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Cytotoxic cells of the immune system may kill infected or transformed host cells via the perforin/granzyme or the Fas ligand (FasL) pathways. The purpose of this study was to determine mRNA expression of perforin, granzyme A and FasL in Eimeria tenella-infected tissues at primary infection and infection of immune chickens as an indirect measure of cytotoxic cell activity. Chickens were rendered immune by repeated E. tenella infections, which were manifested as an absence of clinical signs or pathological lesions and significantly reduced oocyst production upon challenge infection. During primary E. tenella infection, perforin, granzyme A and FasL mRNA expression in caecal tissue was significantly increased at 10 days after infection, compared to uninfected birds. In contrast, at infection of immune birds, perforin and granzyme A mRNA expression in caecal tissue was significantly increased during the early stages of E. tenella challenge infection, days 1-4, which coincided with a substantial reduction of parasite replication in these birds. These results indicate the activation of cytotoxic pathways in immune birds and support a role for cytotoxic T cells in the protection against Eimeria infections. [ABSTRACT FROM AUTHOR]

The purpose of this study was to monitor local cytokine responses to Dictyocaulus viviparus in calves during primary infection and re-infection. Bronchoalveolar lavage fluid (BALF) was collected weekly from experimentally infected calves and interleukin-2 (IL-2), IL-4, IL-5, IL-10 and IFN-γ m RNA expression was quantified in BALF cells. The major finding was a prominent transient increase in IL-4 m RNA expression, compared with that of uninfected calves, observed in BALF cells collected 2-3 weeks post-primary D. viviparus infection. At 2 weeks post-infection, macroscopic worms were also first observed in BALF. Calves re-infected after 10 weeks were partially immune which was evident at slaughter 5 weeks post-infection as a lower worm burden than in previously naïve calves infected at the same time. IL-4 m RNA expression in BALF cells 2 weeks post-re-infection was increased compared with that of uninfected animals but not as high as that of primarily infected calves. BALF cell expression of the other cytokines tested for was not as clearly effected by the D. viviparus infection. It seems likely that the strong IL-4 response observed during primary infection reflects an innate response to the worms that may initiate an ensuing Th2 response, which confers protective immunity. [ABSTRACT FROM AUTHOR]

Outbreaks of erysipelas, a disease caused by infection with Erysipelothrix rhusiopathiae (ER), is a re-emerging problem in cage-free laying hen flocks. The source of ER infection in hens is usually unknown and serological evidence has also indicated the presence of ER or other antigenically related bacteria in healthy flocks. The aim of the present study was to evaluate sample collection, culture methods and DNA-based methodology to detect ER and other Erysipelotrichales in samples from healthy chickens and their environment. We used samples from a research facility with conventionally reared chickens with no history of erysipelas outbreaks where hens with high titres of IgY recognising ER previously have been observed. Microbial DNA was extracted from samples either directly or after pre-culture in nonselective or ER-selective medium. Real-time PCR was used for detection of Erysipelothrix spp. and high-throughput amplicon sequencing of 16S rRNA sequencing was used for detection of Erysipelotrichales. A pilot serological analysis of some Erysipelotrichales members with IgY from unvaccinated and ER-vaccinated high-biosecurity chickens, as well as conventionally reared chickens, was also performed. All samples were negative for ER, E. tonsillarum and E. piscisicarius by PCR analysis. However, 16S rRNA community profiling indicated the presence of several Erysipelotrichales genera in both environmental samples and chicken intestinal samples, including Erysipelothrix spp. that were detected in environmental samples. Sequences from Erysipelothrix spp. were most frequently detected in samples pre-cultured in ER-selective medium. At species level the presence of Erysipelothrix anatis and/or Erysipelothrix aquatica was indicated. Serological results indicated that IgY raised to ER showed some cross-reactivity with E. anatis . Hence, environmental samples pre-cultured in selective medium and analysis by 16S rRNA sequencing proved a useful method for detection of Erysipelotrichales, including Erysipelothrix spp., in chicken flocks. The observation of such bacteria in environmental samples offers a possible explanation for the observation of high antibody titres to ER in flocks without a history of clinical erysipelas., Competing Interests: The authors declare there are no conflicts of interest., (Copyright © 2024 The Authors.)

