Swine Backyard Production Systems in Central Chile: Characterizing Farm Structure, Animal Management, and Production Value Chain.

Backyard production systems (BPS) are highly distributed in central Chile. While poultry BPS have been extensively characterized, there remains a notable gap in the characterization of swine BPS in central Chile. In addition, there is evidence that zoonotic pathogens, such as influenza A virus and Salmonella spp., are circulating in backyard poultry and pigs. A total of 358 BPS located in central Chile were evaluated between 2013 and 2015 by interviewing farm owners. Severe deficiencies in biosecurity measures were observed. The value chain of swine backyard production identified food, veterinary care (visits and products), and replacement or breeding animals as the primary inputs to the backyard. The most common origin of swine replacements was from outside the BPS (63%). The main outputs of the system were identified as meat and live animals, including piglets and breeding animals. In 16% of BPS, breeding animals were lent to other BPS, indicating the existence of animals and animal product movement in and out of backyard farms. Results from this study indicate that swine BPS in central Chile represents an animal-human interface that demands special attention for implementing targeted preventive measures to prevent the introduction and spread of animal pathogens and the emergence of zoonotic pathogens.

Remote Sensing and Ecological Variables Related to Influenza A Prevalence and Subtype Diversity in Wild Birds in the Lluta Wetland of Northern Chile.

The Lluta River is the northernmost coastal wetland in Chile, representing a unique ecosystem and an important source of water in the extremely arid Atacama Desert. During peak season, the wetland is home to more than 150 species of wild birds and is the first stopover point for many migratory species that arrive in the country along the Pacific migratory route, thereby representing a priority site for avian influenza virus (AIV) surveillance in Chile. The aim of this study was to determine the prevalence of influenza A virus (IAV) in the Lluta River wetland, identify subtype diversity, and evaluate ecological and environmental factors that drive the prevalence at the study site. The wetland was studied and sampled from September 2015 to October 2020. In each visit, fresh fecal samples of wild birds were collected for IAV detection by real-time RT-PCR. Furthermore, a count of wild birds present at the site was performed and environmental variables, such as temperature, rainfall, vegetation coverage (Normalized Difference Vegetation Index-NDVI), and water body size were determined. A generalized linear mixed model (GLMM) was built to assess the association between AIV prevalence and explanatory variables. Influenza positive samples were sequenced, and the host species was determined by barcoding. Of the 4349 samples screened during the study period, overall prevalence in the wetland was 2.07% (95% CI: 1.68 to 2.55) and monthly prevalence of AIV ranged widely from 0% to 8.6%. Several hemagglutinin (HA) and neuraminidase (NA) subtypes were identified, and 10 viruses were isolated and sequenced, including low pathogenic H5, H7, and H9 strains. In addition, several reservoir species were recognized (both migratory and resident birds), including the newly identified host Chilean flamingo (Phoenicopterus chilensis). Regarding environmental variables, prevalence of AIV was positively associated with NDVI (OR = 3.65, p < 0.05) and with the abundance of migratory birds (OR = 3.57, p < 0.05). These results emphasize the importance of the Lluta wetland as a gateway to Chile for viruses that come from the Northern Hemisphere and contribute to the understanding of AIV ecological drivers.

Detection and phylogenetic analysis of highly pathogenic A/H5N1 avian influenza clade 2.3.4.4b virus in Chile, 2022.

Highly pathogenic avian influenza (HPAI) A/H5N1 viruses continue to pose a significant threat to animal and human health worldwide. In late 2022, the first confirmed case of HPAI A/H5N1 infection in wild birds in Chile near the Chilean-Peruvian border was reported. Active surveillance by our group in the adyacent Lluta river estuary revealed an increase in A/H5N1 prevalence coinciding with the arrival of migratory birds from the Northern Hemisphere. Genomic analysis of A/H5N1-positive samples demonstrated a close genetic relationship to strains detected in Peru during the same period, which originated from A/H5N1 viruses causing outbreaks in North America. Notably, we identified genetic mutations that did not correlate with known enhanced transmission or binding traits to mammalian receptors. In summary, this study provides valuable genomic insights into the A/H5N1 Clade 2.3.4.4b viruses in wild birds in Chile, emphasizing the need for enhanced surveillance and response strategies to mitigate the threat posed by these highly pathogenic avian influenza viruses in South America.

