An avian influenza (AI) surveillance was undertaken in Maharashtra state, India during the period 2010-2011. There are no reports of AI surveillance in emus from India. A total of 202 blood samples and 467 tracheal and cloacal swabs were collected from eight emu farms. A hemagglutination inhibition (HI) assay was performed for detection of antibodies against AI H5N1, H7N1, H9N2, and avian paramyxovirus type 1 (APMV-1) viruses. A microneutralization (MN) assay was performed to confirm the presence of neutralizing antibodies against AI H9N2 and to compare with HI assays. A total of 28.2% and 28.7% of samples were positive for antibodies against AI H9N2 by HI and MN assays, respectively, using > or = 1:40 as a cut-off titer; 15.3% samples were positive for APMV-1 by HI assay using a > or = 1:10 cut-off titer. Seropositivity of AI H9N2 was nil in the grower (<1 yr) age group and highest (78%) in the breeder (2-3 yr) age group, whereas seropositivity against APMV-1 was observed in all age groups. Performance of both HI and MN assays was similar, suggesting the utility of using the MN assay along with HI assay for surveillance studies. This is the first report of the seroprevalence of AI H9N2 and APMV-1 in emus in India.