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The aggregation of non-serotypable Haemophilus influenzae (NTHI) by whole saliva from patients with chronic obstructive lung disease (COLD) was investigated. Significant differences were observed between salivary aggregating activity of a control and COLD population (P < 0.001). Saliva from patients less prone to acute exacerbations had a greater capacity to aggregate bacteria compared with saliva from patients with a predilection to infection. The mechanism of saliva-mediated aggregation of NTHI was investigated and shown to be related to lysozyme content. Lysozyme activity in saliva was measured by the turbidimetric technique and results showed that patients with chronic bronchitis had increased levels of salivary lysozyme, with a subpopulation within the non-infection-prone group having greater amounts. A significant difference was observed in salivary lysozyme between controls and non-infection-prone (P < 0.005) and infection-prone (P < 0.05) patients, respectively: the non-infection-prone patients having significantly (P < 0.005) more than the infection-prone patients. There was significant correlation (r = 0.742, P < 0.001) between salivary aggregation of NTHI and lysozyme activity. Chromatographically purified human lysozyme had a similar aggregation profile to that of saliva. There was no difference in serum and saliva lactoferrin concentrations between groups, but there was a significant increase (P < 0.05) in serum lysozyme concentration in the non-infection-prone group. This study suggests that the level of salivary lysozyme derived from macrophages may play an important role in determining resistance or susceptibility to acute bronchitis.

This review focuses on saliva as a measure of mucosal immunity in man. The review will cover studies of parameters that modify the early ontogeny patterns of mucosal immunity and the impact of infections and physiological variables on the human mucosal immune system. The most significant modifiers of human mucosal immunity are events that occur in the neonatal maturation period and, later in life, the interplay between the immune system and the neuroendocrine systems. IgA antibodies are the predominant isotype involved in the human mucosal immune response and are important for protection at mucosal surfaces. The level of IgA in mucosal secretions is modified by antigenic stimulation as well as by many physiological variables. Studies have also revealed that IgM plays a significant immunoregulatory role at mucosal surfaces, particularly during episodes of infection or stress. The detection patterns of IgD in saliva of neonates suggests a role for IgD in the initial maturation process of mucosal immunity. The role of IgG at mucosal surfaces is unclear and although IgG may play a compensatory role in IgA deficiency, the detection of high levels of IgG in saliva appears to be associated with periods of increased membrane permeability.

The impact of long-term training on systemic and mucosal immunity was assessed prospectively in a cohort of elite swimmers over a 7-month training season in preparation for national championships. The results indicated significant suppression (P < 0.05) of serum IgA, IgG and IgM and salivary IgA concentration in athletes associated with long-term training at an intensive level. There was also a trend towards lower IgG2 subclass levels in serum in athletes compared with controls (P = 0.07). There were no significant changes in numbers or percentages of B or T cell subsets, but there was a significant fall in natural killer (NK) cell numbers and percentages in athletes over the training season (P < 0.05). After individual training sessions there was a significant decrease in salivary IgA levels for athletes compared with controls (P = 0.002). In athletes there was a downward trend in salivary IgA levels over the 7-month training period in both the pre-exercise (P = 0.06) and post-exercise samples (P = 0.04). There were no significant trends in salivary IgG levels over the study period in either athletes or controls. The only significant change in salivary IgM levels was an increase in detection rate in the pre-competition phase in athletes (P = 0.03). The study suggests that training of elite athletes at an intensive level over both short- and long-time frames suppresses both systemic and mucosal immunity. Protracted immune suppression linked with prolonged training may determine susceptibility to infection, particularly at times of major competitions.

Pseudomonas aeruginosa, an opportunistic bacterial pathogen, is a major course of morbidity and mortality in subjects with compromised respiratory function despite the significant advances in therapeutic practices. The bacteria produces an armoury of products which modify its infective niche to ensure bacterial survival. The role of antibody in protection against pulmonary infection remains poorly defined. Protection appears to be associated with opsonizing antibody whilst some other antibody responses may be deleterious and promote further lung damage. Cell mediated responses are clearly important in protection against infection. This review proposes a vaccine strategy aimed at enhancing specific T cell responses in the lung which, though T cell-derived cytokines, drive the recruitment of neutrophils to the lung and the subsequent activation of these cells results in the clearance of bacteria from the lung.

The ability of mucosally administered antigen to provide protection against Streptococcus equi ('Strangles') infections in horses was examined. First, an enzyme linked immunosorbent assay (ELISA) was developed to detect the immune status of horses to S. equi. This assay was used to select Strangles-naive horses for the study and also to monitor their response to immunisation. Potential vaccine candidates were: (a) orally administered paraformaldehyde killed S. equi; (b) intraperitoneally (IP) administered paraformaldehyde killed S. equi in a non-inflammatory adjuvant; (c) orally administered live avirulent S. equi; (d) orally administered microencapsulated streptococcal M protein. The latter three preparations were first assessed in a rat model, using rate of lung bacterial clearance following intratracheal inoculation of live virulent bacteria as an indication of efficacy. Candidates (a) and (b) were then assessed in an equine model. IP immunisation of horses was shown to effectively induce production of specific antibody in mucosal and systemic sites. Four weeks after initial immunisation, horses were challenged intranasally with live virulent S. equi. Both groups of immunised horses demonstrated partial protection following vaccination. Of the IP immunised horses, only two out of four developed clinical signs of Strangles following live challenge. The orally immunised horses all developed submandibular abscesses containing S. equi. However, none of the immunised horses became as ill as the control horses in terms of fever, anorexia, loss of condition and general malaise.

