Background: The herpes simplex virus ICP34.5 null mutant HSV1716 replicates selectively in actively dividing cells and is currently being assessed as a therapy for gliomas. The enzyme HSV thymidine kinase phosphorylates the thymidine analogue FIAU (5-iodo-1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)uracil) and traps it in infected cells. This study aims to determine whether SPECT imaging with 123I-FIAU can provide evidence of increased thymidine kinase activity following HSV-tk gene expression and hence evidence of viral replication in brain tumours injected with the HSV1716 virus. Method: Brain SPECT images were acquired on a Neurofocus camera at 1 and 24 h after intravenous administration of 123I-FIAU (150 MBq or 300 MBq) in 7 patients with glioma. Scans were obtained pre- and post- virus administration into the tumour. Whole-body images (0.25 h, 5 h, 25 h p.i.) were obtained in three subjects. Results: Initial uptake of FIAU in the tumour was relatively low making assessment of retention difficult. However, the retention index in 2 patients increased post-viral administration which may be consistent with viral replication. Whole-body imaging revealed predominantly renal excretion of the tracer. These results suggest that the distribution of replicating HSV 1716 virus may be imaged using 123I-FIAU, although strategies to increase brain uptake will be essential for routine use.
Abstract SARS-CoV-2 is a recently emerged, highly contagious virus and the cause of the current pandemic. It is a zoonotic virus, although its animal origin is not clear yet. Person-to-person transmission occurs by inhalation of infected droplets and aerosols, or by direct contact with contaminated fomites. Arthropods transmit numerous viral, parasitic, and bacterial diseases; however, the potential role of arthropods in SARS-CoV-2 transmission is not fully understood. Thus far, a few studies have demonstrated that SARS-CoV-2 replication is not supported in cells from certain insect species nor in certain species of mosquitoes after intrathoracic inoculation. In this study, we expanded the work of SARS-CoV-2 susceptibility to biting insects after ingesting a SARS-CoV-2infected blood meal. Species tested included Culicoides sonorensis biting midges, as well as Culex tarsalis and Culex quinquefasciatus mosquitoes, all known biological vectors for numerous RNA viruses. Arthropods were allowed to feed on SARS-CoV-2 spiked blood and at various time points post infection analyzed for the presence of viral RNA and infectious virus. Additionally, cell lines derived from C. sonorensis (W8a), Ae. aegypti (C6/36), Cx. quinquefasciatus (HSU), and Cx. tarsalis (CxTrR2) were tested for SARS-CoV-2 susceptibility. Our results indicate that none of the biting insects, nor the insect cell lines support SARS-CoV-2 replication. We conclude, that biting insect do not pose a risk for transmission of SARS-CoV-2 to humans or animals following a SARS-CoV-2 infected blood meal.
Japanese encephalitis virus (JEV) is a zoonotic mosquito-borne pathogen that regularly causes severe neurological disease in humans in Southeast Asia and the Western Pacific region. Pigs are one of the main amplifying hosts of JEV and play a central role in the virus transmission cycle. The objective of this study was to identify in vitro cell systems to investigate early effects of JEV infection including viral replication and host cell death. Here, we demonstrate the susceptibility of several porcine cell lines to the attenuated genotype III JEV strain SA14-14-2. Monolayers of porcine nasal turbinate (PT-K75), kidney (SK-RST), testis (ST), and monocyte-derived macrophage (CΔ2+) cells were infected with SA14-14-2 for up to five days at a multiplicity of infection (MOI) of 0.1. The hamster kidney cell line BHK-21, previously shown to be susceptible to SA14-14-2, was used as a positive control. Culture supernatants and cells were collected between 0 and 120 h post infection (hpi), and monolayers were observed for cytopathic effect (CPE) using brightfield microscopy. The number of infectious virus particles was quantified by plaque assay and cell viability was determined using trypan blue staining. An indirect immunofluorescence assay was used to detect the presence of JEV NS1 antigens in cells infected at 1 MOI. All four porcine cell lines demonstrated susceptibility to SA14-14-2 and produced infectious virus by 12 hpi. Virus titers peaked at 48 hpi in CΔ2+, BHK-21, and SK-RST cells, at 72 hpi in PT-K75, and at 120 hpi in ST cells. CPE was visible in infected CΔ2+ and BHK-21 cells, but not the other three cell lines. The proportion of viable cells, as measured by trypan blue exclusion, declined after 24 hpi in BHK-21 and 48 hpi in CΔ2+ cells, but did not substantially decline in SK-RST, PT-K75 or ST cells. At 48 hpi, JEV NS1 was detected in all infected cell lines by fluorescence microscopy. These findings demonstrate several porcine cell lines which have the potential to serve as useful research tools for investigating JEV infection dynamics and host cell mechanisms in a natural amplifying host species, such as pigs, in vitro.
Abstract 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.
Abstract SARS-CoV-2 is a recently emerged, highly contagious virus and the cause of the current COVID-19 pandemic. It is a zoonotic virus, although its animal origin is not clear yet. Person-to-person transmission occurs by inhalation of infected droplets and aerosols, or by direct contact with contaminated fomites. Arthropods transmit numerous viral, parasitic, and bacterial diseases; however, the potential role of arthropods in SARS-CoV-2 transmission is not fully understood. Thus far, a few studies have demonstrated that SARS-CoV-2 replication is not supported in cells from certain insect species nor in certain species of mosquitoes after intrathoracic inoculation. In this study, we expanded the work of SARS-CoV-2 susceptibility to biting insects after ingesting a SARS-CoV-2-infected bloodmeal. Species tested included Culicoides sonorensis (Wirth & Jones) (Diptera: Ceratopogonidae) biting midges, as well as Culex tarsalis (Coquillett) and Culex quinquefasciatus (Say) mosquitoes (Diptera: Culicidae), all known biological vectors for numerous RNA viruses. Arthropods were allowed to feed on SARS-CoV-2-spiked blood and at a time point postinfection analyzed for the presence of viral RNA and infectious virus. Additionally, cell lines derived from C. sonorensis (W8a), Aedes aegypti (Linnaeus) (Diptera: Culicidae) (C6/36), Cx. quinquefasciatus (HSU), and Cx. tarsalis (CxTrR2) were tested for SARS-CoV-2 susceptibility. Our results indicate that none of the biting insects, nor the insect cell lines evaluated support SARS-CoV-2 replication, suggesting that these species are unable to be biological vectors of SARS-CoV-2.
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