Tick‐borne viruses: Current trends in large‐scale viral surveillance

Ticks are ectoparasites that transmit pathogens, such as tick‐borne viruses, to their hosts. Tick‐borne viruses are diverse: they can be categorized into two orders, nine families, and at least 12 genera. Almost 25% of these viruses are infectious to humans and some are a serious threat to public health. The global rise in tick‐borne virus diseases has been linked to climate change which has reduced tick mortality in the winter and extended their active period. The spread of tick‐borne viral diseases to humans has received significant interest due to the increased threat to human life; epidemiological monitoring of tick‐borne viruses using molecular, immunological, and environmental methods is now a priority. Nevertheless, many tick‐borne diseases remain undiagnosed, which poses a challenge to public administration and health care officials. This review discusses three major tick‐borne RNA viruses that cause serious infection in humans: severe fever with thrombocytopenia syndrome (SFTS) virus, tick‐borne encephalitis (TBE), and Crimean–Congo hemorrhagic fever (CCHF) virus. Specifically, we discuss the epidemiological monitoring, vector control measures, molecular diagnostics, vaccines, and environmental determinants related to these viruses. Furthermore, we review the current surveillance of these tick‐borne viruses with a specific focus on diagnostic approaches that employ molecular interventions such as viral nucleic acid isolation, PCR‐based diagnostics, and high‐throughput sequencing technologies.     

我国西尼罗病毒和Tahyna病毒的发现与流行

已发现100余种蚊传虫媒病毒在世界各地流行,其引发的人兽共患病是全世界关注的公共卫生问题。长期以来我国仅发现乙型脑炎和登革热两种蚊传虫媒病毒病,但近年来新发现西尼罗病毒和Tahyna病毒及其感染疾病流行。从我国新疆维吾尔自治区采集的蚊虫标本中分离到西尼罗病毒,大量血清学研究证明当地不仅存在西尼罗病毒感染所致疾病,还发生过西尼罗病毒感染引发的病毒性脑炎流行。目前已从新疆维吾尔自治区、青海省和内蒙古自治区采集的蚊虫标本中分离到Tahyna病毒,并发现其在自然界动物中的循环和导致的人类感染流行。西尼罗病毒和Tahyna病毒及其相关感染性疾病的发现为我国虫媒病毒及虫媒病毒病的预防与控制提出了新的挑战。     

Arbovirus and insect‐specific virus discovery in Kenya by novel six genera multiplex high‐resolution melting analysis

A broad diversity of arthropod‐borne viruses (arboviruses) of global health concern are endemic to East Africa, yet most surveillance efforts are limited to just a few key viral pathogens. Additionally, estimates of arbovirus diversity in the tropics are likely to be underestimated as their discovery has lagged significantly over past decades due to limitations in fast and sensitive arbovirus identification methods. Here, we developed a nearly pan‐arbovirus detection assay that uses high‐resolution melting (HRM) analysis of RT–PCR products from highly multiplexed assays to differentiate broad diversities of arboviruses. We differentiated 15 viral culture controls and seven additional synthetic viral DNA sequence controls, within Flavivirus, Alphavirus, Nairovirus, Phlebovirus, Orthobunyavirus and Thogotovirus genera. Among Bunyamwera, sindbis, dengue and Thogoto virus serial dilutions, detection by multiplex RT–PCR‐HRM was comparable to the gold standard Vero cell plaque assays. We applied our low‐cost method for enhanced broad‐range pathogen surveillance from mosquito samples collected in Kenya and identified diverse insect‐specific viruses, including a new clade in anopheline mosquitoes, and Wesselsbron virus, an arbovirus that can cause viral haemorrhagic fever in humans and has not previously been isolated in Kenya, in Culex spp. and Anopheles coustani mosquitoes. Our findings demonstrate how multiplex RT–PCR‐HRM can identify novel viral diversities and potential disease threats that may not be included in pathogen detection panels of routine surveillance efforts. This approach can be adapted to other pathogens to enhance disease surveillance and pathogen discovery efforts, as well as the study of pathogen diversity and viral evolutionary ecology.     