Background: Single-cell transcriptomics provides means to study cell populations at the level of individual cells. In leukocyte biology this approach could potentially aid the identification of subpopulations and functions without the need to develop species-specific reagents. The present study aimed to evaluate single-cell RNA-seq as a tool for identification of chicken peripheral blood leukocytes. For this purpose, purified and thrombocyte depleted leukocytes from 4 clinically healthy hens were subjected to single-cell 3' RNA-seq. Bioinformatic analysis of data comprised unsupervised clustering of the cells, and annotation of clusters based on expression profiles. Immunofluorescence phenotyping of the cell preparations used was also performed., Results: Computational analysis identified 31 initial cell clusters and based on expression of defined marker genes 28 cluster were identified as comprising mainly B-cells, T-cells, monocytes, thrombocytes and red blood cells. Of the remaining clusters, two were putatively identified as basophils and eosinophils, and one as proliferating cells of mixed origin. In depth analysis on gene expression profiles within and between the initial cell clusters allowed further identification of cell identity and possible functions for some of them. For example, analysis of the group of monocyte clusters revealed subclusters comprising heterophils, as well as putative monocyte subtypes. Also, novel aspects of TCRγ/δ + T-cell subpopulations could be inferred such as evidence of at least two subtypes based on e.g., different expression of transcription factors MAF, SOX13 and GATA3. Moreover, a novel subpopulation of chicken peripheral B-cells with high SOX5 expression was identified. An overall good correlation between mRNA and cell surface phenotypic cell identification was shown., Conclusions: Taken together, we were able to identify and infer functional aspects of both previously well known as well as novel chicken leukocyte populations although some cell types. e.g., T-cell subtypes, proved more challenging to decipher. Although this methodology to some extent is limited by incomplete annotation of the chicken genome, it definitively has benefits in chicken immunology by expanding the options to distinguish identity and functions of immune cells also without access to species specific reagents., (© 2024. The Author(s).)

Introduction. Coccidiosis, caused by protozoan parasites of genus Eimeria , is a disease with large impact on poultry production worldwide. It is well known that Eimeria immunity is dependent on Th1-type responses. Gap Statement. In vitro assessment of Eimeria -specific T-cell activity would therefore be a valuable research tool but has so far proven difficult to establish. Aim. The present study aimed to evaluate in vitro induced blast transformation and CD25 expression in defined chicken T-cell populations as a measure of Eimeria immunity. Methodology. Three E. tenella infection experiments were performed and PBMC and/or spleen cells were collected between 6 and 16 days after infection of chickens. Cells were stimulated in vitro with E. tenella antigens and T-cell activation was assessed by immunofluorescence labelling and flow cytometry. Results. The results consistently showed statistically significant E. tenella specific activation of TCRα/β+T cells within a 'window' from 8 to 14 days after infection for both spleen cells and PBMC. Responding T-cells were identified as CD4+CD8-, CD4+CD8αα+ and CD4-CD8αβ+ where the CD4+CD8αα+ cells generally showed the highest responses. All three of these TCRα/βT-cell subsets showed significant E. tenella induced blast transformation and/or CD25 expression albeit not always in concert on the same days after infection indicating complex kinetics of T-cell responses. In general, responses were higher for spleen cells compared to PBMC for all responding T-cell populations. Conclusions. -specific T-cells, e.g. to evaluate vaccine responses. Results indicated that a Th1-type response was induced and suggested a role for CD4+CD8αα+ cells in Eimeria -specific T-cells, e.g. to evaluate vaccine responses. Results indicated that a Th1-type response was induced and suggested a role for CD4+CD8αα+ cells in Eimeria immunity.

