Environmental surveillance of SARS-CoV-2 RNA in wastewater systems and related environments in Wuhan: April to May of 2020.

Wastewater-based epidemiology (WBE) has emerged as an effective environmental surveillance tool in monitoring fecal-oral pathogen infections within a community. Congruently, SARS-CoV-2, the etiologic agent of COVID-19, has been demonstrated to infect the gastrointestinal tissues, and be shed in feces. In the present study, SARS-CoV-2 RNA was concentrated from wastewater, sludge, surface water, ground water, sediment, and soil samples of municipal and hospital wastewater systems and related environments in Wuhan during the COVID-19 middle and low risk periods, and the viral RNA copies quantified using reverse transcription quantitative polymerase chain reaction (RT-qPCR). From the findings of this study, during the middle risk period, one influent sample and three secondary effluents collected from waste water treatment plant 2, as well as two samples from Jinyintan Hospital wastewater system influent were SARS-CoV-2 RNA positive. One sludge sample collected from Guanggu Branch of Tongji Hospital, which was obtained during the low risk period, was also positive for SARS-CoV-2 RNA. These study findings demonstrate the significance of WBE in continuous surveillance of SARS-CoV-2 at the community level, even when the COVID-19 prevalence is low. Overall, this study can be used as an important reference for contingency management of wastewater treatment plants and COVID-19 prevention and control departments of Wuhan.

The discovery and global distribution of novel mosquito-associated viruses in the last decade (2007-2017).

In the last decade, virus hunting and discovery has gained pace. This achievement has been driven by three major factors: (a) advancements in sequencing technologies, (b) scaled-up routine arbovirus surveillance strategies, and (c) the "hunt" for emerging pathogens and novel viruses. Many novel viruses have been discovered from a myriad of hosts, vectors, and environmental samples. To help promote understanding of the global diversity and distribution of mosquito-associated viruses and facilitate future studies, we review mosquito-associated viruses discovered between years 2007 and 2017, across the world. In the analyzed period, novel mosquito-associated viruses belonging to 25 families and a general group of unclassified viruses were categorized. The top three discovered novel mosquito-associated viruses belonged to families Flaviviridae (n=32), Rhabdoviridae (n=16), and Peribunyaviridae (n=14). Also, 67 unclassified viruses were reported. Majority of these novel viruses were identified from Culex spp, Anopheles spp, Aedes spp, and Mansonia spp mosquitoes, respectively. Notably, the number of these discovered novels is not representative of intercontinental virus diversity but rather is influenced by the number of studies done in the study period. Some of these newly discovered mosquito-associated viruses have medical significance, either directly or indirectly. For instance, in the study period, 14 novel mosquito-borne viruses that infect mammalian cells in vitro were reported. These viruses pose a danger to the global health security on emerging viral diseases. On the other hand, some of the newly discovered insect specific viruses described herein have potential application as future biocontrol and vaccine agents against known pathogenic arboviruses. Overall, this review outlines the crucial role played by mosquitoes as viral vectors in the global virosphere.

A global dataset of sequence, diversity and biosafety recommendation of arbovirus and arthropod-specific virus.

Arthropod-borne virus (arbovirus) and arthropod-specific virus (ASV) are viruses circulating amongst hematophagous arthropods that are broadly transmitted in ecological systems. Arbovirus may replicate in both vertebrates and invertebrates and some are known to be pathogenic to animals or humans. ASV only replicate in invertebrate arthropods yet they are basal to many types of arboviruses. We built a comprehensive dataset of arbovirus and ASV by curating globally available data from the Arbovirus Catalog, the arbovirus list in Section VIII-F of the Biosafety in Microbiological and Biomedical Laboratories 6th edition, Virus Metadata Resource of International Committee on Taxonomy of Viruses, and GenBank. Revealing the diversity, distribution and biosafety recommendation of arbovirus and ASV at a global scale is essential to the understanding of potential interactions, evolution, and risks associated with these viruses. Moreover, the genomic sequences associated with the dataset will enable the investigation of genetic patterns distinguishing the two groups, as well as aid in predicting the vector/host relationships of the newly discovered viruses.