Identification of Swedish mosquitoes based on molecular barcoding of the COI gene and SNP analysis

Mosquito‐borne infectious diseases are emerging in many regions of the world. Consequently, surveillance of mosquitoes and concomitant infectious agents is of great importance for prediction and prevention of mosquito‐borne infectious diseases. Currently, morphological identification of mosquitoes is the traditional procedure. However, sequencing of specified genes or standard genomic regions, DNA barcoding, has recently been suggested as a global standard for identification and classification of many different species. Our aim was to develop a genetic method to identify mosquitoes and to study their relationship. Mosquitoes were captured at collection sites in northern Sweden and identified morphologically before the cytochrome c oxidase subunit I (COI) gene sequences of 14 of the most common mosquito species were determined. The sequences obtained were then used for phylogenetic placement, for validation and benchmarking of phenetic classifications and finally to develop a hierarchical PCR‐based typing scheme based on single nucleotide polymorphism sites (SNPs) to enable rapid genetic identification, circumventing the need for morphological characterization. The results showed that exact phylogenetic relationships between mosquito taxa were preserved at shorter evolutionary distances, but at deeper levels, they could not be inferred with confidence using COI gene sequence data alone. Fourteen of the most common mosquito species in Sweden were identified by the SNP/PCR‐based typing scheme, demonstrating that genetic typing using SNPs of the COI gene is a useful method for identification of mosquitoes with potential for worldwide application.     

Dengue and human health: A global scenario of its occurrence,diagnosis and therapeutics

Dengue is one of the highest and rapidly spreading vector-borne viral diseases with high mortality rates. The infection causes acute febrile illness, a major public health concern in the tropics and subtropics globally. The disease is caused by an RNA virus that belongs to the Flaviviridae family. The virus is transferred to humans by the mosquito vector called Aedvrves aegypti, which is the cause of new prevalent sicknesses worldwide. These vector-borne viral diseases spread very fast and pose public health and economic challenges that deemed various prevention and control techniques. The Flavivirus genus consists of five different types of viruses starting from DENV-1 to DENV-5. Thus, the present review focuses on the origin of the virus, how the Dengue virus can be detected, infection, the morphology of the virus, its classifications as proposed by ICTV, the replication and genome of the dengue virus, translation, receptor binding, and some vaccine trial volunteers. In addition, it highlights the current challenges and limitations of effective dengue treatment.     

Development and validation of an arthropod maceration protocol for zoonotic pathogen detection in mosquitoes and fleas

Arthropod‐borne diseases remain a pressing international public health concern. While progress has been made in the rapid detection of arthropod‐borne pathogens via quantitative real‐time (qPCR), or even hand‐held detection devices, a simple and robust maceration and nucleic acid extraction method is necessary to implement biosurveillance capabilities. In this study, a comparison of maceration techniques using five types of beads followed by nucleic acid extraction and detection were tested using two morphologically disparate arthropods, the Aedes aegypti mosquito and Xenopsylla spp. flea, to detect the zoonotic diseases dengue virus serotype‐1 and Yersinia pestis. Post‐maceration nucleic acid extraction was carried out using the 1‐2‐3 Platinum‐Path‐Sample‐Purification (PPSP) kit followed by qPCR detection using the Joint Biological Agent Identification and Diagnostic System (JBAIDS). We found that the 5mm stainless steel beads added to the beads provided in the PPSP kit were successful in macerating the exoskeleton for both Ae. aegypti and Xenopsylla spp. Replicates in the maceration/extraction/detection protocol were increased in a stepwise fashion until a final 128 replicates were obtained. For dengue virus detection there was a 99% positivity rate and for Y. pestis detection there was a 95% positive detection rate. In the examination of both pathogens, there were no significant differences between qPCR instruments, days ran, time of day ran, or operators.     

