Evaluation of two inoculation routes of an adenovirus-mediated viral protein inhibitor in a Crimean-Congo hemorrhagic fever mouse model.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne nairovirus with a wide geographic spread that can cause severe and lethal disease. No specific medical countermeasures are approved to combat this illness. The CCHFV L protein contains an ovarian tumor (OTU) domain with a cysteine protease thought to modulate cellular immune responses by removing ubiquitin and ISG15 post-translational modifications from host and viral proteins. Viral deubiquitinases like CCHFV OTU are attractive drug targets, as blocking their activity may enhance cellular immune responses to infection, and potentially inhibit viral replication itself. We previously demonstrated that the engineered ubiquitin variant CC4 is a potent inhibitor of CCHFV replication in vitro. A major challenge of the therapeutic use of small protein inhibitors such as CC4 is their requirement for intracellular delivery, e.g., by viral vectors. In this study, we examined the feasibility of in vivo CC4 delivery by a replication-deficient recombinant adenovirus (Ad-CC4) in a lethal CCHFV mouse model. Since the liver is a primary target of CCHFV infection, we aimed to optimize delivery to this organ by comparing intravenous (tail vein) and intraperitoneal injection of Ad-CC4. While tail vein injection is a traditional route for adenovirus delivery, in our hands intraperitoneal injection resulted in higher and more widespread levels of adenovirus genome in tissues, including, as intended, the liver. However, despite promising in vitro results, neither route of in vivo CC4 treatment resulted in protection from a lethal CCHFV infection.

An Immunoinformatics Prediction of Novel Multi-Epitope Vaccines Candidate Against Surface Antigens of Nipah Virus.

Nipah virus (NiV) is an emerging zoonotic virus causing outbreaks of encephalitis and respiratory illnesses in humans, with high mortality. NiV is considered endemic in Bangladesh and Southeast Asia. There are no licensed vaccines against NiV. This study aimed at predicting a dual-antigen multi-epitope subunit chimeric vaccine against surface-glycoproteins G and F of NiV. Targeted proteins were subjected to immunoinformatics analyses to predict antigenic B-cell and T-cell epitopes. The proposed vaccine designs were implemented based on the conservancy, population coverage, molecular docking, immune simulations, codon adaptation, secondary mRNA structure, and in-silico cloning. Total 40 T and B-cell epitopes were found to be conserved, antigenic (vaxijen-value > 0.4), non-toxic, non-allergenic, and human non-homologous. Of 12 hypothetical vaccines, two (NiV_BGD_V1 and NiV_BGD_V2) were strongly immunogenic, non-allergenic, and structurally stable. The proposed vaccine candidates show a negative Z-score (- 6.32 and - 6.67) and 83.6% and 89.3% of most rama-favored regions. The molecular docking confirmed the highest affinity of NiV_BGD_V1 and NiV_BGD_V2 with TLR-4 (ΔG = - 30.7) and TLR8 (ΔG = - 20.6), respectively. The vaccine constructs demonstrated increased levels of immunoglobulins and cytokines in humans and could be expressed properly using an adenoviral-based pAdTrack-CMV expression vector. However, more experimental investigations and clinical trials are needed to validate its efficacy and safety. Supplementary Information: The online version contains supplementary material available at 10.1007/s10989-022-10431-z.

Broad-Spectrum In Vitro Antiviral Activity of ODBG-P-RVn: An Orally-Available, Lipid-Modified Monophosphate Prodrug of Remdesivir Parent Nucleoside (GS-441524).

The necessity for intravenous administration of remdesivir confines its utility for treatment of coronavirus disease 2019 (COVID-19) to hospitalized patients. We evaluated the broad-spectrum antiviral activity of ODBG-P-RVn, an orally available, lipid-modified monophosphate prodrug of the remdesivir parent nucleoside (GS-441524), against viruses that cause diseases of human public health concern, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). ODBG-P-RVn showed 20-fold greater antiviral activity than GS-441524 and had activity nearly equivalent to that of remdesivir in primary-like human small airway epithelial cells. Our results warrant in vivo efficacy evaluation of ODBG-P-RVn. IMPORTANCE While remdesivir remains one of the few drugs approved by the FDA to treat coronavirus disease 2019 (COVID-19), its intravenous route of administration limits its use to hospital settings. Optimizing the stability and absorption of remdesivir may lead to a more accessible and clinically potent therapeutic. Here, we describe an orally available lipid-modified version of remdesivir with activity nearly equivalent to that of remdesivir against emerging viruses that cause significant disease, including Ebola and Nipah viruses. Our work highlights the importance of such modifications to optimize drug delivery to relevant and appropriate human tissues that are most affected by such diseases.

