@article{criollo_john_gaghan_fletcher_thachil_crespo_kulkarni_2024, title={Characterization of immune responses and immunopathology in turkeys experimentally infected with clostridial dermatitis-producing strains of Clostridium septicum}, volume={269}, ISSN={["1873-2534"]}, DOI={10.1016/j.vetimm.2024.110717}, abstractNote={Clostridium septicum is one of the major causative agents of clostridial dermatitis (CD), an emerging disease of turkeys, characterized by sudden deaths and necrotic dermatitis. Despite its economic burden on the poultry industry, the immunopathological changes and pathogen-specific immune responses are poorly characterized. Here, we used three strains of C. septicum, namely Str. A1, Str. B1 and Str. C1, isolated from CD field outbreaks, to experimentally infect turkeys to evaluate local (skin and muscle) and systemic (spleen) pathological and immunological responses. Results showed that while all three strains produced an acute disease, Str. A1 and B1 caused significantly higher mortality when compared to Str. C1. Gross and histopathology evaluation showed that birds infected with Str. A1 and B1 had severe inflammatory, edematous, granulomatous and necrotic lesions in the skin, muscle and spleen, while these lesions produced by Str. C1 were relatively less severe and mostly confined to skin and/or muscle. Immune gene expression in these tissues showed that Str. B1-infected birds had significantly higher expression of interleukin (IL)−1β, IL-6 and interferon (IFN)γ genes compared to uninfected control, suggesting a robust inflammatory response both locally as well as systemically. The transcription of IL-1β and IFNγ in the muscle or spleen of Str. A1-infected birds and IL-1β in the skin of Str. C1-infected group was also significantly higher than control. Additionally, Str. A1 or B1-infected groups also had significantly higher IL-4 transcription in these tissues, while birds infected with all three strains developed C. septicum-specific serum antibodies. Furthermore, splenic cellular immunophenotyping in the infected turkeys showed a marked reduction in CD4+ cells. Collectively, it can be inferred that host responses against C. septicum involve an acute inflammatory response along with antibody production and that the disease severity seem to depend on the strain of C. septicum involved in CD in turkeys.}, journal={VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY}, author={Criollo, Valeria and John, Feba Ann and Gaghan, Carissa and Fletcher, Oscar J. and Thachil, Anil and Crespo, Rocio and Kulkarni, Raveendra R.}, year={2024}, month={Mar} }
 @article{boone_kulkarni_cortes_gaghan_mohammed_villalobos_esandi_gimeno_2024, title={Evaluation of Adjuvant Effect of Cytosine-Guanosine-Oligodeoxynucleotide in Meat-Type Chickens Coadministered <i>In Ovo</i> with Herpesvirus of Turkey Vaccine}, volume={2}, ISSN={["1557-8976"]}, url={https://doi.org/10.1089/vim.2023.0125}, DOI={10.1089/vim.2023.0125}, abstractNote={Herpesvirus of turkey (HVT) increases activation of T cells in 1-day-old chickens when administered in ovo. This study evaluated whether adding cytosine-guanosine oligodeoxynucleotides (CpG ODNs) to the HVT vaccine could enhance the adjuvant effect of HVT. We used a CpG ODN dose of 10 μg per egg. The experimental groups were (1) diluent-only control (sham), (2) HVT, (3) HVT+CpG ODN, (4) HVT+non-CpG ODN, (5) CpG ODN, and (6) non-CpG ODN control. Cellular response evaluation included measuring the frequencies of macrophages (KUL01+MHC-II+), gamma delta T cells (γδTCR+MHC-II+), CD4+, and CD8+ T cell subsets, including double-positive (DP) cells. In addition, CD4+ and CD8+ T cell activation was evaluated by measuring the cellular expression of major histocompatibility complex class II (MHC-II), CD44 or CD28 costimulatory molecules. An adjuvant effect was considered when HVT+CpG ODN, but not HVT+non CpG ODN, or CpG ODN, or non-CpG ODN, induced significantly increased effects on any of the immune parameters examined when compared with HVT. The findings showed that (1) HVT vaccination induced significantly higher frequencies of γδ+MHC-II+ and CD4+CD28+ T cells when compared with sham chickens. Frequencies of DP and CD4+CD28+ T cells in HVT-administered birds were significantly higher than those observed in the non-CpG ODN group. (2) Groups receiving HVT+CpG ODN or CpG ODN alone were found to have significantly increased frequencies of activated CD4+ and CD8+ T cells, when compared with HVT. Our results show that CpG ODN administration in ovo with or without HVT significantly increased frequencies of activated CD4+ and CD8+ T cells.}, journal={VIRAL IMMUNOLOGY}, author={Boone, Allison C. and Kulkarni, Raveendra R. and Cortes, Aneg L. and Gaghan, Carissa and Mohammed, Javid and Villalobos, Tarsicio and Esandi, Javier and Gimeno, Isabel M.}, year={2024}, month={Feb} }
 @article{sharma_kulkarni_sharif_hassan_alizadeh_pratt_abdelaziz_2024, title={In ovo feeding of probiotic lactobacilli differentially alters expression of genes involved in the development and immunological maturation of bursa of Fabricius in pre-hatched chicks}, volume={103}, ISSN={["1525-3171"]}, url={https://doi.org/10.1016/j.psj.2023.103237}, DOI={10.1016/j.psj.2023.103237}, abstractNote={Compelling evidence indicates that immunological maturation of the gut-associated lymphoid tissues, including the bursa of Fabricius, is dependent upon antigenic stimulation post-hatch. In view of these data, the present study investigated the impact of exposing the immune system of chick embryos to antigenic stimuli, via in ovo delivery of poultry-specific lactobacilli, on the expression of genes associated with early bursal development and maturation. Broiler line embryonated eggs were inoculated with 106 and 107 colony-forming units (CFUs) of an individual or a mixture of Lactobacillus species, including L. crispatus (C25), L. animalis (P38), L. acidophilus (P42), and L. reuteri (P43), at embryonic day 18 (ED18). The bursa of Fabricius was collected from pre-hatched chicks (ED20) to measure the expression levels of various immune system genes. The results revealed that L. acidophilus and the mixture of Lactobacillus species at the dose of 106 CFU consistently elicited higher expression of genes responsible for B cell development, differentiation, and survival (B cell activating factor (BAFF), BAFF-receptor (BAFF-R)), and antibody production (interleukin (IL)-10) and diversification (TGF-β). Similar expression patterns were also noted in T helper (Th) cell-associated cytokine genes, including Th1-type cytokines (interferon (IFN)-γ and IL-12p40), Th2-type cytokines (IL-4 and IL-13) and Th17 cytokine (IL-17). Overall, these results suggest that the supplementation of poultry-specific lactobacilli to chick embryos might be beneficial for accelerating the development and immunological maturation of the bursa of Fabricius. However, further studies are required to determine if the changes in gene expression are associated with the developmental trajectory and phenotypes of bursal cells.}, number={1}, journal={POULTRY SCIENCE}, author={Sharma, Shreeya and Kulkarni, Raveendra R. and Sharif, Shayan and Hassan, Hosni and Alizadeh, Mohammadali and Pratt, Scott and Abdelaziz, Khaled}, year={2024}, month={Jan} }
 @article{abdelaziz_nixon_joye_hassan_alizadeh_sharif_kulkarni_2024, title={Modulation of functional activity of heat-stressed chicken macrophages by poultry-derived probiotic lactobacilli}, volume={4}, ISSN={["1918-1825"]}, DOI={10.1139/cjas-2023-0124}, abstractNote={This study investigated the potential role of lactobacilli in mitigating the negative effects of heat stress on the functional activity of chicken macrophages. Macrophage-like MQ-NCSU cells were incubated at 40oC or 44oC in the presence or absence of a single or a mixture of different poultry-derived Lactobacillus species, including L. animalis, L. acidophilus, L. reuteri, and L. crispatus. Macrophage activation was evaluated by measuring nitric oxide (NO) production, phagocytic activity, and the transcription levels of cytokines, chemokine, and Toll-like receptors (TLRs). Macrophages exposed to heat stress exhibited increased production of NO, diminished expression of interleukin (IL)-1β and IL-12p40, and elevated expression of TLR2 and TLR4, whereas no significant alterations in the phagocytic activity of macrophages were observed. Conversely, treatment of macrophages with probiotic lactobacilli counteracted the effects associated with heat stress. This was evidenced by a notable enhancement in macrophage phagocytic activity, NO production and expression of IL-1β, IL-12p40, IL-18, and CXCL8, coupled with a reduction in TLR2 and TLR4 expression. These findings suggest that probiotic lactobacilli could be given to chickens to mitigate the negative effects of heat stress on the innate immune system. However, further studies are required to validate the observed effects in an in vivo model.}, journal={CANADIAN JOURNAL OF ANIMAL SCIENCE}, author={Abdelaziz, Khaled and Nixon, Thandi and Joye, Annie and Hassan, Hosni and Alizadeh, Mohammadali and Sharif, Shayan and Kulkarni, Raveendra R.}, year={2024}, month={Apr} }
 @article{kulkarni_gaghan_mohammed_sharif_taha-abdelaziz_2023, title={Cellular Immune Responses in Lymphoid Tissues of Broiler Chickens Experimentally Infected with Necrotic Enteritis–Producing Clostridium perfringens Strains}, volume={67}, url={http://dx.doi.org/10.1637/aviandiseases-d-23-00012}, DOI={10.1637/aviandiseases-d-23-00012}, abstractNote={SUMMARY. Host cellular responses against Clostridium perfringens (CP), the causative agent of necrotic enteritis (NE) in chickens, are poorly understood. In the present study, we first tested the NE-producing ability of seven netB+ CP strains (CP5, CP18, CP26, CP64, CP67, CP68, and NCNE-1), using an experimental infection model of broiler chickens. Evaluation of intestinal gross lesions showed that all the strains, except CP5, were able to produce NE, while CP26 and CP64 strains produced relatively more severe lesions when compared with other groups. Next, cellular responses in the cecal tonsil (CT), bursa of Fabricius, and spleen were evaluated in chickens infected with strains representing variation in the level of virulence, namely, avirulent CP5, virulent CP18, and a relatively more virulent CP26 strain. Immunophenotyping analysis showed that CT or splenic macrophage frequencies were significantly higher in CP18- and CP26-infected chickens compared with uninfected controls, while the frequencies of γδ T-cells and B-cells in the CT of CP26-infected chickens were significantly higher than those in the uninfected, CP5- or CP18-infected groups. The T-cell analysis showed that chickens infected with CP18 and CP26 had a significantly higher number of splenic CD4+ and CD8+ T-cells expressing CD44 and CD28 activation molecules, while CP26-infected chickens also had significantly increased CT frequency of these activated CD4+ and CD8+ T-cells when compared with uninfected or CP5-infected groups. Collectively, our findings suggested that cellular responses, including activation of T-cells, are selectively induced against virulent CP strains and that the NE-producing characteristics of this pathogen may influence the outcome of immunity to NE. RESUMEN. Respuestas inmunes celulares en tejidos linfoides de pollos de engorde infectados experimentalmente con cepas de Clostridium perfringens productoras de enteritis necrótica. Las respuestas celulares del huésped contra Clostridium perfringens (CP), el agente causante de la enteritis necrótica (NE) en pollos, son poco conocidas. En el presente estudio, primero se analizó la capacidad de producción de enteritis necrótica de siete cepas de C. perfringens netB+ (CP5, CP18, CP26, CP64, CP67, CP68 y NCNE-1), utilizando un modelo de infección experimental de pollos de engorde. La evaluación de las lesiones macroscópicas intestinales mostró que todas las cepas, excepto CP5, podían producir enteritis necrótica, mientras que las cepas CP26 y CP64 produjeron lesiones relativamente más severas en comparación con los otros grupos. Posteriormente, se evaluaron las respuestas celulares en las tonsilas cecales (CT), la bolsa de Fabricio y en el bazo de pollos infectados con cepas que representan variaciones en el nivel de virulencia, por ejemplo las cepas CP5 avirulenta, CP18 virulenta y la cepa CP26 relativamente más virulenta. El análisis de inmunofenotipado mostró que las frecuencias de los macrófagos esplénicos o de las tonsilas cecales fueron significativamente más altas en los pollos infectados con las cepas CP18 y CP26 en comparación con los controles no infectados, mientras que las frecuencias de células T γd y células B en tonsilas cecales de los pollos infectados con la cepa CP26 fueron significativamente más altas que las de los pollos de los grupos no infectados, o infectados con las cepas CP5 o CP18. El análisis de células T mostró que los pollos infectados con las cepas CP18 y CP26 tenían un número significativamente mayor de células esplénicas T CD4+ y CD8+ que expresaban moléculas de activación CD44 y CD28, mientras que los pollos infectados con la cepa CP26 también tenían una frecuencia significativamente mayor en las tonsilas cecales de estas células T CD4+ y CD8+ activadas en comparación con grupos no infectados o infectados con la cepa CP5. En conjunto, estos hallazgos sugirieron que las respuestas celulares, incluida la activación de las células T, se inducen selectivamente contra las cepas virulentas de C. perfringens y que las características productoras de enteritis necrótica de este patógeno pueden influir en el resultado de la inmunidad contra la enteritis necrótica.}, number={2}, journal={Avian Diseases}, publisher={American Association of Avian Pathologists (AAAP)}, author={Kulkarni, Raveendra R. and Gaghan, Carissa and Mohammed, Javid and Sharif, Shayan and Taha-Abdelaziz, Khaled}, year={2023}, month={Jun} }
