Insights into the possible role of IFNG and IFNGR1 in Kala-azar and Post Kala-azar Dermal Leishmaniasis in Sudanese patients.

BACKGROUND: Little is known about the parasite/host factors that lead to Post Kala-azar Dermal Leishmaniasis (PKDL) in some visceral leishmaniasis (VL) patients after drug-cure. Studies in Sudan provide evidence for association between polymorphisms in the gene (IFNGR1) encoding the alpha chain of interferon-γ receptor type I and risk of PKDL. This study aimed to identify putative functional polymorphisms in the IFNGR1 gene, and to determine whether differences in expression of interferon-γ (IFNG) and IFNGR1 at the RNA level are associated with pathogenesis of VL and/or PKDL in Sudan. METHODS: Sanger sequencing was used to re-sequence 841 bp of upstream, exon1 and intron1 of the IFNGR1 gene in DNA from 30 PKDL patients. LAGAN and SYNPLOT bioinformatics tools were used to compare human, chimpanzee and dog sequences to identify conserved noncoding sequences carrying putative regulatory elements. The relative expression of IFNG and IFNGR1 in paired pre- and post-treatment RNA samples from the lymph nodes of 24 VL patients, and in RNA samples from skin biopsies of 19 PKDL patients, was measured using real time PCR. Pre- versus post-treatment expression was evaluated statistically using the nonparametric Wilcoxon matched pairs signed-rank test. RESULTS: Ten variants were identified in the 841 bp of sequence, four of which are novel polymorphisms at -77A/G, +10 C/T, +18C/T and +91G/T relative to the IFNGR1 initiation site. A cluster of conserved non-coding sequences with putative regulatory variants was identified in the distal promoter of IFNGR1. Variable expression of IFNG was detected in lymph node aspirates of VL patients before treatment, with a marked reduction (P = 0.006) in expression following treatment. IFNGR1 expression was also variable in lymph node aspirates from VL patients, with no significant reduction in expression with treatment. IFNG expression was undetectable in the skin biopsies of PKDL cases, while IFNGR1 expression was also uniformly low. CONCLUSIONS: Uniformly low expression of IFN and IFNGR1 in PKDL skin biopsies could explain parasite persistence and is consistent with prior demonstration of genetic association with IFNGR1 polymorphisms. Identification of novel potentially functional rare variants at IFNGR1 makes an important general contribution to knowledge of rare variants of potential relevance in this Sudanese population.

Fatal Nocardia paucivorans Lower Respiratory Tract Infection in a Tuberculosis Suspect Who Worked as Gold Miner.

Nocardiosis is an opportunistic infection that affects both immunocompromised as well as immunocompetent patients. The main infections occur as soft tissue and lung infections although they might disseminate to various organs. This is a case study aimed to reflect the severity of the disease and the patient's risk factors associated with the infection. A sputum sample was collected from tuberculosis (TB) suspects for culture. Nocardia-like colonies were isolated, purified, and sent to BGI Company (Hongkong, China). Standard forward sequencing of 16S rRNA was done by ABI Genetic Analyzer (Applied Biosystems). Sequence alignment and nucleotide basic local alignment search tool (BLAST) were done using National Center for Bioinformatics (NCBI) Nucleotide BLAST. Biochemical identification to the colonies was done using an automation system (BD Phoenix™) to confirm the identification. Nocardia paucivorans was identified from the TB suspect. Risk factors were identified as extensive contact to dust, absence of primary care units with complete facilities, and old age. Since the infection of the lungs caused by Nocardia might be similar to pulmonary TB, this case report highlights the importance of accurate diagnosis and identification procedures to differentiate between the two.

Molecular surveillance of putative drug resistance markers of antifolate and artemisinin among imported Plasmodium falciparum in Qatar.

Malaria remains a significant public health challenge and is of global importance. Imported malaria is a growing problem in non-endemic areas throughout the world and also in Qatar due to a massive influx of migrants from endemic countries. Antimalarial drug resistance is an important deterrent in our fight against malaria today. Molecular markers mirror intrinsic antimalarial drug resistance and their changes precede clinical resistance. Thus, in the present study, molecular markers of sulphadoxine-pyrimethamine (Pfdhfr and Pfdhps) and artemisinin (PfATPase6 and Pfk13) were sequenced to determine the drug resistance genotypes among 118 imported P. falciparum isolates in Qatar, between 2013 and 2016. All the isolates had mutant Pfdhfr alleles, with either double mutant (51I/108N) (59.3%) or triple mutant (51I, 59R and 108N) (30.6%) genotypes. I164L substitution was not found in this study. In case of Pfdhps, majority of the samples were carriers of either single (S436A/ A437G/ K540E) mutant (47.2%) or double (S436A/K540E, A437G/K540E, K540E/A581G) mutant (39.8%). A single novel point mutation (431V) was observed in the samples originated from Nigeria and Ghana. Polymorphisms in PfATPase6 were absent and only one non-synonymous mutation in Pfk13 was found at codon G453A from a sample of Kenyan origin. High levels of sulphadoxine-pyrimethamine resistance in the present study provide potential information about the spread of antimalarial drug resistance and will be beneficial for the treatment of imported malaria cases in Qatar.

