Hyperimmunized Chickens Produce Neutralizing Antibodies against SARS-CoV-2.

The novel severe acute respiratory syndrome (SARS) coronavirus, SARS-CoV-2, is responsible for the global COVID-19 pandemic. Effective interventions are urgently needed to mitigate the effects of COVID-19 and likely require multiple strategies. Egg-extracted antibody therapies are a low-cost and scalable strategy to protect at-risk individuals from SARS-CoV-2 infection. Commercial laying hens were hyperimmunized against the SARS-CoV-2 S1 protein using three different S1 recombinant proteins and three different doses. Sera and egg yolk were collected at three and six weeks after the second immunization for enzyme-linked immunosorbent assay and plaque-reduction neutralization assay to determine antigen-specific antibody titers and neutralizing antibody titers, respectively. In this study we demonstrate that hens hyperimmunized against the SARS-CoV-2 recombinant S1 and receptor binding domain (RBD) proteins produced neutralizing antibodies against SARS-CoV-2. We further demonstrate that antibody production was dependent on the dose and type of antigen administered. Our data suggests that antibodies purified from the egg yolk of hyperimmunized hens can be used as immunoprophylaxis in humans at risk of exposure to SARS-CoV-2.

Comparison of cellular immune responses to avian influenza virus in two genetically distinct, highly inbred chicken lines.

Low pathogenicity avian influenza causes mild disease involving the respiratory, gastrointestinal, and reproductive systems of wild and domestic birds. Avian influenza research often emphasizes the effect of the virus genetics on disease, but the influence of host genetics on resistance to infection is not well understood. The genetic determinants of enhanced resistance to influenza can be explored by using genetically distinct, highly inbred chicken lines that differ in susceptibility to influenza. In this study, we compared the mucosal cellular immune responses between the relatively resistant Fayoumi M43 chicken line and the relatively susceptible Leghorn GB2 chicken line after challenging with low pathogenicity avian influenza virus (LPAIV) H6N2. The birds were inoculated at 21 days of age with 107 50 % egg infective dose (EID50) LPAIV H6N2 via nasal and tracheal routes in two separate experiments. Clinical signs were recorded, tracheal swabs were collected to measure viral titer, and tracheas and lungs were harvested for flow cytometric analysis of macrophage, B cell, and T cell populations at 4 days post-infection (dpi) (Experiments 1 and 2) and 6 dpi (Experiment 2). Blood and tears were also collected at 7 and 14 dpi (Experiment 1) to measure antibody levels. Compared to both the non-challenged Fayoumis and the relatively susceptible Leghorn chickens, relatively resistant Fayoumi chickens challenged with LPAIV demonstrated enhanced MHC class I expression on antigen-presenting cells and increased macrophage, B cell, and T cell frequencies in the trachea, which were associated with reduced tracheal viral titers at 4 dpi. In contrast, MHC class I expression and immune cell frequencies in the trachea were not different between challenged Leghorns and non-challenged Leghorns. Furthermore, Leghorns shed higher virus titers in their trachea compared to Fayoumis. Challenged Fayoumis and Leghorns both produced AIV-specific IgY detected in the serum and tears, but AIV-specific IgA was not detected in the tears. In this study, we provide new insight into immune mechanisms of enhanced resistance to avian influenza in chickens, which may lead to improved vaccination strategies and breeding programs.

Molecular Characterization of Newcastle Disease Viruses Isolated from Chickens in Tanzania and Ghana.

Newcastle disease (ND) is one of the most challenging infectious diseases affecting poultry production in Africa, causing major economic losses. To date, Newcastle disease virus isolates from several African countries have been grouped into class II NDV genotypes I, IV, V, VI, VII, XI, XIII, XIV, XVII, XVIII and XXI. Although ND is endemic in many African countries, information on circulating genotypes is still scarce. In Tanzania, outbreaks with genotypes V and XIII have been reported. In West and Central Africa, genotypes XIV, XVII, and XVIII are the most predominant. To investigate other genotypes circulating in Tanzania and Ghana, we performed molecular genotyping on isolates from Tanzania and Ghana using the MinION, a third-generation portable sequencing device from Oxford Nanopore Technologies. Using the MinION, we successfully sequenced the NDV F gene hypervariable region of 24 isolates from Tanzania and four samples from Ghana. In Tanzania, genotypes V, VII and XIII were detected. All isolates from Ghana belonged to genotype XVIII. The data obtained in this study reflect the genetic diversity of NDV in Africa and highlight the importance of surveillance for monitoring the distribution of NDV genotypes and viral evolution.

