Assessing the impact of colostrum feeding delay on serum immunoglobulin G and total protein in dairy goat kids.

This experiment was motivated by the need to understand the impacts of delaying the first colostrum feeding on the prevalence of failed transfer of passive immunity (FTPI). A cohort of 216 kids was stratified into groups based on the colostrum feeding delay postbirth: 0-4 h, 4-8 h, 8-12 h, and 12-16 h. All kids received a single colostrum meal of 300 mL, and blood samples were collected approximately 36 h after feeding. Serum immunoglobulin G (SIgG) was measured using ELISA, and serum total protein (STP) was assessed using the Bradford method and refractometry (STPb and STPr). Statistical methods like Pearson correlations, Bland-Altman plots, and Lin's concordance coefficient were employed to assess associations and agreements between SIgG, STPb and STPr. Receiver operator characteristic analysis was employed to determine optimal STPb and STPr thresholds for predicting FTPI (SIgG < 12 g/L). Subsequently, areas under the curve, sensitivity, and specificity were examined to assess the accuracy of these thresholds. Our results showed that for each hour's delay from birth to colostrum intake (up to 16 h), IgG apparent efficiency of absorption (AEA) decreases at an approximate rate of 2.0% per hour, and SIgG decreases at an approximate rate of 1.0 g/L per hour. However, this decline is not constant over time but intensifies progressively with increased feeding delay. Specifically, reductions in IgG AEA were 1.3, 2.9, and 5.9% per hour, and decreases in SIgG were 0.2, 0.3, and 0.7 g per hour for SIgG across the time intervals of 0-4 to 4-8 h, 4-8 to 8-12 h, and 8-12 to 12-16 h, respectively. Additionally, there was an increase in SIgG of 1.2 g/dL but a decrease in IgG AEA of 1.9% for each gram per kg of BW increase in IgG intake. The correlations between SIgG and STPr and STPb were 0.62, and 0.36, respectively. Optimal STPr and STPb thresholds predicting FTPI were determined to be 4.6 and 6.2 g/dL. The prevalence of FTPI, according to SIgG, STPr, and STPb thresholds were 63, 62, and 45%. Overall, STPr showed higher values for key performance metrics (i.e., sensitivity, likelihood ratio of positive tests, overall accuracy, and Youden's index), indicating better prediction ability than STPb. Our findings corroborate the critical importance of swift colostrum administration, ideally occurring no later than 12 h postbirth. Moreover, our research validates the effectiveness of Brix refractometry as a practical, on-farm method for assessing FTPI in goat kids.

Facilitated introgression from domestic goat into Alpine ibex at immune loci.

Hybridization can result in the transfer of adaptive genetic material from one species to another, known as adaptive introgression. Bottlenecked (and hence genetically depleted) species are expected to be particularly receptive to adaptive introgression, since introgression can introduce new or previously lost adaptive genetic variation. The Alpine ibex (Capra ibex), which recently recovered from near extinction, is known to hybridize with the domestic goat (Capra aegagrus hircus), and signals of introgression previously found at the major histocompatibility complex were suggested to potentially be adaptive. Here, we combine two ancient whole genomes of Alpine ibex with 29 modern Alpine ibex genomes and 31 genomes representing six related Capra species to investigate the genome-wide patterns of introgression and confirm the potential relevance of immune loci. We identified low rates of admixture in modern Alpine ibex through various F statistics and screening for putative introgressed tracts. Further results based on demographic modelling were consistent with introgression to have occurred during the last 300 years, coinciding with the known species bottleneck, and that in each generation, 1-2 out of 100 Alpine ibex had a domestic goat parent. The putatively introgressed haplotypes were enriched at immune-related genes, where the adaptive value of alternative alleles may give individuals with otherwise depleted genetic diversity a selective advantage. While interbreeding with domestic species is a prevalent issue in species conservation, in this specific case, it resulted in putative adaptive introgression. Our findings highlight the complex interplay between hybridization, adaptive evolution, and the potential risks and benefits associated with anthropogenic influences on wild species.

Differences in proteomic profiles and immunomodulatory activity of goat and cow milk fat globule membrane.

