Development of novel systems for bioconversion of cellulosic biomass to useful products

There is increasing concern regarding alternative, sustainable energy sources, such as biofuels, to replace declining oil reserves. The abundance of lignocellulosic biomass makes it the only imaginable resource that can potentially substitute a substantial portion of the fossil fuels we use today, but current methods for producing biofuels from non-food crops are cost intensive and not economically viable. Synthetic biology provides several potential approaches for developing biologically mediated processes for the conversion of lignocellulosic biomass into biofuels. Such systems are based on engineered microbes that produce enzymes for catalysing the conversion of cellulose into fermentable sugars and subsequently into high value products. Effective degradation of cellulose requires multiple classes of enzyme working together. In naturally occurring cellulose degrading microbes, bioconversion is catalysed by a battery of enzymes with different catalytic properties. However, naturally occurring cellulases with multiple catalytic domains seem to be rather rare in known cellulose-degrading organisms. Using synthetic biology approaches, seven cellulases with multiple catalytic domains were engineered and tested to determine the usefulness of such chimeric enzymes to replace cloning of multiple enzymes for biomass conversion. Catalytic domains were taken from Cellulomonas fimi endoglucanases CenA, CenB and CenD, exoglucanase Cex, and β-glucosidase, Cfbglu as well as Cytophaga hutchinsonii cellodextrinase CHU2268. All fusions retained both catalytic activities of the parental enzymes. To investigate the benefits of fusion, Citrobacter freundii NCIMB11490 was transformed with either fused or non-fused enzymes and cultured with cellulose blotting papers as main carbon source. Cells expressing fusions of Cex with CenA or CenD reproducibly showed higher growth than cells expressing non-fused versions, as well as more rapid physical destruction of paper. The opposite was observed for the other combinations. Comparing two different Cex and CenA fusions, CxnA2, which contains two carbohydrate binding modules (CBMs), degraded filter paper faster and led to better growth than CxnA1, which contains only one CBM. It was observed that CxnA1 was exported to the supernatant of E. coli and C. freundii cultures, as also seen for Cex and CenA, although there is no clear biological mechanism for this. Monitoring of growth using colony counts is laborious, but the use of optical density is not possible for cellulose-based cultures as it is affected by the insoluble cellulose particles. The SYBR Green I/propidium iodide live/dead staining protocol was therefore evaluated for growth measurements and was found to allow rapid measurements of large numbers of samples. In conclusion, these studies have demonstrated a simple and useful method for making chimeric proteins from libraries of multiple parts. The results demonstrate that use of fusion proteins can improve biomass conversion in vivo, and could potentially reduce the necessity for cloning of multiple enzymes and improve product yields. A simple and effective method for monitoring growth of bacteria in turbid cultures using a fluorimeter has also been developed

Low seroprevalence of hepatitis C among children at the Princess Marie Louis Children´s Hospital in Accra, Ghana

Hepatitis C is a leading cause of chronic hepatitis and causes severe health problems in areas where prevalence is high. Ghana is noted for a relatively high sero-prevalence of hepatitis C virus infection. However, there is very little data on prevalence of hepatitis C virus (HCV) among children in Ghana, and what data is available indicates very low prevalence rate. We conducted a cross-sectional study to determine the sero-prevalence and associated pre-disposing risk factor for HCV infection among children attending the Princes Marie Louis Children´s Hospital in Accra. Two hundred archived blood samples from a previous study were retrieved and tested for the presence of HCV antibodies using a dipstick test kit. Out of the 200 samples tested, one (1) tested positive for HCV antibodies giving a prevalence of 0.5% among the study group. The results show that there is potentially a very low prevalence of hepatitis C among Ghanaian children. Hence, the higher prevalence among adults usually seen is often due to infection later in life. Obtaining an appropriate vaccine early in life could thus help prevent people from getting infected in later life

Active tuberculosis case findings in Ghanaian health facilities: effectiveness and sensitivity of the symptoms-based screening tool

Introduction: the National Tuberculosis Programme (NTP), Ghana, introduced Symptoms-Based Screening (SBS) Tool for TB case finding. This study aimed to determine the challenges and limitations associated with the use of the SBS Tool for active tuberculosis case finding in Ghanaian health facility settings. Methods: this study targeted suspected TB patients attending two health facilities in the Ho Municipality, Ghana. Initially, suspected TB patients were screened with the SBS tool and presumptive patients subsequently tested for M. tuberculosis using microscopy and geneXpert assay. Additionally, health personnel were interviewed to assess the user-friendliness, challenges, and limitations associated with the tool. Results: of 636 presumptive TB patients identified, 1.73% had tuberculosis. Coughing for > 2 weeks (χ²=24.8; p<0.05); chest pain (χ²=28.3; p<0.01) and night sweat (χ²=34.8; p<0.05) associated significantly with M. tuberculosis infection status. The health personnel found the tool to be not user-friendly and it also lacked indicators to identify other vulnerable individuals such as diabetics, cigarette smokers, alcoholics, immunocompromised, and malnourished individuals. Therefore, the SBS tool was found not to be sensitive enough to identify probable cases. Conclusion: the SBS tool is useful for detecting active TB cases, however, it must be improved to identify vulnerable individuals such as diabetics, immunosuppressed, and malnourished

Occurrence and distribution of extended-spectrum β-lactamase in clinical Escherichia coli isolates at Ho Teaching Hospital in Ghana.

