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ABSTRACT The onset of the coronavirus disease 2019 (COVID-19) pandemic resulted in hundreds of in vitro diagnostic devices (IVDs) coming to market, facilitated by regulatory authorities allowing “emergency use” without a comprehensive evaluation of performance. The World Health Organization (WHO) released target product profiles (TPPs) specifying acceptable performance characteristics for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) assay devices. We evaluated 26 rapid diagnostic tests and 9 enzyme immunoassays (EIAs) for anti-SARS-CoV-2, suitable for use in low- and middle-income countries (LMICs), against these TPPs and other performance characteristics. The sensitivity and specificity ranged from 60.1 to 100% and 56.0 to 100%, respectively. Five of 35 test kits reported no false reactivity for 55 samples with potentially cross-reacting substances. Six test kits reported no false reactivity for 35 samples containing interfering substances, and only one test reported no false reactivity with samples positive for other coronaviruses (not SARS-CoV-2). This study demonstrates that a comprehensive evaluation of the performance of test kits against defined specifications is essential for the selection of test kits, especially in a pandemic setting. IMPORTANCE The markets have been flooded with hundreds of SARS-CoV-2 serology tests, and although there are many published reports on their performance, comparative reports are far fewer and tend to be limited to only a few tests. In this report, we comparatively assessed 35 rapid diagnostic tests or microtiter plate enzyme immunoassays (EIAs) using a large set of samples from individuals with a history of mild to moderate COVID-19, commensurate with the target population for serosurveillance, which included serum samples from individuals previously infected, at undetermined time periods, with other seasonal human coronaviruses, Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-1. The significant heterogeneity in their performances, with only a few tests meeting WHO target product profile performance requirements, highlights the importance of independent comparative assessments to inform the use and procurement of these tests for both diagnostics and epidemiological investigations.

We detected Zika virus RNA in rectal swab samples from 10 patients by using real-time reverse transcription PCR, and we isolated the virus from 1 patient. The longest interval from symptom onset to detection was 14 days. These findings are applicable to diagnosis and infection prevention recommendations.

Marburg virus (MARV) is a negative-sense, single-stranded RNA virus that belongs to the Filoviridae family. Despite having caused numerous outbreaks of severe hemorrhagic fever with high case fatality rates, there are still no clinically approved therapeutics or vaccines to treat or prevent MARV disease. Recombinant vesicular stomatitis viruses (rVSVs) expressing heterologous viral glycoproteins have shown remarkable promise as live-attenuated vaccine vectors, with an rVSV-based Ebola virus vaccine having received regulatory approval in the United States and numerous other countries. Analogous rVSV vaccine vectors have also been developed for MARV and have shown efficacy in several preclinical studies conducted in nonhuman primates. Here, we used a guinea pig model to confirm the protective efficacy of a cloned, rVSV-based candidate vaccine, termed PHV01, expressing the MARV variant Angola glycoprotein. Our results demonstrated that a single dose (2 × 106 PFU) of vaccine administered 28 days prior to challenge with a uniformly lethal dose of guinea-pig-adapted MARV variant Angola provided complete protection from death and disease. Moreover, protection was robust, with as little as 200 PFU of vaccine conferring significant protection. Not only does this study highlight the potential predictive value of the guinea pig model in the evaluation of MARV countermeasures, but it also demonstrates consistent and reproducible protection afforded by a clonal vaccine candidate. Indeed, this study identifies PHV01 as a suitable vaccine candidate for advanced development.

We describe a strain of Lassa virus representing a putative new lineage that was isolated from a cluster of human infections with an epidemiologic link to Togo. This finding extends the known range of Lassa virus to Togo.

Nipah virus (NiV) has been transmitted from patient to caregivers in Bangladesh presumably through oral secretions. We aimed to detect whether NiV-infected patients contaminate hospital surfaces with the virus. During December 2013–April 2014, we collected 1 swab sample from 5 surfaces near NiV-infected patients and tested surface and oral swab samples by real-time reverse transcription PCR for NiV RNA. We identified 16 Nipah patients; 12 cases were laboratory-confirmed and 4 probable. Of the 12 laboratory-confirmed cases, 10 showed NiV RNA in oral swab specimens. We obtained surface swab samples for 6 Nipah patients; 5 had evidence of NiV RNA on >1 surface: 4 patients contaminated towels, 3 bed sheets, and 1 the bed rail. Patients with NiV RNA in oral swab samples were significantly more likely than other Nipah patients to die. To reduce the risk for fomite transmission of NiV, infection control should target hospital surfaces.

