458Y, C469Y, and F495L) identified in parasites from the China-Myanmar border area and characterized the in vitro artemisinin response phenotypes of the mutants. We found that all the parasites with the introduced PfK13 mutations showed higher survival rates in the ring-stage survival assay (RSA) than the wild-type (WT) control, but only parasites with N458Y displayed a significantly higher RSA value (26.3%) than the WT control. After these PfK13 mutations were reverted back to the WT in field parasite isolates, all revertant parasites except those with the C469Y mutation showed significantly lower RSA values than their respective parental isolates. Although the 3D7 parasites with introduced F446I, the predominant PfK13 mutation in northern Myanmar, did not show significantly higher RSA values than the WT, they had prolonged ring-stage development and showed very little fitness cost in in vitro culture competition assays. In comparison, parasites with the N458Y mutations also had a prolonged ring stage and showed upregulated resistance pathways in response to artemisinin, but this mutation produced a significant fitness cost, potentially leading to their lower prevalence in the Greater Mekong subregion.IMPORTANCE Artemisinin resistance has emerged in Southeast Asia, endangering the substantial progress in malaria elimination worldwide. It is associated with mutations in the PfK13 protein, but how PfK13 mediates artemisinin resistance is not completely understood. Here we used a new antibody against PfK13 to show that the PfK13 protein is expressed in all stages of the asexual intraerythrocytic cycle as well as in gametocytes and is partially localized in the endoplasmic reticulum. By introducing four PfK13 mutations into the 3D7 strain and reverting these mutations in field parasite isolates, we determined the impacts of these mutations identified in the parasite populations from northern Myanmar on the ring stage using the in vitro ring survival assay. The introduction of the N458Y mutation into the 3D7 background significantly increased the survival rates of the ring-stage parasites but at the cost of the reduced fitness of the parasites. Introduction of the F446I mutation, the most prevalent PfK13 mutation in northern Myanmar, did not result in a significant increase in ring-stage survival after exposure to dihydroartemisinin (DHA), but these parasites showed extended ring-stage development. Further, parasites with the F446I mutation showed only a marginal loss of fitness, partially explaining its high frequency in northern Myanmar. Conversely, reverting all these mutations, except for the C469Y mutation, back to their respective wild types reduced the ring-stage survival of these isolates in response to in vitro DHA treatment.

Evolution of the Plasmodium vivax multidrug resistance 1 gene in the Greater Mekong Subregion during malaria elimination.

BACKGROUND: The malaria elimination plan of the Greater Mekong Subregion (GMS) is jeopardized by the increasing number of Plasmodium vivax infections and emergence of parasite strains with reduced susceptibility to the frontline drug treatment chloroquine/primaquine. This study aimed to determine the evolution of the P. vivax multidrug resistance 1 (Pvmdr1) gene in P. vivax parasites isolated from the China-Myanmar border area during the major phase of elimination. METHODS: Clinical isolates were collected from 275 P. vivax patients in 2008, 2012-2013 and 2015 in the China-Myanmar border area and from 55 patients in central China. Comparison was made with parasites from three border regions of Thailand. RESULTS: Overall, genetic diversity of the Pvmdr1 was relatively high in all border regions, and over the seven years in the China-Myanmar border, though slight temporal fluctuation was observed. Single nucleotide polymorphisms previously implicated in reduced chloroquine sensitivity were detected. In particular, M908L approached fixation in the China-Myanmar border area. The Y976F mutation sharply decreased from 18.5% in 2008 to 1.5% in 2012-2013 and disappeared in 2015, whereas F1076L steadily increased from 33.3% in 2008 to 77.8% in 2015. While neutrality tests suggested the action of purifying selection on the pvmdr1 gene, several likelihood-based algorithms detected positive as well as purifying selections operating on specific amino acids including M908L, T958M and F1076L. Fixation and selection of the nonsynonymous mutations are differently distributed across the three border regions and central China. Comparison with the global P. vivax populations clearly indicated clustering of haplotypes according to geographic locations. It is noteworthy that the temperate-zone parasites from central China were completely separated from the parasites from other parts of the GMS. CONCLUSIONS: This study showed that P. vivax populations in the China-Myanmar border has experienced major changes in the Pvmdr1 residues proposed to be associated with chloroquine resistance, suggesting that drug selection may play an important role in the evolution of this gene in the parasite populations.

