2020 Ebola virus disease outbreak in Équateur Province, Democratic Republic of the Congo: a retrospective genomic characterisation.

BACKGROUND: The Democratic Republic of the Congo has had 15 Ebola virus disease (EVD) outbreaks, from 1976 to 2023. On June 1, 2020, the Democratic Republic of the Congo declared an outbreak of EVD in the western Équateur Province (11th outbreak), proximal to the 2018 Tumba and Bikoro outbreak and concurrent with an outbreak in the eastern Nord Kivu Province. In this Article, we assessed whether the 11th outbreak was genetically related to previous or concurrent EVD outbreaks and connected available epidemiological and genetic data to identify sources of possible zoonotic spillover, uncover additional unreported cases of nosocomial transmission, and provide a deeper investigation into the 11th outbreak. METHODS: We analysed epidemiological factors from the 11th EVD outbreak to identify patient characteristics, epidemiological links, and transmission modes to explore virus spread through space, time, and age groups in the Équateur Province, Democratic Republic of the Congo. Trained field investigators and health professionals recorded data on suspected, probable, and confirmed cases, including demographic characteristics, possible exposures, symptom onset and signs and symptoms, and potentially exposed contacts. We used blood samples from individuals who were live suspected cases and oral swabs from individuals who were deceased to diagnose EVD. We applied whole-genome sequencing of 87 available Ebola virus genomes (from 130 individuals with EVD between May 19 and Sept 16, 2020), phylogenetic divergence versus time, and Bayesian reconstruction of phylogenetic trees to calculate viral substitution rates and study viral evolution. We linked the available epidemiological and genetic datasets to conduct a genomic and epidemiological study of the 11th EVD outbreak. FINDINGS: Between May 19 and Sept 16, 2020, 130 EVD (119 confirmed and 11 probable) cases were reported across 13 Équateur Province health zones. The individual identified as the index case reported frequent consumption of bat meat, suggesting the outbreak started due to zoonotic spillover. Sequencing revealed two circulating Ebola virus variants associated with this outbreak-a Mbandaka variant associated with the majority (97%) of cases and a Tumba-like variant with similarity to the ninth EVD outbreak in 2018. The Tumba-like variant exhibited a reduced substitution rate, suggesting transmission from a previous survivor of EVD. INTERPRETATION: Integrating genetic and epidemiological data allowed for investigative fact-checking and verified patient-reported sources of possible zoonotic spillover. These results demonstrate that rapid genetic sequencing combined with epidemiological data can inform responders of the mechanisms of viral spread, uncover novel transmission modes, and provide a deeper understanding of the outbreak, which is ultimately needed for infection prevention and control during outbreaks. FUNDING: WHO and US Centers for Disease Control and Prevention.

Seroepidemiological investigation of Crimean Congo hemorrhagic fever virus in livestock in Uganda, 2017.

Crimean-Congo Hemorrhagic fever (CCHF) is an important zoonotic disease transmitted to humans both by tick vectors and contact with fluids from an infected animal or human. Although animals are not symptomatic when infected, they are the main source of human infection. Uganda has reported sporadic human outbreaks of CCHF in various parts of the country since 2013. We designed a nationwide epidemiological study to investigate the burden of CCHF in livestock. A total of 3181 animals were sampled; 1732 cattle (54.4%), 1091 goats (34.3%), and 358 sheep (11.3%) resulting in overall livestock seropositivity of IgG antibodies against CCHF virus (CCHFV) of 31.4% (999/3181). Seropositivity in cattle was 16.9% and in sheep and goats was 48.8%. Adult and juvenile animals had higher seropositivity compared to recently born animals, and seropositivity was higher in female animals (33.5%) compared to male animals (24.1%). Local breeds had higher (36.8%) compared to exotic (2.8%) and cross breeds (19.3%). Animals that had a history of abortion or stillbirth had higher seropositivity compared to those without a history of abortion or stillbirth. CCHFV seropositivity appeared to be generally higher in northern districts of the country, though spatial trends among sampled districts were not examined. A multivariate regression analysis using a generalized linear mixed model showed that animal species, age, sex, region, and elevation were all significantly associated with CCHFV seropositivity after adjusting for the effects of other model predictors. This study shows that CCHFV is actively circulating in Uganda, posing a serious risk for human infection. The results from this study can be used to help target surveillance efforts for early case detection in animals and limit subsequent spillover into humans.

