Serum Granulysin as a Possible Key Marker of Vitiligo Activity and Severity.

Background: Vitiligo is an immune-mediated, chronic skin condition that affects both the innate and adaptive immune systems. Antimicrobial peptide overexpression is one of its defining characteristics. Granulysin (GNLY), an antimicrobial peptide, may play a role in the pathogenesis of various autoimmune diseases. Objectives: To estimate the serum GNLY levels in vitiligo patients and to correlate those levels with the severity and activity of the disease. Materials and Methods: This case-control study included 60 non-segmental vitiligo patients (Group A) and a control group of 60 people who were matched for age and sex, appeared to be in good health, and were not suffering from vitiligo (Group B). The serum granulysin levels of all subjects were measured using an enzyme-linked immunosorbent assay. Results: When compared to the control group, vitiligo patients had significantly higher serum GNLY levels (P = 0.001). When compared to patients with stable disease, those with active vitiligo had significantly higher serum GNLY levels (P = 0.008). Additionally, there was a positive correlation between the serum GNLY levels and the vitiligo area severity index and vitiligo disease activity scores (P = 0.004 and <0.001, respectively). Limitations: Study population was relatively small. Evaluation of serum granulysin before and after treatment could have been more beneficial. Conclusions: Blood granulysin levels could contribute to the pathogenesis of vitiligo. A higher serum granulysin level may also be a trustworthy predictor of the severity and progression of a disease.

Human immune response against salivary antigens of Simulium damnosum s.l.: A new epidemiological marker for exposure to blackfly bites in onchocerciasis endemic areas.

BACKGROUND: Simulium damnosum sensu lato (s.l.) blackflies transmit Onchocerca volvulus, a filarial nematode that causes human onchocerciasis. Human landing catches (HLCs) is currently the sole method used to estimate blackfly biting rates but is labour-intensive and questionable on ethical grounds. A potential alternative is to measure host antibodies to vector saliva deposited during bloodfeeding. In this study, immunoassays to quantify human antibody responses to S. damnosum s.l. saliva were developed, and the salivary proteome of S. damnosum s.l. was investigated. METHODOLOGY/PRINCIPAL FINDINGS: Blood samples from people living in onchocerciasis-endemic areas in Ghana were collected during the wet season; samples from people living in Accra, a blackfly-free area, were considered negative controls and compared to samples from blackfly-free locations in Sudan. Blackflies were collected by HLCs and dissected to extract their salivary glands. An ELISA measuring anti-S. damnosum s.l. salivary IgG and IgM was optimized and used to quantify the humoral immune response of 958 individuals. Both immunoassays differentiated negative controls from endemic participants. Salivary proteins were separated by gel-electrophoresis, and antigenic proteins visualized by immunoblot. Liquid chromatography mass spectrometry (LC-MS/MS) was performed to characterize the proteome of S. damnosum s.l. salivary glands. Several antigenic proteins were recognized, with the major ones located around 15 and 40 kDa. LC-MS/MS identified the presence of antigen 5-related protein, apyrase/nucleotidase, and hyaluronidase. CONCLUSIONS/SIGNIFICANCE: This study validated for the first time human immunoassays that quantify humoral immune responses as potential markers of exposure to blackfly bites. These assays have the potential to facilitate understanding patterns of exposure as well as evaluating the impact of vector control on biting rates. Future studies need to investigate seasonal fluctuations of these antibody responses, potential cross-reactions with other bloodsucking arthropods, and thoroughly identify the most immunogenic proteins.

A snapshot of Plasmodium falciparum malaria drug resistance markers in Sudan: a pilot study.