Nontypeable Haemophilus influenzae (NTHi) is a common cause of infection of the respiratory tract in children and adults. The search for an effective vaccine against this pathogen has focused on components of the outer membrane, and peptidoglycan-associated lipoprotein P6 is among the proposed candidates. This study investigated the immunogenicity of P6 in a rat respiratory model. P6 was purified from two strains of NTHi, one capsule-deficient strain and an H. influenzae type b strain, and assessed for clearance of both homologous and heterologous bacterial strains following mucosal immunization. A protective immune response was determined by enhancement of pulmonary clearance of live bacteria and an increased rate of recruitment of phagocytic cells to the lungs. This was most effective when Peyer's patch immunization was accompanied by an intratracheal (IT) boost. However, the rate of bacterial clearance varied between strains, which suggests some differences in anti-P6 immunological defenses recognizing the expression of the highly conserved P6 lipoprotein on the bacterial surface in some strains. P6-specific antibodies in both serum and bronchoalveolar lavage fluid were cross-reactive and did not differ significantly in strain specificity, demonstrating that difference in clearance was unlikely due to differences in P6-specific antibody levels. Serum homologous and heterologous P6-antibody was bactericidal against NTHi even when enhanced clearance had not been observed. Peyer's patch immunization induced P6-specific CD4+ T-helper cell proliferation in lymphocytes isolated from the mesenteric lymph nodes. An IT boost increased the level of P6-specific antibodies in serum and bronchoalveolar lavage fluid, and P6-specific mesenteric node lymphocyte proliferation. Cells from rats immunized with P6 demonstrated proliferation following stimulation with P6 from nonhomologous strains; however, there was some variation in proliferative responses to P6 from different strains in lymphocytes isolated from animals immunized with killed bacteria. The increase in P6-specific antibodies and T-helper cell responses following an IT boost correlated with an increased rate of recruitment of phagocytic cells and enhanced bacterial clearance of both homologous and heterologous bacteria in the lungs. The data suggests that P6 has the potential to afford protection against pulmonary infection by NTHi following the induction of effective antigen-specific B- and T-cell responses in mucosal tissues.

Rats intestinally immunized with whole killed non-typable Haemophilus influenzae clear this organism from the lungs faster than non-immunized rats. This study investigated the role of antibody and phagocytes in the clearance mechanism. First, dose-response experiments demonstrated that while lowering the dose of non-typable H. influenzae reduced the level of detectable specific antibody in bronchial washings, the ability to accelerate bacterial clearance persisted to much lower doses. Second, specificity experiments showed that intestinal immunization with non-typable H. influenzae cross-protected against Pseudomonas aeruginosa, even though antibodies were not absorbed out of serum by incubation with P. aeruginosa. Third, serum antibody was shown to be bactericidal for non-typable H. influenzae in the presence of complement (P < 0.05), while bronchial washings antibody was not. The bactericidal effect of the serum was abrogated by the addition of bronchial washings. Fourth, an ELISA quenching assay demonstrated that neutrophils from intestinally immunized rats were able to phagocytose more bacteria in a given time period (P < 0.05) than unimmunized rats and rats immunized by other routes. In the fifth experiment, the chemotactic response of neutrophils to casein was shown to be significantly depressed by the addition of bronchial washings obtained from immunized rats (P < 0.01). It is proposed that specific antibody in bronchial washings does not have a direct role in opsonizing bacteria for killing or phagocytosis, but instead has an anti-inflammatory effect. Non-specific effectors such as neutrophils driven by specific immune cells are a likely means of clearance of bacteria following intestinal immunization and acute challenge.

Previous studies have suggested that, in vivo, activated T lymphocytes and neutrophils are important in immunity to nontypable Haemophilus influenzae. We now extend this work by showing that neutrophils pretreated with products of activated T lymphocytes or activated macrophages show significantly enhanced killing of nontypable H. influenzae. Lymphotoxin, a product of activated T lymphocytes, significantly enhanced the neutrophil-mediated killing of nontypable H. influenzae, and tumor necrosis factor, produced by activated T lymphocytes as well as macrophages stimulated by activated T lymphocytes, also significantly increased the bactericidal activity of neutrophils. These cytokine-induced effects were seen with short pretreatment times of neutrophils and were maximal by 30 min. The killing of H. influenzae by neutrophils required the presence of heat-labile opsonins. In the absence of these opsonins, both tumor necrosis factor and lymphotoxin were unable to promote the killing of the bacteria by neutrophils. Furthermore, the results showed that tumor necrosis factor-primed neutrophils displayed significantly increased expression of CR3 and CR4 that was associated with increased phagocytosis of complement-opsonized nontypable H. influenzae. These cytokines may play an important role in immunity toward nontypable H. influenzae by stimulating neutrophil bactericidal activity.

