Accuracy of respiratory inductive plethysmographic cross-sectional areas.

The present study was undertaken to evaluate whether the respiratory inductive plethysmograph (RIP) 1) reflects changes of cross-sectional area enclosed by its transducer band in the presence of deformations of shape or whether it 2) has a stable base line. Testing of RIP was carried out with a device incorporating a thermally compensated oscillator and digital demodulatory circuitry. This system, introduced to commerce in 1983, superceded the nonthermal compensated oscillatory and analog demodulator circuitry first used in 1977. Testing the effects of changing cross-sectional area was accomplished by stretching a standard RIP transducer band around wooden dowels placed in holes on a peg board grid to form 23 curved and 5 rectangular shapes. The output voltage from RIP was linear for both the curved and rectangular shapes for changes of cross-sectional area within a physiological range. However, the regression line of voltage vs. cross-sectional area for the rectangular shapes was parallel and slightly displaced from the regression line for the curved shapes due to mutual coupling of inductance in the corners. Base-line drift from a RIP transducer band stretched to enclose an elliptical shape was less than 2.5 mV over a 12-h observation period. Current RIP technology accurately reflects changes of cross-sectional area of physiological shapes and has a stable base line.

No serologic evidence of an association found between Gulf War service and Mycoplasma fermentans infection.

Occult occupational infection with Mycoplasma fermentans has been proposed as a cause for illness among Persian Gulf War veterans. Symptom data and sera from a 1994-1995 cross-sectional survey of Navy Seabees were used to select symptomatic and asymptomatic Gulf War veterans and nondeployed veterans to evaluate this hypothesis. Survey sera from 96 Seabees were matched to prewar (before September 1990) archived sera. Immunoblot serologic analyses were performed for M. fermentans in a controlled, blinded fashion. Both Gulf War veterans and nondeployed veterans had prewar and postwar serologic evidence of M. fermentans infection consistent with natural infection data. Among study subjects collectively, and stratified by Gulf War service, none of the immunoblot banding profiles (prewar or postwar) or their changes over time were associated with postwar symptoms. These serologic data do not support the hypothesis that Gulf War veterans have experienced Gulf War-related morbidity from M. fermentans infection.

Characterization of a major Mycoplasma penetrans lipoprotein and of its gene.

A novel mycoplasmal species designated as Mycoplasma penetrans has been isolated recently from patients infected with human immunodeficiency virus. p35, a major antigen extracted from the membrane of this mycoplasma using Triton X-114 has been found to be a lipoprotein. After proteolytic treatment of p35, the sequence of one of the resulting peptides was determined and a corresponding oligonucleotide was deduced. Using this oligonucleotide as a probe the p35 gene was cloned and sequenced. Sequence analysis revealed an amino-terminal signal peptide with a potential acylation site which would result in a 35.3 kDa mature product. In addition, the p35 gene was followed by an open reading frame with a corresponding polypeptide partially homologous to p35, in particular to the N-terminus region.

Dopaminergic drugs in the medial preoptic area and nucleus accumbens: effects on motor activity, sexual motivation, and sexual performance.

In two experiments, dopamine agonists and/or antagonists were injected into the medial preoptic area (MPOA) or the nucleus accumbens (NAcc) of male rats. The animals were then tested in an X-mase with four goal boxes, which contained a receptive female, a male, or were empty. In Experiment 1, the D1 antagonist SCH-23390 and the D2 antagonist raclopride in the MPOA decreased the percentage of trials on which the female's chamber was chosen, a measure of sexual motivation. Raclopride also decreased the number of animals that copulated after choosing the female's chamber. The 10-micrograms dose of the D3/D2 agonist quinelorane increased the latency to reach the female's chamber, slowed the onset of copulation, and decreased the number of intromissions preceding an ejaculation. In Experiment 2, 1- and 5-micrograms doses of quinelorane and of the mixed D1/D2 agonist apomorphine were injected bilaterally into the NAcc. Both doses of quinelorane increased the number of times that the subject did not select a chamber within 60 s. No drug in the NAcc affected specifically sexual motivation or performance. The results are consistent with differential influence of the MPOA and the NAcc on motor activity, sexual motivation, and sexual performance.