OBJECTIVES: Malaria infection is still known to be a worldwide public health problem, especially in tropical and sub-tropical African countries like Sudan. A pilot study conducted to describe the trend of P. falciparum drug resistance markers in 2017-2018 in comparison to CQ and AS/SP eras in Sudan. The Pfcrt, Pfmdr-1, Pfdhfr, and Pfdhps genes were investigated. Data deposited by the worldwide antimalarial resistance network was consulted, and the molecular markers previously reported from Sudan were analyzed. RESULTS: Drug molecular markers analysis was successfully done on 20 P. falciparum isolates. The Pfcrt K76 showed high frequency; 16 (80%). For the Pfmdr-1, 9 (45%) isolates were carrying the N86 allele, and 11 (55%) were 86Y allele. While the Y184F of the Pfmdr-1 showed a higher frequency of 184F compared to Y184; 16 (80%) and 4 (20%), respectively. In the Pfdhfr, 51I allele showed higher frequency compared to N51; 18 (90%) and 2 (10%), respectively. For S108N, 18 (90%) were 108 N and 2 (10%) were S108. In the Pfdhps, all isolates were carrying the mutant alleles; 437G and 540E. The frequency distribution of the Pfcrt, Pfmdr-1, Pfdhfr, Pfdhps was significantly different across the whole years in Sudan.

Frequency distribution of IL-17A G197A (rs2275913) and IL-17F A7488G (rs763780) polymorphisms among healthy Sudanese population.

OBJECTIVES: IL-17A G197A and IL-17F A7488G polymorphisms has been identified to be associated with the susceptibility to many diseases. This study aimed to investigate the frequency distribution of IL-17A G197A and IL-17F A7488G polymorphisms among healthy Sudanese population. A descriptive cross-sectional hospital-based molecular study conducted in different sites throughout Sudan. Two ml blood samples were collected from 717 healthy participants. Demographic data and the medical history of the participants were collected. RESULTS: Of the 717 participants, 355 (49.5%) were males and 362 (50.5%) were females, their mean age was 30.2 ± 17.2 and 32.2 ± 16.5, respectively. For IL-17A, the most frequent genotype detected among males and females was IL-17A heterozygote allele (AG); 215 (60.6%) and 194 (53.6%), respectively. Whereas, for IL-17F, the most frequent allele among males and females was the homozygote allele (AA); 298 (83.9%) for males and 322 (89.0%) for females. HWE for genotype distributions of IL-17A was showing statistical insignificance for IL-17A among males and females, P value 0.614. While HWE for IL-17F reached the equilibrium level, P value 0.048. The most frequent age group was those aged between 21 to 40 years; 281 (39.2%). Arab constituted the major ethnicity of the study participants; 418 (58.3%), P value 0.034.

Human antibody reaction against recombinant salivary proteins of Phlebotomus orientalis in Eastern Africa.

BACKGROUND: Phlebotomus orientalis is a vector of Leishmania donovani, the causative agent of life threatening visceral leishmaniasis spread in Eastern Africa. During blood-feeding, sand fly females salivate into the skin of the host. Sand fly saliva contains a large variety of proteins, some of which elicit specific antibody responses in the bitten hosts. To evaluate the exposure to sand fly bites in human populations from disease endemic areas, we tested the antibody reactions of volunteers' sera against recombinant P. orientalis salivary antigens. METHODOLOGY/PRINCIPAL FINDINGS: Recombinant proteins derived from sequence data on P. orientalis secreted salivary proteins, were produced using either bacterial (five proteins) or mammalian (four proteins) expression systems and tested as antigens applicable for detection of anti-P. orientalis IgG in human sera. Using these recombinant proteins, human sera from Sudan and Ethiopia, countries endemic for visceral leishmaniasis, were screened by ELISA and immunoblotting to identify the potential markers of exposure to P. orientalis bites. Two recombinant proteins; mAG5 and mYEL1, were identified as the most promising antigens showing high correlation coefficients as well as good specificity in comparison to the whole sand fly salivary gland homogenate. Combination of both proteins led to a further increase of correlation coefficients as well as both positive and negative predictive values of P. orientalis exposure. CONCLUSIONS/SIGNIFICANCE: This is the first report of screening human sera for anti-P. orientalis antibodies using recombinant salivary proteins. The recombinant salivary proteins mYEL1 and mAG5 proved to be valid antigens for screening human sera from both Sudan and Ethiopia for exposure to P. orientalis bites. The utilization of equal amounts of these two proteins significantly increased the capability to detect anti-P. orientalis antibody responses.

