Ability to detect antibodies to beak and feather disease virus in blood on filter paper decreases with duration of storage

Background Beak and feather disease virus (BFDV) is a circovirus that infects captive and wild psittacine birds, and is of conservation concern. The haemagglutination inhibition (HI) assay is used to determine antibody titres against BFDV, and the use of dried blood spots (DBS) on filter paper stored at room temperature has been suggested to be an equally valid technique to the use of frozen serum. However, research on other pathogens has found variable results when investigating the longevity of antibodies stored on DBS at room temperature. Consequently, we aimed to test the temporal stability of antibodies to BFDV in DBS samples stored long-term at room temperature. A further goal was to add to the current knowledge of antibody response to naturally acquired BFDV infection in crimson rosellas (Platycercus elegans). Methods Blood was collected from wild P. elegans in Victoria, Australia, that had been live-trapped (n = 9) or necropsied (n = 11). BFDV virus load data were obtained from blood stored in ethanol by real-time quantitative PCR (qPCR); antibody titres were obtained by HI assay from either DBS or serum samples, which had been collected concurrently. All HI assays were performed commercially by the Veterinary Diagnostic Laboratory (VDL) in Charles Sturt University, Australia, who were blind to BFDV blood status. Results HI titres from DBS stored at room temperature declined significantly over time (~80 weeks). By contrast, frozen serum samples assayed after 80 weeks in storage all had high HI titres, only varying up to one dilution step from the initial HI titres obtained from DBS at 3–6 weeks after sampling. Weak HI titres from DBS samples all came back negative when the test was repeated only nine weeks later. Novel high HI titres were reported in P. elegans, and while most birds with high antibody titres had corresponding negative qPCR results, a single subadult presented with high HI titres and virus load simultaneously. Conclusion Detection of antibodies on filter paper stored at room temperature decreases over time, increasing the chances of false negatives in these samples, and in repeated testing of samples with weak HI titres. Consequently, serum should be the preferred sample type to use for seroepidemiological studies on BFDV in parrots and other bird species. When not possible, it may help to store DBS on filter paper at −20 °C or lower. However, prompt testing of DBS samples (e.g., <6 weeks in storage) is recommended pending further research on antibody temporal stability. We also show that P. elegans, especially adults, can produce high antibody titres against BFDV, which may help them resist infection.

Effect of Relative Humidity in Air on the Transmission of Respiratory Viruses

Viral respiratory infections have plagued mankind over its known history. Unfortunately, there has been a lack of meaningful progress in preventing the spread of viral respiratory infections globally. The central dogma appears to be that viruses are the villains. This framing focuses on a viral load balance (VLB) in the air. It follows that physical dilution through various means have been the primary focus of attempts to reduce the spread of infections. The problem of obesity provides a good example of how paradigm blindness can slow down progress in a field. Obesity has been framed as an energy balance disorder that blames overeating and lack of exercise for weight gain. Reframing obesity as a disorder of fat metabolism and storage caused by the quantity and quality of carbohydrates in the diet, referred to as the carbohydrate-insulin model (CIM), opened an alternative line of questioning with a testable hypothesis. Similarly, we postulate an alternative way to frame the spread of viral respiratory infections that would lead to new insights and potentially new ways to prevent infections. It has long been recognized that viral respiratory infections show a pronounced seasonal variation, referred to as seasonal forging, such that they increase in the winter but decrease or virtually disappear in the summer. In temperate regions, people spend over 90% of their time indoors. This is, therefore, where most respiratory infections are expected to occur. Evidence has been accumulating for decades on the strong correlation between variations in indoor relative humidity (RH) and variations in infection rates. Within a RH Goldilocks zone of 40%-60%, encapsulated viruses like influenza and SARS are optimally inactivated outside the infected host. Below 40% and above 80%, viruses can survive for extended periods in the air or on surfaces. This may explain in part the seasonality of infections as the indoor level of RH in winter is typically about 20% and above 40% in summer in temperate regions. However, the mechanism for the inactivation at midrange RH (in summer) is not well understood. This paper offers a hypothesis that could explain these observations. We have demonstrated that H2O2 and other reactive oxygen species (ROS) are formed spontaneously at the water-air interface of pure water microdroplets. Using only water and a nebulizing gas in the presence of oxygen, we have demonstrated the significant disinfectant potential of pure water microdroplets caused by the activity of H2O2 and other ROS. We postulate that spontaneous H2O2 and ROS formation in viruses containing exhaled microdroplets have a similar virucidal effect at mid-range RH. The droplet evaporation rate is sufficient to concentrate the solutes and provide enough time for reactions to occur at significantly higher rates than in bulk solutions. The concentration of H2O2 has also been shown to be positively correlated to RH. In addition, several other ROS/RNS may be present or formed through interactions with H2O2 that may act as even more effective virucide disinfectants to inactivate the virus. Below RH 40% evaporation happens too rapidly for these reactions to make an impact before the droplet is desiccated, and above RH 80% the solutes remain too diluted. Rapid inactivation of viruses at midrange RH may therefore play a greater role in preventing infections than physical dilution of virus load in the air through excessive mechanical ventilation. Similar to obesity, we suggest that a new paradigm that considers virus infectivity outside the host rather than the virus load balance in the air alone could greatly contribute to our understanding of respiratory infections. The proposed new “Relative Humidity Infectivity” RHI paradigm could explain the causal mechanisms underlying seasonal respiratory infections. This can point to better prevention strategies that avoid further distortion of our indoor environment and create conditions within which humans can thrive and be optimally protected. We need more focus on testing the various hypotheses and more data to determine which of the two paradigms will lead us in the right direction or how to use the best of both in an optimal combination. The stakes cannot be higher, and the potential for eradicating future viral respiratory pandemics with nature-based solutions may be right under our noses, literally.

