Sero-survey of Avian Influenza in backyard poultry and wild bird species in Iran—2014

Accepted Manuscript Title: Sero-survey of Avian Influenza in backyard poultry and wild bird species in Iran-2014 Author: M.H. Fallah Mehrabadi A.R. Bahonar M. Vasfi Marandi A. Sadrzadeh F. Tehrani M.D. Salman PII: DOI: Reference:

S0167-5877(16)30048-4 http://dx.doi.org/doi:10.1016/j.prevetmed.2016.01.031 PREVET 3979

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Received date: Revised date: Accepted date:

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Please cite this article as: Fallah Mehrabadi, M.H., Bahonar, A.R., Vasfi Marandi, M., Sadrzadeh, A., Tehrani, F., Salman, M.D., Sero-survey of Avian Influenza in backyard poultry and wild bird species in Iran-2014.Preventive Veterinary Medicine http://dx.doi.org/10.1016/j.prevetmed.2016.01.031 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Sero-survey of Avian Influenza in backyard poultry and wild bird species in Iran-2014

Fallah Mehrabadi M.Ha, Bahonar A.R*b, Vasfi Marandi Mc, Sadrzadeh Ad,Tehrani Fe , Salman M.Df

a

Department of Poultry Viral Diseases, Razi Vaccine and Serum Research Institute,

Karaj-Iran b

Department of Food Hygiene & Quality Control, Faculty of Veterinary Medicine,

University of Tehran, Tehran, Iran c

Department of Poultry Diseases, Faculty of Veterinary Medicine, University of Tehran,

Tehran, Iran. d

Department of Poultry Diseases, School of Veterinary Medicine, Azad University,

Garmsar, Iran. e

Department of Health and Management of Poultry Diseases, Iranian Veterinary

Organization. Tehran, Iran. f

Animal Population Health Institute of College of Veterinary Medicine and Biomedical

Sciences, Colorado State University, Fort Collins, CO 80523-1644, USA.

*Corresponding author at: Department of Food Hygiene & Quality Control, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran. Tel: +982161117056. E-mail address: [email protected]

   

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Abstract: In almost all villages in Iran backyard birds, especially chickens, are kept for egg and meat production. AI H9N2 subtype is endemic in Iran. Therefore, estimation of AI prevalence among these birds is important to determine the risk of transmission of infection to commercial farms. The aim of this study was to estimate subclinical infections or previous exposure to H5, H7, and H9 subtypes and to identify potentially important determinants of prevalence of this infectious at premises level in backyard poultry, bird gardens, zoos, and wild bird markets in Iran. A survey was conducted using a cross-sectional design throughout the entire country. A total of 329 villages, seven bird gardens, three zoos and five wild bird markets were included. In each village four families that kept birds were included in the collection of biological samples and background information. The Enzyme-Linked Immunosorbent Assay (ELISA) was used as the screening test and all ELISA-positive samples were examined with the HI test to differentiate H5, H7, and H9. Among the bird gardens, eight of 15 premises (53.3%) were positive in both the ELISA test and HI for H9N2. Testing of samples collected in the villages revealed that 296 out of 329 villages (90%) had positive ELISA tests and also HI tests for H9. The HI-H9 mean titers in positive units were significantly higher than negative units (P<0.001). This study revealed no significant statistical differences between risk variables in seropositive and seronegative bird gardens in the case of H9 (P > 0.05). The results of this study showed that among the risk variables, mountainous area was a protective factor and lack of hygienic disposal of dead birds was a risk factor for AI; this was also observed in rural poultry.

   

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The high sero-prevalence of influenza H9N2 in rural domestic poultry indicates that the disease is endemic. It is necessary to include backyard poultry in any surveillance system and control strategy due to the existence of AIV in backyard poultry and the possibility of transmission of infection to commercial poultry farms. Implementation of an AI surveillance program and biosecurity measures can be useful to control this infection and prevent AI from spreading to commercial farms. Furthermore in Iran there is no program for destruction of birds infected with the H9N2, so an effective vaccination program with regard to issues such as acceptability and cost-benefit must play an important role in reducing infections in backyard poultry.

