Anti-smooth muscle antibody in clinical human and experimental animal Mycoplasma pneumoniae infection.

Autoantibody formation is possibly integral to the development of non-respiratory manifestations of acute Mycoplasma pneumoniae infection. We sought to confirm the occurrence of smooth muscle antibodies (SMA) in humans with acute Myc. pneumoniae respiratory infection and furthermore to assess whether similar autoantibodies would develop in a hamster model of respiratory infection. Paired sera from 21 patients with acute infection were assayed for SMA by immunofluorescence on mouse kidney/stomach substrates. The frequency of SMA was then determined for 52 paediatric patients with acute Myc. pneumoniae infection and 16 controls, and for sera from a hamster model of infection. Five of 21 paired sera had an increment in SMA between acute and convalescent specimens. At a screening dilution of 1:40, 18/52 infected and 0/16 controls had positive sera (P = 0.003); positive specimens demonstrated IgG rather than IgM SMA. In the hamster model of Myc. pneumoniae respiratory infection, significant IgG SMA increases occurred in 7/19 infections but not in 11 controls (P = 0.02). Immunoblotting did not identify actin as the substrate for SMA. Smooth muscle antibody increases are found in a significant minority of Myc. pneumoniae-infected humans and hamsters. A role for SMA in the pathogenesis of Myc. pneumoniae infection remains to be defined.

Markers of virulence among prospectively acquired putative enteropathogenic Escherichia coli serogroups.

We assessed the frequency of proposed enteropathogenic virulence factor genes (eaeA and eaf) by genetic amplification for a series of prospectively collected putative enteropathogenic Escherichia coli serogroup isolates that were acquired from the stool specimens of children. Among 102 isolates, eaeA and eaf markers were determined among 27.5% and 4.9%, respectively. Eaf positivity was found to be coexisting in only a minority of eaeA+ E. coli; the eaeA+/eaf- genotype was most common among strains that had evidence of at least one virulence marker. When clinical variables were compared for two groups of patients whose strains did or did not possess eaeA, the eaeA+ group was more likely to have had an acute diarrheal illness (P = .05) and less likely to have had an underlying chronic illness (P = .03). Localized adherence in vitro was easily recognized for eaeA+/eaf+ E. coli but eaeA+/eaf- isolates were less consistent in manifesting this phenotype. The availability of genetic amplification technologies has the potential to rekindle diagnostic interests in this area, although a rational approach has yet to be defined.

Mycoplasma pneumoniae reinfection and vaccination: protective oral vaccination and harmful immunoreactivity after re-infection and parenteral immunization.

Animal model studies of Mycoplasma pneumoniae infection after live respiratory challenge were conducted to investigate the issues of challenge-rechallenge associated accentuated pathology, postparenteral vaccination associated accentuated pathology, and oral vaccination. Live M. pneumoniae inocula were grown in hamster serum-based medium in order to reduce the potential for the serum growth component to participate in the hyperaccentuated histopathological response as seen with challenge-rechallenge experiments which have used horse serum-based growth media. Despite the use of homologous animal serum, an early hyperaccentuated response occurred (day 3 score 13.3 vs day 10 score 7.7; P = 0.02) which included perivascular infiltrates, and histopathological scores for early (day 3) and late (day 10) disease were similar (P > 0.10) between experiments of challenge-rechallenge when either homologous or heterologous sera were used in inoculum growth media. Parenteral vaccination with heat-killed bacteria also led to an early hyperaccentuated histopathological response after live respiratory challenge (scores on day 3: vaccinated 18.3, unvaccinated 6.2; P < 0.01) and this response was not significantly diminished when inocula were cleaned of growth medium components. An early accentuated response did not follow oral vaccination with heat-killed bacteria (score on day 3: vaccinated 5.7) and the late reaction was significantly less after challenge (scores on day 10: vaccinated 10.3, unvaccinated 14.6; P = 0.011). Studies of parenteral vaccination should include analyses for early disease after live challenge. Oral vaccination offers a promising route for stimulating protective immunity while minimizing undesirable recall immune events.

An assessment of a new diagnostic indirect enzyme immunoassay for the detection of anti-Mycoplasma pneumoniae IgM.

