Dynamics of host blood effects inGlossina morsitans sspp. infected withTrypanosoma congolense andT. brucei

The pattern of infection inGlossina morsitans morsitans andG. m. centralis membrane-fed on eland, buffalo or goat blood mixed withTrypanosoma congolense orT. brucei was studied from day 1 to day 10. Tsetse were initially permissive vectors, with most flies harbouring infections of 10⁴–10⁵ parasites on day 3. However, after a second blood meal on day 3, flies cleared many infections withG. m. morsitans clearing more infections thanG. m. centralis. Infective feeds of goat blood consistently increased final infection rates by limiting the number of infections lost between days 3 and 6. In further experiments withG. m. morsitans only, this effect was replicated by feeding flies on erythrocytes but not on serum. These results suggest that compounds from some mammalian erythrocytes match the target specificity ofG. m. morsitans midgut lectins and, hence, have a protective effect on trypanosome establishment in the fly.     

Influence of D(+)-glucosamine on infection rates and parasite loads in tsetse flies (Glossina spp.) infected with Trypanosoma brucei.

Teneral Glossina morsitans centralis, G. m. morsitans and G. pallidipes were infected with three different clones of Trypanosoma brucei in blood containing D(+)-glucosamine, an inhibitor of tsetse midgut lectin. On average, 5 days of D(+)-glucosamine treatment tripled infection rates, without affecting the proportion of infections that matured. Total infection rates were equal in males and females, but twice as many infections matured in males. Counts of parasites in the guts and salivary glands of 277 flies revealed order of magnitude differences among flies, with females consistently having 2-3-times as many parasites as males. Parasite numbers varied in a sex-specific manner among tsetse-clone combinations, but these differences were not correlated with similar large differences in infection rates. D(+)-glucosamine treatment had no significant effect on parasite loads.     

Relationship between Plasma Interleukin-12 (IL-12) and IL-18 Levels and Severe Malarial Anemia in an Area of Holoendemicity in Western Kenya

In this study, we investigated whether levels of interleukin-12 (IL-12) and IL-18 in plasma are associated with severe malarial anemia outcomes in an area of holoendemicity in western Kenya. We compared plasma IL-12 and IL-18 levels in six groups of children grouped into the categories aparasitemic, asymptomatic, mild malaria, high-density uncomplicated malaria (UC), moderate malarial anemia (MMA), or severe malarial anemia (SMA). IL-12 levels were significantly reduced in children with SMA (P < 0.05) but not in other groups compared to children in the aparasitemic control group. IL-18, a cytokine known to be critical for the induction of gamma interferon along with IL-12, was produced more frequently (70%) in children with UC (P = 0.06) than in children in the aparasitemic control group (32%). However, in the SMA group the IL-18 response rate declined to 30%, which was similar to that in the aparasitemic control group, which showed a 32% response rate. This finding suggests that the IL-18 response may be impaired in children with SMA. In summary, the results from this study support the hypothesis that impairment of IL-12 and/or IL-18 response may contribute to the development of severe malarial anemia in areas of holoendemicity for malaria.     

Predominance of null mutations in ataxia-telangiectasia

Ataxia-telangiectasia (A-T) is an autosomal recessive disorder involving cerebellar degeneration, immunodeficiency, chromosomal instability, radiosensitivity and cancer predisposition. The responsible gene, ATM, was recently identified by positional cloning and found to encode a putative 350 kDa protein with a Pl 3-kinase-like domain, presumably involved in mediating cell cycle arrest in response to radiation-induced DNA damage. The nature and location of A-T mutations should provide insight into the function of the ATM protein and the molecular basis of this pleiotropic disease. Of 44 A-T mutations identified by us to date, 39 (89%) are expected to inactivate the ATM protein by truncating it, by abolishing correct initiation or termination of translation, or by deleting large segments. Additional mutations are four smaller in-frame deletions and insertions, and one substitution of a highly conserved amino acid at the Pl 3-kinase domain. The emerging profile of mutations causing A-T is thus dominated by those expected to completely inactivate the ATM protein. ATM mutations with milder effects may result in phenotypes related, but not identical, to A-T.      

High throughput parallel analysis of hundreds of patient samples for more than 100 mutations in multiple disease genes

As more mutations are identified in genes of known sequence, there is a crucial need in the areas of medical genetics and genome analysis for rapid, accurate and cost-effective methods of mutation detection. We have developed a multiplex allele-specific diagnostic assay (MASDA) for analysis of large numbers of samples (> 500) simultaneously for a large number of known mutations (> 100) in a single assay. MASDA utilizes oligonucleotide hybridization to interrogate DNA sequences. Multiplex DNA samples are immobilized on a solid support and a single hybridization is performed with a pool of allele-specific oligonucleotide (ASO) probes. Any probes complementary to specific mutations present in a given sample are in effect affinity purified from the pool by the target DNA. Sequence-specific band patterns (fingerprints), generated by chemical or enzymatic sequencing of the bound ASO(s), easily identify the specific mutation(s). Using this design, in a single diagnostic assay, we tested samples for 66 cystic fibrosis (CF) mutations, 14 beta-thalassemia mutations, two sickle cell anemia (SCA) mutations, three Tay-Sachs mutations, eight Gaucher mutations, four mutations in Canavan disease, four mutations in Fanconi anemia, and five mutations in BRCA1. Each mutation was correctly identified. Finally, in a blinded study of 106 of these mutations in > 500 patients, all mutations were properly identified. There were no false positives or false negatives. The MASDA assay is capable of detecting point mutations as well as small insertion or deletion mutations. This technology is amenable to automation and is suitable for immediate utilization for high-throughput genetic diagnostics in clinical and research laboratories.      

Markers in HIV associated tuberculosis in Kenya

Tuberculosis is one of the most frequent opportunistic infections in HIV-infected patients in developing countries. In some instances, manifestations of pulmonary tuberculosis precede all other HIV related signs and symptoms because of the high virulence of M. tuberculosis. In order to characterise the interaction between these two pathogens, clinical and immunological parameters in pulmonary tuberculosis patients with and without HIV infection were compared. Amongst newly diagnosed pulmonary tuberculosis patients the association of some of these changes with the clinical outcome were evaluated. Of these, 44% were co-infected with HIV. Pulmonary tuberculosis patients with HIV-1 presented more frequently with lymphadenopathy and diarrhoea than those without HIV-1. Peripheral blood CD4+ counts were significantly lower in patients with pulmonary tuberculosis with HIV-1 than those with pulmonary tuberculosis alone, P= 0.0292. Low CD4+ lymphocyte counts, lymphadenopathy and BCG scar absence could serve as indicators of HIV-1 infection in pulmonary tuberculosis (PTB) patients.