Erysipelas, caused by infection with Erysipelothrix rhusiopathiae (ER) is an important emerging disease in laying hens. We have earlier observed prominent mannose-binding lectin (MBL) acute phase responses in experimentally ER infected chickens. The present study aimed to further examine immune responses to ER by using chickens selectively bred for high (L10H) and low (L10L) serum MBL levels. Chickens were infected with ER at 3 weeks of age and immune parameters and bacterial load were monitored in blood until day 18 after infection. Blood and spleen leukocytes collected on day 18 were stimulated in vitro with ER antigens and blast transformation of different T-cell populations was assessed. The ER infection gave a very varied outcome and no clear differences were observed between L10H and L10L chickens with respect to leukocyte counts, bacterial load or clinical outcome. Nonetheless, rapid innate responses, e.g., heterophilia and increased serum MBL levels were noted in bacteraemic chickens. All ER infected chickens also showed transient increased expression of mannose receptor MRC1L-B and decreased expression of major histocompatibility complex II on monocytes day 1 after infection indicating monocyte activation or relocation. In vitro ER stimulation showed antigen specific blast transformation of CD4+, TCRγ/δ-CD8αβ+ and TCRγ/δ+CD8αβ+ spleen cells from all infected chickens. For CD4+ and TCRγ/δ-CD8αβ+ cells the proportions of blast transformed cells were significantly higher for samples from L10L chickens than those for samples from L10H chickens. This is the first observation of ER-specific T-cells in chickens and interestingly a Th1-type response comprising cytotoxic T-cells was indicated., (© 2022. The Author(s).)

Hatching concepts such as on-farm hatching provide an opportunity to supply newly hatched chickens with optimal nutrition that support growth and development of a healthy gut. Brown algae contain bioactive compounds, especially laminarin and fucoidan that may improve intestinal health and immune responses. This study aimed to examine the effects of early access to feed and water posthatch and feed supplementation with algal extract rich in laminarin from Laminaria digitata, on growth performance, organ and microbiota development and antibody production. A total of 432 Ross 308 chicks were allotted to 36 rearing pens in a 2 × 3 factorial design with two hatching treatments and three dietary treatments. During chick placement, half of the pens were directly provided access to feed and water (Early) while half of the pens were deprived of feed and water for 38 h (Late). The chicks were fed three different starter diets until day 6; a wheat-soybean meal-based control diet, a diet with low inclusion of algal extract (0.057%) and a diet with high inclusion of algal extract (0.114%). Feed intake and BW were registered on pen basis at placement, days 1, 6, 12, 19, 26, 33 and 40. To induce antibody responses, all chicks were vaccinated against avian pneumovirus on day 10. Three chicks per pen were selected as focal animals and used for blood sampling on days 10 and 39. On days 6, 19, and 40, two birds per pen were killed and used for organ measurement and caecal digesta sampling for gut microbiota analysis using the Illumina Miseq PE 250 sequencing platform. Results showed that algal extract did not influence gut microbiota, gut development or vaccine-induced antibody responses. However, during the first 38 h, early-fed chicks consumed on average 19.6 g of feed and gained 27% in BW, while late-fed chicks lost 9.1% in BW which lowered BW and feed intake throughout the study (P < 0.05). Late chicks also had longer relative intestine, higher relative (g/kg BW) weight of gizzard and proventriculus but lower relative bursa weight on day 6 (P < 0.05). No effects of hatching treatment on microbiota or antibody response were detected. The microbiota was affected by age, where alpha diversity increased with age. In conclusion, this study showed that early access to feed but not algal extract improved the growth performance throughout the 40-day growing period, and stimulated early bursa development., Competing Interests: Declaration of interest None., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

A comprehensive analysis of T cell activation markers in chicken is lacking. Kinetics of T cell activation markers (CD25, CD28, CD5, MHC-II, CD44, and CD45) in response to in vitro stimulation of peripheral blood mononuclear cells with concanavalin A (Con A) were evaluated between two chicken lines selected for high and low levels of mannose-binding lectin in serum (L10H and L10L, respectively) by flow cytometry. L10H chickens showed a stronger response to Con A based on the frequency of T cell blasts in both the CD4 + and CD8 + compartment. The majority of the proliferating CD4 + and CD8 + T cells expressed CD25. Proliferating T cells were seen both in the CD4 +  MHC-II +/- and CD8 +  MHC-II +/- population. For both CD4 + and CD8 + T cells, frequencies of CD25 + and MHC-II + T cells were increased 24 h after stimulation. CD28 + frequencies were only increased on CD8 + T cells 48 h after stimulation. An increase in the relative surface expression based on mean fluorescence intensity (MFI) upon activation was observed for most markers except CD5. For CD4 + T cells, CD28 expression increased 24 h after stimulation whereas MHC-II expression increased after 48 h. For CD8 + T cells, a tendency toward an increase in CD25 expression was observed. CD28 expression started to increase 24 h after stimulation and only a transient peak in MHC-II expression on CD8 + T cells was observed after 24 h. CD44 and CD45 expressed on CD4 + and CD8 + T cells increased 24-72 h after stimulation. In summary, the frequency of CD25 + and MHC-II + T cells were shown to be early markers (24 h) for in vitro activation of both CD4 + and CD8 + T cells. Frequency of CD28 + T cells was a later marker (48 h) and only for CD8 + T cells. Surface expression of all markers (MFI) increased permanently or transiently upon activation except for CD5., (© 2021 International Society for Advancement of Cytometry.)

