Pneumococcal antibody responses in elite swimmers

The ability of elite swimmers to mount an antibody response to the pneumococcal vaccine, Pneumovax 23, was assessed at the end of an intensive 12-week training programme. Antibody titres to six pneumococcal polysaccharide types were measured in 20 elite swimmers (10 male, 10 female) aged 17–23 years and 19 sedentary age- and sex-matched students (eight male, 11 female) aged 18–23 years. Blood samples were tested 14 days apart to assess the magnitude of the antibody response and changes in serum immunoglobulin isotypes and IgG subclasses. There were no significant differences in any of the pneumococcal antibody responses to the Pneumovax between swimmers and controls, and no gender effect, either before or after vaccination. The clinically adequate response to the vaccine was greatest for the pneumococcal serotype 4, which was 97% for the total study population. There were no significant correlations between the magnitude of any of the pneumococcal antibody responses and (i) changes in the scores for the swimmers’ international performance; (ii) infection rates in either swimmers or controls; (iii) any psychological variables, assessed by the Profile of Mood States (POMS) questionnaire for either swimmers or controls. Swimmers had significantly lower concentrations of serum IgG2 (P = 0·04) and IgG3 (P = 0·002) before pneumococcal vaccination. The swimmers had an increase in all immunoglobulin isotypes and IgG subclasses post-vaccination, suggesting a polyclonal response to the vaccine that was not observed in control subjects. The magnitude of the subclass responses after vaccination was significantly greater in swimmers compared with controls for IgG1 (P = 0·04), IgG3 (P = 0·04) and IgG4 (P = 0·01). The data indicated that elite swimmers undertaking an intensive training programme were capable of mounting an antibody response to pneumococcal antigens equivalent to that of age- and sex-matched sedentary control subjects, despite the swimmers having lower prevaccination levels of serum immunoglobulins.     

Systemic and Cutaneous Mucus Antibody Responses of Channel Catfish Immunized against the Protozoan Parasite Ichthyophthirius multifiliis

Fish acquire protective immunity against the ciliated protozoan parasite Ichthyophthirius multifiliis following sublethal infection or inoculation with I. multifiliis immobilization antigens (i-antigens). In both cases, parasite-immobilizing antibodies have been identified in sera and mucosal secretions. To investigate the kinetics of this immune response, antibody levels were determined by enzyme-linked immunosorbent assay (ELISA) in the sera and cutaneous mucus of channel catfish (Ictalurus punctatus) that were either infected with parasites or given a single injection of purified i-antigen (5.0 μg/fish) in Freund's incomplete adjuvant. At 5 weeks, infected and inoculated fish had a mean serum (1:80 dilution) antibody absorbance (A₄₀₅) value of 0.54 ± 0.17 and 0.35 ± 0.03, respectively, which were significantly higher (α = 0.05) than the pretreatment serum (1:80 dilution) antibody absorbance value of 0.24 ± 0.05. At 14 weeks, mean serum (1:80 dilution) ELISA absorbance values in the teo groups of fish increased to 0.79 ± 0.30 and 0.71 ± 0.24, respectively. In both groups of fish, antibody levels in cutaneous mucus (undiluted) were much lower than those in sera. Infected fish had detectable mucus (undiluted) antibody levels from 3 to 9 weeks, with the highest mean value (0.30 ± 0.07) occurring at 7 weeks. Although individual inoculated fish produced serum antibody absorbance values comparable to those seen in infected fish, the mean mucus antibody values in this group did not rise above pretreatment levels. I. multifiliis infection induced a transient mucosal antibody response that coincided with the resolution of infection. Whether elicited by infection or intraperitoneal injection of i-antigen, the serum and mucus antibody responses of channel catfish immunized against I. multifiliis did not occur synchronously.     

