The quaternary structure of the amidase from Geobacillus pallidus RAPc8 is revealed by its crystal packing

The amidase from Geobacillus pallidus RAPc8, a moderate thermophile, is a member of the nitrilase enzyme superfamily. It converts amides to the corresponding acids and ammonia and has application as an industrial catalyst. RAPc8 amidase has been cloned and functionally expressed in Escherichia coli and has been purified by heat treatment and a number of chromatographic steps. The enzyme was crystallized using the hanging-drop vapour-diffusion method. Crystals produced in the presence of 1.2 M sodium citrate, 400 mM NaCl, 100 mM sodium acetate pH 5.6 were selected for X-ray diffraction studies. A data set having acceptable statistics to 1.96 Å resolution was collected under cryoconditions using an in-house X-ray source. The space group was determined to be primitive cubic P4232, with unit-cell parameter a = 130.49 (±0.05) Å. The structure was solved by molecular replacement using the backbone of the hypothetical protein PH0642 from Pyrococcus horikoshii (PDB code 1j31 ) with all non-identical side chains substituted with alanine as a probe. There is one subunit per asymmetric unit. The subunits are packed as trimers of dimers with D3 point-group symmetry around the threefold axis in such a way that the dimer interface seen in the homologues is preserved

GENETIC DIVERSITY AND ECO-GEOGRAPHICAL DISTRIBUTION OF ELEUSINE SPECIES COLLECTED FROM ETHIOPIA

Eleusine is a small grass genus with three basic chromosome numbers (x=8, 9 and 10) and comprises of eight species including E. coracana subsp. coracana, (finger millet), which is an important subsistence crop in Africa and India. Research on these species could assist the development of high yielding and multiple stress tolerant variety(s) of the cultivable species, and also guide development of strategic genetic resource management and utilisation of the genus. A total of 72 accessions, sampled from five major species, E. coracana (including both E. coracana subsp. coracana and E. coracana subsp. africana), E. intermedia, E. indica, E. multiflora and E. floccifolia were analysed for genetic variation and inter-relationships using 20 microsatellite markers. All the SSR markers displayed high genetic polymorphism, with polymorphic information content ranging from 0.46 (UGEP110) to 0.91 (UGEP66). A total of 286 alleles were observed with an average of 14.3 alleles per locus. Classic F-statistics revealed the highest intra-specific polymorphism recorded for E. africana (32.45%), followed by E. coracana (16.83%); implying that genetic polymorphism is higher in the cultivable subspecies and its wild relatives, than the other species. Allelic frequency based inter-species genetic distance analysis, showed wider genetic distance between E. indica and E. multiflora (0.719); a narrow genetic distance between E. coracana sub-species africana and E. coracana subspecies coracana (0.3297). The weighted neighbor joining-based clustering revealed that the majority of the accessions in a species share strong similarity and are grouped together than do accessions of inter species.Eleusine est une herbe avec trois nombres de chromosomes de base (x=8, 9 et 10) et comprend huit esp\ue8ces dont E. coracana subsp. coracana, (finger millet), qui est une culture de subsistence importante en Afrique et en Inde. La recherche sur ces esp\ue8ces pourrait aider dans le d\ue9veloppement des vari\ue9t\ue9s d\u2019esp\ue8ces cultivables \ue0 rendement \ue9lev\ue9 et de tol\ue9rance aux stress multiples, et guider le d\ue9veloppement de la gestion des ressources g\ue9n\ue9tiques strat\ue9giques et l\u2019utilisation du genus. Un total de 72 accessions \ue9chantillonn\ue9es de cinq esp\ue8ces majeurs \ue0 savoir E. coracana (incluant E. coracana subsp. coracana et E. coracana subsp. africana), E. intermedia, E. indica, E. multiflora et E. floccifolia \ue9taient analys\ue9es pour variation g\ue9n\ue9tique et relations mutuelles utilisant 20 marqueurs microsatellites. Tous les marquers SSR ont manifest\ue9 un polymorphisme g\ue9n\ue9tique \ue9lev\ue9, avec un contenu d\u2019information polymorphique allant de 0.46 (UGEP110) \ue0 0.91 (UGEP66). Un total de 286 all\ue8les \ue9tait observ\ue9 avec une moyenne de 14.3 all\ue8les par locus. Les statistiqies classiques F ont r\ue9v\ue9l\ue9 le polymorphisme intrasp\ue9cifique le plus \ue9lev\ue9 enregistr\ue9 pour le E. africana (32.45%), suivi de E. coracana (16.83%), ce qui implique que le polymorphisme g\ue9n\ue9tique est le plus \ue9lev\ue9 dans les sous esp\ue8ces cultivables et ses homologues sauvages que les autres esp\ue8ces. L\u2019analyse de la fr\ue9quence all\ue9lique de la distance g\ue9n\ue9tique entre esp\ue8ces a montr\ue9 une plus large distance g\ue9n\ue9tique entre E. indica et E. multiflora (0.719); une \ue9troite distance g\ue9n\ue9tique entre les sous esp\ue8ces Africana de E. coracana et les sous esp\ue8ces coracana de E. coracana (0.3297). La pond\ue9ration des groupements a r\ue9v\ue9l\ue9 que la majorit\ue9 des accessions au sein d\u2019une esp\ue8ce partage une forte similarit\ue9 et sont group\ue9es ensemble en comparaison aux accessions des intra-esp\ue8ces

