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HIV serological diagnosis has evolved during the last decade to give rise to rapid testing using biological materials, such as blood or oral mucosal transudate (OMT). However, blood collection is not always welcomed, justifying the evaluation of OMT-based devices. In a cross sectional study carried out in May 2011 aimed at evaluating the level of awareness about OMT based HIV tests, questionnaires were administered to participants who consented to take part in the study. Eighty-five percent (n = 1520) of participants reported a lack of awareness of HIV oral screening before the study, and surprisingly, no association was found between the awareness of participants and their educational level (p = 0.768). There was also no association (p = 0.743) found between having had previous screening tests and awareness of oral testing. The percentage of participants who accepted the oral test before being informed about it was 31.3% (n = 1520). After sensitization, 76.3% (n = 1520)preferred oral screening for future tests (p = 0). These results reveal that if the OMT based test is affordable, its implementation as a screening tool in the general population could greatly increase participation in screening campaigns and is welcomed by those who want to self-test in a non-invasive way. This will create a better estimation of the national HIV prevalence. Its use could then have a significant public health impact on HIV prevention and clinical management.

A culture-independent approach was used to elucidate the microbial diversity and structure in the anaerobic-aerobic reactors integrated with a constructed wetland for the treatment of tannery wastewater in Modjo town, Ethiopia. The system has been running with removal efficiencies ranging from 94%-96% for COD, 91%-100% for SO4(2-) and S(2-), 92%-94% for BOD, 56%-82% for total Nitrogen and 2%-90% for NH3-N. 16S rRNA gene clone libraries were constructed and microbial community assemblies were determined by analysis of a total of 801 unique clone sequences from all the sites. Operational Taxonomic Unit (OTU)--based analysis of the sequences revealed highly diverse communities in each of the reactors and the constructed wetland. A total of 32 phylotypes were identified with the dominant members affiliated to Clostridia (33%), Betaproteobacteria (10%), Bacteroidia (10%), Deltaproteobacteria (9%) and Gammaproteobacteria (6%). Sequences affiliated to the class Clostridia were the most abundant across all sites. The 801 sequences were assigned to 255 OTUs, of which 3 OTUs were shared among the clone libraries from all sites. The shared OTUs comprised 80 sequences belonging to Clostridiales Family XIII Incertae Sedis, Bacteroidetes and unclassified bacterial group. Significantly different communities were harbored by the anaerobic, aerobic and rhizosphere sites of the constructed wetland. Numerous representative genera of the dominant bacterial classes obtained from the different sample sites of the integrated system have been implicated in the removal of various carbon- containing pollutants of natural and synthetic origins. To our knowledge, this is the first report of microbial community structure in tannery wastewater treatment plant from Ethiopia.

In this study, swine fecal specimens (n = 251) collected from nursing and weaned piglets raised under smallholder production systems were screened for the presence of kobuviruses by RT-PCR. Porcine kobuviruses were detected in 13.1 % (33/251) of the samples. We demonstrated that porcine kobuvirus infections exist in indigenous pigs in Kenya and Uganda and that the prevalence was higher in young piglets than older pigs: nursing piglets (15 %), post-weaning (3-month-old) pigs (17 %), 4-month-old pigs (10 %). Genetic analysis of the partial RNA-dependent RNA polymerase (RdRp) region (690 nt) revealed that kobuviruses circulating in East Africa are diverse, sharing nucleotide sequence identities ranging from 89.7 to 99.1 % and 88 to 92.3 % among them and with known porcine kobuviruses, respectively. The nucleotide sequence identities between our kobuvirus strains and those of human, bovine and canine kobuviruses were 69.4-70.7 %, 73.1-74.4 % and 67-70.7 %, respectively. Additionally, upon sequencing selected samples that showed consistent 720-bp RT-PCR bands while using the same primer set, we detected porcine astroviruses in our samples belonging to type 2 and type 3 mamastroviruses. To our knowledge, this study reports the first detection and molecular analysis of both porcine kobuviruses and astroviruses in an African region. Further studies are required to determine the role of these viruses in gastrointestinal infections of pigs in this region and to determine the genetic diversity of the circulating strains to develop accurate diagnostic tools and implement appropriate control strategies.

