Prolonged Viability of Senecavirus A in Exposed House Flies (Musca domestica).

House flies (Musca domestica) are often present in swine farms worldwide. These flies utilize animal secretions and waste as a food source. House flies may harbor and transport microbes and pathogens acting as mechanical vectors for diseases. Senecavirus A (SVA) infection in pigs occurs via oronasal route, and animals shed high virus titers to the environment. Additionally, SVA possesses increased environmental resistance. Due to these reasons, we investigated the tenacity of SVA in house flies. Five groups of flies, each composed of ten females and ten males, were exposed to SVA, titer of 109.3 tissue culture infectious dose (TCID50/mL). Groups of male and female flies were collected at 0, 6, 12, 24, and 48 h post-exposure. For comparison purposes, groups of flies were exposed to Swinepox virus (SwPV). Infectious SVA was identified in all tested groups. Successful isolation of SVA demonstrated the titers varied between 106.8 and 102.8 TCID50/mL in female groups and varied from 105.85 to 103.8 TCID50/mL in male groups. In contrast, infectious SwPV was only detected in the female group at 6 h. The significant SVA infectious titer for prolonged periods of time, up to 48 h, indicates a potential role of flies in SVA transmission.

Bacteriophage: A Useful Tool for Studying Gut Bacteria Function of Housefly Larvae, Musca domestica.

Beneficial symbiotic bacteria have positive effects on some insects' (such as mosquitoes, cockroaches, and flies) biological activities. However, the effects of a lack of one specific symbiotic bacterium on the life activities of some insects and their natural gut microbiota composition remain unclear. Bacteriophages are viruses that specifically target and kill bacteria and have the potential to shape gut bacterial communities. In previous work, Pseudomonas aeruginosa that naturally colonized the intestines of housefly larvae was shown to be essential to protect housefly larvae from entomopathogenic fungal infections, leading us to test whether a deficiency in Pseudomonas aeruginosa strains in housefly larvae that was specifically caused using bacteriophages could remold the composition of the intestinal bacteria and affect the development of housefly larvae. Our research revealed that the phage, with a titer of 108 PFU/ml, can remove 90% of Pseudomonas aeruginosa in the gut. A single feeding of low-dose phage had no effect on the health of housefly larvae. However, the health of housefly larvae was affected by treatment with phage every 24 h. Additionally, treating housefly larvae with bacteriophages every 24 h led to bacterial composition changes in the gut. Collectively, the results revealed that deficiency in one symbiotic gut bacteria mediated by precise targeting using bacteriophages indirectly influences the intestinal microbial composition of housefly larvae and has negative effects on the development of the host insect. Our results indicated the important role of symbiotic gut bacteria in shaping the normal gut microbiota composition in insects. IMPORTANCE The well-balanced gut microbiota ensures appropriate development of the host insect, such as mosquitoes, cockroaches, and flies. Various intestinal symbiotic bacteria have different influences on the host gut community structure and thus exert different effects on host health. Therefore, it is of great importance to understand the contributions of one specific bacterial symbiont to the gut microbiota community structure and insect health. Bacteriophages that target certain bacteria are effective tools that can be used to analyze gut bacterial symbionts. However, experimental evidence for phage efficacy in regulating insect intestinal bacteria has been little reported. In this study, we used phages as precision tools to regulate a bacterial community and analyzed the influence on host health after certain bacteria were inhibited by bacteriophage. The ability of phages to target intestinal-specific bacteria in housefly larvae and reduce the levels of target bacteria makes them an effective tool for studying the function of gut bacteria.

First molecular-based detection of SARS-CoV-2 virus in the field-collected houseflies.

This is the first report of SARS-CoV-2 detection on field-collected Musca domestica housefly surface and tissue samples using the high-sensitive PCR assay which suggests the possible insect-borne transmission. The study was conducted in Shiraz city, southern Iran, in May and Jun 2020. Adult flies were sampled at the outdoor areas of two hospitals treating COVID-19 patients. Fly samples were first washed twice to remove the insect surface attached to SARS-CoV-2 virions. After that, the disinfected fly samples were homogenized. Fly surface washout and homogenate samples were tested using Taq Man real-time PCR assay for the SARS-CoV-2 virus. In a total of 156 houseflies, 75% of samples from the body washout samples were positive for SARS-CoV-2. Strikingly, 37% of the homogenized specimens were positive for the SARS-CoV-2, suggesting the possible infection of the insects or uptake of the virion to the insect metabolism. The other possibility is the houseflies up took the blood or blood fluids of the patients and the RNA of the SARS-CoV-2 survived in the insect body without replicating. Our preliminary findings suggest that the houseflies could transmit SARS-CoV-2 as a mechanical or biological vector especially during the warm seasons while increasing the population and activity of houseflies.