Rift Valley fever in Kenyan pastoral livestock: Individual-based demographic model to analyse the impact of Rift Valley fever

Background Rift Valley Fever (RVF) is a viral zoonosis and a mosquito-borne disease caused by a phlebovirus in the family Bunyaviridae. It affects livestock, humans and wildlife. Epidemic outbreaks of RVF in East Africa, which occur after heavy rainfalls in cycles of 5-15 years, have caused next to human morbidity and mortality considerable economic losses throughout the livestock production and market chain. Objective Establishment a pastoral livestock demographic model to simulate 10 year alternating normal and drought periods and RVF epidemics. Methods We developed an individual-based C++ language with Borland C++ builder 6 model, to simulate livestock dynamics in North Eastern-Province during normal and drought periods, tracked over days and years. During RVF epidemics and with different control measures, animals were stratified into susceptible, exposed, infectious and recovered. The following scenarios were modeled (i) the demographic dynamics of cattle, camels, sheep and goats; (ii) an RVF outbreak in livestock and (iii) impacts of control measures (combinations of vaccination, sanitary measures, surveillance and vector control). Results/Conclusions Sheep and goat populations increase fastest (9-23%) annually during normal years while cattle and sheep populations show fastest decline during drought. In infected areas, mainly sheep (59%) are infected followed by goats (44%), cattle (31%), and camels (5%). Sheep and goats are most likely to spread the RVF through livestock trade. Slaughtered infected sheep are an important risk factor to human RVF infection. After the 2006/2007 outbreak, 2%, 40%, 30% and less than 1% of cattle, sheep, goats and camels acquired immunity. After seven years, only 4%, of sheep and goats remain immune. Our results will assist in the assessment of cost-benefit and cost-effectiveness of interventions which should improve future intersectoral livestock – public health contingency planning

Including PrEP for key populations in combination HIV prevention: a mathematical modelling analysis of Nairobi as a case-study

