Facilitators and barriers to adolescent participation in a TB clinical trial.

The inclusion of adolescents in TB drug trials is essential for the development of safe, child-friendly regimens for the prevention and treatment of TB. TB Trials Consortium Study 31/AIDS Clinical Trials Group A5349 (S31/A5349) enrolled adolescents as young as 12 years old. We assessed investigator and coordinator described facilitators and barriers to adolescent recruitment, enrollment, and retention.Interviews were conducted with six investigators from sites that enrolled adolescent participants and six investigators from non-enrolling sites. Additionally, two focus groups were conducted with study coordinators from enrolling sites and two focus groups with non-enrolling sites. Discussions were transcribed, analyzed, summarized, and summaries were reviewed by Community Research Advisors Group members and research group representatives for content validity.Investigators and coordinators attributed the successful enrollment of adolescents to the establishment and cultivation of external partnerships, flexibility to accommodate adolescents' schedules, staff engagement, recruitment from multiple locations, dedicated recruitment staff working onsite to access potential participants, creation of youth-friendly environments, and effective communications. Non-enrolling sites were mainly hindered by regulations. Suggestions for improvement in future trials focused on study planning and site preparations.Proactive partnerships and collaboration with institutions serving adolescents helped identify and reduce barriers to their inclusion in this trial..

A human IFNGR1 small deletion hotspot associated with dominant susceptibility to mycobacterial infection.

The immunogenetic basis of severe infections caused by bacille Calmette-Guérin vaccine and environmental mycobacteria in humans remains largely unknown. We describe 18 patients from several generations of 12 unrelated families who were heterozygous for 1 to 5 overlapping IFNGR1 frameshift small deletions and a wild-type IFNGR1 allele. There were 12 independent mutation events at a single mutation site, defining a small deletion hotspot. Neighbouring sequence analysis favours a small deletion model of slipped mispairing events during replication. The mutant alleles encode cell-surface IFNgamma receptors that lack the intra-cytoplasmic domain, which, through a combination of impaired recycling, abrogated signalling and normal binding to IFNgamma exert a dominant-negative effect. We thus report a hotspot for human IFNGR1 small deletions that confer dominant susceptibility to infections caused by poorly virulent mycobacteria.

Neutrophil-specific granule deficiency results from a novel mutation with loss of function of the transcription factor CCAAT/enhancer binding protein epsilon.

Neutrophil-specific granule deficiency (SGD) is a rare disorder characterized by recurrent pyogenic infections, defective neutrophil chemotaxis and bactericidal activity, and lack of neutrophil secondary granule proteins. CCAAT/enhancer binding protein (C/EBP)epsilon, a member of the leucine zipper family of transcription factors, is expressed primarily in myeloid cells, and its knockout mouse model possesses distinctive defects, including a lack of neutrophil secondary granule proteins. Sequence analysis of the genomic DNA of a patient with SGD revealed a five-basepair deletion in the second exon of the C/EBPepsilon locus. The predicted frame shift results in a truncation of the 32-kD major C/EBPepsilon isoform, with loss of the dimerization domain, DNA binding region, and transcriptional activity. The multiple functional defects observed in these early neutrophil progenitor cells, a consequence of C/EBPepsilon deficiency, define SGD as a defect in myelopoiesis and establish the requirement for C/EBPepsilon for the promyelocyte-myelocyte transition in myeloid differentiation.

Mutation in the signal-transducing chain of the interferon-gamma receptor and susceptibility to mycobacterial infection.

