Mechanisms of latency in Mycobacterium tuberculosis.

Mycobacterium tuberculosis can persist within the human host for years without causing disease, in a syndrome known as latent tuberculosis (TB). As one-third of the world population has latent TB, placing them at risk for active TB, the mechanisms by which M. tuberculosis establishes a latent metabolic state, eludes immune surveillance and responds to triggers that stimulate reactivation are a high priority for the future control of TB.

Empiric use of vancomycin during prolonged treatment-induced granulocytopenia. Randomized, double-blind, placebo-controlled clinical trial in patients with acute leukemia.

Because gram-positive infections cause morbidity following intensive antileukemic chemotherapy, the effects of vancomycin versus placebo were evaluated in a randomized, double-blind, placebo-controlled trial in 60 adult patients with acute leukemia and first infectious fever during prolonged (mean of 32 days) granulocytopenia. Gram-positive sepsis was associated with first fever in 17 (28 percent) of the 60 patients. None of 31 patients randomly assigned to receive vancomycin demonstrated gram-positive infection, whereas 16 of 22 patients randomly assigned to receive placebo subsequently had gram-positive infection (seven had sepsis, and nine had local infections; p less than 0.005). All patients with gram-positive infection were then given vancomycin, and all showed prompt clinical responses. The predominant gram-positive organism causing infection was beta-lactam-resistant Staphylococcus epidermis (19 of 44 isolates). Patients randomly assigned to receive vancomycin had more rapid resolution of first infectious fever and fewer total febrile days during the granulocytopenic course than did patients randomly assigned to receive placebo. Although vancomycin had no effect on the presence or absence of documented fungal infection, patients treated with vancomycin received empiric amphotericin B for recurrent or persistent fever later (mean of 14 days after initial antibiotic coverage was begun) than did patients receiving placebo (mean of 9.9 days; p less than 0.005), and thus received fewer total days of empiric amphotericin B therapy (mean of 16.3 days) than did patients given placebo (mean of 24.6 days; p less than 0.01). These data demonstrate that empiric use of vancomycin reduces the morbidity of gram-positive infections following intensive antileukemic therapy and decreases the need for empiric use of toxic amphotericin B.

A new class of antituberculosis agents.

Long-chain lipid envelopes are characteristic of mycobacteria such as those that cause tuberculosis and leprosy. Inhibition of fatty acid synthesis or elongation is a strategy demonstrated to be clinically effective against M. tuberculosis. A new class of compounds designed to inhibit the beta-ketoacyl synthase reaction of fatty acid synthesis has been developed. Of >30 compounds described, the most active were acetamides containing alkylsulfonyl substituents. Inhibitory activities were acutely sensitive to net charge, chain length, and degree of unsaturation. The most active compound 5 (alkyl = C(10)) contained a single methylene spacer between the sulfone and carboxamide and exhibited an MIC of 0.75-1.5 microg/mL, comparable to first-line antituberculosis drugs. These compounds are species-specific, exhibiting no significant activity against bacterial species other than M. tuberculosis and closely related strains. The synthesis, biological activity, and specificity of these compounds are described.

Colonization by Streptococcus penumoniae in human immunodeficiency virus-infected children.

OBJECTIVE: Children with HIV infection are particularly susceptible to invasive pneumococcal disease, yet the effect of HIV infection and its medical management on colonization and resistance to antibiotics are poorly described. To provide a basis for medical practice, we determined the prevalence of nasopharyngeal colonization and antibiotic resistance of Streptococcus pneumoniae in children with HIV infection. METHODS: Cross-sectional prevalence sample of children attending the pediatric HIV and pulmonary clinics to examine nasopharyngeal colonization with S. pneumoniae and antibiotic resistance to beta-lactams and trimethoprim-sulfamethoxazole (T/S). Subjects were matched by age and date of clinic visit. RESULTS: The colonization rate with S. pneumoniae of HIV-infected and -indeterminate children was equal to that of controls (20% vs. 19%). HIV infection, CDC staging or receipt of oral antibiotic therapy did not affect colonization. Isolates from HIV-infected and -indeterminate children were less likely to be penicillin-resistant than those from controls (18% vs. 50%). There was no difference in pneumococcal resistance to T/S among isolates from subjects and controls, despite 72% T/S use in the HIV clinic. CONCLUSION: Colonization with S. pneumoniae in HIV disease is no different from that of comparable children. The high incidence of pneumococcal disease and prophylaxis with T/S are not related to nasopharyngeal colonization. Antibiotic prophylaxis of HIV-infected children does not necessarily lead to increased resistance of S. pneumoniae.

