Serological Detection of SARS-CoV-2 Antibodies in Naturally-Infected Mink and Other Experimentally-Infected Animals

The recent emergence of SARS-CoV-2 in humans from a yet unidentified animal reservoir and the capacity of the virus to naturally infect pets, farmed animals and potentially wild animals has highlighted the need for serological surveillance tools. In this study, the luciferase immunoprecipitation systems (LIPS), employing the spike (S) and nucleocapsid proteins (N) of SARS-CoV-2, was used to examine the suitability of the assay for antibody detection in different animal species. Sera from SARS-CoV-2 naturally-infected mink (n = 77), SARS-CoV-2 experimentally-infected ferrets, fruit bats and hamsters and a rabbit vaccinated with a purified spike protein were examined for antibodies using the SARS-CoV-2 N and/or S proteins. From comparison with the known neutralization status of the serum samples, statistical analyses including calculation of the Spearman rank-order-correlation coefficient and Cohen’s kappa agreement were used to interpret the antibody results and diagnostic performance. The LIPS immunoassay robustly detected the presence of viral antibodies in naturally infected SARS-CoV-2 mink, experimentally infected ferrets, fruit bats and hamsters as well as in an immunized rabbit. For the SARS-CoV-2-LIPS-S assay, there was a good level of discrimination between the positive and negative samples for each of the five species tested with 100% agreement with the virus neutralization results. In contrast, the SARS-CoV-2-LIPS-N assay did not consistently differentiate between SARS-CoV-2 positive and negative sera. This study demonstrates the suitability of the SARS-CoV-2-LIPS-S assay for the sero-surveillance of SARS-CoV-2 infection in a range of animal species.     

Monitoring Natural SARS-CoV-2 Infection in Lions (Panthera leo) at the Barcelona Zoo: Viral Dynamics and Host Responses

To date, no evidence supports the fact that animals play a role in the epidemiology of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the coronavirus infectious disease 2019 (COVID-19). However, several animal species are naturally susceptible to SARS-CoV-2 infection. Besides pets (cats, dogs, Syrian hamsters, and ferrets) and farm animals (minks), different zoo animal species have tested positive for SARS-CoV-2 (large felids and non-human primates). After the summer of 2020, a second wave of SARS-CoV-2 infection occurred in Barcelona (Spain), reaching a peak of positive cases in November. During that period, four lions (Panthera leo) at the Barcelona Zoo and three caretakers developed respiratory signs and tested positive for the SARS-CoV-2 antigen. Lion infection was monitored for several weeks and nasal, fecal, saliva, and blood samples were taken at different time-points. SARS-CoV-2 RNA was detected in nasal samples from all studied lions and the viral RNA was detected up to two weeks after the initial viral positive test in three out of four animals. The SARS-CoV-2 genome was also detected in the feces of animals at different times. Virus isolation was successful only from respiratory samples of two lions at an early time-point. The four animals developed neutralizing antibodies after the infection that were detectable four months after the initial diagnosis. The partial SARS-CoV-2 genome sequence from one animal caretaker was identical to the sequences obtained from lions. Chronology of the events, the viral dynamics, and the genomic data support human-to-lion transmission as the origin of infection.     

First Detection of SARS-CoV-2 B.1.1.7 Variant of Concern in an Asymptomatic Dog in Spain

Natural SARS-CoV-2 infection in pets has been widely documented during the last year. Although the majority of reports suggested that dogs’ susceptibility to the infection is low, little is known about viral pathogenicity and transmissibility in the case of variants of concern, such as B.1.1.7 in this species. Here, as part of a large-scale study on SARS-CoV-2 prevalence in pets in Spain, we have detected the B.1.1.7 variant of concern (VOC) in a dog whose owners were infected with SARS-CoV-2. The animal did not present any symptoms, but viral loads were high in the nasal and rectal swabs. In addition, viral isolation was possible from both swabs, demonstrating that the dog was shedding infectious virus. Seroconversion occurred 23 days after the first sampling. This study documents the first detection of B.1.1.7 VOC in a dog in Spain and emphasizes the importance of performing active surveillance and genomic investigation on infected animals.     