Prenatal diagnosis of fetal isolated right coronary artery to left ventricle fistula.

Coronary artery fistula (CAF) is a rare malformation and is seldom reported during pregnancy. Right coronary artery fistula commonly drains into the right ventricle, right atrium, or pulmonary artery. We describe here a rare case of fetal CAF draining into the left ventricle using cross-sectional and color Doppler echocardiography. We also summarized our experience in the diagnosis of this uncommon malformation, in which tracing the origin, course, and outlet of the abnormal intra-cardiac flow played a key role.

Colorectal follicular lymphoma: A case report.

RATIONALE: Considering the low incidence of colorectal follicular lymphoma (FL) and its clinical features in endoscopic views, only a few studies have described the pathological diagnosis and treatment of this disease. This study aimed to reveal the overall process of clinical diagnosis and treatment of colorectal FL by conducting a case review. PATIENT CONCERNS: A 27-year-old female presented to our department because of "severe bloody stool" lasting for more than 1 month. Her primary symptom was melena. Colonoscopy revealed widespread flat polyps with various immunophenotypes (CD10+, BCL2+, BCL6+, cyclin D1-, CD5-) in the colorectal area. DIAGNOSIS: In accordance with manifestations on positron emission tomography-computed tomography (PET/CT), the patient was diagnosed with stage IV colorectal FL. INTERVENTIONS: PET/CT reexamination after 2 courses of rituximab, cyclophosphamide, liposomal doxorubicin, vincristine sulfate, and hydroprednisone (R-CHOP) regimen and 3 courses of R-CHOP plus etoposide regimen for chemotherapy indicated a significant reduction in tumor burden. Subsequently, rituximab was administered alone in 2 treatment courses. OUTCOMES: Lesions on PET/CT disappeared after reexamination. No recurrence was observed within the 12-month follow-up period. LESSONS: Colorectal FL is a rare disease with an inert clinical course and is common in the ileocecal area. Endoscopic views show multiple polyps. Interventional treatment is usually provided after observation of clinical symptoms or during disease progression. The disease has a relatively good prognosis.

Strategies for Accurate Diagnosis of Fetal Aortic Arch Anomalies: Benefits of Three-Dimensional Sonography With Spatiotemporal Image Correlation and a Novel Algorithm for Volume Analysis.

BACKGROUND: Obtaining an accurate diagnosis of fetal aortic arch anomalies is often difficult with traditional two-dimensional (2D) sonography. Thus, the aim of this study was to assess the value of three-dimensional (3D) sonography with spatiotemporal image correlation for diagnosing fetal aortic arch anomalies using a novel algorithm for 3D volume analysis. METHODS: Two-dimensional and 3D echocardiographic image data from 300 normal fetuses and 30 fetuses with aortic arch anomalies were retrospectively reviewed. Grayscale and high-definition flow imaging data were available for 2D echocardiography. Three-dimensional volumes were acquired in parasagittal views with high-definition flow imaging information. Volume postprocessing was performed using a novel algorithm to obtain 3D tomographic ultrasound imaging slices and color-rendered images. Detection of aortic arch positions, aberrant brachiocephalic patterns, and aortic arch anomalies was compared for 2D and 3D modalities. Postnatal echocardiography was used as the truth standard in assessing aortic anatomy. RESULTS: In total, four cases of double aortic arch, 21 cases of right aortic arch, one case of retroesophageal aortic arch, and four cases of left aortic arch with aberrant right subclavian arteries were included. Inter- and intraobserver agreement were excellent for 2D and 3D modalities. The 3D modality offered better sensitivity and accuracy compared with 2D imaging for the detection of brachiocephalic anomalies (P < .01) and arch anomalies (P < .01) but comparable sensitivity for arch position. There was no difference in specificity for both modalities. CONCLUSIONS: The proposed novel algorithm for volume postprocessing ensures that 3D reconstructed images are obtained with high repeatability. The addition of 3D spatiotemporal image correlation to fetal echocardiography may offer a more accurate diagnosis for arch anomalies.

A meta-analysis of adoptive immunotherapy in postoperative hepatocellular carcinoma.

