Synthesis and distribution of CARDS toxin during Mycoplasma pneumoniae infection in a murine model.

Mice were infected with Mycoplasma pneumoniae and monitored for the synthesis and distribution of the unique adenosine diphosphate-ribosylating and vacuolating Community Acquired Respiratory Distress Syndrome (CARDS) toxin in bronchiolar lavage fluid (BALF) and lung. We noted direct relationships between the concentration of CARDS toxin and numbers of mycoplasma genomes in BALF and the degree of histologic pulmonary inflammation. Immunostaining of lungs revealed extensive colonization by mycoplasmas, including the detection of CARDS toxin in the corresponding inflamed airways. Lung lesion scores were higher during the early stages of infection, decreased gradually by day 14 postinfection, and reached substantially lower values at day 35. Infected mouse immunoglobulin (Ig) M and IgG titers were positive for CARDS toxin as well as for the major adhesin P1 of M. pneumoniae. These data reinforce the proposed pathogenic role of CARDS toxin in M. pneumoniae-mediated pathologies.

Variation in colonization, ADP-ribosylating and vacuolating cytotoxin, and pulmonary disease severity among mycoplasma pneumoniae strains.

RATIONALE: Mycoplasma pneumoniae was recently discovered to produce an ADP-ribosylating and vacuolating cytotoxin, designated CARDS toxin, which is hypothesized to be a primary pathogenic mechanism responsible for M. pneumoniae-induced pulmonary inflammation. It is unknown if cytotoxin production varies with M. pneumoniae strain or if variation in cytotoxin production affects pulmonary disease severity. OBJECTIVES: To examine the production of CARDS toxin by various strains of M. pneumoniae and compare the disease manifestations elicited by these strains in an experimental model of M. pneumoniae respiratory infection. METHODS: BALB/c mice were inoculated once intranasally with SP4 broth (negative control) or three different M. pneumoniae strains: M129-B7, M129-B9, or S1. Mice were assessed at 1, 2, 4, 7, 10, and 14 days after inoculation. Outcome variables included comparisons among M. pneumoniae strains relative to bronchoalveolar lavage (BAL) M. pneumoniae quantitative culture, CARDS toxin-based PCR, and CARDS toxin protein determinations, as well as cytokine and chemokine concentrations. Graded lung histopathologic score (HPS) was also assessed. MEASUREMENTS AND MAIN RESULTS: CARDS toxin concentrations were significantly increased in mice inoculated with strain S1 compared with mice inoculated with M129-B7 or M129-B9 strains. Quantitative M. pneumoniae culture and polymerase chain reaction were also significantly greater in mice infected with S1 strain compared with the other two strains, as were lung HPS and concentrations of IFN-γ, IL-12, IL-1α, macrophage inflammatory protein-1α, and keratinocyte-derived chemokine. In addition, a significant positive correlation was found between CARDS toxin concentration and lung HPS. CONCLUSIONS: CARDS toxin concentrations in BAL are directly linked to the ability of specific M. pneumoniae strains to colonize, replicate, and persist, and elicit lung histopathology. This variation among strains may predict the range in severity of pulmonary disease observed among patients.

DMSA renal scans and the top-down approach to urinary tract infection.

Assessment of first febrile urinary tract infection in young children has usually involved 2 imaging techniques: renal ultrasonogram and voiding cystourethrography. Currently, there is growing interest in using the dimercaptosuccinic acid (DMSA) scan labeled with technetium-99m as an alternative initial study, in the evaluation of UTI. DMSA renal scanning is the most sensitive radiologic study to detect acute pyelonephritis. Early DMSA renal scanning has been called the top-down approach, because the focus is the identification of kidney injury rather than reflux. Positive and negative aspects of DMSA renal scans are discussed by a pediatric urologist.

Systemic cytokine profile in children with community-acquired pneumonia.

