Hyperspectral Imaging and Machine Learning as a Nondestructive Method for Proso Millet Seed Detection and Classification.

Millet is a small-seeded cereal crop with big potential. There are many different cultivars of proso millet (Panicum miliaceum L.) with different characteristics, bringing forth the issue of sorting which are important for growers, processors, and consumers. Current methods of grain cultivar detection and classification are subjective, destructive, and time-consuming. Therefore, there is a need to develop nondestructive methods for sorting the cultivars of proso millet. In this study, the feasibility of using near-infrared (NIR) hyperspectral imaging (900-1700 nm) to discriminate between different cultivars of proso millet seeds was evaluated. A total of 5000 proso millet seeds were randomly obtained and investigated from the ten most popular cultivars in the United States, namely Cerise, Cope, Earlybird, Huntsman, Minco, Plateau, Rise, Snowbird, Sunrise, and Sunup. To reduce the large dimensionality of the hyperspectral imaging, principal component analysis (PCA) was applied, and the first two principal components were used as spectral features for building the classification models because they had the largest variance. The classification performance showed prediction accuracy rates as high as 99% for classifying the different cultivars of proso millet using a Gradient tree boosting ensemble machine learning algorithm. Moreover, the classification was successfully performed using only 15 and 5 selected spectral features (wavelengths), with an accuracy of 98.14% and 97.6%, respectively. The overall results indicate that NIR hyperspectral imaging could be used as a rapid and nondestructive method for the classification of proso millet seeds.

Inactivation of SARS CoV-2 on porous and nonporous surfaces by compact portable plasma reactor.

We report the inactivation of SARS CoV-2 and its surrogate-Human coronavirus OC43 (HCoV-OC43), on representative porous (KN95 mask material) and nonporous materials (aluminum and polycarbonate) using a Compact Portable Plasma Reactor (CPPR). The CPPR is a compact (48 cm3), lightweight, portable and scalable device that forms Dielectric Barrier Discharge which generates ozone using surrounding atmosphere as input gas, eliminating the need of source gas tanks. Iterative CPPR exposure time experiments were performed on inoculated material samples in 3 operating volumes. Minimum CPPR exposure times of 5-15 min resulted in 4-5 log reduction of SARS CoV-2 and its surrogate on representative material samples. Ozone concentration and CPPR energy requirements for virus inactivation are documented. Difference in disinfection requirements in porous and non-porous material samples is discussed along with initial scaling studies using the CPPR in 3 operating volumes. The results of this feasibility study, along with existing literature on ozone and CPPR decontamination, show the potential of the CPPR as a powerful technology to reduce fomite transmission of enveloped respiratory virus-induced infectious diseases such as COVID-19. The CPPR can overcome limitations of high temperatures, long exposure times, bulky equipment, and toxic residuals related to conventional decontamination technologies.

Retrospective Analysis of Wastewater-Based Epidemiology of SARS-CoV-2 in Residences on a Large College Campus: Relationships between Wastewater Outcomes and COVID-19 Cases across Two Semesters with Different COVID-19 Mitigation Policies.

Wastewater-based epidemiology (WBE) has been utilized for outbreak monitoring and response efforts in university settings during the coronavirus disease 2019 (COVID-19) pandemic. However, few studies examined the impact of university policies on the effectiveness of WBE to identify cases and mitigate transmission. The objective of this study was to retrospectively assess relationships between severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) wastewater outcomes and COVID-19 cases in residential buildings of a large university campus across two academic semesters (August 2020-May 2021) under different COVID-19 mitigation policies. Clinical case surveillance data of student residents were obtained from the university COVID-19 response program. We collected and processed building-level wastewater for detection and quantification of SARS-CoV-2 RNA by RT-qPCR. The odds of obtaining a positive wastewater sample increased with COVID-19 clinical cases in the fall semester (OR = 1.50, P value = 0.02), with higher odds in the spring semester (OR = 2.63, P value < 0.0001). We observed linear associations between SARS-CoV-2 wastewater concentrations and COVID-19 clinical cases (parameter estimate = 1.2, P value = 0.006). Our study demonstrated the effectiveness of WBE in the university setting, though it may be limited under different COVID-19 mitigation policies. As a complementary surveillance tool, WBE should be accompanied by robust administrative and clinical testing efforts for the COVID-19 pandemic response.

Characterization of two novel reassortant bluetongue virus serotype 1 strains isolated from farmed white-tailed deer (Odocoileus virginianus) in Florida, USA.

