A novel high-throughput assay to quantify the vaccine-induced inhibition of Bordetella pertussis adhesion to airway epithelia.

BACKGROUND: Pertussis or whooping cough is an acute respiratory illness caused by the Gram-negative pathogen Bordetella pertussis. Despite high vaccination coverage whooping cough is currently re-emerging in many developed countries. Although the causes of pertussis resurgence are matter of debate, emerging evidences suggest that acellular vaccines efficiently protect against the hallmark symptoms of pertussis disease but fail to prevent colonization. This presumably impacts on increased risk of bacterial transmission and consequent spread throughout the population. These evidences suggest that improved vaccines may be required for efficient bacterial clearance in the upper respiratory tract. Consequently, there is a need for novel bioassays to evaluate at pre-clinical or clinical level the impact of different vaccines on B. pertussis colonization. RESULTS: We developed a high-throughput bacterial adhesion inhibition (BAI) assay based on human respiratory cell lines and on live bacteria chemically conjugated to a fluorescent dye. Employing A549 cells as model, we evaluated the impact of antibodies elicited by acellular (aP) and whole cell (wP) vaccines on B. pertussis adhesion in vitro. Moreover, we settled the method also on polarized Calu-3 cells grown at air-liquid interface (ALI), showing that this assay can be extended to more complex cell models mimicking the airway epithelium. CONCLUSIONS: We proved that this method is a sensitive, rapid and reproducible system to evaluate the anti-adhesive properties of vaccine-induced antibodies and can be employed to assess improved pertussis vaccines.

No protective effects of high-dosage dietary zinc oxide on weaned pigs infected with Salmonella enterica serovar typhimurium DT104.

Twenty-eight-day-old weaned pigs were fed diets with a low (LZn), medium (MZn), or high (MZn) Zn concentration (50 to 80, 150, or 2,500 mg Zn/kg of diet, respectively) provided as zinc oxide (ZnO)(24 pigs per group). They were infected orally with Salmonella enterica serovar Typhimurium DT104 on day 32. Salmonellae were cultivated from feces (up to 42 days postinfection [dpi]) and organs (2 and 42 dpi). Activation of the adaptive systemic and mucosal immune systems was investigated by recording anti-Salmonella IgG levels and levels of B and T lymphocyte subpopulations in blood and gut-associated lymphatic tissue. Growth performance was recorded as well. Salmonellae were shed at higher levels and for longer periods in the HZn group (P < 0.05), with no differences in the tissues. At 2 dpi, the relative percentages of CD4(+) T helper cells (P < 0.01) and of CD2(+) T and NK cells (P < 0.01) in blood were reduced from the relative cell counts obtained at 0 dpi, irrespective of the Zn group. The lowest percentage of cytotoxic T cells was found 14 dpi in the HZn group relative to the MZn (P < 0.05) and LZn (P < 0.01) groups. Supplementation of the feed with 2,500 mg Zn/kg of diet immediately after weaning could positively affect the immune responses of piglets infected with Salmonella Typhimurium, but for a short period only. After 2 weeks, all positive effects disappeared, and rather negative effects, such as higher shedding of salmonellae, lower T cell frequencies, and worse performance, occurred. Thus, supplementation with ZnO at high levels in the pig industry should be limited to 2 to 3 weeks.

Immunogenicity and Efficacy of Pneumococcal Conjugate Vaccine (Prevenar13®) in Preventing Acquisition of Carriage of Pneumococcal Vaccine Serotypes in Tanzanian Children With HIV/AIDS.