COVID-19 pandemic: Water, sanitation and hygiene (WASH) as a critical control measure remains a major challenge in low-income countries.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for the deadly respiratory disease called coronavirus disease of 2019 (COVID-19), an ongoing global public health emergency that has been declared a pandemic by the World Health Organization. We review literature on the transmission and control of SARS-CoV-2 and discuss the challenges of focusing on water, sanitation and hygiene (WASH) as critical control measures in low-income countries. A significantly higher prevalence of SARS-CoV-2 infection and COVID-19 related deaths has been reported for the United States of America and other high-income countries in Europe and Asia, regardless of advanced medical facilities in those countries. In contrast, much lower COVID-19 related morbidity and mortality rates have been documented in many low-income countries, despite having comparatively higher socioeconomic burdens and suboptimal medical facilities. By September 29, 2020 over one million deaths have been reported. On the same day, the cumulative total of COVID-19 related morbidity for Africa was 35,954 with 3.5% of the global COVID-19 related deaths. We present arguments for the relatively low COVID-19 morbidity and mortality rates in many low-income countries and discuss the critical importance of WASH for preventing the spread of infectious diseases like COVID-19. We observe that the key recommendations put forward by the World Health Organization to effectively control the pandemic have been difficult to implement in low-income countries. We conclude that the pandemic reinforces previous pronouncements that adequate and effective WASH measures are crucial for public health and recommend closer coordination between public health and WASH sectors.

Zika virus pathogenesis and current therapeutic advances.

Zika virus (ZIKV) is an emerging arthropod-borne flavivirus that, upon infection, results in teratogenic effects and neurological disorders. ZIKV infections pose serious global public health concerns, prompting scientists to increase research on antivirals and vaccines against the virus. These efforts are still ongoing as the pathogenesis and immune evasion mechanisms of ZIKV have not yet been fully elaborated. Currently, no specific vaccines or drugs have been approved for ZIKV; however, some are undergoing clinical trials. Notably, several strategies have been used to develop antivirals, including drugs that target viral and host proteins. Additionally, drug repurposing is preferred since it is less costly and takes less time than other strategies because the drugs used have already been approved for human use. Likewise, different platforms have been evaluated for the design of vaccines, including DNA, mRNA, peptide, protein, viral vectors, virus-like particles (VLPSs), inactivated-virus, and live-attenuated virus vaccines. These vaccines have been shown to induce specific humoral and cellular immune responses and reduce viremia and viral RNA both in vitro and in vivo. Importantly, most of these vaccines have entered clinical trials. Understanding the viral disease mechanism will provide better strategies for developing therapeutic agents against ZIKV. This review provides a comprehensive summary of the viral pathogenesis of ZIKV and current advancements in the development of vaccines and drugs against this virus.

A dataset of distribution and diversity of mosquito-associated viruses and their mosquito vectors in China.

Mosquito-borne viruses such as Zika virus, Japanese Encephalitis virus and Dengue virus present an increasing global health concern. However, in-depth knowledge of the distribution and diversity of mosquito-associated viruses and their related vectors remains limited, especially for China. To promote their understanding, we present the first comprehensive dataset of the distribution and diversity of these viruses and their related vectors in China (including Taiwan, Hong Kong and Macau). Data was drawn from peer-reviewed journal articles, conference papers and thesis publications in both English and Chinese. Geographical data on mosquito-associated viruses' occurrence and related mosquito vector species was extracted, and quality-control processes employed. This dataset contains 2,428 accounts of mosquito-associated viruses' and mosquito species geo-referenced occurrences at various administrative levels in China. The prevalent mosquito-associated virus includes Japanese encephalitis virus, Dengue virus, Banna virus and Culex flavivirus, whereas the abundant mosquito vectors are Culex tritaeryohynchus, Aedes albopictus and Culex pipiens pallens. This geographical dataset delivers a distribution and diversity outline of mosquito-associated viruses in China, and also applicable in various spatial and risk-assessment analysis.

Author Correction: A dataset of distribution and diversity of mosquito-associated viruses and their mosquito vectors in China.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Stability of the Virome in Lab- and Field-Collected Aedes albopictus Mosquitoes across Different Developmental Stages and Possible Core Viruses in the Publicly Available Virome Data of Aedes Mosquitoes.