Novel Low Pathogenic Avian Influenza H6N1 in Backyard Chicken in Easter Island (Rapa Nui), Chilean Polynesia.

Little is known about the prevalence of avian influenza viruses (AIV) in wildlife and domestic animals in Polynesia. Here, we present the results of active AIV surveillance performed during two sampling seasons in 2019 on Easter Island (Rapa Nui). Tracheal and cloacal swabs as well as sera samples were obtained from domestic backyard poultry, while fresh faeces were collected from wild birds. In addition to detecting antibodies against AIV in 46% of the domestic chickens in backyard production systems tested, we isolated a novel low pathogenic H6N1 virus from a chicken. Phylogenetic analysis of all genetic segments revealed that the virus was closely related to AIV's circulating in South America. Our analysis showed different geographical origins of the genetic segments, with the PA, HA, NA, NP, and MP gene segments coming from central Chile and the PB2, PB1, and NS being closely related to viruses isolated in Argentina. While the route of introduction can only be speculated, our analysis shows the persistence and independent evolution of this strain in the island since its putative introduction between 2015 and 2016. The results of this research are the first evidence of AIV circulation in domestic birds on a Polynesian island and increase our understanding of AIV ecology in region, warranting further surveillance on Rapa Nui and beyond.

Evidence of influenza infection in dogs and cats in central Chile.

As companion animals, dogs and cats live in close contact with humans, generating the possibility of interspecies pathogen transmission events. Equine origin H3N8 and avian origin H5N1 influenza virus have been reported in dogs and cats respectively since 2004 with outbreaks associated with different strains recorded for both species in Asia and North America. To date, there have been no reports of influenza viruses from companion animals in South America. To fill this gap in knowledge, we performed active epidemiological surveillance in shelters that received abandoned animals, backyard production systems and veterinary clinics between May 2017 and January 2019 to estimate the burden of influenza infection in cats and dogs in the central region of Chile. Blood samples, oropharyngeal swabs or both were collected for influenza A virus detection by RT-qPCR, NP-ELISA, and hemagglutination inhibition assay. Logistic regression models were performed to assess the association between NP-ELISA-positivity and variables including sex and animal origin. The percentage of ELISA-positive samples was 43.5 % (95 % CI: 37.0-50.1) and 23.3 % (95 % CI: 10.6-42.7) for dogs and cats, respectively. No association was found between NP-ELISA results and sex or animal origin for either dogs or cats. Two ELISA positive samples showed hemagglutination inhibition titers against pandemic H1N1 influenza. One dog sample tested positive by RT-qPCR, indicating an overall RT-qPCR positivity in dogs of 1.1 % (95 % CI: 0.05-6.7). None of the tested cat samples were positive by this assay.

Temporal dynamics and the influence of environmental variables on the prevalence of avian influenza virus in main wetlands in central Chile.