In every year, up to one million children die due to pneumococcal disease. Children infected with Human Immunodeficiency Virus (HIV) are mostly affected, as they appear to have higher rates of pneumococcal carriage and invasive disease. Successful immunity is dependent on mounting a sufficient immune response to the vaccine. We conducted a double blinded crossover randomised controlled trial to determine the serum antibody response (≥4-fold and geometric mean concentration) to pneumococcal vaccine (PCV13) serotypes at 3 months after second vaccination. We also determined the number and proportion of children carrying new (not present at baseline) vaccine serotypes of S. pneumoniae isolated from nasopharynx at 6 months post initial vaccination in recipients of Prevenar13® compared with those given Haemophilus influenzae-type b (Hib) vaccine (control). The study was conducted at St Augustine's also known as Teule Hospital in Muheza, Tanga Tanzania. 225 HIV infected children aged 1-14 years were enrolled from Jan 2013 to Nov 2013 and randomised to Prevenar13® or Hib vaccines each given at baseline and 2-3 months later. Nasopharyngeal and serum samples were collected at baseline and 4-6 months later. Serotyping was done by Quellung Reaction using Staten antisera. Serum antibodies were ELISA quantified. The study revealed a non-significant reduction in the acquisition of new vaccine serotypes of S. pneumoniae in the recipients of PCV13 by nearly a third compared to those who received Hib vaccine. The vaccine efficacy was 30.5% (95% confidence interval [CI] -6.4-54.6%, P = 0.100)]. The antibody response was not enough to induce a 4-fold rise in GMC in 7 of the 13 vaccine serotypes. When combining the effects of preventing new acquisition and clearing existing vaccine type carriage, the overall efficacy was 31.5% (95% CI 1.5-52.4%, P = 0.045). In the PCV13 group, the proportion of participants carrying vaccine serotype was significantly lower after 2 doses of PCV13 (30%; 32/107), compared with the baseline proportion (48%; 51/107). The introduction of PCV13 targeting HIV-positive children in a setting similar to Tanzania is likely to be associated with appreciable decrease in the acquisition and carriage of pneumococci, which is an important marker of the likely effect of the vaccine on pneumococcal disease. Clinical Trial Registration: https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=335579, identifier ACTRN12610000999033.

The effect of Mass Drug Administration for trachoma on antibodies to Chlamydia trachomatis pgp3 in children.

A serologic test for antibodies to chlamydia may be a useful tool for trachoma surveillance. However, little is known about the longitudinal stability of antibody status, especially following Mass Drug Administration (MDA), which is critical to understanding serostatus in trachoma-endemic areas. A longitudinal cohort of 1908 children ages 1-9 years in Tanzania from 50 communities were followed at baseline and for 6 months after MDA. They were evaluated for clinical trachoma, conjunctival swabs were tested for chlamydial infection using GeneXpert platform, and blood spots were collected on filter paper and dried to test for antibodies to Chlamydia trachomatis pgp3 using the Luminex platform. 6.3% of children in the study had infection, and coverage with MDA was 97%. 670 (35%) were sero-positive for pgp3 antibodies at baseline, and 4.0% of these seroreverted to negative following MDA. Of those seronegative at baseline, 3.6% seroconverted. The individual change in log median fluorescence intensity(MFI-BG) values was -0.15 overall (p < .001). Seroconversion rates were lower following MDA and seroreversion rates were slightly higher compared to rates in this same cohort in the absence of MDA. MDA has a small effect on reduction of MFI-BG.

Effects of anticoagulants and Ficoll on human serum antibody reactivities and functions against Mycobacterium tuberculosis.

The ability to utilize leftover samples containing anticoagulants or Ficoll would provide substantial opportunities for future antibody and biomarker studies. Some anticoagulants might influence antibody reactivity against pathogens, but comprehensive studies investigating effects in the context of TB are lacking. We enrolled 24 individuals with and without history of M. tuberculosis and/or HIV-infection and investigated TB antibody reactivities, function, and other host protein biomarkers in simultaneously obtained serum and plasma from serum separation, EDTA, heparin, acid citrate dextrose (ACD), or mononuclear cell preparation (CPT™) tubes which contain heparin and Ficoll. Antibody isotype reactivities to two mycobacterial antigens, as well as phagocytosis of M. tuberculosis, correlated strongly and significantly between serum and plasma, irrespective of type of anticoagulant or Ficoll present (r ≥ 0.85, p < 0.0001). However, the presence of ACD resulted in slightly lower values than those obtained with serum in both indirect (antibody reactivities to mycobacterial antigens) and Sandwich ELISAs (soluble CD14 measurements). Our data demonstrate that leftover plasma, regardless of containing anticoagulants or Ficoll, can be used in TB antibody or other host protein biomarker studies but suggest the value of a correction factor when using ACD plasma interchangeably with serum in antibody binding studies.

