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Antimicrobial Therapy

Article Reference Comparative efficacies of different antibiotic treatments to eradicate nontypeable Haemophilus influenzae infection.
Nonencapsulated and nontypeable Haemophilus influenzae (NTHi) is a major cause of human respiratory tract infections. Some strains of NTHi can cause invasive diseases such as septicemia and meningitis, even if H. influenzae is not generally considered to be an intracellular pathogen. There have been very few reports about the therapeutic efficacy of antibiotics against respiratory tract infection caused by NTHi in mice because it is difficult for H. influenzae to infect mice. Therefore, we evaluated the efficacy of antibiotics against NTHi in both a cell culture model and a mouse model of infection.
Article Reference Outer membrane protein analysis of ampicillin-resistant isolates of Haemophilus influenzae from Saudi Arabia.
Haemophilus influenzae isolates characterized in a previously published study from the Kingdom of Saudi Arabia were analysed by outer membrane protein (OMP) profiling. Isolates from patients with confirmed respiratory tract infections were investigated. Antibiotic susceptibility tests in vitro showed 25/129 (19.4%) had various degrees of reduced susceptibility to ampicillin although all were fully susceptible to ceftazidime and ciprofloxacin. OMP profiles of the beta-lactamase mediated ampicillin-resistant and beta-lactamase negative; ampicillin intermediate resistant strains (BLNAI) isolated were investigated. Dendrograms of scanned SDS-PAGE of these strains showed 15 different groupings from the 15 non-typable (NTHi) isolates tested demonstrating a high degree of heterogeneity whereas the 5 Hib isolates demonstrated significantly closer relatedness and were probably clonal. The present study demonstrates the groupings of H. influenzae strains by OMP profile analysis which did not correlate with the beta-lactamase production ability, BLNAI isolates, geographical origin or biotype.
Article Reference Purulent pericarditis caused by nontypeable Haemophilus influenzae in a pediatric patient.
We report the 1st case of purulent pericarditis due to nontypeable Haemophilus influenzae in an immunocompetent 2-year-old male, which was successfully treated with surgical drainage, debridement, and antimicrobial therapy. The shifting microbial epidemiology of this disease, associated with changes in community health care practices, and the increasing virulence of this organism are discussed.
Article Reference Subinhibitory concentrations of azithromycin decrease nontypeable Haemophilus influenzae biofilm formation and Diminish established biofilms.
Nontypeable Haemophilus influenzae (NTHi) commonly causes otitis media, chronic bronchitis in emphysema, and early airway infections in cystic fibrosis. Long-term, low-dose azithromycin has been shown to improve clinical outcomes in chronic lung diseases, although the mechanism of action remains unclear. The inhibition of bacterial biofilms by azithromycin has been postulated to be one mechanism mediating these effects. We hypothesized that subinhibitory concentrations of azithromycin would affect NTHi biofilm formation. Laboratory strains of NTHi expressing green fluorescent protein and azithromycin-resistant clinical isolates were grown in flow-cell and static-culture biofilm models. Using a range of concentrations of azithromycin and gentamicin, we measured the degree to which these antibiotics inhibited biofilm formation and persistence. Large biofilms formed over 2 to 4 days in a flow cell, displaying complex structures, including towers and channels. Subinhibitory concentrations of azithromycin significantly decreased biomass and maximal thickness in both forming and established NTHi biofilms. In contrast, subinhibitory concentrations of gentamicin had no effect on biofilm formation. Furthermore, established NTHi biofilms became resistant to gentamicin at concentrations far above the MIC. Biofilm formation of highly resistant clinical NTHi isolates (azithromycin MIC of > 64 microg/ml) was similarly decreased at subinhibitory azithromycin concentrations. Clinically obtainable azithromycin concentrations inhibited biofilms in all but the most highly resistant isolates. These data show that subinhibitory concentrations of azithromycin have antibiofilm properties, provide mechanistic insights, and supply an additional rationale for the use of azithromycin in chronic biofilm infections involving H. influenzae.