Acute Kidney Injury in Sub-Sahara Africa: A Single-Center Experience from Khartoum, Sudan.

BACKGROUND: The burden of acute kidney injury (AKI) is high in Africa. While there are no reliable statistics about AKI in Africa, the Global Snapshot Study of the 0by25 initiative of the International Society of Nephrology has determined dehydration, infections, animal envenomation, and complications during pregnancy as the main causes. METHODS: This study was conducted at the Soba University Hospital (SUH), Khartoum, Sudan, a tertiary referral center. We included all hemodialysis patients treated for AKI at SUH between -January 1, 2013 and December 31, 2014 in the study. We reviewed patients' hospital records and characterized pathogenesis, treatment, and patient outcomes. In addition, we investigated survival by Kaplan-Meier and Cox regression analysis. RESULTS: Out of 520 patients who received emergency HD, 71 patients (14%) had AKI (age 40.6 ± 17.3 years, 56.5% were males). Glomerular and tubular-interstitial diseases were the leading cause of AKI, followed by envenomation and intoxication by hair dye. Patients received a median of 5 dialysis sessions for a median of 8 days. In 32 patients (45%) renal function recovered, 10 patients (14%) died, and 29 patients (41%) remained dialysis-dependent. Mortality was significantly higher in females compared to men (hazard ratio 4.1 [95% CI 1.02-16.67]). Outcomes were worse in patients with pre-renal AKI and intoxications. CONCLUSION: Our results indicate a higher mortality in females and in patients with pre-renal AKI and intoxications. Awareness of factors associating with poor outcomes is central to diagnostic and therapeutic efforts, and must be considered in the design of initiatives to reduce risk factors and improve outcomes of AKI in developing countries.

Plasmodium falciparum genotypes in matched peripheral, placental and umbilical cord blood in an area characterised by unstable malaria transmission in eastern Sudan.

BACKGROUND: There are few published studies on Plasmodium falciparum genotypes in peripheral, placental and umbilical cord blood in areas characterised by unstable malaria transmission. METHOD: A cross-sectional study was conducted to investigate P. falciparum genotypes in matched peripheral, placental and umbilical cord blood in eastern Sudan. Thick blood smears and P. falciparum merozoite surface protein 1 (MSP1) and 2 (MSP2) genes as polymorphic markers in polymerase chain reactions were investigated in 3 kinds of samples of 153 pregnant women at delivery. RESULTS: There was no significant difference in the prevalence of blood film-detected P. falciparum in which 5 (3.3%), 7 (4.6%) and 3 (2.0%) (P = 0.437) of the 153 samples were determined to be P. falciparum-positive by microscopy for maternal peripheral, placental and cord blood samples, respectively. Out of these 145 samples, 24 (16.6%), 39 (26.9%) and 24 (16.6%) (P = 0.039) of the peripheral, placental and cord samples, respectively, had submicroscopic parasitaemia (blood films were negative). There was no association between submicroscopic parasitaemia and age or parity. RO33 and K1 (MSP1 alleles) were detected in 21/29 (72.4%), 42/46 (85.7%), 26/27 (92.2%) and 6/29 (20.6), 16/46 (32.6) and 0(0) (P < 0.001) of the maternal, placental and cord samples, respectively. MAD20 was not detected in any of the samples. While the 3D7/IC1 allele was detected in 12 (41.3%), 30 (65.2%) and 4 (14.8%) (P < 0.001) of the peripheral, placental and cord samples, respectively, the FC (MSP2) allele was detected in only the 6 (20.6) placental samples. Multi-clonal infection was detected in 10 (34.4), 27 (58.6) and 3 (11.1) (P < 0.001) of the maternal placental and cord samples, respectively. CONCLUSION: Compared with the peripheral and cord samples, placental samples had a higher prevalence of submicroscopic parasitaemia. MSP1 alleles were predominant in the cord, while MSP2 alleles were predominant in the placental samples, which had a significant higher multiplicity of the infection.

High residual chloroquine blood levels in African children with severe malaria seeking healthcare.