OBJECTIVES: Characterization of the systemic cytokine response in community-acquired pneumonia (CAP) may facilitate our understanding of the host immune response and provide a prognostic as well as diagnostic tool. Systemic cytokine characterization of CAP has been limited largely to a few integral cytokines in adults. METHODS: Analyses were performed to investigate whether significant relationships existed between an expanded serum cytokine profile and etiologies, manifestations, and outcomes of pediatric CAP. The serum concentrations of 15 cytokines were investigated in 55 hospitalized children with well-characterized CAP. RESULTS: Comparison of median cytokine concentrations among patients with CAP caused by Mycoplasma pneumoniae or Chlamydophila pneumoniae, Streptococcus pneumoniae, viruses, mixed infections, or unidentified pathogens revealed significant differences in IFN-alpha, IL-6, IL-17, GM-CSF, and TNF-alpha concentrations. The mixed infections category had significantly elevated concentrations of IFN-alpha, IL-6, GM-CSF, and TNF-alpha. There were significant correlations between concentrations of IL-6 and markers of disease severity (white blood cell band-forms, procalcitonin, and unequivocal consolidation). No single cytokine could reliably differentiate the etiologic cause of pneumonia. CONCLUSIONS: IL-6 is the only one of 15 serum cytokines studied that correlated with indicators of disease severity in childhood CAP. The applicability of cytokine profiles to identify microbiologic etiologies of pneumonia remains to be defined.

Treatment of experimental chronic pulmonary mycoplasmosis.

Mycoplasma pneumoniae infection has been associated with chronic lung disease. Treatment of chronic pulmonary mycoplasmosis has not been well investigated. BALB/c mice were intranasally inoculated once with M. pneumoniae or with sterile media (uninfected controls). Infected mice were treated with telithromycin or placebo daily for 10 days in the chronic phase of disease (18 months after inoculation). Mice (n=43) were evaluated before therapy and 1 day after completion of telithromycin. Treatment of infected mice with telithromycin at 18 months after infection significantly reduced chronic pulmonary histological inflammation compared with infected mice given placebo; however, this treatment did not improve airway obstruction or airway hyperresponsiveness. Therapy longer than 10 days may be necessary to improve pulmonary function.

Management and outcome of children with skin and soft tissue abscesses caused by community-acquired methicillin-resistant Staphylococcus aureus.

BACKGROUND: Although the epidemiology of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) has been explored in many investigations, management of this emerging infection has not been well-studied. For non-methicillin-resistant Staphylococcus aureus skin and soft tissue abscesses, incision and drainage is generally adequate therapy without the use of antibiotics, but this has not been established for CA-MRSA. METHODS: Children presenting to Children's Medical Center of Dallas for management of skin and soft tissue abscesses caused by culture-proved CA-MRSA were prospectively followed. We analyzed data from the initial evaluation and from two follow-up visits that focused on the management and outcome of CA-MRSA infection. Retrospective chart review was performed 2 to 6 months after the initial visit. RESULTS: Sixty-nine children were identified with culture-proved CA-MRSA skin and soft tissue abscess. Treatment consisted of drainage in 96% of patients and wound packing in 65%. All children were treated with antibiotics. Five patients (7%) were prescribed an antibiotic to which their CA-MRSA isolate was susceptible before culture results were known. Four patients (6%) required hospital admission on the first follow-up; none of these patients had received an antibiotic effective against CA-MRSA. A significant predictor of hospitalization was having a lesion initially >5 cm (P = 0.004). Initial ineffective antibiotic therapy was not a significant predictor of hospitalization (P = 1.0). Of the 58 patients initially given an ineffective antibiotic and managed as outpatients, an antibiotic active against CA-MRSA was given to 21 (36%) patients after results of cultures were known. No significant differences in response were observed in those who never received an effective antibiotic than in those who did. CONCLUSIONS: Incision and drainage without adjunctive antibiotic therapy was effective management of CA-MRSA skin and soft tissue abscesses with a diameter of <5 cm in immunocompetent children.

