The CDE region of feline Calicivirus VP1 protein is a potential candidate subunit vaccine.

BACKGROUND: Feline calicivirus (FCV) infection causes severe upper respiratory disease in cats, but there are no effective vaccines available for preventing FCV infection. Subunit vaccines have the advantages of safety, low cost and excellent immunogenicity, but no FCV subunit vaccine is currently available. The CDE protein is the dominant neutralizing epitope region of the main antigenic structural protein of FCV, VP1. Therefore, this study evaluated the effectiveness of the CDE region as a truncated FCV VP1 protein in preventing FCV infection to provide a strategy for developing potential FCV subunit vaccines. RESULTS: Through the prediction of FCV VP1 epitopes, we found that the E region is the dominant neutralizing epitope region. By analysing the spatial structure of VP1 protein, 13 amino acid sites in the CD and E regions were found to form hydrogen bonding interactions. The results show the presence of these interaction forces supports the E region, helping improve the stability and expression level of the soluble E protein. Therefore, we selected the CDE protein as the immunogen for the immunization of felines. After immunization with the CDE protein, we found significant stimulation of IgG, IgA and neutralizing antibody production in serum and swab samples, and the cytokine TNF-α levels and the numbers of CD4+ T lymphocytes were increased. Moreover, a viral challenge trial indicated that the protection generated by the CDE subunit vaccine significantly reduced the incidence of disease in animals. CONCLUSIONS: For the first time, we studied the efficacy of the CDE protein, which is the dominant neutralizing epitope region of the FCV VP1 protein, in preventing FCV infection. We revealed that the CDE protein can significantly activate humoral, mucosal and cellular immunity, and the resulting protective effect can significantly reduce the incidence of animal disease. The CDE region of the FCV capsid is easy to produce and has high stability and excellent immunogenicity, which makes it a candidate for low-cost vaccines.

Ultrasensitive and visual detection of Feline herpesvirus type-1 and Feline calicivirus using one-tube dRPA-Cas12a/Cas13a assay.

BACKGROUND: Feline herpesvirus type 1 (FHV) and Feline calicivirus (FCV) are the primary co-infecting pathogens that cause upper respiratory tract disease in cats. However, there are currently no visual detection assays available for on-site testing. Here, we develop an ultrasensitive and visual detection method based on dual recombinase polymerase amplification (dRPA) reaction and the hybrid Cas12a/Cas13a trans-cleavage activities in a one-tube reaction system, referred to as one-tube dRPA-Cas12a/Cas13a assay. RESULTS: The recombinant plasmid DNAs, crRNAs, and RPA oligonucleotides targeting the FCV ORF1 gene and FHV-1 TK gene were meticulously prepared. Subsequently, dual RPA reactions were performed followed by screening of essential reaction components for hybrid CRISPR-Cas12a (targeting the FHV-1 TK gene) and CRISPR-Cas13a (targeting the FCV ORF1 gene) trans-cleavage reaction. As a result, we successfully established an ultra-sensitive and visually detectable method for simultaneous detection of FCV and FHV-1 nucleic acids using dRPA and CRISPR/Cas-powered technology in one-tube reaction system. Visual readouts were displayed using either a fluorescence detector (Fluor-based assay) or lateral flow dipsticks (LDF-based assay). As expected, this optimized assay exhibited high specificity towards only FHV-1 and FCV without cross-reactivity with other feline pathogens while achieving accurate detection for both targets with limit of detection at 2.4 × 10- 1 copies/µL for the FHV-1 TK gene and 5.5 copies/µL for the FCV ORF1 gene, respectively. Furthermore, field detection was conducted using the dRPA-Cas12a/Cas13a assay and the reference real-time PCR methods for 56 clinical samples collected from cats with URTD. Comparatively, the results of Fluor-based assay were in exceptional concordance with the reference real-time PCR methods, resulting in high sensitivity (100% for both FHV-1 and FCV), specificity (100% for both FHV-1 and FCV), as well as consistency (Kappa values were 1.00 for FHV-1 and FCV). However, several discordant results for FHV-1 detection were observed by LDF-based assay, which suggests its prudent use and interpretaion for clinical detection. In spite of this, incorporating dRPA-Cas12a/Cas13a assay and visual readouts will facilitate rapid and accurate detection of FHV-1 and FCV in resource-limited settings. CONCLUSIONS: The one-tube dRPA-Cas12a/Cas13a assay enables simultaneously ultrasensitive and visual detection of FHV-1 and FCV with user-friendly modality, providing unparalleled convenience for FHV-1 and FCV co-infection surveillance and decision-making of URTD management.