Background: The role of PrEP in combination HIV prevention remains uncertain. We aimed to identify an optimal portfolio of interventions to reduce HIV incidence for a given budget, and to identify the circumstances in which PrEP could be used in Nairobi, Kenya. Methods: A mathematical model was developed to represent HIV transmission among specific key populations (female sex workers (FSW), male sex workers (MSW), and men who have sex with men (MSM)) and among the wider population of Nairobi. The scale-up of existing interventions (condom promotion, anti-retroviral therapy (ART) and male circumcision) for key populations and the wider population as have occurred in Nairobi is represented. The model includes a detailed representation of a Pre-Exposure Prophylaxis (PrEP) intervention and is calibrated to prevalence and incidence estimates specific to key populations and the wider population. Findings: In the context of a declining epidemic overall but with a large sub-epidemic among MSM and MSW, an optimal prevention portfolio for Nairobi should focus on condom promotion for MSW and MSM in particular, followed by improved ART retention, earlier ART, and male circumcision as the budget allows. PrEP for MSW could enter an optimal portfolio at similar levels of spending to when earlier ART is included, however PrEP for MSM and FSW would be included only at much higher budgets. If PrEP for MSW cost as much $500, average annual spending on the interventions modelled would need to be less than$3·27 million for PrEP for MSW to be excluded from an optimal portfolio. Estimated costs per infection averted when providing PrEP to all FSW regardless of their risk of infection, and to high risk FSW only, are $65,160 (95$43,520 - $90,250) and$10,920 (95% credible interval: $4,700 -$51,560) respectively. Interpretation: PrEP could be a useful contribution to combination prevention, especially for underserved key populations in Nairobi. An ongoing demonstration project will provide important information regarding practical aspects of implementing PrEP for key populations in this setting

Natural Products-Based Drug Design against SARS-CoV-2 Mpro 3CLpro

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has received global attention due to the serious threat it poses to public health. Since the outbreak in December 2019, millions of people have been affected and its rapid global spread has led to an upsurge in the search for treatment. To discover hit compounds that can be used alone or in combination with repositioned drugs, we first analyzed the pharmacokinetic and toxicological properties of natural products from Brazil’s semiarid region. After, we analyzed the site prediction and druggability of the SARS-CoV-2 main protease (Mpro), followed by docking and molecular dynamics simulation. The best SARS-CoV-2 Mpro complexes revealed that other sites were accessed, confirming that our approach could be employed as a suitable starting protocol for ligand prioritization, reinforcing the importance of catalytic cysteine-histidine residues and providing new structural data that could increase the antiviral development mainly against SARSCoV-2. Here, we selected 10 molecules that could be in vitro assayed in response to COVID-19. Two compounds (b01 and b02) suggest a better potential for interaction with SARS-CoV-2 Mpro and could be further studied.Research Dean and Graduate Studies of the Federal University of Pará (PROPESP/UFPA)Brazilian National Council for Scientific and Technological Development (CNPq)Brazilian Coordination for Improvement of Personnel Higher Education (CAPES)Bahia Research Foundation (FAPESB, grant numbers APP071/2011, JCB-0039/2013, and RED-008/2013

SENSITIVITY OF MICROSCOPY COMPARED TO MOLECULAR DIAGNOSIS OF P. FALCIPARUM: IMPLICATIONS ON MALARIA TREATMENT IN EPIDEMIC AREAS IN KENYA

Detection of Plasmodium species by microscopy has been the gold standard for diagnosis of malaria for more than a century. Despite the fact that there is a significant decline in the number of positive cases reported from microscopy, anti-malarial drugs prescriptions are on continuous increase as patients present with symptoms of malaria. This makes it difficult to establish accuracy, sensitivity and specificity of light microscopy in diagnosis of malaria in epidemic areas. This study was designed to compare microscopy with polymerase chain reaction as diagnostic methods for malaria in three epidemic areas in Kenya. A total of 356 patients presenting with malaria symptoms were diagnosed by microscopy and dried blood filter paper spots were collected from patient in Kisii, West Pokot and Narok districts. Plasmodium falciparum DNA was extracted from the dried blood filter samples. Primers specific for the Plasmodium Species were designed and used in a two step amplification of the Pfmdr gene. The PCR products were analyzed in ethidium bromide stained 1.5% agarose gel. It was found that 72 out of 350 specimens diagnosed as negative were positive for P. falciparum by nested PCR, while 6 which were microscopy positive were confirmed so by nested PCR. This study demonstrates that there is a high level of misdiagnosis which may either lead to denial for deserved treatment or undeserved treatment. Nested PCR detection of malaria parasites is a very useful complement to microscopy although it is expensive and takes long time. Additionally, smear negative patients suspected to have malaria should be subjected to PCR diagnosis to improve rational drug use. The economic burden of misdiagnosis and mistreatment of malaria outweighs that of PCR diagnosis, hence this diagnostic mode could be tenable in the long run even in rural areas