Development of a Novel In Vitro Assay (ALS Assay) for Evaluation of Vaccine-Induced Antibody Secretion from Circulating Mucosal Lymphocytes

We describe here a novel method for measuring in vitro antibody secretion from the tissue culture of human B lymphocytes in peripheral blood mononuclear cells (PBMC) after oral vaccination with a killed cholera vaccine. Enzyme-linked immunosorbent assay (ELISA) titers of the antibody secreted in the cell supernatant were determined. The validation results demonstrated that human PBMC remained viable and continued to secrete antibodies (total immunoglobulin A [IgA] and IgG) for up to 4 days of incubation at 37°C with 5% CO₂ in cell cultures. The secreted antibody concentration correlated positively with the PBMC concentration and incubation time in the tissue culture and correlated negatively with the storage time of the whole blood at room temperature. In vitro assay of secreting antibody in the lymphocyte supernatant (i.e., the ALS assay) is capable of the detecting specific antibody response after oral vaccination with a killed whole-cell-plus-B-subunit cholera vaccine (WC-B) in healthy adults in a phase I clinical trial. Postimmunization PBMC secreted antibodies to cholera toxin in the cell supernatants. Antibody production did not require any in vitro antigen stimulation. In the ALS assay, antigen-specific antibody titers of prevaccination samples were barely detectable, whereas serum antitoxin ELISA titers in background of prevaccine samples were significantly higher than the ALS titers. We conclude that, without any in vitro antigen stimulation after vaccination, PBMC secrete antibodies into the supernatants in the ALS assay. This assay can quantitatively measure the antigen-specific antibody production from the PBMC culture in postvaccination blood samples.     

Acute cytomegalovirus infection and the host immune response. I. Development and maintenance of cytomegalovirus (CMV) induced in vitro lymphocyte reactivity and its relationship to the production of CMV antibodies

Cytomegalovirus-induced lymphocyte reactivity (CMV-LR) of peripheral blood lymphocytes and antibodies against CMV-early (EA) and late (LA) antigens were studied in eighteen patients with documented acute CMV infections and seventy-six healthy persons. The development of a positive CMV-LR test lagged far behind the appearance of virus-specific antibodies. Positive CMV-LR tests were shown in all fourteen patients who could be tested twice or more. The median value of twenty-two tests in the acute phase (<50 days) was 269 d/min and the median of thirty tests in the post-illness phase (>50 days) was 1301 d/min (P<0·02). Once positive CMV-LR remained so during the follow-up period, up to 479 days after the onset of illness symptoms. In the meantime the LA and EA antibody response remained positive. Only two patients studied once showed negative CMV-LR responses while their serum contained high CMV-EA antibody titres. In CMV-LA seropositive healthy individuals who also possessed CMV-EA antibodies (LA+EA+) CMV-LR was higher (P<0·01) than in the LA seropositives who lacked EA antibodies (LA+EA−). The young LA+EA+ individuals, however, showed better (P<0·02) CMV−LR test results than the aged ones while their CMV antibody levels—especially the EA antibodies (P<0·02)—were lower. This phenomenon of increased CMV antibody production in relation to depressed CMV-LR is possibly caused by age-associated impairment of T lymphocyte function. The combined CMV-LR and EA antibody tests may provide useful means to study the specific immunological host/virus relationship in different clinical situations.     

Long-Term Survival of Streptococcus pneumoniae at Room Temperature on Dorset Egg Medium

Forty-five isolates of Streptococcus pneumoniae were inoculated on Dorset egg and supplemented Columbia agar base media, incubated overnight at 37°C, and then kept at room temperature (RT; 21°C) or 4°C. Long-term viability was best at RT for both media, with all isolates remaining viable on Dorset egg medium for 44 days; viability was 90 and 57% on Columbia agar base medium after 7 and 30 days. We recommend the use of Dorset egg medium for the maintenance of pneumococci at RT.     