Analysis of pulmonary inflammation and function in the mouse and baboon after exposure to Mycoplasma pneumoniae CARDS toxin.

Mycoplasma pneumoniae produces an ADP-ribosylating and vacuolating toxin known as the CARDS (Community Acquired Respiratory Distress Syndrome) toxin that has been shown to be cytotoxic to mammalian cells in tissue and organ culture. In this study we tested the ability of recombinant CARDS (rCARDS) toxin to elicit changes within the pulmonary compartment in both mice and baboons. Animals responded to a respiratory exposure to rCARDS toxin in a dose and activity-dependent manner by increasing the expression of the pro-inflammatory cytokines IL-1alpha, 1beta, 6, 12, 17, TNF-alpha and IFN-gamma. There was also a dose-dependent increase in several growth factors and chemokines following toxin exposure including KC, IL-8, RANTES, and G-CSF. Increased expression of IFN-gamma was observed only in the baboon; otherwise, mice and baboons responded to CARDS toxin in a very similar manner. Introduction of rCARDS toxin to the airways of mice or baboons resulted in a cellular inflammatory response characterized by a dose-dependent early vacuolization and cytotoxicity of the bronchiolar epithelium followed by a robust peribronchial and perivascular lymphocytic infiltration. In mice, rCARDS toxin caused airway hyper-reactivity two days after toxin exposure as well as prolonged airway obstruction. The changes in airway function, cytokine expression, and cellular inflammation correlate temporally and are consistent with what has been reported for M. pneumoniae infection. Altogether, these data suggest that the CARDS toxin interacts extensively with the pulmonary compartment and that the CARDS toxin is sufficient to cause prolonged inflammatory responses and airway dysfunction.

Prospective randomized trial of empiric therapy with trimethoprim-sulfamethoxazole or doxycycline for outpatient skin and soft tissue infections in an area of high prevalence of methicillin-resistant Staphylococcus aureus.

To evaluate empirical therapy with trimethoprim-sulfamethoxazole or doxycycline for outpatient skin and soft tissue infections in an area of high prevalence of methicillin-resistant Staphylococcus aureus, a randomized, prospective, open-label investigation was performed. The overall clinical failure rate was 9%, with all failures occurring in the trimethoprim-sulfamethoxazole group. However, there was no significant difference between the clinical failure rate of empirical trimethoprim-sulfamethoxazole therapy and that of doxycycline therapy.

Pneumonia and septicemia caused by Burkholderia thailandensis in the United States.

Burkholderia thailandensis is closely related to Burkholderia pseudomallei, the causative agent of melioidosis. It is generally considered avirulent and previously has been reported to occur only in Southeast Asia. We report the first case of pneumonia and septicemia caused by B. thailandensis in the United States.

Effect of clarithromycin on cytokines and chemokines in children with an acute exacerbation of recurrent wheezing: a double-blind, randomized, placebo-controlled trial.

BACKGROUND: Clarithromycin is postulated to possess immunomodulatory properties in addition to its antimicrobial activity. OBJECTIVE: To evaluate the effect of clarithromycin on serum and nasopharyngeal cytokine and chemokine concentrations in children with an acute exacerbation of recurrent wheezing. METHODS: Children with a history of recurrent wheezing or asthma and who presented with an acute exacerbation of wheezing were enrolled in a double-blind, randomized trial of clarithromycin vs placebo. Concentrations of tumor necrosis factor alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-1beta (IL-1beta), IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, granulocyte-macrophage colony-stimulating factor, RANTES, eotaxin, macrophage inflammatory protein 1alpha, macrophage inflammatory protein 1beta, and monocyte chemoattractant protein 1 were measured in serum and/or nasopharyngeal aspirates before, during, and after therapy. Mycoplasma pneumoniae and Chlamydophila pneumoniae infection were evaluated for by polymerase chain reaction and serologic testing. RESULTS: Nasopharyngeal concentrations of TNF-alpha, IL-1beta, and IL-10 were significantly and persistently lower in children treated with clarithromycin compared with placebo. There tended to be a greater effect of clarithromycin on nasopharyngeal cytokine concentrations in patients with evidence of M. pneumoniae or C. pneumoniae infection. No significant differences were detected in serum cytokines for children treated with clarithromycin compared with placebo. CONCLUSION: Clarithromycin therapy reduces mucosal TNF-alpha, IL-1beta, and IL-10 concentrations in children with an acute exacerbation of recurrent wheezing.