Keywords: Avian Influenza, Sero-survey, Iran, Backyard birds                        

   

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Introduction In almost all villages in Iran backyard birds (especially chickens) are kept for egg and meat production. Avian Influenza (AI) H9N2 subtype is endemic in Iran, so detection of AI prevalence (H9N2, H5 and H7 subtypes) among these birds is important due to risk of transmission of infection to commercial farms (Hadipour, M.M., 2010). The low pathogenic subtypes of Avian Influenza (LPAI), consisting of different combinations of haemagglutinin(H) and neuraminidase (N) glycoproteins, cause little harm in the wild (Webster et al., 1992; Fouchier et al.,2005). Certain subtypes, however, particularly H5 andH7 may become highly pathogenic if they are transmitted to domesticated birds (Shortridge et al., 1998; Spackmanet al., 2003; Aubin et al., 2005) and include the potential to be zoonotic agents. Detection of specific antibodies, viral isolation and molecular assays are used to discover whether there has been exposure to Avian Influenzavirus (AIV)[Adams, 2010]. Among serological assays, the haemagglutination inhibition (HI) test is commonly used in surveillance activities and detection of infection. Annual migration patterns of wild birds have been considered as avenues for the introduction of new viruses and exotic subtypes of AI into a country and its domestic poultry populations. The interface between wild migratory birds and domestic poultry populations provides conditions that encourage genetic mutation of this virus. Migratory birds can also transfer exotic viruses to nearby commercial units, causing new outbreaks in the commercial poultry farms if biosecurity is inadequate (OIE, 2012). Thus, there is a need to incorporate monitoring of the circulation of viruses among migratory birds and their interaction with free-ranging poultry populations such as backyard flocks. Such

   

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monitoring is an essential component of AI surveillance for any country wishing to enhance biosecurity and reduce the impact of AIV. The aim of this study was to estimate subclinical infections or previous exposure to H5, H7, and H9 subtypes and to identify potentially important determinants of prevalence of this infectious at premises level in backyard poultry, bird gardens, zoos, and wild bird markets in Iran.

Material and Methods Survey design and sampling procedures This survey was cross-sectional in design and was conducted from September to December 2014 throughout the country of Iran. All premises where birds are kept or sold are registered in the veterinary disease database, and every premise has a unique 11-digit code. At the study the premises which were registered in this database were used. Premises were selected from bird gardens, zoos, and markets where domestic and wild birds were kept or sold, as well as in villages where domestic poultry were kept or bred. There were 20 premises where wild birds were kept or sold for showing, and these premises are registered in the veterinary disease database, and at the time of study 15 of those had bird were sampled including seven bird gardens, three zoos and five wild bird markets. Domestic poultry from villages (backyard poultry) were sampled using random sampling design. The sampling and analysis unit was the village. In Iran there are 32 provinces with approximately 65,000 villages where families keep birds for different purposes. Villages across the country were listed by province, and then 0.5 percent of villages with poultry were selected for sampling. The number of villages to be sampled

   

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was determined assuming 30% prevalence at the village level, with 5% precision and 95% confidence level. A total of 329 villages were selected, and in each village four families that kept birds were included in the study. According to our last survey (Fallah Mehrabadi et al., 2015) the prevalence of H9N2 within the villages was 50% so the number of families to sample in each village was based on the assumption of 50% prevalence and 95% confidence level to identifying at least one positive family (Salman, 2003). Eleven birds, according to the proportion of present birds types, were sampled from each family so as to ensure 95% probability of identifying at least one positive bird if the expected prevalence of sero-positive birds is ≥ 30% (EC 2010, 2011/367/EU). In cases where the number of birds in the family flock was fewer than 11, all the birds were sampled. To include species diversity, sampling was performed for all species present in the village. All villages where birds are kept are registered in the veterinary disease database, and every village has a unique 11-digit code. For the selection of villages for this survey a list of villages was transferred to Excel® and for selection of families in each village the list of families that kept birds were made by Excel®. Using the random selection function of Excel®, the final selection list was generated. Premises that were positive for H5 and H7 had follow-up sampling for virus detection; a total of 60 cloacal swabs were taken from each premise that were positive and within a radius of 3 km. All birds in the randomly selected villages had not been vaccinated against AI, as stated by provincial veterinary officials.