The Effect of the Hypertrophy Virus (MdSGHV) on the Ultrastructure of the Salivary Glands of Musca domestica (Diptera: Muscidae).

The salivary glands of insects play a key role in the replication cycle and vectoring of viral pathogens. Consequently, Musca domestica (L.) (Diptera: Muscidae) and the Salivary Gland Hypertrophy Virus (MdSGHV) serve as a model to study insect vectoring of viruses. A better understanding of the structural changes of the salivary glands by the virus will help obtain a better picture of the pathological impact the virus has on adult flies. The salivary glands are a primary route for viruses to enter a new host. As such, studying the viral effect on the salivary glands is particularly important and can provide insights for the development of strategies to control the transmission of vector-borne diseases, such as dengue, malaria, Zika, and chikungunya virus. Using scanning and transmission electron microscopic techniques, researchers have shown the effects of infection by MdSGHV on the salivary glands; however, the exact location where the infection was found is unclear. For this reason, this study did a close examination of the effects of the hypertrophy virus on the salivary glands to locate the specific sites of infection. Here, we report that hypertrophy is present mainly in the secretory region, while other regions appeared unaffected. Moreover, there is a disruption of the cuticular, chitinous lining that separates the secretory cells from the lumen of the internal duct, and the disturbance of this lining makes it possible for the virus to enter the lumen. Thus, we report that the chitinous lining acts as an exit barrier of the salivary gland.

Mechanical transmission of SARS-CoV-2 by house flies.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a recently emerged coronavirus that is the causative agent of the coronavirus disease 2019 (COVID-19) pandemic. COVID-19 in humans is characterized by a wide range of symptoms that range from asymptomatic to mild or severe illness including death. SARS-CoV-2 is highly contagious and is transmitted via the oral-nasal route through droplets and aerosols, or through contact with contaminated fomites. House flies are known to transmit bacterial, parasitic and viral diseases to humans and animals as mechanical vectors. Previous studies have shown that house flies can mechanically transmit coronaviruses, such as turkey coronavirus; however, the house fly's role in SARS-CoV-2 transmission has not yet been explored. The goal of this work was to investigate the potential of house flies to mechanically transmit SARS-CoV-2. For this purpose, it was determined whether house flies can acquire SARS-CoV-2, harbor live virus and mechanically transmit the virus to naive substrates and surfaces. METHODS: Two independent studies were performed to address the study objectives. In the first study, house flies were tested for infectivity after exposure to SARS-CoV-2-spiked medium or milk. In the second study, environmental samples were tested for infectivity after contact with SARS-CoV-2-exposed flies. During both studies, samples were collected at various time points post-exposure and evaluated by SARS-CoV-2-specific RT-qPCR and virus isolation. RESULTS: All flies exposed to SARS-CoV-2-spiked media or milk substrates were positive for viral RNA at 4 h and 24 h post-exposure. Infectious virus was isolated only from the flies exposed to virus-spiked milk but not from those exposed to virus-spiked medium. Moreover, viral RNA was detected in environmental samples after contact with SARS-CoV-2 exposed flies, although no infectious virus was recovered from these samples. CONCLUSIONS: Under laboratory conditions, house flies acquired and harbored infectious SARS-CoV-2 for up to 24 h post-exposure. In addition, house flies were able to mechanically transmit SARS-CoV-2 genomic RNA to the surrounding environment up to 24 h post-exposure. Further studies are warranted to determine if house fly transmission occurs naturally and the potential public health implications of such events.

The Effect of the Musca domestica Salivary Gland Hypertrophy Virus on Food Consumption in Its Adult Host, the Common House Fly (Diptera: Muscidae).