Screening and Identification of Lujo Virus Inhibitors Using a Recombinant Reporter Virus Platform.

Lujo virus (LUJV), a highly pathogenic arenavirus, was first identified in 2008 in Zambia. To aid the identification of effective therapeutics for LUJV, we developed a recombinant reporter virus system, confirming reporter LUJV comparability with wild-type virus and its utility in high-throughput antiviral screening assays. Using this system, we evaluated compounds with known and unknown efficacy against related arenaviruses, with the aim of identifying LUJV-specific and potential new pan-arenavirus antivirals. We identified six compounds demonstrating robust anti-LUJV activity, including several compounds with previously reported activity against other arenaviruses. These data provide critical evidence for developing broad-spectrum antivirals against high-consequence arenaviruses.

Remdesivir targets a structurally analogous region of the Ebola virus and SARS-CoV-2 polymerases.

Remdesivir is a broad-spectrum antiviral nucleotide prodrug that has been clinically evaluated in Ebola virus patients and recently received emergency use authorization (EUA) for treatment of COVID-19. With approvals from the Federal Select Agent Program and the Centers for Disease Control and Prevention's Institutional Biosecurity Board, we characterized the resistance profile of remdesivir by serially passaging Ebola virus under remdesivir selection; we generated lineages with low-level reduced susceptibility to remdesivir after 35 passages. We found that a single amino acid substitution, F548S, in the Ebola virus polymerase conferred low-level reduced susceptibility to remdesivir. The F548 residue is highly conserved in filoviruses but should be subject to specific surveillance among novel filoviruses, in newly emerging variants in ongoing outbreaks, and also in Ebola virus patients undergoing remdesivir therapy. Homology modeling suggests that the Ebola virus polymerase F548 residue lies in the F-motif of the polymerase active site, a region that was previously identified as susceptible to resistance mutations in coronaviruses. Our data suggest that molecular surveillance of this region of the polymerase in remdesivir-treated COVID-19 patients is also warranted.

Tuberculosis case finding using population-based disease surveillance platforms in urban and rural Kenya.

BACKGROUND: Tuberculosis (TB) case finding is an important component of TB control because it can reduce transmission of Mycobacterium tuberculosis (MTB) through prompt detection and treatment of infectious patients. METHODS: Using population-based infectious disease surveillance (PBIDS) platforms with links to health facilities in Kenya we implemented intensified TB case finding in the community and at the health facilities, as an adjunct to routine passive case finding conducted by the national TB program. From 2011 to 2014, PBIDS participants ≥15 years were screened either at home or health facilities for possible TB symptoms which included cough, fever, night sweats or weight loss in the preceding 2 weeks. At home, participants with possible TB symptoms had expectorated sputum collected. At the clinic, HIV-infected participants with possible TB symptoms were invited to produce sputum. Those without HIV but with symptoms lasting 7 days including the visit day had chest radiographs performed, and had sputum collected if the radiographs were abnormal. Sputum samples were tested for the presence of MTB using the Xpert MTB/RIF assay. TB detection rates were calculated per 100,000 persons screened. RESULTS: Of 11,191 participants aged ≥15 years screened at home at both sites, 2695 (23.9%) reported possible TB symptoms, of whom 2258 (83.8%) produced sputum specimens. MTB was detected in 32 (1.4%) of the specimens resulting in a detection rate of 286/100,000 persons screened. At the health facilities, a total of 11,762 person were screened, 7500 (63.8%) had possible TB symptoms of whom 1282 (17.1%) produced sputum samples. MTB was detected in 69 (5.4%) of the samples, resulting in an overall detection rate of 587/100,000 persons screened. The TB detection rate was higher in persons with HIV compared to those without at both home (HIV-infected - 769/100,000, HIV-uninfected 141/100,000, rate ratio (RR) - 5.45, 95% CI 3.25-22.37), and health facilities (HIV-infected 3399/100,000, HIV-uninfected 294/100,000, RR 11.56, 95% CI 6.18-18.44). CONCLUSION: Facility-based intensified TB case finding detected more TB cases per the number of specimens tested and the number of persons screened, including those with HIV, than home-based TB screening and should be further evaluated to determine its potential programmatic impact.