Recombinant Sudan virus and evaluation of humoral cross-reactivity between Ebola and Sudan virus glycoproteins after infection or rVSV-ΔG-ZEBOV-GP vaccination.

Ebola disease outbreaks are major public health events because of human-to-human transmission and high mortality. These outbreaks are most often caused by Ebola virus, but at least three related viruses can also cause the disease. In 2022, Sudan virus re-emerged causing more than 160 confirmed and probable cases. This report describes generation of a recombinant Sudan virus and demonstrates its utility by quantifying antibody cross-reactivity between Ebola and Sudan virus glycoproteins after human infection or vaccination with a licensed Ebola virus vaccine.

A Countrywide Seroepidemiological Survey of Rift Valley Fever in Livestock, Uganda, 2017.

In 2016, an outbreak of Rift Valley fever was reported in the Kabale District in Uganda for the first time in 48 years. Three human cases were confirmed by polymerase chain reaction, and subsequent serological investigations revealed an overall IgG seropositivity of 13% in humans and 13% in animals. In response to this reemergence, we designed a countrywide survey to determine the seropositivity of anti-Rift Valley fever virus (RVFV) IgG antibodies in livestock. Samples were collected from 27 districts and tested for RVFV anti-IgG antibodies. A total of 3,181 livestock samples were tested, of which 54.4% were cattle (1,732 of 3,181), 34.3% were goats (1,091 of 3,181), and 11.3% were sheep (358 of 3,181). Overall RVFV seropositivity was 6.9% (221 of 3,181). Seroprevalence was greater in cattle (10.7%) compared with goats (2.6%) and sheep (2.0%), among females (7.5%) compared with males (5.2%), and among adults (7.6%) compared with juveniles (4.9%) and nurslings (6.4%). Exotic breeds and animals with a history of abortion or stillbirth also had greater odds of RVFV seropositivity. Animals grazed under tethering and paddocking had greater RVFV seropositivity compared with animals that grazed communally, and livestock in the western and eastern regions had the greatest seroprevalence. In a multivariate regression model, animal species (odds ratio [OR], 6.4; 95% CI, 3.5-11.4) and age (OR, 2.3; 95% CI, 1.4-3.6) were associated significantly with RVFV seropositivity. This study could be important in developing risk-based surveillance for early outbreak detection to limit the spread of RVFV in both human and animal populations.

Hantavirus Pulmonary Syndrome: 1993-2018.

OBJECTIVES: From 1993 to 2018, hantavirus infections were reported in 39 states, with hantavirus pulmonary syndrome (HPS) as the most common and fatal manifestation. To identify differences in the presentation of HPS between children and adults, we hypothesized that children with HPS would be diagnosed later in their illness course given the nonspecific clinical features of HPS. METHODS: This was an evaluation of the clinical and demographic characteristics of national HPS cases from 1993 to 2018. Data were from the Centers for Disease Control and Prevention database and 1 state department of health, comprising 97% of US cases. We compared children (0 to 12 years), adolescents (13 to 18 years), and adults using nonparametric and parametric analyses, with additional exploratory analyses to identify clinical variables associated with mortality. RESULTS: Among 719 HPS patients, 22 (3.0%) were aged ≤12 years, 47 (6.5%) were 13 to 18 years old, and the remaining 650 (90.4%) were adults. Overall mortality was 35.4% and did not differ between age groups (P = .8). The time between symptom onset and death differed by age group, with children living a median of 2 days (interquartile range [IQR] 2 to 3), adolescents 4 days (IQR 3 to 5), and adults 5 days (IQR 4 to 8; P = .001). The mean highest hematocrit and median highest creatinine level were significantly associated with mortality in those 0 to 18 years old but not adults. CONCLUSIONS: In our dataset representing the largest study of HPS in the United States, we found that children with HPS died more quickly than adults and that highest hematocrit and creatinine levels were associated with death only among those <19 years old.

Detection of Sporadic Outbreaks of Rift Valley Fever in Uganda through the National Viral Hemorrhagic Fever Surveillance System, 2017-2020.