Summary: Following cessation of continuous Ebola virus (EBOV) transmission within Western Africa, sporadic EBOV disease (EVD) cases continued to re-emerge beyond the viral incubation period. Epidemiological and genomic evidence strongly suggests that this represented transmission from EVD survivors. To investigate whether persistent infections are characterized by ongoing viral replication, we sequenced EBOV from the semen of nine EVD survivors and a subset of corresponding acute specimens. EBOV evolutionary rates during persistence were either similar to or reduced relative to acute infection rates. Active EBOV replication/transcription continued during convalescence, but decreased over time, consistent with viral persistence rather than viral latency. Patterns of genetic divergence suggest a moderate relaxation of selective constraints within the sGP carboxy-terminal tail during persistent infections, but do not support widespread diversifying selection. Altogether, our data illustrate that EBOV persistence in semen, urine, and aqueous humor is not a quiescent or latent infection. : Whitmer et al. find that Ebola virus continues replication/transcription within the eye and male genital tract of Ebola virus disease survivors. They describe viral replication, evolutionary rates, and selective pressures experienced during acute and persistent infection. Keywords: Ebola virus, EVD survivors, persistent viral infection, evolutionary pressure, evolutionary rates, RNA hyper-editing

In September 2014, a single fatal case of Marburg virus was identified in a healthcare worker in Kampala, Uganda. The source of infection was not identified, and no secondary cases were identified. We describe the rapid identification, laboratory diagnosis, and case investigation of the third Marburg virus outbreak in Uganda.

Many of the survivors of the 2014–2015 epidemic of Ebola virus disease (EVD) in West Africa were women of childbearing age. Limited clinical and laboratory data exist that describe these women’s pregnancies and outcomes. We report the case of an EVD survivor who became pregnant and delivered her child in the United States, and we discuss implications of this case for infection control practices in obstetric services. Hospitals in the United States must be prepared to care for EVD survivors.

Nipah virus (NiV) is a paramyxovirus, and Pteropus spp. bats are the natural reservoir. From December 2010 through March 2014, hospital-based encephalitis surveillance in Bangladesh identified 18 clusters of NiV infection. The source of infection for case-patients in 3 clusters in 2 districts was unknown. A team of epidemiologists and anthropologists investigated these 3 clusters comprising 14 case-patients, 8 of whom died. Among the 14 case-patients, 8 drank fermented date palm sap (tari) regularly before their illness, and 6 provided care to a person infected with NiV. The process of preparing date palm trees for tari production was similar to the process of collecting date palm sap for fresh consumption. Bat excreta was reportedly found inside pots used to make tari. These findings suggest that drinking tari is a potential pathway of NiV transmission. Interventions that prevent bat access to date palm sap might prevent tari-associated NiV infection.

BACKGROUND:The 2013-2016 West African Ebola virus disease epidemic was unprecedented in terms of the number of cases and survivors. Prior to this epidemic there was limited data available on the persistence of Ebola virus in survivors' body fluids and the potential risk of transmission, including sexual transmission. METHODOLOGY/PRINCIPAL FINDINGS:Given the urgent need to determine the persistence of Ebola virus in survivors' body fluids, an observational cohort study was designed and implemented during the epidemic response operation in Sierra Leone. This publication describes study implementation methodology and the key lessons learned. Challenges encountered during implementation included unforeseen duration of follow-up, complexity of interpreting and communicating laboratory results to survivors, and the urgency of translating research findings into public health practice. Strong community engagement helped rapidly implement the study during the epidemic. The study was conducted in two phases. The first phase was initiated within five months of initial protocol discussions and assessed persistence of Ebola virus in semen of 100 adult men. The second phase assessed the persistence of virus in multiple body fluids (semen or vaginal fluid, menstrual blood, breast milk, and urine, rectal fluid, sweat, saliva, tears), of 120 men and 120 women. CONCLUSION/SIGNIFICANCE:Data from this study informed national and global guidelines in real time and demonstrated the need to implement semen testing programs among Ebola virus disease survivors. The lessons learned and study tools developed accelerated the implementation of such programs in Ebola virus disease affected countries, and also informed studies examining persistence of Zika virus. Research is a vital component of the public health response to an epidemic of a poorly characterized disease. Adequate resources should be rapidly made available to answer critical research questions, in order to better inform response efforts.