The aims of this study were to assess the role played by alveolar macrophages, tumor necrosis factor alpha (TNF-alpha), and interleukin-1 alpha (IL-1 alpha) in pulmonary immunity against Pseudomonas aeruginosa in animals that have been immunized via the gut-associated lymphoid tissue. Following intra-Peyer's patch immunization and subsequent intratracheal challenge with live bacteria, significantly enhanced bacterial clearance from the lungs correlated with an increase in bronchoalveolar neutrophils, increased recruitment and phagocytic activity of alveolar macrophages, and accelerated production of TNF-alpha in the bronchoalveolar space, while levels of IL-1 alpha remained low. Administration of recombinant TNF-alpha in physiological concentrations did not affect the proliferation of P. aeruginosa in vitro, but when given intratracheally to rats at the time of infection, recombinant TNF-alpha significantly increased bacterial clearance from the lungs. In these animals, phagocytic activity of bronchoalveolar neutrophils was enhanced, while the recruitment of alveolar macrophages and neutrophils remained unchanged. In acutely infected nonimmune animals, bronchoalveolar concentrations of soluble IL-1 alpha and TNF-alpha increased until the time of death. Levels of prostaglandin E2 and thromboxane B2 were similar in each experimental group. These results indicate that infection in immune animals enhanced both recruitment and phagocytic activity of alveolar macrophages as well as induced an accelerated production of TNF-alpha. In immune challenged animals, this cytokine enhanced the phagocytic activity of neutrophils and improved bacterial clearance from the lung. Levels of soluble IL-1 alpha and TNF-alpha in nonimmune rats increased consistently following infection until the time of death, thus implicating these cytokines in the pathogenesis of acute P. aeruginosa pneumonia.

Outer membrane proteins P2, P4, and P6 and two with molecular masses of 26 and 28 kDa have been purified from a strain of nontypeable Haemophilus influenzae by a preparative form of polyacrylamide gel electrophoresis (PAGE). Outer membrane protein P6, with a molecular mass of 16 kDa (determined by sodium dodecyl sulfate [SDS]-PAGE) was purified by both native PAGE and SDS-PAGE from three strains of nontypeable H. influenzae and one strain of type b H. influenzae. The same conditions were required for purification from each strain. The suitability of proteins isolated by these methods was assessed by studying the immune response of rats immunized with P6 in incomplete Freund's adjuvant into the Peyer's patches. P6 purified by either native PAGE or SDS-PAGE did not differ significantly from P6 purified by gel filtration and anion-exchange chromatography in the ability to enhance pulmonary clearance of live bacteria. This study also investigated the effects of SDS on P2 immunological responses in vivo and the effects of the reagents Zwittergent and sodium lauryl sarcosinate on outer membrane protein lymphocyte-proliferative responses in vitro. It was found that the presence of SDS in the immunization emulsion enhanced the antigen-specific cell-mediated response but suppressed the antigen-specific antibody responses. The presence of residual traces of Zwittergent in an outer membrane protein preparation inhibited antigen-specific cell-mediated proliferation, whereas extraction of outer membrane proteins with sodium lauryl sarcosinate did not inhibit antigen-specific proliferation. These results demonstrate that preparative PAGE is a suitable method for the purification of proteins from the outer membrane of H. influenzae required for investigation of their immunological significance as vaccine candidates and that traces of reagents used during protein purification may play an important role in determining the success of in vivo and in vitro studies.

The role of gut-derived CD4+ T cells in clearance of Pseudomonas aeruginosa from the lung was studied by cell transfer experiments. Mesenteric lymph node cells from unimmunized rats, or rats orally immunized with either killed P. aeruginosa or Helicobacter pylori, were transferred to naive rats which were subsequently challenged intra-tracheally with live P. aeruginosa. Recipients of unseparated mesenteric lymph node cells, purifed T cells or CD4+ T cells, from P. aeruginosa-immunized donors, all exhibited enhanced bacterial clearance from the airways compared to recipients of cells from unimmunized donors. Enhanced clearance by T cells was antigen-specific as no enhanced clearance was observed by transfer of cells from donors immunized with H. pylori.

During a prospective study of the ontogeny of the mucosal immune system using saliva, one subject acquired a selective IgA deficiency at 3 years 6 months of age. Prior to this time the infant had normal ontogeny patterns for salivary immunoglobulins and the salivary IgA was confirmed to be dimeric IgA containing secretory component. Two respiratory tract infections at 3 years 4 months and 3 years 5 months were reported prior to the collection of a saliva sample which was deficient in IgA. All subsequent saliva collections remained IgA deficient. Serum and saliva collected at 11 years of age confirmed persistent IgA deficiency. There was a family history of organ-specific autoimmune disease. The prospectively collected data indicate in this subject that the IgA deficiency was not congenital, but was acquired closely associated with two episodes of respiratory tract infections, against a genetic background of disturbed immune regulation.