Associated factors for fear of COVID-19 scale in long-term care settings in Spain.

OBJECTIVE: To examine the relationship between the Fear of COVID-19 Scale (FCV-19S) score and sociodemographic, health, emotional and behavioural factors, in a cross-sectional observational study in 447 older adults living in long-term care (LTC) settings in Madrid (Spain). METHODS: The sample was stratified by nursing home ownership, geographical location, and size. Multiple linear regression analysis was used using backward elimination to identify factors that explained associations with fear, and logistic regression models were used to examine its role as a predictor of adherence to preventive measures. RESULTS: The mean age of the participants was 83.8 years, most were female, had had COVID-19, and were worried about the pandemic. The average score of the FCV-19S was 18.36 (SD: 8.28; range: 7-35), and the variables associated in the multiple linear regression model (explained variance: 34.00%) were being female, lower level of education, satisfaction with life and the residential home, and higher worry about the pandemic. The logistic regression models showed that fear of COVID-19 was a predictor of adherence to preventive measures like wearing facemasks, washing hands, and avoiding physical contact. CONCLUSIONS: fear of COVID-19 was significantly related with sex and subjective factors as life satisfaction and worry; and it influences older people's preventive behaviour. Interventions aimed at reducing fear and promoting adherence to preventive measures would improve their mental health and well-being.

Development and application of a dual ERA method for the detection of Feline Calicivirus and Feline Herpesvirus Type I.

Feline calicivirus (FCV) and feline herpesvirus type I (FHV-1) are the most common viral pathogens responsible for cat respiratory diseases, and coinfection with these two pathogens is often found. In veterinary clinics, the main diagnostic methods for FCV and FHV-1 are test strips and polymerase chain reaction (PCR). However, the sensitivity of test strips are not sufficient, and PCR is time-consuming. Therefore, developing a rapid and high-performance clinical diagnostic test is imperative for the prevention and treatment of these diseases. Enzymatic recombinase amplification (ERA) is an automated isothermal nucleic acid amplification technique that maintains a constant temperature, and is both rapid and highly accurate. In this study, a dual ERA method was developed using the Exo probe for a differential detection of FCV and FHV-1. This dual ERA method demonstrated high performance with the detection limit of 101 copies for both viruses, and no cross-reactions with feline parvovirus virus and F81 cells. To test the utility of the method for clinical applications, 50 nasopharyngeal swabs from cats with respiratory symptoms were collected and tested. The positive rates of FCV and FHV-1 were 40% (20/50, 95% confidence interval [CI], 26.4 to 54.8%) and 14% (7/50, 95% CI, 5.8 to 26.7%), respectively. The rate of coinfection with FCV and FHV-1 was 10% (5/50, 95% CI, 3.3 to 21.8%). These results were in agreement with those found using quantitative real-time PCR. Therefore, this dual ERA method is a novel and efficient clinical diagnostic tool for FCV and FHV-1 detection.

Induction of COX-2 by feline calicivirus via activation of the MEK1-ERK1/2 pathway, and attenuation of feline lung inflammation and injury by MEK1 inhibitor AZD6244 (selumetinib).