An electronic simulator for testing infant apnoea monitors that uses actual physiologic data.

An electronic simulator of physiologic signals used in infant monitoring has been designed, constructed and applied in the Collaborative Home Infant Monitor Evaluation (CHIME). A unique feature of the simulator is that it contains actual physiologic waveforms recorded from infants rather than artificial, idealized signals. The simulator stores breathing waveforms that can be used to test transthoracic-impedance- and inductance-plethysmography-based monitors, and heart rate channels are tested by playing a neonatal QRS complex at preset fixed rates or a variable rate as determined from infant recordings. The transfer characteristics of the simulator are constant over frequencies ranging from 0.5 to 8 Hz for the respiration channels. Data stored in memory are divided into 60 second epochs that can be presented to the monitor being tested in a programmable sequence. A group of 66 CHIME monitors was tested using a simulator programmed with 17 apnoea and bradycardia waveforms. The agreement between monitors as to the duration of detected apnoea decreases as the amount of artefact in the signal increases. Discrepancies between monitors in detecting apnoea duration were found to be similar to inconsistencies between CHIME investigators manually scoring similar waveforms.

Effects on patients with asthma of eradicating visible indoor mould: a randomised controlled trial.

BACKGROUND: It is not clear whether associations between respiratory symptoms and indoor mould are causal. A randomised controlled trial was conducted to see whether asthma improves when indoor mould is removed. METHODS: Houses of patients with asthma were randomly allocated into two groups. In one group, indoor mould was removed, fungicide was applied and a fan was installed in the loft. In the control group, intervention was delayed for 12 months. Questionnaires were administered and peak expiratory flow rate was measured at baseline, 6 months and 12 months. RESULTS: Eighty-one houses were allocated to the intervention group and 83 to the control group; 95 participants in 68 intervention houses and 87 in 63 control houses supplied follow-up information. Peak expiratory flow rate variability declined in both groups, with no significant differences between them. At 6 months, significantly more of the intervention group showed a net improvement in wheeze affecting activities (difference between groups 25%, 95% CI 3% to 47%; p = 0.028), perceived improvement of breathing (52%, 95% CI 30% to 74%; p<0.0001) and perceived reduction in medication (59%, 95% CI 35% to 81%; p<0.0001). By 12 months the intervention group showed significantly greater reductions than the controls in preventer and reliever use, and more improvement in rhinitis (24%, 95% CI 9% to 39%; p = 0.001) and rhinoconjunctivitis (20%, 95% CI 5% to 36%; p = 0.009). CONCLUSIONS: Although there was no objective evidence of benefit, symptoms of asthma and rhinitis improved and medication use declined following removal of indoor mould. It is unlikely that this was entirely a placebo effect.

Protein and antigen variability among strains of Mycoplasma arthritidis.

Six strains of Mycoplasma arthritidis isolated from different host tissues were examined for differences in their proteins and antigens by using one- and two-dimensional electrophoretic techniques as well as immunoblotting. One-dimensional electrophoresis revealed differences in concentrations of individual bands, but not differences in the overall banding pattern. By two-dimensional electrophoretic analysis, 25 proteins were identified as strain-variable whereas the majority of protein spots was strain-constant (about 195 after IEF-2D-PAGE and 145 after NEPHGE-2D-PAGE). Immunoblot analysis using an antiserum against the type-strain of Mycoplasma arthritidis (PG 6) revealed size-heterogeneity of antigens of all six strains. An epitopic relationship between these size-variant antigens could be demonstrated by using monospecific antibodies produced against some of these antigens of Mycoplasma arthritidis. Furthermore, we describe a highly variable antigen of Mycoplasma arthritidis similar to that shown previously in Mycoplasma pulmonis.

Structural variations and phenotypic switching of mycoplasmal antigens.