Aedes mosquitoes can efficiently transmit many pathogenic arboviruses, placing a great burden on public health worldwide. In addition, they also carry a number of insect-specific viruses (ISVs), and it was recently suggested that some of these ISVs might form a stable species-specific "core virome" in mosquito populations. However, little is known about such a core virome in laboratory colonies and if it is present across different developmental stages. In this study, we compared the viromes in eggs, larvae, pupae, and adults of Aedes albopictus mosquitoes collected from a lab colony and compared each to the virome of different developmental stages collected in the field. The virome in lab-derived A. albopictus was very stable across all stages, consistent with a vertical transmission route of these viruses, and formed a possible "vertically transmitted core virome." The different stages of field-collected A. albopictus mosquitoes also contained this stable vertically transmitted core virome, as well as another set of viruses (e.g., viruses distantly related to Guadeloupe mosquito virus, Hubei virga-like virus 2, and Sarawak virus) shared by mosquitoes across different stages, which might represent an "environment-derived core virome." To further study this core set of ISVs, we screened 48 publicly available SRA viral metagenomic data sets of mosquitoes belonging to the genus Aedes, showing that some of the identified ISVs were identified in the majority of SRAs and providing further evidence supporting the core-virome concept.IMPORTANCE Our study revealed that the virome was very stable across all developmental stages of both lab-derived and field-collected Aedes albopictus The data representing the core virome in lab A. albopictus proved the vertical transmission route of these viruses, forming a "vertically transmitted core virome." Field mosquitoes also contained this stable vertically transmitted core virome as well as additional viruses, which probably represented "environment-derived core virome" and which therefore were less stable over time and geography. By further screening publicly available SRA viral metagenomic data sets from mosquitoes belonging to the genus Aedes, some of the identified core ISVs were shown to be present in the majority of SRAs, such as Phasi Charoen-like phasivirus and Guadeloupe mosquito virus. How these core ISVs influence the biology of the mosquito host and arbovirus infection and evolution deserves to be further explored.

Pathogenesis and Immune Response of Ebinur Lake Virus: A Newly Identified Orthobunyavirus That Exhibited Strong Virulence in Mice.

Orthobunyaviruses are a group of viruses with significant public and veterinary health importance. These viruses are mainly transmitted through mosquito-, midge-, and tick-vectors, and are endemic to various regions of the world. Ebinur Lake virus (EBIV), a newly identified member of Orthobunyavirus, was isolated from Culex mosquitoes in Northwest China. In the present study, we aimed to characterize the pathogenesis and host immune responses of EBIV in BALB/c mice, as an animal model. Herein, we determined that BALB/c mice are highly susceptible to EBIV infection. The infected mice exhibited evident clinical signs including weight loss, mild encephalitis, and death. High mortality of mice was observed even with inoculation of one plaque-forming unit (PFU) of EBIV, and the infected mice succumbed to death within 5-9 days. After EBIV challenge, rapid viremic dissemination was detected in the peripheral tissues and the central nervous system, with prominent histopathologic changes observed in liver, spleen, thymus, and brain. Blood constituents' analysis of EBIV infected mice exhibited leukopenia, thrombocytopenia, and significantly elevated ALT, LDH-L, and CK. Further, EBIV infection induced obvious cytokines changes in serum, spleen, and brain in mice. Collectively, our data describe the first study that systematically examines the pathogenesis of EBIV and induced immune response in an immunocompetent standard mouse model, expanding our knowledge of this virus, which may pose a threat to One Health.

Mosquitoes of Etiological Concern in Kenya and Possible Control Strategies.

Kenya is among the most affected tropical countries with pathogen transmitting Culicidae vectors. For decades, insect vectors have contributed to the emergence and distribution of viral and parasitic pathogens. Outbreaks and diseases have a great impact on a country's economy, as resources that would otherwise be used for developmental projects are redirected to curb hospitalization cases and manage outbreaks. Infected invasive mosquito species have been shown to increasingly cross both local and global boarders due to the presence of increased environmental changes, trade, and tourism. In Kenya, there have been several mosquito-borne disease outbreaks such as the recent outbreaks along the coast of Kenya, involving chikungunya and dengue. This certainly calls for the implementation of strategies aimed at strengthening integrated vector management programs. In this review, we look at mosquitoes of public health concern in Kenya, while highlighting the pathogens they have been linked with over the years and across various regions. In addition, the major strategies that have previously been used in mosquito control and what more could be done to reduce or combat the menace caused by these hematophagous vectors are presented.

