Evaluation of peripheral lymphocytes after weaning and vaccination for Mycoplasma hyopneumoniae

This study evaluated immune cell populations in pigs following weaning and vaccination for Mycoplasma hyopneumoniae. Piglets (n = 24) were weaned (day 0) at 16 (±1) days of age, and randomly assigned to the vaccination group (n = 16) or control group (n = 8). Complete blood cell counts, flow cytometry and serology were completed for blood samples collected on days 0 (within hours of weaning), 3, 7, 14, 30 and 60. The M. hyopneumoniae S:P ratios (sample optical density: positive control optical density) were negative in the vaccination group until days 30 and 60, when the S:P ratios were 1.3 and 1.0, respectively. Control animals remained serologically negative. The percentage of CD4⁺ T cells was less (P < 0.01) in control pigs than vaccinated pigs at day 3. In contrast, numbers of CD8⁺ and CD4⁺CD8⁺ T cells were greater (P < 0.01) in control pigs than in vaccinated pigs at days 3 and 7. After day 7, few differences in immune cell types were evident between the groups. Differences in lymphocyte populations could not be solely attributed to vaccination, due at least in part, to the confounding influence of weaning. It was difficult to distinguish the influence of vaccination from the impact of weaning on peripheral immune cell populations.     

Rapid NK-cell activation in chicken after infection with infectious bronchitis virus M41

Natural killer (NK) cells are cytotoxic lymphocytes and play an important role in the early defence against viruses. In this study we focussed on NK cell and interferon (IFN) responses after infection with infectious bronchitis virus (IBV). Based on surface expression of CD107+, enhanced activation of lung NK cells was observed at 1 dpi, whereas in blood prolonged NK-cell activation was found. IFN-α and IFN-β mRNA and proteins were not rapidly induced whereas IFN-γ production in lung, measured by Elispot assay, increased over time at 2 and 4 dpi. In contrast, IFN-γ production in blood was highest at 1 dpi and decreased over time down to levels comparable to uninfected birds at 4 dpi. Collectively, infection with IBV-M41 resulted in activation of NK cells in the lung and blood and rapid production of IFN-γ and not IFN-α and IFN-β compared to uninfected birds.     

Antibody and T cell responses induced in chickens immunized with avian influenza virus N1 and NP DNA vaccine with chicken IL-15 and IL-18

We had examined the immunogenicity of a series of plasmid DNAs which include neuraminidase (NA) and nucleoprotein (NP) genes from avian influenza virus (AIV). The interleukin-15 (IL-15) and interleukin-18 (IL-18) as genetic adjuvants were used for immunization in combination with the N1 and NP AIV genes. In the first trial, 8 groups of chickens were established with 10 specific-pathogen-free (SPF) chickens per group while, in the second trial 7 SPF chickens per group were used. The overall N1 enzyme-linked immunosorbent assay (ELISA) titer in chickens immunized with the pDis/N1 + pDis/IL-15 was higher compared to the chickens immunized with the pDis/N1 and this suggesting that chicken IL-15 could play a role in enhancing the humoral immune response. Besides that, the chickens that were immunized at 14-day-old (Trial 2) showed a higher N1 antibody titer compared to the chickens that were immunized at 1-day-old (Trial 1). Despite the delayed in NP antibody responses, the chickens co-administrated with IL-15 were able to induce earlier and higher antibody response compared to the pDis/NP and pDis/NP + pDis/IL-18 inoculated groups. The pDis/N1 + pDis/IL-15 inoculated chickens also induced higher CD8+ T cells increase than the pDis/N1 group in both trials (P < 0.05). The flow cytometry results from both trials demonstrated that the pDis/N1 + pDis/IL-18 groups were able to induce CD4+ T cells higher than the pDis/N1 group (P < 0.05). Meanwhile, pDis/N1 + pDis/IL-18 group was able to induce CD8+ T cells higher than the pDis/N1 group (P < 0.05) in Trial 2 only. In the present study, pDis/NP was not significant (P > 0.05) in inducing CD4+ and CD8+ T cells when co-administered with the pDis/IL-18 in both trials in comparison to the pDis/NP. Our data suggest that the pDis/N1 + pDis/IL-15 combination has the potential to be used as a DNA vaccine against AIV in chickens.     

