Generation of avian influenza virus (AIV) contaminated fecal fine particulate matter (PM_2.5): Genome and infectivity detection and calculation of immission

Abstract

As a model for aerosol transmission, chicken feces was spiked with avian influenza virus (AIV) subtype H10N7 and used to generate a fine particulate matter aerosol. For this an innovative aerosol chamber was developed, that collected PM_2.5 on quartz microfiber filters. With AIV contaminated PM_2.5 dust-coated filters different incubation times ranging from 0 to 4 days and storage mainly at +4 and +20 °C and at different relative humidity (RH) were performed. Embryonic death in inoculated hen's eggs with filter elute was the AIV infectivity read out. To determine viral genome presence quantitative real time RT-PCR was applied.

The filter elutes contained AIV genome as well as viable virus whereby +20 °C indicated a borderline temperature for infectious virus stability. In addition, high relative humidity was critical for AIV viability in PM_2.5. The results allowed a dispersion calculation of infectious AIV in aerosols assuming a worst case scenario for an AIV outbreak in poultry farms. Thus exposure to AIV associated with PM_2.5 is possible near to infected farms and may be a serious risk for fatal influenza disease in both man and animals. Airborne transmission should be effectively preventable by dispersion of water combined with disinfection into the inside air as well as the exhaust air stream of AIV infected farms.

Introduction

Some of the most highly contagious viruses of human and veterinary concern are airborne pathogens, e.g. human and avian influenza virus (AIV) and foot-and-mouth disease virus (FMDV). The highly pathogenic avian influenza viruses (HPAI) have been a threat to the poultry industry for many years. However, when the Asian H5N1 subtype emerged, a trans-boundary spread became evident and a new dimension of zoonotic danger and fear for evolution of a pandemic virus was raised (Belshe, 2005, Peiris et al., 2007, World Health Organization, 2007). After the first localized Asian H5N1 epidemic in Germany, the virus was spread rapidly and caused many small epidemics in wild birds. In 2007, the virus also was detected on duck raising farms (Rinder et al., 2007, Starick et al., 2007, Weber et al., 2007, Harder et al., 2009). The routes of transmission of Asian H5N1 and other HPAI subtypes among susceptible mammals and birds is still a matter of debate. Air borne transmission of infection cannot be excluded and might be an effective way of infection transmission when virus is inhaled directly in the deeper respiratory tract (Shinya et al., 2006, Tellier, 2006, Brankston et al., 2007, Nicholls et al., 2007). Fine particulate matter from AIV contaminated fecal dust can play an important role in the transmission and onset of infection since particles as small as PM_2.5 (particle size <2.5 μm) can directly invade the lower part of the respiratory tract (DFG, 2008). Rapid and direct transport of AIV into the lungs will be in favor for onset of infection (Shinya et al., 2006, Van Riel et al., 2006). The indoor exposure to aerosolized HPAI-AIV is equally given for humans working in poultry stables and susceptible birds. As workers can prevent virus inhalation by using appropriate masks birds are most endangered by aerosolized AIV within the flocks and possibly in nearby holdings.

Recently the stability and transmission of aerosolized Newcastle Disease Virus has been tested under practical conditions with chickens (Li et al., 2009). The results indicated that virus shed from infected animals readily aerosolized and airborne transmission to susceptible chickens was very efficient. We therefore wanted to know, (i) whether AIV in contaminated fecal dust can be found in the PM_2.5 fraction and, (ii) if this is the case, how long viable virus is present under environmental conditions.