Risk factors of hypertension among adults aged 35-64 years living in an urban slum Nairobi, Kenya.

BACKGROUND: Hypertension is an emerging public health problem in Sub Saharan Africa (SSA) and urbanization is considered to favor its emergence. Given a paucity of information on hypertension and associated risk factors among urban slum dwellers in SSA, we aimed to characterize the distribution of risk factors for hypertension and investigate their association with hypertension in an urban slum in Kenya. METHODS: We conducted a community based cross-sectional survey among adults 35 years and older living in Kibera slum Nairobi, Kenya. Trained interviewers collected data on socio demographic characteristics and self reported health behaviours using modified World Health Organization stepwise surveillance questionnaire for chronic disease risk factors. Anthropometric and blood pressure measurements were performed following standard procedures. Multiple logistic regression was used for analysis and odds ratios with 95 % confidence intervals were calculated to identify risk factors associated with hypertension. RESULT: A total of 1528 adults were surveyed with a mean age of 46.7 years. The age-standardized prevalence of hypertension was 29.4 % (95 % CI 27.0-31.7). Among the 418 participants classified as hypertensive, over one third (39.0 %) were unaware they had hypertension. Prevalence of current smoking and alcohol consumption was 8.5 and 13.1 % respectively. Over one quarter 26.2 % participants were classified as overweight (Body Mass Index [BMI] ≥25 to ≤29.9 kg/m(2)), and 17 % classified as obese (BMI ≥30 kg/m(2)). Overweight, obesity, current smoking, some level of education, highest wealth index, moderate physical activity, older age and being widowed were each independently associated with hypertension. When fit in a multivariable logistic regression model, being a widow [AOR = 1.7; (95 % CI, 1.1-2.6)], belonging to the highest wealth index [AOR = 1.6; (95 % CI, 1.1-2.5)], obesity [AOR = 1.8; 95 % CI, 1.1-3.1)] and moderate physical activity [AOR = 1.9; (95 % CI, 1.2-3.0)], all remained significantly associated with hypertension. CONCLUSION: Hypertension in the slum is a public health problem affecting at least one in three adults aged 35-64 years. Age, marital status, wealth index, physical inactivity and body mass index are important risk factors associated with hypertension. Prevention measures targeting the modifiable risk factors associated with hypertension are warranted to curb hypertension and its progressive effects.

Wide distribution and ancient evolutionary history of simian foamy viruses in New World primates.

BACKGROUND: Although simian foamy viruses (SFV) are the only exogenous retroviruses to infect New World monkeys (NWMs), little is known about their evolutionary history and epidemiology. Previous reports show distinct SFVs among NWMs but were limited to small numbers of captive or wild monkeys from five (Cebus, Saimiri, Ateles, Alouatta, and Callithrix) of the 15 NWM genera. Other studies also used only PCR testing or serological assays with limited validation and may have missed infection in some species. We developed and validated new serological and PCR assays to determine the prevalence of SFV in blood specimens from a large number of captive NWMs in the US (n = 274) and in captive and wild-caught NWMs (n = 236) in Peruvian zoos, rescue centers, and illegal trade markets. Phylogenetic and co-speciation reconciliation analyses of new SFV polymerase (pol) and host mitochondrial cytochrome B sequences, were performed to infer SFV and host co-evolutionary histories. RESULTS: 124/274 (45.2 %) of NWMs captive in the US and 59/157 (37.5 %) of captive and wild-caught NWMs in Peru were SFV WB-positive representing 11 different genera (Alouatta, Aotus, Ateles, Cacajao, Callithrix, Cebus, Lagothrix, Leontopithecus, Pithecia, Saguinus and Saimiri). Seroprevalences were lower at rescue centers (10/53, 18.9 %) compared to zoos (46/97, 47.4 %) and illegal trade markets (3/7, 8/19, 42.9 %) in Peru. Analyses showed that the trees of NWM hosts and SFVs have remarkably similar topologies at the level of species and sub-populations suggestive of co-speciation. Phylogenetic reconciliation confirmed 12 co-speciation events (p < 0.002) which was further supported by obtaining highly similar divergence dates for SFV and host genera and correlated SFV-host branch times. However, four ancient cross-genus transmission events were also inferred for Pitheciinae to Atelidae, Cacajao to ancestral Callithrix or Cebus monkeys, between Callithrix and Cebus monkeys, and Lagothrix to Alouatta. CONCLUSIONS: We demonstrate a broad distribution and stable co-speciation history of SFV in NWMs at the species level. Additional studies are necessary to further explore the epidemiology and natural history of SFV infection of NWMs and to determine the zoonotic potential for persons exposed to infected monkeys in captivity and in the wild.