Donor variability in HIV binding to peripheral blood mononuclear cells

Background

Currently applied indicator organism systems, such as coliforms, are not fully protective of public health from enteric viruses in water sources. Waterborne disease outbreaks have occurred in systems that tested negative for coliforms, and positive coliform results do not necessarily correlate with viral risk. It is widely recognized that bacterial indicators do not co-occur exclusively with infectious viruses, nor do they respond in the same manner to environmental or engineered stressors. Thus, a more appropriate indicator of health risks from infectious enteric viruses is needed.

Presentation of the hypothesis

Torque teno virus is a small, non-enveloped DNA virus that likely exhibits similar transport characteristics to pathogenic enteric viruses. Torque teno virus is unique among enteric viral pathogens in that it appears to be ubiquitous in humans, elicits seemingly innocuous infections, and does not exhibit seasonal fluctuations or epidemic spikes. Torque teno virus is transmitted primarily via the fecal-oral route and can be assayed using rapid molecular techniques. We hypothesize that Torque teno virus is a more appropriate indicator of viral pathogens in drinking waters than currently used indicator systems based solely on bacteria.

Testing the hypothesis

To test the hypothesis, a multi-phased research approach is needed. First, a reliable Torque teno virus assay must be developed. A rapid, sensitive, and specific PCR method using established nested primer sets would be most appropriate for routine monitoring of waters. Because PCR detects both infectious and inactivated virus, an in vitro method to assess infectivity also is needed. The density and occurrence of Torque teno virus in feces, wastewater, and source waters must be established to define spatial and temporal stability of this potential indicator. Finally, Torque teno virus behavior through drinking water treatment plants must be determined with co-assessment of traditional indicators and enteric viral pathogens to assess whether correlations exist.

Implications of the hypothesis

If substantiated, Torque teno virus could provide a completely new, reliable, and efficient indicator system for viral pathogen risk. This indicator would have broad application to drinking water utilities, watershed managers, and protection agencies and would provide a better means to assess viral risk and protect public health.     

Advances in microfluidic PCR for point-of-care infectious disease diagnostics

Global burdens from existing or emerging infectious diseases emphasize the need for point-of-care (POC) diagnostics to enhance timely recognition and intervention. Molecular approaches based on PCR methods have made significant inroads by improving detection time and accuracy but are still largely hampered by resource-intensive processing in centralized laboratories, thereby precluding their routine bedside- or field-use. Microfluidic technologies have enabled miniaturization of PCR processes onto a chip device with potential benefits including speed, cost, portability, throughput, and automation. In this review, we provide an overview of recent advances in microfluidic PCR technologies and discuss practical issues and perspectives related to implementing them into infectious disease diagnostics.     

Mosquito symbioses: from basic research to the paratransgenic control of mosquito‐borne diseases

Mosquito‐borne diseases pose significant concerns in public health. Microbial symbionts of mosquitoes are attracting quite a lot of interest in relation to the development of novel strategies aimed to reduce mosquito vectorial capacity with particular regard to paratransgenesis that relies on genetically modified mosquito symbionts to express molecules within the vector able to interfere with parasite development and transmission. Here, we review the present status of the knowledge of mosquito–symbionts relationships, focusing on perspective in the application of symbiotic control in developing an efficient management of mosquito‐borne diseases.     

DNA microarrays in the clinic: infectious diseases

We argue that the most-promising area of clinical application of microarrays in the foreseeable future is the diagnostics and monitoring of infectious diseases. Microarrays for the detection and characterization of human pathogens have already found their way into clinical practice in some countries. After discussing the persistent, yet often underestimated, importance of infectious diseases for public health, we consider the technologies that are best suited for the detection and clinical investigation of pathogens. Clinical application of microarray technologies for the detection of mycobacteria, Bacillus anthracis, HIV, hepatitis and influenza viruses, and other major pathogens, as well as the analysis of their drug-resistance patterns, illustrate our main thesis.     

