| Animal airborne transmission viral diseases ever known are more than 20, such as Newcastle disease (ND), Avian Influenza (AI) and Foot-and-mouth disease (FMD), Porcine reproductive and respiratory syndrome (PRRS) and so on. AI is included in the World Organization (OIE) for Animal Health list of notifiable diseases. Direct contact and indirect contact—including aerosols or droplets and contaminants—are two important routes through which AI spread. However, the occurrence and transmission mechanisms of AIV aerosol remained unknown, and lack of experimental testify. In this study, an infection model was established to determine the process of direct contact and aerosol infection of specific pathogen free (SPF) chickens after inoculation. We have not only established the time of aerosol occurrence but also determined the aerosol concentrations, dynamic changes and their relationship with virus shedding. Infection doses of AIV through different routes for SPF chickens were quantified and compared. Lesions of SPF chickens infected with AIV aerosol and effection on immune function were monitored dynamically. Furthermore, air samples from chicken farms were collected and airborne AIV were detected by fluorescent RT-PCR.Not only were these results important in better understanding the transmission mechanism of AI, but also had implications in controlling ND, FMD and other contagious diseases.This study consists of four parts:1 The occurrence and transmission characteristics of airborne H9N2 avian influenza virusTo better understand the transmission route of H9N2 avian influenza virus (AIV), an infection model of H9N2 subtype AIV was established and two duplicate trials (T1 and T2) were conducted to observe the process of aerosol infection and direct contact in SPF chickens. Fifteen chickens (inoculation group, G1) were inoculated with H9N2 AIV and housed together with another 15 chickens (direct contact group, G2) in the same positive-negative-pressure isolator (A). Fifteen chickens (aerosol challenged group, G3) were bred in another isolator (B) which was connected with A. The SPF chickens of G1 were inoculated with AIV ocularly and nasally, and the SPF chickens of G2 and G3 were not inoculated with virus, but infected through direct contact and aerosols, respectively. Air samples in isolator A were collected with AGI-30 for the detection of aerosol forming. The seroconversion was assessed by the hemagglutination inhibition (HI) test and viral shedding was detected by RT-PCR and HI-Test. AIV aerosols were initially detected in chickens of T1 and T2 at day 3 and 2 post inoculation (dpi), respectively, reaching their peak concentrations of 7,200 PFU/m3 air and 4,800 PFU/m3 air at 7 dpi, respectively. AIV shedding was detected in chickens of T1G1 and T2G1 at 3 and 2 dpi, respectively. Virus shedding was detected in all chickens of G2, but only in 80-87% chickens of G3. Antibodies were initially detected at 5 and 4 dpi in chickens of T1G1 and T2G1, respectively, reaching their peak levels of 7.07 and 7.20 at 21 and 14 dpi, respectively. The results showed that after being infected with AIV, SPF chickens could excrete viruses via trachea and cloaca, thus leading to contamination of feedstuff and drinking water, as well as viral transmission; infected chickens may excrete viruses to form aerosols which may be transmitted by air flow. Therefore, it is obvious that in addition to direct contact infection due to contaminants, chickens in the same or adjacent henhouses could also be infected mutually with AI, despite no direct contact.2 Quantification and comparison of infection doses of H9N2 subtype avian influenza virus through different routes for SPF chickensIn this study, infection doses of H9N2 subtype avian influenza virus through different routes for SPF chickens were quantified and compared. SPF chickens were randomly divided into 3 groups:aerosol infection group, intranasal infection group and gastrointestinal tract infection group. Each chicken was inoculated with the same volume of different dilution of AIV. Infection was determined by detecting of the specific antibody to AIV. Infection dose 50 percent (ID50) were determined by the method of Reed-Muench. The results showed that ID50 of aerosol infection was 212 TCID50, of intranasal route was 398 TCID50, and of gastrointestinal tract infection was 23988 TCID50. It indicated that the efficiency was different when SPF chickens infected with AIV through different routes, aerosol infection was stronger than that of intranasal route, and the gastrointestinal tract infection was the weakest.3 Impact on immunologic function of SPF chicken experimentally infected with H9N2 AIV aerosolThe pathological changes of SPF chickens infected with H9N2 AIV aerosol were observed dynamically, and immune organ index and weight were measured. The results showed that the thymus index was significantly lower than the control group (p<0.05) since 5 dpi, but the bursa and spleen index showed no significant (p>0.05). At 2 dpi, histopathological changes of heart, liver, lung, kidney, pancreas, thymus, bursa and spleen showed hyperemia, hemorrhage and tumefaction. Atrophy of the thymus, bursa and spleen was detected at 5 dpi, and renal tubular filled with white urate deposition, then gradually recover. Pathology showed heart, lung and pancreas appeared inflammatory cell infiltration, and lung, pancreas, thymus, bursa, spleen, liver showed hemorrhage, lymphocytes decreased, necrosis and/or collapse. Antibodies reactions to ND and IBD vaccines were significantly lower compared with the control group (p<0.01). This showed that the H9N2 subtype AIV infection could cause multiple organs damages, in particular the immune organs, and thus led to immunosuppression to some extent.4 Detection of airborne H9 subtype avian influenza virus in chicken housesAir samples from 6 chicken farms of Shandong Province were collected by AGI-30 air sampler, and oropharyngeal and cloacal swabs were sampled simultaneously. Fifteen air samples were collected in each chicken house. Fluorescent RT-PCR was used for the detection of AIV in air and swab samples. Airborne H9 AIV was detected in four (A, B, C and E) out of 6 farms, and no AIV was detected in swabs. The results showed that airborne H9 AIV was detectable in farms after an outbreak. The AIV aerosol would do great harm to human and animal health and should be paid more attention to.
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