Understanding the persistence of Influenza A virus in the external environment.

The pandemic potential and transmissibility of influenza A viruses (IAV) are contingent on their capacity to replicate effectively within an infected host, maintain their viability and their survival in the environment, and subsequently initiate infection in a new host [1]. Despite the fact that IAV are responsible for seasonal influenza and sporadic pandemics, there remains a significant lack of understanding regarding the persistence of influenza viruses in the external environment. There are existing disparities and gaps in knowledge when it comes to identifying the environmental factors that promote the survival and viability of IAV, as well as those that trigger the inactivation of these viruses. Assessing the risk of sustained human-to-human transmission of IAV and their survival in the external environment is vital for comprehending their potential to cause a pandemic and the associated implications for public health [1].

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1. Understanding IAV transmission

?To gain a better understanding of how the virus circulates and remains viable in the external environment, it is essential to comprehend the modes of influenza transmission. Influenza transmission can be categorized into short- range and long-range transmission [2]. In short-range transmission, an infected individual can transmit the virus to a susceptible person through various means, including direct contact, exhaled droplets, aerosols, and shared objects like phones. In long-range transmission scenarios, susceptible individuals can become exposed to the IAV either by inhaling aerosol particles released by the infected person or by coming into contact with contaminated fomites [2]. It is of utmost importance to identify IAV airborne particles. According to the definition provided by the Infectious Diseases Society of America [3], "respirable" particles are those smaller than 10μm, which can settle in both the lower and upper airways, while "inspirable" particles are those ranging from 10μm to 100μm, predominantly depositing in the upper airways situated in the head. Therefore, based on these criteria, aerosols represent infectious particles smaller than 100μm, while droplets represent infectious particles larger than 100μm [4]. IAV spread through contact, droplet spray, and aerosol transmission [3]. Contact transmission has two forms: direct, where the virus is passed through physical contact (e.g., touching), and indirect, where it is transferred via contaminated objects (e.g., banknotes) [3,5]. Droplet spray transmission occurs when individuals in close proximity release viral droplets into the air, such as when sneezing. These droplets cannot be inhaled; however, they can impact exposed mucous membranes [3,4]. Aerosol transmission involves person-to-person spread through virus-carrying aerosols of "inspirable" size or smaller. These tiny particles can be inhaled into the respiratory tract, reaching the trachea and lungs [3,4]. The “inspirable” aerosols (10-100 μm) tend to settle in the nasopharyngeal region, while only “respirable” aerosols that are small enough (<10 μm) can access and settle in the bronchial and alveolar region [4].

2. Factors that affect IAV persistence in the environment

2.1 Viral determinants

The persistence of IAV relies significantly on the stability of viral morphology characteristics, including the virus envelope, capsid, internal proteins, genomes, and the formation of viral aggregates. The influenza virus possesses a specific configuration, including two envelope proteins known as haemagglutinin (HA) and neuraminidase (NA). HA's role is to attach to sialic acid receptors found on respiratory epithelial cells, facilitating the virus's entry into these cells. On the other hand, NA assists the virus in navigating through the mucin layer in the respiratory tract to reach its target epithelial cells [6]. IAV RNA genomes undergo elevated mutation rates due to both antigenic drift and antigenic shift, resulting in a wide range of genomic variants [2,6]. This diversity may facilitate the virus's rapid adaptation to the host and its efficient transmission through respiratory droplets and aerosols [2]. Two viral envelope proteins, haemagglutinin (HA) and neuraminidase (NA), contribute to preserving viral infectivity and stability in the external environment [1,7]. In one study [7], researchers attempted to determine the viral molecular factors that impact the survival of IAV in the external environment. They observed the survival of two H1N1 strains, the 2009 pandemic strain and a seasonal strain, in saline water at 35°C. The pandemic strain, which had an additional amino acid at position 147 in its HA protein, displayed greater resilience in the external environment. It was noted that viral strains exhibiting elevated surface levels of HA also demonstrated enhanced stability in the external environment. Singanayagam et al. (2020) [1] made comparable findings in their study employing a ferret animal model. When animals were infected with the more stable H1N1 mutant (HA mutation E21K) in contrast to the less stable mutant (HA mutation Y17H), they produced a higher number of viral particles. These viral particles, when released as aerosols into the external environment, exhibited greater efficiency in infecting MDCK culture plates. Furthermore, Labadie et al. (2018) [7] noted that NA plays a role in the stability of IAV in the external environment. NA contains three calcium binding sites. Strains with mutations at calcium binding sites D341N and D382N exhibited heightened viral stability. These findings indicate that mutations in the molecular structure of HA and NA have a direct impact on the stability of IAV in the external environment.