Characterization of a Novel Tanay Virus Isolated From Anopheles sinensis Mosquitoes in Yunnan, China.

Globally, mosquitoes are known to be competent vectors to various arboviruses that cause serious and debilitating diseases to humans and animals. Conversely, mosquitoes harbor a wide array of insect specific viruses (ISVs) that are generally neglected. Extensive characterization of these ISVs is important in understanding their persistence infection effect on host behavior and arbovirus transmission. Herein, we report first time isolation of Tanay virus (TANAV) isolate YN15_103_01 in Anopheles sinensis mosquitoes from Yunnan Province, China. Phylogenetically, the isolate's nucleotide identity had more than 14.47% variance compared to previous TANAV isolates, and it clustered into an independent branch within the genus Sandewavirus in the newly proposed taxon Negevirus. TANAV growth and high titers was attained in Aag2 cells (107 PFU/mL) but with no CPE observed up to 7 days.p.i. compared to C6/36 cells that exhibited extensive CPE at 48 h.p.i. with titers of 107 PFU/mL. Contrarywise, the viral isolate did not replicate in vertebrate cell lines. Electron microscopy analyses showed that its final maturation process takes place in the cell cytoplasm. Notably, the predicted viral proteins were verified to be corresponding to the obtained SDS-PAGE protein bands. Our findings advance forth new and vital knowledge important in understanding insect specific viruses, especially TANAV.

Characterization of Ebinur Lake Virus and Its Human Seroprevalence at the China-Kazakhstan Border.

In recent years, rapidly increasing trade and travel across the China-Kazakhstan border has increased the potential risk of the introduction and exportation of vectors and their related diseases. The Ebinur Lake Nature Reserve is located in Xinjiang Uygur Autonomous Region, near the China-Kazakhstan border, with a suitable ecosystem for mosquito breeding. In our previous work, a novel Orthobunyavirus species named Ebinur Lake virus (EBIV) was isolated in the reserve. To gain insights into the potential risk of EBIV in this region, we conducted a study that aimed to clearly outline EBIV's biological characteristics and its human seroprevalence in this region. Phylogenetically, the analysis of all three segments of EBIV demonstrated that it belongs to the genus Orthobunyavirus, which is clustered in the Bunyamwera serogroup. EBIV replicated efficiently and caused cytopathic effects (CPEs) in vertebrate cells. The survival rates of the EBIV-challenged mice were 0 and 20% when inoculated with viral concentrations ≥104 or 102 plaque-forming units, respectively. For EBIV-infected mice, internal bleeding and pathological changes were observed. In addition, the overall immunoglobulin M (IgM) antibody [1:4 by immunofluorescence assay (IFA)], immunoglobulin G (IgG) antibody (1:10 by IFA), and neutralizing antibody [90% plaque reduction neutralization test (PRNT)] prevalence was 8.05, 12.3, and 0.95%, respectively, in the studied residents. In summary, EBIV is a new member of the Bunyamwera serogroup and is able to competently infect cells derived from mosquitoes, rodents, monkeys, or humans. Furthermore, EBIV caused severe disease and even death in challenged Kunming mice, and the antibodies against EBIV have been detected in local residents, indicating that the virus is a potential animal or human pathogen.

First Isolation and Characterization of a Group C Banna Virus (BAV) from Anopheles sinensis Mosquitoes in Hubei, China.

Banna virus (BAV) is considered to be an emerging human pathogen that is transmitted by blood-sucking insects. BAV was isolated from various species of mosquitoes, midges, and livestock. It is widely distributed geographically, since it was identified in China, Vietnam, and Indonesia. Previously reported evolution studies of BAV indicated that BAV can be divided into two groups, including isolates from China and Vietnam clustered in group A, and Indonesian isolates in group B. In this study, we report the isolation of a new strain of BAV named HB14-71-01 from Anopheles sinensis mosquitoes from Hubei, China. An in vitro comparison study of the HB14-71-01 isolate and the group A BAV revealed differences based on observed cytopathic effect, plaque size, and viral growth rates. Additionally, the phylogenetic analysis indicated that the Hubei isolate belongs to a novel genotype of BAV and emerged nearly 105 years ago (95% highest posterior density (HPD): 35⁻434), unlike the two previously reported genotypes A and B. Our findings extend the knowledge about the genomic diversity and potential vectors/hosts of BAVs and will improve understanding of the relationships between genetic variation and pathogenicity.