Feline calicivirus (FCV) is an important and highly prevalent pathogen of cats that causes acute infectious respiratory disease. Here it is shown in vitro that FCV induces the production of cyclooxygenase-2 (COX-2) through the MEK1-ERK1/2 signaling pathway. Screening of FCV proteins revealed that FCV non-structural protein VPg enhanced COX-2 mRNA expression and protein production in CRFK cells in a concentration-dependent manner. Regions 24-54aa and 84-111aa in FCV VPg were essential for up-regulation. In vivo, COX-2 and IL-6 production caused by FCV infection of kittens was significantly suppressed by the MEK1 inhibitor AZD6244 (selumetinib) and lung inflammation and injury were practically eliminated, with body temperature being returned to normal. AZD6244 may therefore find application as an effective therapeutic agent for the treatment of FCV infection.

Feline Calicivirus Proteinase-Polymerase Protein Degrades mRNAs To Inhibit Host Gene Expression.

To replicate efficiently and evade the antiviral immune response of the host, some viruses degrade host mRNA to induce host gene shutoff via encoding shutoff factors. In this study, we found that feline calicivirus (FCV) infection promotes the degradation of endogenous and exogenous mRNAs and induces host gene shutoff, which results in global inhibition of host protein synthesis. Screening assays revealed that proteinase-polymerase (PP) is a most effective factor in reducing mRNA expression. Moreover, PP from differently virulent strains of FCV could induce mRNA degradation. Further, we found that the key sites of the PP protein required for its proteinase activity are also essential for its shutoff activity but also required for viral replication. The mechanism analysis showed that PP mainly targets Pol II-transcribed RNA in a ribosome-, 5' cap-, and 3' poly(A) tail-independent manner. Moreover, purified glutathione S-transferase (GST)-PP fusion protein exhibits RNase activity in vitro in assays using green fluorescent protein (GFP) RNA transcribed in vitro as a substrate in the absence of other viral or cellular proteins. Finally, PP-induced shutoff requires host Xrn1 to complete further RNA degradation. This study provides a newly discovered strategy in which FCV PP protein induces host gene shutoff by promoting the degradation of host mRNAs. IMPORTANCE Virus infection-induced shutoff is the result of targeted or global manipulation of cellular gene expression and leads to efficient viral replication and immune evasion. FCV is a highly contagious pathogen that persistently infects cats. It is unknown how FCV blocks the host immune response and persistently exists in cats. In this study, we found that FCV infection promotes the degradation of host mRNAs and induces host gene shutoff via a common strategy. Further, PP protein for different FCV strains is a key factor that enhances mRNA degradation. An in vitro assay showed that the GST-PP fusion protein possesses RNase activity in the absence of other viral or cellular proteins. This study demonstrates that FCV induces host gene shutoff by promoting the degradation of host mRNAs, thereby introducing a potential mechanism by which FCV infection inhibits the immune response.

The Fear of COVID-19 Infection in Older People.

OBJECTIVE: To determine the levels of COVID-19-related fear and to investigate fear-associated factors among older people. METHODS: This study was conducted with patients aged 65 years and older from the Family Medicine Clinic of Health Sciences University Sisli Hamidiye Etfal Hospital over a 1-month period. A telephone survey was administered to evaluate patients' sociodemographic data and knowledge level on the COVID-19 pandemic and the degree of its impact. The fear levels of participants were determined using the fear of COVID-19 scale (FCV-19 S). Statistical analysis was performed using SPSS 15.0. A p value of < 0.05 was considered to be statistically significant. RESULTS: The study included 315 participants: 178 were female and 137 were male. The mean age was 71.5 ± 5.6 (min: 65, max: 94) years, and 26 participants were living alone. Moreover, 47.6% participants considered they had sufficient information about the COVID-19 outbreak, and 61.6% received information about the pandemic from television and 22.2% from their inner circle. While 11.7% participants considered they would require psychological support after the COVID-19 pandemic, 30.8% had sleep disorders for the last month. The mean FCV-19 S score was 16.0 ± 6.4; the FCV-19 S scores were statistically higher in participants who were women, living alone, had partial information about the COVID-19 pandemic, had sleep disorders for the last month, and were in requirement of psychological support after the pandemic. CONCLUSION: It is necessary to screen the older people for the COVID-19-related fear and accompanying psychological disorders and to develop appropriate intervention programs for individuals at risk.