Cryptococcal immune reconstitution inflammatory syndrome: report of four cases in three patients and review of the literature.

Immune reconstitution inflammatory syndromes (IRIS) in patients with AIDS are characterized by atypical manifestations of opportunistic pathogens in patients experiencing improvement in CD4 cell counts following receipt of highly active anti-retroviral therapy (HAART). We report four cases of IRIS due to Cryptococcus neoformans in three patients and review the literature of cryptococcal IRIS in AIDS (an additional 21 episodes). The IRIS presentation was lymphadenitis in all three patients; one patient also had meningeal IRIS. Combining our patients with the literature review revealed the following IRIS presentations: lymphadenitis (n=14), central nervous system (CNS) IRIS (n=10): meningitis in six and mass lesions in four, and pulmonary cavities (n=1). The median CD4 count of cases at the time of initial cryptococcal diagnosis and prior to the start of HAART was 25 cells/microl and the median HIV viral load was 439,053 copies/ml. At time of presentation of the IRIS, the median CD4 count had increased by 197 cells/microl. The median time from initial cryptococcal diagnosis and the start of HAART to the IRIS was 11 months (range 7 weeks to 3 years) and 7 months (range <2 weeks to 22 months), respectively. Patients with CNS IRIS tended to have shorter intervals from initiation of HAART to presentation compared to patients with lymphadenitis: median 3.5 months compared to 7 months. In 24 of 25 cases, the clinical manifestations of the IRIS resolved (range: days to months). Only four patients were given anti-inflammatory medications: corticosteroids in two and non-steroidal anti-inflammatory drugs in two, thus precluding assessment of efficacy. Patients with cryptococcal disease who initiate HAART are at risk for cryptococcal IRIS.

Evaluation of LBM415 (NVP PDF-713), a novel peptide deformylase inhibitor, for treatment of experimental Mycoplasma pneumoniae pneumonia.

Mycoplasma pneumoniae is a major cause of community-acquired pneumonia. We evaluated the efficacy of LBM415, a novel peptide deformylase inhibitor antimicrobial agent, for the treatment of M. pneumoniae pneumonia in a mouse model. Eight-week-old BALB/c mice were intranasally inoculated once with 10(7) CFU of M. pneumoniae. Groups of mice were treated with LBM415 (50 mg/kg of body weight) or placebo subcutaneously daily for 13 days, starting 24 h after inoculation. Groups of mice were evaluated at the baseline; at days of treatment 1, 3, 6, and 13; and at 7 days after treatment. The MIC of LBM415 against M. pneumoniae was <0.005 microg/ml. LBM415-treated mice had significantly lower bronchoalveolar lavage fluid M. pneumoniae concentrations than placebo-treated mice on days 6 and 13 of treatment. Compared with placebo treatment, therapy with LBM415 significantly decreased lung histopathology scores at days 3, 6, and 13 of treatment and at 7 days after treatment. Airway obstruction was significantly lower in LBM415-treated mice than in placebo-treated mice on days 1, 3, and 6 of treatment and after 7 days of therapy, while airway hyperresponsiveness was significantly lower only on day 3 of therapy. The bronchoalveolar lavage fluid concentrations of tumor necrosis factor alpha, gamma interferon (IFN-gamma), interleukin-6 (IL-6), IL-12, KC (functional IL-8), monocyte chemotactic protein 1, macrophage inflammatory protein 1alpha, monokine induced by IFN-gamma, and IFN-inducible protein 10 were significantly reduced in LBM415-treated mice compared with the levels in placebo-treated mice. There were no differences in the bronchoalveolar lavage fluid concentrations of granulocyte-macrophage colony-stimulating factor, IL-1beta, IL-2, IL-4, IL-5, and IL-10 between the two groups of mice. LBM415 therapy had beneficial microbiologic, histologic, respiratory, and immunologic effects on acute murine M. pneumoniae pneumonia.