2.2 Environmental determinants

2.2.1 Temperature and humidity

Influenza exhibits distinct transmission patterns across the globe. A study conducted by Tamerius et al. (2013) [8] gathered epidemiological data from 78 locations worldwide. The researchers discovered that in temperate climates, seasonal influenza epidemics tend to coincide with cold-dry weather conditions, characterized by specific humidity below 12 g/kg and temperatures below 18°C. In contrast, in tropical climates (within 12°N/S latitude), epidemics primarily occur during the humid-rainy season when specific humidity exceeds 20 g/kg [8]. Similar observations were noted in a study by Niazi et al. (2021) [9] that examined aerosols containing IAV within different relative humidity (RH) zones. In high RH conditions (>73%), IAV had a survival time of approximately 15 minutes, while infectious aerosols persisted three times longer in dry conditions (<43%). Nevertheless, under elevated RH conditions, the infectivity of IAV is greater than in low RH conditions. The study revealed that IAV's survival in various RH conditions was influenced by salt crystallization within the aerosols. In higher RH conditions, salts concentrated and adversely affected the viral lipid bilayer, nucleocapsid, and viral genome, resulting in shorter persistence in the external environment. This information can offer a practical solution for room air conditioning. One approach could involve configuring the air conditioning system to maintain humidity within the range of 40-60%, thus establishing unfavourable humidity conditions for IAV [9]. However, it is worth noting that this study had certain limitations. The carrier aerosol fluid used contained only PBS, FBS, and allantoic egg fluid, while in real-life scenarios, human respiratory mucus serves as a protective layer for IAV [5]. Additionally, the aerosol particles in the study were smaller than 5μm, making it impossible to assess the survival of larger aerosols. Considering that in subtropical zones (between 12°N/S and 25°N/S latitude), influenza epidemic dynamics appear to be more influenced by human migration patterns than by temperature and humidity [8] further research is needed to evaluate the impact of temperature and humidity on influenza persistence.

2.2.2 pH

The acidic pH has a notable impact on IAV survival in the air. Luo et al. (2023) [10] study made intriguing observations in this regard. They found that an acidic pH (≤5) decreased IAV infectivity and overall survival. However, aerosol particles containing synthetic lung fluid (SLF) or nasal mucus exhibited extended viability in acidic air. The study concluded that SLF and nasal mucus slowed the diffusion of Na+ and Cl+ ions, resulting in reduced water loss from aerosol particles and an extended its lifespan. Larger aerosol particles and droplets also took longer to reach low pH levels, contributing to their extended infectivity and persistence. The research revealed that a tenfold increase in particle size correlated with a tenfold increase in IAV inactivation time [10]. Furthermore, it was observed that in outdoor air, which tends to be more acidic, IAV inactivation was more pronounced compared to less acidic indoor air and purified air. This underscores the importance of regular room ventilation in reducing IAV infectivity and survival. In facilities like hospitals or museums where the use of purified air is necessary, and outdoor ventilation might not be feasible, acidifying the air could be a potential strategy to reduce IAV infectivity and persistence [10]. Acidic pH also influences the functional structure of HA. IAV strains with less stable HA mutations, when subjected to an acidic environment, exhibited structural unfolding of HA in the external environment. As a consequence, IAV lost its ability to infect MDCK cells in culture plates. These findings explain how the increased acidity of outdoor air can lead to a rapid inactivation of IAV [1,7].