 @article{gaghan_browning_cortes_gimeno_kulkarni_2023, title={Effect of CpG-Oligonucleotide in Enhancing Recombinant Herpes Virus of Turkey-Laryngotracheitis Vaccine-Induced Immune Responses in One-Day-Old Broiler Chickens}, volume={11}, ISSN={["2076-393X"]}, url={https://doi.org/10.3390/vaccines11020294}, DOI={10.3390/vaccines11020294}, abstractNote={Infectious laryngotracheitis (ILT) is an economically important disease of chickens. While the recombinant vaccines can reduce clinical disease severity, the associated drawbacks are poor immunogenicity and delayed onset of immunity. Here, we used CpG-oligonucleotides (ODN) as an in ovo adjuvant in boosting recombinant herpesvirus of turkey-laryngotracheitis (rHVT-LT) vaccine-induced responses in one-day-old broiler chickens. Two CpG-ODN doses (5 and 10 μg/egg) with no adverse effect on the vaccine-virus replication or chick hatchability were selected for immune-response evaluation. Results showed that while CpG-ODN adjuvantation induced an increased transcription of splenic IFNγ and IL-1β, and lung IFNγ genes, the IL-1β gene expression in the lung was significantly downregulated compared to the control. Additionally, the transcription of toll-like receptor (TLR)21 in the spleen and lung and inducible nitric oxide synthase (iNOS) in the spleen of all vaccinated groups was significantly reduced. Furthermore, splenic cellular immunophenotyping showed that the CpG-ODN-10μg adjuvanted vaccination induced a significantly higher number of macrophages, TCRγδ+, and CD4+ T cells as well as a higher frequency of activated T cells (CD4+CD44+) when compared to the control. Collectively, the findings suggested that CpG-ODN can boost rHVT-LT-induced immune responses in day-old chicks, which may help in anti-ILT defense during their later stages of life.}, number={2}, journal={VACCINES}, author={Gaghan, Carissa and Browning, Matthew and Cortes, Aneg L. and Gimeno, Isabel M. and Kulkarni, Raveendra R.}, year={2023}, month={Feb} }
 @article{alizadeh_shojadoost_astill_taha-abdelaziz_karimi_bavananthasivam_kulkarni_sharif_2023, title={Effects of <i>in ovo</i> inoculation of multi-strain lactobacilli on cytokine gene expression and antibody-mediated immune responses in chickens (vol 10, 1252518, 2023)}, volume={10}, ISSN={["2297-1769"]}, DOI={10.3389/fvets.2023.1252518}, abstractNote={[This corrects the article DOI: 10.3389/fvets.2020.00105.].}, journal={FRONTIERS IN VETERINARY SCIENCE}, author={Alizadeh, Mohammadali and Shojadoost, Bahram and Astill, Jake and Taha-Abdelaziz, Khaled and Karimi, Seyed Hossein and Bavananthasivam, Jegarubee and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2023}, month={Oct} }
 @article{criollo_gaghan_john_orozco_thachil_crespo_kulkarni_2023, title={Immune Response Evaluation in Commercial Turkeys Affected with Clostridial Dermatitis}, volume={67}, ISSN={["1938-4351"]}, DOI={10.1637/aviandiseases-D-22-00089}, abstractNote={SUMMARY. Clostridial dermatitis (CD), caused by Clostridium septicum and Clostridium perfringens, is an economically important emerging disease of turkeys characterized by sudden deaths and necrotic dermatitis. Immune responses in CD-affected commercial turkeys are poorly understood. In the present study, C. septicum was isolated from CD-affected commercial turkeys during a recent outbreak, and the tissues (skin, muscle, and spleen) were collected and analyzed for immune gene expression, along with samples from clinically healthy birds. The results showed that CD-affected turkeys had significantly higher levels of IL-1β, IL-6, IFNγ, and iNOS transcripts in the skin, muscle, and spleen tissues compared to healthy birds. Affected turkeys also had a significantly elevated transcription of toll-like receptor (TLR21) gene in the skin and spleen tissues, suggesting a role for this receptor in the immune recognition. The expression of IL-4 and IL-13 genes in the spleen and muscle was also significantly higher in the affected birds. Additional birds from the same affected and healthy farms examined for serology revealed that the CD-affected turkeys had significantly higher levels of serum IgM and IgY antibodies. Furthermore, in vitro stimulation of MQ-NCSU macrophages with C. septicum led to a significant transcriptional upregulation of IL-1β and IFNγ genes, while the IL-10 gene expression was downregulated. The surface expression of MHC-II protein and cellular production of nitric oxide were also significantly increased in the C. septicum-stimulated macrophages, indicating cellular activation. Collectively, our findings suggest that the host responses in CD-affected turkeys involve a robust inflammatory response as well as a response mediated by IL4/IL-13 cytokines that may aid in antibody-mediated immunity. RESUMEN. Evaluación de la respuesta inmune en pavos comerciales afectados por dermatitis clostridial. La dermatitis clostridial (CD), causada por Clostridium septicum y Clostridium perfringens, es una enfermedad emergente económicamente importante de los pavos caracterizada por muerte súbitas y dermatitis necrótica. Se conoce poco acerca de las respuestas inmunitarias en pavos comerciales afectados por dermatitis clostridial. En el presente estudio, se aisló C. septicum de pavos comerciales afectados por dermatitis clostridial durante un brote reciente, y los tejidos (piel, músculo y bazo) se recolectaron y analizaron para determinar la expresión de genes inmunitarios junto con muestras de aves clínicamente sanas. Los resultados mostraron que los pavos afectados por dermatitis clostridial tenían niveles significativamente más altos de transcritos de IL-1β, IL-6, IFNγ, and iNOS en los tejidos de la piel, los músculos y el bazo en comparación con las aves sanas. Los pavos afectados también tenían una transcripción significativamente elevada del gene del receptor tipo toll (TLR21) en los tejidos de la piel y el bazo, lo que sugiere un papel de este receptor en el reconocimiento inmunitario. La expresión de los genes IL-4 e IL-13 en el bazo y el músculo también fue significativamente mayor en las aves afectadas. Aves adicionales de las mismas granjas afectadas y sanas que fueron examinadas por serología revelaron que los pavos afectados por dermatitis clostridial tenían niveles significativamente más altos de anticuerpos séricos IgM e IgY. Además, la estimulación in vitro de los macrófagos MQ-NCSU con C. septicum condujo a una regulación transcripcional significativamente al alza de los genes IL-1β and IFNγ, mientras que la expresión del gene IL-10 se reguló a la baja. La expresión superficial de la proteína MHC-II y la producción celular de óxido nítrico también aumentaron significativamente en los macrófagos estimulados por C. septicum, lo que indica activación celular. Colectivamente, estos hallazgos sugieren que las respuestas del huésped en pavos afectados por dermatitis clostridial implican una respuesta inflamatoria robusta, así como una respuesta mediada por citoquinas IL4/IL-13 que pueden ayudar en la inmunidad mediada por anticuerpos.}, number={1}, journal={AVIAN DISEASES}, author={Criollo, Valeria and Gaghan, Carissa and John, Feba and Orozco, Eric and Thachil, Anil and Crespo, Rocio and Kulkarni, Raveendra R.}, year={2023}, month={Mar}, pages={80–88} }
 @article{gaghan_browning_fares_abdul-careem_gimeno_kulkarni_2023, title={In Ovo Vaccination with Recombinant Herpes Virus of the Turkey-Laryngotracheitis Vaccine Adjuvanted with CpG-Oligonucleotide Provides Protection against a Viral Challenge in Broiler Chickens}, volume={15}, ISSN={["1999-4915"]}, url={https://www.mdpi.com/1999-4915/15/10/2103}, DOI={10.3390/v15102103}, abstractNote={Infectious laryngotracheitis (ILT) is an economically important disease in chickens. We previously showed that an in ovo adjuvantation of recombinant herpesvirus of the turkey-Laryngotracheitis (rHVT-LT) vaccine with CpG-oligonucleotides (ODN) can boost vaccine-induced responses in one-day-old broiler chickens. Here, we evaluated the protective efficacy of in ovo administered rHVT-LT + CpG-ODN vaccination against a wild-type ILT virus (ILTV) challenge at 28 days of age and assessed splenic immune gene expression as well as cellular responses. A chicken-embryo-origin (CEO)-ILT vaccine administered in water at 14 days of age was also used as a comparative control for the protection assessment. The results showed that the rHVT-LT + CpG-ODN or the CEO vaccinations provided significant protection against the ILTV challenge and that the level of protection induced by both the vaccines was statistically similar. The protected birds had a significantly upregulated expression of interferon (IFN)γ or interleukin (IL)-12 cytokine genes. Furthermore, the chickens vaccinated with the rHVT-LT + CpG-ODN or CEO vaccine had a significantly higher frequency of γδ T cells and activated CD4+ or CD8+ T cells, compared to the unvaccinated-ILTV challenge control. Collectively, our findings suggest that CpG-ODN can be used as an effective adjuvant for rHVT-LT in ovo vaccination to induce protective immunity against ILT in broiler chickens.}, number={10}, journal={VIRUSES-BASEL}, author={Gaghan, Carissa and Browning, Matthew and Fares, Abdelhamid M. and Abdul-Careem, Mohamed Faizal and Gimeno, Isabel M. and Kulkarni, Raveendra R.}, year={2023}, month={Oct} }
 @article{boone_kulkarni_cortes_villalobos_esandi_gimeno_2023, title={In ovo HVT vaccination enhances cellular responses at hatch and addition of poly I:C offers minimal adjuvant effects}, volume={41}, ISSN={["1873-2518"]}, url={https://doi.org/10.1016/j.vaccine.2023.02.076}, DOI={10.1016/j.vaccine.2023.02.076}, abstractNote={In ovo vaccination with herpesvirus of turkey (HVT) hastens immunocompetence in chickens and the recommended dose (RD) of 6080 plaque-forming-units (PFU) offers the most optimal effects. In previous studies conducted in egg-type chickens, in ovo vaccination with HVT enhanced lymphoproliferation, wing-web thickness with phytohemagglutinin-L (PHA-L), and increased spleen and lung interferon-gamma(IFN-γ) andToll-like receptor 3 (TLR3) transcripts. Here, we evaluated the cellular mechanisms by which HVT-RD can hasten immunocompetence in one-day-old meat-type chickens, and also determined if HVT adjuvantation with a TLR3 agonist, polyinosinic-polycytidylic acid (poly(I:C)), could enhance vaccine-induced responses and provide dose-sparing effects. Compared to sham-inoculated chickens, HVT-RD significantly increased transcription of splenic TLR3 and IFN γ receptor 2 (R2), and lung IFN γ R2, while the splenic IL-13 transcription was found decreased. Additionally, these birds showed increased wing-web thickness following PHA-L inoculation. The thickness was due to an innate inflammatory cell population, CD3+ T cells, and edema. In another experiment, HVT-1/2 (3040 PFU) supplemented with 50 μg poly(I:C) [HVT-1/2 + poly(I:C)] was administered in ovo and immune responses were compared with those produced by HVT-RD, HVT-1/2, 50 μg poly(I:C), and sham-inoculated. Immunophenotyping of splenocytes showed HVT-RD induced a significantly higher frequency of CD4+, CD4+MHC-II+, CD8+CD44+, and CD4+CD28+ T cells compared to sham-inoculated chickens, and CD8+MHC-II+, CD4+CD8+, CD4+CD8+CD28+, and CD4+CD8+CD44+ T cells compared to all groups. Treatment groups, except HVT-1/2 + poly(I:C), had significantly higher frequencies of γδ T cells and all groups induced significantly higher frequencies of activated monocytes/macrophages, compared to sham-inoculated chickens. Poly(I:C)-induced dose-sparing effect was only observed in the frequency of activated monocytes/macrophages. No differences in the humoral responses were observed. Collectively, HVT-RD downregulated IL-13 transcripts (Th2 immune response) and had strong immunopotentiation effects on innate immune responses and the activation of T cells. However addition of poly(I:C) offered a minimal adjuvant/dose-sparing effect.}, number={15}, journal={VACCINE}, author={Boone, Allison C. and Kulkarni, Raveendra R. and Cortes, Aneg L. and Villalobos, Tarsicio and Esandi, Javier and Gimeno, Isabel M.}, year={2023}, month={Apr}, pages={2514–2523} }
 @misc{taha-abdelaziz_singh_sharif_sharma_kulkarni_alizadeh_yitbarek_helmy_2023, title={Intervention Strategies to Control Campylobacter at Different Stages of the Food Chain}, volume={11}, ISSN={["2076-2607"]}, url={https://doi.org/10.3390/microorganisms11010113}, DOI={10.3390/microorganisms11010113}, abstractNote={Campylobacter is one of the most common bacterial pathogens of food safety concern. Campylobacter jejuni infects chickens by 2–3 weeks of age and colonized chickens carry a high C. jejuni load in their gut without developing clinical disease. Contamination of meat products by gut contents is difficult to prevent because of the high numbers of C. jejuni in the gut, and the large percentage of birds infected. Therefore, effective intervention strategies to limit human infections of C. jejuni should prioritize the control of pathogen transmission along the food supply chain. To this end, there have been ongoing efforts to develop innovative ways to control foodborne pathogens in poultry to meet the growing customers’ demand for poultry meat that is free of foodborne pathogens. In this review, we discuss various approaches that are being undertaken to reduce Campylobacter load in live chickens (pre-harvest) and in carcasses (post-harvest). We also provide some insights into optimization of these approaches, which could potentially help improve the pre- and post-harvest practices for better control of Campylobacter.}, number={1}, journal={MICROORGANISMS}, author={Taha-Abdelaziz, Khaled and Singh, Mankerat and Sharif, Shayan and Sharma, Shreeya and Kulkarni, Raveendra R. and Alizadeh, Mohammadali and Yitbarek, Alexander and Helmy, Yosra A.}, year={2023}, month={Jan} }