Establishment and application of ERA-LFD method for rapid detection of feline calicivirus.

Feline calicivirus (FCV) has a single-stranded, positive-sense RNA genome, and it is responsible for many infectious respiratory diseases in cats. In addition, more worryingly, highly virulent strains of FCV can cause high mortality in felines. Therefore, a rapid and reliable diagnosis tool plays an important role in controlling the outbreak of FCV. In this study, enzymatic recombinase amplification (ERA) assay combined with lateral flow dipstick (LFD) was developed for the detection of FCV, targeting a relatively conversed position of FCV-ORF1. The results showed that the optimal reaction condition was at 40 °C for 30 min. ERA-LFD method was highly sensitive with the detection limit as low as 3.2 TCID50 of FCV RNA per reaction. The specificity analysis demonstrated no cross-reactivity with feline parvovirus (FPV), feline herpesvirus (FHV) and feline infectious peritonitis virus (FIPV). ERA-LFD was highly repeatable and reproducible, with the intra-assay and inter-assay coefficients of variation for this method both less than 7%. The general test showed that all the recombinant plasmids with known mutant sites and FCV strains with different mutant sites stored in our laboratory were all detected by this method. Of the 23 samples, 14 samples were tested positive for FCV by ERA-LFD and RT-qPCR, respectively. In summary, ERA-LFD assay was a fast, accurate and convenient diagnosis tool for the detection of FCV. KEY POINTS: • The detection principle of ERA-LFD was introduced. • Almost all the currently known FCV strains can be detected. • ERA-LFD is easy to operate and can be used for field detection.

Development of a triple NanoPCR method for feline calicivirus, feline panleukopenia syndrome virus, and feline herpesvirus type I virus.

BACKGROUND: Feline calicivirus (FCV), Feline panleukopenia virus (FPV), and Feline herpesvirus type I (FHV-1) are the three most common pathogens in cats, and also are the main pathogens leading to the death of kittens. Here, by a combination of gold nanoparticles and conventional PCR, we established a novel triple NanoPCR molecular detection method for clinical detection. RESULTS: The triple NanoPCR molecular detection is able to detect 2.97 × 101copies/µL FCV recombinant copies plasmid per reaction, 2.64 × 104copies/µL FPV recombinant copies plasmid per reaction, and 2.85copies/µL FHV-1 recombinant copies plasmid per reaction at the same time. The sensitivity of each plasmid is 100 times, 10 times, and 100 times higher than conventional PCR, respectively. The clinical results showed that among the 38 samples, the positive rates of FCV, FPV, and FHV-1 in a NanoPCR test were 63.16, 31.58, and 60.53%, while in a conventional PCR were 39.47, 18.42, and 34.21%. CONCLUSIONS: In this report, it is the first time that NanoPCR assays are applied in the detection of FCV, FPV, and FHV-1 as well. This sensitive and specific NanoPCR assay can be widely used in clinical diagnosis and field monitoring of FCV, FPV, and FHV-1 infections.

Parallel exploratory and confirmatory factor analysis of the Hungarian Fear of COVID-19 Scale in a large general population sample: a psychometric and dimensionality evaluation.