SARS CoV-2 (Delta Variant) Infection Kinetics and Immunopathogenesis in Domestic Cats

Continued emergence of SARS-CoV-2 variants highlights the critical need for adaptable and translational animal models for acute COVID-19. Limitations to current animal models for SARS CoV-2 (e.g., transgenic mice, non-human primates, ferrets) include subclinical to mild lower respiratory disease, divergence from clinical COVID-19 disease course, and/or the need for host genetic modifications to permit infection. We therefore established a feline model to study COVID-19 disease progression and utilized this model to evaluate infection kinetics and immunopathology of the rapidly circulating Delta variant (B.1.617.2) of SARS-CoV-2. In this study, specific-pathogen-free domestic cats (n = 24) were inoculated intranasally and/or intratracheally with SARS CoV-2 (B.1.617.2). Infected cats developed severe clinical respiratory disease and pulmonary lesions at 4- and 12-days post-infection (dpi), even at 1/10 the dose of previously studied wild-type SARS-CoV-2. Infectious virus was isolated from nasal secretions of delta-variant infected cats in high amounts at multiple timepoints, and viral antigen was co-localized in ACE2-expressing cells of the lungs (pneumocytes, vascular endothelium, peribronchial glandular epithelium) and strongly associated with severe pulmonary inflammation and vasculitis that were more pronounced than in wild-type SARS-CoV-2 infection. RNA sequencing of infected feline lung tissues identified upregulation of multiple gene pathways associated with cytokine receptor interactions, chemokine signaling, and viral protein–cytokine interactions during acute infection with SARS-CoV-2. Weighted correlation network analysis (WGCNA) of differentially expressed genes identified several distinct clusters of dysregulated hub genes that are significantly correlated with both clinical signs and lesions during acute infection. Collectively, the results of these studies help to delineate the role of domestic cats in disease transmission and response to variant emergence, establish a flexible translational model to develop strategies to prevent the spread of SARS-CoV-2, and identify potential targets for downstream therapeutic development.     

Inter‐ and intraspecies transmission of canine influenza virus (H3N2) in dogs,cats, and ferrets

Background The emergence of zoonotic viruses in domestic animals is a significant public health concern. Canine influenza virus (CIV) H3N2 is a virus that can infect companion animals and is, therefore, a potential public health concern. Objective This study investigated the inter‐ and intraspecies transmission of CIV among dogs, cats, and ferrets, under laboratory conditions, to determine whether transmission of the virus was possible between as well as within these domestic animal species. Method The transmission routes for inter‐ and intraspecies transmission were airborne and direct contact, respectively. Transmission was conducted through intranasal infection of dogs followed by exposure to either cats or ferrets and by comingling infected and naïve animals of the same species. Results The interspecies transmission of CIV H3N2 via airborne was only observed from dogs to cats and not from dogs to ferrets. However, direct intranasal infection of either cats or ferrets with CIV could induce influenza‐like clinical signs, viral shedding, and serological responses. Additionally, naïve cats and ferrets could be infected by CIV via direct contact with infected animals of the same species. Conclusion Cats appear to be another susceptible host of CIV H3N2, whereas ferrets are not likely natural hosts. The molecular‐based mechanism of interspecies and intraspecies transmission of CIV H3N2 should be further studied.     

An Updated Review on SARS-CoV-2 Infection in Animals

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has lasted for two years and caused millions of infections and deaths in humans. Although the origin of SARS-CoV-2 infection in humans remains unknown, infection in animals has been frequently reported in varieties of animals all over the world. Both experimental and natural infections of SARS-CoV-2 in different animal species provide useful information on viral host range and pathogenicity. As the pandemic continues to evolve, SARS-CoV-2 infection in animals will be expanding. In this review, we summarized SARS-CoV-2 testing and infection in animals as well as SARS-CoV-2 strains and transmission in animals. Current data showed that at least 18 different animal species tested positive for SARS-CoV-2. These 18 animal species belong to pet, captive, farmed, and wild animals. Fifteen of the eighteen animal species were known to be positive for the Delta variant and ten animal species were infected with two different types of variants. Human-to-animal, animal-to-animal, and animal-to-human transmission events were suggested in different outbreaks involved in animal infection with SARS-CoV-2. Continued testing, immunization, and surveillance are warranted.     