Microbiologic and immunologic evaluation of a single high dose of azithromycin for treatment of experimental Mycoplasma pneumoniae pneumonia.

We evaluated the efficacy of azithromycin therapy given as a single high dose or divided over 5 days for the treatment of mild experimental Mycoplasma pneumoniae pneumonia. Although both azithromycin regimens significantly reduced quantitative cultures, lung histopathology, and pulmonary cytokines and chemokines, there were no significant differences between the two regimens.

Mycoplasma pneumoniae induces host-dependent pulmonary inflammation and airway obstruction in mice.

Respiratory tract infections result in wheezing in a subset of patients. Mycoplasma pneumoniae is a common etiologic agent of acute respiratory infection in children and adults that has been associated with wheezing in 20-40% of individuals. The current study was undertaken to elucidate the host-dependent pulmonary and immunologic response to M. pneumoniae respiratory infection by studying mice with different immunogenetic backgrounds (BALB/c mice versus C57BL/6 mice). After M. pneumoniae infection, only BALB/c mice developed significant airway obstruction (AO) compared with controls. M. pneumoniae-infected BALB/c mice manifested significantly elevated airway hyperresponsiveness (AHR) compared with C57BL/6 mice 4 and 7 d after inoculation as well as BALB/c control mice. Compared with C57BL/6 mice, BALB/c mice developed worse pulmonary inflammation, including greater peribronchial infiltrates. Infected BALB/c mice had significantly higher concentrations of tumor necrosis factor-alpha, interferon-gamma, interleukin (IL)-1beta, IL-6, IL-12, KC (functional IL-8), and macrophage inflammatory protein 1alpha in the bronchoalveolar lavage fluid compared with infected C57BL/6 mice. No differences in IL-2, IL-4, IL-5, IL-10, and granulocyte/macrophage colony-stimulating factor concentrations were found. The mice in this study exhibited host-dependent infection-related AO and AHR associated with chemokine and T-helper type (Th)1 pulmonary host response and not Th2 response after M. pneumoniae infection.

IL-4-independent pathways exacerbate methacholine-induced airway hyperreactivity during mycoplasma respiratory disease.

BACKGROUND: Mycoplasma respiratory infections are associated with wheezing and exacerbation of airway hyperresponsiveness (AHR) in asthmatic patients. IL-4 is a key cytokine in the development of AHR and airway reconstruction in asthmatic patients and might be an important component in exacerbation of AHR during pulmonary mycoplasma infection. OBJECTIVE: This study evaluates the effect of IL-4 on the severity of methacholine-induced AHR associated with mycoplasma pulmonary mycoplasma infection. METHODS: BALB/c and IL-4 knockout (KO) mice were infected with Mycoplasma pulmonis, and their enhanced pause scores were monitored before and after methacholine inhalation with whole-body plethysmography. RESULTS: IL-4 KO mice showed no difference in histopathology of the lungs before or after Mycoplasma pulmonis infection when compared with BALB/c mice. There was an increase in airway obstruction from days 7 to 21 after infection in both strains of mice, but there was no strain difference in airway resistance-associated mycoplasma disease. However, IL-4 KO mice had significantly higher methacholine-induced AHR after M pulmonis infection when compared with BALB/c mice. There was no difference in AHR between uninfected IL-4 KO and control mice. CONCLUSION: In contrast to our hypothesis, IL-4-independent pathways exacerbate methacholine-induced AHR and promote airway obstruction during the pathogenesis of mycoplasma respiratory disease.