Although wild birds are considered the main reservoir of the influenza A virus (IAV) in nature, empirical investigations exploring the interaction between the IAV prevalence in these populations and environmental drivers remain scarce. Chile has a coastline of more than 4000 kilometres with hundreds of wetlands, which are important habitats for both resident and inter-hemispheric migratory species. The aim of this study was to characterize the temporal dynamics of IAV in main wetlands in central Chile and to assess the influence of environmental variables on AIV prevalence. For that purpose, four wetlands were studied from September 2015 to June 2018. Fresh faecal samples of wild birds were collected for IAV detection by real-time RT-PCR. Furthermore, a count of wild birds present at the site was performed and environmental variables, such as temperature, rainfall, vegetation coverage (Normalized Difference Vegetation Index (NDVI)) and water body size, were determined. A generalized linear mixed model was built to assess the association between IAV prevalence and explanatory variables. An overall prevalence of 4.28% ± 0.28% was detected with important fluctuations among seasons, being greater during summer (OR = 4.87, 95% CI 2.11 to 11.21) and fall (OR = 2.59, 95% CI 1.12 to 5.97). Prevalence was positively associated with minimum temperature for the month of sampling and negatively associated with water body size measured two months before sampling, and NDVI measured three months before sampling. These results contribute to the understanding of IAV ecological drivers in Chilean wetlands providing important considerations for the global surveillance of IAV.

Equine-Like H3 Avian Influenza Viruses in Wild Birds, Chile.

Since their discovery in the United States in 1963, outbreaks of infection with equine influenza virus (H3N8) have been associated with serious respiratory disease in horses worldwide. Genomic analysis suggests that equine H3 viruses are of an avian lineage, likely originating in wild birds. Equine-like internal genes have been identified in avian influenza viruses isolated from wild birds in the Southern Cone of South America. However, an equine-like H3 hemagglutinin has not been identified. We isolated 6 distinct H3 viruses from wild birds in Chile that have hemagglutinin, nucleoprotein, nonstructural protein 1, and polymerase acidic genes with high nucleotide homology to the 1963 H3N8 equine influenza virus lineage. Despite the nucleotide similarity, viruses from Chile were antigenically more closely related to avian viruses and transmitted effectively in chickens, suggesting adaptation to the avian host. These studies provide the initial demonstration that equine-like H3 hemagglutinin continues to circulate in a wild bird reservoir.

Movement Restriction and Increased Surveillance as Efficient Measures to Control the Spread of Highly Pathogenic Avian Influenza in Backyard Productive Systems in Central Chile.

During the last 5 years there has been an alarming number of reports of highly pathogenic avian influenza worldwide. However, little is known about the status of this disease in South America. Chile has been the only country in South America where an HPAI outbreak was reported. This outbreak occurred in 2002 and was due to an H7N3 HPAI, where the most plausible hypothesis that explained the entrance of the disease to the country, had relation to migratory wild birds. Commercial poultry farms in Chile are highly integrated and have high biosecurity standards. Nevertheless, poultry backyard production systems lack biosecurity measures and are widely distributed. Since 2002 outbreak, avian influenza viruses have been identified in wild birds and different animal species kept in backyard productive systems (BPS) in Chile. The aim of this study was to simulate the possible natural history of HPAI after its introduction to BPS in central Chile and to simulate different intervention strategies. To do so, the North American Animal Disease Spread Model version 3.3 was used. The results showed that a median of 15,930 BPS would be affected if HPAI spread among BPS in central Chile, representing 97.8% of the current amount of BPS existing in study zone. Movement restrictions, pre-emptive destruction, passive surveillance, tracing of infected premises and combinations of the three, where the intervention strategies tested in the simulation model. From all the interventions simulated, movement restrictions together with increasing surveillance (through increasing passive surveillance and good tracing of infected premises) had the biggest effect, reducing the median number of infected BPS in 90.8%. However, more studies are needed to more accurately estimate local contact rates. These results can guide the official veterinary services to consider potential mechanisms to control or prevent an HPAI emergency situation.

