Poultry (Gallus domesticus) farming plays an important role as an income generating enterprise in a developing country like Nepal, contributing more than 4% to the national GDP. It is also one of the major sources of protein for growing population. Newcastle Disease (ND) is a major poultry disease affecting both commercial and backyard poultry production worldwide. There were more than 90 reported cases of ND outbreaks in Nepal in 2018, with over 74,986 birds being affected. ND might be responsible for over 7% of total poultry mortality in the country. Recent outbreak of ND in 2021 affected many farms throughout Nepal, and caused massive poultry production loss. ND is caused by a single stranded RNA virus which presents very similar clinical symptoms as Influenza A (commonly known as Bird flu), adding much complexity to clinical disease identification and intervention.We conducted a nationwide ND and Influenza A prevalence study, collecting samples from commercial and backyard poultry farms from across the major poultry production hubs of Nepal, and conducted both serological and molecular assessments-giving us disease exposure history and identification of floating strains of ND Virus (NDV). Of 600 commercial chickens tested from various farms, both NDV (n=381, 64%) and IA (n=125, 21%) antibodies were detected in the majority of the samples. In backyard chicken (n=108, 39 farms), sero-prevalence was also relatively high for both NDV (n=38, 35%) and IA (n=17, 16%). Out of the 40 commercial farms, majority had detectable NDV (n=31, 78%) and IA (n=15, 38%) virus present. In backyard farms (n=36), we also detected NDV (n=6, 16%) and IA (n=1, 3%) virus. We Genotyped (strain) detected NDV, and found Genotype II to be present in most of the commercial farms (which might be coming from live vaccine usage) and Genotype I in some backyard poultry samples. The identified Genotype I strain is reported for the first time, and hence could be an endemic NDV strain found in Nepal. Our 2021 ND outbreak investigation identified Genotype VII c as the causative strain.Additionally, we have developed a thermostable I-2 NDV vaccine (Ranigoldunga™) in tablet formulation and tested on various (mixed) breeds of chicken (G. domesticus). This vaccine seems to be highly effective against NDV, including a virulent 2021 outbreak strain (Genotype VII c). The I-2 Tablet ND vaccine showed more than 85% efficacy when administered either ocularly or in water, and has a stability of 30 days in room temperature.

Poultry (Gallus domesticus) farming plays an important role as an income generating enterprise in a developing country like Nepal, contributing more than 4% to the national Gross Domestic Product (GDP). Newcastle Disease (ND) is a major poultry disease affecting both commercial and backyard poultry production worldwide. There were more than 90 reported ND outbreaks in Nepal in 2018 with over 74,986 birds being affected. ND is responsible for over 7% of total poultry mortality in the country. Recent outbreaks of ND in 2021 affected many farms throughout Nepal and caused massive loss in poultry production. ND is caused by a single-stranded ribonucleic acid (RNA) virus that presents very similar clinical symptoms as Influenza A (commonly known as bird flu) adding much complexity to clinical disease identification and intervention. We conducted a nationwide ND and Influenza A (IA) prevalence study, collecting samples from representative commercial and backyard poultry farms from across the major poultry production hubs of Nepal. We used both serological and molecular assessments to determine disease exposure history and identification of strains of ND Virus (NDV). Of the 40 commercial farms tested, both NDV (n = 28, 70%) and IAV (n = 11, 27.5%) antibodies were detected in majority of the samples. In the backyard farms (n = 36), sero-prevalence of NDV and IAV were 17.5% (n = 7) and 7.5% (n = 3) respectively. Genotype II NDV was present in most of the commercial farms, which was likely due to live vaccine usage. We detected never reported Genotype I NDV in two backyard farm samples. Our investigation into 2021 ND outbreak implicated Genotype VII.2 NDV strain as the causative pathogen. Additionally, we developed a Tablet formulation of the thermostable I2-NDV vaccine (Ranigoldunga™) and assessed its efficacy on various (mixed) breeds of chicken (Gallus domesticus). Ranigoldunga™ demonstrated an overall efficacy >85% with a stability of 30 days at room temperature (25°C). The intraocularly administered vaccine was highly effective in preventing ND, including Genotype VII.2 NDV strain.