   

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Questionnaire The questionnaire for independent variables was designed according to advice from veterinary experts regarding likely risk factors of influenza in Iran (including weather, existence of a river, lagoon, or lake, a poultry slaughterhouse, live bird market, duck farm or other commercial poultry farm within up to a 3 km radius; locating near a general transportation route; lack of hygienic disposal of dead birds and using them as food for carnivores; and the village’s water source). The questionnaire was completed through a direct interview of the bird owners. Prior to its administration for this study the questionnaire was tested on 397 interviews the previous year (Fallah Mehrabadi et al., 2015). The questionnaire is available from the senior author upon request. Laboratory tests Up to 11 birds of various species of domestic birds (poultry, duck, goose, turkey, Pigeon, and others) were sampled from each premise. One ml of blood was taken from the wing vein and preserved at room temperature for one hour to separate the serum. Then each serum sample was separated and placed in a 1.5 ml micro tube. The samples were coded, the premise and bird species were recorded, and the tubes were transferred to the laboratory with cold-chain, and stored in a freezer (-20 C) for HI and ELISA testing. The Enzyme Linked Immunosorbent Assay (ELISA) A-Type was performed using a commercial kit (BioChek AI Ab test kit, solid plate format CK121 480 Tests/ 5 plates OIE Certified). This kit includes a complete set of chemical and immunological reagents that allows the detection of antibodies to the virus causing AI. The test is able to determine from blood or serum of chickens and turkeys the presence of antibodies that may have been produced through a field infection or from vaccination. The test kit

   

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detects antibodies to all known H- serotypes (H1–H15) of Influenza Type A. All ELISApositive samples were examined with the HI test to differentiate H5, H7, and H9. For H5 and H7 two serial HI tests were done. Sera that were positive in the first test (H5N2 and H7N1) were examined with another antigen (H5N1 and H7N7) for confirmation in the central laboratory of the Iranian Veterinary Organization (IVO). Two-fold serial dilutions of sera were made and sera with titers ≥4 (based on log2) from the negative control were considered positive; those with a titer ≥4 and premises that had at least one positive bird were considered positive. The H9N2 antigen was used to test for the presence of H9. For detection of Influenza Virus of H5 and H7 heamagglutinin subtype from the cloacal swabs, Real Time RT-PCR was used according to the protocol of OIE/FAO Reference Laboratory in Italy (IZS Ve. Legnaro-Padova) [Monne et al., 2008]. Data collection and data analysis Questionnaire data was collected for each premise; these data and the results of the laboratory tests were recorded in the IVO disease monitoring and surveillance system (GIS). The reliability of data about the questions which could have different answers in repeated interview, based on the interviewees' opinion, was examined using Cronbach's Alpha statistical procedure for reliability (Tavakol, 2011). For statistical description of the data, central tendency mean was used to describe quantitative variables and frequency distribution for qualitative variables. Qualitative variables were then examined for their association using chi-square and logistic regression tests and presented with Odds Ratios (OR), 95% CI for OR and P-values. And association of quantitative variables that haven't normal distribution were examined using Wilcoxon-Mann-Whitney test (Salman, 2003).

   

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A P<0.05 was considered to be statistically significant. All statistical procedures were conducted using SPSS version 16, and ArcGIS version 9.2 was used for mapping. Data description The Cronbach's Alpha of the questionnaire data was 0.724. Most of the bird species kept in bird gardens and zoos are chickens, various waterfowl (ducks, geese, and swans), and pigeons. In the villages birds are kept for various purposes: chickens (the most common species) for egg production, turkey and duck for meat production, and pet pigeons. For this study 468 birds from 15 bird gardens and zoos and 8901 birds from 329 villages were sampled. The number of birds sampled and the frequency of HI (H9) positive birds are shown in Table 1. Results Among the bird gardens and zoos, eight of 15 premises (53.3%) were positive in both the ELISA test and H9. In addition, three waterfowl samples were positive for H5, and all samples were negative for H7. All PCR tests on the cloacal swabs were negative for H5 and H7. Testing of samples collected in the villages revealed that 296 out of 329 villages (90%) had positive ELISA tests and also HI tests for H9. Three samples were positive for H5, two chickens and one turkey. All the samples were negative for H7. All the collected swab samples were PCR negative for detection of H5 and H7. More than 95% of villages in 16 provinces were seropositive; Lorestan and Hormozgan had no seropositive villages (Figure 1). The H9 mean titers in positive units were significantly higher than those with negative serum (p<0.001).

