Maowia M. Mukhtar

Autoclaved Leishmania major vaccine for prevention of visceral leishmaniasis: a randomised, double-blind, BCG-controlled trial in Sudan

BACKGROUND Visceral leishmaniasis is a major cause of morbidity and mortality in the Sudan. Drug treatment is expensive, and drug resistance is becoming increasingly common. Safe, effective, and cheap vaccines are needed. We report the results of a vaccine trial against human visceral leishmaniasis. METHODS We undertook a double-blind randomised trial to test the safety and efficacy of an autoclaved Leishmania major (ALM) promastigote vaccine (1 mg per dose). Of 5093 volunteers screened, 2306 had negative leishmanin skin tests and reciprocal titres of less than 6400 in the direct agglutination test. They were randomly assigned two doses of ALM mixed with BCG or BCG alone. Volunteers were followed up for 2 years. The primary endpoint was clinical visceral leishmaniasis or post-kala-azar dermal leishmaniasis. Analyses were by intention to treat. FINDINGS Side-effects were confined to the injection site. The cumulative frequency of visceral leishmaniasis at 2 years did not differ significantly between the group assigned ALM plus BCG and that assigned BCG alone (133/1155 [11.5%] vs 141/1151 [12.3%], p=0.6). The vaccine efficacy was 6% (95% CI -18 to 25). The proportion of individuals showing leishmanin skin conversion was significantly higher in the ALM plus BCG group than in the BCG alone group throughout follow-up (303 [30%] vs 72 [7%] at 42 days). Individuals whose leishmanin test converted after vaccination (induration > or =5 mm) had a significantly lower frequency of visceral leishmaniasis than non-responders (27/375 [7.2%] vs 210/1660 [12.7%], p=0.003). INTERPRETATION We found no evidence that two doses of ALM plus BCG offered significant protective immunity against visceral leishmaniasis compared with BCG alone. Leishmanin skin conversion with an induration of 5 mm or more in either group was associated with protection from the disease.

C-terminal titin deletions cause a novel early-onset myopathy with fatal cardiomyopathy

The giant protein titin is essential for striated muscle development, structure, and elasticity. All titin mutations reported to date cause late‐onset, dominant disorders involving either skeletal muscle or the heart. Our aim was to delineate the phenotype and determine the genetic defects in two consanguineous families with an early‐onset, recessive muscle and cardiac disorder.

A Global Comparative Evaluation of Commercial Immunochromatographic Rapid Diagnostic Tests for Visceral Leishmaniasis

Accuracy of rapid diagnostic tests was high in the Indian subcontinent; however, in Brazilian and East African samples, reduced sensitivity suggests that several cannot be used alone to exclude visceral leishmaniasis. Data on ease of use and performance using whole blood and in human immunodeficiency virus coinfections is needed.

Design, Development and Evaluation of rK28-Based Point-of-Care Tests for Improving Rapid Diagnosis of Visceral Leishmaniasis

Background Visceral leishmaniasis (VL) is diagnosed by microscopic confirmation of the parasite in bone marrow, spleen or lymph node aspirates. These procedures are unsuitable for rapid diagnosis of VL in field settings. The development of rK39-based rapid diagnostic tests (RDT) revolutionized diagnosis of VL by offering high sensitivity and specificity in detecting disease in the Indian subcontinent; however, these tests have been less reliable in the African subcontinent (sensitivity range of 75–85%, specificity of 70–92%). We have addressed limitations of the rK39 with a new synthetic polyprotein, rK28, followed by development and evaluation of two new rK28-based RDT prototype platforms. Methodology/Principal Findings Evaluation of 62 VL-confirmed sera from Sudan provided sensitivities of 96.8% and 93.6% (95% CI = K28: 88.83–99.61%; K39: 84.30–98.21%) and specificities of 96.2% and 92.4% (95% CI = K28: 90.53–98.95%; K39: 85.54–96.65%) for rK28 and rK39, respectively. Of greater interest was the observation that individual VL sera with low rK39 reactivity often had much higher rK28 reactivity. This characteristic of the fusion protein was exploited in the development of rK28 rapid tests, which may prove to be crucial in detecting VL among patients with low rK39 antibody levels. Evaluation of two prototype lateral flow-based rK28 rapid tests on 53 VL patients in Sudan and 73 VL patients in Bangladesh provided promisingly high sensitivities (95.9% [95% CI = 88.46–99.1 in Sudan and 98.1% [95% CI = 89.93–99.95%] in Bangladesh) compared to the rK39 RDT (sensitivities of 86.3% [95% CI = 76.25–93.23%] in Sudan and 88.7% [95% CI = 76.97–95.73%] in Bangladesh). Conclusions/Significance Our study compares the diagnostic accuracy of rK39 and rK28 in detecting active VL cases and our findings indicate that rK28 polyprotein has great potential as a serodiagnostic tool. A new rK28-based RDT will prove to be a valuable asset in simplifying VL disease confirmation at the point-of-care.

A study of the urban malaria transmission problem in Khartoum

A study of malaria prevalence and transmission was carried out in Khartoum, the capital of Sudan. The sentinel sites were El manshia, an urban area on the Blue Nile and Ed dekheinat, a lower-income peri-urban area bordering the White Nile. Anopheles arabiensis, the only malaria vector encountered, was present throughout the year although vector density varied seasonally. Plasmodium falciparum was the only species found in El manshia. In Ed dekheinat P. falciparum, Plasmodium ovale and Plasmodium vivax constituted 84.9, 8.2 and 6.9% of the cases, respectively. Plasmodium ovale appears to have recently spread into Khartoum since it has not previously been reported there. We conclude that focal transmission of malaria in the districts bordering both Niles has become established and that the reservoir of human infections has increased in recent years leading to increased risk of malaria epidemics, particularly in the aftermath of seasonal flooding.