Background: Commercial hatcheries play an important role in the overall poultry value chain- providing small to large poultry farmers with day old chicks. Any outbreak in such hatcheries can spread diseases to other farms. Regular screening of major avian pathogens, along with strict bio-security measures, can prevent spread of diseases in hatcheries. Newcastle Disease Virus (NDV), Infectious Bronchitis Virus (IBV), Mycoplasma gallisepticum (MG), Mycoplasma synoviae (MS), Infectious Bursal Disease Virus (IBDV) and Influenza A Virus (IAV) are among the most prevalent poultry diseases which can be detected in egg albumin. Method: We retrospectively (August 2020- August 2021, except October 2020) analyzed diagnostic results for six selected avian pathogens (NDV, IBV, MS, MG, IBDV and IAV) on eggs (n=4343) received from eleven major commercial poultry hatcheries located in the five adjoining districts of Kathmandu, Nepal. Albumin from 10% randomly selected eggs from each hatchery were tested for the six avian pathogens using multiplex PCR. Result: Majority (7/11, 64%) of the poultry hatcheries had at least one of the six pathogens present. We detected at least one avian pathogen in nine out of eleven months (82%) of screening. Except for IBDV, we found one or more of the other major avian pathogens- Influenza A (IAV) (n=4 times) and Mycoplasma gallisepticum (MG) (n=4 times) were detected the most, followed by Newcastle Virus (NDV) (n=3 times). Infectious bronchitis virus (IBV) were detected twice, and Mycoplasma synoviae (MS) was detected once. Conclusion: In a resource strapped country like Nepal, poultry disease outbreak investigation in particular and surveillance in general are challenging. Meanwhile, poultry production is highly impacted by disease outbreaks often triggered by poor bio-security and lack of pathogen screening practices. Our molecular screening tests have picked up major poultry pathogens present throughout the year in eggs collected from hatcheries. Influenza A was detected at 4 different incidences throughout the year, which is of concern to both human and animal health. Quick systematic screening of eggs at key distribution points (hatcheries) for major avian pathogens is an effective surveillance tool for early disease detection and containment of outbreaks. Keywords: eggs, avian pathogens, disease screening, surveillance, poultry health

Poultry industry in Nepal has experienced remarkable growth in the last decade, but farm biosafety and biosecurity measures are often overlooked by farmers due to lack of knowledge or to save cost. As a result, farms often suffer from sporadic and regular outbreaks of many zoonotic diseases such as highly pathogenic avian influenza (HPAI), impacting production and creating public health challenges. Poor farm management practices, including overuse of antibiotics for prophylaxis and therapeutics, can complicate the spread of poultry diseases by creating and enhancing antimicrobial resistance (AMR) that is threating to both, poultry, and human health. We assessed biosafety, biosecurity risks and AMR stewardship in sixteen poultry farms located in four districts (Ramechhap, Nuwakot, Sindhupalchowk, and Kavre) surrounding densely populated Kathmandu valley. Risk assessment and AMR stewardship evaluation questionnaire were administered to formulate biosafety and biosecurity compliance matrix (BBCM). Risk assessment checklist assessed facility operations, personnel and standard operating procedures, water supply, cleaning and maintenance, rodent/pest control and farm record keeping. Oral and cloacal samples from the poultry were collected, pooled, and screened for eight poultry pathogens using Polymerase Chain Reaction (PCR) tests. Based on BBCM, we identified one of the farms in Sindhupalchowk (Farm 4) having the most (BBCM score= 67%) and a farm in Kavre (Farm 3) to having the least (BBCM score= 12%) biosafety and biosecurity compliance. Although most of the farms (61.6%) followed general poultry farming practices, only half had clean and well-maintained farms. Personal safety standard procedure compliance (BBCM score = 42.4%) and rodent control (BBCM score = 3.1%) were the biggest gaps. At least one of either bacterial or viral pathogen was detected in all farms. Mycoplasma gallisepticum was the most common disease detected in all but one farm, followed by Mycoplasma synoviae. Although more than half of the farmers considered AMR a threat, over 26% of them used antibiotics as a preventive measure and 81% did not consider withdrawal period for antibiotics prior to processing of their meat products. Additionally, antibiotics classified as Watch and Restrict by the WHO were frequently used by the farmers to treat bacterial infections in their farms. Lack of awareness and inadequate enforcement of regulations have exacerbated the risk of disease transmission in farms and compromised antimicrobial stewardship.