IFN-gamma is critical in the immune response to mycobacterial infections, and deficits in IFN-gamma production and response have been associated with disseminated nontuberculous mycobacterial infections. Mutations in the IFN-gamma receptor ligand-binding chain (IFNgammaR1) have been shown to confer susceptibility to severe infection with nontuberculous mycobacteria. However, mutations in the IFN-gamma receptor signal-transducing chain (IFNgammaR2) have not been described. We describe a child with disseminated Mycobacterium fortuitum and M. avium complex infections and absent IFN-gamma signaling due to a mutation in the extracellular domain of IFNgammaR2. In vitro cytokine production by patient PBMCs showed 75% less PHA-induced IFN-gamma production than in normal cells, while patient PHA-induced TNF-alpha production was normal. The normal augmentation of TNF-alpha production when IFN-gamma was added to endotoxin was absent from patient cells. Expression of IFNgammaR1 was normal, but there was no phosphorylation of Stat1 in response to IFN-gamma stimulation. DNA sequence analysis of the gene for IFNgammaR2 showed a homozygous dinucleotide deletion at nucleotides 278 and 279, resulting in a premature stop codon in the protein extracellular domain. This novel gene defect associated with disseminated nontuberculous mycobacterial infection emphasizes the critical role that IFN-gamma plays in host defense against mycobacteria.

Interferon-gamma and interleukin-12 pathway defects and human disease.

A genetic component to human mycobacterial disease susceptibility has long been postulated. Over the past five years, mutations in the interferon-gamma (IFNgamma) receptor, IL-12 receptor beta1 (IL-12Rbeta1), and IL-12 p40 genes have been recognized. These mutations are associated with heightened susceptibility to disease caused by intracellular pathogens including nontuberculous mycobacteria, vaccine-associated bacille Calmette Guerin (BCG), Salmonella species, and some viruses. We describe the genotype-phenotype correlations in IFNgamma receptor, IL-12Rbeta1, and IL-12 p40 deficiency, and discuss how study of these diseases has enhanced knowledge of human host defense against mycobacteria and other intracellular pathogens.

Defining the optimal dose of rifapentine for pulmonary tuberculosis: Exposure-response relations from two phase II clinical trials.

Rifapentine is a highly active antituberculosis antibiotic with treatment-shortening potential; however, exposure-response relations and the dose needed for maximal bactericidal activity have not been established. We used pharmacokinetic/pharmacodynamic data from 657 adults with pulmonary tuberculosis participating in treatment trials to compare rifapentine (n = 405) with rifampin (n = 252) as part of intensive-phase therapy. Population pharmacokinetic/pharmacodynamic analyses were performed with nonlinear mixed-effects modeling. Time to stable culture conversion of sputum to negative was determined in cultures obtained over 4 months of therapy. Rifapentine exposures were lower in participants who were coinfected with human immunodeficiency virus, black, male, or fasting when taking drug. Rifapentine exposure, large lung cavity size, and geographic region were independently associated with time to culture conversion in liquid media. Maximal treatment efficacy is likely achieved with rifapentine at 1,200 mg daily. Patients with large lung cavities appear less responsive to treatment, even at high rifapentine doses.

Two-stage activity-safety study of daily rifapentine during intensive phase treatment of pulmonary tuberculosis.

BACKGROUND: Rifapentine (RPT) has potent activity against Mycobacterium tuberculosis; however, the optimal dose for anti-tuberculosis treatment is unknown. OBJECTIVE: To determine the antimicrobial activity, safety and tolerability of RPT 450 mg or 600 mg administered daily during the first 8 weeks of treatment for pulmonary tuberculosis (TB). DESIGN: In a two-stage, randomised open-label study, adults with sputum smear-positive TB were randomised to receive RPT 450 mg, RPT 600 mg or rifampicin (RMP) 600 mg daily for 8 weeks with isoniazid, pyrazinamide and ethambutol. The primary endpoint was sputum culture status on Löwenstein-Jensen (LJ) medium at completion of 8 weeks of treatment. RESULTS: A total of 153 participants were enrolled. Both RPT regimens met pre-specified criteria to advance to stage 2. At completion of 8 weeks of treatment, LJ culture conversion occurred in 85% (35/41), 96% (43/45) and 94% (34/36) of participants in the RPT 450 mg, RPT 600 mg and RMP groups, respectively. The proportions of participants discontinuing treatment were similar (respectively 1/54 [2.0%], 1/51 [2.0%] and 4/48 [8.3%] in the RPT 450 mg, RPT 600 mg and RMP groups), as were ⩾grade 3 adverse events (0/54 [0%], 1/51 [2.0%] and 4/48 [8.3%]). CONCLUSIONS: There was a trend towards greater efficacy with RPT 600 mg than with RPT 450 mg. Daily RPT was safe and well-tolerated.