综述与专论: 树鼩病毒感染模型的研究新进展

病毒感染引起的疾病接近中国主要传染病发病率的一半,也是传染病致死的主要病因。建立与人类亲缘关系较近、方便有效的感染人类病毒的动物模型,对了解病毒的生物学特性、感染致病机理及制定有效防控措施具有重要意义。树鼩作为灵长类动物的近亲,与人类在生理生化、基因组学等方面的相似性远高于大鼠、小鼠等常用啮齿类实验动物,并具有个体小、便于实验操作、饲养成本低、能感染多种人类病毒等特点,作为动物模型在病毒感染性疾病研究中突显优势和潜能。本文从地区分布、进化、生物学特性等方面,阐述了树鼩作为动物模型应用于病毒感染性疾病研究的优势,包括在乙型肝炎病毒、甲型肝炎病毒,及其他病毒感染疾病研究中的进展。     

Flaviviral disease mosquito vector surveillance in Incheon Metropolitan City and the Hwaseong area,Gyeonggi‐Do,Republic of Korea,in 2015

Owing to global climate change, the global resurgence of vector‐borne infectious diseases and their potential to inflict widespread casualties among human populations has emerged as a pivotal burden on public health systems. In this study, the prevalence of flaviviral diseases transmitted by mosquitoes, and their target vector diversity, abundance, and distribution was investigated to enable the mapping of hotspots for these diseases. For the surveillance of the vector mosquitoes carrying flaviviruses during April to November 2015, female mosquitoes were collected to study whether they carried pathogens from abroad at seven locations in Incheon Metropolitan City (Incheon) as a typical urban area and Hwaseong‐si (= city, Hwaseong) of Gyeonggi‐do (= province) as a rural area. A total of 15 species belonging to seven genera (29,102 female mosquitoes) were collected with black‐light and BG‐Sentinel? traps at a collection rate of 260 per trap/night from whole collection locations. The most collected mosquito species in Incheon were Aedes vexans nipponii (species ratio (SR), 29.9%) and the Culex pipiens complex (SR, 28.8%), followed by Anopheles sinensis s.l. (SR, 27.9%) and Ochlerotatus koreicus (SR, 7.1%). From the results of viral RNA detection, five flaviviruses were found in 20,981 individuals (excluding An. sinensis; 696 pools) in the Cx. pipiens complex and Ae. vexans nipponii. Three Japanese encephalitis virus (JEV)‐positive pools were from the Cx. pipiens complex, a Chaoyang virus pool was found from Ae. vexans nipponii, and the remaining unidentified flavivirus pool was from Cx. pipiens. The three JEV‐positive pools were phylogenetically grouped as genotype V. The results of our study demonstrate that enhanced monitoring and long‐term surveillance of these vector viruses are of great public health importance.     

Update on the proteomics of major arthropod vectors of human and animal pathogens

Vector‐borne diseases (VBDs) are defined as infectious diseases of humans and animals caused by pathogenic agents such as viruses, protists, bacteria, and helminths transmitted by the bite of blood‐feeding arthropod (BFA) vectors. VBDs represent a major public health threat in endemic areas, generally subtropical zones, and many are considered to be neglected diseases. Genome sequencing of some arthropod vectors as well as modern proteomic and genomic technologies are expanding our knowledge of arthropod–pathogen interactions. This review describes the proteomic approaches that have been used to investigate diverse biological questions about arthropod vectors, including the interplay between vectors and pathogens. Proteomic studies have identified proteins and biochemical pathways that may be involved in molecular crosstalk in BFA‐pathogen associations. Future work can build upon this promising start and functional analyses coupled with interactome bioassays will be carried out to investigate the role of candidate peptides and proteins in BFA‐human pathogen associations. Dissection of the host–pathogen interactome will be key to understanding the strategies and biochemical pathways used by BFAs to cope with pathogens.     