Background: Coccidiosis is an infectious disease with large negative impact on the poultry industry worldwide. It is an enteric infection caused by unicellular Apicomplexan parasites of the genus Eimeria. The present study aimed to gain more knowledge about interactions between parasites and the host immune system during the early asexual replication phase of E. tenella in chicken caeca. For this purpose, chickens were experimentally infected with E. tenella oocysts, sacrificed on days 1-4 and 10 after infection and mRNA from caecal tissues was extracted and sequenced., Results: Dual RNA-seq analysis revealed time-dependent changes in both host and parasite gene expression during the course of the infection. Chicken immune activation was detected from day 3 and onwards with the highest number of differentially expressed immune genes recorded on day 10. Among early (days 3-4) responses up-regulation of genes for matrix metalloproteinases, several chemokines, interferon (IFN)-γ along with IFN-stimulated genes GBP, IRF1 and RSAD2 were noted. Increased expression of genes with immune suppressive/regulatory effects, e.g. IL10, SOCS1, SOCS3, was also observed among early responses. For E. tenella a general up-regulation of genes involved in protein expression and energy metabolism as well as a general down-regulation genes for DNA and RNA processing were observed during the infection. Specific E. tenella genes with altered expression during the experiment include those for proteins in rhoptry and microneme organelles., Conclusions: The present study provides novel information on both the transcriptional activity of E. tenella during schizogony in ceacal tissue and of the local host responses to parasite invasion during this phase of infection. Results indicate a role for IFN-γ and IFN-stimulated genes in the innate defence against Eimeria replication., (© 2021. The Author(s).)

The study aimed to monitor parasite and host gene expression during the early stages of Eimeria tenella infection of chicken cells using dual RNA-Seq analysis. For this, we used chicken macrophage-like cell line HD11 cultures infected in vitro with purified E. tenella sporozoites. Cultures were harvested between 2 and 72 h post-infection and mRNA was extracted and sequenced. Dual RNA-Seq analysis showed clear patterns of altered expression for both parasite and host genes during infection. For example, genes in the chicken immune system showed upregulation early (2–4 h), a strong downregulation of genes across the immune system at 24 h and a repetition of early patterns at 72 h, indicating that invasion by a second generation of parasites was occurring. The observed downregulation may be due to immune self-regulation or to immune evasive mechanisms exerted by E. tenella. Results also suggested pathogen recognition receptors involved in E. tenella innate recognition, MRC2, TLR15 and NLRC5 and showed distinct chemokine and cytokine induction patterns. Moreover, the expression of several functional categories of Eimeria genes, such as rhoptry kinase genes and microneme genes, were also examined, showing distinctive differences which were expressed in sporozoites and merozoites.

Background: Erysipelas, caused by Erysipelothrix rhusiopathiae (ER), is an important emerging disease in free-range and organic egg-production. The aim of the present study was to assess if quantification of ER specific IgY titers may aid the understanding of erysipelas in commercial laying hens. The methodology was validated with sequentially collected sera from experimentally ER infected SPF-chickens and subsequently applied on sera from Swedish commercial laying hens collected during and after outbreaks of erysipelas or collected at slaughter from healthy hens housed in furnished cages, barn production or in organic production (with outdoor access)., Results: In experimentally infected SPF-chickens, titers to ER were significantly increased approximately one week after infection while IgY to ER in uninfected age-matched controls remained low. Also chickens infected with low doses of ER, not displaying clinical signs of disease and with low recovery of ER in blood samples showed high titers of IgY to ER. For laying hens during and after erysipelas outbreaks the majority of samples were considered positive for antibodies to ER with a large variation in levels of IgY titers to ER between individuals. For healthy laying hens at slaughter all samples were deemed positive for antibodies to ER. An influence of flock on levels of IgY titers to ER was observed for both healthy hens and hens during erysipelas outbreaks. For healthy laying hens at slaughter no influence of the housing systems included in the study, history of erysipelas outbreaks at the farm or vaccination on levels of IgY titers to ER was noticed., Conclusions: Taken together, these results show that high numbers of commercial laying hens showed high IgY titers to ER, comparable to those elicited by experimental ER infection, indicating that ER or bacteria that raises antibodies that cross-react with ER are common in this environment.