This study aimed to clarify the composition and bioactivity differences between goat and cow milk fat globule membrane (MFGM) protein by proteomic, and the immunomodulatory activity of MFGM proteins was further evaluated by using mouse splenic lymphocytes in vitro. A total of 257 MFGM proteins showed significant differences between goat and cow milk. The upregulated and unique MFGM proteins in goat milk were significantly enriched in the positive regulation of immune response, negative regulation of Interleukin-5 (IL-5) secretion, and involved in nucleotide-binding oligomerization domain (NOD)-like receptor signaling. The contents of IL-2 and Interferon-γ in the supernatant of spleen lymphocytes treated with goat MFGM proteins were much higher than those of IL-4 and IL-5, suggesting a Th1-skewed immune response. These results revealed that goat MFGM proteins could possess better immunomodulatory effects as compared to cow milk. Our findings may provide new insights to elucidate the physiological functions and nutritional of goat milk.

Short communication: Goat mastitis and the formation of neutrophil extracellular traps (NETs).

Mastitis, an inflammation of the mammary gland affecting milk production and quality in dairy herds, is often associated with Staphylococcus spp. in goats. Neutrophils are crucial in combating infections by migrating into milk and deploying various defense strategies, including the release of neutrophil extracellular traps (NETs) composed of DNA, histones, and bactericidal proteins. This study investigated whether NETs are released by goat neutrophils stimulated in vitro by Staphylococcus aureus and Staphylococcus warneri, two common pathogens of goat mastitis. PMNs were isolated from blood from healthy adult goats. We evaluated goat NET formation by stimulating cells with: phorbol 12-myristate 13-acetate (PMA) as a positive control, cytochalasin for inhibition of actin polymerization, S. aureus, and S. warneri. NET formation was observed in response to chemical stimulation and bacterial presence, effectively trapping pathogens. Variations in NET formation between S. aureus and S. warneri suggest pathogen-specific responses. These findings suggest that the formation of NETs may be an important complementary mechanism in the defense against mastitis in goats. In conclusion, this study unveils a novel defense mechanism in goats, indicating the role of NETs against S. aureus and S. warneri in mastitis.

Altering the competitive environment of B cell epitopes significantly extends the duration of antibody production.

Persistent immunoglobulin G (IgG) production (PIP) provides long-term vaccine protection. While variations in the duration of protection have been observed with vaccines prepared from different pathogens, little is known about the factors that determine PIP. Here, we investigated the impact of three parameters on the duration of anti-peptide IgG production, namely amino acid sequences, protein carriers, and immunization programs. We show that anti-peptide IgG production can be transformed from transient IgG production (TIP) to PIP, by placing short peptides (Pi) containing linear B cell epitopes in different competitive environments using bovine serum albumin (BSA) conjugates instead of the original viral particles. When goats were immunized with the peste des petits ruminants (PPR) live-attenuated vaccine (containing Pi as the constitutive component) and BSA-Pi conjugate, anti-Pi IgG production exhibited TIP (duration < 60 days) and PIP (duration > 368 days), respectively. Further, this PIP was unaffected by subsequent immunization with the PPR live-attenuated vaccine in the same goat. When goats were coimmunized with PPR live-attenuated vaccine and BSA-Pi, the induced anti-Pi IgG production showed a slightly extended TIP (from ~60 days to ~100 days). This discovery provides new perspectives for studying the fate of plasma cells in humoral immune responses and developing peptide vaccines related to linear neutralizing epitopes from various viruses.

Impact of cow, buffalo, goat or camel milk consumption on oxidative stress, inflammation and immune response post weaning time.

Milk is a whitish liquid that is secreted from mammary glands; and considered as the primary source of nutrition for newborns since they are not able to digest solid food. However, it contains primary nutrients, as well as growth and immune factors. Early weaning is a critical issue that face women and their babies in developing countries. To avoid infant malnutrition, they tend to use other milk types instead of baby formula. Therefore, the present study aimed to evaluate the impact of cow, buffalo, goat or camel milk consumption on oxidative stress, inflammation and immune response in male and female Sprague Dawley rats post weaning time. The amino acids, fatty acids, minerals and vitamins in the tested milk types were evaluated. Animals were divided into 5 groups (control, cow, buffalo, goat and camel milk administrated groups) (10 rats/group); each animal was administrated by 3.4 ml/day. Rats were administered with milk for 6 weeks; at the end of the 5th week, five animals of each group were isolated and the remaining five animals were immunized with sheep red blood cells (SRBCs) and kept for another week to mount immune response. The effect of different milk types on rats' immune response towards SRBCs was evaluated through pro-inflammatory cytokines, antioxidants, ESR and CRP measurement; together, with the histopathological examination of spleen samples and hemagglutination assay. Camel milk consumption reduced oxidative stress and inflammation in spleen that resulted from SRBCs immunization; in addition to, B cell stimulation that was apparent from the high level of anti-SRBCs antibodies. Camel milk is recommended for newborn consumption, due to its high-water content, unsaturated fatty acids, and vitamin C, as well as low lactose and fat content.