Isolated intracranial infection with Mycobacterium avium complex.

UNLABELLED: Mycobacterium avium-M. complex (MAC) has been linked to devastating respiratory and systemic illnesses in patients, especially in those who are immunosuppressed. The purpose of this study is to describe a case of isolated central nervous system (CNS) infection with MAC. This is a single case report of a patient with isolated intracranial mycobacterial infection. SETTING: the patient was treated and the immunohistochemical investigations were undertaken at the National Institutes of Health in Bethesda, Maryland, USA. INTERVENTION: the patient initially was treated with a cocktail of antimycobacterial medications. However, because his disease was refractory, he underwent a suboccipital craniotomy and evacuation of his cerebellar mass. The patient was determined to have a low production of interferon-gamma (INF-gamma) and tumor necrosis factor-alpha (TNF-alpha) when compared to normal values. Despite extensive radiographic imaging studies and biopsies, there was no evidence of another focus of MAC infection in this patient. We conclude that intracranial infectious lesions in patients such as ours should be treated with conventional systemic antibiotic regimens as the first-line of therapy. We suggest neurosurgical intervention in medically refractory cases of intracranial infections.

Cost-effectiveness of routine diagnostic evaluation of pulmonary tuberculosis in a primary care unit in Brazil.

SETTING: Primary health care unit in Rio de Janeiro City, Brazil. OBJECTIVE: To estimate and compare the cost-effectiveness of strategies used for passive case finding of pulmonary tuberculosis (PTB) cases using tests available at the primary care level. DESIGN: Data on PTB suspects were reviewed, and a decision model was developed using sputum smear microscopy and chest radiography (CXR) according to three different strategies for PTB detection. A cost-effectiveness analysis was performed to estimate the cost per correct PTB diagnosis. Mycobacterial culture was used to calculate the effectiveness of the strategies. Unit costs of health resource utilisation were obtained from the payer's perspective (the Brazilian Public Health System). RESULTS: For the evaluation of 254 PTB suspects, the total costs of strategies ranged from US$5369 to US$5944; the probability of a correct PTB diagnosis ranged from 0.66 to 0.86; the number of visits required to complete the diagnostic process ranged from two to three, and cost per PTB case identified ranged from US$47.93 to US$53.07. The cost-effectiveness of the three strategies studied varied between US$56.69 and US$72.55 per correct PTB case detected. CONCLUSION: A strategy in which sputum smears and CXR were requested for all PTB suspects at the initial evaluation was cost-effective, had a high probability of correct PTB diagnosis and could be accomplished in two visits.

Safety and pharmacokinetics of escalating daily doses of the antituberculosis drug rifapentine in healthy volunteers.

Rifapentine (RP T) is an antituberculosis drug that may shorten treatment duration when substituted for rifampin (RI F).The maximal tolerated daily dose of RP T and its potential for cytochrome 3A4 induction and autoinduction at clinically relevant doses are unknown. In this phase I, dose-escalation study among healthy volunteers, daily doses as high asa prespecified maximum of 20 mg/kg/day were well tolerated. Steady-state RP T concentrations increased with dose from 5 to 15 mg/kg, but area under the plasma concentration­time curve (AU C0­24) and maximum concentration (Cmax)were similar in the 15- and 20-mg/kg cohorts. Although RP T pharmacokinetics (PK) appeared to be time-dependent,accumulation occurred with daily dosing. The mean AU C0­12 of oral midazolam (MDZ), a cytochrome 3A (CYP 3A) probe drug, was reduced by 93% with the coadministration of RPT and by 74% with the coadministration of RIF (P < 0.01).Changes in the oral clearance of MDZ did not vary by RP T dose. In conclusion, RP T was tolerated at doses as high as20 mg/kg/day, its PK were less than dose-proportional, and its CYP 3A induction was robust.