Acute episodes of bronchitis have been shown to be unequally distributed within a population of subjects with chronic bronchitis. Two groups were identified based on incidence of acute bronchitis--subjects who were 'infection-prone' (2-5 infections per year) and those who were 'non-infection-prone' (0-1 infections per year). Minor differences in clinical parameters existed, except for smoking experience. The non-infection-prone group included more current smokers, and the total smoking experience (in 'pack years') was significantly greater in this group. Between-year analysis demonstrated a stability of classification, established after a minimum of two years' prospective observation. Parameters of the host-parasite relationship were assessed in both groups. A significantly greater polybacterial colonization of the oropharynx was observed for chronic bronchitics, both infection-prone (P < 0.0001) and non-infection-prone (P < 0.001), compared with control subjects. Infection-prone chronic bronchitics had significantly greater total bacteria cultured from the oropharynx compared to the non-infection-prone group (P < 0.05); adherence of indigenous microflora to buccal epithelial cells, in particular Gram-positive cocci (P < 0.01) and in vitro adherence of non-serotypable Haemophilus influenzae to buccal cells (P < 0.05) compared with the control and non-infection-prone groups. These studies suggest that an important variation in subjects with chronic bronchitis is the binding capacity of epithelial cells for bacteria, which when increased enhances susceptibility to colonization and clinical infection.

The aim of this study was to determine the efficacies of prior mucosal (oral, intra-Peyer's patch, or intratracheal) and systemic (subcutaneous) immunizations with killed Pseudomonas aeruginosa in clearance of an acute P. aeruginosa respiratory infection in rats. Rats were immunized with paraformaldehyde-killed P. aeruginosa at various doses, and 2 weeks later, the rats were challenged with a log10 dose of 8.7 live bacteria. This dose was fatal for unimmunized rats, with death occurring approximately 12 h after challenge. The numbers of surviving bacteria in the airways and lung tissue were determined by analyses of bronchoalveolar lavage fluid (BAL) and lung homogenate samples, respectively. Enhanced bacterial clearance was associated with survival of intra-Peyer's patch-immunized rats. Determination of bacterial clearance in BAL 4 h after challenge demonstrated that the use of all immunization routes led to significant clearance compared with the bacterial levels in unimmunized controls (the order of effectiveness was intra-Peyer's patch > oral-intratracheal > intratracheal > subcutaneous > oral). Bacterial clearance in the lung homogenate was also significantly greater for all immunization routes than in the unimmunized controls (the order of effectiveness was intra-Peyer's patch > subcutaneous > oral-intratracheal > oral = intratracheal). Prior oral immunization with killed P. aeruginosa also induced enhanced bacterial clearance of heterologous strains of P. aeruginosa, Haemophilus influenzae, and to a lesser extent, Klebsiella pneumoniae. Because of the ease of antigen delivery, oral immunization with killed P. aeruginosa may be an important route of immunization for induction of enhanced bacterial clearance of subsequent acute respiratory infection with P. aeruginosa and other gram-negative bacteria.

A prospective study to define the normal patterns of development of mucosal immunity in 263 children provided a unique opportunity to study the mucosal immune response in an infant who unexpectedly died from sudden infant death syndrome. The subject initially had a normal pattern of mucosal immune function, which was perturbed after a transient mild upper respiratory tract infection at 3 1/2 wk of age. After the upper respiratory tract infection, there was an increase in mucosal permeability and the appearance of IgA and IgM in saliva. The unusual features in this case were the degree and the duration of the increases in salivary IgA and IgM after resolution of the illness. The marked abnormalities suggested a persistent stimulation of the mucosal immune response. The case provides informative data on potential mechanisms of sudden infant death syndrome and supports a role for involvement of upper respiratory tract infection.

This paper investigates the effector mechanisms of immune clearance in the lungs of rats immunized against mucoid Pseudomonas aeruginosa. After the gut-associated lymphoid tissue was primed and after a subsequent pulmonary challenge with live bacteria, significantly accelerated bacterial clearances from the lung and raised levels of anti-P. aeruginosa antibodies in sera (immunoglobulin G [IgG], IgA, and IgM) and bronchoalveolar lavages (IgG and IgA) were observed for all immune animals. These changes were associated with enhanced recruitment, chemotaxis, chemokinesis, phagocytic indices, and chemiluminescence of pulmonary polymorphonuclear neutrophils (PMN). In the alveolar spaces of immune animals, an increase in the level of PMN recruitment was not associated with higher levels of leukotriene B4 (LTB4). In contrast, in nonimmune animals that were intratracheally infected with P. aeruginosa, the levels of recruitment and activity of alveolar PMN were lower than those in immune rats but PMN infiltration correlated with a significant increase in the synthesis of LTB4 in the alveolar space. In pulmonary tissue, LTB4 synthesis for both groups was elevated. These findings suggest that accelerated clearance of mucoid P. aeruginosa from the lungs of intestinally immunized rats is due at least in part to factors that induce the enhancement of PMN recruitment and activity in the alveolar space. The mediators that regulate this enhanced response remain unknown but do not seem to include LTB4. The high levels of LTB4 measured in the bronchoalveolar lavages and pulmonary tissues from nonimmune animals infected with live bacteria implicate LTB4 as an important amplifier of the inflammatory response during acute pulmonary infections with mucoid P. aeruginosa in unimmunized hosts.