Leukocytic responses and intestinal mucin dynamics of broilers protected with Enterococcus faecium EF55 and challenged with Salmonella Enteritidis

The protective effect of Enterococcus faecium EF55 in chickens challenged with Salmonella enterica serovar Enteritidis phage type 4 (SE PT4) was assessed. The antibacterial effect on the bacterial microflora in the small intestine in relation to white blood cell count, phenotyping of peripheral blood and intestinal lymphocytes, functional activity of lymphocytes and phagocytes and mucin quantitation were investigated. Day-old chicks (85) were randomly divided into four groups. The probiotic group (EF) and Salmonella + probiotic group (EFSE) received E. faecium EF55 (10⁹ CFU – 3 g/group/day) for 21 days. The Salmonella group (SE) and EFSE group were infected with Salmonella Enteritidis (10⁸ CFU in 0.2 ml PBS) in a single dose per os on day four of the experiment. The control group chicks (C) were fed a commercial diet without added bacteria. Supplementation of EF55 in the diet of the chickens in the EFSE group, challenged with S. Enteritidis, caused the density of the intestinal mucin layer to increase significantly in non-specific regions (duodenum and jejunum), but decrease significantly in target regions (caeca) for S. Enteritidis. Probiotic treatment also appeared to result in a significantly higher number of lymphocytes in peripheral blood and a tendency to increase CD3, CD4, CD8, and IgM positive cells 3 days post-infection with S. Enteritidis. The results demonstrated an antibacterial effect and suggested that EF55 had a moderating effect on intestinal mucin production and leukocytic response in the early phase of S. Enteritidis infection.     

Characterization of tuberculous granulomas in different stages of progression and associated tertiary lymphoid tissue in goats experimentally infected with Mycobacterium avium subsp. hominissuis

Oral infection of goats with Mycobacterium avium subsp. hominissuis (MAH) resulted in a large variety of granulomas in organized gut-associated lymphatic tissues and intestinal lymph nodes. To characterize the cellular composition of granulomas, CD4⁺, CD8⁺, γδ, B lymphocytes and plasma, CD25⁺, CD68⁺, MHC-II⁺, Ki67⁺ and endothelial cells were labeled in consecutive frozen sections by immunohistochemistry and acid fast bacilli (AFB) by Kinyoun stain. Granulomas with extensive necrosis, little mineralization and variable numbers of AFB surrounded by many CD4⁺ T cells, but only few epitheloid macrophages were observed in severely sick goats at 2–3 mpi. They were interpreted as exuberant immune reaction. Organized granulomas with very few AFB were seen in clinically healthy goats at 13 mpi. The necrotic cores were surrounded by a zone of granulomatous infiltrate with many epitheloid macrophages and few lymphocytes. This zone was initially wide and highly vascularized and became progressively smaller. It was enclosed by an increasing layer of connective tissue. All organized granulomas were surrounded by compartimentalized tertiary lymphoid tissue. The granulomas in experimental infection of goats with MAH reflect the heterogeneity of lesions seen in mycobacterial infections of humans and ruminants and are therefore valuable for comparative research.     

Expansion of intracellular IFN-γ positive lymphocytes during Mycoplasma agalactiae infection in sheep

A method to assess the expansion of antigen-specific intracellular IFN-γ positive T cell subsets during the infection will be helpful for a better understanding of mycoplasmal infections physiopathology in the sheep. We analysed the percentage of antigen-specific lymphocytes positive for intracellular IFN-γ during the infection of sheep with Mycoplasma agalactiae by culturing peripheral blood mononuclear cells of infected or uninfected animals with irradiated M. agalactiae. The expansion of antigen-specific IFN-γ positive lymphocytes in infected sheep was initially sustained by CD4⁺ T cells at day 15 after infection, when antigen specific IgG start to be detectable, followed by CD8/IFN-γ double positive cells. γδ T-cells were not expanded at any time point analysed. IFNγ⁺ T cells disappear 60 days after infection, suggesting that antigen specific IFNγ⁺ T cells, mainly detected in the early phase of the disease, could be useful to understand the role of cell-mediated immunity during M. agalactiae infection.     