Streptococcus pneumoniae antibody titres in patients with primary antibody deficiency receiving intravenous immunoglobulin (IVIG) compared to subcutaneous immunoglobulin (SCIG)

Intravenous immunoglobulin (IVIG) and subcutaneous immunoglobulin (SCIG) are effective in the treatment of patients with primary antibody deficiency disorders (PAD). The purpose of this study was to evaluate Streptococcus pneumoniae (Spn) antibody titres to 14 serotypes in patients receiving IVIG compared to SCIG and to correlate Spn antibody levels to clinical outcome. The doses of immunoglobulin (Ig)G/kg/month were similar in both IVIG and SCIG groups. In 11 patients treated with IVIG, Spn antibody titres were ≥ 1·3 μg/ml to 99·4 ± 2·1% of the 14 serotypes at peak IVIG but decreased to 66·9 ± 19·8% at trough IVIG. Loss of Spn titres ≥ 1·3 μg/ml was most frequent for Spn serotypes 1, 4, 9V and 23. This correlated with lower Spn antibody titres to these serotypes at peak IVIG compared to the other serotypes. In 13 patients treated with SCIG, Spn antibody titres were protective to 58·2 ± 23·3% of the serotypes 3–5 days after infusion, similar to trough IVIG. Similarly, the Spn serotypes with the least protective percentages were the same as the ones observed in trough IVIG. There were no annualized serious bacterial infections (aSBI) in either group. However, there were significantly decreased annualized other infections (aOI) in the SCIG group compared to the IVIG-treated group, 0·8 ± 0·7 versus 2·2 ± 1·2 infections/patient/year (P = 0·004). Breakthrough aOI did not correlate with protective or higher serum Spn antibody titres.     

Molecular Characterization of the Aeromonas hydrophila aroA Gene and Potential Use of an Auxotrophic aroA Mutant as a Live Attenuated Vaccine

The aroA gene of Aeromonas hydrophila SO2/2, encoding 5-enolpyruvylshikimate 3-phosphate synthase, was cloned by complementation of the aroA mutation in Escherichia coli K-12 strain AB2829, and the nucleotide sequence was determined. The nucleotide sequence of the A. hydrophila aroA gene encoded a protein of 440 amino acids which showed a high degree of homology to other bacterial AroA proteins. To obtain an effective attenuated live vaccine against A. hydrophila infections in fish, the aroA gene was inactivated by the insertion of a DNA fragment containing a kanamycin resistance determinant and reintroduced by allelic exchange into the chromosome of A. hydrophila AG2 by means of the suicide vector pSUP202. The A. hydrophila mutant AG2 aroA::Ka^r was highly attenuated when inoculated intraperitoneally into a rainbow trout, with a 50% lethal dose of >2 × 10⁸ CFU. The mutants were not recoverable from the internal organs after 48 h postinoculation. Immunohistochemical studies demonstrated that immunopositive materials, but not whole cells, reacting with a polyclonal antiserum against A. hydrophila were present in the kidney and spleen 9 days postinjection. Vaccination of rainbow trout with the AroA mutant as a live vaccine conferred significant protection against the wild-type strain of A. hydrophila.     

Tumor growth reduction is regulated at the gene level in Walker 256 tumor-bearing rats supplemented with fish oil rich in EPA and DHA

We investigated the effect of fish oil (FO) supplementation on tumor growth, cyclooxygenase 2 (COX-2), peroxisome proliferator-activated receptor gamma (PPARγ), and RelA gene and protein expression in Walker 256 tumor-bearing rats. Male Wistar rats (70 days old) were fed with regular chow (group W) or chow supplemented with 1 g/kg body weight FO daily (group WFO) until they reached 100 days of age. Both groups were then inoculated with a suspension of Walker 256 ascitic tumor cells (3×10⁷ cells/mL). After 14 days the rats were killed, total RNA was isolated from the tumor tissue, and relative mRNA expression was measured using the 2^-ΔΔCT method. FO significantly decreased tumor growth (W=13.18±1.58 vs WFO=5.40±0.88 g, P<0.05). FO supplementation also resulted in a significant decrease in COX-2 (W=100.1±1.62 vs WFO=59.39±5.53, P<0.001) and PPARγ (W=100.4±1.04 vs WFO=88.22±1.46, P<0.05) protein expression. Relative mRNA expression was W=1.06±0.022 vs WFO=0.31±0.04 (P<0.001) for COX-2, W=1.08±0.02 vs WFO=0.52±0.08 (P<0.001) for PPARγ, and W=1.04±0.02 vs WFO=0.82±0.04 (P<0.05) for RelA. FO reduced tumor growth by attenuating inflammatory gene expression associated with carcinogenesis.     