   

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Risk factor analysis This study revealed no significant statistical differences in potential risk factors between seropositive and sero-negative bird gardens in the case of H9 (P>0.05). Table 2 shows the distribution of the investigated variables among AI-positive and AInegative villages in Iran. Table 2 also shows the univariable analysis by chi-square that revealed four variables (including weather type, lack of hygienic disposal of dead birds, live bird marketing nearby, and waterfowl breeding unit nearby) with P<0.05 which were then offered to the multivariable logistic model shown in Table 3 and results revealed that the cold and wet weather was a protective factor and a lack of hygienic disposal of dead birds was a risk factor. Discussion H9N2 is endemic in several countries, including Iran (Norouzian, 2012). For example, the range of AI prevalence rates in Pakistan commercial poultry is reported to be 10-20 percent (Naeem et al. 2003). In previous years, AI outbreaks in commercial and backyard poultry in some provinces caused by influenza subtypes have raised the level of concern regarding the possibility of circulation of avian influenza virus of different subtypes in rural and wild birds. In this study 90% of sampled villages were positive for H9. This high prevalence was the same as that previously reported in domestic poultry in Iran (Fallah Mehrabadi et al., 2015), which is slightly higher than other studies that have been conducted in the country (Hadipour, 2010, Hadipour, 2011). In a study reported by Van Kammen (1982), AI antibody titers were detected in backyard birds.

   

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Ghaniei et al. (2013) showed a sero-prevalence of 40.6% for H9N2 in broiler chickens in northwest Iran, and the authors concluded that the high prevalence of AIV (H9N2) antibodies in the serum of birds emphasized that H9N2 plays an important role in respiratory complexes in broiler chickens in that region and probably throughout Iran. Results of a study on slaughtered poultry in the city of Kashan by (Nikkhah, 2013) showed that 17 out of 20 unvaccinated flocks were positive for influenza H9N2 subtype. In the city of Shahrekord Fathi et al. (2010) found that 53.3% of flocks were positive for H9N2. The results of this study showed that among the variables studied, cold and wet weather (mountainous area) was a protective factor. The prevalence in villages located in mountainous regions was significantly lower than for other villages. This result is contrary to a study done by Fallah et al. (2015) that indicated that the prevalence in moderate weather was the lowest. A European study by Wang et al. (2010) using a logistic regression model to analyze risk factors related to AI showed that a normalized difference vegetation index (NDVI) in December, an average of this index in March, low latitude, increasing minimum temperature in January, and a decrease in rainfall in January were risk factors related to diseases in migratory birds. Risk maps also show that this disease is connected to food (source) availability, rising temperatures, and reduction in rainfall (Wang et al. 2010). Our results also showed that a lack of hygienic disposal of dead birds was a risk factor for AI, and this was also observed in rural poultry. Poor biosecurity practices have been shown in other studies as well. Vasfi-Mirandi et al. (2002) showed that manure was delivered by truck from Tehran and Qazvin provinces to the southern part of the country, and then feed was delivered from the southern ports to other provinces using the same

   

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trucks without proper disinfection. This was a significant risk factor for the spread of H9N2 infection through the whole country (Vasfi-Marandi 2002). Conclusion The high sero-prevalence of influenza H9N2 in rural domestic poultry indicates that the disease is probably endemic for some years. It is necessary to include backyard poultry in any surveillance system and control strategy for this disease due to the existence of the infection in backyard poultry and the possibility of transmission of infection to commercial poultry farms. Implementation of an AI surveillance program and biosecurity measures can be useful to control this infection (Saadat 2014). Furthermore in Iran there is no program for destruction of birds infected with the H9N2 AI subtype, so an effective vaccination program with regard to issues such as acceptability and cost-benefit must play an important role in reducing infections in backyard poultry. Acknowledgements The authors wish to thank the department of health and management of poultry diseases of Iranian Veterinary Organization and the provinces’ veterinary branches for their support and cooperation.

   

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Spackman, E., Stöhr, K., Suarez, D.L., Sung, H.W., Swayne, D.E., Tardy-Panit, M., Tashiro, M., Thawatsupha, P., Tumpey, T., Uyeki, T., Tu, P.V., Van der Werf, S., Vong, S., Webby, R., Webster, R., Wood, J., Xu, X., Yi, G., and Zhang, w., 2005. Evolution of H5N1 avian influenza viruses in Asia. Emerg. Infect. Dis. 11, 1515–1521. EC, 2010. Commission Decision 2011/367/EU of 25 June 2010 on the implementation by Member States of surveillance programs for avian influenza in poultry and wild birds, Official J. of the European Union. L 166, 1.7.2010, p.7.