It is becoming apparent that a high rate of variability of surface structures is a ubiquitous property among mycoplasmas. The present study demonstrates how variations in the size of the V-1 antigen (a major surface antigen of Mycoplasma pulmonis thought to be associated with virulence) are reflected by phenotypic differences (cytadherence) that may play a role in virulence of the organism. Furthermore, a similar antigen is described for the human pathogen Urea-plasma urealyticum, and data are presented on the analysis of clinical isolates that demonstrate the potential for variation in the size of this antigen in vivo. Although no direct connection of antigen variation to natural disease has yet been presented, the data further document the tremendous potential for virulence-related diversity possessed by these organisms and emphasize the importance of a valid animal model for discerning the true relationship between variation and virulence.

Variable antigens of Ureaplasma urealyticum containing both serovar-specific and serovar-cross-reactive epitopes.

Currently there are 14 recognized serovars of Ureaplasma urealyticum, and it has been postulated that only certain ones may be associated with disease and that lack of serovar-specific antibody may be an important risk factor. Unfortunately, ureaplasma antigens important in the human immune response and disease pathogenesis are poorly defined. By using sera from ureaplasma-infected patients and antiureaplasma monoclonal antibodies, the present study has demonstrated, for serovars 3, 8, and 10, antigens which (i) are species specific, (ii) contain both serovar-specific and cross-reactive epitope(s), (iii) are produced not only in vitro but also in vivo, (iv) undergo a high rate of structural variation in vitro, (v) are present and structurally variable on invasive ureaplasma isolates (i.e., those from placenta, lung, and cerebrospinal fluid), and (vi) are among the predominant antigens recognized during infections in humans. Furthermore, we have shown that monoclonal antibodies to these antigens can inhibit the growth of the organisms in vitro, indicating the potential for these antigens to be important for host defense.

Ureaplasma urealyticum intrauterine infection: role in prematurity and disease in newborns.

Ureaplasma urealyticum, a common commensal of the urogenital tract of sexually mature humans, is gaining recognition as an important opportunistic pathogen during pregnancy. While its etiologic significance in many aspects of adverse pregnancy remains controversial, recent evidence indicates that U. urealyticum in the absence of other organisms is a cause of chorioamnionitis. Furthermore, ureaplasmal infection of the chorioamnion is significantly associated with premature spontaneous labor and delivery. In at least some cases, it appears to be causal. Present evidence indicates that U. urealyticum is a cause of septicemia, meningitis, and pneumonia in newborn infants, particularly those born prematurely. There is strong but not definitive evidence that ureaplasmal infection of the lower respiratory tract can lead to development of chronic lung disease in very low-birth-weight infants. Although risk factors for colonization of the lower genitourinary tract have been identified, little information is available concerning risk factors for intrauterine infection and host immune responses to invasive infection. Recent establishment of animal models of respiratory and central nervous system diseases should provide an opportunity to evaluate risk factors, pathogenic mechanisms, and operative immune mechanisms. However, the most critical need is additional information concerning indications for diagnosis and treatment as well as efficacy of treatment.

Cytopathic effects of Mycoplasma pulmonis in vivo and in vitro.

This study was performed to evaluate the cytopathic features resulting from Mycoplasma pulmonis infection of tracheal organ cultures compared to with those seen in in vivo infection and to use this system to determine possible differences in cytopathic effects in two M. pulmonis variants found to cause different diseases in vivo. The attachment of M. pulmonis to respiratory epithelium was similar in vivo and in vitro. Cytopathic effects seen in both systems were also similar in loss of tight junctions between cells and exfoliation of respiratory cells, resulting in exposure of the subepithelial layer. These similarities indicate that the observed tissue damage is initiated by the mycoplasmas rather than by immunologic host responses but does not exclude the possibility that host responses may subsequently contribute to the cytopathological events. Comparison of the effects of the two variants (one known to cause death in vivo) did not reveal differences in vitro. This suggests that host factors (not present in vitro) may account for differences in virulence. Detailed in vitro studies allowed the identification of the time frame corresponding to the in vivo infection and also revealed the limitations of the in vitro system.

Protein and antigen heterogeneity among strains of Mycoplasma fermentans.