Ethical Issues in Field Trials of Genetically Modified Disease‐Resistant Mosquitoes

Mosquito‐borne diseases take a tremendous toll on human populations, especially in developing nations. In the last decade, scientists have developed mosquitoes that have been genetically modified to prevent transmission of mosquito‐borne diseases, and field trials have been conducted. Some mosquitoes have been rendered infertile, some have been equipped with a vaccine they transmit to humans, and some have been designed to resist diseases. This article focuses on ethical issues raised by field trials of disease‐resistant, genetically modified mosquitoes. Some of these issues include: protecting the public and the environment from harm, balancing benefits and risks, collaborating with the local community, avoiding exploitation, and safeguarding the rights and welfare of research subjects. One of the most difficult problems involves protecting the welfare of community members who will be impacted by the release of mosquitoes but who are not enrolled in the study as research subjects. To address this concern, field trials should take place only when the targeted disease is a significant public health problem in an isolated area, the benefits of the trial for the community are likely to outweigh the risks, community leaders approve of the trial, and there are measures in place to protect the welfare of un‐enrolled community members, such as informing the community about the study and offering free treatment to people who contract mosquito‐borne diseases. Since the justification of any field trial depends on a careful examination of the scientific and ethical issues, proposed studies should be evaluated on a case‐by‐case basis.     

Investigation on the virologic vector‐borne diseases by mosquito vector in Yeongnam,Korea

To prevent the influx, propagation and spread of epidemic vectors (vector borne disease) that are increasing due to global warming and climate change, we kept the mosquito vectors under constant surveillance from April (2nd week) to September (3rd week), 2011 in Yeongnam area. Samples were collected at Youngdo (Busan) for Dengue fever and Yellow fever and by support of Gimcheon, Gyeongju, Ulju(Ulsan), Jinju, Hapcheon public health center for Japanese encephalitis and malaria, and also at two sites in Eulsook‐do for Westnile. A BG‐sentinel trap was used to collect samples for Dengue fever and Yellow fever, whereas black light traps were used for Westnile and Japanese encephalitis. At first, mosquito vectors were classified according to a standard classification method and then identified as to whether or not each of the collected samples had the pathogens by using the Real‐time PCR method. Two species of mosquito vectors, Culex pipiens pallens and Ochlerotatus togoi were mostly collected by BG‐sentinel traps. On the other hand, Aedes vexans nipponii and Anopheles sinensis were mainly collected by black light traps (until May). Since then, the number of Culex tritaeniorhynchus gradually increased and then declined in September. The largest number, a total of 12,624 mosquito vectors were collected in the Jinju region during collection. As the result of the Real‐time PCR experiment to detect the pathogens, no viruses had been detected, until now. But it requires sustainable and long‐term monitoring because infectious disease is expected to be spread in the long term.     

植物病毒资源属性和应用基础研究进展

病毒作为地球上最简单的生命形式,通过感染人、动物和植物等寄主产生传染性疾病。与其他微生物相比,病毒具有基因组小、复制量大、遗传操作简单等特点,具有很强的生物资源属性。过去几十年,对植物病毒的研究主要集中于解析其致病机制、植物的抗性机制及如何防控植物病害。但是随着研究的深入及概念的革新,人们发现植物病毒还具有很强的生物资源属性。随着分子生物学以及基因组、转录组、蛋白组学等技术的发展,越来越多的植物病毒被发现、改造和利用。本综述着重围绕植物病毒的资源属性与病毒载体的改造利用及其在生物工程方面的应用等最新研究进展,讨论其广泛的应用前景,挖掘其资源化的潜力。     

Unravelling virus community ecology in bats through the integration of metagenomics and community ecology

The spillover of viruses from wildlife into agricultural animals or humans has profound socioeconomic and public health impact. Vampire bats, found throughout South America, feed directly on humans and other animals and are an important reservoir for zoonotic viruses, including rabies virus. This has resulted in considerable effort in understanding both the ecology of bat‐borne viruses and the composition and associated correlates of the structure of entire virus communities in wildlife, particularly in the context of disease control interventions. In a From the Cover article in this issue of Molecular Ecology, Bergner et al. (2019) set out to reveal virus community dynamics in vampire bats by interrogating factors that affect the structure, diversity and richness of these communities. Due to the linkage of metagenomic sequence data with community ecology, this study represents an important advance in the field of virus ecology.     