   

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Fallah Mehrabadi, M.H., Bahonar A.R., Tehrani, F., Vasfimarandi, M., Sadrzadeh, A., Ghafouri, S.A., Meshkat, M. and Masror, F., 2015. Avian Influenza H9N2 Seroepidemiological Survey in Rural Domestic Poultry of Iran. Iranian J. Epi. 10(4): 1-8. FathiHafashgani, E., Dosti, I., Gholami, M., ZamaniMoghadam, A., 2010. Survey of the synchronous prevalence of Newcastle and influenza H9N2 in broiler in Shahr e Kord. Vet. Modern Res. Mag. 2:35-40. Fouchier, R.A.M., Munster, V., Wallensten, A., Bestebroer, T.M., Herfst, S., Smith, D., Rimmelzwaan, G.F., Olsen, B., Osterhaus, A.D.M.E., 2005. Characterization of a novel influenza a virus hemagglutinin subtype (H16) obtained from black-headed gulls. J. Virol. 79, 2814–2822. (doi:10.1128/JVI.79.5.2814-2822.2005). Ghaniei, A., Allymehr, M., Moradschend, A., 2013. Seroprevalence of avian influenza (H9N2) in broiler chickens in North West of Iran. Asian Pac. J. of Trop. Biomed. 3(10), 822–824. Hadipour, M.M., 2010.Seroprevalence survey of H9N2 avian influenzavirus in backyard chickens around the Caspian Sea in Iran. Rev. Bras. Cienc. Avic. 12 (1), 1-4. Hadipour, M.M., 2011. Serological Evidence of Inter-Species Transmission of H9N2 AvianInfluenza Virus in Poultry. Iran.Int. J. of Animal and Vet. Advances.3(1), 29-32. Monne, I., Ormelli, S., Salviato, A., De Battisti, C., Bettini, F., Salomoni, A., Drago, A., Zecchin, B., Capua, I., and Cattoli, G., 2008. Development and Validation of a OneStep Real-Time PCR Assay for Simultaneous Detection of Subtype H5, H7, and H9 Avian Influenza Viruses. J. Clin. Microbiol. 46(5): 1769–1773.

   

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Naeem, K., Naurin, M., Rashid, S., and Bano, S., 2003. Seroprevalence of Avian Influenza Virus and its relationship with increased mortality and decreased egg production. Avian Pathol. 32: 285- 289.109-119. Nicoll, A., Danielsson, N., 2013. A novel reassortant avian influenza A (H7N9) virus in China-what are the implications for Europe. Euro Surveil. 18(15): 1-5. NikkhahGhamsary, M., 2009. Seroprevalence of Avian Pneumovirus in broiler chickens in Kashan City, D.V.M. Thesis, Islamic Azad University, Shahrekord Branch. Norouzian H., Gholami S.J., VasfiMarandi M., 2012. Phylogenetic analysis of hemagglutinin gene of H9N2 avian influenza viruses isolated from chickens in Iran in 2010-2011: Emerging of a new subgroup. Iran J Virol. 6(1): 18-26. OIE, 2012. Terrestrial Animal Health Code. 7th Edition. Chapter 2.3.4.OIE, Paris. Saadat, Y., Ghafouri, S.A., Tehrani, F., GhalyanchiLangeroudi, A., 2014. An active serological survey of antibodies to Newcastle disease and avian influenza (H9N2) viruses in the unvaccinated backyard poultry in Bushehr Province,Iran, 2012–2013. Asian Pac J Trop Biomed. 4(Suppl 1): S213–S216.doi: 10.12980/APJTB.4.2014C1293, 1-5. Saif, Y.M., 2013. Diseases of Poultry. 13th Edition, Blackwell Publishing Ltd. pp. 181213. Salman, M.D., 2003. Animal Disease Surveillance and Survey Systems, Methods and Applications. First edition, Blackwell Publishing. Chapter 4. Salman, M.D., 2003. Animal Disease Surveillance and Survey Systems, Methods and Applications. First Edition, Blackwell Publishing. Chapter 5. Shortridge, K.F., Zhou, N.N., Guan, Y., Gao, P., Ito, T., Kawaoka, Y.,Kodihalli, S., Krauss, S., Markwell, D., Murti, K.G., Norwood, M., Senne, D., Sims, L., Takada, A.,

   

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Webster, R.G., 1998. Characterization of avian H5N1 influenza viruses from poultry in Hong Kong. Virology 252, 331–342. (doi:10.1006/viro.1998.9488). Spackman, E., Senne, D.A., Davison, S., Suarez, D.L., 2003. Sequence analysis of recent H7 avian influenza viruses associated with three different outbreaks in commercial poultry in the United States. J. Virol. 77, 13 399-13 402. (doi:10.1128/JVI.77.24.1339913402.2003). Tavakol, M., Dennnick, R., 2011. Making sense of Cronbach’s Alpha. Int. I. Med. Ed. 2:53-55. Van Kammen, A., 1982. Survey of some poultry viruses in Papua New Guinea. Trop. Anim. Health Prod. p. 14. Vasfi-Marandi,

M.,

Bozorgmehrifard,

M.H.,

Tabatabaei,

S.M.,

2002.