 @article{gaghan_gorrell_taha-abdelaziz_sharif_kulkarni_2023, title={Intracloacal Inoculation of Broiler Chickens with Clostridium perfringens Strains: Evaluation of Necrotic Enteritis Disease Development and Lymphoid Immune Responses}, volume={11}, ISSN={["2076-2607"]}, url={https://doi.org/10.3390/microorganisms11030771}, DOI={10.3390/microorganisms11030771}, abstractNote={Necrotic enteritis (NE) is an economically important disease of chickens. We have recently shown that inflammatory responses in chickens inoculated orally with virulent Clostridium perfringens were spatially regulated. Here, we used previously virulence-characterized netB+ C. perfringens strains, avirulent CP5 and virulent CP18 and CP26, to assess the severity of NE and immune responses in broiler chickens when inoculated intracloacally. The results showed that CP18- and CP26-infected birds had a reduced weight gain and developed milder/less severe NE lesions, as determined by the gross lesions scores, suggesting a subclinical-grade infection. Gene expression analysis in infected birds revealed three statistically significant observations compared to uninfected-control: (1) Increased expression of anti-inflammatory/immunoregulatory interleukin (IL)-10/transforming growth factor (TGF)β in cecal tonsil (CT) and bursa of Fabricius in the CP18/CP26-infected groups. (2) Increased CT transcription of pro-inflammatory IL-1β, IL-6 and interferon (IFN)γ and decreased Harderian gland (HG) expression of IFNγ in the CP18/CP26-infected birds. (3) Increased HG or bursal expression of IL-4 and IL-13 in CP5-infected birds. Collectively, intracloacal C. perfringens inoculation seems to induce a highly regulated inflammatory response in the CT and other mucosal lymphoid organs and an intracloacal infection model may be useful in evaluating immune responses in chickens with subclinical NE.}, number={3}, journal={MICROORGANISMS}, author={Gaghan, Carissa and Gorrell, Kaitlin and Taha-Abdelaziz, Khaled and Sharif, Shayan and Kulkarni, Raveendra R.}, year={2023}, month={Mar} }
 @article{kulkarni_gaghan_gorrell_fletcher_2023, title={Mucosal and systemic lymphoid immune responses against Clostridium perfringens strains with variable virulence in the production of necrotic enteritis in broiler chickens}, volume={1}, ISSN={["1465-3338"]}, DOI={10.1080/03079457.2022.2154195}, abstractNote={ABSTRACT Necrotic enteritis (NE), caused by Clostridium perfringens, is an economically important disease of chickens. Although NE pathogenesis is moderately well studied, the host immune responses against C. perfringens are poorly understood. The present study used an experimental NE model to characterize lymphoid immune responses in the caecal tonsils (CT), bursa of Fabricius, Harderian gland (HG) and spleen tissues of broiler chickens infected with four netB+ C. perfringens strains (CP1, CP5, CP18, and CP26), of which CP18 and CP26 strains also carried the tpeL gene. The gross and histopathological lesions in chickens revealed CP5 to be avirulent, while CP1, CP18, and CP26 strains were virulent with CP26 being “very virulent”. Gene expression analysis showed that, while the virulent strains induced a significantly upregulated expression of pro-inflammatory IL-1β gene in CT, the CP26-infected birds had significantly higher CT transcription of IFNγ and IL-6 pro-inflammatory genes compared to CP5-infected or uninfected chickens. Furthermore, CP26 infection also led to significantly increased bursal and HG expression of the anti-inflammatory/regulatory genes, IL-10 or TGFβ, compared to control, CP5 and CP1 groups. Additionally, the splenic pro- and anti-inflammatory transcriptional changes were observed only in the CP26-infected chickens. An antibody-mediated response, as characterized by increased IL-4 and/or IL-13 transcription and elevated IgM levels in birds infected with virulent strains, particularly in the CP26-infected group compared to uninfected controls, was also evident. Collectively, our findings suggest that lymphoid immune responses during NE in chickens are spatially regulated such that the inflammatory responses against C. perfringens depend on the virulence of the strain.}, journal={AVIAN PATHOLOGY}, author={Kulkarni, Raveendra R. and Gaghan, Carissa and Gorrell, Kaitlin and Fletcher, Oscar J.}, year={2023}, month={Jan} }
 @article{adams_kulkarni_mohammed_crespo_2022, title={A flow cytometric method for enumeration and speciation of coccidia affecting broiler chickens}, volume={301}, ISSN={["1873-2550"]}, url={https://doi.org/10.1016/j.vetpar.2021.109634}, DOI={10.1016/j.vetpar.2021.109634}, abstractNote={Production losses, mortality, and control measures associated with coccidiosis, caused by Eimera species, cost the broiler industry over $14 billion a year. Current means to distinguish Eimeria species such as oocyst morphology, pre-patent period and site of infection are subjective, labor intensive or unsuitable for high-throughput applications. Although Polymerase Chain Reaction (PCR) techniques have been validated, the target gene cannot differentiate relative abundance of each species in mixed infections. In this study, we developed a non-antibody-based flow cytometry high throughput method to simultaneously enumerate and speciate four Eimeria species, E. acervulina, E. mitis, E. maxima, and E. tenella, using commercial coccidia vaccine as well as field fecal samples. Our findings showed that the four Eimeria oocyst populations could be distinctly speciated based on their size and granularity (shape) via scatter plotting. These distinct populations were sorted and confirmed by quantitative real-time PCR assay. Finally, the flow cytometry findings were applied to enumerate and speciate oocysts from fecal samples collected from commercial broiler flocks vaccinated for coccidiosis at day of hatch and the results were validated against the conventional manual method of floatation and microscopic examination. Collectively, the findings of this study suggested that non-antibody based Flow Cytometry technique can be successful in the simultaneous enumeration and speciation of coccidia. Further development and validation is needed to make this diagnostic tool useful for field applications at a much larger scale as well as to speciate other Eimeria species.}, journal={VETERINARY PARASITOLOGY}, publisher={Elsevier BV}, author={Adams, Daniel S. and Kulkarni, Raveendra R. and Mohammed, Javid P. and Crespo, Rocio}, year={2022}, month={Jan} }
 @article{lieshout_rghei_cao_he_soule_zhu_thomas_sorensen_frost_tierney_et al._2022, title={AAV-monoclonal antibody expression protects mice from Ebola virus without impeding the endogenous antibody response to heterologous challenge}, volume={26}, ISSN={["2162-2531"]}, DOI={10.1016/j.omtm.2022.08.003}, abstractNote={Filoviruses cause severe hemorrhagic fever with case fatality rates as high as 90%. Filovirus-specific monoclonal antibodies (mAbs) confer protection in nonhuman primates as late as 5 days after challenge, and FDA-approved mAbs REGN-EB3 and mAb114 have demonstrated efficacy against Ebola virus (EBOV) infection in humans. Vectorized antibody expression mediated by adeno-associated virus (AAV) can generate protective and sustained concentrations of therapeutic mAbs in animal models for a variety of infectious diseases, including EBOV. Here we demonstrate that AAV6.2FF-mediated expression of murine IgG2a EBOV mAbs, 2G4 and 5D2, protects from mouse-adapted (MA)-EBOV infection with none of the surviving mice developing anti-VP40 antibodies above background. Protective serum concentrations of AAV6.2FF-2G4/AAV6.2FF-5D2 did not alter endogenous antibody responses to heterologous virus infection. AAV-mediated expression of EBOV mAbs 100 and 114, and pan-ebolavirus mAbs, FVM04, ADI-15878, and CA45, as human IgG1 antibodies conferred protection against MA-EBOV at low serum concentrations, with minimum protective serum levels as low as 2 μg/mL. Vectorized expression of murine IgG2a or human IgG1 mAbs led to sustained expression in the serum of mice for >400 days or for the lifetime of the animal, respectively. AAV6.2FF-mediated mAb expression offers an alternative to recombinant antibody administration in scenarios where long-term protection is preferable to passive immunization.}, journal={MOLECULAR THERAPY-NUCLEIC ACIDS}, author={Lieshout, Laura P. and Rghei, Amira D. and Cao, Wenguang and He, Shihua and Soule, Geoff and Zhu, Wenjun and Thomas, Sylvia P. and Sorensen, Debra and Frost, Kathy and Tierney, Kevin and et al.}, year={2022}, month={Sep}, pages={505–518} }
 @article{kulkarni_gaghan_mohammed_2022, title={Avian Macrophage Responses to Virulent and Avirulent Clostridium perfringens}, volume={11}, ISSN={["2076-0817"]}, url={https://doi.org/10.3390/pathogens11010100}, DOI={10.3390/pathogens11010100}, abstractNote={The present study evaluated the avian macrophage responses against Clostridium perfringens that varied in their ability to cause necrotic enteritis in chickens. Strains CP5 (avirulent-netB+), CP1 (virulent-netB+), and CP26 (highly virulent-netB+tpeL+) were used to evaluate their effect on macrophages (MQ-NCSU cells) and primary splenic and cecal tonsil mononuclear cells. The bacilli (whole cells) or their secretory products from all three strains induced a significant increase in the macrophage transcription of Toll-like receptor (TLR)21, TLR2, interleukin (IL)-1β, inducible nitric oxide synthase (iNOS), and CD80 genes as well as their nitric oxide (NO) production and major histocompatibility complex (MHC)-II surface expression compared to an unstimulated control. The CP1 and CP26-induced expression of interferon (IFN)γ, IL-6, CD40 genes, MHC-II upregulation, and NO production was significantly higher than that of CP5 and control groups. Furthermore, splenocytes and cecal tonsillocytes stimulated with bacilli or secretory products from all the strains showed a significant increase in the frequency of macrophages, their surface expression of MHC-II and NO production, while CP26-induced responses were significantly higher for the rest of the groups. In summary, macrophage interaction with C. perfringens can lead to cellular activation and, the ability of this pathogen to induce macrophage responses may depend on its level of virulence.}, number={1}, journal={PATHOGENS}, author={Kulkarni, Raveendra R. and Gaghan, Carissa and Mohammed, Javid}, year={2022}, month={Jan} }
 @article{gaghan_adams_mohammed_crespo_livingston_kulkarni_2022, title={Characterization of vaccine-induced immune responses against coccidiosis in broiler chickens}, volume={40}, ISSN={["1873-2518"]}, url={https://doi.org/10.1016/j.vaccine.2022.05.043}, DOI={10.1016/j.vaccine.2022.05.043}, abstractNote={Coccidiosis, caused by Eimeria protozoan species, is an economically important enteric disease of poultry. Although commercial live vaccines are widely used for disease control, the vaccine-induced protective immune mechanisms are poorly characterized. The present study used a commercial broiler vaccine containing a mixture of E. acervulina, E. maxima, and E. tenella. One-day-old chicks were vaccinated by spray followed by a challenge at 21 days of age with a mixture of wild type Eimeria species via oral gavage. Oocyst shedding, immune gene expression and cellular responses in the spleen and cecal tonsils were measured at pre- (days 14 and 21) and post-challenge (days 24, 28 and 35) time points. Results showed that the oocyst counts were significantly reduced in the vaccinated chickens at post-challenge compared to unvaccinated control group. While the vaccinated birds had a significantly increased toll-like receptor (TLR) 21 gene expression at pre-challenge, the transcription of interferon (IFN)γ, Interleukin (IL)-12 and CD40 genes in spleen and cecal tonsils of these birds was significantly higher at post-challenge compared to unvaccinated chickens. Cellular immunophenotyping analysis found that vaccination led to increased frequency of macrophages and activated T cells (CD8+CD44+ and CD4+CD44+) in the spleen and cecal tonsils at post-challenge. Furthermore, in vitro stimulation of chicken macrophages (MQ-NCSU cells) with purified individual species of E. acervulina, E. maxima, and E. tenella showed a significantly increased expression of TLR21, TLR2 and IFNγ genes as well as nitric oxide production. Collectively, these findings suggest that TLR21 and TLR2 may be involved in the immune cell recognition of Eimeria parasites and that the vaccine can induce a robust macrophage activation leading to a T helper-1 dominated protective response at both local and systemic lymphoid tissues.}, number={29}, journal={VACCINE}, publisher={Elsevier BV}, author={Gaghan, Carissa and Adams, Daniel and Mohammed, Javid and Crespo, Rocio and Livingston, Kimberly and Kulkarni, Raveendra R.}, year={2022}, month={Jun}, pages={3893–3902} }