Influence of age on bronchoscopic findings in healthy beagle dogs

The aims of this study were to evaluate the effect of age on bronchoscopic features and bronchoalveolar lavage fluid (BALF) cellularity in dogs. Thirty healthy beagle dogs from three age groups were included: young dogs (10 months to 4.5 years of age; n = 8), middle-aged dogs (5–8 years old; n = 13) and older dogs (>8 years; n = 9). Haematology, thoracic radiography, bronchoscopy and bronchoalveolar lavage were performed; bronchoscopic findings were scored and BALF total and differential cell counts were determined. The total bronchoscopic score was higher in older dogs; these dogs had more irregular bronchial mucosa, more prominent mucosal vessels and bronchiectasis. Younger dogs had a higher percentage of neutrophils in BALF compared with middle-aged and old dogs and a higher percentage of lymphocytes in BALF compared with middle-aged dogs. The results show that age has an effect on bronchoscopic features of airways and the composition of BALF in the dog.     

Bovine natural killer cells are present in Escherichia coli infected mammary gland tissue and show antimicrobial activity in vitro

Natural killer (NK) cells are early responders in bacterial infections but their role in bovine mastitis has not been characterized. For the first time, we show the presence of NK cells (NKp46⁺/CD3⁻) in bovine mammary gland tissue after an intramammary challenge with Escherichia (E.) coli. A small number of NK cells was detected in milk from quarters before and during an E. coli challenge. In vitro cultures of primary bovine mammary gland epithelial cells stimulated with UV irradiated E. coli induced significant migration of peripheral blood NK cells (pbNK) within 2 h. Furthermore, pbNK cells significantly reduced counts of live E. coli in vitro within 2 h of culture. The results show that bovine NK cells have the capacity to migrate to the site of infection and produce antibacterial mediators. These findings introduce NK cells as a leukocyte population in the mammary gland with potential functions in the innate immune response in bovine mastitis.     

Diet selection by calves facing pairs of nutritionally complementary foods

Experimental evidence suggests that ruminant animals are capable of selecting a nutritionally balanced diet (i.e. a diet appropriate to their metabolic needs) from imbalanced but complementary foods. We tested this hypothesis by offering calves different combinations of the same complementary foods. A 5 × 5 Latin square design was used to assess the effect on diet selection of different combinations (termed as food A and food B) of two complementary foods (alfalfa-grass hay and maize grain) offered in separate feed bunks. In Treatment 1, food A and food B had the same composition, 50.0% hay and 50.0% grain (T50:50). In the rest of the treatments, hay–grain combinations in foods A and B were, respectively: 62.5–37.5% and 37.5–62.5% (T63:37); 75.0–25.0% and 25.0–75.0% (T75:25); 87.5–12.5% and 12.5–87.5% (T88:12); and 100.0–0.0% and 0.0–100.0% (T100:0). Daily intake data were analysed through repeated measures analysis. Crude protein intake was similar across treatments (P = 0.28). An increasing tendency (P = 0.01) in food B intake (the food with more grain content) from T50:50 to T100:0, however, led into a decreasing tendency (P < 0.001) in the protein: energy ratio of the diet across treatments. Grain consumption in treatments T75:25, T88:12 and T100:0 was higher than expected, leading to protein levels in the diet below those required for a balanced diet (predicted vs. observed: 19.38 vs. 16.67, P = 0.004; 20.53 vs. 16.10, P = 0.001; and 20.23 vs. 16.79 g kgMW^? 1 d^? 1, P = 0.015, for T75:25, T88:12 and T100:0, respectively). Although calves were potentially able to select a balanced diet in all feeding treatments, they failed when offered high grain foods at choice (75% of maize grain or more). Evolved mechanisms to regulate macronutrient intake under natural conditions may fail to adequately operate in artificial environments involving high-energy density foods (e.g. maize grain).     