Antibodies to Helicobacter pylori and CagA protein are associated with the response to antibacterial therapy in patients with H. pylori-positive API2-MALT1-negative gastric MALT lymphoma.

The aim of this study was to clarify predictive factors for response to eradication therapy in cases of Helicobacter pylori (H. pylori)-positive API2-MALT1-negative gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Sixty-six patients who were examined for H. pylori infection and the presence of the API2-MALT1 chimeric transcript and who underwent H. pylori eradication therapy as first-line therapy, were enrolled in this study. Immunohistochemical markers (p53, Ki-67, and BCL10), microsatellite instability, loss of heterozygosity, serum levels of antibodies (anti-H. pylori and anti-CagA), and markers for gastritis (gastrin and pepsinogens) were examined, and the results were compared between patients whose tumors regressed completely after eradication therapy (responders) and patients whose tumors did not regress (non-responders). Of the 66 patients with localized gastric MALT lymphoma, 47 (71.2%) showed complete remission after eradication therapy. None of the H. pylori-negative (n = 9) and/or API2-MALT1-positive (n = 7) patients responded to antibacterial treatment. Of 44 patients with H. pylori-positive API2-MALT1-negative gastric MALT lymphoma, 38 (86.4%) showed complete remission after eradication therapy. Titers of antibodies against H. pylori and CagA protein were significantly higher in the responders than in the non-responders (P = 0.0235 and 0.0089, respectively). No significant difference between the groups was observed for the other factors. In conclusion, measurement of titers of serum antibodies to H. pylori and CagA protein may be useful for predicting the response to eradication therapy in patients with H. pylori-positive API2-MALT1-negative gastric MALT lymphoma.

Mycoplasma pneumoniae as a cause of vulvar ulcers in a non-sexually active girl: a case report.

BACKGROUND: Non-sexually active young females very rarely develop genital ulcers. Such ulcers pose a diagnostic challenge as well as physical and emotional distress for patients and family; therefore, the search for their etiology requires exhaustive investigation. Several viruses such as Epstein-Barr virus have been associated with this entity; however, Mycoplasma pneumoniae has rarely been linked to such ulcers in the literature. We present a case of vulvar ulcers in a non-sexually active young girl during the course of pneumonia caused by Mycoplasma pneumoniae. CASE PRESENTATION: A 10-year-old non-sexually active girl of cypriot origin presented at a hospital with fever, dry cough, and acute vulvar ulcers. Laboratory investigations as well as imaging studies revealed Mycoplasma pneumoniae as the cause of her pneumonia and acute vulvar ulcers. CONCLUSIONS: Although a rare cause of vulvar ulcers, Mycoplasma pneumoniae should be considered in the differential diagnosis of acute vulvar ulcers coexisting with respiratory symptoms.

Monensin might protect Ontario, Canada dairy cows from paratuberculosis milk-ELISA positivity.

Our objective was to define the role of monensin sodium in protecting cows from being milk-ELISA positive for paratuberculosis in Ontario, Canada dairy herds. In total, 4933 dairy cows from 94 herds were enrolled in this cross-sectional study. Forty-four of the enrolled herds were selected purposively by their herd veterinarian and another 50 herds were randomly selected from a local milk production-recording agency. A herd-management survey was completed on each farm during the months of May through August 2003. During this same time-period, composite milk samples were collected from all lactating cows and tested with a milk-ELISA for antibodies to Mycobacterium avium subspecies paratuberculosis. Analyses were stratified according to the paratuberculosis history of the herds. In the 48 herds in which paratuberculosis had not been diagnosed before, the use of calf hutches and monensin in milking cows were both associated with reduced odds of a cow testing positive (OR=0.19 and 0.21, respectively). In the 46 herds with a prior history of paratuberculosis, feeding monensin to the breeding-age heifers was associated with decreased odds of a cow testing positive (OR=0.54). Monensin use might be associated with milk-ELISA positivity, but its impact on the transmission of paratuberculosis remains unknown.

Effect of azithromycin treatment on endothelial function in patients with coronary artery disease and evidence of Chlamydia pneumoniae infection.