Strategies to prevent or control infections after bone marrow transplants.

Patients receiving bone marrow transplants are at risk of life-threatening infections early post-transplant. This predisposition results from extensive mucosal damage and severe granulocytopenia. Common causes of infection include bacteria and fungi. Infections with opportunistic pathogens occur later and are associated with defects in cellular and/or humoral immunity. The most common sites of infections are the gastrointestinal tract, oropharynx, lung, skin and indwelling vascular catheters. Empiric approaches designed to treat common bacterial and fungal pathogens are generally effective as are measures designed to prevent dissemination of infections. These approaches are also used to prevent fungal infections.

Topical ciprofloxacin treatment of Pseudomonas keratitis in rabbits.

Ciprofloxacin is a new quinolone antibiotic that is highly active in vitro against Pseudomonas aeruginosa. A rabbit model of bacterial keratitis was used to assess the in vivo efficacy of topical ciprofloxacin. Albino rabbits received intrastromal injections of 5 X 10(2) aminoglycoside-resistant P aeruginosa organisms. At five hours after inoculation, ciprofloxacin (3 mg/mL) therapy was initiated (one drop every 30 minutes for 12 hours). Corneal tissue was then excised for bacterial colony counts. No organisms were recovered from ciprofloxacin-treated eyes, compared with an average of 3.1 X 10(7) organisms per milliliter recovered from untreated controls. This model suggests that topical ciprofloxacin may be clinically useful in the treatment of aminoglycoside-resistant P aeruginosa keratitis.

Use of collagen corneal shields in the treatment of bacterial keratitis.

We used an animal model of Pseudomonas keratitis to compare treatment by topical tobramycin with and without the presence of a commercially available collagen corneal shield. Pilot studies showed a significant, 30-fold increase in penetration of tobramycin into the anterior chamber in eyes with a collagen shield in place. Twenty albino rabbit eyes were inoculated with P. aeruginosa to produce stromal keratitis. After 12 hours of topical tobramycin dosing, eyes with a collagen corneal shield in place had a statistically significant (P less than .01) decrease in colony forming unit counts in comparison to treated eyes without a shield and control eyes.

Topical imipenem therapy of aminoglycoside-resistant Pseudomonas keratitis in rabbits.

We used a rabbit model of bacterial keratitis to assess in vivo efficacy of topical imipenem, a highly potent beta-lactam antibiotic with an unusually broad spectrum of activity, including aminoglycoside-resistant Pseudomonas aeruginosa. Albino rabbits received intrastromal injections of 5 x 10(2) organisms of an aminoglycoside-resistant strain of P. aeruginosa. At five hours postinoculation, imipenem (5 mg/ml) therapy was initiated using one drop per 30 minutes for 12 hours. Corneal tissue was then excised for colony forming unit counts. Imipenem was highly effective in reducing colony forming unit counts to zero in comparison to 4.1 x 10(5) organisms for untreated controls. A second regimen beginning 24 hours postinoculation of one drop per hour for 24 hours was also successful in significantly reducing colony forming units vs controls (P less than .05). These data suggest that topical imipenem may have clinical applicability in the treatment of P. aeruginosa keratitis.

Contaminated fractures of the tibia: a comparison of treatment modalities in an animal model.