Plasminogen activator urokinase interacts with the fusion protein and antagonizes the growth of Peste des petits ruminants virus.

Peste des petits ruminants is an acute and highly contagious disease caused by the Peste des petits ruminants virus (PPRV). Host proteins play a crucial role in viral replication. However, the effect of fusion (F) protein-interacting partners on PPRV infection is poorly understood. In this study, we found that the expression of goat plasminogen activator urokinase (PLAU) gradually decreased in a time- and dose-dependent manner in PPRV-infected goat alveolar macrophages (GAMs). Goat PLAU was subsequently identified using co-immunoprecipitation and confocal microscopy as an F protein binding partner. The overexpression of goat PLAU inhibited PPRV growth and replication, whereas silencing goat PLAU promoted viral growth and replication. Additionally, we confirmed that goat PLAU interacted with a virus-induced signaling adapter (VISA) to antagonize F-mediated VISA degradation, increasing the production of type I interferon. We also found that goat PLAU reduced the inhibition of PPRV replication in VISA-knockdown GAMs. Our results show that the host protein PLAU inhibits the growth and replication of PPRV by VISA-triggering RIG-I-like receptors and provides insight into the host protein that antagonizes PPRV immunosuppression.IMPORTANCEThe role of host proteins that interact with Peste des petits ruminants virus (PPRV) fusion (F) protein in PPRV replication is poorly understood. This study confirmed that goat plasminogen activator urokinase (PLAU) interacts with the PPRV F protein. We further discovered that goat PLAU inhibited PPRV replication by enhancing virus-induced signaling adapter (VISA) expression and reducing the ability of the F protein to degrade VISA. These findings offer insights into host resistance to viral invasion and suggest new strategies and directions for developing PPR vaccines.

Successful transfer of passive immunity: the natural alternative to antibiotics for boosting the survival of intensively reared dairy goat kids.

In dairy operations, antibiotics have traditionally been used to treat, prevent, and control diseases. However, given the mounting global crisis of antimicrobial resistance (AMR), farmers are urged to re-assess and reduce their reliance on antibiotics. Thus, this randomized, double-blinded cohort study aimed to estimate the prevalence of failed and successful transfer of passive immunity (FTPI and STPI) in dairy goat kids reared under commercial conditions, and the effects of antibiotic metaphylaxis on the pre-weaning (≤42 d old) mortality in FTPI and STPI kids. Plasma concentration of immunoglobulin G at 1d old (pIgG-24 h) was measured in 747 male Saanen kids for the determination of FTPI and STPI (pIgG-24 h < 12 and ≥12 g/L, respectively). Kids were then randomly divided into two groups: those receiving a single penicillin injection at 1 d old (PEN), and those receiving no treatment (CTR). The mean (±SD) pIgG-24 h and initial BW (IBW) were 17 ± 9.8 g/L and 4.1 ± 0.64 kg. The prevalence of FTPI was 29% (220/747 kids). Gastrointestinal complications were the primary cause of death (41%), followed by septicemia (22%) and arthritis (17%). A single penicillin injection reduced preweaning mortality by 55% (10 vs 22%, PEN vs CTR). However, results suggest that such a decline was mainly driven by the improved survival rates among FTPI kids, which increased by 19% (from 62% in CTR-FTPI to 82% in PEN-FTPI), as opposed to an 8% increase among STPI kids (from 85% in CTR-STPI to 93% in PEN-STPI). Additionally, the odds of mortality ≤ 42 d old were threefold higher in the CTR-FTPI group when compared to both the CTR-STPI and PEN-FTPI groups, suggesting a potential parity between STPI and PEN for mortality rate reduction. Taken together, the results indicate that although metaphylactic antibiotics can halve preweaning mortality, similar improvements are likely to be achieved via increased STPI rates. Furthermore, by targeting metaphylactic interventions to high-risk groups (i.e., those displaying signs of inadequate colostrum intake and/or low birth BW), farmers could reduce treatment costs and mitigate AMR risks. While these findings carry considerable weight for commercial dairy goat practices, their applicability to other systems (i.e., extensive, semi-intensive, mohair, meat systems) warrants further investigation.