The Musca domestica salivary gland hypertrophy virus (MdSGHV) substantially enlarges the house fly's salivary glands and prevents or delays ovarian development in its adult host, but the effect that MdSGHV has on the house fly's food consumption is currently unknown. Using house flies from a laboratory-reared colony, we evaluated the effect of MdSGHV infection on food consumption over a 7-d period. Both treatment (virus-infected) and control (saline-injected) flies were provided with a choice of 8% sucrose solution and 4% powdered milk solution to determine food preferences. Quantities of each solution consumed were measured every 24 h for each fly to measure food consumptions. Infected house flies were shown to consume less overall of both solutions than house flies injected with saline. The largest consumption discrepancy was seen between female house flies. Healthy female flies with developing ovaries continued to consume a sugar and protein diet, whereas infected female flies fed predominantly on a sugar diet. Additionally, infected male and female flies consumed significantly lower quantities of protein and sucrose than control flies. This suggests that MdSGHV has a negative consumption effect (e.g., hunger, starvation) on its host. Thus, differences in food consumption of infected and control flies probably represent differences in the nutritional requirements of flies resulting from viral infection.

Trace amounts of African swine fever virus DNA detected in insects collected from an infected pig farm in Estonia.

BACKGROUND: African swine fever (ASF), a severe multi-systemic disease in pigs, was introduced into Estonia in 2014. The majority of outbreaks have occurred during the summer months. Given that ASFV is transmitted in a sylvatic cycle that includes the transmission by African soft ticks and that mechanical transmission by flying insects was shown, transmission by other arthropod vectors need to be considered. OBJECTIVES: Here, we report the results of a pilot study on flying insects caught on an outbreak farm during epidemiological investigations. METHODS: In brief, 15 different insect species (flies and mosquitoes) were collected by random catch using an aerial net. Nucleic acids derived from these samples or their pools were tested for African swine fever virus (ASFV) DNA by real-time PCR. RESULTS AND CONCLUSIONS: Viral DNA was detected in small quantities in two samples from flies and mosquitoes. Given the slow spread of virus within the farm, the impact of these findings seems rather low, but a role in local transmission cannot be ruled out. However, given the very low number of insects sampled, and taken into the account that viral isolation was not performed and insects outside the farm were not investigated, future investigations are needed to assess the true impact of insects as mechanical vectors.

In vitro Acquisition and Retention of Low-Pathogenic Avian Influenza H9N2 by Musca domestica (Diptera: Muscidae).

Avian influenza virus (AIV) H9N2 emerged in the 1990s as an economically important disease in poultry and occasionally infects humans and other mammals. The aim of this study was to evaluate the acquisition and retention of H9N2 AIV on and within the house fly, Musca domestica (Linnaeus 1758), under laboratory conditions. In first experiment, 100 adult house flies were divided into control and treatment groups equally. Treatment group was fed with a meal containing H9N2 virus, while control group was supplied with an identical meal without virus. Fifteen minutes after exposure in each group, flies were washed twice to remove surface particles, disinfected and then homogenized for testing. The two external body surface washes and the homogenate samples were tested for H9N2 to distinguish exterior from interior viral load. Second experiment was performed likewise but five flies from each group were taken at 0, 6, 24, 48, 72, 96, and 120 h post-exposure. All samples were subjected to real-time reverse-transcription polymerase chain reaction (RRT-PCR) for detecting H9-Specific viral RNA. Results of the first experiment showed that viral RNA was detectable in both of external surface and homogenates samples. Second experiment revealed that persistence of H9N2 AIVs on external body surface and within the body of M. domestica were 24 and 96 h, respectively. Moreover, viral RNAs concentration declined during the time after exposure to AIV H9N2 either outside or within house flies. Overall, house fly was able to acquire and preserve H9N2 AIV experimentally, which may contribute the spread of virus among poultry farms.

Transmission of lumpy skin disease virus: A short review.