BACKGROUND: It has been suggested that infection with Chlamydia pneumoniae(CPn) can trigger inflammatory mechanisms that may in turn impair vascular endothelial function. The aim of the present study was to assess whether treatment with the macrolide antibiotic azithromycin improves endothelial function in patients with coronary artery disease and antibodies positive to CPn. METHODS AND RESULTS: We carried out a randomized, prospective, double-blind, placebo-controlled trial in 40 male patients (mean age, 55+/-9 years) with documented coronary artery disease and positive CPn-IgG antibody titers. After baseline evaluation, patients were randomized to receive either azithromycin or placebo for 5 weeks. Flow-mediated dilation (FMD) of the brachial artery and E-selectin, von Willebrand factor, and C-reactive protein (CRP) levels were assessed at study entry and at the end of the treatment period. Our results showed that patients who received azithromycin had a significant improvement in FMD (mean change, 2.1+/-1.1%; P<0.005). In contrast, FMD was not significantly changed in the placebo group (mean change, -0.02+/-0.2%, P=0.64). Azithromycin therapy also resulted in a significant decrease of E-selectin and von Willebrand factor levels. CRP levels were not significantly altered by treatment with either azithromycin or placebo. Beneficial effects of azithromycin treatment were independent from the presence of low (<1:32) or high (> or =1:32) CPn antibody titers. CONCLUSIONS: Our findings indicate that treatment with azithromycin has a favorable effect on endothelial function in patients with documented coronary artery disease and evidence of CPn infection irrespective of antibody titer levels. Whether these favorable actions of antibiotic treatment will translate into a beneficial effect on atherogenesis and cardiac events needs further investigation.

Sex and low-level sampling stress modify the impacts of sewage effluent on the rainbow trout (Oncorhynchus mykiss) immune system.

The objective of the present study was to investigate the influence of chronic exposure to municipal sewage treatment effluent at environmentally relevant concentrations on immune parameters in rainbow trout (Oncorhynchus mykiss), including the assessment of potential differences in reactivity between sexually mature male and female fish. Trout were exposed to 1.5 and 15% (v/v) secondary treated municipal sewage effluent for 32 weeks. Fish were injected intra-peritoneally either with inactivated Aeromonas salmonicida to simulate an infection or with PBS as control for this immune challenge 6 weeks prior to sampling. Exposure to effluent resulted in a decrease in A. salmonicida-specific serum antibody level and blood lymphocyte numbers in mature females, but not in male fish. Injection of A. salmonicida resulted in enhanced serum lysozyme activity in mature male trout, which were not exposed to effluent. This stimulating effect of A. salmonicida could not be found in effluent-exposed trout, again potentially revealing a suppressive effect of the effluent. An influence of sampling fish on two consecutive days was observed in many immune parameters, most likely reflecting handling stress. Leucocyte and lymphocyte numbers in peripheral blood were consistently lower in male and female fish on the second sampling day. Phagocytosis in head kidney macrophages from male trout was also influenced by sampling day, whereby a stimulation of this reaction occurred on the second day of sampling. Liver mixed function oxygenase activity was found to be enhanced in mature male trout exposed to 15% effluent. In conclusion, the study showed, that exposure to sewage treatment plant effluent, in surface water relevant concentrations, can lead to potentially adverse effects on selected immune reactions in rainbow trout. However, this study also demonstrated that both handling stress and the sex of mature fish have distinct influences on the immune response detected in male and female fish and are likely to influence measured immune parameters to the extent that subtle effluent induced changes may be difficult to detect.

Association of Chlamydia pneumoniae antibody with the cholesterol-lowering effect of statins.

To investigate the association between Chlamydia pneumoniae (C. pneumoniae) infection and atherosclerosis, we compared the effect of lipid-lowering drugs on carotid intima-media thickness (IMT) between patients who were positive and negative for C. pneumoniae antibodies. A total of 165 asymptomatic hypercholesterolemic patients were randomized to probucol (500 mg per day, n = 82) or pravastatin (10 mg per day, n = 83) and followed for 2 years. The 2-year change of IMT in the common carotid artery was the primary endpoint, while mean IMT change and major cardiovascular events were secondary endpoints. C. pneumoniae antibodies (IgA and IgG) were measured by enzyme-linked immunosorbent assay. The 50 patients without C. pneumoniae antibodies showed significant reduction of IMT progression (-19%), while no significant change of IMT was noted in the 115 antibody-positive patients (-6%). Significant inverse associations were found between the reduction of IMT progression and the C. pneumoniae IgA- and IgG-antibody index (P < 0.01 and 0.01, respectively). No significant differences in the reduction of serum total-cholesterol and LDL-cholesterol were found between antibody-positive and -negative patients. There was no significant difference of efficacy between probucol and pravastatin. These observations suggest that C. pneumoniae infection reduces the effect of lipid-lowering therapy on carotid atherosclerosis and that this organism may play a role in the progression of atherosclerosis.

