Molecular Characterization of Circulating Yellow Fever Viruses from Outbreak in Ghana, 2021-2022.

Yellow fever virus, transmitted by infected Aedes spp. mosquitoes, causes an acute viral hemorrhagic disease. During October 2021-February 2022, a yellow fever outbreak in some communities in Ghana resulted in 70 confirmed cases with 35 deaths (case-fatality rate 50%). The outbreak started in a predominantly unvaccinated nomadic community in the Savannah region, from which 65% of the cases came. The molecular amplification methods we used for diagnosis produced full-length DNA sequences from 3 confirmed cases. Phylogenetic analysis characterized the 3 sequences within West Africa genotype II; strains shared a close homology with sequences from Cote d'Ivoire and Senegal. We deployed more sensitive advanced molecular diagnostic techniques, which enabled earlier detection, helped control spread, and improved case management. We urge increased efforts from health authorities to vaccinate vulnerable groups in difficult-to-access areas and to educate the population about potential risks for yellow fever infections.

SARS-CoV-2 Molecular Evolutionary Dynamics in the Greater Accra Region, Ghana.

To assess dynamics of SARS-CoV-2 in Greater Accra Region, Ghana, we analyzed SARS-CoV-2 genomic sequences from persons in the community and returning from international travel. The Accra Metropolitan District was a major origin of virus spread to other districts and should be a primary focus for interventions against future infectious disease outbreaks.

Hepatitis C virus (HCV) infection among patients with sickle cell disease at the Korle-Bu teaching hospital.

BACKGROUND: Hepatitis C virus (HCV) infection is a blood borne infection that remains potentially transmissible through blood transfusions. Sickle cell disease (SCD) is a common inheritable haemoglobinopathy in Ghana that requires multiple blood transfusions as part of its management. The SCD patient is therefore at a high risk of HCV infection; however, data on the occurrence of HCV in SCD patients has not been documented in Ghana. This study sought to determine the prevalence and genotypes of HCV infection in SCD patients. MATERIALS AND METHODS: This was a cross-sectional study which enrolled 141 sickle-cell disease patients from the Ghana Institute for Clinical Genetics, Korle-Bu Teaching Hospital (KBTH). Patient information was obtained through a structured questionnaire. Aliquots of the plasma obtained was used for both serology with Advanced Quality Rapid Anti-HCV Test Strip and molecular testing by RT-PCR with primers targeting the HCV core gene. The amplified DNA were purified and subjected to phylogenetic analysis to characterize HCV genotypes. RESULTS: Twelve (9%) out of the 141 patients were sero-positive for HCV total antibodies. HCV RNA was amplified from 8 (6%) out of the total number of patients' samples. One of the 12 sero-positives was HCV RNA positive. Five (63%) out of the 8 HCV RNA positive samples were successfully sequenced. The phylogenetic tree constructed with the study and GenBank reference sequences, clustered all five study sequences into HCV genotype 1. CONCLUSION: The HCV seroprevalence of 9% among sickle cell disease patients is higher than reported for the general Ghanaian population which is 3%. Genotype 1 is the common HCV genotype infecting SCD patients. Sickle cell disease is likely to be a high-risk group for HCV inapparent infections in Ghana as seroprevalence does not correlate with viremia. However, even with higher seroprevalence, the group must be given priority in resource allocation for preventive, diagnostic and therapeutic strategies.