 @article{shojadoost_alizadeh_boodhoo_astill_karimi_doost_taha-abdelaziz_kulkarni_sharif_2022, title={Effects of Treatment with Lactobacilli on Necrotic Enteritis in Broiler Chickens}, volume={1}, ISSN={["1867-1314"]}, DOI={10.1007/s12602-021-09901-5}, abstractNote={Growth promoter antibiotics have been commonly used for the control of necrotic enteritis (NE) in broilers for decades. However, due to a ban on the use of these antibiotics, alternatives such as probiotics have been tested widely for NE control. The present study tested the efficacy of four different species of lactobacilli (two isolates of Lactobacillus johnsonii and one of Ligilactobacillus (L.) salivarius, Limosilactobacillus (L.) reuteri, and L. crispatus) against NE. Day-old male broiler chickens were divided into six groups and orally inoculated with 1 × 10 7  or 1 × 10 8  colony-forming units (CFU) of lactobacilli on 1, 7, 14, and 20 days of age. While negative and positive control groups did not receive lactobacilli, the latter was challenged with Clostridium perfringens (CP). Chickens, at 21 days old, were challenged for 3 days with 3 × 10 8  CFU of a virulent strain of CP. Tissues were collected for immune system gene expression, immunophenotyping, intestinal histomorphometry, and microbiota analysis. Lactobacilli inoculation conferred partial protection in chickens against NE, marked by lowered lesion scores and improved villus:crypt ratio. Immunomodulatory effects were demonstrated by the significant alteration of interferon (IFN)-γ, interleukin (IL)-1β, IL-2, IL-12p35, IL-17, and transforming growth factor beta (TGF-β) gene transcription in the duodenum and jejunum as well as subtle changes in the frequency of CD8 + T cells and B cells in the cecal tonsil of the treated chickens. Microbiota analysis showed increased levels of some bacterial phyla including Actinobacteria, Lactobacillaceae, and Firmicutes. In conclusion, these findings suggest that the use of certain lactobacilli can reduce NE severity and modulate immune responses and intestinal microbiota composition in chickens.}, journal={PROBIOTICS AND ANTIMICROBIAL PROTEINS}, author={Shojadoost, Bahram and Alizadeh, Mohammadali and Boodhoo, Nitish and Astill, Jake and Karimi, Seyed Hossein and Doost, Janan Shoja and Taha-Abdelaziz, Khaled and Kulkarni, Raveendra and Sharif, Shayan}, year={2022}, month={Jan} }
 @article{kairmi_taha-abdelaziz_yitbarek_sargolzaei_spahany_astill_shojadoost_alizadeh_kulkarni_parkinson_et al._2022, title={Effects of therapeutic levels of dietary antibiotics on the cecal microbiome composition of broiler chickens}, volume={101}, ISSN={["1525-3171"]}, DOI={10.1016/j.psj.2022.101864}, abstractNote={Dietary antibiotics, including antibiotic growth promoters (AGPs), have been commonly used to improve health and growth of poultry. The present study investigated the effects of therapeutic doses of dietary antibiotics, including bacitracin methylene disalicylate (BMD), penicillin G potassium (PP) and an ionophore (salinomycin, SA), on the cecal microbiome of chickens. BMD and SA treatments were given as dietary supplements from d 1 to 35 of age. The SAPP (salinomycin+ penicillin G potassium) group was given SA as a dietary supplement from d 1 to 35 of age and PP was added to drinking water from d 19 to 24 of age to simulate common practices for control of necrotic enteritis in broilers. The cecal contents were collected from all treatment groups on d 10, 24, and 35 of age and DNA was extracted for metagenomic analysis of the cecal microbiome. The results revealed that dietary or water supplementation of therapeutic levels of antibiotics and ionophores to chickens significantly altered the cecal microbial homeostasis during different stages of the chicken life. The alpha diversity analysis showed that BMD, SA, and SAPP treatments decreased diversity and evenness of the cecal microbiome of treated chickens on d 10 of age. Species richness was also reduced on d 35 following treatment with BMD. Beta diversity analyses revealed that SAPP and BMD induced significant changes in the relative abundance of Gram-positive and -negative bacteria on d 10, while no significant differences were observed on d 24. On d 35, the non-treated control group had higher relative abundance of unclassified Gram-positive and -negative bacteria compared to SA, SAPP, and BMD treatment groups. Overall, despite their beneficial role in controlling necrotic enteritis outbreaks, the findings of this study highlight the potential negative effects of dietary supplementation of therapeutic levels of antibiotics on the gut microbiome and suggest that adjusting gut bacteria may be required to restore microbial richness and diversity of the gut microbiome following treatment with these antibiotics.}, number={6}, journal={POULTRY SCIENCE}, author={Kairmi, Seyed Hossien and Taha-Abdelaziz, Khaled and Yitbarek, Alexander and Sargolzaei, Mehdi and Spahany, Heidi and Astill, Jake and Shojadoost, Bahram and Alizadeh, Mohammadali and Kulkarni, Raveendra R. and Parkinson, John and et al.}, year={2022}, month={Jun} }
 @article{boyett_crespo_vinueza_gaghan_mohammed_kulkarni_2022, title={Enumeration and speciation of coccidia affecting turkeys using flow cytometry method}, volume={31}, ISSN={["1537-0437"]}, url={https://doi.org/10.1016/j.japr.2022.100270}, DOI={10.1016/j.japr.2022.100270}, abstractNote={Enumeration of Eimeria oocysts is a common practice in monitoring coccidiosis in turkeys; however, the conventional method of manual microscopic examination of Eimeria oocysts is time-consuming. Previously, we used flow cytometry (FCM) to enumerate and speciate coccidia affecting chickens and here, we extended those findings to turkey coccidia species, E. adenoides and E. meleagrimitis. Using FCM, a commercial vaccine containing these species was used to optimize the scatter-plot parameters, including Forward-(size) and Side-(shape/granularity) scatter Area, Height, and Width patterns for Eimeria. The two Eimeria species populations in the vaccine were accurately phenotyped and the gated populations were then sorted using a Cell sorter instrument to obtain pure oocyst suspensions. The individual Eimeria species identity of sorted oocysts was confirmed by PCR using species-specific primers. A significant (P = 0.0466) correlation (R = 0.9893) in the total oocyst count between FCM and manual methods were observed. Furthermore, when FCM was employed to analyze farm fecal samples, the close similarities in the oocyst morphologies coupled with organic debris particulate interference prevented a precise separation of these 2 species resulting in a lack of oocyst count (OPG) correlation between the 2 methods. The OPG counts by FCM were much lower than the manual method; however, a partial OPG trend between the two methods was observed only at the early timepoint collections during a period of 35 d. Collectively, our findings showed that FCM can be used in the enumeration of turkey Eimeria oocysts with a potential scope for a more precise enumeration and speciation in field samples.}, number={3}, journal={JOURNAL OF APPLIED POULTRY RESEARCH}, publisher={Elsevier BV}, author={Boyett, Taylor and Crespo, Rocio and Vinueza, Valeria C. and Gaghan, Carissa and Mohammed, Javid P. and Kulkarni, Raveendra R.}, year={2022}, month={Sep} }
 @article{boodhoo_shojadoost_alizadeh_kulkarni_sharif_2022, title={Ex Vivo Differential Responsiveness to Clostridium perfringens and Lactococcus lactis by Avian Small Intestine Macrophages and T Cells}, volume={13}, ISSN={["1664-3224"]}, DOI={10.3389/fimmu.2022.807343}, abstractNote={Tissue resident immune system cells in the chicken intestine play a significant role in the protection against pathogens. However, very little is known about these cells. The current study was conducted to further characterize chicken intestinal immune system cells. Furthermore, this study aimed to assess the immune modulatory action of a highly virulentClostridium perfringens, a commonly found chicken intestinal microbe, in comparison with a non-commensal,Lactococcus lactis, on intestine-derived immune system cells. The results demonstrated varying distribution of innate and adaptive immune cells along the avian gut-associated lymphoid tissue (GALT) in the duodenum, jejunum, ileum, and cecal tonsils. In addition, steady-state and tissue-specific presence of CD25+ cells among αβ and γδ T-cell subsets was assessed along the intestine.Ex vivostimulation withC. perfringensorL. lactisresulted in a significant increase in the frequency of CD25+ T cells (γδ and αβ T cells). In addition, significantly more cell death was observed inex vivostimulation withC. perfringens, which was indirectly correlated with a decrease in macrophage activation based on nitric oxide (NO) production with no effect on lymphoid cell responsiveness as per intracellular interferon (IFN)-gamma (γ) staining.Ex vivostimulation withL. lactisactivated γδ T cells and αβ T cells, based on intracellular IFN-γ staining, while it had limited effect on macrophages. However, the ability of γδ and αβ T cells to produce IFN-γ and the ability of macrophages production of NO was rescued in the presence ofL. lactis.These results demonstrate the potential application ofL. lactis, as a probiotic, against virulentC. perfringensinfection in chicken.}, journal={FRONTIERS IN IMMUNOLOGY}, author={Boodhoo, Nitish and Shojadoost, Bahram and Alizadeh, Mohammadali and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2022}, month={Feb} }
 @article{daneshmand_kermanshahi_mohammed_sekhavati_javadmanesh_ahmadian_alizadeh_razmyar_kulkarni_2022, title={Intestinal changes and immune responses during Clostridium perfringens-induced necrotic enteritis in broiler chickens}, volume={101}, ISSN={["1525-3171"]}, url={https://doi.org/10.1016/j.psj.2021.101652}, DOI={10.1016/j.psj.2021.101652}, abstractNote={Clostridium perfringens-induced necrotic enteritis (NE) is an economically important disease of broiler chickens. The present study evaluated the effect of C. perfringens on the intestinal histomorphometry, enteric microbial colonization, and host immune responses using 3 experimental NE reproduction methods. The experimental groups consisted of 1) unchallenged Control diet (corn-soybean meal), 2) Control diet + Eimera inoculation at d 11 followed by C. perfringens challenge at d 15 (ECp), 3) Wheat-based diet + C. perfringens challenge (WCp), and 4) Wheat-based diet + Eimeria inoculation followed by C. perfringens challenge (WECp). The results showed that chickens receiving ECp and WECp had reduced (P < 0.05) bird performance coupled with enteric gross lesions and epithelial damage at d 17 and 24 of age compared to unchallenged control birds. These ECp and WECp administered birds also had increased (P < 0.05) ileal colonization by clostridia and E. coli at d 17 and 24, while the resident Lactobacillus counts were reduced (P < 0.05) at d 24 of age. Furthermore, at d 24, jejunal transcription of IL-6, IL-10, annexin-A1 and IL-2 genes was upregulated (P < 0.05) in the ECp group, whereas the transcription of TNF receptor associated factor (TRAF)-3 gene was increased (P < 0.05) in WECp treated birds when compared to unchallenged control group. Additionally, stimulation of chicken splenocytes and cecal tonsilocytes with virulent C. perfringens bacilli or their secretory proteins resulted in a higher (P < 0.05) frequency of T cells and their upregulation of MHC-II molecule, as determined by flow cytometry. These findings suggest that C. perfringens, while inducing epithelial damage and changes in microbiota, can also trigger host immune responses. Furthermore, NE reproduction methods using coccidia with or without the wheat-based dietary predisposition seem to facilitate an optimal NE reproduction in broiler chickens and thus, may provide better avenues for future C. perfringens research.}, number={3}, journal={POULTRY SCIENCE}, publisher={Elsevier BV}, author={Daneshmand, Ali and Kermanshahi, Hassan and Mohammed, Javid and Sekhavati, Mohammad Hadi and Javadmanesh, Ali and Ahmadian, Monireh and Alizadeh, Marzieh and Razmyar, Jamshid and Kulkarni, Raveendra R.}, year={2022}, month={Mar} }
 @misc{kulkarni_gaghan_gorrell_sharif_taha-abdelaziz_2022, title={Probiotics as Alternatives to Antibiotics for the Prevention and Control of Necrotic Enteritis in Chickens}, volume={11}, ISSN={["2076-0817"]}, url={https://doi.org/10.3390/pathogens11060692}, DOI={10.3390/pathogens11060692}, abstractNote={Necrotic enteritis (NE) in poultry is an economically important disease caused by Clostridium perfringens type A bacteria. A global trend on restricting the use of antibiotics as feed supplements in food animal production has caused a spike in the NE incidences in chickens, particularly in broiler populations. Amongst several non-antibiotic strategies for NE control tried so far, probiotics seem to offer promising avenues. The current review focuses on studies that have evaluated probiotic effects on C. perfringens growth and NE development. Several probiotic species, including Lactobacillus, Enterococcus, Bacillus, and Bacteroides bacteria as well as some yeast species have been tested in chickens against C. perfringens and NE development. These findings have shown to improve bird performance, reduce C. perfringens colonization and NE-associated pathology. The underlying probiotic mechanisms of NE control suggest that probiotics can help maintain a healthy gut microbial balance by modifying its composition, improve mucosal integrity by upregulating expression of tight-junction proteins, and modulate immune responses by downregulating expression of inflammatory cytokines. Collectively, these studies indicate that probiotics can offer a promising platform for NE control and that more investigations are needed to study whether these experimental probiotics can effectively prevent NE in commercial poultry operational settings.}, number={6}, journal={PATHOGENS}, publisher={MDPI AG}, author={Kulkarni, Raveendra R. and Gaghan, Carissa and Gorrell, Kaitlin and Sharif, Shayan and Taha-Abdelaziz, Khaled}, year={2022}, month={Jun} }