BACKGROUND: Adoptive immunotherapy (AIT) has been adopted as an adjuvant treatment for hepatocellular carcinoma (HCC) patients after curative therapy. However, the outcomes of AIT remain controversial. PURPOSE: The purpose of this study is to analyze the safety and efficacy of AIT with the recurrence rate and mortality. MATERIALS AND METHODS: We identified eight randomized controlled trials (RCTs) that adopted AIT to HCC after curative treatments. A meta-analysis was carried out to assess the recurrence rate and mortality. RESULTS: Eight RCTs with 964 patients were included in the study. The overall analysis showed that AIT treatment can not only decrease the 1-year (risk ratio [RR] =0.59, 95% confidence interval [95% CI] = 0.48-0.72, P < 0.00001), 2-year (RR = 0.69, 95% CI = 0.60-0.79, P < 0.00001), and 3-year (RR = 0.82, 95% CI = 0.74-091, P = 0.0001) recurrence, but also decrease the 1-year (RR = 0.43, 95% CI = 0.30-0.62, P = 0.00001), 2-year (RR = 0.56, 95% CI = 0.46-0.74, P < 0.00001), and 3-year (RR = 0.85, 95% CI = 0.73-0.99, P = 0.03) mortality. The results also indicate that the group of lymphokine-activated killer (LAK) cells showed lower pooled RR values compared to the group of cytokine-induced killer cells among every subgroups. However, the AIT treatment failed to affect the 5-year recurrence rate and mortality (P > 0.05). CONCLUSIONS: This review provides available evidences that AIT, especially the treatment of LAK, can be used to decrease the early recurrence and mortality of postoperative HCC but may not the long term.

Natural deep eutectic solvents as eco-friendly and sustainable dilution medium for the determination of residual organic solvents in pharmaceuticals with static headspace-gas chromatography.

Reported here is a simple and rapid static headspace gas chromatography (SHS-GC) method for the determination of trace solvents including ethanol, isopropanol, n-butanol, 1,4-dioxane, tetrahydrofuran, acetonitrile, methanol and acetone which commonly used in drug production process. Natural deep eutectic solvents (NADESs) are firstly used as the matrix medium for this method, which provided high sensitivity for residual solvents detection. With the optimized method, validation experiments were performed and the data showed excellent linearity for all the solvents (R2 ≥ 0.999, n = 7). The limits of detection (LOD) for ethanol, isopropanol, n-butanol, 1,4-dioxane, tetrahydrofuran, acetonitrile, methanol and acetone are 0.09, 0.08, 0.07, 0.11, 0.06, 0.10, 0.12 and 0.08 µg g-1, respectively. Accuracy was checked by a recovery experiment at three different levels, and the recoveries of the tested solvents were ranged from 94.3% to 105.4%. The relative standard deviation (RSD) of each solvent for intra- and inter-day precision is in the range of 0.85 to 3.65 and 1.51 to 4.53, respectively. The developed approach can be readily used for determination of the residual solvents in six active pharmaceutical ingredients including pramipexole dihydrochloride, rivaroxaban, lisinopril, ramipril, imatinib mesylate and sitagliptin.

Characterization of Plasmodium berghei Pbg37 as Both a Pre- and Postfertilization Antigen with Transmission-Blocking Potential.

Transmission-blocking vaccines (TBVs) interrupting malaria transmission are an integrated tool for malaria eradication. We characterized a sexual-stage-specific gene (PBANKA_060330) from Plasmodium berghei and studied its potential for use as a TBV. This gene, referred to as pbg37, encodes a protein of 37 kDa with a signal peptide and multiple transmembrane domains and is preferentially expressed in gametocytes. A recombinant Pbg37 (rPbg37) protein targeting the N-terminal 63 amino acids (amino acids 26 to 88) expressed in bacteria elicited strong antibody responses in mice. Western blotting demonstrated Pbg37 expression in gametocytes, zygotes, and, to a lesser extent, ookinetes and its predominant association with the membranes of gametocytes. Indirect immunofluorescence assay showed an abundant surface localization of Pbg37 on gametes and zygotes but reduced amounts on retorts and ookinetes. Knockout of pbg37 (Δpbg37) led to a considerable reduction in gametocytemia, which translated into a ~92.1% decrease in the oocyst number in mosquitoes. Deletion of pbg37 had a more substantial influence on the development and maturation of microgametocytes. As a result, the Δpbg37 lines exhibited a higher female/male gametocyte ratio, fewer mature male gametocytes, and defects in the exflagellation of mature microgametocytes. To test the transmission-blocking potential of Pbg37, an in vitro ookinete assay showed that the major inhibitory effects of anti-Pbg37 antiserum were on the exflagellation and fertilization processes. Direct feeding of mosquitoes on mice immunized with rPbg37 or a control protein showed that rPbg37-immunized and P. berghei-infected mice had a significant reduction (49.1%) in oocyst density compared to the controls. The conservation of this gene in Plasmodium warrants further investigations in human malaria parasites.

