Bronchoalveolar lavage findings in pulmonary paracoccidioidomycosis

Bronchoalveolar fluid cytology from six progressive pulmonary paracoccidioidomycotic patients showed an alveolitis of neutrophilic pattern which was independent of the of the duration of the chronic fungal disease. The percentage of neutrophils in paracoccidioidomycotic alveolitis was higher than in other neutrophilic alveolitis conditions.     

Bronchoalveolar lavage in the assessment of pulmonary Hodgkin's disease

Abnormal chest radiographs in patients with Hodgkin's disease are occasionally due to pulmonary Hodgkin's disease. The fluids recovered from bronchoalveolar lavages (BALs) from 50 patients prior to autologous bone marrow transplantation for advanced Hodgkin's disease were examined. Abnormal chest roentgenograms were present in 24 patients (48%); 4 (17%) of these had Reed-Sternberg cells or their mononucleated variants in the lavage fluid and an alveolar lymphocytosis averaging 31.4% (normal: 11.5%). The lymphocytes were small and monotonous. Of the 20 patients with abnormal chest roentgenograms but no Reed-Sternberg cells in the lavage fluid, the lymphocyte count was 10.88%, with only 3 patients exceeding 17%. Two patients with normal chest roentgenograms had Reed-Sternberg-like cells in their lavage fluids and averaged 23% lymphocytes in their lavage differential count. Eosinophils averaged 1% or less of the lavage differential and were not predictive of pulmonary Hodgkin's disease. This experience suggests that pulmonary Hodgkin's disease can be diagnosed by BAL. Reed-Sternberg cells and their mononucleated variants can be recognized by their characteristic cytomorphologic features, although care must be taken not to misinterpret reactive binucleated macrophages as neoplastic cells. In patients with Hodgkin's disease, Reed-Sternberg cells should be sought when an alveolar lymphocytosis is present.     

Mycobacterium avium infection in a silicone-injected breast

A case of atypical mycobacterial infection (M. avium intracellulare) in a silicone-injection augmented breast is described. The silicone injection may have been a contributing factor to the development of this unusual infection. Disseminated M. avium was present in this patient, with breast involvement being suggested by the rapid appearance and disappearance of localized areas of erythema and tenderness. Aggressive treatment of these breast infections while they are still localized may prevent systemic spread. Conventional incision and drainage of the breast abscess combined with multidrug treatment directed against M. avium is the recommended therapy.     

Bronchoalveolar lavage in an immunosuppressed rabbit model of invasive pulmonary aspergillosis

A rabbit model of invasive aspergillosis has been used to investigate the pathogenesis of Aspergillus infection in the immunosuppressed host. The animals received hydrocortisone daily and a single dose of cyclophosphamide 2 days prior to intratracheal instillation of conidia from Aspergillus fumigatus. Bronchoalveolar lavage (BAL) was performed in 3 infected and 2 control saline treated animals sacrificed on days 1, 2, 4, 7 and 10 following inoculation. Infective load within the lung was quantified using an assay for chitin which is an important component of fungal cell walls (in particular the hyphal cell wall) and is not present in vertebrate tissue. The total BAL white cell count did not discriminate between infected and saline treated animals and Aspergillus was cultured from one lavage specimen only. Infected animals developed a marked neutrophil alveolitis by day 2 in contrast to a near total absence of neutrophils in the lavages of the control animals. Phagocytosis of conidia by alveolar macrophages was prominent but did not prevent progressive infection as confirmed by measurement of lung chitin. This pattern of cellular response within the alveolar airspace reflects the complex nature of the response to Aspergillus infection in the immunosuppressed host.     