Lumpy skin disease (LSD) is a viral transboundary disease endemic throughout Africa and of high economic importance that affects cattle and domestic water buffaloes. Since 2012, the disease has spread rapidly and widely throughout the Middle Eastern and Balkan regions, southern Caucasus and parts of the Russian Federation. Before vaccination campaigns took their full effect, the disease continued spreading from region to region, mainly showing seasonal patterns despite implementing control and eradication measures. The disease is capable of appearing several hundred kilometers away from initial (focal) outbreak sites within a short time period. These incursions have triggered a long-awaited renewed scientific interest in LSD resulting in the initiation of novel research into broad aspects of the disease, including epidemiology, modes of transmission and associated risk factors. Long-distance dispersal of LSDV seems to occur via the movement of infected animals, but distinct seasonal patterns indicate that arthropod-borne transmission is most likely responsible for the swift and aggressive short-distance spread of the disease. Elucidating the mechanisms of transmission of LSDV will enable the development of more targeted and effective actions for containment and eradication of the virus. The mode of vector-borne transmission of the disease is most likely mechanical, but there is no clear-cut evidence to confirm or disprove this assumption. To date, the most likely vectors for LSDV transmission are blood-sucking arthropods such as stable flies (Stomoxys calcitrans), mosquitoes (Aedes aegypti), and hard ticks (Rhipicephalus and Amblyomma species). New evidence suggests that the ubiquitous, synanthropic house fly, Musca domestica, may also play a role in LSDV transmission, but this has not yet been tested in a clinical setting. The aim of this review is to compile and discuss the earlier as well as the most recent research data on the transmission of LSDV.

Coevolution of hytrosaviruses and host immune responses.

BACKGROUND: Hytrosaviruses (SGHVs; Hytrosaviridae family) are double-stranded DNA (dsDNA) viruses that cause salivary gland hypertrophy (SGH) syndrome in flies. Two structurally and functionally distinct SGHVs are recognized; Glossina pallidipes SGHV (GpSGHV) and Musca domestica SGHV (MdSGHV), that infect the hematophagous tsetse fly and the filth-feeding housefly, respectively. Genome sizes and gene contents of GpSGHV (~ 190 kb; 160-174 genes) and MdSGHV (~ 124 kb; 108 genes) may reflect an evolution with the SGHV-hosts resulting in differences in pathobiology. Whereas GpSGHV can switch from asymptomatic to symptomatic infections in response to certain unknown cues, MdSGHV solely infects symptomatically. Overt SGH characterizes the symptomatic infections of SGHVs, but whereas MdSGHV induces both nuclear and cellular hypertrophy (enlarged non-replicative cells), GpSGHV induces cellular hyperplasia (enlarged replicative cells). Compared to GpSGHV's specificity to Glossina species, MdSGHV infects other sympatric muscids. The MdSGHV-induced total shutdown of oogenesis inhibits its vertical transmission, while the GpSGHV's asymptomatic and symptomatic infections promote vertical and horizontal transmission, respectively. This paper reviews the coevolution of the SGHVs and their hosts (housefly and tsetse fly) based on phylogenetic relatedness of immune gene orthologs/paralogs and compares this with other virus-insect models. RESULTS: Whereas MdSGHV is not vertically transmitted, GpSGHV is both vertically and horizontally transmitted, and the balance between the two transmission modes may significantly influence the pathogenesis of tsetse virus. The presence and absence of bacterial symbionts (Wigglesworthia and Sodalis) in tsetse and Wolbachia in the housefly, respectively, potentially contributes to the development of SGH symptoms. Unlike MdSGHV, GpSGHV contains not only host-derived proteins, but also appears to have evolutionarily recruited cellular genes from ancestral host(s) into its genome, which, although may be nonessential for viral replication, potentially contribute to the evasion of host's immune responses. Whereas MdSGHV has evolved strategies to counteract both the housefly's RNAi and apoptotic responses, the housefly has expanded its repertoire of immune effector, modulator and melanization genes compared to the tsetse fly. CONCLUSIONS: The ecologies and life-histories of the housefly and tsetse fly may significantly influence coevolution of MdSGHV and GpSGHV with their hosts. Although there are still many unanswered questions regarding the pathogenesis of SGHVs, and the extent to which microbiota influence expression of overt SGH symptoms, SGHVs are attractive 'explorers' to elucidate the immune responses of their hosts, and the transmission modes of other large DNA viruses.

Lesser housefly (Fannia canicularis) as possible mechanical vector for Aleutian mink disease virus.