Effect of Pullet Vaccination on Development and Longevity of Immunity.

Avian respiratory disease causes significant economic losses in commercial poultry. Because of the need to protect long-lived poultry against respiratory tract pathogens from an early age, vaccination programs for pullets typically involve serial administration of a variety of vaccines, including infectious bronchitis virus (IBV), Newcastle disease virus (NDV), and infectious laryngotracheitis virus (ILTV). Often the interval between vaccinations is only a matter of weeks, yet it is unknown whether the development of immunity and protection against challenge when vaccines are given in short succession occurs in these birds, something known as viral interference. Our objective was to determine whether serially administered, live attenuated vaccines against IBV, NDV, and ILTV influence the development and longevity of immunity and protection against challenge in long-lived birds. Based on a typical pullet vaccination program, specific-pathogen-free white leghorns were administered multiple live attenuated vaccines against IBV, NDV, and ILTV until 16 weeks of age (WOA), after which certain groups were challenged with IBV, NDV, or ILTV at 20, 24, 28, 32, and 36 WOA. Five days post-challenge, viral load, clinical signs, ciliostasis, tracheal histopathology, and antibody titers in serum and tears were evaluated. We demonstrate that pullets serially administered live attenuated vaccines against IBV, NDV, and ILTV were protected against homologous challenge with IBV, NDV, or ILTV for at least 36 weeks, and conclude that the interval between vaccinations used in this study (at least 2 weeks) did not interfere with protection. This information is important because it shows that a typical pullet vaccination program consisting of serially administered live attenuated vaccines against multiple respiratory pathogens can result in the development of protective immunity against each disease agent.

Evaluation of a coccidia vaccine using spray and gel applications.

Coccidiosis is an economically significant disease of poultry caused by species of Eimeria, a parasitic protozoan. Disease can result in poor feed conversion, reduced weight gain, and can lead to the development of necrotic enteritis. For prevention of coccidiosis, poultry are commonly vaccinated with a live, sporulated oocysts mass applied with a vaccination cabinet in the hatchery. Traditionally, coccidia vaccines have been applied by coarse spray in a water based diluent, however, new technology using gel diluents has entered the US market. Gel diluents can have variable viscosities and are "dropped" onto chicks with an applicator bar. It is thought that gel droplets remain intact on the birds for longer than water based droplets, allowing more time for preening and ingestion of oocysts. In this experiment, the efficacy of a commercial coccidia vaccine applied with a water based diluent, a more viscous gel diluent, and a less viscous gel diluent was compared. Fecal samples were collected at multiple time points post-vaccination to quantify vaccine oocyst shedding. Shedding in the first cycle (days 5 to 8 post-vaccination) was related to the number of oocysts received from each application method, where the groups receiving higher doses shed more oocysts. However, a decrease in shedding was seen for the more viscous gel group in the second cycle (days 12 to 15 post-vaccination). Chickens were challenged with Eimeria maxima oocysts and 7 days post-challenge body weight gains and gross and microscopic lesions were recorded to evaluate protection levels for the different vaccine applications. All vaccinated groups appeared to be protected based on body weight gain and lesion scoring. The results of this project indicate that all vaccine applications are effective at protecting against Eimeria maxima challenge when using a proper dose of vaccine that allows for repeated oocyst cycling in the litter post-vaccination.

Use of filter papers to determine seroprevalence of Toxoplasma gondii among hunted ungulates in remote Peruvian Amazon.

Toxoplasmosis is a zoonosis caused by the protozoan Toxoplasma gondii, and it is found worldwide. To determine whether ungulates are reservoirs of T. gondii in an isolated and remote region of the northeastern Peruvian Amazon, antibodies to T. gondii were determined in 5 species of ungulates by the modified agglutination test (MAT). These animals were hunted by subsistence hunters along the Yavarí-Mirín River, in the northeastern Peruvian Amazon. Blood samples were collected by hunters on filter papers. For determination of T. gondii antibodies, blood was eluted from filter papers, and a titer of 1:25 was considered indicative of exposure to T. gondii. Antibodies to T. gondii were found in 26 (31.0%) peccaries (Pecari tajacu, Tayassu pecari), six (17.1%) brocket deer (Mazama americana, Mazama gouazoubira), and four (40.0%) lowland tapir (Tapirus terrestris). We also introduced a modification to the MAT protocol that allows the extraction of fluid samples from several types of laboratory-grade filter paper, thus enabling researchers to easily adapt their approaches to the materials presented to them.