Microbiological evaluation of antibiotics for empirical therapy of community-acquired infections of the lower respiratory tract]

During first 3 days after patient hospitalization with pneumonia or chronic obstruction pulmonary disease (COPD) pathogens in sputum were studied according NCCLS standards (for 1999 year). Among 93 pathogens isolated in pneumonia the most frequent were S. pneumoniae (41.9%), H. influenzae (21.5%). Among 232 pathogens isolated in COPD the most frequent were S. pneumoniae (35.5%), H. influenzae (16.8%). Other pathogens were staphylococci, moraxella, gram-negative bacteria. No penicillin-resistant S. pneumoniae, were isolated, the strains with moderate penicillin resistance were less than 3% in both groups. Among H. influenzae isolated from patients with pneumonia 25% were beta-lactamase producers, from COPD patients 21% strains produced beta-lactamase. Totally among all studied pathogens only 58% were sensitive to ampicillin in pneumonia groups and 48% in COPD groups, for azithromycin 70.7% and 71% respectively, for cefuroxime 84.5% and 85% respectively. Ampicillin efficacy for empirical treatment of community-acquired low respiratory tract infections was substantially less than that of modern antibiotics.     

Synthesis and antibacterial activity of HMR 3647 a new ketolide highly potent against erythromycin-resistant and susceptible pathogens

In the search for new ketolides with improved activities against erythromycin-resistant S. pneumoniae and H. influenzae we synthesized a new 11,12 carbamate ketolide substituted by an imidazo-pyridyl side chain: HMR 3647. This compound demonstrated a potent activity against erythromycin susceptible and resistant pathogens, including penicillin G/erythromycin A-resistant S. pneumoniae and H. influenzae. In vivo, HMR 3647 displayed good pharmacokinetic parameters (Cmax = 2.9 microg/ml, bioavailability=49%, AUC0.8 = 17.2 microg.h/l, t1/2=1h) and was shown to have a high therapeutic efficacy in mice infected by various respiratory pathogens, including multi-resistant S. pneumoniae and Gram negative bacteria such as H. influenzae. HMR 3647 appears to be a very promising agent for the treatment of respiratory infections and is currently in clinical trials.     

Nosopharyngeal microflora in ambulatory treated children and adults with upper respiratory tract infections

Upper respiratory tract consists resident and transient bacterial microflora, which in appropriate condition can cause infection. Bacteriological study was performed among 201 patients with upper respiratory tract infections treated in ambulatory. From nasal and pharyngeal swabs Staphylococcus aureus, Haemophilus influenzae, Streptococcus pneumoniae, Moraxella catarrhalis, and Streptococci group A, B, C, G were isolated. Antibiotic susceptibility testing of isolated strains was performed using CLSI criteria. All isolated strains of streptococci were susceptible to penicillin; some of them demonstrated resistance to macrolides and lincosamides. Few isolated strains of H. influenzae demonstrated resistance to penicillin and cotrimoxazole. Azitromycin resistant strains were not detected. All isolated strains of M. catarrhalis were beta-lactamase positive and demonstrated resistance to penicillin. Strains of methicillin sensitive S. aureus (MSSA) were isolated most frequently from pharyngeal swabs (35.4%) and S. pneumoniae (33.3)--from nasal swabs.     

Induction of stable L-forms of Staphylococcus aureus and Streptococcus faecalis

Stable L-forms were induced from Staphylococcus aureus and Streptococcus faecalis. These formed typical foamy L-colonies and showed large and small round bodies. They grew continuously on routine antibiotic-free nutrient broth and blood agar media for 12 passages without reversion to their parental forms. At different concentrations of penicillin various morphological forms were observed. Effect of sucrose, normal horse serum and penicillin on their adaptation and stabilization is discussed.     

The occurrence of denitrification in extremely halophilic bacteria

Abstract A total of 97 aerobic and facultatively anaerobic bacteria, and 3 Candida albicans , were tested for their ability to inhibit the growth of Haemophilus influenzae . Only strains of Pseudomonas aeruginosa showed any inhibitory effect and all 5 strains tested clearly inhibited the growth of all 10 strains of H. influenzae . The inhibition of H. influenzae . by Ps. aeruginosa may play a role in the establishment of chronic Ps. aeruginosa infections in the respiratory tracts of patients with bronchiectasis and cystic fibrosis (CF).     