Flies are known vectors for a variety of infectious diseases in animals. In fur mink farming, one of the most severe diseases is Aleutian disease, which is caused by the Aleutian mink disease virus (AMDV). The presence of large fly populations is a frequent issue in mink farms; however, no studies assessing their role as AMDV carrier vectors have been conducted to-date. In order to determine the presence of AMDV in aerial flies from an infected mink farm, flies (n = 254) and environmental swab samples (n = 4) were collected from two farm barns. Fannia canicularis (L.) (Diptera: Muscidae) represented more than 99% of the fly population. One hundred and fifty specimens of this species were divided into pools of ten flies and analysed by qPCR, and positive samples were further sequenced. All fly pools and environmental samples tested positive for AMDV, and sequence analysis revealed identical genotypes in both types of samples. This is the first report of AMDV contamination in flies from mink farms, suggesting that F. canicularis may act as an AMDV vector. These results may be of interest for epidemiological studies and also for the improvement of control measures against this virus in mink farms.

Detection of vaccine-like lumpy skin disease virus in cattle and Musca domestica L. flies in an outbreak of lumpy skin disease in Russia in 2017.

Since 2012, lumpy skin disease virus (LSDV) has been spreading from the Middle East to south-east Europe and Russia. Although vaccination campaigns have managed to contain LSDV outbreaks, the risk of further spread is still high. The most likely route of LSDV transmission in short distance spread is vector-borne. Several arthropod species have been suggested as potential vectors, but no proven vector has yet been identified. To check whether promiscuous-landing synanthropic flies such as the common housefly (Musca domestica) could be involved, we carried out entomological trapping at the site of a recent LSDV outbreak caused by a vaccine-like LSDV strain. The presence of vaccine-like LSDV DNA was confirmed by the assay developed herein, the assay by Agianniotaki et al. (2017) and RPO30 gene sequencing. No evidence of field LSDV strain circulation was revealed. In this study, we discovered that M. domestica flies carried vaccine-like LSDV DNA (Ct  > 25.5), whereas trapped stable flies from the same collection were negative for both field and vaccine LSDV. To check whether flies were contaminated internally and externally, 50 randomly selected flies from the same collection were washed four times and tested. Viral DNA was mainly detected in the 1st wash fluid, suggesting genome or even viral contamination on the insect cadaver. In this study, internal contamination in the insect bodies without differentiation between the body locations was also revealed; however, the clinical relevance for mechanical transmission is unknown. Further work is needed to clarify a role of M. domestica in the transmission of LSDV. To our knowledge, this is the first report demonstrating that an attenuated LSD vaccine strain has been identified in Russian cattle given the ban on the use of live attenuated vaccines against LSDV.

Hytrosaviruses: current status and perspective.

Salivary gland hytrosaviruses (SGHVs) are entomopathogenic dsDNA, enveloped viruses that replicate in the salivary glands (SGs) of the adult dipterans, Glossina spp (GpSGHV) and Musca domestica (MdSGHV). Although belonging to the same virus family (Hytrosaviridae), SGHVs have distinct morphologies and pathobiologies. Two GpSGHV strains potentially account for the differential pathologies in lab-bred tsetse. New data suggest incorporation of host-derived cellular proteins and lipids into mature SGHVs. In addition to within the SGs, MdSGHV undergoes limited replication in the corpora allata, potentially disrupting hormone biosynthesis, and GpSGHV replicates in the milk glands providing a transmission conduit to progeny tsetse. Whereas MdSGHV is a potential biocontrol agent, the vertically transmitted GpSGHV is unsuitable for tsetse vector control but does jeopardize tsetse mass rearing.

Low potential for mechanical transmission of Ebola virus via house flies (Musca domestica).

BACKGROUND: Ebola virus (EBOV) infection results in high morbidity and mortality and is primarily transmitted in communities by contact with infectious bodily fluids. While clinical and experimental evidence indicates that EBOV is transmitted via mucosal exposure, the ability of non-biting muscid flies to mechanically transmit EBOV following exposure to the face had not been assessed. RESULTS: To investigate this transmission route, house flies (Musca domestica Linnaeus) were used to deliver an EBOV/blood mixture to the ocular/nasal/oral facial mucosa of four cynomolgus macaques (Macaca fascicularis Raffles). Following exposure, macaques were monitored for evidence of infection through the conclusion of the study, days 57 and 58. We found no evidence of systemic infection in any of the exposed macaques. CONCLUSIONS: The results of this study indicate that there is a low potential for the mechanical transmission of EBOV via house flies - the conditions in this study were not sufficient to initiate infection.

Detection of the Emerging Picornavirus Senecavirus A in Pigs, Mice, and Houseflies.