Differential expression of pro-inflammatory and anti-inflammatory cytokines during experimental infection with low or high virulence bovine viral diarrhea virus in beef calves

The objective was to compare the mRNA expression of pro-inflammatory (TNF-α, IL-1β, IFN-γ, IL-2, IL-12, IL-15) and anti-inflammatory (IL-4, IL-10, TGF-β) cytokines, after experimental infection with low or high virulence noncytopathic (ncp) bovine viral diarrhea virus (BVDV). Thirty BVDV-naïve, beef calves were intranasally inoculated with low (LV; n = 10, SD-1) or high (HV; n = 10, 1373) virulence ncp BVDV or with BVDV-free cell culture medium (Control, n = 10). Calves were euthanized on day 5 post-inoculation, and tracheo-bronchial lymph node and spleen samples were collected for mRNA expression through quantitative-RT-PCR. mRNA levels of pro-inflammatory (TNF-α, IL-1β, IL-2, IFN-γ) and anti-inflammatory (IL-4 and IL-10) cytokines were up-regulated in tracheo-bronchial lymph nodes of HV, but not in LV, compared to the control group (P < 0.05). IL-12 mRNA level was up-regulated in tracheo-bronchial lymph nodes of both LV and HV groups (P ≤ 0.05). A significant up-regulation of IL-15 mRNA was observed in tracheo-bronchial lymph nodes for LV calves (P < 0.002), but not for HV calves. Experimental inoculation with BVDV-2 1373 stimulated significant mRNA expression of pro-inflammatory and anti-inflammatory cytokines. In contrast, inoculation with BVDV-1a SD-1 only resulted in up-regulation of IL-12 and IL-15 mRNA, which is associated with activation of macrophages and NK cells during innate immune response.     

Pharmacokinetics of oral gabapentin in greyhound dogs

The purpose of this study was to assess the pharmacokinetics of gabapentin in healthy greyhound dogs after single oral doses targeted at 10 and 20 mg/kg PO. Six healthy greyhounds were enrolled (3 males, 3 females). Blood was obtained at predetermined times for the measurement of gabapentin plasma concentrations by liquid chromatography/mass spectrometry. Pharmacokinetic parameters were determined with computer software.The actual mean (and range) doses administered were 10.2 (9.1–12.0) mg/kg and 20.5 (18.2–24) mg/kg for the 10 mg/kg and 20 mg/kg targeted dose groups. The mean C_MAX for the 10 and 20 mg/kg groups were 8.54 and 13.22 μg/mL at 1.3 and 1.5 h, and the terminal half-lives were 3.3 and 3.4 h, respectively. The relative bioavailability of the 10 mg/kg group was 1.13 compared to the 20 mg/kg group. Gabapentin was rapidly absorbed and eliminated in dogs, indicating that frequent dosing is needed to maintain minimum targeted plasma concentrations.     

The effect of sea transport from Ireland to the Lebanon on inflammatory,adrenocortical, metabolic and behavioural responses of bulls

The objective was to investigate the effect of sea transport on the physiological, behavioural and performance responses of bulls. One-hundred and eleven bulls (mean body weight (standard error of the mean) 429 (5.7 kg)) were randomly assigned to one of three treatments; control (C; n = 54) bulls were housed in 6 pens at Teagasc, Grange Research Centre at a stocking density of (1), 1.7 m²/head (C1.7; 3 pens) and (2), 3.4 m²/head (C3.4; 3 pens) and (3), transported (T) bulls (n = 57) were penned at a space allowance of 1.7 m²/head (6 pens) and allocated to one of five decks on the shipping vessel. C and T bulls were subjected to the same live weight (d −2), blood sampling and rectal temperature (d −1) measurements pre-transport and on d 3, d 6, d 9 and d 11 of the study. T bulls had greater (P < 0.05) live weight gain (+4.4%) compared with C1.7 bulls (−2.0%) and C3.4 (+0.13%)). Time spent lying was greater (P < 0.05) among C1.7 and C3.4 bulls (9.9% and 53.3%, respectively) compared with T bulls (45.8%). Rectal body temperature was not different (P > 0.05) among treatment groups throughout the study. At d 11, neutrophil % was greater (P < 0.05) in transported bulls on decks 1, 2, 4 and 5 compared with C1.7 and C3.4 treatments. Plasma cortisol concentrations were not different (P > 0.05) between control and transported bulls. Plasma creatine kinase (CK) activity was lower (P < 0.05) among C3.4 and T bulls on decks 2, 3, 4 and 5 compared with d 3 values. In conclusion, the welfare of bulls transported by sea on the sea journey was not adversely affected. Housing control bulls at a reduced space allowance (1.7 m²) had a negative effect on live weight gain.     