Noninvasive stool antigen assay can effectively screen Helicobacter pylori Infection and assess success of eradication therapy in hemodialysis patients.

Helicobacter pylori (H pylori) stool antigen (HpSA), serological antibody against H pylori (immunoglobulin G [IgG]), and urea breath test (UBT) are noninvasive methods used to detect H pylori infection that can allow a patient to avoid the discomfort and risk of invasive endoscopy. However, because the UBT has proven not highly reliable in patients with end-stage renal disease (ESRD), this study attempts to investigate the diagnostic efficacy of HpSA and IgG for H pylori detection in 80 patients with ESRD and 80 dyspeptic patients without renal function impairment as a control group. All patients in both study groups underwent panendoscopy to obtain gastric biopsy specimens for histological examination and H pylori culture. With H pylori infection defined as a positive result on either histological examination or culture, we evaluated the reliability of HpSA and serum IgG in detecting H pylori infection. Forty of the patients with ESRD (50%) and 48 patients in the control group (60%) were proven to be infected with H pylori. To eradicate H pylori infection, these patients were administered a 1-week course of triple therapy. To evaluate the success of H pylori eradication, 38 patients in the ESRD group and 44 patients in the control group underwent a follow-up endoscopy and provided stool samples for HpSA 6 to 8 weeks later. Success of H pylori eradication was found in 86.8% of the patients with ESRD (33 of 38 patents) and 84.1% of the control patients (37 of 44 patients). Before therapy, HpSA for H pylori detection was 97.5% sensitive and 97.5% specific in patients with ESRD, as effective as that in the control group. After therapy, HpSA was 100% sensitive and more than 96% specific to detect the failure of H pylori eradication therapy in both the ESRD and control groups. Conversely, the use of IgG as a screening method for H pylori infection proved to be less effective because it showed a sensitivity of 87.5% and specificity of 80% in this study. Monitoring the success of triple therapy, IgG had a specificity of only 21.9% in the ESRD group and 24.3% in the control group. In summary, HpSA is a noninvasive and reliable tool to screen H pylori infection before therapy and assess the success of eradication therapy in patients with ESRD.

Effect of beta propiolactone on specific antibody measurements by ELISA.

Serum samples from 30 HIV seronegative patients were treated with beta propiolactone (BPL) to determine whether BPL interfaces with ELISA for specific antibodies against protein and carbohydrate antigens. BPL had no discernible effect on specific antibody measurements by ELISA. With the measuring need for specific antibody measurements in the management of HIV seropositive patients, it is reassuring that this laboratory safety measure does not impair the reliability of results.

Vomitoxin-induced dysregulation of serum IgA, IgM and IgG reactive with gut bacterial and self antigens.

The effect of dietary vomitoxin exposure on immunoglobulins that react with naturally occurring gut bacterial and self antigens was assessed in the B6C3F1 mouse. Ingestion of 25 ppm vomitoxin for 4 and 8 wk resulted in significantly elevated total IgA but depressed total IgG and IgM in serum when compared with control mice fed semi-purified diet only. IgA specific for phosphorylcholine (PC) and inulin (haptens associated with intestinal bacteria) increased significantly in mice fed vomitoxin whereas IgM with the identical specificity decreased. When sera were assessed for autoantibodies recognizing DNA and bromelated mouse red blood cells (MRBC), vomitoxin-exposed mice exhibited elevated specific IgA as compared with controls. This occurred together with decreases in DNA-specific IgG and IgM, and decreases in MRBC-specific IgM. Additionally, vomitoxin exposure did not enhance the specific serum IgA response to orally administered trinitrophenylated sheep red blood cells (TNP-SRBC), but significantly depressed TNP-specific serum IgG. The results suggest that hyperelevation of total and specific serum IgA for oral and self antigens occurs during vomitoxin feeding and that may be coupled with down-regulation of total and specific IgM or IgG. These effects could be contributory to the capacity of vomitoxin to induce IgA immune complex glomerulonephritis.