Hemorrhagic diseases caused by epizootic hemorrhagic disease virus or by bluetongue virus (BTV) are the most important orbivirus diseases affecting ruminants, including white-tailed deer (WTD). Bluetongue virus is of particular concern for farmed WTD in Florida, given its lethality and its wide distribution throughout the state. This study reports the clinical findings, ancillary diagnostics, and genomic characterization of two BTV serotype 1 strains isolated from two farmed WTD, from two different farms in Florida in 2019 and 2022. Phylogenetic and genetic analyses indicated that these two novel BTV-1 strains were reassortants. In addition, our analyses reveal that most genome segments of these strains were acquired from BTVs previously detected in ruminants in Florida, substantiating their endemism in the Southeastern U.S. Our findings underscore the need for additional research to determine the genetic diversity of BTV strains in Florida, their prevalence, and the potential risk of new BTV strains to WTD and other ruminants.

Lack of SARS-CoV-2 in environmental samples collected from September 2020-February 2021 in a university that followed CDC reopening guidance.

This study aimed to provide environmental surveillance data for evaluating the risk of acquiring SARS-CoV-2 in public areas with high foot traffic in a university. Air and surface samples were collected at a university that had the second highest number of COVID-19 cases among public higher education institutions in the U.S. during Fall 2020. A total of 60 samples were collected in 16 sampling events performed during Fall 2020 and Spring 2021. Nearly 9800 students traversed the sites during the study period. SARS-CoV-2 was not detected in any air or surface samples. The university followed CDC guidance, including COVID-19 testing, case investigations, and contact tracing. Students, faculty, and staff were asked to maintain physical distancing and wear face coverings. Although COVID-19 cases were relatively high at the university, the possibility of acquiring SARS-CoV-2 infections at the sites tested was low.

Comparison of oseltamivir and α-galactosylceramide for reducing disease and transmission in pigs infected with 2009 H1N1 pandemic influenza virus.

Influenza virus infections are a major cause of respiratory disease in humans. Neuraminidase inhibitors (NAIs) are the primary antiviral medication used to treat ongoing influenza infections. However, NAIs are not always effective for controlling virus shedding and lung inflammation. Other concerns are the emergence of NAI-resistant virus strains and the risk of side effects, which are occasionally severe. Consequently, additional anti-influenza therapies to replace or combine with NAIs are desirable. Here, we compared the efficacy of the NAI oseltamivir with the invariant natural killer T (iNKT) cell superagonist, α-galactosylceramide (α-GalCer), which induces innate immune responses that inhibit influenza virus replication in mouse models. We show that oseltamivir reduced lung lesions and lowered virus titers in the upper respiratory tract of pigs infected with A/California/04/2009 (CA04) pandemic H1N1pdm09. It also reduced virus transmission to influenza-naïve contact pigs. In contrast, α-GalCer had no impact on virus replication, lung disease, or virus transmission, even when used in combination with oseltamivir. This is significant as iNKT-cell therapy has been studied as an approach for treating humans with influenza.

Assessment of a mass balance equation for estimating community-level prevalence of COVID-19 using wastewater-based epidemiology in a mid-sized city.

Wastewater-based epidemiology (WBE) has emerged as a valuable epidemiologic tool to detect the presence of pathogens and track disease trends within a community. WBE overcomes some limitations of traditional clinical disease surveillance as it uses pooled samples from the entire community, irrespective of health-seeking behaviors and symptomatic status of infected individuals. WBE has the potential to estimate the number of infections within a community by using a mass balance equation, however, it has yet to be assessed for accuracy. We hypothesized that the mass balance equation-based approach using measured SARS-CoV-2 wastewater concentrations can generate accurate prevalence estimates of COVID-19 within a community. This study encompassed wastewater sampling over a 53-week period during the COVID-19 pandemic in Gainesville, Florida, to assess the ability of the mass balance equation to generate accurate COVID-19 prevalence estimates. The SARS-CoV-2 wastewater concentration showed a significant linear association (Parameter estimate = 39.43, P value < 0.0001) with clinically reported COVID-19 cases. Overall, the mass balance equation produced accurate COVID-19 prevalence estimates with a median absolute error of 1.28%, as compared to the clinical reference group. Therefore, the mass balance equation applied to WBE is an effective tool for generating accurate community-level prevalence estimates of COVID-19 to improve community surveillance.