Effects of Sample Collection and Storage Methods on Antipneumococcal Immunoglobulin A in Saliva

Saliva contains components of both the mucosal and systemic immune systems. Variable flow rates, immunoglobulin proteases, and variation in collection and storage methods all introduce differences in the estimated concentrations of antibodies. We evaluated the effect of four collection methods and three storage protocols on the concentrations of immunoglobulin A (IgA) antibodies to pneumococcal capsular antigens 1, 5, 6B, and 14 and to pneumococcal surface adhesin A (PsaA) in saliva. Specimens were collected from 30 healthy Kenyan adults by collecting drool, by pipette suction, and with two commercial kits, OraSure and Oracol. Aliquots from each specimen were snap-frozen with glycerol in liquid nitrogen or stored for 4 to 8 h at +4°C either with or without the addition of protease enzyme inhibitors prior to storage at −70°C. Anticapsular IgA concentrations were not significantly different with different collection methods, but snap-freezing the specimens in liquid nitrogen led to concentrations 41 to 47% higher than those of specimens stored by the other methods (P < 0.0005).     

Phenotypic Differences of Cryptococcus Molecular Types and Their Implications for Virulence in a Drosophila Model of Infection

Compared to Cryptococcus neoformans, little is known about the virulence of the molecular types in Cryptococcus gattii. We compared in vitro virulence factor production and survival data using a Drosophila model of infection to further characterize the phenotypic features of different cryptococcal molecular types. Forty-nine different isolates were inoculated into wild-type flies and followed for survival. In vitro, isolates were assessed for growth at 30 and 37°C, melanin production, capsule size, resistance to H₂O₂, and antifungal susceptibility. A mediator model was used to assess molecular type and virulence characteristics as predictors of survival in the fly model. VGIII was the most virulent molecular type in flies (P < 0.001). At 30°C, VGIII isolates grew most rapidly; at 37°C, VNI isolates grew best. C. gattii capsules were larger than those of C. neoformans (P < 0.001). Mediator model analysis found a strong correlation of Drosophila survival with molecular type and with growth at 30°C. We found molecular-type-specific differences in C. gattii in growth at different temperatures, melanin production, capsule size, ability to resist hydrogen peroxide, and antifungal susceptibility, while growth at 30°C and the VGIII molecular type were strongly associated with virulence in a Drosophila model of infection.     

Effectiveness of low‐dose intravenous immunoglobulin therapy in minor primary antibody deficiencies: A 2‐year real‐life experience

Primary antibody deficiencies (PAD) are the most prevalent group of primary immunodeficiencies (PID) in adults and immunoglobulin replacement therapy (IRT) is the mainstay therapy to improve clinical outcomes. IRT is, however, expensive and, in minor PAD, clear recommendations concerning IRT are lacking. We conducted a retrospective real‐life study to assess the effectiveness of low‐dose IRT in minor PAD on 143 patients fulfilling European Society for Immunodeficiencies (ESID) diagnostic criteria for immunoglobulin (Ig)G subclass deficiency (IgGSD) or unclassified antibody deficiency (UAD). All patients were treated with intravenous low‐dose IRT (0.14 ± 0.06 g/kg/month). Immunoglobulin (Ig) classes and IgG subclasses were measured at baseline and after 1 year of IRT. The annual rate of total infections, upper respiratory tract infections (URTI), lower respiratory tract infections (LRTI) and hospitalizations was measured at baseline and after 1 and 2 years of IRT. After 1 year of IRT significant improvement was demonstrated in: (a) serum IgG (787.9 ± 229.3 versus 929.1 ± 206.7 mg/dl; p < 0.0001); (b) serum IgG subclasses (IgG1 = 351.4 ± 109.9 versus 464.3 ± 124.1, p < 0.0001; IgG2 = 259.1 ± 140 versus 330.6 ± 124.9, p < 0.0001; IgG3 = 50.2 ± 26.7 versus 55.6 ± 28.9 mg/dl, p < 0.002); (c) annual rate of total infections (5.75 ± 3.87 versus 2.13 ± 1.74, p < 0.0001), URTI (1.48 ± 3.15 versus 0.69 ± 1.27; p < 0.005), LRTI (3.89 ± 3.52 versus 1.29 ± 1.37; p < 0.0001) and hospitalizations (0.37 ± 0.77 versus 0.15 ± 0.5; p < 0.0002). The improvement persisted after 2 years of IRT. No significant improvement in URTI annual rate was noted in UAD and in patients with bronchiectasis. In conclusion, low‐dose IRT can improve clinical outcomes in UAD and IgGSD patients, providing a potential economical advantage over the standard IRT dose.     