Despite widespread resistance, chloroquine remains widely used in West Africa, particularly in home treatment. We examined chloroquine blood levels on admission to a referral hospital with respect to the manifestation of severe malaria in 290 Ghanaian children. Of the patients, 78% exhibited chloroquine concentrations (subtherapeutic, 35%; therapeutic, 37%; supratherapeutic, 6%) and 11% died. Most parasites (78%) carried the pfcrt-T76 chloroquine resistance mutation. High drug concentrations correlated with reduced parasitaemia but also with selection of resistant parasites, lower respiratory and heart rates, increased plasma lactate levels and impaired consciousness. Geometric mean chloroquine concentrations tended to be higher in children who died than in survivors (1.135 vs. 778nmol/l; P=0.09). Supratherapeutic drug levels (>5000nmol/l) were associated with fatal outcome (odds ratio 8.6; 95% CI 1.4-51.7). Residual chloroquine concentrations were found to be abundant in children with severe malaria and to be associated with alterations in the clinical manifestation of the disease and its case fatality. This may result from toxic effects of the drug and/or reflect preceding overtreatment in children with acute life-threatening disease. In areas of intense chloroquine resistance and frequent pre-treatment, additional administration of chloroquine at hospital admission is not only ineffective but may even further endanger patients.

Field-based evidence for linkage of mutations associated with chloroquine (pfcrt/pfmdr1) and sulfadoxine-pyrimethamine (pfdhfr/pfdhps) resistance and for the fitness cost of multiple mutations in P. falciparum.

Mutations in the Plasmodium falciparum pfcrt gene on chromosome 7 and possibly mutations in pfmdr1 on chromosome 5 have a role in conferring resistance against chloroquine (CQ), as do mutations of pfdhfr on chromosome 4 and pfdhps on chromosome 8 in terms of resistance against sulfadoxine/pyrimethamine (SP). The additive role of multiple mutations in the development of resistance to each drug suggests a non-random occurrence. In this study, parasite isolates were obtained from 50 patients with uncomplicated P. falciparum malaria from rural Eastern Sudan, an endemic setting with minimal overlap of infection. The parasite isolates were genotyped for detection of 12 alleles in CQ and SP resistance genes. Our main findings were: (1) the frequency of mutant alleles, pfcrt K76T, pfmdr1 N86Y, pfdhfr N51I, pfdhfr S108N, pfdhps K540E and pfdhps A581G were; 0.90, 0.86, 0.84, 0.84, 0.80 and 0.20, respectively. (2) No mutations were detected for the pfdhfr loci A16V, C59R and I164L, and for pfdhps loci S436A, A437G and A613S. (3) There was a statistically significant association between the mutations in: (i) the CQ resistance (CQR) genes, pfcrt T76 and pfmdr1 Y86 (P< or =0.001), (ii) the SP resistance (SPR) genes, pfdhfr I51, pfdhfr N108 and pfdhps E540 (P< or =0.001-0.04) and (iii) the CQ "i" and SP "ii" resistance genes (P=0.001) 4. The fitness cost of multiple mutations was revealed by a significantly reduced parasite density of isolates bearing the mutant alleles (P=0.048). However, the significantly higher gametocyte carriage rate among isolates with resistance mutations (P=0.001) is possibly an evolutionary mechanism for survival of mutant parasites.

Convergent adaptation of human lactase persistence in Africa and Europe.

A SNP in the gene encoding lactase (LCT) (C/T-13910) is associated with the ability to digest milk as adults (lactase persistence) in Europeans, but the genetic basis of lactase persistence in Africans was previously unknown. We conducted a genotype-phenotype association study in 470 Tanzanians, Kenyans and Sudanese and identified three SNPs (G/C-14010, T/G-13915 and C/G-13907) that are associated with lactase persistence and that have derived alleles that significantly enhance transcription from the LCT promoter in vitro. These SNPs originated on different haplotype backgrounds from the European C/T-13910 SNP and from each other. Genotyping across a 3-Mb region demonstrated haplotype homozygosity extending >2.0 Mb on chromosomes carrying C-14010, consistent with a selective sweep over the past approximately 7,000 years. These data provide a marked example of convergent evolution due to strong selective pressure resulting from shared cultural traits-animal domestication and adult milk consumption.