Analysis of Zika Virus Sequence Data Associated with a School Cohort in Haiti.

Zika virus (ZIKV) infections occurred in epidemic form in the Americas in 2014-2016, with some of the earliest isolates in the region coming from Haiti. We isolated ZIKV from 20 children with acute undifferentiated febrile illness who were part of a cohort of children seen at a school clinic in the Gressier region of Haiti. The virus was also isolated from three pools of Aedes aegypti mosquitoes collected at the same location. On phylogenetic analysis, three distinct ZIKV clades were identified. Strains from all three clades were present in Haiti in 2014, making them among the earliest isolates identified in the Western Hemisphere. Strains from all three clades were also isolated in 2016, indicative of their persistence across the time period of the epidemic. Mosquito isolates were collected in 2016 and included representatives from two of the three clades; in one instance, ZIKV was isolated from a pool of male mosquitoes, suggestive of vertical transmission of the virus. The identification of multiple ZIKV clades in Haiti at the beginning of the epidemic suggests that Haiti served as a nidus for transmission within the Caribbean.

Genome Sequence of Genotype 1A Hepatovirus A Isolated from Plasma from a Haitian Child.

Genotype 1A hepatovirus A was identified by quantitative reverse transcription-PCR and isolated from plasma from a Haitian child with acute undifferentiated febrile illness and malaise. The strain was most closely related to Brazilian strains, consistent with recognized patterns of virus movement in the Caribbean region.

Characterization of a Novel Reassortant Epizootic Hemorrhagic Disease Virus Serotype 6 Strain Isolated from Diseased White-Tailed Deer (Odocoileus virginianus) on a Florida Farm.

We report an outbreak of a novel reassortant epizootic hemorrhagic disease virus serotype 6 (EHDV-6) in white-tailed deer (WTD) on a Florida farm in 2019. At necropsy, most animals exhibited hemorrhagic lesions in the lung and heart, and congestion in the lung, liver, and spleen. Histopathology revealed multi-organ hemorrhage and congestion, and renal tubular necrosis. Tissues were screened by RT-qPCR and all animals tested positive for EHDV. Tissues were processed for virus isolation and next-generation sequencing was performed on cDNA libraries generated from the RNA extracts of cultures displaying cytopathic effects. Six isolates yielded nearly identical complete genome sequences of a novel U.S. EHDV-6 strain. Genetic and phylogenetic analyses revealed the novel strain to be most closely related to a reassortant EHDV-6 strain isolated from cattle in Trinidad and both strains received segment 4 from an Australian EHDV-2 strain. The novel U.S. EHDV-6 strain is unique in that it acquired segment 8 from an Australian EHDV-8 strain. An RNAscope® in situ hybridization assay was developed against the novel U.S. EHDV-6 strain and labeling was detected within lesions of the heart, kidney, liver, and lung. These data support the novel U.S. reassortant EHDV-6 strain as the cause of disease in the farmed WTD.

Low-frequency variants in mildly symptomatic vaccine breakthrough infections presents a doubled-edged sword.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOC) has raised questions regarding vaccine protection against SARS-CoV-2 infection, transmission, and ongoing virus evolution. Twenty-three mildly symptomatic "vaccination breakthrough" infections were identified as early as January 2021 in Alachua County, Florida, among individuals fully vaccinated with either the BNT162b2 (Pfizer) or the Ad26 (Janssen/J&J) vaccines. SARS-CoV-2 genomes were successfully generated for 11 of the vaccine breakthroughs, and 878 individuals in the surrounding area and were included for reference-based phylogenetic investigation. These 11 individuals were characterized by infection with VOCs, but also low-frequency variants present within the surrounding population. Low-frequency mutations were observed, which have been more recently identified as mutations of interest owing to their location within targeted immune epitopes (P812L) and association with increased replicative capacity (L18F). We present these results to posit the nature of the efficacy of vaccines in reducing symptoms as both a blessing and a curse-as vaccination becomes more widespread and self-motivated testing reduced owing to the absence of severe symptoms, we face the challenge of early recognition of novel mutations of potential concern. This case study highlights the critical need for continued testing and monitoring of infection and transmission among individuals regardless of vaccination status.

Environmental Surveillance for SARS-CoV-2 in Two Restaurants from a Mid-scale City that Followed U.S. CDC Reopening Guidance.