Value of allohaemagglutinins in the diagnosis of a polysaccharide antibody deficiency

Polysaccharide antibody deficiency is characterized by a poor or absent antibody response after vaccination with an unconjugated pneumococcal polysaccharide vaccine. Allohaemagglutinins (AHA) are antibodies to A or B polysaccharide antigens on the red blood cells, and are often used as an additional or alternative measure to assess the polysaccharide antibody response. However, few studies have been conducted to establish the clinical significance of AHA. To investigate the value of AHA to diagnose a polysaccharide antibody deficiency, pneumococcal polysaccharide antibody titres and AHA were studied retrospectively in 180 subjects in whom both tests had been performed. Receiver operating characteristic curves for AHA versus the pneumococcal vaccine response as a marker for the anti-polysaccharide immune response revealed an area under the curve between 0·5 and 0·573. Sensitivity and specificity of AHA to detect a polysaccharide antibody deficiency, as diagnosed by vaccination response, were low (calculated for cut-off 1/4–1/32). In subjects with only low pneumococcal antibody response, the prevalence of bronchiectasis was significantly higher than in subjects with only low AHA (45·5 and 1·3%, respectively) or normal pneumococcal antibody response and AHA (2·4%). A logistic regression model showed that low pneumococcal antibody response but not AHA was associated with bronchiectasis (odds ratio 46·2). The results of this study do not support the routine use of AHA to assess the polysaccharide antibody response in patients with suspected immunodeficiency, but more studies are warranted to clarify the subject further.     

The relationship between the morphology of rosette-forming cells and their mode of rosette formation

Rats were injected in the footpad with sheep erythrocytes. The proportion of each morphological type of rosette-forming cell in the popliteal lymph node was measured, in vitro, performing the immunocyto-adherence technique at 4° and 37°. Among the different morphological types of rosette-forming cell, which appear after immunization, few plasmacytes form rosettes at 4°.

The number of rosettes formed at 37° was increased by the addition of anti-rat immunoglobulin (Ig) to aliquots of lymph node preparations from rats 7 days after immunization. This increase was accounted for by an increase in the number of rosette-forming plasmacytes and rosette-forming cells intermediate between small blasts and plasmacytes, and was thought to be due to the detection of cells secreting non-haemagglutinating antibody. The enhancing effect of anti-rat Ig at 37° contrasted with its inhibitory effect at 4°. The presence of 10^-3 M puromycin only slightly reduced the number of rosettes as compared with untreated control aliquots, but it prevented or reduced the enhancing effect of anti-rat Ig. The addition of anti-rat Ig to aliquots of lymph node preparations from rats 4 days after immunization reduced the number of rosettes formed as compared with control aliquots. It is considered that plasmacytes lack a surface associated antigen-binding receptor and depend for their rosette-forming ability on the secretion of antibody.