SARS-CoV-2 Variant-Specific Infectivity and Immune Profiles Are Detectable in a Humanized Lung Mouse Model

Small animal models that accurately model pathogenesis of SARS-CoV-2 variants are required for ongoing research efforts. We modified our human immune system mouse model to support replication of SARS-CoV-2 by implantation of human lung tissue into the mice to create TKO-BLT-Lung (L) mice and compared infection with two different variants in a humanized lung model. Infection of TKO-BLT-L mice with SARS-CoV-2 recapitulated the higher infectivity of the B.1.1.7 variant with more animals becoming infected and higher sustained viral loads compared to mice challenged with an early B lineage (614D) virus. Viral lesions were observed in lung organoids but no differences were detected between the viral variants as expected. Partially overlapping but distinct immune profiles were also observed between the variants with a greater Th1 profile in VIDO-01 and greater Th2 profile in B.1.1.7 infection. Overall, the TKO-BLT-L mouse supported SARS-CoV-2 infection, recapitulated key known similarities and differences in infectivity and pathogenesis as well as revealing previously unreported differences in immune responses between the two viral variants. Thus, the TKO-BLT-L model may serve as a useful animal model to study the immunopathobiology of newly emerging variants in the context of genuine human lung tissue and immune cells.     

Protective Effects of Astodrimer Sodium 1% Nasal Spray Formulation against SARS-CoV-2 Nasal Challenge in K18-hACE2 Mice

Strategies to combat COVID-19 require multiple ways to protect vulnerable people from infection. SARS-CoV-2 is an airborne pathogen and the nasal cavity is a primary target of infection. The K18-hACE2 mouse model was used to investigate the anti-SARS-CoV-2 efficacy of astodrimer sodium formulated in a mucoadhesive nasal spray. Animals received astodrimer sodium 1% nasal spray or PBS intranasally, or intranasally and intratracheally, for 7 days, and they were infected intranasally with SARS-CoV-2 after the first product administration on Day 0. Another group was infected intranasally with SARS-CoV-2 that had been pre-incubated with astodrimer sodium 1% nasal spray or PBS for 60 min before the neutralisation of test product activity. Astodrimer sodium 1% significantly reduced the viral genome copies (>99.9%) and the infectious virus (~95%) in the lung and trachea vs. PBS. The pre-incubation of SARS-CoV-2 with astodrimer sodium 1% resulted in a significant reduction in the viral genome copies (>99.9%) and the infectious virus (>99%) in the lung and trachea, and the infectious virus was not detected in the brain or liver. Astodrimer sodium 1% resulted in a significant reduction of viral genome copies in nasal secretions vs. PBS on Day 7 post-infection. A reduction in the viral shedding from the nasal cavity may result in lower virus transmission rates. Viraemia was low or undetectable in animals treated with astodrimer sodium 1% or infected with treated virus, correlating with the lack of detectable viral replication in the liver. Similarly, low virus replication in the nasal cavity after treatment with astodrimer sodium 1% potentially protected the brain from infection. Astodrimer sodium 1% significantly reduced the pro-inflammatory cytokines IL-6, IL-1α, IL-1β, TNFα and TGFβ and the chemokine MCP-1 in the serum, lung and trachea vs. PBS. Astodrimer sodium 1% nasal spray blocked or reduced SARS-CoV-2 replication and its sequelae in K18-hACE2 mice. These data indicate a potential role for the product in preventing SARS-CoV-2 infection or for reducing the severity of COVID-19.     

Investigations on SARS-CoV-2 Susceptibility of Domestic and Wild Animals Using Primary Cell Culture Models Derived from the Upper and Lower Respiratory Tract

Several animal species are susceptible to SARS-CoV-2 infection, as documented by case reports and serological and in vivo infection studies. However, the susceptibility of many animal species remains unknown. Furthermore, the expression patterns of SARS-CoV-2 entry factors, such as the receptor angiotensin-converting enzyme 2 (ACE2), as well as transmembrane protease serine subtype 2 (TMPRSS2) and cathepsin L (CTSL), cellular proteases involved in SARS-CoV-2 spike protein activation, are largely unexplored in most species. Here, we generated primary cell cultures from the respiratory tract of domestic and wildlife animals to assess their susceptibility to SARS-CoV-2 infection. Additionally, the presence of ACE2, TMPRSS2 and CTSL within respiratory tract compartments was investigated in a range of animals, some with unknown susceptibility to SARS-CoV-2. Productive viral replication was observed in the nasal mucosa explants and precision-cut lung slices from dogs and hamsters, whereas culture models from ferrets and multiple ungulate species were non-permissive to infection. Overall, whereas TMPRSS2 and CTSL were equally expressed in the respiratory tract, the expression levels of ACE2 were more variable, suggesting that a restricted availability of ACE2 may contribute to reduced susceptibility. Summarized, the experimental infection of primary respiratory tract cell cultures, as well as an analysis of entry-factor distribution, enable screening for SARS-CoV-2 animal reservoirs.     