A total of 2401 isolates of Haemophilus parainfluenzae was isolated from respiratory secretions of 36 healthy adults and 128 patients with chronic bronchitis over a period of 1 year. The isolates were allocated to eight biotypes, by their production of indole, urease and ornithine decarboxylase. Biotypes I and II constituted most of the isolates of H. parainfluenzae from the oropharynx of controls (75%) and chronic bronchitics (c. 90%). Among the patients, there was no difference in the isolation rate between oropharyngeal swabs and sputum specimens. Biotypes III, IV, VI, VII and VIII were isolated less frequently, as was a new taxon defined here as biotype V which does not produce indole, urease or ornithine decarboxylase. Biotype III was isolated significantly less frequently from cases of chronic bronchitis than from controls, whereas biotype II was isolated somewhat more frequently from the patients, especially during acute episodes.

The immunogenicity of a novel particulate oral influenza vaccine was examined in terms of antibody response and protection in mice. Oral immunization with chicken erythrocytes (CRBC) adsorbed with gamma-irradiated influenza A virus induced high levels of immunoglobulin G antibodies and protection in the lung compared with gamma-irradiated virus administered alone or CRBC. Immunoglobulin A antibodies were the predominant antibodies in nasal washings, and their presence did not correlate with protection as well as immunoglobulin G antibodies. Immunity was not specific for the immunizing virus subtype, as antibodies and enhanced lung clearance of virus were demonstrated with different virus subtypes. However, mice were not protected when challenged with live influenza B virus. The antibody response and the degree of protection were dependent on both the concentration of virus adsorbed to CRBC and number of CRBC adsorbed to virus. Virus-adsorbed CRBC given subcutaneously failed to induce antibodies or protection. Oral immunization with A/Qld/6/72 (H3N2) virus gave a high level of protection over 12 weeks, which could be demonstrated with different subtypes. Protection correlated with antibody levels in the lung determined by both enzyme-linked immunosorbent and hemagglutination inhibition assays, although the levels detected by the latter declined over time.

In this study an enzyme-linked immunosorbent assay has been developed for the determination of lysozyme in saliva, serum and urine. The assay relies on the detection of specific protein rather than lytic activity, a property which has been shown to be most suitable for the quantitation of lysozyme in mucin containing substances. Our results indicate that no pretreatment is necessary for the immunochemical method. The assay is sensitive to concentrations as low as 1 microgram lysozyme/l. The intra-assay and inter-assay coefficients of variation were 5.9% and 15.8% respectively. The lysozyme level in whole saliva was 55.53 +/- 30.35 mg/l, in serum the level was 0.64 +/- 0.15 mg/l and in urine it was 0.17 +/- 0.22 mg/l. Comparisons between immunochemical determination and lytic assays showed a good correlation (serum, r = 0.79, P less than 0.01; saliva, r = 0.85, P less than 0.005; treated saliva, r = 0.96, P less than 0.001).

Rats immunized by intra-Peyer's patch (IPP) injection with non-typable Haemophilus influenzae (NTHI) have been shown to clear this organism from the respiratory tract faster than non-immunized rats. We therefore performed a series of experiments in order to determine the mechanism of action of the enhanced pulmonary clearance. The experiments show that homing of intestinal T cells to the respiratory tract is an important component in the observed immunity, while specific antibody adsorbed to bacteria does not influence pulmonary bacterial clearance rate. Mucosally derived lymphocytes were collected from the thoracic duct of rats primed by IPP inoculation with NTHI, and intravenously transfused to recipient rats. These rats were shown to clear bacteria from bronchial spaces faster than non-transfused rats, or rats transfused with non-immune lymphocytes. Lymphocytes collected from the spleens of immunized rats were also capable of conferring the ability to accelerate pulmonary clearance. When thoracic duct lymphocytes (TDL) purified for T lymphocytes were transferred to recipients, the NTHI clearance rate was accelerated. In experiments to evaluate the activity of specific antibody, it was demonstrated that NTHI opsonized with antibody from bronchial washings of immunized rats was not cleared from the respiratory tract of naive rats faster than non-opsonized controls. These data indicate that immune clearance of NTHI from the respiratory tract following gut immunization is dependent upon antigen-primed lymphocytes, that primed T cells are capable of conferring this protection, and that a primary role for specific antibody in the process cannot be established.

Following the administration of a standardized questionnaire, 62 adult patients with chronic bronchitis were enrolled into a double-blind controlled trial of an oral killed Haemophilus influenzae vaccine in the highlands of Papua New Guinea. A 3-day course of vaccine or placebo was given monthly for 3 consecutive months. Participants were monitored weekly over 12 months for acute exacerbations; early morning sputum specimens were collected monthly and during acute exacerbations. Density of colonization by H. influenzae and H. parainfluenzae was determined by standard quantitative and semiquantitative techniques, and the latter method (quadrant score) was used to determine the density of growth of pneumococci. A total of 30 patients received vaccine and 32 placebo. The incidence rate of acute bronchitis in the vaccine group (0.011 episodes/person-weeks) was significantly lower than that in the placebo group (0.021 episodes/person-weeks), but there was no difference between the two groups in the incidence rates of more severe disease. Vaccine efficacy was maximal at times of peak incidence of disease. There was no evidence of a decline in vaccine efficacy for acute bronchitis over the 12-month follow-up period. The number of viable H. influenzae in the sputum declined in both vaccine and placebo groups over the 12-month follow-up period. The average concentration of H. influenzae in the vaccine group fell below that in the placebo group within 1 to 2 months after first immunization and remained so for 12 months, although the difference between the two groups narrowed during the follow-up period.(ABSTRACT TRUNCATED AT 250 WORDS)