QuantiFERON-TB Gold In-Tube for the detection of Mycobacterium tuberculosis infection in children with household tuberculosis contact.

SETTING: Improved strategies are needed for detecting Mycobacterium tuberculosis infection in children in TB-endemic settings. OBJECTIVE: To determine the prevalence of M. tuberculosis infection by tuberculin skin testing (TST) and by the QuantiFERON-TB Gold In-Tube (QFT-GIT) test in children with an adult household contact with pulmonary TB in South Africa. DESIGN: Cross-sectional study. RESULTS: A total of 167 adult pulmonary TB cases (153/167, 92% human immunodeficiency virus [HIV] infected) and 270 pediatric contacts (median age 6 years, 14/270, 5% HIV-infected) were enrolled. All children completed QFT-GIT testing and 254 (94.1%) completed TST testing. Prevalence of M. tuberculosis infection was 28% (71/254, 95%CI 23-34) using TST (5 mm cut-off) and 29% (79/270, 95%CI 24-35) using QFT-GIT (P = 0.49). Agreement between TST and QFT-GIT was 81% (kappa 0.58). Nineteen (7%) QFT-GIT results were indeterminate. Children aged <2 years were more likely than older children to have indeterminate QFT-GIT results (aOR 5.7, 95%CI 1.5-22, P = 0.01) and discordant QFT-GIT and TST results (aOR 3.5, 95%CI 1.7-7.6, P = 0.001). CONCLUSION: Prevalence of M. tuberculosis infection in pediatric contacts was high regardless of the diagnostic method used. TST should not be excluded for the detection of pediatric M. tuberculosis infection in this setting, but QFT-GIT may be a feasible alternative in children aged ≥ 2 years.

Alternative anti-tuberculosis regimen including ofloxacin for the treatment of patients with hepatic injury.

SETTING: Tuberculosis (TB) clinic of a university-based public hospital in Rio de Janeiro city, Brazil. OBJECTIVE: To describe treatment outcomes for TB patients with liver injury who received a 12-month regimen of ethambutol (E, EMB) and ofloxacin (O, OFL), including streptomycin (S, SM) for the first 3 months (3SEO/9EO) under routine clinical care conditions. DESIGN: A retrospective study of a cohort of TB patients prescribed 3SEO/9EO was conducted over a 66-month period. Data were obtained by review of existing medical records. Primary outcomes assessed were cure, treatment failure, treatment default, TB relapse and death. RESULTS: Outcomes were assessed for 40 patients with hepatic injury who met study criteria. Twenty-three (58%) were male and 13 (33%) were human immunodeficiency virus seropositive. Thirty-four (85%) patients were cured. Three patients (7.5%) defaulted from treatment, and three other patients died (7.5%). There were no treatment failures or relapses during 2 years of follow-up. Clinically recognized drug toxicity occurred in five patients (12.5%), and in each case was attributed to SM. CONCLUSION: In this series of TB patients with serious liver injury, 3SEO/9EO was well-tolerated, and it was effective in 85% of patients when used under routine clinical care conditions.

561del4 defines a novel small deletion hotspot in the interferon-gamma receptor 1 chain.

Patients with a dominant small deletion (818del4, hotspot) in the interferon-gamma receptor 1 (IFNGR1) gene (6q23-q24) and increased susceptibility to mycobacterial infections have been recently reported. We describe a female patient homozygous for a 4-bp deletion in exon 5 of IFNGR1 (561del4) who developed postvaccinal disseminated Bacille Calmette-Guerin infection. She was born to unrelated Argentinean parents, each of whom was heterozygous for this mutation. 561del4 has been previously described as a maternally inherited mutation in a compound heterozygous German patient. By single nucleotide polymorphism analysis of the areas surrounding the deletion, we showed the independent inheritance of 561del4 in three heterozygous carriers. Polypurine runs and "direct repeats," previously shown to be associated with areas of recurrent small deletions, were found in the flanking region of 561del4. The independent inheritance of three identical mutational events defines 561del4 as a new hotspot in the IFNGR1 gene.