Development and Characterization of New Monoclonal Antibodies Against Porcine Interleukin-17A and Interferon-Gamma.

Current research efforts require a broad range of immune reagents, but those available for pigs are limited. The goal of this study was to generate priority immune reagents for pigs and pipeline them for marketing. Our efforts were aimed at the expression of soluble swine cytokines and the production of panels of monoclonal antibodies (mAbs) to these proteins. Swine interleukin-17A (IL-17A) and Interferon-gamma (IFNγ) recombinant proteins were produced using yeast expression and used for monoclonal antibody (mAb) production resulting in panels of mAbs. We screened each mAb for cross-species reactivity with orthologs of IL-17A or IFNγ and checked each mAb for inhibition by other related mAbs, to assign mAb antigenic determinants. For porcine IL-17A, the characterization of a panel of 10 mAbs identified eight different antigenic determinants; interestingly, most of the mAbs cross-reacted with the dolphin recombinant ortholog. Likewise, the characterization of a panel of nine anti-PoIFNγ mAbs identified four different determinants; most of the mAbs cross-reacted with dolphin, bovine, and caprine recombinant orthologs. There was a unique reaction of one anti-PoIFNγ mAb that cross-reacted with the zebrafish recombinant ortholog. The αIL-17A mAbs were used to develop a quantitative sandwich ELISA detecting the yeast expressed protein as well as native IL-17A in stimulated peripheral blood mononuclear cell (PBMC) supernatants. Our analyses showed that phorbol myristate acetate/ionomycin stimulation of PBMC induced significant expression of IL-17A by CD3+ T cells as detected by several of our mAbs. These new mAbs expand opportunities for immunology research in swine.

Identification of Differential Responses of Goat PBMCs to PPRV Virulence Using a Multi-Omics Approach.

Peste des petits ruminants (PPR) is an acute transboundary infectious viral disease of small ruminants, mainly sheep and goats. Host susceptibility varies considerably depending on the PPR virus (PPRV) strain, the host species and breed. The effect of strains with different levels of virulence on the modulation of the immune system has not been thoroughly compared in an experimental setting so far. In this study, we used a multi-omics approach to investigate the host cellular factors involved in different infection phenotypes. Peripheral blood mononuclear cells (PBMCs) from Saanen goats were activated with a T-cell mitogen and infected with PPRV strains of different virulence: Morocco 2008 (high virulence), Ivory Coast 1989 (low virulence) and Nigeria 75/1 (live attenuated vaccine strain). Our results showed that the highly virulent strain replicated better than the other two in PBMCs and rapidly induced cell death and a stronger inhibition of lymphocyte proliferation. However, all the strains affected lymphocyte proliferation and induced upregulation of key antiviral genes and proteins, meaning a classical antiviral response is orchestrated regardless of the virulence of the PPRV strain. On the other hand, the highly virulent strain induced stronger inflammatory responses and activated more genes related to lymphocyte migration and recruitment, and inflammatory processes. Both transcriptomic and proteomic approaches were successful in detecting viral and antiviral effectors under all conditions. The present work identified key immunological factors related to PPRV virulence in vitro.

Species-Specific Humoral Immune Responses in Sheep and Goats upon Small Ruminant Lentivirus Infections Inversely Correlate with Protection against Virus Replication and Pathological Lesions.