First Molecular Detection of SARS-CoV-2 in Sewage and Wastewater in Ghana.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is shed in the stool of infected individuals and can be detected in sewage and wastewater contaminated with infected stool. This study is aimed at detecting the virus and its potential survival in sewage and wastewater in Ghana. The cross-sectional study included samples from 16 validated environmental surveillance sites in 7 regions of Ghana. A total of 354 samples composed of wastewater (280) and sewage (74) were collected from November 2020 to November 2022. Overall, 17% of the samples were positive for SARS-CoV-2 by real-time PCR, with 6% in sewage and 11% in wastewater. The highest number of positive samples was collected from the Greater Accra Region (7.3%) with the least recorded in the Bono East Region (0.6%). Further characterization of the positive samples using the next-generation sequencing (NGS) approach yielded two variants: Alpha (B.1.1.7) and Delta (AY.36). Attempts to isolate SARS-CoV-2 in the Vero cell line were not successful probably due to the low viral load concentrations (Ct values > 35) or prolonged exposure to high temperatures rendering the virus noninfectious. Our findings suggest that SARS-CoV-2 RNA in sewage and wastewater may not be infectious, but the prevalence shows that the virus persists in the communities within Ghana.

Unsaturated fatty acid perturbation combats emerging triazole antifungal resistance in the human fungal pathogen Aspergillus fumigatus.

Contemporary antifungal therapies utilized to treat filamentous fungal infections are inhibited by intrinsic and emerging drug resistance. Consequently, there is an urgent need to develop novel antifungal compounds that are effective against drug-resistant filamentous fungi. Here, we utilized an Aspergillus fumigatus cell-based high-throughput screen to identify small molecules with antifungal activity that also potentiated triazole activity. The screen identified 16 hits with promising activity against A. fumigatus. A nonspirocyclic piperidine, herein named MBX-7591, exhibited synergy with triazole antifungal drugs and activity against pan-azole-resistant A. fumigatus isolates. MBX-7591 has additional potent activity against Rhizopus species and CO2-dependent activity against Cryptococcus neoformans. Chemical, genetic, and biochemical mode of action analyses revealed that MBX-7591 increases cell membrane saturation by decreasing oleic acid content. MBX-7591 has low toxicity in vivo and shows good efficacy in decreasing fungal burden in a murine model of invasive pulmonary aspergillosis. Taken together, our results suggest MBX-7591 is a promising hit with a novel mode of action for further antifungal drug development to combat the rising incidence of triazole-resistant filamentous fungal infections.IMPORTANCEThe incidence of infections caused by fungi continues to increase with advances in medical therapies. Unfortunately, antifungal drug development has not kept pace with the incidence and importance of fungal infections, with only three major classes of antifungal drugs currently available for use in the clinic. Filamentous fungi, also called molds, are particularly recalcitrant to contemporary antifungal therapies. Here, a recently developed Aspergillus fumigatus cell reporter strain was utilized to conduct a high-throughput screen to identify small molecules with antifungal activity. An emphasis was placed on small molecules that potentiated the activity of contemporary triazole antifungals and led to the discovery of MBX-7591. MBX-7591 potentiates triazole activity against drug-resistant molds such as A. fumigatus and has activity against Mucorales fungi. MBX-7591's mode of action involves inhibiting the conversion of saturated to unsaturated fatty acids, thereby impacting fungal membrane integrity. MBX-7591 is a novel small molecule with antifungal activity poised for lead development.

Occurrence of influenza and bacterial infections in cancer patients receiving radiotherapy in Ghana.