Factores ambientales relacionados con la presentación de virus influenza A en aves silvestres / Environmental factors related to Influenza A virus occurrence in wild birds

RESUMEN La investigación y el interés por el virus influenza aviar han aumentado considerablemente en las últimas décadas en respuesta a los brotes de influenza aviar de alta patogenicidad en aves de corral y a su potencial zoonótico. Las aves silvestres acuáticas son el principal reservorio del virus en la naturaleza, por lo tanto, la comprensión de la dinámica de infección del virus influenza A (VIA) en estas poblaciones es fundamental para entender su potencial de persistencia en el ambiente y sus posibilidades de transmisión hacia aves domésticas y humanos. Se ha identificado que factores ambientales (como temperatura, precipitaciones, vegetación y características del paisaje, entre otros) pueden tener un importante rol en el mantenimiento y diseminación del virus en las zonas de concentración de aves silvestres. Sin embargo, los estudios que incluyen aspectos ecológicos del virus y que exploran la interacción entre la prevalencia del VIA en aves silvestres y el ambiente, continúan siendo escasos. En esta revisión se resumen los esfuerzos de investigación que se han realizado para identificar a los factores ambientales involucrados en la persistencia y transmisión del VIA en lugares de concentración de aves silvestres y cómo estos factores pueden incidir en la prevalencia del virus en estas poblaciones, generando diferencias en la presentación de la infección entre distintas zonas geográficas.

ABSTRACT Research and interest in avian influenza virus have increased considerably in recent decades in response to highly pathogenic avian influenza outbreaks in poultry and its zoonotic potential. Wild waterfowl are the main reservoir of the virus, therefore studying the dynamics of influenza A virus (IAV) infection in these populations is essential in order to understand its potential persistence in the environment and transmission to poultry and humans. It has been identified that environmental factors (such as temperature, rainfall, vegetation and landscape characteristics, among others) can play an important role in the maintenance and dissemination of the virus in the areas of concentration of wild birds. However, studies that include ecological aspects of the virus and explore the interaction between the prevalence of IAV in wild birds and environmental factors remain scarce. This review summarizes research efforts that have been made to identify the environmental factors involved in the persistence and transmission of IAV in areas of wild bird concentration and how these factors may influence the prevalence of the virus in these populations, generating differences in the presentation of the infection among different geographical areas.

Assessing Antibiotic Residues in Poultry Eggs from Backyard Production Systems in Chile, First Approach to a Non-Addressed Issue in Farm Animals.

Eggs are the main product generated from backyard poultry production systems (BPS) because they can quickly be consumed and sold to meet essential family needs. Nevertheless, antimicrobial residues can accumulate in this product. The objective of this study was to evaluate the presence of antimicrobial residues in eggs produced by poultry kept in BPS in central Chile. To assess this, eggs were obtained from 83 BPS and analysed to evaluate the presence of antibiotic residues (families: tetracyclines, beta-lactams, aminoglycosides and macrolides), using a Four-Plate Test screening method for the detection, based on a bacterial growth inhibition method. Results show a lack of biosecurity procedures at BPS level, making these systems susceptible to the dissemination of antimicrobial residues. These include intensive animal production units in the proximity, and the presence of shared watercourses with other farms. Furthermore, 66% of the surveyed owners are indicated as giving pharmacological treatments to their chickens. Eggs from 61 BPS were positive for at least one antimicrobial. Fifty-three BPS were positive for more than one antimicrobial, and one BPS was positive for all four antimicrobials tested. Consequently, there is a risk that poultry eggs produced in BPS in central Chile carry residues of different families of antimicrobials.

Risk factors and spatial relative risk assessment for influenza A virus in poultry and swine in backyard production systems of central Chile.