 @article{bavananthasivam_alqazlan_alizadeh_matsuyama-kato_astill_kulkarni_sharif_2022, title={The Regulatory Microenvironment in Feathers of Chickens Infected with Very Virulent Marek's Disease Virus}, volume={14}, ISSN={["1999-4915"]}, DOI={10.3390/v14010112}, abstractNote={Vaccines against Marek’s disease can protect chickens against clinical disease; however, infected chickens continue to propagate the Marek’s disease virus (MDV) in feather follicles and can shed the virus into the environment. Therefore, the present study investigated if MDV could induce an immunoregulatory microenvironment in feathers of chickens and whether vaccines can overcome the immune evasive mechanisms of MDV. The results showed an abundance of CD4+CD25+ and CD4+ transforming growth factor-beta (TGF-β)+ T regulatory cells in the feathers of MDV-infected chickens at 21 days post-infection. In contrast, vaccinated chickens had a lower number of regulatory T cells. Furthermore, the expression of TGF-β and programmed cell death receptor (PD)-1 increased considerably in the feathers of Marek’s disease virus-infected chickens. The results of the present study raise the possibility of an immunoregulatory environment in the feather pulp of MDV-infected chickens, which may in turn favor replication of infectious MDV in this tissue. Exploring the evasive strategies employed by MDV will facilitate the development of control measures to prevent viral replication and transmission.}, number={1}, journal={VIRUSES-BASEL}, author={Bavananthasivam, Jegarubee and Alqazlan, Nadiyah and Alizadeh, Mohammadali and Matsuyama-Kato, Ayumi and Astill, Jake and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2022}, month={Jan} }
 @misc{shojadoost_yitbarek_alizadeh_kulkarni_astill_boodhoo_sharif_2021, title={IMMUNOLOGY, HEALTH AND DISEASE}, volume={100}, ISSN={["1525-3171"]}, url={https://doi.org/10.1016/j.psj.2020.12.027}, DOI={10.1016/j.psj.2020.12.027}, abstractNote={Vitamins are nutritional elements which are necessary for essential activities such as development, growth, and metabolism of cells. In addition to these conventional functions, vitamins A, D, E, and C have vital roles in normal function of the immune system as their deficiency is known to impair innate and adaptive host responses. By altering transcription of multiple immune system genes and contributing to antioxidant activities, these vitamins influence the immune system in different ways including modulation of cell-mediated and antibody-mediated responses, immunoregulation, and antiinflammatory effects. Furthermore, supplementation of these vitamins to poultry may assist the immune system to combat microbial pathogens while reducing detrimental effects associated with stress and enhancing responses to vaccines. In this article, the relationship between the chicken immune system and vitamins A, D, E, and C is reviewed, and evidence from the literature pertaining to how these vitamins exert their antiinflammatory, regulatory, and antimicrobial effects is discussed.}, number={4}, journal={POULTRY SCIENCE}, publisher={Elsevier BV}, author={Shojadoost, Bahram and Yitbarek, Alexander and Alizadeh, Mohammadali and Kulkarni, Raveendra R. and Astill, Jake and Boodhoo, Nitish and Sharif, Shayan}, year={2021}, month={Apr} }
 @misc{alizadeh_shojadoost_boodhoo_astill_taha-abdelaziz_hodgins_kulkarni_sharif_2021, title={Necrotic enteritis in chickens: a review of pathogenesis, immune responses and prevention, focusing on probiotics and vaccination}, volume={22}, ISSN={["1475-2654"]}, DOI={10.1017/S146625232100013X}, abstractNote={Abstract}, number={2}, journal={ANIMAL HEALTH RESEARCH REVIEWS}, author={Alizadeh, Mohammadali and Shojadoost, Bahram and Boodhoo, Nitish and Astill, Jake and Taha-Abdelaziz, Khaled and Hodgins, Douglas C. and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2021}, month={Dec}, pages={147–162} }
 @article{alizadeh_shojadoost_astill_taha-abdelaziz_karimi_bavananthasivam_kulkarni_sharif_2020, title={Effects of in ovo Inoculation of Multi-Strain Lactobacilli on Cytokine Gene Expression and Antibody-Mediated Immune Responses in Chickens}, volume={7}, ISBN={2297-1769}, DOI={10.3389/fvets.2020.00105}, abstractNote={This study was conducted to investigate the effects of various doses of a multi-strain lactobacilli mixture (Lactobacillus salivarius, Lactobacillus reuteri, Lactobacillus crispatus, and Lactobacillus johnsonii) on the innate and adaptive immune responses in broiler chickens. At embryonic day eighteen, 200 eggs were injected with PBS, or three different doses of a multi-strain lactobacilli mixture (1 × 105, 1 × 106, and 1 × 107 CFU/egg, P1, P2, and P3 respectively) along with a group of negative control. On days 5 and 10 post-hatch, cecal tonsil, bursa of fabricius, and spleen were collected for gene expression and cellular analysis. On days 14 and 21 post-hatch, birds were immunized intramuscularly with both sheep red blood cells (SRBC) and keyhole limpet hemocyanin (KLH). Serum samples were collected on days 0, 7, 14, and 21 after primary immunization. The results demonstrated that lactobacilli inoculation increased the splenic expression of cytokines, including interferon (IFN)-α, IFN-β, IFN-γ, interleukin (IL)-8, and IL-12 on day 5 post-hatch compared to the control group (PBS). However, in cecal tonsils, lactobacilli treatment downregulated the expression of IL-6 on day 5 post-hatch and IL-2 and IL-8 on day 10 post-hatch. No significant differences were observed in the expression of cytokine genes in the bursa except for IL-13 which was upregulated in lactobacilli-treated groups P2 and P3 on days 5 and 10 post-hatch. Flow cytometry analysis showed that the percentage of KUL01, CD4+ and CD8+ splenocytes was not affected by treatments. In addition, no significant differences were observed for antibody titers against SRBC. However, lactobacilli treatment (P1, P2, and P3) was found to increase IgM titers on day 21 post-primary immunization compared to controls. Furthermore, in ovo injection of the highest dose of probiotics (1 × 107, P3) increased serum IgG titers against KLH on day 7 post-primary immunization. In conclusion, this study demonstrated that that in ovo administration of lactobacilli can improve antibody-mediated immune responses and differentially modulate cytokine expression in mucosal and systemic lymphoid tissues of chickens.}, journal={FRONTIERS IN VETERINARY SCIENCE}, author={Alizadeh, Mohammadali and Shojadoost, Bahram and Astill, Jake and Taha-Abdelaziz, Khaled and Karimi, Seyed Hossein and Bavananthasivam, Jegarubee and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2020} }
 @article{kulkarni_taha-abdelaziz_shojadoost_astill_sharif_2020, title={Gastrointestinal diseases of poultry: causes and nutritional strategies for prevention and control}, volume={73}, ISBN={["978-1-78676-304-4"]}, ISSN={["2059-6944"]}, DOI={10.19103/AS.2019.0059.11}, journal={IMPROVING GUT HEALTH IN POULTRY}, author={Kulkarni, Raveendra R. and Taha-Abdelaziz, Khaled and Shojadoost, Bahram and Astill, Jake and Sharif, Shayan}, year={2020}, pages={205–236} }
 @article{boone_kaser_cortes_kulkarni_abad_villalobos_esandi_perozo_lemiere_gimeno_2020, title={In ovo vaccination with herpesvirus of turkey enhances innate and cellular responses in meat-type chickens: Effect of vaccine dose and strain}, volume={38}, ISSN={["1873-2518"]}, DOI={10.1016/j.vaccine.2020.05.050}, abstractNote={In ovo vaccination with herpesvirus of turkey (HVT) or recombinant HVT (rHVT) is commonly used in meat-type chickens. Previous studies showed that in ovo vaccination with HVT enhances innate, cellular, and humoral immune responses in egg-type chicken embryos. This study evaluated if in ovo vaccination with HVT hastens immunocompetence of commercial meat-type chickens and optimized vaccination variables (dose and strain of HVT) to accelerate immunocompetence. A conventional HVT vaccine was given at recommended dose (RD), HVT-RD = 6080 plaque forming units (PFU), double-dose (2x), half-dose (1/2), or quarter-dose (1/4). Two rHVTs were given at RD: rHVT-A = 7380 PFU, rHVT-B = 8993 PFU. Most, if not all, treatments enhanced splenic lymphoproliferation with Concanavalin A and increased the percentage of granulocytes at day of age. Dose had an effect and HVT-RD was ideal. An increase of wing-web thickness after exposure to phytohemagglutinin-L was only detected after vaccination with HVT-RD. Furthermore, compared to sham-inoculated chickens, chickens in the HVT-RD had an increased percentage of CD3+ T cells and CD4+ T-helper cells, and increased expression of major histocompatibility complex (MHC)-II on most cell subsets (CD45+ cells, non-T leukocytes, T cells and the CD8+ and T cell receptor γδ T-cell subsets). Other treatments (HVT-1/2 and rHVT-B) share some of these features but differences were not as remarkable as in the HVT-RD group. Expression of MHC-I was reduced, compared to sham-inoculated chickens, in most of the cell phenotypes evaluated in the HVT-RD, HVT-2x and rHVT-A groups, while no effect was observed in other treatments. The effect of in ovo HVT on humoral immune responses (antibody responses to keyhole limpet hemocyanin and to a live infectious bronchitis/Newcastle disease vaccine) was minimal. Our study demonstrates in ovo vaccination with HVT in meat-type chickens can accelerate innate and adaptive immunity and we could optimize such effect by modifying the vaccine dose.}, number={31}, journal={VACCINE}, author={Boone, Allison C. and Kaser, Tobias and Cortes, Aneg L. and Kulkarni, Raveendra R. and Abad, Blanca A. Lopez de Juan and Villalobos, Tarsicio and Esandi, Javier and Perozo, Francisco and Lemiere, Stephane and Gimeno, Isabel M.}, year={2020}, month={Jun}, pages={4837–4845} }
 @misc{alkie_yitbarek_hodgins_kulkarni_taha-abdelaziz_sharif_2019, title={Development of innate immunity in chicken embryos and newly hatched chicks: a disease control perspective}, volume={48}, ISSN={["1465-3338"]}, DOI={10.1080/03079457.2019.1607966}, abstractNote={ABSTRACT Newly hatched chickens are confronted by a wide array of pathogenic microbes because their adaptive immune defences have limited capabilities to control these pathogens. In such circumstances, and within this age group, innate responses provide a degree of protection. Moreover, as the adaptive immune system is relatively naïve to foreign antigens, synergy with innate defences is critical. This review presents knowledge on the ontogeny of innate immunity in chickens pre-hatch and early post-hatch and provides insights into possible interventions to modulate innate responses early in the life of the bird. As in other vertebrate species, the chicken innate immune system which include cellular mediators, cytokine and chemokine repertoires and molecules involved in antigen detection, develop early in life. Comparison of innate immune systems in newly hatched chickens and mature birds has revealed differences in magnitude and quality, but responses in younger chickens can be boosted using innate immune system modulators. Functional expression of pattern recognition receptors and several defence molecules by innate immune system cells of embryos and newly hatched chicks suggests that innate responses can be modulated at this stage of development to combat pathogens. Improved understanding of innate immune system ontogeny and functionality in chickens is critical for the implementation of sound and safe interventions to provide long-term protection against pathogens. Next-generation tools for studying genetic and epigenetic regulation of genes, functional metagenomics and gene knockouts can be used in the future to explore and dissect the contributions of signalling pathways of innate immunity and to devise more efficacious disease control strategies.}, number={4}, journal={AVIAN PATHOLOGY}, author={Alkie, Tamiru N. and Yitbarek, Alexander and Hodgins, Douglas C. and Kulkarni, Raveendra R. and Taha-Abdelaziz, Khaled and Sharif, Shayan}, year={2019}, month={Jul}, pages={288–310} }