Erysipelas, a disease caused by Erysipelothrix rhusiopathiae (ER), is an increasing problem in laying hens housed in cage-free systems. This study aimed to monitor immune responses during ER infection of naïve chickens and chickens vaccinated intra muscularly with a commercial inactivated ER vaccine. Chickens were infected intra muscularly with ER at 30 days of age and blood leukocyte counts, serum levels of mannose binding lectin (MBL) and ER-specific IgY were monitored until the experiment was terminated at day 15 after infection. ER was detected in blood from more chickens and at higher bacterial counts in the naïve group (day 1: 1 of 7 chickens; day 3: 6 of 6 chickens) than in the vaccinated group (day 1: 0 of 7 chickens; day 3: 1 of 6 chickens). During the acute phase of infection transient increases in circulating heterophil numbers and serum MBL levels were detected in all ER infected chickens but these responses were prolonged in chickens from the naïve group compared to vaccinated chickens. Before infection IgY titers to ER in vaccinated chickens did not differ significantly from those of naïve chickens but vaccinated chickens showed significantly increased IgY titers to ER earlier after infection compared to chickens in the naïve group. In conclusion, the ER infection elicited prompt acute innate responses in all chickens. Vaccinated chickens did not have high IgY titers to ER prior to infection but did however show lower levels of bacteraemia and their acute immune responses were of shorter duration.

The disease erysipelas caused by Erysipelothrix rhusiopathiae (ER) is a major concern in pig production. In the present study the genomes of ER from pigs ( n =87), wild boars ( n =71) and other sources ( n =85) were compared in terms of whole-genome SNP variation, accessory genome content and the presence of genetic antibiotic resistance determinants. The aim was to investigate if genetic features among ER were associated with isolate origin in order to better estimate the risk of transmission of porcine-adapted strains from wild boars to free-range pigs and to increase our understanding of the evolution of ER. Pigs and wild boars carried isolates representing all ER clades, but clade one only occurred in healthy wild boars and healthy pigs. Several accessory genes or gene variants were found to be significantly associated with the pig and wild boar hosts, with genes predicted to encode cell wall-associated or extracellular proteins overrepresented. Gene variants associated with serovar determination and capsule production in serovars known to be pathogenic for pigs were found to be significantly associated with pigs as hosts. In total, 30 % of investigated pig isolates but only 6 % of wild boar isolates carried resistance genes, most commonly tetM (tetracycline) and lsa (E) together with lnu (B) (lincosamides, pleuromutilin and streptogramin A). The incidence of variably present genes including resistance determinants was weakly linked to phylogeny, indicating that host adaptation in ER has evolved multiple times in diverse lineages mediated by recombination and the acquisition of mobile genetic elements. The presented results support the occurrence of host-adapted ER strains, but they do not indicate frequent transmission between wild boars and domestic pigs. This article contains data hosted by Microreact.

Purpose: The present study aimed to establish pretreatment protocols as well as real-time and droplet digital polymerase chain reaction (PCR) methodologies to detect and quantify Erysipelothrix rhusiopathiae (ER) DNA in blood samples from infected chickens, as tools for routine diagnostics and monitoring of experimental infections. Chicken blood is a problematic matrix for PCR analysis because nucleated erythrocytes contribute large amounts of host DNA that inhibit amplification., Methodology: Using artificially spiked samples of fresh chicken blood, as well as blood samples from three experimental infection studies, the performance of pretreatment protocols, including choice of blood stabilization agent, centrifugation speeds and Ficoll gradient separation, was evaluated. The results were compared with those from traditional culture-based protocols combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS).Results/Key findings. Simple preparations producing cell-free samples performed well on artificial spike-in samples, providing high sensitivity. However, performance was poor in clinical samples or artificial samples where the bacteria were incubated for 4 h or more in fresh blood prior to DNA extraction. In these samples, a Ficoll separation protocol that creates samples rich in lymphocytes, monocytes and thrombocytes prior to DNA extraction was far more effective., Conclusions: Our results indicate that ER bacteria undergo rapid phagocytosis in chicken blood and that analysis of a blood fraction enriched for phagocytic cells is necessary for reliable detection and quantification. The presented results explain the poor performance of PCR detection reported in previously published experimental ER infection studies, and the proposed solutions are likely to have broader implications for PCR-based veterinary diagnostics in non-mammalian host species such as poultry and fish.