Article Reference Recurrent posttraumatic meningitis due to nontypable Haemophilus influenzae: case report and review of the literature.
We report a case of relapsing Haemophilus influenzae meningitis in a boy at the age of nearly 3 years and 4.2 years who had been successfully vaccinated against H. influenzae serotype b (Hib). The pathogen was a nonencapsulated (nontypable) H. influenzae strain of biotypes III and VI, respectively. A rhinobasal impalement injury with development of a posttraumatic encephalocele is considered to be the predisposing condition. Review of the literature reveals that in patients systemically infected by nonencapsulated H. influenzae strains predisposing factors such as cerebrospinal fluid-shunts, implants and traumas are often found. To obtain further information on potential new disease patterns H. influenzae isolates from cerebrospinal fluid should be examined for capsule production and, if relevant, further characterized by capsular typing.
Article Reference First characterization of heterogeneous resistance to imipenem in invasive nontypeable Haemophilus influenzae isolates.
This study describes the first two reported invasive nontypeable Haemophilus influenzae (NTHI) isolates (strains 183 and 184) with heterogeneous resistance to imipenem. For both isolates, Etest showed imipenem MICs of > or =32 microg/ml. When the two strains were examined by the quantitative method of population analysis, both strain populations were heterogeneously resistant to imipenem and contained subpopulations growing in the presence of up to 32 microg of imipenem/ml at frequencies of 1.7 x 10(-5) and 1.5 x 10(-7), respectively. By pulsed-field gel electrophoresis analysis, the two isolates appeared to be genetically closely related. The sequencing of the ftsI gene encoding penicillin-binding protein 3 (PBP 3) and comparison with the sequence of the imipenem-susceptible H. influenzae strain Rd identified a pattern of six amino acid substitutions shared between strains 183 and 184; an additional change was unique to strain 183. No relationship between mutations in the dacB gene encoding PBP 4 and imipenem resistance was found. The replacement of the ftsI gene in the imipenem-susceptible strain Rd (for which the MIC of imipenem is 0.38 to 1 microg/ml) with ftsI from strain 183 resulted in a transformant for which the MIC of imipenem ranged from 4 to 8 microg/ml as determined by Etest. The Rd/183 transformant population showed heterogeneous resistance to imipenem; it contained subpopulations growing in the presence of up to 32 mug of imipenem/ml at a frequency of 3.3 x10(-8). The presence of additional resistance mechanisms, such as the overexpression of the AcrAB efflux pump, was investigated and does not seem to be involved. These data indicate that the heterogeneous imipenem resistance phenotype of our NTHI clone depends largely on the PBP 3 amino acid substitutions. We speculated that bacterial regulatory networks may play a role in the control of the heterogeneous expression of the resistance phenotype.
Article Reference Nontypeable Haemophilus influenzae meningitis in children: phenotypic and genotypic characterization of isolates.
With the decline in the incidence of invasive Haemophilus influenzae type b disease as result of routine immunization of infants, the potential emergence of nontypeable H. influenzae (NTHi) strains as important pathogens has been suggested.
Article Reference Comparative activities of antibiotics against intracellular non-typeable Haemophilus influenzae.
Non-typeable Haemophilus influenzae (NTHi) is a major bacterial pathogen of community-acquired respiratory tract infection and is usually found extracellularly, although studies have revealed that NTHi may possess the ability to invade human epithelial cells where it is then protected against attack by the local immune system and partly against the effect of antibiotics. The aim of the present study was to assess the ability of ampicillin, azithromycin, telithromycin, ciprofloxacin and moxifloxacin, five antibiotics in common clinical use, to kill NTHi within bronchial epithelial cells.
Article Reference Oral beta-lactams in the treatment of acute otitis media.
