An apicoplast-resident folate transporter is essential for sporogony of malaria parasites

Malaria parasites are fast replicating unicellular organisms and require substantial amounts of folate for DNA synthesis. Despite the central role of this critical co-factor for parasite survival, only little is known about intraparasitic folate trafficking in Plasmodium. Here, we report on the expression, subcellular localisation and function of the parasite's folate transporter 2 (FT2) during life cycle progression in the murine malaria parasite Plasmodium berghei. Using live fluorescence microscopy of genetically engineered parasites, we demonstrate that FT2 localises to the apicoplast. In invasive P. berghei stages, a fraction of FT2 is also observed at the apical end. Upon genetic disruption of FT2, blood and liver infection, gametocyte production and mosquito colonisation remain unaltered. But in the Anopheles vector, FT2 deficient parasites develop inflated oocysts with unusual pulp formation consisting of numerous single-membrane vesicles, which ultimately fuse to form large cavities. Ultrastructural analysis suggests that this defect reflects aberrant sporoblast formation caused by abnormal vesicular traffic. Complete sporogony in FT2-deficient oocysts is very rare, and mutant sporozoites fail to establish hepatocyte infection, resulting in a complete block of parasite transmission. Our findings reveal a previously unrecognised organellar folate transporter that exerts critical roles for pathogen maturation in the arthropod vector.Deutsche Forschungsgemeinschaft, Grant/ Award Numbers: 419345764, IRTG2290; Francis Crick Institute, Grant/Award Number: FC001043; Wellcome Trust, Grant/Award Number: 210918/Z/18/

Mansonella perstans, Onchocerca volvulus and Strongyloides stercoralis infections in rural populations in central and southern Togo

Background: Mansonella perstans, Onchocerca volvulus and Strongyloides stercoralis are widespread helminth parasites in the tropics. Their distribution remains difficult to determine as it may change during national disease control programs and with regional mass drug administration (MDA). Epidemiological surveys are of importance to evaluate the geographical distribution of these helminth parasites and the diseases they may cause, however, up to date epidemiological evaluations on M. perstans and S. stercoralis in Togo are rare, and surveys on O. volvulus are important especially under the aspect of MDA of ivermectin which is performed since decades. Methods: Dry blood samples (n = 924) were collected from rural populations in the Régions Central and Plateaux in Togo, and analyzed by parasite-specific real-time PCR and ELISA techniques. Results: Dry blood samples from 733 persons where investigated by real-time PCR tested for DNA of blood-circulating M. perstans microfilaria, and a prevalence of 14.9% was detected. Distinct differences were observed between genders, positivity was higher in men increasing with age, and prevalence was highest in the Région Plateaux in Togo. IgG4 responses to O. volvulus antigen (OvAg) were studied in 924 persons and 59% were found positive. The distribution of parasite infestation between age and gender groups was higher in men increasing with age, and regional differences were detected being highest in the Région Plateaux. The diagnostic approach disclosed 64,5% positive IgG4 responses to S. stercoralis infective third-stage larvae-specific antigen (SsL3Ag) in the surveyed regions. Antigen cross reactivity of SsL3Ag with parasite co-infections may limit the calculated prevalence. Singly IgG4 positive for SsL3Ag were 13.9%, doubly positive for OvAg and SsL3Ag were 35.5% and triply positive for M. perstans, O. volvulus and S. stercoralis were 9.9%. Conclusions: Mansonelliasis, onchocerciasis and strongyloidiasis remain prevalent in the surveyed regions, yet with local differences. Our observations suggest that transmission of M. perstans, O. volvulus and S. stercoralis may be ongoing. The degree of positive test results in the examined rural communities advocate for the continuation of MDA with ivermectin and albendazole, and further investigations should address the intensity of transmission of these parasites. Keywords: Onchocerca volvulus, Mansonella perstans, Strongyloides stercoralis, Onchocerciasis, Mansonelliasis, Strongyloidiasis, Togo, Prevalenc

<i>Onchocerca volvulus</i> infection and serological prevalence, ocular onchocerciasis and parasite transmission in northern and central Togo after decades of <i>Simulium damnosum</i> s.l. vector control and mass drug administration of ivermectin

<div><p>Background</p><p>Mass drug administration (MDA) of ivermectin has become the main intervention to control onchocerciasis or “river blindness”. In Togo, after many years of MDA, <i>Onchocerca volvulus</i> infection has declined dramatically, and elimination appears achievable, but in certain river basins the current situation remains unknown. We have conducted parasitological, serological, ophthalmological, and entomological assessments in northern and central Togo within the river basins of Ôti, Kéran and Mô.</p><p>Methodology/Principal findings</p><p>Examinations were completed in 1,455 participants from 11 onchocerciasis sentinel villages, and <i>O</i>. <i>volvulus</i> transmission by <i>Simulium damnosum</i> sensu lato (s.l.) was evaluated. In children (aged 1–10 years), the prevalence of microfilariae (Mf) was 2.3% and in adults it ranged from 5.1 to 13.3%. Positive IgG4 responses to <i>O</i>. <i>volvulus</i> adult (crude) worm antigen (OvAg) and the recombinant Ov16 antigen were in all-ages 48.7% and 34.4%, and 29.1% and 14.9% in children, respectively. In the river basin villages of Kéran, Mô and Ôti, the IgG4 seroprevalences to OvAg in children were 51.7%, 23.5% and 12.7%, respectively, and to the Ov16 antigen 33.3% (Kéran) and 5.2% (Ôti). Onchocerciasis ocular lesions (punctate keratitis, evolving iridocyclitis and chorioretinitis) were observed in children and young adults. <i>O</i>. <i>volvulus</i>-specific DNA (Ov150) was detected by poolscreen in vector samples collected from Tchitchira/Kéran(22.8%), Bouzalo/Mô(11.3%), Baghan/Mô(2.9%) and Pancerys/Ôti(4.9%); prevalences of <i>O</i>. <i>volvulus</i> infection in <i>S</i>. <i>damnosum</i> s.l. were, respectively, 1%, 0.5%, 0.1% and 0.2%.</p><p>Conclusions/Significance</p><p>In the northern and central river basins in Togo, interruption of <i>O</i>. <i>volvulus</i> transmission has not yet been attained. Patent <i>O</i>. <i>volvulus</i> infections, positive antibody responses, progressive ocular onchocerciasis were diagnosed, and parasite transmission by <i>S</i>. <i>damnosum</i> s.l. occurred close to the survey locations. Future interventions may require approaches selectively targeted to non-complying endemic populations, to the seasonality of parasite transmission and national onchocerciasis control programs should harmonize cross-border MDA as a coordinated intervention.</p></div