Right heart dilatation in a fetus with an abnormal foramen ovale valve: an indicator of interatrial communication restriction.

AIMS: Foramen ovale (FO) valve with a shape or motion abnormality is frequently detected during routine obstetric ultrasonic examinations. However, the hemodynamics mechanism of this entity remains unclear. The purpose of the study is to determine the relevance of interatrial communication restriction and resultant morphological modifications. MATERIALS AND METHODS: We reviewed the echocardiographic records of fetuses with isolated abnormal FO valve evaluated between January of 2010 and december of 2016. The size (DFO) of the FO orifice, opening angle (α) of the FO valve, and dimensions of cardiac chambers, FO channel outlet (DOUT) and inferior vena cava (DIVC) were measured. We evaluated their (DFO, DOUT, α) relationships to the diameters of RA and DIVC. Five hundred and seventy normal fetuses were selected to establish the normal range of the DOUT/DIVC ratio so as to provide a criterion for restriction. RESULTS: An abnormal FO valve was identified in 89 fetuses without congenital heart disease, with restriction noted in 62 fetuses (45 fetuses with RA dilatation, 12 fetuses with RA and RV dilatation, and 5 fetuses with no RA dilatation). There were no significant correlations between RA/LA and DFO/DIVC, RA/ LA and α. RA/LA was negatively correlated with DOUT/DIVC (R2=0.97, p<0.01). CONCLUSIONS: For a fetus with an abnormal FO valve, right heart dilatation could be considered as an indicator of interatrial communication restriction, which could be assessed by evaluating the FO channel outlet. The degree of right atrium dilatation indicates the severity of the restriction.

Plasmodium falciparum Falcipain-2a Polymorphisms in Southeast Asia and Their Association With Artemisinin Resistance.

Background: Falcipain-2a ([FP2a] PF3D7_1115700) is a Plasmodium falciparum cysteine protease and hemoglobinase. Functional FP2a is required for potent activity of artemisinin, and in vitro selection for artemisinin resistance selected for an FP2a nonsense mutation. Methods: To investigate associations between FP2a polymorphisms and artemisinin resistance and to characterize the diversity of the enzyme in parasites from the China-Myanmar border, we sequenced the full-length FP2a gene in 140 P falciparum isolates collected during 2004-2011. Results: The isolates were grouped into 8 different haplotype groups. Haplotype group I appeared in samples obtained after 2008, coinciding with implementation of artemisinin-based combination therapy in this region. In functional studies, compared with wild-type parasites, the FP2a haplotypes demonstrated increased ring survival, and all haplotype groups exhibited significantly reduced FP2a activity, with group I showing the slowest protease kinetics and reduced parasite fitness. Conclusions: These results suggest that altered hemoglobin digestion due to FP2a mutations may contribute to artemisinin resistance.

Prenatal screening of fetal ventriculoarterial connections: benefits of 4D technique in fetal heart imaging.