The variation of concentrations of immunoglobulins and albumin in consecutive daily collections of saliva was studied in 33 infants, aged 6 months to 5 years, for periods ranging from 16 to 26 days. The concentration and the within-child variability of IgA and albumin and the detection of IgG and IgM in saliva increased with age. Between-child variances were greater than the within-child variances by a factor of 2.8 for log (IgA) and 1.3 for log (albumin). The geometric mean IgA levels were consistently higher and IgG was detected more frequently during upper respiratory tract infections compared with periods of non-infection. There were no changes in albumin levels between infection and non-infection periods, suggesting a local immune response rather than serum leakage. There were significant within-child correlations (autocorrelations) between levels of IgA in saliva collected on consecutive days and samples collected up to 3 days apart. The autocorrelations between levels of albumin were significant for samples collected up to 2 days apart. The autocorrelation for IgA was significantly greater during infection periods compared with non-infection periods for samples collected on consecutive days.

Sixty-six volunteers who considered themselves to have wheat dust-related respiratory disease, were reviewed from a farming community. A spectrum of disease was described ranging from a mild allergic rhinitis limited to periods of wheat dust exposure through to perennial asthma in which wheat dust was but one of several precipitating factors. Approximately half of those with asthma had symptoms limited to times of wheat dust exposure. Results of a limited survey of farmers showed patterns of symptoms similar to those in the volunteer group. Most subjects were atopic with elevated IgE levels, and had positive skin prick tests to environmental allergens. All but one of 65 subjects tested had a positive RAST test to wheat dust antigen. It is concluded that wheat dust associated respiratory tract disease is a significant problem in Australian farming communities, that a particular pattern of asthma can be described, and that IgE mediated reactions contribute to this disease pattern.

We have reviewed the literature to determine the value of C-reactive protein (CRP) measurements in the diagnosis and management of a wide range of conditions. CRP levels are of value in 6 clinical situations: (a) monitoring the response to antibiotic treatment in patients with known bacterial infections, (b) in obstetric patients with premature rupture of membranes, a rise in CRP can give early warning of intrauterine infections, (c) differentiation between active disease and infections in patients with systemic lupus and ulcerative colitis where the level of response to active disease has been previously established, (d) as a measure of disease activity and response to disease-modifying drugs in rheumatoid arthritis, (e) early detection of complications in postoperative patients, (f) in differentiating between infection and graft-versus-host-disease in bone marrow transplant patients. CRP levels have been used in an attempt to differentiate between bacterial and viral infections in various clinical situations, however the published literature does not support this role.

The ability of salivary secretions and sputum sol from patients with chronic bronchitis to inhibit the adhesion of non-serotypable Haemophilus influenzae to human buccal epithelial cells has been examined. Specific H. influenzae antibodies were measured by an enzyme-linked immunosorbent assay (ELISA). Saliva and sputum levels of anti-H. influenzae immunoglobulin A (IgA) antibody were related inversely to buccal cell adherence (r = -0.92). Absorption of saliva and sputum sol with H. influenzae removed antibody detected by ELISA but only partially removed inhibition of attachment of bacteria. In patients with a history of recurrent bronchitis significantly more inhibitory activity remained after absorption of the secretory IgA (sIgA) specific antibody compared with patients who were resistant to acute episodes of infection (P less than 0.05). These observations suggest that factors, other than sIgA, are present in respiratory secretions and are capable of modulating H. influenzae attachment to buccal epithelial cells.

Respiratory tract specimens from chronic bronchitic patients were cultured for Haemophilus species on conventional chocolate agar and a modified sucrose medium in order to determine the accuracy of the new medium in differentiating Haemophilus influenzae from Haemophilus parainfluenzae strains. Haemophilus influenzae biotypes II and III and Haemophilus parainfluenzae biotypes I and II were found to be the predominant strains isolated from the respiratory tract. The modified sucrose medium was found to be a rapid and reliable means of differentiating Haemophilus influenzae from Haemophilus parainfluenzae by sucrose fermentation, on initial isolation.

Subjects prone to recurrent acute bronchitis were admitted to a six-month double-blind clinical study, in which the value of oral immunization with a preparation of killed Haemophilus influenzae was tested. Most subjects had early smoking-related chronic lung disease, unrecognized by either the patient or his/her doctor. Subjects taking the active agent had a 41% reduction in the total number of episodes of acute bronchitis (P = 0.16), a 60% reduction in the number of episodes of acute wheezy bronchitis (P = 0.02) and a 58% reduction in antibiotic use (P = 0.07). The power of analysis was restricted by the small study group. Parameters of episode severity favoured the test group, suggesting that individual episodes of acute bronchitis in subjects taking an oral preparation of killed H. influenzae were less severe than in those taking placebo tablets. Oral immunization with H. influenzae selectively reduced the increase in colonization of the oropharynx with H. influenzae which occurred in the group taking placebo. This is the first demonstration that the common mucosal system can be activated to modify a colonization pattern at a distant site.