Objective: This study determined the occurrence and distribution of Extended Spectrum β-Lactamase (ESBL) genotypes of E. coli isolates in Ho Teaching Hospital, Ghana.Design: A cross-sectional study.Setting: A single centre study was conducted at Ho Teaching Hospital of Ghana.Participants: Patients who visited Ho Teaching Hospital Laboratory with the request for culture and susceptibility testing.Main outcome measure: Escherichia coli were isolated, and Extended-Spectrum β-Lactamase genes were detected.Results: Of the 135 isolates, 56(41.5%,95% CI: 33.1% – 50.3%) were ESBL producers. More males, 14(58.3%), produced ESBL than females, 42(37.8%). The ESBL prevalence was highest among the elderly who were 80 years and above 3(100.0%), with the least prevalence among patients within 50-59 years and 0-9 years age bracket, representing 4(25.0%) and 3(27.3%), respectively. The total prevalence of ESBL was marginally higher among out-patients (41.8% 95% CI: 31.9% - 52.2%) compared to in-patients [40.5% 95% CI: 24.8% - 57.9]. BlaTEM-1 was the predominant ESBL genotype obtained from 83.9% (47/56) of the confirmed ESBL producing isolates, with the least being TOHO-1 4(7.1%). The co-existence of 2 different ESBL genes occurred in 19(33.9%) of the isolates. The single and quadruple carriage were 16(28.6%) and 3(5.4%), respectively. The highest co-existence of the ESBL genotypes was recorded for blaTEM-1 and blaCTXM-1 15(26.8%), followed by blaTEM-1, blaCTXM-1 and blaSHV-73 [12(21.4%)].Conclusion: The high prevalence of ESBL-producing E. coli isolates with multiple resistant gene carriage is a threat to healthcare in the study area

Short-term treatment outcomes in human immunodeficiency virus type-1 and hepatitis B virus co-infections

BACKGROUND: Co-infection of HIV with HBV is common in West Africa but little information is available on the effects of HBV on short-term therapy for HIV patients. A 28 day longitudinal study was conducted to examine short-term antiretroviral therapy (ART) outcomes in HIV infected individuals with HBV co-infection. METHODS: Plasma from 18 HIV infected individuals co-infected with HBV and matched controls with only HIV infection were obtained at initiation, and 7 and 28 days after ART. HIV-1 viral load changes were monitored. Clinical and demographic data were also obtained from patient folders, and HIV-1 drug resistance mutation and subtype analysis performed. RESULTS: The presence of HBV co-infection did not significantly affect HIV-1 viral load changes within 7 or 28 days. The CD4(+) counts on the other hand of patients significantly affected the magnitude of HIV-1 viral load decline after 7 days (ρ = −0.441, p = 0.040), while the pre-ART HIV-1 VL (ρ = 0.844, p = <0.001) and sex (U = 19.0, p = 0.020) also determined HIV-1 viral load outcomes after 28 days of ART. Even though the geometric sensitivity score of HIV-1 strains were influenced by the HIV-1 subtypes (U = 56.00; p = 0.036), it was not a confounder for ART outcomes. CONCLUSIONS: There may be the need to consider the confounder effects of sex, pre-ART CD4(+), and pre-ART HIV-1 viral load in the discourse on HIV and HBV co-infection

The global inequity of COVID-19 diagnostics : challenges and opportunities

Diagnostics for COVID-19 have advanced at an unprecedented pace over the last two years. Testing is a critical pillar of pandemic control, and is required for epidemiological tracking, treatment, and surveillance. Despite high quality SARS-CoV-2 viral diagnostic capability, there are vast global inequities in access. The Virology, Immunology, and Diagnostics Working Group(WG) of the COVID-19 Clinical Research Coalition (CRC) brings together experts in immunology, infectious diseases, and microbiology to advocate for equity-based COVID-19 research, prioritising solutions driven by communities in low-income and lower middle-income countries (LMICs).1 This commentary shares the unique perspective of the WG on the asymmetry in COVID-19 diagnostic access between low-income and high-income settings, the barriers to these disparities, and highlights opportunities to remedy these inequities.PostprintPeer reviewe

Is malaria immunity a possible protection against severe symptoms and outcomes of COVID-19?

Malaria-endemic areas of the world are noted for high morbidity and mortality from malaria. Also noted in these areas is the majority of persons in the population having acquired malaria immunity. Though this acquired malaria immunity does not prevent infection, it resists the multiplication of Plasmodium parasites, restricting disease to merely uncomplicated cases or asymptomatic infections. Does this acquired malaria immunity in endemic areas protect against other diseases, especially outbreak diseases like COVID-19? Does malaria activation of innate immunity resulting in trained or tolerance immunity contribute to protection against COVID-19? In an attempt to answer these questions, this review highlights the components of malaria and viral immunity and explores possible links with immunity against COVID-19. With malaria-endemic areas of the world having a fair share of cases of COVID-19, it is important to direct research in this area to evaluate and harness any benefits of acquired malaria immunity to help mitigate the effects of COVID-19 and any possible future outbreaks. FUNDING: None declared