BACKGROUND: Influenza co-infection with bacteria is a leading cause of influenza-related deaths and severe respiratory infections, especially among high-risk groups like cancer patients undergoing treatment. However, acute respiratory infection (ARI)-like symptoms developed by upper-torso cancer (UTC) patients receiving radiotherapy are considered as side-effects of the radiation. Hence influenza and bacterial pathogens implicated in ARI are not investigated. METHODS: This prospective cohort study examined 85 in-patients with upper-torso cancers undergoing radiotherapy at the National Radiotherapy, Oncology and Nuclear Medicine Centre (NRONMC) of Korle-Bu Teaching Hospital (KBTH) in Accra, Ghana. Eligible patients who consented were recruited into the study from September 2018 to April 2019. Influenza viruses A and B in addition to the following bacteria species Streptococcus pneumonia, Haemophilus influenzae, Neisseria meningitidis and Staphylococcus aureus were detected from oropharyngeal and nasopharyngeal swab specimens collected at three different time points. Presence of respiratory pathogens were investigated by influenza virus isolation in cell culture, bacterial culture, polymerase chain reaction (PCR) and next generation sequencing (NGS) assays. RESULTS: Of the 85 eligible participants enrolled into the study, 87% were females. Participants were 17 to 77 years old, with a median age of 49 years. Most of the participants (88%) enrolled had at least one pathogen present. The most prevalent pathogen was N. meningitidis (63.4%), followed by H. influenzae (48.8%), Influenza viruses A and B (32.9%), S. pneumoniae (32.9%) and S. aureus (12.2%). Approximately, 65% of these participants developed ARI-like symptoms. Participants with previous episodes of ARI, did not live alone, HNC and total radiation less than 50 Gy were significantly associated with ARI. All treatment forms were also significantly associated with ARI. CONCLUSION: Data generated from the study suggests that ARI-like symptoms observed among UTC patients receiving radiotherapy in Ghana, could be due to influenza and bacterial single and co-infections in addition to risk factors and not solely the side-effects of radiation as perceived. These findings will be prime importance for diagnosis, prevention, treatment and control for cancer patients who present with such episodes during treatment.

Circulation of hepatitis delta virus and occult hepatitis B virus infection amongst HIV/HBV co-infected patients in Korle-Bu, Ghana.

BACKGROUND: Within HIV/HBV infected patients, an increase in HDV infection has been observed; there is inadequate information on HDV prevalence as well as virologic profile in Ghana. This study sought to determine the presence of HDV in HIV/HBV co-infected patients in Ghana. METHODS: This was a longitudinal purposive study which enrolled 113 HIV/HBV co-infected patients attending clinic at Korle-Bu Teaching Hospital (KBTH) in Accra, Ghana. After consenting, 5 mL whole blood was collected at two-time points (baseline and 4-6 months afterwards). The sera obtained were tested to confirm the presence of HIV, HBV antibodies and/or antigens, and HBV DNA. Antibodies and viral RNA were also determined for HDV. Amplified HBV DNA and HDV RNA were sequenced and phylogenetic analysis carried out with reference sequences from the GenBank to establish the genotypes. RESULTS: Of the 113 samples tested 63 (55.7%) were females and 50 (44.25%) were males with a median age of 45 years. A total of 100 (88.5%) samples had detectable HBV surface antigen (HBsAg), and 32 out of the 113 had detectable HBV DNA. Nucleotide sequences were obtained for 15 and 2 samples of HBV and HDV, respectively. Phylogenetic analysis was predominantly genotype E for the HBVs and genotype 1 for the HDVs. Of the 13 samples that were HBsAg unreactive, 4 (30.8%) had detectable HBV DNA suggesting the incidence of occult HBV infections. The percentage occurrence of HDV in this study was observed to be 3.54. CONCLUSION: Our data suggest the presence and circulation of HDV and incidence of occult HBV infection in HIV/HBV co-infected patients in Ghana. This informs health staff and makes it imperative to look out for the presence of HDV and occult HBV in HIV/HBV co-infected patients presenting with potential risk of liver cancers and HBV transmission through haemodialysis and blood transfusions.

Detection of vaccine-derived poliovirus circulation by environmental surveillance in the absence of clinical cases.