The proteins and antigens of three strains of Mycoplasma fermentans were compared with those of a mycoplasma, designated "Mycoplasma incognitus," recently identified in tissues of AIDS patients. Previous studies have shown that "M. incognitus" is most likely not a new species but rather a strain of M. fermentans. In the present study, one- and two-dimensional electrophoretic analysis demonstrated the expected similarity between these mycoplasmas, but it also demonstrated several distinct protein differences. Nine proteins were identified as strain variable by two-dimensional gel electrophoresis. Also, immunoblot analysis using rabbit antiserum against the type strain of M. fermentans (strain PG 18) documented the occurrence of size heterogeneity in at least one and possibly two other antigens.

Cytopathogenicity of Mycoplasma fermentans (including strain incognitus).

Mycoplasma fermentans strain incognitus, an organism recently identified in tissues of patients with AIDS and in tissues of otherwise healthy adults with an acute fatal respiratory disease, was evaluated for cytopathogenicity for tracheal tissue in vivo and in vitro. In this study, the organism produced a chronic infection of the lower respiratory tract in LEW rats following intranasal inoculation and induced both ciliostasis and cytopathology in experimentally infected tracheal explants from rats. The time of onset of ciliostasis, type of cytopathogenicity, and localization of organism in strain incognitus were different from those in other strains of M. fermentans as well as other species of mycoplasmas isolated from humans. The results strongly support, but do not prove, that M. fermentans strain incognitus is an unusually invasive mycoplasma, as it was the only strain found within respiratory epithelial cells both in vivo and in vitro. Detection of the organism within the lamina propria also supported the organism's invasive potential. Further study of both the in vivo and in vitro models should provide insights into this potentially unique mycoplasma-host relationship.

Adsorption of mycoplasma virus P1 to host cells.

The adsorption of mycoplasma virus P1, a virus which infects some strains of Mycoplasma pulmonis, to host cells was examined. Mutants of M. pulmonis to which P1 virus did not adsorb were isolated. Proteins from the mutants and from wild-type cells were compared by two-dimensional polyacrylamide gel electrophoresis, and the only observed difference was in the surface antigen V-1. The electrophoretic properties of V-1 also correlated with the host range of the virus. These data strongly suggest that the V-1 antigen affects adsorption of P1 virus to host cells.

Serotype diversity and antigen variation among invasive isolates of Ureaplasma urealyticum from neonates.

Ureaplasma urealyticum has previously been isolated from the cultured cerebrospinal fluid of 13 of 418 newborn infants; additional bloodstream isolates were obtained from the same population. Ten of the 13 cerebrospinal fluid and 3 bloodstream isolates were available for serotyping in the present study. By the use of serotype-specific reagents, including monoclonal antibodies, 70% of the cerebrospinal fluid isolates were identifiable as serotype 1, 3, 6, 8, or 10; i.e., they represented 5 of the 14 established serotypes or both presently defined genomic clusters. One of the bloodstream isolates was identified as serotype 3. Our data support the hypothesis that the property of invasiveness for unreaplasmas is likely not limited to one or a few particular serotypes among the 14 established serovars. Additionally, our study has shown that even in isolates of the same serotype, there can be size variation in the antigens expressed. Therefore, it would appear that many serotypes are invasive and that perhaps antigen variability and host factors may be more important determinants for ureaplasma infections than different serotypes per se.

Evaluation of intraspecies genetic variation within the 16S rRNA gene of Mycoplasma hominis and detection by polymerase chain reaction.

Mycoplasma hominis is a heterogeneous species with DNA-DNA hybridization values ranging from 51 to 100%. We report here the sequencing of the 16S rRNA gene of a strain (183) that greatly differs from the type strain (PG21) of this species. Comparison of 16S rDNA sequences from these two strains showed limited differences, indicating that the two strains belong to the same rRNA species complex. Using these nucleotide sequence data, we established a rapid method for the detection of M. hominis by using polymerase chain reaction. This method was shown to be sensitive and specific when tested with reference strains and clinical isolates.

Epitope mapping of the variable repetitive region with the MB antigen of Ureaplasma urealyticum.