 @article{shojadoost_kulkarni_yitbarek_laursen_taha-abdelaziz_negash alkie_barjesteh_quinteiro-filho_smith_sharif_et al._2019, title={Dietary selenium supplementation enhances antiviral immunity in chickens challenged with low pathogenic avian influenza virus subtype H9N2}, volume={207}, ISSN={0165-2427}, url={http://dx.doi.org/10.1016/j.vetimm.2018.12.002}, DOI={10.1016/j.vetimm.2018.12.002}, abstractNote={Selenium supplementation in poultry feeds has been known to have beneficial effects on the bird health and performance; however antiviral effects of selenium have remained largely unknown. In this study, we have evaluated the effects of supplementation of chicken diets with organic (Selenium Enriched Yeast; SEY) and inorganic selenium (Sodium Selenite; SS) on low pathogenicity avian influenza virus (H9N2) shedding in the cloacal and oropharyngeal swab samples as well as examined the expression of immune related genes. Chickens were fed two doses (High- 0.30 mg/kg of feed; Low- 0.15 mg/kg of feed) of selenium supplementation for 2 weeks followed by low pathogenicity avian influenza virus challenge. Our results showed that the cloacal shedding of virus in all the selenium supplemented groups was significantly lower when compared to the non-supplemented control groups. In addition, the oropharyngeal shedding of virus in chickens fed with organic selenium supplementation was significantly lower than that in the chickens that received either inorganic selenium supplemented feed or controls. Furthermore, the expression of interferon stimulated genes (Viperin, OAS: 2′-5′ oligoadenylate synthetase and MDA5: melanoma differentiation-associated gene) in the cecal tonsils was significantly elevated in the selenium treated groups when compared to controls. Additionally, a significantly higher transcription of interferon (IFN)-α, IFN-β and IFN-γ genes in the cecal tonsils and spleens of chickens receiving SEY-L and SS-H supplemented feed was also observed at post virus challenge time points compared to untreated controls. The results of this study demonstrated that supplementation of chicken diets with selenium, can enhance antiviral defense and thus, may have a beneficial effect in controlling viral infections in poultry.}, journal={Veterinary Immunology and Immunopathology}, publisher={Elsevier BV}, author={Shojadoost, Bahram and Kulkarni, Raveendra R. and Yitbarek, Alexander and Laursen, Adrianna and Taha-Abdelaziz, Khaled and Negash Alkie, Tamiru and Barjesteh, Neda and Quinteiro-Filho, Wanderly M. and Smith, Trevor K. and Sharif, Shayan and et al.}, year={2019}, month={Jan}, pages={62–68} }
 @article{taha-abdelaziz_astill_kulkarni_read_najarian_farber_sharif_2019, title={In vitro assessment of immunomodulatory and anti-Campylobacter activities of probiotic lactobacilli}, volume={9}, ISSN={["2045-2322"]}, DOI={10.1038/s41598-019-54494-3}, abstractNote={Abstract}, journal={SCIENTIFIC REPORTS}, author={Taha-Abdelaziz, Khaled and Astill, Jake and Kulkarni, Raveendra R. and Read, Leah R. and Najarian, Afsaneh and Farber, Jeffrey M. and Sharif, Shayan}, year={2019}, month={Nov} }
 @article{barjesteh_taha-abdelaziz_kulkarni_sharif_2019, title={Innate antiviral responses are induced by TLR3 and TLR4 ligands in chicken tracheal epithelial cells: Communication between epithelial cells and macrophages}, volume={534}, ISSN={0042-6822}, url={http://dx.doi.org/10.1016/j.virol.2019.06.003}, DOI={10.1016/j.virol.2019.06.003}, abstractNote={The chicken upper respiratory tract is the portal of entry for respiratory pathogens including avian influenza virus (AIV). There is a paucity of information about the role of airway epithelial cells in the induction of antiviral responses in the chicken trachea. A better understanding of the role of these cells in the initiation of innate responses may improve prophylactic or therapeutic strategies for control of viral infections. The present study aimed to characterize antiviral innate responses in chicken tracheal epithelial cells (cTECs) induced by TLR ligands. The results demonstrated that stimulation of cTECs with TLR ligands induced antiviral responses, and subsequently reduced the replication of AIV in cTECs. Additionally, stimulated cTECs were able to influence the function of other cells such as macrophages. Overall, these results provided evidence that cTECs mount antiviral responses after stimulation with TLR ligands through IRF7 and NF-κB signaling pathways, leading to activation of other cells, such as macrophages.}, journal={Virology}, publisher={Elsevier BV}, author={Barjesteh, Neda and Taha-Abdelaziz, Khaled and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2019}, month={Aug}, pages={132–142} }
 @article{shojadoost_kulkarni_brisbin_quinteiro-filho_alkie_sharif_2019, title={Interactions between lactobacilli and chicken macrophages induce antiviral responses against avian influenza virus}, volume={125}, ISSN={0034-5288}, url={http://dx.doi.org/10.1016/j.rvsc.2017.10.007}, DOI={10.1016/j.rvsc.2017.10.007}, abstractNote={Macrophages are an important cell type of the innate immune system that upon activation, can exert antiviral functions and also can induce virus-specific adaptive immune responses. Macrophage interaction with certain probiotic bacteria such as lactobacilli can enhance antiviral functions of these cells. We have previously shown that administration of lactobacilli to chickens can effectively augment immune response to vaccine antigens. Here, we investigated the effects of representative strains of three Lactobacillus species, L. acidophilus, L. reuteri and L. salivarius used alone or in combination, in enhancing antiviral activity of chicken macrophages against avian influenza virus in an in vitro model using MQ-NCSU cells. Treatment of macrophages with probiotic lactobacilli significantly enhanced the antiviral functions, as determined by the virus titration assay. We also found that lactobacilli stimulation of macrophages induced significantly higher expression of interleukin (IL)-1β, interferon (IFN)- γ and IFN-α cytokine genes as well as interferon regulatory factor-7 (IRF7), 2′,5′-oligoadenylate synthetase (OAS) and interferon-inducible transmembrane protein M3 (IFITM3) genes. Furthermore, macrophages that were treated with lactobacilli had significantly enhanced production of nitric oxide (NO) and IFN-γ protein as well as surface expression of the costimulatory molecule CD40. However, the antiviral and immunostimulatory effects of probiotic lactobacilli largely depended on the Lactobacillus species studied. Collectively, the results from our study using an in vitro model showed that certain Lactobacillus species can effectively augment antiviral responses in chicken macrophages.}, journal={Research in Veterinary Science}, publisher={Elsevier BV}, author={Shojadoost, Bahram and Kulkarni, Raveendra R. and Brisbin, Jennifer T. and Quinteiro-Filho, Wanderley and Alkie, Tamiru N. and Sharif, Shayan}, year={2019}, month={Aug}, pages={441–450} }
 @article{laursen_kulkarni_taha-abdelaziz_plattner_read_sharif_2018, title={Characterizaton of gamma delta T cells in Marek’s disease virus (Gallid herpesvirus 2) infection of chickens}, volume={522}, ISSN={0042-6822}, url={http://dx.doi.org/10.1016/j.virol.2018.06.014}, DOI={10.1016/j.virol.2018.06.014}, abstractNote={Immunity against Marek's disease (MD), caused by Gallid herpesvirus 2 (GaHV-2), in chickens is mediated by both innate and adaptive responses. The present study evaluated the effects of GaHV-2 infection on distribution and frequency of γδ T cells in tissues, as well as their expression of cytokines. We found that the infected chickens had significantly higher number of γδ T cells in their spleens by 10 and 21 days post-infection (d.p.i.) and nearly 100% of these γδ T cells were CD8+ at 21 d.p.i. Conversely, the number of γδ T cells in the cecal tonsils of GaHV-2-infected birds decreased compared to uninfected birds. Splenic γδ T cells had up-regulated expression of interferon-γ early in infection followed by simultaneous gene expression of interleukin-10 during the later phases. In conclusion, these results suggest a potential role for γδ T cells in host response to GaHV-2 and further elucidate the underlying immunological mechanisms of interactions between this virus and its host.}, journal={Virology}, publisher={Elsevier BV}, author={Laursen, Adrianna M.S. and Kulkarni, Raveendra R. and Taha-Abdelaziz, Khaled and Plattner, Brandon L. and Read, Leah R. and Sharif, Shayan}, year={2018}, month={Sep}, pages={56–64} }
 @article{bhaumik_kulkarni_weldon_silveira_ahmed_gunisetty_chandele_antia_verma_sutter_et al._2018, title={Immune Priming and Long-term Persistence of Memory B Cells After Inactivated Poliovirus Vaccine in Macaque Models: Support for at least 2 Doses}, volume={67}, ISSN={1058-4838 1537-6591}, url={http://dx.doi.org/10.1093/cid/ciy634}, DOI={10.1093/cid/ciy634}, abstractNote={Abstract Background As a risk-mitigation strategy to minimize paralytic polio following withdrawal of Sabin type 2 from the oral poliovirus vaccine in April 2016, a single full dose or 2 fractional doses of inactivated poliovirus vaccine (IPV) are recommended. However, limited knowledge exists on long-term persistence of immune memory following 1- or 2-dose IPV schedules. Methods We examined induction and maintenance of immune memory following single- vs 2-dose IPV schedules, either full-dose intramuscular or fractional-dose intradermal, in rhesus macaques. Humoral responses, bone marrow–homing antibody-secreting plasma cells, and blood-circulating/lymph node–homing memory B cells were examined longitudinally. Results A single dose of IPV, either full or fractional, induced binding antibodies and memory B cells in all vaccinated macaques, despite failing to induce neutralizing antibodies (NT Abs) in many of them. However, these memory B cells declined rapidly, reaching below detection in the systemic circulation by 5 months; although a low frequency of memory B cells was detectable in draining lymph nodes of some, but not all, animals. By contrast, a 2-dose vaccination schedule, either full or fractional, efficiently induced NT Abs in all animals along with bone marrow–homing plasma cells and memory B cells. These memory B cells persisted in the systemic circulation for up to 16 months, the maximum duration tested after the second dose of vaccination. Conclusions Two doses of IPV, regardless of whether fractional or full, are more effective than a single dose for inducing long-lasting memory B cells.}, number={suppl_1}, journal={Clinical Infectious Diseases}, publisher={Oxford University Press (OUP)}, author={Bhaumik, Siddhartha Kumar and Kulkarni, Raveendra R and Weldon, William C and Silveira, Eduardo L V and Ahmed, Hasan and Gunisetty, Sivaram and Chandele, Anmol and Antia, Rustom and Verma, Harish and Sutter, Roland and et al.}, year={2018}, month={Oct}, pages={S66–S77} }
 @article{bavananthasivam_kulkarni_read_sharif_2018, title={Reduction of Marek's Disease Virus Infection by Toll-Like Receptor Ligands in Chicken Embryo Fibroblast Cells}, volume={31}, ISSN={0882-8245 1557-8976}, url={http://dx.doi.org/10.1089/vim.2017.0195}, DOI={10.1089/vim.2017.0195}, abstractNote={Evolutionarily conserved pattern recognition receptors, including Toll-like receptors (TLRs) recognize pathogen-associated molecular patterns (PAMPs) that are present in microbes. PAMPs induce several pathways downstream of TLRs that lead to induction of antiviral responses. The objective of this study was to investigate the stimulatory effect of various PAMPs (in the form of TLR ligands) in reducing Marek's disease virus (MDV) infection in chicken embryo fibroblast cells (CEFs). To this end, CEFs were pretreated with Pam3CSK4, Poly(IC), lipopolysaccharide (LPS), and CpG ODN as TLR2, TLR3, TLR4, and TLR21 ligands, respectively for 24 h followed by infection with MDV. The results indicated that pretreatment with Poly(IC) resulted in a robust reduction (by about 81%) of MDV infection in CEFs at 96 h postinfection while a moderate reduction was observed with treatment of Pam3CSK4 (35%), LPS (26%), and CpG ODN (23%) PAMPs. Transcriptional analysis of gene expression in CEFs demonstrated that all TLR ligand treatments and MDV infection significantly increased the expression of type I interferons, interleukin (IL)-1β, interferon regulatory factor 7 (IRF7), interferon induced protein with tetratricopeptide repeats 5 (IFIT5), and myxoma-resistance protein (Mx). Further studies are needed to explore the mechanism by which PAMPs, particularly the TLR3 ligands could reduce MDV infection in CEFs, which may play an important role in controlling the replication of MDV in chicken.}, number={5}, journal={Viral Immunology}, publisher={Mary Ann Liebert Inc}, author={Bavananthasivam, Jegarubee and Kulkarni, Raveendra R. and Read, Leah and Sharif, Shayan}, year={2018}, month={Jun}, pages={389–396} }
 @article{kulkarni_villanueva_read_brisbin_bhaumik_lamarre_murali-krishna_sharif_2017, title={CpG oligonucleotide-mediated co-stimulation of mouse invariant natural killer T cells negatively regulates their activation status}, volume={369}, ISSN={0302-766X 1432-0878}, url={http://dx.doi.org/10.1007/s00441-017-2631-y}, DOI={10.1007/s00441-017-2631-y}, abstractNote={Invariant natural killer T (iNKT) cells play important roles in antimicrobial defense and immune-regulation. We have previously shown that iNKT cells express certain toll-like receptors (TLR), and that TLR co-stimulation of iNKT cells in the presence of suboptimal concentrations of T cell receptor (TCR) agonists enhances cellular activation. In the present study, we investigated the regulatory effects of CpG oligonucleotides in mouse primary hepatic and splenic iNKT cells and in DN32.D3 iNKT cells. We show that CpG treatment of iNKT cells in the presence of higher concentrations of TCR agonists (α-GalCer or anti-CD3 mAb) results in the up-regulation of TLR9 in iNKT cells with a concurrent reduction in their cellular activation, as assessed by their production of IL-2, IL-4 and IFN-γ compared with controls. CpG-mediated down-regulation of iNKT cell activation has been found to depend, at least in part, on signaling by MyD88, a critical adapter moiety downstream of TLR9 signaling. Mechanistically, iNKT cells treated with CpG in the presence of TCR agonists show inhibition of MAPK signaling as determined by the levels of ERK1/2 and p38 MAPKs. Furthermore, CpG treatment leads to an increased induction of phosphatases, DUSP1 and SHP-1, that seem to impede MAPK and TCR signaling, resulting in the negative regulation of iNKT cell activation. Our findings therefore suggest a novel regulatory role for CpG in iNKT cells in the mediation of a negative feedback mechanism to control overactive iNKT cell responses and hence to avoid undesirable excessive immunopathology.}, number={3}, journal={Cell and Tissue Research}, publisher={Springer Nature}, author={Kulkarni, Raveendra R. and Villanueva, Alexander Ian and Read, Leah R. and Brisbin, Jennifer T. and Bhaumik, Siddhartha Kumar and LaMarre, Jonathan and Murali-Krishna, Kaja and Sharif, Shayan}, year={2017}, month={May}, pages={541–554} }