Backyard production systems (BPS) are a common form of poultry and swine production worldwide. The limited implementation of biosecurity standards in these operations makes BPS a potential source for the emergence of pathogens that have an impact on both animal and public health. Information regarding circulation of influenza A virus (IAV) in poultry and swine raised in BPS is scarce; particularly in South American countries. The objective of this study was to estimate prevalence and seroprevalence of IAV in BPS in central Chile, identify subtype diversity, evaluate risk factors and spatial relative risk for IAV. Samples were collected from 329 BPS from central Chile. Seroprevalence at BPS level was 34.7% (95% CI: 23.1%-46.2%), 19.7% (95% CI: 9.9%-30.6%) and 11.7% (95% CI: 7.2%-16.4%), whereas prevalence at BPS level was 4.2% (95% CI: 0.0%-8.8%), 8.2% (95% CI: 0.8%-14.0%) and 9.2% (95% CI: 4.8%-13.1%), for the Metropolitan, Valparaiso and LGB O'Higgins regions, respectively. Spatial analysis revealed that central-western area of Metropolitan region and the southern province of Valparaiso region could be considered as high-risk areas for IAV (spatial relative risk = 2.2, p < .05). Logistic regression models identified the practice of breeding both poultry and pigs at the BPS as a risk factor (95% CI 1.06-3.75). From 75 IAV ELISA-positive sera, 20 chicken sera had haemagglutination inhibition titres ranging from 20 to 160, and of these, 11 had microneutralization titres ranging from 40 to 960 for one or more IAV subtypes. Identified subtypes were H1, H3, H4, H9, H10 and H12. Results from this study highlight the need for further IAV surveillance programmes in BPS in Chile. Early detection of IAV strains circulating in backyard animals, especially in regions with large human populations, could have an enormous impact on animal and public health.

Low pathogenic avian influenza (H7N6) virus causing an outbreak in commercial Turkey farms in Chile.

In late 2016, an H7N6 low pathogenic avian influenza virus outbreak occurred in domestic turkeys in Central Chile. We characterized the genetic and antigenic properties of the outbreak virus and its experimental transmission in chickens. Our studies demonstrate that the outbreak virus is a reassortment of genes identified from Chilean wild bird viruses between 2013 and 2017 and displays molecular adaptations to poultry and antiviral resistance to adamantanes. Further, these wild bird viruses are also able to transmit in experimentally infected chickens highlighting the need for continued surveillance and improvement of biosecurity in poultry farms.

Backyard poultry production in Chile: animal health management and contribution to food access in an upper middle-income country.

Backyard production systems (BPS) that involve poultry are a good way to improve food security and poverty alleviation. Few studies have been carried out to quantify the contribution of poultry production to these households and the constraints they might face if a priority animal disease enters these systems. This study aims to characterize the poultry-rearing BPS in central Chile and to identify socio-economic factors associated to households' consumption of poultry. Data was collected from 384 BPS through a face-to-face semi-structured questionnaire. Value chain framework associated with BPS poultry rearing and cash flow analysis of BPS was done to identify the inputs/outputs of the system and to know the profitability of the system. Multiple linear regression was performed to identify the BPS and household factors associated to poultry consumption. The results of this study suggest that BPS in central Chile have biosecurity deficiencies such as: lack of confinement, lack of veterinary assistance and incorrect handling of dead animals. Cash flow analysis indicated that 62% of the BPS had a positive balance from production. Distance to closest market and per capita income were factors associated to poultry value to farmers. Different factors were significant predictors of household poultry consumption. Positive predictors of consumption were identified as: (i) older owners, (ii) higher transportation price to closest market, (iii) larger flock size (iv) birds raised by women and (v) owning a car. On the contrary, (i) higher per capita income and (ii) bigger household size predicted a reduction in consumption. The results indicate the importance of BPS to low-income families and those living in remote areas while also highlighting the vulnerability of these systems to disease risks.

Circulation of influenza in backyard productive systems in central Chile and evidence of spillover from wild birds.