Low Dose Pig Anti-Influenza Virus Monoclonal Antibodies Reduce Lung Pathology but Do Not Prevent Virus Shedding

We have established the pig, a large natural host animal for influenza, with many physiological similarities to humans, as a robust model for testing the therapeutic potential of monoclonal antibodies (mAbs). In this study we demonstrated that prophylactic intravenous administration of 15 mg/kg of porcine mAb pb18, against the K160–163 site of the hemagglutinin, significantly reduced lung pathology and nasal virus shedding and eliminated virus from the lung of pigs following H1N1pdm09 challenge. When given at 1 mg/kg, pb18 significantly reduced lung pathology and lung and BAL virus loads, but not nasal shedding. Similarly, when pb18 was given in combination with pb27, which recognized the K130 site, at 1 mg/kg each, lung virus load and pathology were reduced, although without an apparent additive or synergistic effect. No evidence for mAb driven virus evolution was detected. These data indicate that intravenous administration of high doses was required to reduce nasal virus shedding, although this was inconsistent and seldom complete. In contrast, the effect on lung pathology and lung virus load is consistent and is also seen at a one log lower dose, strongly indicating that a lower dose might be sufficient to reduce severity of disease, but for prevention of transmission other measures would be needed.

Effects of an alphasatellite on life cycle of the nanovirus Faba bean necrotic yellows virus

Nanoviruses are plant viruses with a multipartite single-stranded DNA (ssDNA) genome. Alphasatellites are commonly associated with nanovirus infections, but their putative impact on their helper viruses is unknown. In this study, we investigated the role of subterranean clover stunt alphasatellite 1 (hereafter named SCSA 1) on various important traits of faba bean necrotic yellows virus (FBNYV) in its host plant Vicia faba and aphid vector Acyrthosiphon pisum , including disease symptoms, viral accumulation and transmission. The results indicate that SCSA 1 does not affect the symptom severity nor the overall FBNYV accumulation in V. faba, but changes the relative amounts of its different genomic segments. Moreover, the association of SCSA 1 with FBNYV increases the rate of plant-to-plant transmission by a process seemingly unrelated to simple increase of the viral accumulation in the vector. These results represent the first study on the impact of an alphasatellite on the biology of its helper nanovirus. They suggest that SCSA 1 may benefit FBNYV, but the genericity of this conclusion is discussed and questioned. Importance Alphasatellites are circular single stranded DNA molecules frequently found in association with natural isolates of nanoviruses and some geminiviruse, the two ssDNA plant infecting virus families. While the implications of alphasatellite presence in geminivirus infections are relatively well documented, comparable studies on alphasatellites associated with nanoviruses are not available. Here we confirm that subterranean clover stunt alphasatellite 1 affects different traits of its helper nanovirus, faba bean necrotic yellows virus, both in the host plant and aphid vector. We show that the frequencies of the virus segments change in the presence of alphasatellite, in both plant and vector. We also confirm that while within-plant virus load and symptom are not affected by alphasatellite, the presence of alphasatellite decreases within-aphid virus load, but significantly increases virus transmission rate, so may confer a possible evolutionary advantage for the helper virus.