One of the major surface structures of Ureaplasma urealyticum recognized by antibodies of patients during infection is the MB antigen. Previously, we showed by Western blot (immunoblot) analysis that any one of the anti-MB monoclonal antibodies (MAbs) 3B1.5, 5B1.1, and 10C6.6 could block the binding of patient antibodies to MB. Subsequent DNA sequencing revealed that a unique six-amino-acid direct tandem repeat region composed the carboxy two-thirds of this antigen. In the present study, using antibody-reactive peptide scanning of this repeat region, we demonstrated that the amino acids defining the epitopes for MAbs 3B1.5 5B1.1 and 10C6.6 are EQP, GK, and KEQPA, respectively. Peptide scanning analysis of an infected patient's serum antibody response showed that the dominant epitope was defined by the sequence PAGK. Mapping of these continuous epitopes revealed overlap between all MAb and patient polyclonal antibody binding sites, thus explaining the ability of a single MAb to apparently block all polyclonal antibody binding sites. We also show that a single amino acid difference in the sequence of the repeats of serovars 3 and 14 accounts for the lack of reactivity with serovar 14 of two of the serovar 3-specific MAbs. Finally, the data demonstrate the need to obtain the sequences of the mba genes of all serovars before an effective serovar-specific antibody detection method can be developed.

Accuracy of respiratory inductive plethysmograph over wide range of rib cage and abdominal compartmental contributions to tidal volume in normal subjects and in patients with chronic obstructive pulmonary disease.

We assessed the accuracy of the respiratory inductive plethysmograph in the supine position to spirometry by the two-body position, least squares calibration and single-body position, isovolume calibration procedures. The comparison was carried out simultaneously in normal subjects breathing naturally and with voluntarily controlled abdominal or thoracic breathing, and in patients with COPD breathing naturally and with voluntarily controlled abdominal breathing patterns. In both groups, there was no significant difference in estimation of tidal volume between the 2 calibration procedures for the various breathing patterns. There was greater deviation from spirometric tidal volume values for both calibration methods in patients with COPD during abdominal than during natural breathing. In the normal subjects, agreement between the rib cage and abdominal partitioning of tidal volume for both calibration methods was good, but in the patients with COPD there was greater variability. In normal subjects, over a wide range of rib cage and abdominal compartmental contributions to tidal volume, either calibration procedure appears satisfactory. For patients with COPD, if large changes occur in the distribution of rib cage and abdominal contributions to tidal volume, then validation of respiratory inductive plethysmography to spirometry must be rechecked.

Genotypic and phenotypic analysis of Mycoplasma fermentans strains isolated from different host tissues.

A correlation was found between the expression of a specific Mycoplasma fermentans surface antigen (Pra, proteinase-resistant antigen) and the site of isolation of the organism from the infected host. Strains which expressed Pra were most frequently associated with cells of bone marrow origin, and strains which lacked expression of Pra were most commonly isolated from the respiratory tract, genital tract, and arthritic joints, i.e., epithelial cell surfaces. Pra was previously shown to be resistant to degradation by proteinases and was hypothesized to play a protective role at the organism surface and perhaps to influence which host tissue site was colonized by the organism. The methods used for this phenotyping scheme required isolation and growth of the mycoplasma in quantities sufficient for immunoblot analysis using monoclonal antibodies. We wanted to determine a more rapid and less cumbersome technique to supplement this method for determining the Pra phenotype directly in clinical specimens. Here we describe PCR studies to investigate the movement of a previously identified M. fermentans insertion sequence (IS)-like element. These data showed a correlation between a specific IS genotype and the Pra+ phenotype. Production of a 160-bp product using a single set of IS-based primers was associated with expression of Pra. The genomic IS location resulting in the 160-bp product was determined by using Southern blot analysis and was found to be a stable insertion site characteristic of genotype I strains. Additional analyses of sequences within and flanking the IS insertion sites revealed another pair of PCR primer sites which resulted in the consistent production of a 450-bp amplicon. The stability of this site was dependent on the absence of the IS-like element between the primer sites. The production of this 450-bp amplicon correlated with the Pra mutant phenotype and was characteristic of genotype II strains. The data showed that the sequence within the IS may be unstable and that reliable genotyping sequences are more easily found in the stable genomic sites which flank the IS element.