Acute otitis media (AOM) is an important infectious disease among children throughout the world. The management of AOM is complicated by difficulties in diagnosis, spontaneous resolution, altered microbiology after pneumococcal conjugate vaccine use, and expanding antimicrobial resistance. The recent introduction of the heptavalent pneumococcal vaccine has resulted in a decrease in AOM due to penicillin-nonsusceptible strains of Streptococcus pneumoniae (PNSP) and an increase in infection due to beta-lactamase-positive nontypeable Haemophilus influenzae. The results of in vitro susceptibility testing in the context of broad surveillance programs provide useful information regarding the potency and spectrum of the oral beta-lactam antibacterial agents used in the treatment of AOM. The application of pharmacokinetic/pharmacodynamic breakpoints provides an effective means of projecting the potential clinical efficacy of the available agents. In view of what appears to be an increase prominence of beta-lactamase-positive Gram-negative AOM pathogens and a decrease in PNSP, it appears that empiric therapy for AOM should now include an agent with activity against beta-lactamase-positive H. influenzae and S. pneumoniae.
Article Reference Effects of macrolides on antigen presentation and cytokine production by dendritic cells and T lymphocytes.
Macrolides are effective therapeutic agents for chronic respiratory tract diseases, such as chronic sinusitis, sinobronchial syndrome and diffuse panbronchiolitis. Although only limited information is available about their mechanisms, suppression of various inflammatory cytokines (IL-8, etc.) and some transcription factors has been reported to be involved. Non-typeable Haemophilus influenzae (NTHI) is one of the most important pathogens of the respiratory tract. P6 is one of the outer membrane proteins of NTHI and the target antigen of protective antibodies. To analyze the influence of macrolides on human dendritic cells (DCs), we treated DCs with macrolides and used them as antigen-presenting cells (APCs). Clarithromycin, roxithromycin and prednisolone suppressed the in vitro proliferative response of CD4+ T cells to P6 and also the production of cytokines. As a control, we also cultured DCs alone and exposed them to the medicament, while conversely culturing T cells without adding any drugs to the cultures. The results showed similar tendencies for suppression of immune responses. These findings suggest that macrolides suppress the antigen-specific immune responses of DCs in vitro.
Article Reference The non-typeable Haemophilus influenzae Sap transporter provides a mechanism of antimicrobial peptide resistance and SapD-dependent potassium acquisition.
We have shown that non-typeable Haemophilus influenzae (NTHI) resists killing by antimicrobial peptides (APs). A mutant defective in expression of the sap (sensitivity to antimicrobial peptides) gene cluster product SapA is sensitive to killing by APs and is significantly attenuated in its ability to survive in a chinchilla model of otitis media compared with the parent strain. In NTHI, SapA is believed to function as the periplasmic solute binding protein of an ABC transporter. Here, we demonstrated that recombinant chinchilla beta defensin-1 specifically interacted with recombinant SapA and that AP exposure increased expression of the sap operon. We further demonstrated that the putative Sap transporter ATPase protein, SapD, was required for AP resistance as well as potassium uptake in NTHI strain 86-028NP. Loss of SapD additionally abrogated NTHI survival in vivo. Complementation of the sapD mutation restored the ability to grow in potassium-limited medium, resistance to AP-mediated killing and survival in vivo. Collectively, these data support a mechanism of Sap system-mediated resistance to APs that depends on Sap-dependent transport of APs and a Sap-dependent restoration of potassium homeostasis. Thus, NTHI required a functional Sap system to mediate bacterial survival and pathogenesis in vivo.
Article Reference Construction of a mutant and characterization of the role of the vaccine antigen P6 in outer membrane integrity of nontypeable Haemophilus influenzae.