 @article{singh_alkie_nagy_kulkarni_hodgins_sharif_2016, title={Delivery of an inactivated avian influenza virus vaccine adjuvanted with poly(D,L-lactic-co-glycolic acid) encapsulated CpG ODN induces protective immune responses in chickens}, volume={34}, ISSN={0264-410X}, url={http://dx.doi.org/10.1016/j.vaccine.2016.08.009}, DOI={10.1016/j.vaccine.2016.08.009}, abstractNote={In poultry, systemic administration of commercial vaccines consisting of inactivated avian influenza virus (AIV) requires the simultaneous delivery of an adjuvant (water-in-oil emulsion). These vaccines are often limited in their ability to induce quantitatively better local (mucosal) antibody responses capable of curtailing virus shedding. Therefore, more efficacious adjuvants with the ability to provide enhanced immunogenicity and protective anti-AIV immunity in chickens are needed. While the Toll-like receptor (TLR) 21 agonist, CpG oligodeoxynucleotides (ODNs) has been recognized as a potential vaccine adjuvant in chickens, poly(D,L-lactic-co-glycolic acid) (PLGA) nanoparticles, successfully tested as vaccine delivery systems in other species, have not been extensively explored. The present study, therefore, assessed both systemic and mucosal antibody-mediated responses following intramuscular vaccination (administered at 7 and 21 days post-hatch) of chickens with PLGA encapsulated H9N2 AIV plus encapsulated CpG ODN 2007 (CpG 2007), and nonencapsulated AIV plus PLGA encapsulated CpG 2007 vaccine formulations. Virus challenge was performed at 2 weeks post-secondary vaccination using the oculo-nasal route. Our results showed that chickens vaccinated with the nonencapsulated AIV vaccine plus PLGA encapsulated CpG 2007 developed significantly higher systemic IgY and local (mucosal) IgY antibodies as well as haemagglutination inhibition antibody titres compared to PLGA encapsulated AIV plus encapsulated CpG 2007 vaccinated chickens. Furthermore, chickens that received CpG 2007 as an adjuvant in the vaccine formulation had antibodies exhibiting higher avidity indicating that the TLR21-mediated pathway may enhance antibody affinity maturation qualitatively. Collectively, our data indicate that vaccination of chickens with nonencapsulated AIV plus PLGA encapsulated CpG 2007 results in qualitatively and quantitatively augmented antibody responses leading to a reduction in virus shedding compared to the encapsulated AIV plus PLGA encapsulated CpG 2007 formulation.}, number={40}, journal={Vaccine}, publisher={Elsevier BV}, author={Singh, Shirene M. and Alkie, Tamiru N. and Nagy, Éva and Kulkarni, Raveendra R. and Hodgins, Douglas C. and Sharif, Shayan}, year={2016}, month={Sep}, pages={4807–4813} }
 @article{hu_yu_kulkarni_sharif_cui_xie_nie_gong_2015, title={Modulation of cytokine gene expression by selected Lactobacillus isolates in the ileum, caecal tonsils and spleen of Salmonella-challenged broilers}, volume={44}, ISSN={0307-9457 1465-3338}, url={http://dx.doi.org/10.1080/03079457.2015.1086725}, DOI={10.1080/03079457.2015.1086725}, abstractNote={Probiotics have been used to control Salmonella colonization in the chicken intestine. Recently, we demonstrated that certain selected Lactobacillus isolates were able to reduce Salmonella infection in the chicken spleen and liver as well as down-regulated Salmonella pathogenicity island 1 virulence gene expression in the chicken caecum. To further understand the mechanisms through which Lactobacillus protected chickens from Salmonella infection, the present study has investigated the Lactobacillus isolate(s)-induced host immune response of chickens to Salmonella enterica serovar Typhimurium infection. A thorough examination of cytokine gene expression in the ileum, caecal tonsils, and spleen on days 1 and 3 post-Salmonella infection showed a dynamic spatial and temporal response to Salmonella infection and Lactobacillus treatments. In most instances, it was evident that treatment of chickens with Lactobacillus isolates could significantly attenuate Salmonella-induced changes in the gene expression profile. These included the genes encoding pro-inflammatory cytokines [lipopolysaccharide-induced TNF factor, interleukin (IL)-6, and IL-8], T helper 1 cytokines [IL-12 and interferon (IFN)-γ], and T helper 2 cytokines (IL-4 and IL-10). Another important observation from the present investigation was that the response induced by a combination of Lactobacillus isolates was generally more effective than that induced by a single Lactobacillus isolate. Our results show that administration of certain selected Lactobacillus isolates can effectively modulate Salmonella-induced cytokine gene expression, and thus help reduce Salmonella infection in chickens.}, number={6}, journal={Avian Pathology}, publisher={Informa UK Limited}, author={Hu, Jie-Lun and Yu, Hai and Kulkarni, Raveendra R. and Sharif, Shayan and Cui, Steve W. and Xie, Ming-Yong and Nie, Shao-Ping and Gong, Joshua}, year={2015}, month={Nov}, pages={463–469} }
 @article{kulkarni_rasheed_bhaumik_ranjan_cao_davis_marisetti_thomas_gangappa_sambhara_et al._2014, title={Activation of the RIG-I Pathway during Influenza Vaccination Enhances the Germinal Center Reaction, Promotes T Follicular Helper Cell Induction, and Provides a Dose-Sparing Effect and Protective Immunity}, volume={88}, ISSN={0022-538X 1098-5514}, url={http://dx.doi.org/10.1128/jvi.02273-14}, DOI={10.1128/jvi.02273-14}, abstractNote={ABSTRACT}, number={24}, journal={Journal of Virology}, publisher={American Society for Microbiology}, author={Kulkarni, R. R. and Rasheed, M. A. U. and Bhaumik, S. K. and Ranjan, P. and Cao, W. and Davis, C. and Marisetti, K. and Thomas, S. and Gangappa, S. and Sambhara, S. and et al.}, year={2014}, month={Sep}, pages={13990–14001} }
 @article{villanueva_haeryfar_mallard_kulkarni_sharif_2014, title={Functions of invariant NK T cells are modulated by TLR ligands and IFN-α}, volume={21}, ISSN={1753-4259 1753-4267}, url={http://dx.doi.org/10.1177/1753425914527327}, DOI={10.1177/1753425914527327}, abstractNote={Invariant NK T (iNKT) cells perform numerous immunoregulatory functions. In mice, they express a unique and invariant Vα14-Jα18 rearrangement of α chain in their TCR recognizing glycolipid Ags presented by CD1d. This recognition results in the rapid release of both Th1- and Th2-type cytokines, making them early mediators of the immune response. Owing to their rapid activation and genetic rigidity of their TCR, iNKT cells share characteristics with innate lymphocytes. Therefore, we investigated whether iNKT cells could be induced to express TLRs, a class of pathogen recognition receptor. Mouse iNKT cells were stimulated with anti-CD3 monoclonal Ab and IFN-α, resulting in an increase in the transcription of TLRs 3, 5, 7 and 9, and increased surface expression of TLR3. These cells were subsequently stimulated with TLR ligands, resulting in an increase in the production of IFN-γ, IL-4 and TNF-α. Supernatants from these cells also increased macrophage production of IL-6 and prostaglandin E2, and increased their phagocytic activity and CD80 expression. These supernatants also reduced vesicular stomatitis virus-GFP replication in fibroblasts. This study demonstrates the role of IFN-α in iNKT cell activation, as well as the direct modulatory effects of TLR ligands on iNKT cell function, including antiviral activity.}, number={3}, journal={Innate Immunity}, publisher={SAGE Publications}, author={Villanueva, A Ian and Haeryfar, SM Mansour and Mallard, Bonnie A and Kulkarni, Raveendra R and Sharif, Shayan}, year={2014}, month={Mar}, pages={275–288} }
 @article{kulkarni_villanueva_elawadli_jayanth_read_mansour haeryfar_sharif_2012, title={Costimulatory activation of murine invariant natural killer T cells by toll-like receptor agonists}, volume={277}, ISSN={0008-8749}, url={http://dx.doi.org/10.1016/j.cellimm.2012.06.002}, DOI={10.1016/j.cellimm.2012.06.002}, abstractNote={Invariant NKT (iNKT) cells are glycolipid-reactive lymphocytes with anti-microbial properties. Toll-like receptor (TLR)-primed antigen-presenting cells are known to activate iNKT cells, however, the expression and function of TLRs in iNKT cells remain largely unknown. Here, we show that TCR-activation of murine iNKT cells by α-GalactosylCeramide (α-GalCer) or anti-CD3 antibodies can result in increased expression of TLR genes. TLR3, 5 and 9-mediated costimulation of TCR-preactivated iNKT cells resulted in enhancement of iNKT cell activation, as determined by their cytokine production. Expression of TLR3 and 9 at protein level was also confirmed in TCR-activated iNKT cells. Furthermore, TCR-preactivation followed by TLR9-costimulation of iNKT cells increased their ability to induce maturation of dendritic cells. Thus, our findings show that iNKT cells can up-regulate their TLR expression upon TCR activation and a subsequent TLR-signaling in these cells can lead to their enhanced activation, suggesting a new possible mode of iNKT cell activation.}, number={1-2}, journal={Cellular Immunology}, publisher={Elsevier BV}, author={Kulkarni, Raveendra R. and Villanueva, Alexander Ian and Elawadli, Inas and Jayanth, Preethi and Read, Leah R. and Mansour Haeryfar, S.M. and Sharif, Shayan}, year={2012}, month={May}, pages={33–43} }
 @article{mallick_kulkarni_st. paul_parvizi_nagy_behboudi_sharif_2012, title={Vaccination with CpG-Adjuvanted Avian Influenza Virosomes Promotes Antiviral Immune Responses and Reduces Virus Shedding in Chickens}, volume={4}, ISSN={0882-8245 1557-8976}, url={http://dx.doi.org/10.1089/vim.2011.0085}, DOI={10.1089/vim.2011.0085}, abstractNote={The use of virosomes as a vaccine platform has proven successful against several viruses. Here we examined the protective efficacy of a virosome-based vaccine consisting of avian influenza virus (AIV) A/Duck/Czech/56/H4N6 in chickens against a homologous AIV challenge. Virosomes adjuvanted with CpG-ODN or recombinant chicken interferon (IFN)-γ significantly reduced virus shedding after virus challenge. Furthermore, immunization with virosomes adjuvanted with CpG-ODN increased hemagglutination inhibition (HI) and virus-specific neutralizing serum antibodies, as well as virus-specific serum IgG and mucosal IgA responses. We also found a significant increase in the expression of type I and II interferon genes in the protected birds following virus challenge. In summary, this study demonstrated the ability of virosomes adjuvanted with CpG-ODN to reduce AIV shedding, and elicit virus-specific protective antibody responses in vaccinated birds.}, journal={Viral Immunology}, publisher={Mary Ann Liebert Inc}, author={Mallick, Amirul I. and Kulkarni, Raveendra R. and St. Paul, Michael and Parvizi, Payvand and Nagy, Éva and Behboudi, Shahriar and Sharif, Shayan}, year={2012}, month={Apr}, pages={120418071632002} }
 @article{mallick_haq_brisbin_mian_kulkarni_sharif_2011, title={Assessment of Bioactivity of a Recombinant Chicken Interferon-Gamma Expressed Using a Baculovirus Expression System}, volume={31}, ISSN={1079-9907 1557-7465}, url={http://dx.doi.org/10.1089/jir.2010.0130}, DOI={10.1089/jir.2010.0130}, abstractNote={The full-length coding sequence of chicken interferon-γ (ChIFN-γ) was cloned into a baculovirus nonfusion vector, pFastBacDual, and expressed in Sf21 insect cells. Recombinant ChIFN-γ (rChIFN-γ) protein was found to be expressed both intracellularly as well as in the culture supernatants. The affinity-purified rChIFN-γ contained 14, 17, and 28 kDa proteins, possibly representing both glycosylated and nonglycosylated protein forms of ChIFN-γ. The bioactivity of rChIFN-γ was confirmed in vitro by production of nitric oxide in a chicken macrophage cell line (HD11) and antiviral activity against vesicular stomatitis virus in primary chicken embryonic fibroblast cells. Further, HD11 cells stimulated with rChIFN-γ showed significant upregulation of inducible nitric oxide synthases, IFN-γ, interleukin-1β, interleukin-12p35, signal transducers and activators of transcription 1, class II, major histocompatibility complex, transactivator, and major histocompatibility complex II-β chain (BL-B) transcripts. In conclusion, the present study provides information on the ability of functionally active rChIFN-γ expressed in a baculovirus system in inducing significant transcriptional upregulation of various immune system-related genes, including those that encode cytokines, antigen-presenting molecules, and transcription factors involved in the major histocompatibility complex and IFN-signaling pathway.}, number={6}, journal={Journal of Interferon & Cytokine Research}, publisher={Mary Ann Liebert Inc}, author={Mallick, Amirul I. and Haq, Kamran and Brisbin, Jennifer T. and Mian, M. Firoz and Kulkarni, Raveendra R. and Sharif, Shayan}, year={2011}, month={Jun}, pages={493–500} }
