Innate recognition of pathogens depends on the interaction between microbial structures known as pathogen-associated molecular patterns (PAMPs) and pattern recognition receptors (PRRs) in host cells. Toll-like receptors (TLR) are among the most important PRRs being expressed on and in a wide range of immune cell types. Studies on the interaction mechanisms between different pathogen species and the immune system of the dromedary camel are still scarce. The present study aimed to investigate the immunomodulatory effect of synthetic bacterial and viral TLR ligands on some phenotypic properties and selected functions of neutrophils purified from dromedary camel blood. Neutrophils were separated from camel blood (n = five animals) and were stimulated in vitro with the TLR ligands LPS, Pam3CSK4, R848 (Resiquimod), and Poly IC or were left without stimulation. Stimulation with the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) was used as a positive control stimulation. Shape change, phagocytosis activity, ROS production, the expression of cell surface markers, and cell vitality were compared between stimulated and non-stimulated cells. With exception of the TLR3 agonist Poly IC, all TLR ligands used showed the potential to stimulate camel neutrophils resulting in increased cell size and the upregulation of CD18 and CD14 on their surface. Similarly, the phagocytosis activity of camel neutrophils was significantly improved after priming with all TLR ligands, except Poly IC, which, in contrast, resulted in a reduced percentage of phagocytosis-positive cells. In contrast to stimulation with PMA, which induced a significant ROS production in camel neutrophils, none of the TLR ligands used stimulated ROS generation in neutrophils. Only stimulation with Pam3CSK4 increased the expression of MHCII molecules on camel neutrophils, resulting in an expanded MHCIIhigh fraction within camel neutrophils. Our study indicates selective immunomodulating effects of TLR agonists on purified camel neutrophils without affecting their vitality.

(1) Toll-like receptors (TLR) are a family of pattern recognition receptors that sense distinct molecular patterns of microbial origin. Although the immune cell composition of camel milk has been recently described, host–pathogen interaction studies in the camel mammary gland are still scarce. The present study aimed to use a whole milk stimulation assay for investigating the modulatory effect of selected Toll-like receptor (TLR) ligands on the phenotype and function of milk immune cells. (2) Methods—camel milk samples (n = 7) were stimulated in vitro with the TLR4 ligand LPS or the TLR2/1 ligand Pam3CSK4, and separated milk cells were evaluated for stimulation-induced shape change, the expression of cell surface markers, phagocytosis, apoptosis, ROS production, and NETosis. Stimulation with PMA was used as a control stimulation. (3) Results—all stimulants induced shape change in milk cells, change in the expression of several cell markers, and increased cell apoptosis and NETosis. In addition, stimulation with Pam3CSK4 and PMA was associated with enhanced ROS production, while only PMA stimulation resulted in enhanced bacterial phagocytosis by milk immune cells. (4) Conclusions—our data indicates selective modulating effects of the TLR ligands LPS and Pam3CSK4 on camel milk phagocytes. These results may have implications for the use of synthetic TLR agonists as immunomodulatory adjuvants of the immune response to intra-mammary vaccines against mastitis pathogens.

Background and Aim: Bovine viral diarrhea virus (BVDV) is one of the most important viral pathogens causing high economic losses in cattle of all ages. Despite the active vaccination campaigns against BVDV, many outbreaks are still detected in various populations of cattle worldwide. Other species of animals such as dromedary camels, sheep, and goats may harbor BVDV infection and cause variable clinical syndromes. Thus, they may act as a source of infection to the cattle population around them. However, little is still known about the roles of these animals in the viral transmission and sustainability of BVDV in the environment. This study aimed to explore if the dromedary camels, sheep, and goats may seroconvert against BVDV and to study some associated risk factors for BVDV in these species of animals. Materials and Methods: We tested 1012 serum samples from dromedary camels, 84 from goats, and 21 from sheep for BVDV antibodies using commercial enzyme-linked immunosorbent assay (ELISA) kits. Meanwhile, we selected 211 serum samples from dromedary camels to be tested for the BVDV antigen using the commercial ELISA kits. Results: Our results show that 49/1117 serum samples were positive for the BVDV antibodies in dromedary camels (46/1012), goats (3/84), and none of the tested sheep samples were positive. However, none of the collected serum samples tested positive for the BVDV antigen. Conclusion: Seroconversion of some dromedary camels, sheep, and goats to the BVDV with no history of vaccination against BVDV strongly suggests the potential roles of these species of animals in the virus transmission cycle. The main limitations of the current study are (1) the lack of samples from other species of animals that lived close by these animals, particularly cattle. (2) lack of follow-up samples from the same animal over a long period. We believe the long-term longitudinal study of BVDV in various species of animals, particularly dromedary camels, goats, and sheep, is one of our future research directions. This will provide more information about the dynamics of BVDV antibodies in these species of animals.