Etiology and Incidence of Viral Acute Respiratory Infections Among Refugees Aged 5 Years and Older in Hagadera Camp, Dadaab, Kenya.

We used the Centers for Disease Control and Prevention-Kenya Medical Research Institute Acute Respiratory Infection (ARI) Surveillance System data to estimate severe acute respiratory infection (SARI) hospitalization rates, viral etiology, and associated complaints of influenza-like illnesses (ILI) and SARI conditions among those aged 5 years and older in Hagadera, Dadaab refugee camp, Kenya, for 2010-2012. A total of 471 patients aged ≥ 5 years met the case definition for ILI or SARI. SARI hospitalization rates per 10,000 person-years were 14.7 (95% confidence interval [CI] = 9.1, 22.2) for those aged 5-14 years; 3.4 (95% CI = 1.6, 7.2) for those aged 15-24 year; and 3.8 (95% CI = 1.6, 7.2) for those aged ≥ 25 years. Persons between the ages of 5 and 14 years had 3.5 greater odds to have been hospitalized as a result of SARI than those aged ≥ 25 years (odds ratio [OR] = 3.5, P < 0.001). Among the 419 samples tested, 169 (40.3%) were positive for one or more virus. Of those samples having viruses, 36.9% had influenza A; 29.9% had adenovirus; 20.2% had influenza B; and 14.4% had parainfluenza 1, 2, or 3. Muscle/joint pain was associated with influenza A (P = 0.002), whereas headache was associated with influenza B (P = 0.019). ARIs were responsible for a substantial disease burden in Hagadera camp.

Needs, acceptability, and value of humanitarian medical assistance in remote Peruvian Amazon riverine communities.

Much debate exists regarding the need, acceptability, and value of humanitarian medical assistance. We conducted a cross-sectional study on 457 children under 5 years from four remote riverine communities in the Peruvian Amazon and collected anthropometric measures, blood samples (1-4 years), and stool samples. Focus groups and key informant interviews assessed perspectives regarding medical aid delivered by foreigners. The prevalence of stunting, anemia, and intestinal parasites was 20%, 37%, and 62%, respectively. Infection with multiple parasites, usually geohelminths, was detected in 41% of children. The prevalence of intestinal parasites both individual and polyparasitism increased with age. Participants from smaller communities less exposed to foreigners expressed lack of trust and fear of them. However, participants from all communities were positive about foreigners visiting to provide health support. Prevalent health needs such as parasitic infections and anemia may be addressed by short-term medical interventions. There is a perceived openness to and acceptability of medical assistance delivered by foreign personnel.

Severe acute respiratory infection in children in a densely populated urban slum in Kenya, 2007-2011.

BACKGROUND: Reducing acute respiratory infection burden in children in Africa remains a major priority and challenge. We analyzed data from population-based infectious disease surveillance for severe acute respiratory illness (SARI) among children <5 years of age in Kibera, a densely populated urban slum in Nairobi, Kenya. METHODS: Surveillance was conducted among a monthly mean of 5,874 (range = 5,778-6,411) children <5 years old in two contiguous villages in Kibera. Participants had free access to the study clinic and their health events and utilization were noted during biweekly home visits. Patients meeting criteria for SARI (WHO-defined severe or very severe pneumonia, or oxygen saturation <90%) from March 1, 2007-February 28, 2011 had blood cultures processed for bacteria, and naso- and oro- pharyngeal swabs collected for quantitative real-time reverse transcription polymerase chain reaction testing for influenza viruses, parainfluenza viruses (PIV), respiratory syncytial virus (RSV), adenovirus, and human metapneumovirus (hMPV). Swabs collected during January 1, 2009 - February 28, 2010 were also tested for rhinoviruses, enterovirus, parechovirus, Mycoplasma pneumoniae, and Legionella species. Swabs were collected for simultaneous testing from a selected group of control-children visiting the clinic without recent respiratory or diarrheal illnesses. RESULTS: SARI overall incidence was 12.4 cases/100 person-years of observation (PYO) and 30.4 cases/100 PYO in infants. When comparing detection frequency in swabs from 815 SARI cases and 115 healthy controls, only RSV and influenza A virus were significantly more frequently detected in cases, although similar trends neared statistical significance for PIV, adenovirus and hMPV. The incidence for RSV was 2.8 cases/100 PYO and for influenza A was 1.0 cases/100 PYO. When considering all PIV, the rate was 1.1 case/100 PYO and the rate per 100 PYO for SARI-associated disease was 1.5 for adenovirus and 0.9 for hMPV. RSV and influenza A and B viruses were estimated to account for 16.2% and 6.7% of SARI cases, respectively; when taken together, PIV, adenovirus, and hMPV may account for >20% additional cases. CONCLUSIONS: Influenza viruses and RSV (and possibly PIV, hMPV and adenoviruses) are important pathogens to consider when developing technologies and formulating strategies to treat and prevent SARI in children.

