Moawia M. Mukhtar

Immunocompetence may be important in the effectiveness of Mectizan® (ivermectin) in the treatment of human onchocerciasis

Mectizan (Ivermectin) has been proved to be central to the control of onchoceriasis through self-sustainable community-based treatment. The possibility of parasitological unresponsiveness to this treatment or selection for drug resistance has emerged recently in many occasions. The reason for the reduced ability of Mectizan to maintain low levels of dermal microfilariae and early recurrent pruritus can only be speculated upon. Here, we report our own findings to address this particular issue.

Alum-precipitated autoclaved Leishmania major plus bacille Calmette-Guérrin, a candidate vaccine for visceral leishmaniasis: safety, skin-delayed type hypersensitivity response and dose finding in healthy volunteers

In a previous efficacy study, autoclaved Leishmania major (ALM) + bacille Calmette-Guérrin (BCG) vaccine was shown to be safe, but not superior to BCG alone, in protecting against visceral leishmaniasis. From June 1999 to June 2000, we studied the safety and immunogenicity of different doses of alum-precipitated ALM + BCG vaccine mixture administered intradermally to evaluate whether the addition of alum improved the immunogenicity of ALM. Twenty-four healthy adult volunteers were recruited and sequentially allocated to receive either 10 microg, 100 microg, 200 microg, or 400 microg of leishmanial protein in the alum-precipitated ALM + BCG vaccine mixture. Side effects were minimal for all doses and confined to the site of injection. All volunteers in the 10 microg, 100 microg, and 400 microg groups had a leishmanin skin test (LST) reaction of > or = 5 mm by day 42 and this response was maintained when tested after 90 d. Only 1 volunteer out of 5 in the 200 microg group had a LST reaction of > or = 5 mm by day 42 and the reasons for the different LST responses in this group are unclear. This is the first time that an alum adjuvant with ALM has been in used in humans and the vaccine mixture was safe and induced a strong delayed type hypersensitivity (DTH) reaction in the study volunteers. On the basis of this study we suggest that 100 1 microg of leishmanial protein in the vaccine mixture is a suitable dose for future efficacy studies, as it induced the strongest DTH reaction following vaccination.

Detection of antibodies to Leishmania donovani in animals in a kala-azar endemic region in eastern Sudan: a preliminary report

The prevalence of antibodies against Leishmania donovani in selected domestic and wild animal species in 2 villages in Sudan with active L. donovani transmission in humans was investigated. Screening of domestic animals (donkeys, cows, sheep, goats, camels and dogs) with the direct agglutination test (DAT) detected reaction rates above the cut-off titres in donkeys (68.7%), cows (21.4%) and goats (8.5%), and which were also found in wild rats (5.5%). Sera of sheep, camels and dogs had a weak agglutination reaction below the cut-off titre. Testing of the same sera by enzyme-linked immunosorbent assay (ELISA), against a lysate of L. donovani promastigotes, showed reaction rates above the cut-off optical density in cows (47.6%), goats (13.6%), and in rats (4.1%). No Leishmania parasite was isolated from spleen, liver, bone-marrow or spleen of Nile rats.

Mastitis in Camelus dromedarius and the somatic cell content of camels' milk

Seven hundred and sixty-three camels from 400 herds of local Sudanese camel breeds were investigated for the prevalence of mastitis, identification of its bacterial causes and determination of the leucocyte contents of camels milk. One hundred and forty-nine (19.5 per cent) of the 763 camels examined were diagnosed as mastitis cases based on clinical signs. One hundred and fifty-nine (47.3 per cent) of the 336 randomly selected milk samples were reactive in a rapid mastitis test and 16 of the 153 tested samples contained Brucella abortus agglutinating antibodies. Streptococcus, Staphylococcus, Micrococcus and Aerobacter species and Escherichia coli were found to be the main causes of mastitis (in descending order). The leucocyte contents of the 757 milk samples ranged from < 5 x 10(5) to > 7.5 x 10(6) leucocyte ml-1 and 42.8 per cent of the samples contained < 5 x 10(5) cells ml-1. Neither significant correlation between the leucocyte content of milk and isolated bacterial species nor significant variation in leucocyte contents during different stages of lactation were detected.