Effect of Baseline HIV Disease Parameters on CD4+ T Cell Recovery After Antiretroviral Therapy Initiation in Kenyan Women

Antiretroviral therapy (ART) for HIV infection reconstitutes the immune system and improves survival. However, the rate and extent of CD4+ T cell recovery varies widely. We assessed the impact of several factors on immune reconstitution in a large Kenyan cohort.HIV-infected female sex workers from a longitudinal cohort, with at least 1 year of pre-ART and 6 months of post-ART follow-up (n = 79), were enrolled in the current study. The median pre-ART follow-up was 4,040 days. CD4 counts were measured biannually and viral loads where available. The median CD4 count at ART initiation was 180 cells/ul, which increased to 339 cells/ul at the most recent study visit. The rate of CD4+ T cell increase on ART was 7.91 cells/month (mean = 13, range -25.92 to 169.4). LTNP status prior to ART initiation did not associate with the rate of CD4 recovery on ART. In univariate analyses, associations were observed for CD4 recovery rate and duration of pre-ART immunosuppression (r = -0.326, p = 0.004) and CD4 nadir (r = 0.284, p = 0.012). In multivariate analysis including age, CD4 nadir, duration of HIV infection, duration of pre-ART immunosuppression, and baseline viral load, only CD4 nadir (p = 0.007) and not duration of immunosuppression (p = 0.87) remained significantly associated with the rate of CD4 recovery.These data suggest that prior duration of immune suppression does not predict subsequent recovery once ART is initiated and confirm the previous observation that the degree of CD4 depletion prior to ART initiation is the most important determinant of subsequent immune reconstitution

Comparison of baseline lymphoma and HIV characteristics in Malawi before and after implementation of universal antiretroviral therapy

Access to antiretroviral therapy (ART) led to epidemiological changes in human immunodeficiency virus (HIV) associated lymphoma in high-income countries such as reductions in diffuse large B-cell lymphoma (DLBCL) and stable or increased Hodgkin lymphoma (HL) and Burkitt lymphoma (BL). In 2016, Malawi implemented a universal ART (UART) policy, expanding ART eligibility to all persons living with HIV (PLWH). We compare the distribution of lymphoma subtypes and baseline HIV and prognostic characteristics for lymphoma patients in Malawi before and after implementation of UART. We enrolled patients with pathologically confirmed incident lymphoproliferative disorders into a observational clinical cohort. At diagnosis, a comprehensive clinicopathological evaluation was performed. Of 412 participants, 156 (38%) were pre-UART (2013-June 2016) and 256 (62%) post-UART (July 2016-2020). HIV prevalence was 50% in both groups. The most common pre-UART diagnoses were DLBCL [75 (48%)], low-grade non-Hodgkin lymphoma (NHL) [19 (12%)], HL [17 (11%)] and, BL [13 (8%)]. For post-UART they were DLBCL [111 (43%)], NHL [28 (11%)], BL [27 11%)] and, HL [20 (8%)]. Among PLWH, 44 (57%) pre-UART initiated ART prior to lymphoma diagnosis compared to 99 (78%) post-UART (p = 0.02). HIV-ribonucleic acid was suppressed <1000 copies/mL in 56% (33/59) pre-UART and 71% (73/103) post-UART (p = 0.05). CD4 T-cell counts were similar for both groups. We observed similar findings in the subset of participants with DLBCL. Overall, there were no significant changes in incident lymphoma subtypes (p = 0.61) after implementation of UART, but HIV was better controlled. Emerging trends bear monitoring and may have implications for prognosis and health system priority setting

methods and tools for the development of adaptive applications

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