One hundred and nine subjects with chronic bronchitis were studied prior to winter and without clinical infection, to determine baseline patterns of bacterial colonisation. Qualitative analysis of cultures of oropharyngeal swabs showed little difference from age matched normal controls (17) except for growth of small numbers of Gram negative coliforms in the chronic bronchitic group. Quantitation of bacteria colonising the oropharynx showed small numbers (mean of 10(5) cfu/ml), with no particular bacteria dominating. Haemophilus influenzae was present in 7.3% of throat swabs from chronic bronchitic patients, but the organism was always less than 10% of the total count. Quantitation of bacteria in sputum showed significantly higher numbers (mean 10(7) cfu/ml). H. influenzae was detected in 25.7% of available specimens, and when present constituted greater than 90% of the total count. Biotyping of H. influenzae isolates demonstrated a separate colonisation of the upper and lower respiratory tracts.

Immunoglobulin and albumin concentrations and Escherichia coli antibody levels were determined in a prospective study of saliva taken from 63 healthy infants during the first year of life. Albumin and IgG were present in high concentrations at birth (57 +/- 6 and 35 +/- 8 mg/l respectively) and decreased in parallel to low values at two months of age. IgA was detected by three weeks of age. The IgA concentration fluctuated until six months of age, after which constant values were observed (14 +/- 3 mg/l). Low levels of salivary IgM (2.4 +/- 1.2 mg/l) were demonstrated in 37% of infants at four weeks of age. No E. coli antibody was detected. There was no significant difference between breast-fed and formula-fed infants. These findings suggest: (i) that mucosal membrane permeability is not restricted to the gut; (ii) that changes in mucosal permeability are non specific and not restricted to the uptake of specific food protein or ingestion of maternal milk; (iii) maternal IgG may contribute to mucosal defence in the neonate; (iv) fluctuations in the concentration of IgA may reflect a balance between intense antigenic stimulation in the gut in the first weeks of life and immune regulatory mechanisms and (v) ingestion of maternal milk does not modify the pattern of ontogeny.

The relative contribution of serum-derived and locally produced IgA has been estimated in sheep mammary secretion at various stages of lactation by comparing the transport of radiolabeled IgA and IgG2 from serum to secretions. During early and mid-lactation, but not during involution, serum IgA was selectively transported to mammary secretion on a large scale; in comparison, there was no transport of serum IgA into intestinal secretion. Estimates of local production indicated that the bulk of the IgA in mammary involution secretion and intestinal secretion was locally produced, whereas there was no evidence for local production of IgA in early or mid-lactation secretion. Our studies reveal an inverse relationship between the extent of local production of IgA by plasma cells underlying secretory epithelia and the selective transport of IgA from serum. This finding suggests that selective transport of serum IgA occurs at a number of mucosal sites, but is dependent on secretory component availability, because where local production is predominant, there are fewer secretory component molecules available for serum IgA transport. For this reason, bile transport of IgA does not represent a unique mechanism for the removal of IgA from serum.

The amounts of immunoglobulins formed in the respiratory tract of sheep were investigated by comparing the distribution of radiolabelled immunoglobulin between plasma and lymph from the caudal mediastinal lymph node efferent duct which drains 50-70% of lung lymph. In addition, immunoglobulin-containing cells in the lymph node lymph and in the respiratory mucosa were counted by immunofluorescence. Whereas 91.6 +/- 5.92% and 67.73 +/- 10.07% of IgG1 and IgG2 respectively were plasma-derived, only 35.89 +/- 7.09% of IgA was plasma-derived. IgM and albumin were wholly plasma-derived. There was no evidence for selective transport of any immunoglobulins from plasma into lymph. It was estimated that mediastinal lymph node lymph contributed 0.12 g IgA to the circulation daily, the bulk of this being of local origin. There were only few cells expressing IgA in the caudal mediastinal lymph node and its efferent lymph indicating that, unlike gut-associated lymphoid tissue, bronchus-associated lymphoid tissue in sheep is not a major site of IgA cell precursor production. Despite this finding, IgA was the most frequent isotype of Ig-expressing cells in the respiratory mucosa and it is concluded that the locally formed IgA in lymph node lymph originated from mucosal plasma cells. IgM- and IgG-expressing cells were much less numerous than IgA-cells in the mucosa, but were the predominant isotype in the caudal mediastinal lymph node and regional lymph nodes and in mediastinal lymph node efferent lymph. These experiments have established that the respiratory tract of sheep is qualitatively similar to the intestine with respect to immunoglobulin synthesis in that the bulk of IgA is locally derived and a smaller but significant local contribution of IgG1 and IgG2 occurs, but that it is a poor source of IgA cell precursors. This implies that the IgA plasma cell population in the respiratory mucosa probably originates from distant mucosal sites.