Adjunctive corticosteroid therapy for patients whose treatment for disseminated Mycobacterium avium complex infection has failed.

Patients with AIDS and disseminated Mycobacterium avium complex (MAC) infection can have progressive disease despite combination antimycobacterial therapy. Our goal was to determine the utility of corticosteroids as adjunctive therapy for AIDS patients with disseminated MAC infection and refractory symptoms despite combination antimycobacterial therapy. We retrospectively reviewed 12 consecutive patients whose therapy for MAC infection clinically failed and who subsequently received low-dose oral corticosteroids in addition to continued combination antimycobacterial therapy. With the addition of corticosteroids, 11 of 12 patients experienced a rapid improvement in symptoms, with diminished or resolved fevers and night sweats, and an increased sense of well-being and energy. Ten of 12 patients gained weight; the differences between weights before and after steroid therapy were statistically significant (P = .003, paired Student's t-test), and the weights achieved were similar to baseline weights prior to the development of MAC infection (P = .2). Mean survival after diagnosis of MAC infection was 17.5 months. New opportunistic processes developed in seven patients during corticosteroid therapy. However, given the severely immunocompromised status of these patients, it was not possible to attribute the development of new opportunistic processes directly to corticosteroid therapy.

Burkholderia pseudomallei infection in a Puerto Rican patient with chronic granulomatous disease: case report and review of occurrences in the Americas.

Burkholderia species, notably Burkholderia cepacia and Burkholderia gladioli, are important pathogens in patients with chronic granulomatous disease (CGD). Burkholderia pseudomallei, the causative agent of melioidosis, is endemic in Southeast Asia and northern Australia but is a rare pathogen in other parts of the world. We describe the occurrence of B. pseudomallei infection in a Puerto Rican patient with CGD. This is one of only a small number of documented cases of melioidosis autochthonous to the Americas and is the first reported case of B. pseudomallei infection in a CGD patient from the Americas. We conclude that B. pseudomallei, like B. cepacia and B. gladioli, should be considered a potential pathogen in patients with CGD and that melioidosis should be considered in the differential diagnosis for ill residents of or travelers to Puerto Rico.

Human immunodeficiency virus-seronegative adults with extrapulmonary tuberculosis have abnormal innate immune responses.

Extrapulmonary tuberculosis is presumably a marker of underlying immunodeficiency, but cytokine response pathways in these patients have not been well studied. Cytokine responses of peripheral blood mononuclear cells from human immunodeficiency virus-seronegative adults with prior culture-confirmed extrapulmonary tuberculosis were compared with those of persons with latent Mycobacterium tuberculosis infection. Mitogen-stimulated interferon (IFN)-gamma production, interleukin (IL)-12 production, and IFN-gamma receptor- and IL-12 receptor-mediated cytokine production did not differ between case patients and control patients. However, median resting IL-8 production was significantly lower in case patients than control patients (8051 vs. 19,290 pg/mL; P=.009). In addition, the median tumor necrosis factor (TNF)-alpha response was lower in case patients than control patients after stimulation with lipopolysaccharide (833 vs. 1149 pg/mL; P=.06) and lipopolysaccharide plus IFN-gamma (3301 vs. 4411 pg/mL; P=.04). These abnormalities in resting IL-8 and lipopolysaccharide-induced TNF-alpha production were not associated with IFN-gamma or IL-12 abnormalities and were detected up to several years after cure of disease, suggesting an abnormality in innate immunity.

Fulminant babesiosis treated with clindamycin, quinine, and whole-blood exchange transfusion.