Comparative Metagenomic Profiling of Viromes Associated with Four Common Mosquito Species in China.

Vast viruses are thought to be associated with mosquitoes. Anopheles sinensis, Armigeres subalbatus, Culex quinquefasciatus, and Culex tritaeniorhynchus are very common mosquito species in China, and whether the virome structure in each species is species-specific has not been evaluated. In this study, a total of 2222 mosquitoes were collected from the same geographic location, and RNAs were sequenced using the Illumina Miseq platform. After querying to the Refseq database, a total of 3,435,781, 2,223,509, 5,727,523, and 6,387,867 paired-end reads were classified under viral sequences from An. sinensis, Ar. subalbatus, Cx. quinquefasciatus, and Cx. tritaeniorhynchus, respectively, with the highest prevalence of virus-associated reads being observed in Cx. quinquefasciatus. The metagenomic comparison analysis showed that the virus-related reads were distributed across 26 virus families, together with an unclassified group of viruses. Anelloviridae, Circoviridae, Genomoviridae, Iridoviridae, Mesoniviridae, Microviridae, Myoviridae, Parvoviridae, Phenuiviridae, and Podoviridae were the top ten significantly different viral families among the four species. Further analysis reveals that the virome is species-specific in four mosquito samples, and several viral sequences which maybe belong to novel viruses are discovered for the first time in those mosquitoes. This investigation provides a basis for a comprehensive knowledge on the mosquito virome status in China.

Mosquito-Associated Viruses in China.

Mosquitoes are classified into approximately 3500 species and further grouped into 41 genera. Epidemiologically, they are considered to be among the most important disease vectors in the world and they can harbor a wide variety of viruses. Several mosquito viruses are considered to be of significant medical importance and can cause serious public health issues throughout the world. Such viruses are Japanese encephalitis virus (JEV), dengue virus (DENV), chikungunya virus (CHIKV), and Zika virus (ZIKV). Others are the newly recognized mosquito viruses such as Banna virus (BAV) and Yunnan orbivirus (YNOV) with unclear medical significance. The remaining mosquito viruses are those that naturally infect mosquitoes but do not appear to infect humans or other vertebrates. With the continuous development and improvement of mosquito and mosquito-associated virus surveillance systems in China, many novel mosquito-associated viruses have been discovered in recent years. This review aims to systematically outline the history, characteristics, distribution, and/or current epidemic status of mosquito-associated viruses in China.

Metagenomic Virome Analysis of Culex Mosquitoes from Kenya and China.

Many blood-feeding arthropods are known vectors of viruses that are a source of unprecedented global health concern. Mosquitoes are an integral part of these arthropod vectors. Advancements in next-generation sequencing and bioinformatics has expanded our knowledge on the richness of viruses harbored by arthropods. In the present study, we applied a metagenomic approach to determine the intercontinental virome diversity of Culex quinquefasciatus and Culex tritaeniorhynchus in Kwale, Kenya and provinces of Hubei and Yunnan in China. Our results showed that viromes from the three locations were strikingly diverse and comprised 30 virus families specific to vertebrates, invertebrates, plants, and protozoa as well as unclassified group of viruses. Though sampled at different times, both Kwale and Hubei mosquito viromes were dominated by vertebrate viruses, in contrast to the Yunnan mosquito virome, which was dominated by insect-specific viruses. However, each virome was unique in terms of virus proportions partly influenced by type of ingested meals (blood, nectar, plant sap, environment substrates). The dominant vertebrate virus family in the Kwale virome was Papillomaviridae (57%) while in Hubei it was Herpesviridae (30%) and the Yunnan virome was dominated by an unclassified viruses group (27%). Given that insect-specific viruses occur naturally in their hosts, they should be the basis for defining the viromes. Hence, the dominant insect-specific viruses in Kwale, Hubei, and Yunnan were Baculoviridae, Nimaviridae and Iflaviridae, respectively. Our study is preliminary but contributes to growing and much needed knowledge, as mosquito viromes could be manipulated to prevent and control pathogenic arboviruses.