It is safe to stop antiretroviral therapy in patients with preantiretroviral CD4 cell counts >250 cells/microL.

OBJECTIVE: To study clinical, immunologic, and virologic outcomes in patients who stop antiretroviral therapy (ART) with relatively preserved CD4 cell counts. DESIGN AND METHODS: Patients with a documented CD4 cell count >250 cells/microL who stopped ART for any reason for at least 5 weeks were studied. Relevant clinical and laboratory data were collected using a standardized data collection form. MAIN OUTCOME MEASURES: Patients were monitored for outcomes including Centers for Disease Control (CDC) category B or C events, time to restarting ART, and time to reaching a CD4 cell count of < or = 250 cells/microL. RESULTS: A total of 107 patients were included. The median time on ART was 45 months and median number of antiretroviral medications was 4. The median pre-ART CD4 cell count and HIV viral load were 463 cells/microL and 4.35 log copies/mL, respectively. The median CD4 cell at time of ART stop was 739 cell/microL. The slope of the CD4 decrease was 65 cells/mo in the first 2 months, which was greater than the subsequent decline of 8 cells/mo thereafter (P < 0.01). Similarly the median viral load increase was 2.54 log copies/mL in the first 2 months after stopping and was unchanged after that point. Two patients experienced the retroviral rebound syndrome after ART cessation but no CDC category B or C events were observed during 10 months of follow-up. The median time from stopping ART to reaching the combined endpoint of CD4 <250 or restarting ART was 8.9 months. In multivariate analysis, pre-ART CD4 cell count >250 was protective of reaching the combined endpoint (odds ratio = 0.156, P = 0.03). Other predictors of reaching the combined endpoint in multivariate analysis were older age and number of prior ART agents. Patients who restarted ART had a favorable virologic and immunologic response. CONCLUSIONS: Patients with relatively high CD4 cell counts prior to starting ART did well after stopping ART. Pre-ART CD4 cell count can be used to predict outcomes after ART cessation.

Diagnostic utility and clinical significance of naso- and oropharyngeal samples used in a PCR assay to diagnose Mycoplasma pneumoniae infection in children with community-acquired pneumonia.

PCR assays of naso- and oropharyngeal samples among hospitalized children appear equally effective for the diagnosis of serologically confirmed community-acquired mycoplasmal pneumonia. However, the combination of results from both sites yields optimal sensitivity (57%), specificity (98%), and positive (92%) and negative (82%) predictive values when compared with Mycoplasma pneumoniae enzyme-linked immunosorbent assay.

A randomized, double-blind, placebo-controlled trial of dexamethasone in severe respiratory syncytial virus (RSV) infection: effects on RSV quantity and clinical outcome.

Forty-one previously healthy children <2 years of age who required mechanical ventilation for respiratory syncytial virus (RSV) infection were randomized to receive dexamethasone (0.5 mg/kg; n=22) or saline placebo (n=19) intravenously every 12 h for 4 days. RSV quantity was measured by quantitative plaque assay in fresh tracheal and nasal aspirates obtained at intervals of 24+/-3 h on days 0, 1, 2, 5, and 7 following entry. Analysis by linear mixed-effects modeling demonstrated a significantly greater decline in mean tracheal RSV quantity in the placebo group than in the dexamethasone group from day 0 to day 1 (0.82 vs. 0.21 log pfu/mL; P=.01) and from day 0 to day 2 (1.45 vs. 0.53 log pfu/mL; P=.03). No differences were found between groups in nasal RSV quantity, white blood cell counts in tracheal or nasal aspirates, serum neutralizing antibody titers during convalescence, or duration of mechanical ventilation, intensive care unit stay, or hospital stay.