Bronchoalveolar lavage in liver transplant patients

Because immunosuppression is required to control rejection, liver allograft recipients are susceptible to a variety of opportunistic pathogens. A total of 191 bronchoalveolar lavage (BAL) specimens from 89 patients (53 adults and 36 children) who underwent orthotopic liver transplantation was reviewed. One case each of cytomegalovirus (CMV), staphylococcal and Enterobacter pneumonia was diagnosed with the aid of pretransplant BAL. The pretransplant BAL in 62 patients showed rare yeasts in 24.2%; these probably represent oropharyngeal contaminants since the patients involved had no symptoms of Candida pneumonia. Among 54 patients who developed respiratory symptoms and underwent posttransplant BAL, 23 (42.6%) were infected with opportunistic pathogens, including Pneumocystis carinii (22.2%), CMV (22.2%) and herpes simplex virus (HSV) (7.4%). Frequently, infection with multiple organisms was present. Adults constituted 100% of the HSV-infected group, 69.2% of the CMV-infected group and 16.6% of the group infected with P carinii. The diagnosis of these infections was aided by a combination of cytology, microbial culture and in situ hybridization techniques. Although BAL permitted the diagnosis and treatment of opportunistic infections, high mortality (62.5%) occurred with CMV and HSV pneumonia. Further studies into methods that permit earlier diagnoses of these infections are necessary.     

A comparison of ovine monocyte-derived macrophage function following infection with Mycobacterium avium ssp. avium and Mycobacterium avium ssp. paratuberculosis

Mycobacterium avium ssp. paratuberculosis causes Johne's disease in ruminants, whereas the antigenically and genetically similar subspecies Mycobacterium avium ssp. avium is less virulent. In this study, we compared one strain of each subspecies for its ability to survive, induce cytokines, suppress MHC class I and II expression and induce apoptosis or necrosis in ovine monocyte-derived macrophages. Both subspecies survived intracellularly and induced the secretion of IL-10. Low levels of TNF-alpha were detected after infection with both subspecies at 4 h. IL-12 was not upregulated after infection. Downregulation of MHC class I and II was evident in response to infection with both M. avium ssp. avium and M. avium ssp. paratuberculosis. No significant cytotoxicity was detectable in ovine macrophages after the addition of bacteria. M. avium ssp. paratuberculosis induced slightly more apoptosis than M. avium ssp. avium. Still the overall rate of apoptosis was very low and both subspecies suppressed LPS-induced macrophage apoptosis.     

Bronchoalveolar lavage in lung transplant patients.

Over a four-month period (November 1991-February 1991), six patients underwent lung transplantation for end-stage pulmonary disease. Twenty-four transbronchial biopsies with concurrent bronchoalveolar lavage were performed at regular intervals to evaluate acute graft rejection and/or infection. A study was performed to evaluate the role of lavage in the cytodiagnosis of acute pulmonary graft rejection. Histologic features of acute rejection were present in 11/24 biopsies (46%), consisting of perivascular and/or peribronchial lymphoid aggregates. Concurrent lavage findings were hyperplastic alveolar lining cells (one specimen, 9%), atypical lymphocytes (one specimen, 9%), and acute and chronic inflammation (nine specimen, 82%). Among the 13 lavages without concurrent histologic evidence of acute graft rejection (54%), atypical lymphocytes were present in one specimen (8%). The majority of the specimens (54%) showed a relative lack of inflammation. Two of the 24 lavages (8%) contained either Candida or cytomegalovirus (CMV), which were not present on the concurrent biopsy, and one biopsy contained CMV, which was absent on the lavage smear. Although a higher percentage of lavages with histologic evidence of acute graft rejection contained acute and chronic inflammatory cells as compared to lavages without concurrent histologic evidence of rejection, the cytodiagnosis of acute graft rejection on bronchoalveolar lavage is indeterminate.     

Bronchoalveolar lavage protein patterns in children with malignancies,immunosuppression, fever and pulmonary infiltrates

Severe respiratory infections are a major cause of morbidity and mortality in children receiving immunosuppressive therapy for malignancies. The goal of this study was to assess the major changes in the protein patterns in these children. Bronchoalveolar lavage (BAL) fluids of seven control children and of ten children with malignancies and fever not responding to broad spectrum antibiotic treatment was separated by horizontal two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the isoelectric point range 3-9. We observed a large increase of alpha(1)-antitrypsin (p = 0.0004) and decreases of the immunoglobulin (Ig) binding factor, transthyretin and cystatin S. Significant changes occurred also in the small acidic proteins. The relative abundance of the IgG heavy and light chains may hinder the separation and identification of many minor protein spots located in the basic area of the gel, suggesting that their removal during sample preparation may be warranted. This study demonstrated significant alterations in BAL fluid proteome in immunosuppressed children with persistent fever and pulmonary infiltrates. Future target regions of interest were identified. Sample prefractionation and the selection of suitable narrow isoelectric point ranges will be necessary for optimized detection and separating conditions.     