BACKGROUND: This study aims to confirm validity and reliability of the Hungarian version of Fear of COVID-19 Scale (FCV-19S) and evaluate its dimensional structure. METHODS: Cross-sectional survey was carried out in 2021 among Hungarian general population. In addition to classical test theory methods, construct dimensionality of FCV-19S was assessed using EFA with principal axis factoring method and CFA with diagonally-weighted least squares estimation. Fear score was compared in age, gender, educational level, vaccination and infection subgroups. RESULTS: Significant differences in FCV-19S mean scores were observed between three subgroups (age, gender, vaccination). Items showed good internal consistency (Cronbach α = 0.88). EFA identified two latent factors (eig = 4.2 and 1.02), though parallel analysis supports the one-factor model. The two-dimensional structure was confirmed by CFA, items 3,4,6,7 correlated with Factor 1 (physiological fear), items 1,2,5 with Factor 2 (emotional fear). CONCLUSION: The Hungarian version of FCV-19S seems valid and reliable. The EFA identified two-latent factors (emotional and physiological fear), that was confirmed by CFA. The two-factor structure had better model fit, though its' acceptance is limited.

Evaluation of COVID-19 fear and quality of life in patients with haematopoietic stem cell transplantation during the COVID-19 pandemic.

OBJECTIVE: We aimed to determine the coronavirus disease 2019 (COVID-19) fear state in haematopoietic stem cell transplant patients and to examine its relationship with quality of life. METHODS: In this prospective study, 64 patients who underwent HSCT during the pandemic were included. The COVID-19 fear situation was evaluated with the Fear of COVID-19 Scale (FCV-19S). Quality of life was evaluated with the European Organisation for Quality of Life Research and Treatment Core Questionnaire (EORTC QLQ-C30) (Version 3). RESULTS: The median FCV-19S score was 16.5 (12.0-22.0). The FCV-19S score was significantly higher in urban residents than rural residents. The general health score was 59.64 ± 20.04. The strongest positive correlation between fear level and life quality was found in emotional function. A weak, significant, positive correlation was observed between role function, nausea-vomiting, pain, appetite loss and fear level. CONCLUSION: FCV-19S is a quick, safe and valid tool that can be used to determine the COVID-19 fear level in vulnerable patient groups such as HSCT patients and to direct them to the necessary psycho-oncological support.

14-day famciclovir treatment significantly reduces the incidence of postherpetic neuralgia in elderly patients with herpes zoster.

WHAT IS KNOWN AND OBJECTIVE: Pain is the main symptom of herpes zoster (HZ), whilst postherpetic neuralgia (PHN) is a long-term unbearable pain, which seriously affects the quality of life of patients and is also the most intractable problem for clinicians. Early antiviral treatment is considered as a key measure to reduce acute pain and PHN. Nevertheless, most patients still have long-term pain after 7 days of antiviral treatment, and some patients will develop PHN. This study aimed to investigate whether prolonged duration of antiviral therapy could reduce HZ acute pain and the occurrence of PHN. METHODS: The outpatient data of HZ patients over 50 years old who visited the Dermatology Department from January 2016 to May 2018 were retrospectively analysed. According to the different courses of treatment of famciclovir (FCV), the patients were divided into 7-day FCV group and 14-day FCV group. The numerical rating scale (NRS) score at the first visit and on the 7th, 14th and 21st days after the start of treatment, the adverse drug reactions and the incidence of PHN were compared between the two groups. RESULTS: A total of 219 patients were involved in the analysis. For acute pain control, the 14-day FCV group was better than the 7-day FCV group. For patients with mild initial pain, there was no significant difference in NRS between the two treatments. For patients with moderate-to-severe initial pain, the NRS in the 14-day FCV group was significantly lower than that of the 7-day FCV group on the 14th and 21st days after starting treatment. PHN occurred in patients with moderate-to-severe initial pain, and the incidence was significantly lower in the 14-day FCV group than in the 7-day FCV group. There was no significant difference in the number of adverse reactions between the two groups. WHAT IS NEW AND CONCLUSION: Compared with the traditional 7-day antiviral therapy, the 14-day course of FCV can reduce the acute pain and the incidence of PHN in elderly patients with HZ, especially in patients with moderate to severe initial pain. Prolonging the course of medication did not increase the side effects.