Since mask use and physical distancing are difficult to maintain when people dine indoors, restaurants are perceived as high risk for acquiring COVID-19. The air and environmental surfaces in two restaurants in a mid-scale city located in north central Florida that followed the Centers for Disease Control and Prevention (CDC) reopening guidance were sampled three times from July 2020 to February 2021. Sixteen air samples were collected for 2 hours using air samplers, and 20 surface samples by using moistened swabs. The samples were analyzed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) for the presence of SARS-CoV-2 genomic RNA. A total of ~550 patrons dined in the restaurants during our samplings. SARS-CoV-2 genomic RNA was not detected in any of the air samples. One of the 20 surface samples (5%) was positive. That sample had been collected from a plastic tablecloth immediately after guests left the restaurant. Virus was not isolated in cell cultures inoculated with aliquots of the RT-PCR-positive sample. The likelihood that patrons and staff acquire SARS-CoV-2 infections may be low in restaurants in a mid-scale city that adopt CDC restaurant reopening guidelines, such as operation at 50% capacity so that tables can be spaced at least 6 feet apart, establishment of adequate mechanical ventilation, use of a face covering except while eating or drinking, and implementation of disinfection measures.

SARS-CoV-2 viroporin encoded by ORF3a triggers the NLRP3 inflammatory pathway.

Heightened inflammatory response is a prominent feature of severe COVID-19 disease. We report that the SARS-CoV-2 ORF3a viroporin activates the NLRP3 inflammasome, the most promiscuous of known inflammasomes. Ectopically expressed ORF3a triggers IL-1ß expression via NFκB, thus priming the inflammasome. ORF3a also activates the NLRP3 inflammasome but not NLRP1 or NLRC4, resulting in maturation of IL-1ß and cleavage/activation of Gasdermin. Notably, ORF3a activates the NLRP3 inflammasome via both ASC-dependent and -independent modes. This inflammasome activation requires efflux of potassium ions and oligomerization between the kinase NEK7 and NLRP3. Importantly, infection of epithelial cells with SARS-CoV-2 similarly activates the NLRP3 inflammasome. With the NLRP3 inhibitor MCC950 and select FDA-approved oral drugs able to block ORF3a-mediated inflammasome activation, as well as key ORF3a amino acid residues needed for virus release and inflammasome activation conserved in the new variants of SARS-CoV-2 isolates across continents, ORF3a and NLRP3 present prime targets for intervention.

SARS-CoV-2 in residential rooms of two self-isolating persons with COVID-19.

Individuals with COVID-19 are advised to self-isolate at their residences unless they require hospitalization. Persons sharing a dwelling with someone who has COVID-19 have a substantial risk of being exposed to the virus. However, environmental monitoring for the detection of virus in such settings is limited. We present a pilot study on environmental sampling for SARS-CoV-2 virions in the residential rooms of two volunteers with COVID-19 who self-quarantined. Apart from standard surface swab sampling, based on availability, four air samplers positioned 0.3-2.2 m from the volunteers were used: a VIable Virus Aerosol Sampler (VIVAS), an inline air sampler that traps particles on polytetrafluoroethylene (PTFE) filters, a NIOSH 2-stage cyclone sampler (BC-251), and a Sioutas personal cascade impactor sampler (PCIS). The latter two selectively collect particles of specific size ranges. SARS-CoV-2 RNA was detected by real-time Reverse-Transcription quantitative Polymerase Chain Reaction (rRT-qPCR) analyses of particles in one air sample from the room of volunteer A and in various air and surface samples from that of volunteer B. The one positive sample collected by the NIOSH sampler from volunteer A's room had a quantitation cycle (Cq) of 38.21 for the N-gene, indicating a low amount of airborne virus [5.69E-02 SARS-CoV-2 genome equivalents (GE)/cm3 of air]. In contrast, air samples and surface samples collected off the mobile phone in volunteer B's room yielded Cq values ranging from 14.58 to 24.73 and 21.01 to 24.74, respectively, on the first day of sampling, indicating that this volunteer was actively shedding relatively high amounts of SARS-CoV-2 at that time. The SARS-CoV-2 GE/cm3 of air for the air samples collected by the PCIS was in the range 6.84E+04 to 3.04E+05 using the LED-N primer system, the highest being from the stage 4 filter, and similarly, ranged from 2.54E+03 to 1.68E+05 GE/cm3 in air collected by the NIOSH sampler. Attempts to isolate the virus in cell culture from the samples from volunteer B's room with the aforementioned Cq values were unsuccessful due to out-competition by a co-infecting Human adenovirus B3 (HAdVB3) that killed the Vero E6 cell cultures within 4 days of their inoculation, although Cq values of 34.56-37.32 were measured upon rRT-qPCR analyses of vRNA purified from the cell culture medium. The size distribution of SARS-CoV-2-laden aerosol particles collected from the air of volunteer B's room was >0.25 µm and >0.1 µm as recorded by the PCIS and the NIOSH sampler, respectively, suggesting a risk of aerosol transmission since these particles can remain suspended in air for an extended time and travel over long distances. The detection of virus in surface samples also underscores the potential for fomite transmission of SARS-CoV-2 in indoor settings.