BACKGROUND: Babesiosis is an increasingly recognized parasitic infection with manifestations that range from a subclinical or mild flu-like illness to life-threatening disease. Risk factors that may be associated with a more severe clinical course include immunosuppression, splenectomy, and advanced age. The most effective chemotherapeutic regimen, clindamycin plus quinine, is sometimes ineffective in cases of severe disease. CASE REPORT: A previously healthy, 58-year-old man was infected by Babesia microti, presumably through a tick bite. He developed fulminant disease characterized by severe hemolytic anemia, disseminated intravascular coagulation, acute renal failure, and respiratory failure. There was no history of splenectomy or immunodeficiency. He was given oral clindamycin (300 mg/4x/day) 2 days before admission. Oral quinine (650 mg/3x/day) was added upon hospitalization. There was no clinical improvement despite antibiotic therapy with clindamycin and quinine. On the second hospital day, a whole-blood exchange transfusion was performed to simultaneously lower the parasite load and replace the patient's plasma. With an automated blood cell separator, 87 percent of the patient's total blood volume was exchanged. As replacement fluid, 6.7 L of packed RBCs reconstituted with FFP (average Hct, 33%) was used. The patient's Hct increased from 26.9 percent before the exchange to 28.3 percent after the exchange. The percentage of parasitized RBCs decreased from 13.8 percent just before exchange to 4.2 percent immediately after exchange. There was rapid clinical improvement after the whole-blood exchange transfusion. The patient's subsequent clinical course was marked by a disappearance of the parasitemia and continued slow, general improvement. Therapy with clindamycin was continued for 14 days after the exchange transfusion and quinine for 17 days. CONCLUSION: In cases of severe babesiosis, prompt institution of whole-blood exchange transfusion, in combination with appropriate antimicrobial therapy, can be life-saving.

Detection of intracellular phosphorylated STAT-1 by flow cytometry.

We have applied flow cytometry to the investigation of interferon-gamma activation of human monocytes. This approach uses monoclonal antibodies that distinguish between the native and phosphorylated forms of STAT-1. It enables rapid and quantitative assessment of STAT-1 phosphorylation on a discrete cell basis and is both more sensitive and less time consuming than immunoblotting. Furthermore, it allows for discrimination between a mixture of cells that differ in their response to interferon-gamma. This approach should allow for the evaluation of different intracellular signaling pathways using a combination of monoclonal reagents that are specific for native and activation modified proteins. Application of this form of testing should prove valuable in screening for signaling defects in selected patients with recurrent infections. In addition, this technique should permit dissection of a full range of cellular signaling pathways at the protein level.

Viral infections in interferon-gamma receptor deficiency.

Interferon-gamma receptor deficiency is a recently described immunodeficiency that is associated with onset of severe mycobacterial infections in childhood. We describe the occurrence of symptomatic and often severe viral infections in 4 patients with interferon-gamma receptor deficiency and mycobacterial disease. The viral pathogens included herpes viruses, parainfluenza virus type 3, and respiratory syncytial virus. We conclude that patients with interferon-gamma receptor deficiency and mycobacterial disease have increased susceptibility to some viral pathogens.

Abnormal regulation of interferon-gamma, interleukin-12, and tumor necrosis factor-alpha in human interferon-gamma receptor 1 deficiency.

Mycobacterial infections are critically controlled by interferon-gamma (IFN-gamma) and the cellular responses it elaborates, as shown by patients with mutations in the IFN-gamma receptor ligand-binding chain (IFN-gamma R1) who have disseminated nontuberculous mycobacterial infections. The immunologic sequelae of IFN-gamma R1 deficiency were characterized in 2 unrelated patients from the Indian subcontinent with novel homozygous recessive IFN-gamma R1 mutations. In vitro, these patients' peripheral blood mononuclear cells produced 10% of normal IFN-gamma and interleukin-12 (IL-12) in response to phytohemagglutinin (PHA) but normal amounts of IFN-gamma in response to PHA plus IL-12. Tumor necrosis factor-alpha (TNF-alpha) production was normal in response to endotoxin and to PHA but was not augmented by the addition of IFN-gamma. An abnormal phenotype was not found in heterozygous patient relatives. These patients demonstrate the critical role that the IFN-gamma receptor plays in the regulation of IFN-gamma, IL-12, and TNF-alpha.