Outer membrane protein P6 is the subject of investigation as a vaccine antigen to prevent infections caused by nontypeable Haemophilus influenzae, which causes otitis media in children and respiratory tract infections in adults with chronic lung disease. P6 induces protective immune responses in animal models and is the target of potentially protective immune responses in humans. P6 is a 16-kDa lipoprotein that shares homology with the peptidoglycan-associated lipoproteins of gram-negative bacteria and is highly conserved among strains of H. influenzae. To characterize the function of P6, an isogenic mutant was constructed by replacing the P6 gene with a chloramphenicol resistance cassette. The P6 mutant showed altered colony morphology and slower growth in vitro than that of the parent strain. By electron microscopy, the P6 mutant cells demonstrated increased size, variability in size, vesicle formation, and fragility compared to the parent cells. The P6 mutant showed hypersensitivity to selected antibiotics with different mechanisms of action, indicating increased accessibility of the agents to their targets. The P6 mutant was more sensitive to complement-mediated killing by normal human serum. Complementation of the mutation in trans completely or partially restored the phenotypes. We concluded that P6 plays a structural role in maintaining the integrity of the outer membrane by anchoring the outer membrane to the cell wall. The observation that the absence of expression of P6 is detrimental to the cell is a highly desirable feature for a vaccine antigen, supporting further investigation of P6 as a vaccine candidate for H. influenzae.
Article Reference Multiple combination antibiotic susceptibility testing of nontypeable Haemophilus influenzae biofilms.
Haemophilus influenzae is a cause of otitis media with effusion (OME). Animal models demonstrate growth of H. influenzae biofilms in OME, which may explain why OME does not respond well to conventional antibiotic therapy. Using a previously developed in vitro model, we performed H. influenzae susceptibility studies to see if H. influenzae biofilm cultures were more resistant to antibiotics than planktonic (broth) cultures, and to determine which antibiotics were most effective against H. influenzae biofilms. H. influenzae isolates were grown as biofilms on polystyrene pins. Biofilm and planktonic minimum inhibitory concentrations (MICs) were measured for 8 antibiotics, and multiple combination testing was performed with 66 groupings of 1, 2, or 3 antibiotics. We found that biofilm cultures were more resistant to antibiotics than planktonic ones. Antibiotic combinations containing rifampin and ciprofloxacin were most effective against biofilms. Biofilm testing reveals differences in effectiveness among antibiotics not apparent from conventional susceptibility testing, and suggests novel antibiotic regimens that could be studied for treatment of OME.
Article Reference Invasive nontypeable Haemophilus influenzae infection in an adult with laryngeal cancer.
We describe the first case of a man diagnosed with laryngeal cancer presenting with nontypeable Haemophilus influenzae bacteremia and dissemination to a gouty joint and review the pertinent literature.
Article Reference Treating acute otitis media post-PCV-7: judicious antibiotic therapy.
Acute otitis media (AOM) is treated with antibiotics in the United States, but the changing distribution of bacterial pathogens that cause the disorder can present physicians with several challenges. Most physicians treat AOM empirically, and their treatment choice should target Streptococcus pneumonia, nontypeable Haemophilus influenzae, and Moraxella catarrhalis, as those bacteria are most often isolated in AOM. First-line treatment for new onset AOM remains amoxicillin (80-90 mg/kg/d, divided twice daily). For persistent or recurrent AOM, guidelines recommend high-dose amoxicillin-clavulanate, cefdinir, cefprozil, cefpodoxime, cefuroxime, or ceftriaxone. Improved diagnosis and optimizing the choice of therapy by considering in vitro and in vivo efficacy of the different antibiotics will improve patient outcomes. Improved patient outcomes will result in fewer AOM episodes, decreased antibiotic resistance, and reduced direct and indirect health care costs.
Article Reference Peptides selected for binding to a virulent strain of Haemophilus influenzae by phage display are bactericidal.