External fixation is the current standard treatment for skeletal stabilization of open tibial fractures, but intramedullary fixation techniques have become increasingly popular. The aim of this study was to compare, in an animal model, the susceptibility to infection of contaminated fractures stabilized with external fixation with that of contaminated fractures fixed with intramedullary locking nails with or without reaming. A unilateral osteotomy of the tibia was performed in 15 goats under general anesthesia. Each osteotomy was stabilized with either (a) a unilateral biplanar external fixator, (b) an 8 mm diameter intramedullary rod inserted without reaming of the medullary cavity, or (c) a 10 mm diameter rod inserted after reaming. A standardized inoculum of Staphylococcus aureus, 10(3) colony forming units per milliliter, was placed at each osteotomy site on a piece of absorbable gelatin sponge, to simulate contamination of an open fracture. Antibiotics were not administered. The animals were allowed full activity after the procedure. Fourteen days postoperatively, the animals were killed, radiographs of the tibiae were taken, and the tibiae were harvested in a sterile manner. Multiple specimens for quantitative microbiological analysis were taken from the fracture site and from sites 3 cm distal and 6 cm proximal to the fracture. Additional specimens of bone were taken for histological study. Clinical, radiographic, and microbiological analysis demonstrated that, in this animal model, there were significantly fewer and less severe infections in fractures fixed with external fixation than in those fixed with an intramedullary nail with or without reaming. There was marked cortical necrosis in tibiae that had been fixed with nailing and reaming.

Use of collagen corneal shields versus soft contact lenses to enhance penetration of topical tobramycin.

We compared the corneal penetration in rabbits of topical tobramycin in the presence of collagen corneal shields and bandage soft contact lenses. A collagen corneal shield was placed on six albino rabbit eyes, while therapeutic soft contact lenses (61.4% poly-2-hydroxyethyl-methacrylate/38.6% water) were placed on six eyes. Four control eyes received no shield or contact lens. Topical tobramycin was applied to all 16 eyes every five minutes for six doses. Samples of aqueous humor were removed at 15 and 60 minutes following the last dose. Collagen corneal shields allowed a significant (P less than .05) increase in tobramycin penetration into the anterior chamber at 60 minutes compared with hydrophilic soft contact lenses or controls.

Management of infectious complications of acute leukemia and antileukemia therapy.

The patient with acute leukemia is predisposed to infection by bone marrow failure that leads to absence of granulocytes, by extramedullary leukemia infiltration that leads to barrier breakdown, and by cytotoxic antileukemia therapy that exaggerates both the hematopoietic and the tissue mucosal defects. Empiric approaches tailored to treat commonly occurring bacterial and fungal infections have successfully decreased the morbidity and mortality from overwhelming infection in these compromised patients. More recently, prophylaxis directed against dissemination of pathogens from specific sites has had a positive impact in preventing the clinical and microbiological manifestations of infection during profound aplasia. The approaches that have been successful in preventing and treating bacterial infections are being applied to the increasingly prevalent fungal infections that occur later during the granulocytopenic course, with encouraging results.

Determination of ticarcillin levels in serum by high-pressure liquid chromatography.

A high-pressure liquid chromatographic method for determining the concentrations of ticarcillin in serum was developed and compared, with 93 patient sera, to a standard agar well diffusion bioassay. For analysis, serum plus temocillin, the internal standard, were extracted with chloroform-n-amyl alcohol and back extracted into phosphate buffer. A reverse-phase C18 column and an ammonium acetate-methanol mobile phase were used with detection at 242 nm. Reproducibility studies yielded coefficients of variation ranging from 2.4 to 4.7% for low, mid, and high controls. Although cefoxitin, cephalothin, and cefuroxime exhibited retention similar to that of ticarcillin, a wide range of commonly administered antibiotics and other drugs did not interfere. The high-pressure liquid chromatographic assay is an accurate, reproducible method for determining the concentration of ticarcillin in serum during multiple antibiotic therapy or when rapid results are required.

Fatal infection after a bee sting.

Life-threatening or even fatal bee infections can rarely develop after bee stings.

Identification of clinical isolates of gram-negative nonfermentative bacteria by an automated cellular fatty acid identification system.