In summer 2012, an outbreak of hantavirus infections occurred among overnight visitors to Yosemite National Park in California, USA. An investigation encompassing clinical, epidemiologic, laboratory, and environmental factors identified 10 cases among residents of 3 states. Eight case-patients experienced hantavirus pulmonary syndrome, of whom 5 required intensive care with ventilatory support and 3 died. Staying overnight in a signature tent cabin (9 case-patients) was significantly associated with becoming infected with hantavirus (p

In 2012, a female wildlife biologist experienced fever, malaise, headache, generalized myalgia and arthralgia, neck stiffness, and a sore throat shortly after returning to the United States from a 6-week field expedition to South Sudan and Uganda. She was hospitalized, after which a maculopapular rash developed and became confluent. When the patient was discharged from the hospital on day 14, arthralgia and myalgia had improved, oropharynx ulcerations had healed, the rash had resolved without desquamation, and blood counts and hepatic enzyme levels were returning to reference levels. After several known suspect pathogens were ruled out as the cause of her illness, deep sequencing and metagenomics analysis revealed a novel paramyxovirus related to rubula-like viruses isolated from fruit bats. Download MP3 Length: 1:15

We investigated the extent of lymphocytic choriomeningitis virus (LCMV) infection in employees and rodents at 3 commercial breeding facilities. Of 97 employees tested, 31 (32%) had IgM and/or IgG to LCMV, and aseptic meningitis was diagnosed in 4 employees. Of 1,820 rodents tested in 1 facility, 382 (21%) mice (Mus musculus) had detectable IgG, and 13 (0.7%) were positive by reverse transcription PCR; LCMV was isolated from 8. Rats (Rattus norvegicus) were not found to be infected. S-segment RNA sequence was similar to strains previously isolated in North America. Contact by wild mice with colony mice was the likely source for LCMV, and shipments of infected mice among facilities spread the infection. The breeding colonies were depopulated to prevent further human infections. Future outbreaks can be prevented with monitoring and management, and employees should be made aware of LCMV risks and prevention. Download MP3 Length: 1:42

During follow-up of a 2012 US outbreak of lymphocytic choriomeningitis virus (LCMV), we conducted a trace-forward investigation. LCMV-infected feeder mice originating from a US rodent breeding facility had been distributed to >500 locations in 21 states. All mice from the facility were euthanized, and no additional persons tested positive for LCMV infection.

We investigated hantaviruses in rodents in the southern Amazon Basin of Peru and identified an Andes virus variant from Neacomys spinosus mice. This finding extends the known range of this virus in South America and the range of recognized hantaviruses in Peru. Further studies of the epizoology of hantaviruses in this region are warranted.

We measured the performance of exposure screening questions to identify Nipah virus encephalitis in hospitalized encephalitis patients during the 2012–13 Nipah virus season in Bangladesh. The sensitivity (93%), specificity (82%), positive predictive value (37%), and negative predictive value (99%) results suggested that screening questions could more quickly identify persons with Nipah virus encephalitis.

Background: Ebola virus has been detected in semen of Ebola virus disease survivors after recovery. Liberia's Men's Health Screening Program (MHSP) offers Ebola virus disease survivors semen testing for Ebola virus. We present preliminary results and behavioural outcomes from the first national semen testing programme for Ebola virus. Methods: The MHSP operates out of three locations in Liberia: Redemption Hospital in Montserrado County, Phebe Hospital in Bong County, and Tellewoyan Hospital in Lofa County. Men aged 15 years and older who had an Ebola treatment unit discharge certificate are eligible for inclusion. Participants' semen samples were tested for Ebola virus RNA by real-time RT-PCR and participants received counselling on safe sexual practices. Participants graduated after receiving two consecutive negative semen tests. Counsellors collected information on sociodemographics and sexual behaviours using questionnaires administered at enrolment, follow up, and graduation visits. Because the programme is ongoing, data analysis was restricted to data obtained from July 7, 2015, to May 6, 2016. Findings: As of May 6, 2016, 466 Ebola virus disease survivors had enrolled in the programme; real-time RT-PCR results were available from 429 participants. 38 participants (9%) produced at least one semen specimen that tested positive for Ebola virus RNA. Of these, 24 (63%) provided semen specimens that tested positive 12 months or longer after Ebola virus disease recovery. The longest interval between discharge from an Ebola treatment unit and collection of a positive semen sample was 565 days. Among participants who enrolled and provided specimens more than 90 days since their Ebola treatment unit discharge, men older than 40 years were more likely to have a semen sample test positive than were men aged 40 years or younger (p=0·0004). 84 (74%) of 113 participants who reported not using a condom at enrolment reported using condoms at their first follow-up visit (p

