Persistence of the 2009 pandemic influenza A (H1N1) virus in water and on non-porous surface

Abstract

Knowledge of influenza A virus survival in different environmental conditions is a key element for the implementation of hygiene and personal protection measures by health authorities. As it is dependent on virus isolates even within the same subtype, we studied the survival of the 2009 H1N1 pandemic (H1N1pdm) virus in water and on non-porous surface. The H1N1pdm virus was subjected to various environmental parameters over time and tested for infectivity. In water, at low and medium salinity levels and 4°C, virus survived at least 200 days. Increasing temperature and salinity had a strong negative effect on the survival of the virus which remained infectious no more than 1 day at 35°C and 270 parts per thousand (ppt) of salt. Based on modeled data, the H1N1pdm virus retained its infectivity on smooth non-porous surface for at least 7 days at 35°C and up to 66 days at 4°C. The H1N1pdm virus has thus the ability to persist in water and on glass surface for extended periods of time, even at 35°C. Additional experiments suggest that external viral structures in direct contact with the environment are mostly involved in loss of virus infectivity.

Figure 1. Effect of the initial change on virus infectivity in water.

Loss of infectivity during the first 30 min in water for A/Paris/2590/2009 (H1N1)v (A) and A/New Caledonia/20/99 (H1N1) (B). The infectivity loss represented the difference between the T_theoretical, i.e. 10^7.28 TCID₅₀/mL, and the mean titre obtained after 30 min (T₀), according to different conditions of temperature and salinity. The mean titre corresponded to the titres made in parallel from the 2 aliquots of water samples. Maximal and minimal values are represented as error bars.

Figure 2. Effect of the initial change on virus infectivity on watch glass.

Loss of infectivity during the first 30 min (wet) and after complete drying of the viral suspension (dry) on watch glass for A/Paris/2590/2009 (H1N1)v (A) and A/New Caledonia/20/99 (H1N1) (B) strains. The infectivity loss represented i) the difference between the T_theoretical and the mean titre observed after 30 min (T_wet), and ii) the difference between the T_theoretical and the mean titre obtained after complete drying of the viral suspension (T₀). The mean titre corresponded to the titres made in parallel from the 2 watch glass samples. Maximal and minimal values are represented as error bars.

Figure 3. Viral survival in water at 4°C.

Viral persistence of A/Paris/2590/2009 (H1N1)v (—) and A/New Caledonia/20/99 (H1N1)(····) in water at 4°C. Linear regression for persistence at 0 ppt (A), 5 ppt (B), 35 ppt (C) and 270 ppt (D) are represented. TCID₅₀ values corresponded to the mean values of the titres made in parallel from the 2 aliquots of water samples.

Figure 4. Viral survival in water at 35°C.

Viral persistence of A/Paris/2590/2009(H1N1)v (—) and A/New Caledonia/20/99 (H1N1) (····) in water at 35°C. Linear regression for persistence of H1N1pdm in water at 0 ppt (A), 5 ppt (B), and 35 ppt (C) are represented. TCID₅₀ values corresponded to the mean values of the titres made in parallel from the 2 aliquots of water samples.

Figure 5. Endpoint RT-PCR targeting the whole M segment.

RT-PCR was performed for A/Paris/2590/2009 (H1N1)v (A) and A/New Caledonia/20/99 (H1N1) (B) and for selected conditions: at different days (designated as dx, x being the number of the day), different temperatures (for 4, 25 and 35°C respectively) and different levels of salinity (for 0, 35 and 270 ppt respectively). +: initial stock of A/Paris/2590/2009 (H1N1)v or A/New Caledonia/20/99 (H1N1) strain as a positive control. –: water as a negative control.

Figure 6. Viral survival kinetic on watch glass.

Viral persistence of A/Paris/2590/2009 (H1N1)v (—) and A/New Caledonia/20/99 (H1N1) (····) at 4 (•;○), 25 (▪;□) and 35°C (▴;Δ). Linear regressions for persistence during drying (A) and after drying (B) are represented. TCID₅₀ values corresponded to the mean values of the titres made in parallel from the 2 watch glass samples.