The protective M proteins of the equine group C streptococci

The group C streptococci are the most commonly isolated bacteria from disease states in the horse. Important virulence factors of S. equi and S. zooepidemicus are the hyaluronic acid capsule and the antiphagocytic fibrillar M protein located on the surface of the cell wall and extending into and through the capsule. The hyaluronic acid capsule is non-antigenic and so is not involved in protective immunity. The M protein, a superantigen, elicits very strong B and T cell responses that may result in protective immunity mediated by opsonic antibodies in plasma and by locally synthesized IgG and IgA on the pharyngeal mucosa. However, vaccines based on acid or mutanolysin extracted M protein do not confer a high level of protection against field exposure. Protective antibodies to S. equi or S. zooepidemicus can in part be assayed by the bactericidal test that measures opsonization for equine neutrophils. A mouse-challenge model has also been used to test immunizing potency of streptococcal extracts and in a passive protection test for protective antibody. There is as yet no means of distinguishing protective opsonic or mucosal antibodies from other antibodies produced against the many epitopes on the M molecule.

Measles in suburban Khartoum: an epidemiological and clinical study

Clinical and epidemiological data were collected from 187 clinically diagnosed measles patients in Haj Yousif area, suburban Khartoum. Laboratory tests confirmed the diagnosis in 141 (75%) of the cases, but demonstrated that in 46 (25%) patients the clinical symptoms were not caused by an acute measles virus (MV) infection. According to their vaccination card, 59% of the laboratory‐confirmed measles cases had been vaccinated for measles. Compared with non‐measles rash disease cases, confirmed measles cases more often had severe illness (P < 0.0001), were dehydrated (P=0.01) at presentation and less likely to recover without complications [OR 0.19 (95% CI 0.09, 0.39)]. There was no difference in death rate (P=0.20). Underweight [weight‐for‐age Z score (WAZ) ≤ −2 SD] was an independent predictor of recovery with complications [OR 0.4 (95% CI 0.2, 0.99)]. Severe measles cases (those who developed diarrhoea, pneumonia, otitis media, encephalitis or haemorrhagic rash) had similar vaccination rates and time intervals since vaccination as uncomplicated measles cases. Although severe measles had lower WAZ‐scores (P=0.004), none of the nutritional parameters studied were predictive of outcome. Mortality was higher in the severe measles group [OR 8.8 (95% CI 1.7, 85.2)]. In 11 of 141 confirmed measles cases serological evidence of a recent infection with another virus was found, most commonly varicella zoster virus and dengue virus; spotted fever and rubella were among the most frequent diagnoses in 17 of 47 cases of the non‐measles cases.

Application of molecular biological techniques for detection of epizootic hemorrhagic disease virus (EHDV-318) recovered from a sentinel calf in central Sudan

Epizootic hemorrhagic disease virus (EHDV), isolate 318 (EHDV-318), an untyped virus recovered from a sentinel calf herd at the Khartoum University farm in central Sudan, was characterized using molecular biological techniques. With dot blot hybridization technique, a cDNA probe derived from genome segment 6 of EHDV-2 (Alberta strain) hybridized with RNA from EHDV-318. Application of serogroup-specific EHDV polymerase chain reaction (PCR) to EHDV-318 RNA resulted in specific amplification of a 387 bp PCR product. Amplification product was visualized on ethidium bromide-stained agarose gel. Specificity of the PCR products was confirmed by chemiluminescent hybridization with a non-radiolabelled internal probe. No amplification product or hybridization signal was detected when the serotype-specific EHDV-1 or EHDV-2 PCR-based assays were applied to RNA from EHDV-318. The scientific data presented in this study indicated that cDNA probes and serogroup-specific PCR-based assay can be used to classify the virus as a member of EHDV serogroup, and as serotypically distinct from EHDV-1 and EHDV-2.

Malaria Risk Mapping for Control in the Republic of Sudan

Evidence shows that malaria risk maps are rarely tailored to address national control program ambitions. Here, we generate a malaria risk map adapted for malaria control in Sudan. Community Plasmodium falciparum parasite rate (PfPR) data from 2000 to 2010 were assembled and were standardized to 2–10 years of age (PfPR2–10). Space-time Bayesian geostatistical methods were used to generate a map of malaria risk for 2010. Surfaces of aridity, urbanization, irrigation schemes, and refugee camps were combined with the PfPR2–10 map to tailor the epidemiological stratification for appropriate intervention design. In 2010, a majority of the geographical area of the Sudan had risk of < 1% PfPR2–10. Areas of meso- and hyperendemic risk were located in the south. About 80% of Sudans population in 2011 was in the areas in the desert, urban centers, or where risk was < 1% PfPR2–10. Aggregated data suggest reducing risks in some high transmission areas since the 1960s.

A new complex homozygous large rearrangement of the PINK1 gene in a Sudanese family with early onset Parkinson’s disease

PARK2 and PINK1 gene mutations are involved in recessive early onset Parkinson’s disease (EOPD). In order to determine the causative mutations in three affected sibs from a consanguineous Sudanese family with EOPD, multiplex ligation-dependent probe amplification was performed and revealed that the patients were homozygous for a deletion of PINK1 exons 4 to 8. Breakpoint analysis revealed a complex rearrangement combining a large deletion and the insertion of a sequence duplicated from the DDOST gene intron 2, located near the PINK1 gene. As breakpoint sequences displayed only three base pairs of homology, this rearrangement may result from Fork Stalling and Template Switching mechanism. This third large rearrangement of PINK1 enlarges the mutation spectrum and, together with recent published data in Tunisian patients with EOPD, points out that PINK1 gene analysis, including search for large rearrangement, should be considered in early onset recessive PD patients, particularly those from Arab origin.