Mutations in the Amiloride-Sensitive Epithelial Sodium Channel in Patients With Cystic Fibrosis-Like Disease

We investigated whether mutations in the genes that code for the different subunits of the amiloride-sensitive epithelial sodium channel (ENaC) might result in cystic fibrosis (CF)-like disease. In a small fraction of the patients, the disease could be potentially explained by an ENaC mutation by a Mendelian mechanism, such as p.V114I and p.F61L in SCNN1A. More importantly, a more than three-fold significant increase in incidence of several rare ENaC polymorphisms was found in the patient group (30% vs. 9% in controls), indicating an involvement of ENaC in some patients by a polygenetic mechanism. Specifically, a significantly higher number of patients carried c.-55+5G > C or p.W493R in SCNN1A in the heterozygous state, with odds ratios (ORs) of 13.5 and 2.7, respectively. The p.W493R-SCNN1A polymorphism was even found to result in a four-fold more active ENaC channel when heterologously expressed in Xenopus laevis oocytes. About I in 975 individuals in the general population will be heterozygous for the hyperactive p.W493R-SCNN1A mutation and a cystic fibrosis transmembrane conductance regulator (CFTR) gene that results in very low amounts (0-10%) functional CFTR. These ENaC/CFTR genotypes may play a hitherto unrecognized role in lung diseases. Hum Mutat 30:1093-1103, 2009. (C) 2009 Wiley-Liss, Inc

Prevalence of microfilariae (Mf) of <i>Onchocerca volvulus</i> in sentinel villages (n = 363) of the National Onchocerciasis Control Programme (NOCP) in Togo.

<p>Regular epidemiological surveys (n = 957) were conducted by the NOCP over a 37-year period (1976–2014) in onchocerciasis endemic villages, and from each participant (n = 193,742) a skin biopsy was taken from the left and right iliac crest and the emerging Mf were counted after snip incubation. The graph shows the microfilarial prevalence (median, minimum, maximum, in %) as detected during the annual surveys. Anti-vectorial interventions were applied since 1976, whereas the central regions were incorporated into the Programme in 1987. Since 1988, vector control measures were supplemented by MDA with ivermectin. Initially MDA was applied mainly by mobile teams; during some years of the early 1990’s, aerial larvicide application was suspended in several river basins. In the northern territories (SIZ) vector control and intensified ivermectin distribution was continued after OCP’s closure in 2002. Special interventions in the post-OCP period included continued aerial larvicide application for five additional years (2003–2007) and biannual ivermectin MDA until the end of 2012.</p

Collection of <i>Simulium damnosum</i> s.l. at the catch point at the Mô river (village of Bouzalo) caught per person (one day per week).

<p>The annual biting rate (ABR) from January 2016 until December 2016 was calculated by multiplying the number of blackflies caught daily by the number of days per week for each month to add up to 12 months. Collections were conducted from 7am to 6pm with alternating fly catchers every two hours as described in Material and Methods.</p

Ocular pathologies in study participants (n = 1,183) from onchocerciasis endemic villages situated in the river basins of Kéran, Mô and Ôti in northern and central Togo.

<p>The village populations have been treated annually via CDTI. The ocular pathologies, their grades of evolution and extent were classified as described previously [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0006312#pntd.0006312.ref012" target="_blank">12</a>] and are indicated for the right and left eye (RLE), the left eye (LE) and the right eye (RE). Chi-Square tests were applied to compare ocular pathologies of the right eye (RE) between female and male patients. One-sided Fisher exact test was used to evaluate differences in the prevalence of ocular pathologies in patients from the Ôti, Kéran and Mô river basins, and significant differences are indicated by p<0.05. Spearman’s rank correlation analyses of ocular pathologies of the right eye (RE) and age were conducted; the correlation coefficient ρ and the significant associations (p<0.05) are shown.</p

Ov150 rt-PCR (Poolscreen) testing of <i>Simulium damnosum</i> s.l. black flies collected at sentinel catch points in northern and central Togo in 2015.

<p>Each pool consists of 25 blackflies and is listed with the total number of tested flies by catch point, and the number of Ov150-positive pools. The prevalence (in % with 95% confidence intervals) of <i>O</i>. <i>volvulus</i> in <i>S</i>. <i>damnosum</i> s.l. is calculated according to Katholi et al. 1995 [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0006312#pntd.0006312.ref016" target="_blank">16</a>]. Fisher’s exact test was applied to evaluate differences in infection rates between pools.</p