Senecavirus A (SVA) is an emerging picornavirus that has been recently associated with an increased number of outbreaks of vesicular disease and neonatal mortality in swine. Many aspects of SVA infection biology and epidemiology remain unknown. Here, we present a diagnostic investigation conducted in swine herds affected by vesicular disease and increased neonatal mortality. Clinical and environmental samples were collected from affected and unaffected herds and were screened for the presence of SVA by real-time reverse transcriptase PCR and virus isolation. Notably, SVA was detected and isolated from vesicular lesions and tissues of affected pigs, environmental samples, mouse feces, and mouse small intestine. SVA nucleic acid was also detected in houseflies collected from affected farms and from a farm with no history of vesicular disease. Detection of SVA in mice and housefly samples and recovery of viable virus from mouse feces and small intestine suggest that these pests may play a role on the epidemiology of SVA. These results provide important information that may allow the development of improved prevention and control strategies for SVA.

Enhancement of the Musca domestica hytrosavirus infection with orally delivered reducing agents.

House flies (Musca domestica L.) throughout the world are infected with the salivary gland hypertrophy virus MdSGHV (Hytrosaviridae). Although the primary route of infection is thought to be via ingestion, flies that are old enough to feed normally are resistant to infection per os, suggesting that the peritrophic matrix (PM) is a barrier to virus transmission. Histological examination of the peritrophic matrix of healthy flies revealed a multilaminate structure produced by midgut cells located near the proventriculus. SEM revealed the PM to form a confluent sheet surrounding the food bolus with pores/openings less than 10nm in diameter. TEM revealed the PM to be multilayered, varying in width from 350 to 900 nm, and generally thinner in male than in female flies. When flies were fed on the reducing agents dithiothetriol (DTT) or tris (2-caboxyethyl)phosphine hydrochloride (TCEP) for 48 h before per os exposure to the virus, infection rates increased 10- to 20-fold compared with flies that did not receive the reducing agent treatments. PM's from flies treated with DTT and TCEP displayed varying degrees of disruption, particularly in the outer layer, and lacked the electron-dense inner layer facing the ectoperitrophic space. Both drugs were somewhat toxic to the flies, resulting in>40% mortality at doses greater than 10mM (DTT) or 5 mM (TCEP). DTT increased male fly susceptibility (55.1% infected) more than that of females (7.8%), whereas TCEP increased susceptibility of females (42.9%) more than that of males (26.2%). The cause for the sex differences in response to oral exposure the reducing agents is unclear. Exposing flies to food treated with virus and the reducing agents at the same time, rather than pretreating flies with the drugs, had no effect on susceptibility to the virus. Presumably, the reducing agent disrupted the enveloped virus and acted as a viricidal agent. In summary, it is proposed that the reducing agents influence integrity of the PM barrier and increase the susceptibility of flies to infection by MdSGHV.

Analysis of the structural proteins from the Musca domestica hytrosavirus with an emphasis on the major envelope protein.

The Musca domestica hytrosavirus (MdHV), a member of the family Hyrosaviridae, is a large, dsDNA, enveloped virus that infects adult house flies and causes a diagnostic hypertrophy of the salivary gland. Herein, studies were directed at identifying key structural components of the viral envelope and nucleocapsid. SDS-PAGE of detergent-treated virus fractions identified protein bands unique to the envelope and nucleocapsid components. Using prior LC-MSMS data we identified the viral ORF associated with the major envelope band, cloned and expressed recombinant viral antigens, and prepared a series of polyclonal sera. Western blots confirmed that antibodies recognized the target viral antigen and provided evidence that the viral protein MdHV96 underwent post-translational processing; antibodies bound to the target high molecular weight parent molecule as well as distinct sets of smaller bands. Immuno gold electron microscopy demonstrated that the anti-MdHV96 sera recognized target antigens associated with the envelope. The nucleocapsids migrated from the virogenic stroma in the nucleus through the nuclear membrane into the cytoplasm, where they acquired an initial envelope that contained MdHV96. This major envelope protein, appeared to incorporate into intracellular membranes of both the caniculi and rough endoplasmic reticulum membranes and mediate binding to the nucleocapsids. Oral infection bioassays demonstrated that the anti-HV96 polyclonal sera acted as neutralizing agents in suppressing the levels of orally acquired infections.

Muscavirus (MdSGHV) disease dynamics in house fly populations--how is this virus transmitted and has it potential as a biological control agent?