Effect of CRTH2 antagonism on the response to experimental rhinovirus infection in asthma: a pilot randomised controlled trial

Background and aimsThe chemoattractant receptor-homologous molecule expressed on T helper type 2 cells (CRTH2) antagonist timapiprant improved lung function and asthma control in a phase 2 study, with evidence suggesting reduced exacerbations. We aimed to assess whether timapiprant attenuated or prevented asthma exacerbations induced by experimental rhinovirus (RV) infection. We furthermore hypothesised that timapiprant would dampen RV-induced type 2 inflammation and consequently improve antiviral immune responses.MethodsAtopic patients with partially controlled asthma on maintenance inhaled corticosteroids were randomised to timapiprant (n=22) or placebo (n=22) and challenged with RV-A16 3 weeks later. The primary endpoint was the cumulative lower respiratory symptom score over the 14 days post infection. Upper respiratory symptoms, spirometry, airway hyperresponsiveness, exhaled nitric oxide, RV-A16 virus load and soluble mediators in upper and lower airways samples, and CRTH2 staining in bronchial biopsies were additionally assessed before and during RV-A16 infection.ResultsSix subjects discontinued the study and eight were not infected; outcomes were assessed in 16 timapiprant-treated and 14 placebo-treated, successfully infected subjects. There were no differences between treatment groups in clinical exacerbation severity including cumulative lower respiratory symptom score day 0–14 (difference 3.0 (95% CI −29.0 to 17.0), p=0.78), virus load, antiviral immune responses, or RV-A16-induced airway inflammation other than in the bronchial biopsies, where CRTH2 staining was increased during RV-A16 infection in the placebo-treated but not the timapiprant-treated group. Timapiprant had a favourable safety profile, with no deaths, serious adverse events or drug-related withdrawals.ConclusionTimapiprant treatment had little impact on the clinicopathological changes induced by RV-A16 infection in partially controlled asthma.

Low dose pig anti-influenza virus monoclonal antibodies reduce lung pathology but do not prevent virus shedding

We have established the pig, a large natural host animal for influenza, with many physiological similarities to humans, as a robust model for testing the therapeutic potential of monoclonal antibodies (mAbs). In this study we demonstrated that prophylactic intravenous administration of 15mg/kg of porcine mAb pb18, against the K160-163 site of the haemagglutinin, significantly reduced lung pathology and nasal virus shedding and eliminated virus from the lung of pigs following H1N1pdm09 challenge. When given at 1mg/kg, pb18 significantly reduced lung pathology and lung and BAL virus loads, but not nasal shedding. Similarly, when pb18 was given in combination with pb27, which recognised the K130 site, at 1mg/kg each, lung virus load and pathology were reduced, although without an apparent additive or synergistic effect. No evidence for mAb driven virus evolution was detected. These data indicate that intravenous administration of high doses was required to reduce nasal virus shedding, although this was inconsistent and seldom complete. In contrast the effect on lung pathology and lung virus load is consistent and is also seen at one log lower doses, strongly indicating that a lower dose might be sufficient to reduce severity of disease, but for prevention of transmission other measures would be needed.

Personalized Virus Load Curves for Acute Viral Infections

We introduce an explicit function that describes virus-load curves on a patient-specific level. This function is based on simple and intuitive model parameters. It allows virus load analysis of acute viral infections without solving a full virus load dynamic model. We validate our model on data from mice influenza A, human rhinovirus data, human influenza A data, and monkey and human SARS-CoV-2 data. We find wide distributions for the model parameters, reflecting large variability in the disease outcomes between individuals. Further, we compare the virus load function to an established target model of virus dynamics, and we provide a new way to estimate the exponential growth rates of the corresponding infection phases. The virus load function, the target model, and the exponential approximations show excellent fits for the data considered. Our virus-load function offers a new way to analyze patient-specific virus load data, and it can be used as input for higher level models for the physiological effects of a virus infection, for models of tissue damage, and to estimate patient risks.