Wastewater surveillance for SARS-CoV-2 in a small coastal community: Effects of tourism on viral presence and variant identification among low prevalence populations.

Wastewater-based epidemiology has been used to measure SARS-CoV-2 prevalence in cities worldwide as an indicator of community health, however, few longitudinal studies have followed SARS-CoV-2 in wastewater in small communities from the start of the pandemic or evaluated the influence of tourism on viral loads. Therefore the objective of this study was to use measurements of SARS-CoV-2 in wastewater to monitor viral trends and variants in a small island community over a twelve-month period beginning May 1, 2020, before the community re-opened to tourists. Wastewater samples were collected weekly and analyzed to detect and quantify SARS-CoV-2 genome copies. Sanger sequencing was used to determine genome sequences from total RNA extracted from wastewater samples positive for SARS-CoV-2. Visitor data was collected from the local Chamber of Commerce. We performed Poisson and linear regression to determine if visitors to the Cedar Key Chamber of Commerce were positively associated with SARS-CoV-2-positive wastewater samples and the concentration of SARS-CoV-2 RNA. Results indicated that weekly wastewater samples were negative for SARS-CoV-2 until mid-July when positive samples were recorded in four of five consecutive weeks. Additional positive results were recorded in November and December 2020, as well as January, March, and April 2021. Tourism data revealed that the SARS-CoV-2 RNA concentration in wastewater increased by 1.06 Log10 genomic copies/L per 100 tourists weekly. Sequencing from six positive wastewater samples yielded two complete sequences of SARS-CoV-2, two overlapping sequences, and two low yield sequences. They show arrival of a new variant SARS-CoV-2 in January 2021. Our results demonstrate the utility of wastewater surveillance for SARS-CoV-2 in a small community. Wastewater surveillance and viral genome sequencing suggest that population mobility likely plays an important role in the introduction and circulation of SARS-CoV-2 variants among communities experiencing high tourism and who have a small population size.

Independent infections of porcine deltacoronavirus among Haitian children.

Coronaviruses have caused three major epidemics since 2003, including the ongoing SARS-CoV-2 pandemic. In each case, the emergence of coronavirus in our species has been associated with zoonotic transmissions from animal reservoirs1,2, underscoring how prone such pathogens are to spill over and adapt to new species. Among the four recognized genera of the family Coronaviridae, human infections reported so far have been limited to alphacoronaviruses and betacoronaviruses3-5. Here we identify porcine deltacoronavirus strains in plasma samples of three Haitian children with acute undifferentiated febrile illness. Genomic and evolutionary analyses reveal that human infections were the result of at least two independent zoonoses of distinct viral lineages that acquired the same mutational signature in the genes encoding Nsp15 and the spike glycoprotein. In particular, structural analysis predicts that one of the changes in the spike S1 subunit, which contains the receptor-binding domain, may affect the flexibility of the protein and its binding to the host cell receptor. Our findings highlight the potential for evolutionary change and adaptation leading to human infections by coronaviruses outside of the previously recognized human-associated coronavirus groups, particularly in settings where there may be close human-animal contact.

A Valve-Enabled Sample Preparation Device with Isothermal Amplification for Multiplexed Virus Detection at the Point-of-Care.

Early and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza viruses at the point-of-care is crucial for reducing disease transmission during the current pandemic and future flu seasons. To prepare for potential cocirculation of these two viruses, we report a valve-enabled, paper-based sample preparation device integrated with isothermal amplification for their simultaneous detection. The device incorporates (1) virus lysis and RNA enrichment, enabled by ball-based valves for sequential delivery of reagents with no pipet requirement, (2) reverse transcription loop-mediated isothermal amplification, carried out in a coffee mug, and (3) colorimetric detection. We have used the device for simultaneously detecting inactivated SARS-CoV-2 and influenza A H1N1 viruses in 50 min, with limits of detection at 2 and 6 genome equivalents, respectively. The device was further demonstrated to detect both viruses in environmental samples.