SARS-CoV-2 Infections in Animals: Reservoirs for Reverse Zoonosis and Models for Study

The recent SARS-CoV-2 pandemic has brought many questions over the origin of the virus, the threat it poses to animals both in the wild and captivity, and the risks of a permanent viral reservoir developing in animals. Animal experiments have shown that a variety of animals can become infected with the virus. While coronaviruses have been known to infect animals for decades, the true intermediate host of the virus has not been identified, with no cases of SARS-CoV-2 in wild animals. The screening of wild, farmed, and domesticated animals is necessary to help us understand the virus and its origins and prevent future outbreaks of both COVID-19 and other diseases. There is intriguing evidence that farmed mink infections (acquired from humans) have led to infection of other farm workers in turn, with a recent outbreak of a mink variant in humans in Denmark. A thorough examination of the current knowledge and evidence of the ability of SARS-CoV-2 to infect different animal species is therefore vital to evaluate the threat of animal to human transmission and reverse zoonosis.     

Development of a Hamster Natural Transmission Model of SARS-CoV-2 Infection

The global pandemic of coronavirus disease (COVID-19) caused by infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has led to an international thrust to study pathogenesis and evaluate interventions. Experimental infection of hamsters and the resulting respiratory disease is one of the preferred animal models since clinical signs of disease and virus shedding are similar to more severe cases of human COVID-19. The main route of challenge has been direct inoculation of the virus via the intranasal route. To resemble the natural infection, we designed a bespoke natural transmission cage system to assess whether recipient animals housed in physically separate adjacent cages could become infected from a challenged donor animal in a central cage, with equal airflow across the two side cages. To optimise viral shedding in the donor animals, a low and moderate challenge dose were compared after direct intranasal challenge, but similar viral shedding responses were observed and no discernible difference in kinetics. The results from our natural transmission set-up demonstrate that most recipient hamsters are infected within the system developed, with variation in the kinetics and levels of disease between individual animals. Common clinical outputs used for the assessment in directly-challenged hamsters, such as weight loss, are less obvious in hamsters who become infected from naturally acquiring the infection. The results demonstrate the utility of a natural transmission model for further work on assessing the differences between virus strains and evaluating interventions using a challenge system which more closely resembles human infection.     

Companion animals likely do not spread COVID-19 but may get infected themselves

Coronavirus disease 2019 (COVID-19) is a highly contagious infectious disease caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). From the epidemiological data, the picture emerges that the more severe etiopathologies among COVID-19 patients are found in elderly people. The risk of death due to COVID-19 increases exponentially with age. Eight out of 10 COVID-19 related deaths occur in people older than 65 years of age. Older patients with comorbid conditions such as hypertension, heart failure, diabetes mellitus, asthma, chronic obstructive pulmonary disease, and cancer have a much higher case fatality rate. Governments and public health authorities all over the world have realized that protections of vulnerable older adults should be a priority during the COVID-19 pandemic. COVID-19 is a zoonotic disease. The SARS-CoV-2 virus was originally transmitted likely from a bat or a pangolin to humans. Recent evidence suggests that SARS-CoV-2, similar to other coronaviruses, can infect several species of animals, including companion animals such as dogs, cats, and ferrets although their viral loads remain low. While the main source of infection transmission therefore is human to human, there are a few rare cases of pets contracting the infection from a SARS-CoV-2-infected human. Although there is no evidence that pets actively transmit SARS-CoV-2 via animal-to-human transmission, senior pet ownership potentially may pose a small risk to older adults by (1) potentially enabling animal-to-human transmission of SARS-CoV-2 in the most vulnerable population and (2) by increasing the exposition risk for the elderly due to the necessity to care for the pet and, in the case of dogs, to take them outside the house several times per day. In this overview, the available evidence on SARS-CoV-2 infection in pets is considered and the potential for spread of COVID-19 from companion animals to older individuals and the importance of prevention are discussed.