Major Capsid Protein Synthesis from the Genomic RNA of Feline Calicivirus.

Caliciviruses have a positive-strand RNA genome with a length of about 7.5 kb that contains 2, 3, or 4 functional open reading frames (ORFs). A subgenomic mRNA (sg-RNA) is transcribed in the infected cell, and both major capsid protein viral protein 1 (VP1) and minor capsid protein VP2 are translated from the sg-RNA. Translation of proteins from the genomic RNA (g-RNA) and from the sg-RNA is mediated by the RNA-linked viral protein VPg (virus protein, genome linked). Most of the calicivirus genera have translation mechanisms leading to VP1 expression from the g-RNA. VP1 is part of the polyprotein for sapoviruses, lagoviruses, and neboviruses, and a termination/reinitiation mechanism was described for noroviruses. Vesiviruses have no known mechanism for the expression of VP1 from the g-RNA, and the Vesivirus genus is the only genus of the Caliciviridae that generates VP1 via a precursor capsid leader protein (LC-VP1). Analyses of feline calicivirus (FCV) g-RNA translation showed a low level of VP1 expression with an initiation downstream of the original start codon of LC-VP1, leading to a smaller, truncated LC-VP1 (tLC-VP1) protein. Deletion and substitution analyses of the region surrounding the LC-VP1 start codon allowed the identification of sequences within the leader protein coding region of FCV that have an impact on VP1 translation frequency from the g-RNA. Introduction of such mutations into the virus showed an impact of strongly reduced tLC-VP1 levels translated from the g-RNA on viral replication.IMPORTANCE Caliciviruses are a cause of important diseases in humans and animals. It is crucial to understand the prerequisites of efficient replication of these viruses in order to develop strategies for prevention and treatment of these diseases. It was shown before that all caliciviruses except vesiviruses have established mechanisms to achieve major capsid protein (VP1) translation from the genomic RNA. Here, we show for the first time that a member of the genus Vesivirus also has a translation initiation mechanism by which a precursor protein of the VP1 protein is expressed from the genomic RNA. This finding clearly points at a functional role of the calicivirus VP1 capsid protein in early replication, and we provide experimental data supporting this hypothesis.

Fear and depression during the COVID-19 outbreak in Cameroon: a nation-wide observational study.

BACKGROUND: The COVID-19 pandemic has been associated with significant psychological and social distress worldwide. We investigated fear and depression among adults in Cameroon during different phases of the COVID-19 outbreak. METHODS: An online survey was conducted in Cameroon from June-December 2020 using a structured questionnaire. Socio-demographic data and information regarding COVID-19 history were obtained. Fear and depressive symptoms were assessed using the Fear of COVID-19 score (FCV-19S) and the Patient Health Questionnaire (PHQ-9), respectively. Responses were clustered in weeks to better appreciate their evolution over time. RESULTS: Overall, 7381 responses from all ten regions of Cameroon were analysed (median age: 30 years, 73.3% male). The prevalence of depression (PHQ-9 score ≥ 10) was 8.4%, and that of high fear of COVID-19 (FCV-19S scores ≥19) was 57.4%. These rates were similar across genders, age-groups, and region of residence. While mean weekly PHQ-9 scores remained fairly stable throughout the study period (range: 2.53-3.21; p = 0.101), mean FCV-19S scores were highest during the early weeks but decreased significantly thereafter (from 20.31 to 18.34; p <  0.001). Multivariate analyses revealed that having a postgraduate degree, a history of quarantine, flu-like symptoms during the past 14 days, and higher FCV-19S scores were associated with more severe depressive symptoms, while obtaining COVID-19 information from various sources reduced the odds for depression. CONCLUSION: Depression amidst the COVID-19 crisis is less prevalent in Cameroon than in other countries. Prompt and widespread dissemination of adequate COVID-19 information may reduce the risks for depression by dispelling fear and anxiety among Cameroonians.