Human enteric viruses–potential indicators for enhanced monitoring of recreational water quality

Recreational waters contaminated with human fecal pollution are a public health concern, and ensuring the safety of recreational waters for public use is a priority of both the Environmental Protection Agency (EPA) and the Centers for Disease Control and Prevention (CDC). Current recreational water standards rely on fecal indicator bacteria (FIB) levels as indicators of human disease risk. However present evidence indicates that levels of FIB do not always correspond to the presence of other potentially harmful organisms, such as viruses. Thus, enteric viruses are currently tested as water quality indicators, but have yet to be successfully implemented in routine monitoring of water quality. This study utilized enteric viruses as possible alternative indicators of water quality to examine 18 different fresh and offshore recreational waters on O‘ahu, Hawai‘i, by using newly established laboratory techniques including highly optimized PCR, real time PCR, and viral infectivity assays. All sample sites were detected positive for human enteric viruses by PCR including enterovirus, norovirus genogroups I and II, and male specific FRNA coliphage. A six time-point seasonal study of enteric virus presence indicated significant variation in virus detection between the rainy and dry seasons. Quantitative PCR detected the presence of norovirus genogroup II at levels at which disease risk may occur, and there was no correlation found between enteric virus presence and FIB counts. Under the present laboratory conditions, no infectious viruses were detected from the samples PCR-positive for enteric viruses. These data emphasize both the need for additional indicators for improved monitoring of water quality, and the feasibility of using enteric viruses as these indicators. Electronic Supplementary MaterialSupplementary material is available for this article at 10.1007/s12250-015-3644-x and is accessible for authorized users.     

Clinical applications of molecular biology for infectious diseases

Molecular biological methods for the detection and characterisation of microorganisms have revolutionised diagnostic microbiology and are now part of routine specimen processing. Polymerase chain reaction (PCR) techniques have led the way into this new era by allowing rapid detection of microorganisms that were previously difficult or impossible to detect by traditional microbiological methods. In addition to detection of fastidious microorganisms, more rapid detection by molecular methods is now possible for pathogens of public health importance. Molecular methods have now progressed beyond identification to detect antimicrobial resistance genes and provide public health information such as strain characterisation by genotyping. Treatment of certain microorganisms has been improved by viral resistance detection and viral load testing for the monitoring of responses to antiviral therapies. With the advent of multiplex PCR, real-time PCR and improvements in efficiency through automation, the costs of molecular methods are decreasing such that the role of molecular methods will further increase. This review will focus on the clinical utility of molecular methods performed in the clinical microbiology laboratory, illustrated with the many examples of how they have changed laboratory diagnosis and therefore the management of infectious diseases.     

Developing arbovirus resistance in mosquitoes

Diseases caused by arthropod-borne viruses are increasingly significant public health problems, and novel methods are needed to control pathogen transmission. The hypothesis underlying the research described here is that genetic manipulation of Aedes aegypti mosquitoes can profoundly and permanently reduce their competence to transmit dengue viruses to human hosts. Recent key findings now allow us to test the genetic control hypothesis. We have identified viral genome-derived RNA segments that can be expressed in mosquito midguts and salivary glands to ablate homologous virus replication and transmission. We have demonstrated that both transient and heritable expression of virus-derived effector RNAs in cultured mosquito cells can silence virus replication, and have characterized the mechanism of RNA-mediated resistance. We are now developing virus-resistant mosquito lines by transformation with transposable elements that express effector RNAs from mosquito-active promoters.