A fixed-dose 24-hour regimen of artesunate plus sulfamethoxypyrazine-pyrimethamine for the treatment of uncomplicated Plasmodium falciparum malaria in eastern Sudan.

BACKGROUND: Artemisinin-based combination therapy is increasingly being adopted as first-line antimalarial therapy. The choice of appropriate therapy depends on efficacy, cost, side effects, and simplicity of administration. METHODS: the efficacy of fixed co-formulated (f) artesunate-sulfamethoxypyrazine-pyrimethamine (AS+SMP f) administered at time intervals of 12 hours for a 24-hour therapy was compared with the efficacy of the same drug given as a loose combination (AS+SMP l) with a dose interval of 24 hours for 3 days for the treatment of uncomplicated Plasmodium falciparum malaria in eastern Sudan. RESULTS: seventy-three patients (39 and 34 in the fixed and the loose regimen of AS+SMP respectively) completed the 28-days of follow-up. On day 3; all patients in both groups were a parasitaemic but one patient in the fixed group of AS+SMP f was still febrile. Polymerase chain reaction genotyping adjusted cure rates on day 28 were 92.3% and 97.1% (P > 0.05) for the fixed and loose combination of AS+SMP respectively. Three (4.1%) patients (one in the fixed and two patients in the loose group of AS+SMP) in the study suffered drug-related adverse effects. Gametocytaemia was not detected during follow-up in any of the patients. CONCLUSION: both regimens of AS+SMP were effective and safe for the treatment of uncomplicated P. falciparum malaria in eastern Sudan. Due to its simplicity, the fixed dose one-day treatment regimen may improve compliance and therefore may be the preferred choice.

High prevalence of drug-resistance mutations in Plasmodium falciparum and Plasmodium vivax in southern Ethiopia.

BACKGROUND: In Ethiopia, malaria is caused by both Plasmodium falciparum and Plasmodium vivax. Drug resistance of P. falciparum to sulfadoxine-pyrimethamine (SP) and chloroquine (CQ) is frequent and intense in some areas. METHODS: In 100 patients with uncomplicated malaria from Dilla, southern Ethiopia, P. falciparum dhfr and dhps mutations as well as P. vivax dhfr polymorphisms associated with resistance to SP and P. falciparum pfcrt and pfmdr1 mutations conferring CQ resistance were assessed. RESULTS: P. falciparum and P. vivax were observed in 69% and 31% of the patients, respectively. Pfdhfr triple mutations and pfdhfr/pfdhps quintuple mutations occurred in 87% and 86% of P. falciparum isolates, respectively. Pfcrt T76 was seen in all and pfmdr1 Y86 in 81% of P. falciparum. The P. vivax dhfr core mutations N117 and R58 were present in 94% and 74%, respectively. CONCLUSION: These data point to an extraordinarily high frequency of drug-resistance mutations in both P. falciparum and P. vivax in southern Ethiopia, and strongly support that both SP and CQ are inadequate drugs for this region.

Risk mapping of visceral leishmaniasis: the role of local variation in rainfall and altitude on the presence and incidence of kala-azar in eastern Sudan.

Visceral leishmaniasis (VL) is a vector-borne disease highly influenced by environmental factors. A model was developed for mapping the distribution and incidence of VL in Gedaref State, eastern Sudan, in relation to different environmental factors. Geographical information systems (GIS) were used to extract and map regression results for environmental variables of 190 villages in Gedaref State, including rainfall, vegetation status, soil type, altitude, distance from river, topography, wetness indexes, and average rainfall estimates. VL incidence in each village was calculated from hospital records. By use of logistic and linear multivariate regression analyses, models were developed to determine which environmental factors explain variability in VL presence and incidence. We found that average rainfall and the altitude were the best predictors of VL incidence. The resulting models were mapped by GIS software predicting both VL presence or absence and incidence at any locality in Gedaref State. The results are discussed in relation to VL control.