     

Susceptibility of Domestic Goat (Capra aegagrus hircus) to Experimental Infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) B.1.351/Beta Variant

A wide range of animal species are susceptible to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Natural and/or experimental infections have been reported in pet, zoo, farmed and wild animals. Interestingly, some SARS-CoV-2 variants, such as B.1.1.7/Alpha, B.1.351/Beta, and B.1.1.529/Omicron, were demonstrated to infect some animal species not susceptible to classical viral variants. The present study aimed to elucidate if goats (Capra aegagrus hircus) are susceptible to the B.1.351/Beta variant. First, an in silico approach was used to predict the affinity between the receptor-binding domain of the spike protein of SARS-CoV-2 B.1.351/Beta variant and angiotensin-converting enzyme 2 from goats. Moreover, we performed an experimental inoculation with this variant in domestic goat and showed evidence of infection. SARS-CoV-2 was detected in nasal swabs and tissues by RT-qPCR and/or immunohistochemistry, and seroneutralisation was confirmed via ELISA and live virus neutralisation assays. However, the viral amount and tissue distribution suggest a low susceptibility of goats to the B.1.351/Beta variant. Therefore, although monitoring livestock is advisable, it is unlikely that goats play a role as SARS-CoV-2 reservoir species, and they are not useful surrogates to study SARS-CoV-2 infection in farmed animals.     

Neuroinvasion and Encephalitis Following Intranasal Inoculation of SARS-CoV-2 in K18-hACE2 Mice

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection can cause neurological disease in humans, but little is known about the pathogenesis of SARS-CoV-2 infection in the central nervous system (CNS). Herein, using K18-hACE2 mice, we demonstrate that SARS-CoV-2 neuroinvasion and encephalitis is associated with mortality in these mice. Intranasal infection of K18-hACE2 mice with 10⁵ plaque-forming units of SARS-CoV-2 resulted in 100% mortality by day 6 after infection. The highest virus titers in the lungs were observed on day 3 and declined on days 5 and 6 after infection. By contrast, very high levels of infectious virus were uniformly detected in the brains of all the animals on days 5 and 6. Onset of severe disease in infected mice correlated with peak viral levels in the brain. SARS-CoV-2-infected mice exhibited encephalitis hallmarks characterized by production of cytokines and chemokines, leukocyte infiltration, hemorrhage and neuronal cell death. SARS-CoV-2 was also found to productively infect cells within the nasal turbinate, eye and olfactory bulb, suggesting SARS-CoV-2 entry into the brain by this route after intranasal infection. Our data indicate that direct infection of CNS cells together with the induced inflammatory response in the brain resulted in the severe disease observed in SARS-CoV-2-infected K18-hACE2 mice.     

SARS-CoV-2 Infections and Viral Isolations among Serially Tested Cats and Dogs in Households with Infected Owners in Texas,USA

Understanding the ecological and epidemiological roles of pets in the transmission of SARS-CoV-2 is critical for animal and human health, identifying household reservoirs, and predicting the potential enzootic maintenance of the virus. We conducted a longitudinal household transmission study of 76 dogs and cats living with at least one SARS-CoV-2-infected human in Texas and found that 17 pets from 25.6% of 39 households met the national case definition for SARS-CoV-2 infections in animals. This includes three out of seventeen (17.6%) cats and one out of fifty-nine (1.7%) dogs that were positive by RT-PCR and sequencing, with the virus successfully isolated from the respiratory swabs of one cat and one dog. Whole-genome sequences of SARS-CoV-2 obtained from all four PCR-positive animals were unique variants grouping with genomes circulating among people with COVID-19 in Texas. Re-sampling showed persistence of viral RNA for at least 25 d-post initial test. Additionally, seven out of sixteen (43.8%) cats and seven out of fifty-nine (11.9%) dogs harbored SARS-CoV-2 neutralizing antibodies upon initial sampling, with relatively stable or increasing titers over the 2–3 months of follow-up and no evidence of seroreversion. The majority (82.4%) of infected pets were asymptomatic. ‘Reverse zoonotic’ transmission of SARS-CoV-2 from infected people to animals may occur more frequently than recognized.     

Intra-Host SARS-CoV-2 Evolution in the Gut of Mucosally-Infected Chlorocebus aethiops (African Green Monkeys)

In recent months, several SARS-CoV-2 variants have emerged that enhance transmissibility and escape host humoral immunity. Hence, the tracking of viral evolutionary trajectories is clearly of great importance. Little is known about SARS-CoV-2 evolution in nonhuman primate models used to test vaccines and therapies and to model human disease. Viral RNA was sequenced from rectal swabs from Chlorocebus aethiops (African green monkeys) after experimental respiratory SARS-CoV-2 infection. Two distinct patterns of viral evolution were identified that were shared between all collected samples. First, mutations in the furin cleavage site that were initially present in the virus as a consequence of VeroE6 cell culture adaptation were not detected in viral RNA recovered in rectal swabs, confirming the necessity of this motif for viral infection in vivo. Three amino acid changes were also identified; ORF 1a S2103F, and spike D215G and H655Y, which were detected in rectal swabs from all sampled animals. These findings are demonstrative of intra-host SARS-CoV-2 evolution and may identify a host-adapted variant of SARS-CoV-2 that would be useful in future primate models involving SARS-CoV-2 infection.     