2.2.3 Ultraviolet radiation (UV)

The understanding of the impact of UV radiation on the persistence of IAV in the outdoor environment is still limited. In a study conducted by Schuit et al. (2020) [11], IAV was subjected to three different lighting conditions, including full light exposure, 50% light exposure, and complete darkness. The findings from this research suggest that IAV exhibit a slower decay rate in conditions with lower light intensity, indicating that reduced light levels contribute to their prolonged survival.

2.2.4 Surface material

As previously mentioned, the transmission of IAV includes the fomite route. In a study by Greatorex et al. (2011) [12], it was observed that IAV could persist on a wide range of commonly used items, such as soft toys. Notably, surfaces that facilitated the virus's survival for up to eight hours included stainless steel, computer keyboards, and light switches made from polyvinyl chloride. Intriguing findings were also reported by Thomas et al. (2008) [5], where it was discovered that IAV could endure on banknotes for as long as 17 days, particularly when the infectious inoculum had a higher concentration (8.9 X 105 TCID50/ml) and when the viruses were coated in human respiratory mucus. These results from both studies emphasize the importance of taking preventive measures against fomite transmission. Implementing options such as contactless payment methods, automatic doors in public spaces, and the availability of hand sanitizers are just a few examples of such measures.

2.2.5 Airflow dynamics

Throughout the COVID-19 pandemic, it became common knowledge that maintaining a minimum distance of 2 meters was advisable in order to prevent the airborne spread of viruses [13]. However, it is now prudent to reassess this public health policy and consider extending the recommended social distance to more than 6 meters. In a study conducted by Dbouk and Drikakis (2020) [13], it was observed that under calm weather conditions, infectious aerosols could travel a maximum distance of 2 meters within 49 seconds, with their height reaching 1 meter at the 5-second timepoint. On the other hand, when wind speed reached 4 km/h during windy weather conditions, these aerosols were found to travel as far as 6 meters within just 5 seconds, and their maximum height (1.63 meters) was consistently reached at various time points between 1 second and 5 seconds. IAV can also affect gastrointestinal tract and spread through the faeces although it is less common route of infection [14]. A noteworthy discovery was reported by Li et al. in 2020 [15], where researchers found that when a toilet is flushed, infectious aerosols can be dispersed to a height of 94-106 cm within a time frame of 35-70 seconds. Based on these findings, several preventive measures should be put in place to reduce the risk of IAV transmission. These measures include the closure of toilet lids, maintaining good hand hygiene practices, and considering the installation of air conditioning systems in public restrooms [15].

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CONCLUSION

The transmission and survival of IAV in hosts and the environment are crucial for understanding their pandemic potential and public health impact. Despite IAV's well- known role in seasonal flu and occasional pandemics, there are significant knowledge gaps regarding its persistence in the environment. This article has identified these knowledge deficiencies and proposed practical non-pharmaceutical measures to reduce IAV transmission. Environmental conditions like temperature, humidity, pH, and UV radiation play a significant role in IAV survival. Recognizing their influence is crucial for outbreak prediction and management. Surface materials and fomite transmission highlight the role of common objects in virus spread, emphasizing the need for hygiene and preventive measures in public spaces. Airflow dynamics affect the transmission of infectious aerosols, prompting the re-evaluation of social distancing guidelines based on wind conditions. Finally, the less common faecal route of IAV transmission underscores the importance of additional preventive measures in public restrooms. In conclusion, our understanding of IAV persistence and transmission is evolving. Ongoing research and vigilance are essential to combat potential outbreaks. Public health measures informed by this knowledge can mitigate IAV transmission and reduce its impact on global health.

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