This study aimed to set up methodology to monitor parasite-specific T-cell activation in vitro using Eimeria tenella-infected chickens. A sonicated E. tenella sporozoite protein preparation was used for the activation of chicken spleen cell cultures. Proliferation assessed by 3H-thymidin incorporation or blast transformation of T-cells assessed by immunofluorescence labelling and flow cytometry were used as read-outs for activation. Results showed that E. tenella-specific proliferation was detected in cultures of spleen cells collected in a 'window' between 8 and 14 days after primary infection. However, due to high variation in proliferative responses between individuals and to high background proliferation, large numbers of observations were needed to obtain significant results. Moreover, the outcome was not improved by increasing the infection dose to chickens or by depletion of T-cell receptor (TCR) γ/δ expressing cells from cultures. An E. tenella-specific blast transformation response was observed for TCRα/β expressing cells within the same 'window', confirming the identity of the responding cells as classic T-cells. Thus, it is possible to study the kinetics of E. tenella-specific T-cell responses in vitro. However, more in-depth phenotypic identification of the responding T-cells could improve the methodology.

Phagocytic activity of leukocytes in whole blood was assessed as a potential immune competence trait in chickens. A flow cytometry based whole blood phagocytosis (WBP) assay was set up and evaluated using blood from chickens homozygous for four different MHC haplotypes, B12, B15, B19 and B21. Fluorescent latex beads and two serotypes of fluorescently labelled heat-killed bacteria (Salmonella Infantis and Salmonella. Typhimurium) were evaluated as phagocytic targets. In addition, the opsonophagocytic potential (OPp) of individual sera from the birds was included in a phagocytosis assay using the HD11 chicken macrophage cell line. Results showed that both serotypes of bacteria but not the latex beads were effectively phagocytosed by leukocytes in the whole blood cultures. Differences were observed in the phagocytic capacity of monocytes and thrombocyte/lymphocytes, respectively between the different MHC lines. No significant differences on the OPp of serum was identified between MHC lines. In addition, for both phagocytic activity of leukocytes and OPp of serum large variations between individuals were observed within MHC haplotypes. No significant relationships were observed between the phagocytic activity of leukocytes and serum OPp or Salmonella-specific IgY levels. In conclusion, our results suggest that the WBP assay, using a no-lyse no-wash single staining method, is a rapid and convenient method to assess phagocytic functions of different leukocyte populations., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Opsonins, an important arm of the innate immune system, are various soluble proteins, which play a critical role in destruction of invading pathogens directly or via engulfment of pathogens through the intermediate of phagocytosis. The diversity of opsonin profiles is under genetic influence and may be associated with variation in disease resistance. The aim of this study was to set up an assay to determine serum opsonophagocytic potential (OPp) for chicken sera by flow cytometry and to evaluate the assay using samples from different chicken lines. Two chicken lines selected for high and low concentrations of mannose-binding lectin, a known opsonin, in serum were used to establish the method. Furthermore, the presumed "robust" Hellevad chickens and two other commercial chicken lines (Hisex and Bovans) were tested to evaluate OPp as a parameter reflecting general immune competence. The results showed that Hellevad and Bovans chickens had higher OPp than Hisex chickens. There were no correlations between concentrations of total IgY or mannose-binding lectin and OPp. However, a strong positive correlation was observed between vaccine-induced infectious bronchitis virus titres and OPp. Moreover, inverse relationships were observed between concentrations of total serum IgM as well as natural antibody levels, and OPp. In conclusion, in vitro opsonophagocytosis assessment and determination of OPp may be of relevance when addressing general innate immunocompetence. RESEARCH HIGHLIGHTS A flow cytometry method was developed to assess poultry serum opsonophagocytosis potential. This method is based on serum-opsonin-coated polystyrene beads and HD11 cell phagocytosis. Serum samples from different commercial chicken lines were compared. Opsonophagocytic potential may be included in assay panels for general immune competence of poultry.