     

Blood monocyte activation in rheumatoid arthritis: increased monocyte adhesiveness, integrin expression, and cytokine release

Infiltration of the synovium by mononuclear cells, namely lymphocytes and monocytes, is one of the main features of rheumatoid arthritis (RA) and is considered to be responsible for the development of the disease. In this study in 31 consecutive patients with RA, we investigated whether peripheral blood monocytes exhibited markers of cellular activation related to cell migration. Using flow cytometry with the respective specific antibodies, we studied the expression of integrins CD11a, CD11b, CD11c, CD49d (VLA-4), and CD49e (VLA-5) on monocytes from patients with RA and from normal (N) subjects. IL-1β, IL-6, and tumour necrosis factor-alpha (TNF-α) production by cultured monocytes was measured by immunoassay. Adhesiveness of monocytes was studied on various surfaces (plastic, human fibronectin, gelatin-coated plasma, subendothelial matrix) and on cultured endothelial cells under basal conditions or after stimulation by IL-1β. An increased number of CD14⁺ monocytes (Mo) from RA patients expressed the CD11b molecule (RA Mo = 90·3%, N Mo = 83·4%, P < 0·005). The expression of CD11b on CD14⁺ monocytes was significantly increased in RA patients (median fluorescence intensity (FI): RA Mo = 145 (range 80–466) units; normal Mo = 95 (range 24–164) units; P < 0·003). Production of extracellular IL-1β and IL-6 by RA monocytes was significantly enhanced compared with monocytes from normal subjects (IL-1β: RA = 2·65 ± 0·91 ng/ml versusN = 1·35 ± 0·85 pg/ml, P < 0·05; IL-6: RA = 4·83 ± 0·90 ng/ml versusN = 2·40 ± 0·95 ng/ml, P < 0·05). Compared with normal monocytes, RA monocytes exhibited increased adhesion to the various surfaces studied (plastic, P < 0·01; fibronectin, P < 0·01; and gelatin-coated normal or RA plasma, P < 0·01) as well as to unstimulated (P < 0·01) and IL-1β-stimulated endothelial cells (IL-1β for 4 h, P < 0·05; IL-1β for 24 h, P < 0·05). In our study, blood monocytes from RA patients exhibited features of activation related to cell adhesion.     

C1q rs292001 polymorphism and C1q antibodies in juvenile lupus and their relation to lupus nephritis

C1q deficiency is related strongly to systemic lupus erythematosus (SLE), but very few and inconsistent studies explored the single nucleotide polymorphisms of the C1q gene in relation to juvenile SLE (jSLE) and lupus nephritis (LN). The objective of this study was to analyse whether C1q rs 292001 polymorphism is associated with SLE and disease phenotype, especially nephritis, and to investigate the relation between this polymorphism and clinical data, treatment outcome, serum level of C1q protein and antibodies. Typing of C1q rs292001 polymorphism using restriction fragment length polymorphism and measuring serum levels of C1q protein and antibodies by enzyme-linked immunosorbent assay (ELISA) were performed for 130 children with SLE and 208 healthy controls. The A allele of C1q rs292001 was associated with jSLE and LN (P = 0·005 and 0·013, respectively) and the AA genotype was associated with jSLE (P = 0·036). Low serum levels of C1q protein were found in jSLE and LN (P < 0·001 and 0·009, respectively), and these levels were increased after treatment in patients with LN (P = 0·009) and active renal disease (P = 0·027). Higher titres of C1q antibodies were found in patients with LN (P = 0·015) and correlated negatively with C1q protein level (P < 0·001) and patient age (P = 0·04). The A allele and AA genotype of C1q rs292001 can be considered a susceptibility risk factor and the GG genotype could be considered protective for jSLE and LN in the studied cohort of Egyptian children. Decreased serum levels of C1q protein and increased titres of C1q antibodies may be involved in the pathogenesis of jSLE, especially LN.     