An automated cellular fatty acid (CFA) bacterial identification system, Microbial Identification System (MIS; Microbial ID, Newark, Del.), was compared with a conventional system for the identification of 573 strains of gram-negative nonfermentative bacteria. MIS identifications were based exclusively on the CFA composition following 22 to 26 h of growth at 28 degrees C on Trypticase soy agar. MIS identifications were listed with a confidence measurement (similarity index [SI]) on a scale of 0 to 1.0. A value of greater than or equal to 0.5 was considered a good match. The MIS correctly listed as the first choice 478 of 532 (90%) strains contained in the data base. However, only 314 (59%) had SI values of greater than or equal to 0.5. Of the 54 strains in which there was not agreement, 37 belonged to the genera Acinetobacter, Moraxella, or Alcaligenes or were Pseudomonas pickettii. Reproducibility studies suggest that SI variation is most likely a function of a difference in culture age at the time of analysis, which is due to the relatively low temperature and time of incubation. Other discrepancies were attributable to insufficiently characterized library entries or an inability to differentiate chemotaxonomically closely related species. The MIS, as the first automated CFA identification system, is an accurate, efficient, and relatively rapid method for the identification of gram-negative nonfermentative bacteria. The development of a CFA library with the media and incubation conditions routinely used for the isolation of clinical pathogens could further decrease the identification time and provide an increase in accuracy.

Determination of norfloxacin and ciprofloxacin concentrations in serum and urine by high-pressure liquid chromatography.

A high-pressure liquid chromatographic method for the determination of norfloxacin or ciprofloxacin concentrations in body fluids was developed and compared with a standard bioassay. The high-pressure liquid chromatographic assay utilizes a reverse-phase C18 column, an internal standard, and fluorescence detection, with reproducibility studies yielding coefficients of variation ranging from 0.6 to 3.7% and 0.9 to 2.7% for norfloxacin and ciprofloxacin, respectively. Correlation coefficients with the bioassay were 0.966 for norfloxacin and 0.952 for ciprofloxacin.

High resolution deuterium NMR studies of bacterial metabolism.

High resolution deuterium NMR spectra were obtained from suspensions of five bacterial strains: Escherichia coli, Clostridium perfringens, Klebsiella pneumoniae, Proteus mirabilis, and Staphylococcus aureus. Deuterium-labeled D-glucose at C-1, C-2, and C-6 was used to monitor dynamically anaerobic metabolism. The flux of glucose through the various bacterial metabolic pathways could be determined by following the disappearance of glucose and the appearance of the major end products in the 2H NMR spectrum. The presence of both labeled and unlabeled metabolites could be detected using 1H NMR spectroscopy since the proton resonances in the labeled species are shifted upfield due to an isotopic chemical shift effect. The 1H-1H scalar coupling observed in both the 2H and 1H NMR spectra was used to assign definitively the resonances of labeled species. An increase in the intensity of natural abundance deuterium signal of water can be used to monitor pathways in which a deuteron is lost from the labeled metabolite. The steps in which label loss can occur are outlined, and the influence these processes have on the ability of 2H NMR spectroscopy to monitor metabolism are assessed.

Systemic infection and colonization with and without prophylactic norfloxacin use over time in the granulocytopenic, acute leukemia patient.

Patients undergoing intensive antileukemic chemotherapy and profound granulocytopenia are susceptible to overwhelming infections, particularly those arising from disease- and therapy-related gastrointestinal tract damage. We have previously demonstrated that the ability to suppress bacterial colonization of this site with oral norfloxacin prophylaxis (400 mg every 12 h) affects the incidence and distribution of aerobic gram-negative bacterial infections and the overall management of infectious complications in this patient population. We have now determined the broad impact of continuous, long-term use of oral norfloxacin on aerobic gram-negative bacterial infection and colonization, overall management of presumed and documented infections during marrow aplasia and emergence of clinically significant antibiotic resistant pathogens during intensive antileukemic chemotherapy. Oral norfloxacin prophylaxis administered throughout the course of induced granulocytopenia continues to afford effective protection by suppressing the development of aerobic gram-negative infections, particularly those arising from the gastrointestinal tract, and preventing the acquisition or emergence of multiply resistant pathogens. A long-range effect of norfloxacin on pathogens colonizing the respiratory tract is also detected, with inhibition of acquired drug resistance occurring at that site as well. For these reasons, norfloxacin continues to be an excellent agent for oral prophylactic use in this patient population.