BACKGROUND: On August 25, 2019, the Noguchi Memorial Institute for Medical Research notified the confirmation of a circulating-vaccine-derived poliovirus type-2 (cVDPV2) from the Agbogbloshie environmental surveillance (AES) site, in the Greater Accra Region. A field investigation of the outbreak was conducted to describe the results of epidemiological and laboratory investigations, and control efforts. METHODS: We conducted a descriptive investigation, records review, and active-case-search. Caregivers were interviewed on the vaccination status of their children; knowledge, attitude, and practices on polio prevention; water, sanitation and hygiene practices, and health-seeking behaviors. Stool from healthy children <5 y and sewage samples were taken for laboratory diagnosis. RESULTS: cVDPV2 genetically similar to the cVDPV2 diagnosed recently in the Northern Region of Ghana and Nigeria was identified. 2019 half-year coverage of OPV and IPV was 22%. Fully immunized children were 49% (29/59). Most health workers (70%) had a fair knowledge of polio and acute flaccid paralysis (AFP). Forty-six percent of care-givers admitted to using the large drain linked to the site where the cVDPV2 was isolated as their place of convenience and disposing of the fecal matter of their children. No AFP case was identified. Stool samples from 40 healthy children yielded non-polio enteroviruses while 75% (3/4) of the additional sewage samples yielded cVDPV2. CONCLUSION: cVDPV2 was isolated from the AES site. No AFP or poliovirus was identified from healthy children. There is a need to improve health workers' knowledge on AFP and to address the dire sanitation conditions in the Agbogbloshie market and its environs.

Strengthening laboratory surveillance of viral pathogens: Experiences and lessons learned building next-generation sequencing capacity in Ghana.

OBJECTIVE: To demonstrate the feasibility of applying next-generation sequencing (NGS) in medium-resource reference laboratories in Africa to enhance global disease surveillance. METHODS: A training program was developed to support implementation of NGS at Noguchi Memorial Institute for Medical Research (NMIMR), University of Ghana. The program was divided into two training stages, first at the Centers for Disease Control and Prevention (CDC) in Atlanta, GA, followed by on-site training at NMIMR for a larger cohort of scientists. RESULTS: Self-assessment scores for topics covered during the NGS training program were higher post-training relative to pre-training. During the NGS Training II session at NMIMR, six enterovirus isolates from acute flaccid paralysis cases in Ghana were successfully sequenced by trainees, including two echovirus 6, two echovirus 11 and one echovirus 13. Another genome was an uncommon type (EV-B84), which has not been reported in Africa since its initial discovery from a Côte d'Ivoire specimen in 2003. CONCLUSIONS: The success at NMIMR provides an example of how to approach transferring of NGS methods to international laboratories. There is great opportunity for collaboration between institutes that have genomics expertise to ensure effectiveness and long-term success of global NGS capacity building programs.

Expanding molecular diagnostics of helminthiasis: Piloting use of the GPLN platform for surveillance of soil transmitted helminthiasis and schistosomiasis in Ghana.

The efforts to control and eradicate polio as a global health burden have been successful to the point where currently only three countries now report endemic polio, and the number of cases of polio continues to decrease. The success of the polio programme has been dependant on a well-developed network of laboratories termed the global polio laboratory network (GPLN). Here we explore collaborative opportunities with the GPLN to target two of the 18 diseases listed as a neglected tropical diseases (NTD) namely soil transmitted helminthiasis (STH) and Schistosomiasis (SCH). These were chosen based on prevalence and the use of faecal materials to identify both polio, STH and SCH. Our study screened 448 faecal samples from the Ghana GPLN using three triplex TaqMan assays to identify Ascaris lumbricoides, Necator americanus, Ancylostoma spp, Trichuris trchiura, Strongyloides stercoralis and Schistosoma spp. Our results found a combined helminth prevalence of 22%. The most common helminth infection was A. lumbricoides with a prevalence of 15% followed by N. americanus (5%), Ancylostoma spp. (2.5%), Schistosoma spp. (1.6%) and S. stercoralis (1%). These results show that it is possible to identify alternative pathogens to polio in the samples collected by the GPLN platform and to introduce new diagnostic assays to their laboratories. The diagnostic methods employed were also able to identify S. stercoralis positive samples, which are difficult to identify using parasitological methods such as Kato-Katz. This study raises the possibility of collaboration with the GPLN for the surveillance of a wider range of diseases which would both benefit the efforts to control the NTDs and also increase the scope of the GPLN as a diagnostic platform.