Rift Valley fever (RVF) is a zoonotic disease of public health and economic importance. Uganda has reported sporadic outbreaks of RVF in both humans and animals across the country, especially in the southwestern part of the "cattle corridor" through an established viral hemorrhagic fever surveillance system. We report 52 human cases of laboratory-confirmed RVF from 2017 to 2020. The case fatality rate was 42%. Among those infected, 92% were males and 90% were adults (≥ 18 years). Clinical symptoms were characterized by fever (69%), unexplained bleeding (69%), headache (51%), abdominal pain (49%), and nausea and vomiting (46%). Most of the cases (95%) originated from central and western districts that are part of the cattle corridor of Uganda, where the main risk factor was direct contact with livestock (P = 0.009). Other predictors of RVF positivity were determined to be male gender (P = 0.001) and being a butcher (P = 0.04). Next-generation sequencing identified the predominant Ugandan clade as Kenya-2, observed previously across East Africa. There is need for further investigation and research into the effect and spread of this neglected tropical disease in Uganda and the rest of Africa. Control measures such as promoting vaccination and limiting animal-human transmission could be explored to reduce the impact of RVF in Uganda and globally.

Use of Ebola Vaccine: Expansion of Recommendations of the Advisory Committee on Immunization Practices To Include Two Additional Populations - United States, 2021.

On December 19, 2019, the Food and Drug Administration (FDA) approved rVSVΔG-ZEBOV-GP Ebola vaccine (ERVEBO, Merck) for the prevention of Ebola virus disease (EVD) caused by infection with Ebola virus, species Zaire ebolavirus, in adults aged ≥18 years. In February 2020, the Advisory Committee on Immunization Practices (ACIP) recommended preexposure vaccination with ERVEBO for adults aged ≥18 years in the United States who are at highest risk for potential occupational exposure to Ebola virus because they are responding to an outbreak of EVD, work as health care personnel at federally designated Ebola treatment centers in the United States, or work as laboratorians or other staff members at biosafety level 4 facilities in the United States (1).

Postmortem Surveillance for Ebola Virus Using OraQuick Ebola Rapid Diagnostic Tests, Eastern Democratic Republic of the Congo, 2019-2020.

After a pilot study, we tested 443 cadavers using OraQuick Ebola rapid diagnostic tests during surveillance after the 10th Ebola outbreak in the Democratic Republic of the Congo. No false negative and 2% false-positive results were reported. Quickly returning results and engaging the community enabled timely public health actions.

Hantavirus Disease and COVID-19.

OBJECTIVES: Navajo Nation is disproportionately affected by hantavirus cardiopulmonary syndrome (HCPS), a severe respiratory disease that can quickly progress to respiratory failure and cardiogenic shock. The initial signs and symptoms of HCPS are indistinguishable from coronavirus disease 2019 (COVID-19). However, this distinction is critical, as the disease course differs greatly, with most patients with COVID-19 experiencing mild to moderate illness. We set out to determine if the evaluation of peripheral blood smears for five hematopathologic criteria previously identified as hallmarks of hantavirus infection, or "the hantavirus 5-point screen," could distinguish between COVID-19 and HCPS. METHODS: The hantavirus 5-point screen was performed on peripheral blood smears from 139 patients positive for COVID-19 seeking treatment from Tséhootsooí Medical Center and two Emory University hospitals. RESULTS: Of these 139 individuals, 136 (98%) received a score of 3/5 or below, indicating low suspicion for HCPS. While thrombocytopenia, one of the key signs of HCPS, was seen in the patients with COVID-19, it was generally mild and remained stable on repeat specimens collected 12 to 24 hours later. CONCLUSIONS: Given these findings, the 5-point screen remains a useful rapid screening tool for potential HCPS cases and may be useful to distinguish early HCPS from COVID-19 in HCPS endemic regions.

Development and implementation of the Ebola Exposure Window Calculator: A tool for Ebola virus disease outbreak field investigations.