 @article{villanueva_kulkarni_sharif_2011, title={Synthetic double-stranded RNA oligonucleotides are immunostimulatory for chicken spleen cells}, volume={35}, ISSN={0145-305X}, url={http://dx.doi.org/10.1016/j.dci.2010.08.001}, DOI={10.1016/j.dci.2010.08.001}, abstractNote={Toll-like receptors (TLRs) are an integral part of the innate immune system that recognize microbe-derived molecular patterns and initiate innate and adaptive defenses against invading pathogens. TLR3 and TLR7 are involved in sensing virus-associated single-stranded and double-stranded RNA (dsRNA) molecules in cellular endosomes to activate the type I interferon pathway in mammals. Although certain synthetic dsRNA molecules have been identified to show immunostimulation in mammalian cells, very little is known about the ability of these sequences to stimulate avian cells. The current study investigated immunostimulatory properties of four synthetic oligonucleotide sequences using chicken splenocytes. Expression of TLR3 and 7, type I interferons and several other cytokines as well as TLR signaling pathway-related genes at different time points post-stimulation was quantified by real-time PCR. A dose-dependent increase in expression of TLR3 was observed in splenocytes treated with poly-UGUGU (poly-UG) and β-galactosidase dsRNA molecules. TLR3 and TLR7 gene expression was significantly up-regulated upon stimulation with all four dsRNA molecules. Furthermore, in a time course study, a significant increase was noted in the expression of TLR3, TLR7, interferon (IFN)-α, IFN-β, interleukin (IL)-1β, IL-6 as well as 2′,5′-OAS in splenocytes treated with poly-UG. In conclusion, the present study demonstrated the immunostimulatory properties of dsRNA oligonucleotides, especially those that contain a poly-UG motif, in chickens.}, number={1}, journal={Developmental & Comparative Immunology}, publisher={Elsevier BV}, author={Villanueva, A.I. and Kulkarni, R.R. and Sharif, S.}, year={2011}, month={Jan}, pages={28–34} }
 @article{jiang_kulkarni_parreira_poppe_roland_prescott_2010, title={Assessment of 2 Salmonella enterica serovar Typhimurium-based vaccines against necrotic enteritis in reducing colonization of chickens by Salmonella serovars of different serogroups}, volume={74}, number={4}, journal={Canadian Journal of Veterinary Research}, author={Jiang, Y.-F. and Kulkarni, R.R. and Parreira, V.R. and Poppe, C. and Roland, K.L. and Prescott, J.F.}, year={2010}, month={Oct}, pages={264–270} }
 @article{kulkarni_behboudi_sharif_2010, title={Insights into the role of Toll-like receptors in modulation of T cell responses}, volume={343}, ISSN={0302-766X 1432-0878}, url={http://dx.doi.org/10.1007/s00441-010-1017-1}, DOI={10.1007/s00441-010-1017-1}, abstractNote={The innate immune receptors, such as Toll-like receptors (TLRs), are intimately involved in the early sensing of invading microorganisms or their structural components. Engagement of TLRs with their ligands results in activation of several downstream intracellular pathways leading to activation of innate and adaptive immune system cells. It was initially thought that TLRs are primarily expressed by antigen-presenting cells (APCs), such as macrophages and dendritic cells, and that interactions between microbial ligands and TLRs in these cells will indirectly result in activation of cells of the adaptive immune system, especially T cells. However, it has now become evident that TLRs are also expressed by various T cell subsets, such as conventional αβT cells, regulatory T cells, and γδT cells as well as natural killer T cells. Importantly, it appears that at least in some of these T cell subsets, TLRs are functionally active, because stimulation of these cells with TLR agonists in the absence of APCs results in exertion of effector or regulatory functions of T cells. The present review attempts to summarize the recent findings related to TLR expression in different T cell subsets and the direct role of TLRs in the induction and regulation of T cell responses, including those responses that occur at mucosal surfaces. In addition, the potential use of TLR agonists for steering T cell responses as a prophylactic or therapeutic strategy in the context of infectious, allergic or autoimmune diseases is explored.}, number={1}, journal={Cell and Tissue Research}, publisher={Springer Science and Business Media LLC}, author={Kulkarni, Raveendra and Behboudi, Shahriar and Sharif, Shayan}, year={2010}, month={Aug}, pages={141–152} }
 @article{kulkarni_haeryfar_sharif_2010, title={The invariant NKT cell subset in anti-viral defenses: a dark horse in anti-influenza immunity?}, volume={88}, ISSN={0741-5400}, url={http://dx.doi.org/10.1189/jlb.0410191}, DOI={10.1189/jlb.0410191}, abstractNote={Abstract}, number={4}, journal={Journal of Leukocyte Biology}, publisher={Wiley}, author={Kulkarni, R. R. and Haeryfar, S. M. and Sharif, S.}, year={2010}, month={Jun}, pages={635–643} }
 @article{kulkarni_parreira_jiang_prescott_2009, title={A Live Oral Recombinant Salmonella enterica Serovar Typhimurium Vaccine Expressing Clostridium perfringens Antigens Confers Protection against Necrotic Enteritis in Broiler Chickens}, volume={17}, ISSN={1556-6811}, url={http://dx.doi.org/10.1128/cvi.00406-09}, DOI={10.1128/cvi.00406-09}, abstractNote={ABSTRACT}, number={2}, journal={Clinical and Vaccine Immunology}, publisher={American Society for Microbiology}, author={Kulkarni, R. R. and Parreira, V. R. and Jiang, Y.- F. and Prescott, J. F.}, year={2009}, month={Dec}, pages={205–214} }
 @article{jiang_kulkarni_parreira_prescott_2009, title={Immunization of Broiler Chickens Against Clostridium perfringens–Induced Necrotic Enteritis Using Purified Recombinant Immunogenic Proteins}, volume={53}, ISSN={0005-2086 1938-4351}, url={http://dx.doi.org/10.1637/8656-021109-reg.1}, DOI={10.1637/8656-021109-reg.1}, abstractNote={Abstract This study identified and assessed secreted proteins of Clostridium perfringens additional to those previously described for their ability to protect broiler chickens against necrotic enteritis (NE). Secreted proteins of virulent and avirulent C. perfringens were electrophoretically separated and reacted with serum of chickens immune to NE. Three immunoreactive protein bands unique to the virulent C. perfringens were identified by mass spectrometry as the toxin C. perfringens large cytotoxin (TpeL), endo-beta-N-acetylglucosaminidase (Naglu), and phosphoglyceromutase (Pgm). The genes encoding Naglu and Pgm proteins were cloned, and their gene products were purified as histidine-tagged recombinant proteins from Escherichia coli and used in immunizing chickens. Immunized and nonimmunized control broiler chickens were then challenged with two different strains (CP1, CP4) of C. perfringens and assessed for the development of NE. Of the two immunogens, Pgm immunization showed significant protection of broiler chickens against experimental NE, although protection reduced as challenge severity increased. However, birds immunized with Naglu were protected from challenge only with strain CP4. Birds immunized with these proteins had antigen-specific antibodies when tested in an enzyme-linked immunosorbent assay. In conclusion, this study demonstrated the partial efficacy of additional secreted proteins in immunity of broiler chickens to NE. The study also showed that there may be differences in the protective ability of immunogens depending on the infecting C. perfringens strain.}, number={3}, journal={Avian Diseases}, publisher={American Association of Avian Pathologists (AAAP)}, author={Jiang, Yanfen and Kulkarni, Raveendra R. and Parreira, Valeria R. and Prescott, John F.}, year={2009}, month={Sep}, pages={409–415} }
 @article{chalmers_bruce_hunter_parreira_kulkarni_jiang_prescott_boerlin_2008, title={Multilocus Sequence Typing Analysis of Clostridium perfringens Isolates from Necrotic Enteritis Outbreaks in Broiler Chicken Populations}, volume={46}, ISSN={0095-1137}, url={http://dx.doi.org/10.1128/jcm.01548-08}, DOI={10.1128/jcm.01548-08}, abstractNote={ABSTRACT}, number={12}, journal={Journal of Clinical Microbiology}, publisher={American Society for Microbiology}, author={Chalmers, G. and Bruce, H. L. and Hunter, D. B. and Parreira, V. R. and Kulkarni, R. R. and Jiang, Y.-F. and Prescott, J. F. and Boerlin, P.}, year={2008}, month={Oct}, pages={3957–3964} }
 @article{kulkarni_parreira_sharif_prescott_2008, title={Oral immunization of broiler chickens against necrotic enteritis with an attenuated Salmonella vaccine vector expressing Clostridium perfringens antigens}, volume={26}, ISSN={0264-410X}, url={http://dx.doi.org/10.1016/j.vaccine.2008.05.079}, DOI={10.1016/j.vaccine.2008.05.079}, abstractNote={Necrotic enteritis (NE) in broiler chickens is caused by Clostridium perfringens but currently no effective vaccine is available. Our previous study showed that certain C. perfringens secreted proteins when administered intramuscularly protected chickens against experimental infection. In the current study, genes encoding three C. perfringens proteins: fructose-biphosphate-aldolase (FBA), pyruvate:ferredoxin-oxidoreductase (PFOR) and hypothetical protein (HP), were cloned into an avirulent Salmonella enterica sv. typhimurium vaccine vector. Broiler chickens immunized orally with recombinant Salmonella expressing FBA or HP proteins were significantly protected against NE challenge. Immunized birds developed serum and mucosal antibodies to both clostridial and Salmonella antigens. This study showed the oral immunizing ability of two C. perfringens antigens against NE in broiler chickens through an attenuated Salmonella vaccine vector.}, number={33}, journal={Vaccine}, publisher={Elsevier BV}, author={Kulkarni, R.R. and Parreira, V.R. and Sharif, S. and Prescott, J.F.}, year={2008}, month={Aug}, pages={4194–4203} }
 @article{kulkarni_parreira_sharif_prescott_2007, title={Immunization of Broiler Chickens against Clostridium perfringens-Induced Necrotic Enteritis}, volume={14}, ISSN={1556-6811}, url={http://dx.doi.org/10.1128/cvi.00162-07}, DOI={10.1128/cvi.00162-07}, abstractNote={ABSTRACT}, number={9}, journal={Clinical and Vaccine Immunology}, publisher={American Society for Microbiology}, author={Kulkarni, R. R. and Parreira, V. R. and Sharif, S. and Prescott, J. F.}, year={2007}, month={Jul}, pages={1070–1077} }
 @article{brisbin_haghighi_kulkarni_sharif_2007, title={Regulation of the intestinal immune system by commensal microbiota in avian and ruminant species}, volume={2}, number={8}, journal={CAB Reviews: Perspectives in Agriculture, Veterinary Science, Nutrition and Natural Resources}, author={Brisbin, J.T. and Haghighi, H. and Kulkarni, R.R. and Sharif, S.}, year={2007}, pages={11–19} }
 @article{kulkarni_parreira_sharif_prescott_2006, title={Clostridium perfringens Antigens Recognized by Broiler Chickens Immune to Necrotic Enteritis}, volume={13}, ISSN={1556-6811}, url={http://dx.doi.org/10.1128/cvi.00292-06}, DOI={10.1128/cvi.00292-06}, abstractNote={ABSTRACT}, number={12}, journal={Clinical and Vaccine Immunology}, publisher={American Society for Microbiology}, author={Kulkarni, R. R. and Parreira, V. R. and Sharif, S. and Prescott, J. F.}, year={2006}, month={Oct}, pages={1358–1362} }
 @article{thompson_parreira_kulkarni_prescott_2006, title={Live attenuated vaccine-based control of necrotic enteritis of broiler chickens}, volume={113}, ISSN={0378-1135}, url={http://dx.doi.org/10.1016/j.vetmic.2005.10.015}, DOI={10.1016/j.vetmic.2005.10.015}, abstractNote={A vaccine for necrotic enteritis (NE) of chickens would reduce the current need to prevent or treat the disease in broiler chickens with antimicrobial drugs. The objective of this study was to understand aspects of immunity to the disease. The first experiment examined the virulence of six strains of Clostridium perfringens isolated from cases of NE in broiler chickens. Using a 5-day experimental oral infection of 2-week-old broiler chickens, four of the six strains were found to be virulent. Pulsed-field gel electrophoresis and PCR showed that virulence was not associated with a plasmid encoding the beta2 toxin gene, cpb2, since this was present in virulent and one of the two avirulent strains. In the second experiment, two virulent and one avirulent strains were tested for their ability to immunize ("infection-immunization") chickens through the oral route. The procedure used experimental infection for 5 days followed by bacitracin treatment for 9 days, and then re-challenge 2 days later with a virulent strain, CP4. Infection-immunization with the virulent isolates protected chickens from subsequent virulent challenge, whereas the infection-immunization with the avirulent isolate did not. In a third experiment, two of four alpha-toxin-negative mutants of CP4 protected birds from experimental NE after oral immunization. These two mutants were also attenuated for virulence. We conclude that it is possible to immunize chickens successfully against NE and that immunogen(s) other than alpha-toxin are important in protective immunity against oral infection.}, number={1-2}, journal={Veterinary Microbiology}, publisher={Elsevier BV}, author={Thompson, D.R. and Parreira, V.R. and Kulkarni, R.R. and Prescott, J.F.}, year={2006}, month={Mar}, pages={25–34} }
 @article{rao_mishra_kulkarni_1999, title={Development of a precocious strain of Eimeria tenella: Preliminary observations}, volume={13}, number={2}, journal={Journal of Veterinary Parasitology}, author={Rao, J.R. and Mishra, A.K. and Kulkarni, R.R.}, year={1999}, pages={107–110} }
 @article{kulkarni_rao_omanwar_singh_1999, title={Eimeria tenella: Isolation, purification and protein profile of merozoite and oocysts wall proteins of a strain of Eimeria tenella selected for precociousness}, volume={13}, number={2}, journal={Journal of Veterinary Parasitology}, author={Kulkarni, R.R. and Rao, J.R. and Omanwar, S. and Singh, R.K.}, year={1999}, pages={119–123} }