Two large outbreaks of Ebola hemorrhagic fever occurred in Uganda in 2000 and 2007. In May 2011, we identified a single case of Sudan Ebola virus disease in Luwero District. The establishment of a permanent in-country laboratory and cooperation between international public health entities facilitated rapid outbreak response and control activities.

Three clusters of organ transplant–associated lymphocytic choriomeningitis virus (LCMV) transmissions have been identified in the United States; 9 of 10 recipients died. In February 2011, we identified a fourth cluster of organ transplant–associated LCMV infections. Diabetic ketoacidosis developed in the organ donor in December 2010; she died with generalized brain edema after a short hospitalization. Both kidneys, liver, and lung were transplanted to 4 recipients; in all 4, severe posttransplant illness developed; 2 recipients died. Through multiple diagnostic methods, we identified LCMV infection in all persons, including in at least 1 sample from the donor and 4 recipients by reverse transcription PCR, and sequences of a 396-bp fragment of the large segment of the virus from all 5 persons were identical. In this cluster, all recipients developed severe illness, but 2 survived. LCMV infection should be considered as a possible cause of severe posttransplant illness.

Seroprevalence of Crimean-Congo hemorrhagic fever virus (CCHFV) is high in some regions of Greece, but only 1 case of disease has been reported. We used 4 methods to test 118 serum samples that were positive for CCHFV IgG by commercial ELISA and confirmed the positive results. A nonpathogenic or low-pathogenicity strain may be circulating.

Next-generation sequencing (NGS) of agents causing idiopathic human diseases has been crucial in the identification of novel viruses. This study describes the isolation and characterization of two novel orthobunyaviruses obtained from a jungle myna and a paddy bird from Karnataka State, India. Using an NGS approach, these isolates were classified as Cat Que and Balagodu viruses belonging to the Manzanilla clade of the Simbu serogroup. Closely related viruses in the Manzanilla clade have been isolated from mosquitos, humans, birds, and pigs across a wide geographic region. Since Orthobunyaviruses exhibit high reassortment frequency and can cause acute, self-limiting febrile illness, these data suggest that human and livestock infections of the Oya/Cat Que/Manzanilla virus may be more widespread and/or under-reported than anticipated. It therefore becomes imperative to identify novel and unknown viruses in order to understand their role in human and animal pathogenesis. The current study is a step forward in this regard and would act as a prototype method for isolation, identification and detection of several other emerging viruses.

BACKGROUND: Marburg virus (MARV), a zoonotic pathogen causing severe hemorrhagic fever in man, has emerged in Angola resulting in the largest outbreak of Marburg hemorrhagic fever (MHF) with the highest case fatality rate to date. METHODOLOGY/PRINCIPAL FINDINGS: A mobile laboratory unit (MLU) was deployed as part of the World Health Organization outbreak response. Utilizing quantitative real-time PCR assays, this laboratory provided specific MARV diagnostics in Uige, the epicentre of the outbreak. The MLU operated over a period of 88 days and tested 620 specimens from 388 individuals. Specimens included mainly oral swabs and EDTA blood. Following establishing on site, the MLU operation allowed a diagnostic response in

Proteolytic processing of the Lassa virus envelope glycoprotein precursor GP-C by the host proprotein convertase site 1 protease (S1P) is a prerequisite for the incorporation of the subunits GP-1 and GP-2 into viral particles and, hence, essential for infectivity and virus spread. Therefore, we tested in this study the concept of using S1P as a target to block efficient virus replication.We demonstrate that stable cell lines inducibly expressing S1P-adapted alpha(1)-antitrypsin variants inhibit the proteolytic maturation of GP-C. Introduction of the S1P recognition motifs RRIL and RRLL into the reactive center loop of alpha(1)-antitrypsin resulted in abrogation of GP-C processing by endogenous S1P to a similar level observed in S1P-deficient cells. Moreover, S1P-specific alpha(1)-antitrypsins significantly inhibited replication and spread of a replication-competent recombinant vesicular stomatitis virus expressing the Lassa virus glycoprotein GP as well as authentic Lassa virus. Inhibition of viral replication correlated with the ability of the different alpha(1)-antitrypsin variants to inhibit the processing of the Lassa virus glycoprotein precursor.Our data suggest that glycoprotein cleavage by S1P is a promising target for the development of novel anti-arenaviral strategies.