During an Ebola virus disease (EVD) outbreak, calculating the exposure window of a confirmed case can assist field investigators in identifying the source of infection and establishing chains of transmission. However, field investigators often have difficulty calculating this window. We developed a bilingual (English/French), smartphone-based field application to assist field investigators in determining the exposure window of an EVD case. The calculator only requires the reported date of symptoms onset and the type of symptoms present at onset or the date of death. Prior to the release of this application, there was no similar electronic capability to enable consistent calculation of EVD exposure windows for field investigators. The Democratic Republic of the Congo Ministry of Health endorsed the application and incorporated it into trainings for field staff. Available for Apple and Android devices, the calculator continues to be downloaded even as the eastern DRC outbreak resolved. We rapidly developed and implemented a smartphone application to estimate the exposure window for EVD cases in an outbreak setting.

Use of Ebola Vaccine: Recommendations of the Advisory Committee on Immunization Practices, United States, 2020.

This report summarizes the recommendations of the Advisory Committee on Immunization Practices (ACIP) for use of the rVSVΔG-ZEBOV-GP Ebola vaccine (Ervebo) in the United States. The vaccine contains rice-derived recombinant human serum albumin and live attenuated recombinant vesicular stomatitis virus (VSV) in which the gene encoding the glycoprotein of VSV was replaced with the gene encoding the glycoprotein of Ebola virus species Zaire ebolavirus. Persons with a history of severe allergic reaction (e.g., anaphylaxis) to rice protein should not receive Ervebo. This is the first and only vaccine currently licensed by the Food and Drug Administration for the prevention of Ebola virus disease (EVD). These guidelines will be updated based on availability of new data or as new vaccines are licensed to protect against EVD.ACIP recommends preexposure vaccination with Ervebo for adults aged ≥18 years in the U.S. population who are at highest risk for potential occupational exposure to Ebola virus species Zaire ebolavirus because they are responding to an outbreak of EVD, work as health care personnel at federally designated Ebola treatment centers in the United States, or work as laboratorians or other staff at biosafety level 4 facilities in the United States. Recommendations for use of Ervebo in additional populations at risk for exposure and other settings will be considered and discussed by ACIP in the future.

Use of Nucleic Acid Amplification Testing for Rapid Detection of Mycobacterium tuberculosis Complex Among US Tuberculosis Patients, 2011‒2017.

Background: Nucleic acid amplification (NAA) tests rapidly detect Mycobacterium tuberculosis complex directly from clinical specimens, providing valuable results for those evaluated for tuberculosis. Methods: We analyzed characteristics of cases with NAA testing performed, compared cases with positive and negative NAA test results, and calculated turnaround time and time to treatment for all verified cases reported to the National Tuberculosis Surveillance System in the United States during 2011-2017. Results: Among 67082 verified tuberculosis cases with NAA testing information, 30820 (45.9%) were reported as not having an NAA test performed; the proportion without NAA testing declined annually, from 60.5% in 2011 to 33.6% in 2017. Of 67082 verified cases, 27912 (41.6%) had positive, 8215 (12.2%) had negative, and 135 (0.2%) had indeterminate NAA test results. Among the 33937 cases with an acid-fast bacilli (AFB) smear-positive result, 24093 (70.9%) had an NAA test performed; 11490 of the 30244 (38.0%) with an AFB smear-negative result had an NAA test performed. Although sputum was the most common specimen type tested, 79.8% (7023/8804) of nonsputum specimen types had a positive NAA test result. Overall, 63.7% of cases with laboratory testing had NAA test results reported <6 days following specimen collection; for 13891 cases not yet on treatment, median time to treatment after the laboratory report date was 2 days. Conclusions: Our analyses demonstrate increased NAA test utilization between 2011 and 2017. However, a large proportion of cases did not have an NAA test performed, reflecting challenges in broader uptake, suggesting an opportunity to expand use of this diagnostic methodology.

Plasmodium vivax Infections in Duffy-Negative Individuals in the Democratic Republic of the Congo.