Persistence of antichlamydial antibodies after treatment of acute salpingitis with doxycycline.

The effect of treatment with doxycycline on serum IgG and IgA antichlamydial antibodies was evaluated in 33 women who had had acute salpingitis associated with high titers of serum IgG (> or = 1:128) and/or IgA (> or = 1:16) antichlamydial antibodies. Overall, 29 women (87.9%) remained with high titers of IgG and/or IgA antibodies. No change or insignificant change in IgG antibody titer was demonstrated in 21 women (63.6%) and in IgA antibody titer in 21 women (63.6%). Positive seroconversion or a significant increase (> or = 4-fold) in IgG antibody titer was demonstrated in eight women (24.2%) and in IgA antibody titer in six women (18.1%). Negative seroconversion or a significant decrease in IgG antibody titer was demonstrated in four women (12.1%) and in IgA antibody titer in six women (18.1%). It is concluded that in most patients who had acute salpingitis associated with pretreatment high titers of serum antichlamydial antibodies, posttreatment titers may remain high even if treatment with doxycycline results in complete resolution of clinical signs and symptoms of the disease.

Immunoadjuvant potential of Asparagus racemosus aqueous extract in experimental system.

The immunoadjuvant potential of Asparagus racemosus (Willd.) Family (Liliaceae) aqueous root extract was evaluated in experimental animals immunized with diphtheria, tetanus, pertussis (DTP) vaccine. Immunostimulation was evaluated using serological and hematological parameters. Oral administration of test material at 100 mg/kg per day dose for 15 days resulted significant increase (P = 0.0052) in antibody titers to Bordtella pertussis as compared to untreated (control) animals. Immunized animals (treated and untreated) were challenged with B. pertussis 18323 strain and the animals were observed for 14 days. Results indicate that the treated animals did show significant increase in antibody titers as compared to untreated animals after challenge (P = 0.002). Immunoprotection against intra-cerebral challenge of live B. pertussis cells was evaluated based on degree of sickness, paralysis and subsequent death. Reduced mortality accompanied with overall improved health status was observed in treated animals after intra-cerebral challenge of B. pertussis indicating development of protective immune response. Present study indicates applications of test material as potential immunoadjuvant that also offers direct therapeutic benefits resulting in less morbidity and mortality.

Increased serum IgM class anti-lipid A antibody and therapeutic effect of ursodeoxycholic acid in primary biliary cirrhosis.

BACKGROUND/AIMS: There is as yet no explanation for the increased serum IgM level observed in patients with primary biliary cirrhosis. METHODOLOGY: The serum IgM class anti-lipid A antibody was determined in patients with primary biliary cirrhosis using Elisa and Western blot techniques. RESULTS: Using enzyme-linked immunosorbent assay, we found the concentration of serum IgM class anti-lipid A antibody was the highest in 21 patients with PBC as compared to 29 patients with other liver diseases and 19 controls. Using western blotting, IgM class anti-lipid A antibody was detected as a clear band in patients with primary biliary cirrhosis. The level of this antibody correlated significantly (p < 0.005) with that of total IgM. Treatment with ursodeoxycholic acid improved not only the routine biochemical profile but also the level of IgM class anti-lipid A antibody (p < 0.005). CONCLUSIONS: Since the lipid A plays a primary role in the biological activities of lipopolysaccharide (a component of gram-negative bacteria), bacterial antigen may participate in the elevation of serum IgM levels in patients with primary biliary cirrhosis. Ursodeoxycholic acid treatment may improve the cholestasis, and this may have altered the response to stimulation by gut derived bacterial antigens.

Intranasal immunization with lipoteichoic acid and cholera toxin evokes specific pharyngeal IgA and systemic IgG responses and inhibits streptococcal adherence to pharyngeal epithelial cells in mice.