Exploring fear of COVID-19 and its correlates among older adults in Bangladesh.

OBJECTIVE: This study was aimed to assess the perceived fear of COVID-19 and its associated factors among older adults in Bangladesh. METHODS: This cross-sectional study was conducted in October 2020 among 1032 older Bangladeshi adults aged ≥60 years. A semi-structured questionnaire was used to collect information on participants' characteristics and COVID-19 related information. Perceived fear of COVID-19 was measured using the seven-item Fear of COVID-19 Scale (FCV-19S), where the cumulative score ranged from 7 to 35. Multiple linear regression was performed to identify factors associated with perceived fear of COVID-19. RESULTS: The mean fear score was 19.4. Participants who were concerned about COVID-19 (ß: 2.75, 95% CI: 1.71 to 3.78) and overwhelmed by COVID-19 (ß: 3.31, 95% CI: 2.33 to 4.29) were significantly more likely to be fearful of COVID-19. Moreover, older adults who felt themselves isolated from others and whose close friends and family members were diagnosed with COVID-19 were more fearful. However, the participants who received COVID-19 related information from the health workers had a lower level of fear (ß: -1.90, 95% CI: - 3.06 to - 0.73). CONCLUSIONS: The presence of overwhelming fear of COVID-19 among the older adults of Bangladesh underlines the psychological needs of these vulnerable groups. Health workers have a key role in addressing these needs and further research is needed to identify the effective strategies for them to use.

Antiviral Potential of Plants against Noroviruses.

Human noroviruses, which belong to the enterovirus family, are one of the most common etiological agents of food-borne diseases. In recent years, intensive research has been carried out regarding the antiviral activity of plant metabolites that could be used for the preservation of fresh food, because they are safer for consumption when compared to synthetic chemicals. Plant preparations with proven antimicrobial activity differ in their chemical compositions, which significantly affects their biological activity. Our review aimed to present the results of research related to the characteristics, applicability, and mechanisms of the action of various plant-based preparations and metabolites against norovirus. New strategies to combat intestinal viruses are necessary, not only to ensure food safety and reduce infections in humans but also to lower the direct health costs associated with them.

Ìn situ inactivation of human norovirus GII.4 by cold plasma: Ethidium monoazide (EMA)-coupled RT-qPCR underestimates virus reduction and fecal material suppresses inactivation.

Cold atmospheric-gaseous plasma (CAP) is an emerging non-thermal technology for decontamination of foodborne bacterial and viral pathogens. We obtained a >5 log10 reduction in the titer (TCID50) of feline calicivirus (FCV) on stainless steel discs and Romaine lettuce leaves after 3 min wet exposure to air plasma generated by a two-dimensional array of integrated coaxial-microhollow dielectric barrier discharge (2D-AICM-DBD). However, when human norovirus (HuNoV GII.4) was treated for 5 min under the same conditions, ~2.6 log10 (>99.5%) reduction in genome copy number was observed as measured by ethidium monoazide-coupled RT-qPCR (EMA-RT-qPCR). To assess this discrepancy, we studied CAP's effect on FCV by the cell culture method and by the EMA-coupled RT-qPCR method. It was found that the molecular titration method (EMA-RT-qPCR) underestimates the level of virus reduction by CAP. Additionally, the fecal matter present in HuNoV samples partially suppressed virucidal activity of CAP. Assuming that the lower virus reduction measured by EMA-RT-qPCR method compared to cell culture method for FCV is the same as for HuNoV, we can conclude that FCV may be used as a surrogate for HuNoV to assess the virucidal effect of CAP. CAP is able to inactivate 3.5 Log10 units of HuNoV at low titers after 2 min of exposure.