Native rodent species are unlikely sources of infection for Leishmania (Viannia) braziliensis along the Transoceanic Highway in Madre de Dios, Peru.

An estimated 2.3 million disability-adjusted life years are lost globally from leishmaniasis. In Peru's Amazon region, the department of Madre de Dios (MDD) rises above the rest of the country in terms of the annual incidence rates of human leishmaniasis. Leishmania (Viannia) braziliensis is the species most frequently responsible for the form of disease that results in tissue destruction of the nose and mouth. However, essentially nothing is known regarding the reservoirs of this vector-borne, zoonotic parasite in MDD. Wild rodents have been suspected, or proven, to be reservoirs of several Leishmania spp. in various ecosystems and countries. Additionally, people who live or work in forested terrain, especially those who are not regionally local and whose immune systems are thus naïve to the parasite, are at most risk for contracting L. (V.) braziliensis. Hence, the objective of this study was to collect tissues from wild rodents captured at several study sites along the Amazonian segment of the newly constructed Transoceanic Highway and to use molecular laboratory techniques to analyze samples for the presence of Leishmania parasites. Liver tissues were tested via polymerase chain reaction from a total of 217 rodents; bone marrow and skin biopsies (ear and tail) were also tested from a subset of these same animals. The most numerous rodent species captured and tested were Oligoryzomys microtis (40.7%), Hylaeamys perenensis (15.7%), and Proechimys spp. (12%). All samples were negative for Leishmania, implying that although incidental infections may occur, these abundant rodent species are unlikely to serve as primary reservoirs of L. (V.) braziliensis along the Transoceanic Highway in MDD. Therefore, although these rodent species may persist and even thrive in moderately altered landscapes, we did not find any evidence to suggest they pose a risk for L. (V.) braziliensis transmission to human inhabitants in this highly prevalent region.

Human aflatoxin exposure in Kenya, 2007: a cross-sectional study.

Aflatoxins contaminate approximately 25% of agricultural products worldwide. They can cause liver failure and liver cancer. Kenya has experienced multiple aflatoxicosis outbreaks in recent years, often resulting in fatalities. However, the full extent of aflatoxin exposure in Kenya has been unknown. Our objective was to quantify aflatoxin exposure across Kenya. We analysed aflatoxin levels in serum specimens from the 2007 Kenya AIDS Indicator Survey - a nationally representative, cross-sectional serosurvey. KAIS collected 15,853 blood specimens. Of the 3180 human immunodeficiency virus-negative specimens with ≥1 mL sera, we randomly selected 600 specimens stratified by province and sex. We analysed serum specimens for aflatoxin albumin adducts by using isotope dilution MS/MS to quantify aflatoxin B1-lysine, and normalised with serum albumin. Aflatoxin concentrations were then compared by demographic, socioeconomic and geographic characteristics. We detected serum aflatoxin B1-lysine in 78% of serum specimens (range =

Recombinant protein-based assays for detection of antibodies to severe acute respiratory syndrome coronavirus spike and nucleocapsid proteins.

Recombinant severe acute respiratory syndrome (SARS) nucleocapsid and spike protein-based immunoglobulin G immunoassays were developed and evaluated. Our assays demonstrated high sensitivity and specificity to the SARS coronavirus in sera collected from patients as late as 2 years postonset of symptoms. These assays will be useful not only for routine SARS coronavirus diagnostics but also for epidemiological and antibody kinetic studies.