Fluoxetine combined with clorazepate dipotassium and behaviour modification for treatment of anxiety-related disorders in dogs

The effectiveness of clorazepate dipotassium combined with fluoxetine and a behaviour modification programme for the treatment of anxiety disorders in dogs was investigated. Forty dogs with anxiety disorders were initially enrolled and 36 dogs completed the trial. Dogs were classified into two behavioural categories (anxious dogs with aggression and anxious dogs without aggression) according to their presenting complaints, and were also subdivided into males, females, juveniles and adults. The dog owners were provided with a behaviour modification plan for their dogs to be commenced in the first week of therapy. Clorazepate dipotassium was administered PO at 1.0 mg/kg every 24 h for 4 weeks, and fluoxetine was administered PO at 1.0 mg/kg every 24 h for 10 weeks. Therapy with both drugs was initiated simultaneously. Improvement was reported in 25/36 dogs. Significant differences in treatment effects were observed between anxious dogs with aggression and anxious dogs without aggression (P < 0.05). Positive correlations between owner compliance with the treatment plan and reported improvement achieved during three periods of study were also noted.     

Pharmacokinetic,metabolism and withdrawal time of orphenadrine in camels (Camelus dromedarius) after intravenous administration

The pharmacokinetics of orphenadrine (ORPH) following a single intravenous (i.v.) dose was investigated in six camels (Camelus dormedarius). Orphenadrine was extracted from the plasma using a simple sensitive liquid–liquid extraction method and determined by gas chromatography/mass spectrometry (GC/MS). Following i.v. administration plasma concentrations of ORPH decline bi-exponentially with distribution half-life (t_1/2α) of 0.50 ± 0.07 h, elimination half-life (t_1/2β) of 3.57 ± 0.55 h, area under the time concentration curve (AUC) of 1.03 ± 0.10 g/h l⁻¹. The volume of distribution at steady state (Vd_ss) 1.92 ± 0.22 l kg⁻¹, volume of the central compartment of the two compartment pharmacokinetic model (V_c) 0.87 ± 0.09 l kg⁻¹, and total body clearance (Cl_T) of 0.60 ± 0.09 l/h kg⁻¹. Three orphenadrine metabolites were identified in urine samples of camels. The first metabolite N-desmethyl-orphenadrine resulted from N-dealkylation of ORPH with molecular ion m/z 255. The second N,N-didesmethyl-orphenadrine, resulted from N-didesmethylation with molecular ion m/z 241. The third metabolite, hydroxyl-orphenadrine, resulted from the hydroxylation of ORPH with molecular ion m/z 285. ORPH and its metabolites in camel were extensively eliminated in conjugated form. ORPH remains detectable in camel urine for three days after i.v. administration of a single dose of 350 mg orphenadrine aspartate.     

Immune and inflammatory responses in pigs infected with Trichuris suis and Oesophagostomum dentatum