A simple method is described for the preparation of highly purified IgA and IgM from small volumes of human serum. Enriched IgM and IgA fractions were prepared by precipitation with 7% (w/v) and 14% (w/v) polyethylene glycol respectively. This was followed by affinity chromatography and gel filtration. The final recovery of both IgA and IgM was approximately 30%. The purified preparations obtained were characterized by immunoelectrophoresis, double immunodiffusion, radial immunodiffusion, radioimmunoassay and gel filtration.

Measurement of early pregnancy factor (EPF) by the rosette inhibition test was performed on serum samples from 14 women using intrauterine devices (IUDs). Serial blood samples taken during the luteal phase of 23 cycles demonstrated in six of the cycles a transient appearance of EPF during the 6- to 8-day period following ovulation. In contrast, no EPF activity was found in sera obtained from women in whom fertilization was prevented, either by sexual abstinence or tubal ligation. These observations support the postulate that the IUD functions by the prevention of implantation of the fertilized ovum, rather than by preventing fertilization.

Involvement of the central nervous system (CNS) in chronic lymphatic leukaemia (CLL) is rare. However, when a patient with CLL presented with meningeal symptoms and a high CSF lymphocyte count in the absence of identifiable infection, meningeal leukaemia was considered the most likely diagnosis. Subsequent quantitation of T and B lymphocytes in the blood and CSF was found to be a most valuable aid in excluding the diagnosis, and later a cryptococcal infection was proven. This case is reported to draw attention to the value of T and B lymphocyte quantitation where difficulty is experienced in interpretation of cells in CSF in patients with leukaemia or lymphoma.

The kinetics of T lymphocyte responses to mucosally presented antigen and the appearance of antigen-reactive T cells in mucosal and systemic tissues have been investigated by measuring secondary mixed lymphocyte culture reactivity after intestinal administration of alloantigens. The data indicate that T lymphocytes in the intestine are able to respond to mucosally presented alloantigens, and that following immunisation the T cell reactivity occurs first in Peyer's patches then sequentially in mesenteric lymph nodes, thoracic duct lymph, gut lamina propria and distant mucosal sites. The response was mucosally restricted and the ultimate location of antigen-reactive cells is influenced by site of antigen administration. Evidence is also presented for the existence of systemic nonreactivity to mucosally presented antigens.

An in vitro system of T-dependent polyclonally activated B lymphocytes has been used to identify the cellular basis of immunosuppression induced by penicillamine. Suppression of immunoglobulin synthesis was dose dependent, and required the drug to be present at the initial stages of the culture. Coculture studies identified T helper cells as the primary target. Study of T cell membrane receptors identified a susceptible subpopulation characterized by its inability to form tight sheep red blood cell rosettes, a delay in the generation of T mu receptors and reduced "help" for polyclonally activated B cells. B lymphocytes were not primarily suppressed by penicillamine.

1. Food consumption, live weight, anatomical measurements on the gut organs and the absorptive capacity of the small intestine for L-leucine and D(plus)-glucose were made on virgin (control), pregnant and lactating albino rats. 2. Food intake increased by approximately 60% during pregnancy and a further 250% during lactation. 3. Pregnancy did not markedly influence the gross anatomy of the gastrointestinal tract. There was evidence for increased villus height and percentage water in the small intestine and for increased length of the colon during pregnancy. 4. During lactation, the alimentary canal progressively increased in weight and size. It partially regressed following weaning. 5. All anatomical measurements, except the length of the small intestine, completely regressed to control values within 20 d of weaning. The increased intestinal length had not completely regressed by day 30 post-weaning. 6. No significant change was observed in absolute absorption of glucose or leucine during pregnancy. 7. Absolute absorption of leucine and of glucose was increased during lactation. Greatest absorption occurred on the 10th day of lactation. 8. Results for absorption of leucine and glucose per unit length indicated that the ability of the mucosal cells to absorb or the number of absorptive cells/mm had changed during lactation and the post-lactation periods.

The authors validated use of the rosette inhibition test for the detection of early pregnancy factor (EPF) in human pregnancy, first by optimizing conditions for the assay, using known pregnant and nonpregnant sera, and second, by examining the performance of this assay in three clinical situations: a "blind" study involving coded first-trimester sera showed 80% correlation with pregnancy status; serial assay of EPF activity in sera collected from normal women attempting to conceive, correlated with human chorionic gonadotropin beta-subunit (beta-hCG) levels and pregnancy status; a longitudinal study of serial serum samples through two normal pregnancies showing the continued presence of EPF until the early third trimester in each case. It was concluded that with the rosette inhibition assay, consistent demonstration of a pregnancy-associated substance (EPF) could be obtained.

This overview of the current state of knowledge of the immune system as it is relevant to the oral cavity in health and disease, is presented to illustrate the significance of immunology to the practice of dentistry. Thus immunological mechanisms contribute in a major way to the pathogenesis of the two most common dental disorders and it may be that the ultimate control of dental caries may be through manipulation of the mucosal immune apparatus. Future advances require a clearer and more quantitative analysis of oral mucosal immunity in relation to health and disease.