A new and rapid IgM enzyme immunoassay (EIA)(Immuno Well, Gen-Bio, San Diego, CA) was evaluated for its ability to accurately establish a serodiagnosis for acute Mycoplasma pneumoniae infection. Case definitions were established with the combination of complement fixation (CF) serology, IgM anti-P1 immunoblotting, and clinical and other laboratory data. In an asymptomatic population of 52 children and adults, the EIA was positive for 3.9%. For 17 serum pairs for which there was evidence of a greater than or equal to 4-fold rise in CF titer, 5 acute sera (29.4%) and 14 convalescent sera (82.3%) were positive. In applying the assay to sera that were acquired from a prospective study of childhood infection where the positive case definition was maintained for 22.3% of patients, the sensitivity, specificity, positive predictive value, and negative predictive value for the EIA was 90.5%, 93.2%, 79.2%, and 97.1%, respectively. When the EIA results of a "low-positive," as defined by the manufacturer, were excluded from the positive group, the respective values were 81.0%, 97.3%, 89.5%, and 94.7%. The accuracy of this new assay will be influenced by the prevalence of true illness in the populations to which it may be applied.

A comparison of whole cell protein profiles for sporadic human isolates of Streptococcus equisimilis and beta-haemolytic group G streptococci.

Whole cell protein profiles were resolved for Streptococcus equisimilis (group C) and large colony human biotype beta-haemolytic group G streptococci by the use of one dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Strains of S. equisimilis (27 in toto) were distributed among eight patterns designated A to H. Pattern A represented 48.2% of the latter isolates. Strains of group G streptococci (59 in toto) were distributed among sixteen patterns designated 1-16, and there were no predominant patterns which represented more than 20% of all strains. Profiles were reproducible, not susceptible to strain passage, but susceptible to variation in growth media. Considerable homology was observed among bacteria in either Lancefield group.

IgM anti-P1 immunoblotting. A standard for the rapid serologic diagnosis of Mycoplasma pneumoniae infection in pediatric care.

STUDY OBJECTIVE: To prospectively evaluate the use of an IgM anti-P1 immunoblotting assay for the rapid diagnosis of Mycoplasma pneumoniae infection in a pediatric setting. PATIENTS AND METHODS: Blood specimens from 107 children representing 108 predominantly respiratory illnesses were obtained for a prospective evaluation of the IgM anti-P1 assay. Primary patient diagnoses were determined by a combination of the complement fixation test and supplementary microbiologic and nonmicrobiologic diagnostic tests. The potential effect of the assay results on antibiotic therapy was assessed by observing concurrent therapy. RESULTS: M pneumoniae was the primary etiologic agent of disease in 19 patients. The sensitivity, specificity, positive predictive value, and negative predictive value of the IgM test to determine a case of primary M pneumoniae disease was 84.2 percent, 95.5 percent, 80.0 percent, and 96.6 percent, respectively. Twenty-seven children may have had antimicrobial therapy appropriately modified if results of the assay were directly utilized. Three of four patients with positive assays, which would have been falsely indicative of primary disease, had evidence of a recent probable M pneumoniae infection shortly preceding the acute illness. CONCLUSION: The rapid IgM anti-P1 assay is reasonably specific for the diagnosis of M pneumoniae infection. Apart from establishing prompt and accurate diagnosis, the results have the potential to change treatment measures in a significant proportion of patients.

Differentiation of species in human beta-haemolytic group G streptococci using immunoglobulin Fc fragment receptor.

AIMS: To assess the ability of human immunoglobulin Fc fragment binding activity to differentiate human biotype large colony group G streptococci from the group G "Streptococcus milleri group". METHODS: Fifty two isolates of large colony group G streptococci and 30 group G "S milleri group" strains were tested for their ability to bind fluorescein conjugated human IgG Fc fragments after acetone fixation. Immunoblotting with peroxidase labelled human Fc fragments after resolution of bacterial polypeptides by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed for six large colony strains. RESULTS: All large colony group G streptococci showed positive Fc fragment binding whereas all "S milleri group" bacteria failed to bind Fc fragments when viewed by fluorescence microscopy. All six large colony strains showed similar immunoblot binding patterns. CONCLUSION: Immunoglobulin Fc fragment receptor content distinguishes the large colony group G streptococci from the group G "S milleri group" and mayhave a role in the rapid laboratory diagnosis of pharyngeal pathogens.