The aim of the present study was to investigate parasite induced immune responses in pigs co-infected with Trichuris suis and Oesophagostomum dentatum as compared to mono-species infected pigs. T. suis is known to elicit a strong immune response leading to rapid expulsion, and a strong antagonistic effect on O. dentatum populations has been observed in co-infected pigs. Forty-eight helminth naïve pigs were allocated into 4 groups in a 2-factorial design. Two groups were trickle inoculated with either 10 T. suis eggs/kg/day (Group T) or 20 O. dentatum L3/kg/day (Group O). Group OT was infected with same levels of both T. suis and O. dentatum (Group OT) and Group C remained uninfected. In each group, six pigs were necropsied after 35 days and the remaining pigs after 71 days. Parasite E/S-antigen specific serum antibodies were quantified by an in-direct ELISA. qPCR was used to measure the expression of immune function related genes in the mucosa of proximal colon and the draining lymph node. Highly significant interactions were identified for O. dentatum specific IgG1 (p < 0.0001) and IgG2 (p < 0.0006) antibodies with a remarkable 2-fold higher antibody response in group OT pigs as compared to group O. These findings indicated that T. suis enhanced the antibody response against O. dentatum in Group OT. The gene expression data confirmed a strong Type 2 response to T. suis (e.g. marked increase in IL-13, ARG1 and CCL11) and clearly weaker in amplitude and/or delayed onset response to O. dentatum in the single infected group. Interactions were found between the two nematodes with regard to several cytokines, e.g. the increase in IL-13 observed in Group T was absent in Group OT (p = 0.06, proximal colon mucosa, 35 and 71 p.i.). Some of these immune response-related interactions may support, or even partially explain, the observed interactions between the two worm populations in co-infected pigs.     

Development of a tandem repeat-based multilocus typing system distinguishing Babesia bovis geographic isolates

Mini- and microsatellite sequences have proven to be excellent tools for the differentiation of strains and populations in several protozoan parasites due to their high variability. In the present work we have searched the genome of the tick-transmitted bovine hemoprotozoon Babesia bovis for tandem repeats (TRs) that could be useful for a multilocus typing system. Hundred and nineteen sequences were shortlisted and tested in five common B. bovis reference isolates originating from distinct geographic locations of North and South America: Texas, USA (T2Bo), Mexico (RAD and Mo7), and Santa Fe and Salta, Argentina (R1A and S2P, respectively). Satellite sequences were PCR-amplified using specific primers, separated by polyacrylamide gel electrophoresis, visualized by silver staining and sized. Fourteen TR sequences could be reliably amplified in all isolates and displayed length polymorphism. All primers used were specific for B. bovis and did not amplify genomic DNA from the bovine host or from Babesia bigemina, the principal co-infecting bovine parasite in the Americas, allowing their future use in field surveys. The 14 satellite markers identified are distributed throughout the four chromosomes of B. bovis as follows: chromosome 1 (n = 3), chromosome 2 (n = 2), chromosome 3 (n = 5), and chromosome 4 (n = 4). Within the five B. bovis isolates we identified nine satellite marker loci with two alleles, three with three alleles, one with four and another with five alleles. In comparison to Theileria parva, a bovine hemoprotozoan that pertains to the same piroplasmida order and owns a genome of similar size, the number of polymorphic TRs and the average number of alleles per TR locus seem to be significantly reduced in the B. bovis genome. Furthermore, the ratio of micro- to minisatellites in both B. bovis and T. parva is considerably lower than in other eukaryotes, as confirmed by bioinformatic analysis. The multilocus genotype of the five B. bovis isolates was assessed and the genetic distance between each other determined followed by cluster analysis based on neighbor joining. The resulting phenogram showed that B. bovis isolates segregated into three clusters according to their geographic origin. The presented marker system is suitable to explore various parameters of B. bovis populations such as genetic diversity, infection dynamics and their structure under different epidemiological situations, which are of crucial importance for improved control strategies.     

The influence of oral bacteria on tissue levels of Toll-like receptor and cytokine mRNAs in feline chronic gingivostomatitis and oral health