OBJECTIVE: Streptococcus (S.) pyogenes is common cause of acute tonsillitis. Lipoteichoic acid (LTA), which is a common constitute of the cell surface of most gram positive bacteria, is known to act as a substance of bacterial site for adherence to epithelium and antiserum to LTA is reported to inhibit bacterial attachment to epithelial cells in vitro. Cholera toxin subunit B (CT-B) is known to be a mucosal adjuvant. The purpose of this study is to investigate whether intranasal immunization with LTA and CT-B may be a possible candidate for vaccine formulation. METHODS: Six-week-old male BALB/c mice were assigned to three experimental groups, mice immunized with LTA and CT-B, with LTA alone and with phosphate buffered saline (PBS) as a control. Immunizations were performed intranasally every 2 days for 2 weeks in every group. At the 21 days after immunization, sera, pharyngeal washings and pharyngeal epithelial cells were taken. The levels of serum IgG and pharyngeal IgA antibodies to LTA were measured by enzyme-linked immunosorbent assay (ELISA). The adherence rates of S. pyogenes pretreated by pharyngeal washings to pharyngeal epithelial cells from the mice were determined by in vitro adherence assay. RESULTS: The serum anti-LTA IgG antibody levels of either mice immunized with LTA and CT-B or mice immunized with LTA alone were significantly higher than those of mice administered with PBS alone. The pharyngeal anti-LTA IgA antibody levels of the mice immunized with LTA and CT-B were significantly higher than those of either mice with LTA alone or mice with PBS alone. The streptococcal adherence rates to pharyngeal epithelial cells were significantly decreased by pretreatment with pharyngeal washings from the mice immunized with LTA and CT-B as compared with pretreatment with those from either mice with PBS or mice with LTA alone. CONCLUSIONS: These data shows that intranasal immunization with LTA and CT-B evokes a good pharyngeal IgA response as well as systemic IgG response to LTA and inhibits streptococcal adherence to pharyngeal epithelial cells, suggesting that intranasal immunization with LTA and CT-B may be an effective approach to prevent streptococcal tonsillitis.

[Effect of antibiotics and antibody on phagocytic bactericidal activity of polymorphonuclear neutrophils of Bordetella pertussis].

The phagocytic bactericidal activity of the polymononucler neutrophils (PMNs) that were collected from healthy volunteer with and without antibody against Bordetella pertussis was investigated. Furthermore, these activity against B. pertussis under observing penicillins or macrolides antibiotics was investigated. Although no efficacy to B. pertussis strain by the PMNs in serum without antibody, but the viable cells of B. pertussis decreased to 1/1,000 1 hr after incubation and was not detected after 4 hrs. In particular, the viable cells of B. pertussis by the PMNs in serum with antibody was markedly reduced when azithromycin was present. These results suggests that the synergistic action of macrolide antibiotics and antibody-mediated phagocytic bactericidal activity on B. pertussis may have clinical relevance.

Anti-idiotype antibodies and circulating immune complexes after immunisation with tetanus-toxoid.

OBJECTIVES: To determine if anti-idiotype antibodies and circulating immune complexes in individuals before and after immunisation with tetanus toxoid play a role in the immune response. DESIGN: A study of individuals who were administered a single dose of tetanus toxoid (TT) and who were unimmunized. SETTING: Out patient departments of a large public hospital in Bombay, India. SUBJECTS: Thirty eight individuals pre-immunisation and forty five individuals post-immunisation with tetanus toxoid, tested at 1, 3, 6, and 12 months. MAIN OUTCOME MEASURE: Development of anti-tetanus anti-idiotype antibodies and circulating immune complexes. RESULTS: Pre-immunisation cases did show presence of anti-tetanus antibodies but in lower titres than post-immunisation up to six months, after which there was a reduction. Specific anti-idiotype antibodies were detected in 19 cases. One and three months after immunisation more cases had high titre antibodies and circulating immune complexes, though after six months, there was a fall in anti-tetanus antibody titres. Circulating immune complexes were seen in those samples having anti-idiotype antibodies. CONCLUSIONS: Though a significant rise in anti-tetanus antibody anti-idiotype antibodies, protective levels in mice and circulating immune complexes are seen after immunisation with TT it lasts for six months. When followed up for a period of one year it is observed that in cases having auto anti-idiotype antibodies, the anti-tetanus antibodies are maintained for a longer period.