Cytotaxonomic and molecular analysis of Simulium (Edwardsellum) damnosum sensu lato (Diptera: Simuliidae) from Abu Hamed, Sudan

Abstract The northernmost focus forOnchocerca volvulusLeuckhart (Nematoda: Onchocercidae), the causative agent of human onchocerciasis, is found along the Nile near the town of Abu Hamed in Sudan. The vector forO. volvulusat this focus is a single monomorphic population ofSimulium (Edwardsellum) damnosumTheobald. This black fly population is limited to a small area between the fourth and fifth cataracts of the Nile River that is isolated geographically from all other populations ofS. damnosumsensu lato. Phylogenies produced from cytological analyses and sequence data derived from the NADH dehydrogenase subunit 4 and 16S rRNA genes indicate that Abu Hamed black flies are similar to, but distinct from, the savanna-dwelling sibling species ofS. damnosums.l.,Simulium (Edwardsellum) damnosumsensu strictu Theobald, andS. (Edwardsellum) sirbanumVajime & Dunbar. The DNA sequence and the cytological data support the hypothesis that the black fly population present in Abu Hamed may represent a new sibling species ofS. damnosums.l. We propose that this population be informally designated as the hamedense form of theSimulium damnosumcomplex.

Epidemiology of schistosomiasis in Gezira area Central Sudan and analysis of cytokine profiles

OBJECTIVE To determine and compare anti-schistosoma IgG, interleukin-10 (IL-10) and interferon-γ(IFN-γ) levels in the serum of patients and endemic controls and to investigate the epidemiological situation of Al-Hebaika village in the northern part of Gezira Agricultural Irrigation Scheme in 2005. METHODS During 2005 survey, serum were collected from 118 villagers. Sixty eight were parasitological positive (patients), and 50 were negative (endemic controls). Indirect ELISA was used to measure and compare the levels of immunoglobulin G (IgG) against Schistsoma mansoni (S. mansoni) soluble worm antigen (SWA) in the patients and endemic control groups from the village and compared with 20 healthy non endemic controls. Sandwich ELISA was also used to measure and compare IL-10 and IFN-γ in the serum of the selected groups. RESULTS The overall prevalence of S. haematobium was 20.0% and 0.9% in the first and the second surveys respectively, while the intensity of infection was the same in the two surveys 1.38 [geometric mean egg count (GMFC)]. The overall prevalence of S. mansoni infection was 68.5% and 15.4%, while the intensity of infection was 2.75 (GMEC) and 1.70 (GMEC) in the two surveys respectively. IgG reactivity against SWA showed no significant difference between Schistosoma positive patients and endemic controls. However, there were high significant differences between each of these two groups and the non endemic control group (P= 0,000). Schistosoma patients and exposed controls had significantly higher IL-10 concentration compared with non endemic controls. While endemic controls showed significantly higher IFN-γ concentration than patients (P = 0.000). Also there was very significant difference between IFN-γ levels of each of patients endemic controls and that of the non endemic controls (P = 0.003). CONCLUSIONS The study concluded that IFN-γ has a role in the natural resistant to schistosoma mansoni infection. The prevalence and intensity of S. mansoni in the Gezira Irrigation Scheme was greatly reduced. S. haematobium has disappeared from the area.

The burden of Onchocerca volvulus in Sudan

Onchocerciasis has been reported in Sudan since 1908 and now prevails in three endemic regions known as the southern, northern and eastern foci. The southern focus is the largest, with nodule and blindness rates exceeding 80% and 12%, respectively, in certain villages. Onchocercal infection in this region causes only a mild skin reaction although microfilarial loads in the skin are high. In contrast, those with onchocerciasis in the northern focus, located between the fourth and fifth Nile cataracts, present with limited but severe skin reactions, low nodule rates (16%), low microfilarial loads in the skin and no ocular involvement. The characteristics of patients from the eastern focus, close to the border with Ethiopian border are similar to those in the north, although most onchocercal skin disease in this area comprises the severe localized pruritus known as sowda.