As Cameroon scales up its national HIV/AIDS control program, evaluating the performance of commercially available tests for accurate and cost effective diagnostics becomes essential. A cross-sectional study assessed the performance of an HIV oral rapid test. A total of 1520 participants consented to participate in the study. After counselling, they were tested for HIV using the national algorithm followed by OraQuick. Results of the national algorithm were compared to those of OraQuick, for sensitivity, specificity, positive predictive and negative predictive values. 62% of participants were male, and 1% was reported HIV-positive following the national algorithm. The OraQuick test had 93% sensitivity, 99% specificity, 99.93% NPV and 90% PPV (95% CI, Kappa 0.965). Though more expensive (2-6x) compared to the national algorithm tests, oral mucosal transudate-based test demonstrated good performance. Therefore, it could be implemented in resource-constrained settings if subsidized and could increase participation since less invasive with no blood accident exposure.

Infection by human rhinovirus (HRV) is a major cause of upper and lower respiratory tract disease worldwide and displays considerable phenotypic variation. We examined diversity by completing the genome sequences for all known serotypes (n = 99). Superimposition of capsid crystal structure and optimal-energy RNA configurations established alignments and phylogeny. These revealed conserved motifs; clade-specific diversity, including a potential newly identified species (HRV-D); mutations in field isolates; and recombination. In analogy with poliovirus, a hypervariable 5' untranslated region tract may affect virulence. A configuration consistent with nonscanning internal ribosome entry was found in all HRVs and may account for rapid translation. The data density from complete sequences of the reference HRVs provided high resolution for this degree of modeling and serves as a platform for full genome-based epidemiologic studies and antiviral or vaccine development.

A comparison of gene content and genome architecture of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major, three related pathogens with different life cycles and disease pathology, revealed a conserved core proteome of about 6200 genes in large syntenic polycistronic gene clusters. Many species-specific genes, especially large surface antigen families, occur at nonsyntenic chromosome-internal and subtelomeric regions. Retroelements, structural RNAs, and gene family expansion are often associated with syntenic discontinuities that-along with gene divergence, acquisition and loss, and rearrangement within the syntenic regions-have shaped the genomes of each parasite. Contrary to recent reports, our analyses reveal no evidence that these species are descended from an ancestor that contained a photosynthetic endosymbiont.

African trypanosomes cause human sleeping sickness and livestock trypanosomiasis in sub-Saharan Africa. We present the sequence and analysis of the 11 megabase-sized chromosomes of Trypanosoma brucei. The 26-megabase genome contains 9068 predicted genes, including approximately 900 pseudogenes and approximately 1700 T. brucei-specific genes. Large subtelomeric arrays contain an archive of 806 variant surface glycoprotein (VSG) genes used by the parasite to evade the mammalian immune system. Most VSG genes are pseudogenes, which may be used to generate expressed mosaic genes by ectopic recombination. Comparisons of the cytoskeleton and endocytic trafficking systems with those of humans and other eukaryotic organisms reveal major differences. A comparison of metabolic pathways encoded by the genomes of T. brucei, T. cruzi, and Leishmania major reveals the least overall metabolic capability in T. brucei and the greatest in L. major. Horizontal transfer of genes of bacterial origin has contributed to some of the metabolic differences in these parasites, and a number of novel potential drug targets have been identified.

Trypanosoma brucei, a flagellate protozoa of the family Trypanosomatidae, has become one of the model systems for unicellular pathogens to study fundamentally important biological phenomena. Currently, the method of choice to examine gene function in these organisms is RNA interference (RNAi). mRNA degradation is triggered by double-stranded RNA (dsRNA) produced in vivo from transgenes transcribed from opposing tetracycline (tet)-inducible T7 RNA polymerase promoters, or hairpin RNA transcribed from the tet-inducible procyclic acidic repetitive protein promoter. In this chapter, we describe some of the methods we employ for ablation of gene expression by RNAi in T. brucei with particular emphasis on transfection and cloning of procyclic cells, induction of dsRNA expression, isolation of RNA and analysis of dsRNA, and target mRNA.

In animals and protozoa gene-specific double-stranded RNA triggers the degradation of homologous cellular RNAs, the phenomenon of RNA interference (RNAi). RNAi has been shown to represent a novel paradigm in eukaryotic biology and a powerful method for studying gene function. Here we discuss RNAi in terms of its mechanism, its relationship to other post-transcriptional gene silencing phenomena in plants and fungi, its connection to retroposon silencing and possibly to translation, and its biological role. Among the organisms where RNAi has been demonstrated the protozoan parasite Trypanosoma brucei represents the most ancient branch of the eukaryotic lineage. We provide a synopsis of what is currently known about RNAi in T. brucei and outline the recent advances that make RNAi the method of choice to disrupt gene function in these organisms.