Although Plasmodium vivax has been assumed to be absent from sub-Saharan Africa because of the protective mutation conferring the Duffy-negative phenotype, recent evidence has suggested that P. vivax cases are prevalent in these regions. We selected 292 dried blood spots from children who participated in the 2013-2014 Demographic and Health Survey of the Democratic Republic of the Congo (DRC), to assess for P. vivax infection. Four P. vivax infections were identified by polymerase chain reaction, each in a geographically different survey cluster. Using these as index cases, we tested the remaining 73 samples from the four clusters. With this approach, 10 confirmed cases, three probable cases, and one possible case of P. vivax were identified. Among the 14 P. vivax cases, nine were coinfected with Plasmodium falciparum. All 14 individuals were confirmed to be Duffy-negative by sequencing for the single point mutation in the GATA motif that represses the expression of the Duffy antigen. This finding is consistent with a growing body of literature that suggests that P. vivax can infect Duffy-negative individuals in Africa. Future molecular and sequencing work is needed to understand the relationship of these isolates with other P. vivax samples from Asia and South America and discover variants linked to P. vivax virulence and erythrocyte invasion.

Pfhrp2-Deleted Plasmodium falciparum Parasites in the Democratic Republic of the Congo: A National Cross-sectional Survey.

Background: Rapid diagnostic tests (RDTs) account for more than two-thirds of malaria diagnoses in Africa. Deletions of the Plasmodium falciparum hrp2 (pfhrp2) gene cause false-negative RDT results and have never been investigated on a national level. Spread of pfhrp2-deleted P. falciparum mutants, resistant to detection by HRP2-based RDTs, would represent a serious threat to malaria elimination efforts. Methods: Using a nationally representative cross-sectional study of 7,137 children under five years of age from the Democratic Republic of Congo (DRC), we tested 783 subjects with RDT-/PCR+ results using PCR assays to detect and confirm deletions of the pfhrp2 gene. Spatial and population genetic analyses were employed to examine the distribution and evolution of these parasites. Results: We identified 149 pfhrp2-deleted parasites, representing 6.4% of all P. falciparum infections country-wide (95% confidence interval 5.1-8.0%). Bayesian spatial analyses identified statistically significant clustering of pfhrp2 deletions near Kinshasa and Kivu. Population genetic analysis revealed significant genetic differentiation between wild-type and pfhrp2-deleted parasite populations (GST = .046, p ≤ .00001). Conclusions: Pfhrp2-deleted P. falciparum is a common cause of RDT-/PCR+ malaria among asymptomatic children in the DRC and appears to be clustered within select communities. Surveillance for these deletions is needed, and alternatives to HRP2-specific RDTs may be necessary.

Low prevalence of Plasmodium malariae and Plasmodium ovale mono-infections among children in the Democratic Republic of the Congo: a population-based, cross-sectional study.

BACKGROUND: In an effort to improve surveillance for epidemiological and clinical outcomes, rapid diagnostic tests (RDTs) have become increasingly widespread as cost-effective and field-ready methods of malaria diagnosis. However, there are concerns that using RDTs specific to Plasmodium falciparum may lead to missed detection of other malaria species such as Plasmodium malariae and Plasmodium ovale. METHODS: Four hundred and sixty six samples were selected from children under 5 years old in the Democratic Republic of the Congo (DRC) who took part in a Demographic and Health Survey (DHS) in 2013-14. These samples were first tested for all Plasmodium species using an 18S ribosomal RNA-targeted real-time PCR; malaria-positive samples were then tested for P. falciparum, P. malariae and P. ovale using a highly sensitive nested PCR. RESULTS: The prevalence of P. falciparum, P. malariae and P. ovale were 46.6, 12.9 and 8.3 %, respectively. Most P. malariae and P. ovale infections were co-infected with P. falciparum-the prevalence of mono-infections of these species were only 1.0 and 0.6 %, respectively. Six out of these eight mono-infections were negative by RDT. The prevalence of P. falciparum by the more sensitive nested PCR was higher than that found previously by real-time PCR. CONCLUSIONS: Plasmodium malariae and P. ovale remain endemic at a low rate in the DRC, but the risk of missing malarial infections of these species due to falciparum-specific RDT use is low. The observed prevalence of P. falciparum is higher with a more sensitive PCR method.

Malaria surveillance in the Democratic Republic of the Congo: comparison of microscopy, PCR, and rapid diagnostic test.

Malaria surveillance is critical for control efforts, but diagnostic methods frequently disagree. Here, we compare microscopy, PCR, and a rapid diagnostic test in 7137 samples from children in the Democratic Republic of the Congo using latent class analysis. PCR had the highest sensitivity (94.6%) and microscopy had the lowest (76.7%).