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seroepidemiologic study of avian influenza (H9N2) in Iran. Scientific-research Iranian Vet. J. 5(8), 23-31. Wang, Si. Y., Skidmore, T. A. K., De Boer, W. F., Li, L., Prins, H. H. T. 2010. Environmental factors influencing the spread of the highly pathogenic avian influenza H5N1 virus in wild birds in Europe. Ecology and Society 15(3): 26. [online] URL: http://www.ecologyandsociety.org/vol15/iss3/art26/ Webster, R.G., Bean, W.J., Gorman, O.T., Chambers, T.M.,Kawaoka, Y., 1992. Evolution and ecology of influenza-A viruses. Microbiol. Rev. 56, 152–179.

   

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Figure 1. Seroprevalence of AI in the different provinces of Iran from September to December 2014. The map was produced using ArcGIS® version 9.2 (ESRI, USA).  

   

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Table 1.Number of birds sampled and number and frequency of HI (H9) seropositive birds from villages and bird gardens and zoos, based on bird species. Data was collected in Iran in September through December 2014. Location Bird gardens and zoos (n=15)

Villages (n=329)

Bird species Number of HI (H9) positive birds

Number of sampled birds

Number of HI (H9) positive birds

Number sampled birds

Chicken

159

37 (23%)

7911

2704 (34%)

Waterfowl

148

10 (7%)

541

20 (4%)

Turkey

13

0 (0%)

412

45 (11%)

Pigeon

13

2 (15%)

6

2 (33%)

Other species*

153

28 (18%)

31

0 (0%)

Total

468

77 (16%)

8901

2771 (31%)

of

*Including (quail, partridge, ostrich)

   

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Table 2.Univariable analysis of investigated independent variables for H9N2 AI in Iran at villages level, based on questionnaire data collected from September to December 2014. Variable

Category

Weather type

Moderate and wet Mountain(cold and wet) Hot and dry

lack of hygienic disposal of dead birds live marketing nearby waterfowl breeding nearby

bird

unit

river nearby pond nearby lake nearby other commercial unit nearby poultry slaughter-house nearby

water supply

public transportation nearby

AIseropositivity

 OR(95% CI)

P*

# neg

17

9

136

15

.121

0.29,.449

.003

75

14

.579

.160,2.088

.403

Hot and wet

11

2

.342

.093,1.254

.105

Moderate No Yes

47 85

3 5 11

.351

0.52,2.361

.282

3.101

1.151,8.353

.025

27

.879

39 4

.173

.408,1.895 .045,.674

.742 .011

38 5

.304

.100,.923

.036

29 14

.876

.441,1.740

.705

37 6

.591

.227,1.535

.280

38 5

.480

.168,1.377

.172

27 16

.667

.341,1.303

.236

41 2 16

.820

.175,3.832

.801

2

1.474

.711,3.056

.297

Unknown

53

No

148 5

Yes

281

No

275

Yes

11

No Yes No Yes No Yes No

201 85 261 25 269 17 205

Yes

81

No

275

Yes well Spring

11 114 21

River

60

6

2.172

.467,10.099

.322

Subterranean canal

4

1

2.069

.776,5.517

.146

Others

87

18

.828

No

91

13

.929

.087,7.846 .463,1.864

.869 .835

Yes

195

30

   

OR

# pos

19

Table 3.Logistic regression analysis of variables associated with AI seropositive, at villages level in Iran, 2014. Variable Constant Weather type

lack of hygienic disposal of dead birds

Category

OR 95% 12.182 1 .124 .660 .355 .267 1 3.336 .824

Moderate and wet Mountain(cold and wet) Hot and dry Hot and wet Moderate No Yes Unknown

   

95% CI OR .029,.525 .180,2.420 .096,1.313 .038,1.878 1.209,9.204 .367,1.853

P < 0.001 .005 .531 .121 .185 .020 .640

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