Pathogenic Mycobacterium avium remodels the phagosome membrane in macrophages within days after infection

As part of their strategy for intracellular survival, mycobacteria prevent maturation of the phagosomes in which they reside inside macrophages. The molecular basis for this inhibition is only now beginning to emerge, by way of the molecular characterisation of the phagosome membrane when it encloses virulent mycobacteria. Our own work has shown that at 15 days after the phagocytic uptake of Mycobacterium avium by mouse bone marrow-derived macrophages, the phagosome membrane is depleted about 4-fold for cell surface-derived membrane glycoconjugates, labelled by exogalactosylation, in comparison to the membrane of early endosomes with which it continues to interact. Here we asked whether this depletion occurred at early or late stages after infection. We found that only about half of the depletion had occurred at about 5 hours after the beginning of phagocytic uptake, with the remainder becoming established thereafter, with a half-time of about 2.5 days. Phagosomes became depleted in relation to early endosomes with which they continued to exchange membrane constituents. Early endosomes themselves became gradually depleted by about 30% during the 15-day post-infection period. In contrast, late endosomes/lysosomes remained unchanged, with a concentration of surface-derived glycoconjugates between that of early endosomes and of phagosomes at day 15 post infection. In view of the slowness of the post-infection change of phagosome membrane composition, we proposed that this change did not play a role in preventing maturation immediately after phagosome formation, but rather correlated with the process of maintaining the phagosomes in an immature state.     

Methylation of GPLs in Mycobacterium smegmatis and Mycobacterium avium

Several species of mycobacteria express abundant glycopeptidolipids (GPLs) on the surfaces of their cells. The GPLs are glycolipids that contain modified sugars including acetylated 6-deoxy-talose and methylated rhamnose. Four methyltransferases have been implicated in the synthesis of the GPLs of Mycobacterium smegmatis and Mycobacterium avium. A rhamnosyl 3-O-methytransferase and a fatty acid methyltransferase of M. smegmatis have been previously characterized. In this paper, we characterize the methyltransferases that are responsible for modifying the hydroxyl groups at positions 2 and 4 of rhamnose and propose the biosynthetic sequence of GPL trimethylrhamnose formation. The analysis of M. avium genes through the creation of specific mutants is technically difficult; therefore, an alternative approach to determine the function of putative methyltransferases of M. avium was undertaken. Complementation of M. smegmatis methyltransferase mutants with M. avium genes revealed that MtfC and MtfB of the latter species have 4-O-methyltransferase activity and that MtfD is a 3-O-methyltransferase which can modify rhamnose of GPLs in M. smegmatis.     

Regulation of toll-like receptor 2 expression by macrophages following Mycobacterium avium infection

Recent studies have implicated Toll-like receptors (TLR), especially TLR2 and TLR4, as sentinel receptors that signal the interaction of macrophages with bacterial pathogens via a NF-kappaB-mediated pathway. The regulation of TLR gene expression, however, has not been intensively studied. Here, we report that TLR2 mRNA was induced following infection of murine macrophages with Mycobacterium avium. The changes in TLR2 mRNA correlated with an increase in TLR2 surface expression. Infection with M. avium resulted in a concomitant decrease in TLR4 mRNA. The effect of M. avium infection on TLR2 mRNA appeared to be mediated, in part, by TLR2 because the induction of the mRNA was partially blocked by preincubation of the macrophages with an anti-human TLR2 Ab. In contrast, the effect of LPS stimulation was mediated via TLR4 because infection of macrophages from LPS(d) mice, which do not express active TLR4, resulted in an increase in TLR2 mRNA, while treatment of macrophages from these mice with LPS failed to induce TLR2 mRNA. Several cytokines, including TNF-alpha, IL-1alpha, and GM-CSF, but not IFN-gamma, induced TLR2 mRNA. M. avium infection resulted in the induction of TLR2 mRNA by macrophages from both TNFRI knockout and NF-kappaB p50 knockout mice.     