BACKGROUND: Zaire ebolavirus (ZEBOV) produces a lethal viral hemorrhagic fever in humans and non-human primates. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrate that the VSVDeltaG/ZEBOVGP vaccine given 28 days pre-challenge either intranasally (IN), orally (OR), or intramuscularly (IM) protects non-human primates against a lethal systemic challenge of ZEBOV, and induces cellular and humoral immune responses. We demonstrated that ZEBOVGP-specific T-cell and humoral responses induced in the IN and OR groups, following an immunization and challenge, produced the most IFN-gamma and IL-2 secreting cells, and long term memory responses. CONCLUSIONS/SIGNIFICANCE: We have shown conclusively that mucosal immunization can protect from systemic ZEBOV challenge and that mucosal delivery, particularly IN immunization, seems to be more potent than IM injection in the immune parameters we have tested. Mucosal immunization would be a huge benefit in any emergency mass vaccination campaign during a natural outbreak, or following intentional release, or for mucosal immunization of great apes in the wild.

Ebola viruses are highly lethal human pathogens that have received considerable attention in recent years due to an increasing re-emergence in Central Africa and a potential for use as a biological weapon. There is no vaccine or treatment licensed for human use. In the past, however, important advances have been made in developing preventive vaccines that are protective in animal models. In this regard, we showed that a single injection of a live-attenuated recombinant vesicular stomatitis virus vector expressing the Ebola virus glycoprotein completely protected rodents and nonhuman primates from lethal Ebola challenge. In contrast, progress in developing therapeutic interventions against Ebola virus infections has been much slower and there is clearly an urgent need to develop effective post-exposure strategies to respond to future outbreaks and acts of bioterrorism, as well as to treat laboratory exposures. Here we tested the efficacy of the vesicular stomatitis virus-based Ebola vaccine vector in post-exposure treatment in three relevant animal models. In the guinea pig and mouse models it was possible to protect 50% and 100% of the animals, respectively, following treatment as late as 24 h after lethal challenge. More important, four out of eight rhesus macaques were protected if treated 20 to 30 min following an otherwise uniformly lethal infection. Currently, this approach provides the most effective post-exposure treatment strategy for Ebola infections and is particularly suited for use in accidentally exposed individuals and in the control of secondary transmission during naturally occurring outbreaks or deliberate release.

BackgroundRecent importation of Lassa fever into Germany, the Netherlands, the United Kingdom, and the United States by travelers on commercial airlines from Africa underscores the public health challenge of emerging viruses. Currently, there are no licensed vaccines for Lassa fever, and no experimental vaccine has completely protected nonhuman primates against a lethal challenge.Methods and findingsWe developed a replication-competent vaccine against Lassa virus based on attenuated recombinant vesicular stomatitis virus vectors expressing the Lassa viral glycoprotein. A single intramuscular vaccination of the Lassa vaccine elicited a protective immune response in nonhuman primates against a lethal Lassa virus challenge. Vaccine shedding was not detected in the monkeys, and none of the animals developed fever or other symptoms of illness associated with vaccination. The Lassa vaccine induced strong humoral and cellular immune responses in the four vaccinated and challenged monkeys. Despite a transient Lassa viremia in vaccinated animals 7 d after challenge, the vaccinated animals showed no evidence of clinical disease. In contrast, the two control animals developed severe symptoms including rashes, facial edema, and elevated liver enzymes, and ultimately succumbed to the Lassa infection.ConclusionOur data suggest that the Lassa vaccine candidate based on recombinant vesicular stomatitis virus is safe and highly efficacious in a relevant animal model that faithfully reproduces human disease.

We report a case of probable Zaire Ebola virus–related ophthalmologic complications in a physician from the United States who contracted Ebola virus disease in Liberia. Uveitis, immune activation, and nonspecific increase in antibody titers developed during convalescence. This case highlights immune phenomena that could complicate management of Ebola virus disease–related uveitis during convalescence.