Maedi-Visna-like genotype A strains and Caprine arthritis encephaltis-like genotype B strains are small ruminant lentiviruses (SRLV) which, for incompletely understood reasons, appear to be more virulent in sheep and goats, respectively. A 9-month in vivo infection experiment using Belgian genotype A and B SRLV strains showed that almost all homologous (genotype A in sheep; genotype B in goats) and heterologous (genotype A in goats; genotype B in sheep) intratracheal inoculations resulted in productive infection. No differences in viremia and time to seroconversion were observed between homologous and heterologous infections. Higher viral loads and more severe lesions in the mammary gland and lung were however detected at 9 months post homologous compared to heterologous infection which coincided with strongly increased IFN-γ mRNA expression levels upon homologous infection. Pepscan analysis revealed a strong antibody response against immune-dominant regions of the capsid and surface proteins upon homologous infection, which was absent after heterologous infection. These results inversely correlated with protection against virus replication in target organs and observed histopathological lesions, and thus require an in-depth evaluation of a potential role of antibody dependent enhancement in SRLV infection. Finally, no horizontal intra- and cross-species SRLV transmission to contact animals was detected.

Fasciola gigantica tegumental calcium-binding EF-hand protein 4 exerts immunomodulatory effects on goat monocytes.

BACKGROUND: The liver fluke Fasciola gigantica secretes excretory-secretory proteins during infection to mediate its interaction with the host. In this study, we investigated the immunomodulatory effects of a recombinant tegumental calcium-binding EF-hand protein 4 of F. gigantica (rFg-CaBP4) on goat monocytes. METHODS: The rFg-CaBP4 protein was induced and purified by affinity chromatography. The immunogenic reaction of rFg-CaBP4 against specific antibodies was detected through western blot analysis. The binding of rFg-CaBP4 on surface of goat monocytes was visualized by immunofluorescence assay. The localization of CaBP4 within adult fluke structure was detected by immunohistochemical analysis. The cytokine transcription levels in response to rFg-CaBP4 were examined using ABI 7500 real-time PCR system. The expression of the major histocompatibility complex (MHC) class-II molecule (MHC-II) in response to rFg-CaBP4 protein was analyzed using Flow cytometry. RESULTS: The isopropyl-ß-D-thiogalactopyranoside-induced rFg-CaBP4 protein reacted with rat sera containing anti-rFg-CaBP4 polyclonal antibodies in a western blot analysis. The adhesion of rFg-CaBP4 to monocytes was visualized by immunofluorescence and laser scanning confocal microscopy. Immunohistochemical analysis localized native CaBP4 to the oral sucker, pharynx, genital pore, acetabulum and tegument of adult F. gigantica. Co-incubation of rFg-CaBP4 with concanavalin A-stimulated monocytes increased the transcription levels of interleukin (IL)-2, IL-4, interferon gamma and transforming growth factor-ß. However, a reduction in the expression of IL-10 and no change in the expression of tumor necrosis factor-α were detected. Additionally, rFg-CaBP4-treated monocytes exhibited a marked increase in the expression of the major histocompatibility complex (MHC) class-II molecule (MHC-II) and a decrease in MHC-I expression, in a dose-dependent manner. CONCLUSIONS: These findings provide additional evidence that calcium-binding EF-hand proteins play roles in host-parasite interaction. Further characterization of the immunomodulatory role of rFg-CaBP4 should expand our understanding of the strategies used by F. gigantica to evade the host immune responses.

The immunogenic maturation of goat monocyte-derived dendritic cells and upregulation of toll-like receptors by five antigens of Haemonchus contortus in-vitro.

Previously, it was found that several proteins of Haemonchus contortus were involved in the stimulation of the host immune system. However, the information about the selection of superlative antigens with immunogenic efficacies on host DCs is lacking. In the current study, the stimulatory effects of five recombinant proteins (elongation factor-1α, arginine kinase, ES-15, ES-24, and ADP-ribosylation factor 1) of H. contortus on the maturation of goat monocyte-derived dendritic cells (md-DCs) were reported. Recombinant proteins were purified separately in E. coli expression and incubated with isolated goat peripheral blood mononuclear cells (PBMC). Immunofluorescence assay (IFA) results confirmed the binding of these molecules to the md-DC's surface as compared to control groups. In the flow cytometry analysis, recombinant proteins induced md-DC stimulation via the up-regulation of the expression of the costimulatory molecule (CD80) and MHC-II. Quantitative RT-PCR data showed a significant increase in the expression of specific genes of the WNT and toll-like receptor (TLR) signaling pathways. The result of ELISA indicated the higher levels of cytokine (IL-10, IL-12, IFN-γ, and TNF-α) secretion in the md-DC compared to the negative (pET-32a His-Tag) and blank (PBS) control groups. The data gives valuable support in the selection of potential antigens for future studies on the immunomodulation of the host against the infection of H. contortus.