Backyard productive systems (BPS) are recognized as the most common form of animal production in the world. However, BPS frequently exhibit inherent biosecurity deficiencies, and could play a major role in the epidemiology of animal diseases and zoonoses. The aim of this study was to determine if influenza A viruses (IAV) were prevalent in backyard poultry and swine BPS in central Chile. Through active surveillance in Valparaiso and Metropolitan regions from 2012 - 2014, we found that influenza virus positivity by real-time RT-PCR (qRT-PCR) ranged from 0% during winter 2012-45.8% during fall 2014 at the farm level. We also obtained an H12 hemagglutinin (HA) sequence of wild bird origin from a domestic Muscovy duck (Cairina moschata), indicating spillover from wild birds into backyard poultry populations. Furthermore, a one-year sampling effort in 113 BPS in the Libertador Bernardo O'Higgins (LGB ÓHiggins) region showed that 12.6% of poultry and 2.4% of swine were positive for IAV by enzyme-linked immunosorbent assay (ELISA), indicative of previous exposure of farm animals to IAV. This study highlights the need for improved IAV surveillance in backyard populations given the close interaction between domestic animals, wild birds and people in these farms, particularly in an understudied region, like South America.

Wild birds in Chile Harbor diverse avian influenza A viruses.

While the circulation of avian influenza viruses (IAV) in wild birds in the northern hemisphere has been well documented, data from South America remain sparse. To address this gap in knowledge, we undertook IAV surveillance in wild birds in parts of Central and Northern Chile between 2012 and 2015. A wide diversity of hemagglutinin (HA) and neuraminidase (NA) subtypes were identified and 16 viruses were isolated including low pathogenic H5 and H7 strains, making this the largest and most diverse collection of Chilean avian IAVs to date. Unlike IAVs isolated from wild birds in other South American countries where the genes were most like viruses isolated from wild birds in either North America or South America, the Chilean viruses were reassortants containing genes like viruses isolated from both continents. In summary, our studies demonstrate that genetically diverse avian IAVs are circulating in wild birds in Chile highlighting the need for further investigation in this understudied area of the world.

Swine Influenza Virus (H1N2) Characterization and Transmission in Ferrets, Chile.

Phylogenetic analysis of the influenza hemagglutinin gene (HA) has suggested that commercial pigs in Chile harbor unique human seasonal H1-like influenza viruses, but further information, including characterization of these viruses, was unavailable. We isolated influenza virus (H1N2) from a swine in a backyard production farm in Central Chile and demonstrated that the HA gene was identical to that in a previous report. Its HA and neuraminidase genes were most similar to human H1 and N2 viruses from the early 1990s and internal segments were similar to influenza A(H1N1)pdm09 virus. The virus replicated efficiently in vitro and in vivo and transmitted in ferrets by respiratory droplet. Antigenically, it was distinct from other swine viruses. Hemagglutination inhibition analysis suggested that antibody titers to the swine Chilean H1N2 virus were decreased in persons born after 1990. Further studies are needed to characterize the potential risk to humans, as well as the ecology of influenza in swine in South America.

Avian H11 influenza virus isolated from domestic poultry in a Colombian live animal market.

Live animal markets (LAMs) are an essential source of food and trade in Latin American countries; however, they can also serve as 'hotbeds' for the emergence and potential spillover of avian influenza viruses (AIV). Despite extensive knowledge of AIV in Asian LAMs, little is known about the prevalence South American LAMs. To fill this gap in knowledge, active surveillance was carried out at the major LAM in Medellin, Colombia between February and September 2015. During this period, overall prevalence in the market was 2.67% and a North American origin H11N2 AIV most similar to a virus isolated from Chilean shorebirds asymptomatically spread through multiple bird species in the market resulting in 17.0% positivity at peak of infection. Phenotypically, the H11 viruses displayed no known molecular markers associated with increased virulence in birds or mammals, had α2,3-sialic acid binding preference, and caused minimal replication in vitro and little morbidity in vivo. However, the Colombian H11N2 virus replicated and transmitted effectively in chickens explaining the spread throughout the market. Genetic similarity to H11 viruses isolated from North and South American shorebirds suggest that the LAM occurrence may have resulted from a wild bird to domestic poultry spillover event. The ability to spread in domestic poultry as well as potential for human infection by H11 viruses highlight the need for enhanced AIV surveillance in South America in both avian species and humans.