Nontypeable Haemophilus influenzae (NTHi) is an obligate parasite of the oropharynx of humans, in whom it commonly causes mucosal infections such as otitis media, sinusitis, and bronchitis. We used a subtractive phage display approach to affinity select for peptides binding to the cell surface of a novel invasive NTHi strain R2866 (also called Int1). Over half of the selected phage peptides tested were bactericidal toward R2866 in a dose-dependent manner. Five of the clones encoded the same peptide sequence (KQRTSIRATEGCLPS; clone hi3/17), while the remaining four clones encoded unique peptides. All of the bactericidal phage peptides but one were cationic and had similar physical-chemical properties. Clone hi3/17 possessed a similar level of activity toward a panel of clinical NTHi isolates and H. influenzae type b strains but lacked bactericidal activity toward gram-positive (Enterococcus faecalis, Staphylococcus aureus) and gram-negative (Proteus mirabilis, Pseudomonas aeruginosa, and Salmonella enterica) bacteria. These data indicate that peptides binding to bacterial surface structures isolated by phage display may prove of value in developing new antibiotics.
Article Reference Bacterial otitis media: a vaccine preventable disease?
Otitis media (OM) is the most common childhood illness for which medical advice is sought. Whilst the disease rarely results in death, there is a significant level of morbidity and economic burden on the community. Although the causes of OM are multifactoral, bacterial and viral infections are the single most important cause. Bacteria responsible for infections of the middle ear are predominantly, nontypeable Haemophilus influenzae, Streptococcus pneumoniae and Moraxella catarrhalis. Antibiotics have been widely used to treat children who present to a medical clinic with OM. However, given the high prevalence of this disease and the increasing incidence of microbial resistance to antibiotics, there is a need to develop alternative therapeutic strategies such as vaccination. Pneumococcal polysaccharide vaccination has produced disappointing results for effectiveness in preventing OM and there is evidence of an increased incidence of disease due to non-vaccine serotypes. An efficacious vaccine for bacterial OM would require combining protective protein antigens from all three causative bacteria. A combined bacterial-viral vaccine formulation would produce the most profound and sustained impact on reducing the global incidence of OM.
Article Reference Antibiotic activity of telithromycin and comparators against bacterial pathogens isolated from 3,043 patients with acute exacerbation of chronic bronchitis.
Antimicrobial therapy is considered an important component in the medical management of most patients with acute exacerbation of chronic bronchitis (AECB). The three predominant bacterial species isolated are nontypeable Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae. Staphylococcus aureus is also frequently isolated while atypical bacteria are thought to cause up to 10% of exacerbations. Antibacterial resistance is increasing worldwide and little surveillance data exist concerning pathogens isolated from patients with AECB.
Article Reference Persistent colonization by Haemophilus influenzae in chronic obstructive pulmonary disease.
Nontypeable Haemophilus influenzae colonizes the respiratory tract of adults with chronic obstructive pulmonary disease (COPD) and causes intermittent exacerbations. Isolates of H. influenzae collected monthly in a prospective study were subjected to molecular typing. During a 7-year study spanning 345 patient-months of observation, 122 episodes of negative cultures lasting 1 month or more, and that were preceded and followed by isolation of an apparently identical strain of H. influenzae, were found. Seventeen such episodes of negative cultures, lasting 6 months or more and spanning 203 patient-months, were studied in detail to test the hypothesis that these periods of negative cultures represented continuous colonization by the same strain of H. influenzae. Molecular typing by three independent methods established that the strains preceding and following the episodes of negative cultures were indeed identical. Strain-specific H. influenzae DNA was detected in some of the sputum samples that had yielded negative cultures. These results indicate that some patients with COPD are persistently colonized with H. influenzae and that sputum cultures underestimate the frequency of colonization of the respiratory tract by H. influenzae in COPD. This observation has a significant impact on understanding bacterial colonization in COPD.
Article Reference Acute mastoiditis in Southern Israel: a twelve year retrospective study (1990 through 2001).
Acute mastoiditis is a serious complication of acute otitis media (AOM) and has been increasingly reported in the last decade.