Immune Responses and Safety after Dart or Booster Vaccination of Bison with Brucella abortus Strain RB51

One alternative for management of brucellosis in Yellowstone National Park bison (Bison bison) is vaccination of calves and yearlings. Although Brucella abortus strain RB51 vaccination protects bison against experimental challenge, the effect of booster vaccinations was unknown. This study characterized immunologic responses after dart or booster vaccination of bison with Brucella abortus strain RB51. In two studies, 8- to 10-month-old female bison were inoculated with saline (n = 14), hand vaccinated with 1.1 × 10¹⁰ to 2.0 × 10¹⁰ CFU of RB51 (n = 21), or dart vaccinated with 1.8 × 10¹⁰ CFU of RB51 (n = 7). A subgroup of hand vaccinates in study 1 was randomly selected for booster vaccination 15 months later with 2.2 × 10¹⁰ CFU of RB51. Compared to single vaccinates, booster-vaccinated bison had greater serologic responses to RB51. However, there was a trend for antigen-specific proliferative responses of peripheral blood mononuclear cells (PBMC) from booster vaccinates to be reduced compared to responses of PBMC from single vaccinates. PBMC from booster vaccinates tended to have greater gamma interferon (IFN-γ) production than those from single vaccinates. In general, dart vaccination with RB51 induced immunologic responses similar to those of hand vaccination. All vaccinates (single hand, dart, or booster) demonstrated greater (P < 0.05) immunologic responses at various times after vaccination than nonvaccinated bison. Booster vaccination with RB51 in early gestation did not induce abortion or fetal infection. Our data suggest that booster vaccination does not induce strong anamnestic responses. However, phenotypic data on resistance to experimental challenge are required to fully assess the effect of booster vaccination on protective immunity.     

Immune Responses of Bison and Efficacy after Booster Vaccination with Brucella abortus Strain RB51

Thirty-one bison heifers were randomly assigned to receive saline or a single vaccination with 10¹⁰ CFU of Brucella abortus strain RB51. Some vaccinated bison were randomly selected for booster vaccination with RB51 at 11 months after the initial vaccination. Mean antibody responses to RB51 were greater (P < 0.05) in vaccinated bison after initial and booster vaccination than in nonvaccinated bison. The proliferative responses by peripheral blood mononuclear cells (PBMC) from the vaccinated bison were greater (P < 0.05) than those in the nonvaccinated bison at 16 and 24 weeks after the initial vaccination but not after the booster vaccination. The relative gene expression of gamma interferon (IFN-γ) was increased (P < 0.05) in the RB51-vaccinated bison at 8, 16, and 24 weeks after the initial vaccination and at 8 weeks after the booster vaccination. The vaccinated bison had greater (P < 0.05) in vitro production of IFN-γ at all sampling times, greater interleukin-1β (IL-1β) production in various samplings after the initial and booster vaccinations, and greater IL-6 production at one sampling time after the booster vaccination. Between 170 and 180 days of gestation, the bison were intraconjunctivally challenged with approximately 1 × 10⁷ CFU of B. abortus strain 2308. The incidences of abortion and infection were greater (P < 0.05) in the nonvaccinated bison after experimental challenge than in the bison receiving either vaccination treatment. Booster-vaccinated, but not single-vaccinated bison, had a reduced (P < 0.05) incidence of infection in fetal tissues and maternal tissues compared to that in the controls. Compared to the nonvaccinated bison, both vaccination treatments lowered the colonization (measured as the CFU/g of tissue) of Brucella organisms in all tissues, except in retropharyngeal and supramammary lymph nodes. Our study suggests that RB51 booster vaccination is an effective vaccination strategy for enhancing herd immunity against brucellosis in bison.     

Effects of in vitro hyperthermia on proliferative responses and lymphocyte activity