Characterization of innate immune response to Brucella melitensis infection in goats with permissive or restrictive phenotype for Brucella intramacrophagic growth.

Caprine brucellosis is a chronic, world-wide distributed disease which causes reproductive failure in goats and Brucella melitensis, its causative agent, bears a great zoonotic potential. There is evidence suggesting that some cattle and pigs have an innate ability to resist Brucella infection, but this has not yet been investigated in goats. In this study, we compared caprine macrophages that exhibit extreme restriction and permissiveness to B. melitensis' intracellular growth in vitro. Monocyte derived macrophages (MDMs) from 110 female goats were cultured and challenged in vitro with B. melitensis 16 M. After initial screening, 18 donor goats were selected based on their macrophages ability to restrict or allow bacterial intracellular growth and some elements of humoral and cellular immunity were studied in depth. MDMs that were able to restrict the pathogen's intracellular growth showed enhanced bacterial internalization, although there were no differences between groups in the production of reactive oxygen and nitrogen intermediates following 48 h treatment with heat-killed B. melitensis. Moreover, there were no differences between groups in the level of antibodies reacting with keyhole limpet hemocyanin (natural antibodies, NAbs) or with Brucella LPS antigens (cross-reacting antibodies, CrAbs), although a strong positive correlation between individual levels of IgM NAbs and IgM CrAbs was detected. Altogether, these results represent an initial step in understanding innate primary host response to B. melitensis, and deciphering which mechanisms may determine a successful outcome of the infection in goats.

Serological and Molecular Investigation of Batai Virus Infections in Ruminants from the State of Saxony-Anhalt, Germany, 2018.

Arthropod-borne Batai virus (BATV) is an Orthobunyavirus widely distributed throughout European livestock and has, in the past, been linked to febrile diseases in humans. In Germany, BATV was found in mosquitoes and in one captive harbor seal, and antibodies were recently detected in various ruminant species. We have, therefore, conducted a follow-up study in ruminants from Saxony-Anhalt, the most affected region in Eastern Germany. A total of 325 blood samples from apparently healthy sheep, goats, and cattle were tested using a BATV-specific qRT-PCR and SNT. Even though viral RNA was not detected, the presence of antibodies was confirmed in the sera of all three species: sheep (16.5%), goats (18.3%), and cattle (41.4%). Sera were further analyzed by a glycoprotein Gc-based indirect ELISA to evaluate Gc-derived antibodies as a basis for a new serological test for BATV infections. Interestingly, the presence of neutralizing antibodies was not directly linked to the presence of BATV Gc antibodies. Overall, our results illustrate the high frequency of BATV infections in ruminants in Eastern Germany.

Comparison of the response of mammary gland tissue from two divergent lines of goat with high and low milk somatic cell scores to an experimental Staphylococcus aureus infection.

Mastitis represents one of the major economic and health threats to the livestock sector associated with reduction in milk quality, loss of production and is a major reason for culling. Somatic cell score (SCS) is used as a criterion in breeding programmes to select cows genetically less susceptible to mastitis. The relevance of SCS as a predictor of udder health and susceptibility to mastitis is still untested in goats. In this study, two lines of French Alpine goats selected for extreme breeding values for somatic cell scores, one line with high SCS (HSCS) and the other with low SCS (LSCS), were used to test the hypothesis that the mammary response and function differed between the lines. The aim of the present study was to investigate differences in the early immune response in caprine mammary gland tissues challenged with Staphylococcus aureus, one of the main pathogens responsible for the intra-mammary infection in small ruminants, using transcriptomic and histopathology analyses. The comparison between HSCS and LSCS goat lines, showed differences in the response at the histological level for inflammation, presence of neutrophils and micro-abscess formation, and at the molecular level in the expression of CXCL8, IL-6, NFKBIZ and IL-1ß. CXCL8 and CXCL2 genes, which showed a higher level of expression in the experimentally infected HSCS line. The molecular data and histopathology both suggested that following S. aureus infection, mobilization, recruitment, infiltration, and chemotaxis of neutrophil, leads to a more severe inflammation in the HSCS compared to LSCS animals. Our results represent an initial basis for further studies to unravel the genetic basis of early mastitis inflammatory responses and the selection of dairy animals more resistant to bacterial mastitis.