Eye Infection with SARS-CoV-2 as a Route to Systemic Immunization?

Infectious diseases of the conjunctiva and cornea usually leave behind both broad local and systemic immunity. Case reports of SARS-CoV-2-positive conjunctivitis with subsequent systemic immunity suggest a new route of immunization preventing the primary infection of the airways. Material and Methods: A total of 24 Syrian field hamsters were treated. In systematic animal experiments, we infected the eyes of n = 8 animals (group 1) and the airways of another n = 8 animals (group 2) with SARS-CoV-2 (Wuhan type); n = 8 hamsters served as controls (group 3). The weight development of the animals was recorded. After two weeks of observation of disease symptoms, all animals were re-exposed to SARS-CoV-2 in the respiratory tract (challenge) to determine whether immunity to the virus had been achieved. Results: The epi-ocularly infected animals (group 1) showed no clinically visible disease during the ocular infection phase. At most, there was a slightly reduced weight gain compared to the control group (group 3), while the respiratory infected animals (group 2) all lost weight, became lethargic, and slowly recovered after two weeks. After the challenge, none of the animals in groups 1 and 2 became ill again. The animals in the negative control (group 3) all became ill. Cytotoxic antibodies were detectable in the blood of the infected groups before and after challenge, with higher titers in the epi-ocularly infected animals. Conclusion: By epi-ocular infection with SARS-CoV-2, the development of systemic immunity with formation of cytotoxic antibodies without severe general disease could be observed in the experimental animals, which did not induce any more disease upon a second infection in the respiratory tract. Therefore, it can be concluded that a purely epi-ocular infection with SARS-CoV2 only induces a weak disease pattern followed by systemic immunity.     

Hepatitis C virus JFH-1 strain infection in chimpanzees is associated with low pathogenicity and emergence of an adaptive mutation

The identification of the hepatitis C virus (HCV) strain JFH-1 enabled the successful development of infectious cell culture systems. Although this strain replicates efficiently and produces infectious virus in cell culture, the replication capacity and pathogenesis in vivo are still undefined. To assess the in vivo phenotype of the JFH-1 virus, cell culture-generated JFH-1 virus (JFH-1cc) and patient serum from which JFH-1 was isolated were inoculated into chimpanzees. Both animals became HCV RNA-positive 3 days after inoculation but showed low-level viremia and no evidence of hepatitis. HCV viremia persisted 8 and 34 weeks in JFH-1cc and patient serum-infected chimpanzees, respectively. Immunological analysis revealed that HCV-specific immune responses were similarly induced in both animals. Sequencing of HCV at various times of infection indicated more substitutions in the patient serum-inoculated chimpanzee, and the higher level of sequence variations seemed to be associated with a prolonged infection in this animal. A common mutation G838R in the NS2 region emerged early in both chimpanzees. This mutation enhances viral assembly, leading to an increase in viral production in transfected or infected cells. CONCLUSION: Our study shows that the HCV JFH-1 strain causes attenuated infection and low pathogenicity in chimpanzees and is capable of adapting in vivo with a unique mutation conferring an enhanced replicative phenotype.     

Impact of a large deletion in the neuraminidase protein identified in a laninamivir‐selected influenza A/Brisbane/10/2007 (H3N2) variant on viral fitness in vitro and in ferrets

Viral fitness of a laninamivir‐selected influenza A/Brisbane/10/2007‐like (H3N2) isolate (LRVp9) containing a 237‐amino acid neuraminidase deletion and a P194L hemagglutinin mutation was evaluated in vitro and in ferrets. LRVp9 and the wild‐type (WT) virus showed comparable replication kinetics in MDCK‐ST6GalI cells. Cultured virus was recovered between days 2 and 5 post‐infection in nasal washes (NW) from the 4 WT‐infected ferrets whereas no virus was recovered from the LRVp9‐infected animals. There was a ≥1 log reduction in viral RNA copies/μl of NW for LRVp9 compared to WT at most time points. The large neuraminidase deletion compromises viral infectivity in vivo.