Bronchoalveolar lavage in interstitial lung disease: effect of volume of fluid infused

To evaluate the effect of varying infusate volume on the results of bronchoalveolar lavage (BAL) in patients with interstitial lung disease, 55 patients underwent 58 BAL during which both a 100- and 250-ml lavage was performed in the same lobe of the lung. Although the percent of the fluid that was returned and the total numbers of cells were greater in the 250- vs. the 100-ml lavage, there were no significant differences in cell differentials or numbers of cells per milliliter between the 100- and 250-ml BAL. We conclude that infusate volume does not affect cell differentials or numbers of cells per milliliter of bronchoalveolar lavage fluid in patients with interstitial lung disease.     

Mycobacterium avium subspecies paratuberculosis and Mycobacterium avium subsp. avium infections in a tule elk (Cervus elaphus nannodes) herd

Between 2 August and 22 September 2000, 37 hunter-killed tule elk (Cervus elaphus nannodes) were evaluated at the Grizzly Island Wildlife Area, California, USA, for evidence of paratuberculosis. Elk were examined post-mortem, and tissue and fecal samples were submitted for radiometric mycobacterial culture. Acid-fast isolates were identified by a multiplex polymerase chain reaction (PCR) that discriminates among members of the Mycobacterium avium complex (MAC). Histopathologic evaluations were completed, and animals were tested for antibodies using a Johne's enzyme-linked immunosorbent assay (ELISA) and agar gel immunodiffusion. In addition, 104 fecal samples from tule elk remaining in the herd were collected from the ground and submitted for radiometric mycobacterial culture. No gross lesions were detected in any of the hunter-killed animals. Mycobacterium avium subsp. paratuberculosis (MAP) was cultured once from ileocecal tissue of one adult elk and was determined to be a strain (A18) found commonly in infected cattle. One or more isolates of Mycobacterium avium subsp. avium (MAA) were isolated from tissues of five additional adult elk. Gastrointestinal tract and lymph node tissues from 17 of the 37 elk (46%) examined had histopathologic lesions commonly seen with mycobacterial infection; however, acid-fast bacteria were not observed. All MAC infections were detected from adult elk (P = 0.023). In adult elk, a statistically significant association was found between MAA infection and ELISA sample-to-positive ratio (S/P) > or = 0.25 (P=0.021); four of five MAA culture-positive elk tested positive by ELISA. Sensitivity and specificity of ELISA S/P > or = 0.25 for detection of MAA in adult elk were 50% and 93%, respectively. No significant associations were found between MAC infection and sex or histopathologic lesions. Bacteriologic culture confirmed infection with MAP and MAA in this asymptomatic tule elk herd. The Johne's ELISA was useful in signaling mycobacterial infection on a population basis but could not discriminate between MAA and MAP antibodies. The multiplex PCR was useful in discriminating among the closely related species belonging to MAC.     

Bronchoalveolar lavage fluid cytology in patients with Pneumocystis carinii pneumonia

OBJECTIVE: To evaluate bronchoalveolar lavage (BAL) cytology and organism burden in patients with Pneumocystis carinii pneumonia (PCP) who were infected with the human immunodeficiency virus (HIV) and in those with other immunodeficiencies. STUDY DESIGN: BAL fluid samples from patients with PCP were selected (HIV-infected patients, n = 15; patients with other immunodeficiencies, n = 11). May-Grünwald-Giemsa-stained cytocentrifuge preparations were evaluated. Foamy alveolar casts (FACs) and P carinii clusters were counted. RESULTS: The numbers of FACs and P carinii clusters in BAL fluid samples of HIV-infected patients were significantly higher as compared to those in samples from patients with other immunodeficiencies. Striking cytologic findings observed in half the samples from both patient groups included the presence of foamy alveolar macrophages, activated lymphocytes, plasma cells and reactive type II pneumocytes. Furthermore, a peculiar cell type, "nonidentified cell" (NIC), was observed almost exclusively in BAL fluid samples from HIV-infected patients. CONCLUSION: BAL fluid samples from HIV-infected patients with PCP displayed higher organism burdens as compared to those from patients with other immunodeficiencies. Moreover, cytologic findings suggestive of noninfectious lung conditions were common in BAL fluid samples obtained from patients with PCP. Further study is required to elucidate the identity of the NIC cell type.