BACKGROUND: Identification of prenatal ventriculoarterial connections in fetuses with conotruncal anomalies (CTA) remains one of the greatest challenges for sonographers performing screening examinations. Herein, we propose a novel protocol of 4D volume analysis that identifies ventriculoarterial connections and evaluate its clinical utility in routine screenings. METHODS: Twenty-nine cases of transposition of the great arteries (TGA), 22 cases of double-outlet right ventricle (DORV), 36 cases of tetralogy of Fallot (TOF), 14 cases of truncus arteriosus (TCA), and randomly selected 70 normal fetuses were reviewed in this study. All cases were evaluated using 2D data alone (2D method), post-processing volumes with no exact algorithm (4D-1 method), or with the proposed algorithm (4D-2 method), or using the 2D and 4D data together (combined method). Comparisons were made to evaluate the detection rate of ventriculoarterial connections for these different methods. RESULTS: During 18-28 gestational weeks, the detection rate of 4D-2 modality was satisfactory. The detection rate of the combined method was significantly higher than 2D method in the identification of TGA, TOF, and TCA. The detection rate of 4D-1 method was significantly lower than 4D -2 modality for CTA fetuses. During late pregnancy, the detection rate for both 4D modalities was very low due to the poor quality of the 4D volumes. CONCLUSIONS: We proposed a detailed protocol, which allowed the examiner to identify fetal ventriculoarterial connections by 4D volumes. Inclusion of blood information into the volumes improved diagnosis. Our findings suggest that the incorporation of 4D STIC into routine screenings could improve the detection for TGA, TOF, and TCA.

Role of four-dimensional echocardiography with high-definition flow imaging and spatiotemporal image correlation in detecting fetal pulmonary veins.

BACKGROUND: Prenatal diagnosis of fetal total anomalous pulmonary vein connection (TAPVC) remains challenging for most screening sonographers. The purpose of this study was to evaluate the use of four-dimensional echocardiography with high-definition flow imaging and spatiotemporal image correlation (4D-HDFI) in identifying pulmonary veins in normal and TAPVC fetuses. MATERIAL & METHODS: We retrospectively reviewed and performed 4D-HDFI in 204 normal and 12 fetuses with confirmed diagnosis of TAPVC. Cardiac volumes were available for postanalysis to obtain 4D-rendered images of the pulmonary veins. For the normal fetuses, two other traditional modalities including color Doppler and HDFI were used to detect the number of pulmonary veins and comparisons were made between each of these traditional methods and 4D-HDFI. RESULTS: For conventional echocardiography, HDFI modality was superior to color Doppler in detecting more pulmonary veins in normal fetuses throughout the gestational period. 4D-HDFI was the best method during the second trimester of pregnancy in identifying normal fetal pulmonary veins. 4D-HDFI images vividly depicted the figure, course, and drainage of pulmonary veins in both normal and TAPVC fetuses. CONCLUSION: HDFI and the advanced 4D-HDFI technique could facilitate identification of the anatomical features of pulmonary veins in both normal and TAPVC fetuses; 4D-HDFI therefore provides additional and more precise information than conventional echocardiography techniques.

Significant Divergence in Sensitivity to Antimalarial Drugs between Neighboring Plasmodium falciparum Populations along the Eastern Border of Myanmar.

Malaria parasites in different areas where malaria is endemic display different levels of resistance to antimalarial drugs as the result of varied drug use histories. To provide updated knowledge of drug sensitivities during the malaria elimination phase in Southeast Asia, an epicenter of multidrug resistance, we determined in vitro susceptibilities of culture-adapted Plasmodium falciparum isolates from two eastern border regions (Wa and Kachin) of Myanmar to 10 drugs. Despite their close proximity, the Kachin parasites displayed higher 50% inhibitory concentrations than the Wa parasites to chloroquine, piperaquine, naphthoquine, mefloquine, quinine, pyrimethamine, pyronaridine, lumefantrine, and dihydroartemisinin. Genotyping of genes associated with drug resistance also showed significant differences in the prevalence rates of mutant alleles between the two regions. Particularly, major pfdhfr mutations mediating pyrimethamine resistance and the pfdhps A437G mutation had significantly higher frequencies in the Kachin parasites (P < 0.005). Moreover, when pfdhfr and pfdhps were considered together, the wild-type allele was found only in the Wa samples (22.6%). In addition, the pfmdr1 Y184F mutation reached 38.7% in the Kachin parasites, compared to 9.7% in the Wa parasites, whereas N86Y was only detected in the Wa parasites, at 22.6%. Furthermore, the F446I mutation and all mutations in the propeller domain of the PfK13 gene were significantly more frequent in the Kachin parasites. Collectively, this work demonstrates that even in spatially closely separated regions, parasites can exhibit drastic differences in drug sensitivities and genetic makeups underlying drug resistance, which may reflect regionally different drug histories and genetic drift of these isolated parasite populations.