Background and Aim: Avian metapneumovirus (aMPV) is a recently discovered respiratory virus in chickens. Avian metapneumovirus has been linked to respiratory syndromes, reproductive failure in affected chickens and turkeys, swollen head syndrome in chickens, and rhinotracheitis in turkeys. Wild birds are considered potential reservoirs of aMPV, particularly aMPV-C. However, little is known about the prevalence of aMPV in Saudi Arabia. Considering the relevance of backyard chickens in the transmission and sustainability of certain avian viral diseases, this study aimed to assess aMPV exposure in backyard chickens and wild birds circulating near selected locations. Materials and Methods: We collected 368 serum samples from unvaccinated backyard chickens in ten locations in Eastern Saudi Arabia. Furthermore, we collected 78 serum samples from species of free-ranging birds belonging to the Columbidae family, such as pigeons and doves, captured from the same areas. Using commercial enzyme-linked immunosorbent assay kits, we tested the sera of domestic backyard chickens and wild birds for antibodies against aMPV. Results: Our results showed that 74/368 birds were positive for aMPV-related antibodies. Conversely, none of the tested wild birds seroconverted to aMPV. Conclusion: The antibody titers detected in the backyard chickens suggested recent exposure to aMPV. Considering these results, further large-scale serological and molecular studies are needed to evaluate the prevalence of aMPV in these birds and characterize the circulating strains of aMPV in this region. Keywords: antibodies, chickens, pigeons, Saudi Arabia.

Neutrophilic granulocytes are vital immune cells of the early response to pathogens. They contribute to the antimicrobial response through phagocytosis, production of reactive oxygen species, cytokine production, degranulation, and NET-formation. Neutrophil extracellular traps (NETs), also known as NETosis, are a critical antibacterial effector mechanism of cells of myeloid effector cells, including neutrophils and macrophages. Toll-like receptors (TLRs) are pattern recognition receptors (PRRs) that mediate pathogen sensing through the recognition of microbial structures known as pathogen-associated molecular patterns (PAMPs). The present study aimed to investigate the potential of several TLR ligands that mimic the sensing of bacterial and viral pathogens to stimulate NET-formation or Ca2+ influx in camel neutrophils. Neutrophils were purified from blood and were stimulated in vitro with ligands to TLR4, TLR2/1, TLR7/8, or TLR3. Net-formation was analyzed using the DNA-sensitive dye SYTOX™ Green and staining with antibodies to the neutrophil's granular enzyme myeloperoxidase. Real-time stimulation-induced Ca2+ influx was measured using the Ca2+-sensitive dye Flou-4 and flow cytometry. Only the TLR4-ligand lipopolysaccharide (LPS) could induce NET-formation in camel neutrophils, while none of the investigated TLR agonists showed a Ca2+ influx-inducing effect in camel neutrophils. The current study represents the first report on the impact of direct activation of TLR on NET-formation and Ca2+ influx in camel neutrophils with a selective effect of LPS on NET-formation induction. Future studies may investigate the molecular mechanisms behind the different responsiveness of bovine and camel neutrophils to TLR stimulation.