Screening and Characterization of RAPD Markers in Viscerotropic Leishmania Parasites

Visceral leishmaniasis (VL) is mainly due to the Leishmania donovani complex. VL is endemic in many countries worldwide including East Africa and the Mediterranean region where the epidemiology is complex. Taxonomy of these pathogens is under controversy but there is a correlation between their genetic diversity and geographical origin. With steady increase in genome knowledge, RAPD is still a useful approach to identify and characterize novel DNA markers. Our aim was to identify and characterize polymorphic DNA markers in VL Leishmania parasites in diverse geographic regions using RAPD in order to constitute a pool of PCR targets having the potential to differentiate among the VL parasites. 100 different oligonucleotide decamers having arbitrary DNA sequences were screened for reproducible amplification and a selection of 28 was used to amplify DNA from 12 L. donovani, L. archibaldi and L. infantum strains having diverse origins. A total of 155 bands were amplified of which 60.65% appeared polymorphic. 7 out of 28 primers provided monomorphic patterns. Phenetic analysis allowed clustering the parasites according to their geographical origin. Differentially amplified bands were selected, among them 22 RAPD products were successfully cloned and sequenced. Bioinformatic analysis allowed mapping of the markers and sequences and priming sites analysis. This study was complemented with Southern-blot to confirm assignment of markers to the kDNA. The bioinformatic analysis identified 16 nuclear and 3 minicircle markers. Analysis of these markers highlighted polymorphisms at RAPD priming sites with mainly 5′ end transversions, and presence of inter– and intra– taxonomic complex sequence and microsatellites variations; a bias in transitions over transversions and indels between the different sequences compared is observed, which is however less marked between L. infantum and L. donovani. The study delivers a pool of well-documented polymorphic DNA markers, to develop molecular diagnostics assays to characterize and differentiate VL causing agents.

Tuberculosis drug resistance isolates from pulmonary tuberculosis patients, Kassala State, Sudan.

BACKGROUND This study was conducted in Kassala Teaching Hospital, Kassala State, Sudan (January 2006-June 2008) to determine the rate of mycobacterium drug resistance to anti-tuberculous treatment and to explore the genotype of Mycobacterium tuberculosis resistant isolates using rpoB gene. METHODS 53 isolates of mycobacterium isolated from pulmonary tuberculosis (PTB) patients from Kassala State were subjected to drug susceptibility testing (DST) to anti-tuberculous drugs; 10 M.tuberculosis complex (MTBC) resistant isolates were subjected to polymerase chain reaction (PCR), and commercially the amplified DNA was sequenced. RESULTS DST detected resistance in 23/53 (43.39%) isolates, among which rifampicin had a high number of resistant isolates (13/23), followed by streptomycin (11/23), and multi-drug resistance was detected in 5 isolates. DNA sequence analysis of 10 MTBC-resistant isolates detected variations within and outside the rifampicin resistant determining region (RRDR). Variation within RRDR was detected at positions 512 (AGC/ATC, Ser/Ile), and 528 (CGC/CTC, Arg/Leu). Outside the RRDR region variations were detected at positions 498 (GTG/GGG, Val/gly), 488 (ACA/ACC, Thr/Thr), which is a silent mutation. Insertions were observed at positions 484, 496 (GTG/GTGA, CGG/CAGG, respectively). Deletion was observed at position 487 (ATC/_TC). DISCUSSION AND CONCLUSION This study revealed that high resistance to rifampicin was associated with various point mutations in and out of the RRDR of the rpoB gene. Molecular methods are needed for early detection of TB disease and drug resistance.