Evolution of Plasmodium falciparum drug resistance genes following artemisinin combination therapy in Sudan.

BACKGROUND: Malaria control efforts in Sudan rely heavily on case management. In 2004, health authorities adopted artemisinin-based combination therapies (ACTs) for the treatment of uncomplicated malaria. However, some recent surveys have reported ACT failure and a prevalent irrational malaria treatment practice. Here we examine whether the widespread use of ACT and failure to adhere to national guidelines have led to the evolution of drug resistance genes. METHODS: We genotyped known drug resistance markers (Pfcrt, Pfmdr-1, Pfdhfr, Pfdhps, Pfk13 propeller) and their flanking microsatellites among Plasmodium falciparum isolates obtained between 2009 and 2016 in different geographical regions in Sudan. Data were then compared with published findings pre-ACT (1992-2003). RESULTS: A high prevalence of Pfcrt76T, Pfmdr-1-86Y, Pfdhfr51I, Pfdhfr108N, Pfdhps37G was observed in all regions, while no Pfk13 mutations were detected. Compared with pre-ACT data, Pfcrt-76T and Pfmdr-1-86Y have decayed, while Pfdhfr-51I, Pfdhfr-108N and Pfdhps-437G strengthened. Haplotypes Pfcrt-CVIET, Pfmdr-1-NFSND/YFSND, Pfdhfr-ICNI and Pfdhps-SGKAA predominated in all sites. Microsatellites flanking drug resistance genes showed lower diversity than neutral ones, signifying high ACT pressure/selection. CONCLUSIONS: Evaluation of P. falciparum drug resistance genes in Sudan matches the drug deployment pattern. Regular monitoring of these genes, coupled with clinical response, should be considered to combat the spread of ACT resistance.

Distribution of Mutations Associated with Antifolate and Chloroquine Resistance among Imported Plasmodium vivax in the State of Qatar.

Plasmodium vivax is the most prevalent parasite worldwide, escalating by spread of drug resistance. Currently, in Qatar, chloroquine (CQ) plus primaquine are recommended for the treatment of P. vivax malaria. The present study examined the prevalence of mutations in dihydrofolate reductase (dhfr), dihydropteroate synthase (dhps) genes and CQ resistance transporter (crt-o) genes, associated with sulphadoxine-pyrimethamine (SP) and chloroquine resistance, among imported P. vivax cases in Qatar. Blood samples were collected from patients positive for P. vivax and seeking medical treatment at Hamad General Hospital, Doha, during 2013-2016. The Sanger sequencing method was performed to examine the single nucleotide polymorphisms in Pvdhfr, Pvdhps, and Pvcrt-o genes. Of 314 examined P. vivax isolates, 247 (78.7%), 294 (93.6%) and 261 (83.1%) were successfully amplified and sequenced for Pvdhfr, Pvdhps, and Pvcrt-o, respectively. Overall, 53.8% (N = 133) carried mutant alleles (58R/117N) in Pvdhfr, whereas 77.2% (N = 227) and 90% (N = 235) isolates possessed wild type allele in Pvdhps and Pvcrt-o genes, respectively. In addition, a total of eleven distinct haplotypes were detected in Pvdhfr/Pvdhps genes. Interestingly, K10 insertion in the Pvcrt-o gene was observed only in patients originating from the Indian subcontinent. The results suggested that CQ remains an acceptable treatment regimen but further clinical data are required to assess the effectiveness of CQ and SP in Qatar to support the current national treatment guidelines. In addition, limited distribution of genetic polymorphisms associated with CQ and SP resistance observed in imported P. vivax infections, necessitates regular monitoring of drug resistant P. vivax malaria in Qatar.

Transmission of Plasmodium vivax in Duffy-negative individuals in central Sudan.

BACKGROUND: Due to the recently observed rise in Plasmodium vivax incidence in Sudan and reported transmission in Duffy-negative individuals; we aimed to assess the possibility of P. vivax transmission in Duffy-negative individuals in Gezira state, central Sudan. METHOD: A total of 126 suspected malaria patients were diagnosed with P. vivax infection using microscopy, RDT and PCR. PCR-RFLP was used to genotype participants Duffy status. RESULTS: Forty eight (38%) were positive for P. vivax infection by PCR. Four patients (8.3%) were homozygous Duffy-negative. CONCLUSION: These results confirm that P. vivax can infect Duffy-negative individuals, suggesting alternative mechanisms to bind and invade erythrocytes.