MOLECULAR DETECTION OF INFECTIOUS PATHOGENS OF THE UPPER RESPIRATORY TRACT IN CAPTIVE NONDOMESTIC FELIDS.

Upper respiratory tract disease is a complex infectious disease process with multiple pathogens involved. Identification of infectious agents in wild animals is of great importance for wildlife conservation. The aim of this study was to evaluate the molecular detection of feline herpesvirus type 1, feline calicivirus (FCV), Bordetella bronchiseptica , Chlamydophila felis , and Mycoplasma felis using ocular and nasal swabs in three species of captive nondomestic felids. Mycoplasma felis was detected in two ocular samples of Puma concolor and in one nasal sample of one Panthera onca . FCV was detected in association with M. felis in one P. concolor . The other pathogens tested were not detected. To the authors' knowledge, this is the first report of M. felis in nondomestic felids from Brazil.

The visualisation and quantification of human gastrointestinal fat distribution with MRI: a randomised study in healthy subjects.

We aimed to study the fate of fat during digestion. For this purpose, we validated and investigated the non-invasive quantification of gastric and duodenal fat emptying and emulsion processing (creaming and phase separation) using the MRI method iterative decomposition with echo asymmetry and least squares estimation (IDEAL). In total, twelve healthy subjects were studied on two separate visits in a single-blind, randomised, cross-over design study. IDEAL was utilised to repeatedly acquire quantitative fat fraction maps of the gastrointestinal tract after infusion of one of two fat emulsions: E1 (acid stable, droplet size 0·33 mm) and E4 (acid unstable, 0·38 mm). In vitro and in vivo validation was carried out using diluted emulsion and gastric content samples, respectively, and resulted in Lin's concordance correlation coefficients of 1·00 (95% CI 0·98, 1·00) and 0·91 (95% CI 0·87, 0·94), respectively. Fat fraction maps and intragastric emulsion profiles enabled the identification of features of intraluminal phase separation and creaming that were not visible in conventional MRI. Gastric fat emptying was faster for E4 compared with E1 with a difference of 2·5 (95% CI 1·9, 3·1) ml/h. Duodenal content volumes were larger for E1 than for E4 with a difference of 4·9 (95% CI 3·9, 8·5) ml. This study demonstrated that with IDEAL it was possible (1) to visualise the intragastric and duodenal fat distribution and (2) to quantify the differences in emptying, phase separation and creaming of an acid-stable and an acid-unstable emulsion. This method has potential to bridge the gap between current in vitro digestive models and in vivo behaviour and to be applied in the development of effective functional foods.

Antiviral effects of grape seed extract against feline calicivirus, murine norovirus, and hepatitis A virus in model food systems and under gastric conditions.

Grape seed extract (GSE) has antiviral activities against hepatitis A virus (HAV) and human norovirus surrogates (feline calicivirus (FCV-F9) and murine norovirus (MNV-1)). The objectives of this study were to determine (1) time and dose-dependence of GSE against FCV-F9, MNV-1, and HAV at room temperature (RT) and 37 °C over 24 h; and (2) GSE effects in model foods (apple juice (AJ) and 2% milk) and simulated gastric conditions at 37 °C. Viruses at ∼5 log PFU/ml were treated with 0.5-8 mg/ml GSE prepared in water, AJ, milk or gastric juices, or water over 24 h at RT or 37 °C. Infectivity of triplicate treatments was evaluated using plaque assays. GSE effects increased with time and concentration. GSE at 1 mg/ml in AJ reduced MNV-1 to undetectable levels after 1 h and by 1 log in milk after 24 h. GSE at 1 and 2 mg/ml in AJ reduced HAV to undetectable levels after 1 h, while 2 and 4 mg/ml GSE in milk caused ∼1 log reduction after 24 h. GSE at 2 mg/ml in intestinal fluid reduced FCV-F9, MNV-1 and HAV to undetectable levels after 6 h. GSE appears to be a suitable natural option for foodborne viral reduction.