Feline chronic gingivostomatitis (FCGS) is an inflammatory disease of the oral cavity that causes severe pain and distress in affected cats. Treatment methods are currently very limited. The aims of this study were to assess the feline innate immune response by investigating the levels of cytokine and Toll-like receptor (TLR) mRNAs in tissue biopsies of cats with and without FCGS, and to relate this to the presence or absence of putative oral pathogens identified previously within these cats. Mucosal biopsies were collected from 28 cats with FCGS and eight healthy cats. The levels of TLR (TLR2, TLR3, TLR4, TLR7, TLR9) and cytokine (IL-1β, IL-4, IL-6, IL-10, IL-12, TNF-α, IFN-γ) mRNA was determined using quantitative PCR. In the FCGS group a statistically significant increase was seen in TLR2, TLR7, TNF-α, IFN-γ, IL-1β and IL-6 mRNA levels compared to the healthy group. In cats where Tannerella forsythia was present, statistically significant increases were seen in TLR2, TLR4, TLR7, TLR9, TNF-α and IL-1β mRNA levels compared to cats where this putative pathogen was absent. Statistically significant increases in mRNA expression were also seen in cats harbouring feline calicivirus (FCV) (TLR2, IL-1β, IL-6, IFN-γ) and Porphyromonas circumdentaria (TLR2, TLR3) compared to cats where these putative pathogens were absent. Pasteurella multocida subsp. multocida and Pseudomonas sp. did not significantly alter the expression of any TLR or cytokine mRNAs when compared to animals who tested negative for these species, while cats colonised with P. multocida subsp. septica demonstrated a statistically significant reduction in the expression of TLR7, TNF-α and IFN-γ mRNAs compared to cats free of this species. The expression of mRNA for several TLRs and cytokines is elevated in FCGS. A positive correlation was observed between clinical disease severity and the presence of FCV (p = 0.001; Rho = 0.58). Although the number of cats harbouring T. forsythia was low by comparison, 80% of samples in which it was present were from cases with the highest clinical disease severity. Positive correlations with clinical disease severity were seen for TLR2 (p = 0.00086), TLR7 (p = 0.049), TNF-α (p = 0.027), IFN-γ (p = 0.0015), IL-1β (p = 0.004) and IL-6 (p = 0.00001) mRNAs. The putative pathogens FCV and T. forsythia may be important in stimulating a host immune response to FCGS and may play a role in the pathogenesis of this disease.     

Characterization of an anti-rainbow trout (Oncorhynchus mykiss) CD3ε monoclonal antibody

This study characterizes a monoclonal antibody (mAb) produced against the cytoplasmic tail region of the epsilon chain of the CD3 (CD3?) transmembrane protein found on T lymphocytes of rainbow trout (Oncorhynchus mykiss). Flow cytometry and fluorescent microscopy conducted on trout leukocytes with the anti-trout CD3? mAb showed a distinctive population of IgM^? CD3e⁺ lymphocytes fitting the expected profile of T-cells. Immunoprecipitation of lysates derived from trout lymphocytes revealed a 19 kDa protein and peptide analysis confirmed its specificity for CD3?. In vitro proliferation assays with T-cell mitogens, ConA and PHA, resulted in a 3 fold increase in the percentage of CD3?+ lymphocytes compared to LPS and control cultures. The mAb characterized in this study will be useful in further elucidation for both the role and distribution of T lymphocytes in the teleost immune system.     

Effects of inoculum size on cell-mediated and humoral immune responses of foals experimentally infected with Rhodococcus equi: A pilot study

The objective of this pilot study was to compare the cytokine profile as well as cell-mediated and antibody responses of foals infected with a low inoculum of virulent Rhodococcus equi resulting in subclinical pneumonia to that of foals infected with a high inoculum resulting in severe clinical pneumonia. The mean (±SD) ratio of post-infection to pre-infection anti-R. equi IgG(T) concentration was significantly (P = 0.002) higher in foals infected with the high inoculum (195 ± 145; range 62–328) compared to foals infected with the low inoculum (3.9 ± 4.5; range 0.5–11). Similarly, mean (±SD) ratio of post-infection to pre-infection IgM concentration was significantly (P = 0.002) higher in foals infected with the high inoculum (12 ± 4.0; range 7.4–14) compared to foals infected with the low inoculum (2.5 ± 1.5; range 1.2–4.7). Proliferative responses to R. equi antigens as well as expression of mRNA for IL-2, IL-4, IL-10, and IFN-γ in BLN were not significantly different between the two groups. There was a tendency (P = 0.073) towards a higher IFN-γ/IL-4 ratio in the low inoculum group. This study demonstrates that the size of inoculum modulates the IgG subisotype response and possibly the cytokine profile of foals.