Ex vivo SARS-CoV-2 infection of human lung reveals heterogeneous host defense and therapeutic responses.

Cell lines are the mainstay in understanding the biology of COVID-19 infection but do not recapitulate many of the complexities of human infection. The use of human lung tissue is one solution for the study of such novel respiratory pathogens. We hypothesized that a cryopreserved bank of human lung tissue would allow for the ex vivo study of the interindividual heterogeneity of host response to SARS-CoV-2, thus providing a bridge between studies with cell lines and studies in animal models. We generated a cryobank of tissues from 21 donors, many of whom had clinical risk factors for severe COVID-19. Cryopreserved tissues preserved 90% cell viability and contained heterogenous populations of metabolically active epithelial, endothelial, and immune cell subsets of the human lung. Samples were readily infected with HCoV-OC43 and SARS-CoV-2 and demonstrated comparable susceptibility to infection. In contrast, we observed a marked donor-dependent heterogeneity in the expression of IL6, CXCL8, and IFNB1 in response to SARS-CoV-2. Treatment of tissues with dexamethasone and the experimental drug N-hydroxycytidine suppressed viral growth in all samples, whereas chloroquine and remdesivir had no detectable effect. Metformin and sirolimus, molecules with predicted but unproven antiviral activity, each suppressed viral replication in tissues from a subset of donors. In summary, we developed a system for the ex vivo study of human SARS-CoV-2 infection using primary human lung tissue from a library of donor tissues. This model may be useful for drug screening and for understanding basic mechanisms of COVID-19 pathogenesis.

Detection of SARS-CoV-2 in the gastrointestinal tract among patients with negative nasopharyngeal COVID-19 testing prior to endoscopy.

Background and study aims The clinical significance of SARS-CoV-2 RNA in the stool remains unclear. We aimed to determine whether SARS-CoV-2 is detected via real-time reverse transcriptase polymerase chain reaction (rRT-PCR) in the gastrointestinal tracts of patients scheduled for endoscopy and if the virus obtained from these clinical specimens could be isolated in culture. Patients and methods All patients underwent symptom screening and had negative nasopharyngeal testing for SARS-CoV-2 within 72 hours of their scheduled procedure. Study samples were collected via nasopharyngeal swab, rectal swab, and fluid from the upper gastrointestinal tract and/or colon based on their endoscopic procedure(s). Samples were tested for SARS-CoV-2 via rRT-PCR. SARS-CoV-2 positive specimens were isolated and cultured in Vero-E6 cells. Results 243 patients (mean age 63.1 years;54.3 % men) were enrolled from July 15, 2020 to September 2, 2020. SARS-CoV-2 testing was performed from 242 (99.6 %) nasopharyngeal, 243 (100 %) rectal, 183 (75.3 %) upper gastrointestinal tract and 73 (30 %) colon samples. SARS-CoV-2 RNA was detected in the nasopharynx and gastrointestinal specimens in one patient (0.4 %). After a 14-day incubation period, there was no evidence of virus growth in cells incubated with any of these specimens. Conclusions SARS-CoV-2 was rarely detected in the gastrointestinal tract of patients with negative nasopharyngeal testing prior to endoscopy. No live virus was detected by culture, further highlighting that presence of viral genome on its own is not sufficient proof of infectivity. PCR-based screening provides limited insight into virus infectivity and its results should be interpreted carefully as to avoid unnecessary delays in clinical care or inadvertent risk exposure.

Orthobunyaviruses in the Caribbean: Melao and Oropouche virus infections in school children in Haiti in 2014.

We report the identification of two orthobunyaviruses, Melao virus (MELV) and Oropouche virus (OROV), in plasma specimens from Haitian children with acute febrile illness who presented during outbreaks caused by alpha- and flaviviruses in 2014. Heretofore not described as a human pathogen, MELV was isolated in cell culture from the plasma of five case patients. OROV RNA was detected in the plasma of an additional child, using an unbiased sequencing approach, with phylogenetic inference suggesting a close relationship with strains from Brazil. Abdominal pain was reported by four case patients with MELV infections, with lymphadenopathy noted in two cases. Our findings document the occurrence of these orthobunyaviruses within the Caribbean region and highlight the critical importance of surveillance with viral genome sequence analyses to identify outbreaks caused by these and other emerging viruses.