Fever is induced by both exogenous products like endotoxin, and endogenous cytokines, most notably IL-1 and IL-6, and tumour necrosis factor (TNF). These mediators are believed to interact with the hypothalamus, to induce enhanced body temperature. However, little is known about the biological effects of fever on the function of the immune system. We here report that a 90-min pulse of mild hyperthermia (40°C) induces enhanced proliferation of peripheral blood mononuclear cells (PBMC). This proliferative response was completely inhibited by antibodies to MHC class II, which had no effect on mitogen-induced proliferation of PBMC. The enzyme-linked immunospot (ELISPOT) assay is a sensitive method for detection of single cells secreting antibodies or cytokines. A 90-min pulse of mild hyperthermia (40°C) induced a significantly enhanced immunoglobulin production in PBMC, as determined by ELISPOT, indicating B cell activation. The T cell cytokine pattern both with and without stimulation with hyperthermia differed between individuals. Enhanced interferon-gamma (IFN-γ) secretion was noted at 39–41°C. This IFN-γ response was inhibited by antibodies to MHC class II and thus was MHC class II-restricted and dependent on antigen-presenting cells. None of the individuals tested showed IL-4 response after stimulation with hyperthermia. These findings favour the notion that fever may play an important role in immune responses, and it is possible that fever may act as a physiological adjuvant, with effects on the immune system both in infection and inflammation of other origins.     

A Novel Solid Medium for Culturing Mycobacterium tuberculosis Isolates from Clinical Specimens

The laboratory diagnosis of tuberculosis usually relies on culture-based isolation of the causative Mycobacterium tuberculosis bacteria. We developed and evaluated the performance of MOD9, a new blood-free derivative of the MOD4 solid medium on which we previously reported for the isolation and culture of mycobacteria. First, inoculation of Lowenstein-Jensen medium with 21 M. tuberculosis isolates at 10⁵, 10³, and 10 CFU yielded colonies in 5.7 ± 1.5 days, 7.6 ± 0.8 days, and 10.8 ± 1.7 days versus 1.5 ± 0.4 days, 3.5 ± 0.6 days, and 4.9 ± 1 days in MOD9 (P < 0.05, Student''s t test). Further, the time to detectable growth of M. tuberculosis was measured on MOD9 and Lowenstein-Jensen media after duplicate inoculation of 250 clinical specimens decontaminated with 0.7% chlorhexidine. The contamination rate was 1.6% (4/250) on MOD9 versus 4.4% (11/250) on Lowenstein-Jensen medium (P = 0.11, Fisher''s exact test). Chlorhexidine-MOD9 yielded 38/250 (15.2%) isolates versus 32/250 (12.8%) isolates for the chlorhexidine-LJ (P = 0.5195, Fisher''s exact test). All together, eight M. tuberculosis isolates were cultured solely from chlorhexidine-MOD9, and two M. tuberculosis isolates were cultured solely from chlorhexidine-LJ. The time to detection was 9.8 ± 3.9 (range, 5 to 18) days for chlorhexidine-MOD9 versus 17.4 ± 5.9 (range, 10 to 35) days for chlorhexidine-LJ (P < 0.05, Student''s t test). These data indicate the superiority of the MOD9 medium over the standard LJ medium following chlorhexidine decontamination for the recovery of M. tuberculosis.     

Immunoglobulin A in serum: an old acquaintance as a new prognostic biomarker in idiopathic pulmonary fibrosis

Immunoglobulin (Ig)A is an important immunoglobulin in mucosal immunity and protects the lungs against invading pathogens. The production of IgA is regulated by transforming growth factor (TGF)-β, a versatile cytokine and key player in the pathogenesis of pulmonary fibrosis. TGF-β is up-regulated in patients with idiopathic pulmonary fibrosis (IPF), but difficult to use as a biomarker. The aim of this study was to evaluate the prognostic value of IgA in serum in patients with IPF. We examined IgA levels at time of diagnosis in 86 patients diagnosed with IPF. Mean serum IgA level in IPF is 3·22 g/l and regression analyses showed a significant association with mortality (hazard ratio = 1·445, P = 0·002). A significantly worse survival was found in patients with IgA serum levels > 2·85 g/l compared to patients with lower IgA serum levels (P = 0·003). These findings were confirmed in a duplication cohort. In conclusion, the level of IgA in blood is a promising prognostic marker in IPF and can be implemented easily in the hospital setting. Future studies are warranted to investigate if repeated measurements of serum IgA can further improve the performance of serum IgA as a prognostic marker.