The newly classified family Hytrosaviridae comprises several double-stranded DNA viruses that have been isolated from various dipteran species. These viruses cause characteristic salivary gland hypertrophy and suppress gonad development in their hosts. One member, Muscavirus or MdSGHV, exclusively infects adult house flies (Musca domestica) and, owing to its massive reproduction in and release from the salivary glands, is believed to be transmitted orally among feeding flies. However, results from recent experiments suggest that additional transmission routes likely are involved in the maintenance of MdSGHV in field populations of its host. Firstly, several hours before newly emerged feral flies begin feeding activities, the fully formed peritrophic matrix (PM) constitutes an effective barrier against oral infection. Secondly, flies are highly susceptible to topical virus treatments and intrahemocoelic injections. Thirdly, disease transmission is higher when flies are maintained in groups with infected conspecifics than when flies have access to virus-contaminated food. We hypothesize that interactions between flies may lead to cuticular damage, thereby providing an avenue to viral particles for direct access to the hemocoel. Based on our current knowledge, two options seem plausible for developing Muscavirus as a sterilizing agent to control house fly populations: The virus may either be formulated with PM-disrupting materials to facilitate oral infection from a feeding bait system, or amended with abrasive materials to enhance infection through a damaged cuticle after topical aerosol applications.

Experimental assessment of houseflies as vectors in avian influenza subtype H5N1 transmission in chickens.

In this study, laboratory-reared houseflies were experimentally exposed to the high pathogenicity avian influenza virus (HPAI) subtype H5N1 virus to evaluate the houseflies as vectors in HPAI-H5N1 virus transmission in chickens. One hundred and fifty houseflies (Musca domestica L.) were equally allocated into three groups. Groups 2 and 3 were exposed to the HPAI-H5N1 virus by allowing the flies to consume food containing the virus for 15 min, while the flies in group 1 were allowed to consume H5N1-free food and would serve as a negative control group. Group 2 flies were euthanatized immediately after H5N1 exposure, while group 3 were held at room temperature for 24 hr and euthanatized. The houseflies in the transmission of the HPAI-H5N1 virus were examined by challenging three groups of housefly homogenates into layer chickens via the oral drop. Morbidity and mortality were observed for 14 days, and virus shedding monitored via oropharyngeal swabs (OS) and cloacal swabs (CS), which were collected daily and determined by real-time reverse transcription-PCR and virus titration. Experimental challenge showed that all the chickens of groups 2 and 3 died within 7 days of inoculation. The OS had higher concentrations of virus than CS. Moreover, the chickens of group 2 had higher concentrations of virus shedding than the chickens of group 3. Immunohistochemistry detected the nucleoprotein of the type A influenza virus in all tissue samples collected, including the trachea, duodenum, pancreas, and brain. In summary, this study demonstrates that houseflies could serve as vectors in HPAI-H5N1 virus transmission in chickens under experimental conditions.

Antiviral, immunomodulatory, and free radical scavenging activities of a protein-enriched fraction from the larvae of the housefly, Musca domestica.

In our previous study, protein-enriched fraction (PEF) that was isolated from the larvae of the housefly, Musca domestica L. (Diptera: Muscidae), showed excellent hepatoprotective activity as well as the potential for clinical application in therapy for liver diseases. In this study, antiviral, immunomodulatory, and free radical scavenging activities of PEF were evaluated. The antiviral results demonstrated that PEF inhibited the infection of avian influenza virus H9N2 and had a virucidal effect against the multicapsid nucleopolyhedrovirus of the alfalfa looper, Autographa californica Speyer (Lepidoptera: Noctuidae) in vitro. The mortality of silkworm larve in a PEF treatment group decreased significantly compared with a negative control. PEF showed excellent scavenging activity for 1,1-diphenyl-2-picrylhydrazyl and superoxide anion radicals, which were similar to those of ascorbic acid. The imunomodulatory results suggested that PEF could effectively improve immune function in experimental mice. Our results indicated that PEF could possibly be used for the prophylaxis and treatment of diseases caused by avian influenza virus infection. In addition, PEF with virucidal activity against insect viruses might provide useful for the development of antimicrobial breeding technology for economically important insects. As a natural product from insects, PEF could be a potential source for the discovery of potent antioxidant and immunomodulatory agents.