Characterization of the domestic goat γδ T cell receptor gene loci and gene usage.

Goats and cattle diverged 30 million years ago but retain similarities in immune system genes. Here, the caprine T cell receptor (TCR) gene loci and transcription of its genes were examined and compared to cattle. We annotated the TCR loci using an improved genome assembly (ARS1) of a highly homozygous San Clemente goat. This assembly has already proven useful for describing other immune system genes including antibody and leucocyte receptors. Both the TCRγ (TRG) and TCRδ (TRD) loci were similarly organized in goats as in cattle and the gene sequences were highly conserved. However, the number of genes varied slightly as a result of duplications and differences occurred in mutations resulting in pseudogenes. WC1+ γδ T cells in cattle have been shown to use TCRγ genes from only one of the six available cassettes. The structure of that Cγ gene product is unique and may be necessary to interact with WC1 for signal transduction following antigen ligation. Using RT-PCR and PacBio sequencing, we observed the same restriction for goat WC1+ γδ T cells. In contrast, caprine WC1+ and WC1- γδ T cell populations had a diverse TCRδ gene usage although the propensity for particular gene usage differed between the two cell populations. Noncanonical recombination signal sequences (RSS) largely correlated with restricted expression of TCRγ and δ genes. Finally, caprine γδ T cells were found to incorporate multiple TRD diversity gene sequences in a single transcript, an unusual feature among mammals but also previously observed in cattle.

Study on immunogenicity and antigenicity of a novel brucella multiepitope recombined protein.

Currently, brucellosis is a reemerged zoonotic infectious disease with an increased incidence in recent years. A simple, rapid and sensitive method for diagnosing brucellosis can help to reduce medical burden and economic loss. Previously, a multiple epitope recombinant protein was constructed based on linear B-cell epitope prediction tools. In this study, the recombinant protein was used as an antigen to study the immune response produced by immunized mice, and goat serum was used to verify its diagnostic accuracy. The production of antibodies was successfully induced in the vaccinated mice. Flow cytometric analysis revealed that the percentage of CD4+, CD8+ and the CD4+/CD8+ ratios were increased by T cell subsets in mouse splenocytes, indicating that the recombinant protein induced a strong immune response had strong immunoreactivity. Using indirect ELISA, the recombinant protein correctly diagnosed positive and negative brucellosis samples. Compared with the whole bacterial antigen, the recombinant protein had a weaker sensitivity but a stronger specificity. Animal experiments showed that the recombinant protein had good antigenicity, and indirect ELISA indicates that it can be used as an antigen to diagnose brucellosis. Therefore, the recombinant protein is a potential candidate antigen for brucellosis vaccine development and serological diagnosis.

The GT1-TPS Structural Domain Protein From Haemonchus contortus Could Be Suppressive Antigen of Goat PBMCs.

Trehalose phosphate synthase (TPS), a key enzyme in trehalose synthesis, is not present in mammals but critical to the viability of a wide range of lower organisms. However, almost nothing is known about the function of Hc-TPS (GT1-TPS structural domain protein from Haemonchus contortus). In this study, Hc-TPS gene was cloned and the recombinant protein (rHc-TPS) was expressed and purified. The quantitative real-time PCR (qPCR) results showed that Hc-TPS was transcribed at different stages of H. contortus, with higher levels of transcription at the molting and embryo stages. Immunofluorescence analysis showed that Hc-TPS was widely distributed in adults, but the expression was mainly localized on the mucosal surface of the intestine as well as in the embryos of female worms. The impacts of rHc-TPS on peripheral blood mononuclear cell (PBMC) proliferation, nitric oxide (NO) generation, transcriptional expression of cytokines, and related pathways were examined by co-incubating rHc-TPS with goat PBMCs. The results showed that rHc-TPS significantly inhibited PBMC proliferation and NO secretion in a dose-dependent manner. We also found that rHc-TPS activated the interleukin (IL)-10/signal transducer and activator of transcription 3/suppressor of cytokine signaling 3 (IL-10/STAT3/SOCS3) axis and significantly promoted SOCS3 expression, while inhibiting interferon-gamma (INF-γ), IL-4, IL-9, and IL-2 pathways. Our findings may contribute to understanding the immune evasion mechanism for the parasite during host-parasite interactions and also help to provide ideas for discovering new drug targets.