Bacterial pericarditis and tamponade due to nonencapsulated Haemophilus influenzae complicating a case of adult community-acquired pneumonia

We report a case of bacterial pericarditis in an immunologically competent adult female caused by nonencapsulated Haemophilus influenzae (H influenzae) that was complicated by the acute development of life-threatening pericardial tamponade. H influenzae is a gram-negative coccobacillus, a pathogen most frequently associated with childhood exanthema (otitis media, meningitis) and, less frequently, adult pneumonia. Encapsulated, type b, or typable H influenzae is the strain implicated in childhood infections. On the other hand, nonencapsulated or nontypable H influenzae is the specific strain most often associated with exacerbation of chronic obstructive airway disease. Bacterial pericarditis caused by either subtype of H influenzae is exceedingly rare. We have located only 15 previously reported cases of H influenzae pericarditis occurring in adults in the world medical literature, the majority of which date back to the pre-antibiotic era. In 12 of these 15 cases (the only cases in which typing could be accomplished), the encapsulated strain of H influenzae was cultured from the pericardial fluid. Thus, to the best of our knowledge, we are reporting here the first case of bacterial pericarditis caused by nonencapsulated H influenzae in an immunologically competent adult.     

Antibiotic sensitivity of Haemophilus influenzae isolated from patients with inflammatory lung diseases

In 1980-1986 the sensitivity of 2,045 H. influenzae strains isolated from the bronchial contents of patients with inflammatory lung diseases were studied. This study revealed that 60-80% of H. influenzae cultures circulating in Leningrad were sensitive to tetracyclin, chloramphenicol and erythromycin. During the period of observation the tendency towards the decrease of the number of highly sensitive H. influenzae cultures and the increase of the number of strains resistant to all antibiotic preparations was followed. Most of H. influenzae strains isolated in Leningrad were sensitive to penicillin, oleandomycin and ampicillin. In 1983 the appearance of H. influenzae strains, multiresistant to antibiotics, was noted. In 1986 these strains constituted 4.5% of all isolated cultures.     

Analysis of gene expression in penicillin G induced persistence of Chlamydia pneumoniae

Chlamydia pneumoniae is responsible for respiratory tract infections and has been associated to chronic diseases such as atherosclerosis. The involvement of C. pneumoniae in chronic diseases may be correlated to its ability to induce persistent forms in which Chlamydiae remain viable but are not cultivable. The aim of our study is to investigate C. pneumoniae specific gene activities associated with the development of Chlamydial persistence in a cell culture system in the presence of penicillin G. Chlamydia-infected HEp 2 cells were incubated with or without penicillin G for up to 72 hours. The relative mRNA expression levels of early and late genes in treated and untreated cell cultures were determined by Real-time RT-PCR. Our results revealed a consistent down-regulation of Chlamydial hctA and hctB genes (p=0.012 and p=0.003 respectively) in association with up-regulation of htrA gene (p=0.002) during penicillin G-induced persistence suggesting these gene sets as leading candidate for in vivo investigation of the development of persistent Chlamydial infection. In conclusion, the Chlamydial expression pattern of hctA, hctB, and htrA genes may be helpful to identify target molecules to diagnose and treat Chlamydia-associated chronic diseases.     

Serotypes and biotypes and antibiotic susceptibility of Haemophilus influenzae encountered in a clinical laboratory in Taiwan.

Forty-four serologically and biochemically typable Haemophilus influenzae isolates from clinical specimens in Taiwan were subjected to analysis in their relationship with source of isolation and age distribution. It was found that all isolates from blood and cerebrospinal fluid were serotype b, biotype I, and all were in children less than 4 years of age. Serotypes b and e, biotypes I and III were encountered to have the highest incidence of infection caused by H. influenzae in this area. All H. influenzae isolates were further tested for susceptibility to several selected antibiotics. All strains of this organism were susceptible to erythromycin and chloramphenicol. All but two strains were susceptible to tetracycline, whereas more strains were resistant to carbenicillin, gentamycin, keflin, and penicillin. Thirty-four percent strains were found to be resistant to ampicillin and all were beta-lactamase producer. No direct correlation between ampicillin resistance and serotypes or biotypes was recognized.     

Etiology of acute and chronic bronchopulmonary diseases in children. II. A study of the specific immune response by counterimmunoelectrophoresis and the complement fixation and passive hemagglutination reactions

Specific immune response to Streptococcus pneumoniae and Haemophilus influenzae has been studied in 158 children with acute pneumonia and pleuritis and 128 children with chronic pneumonia by countercurrent immunoelectrophoresis (CIE) and in the complement fixation (CFT) and passive hemagglutination (PHA) tests. The use of CIE leads to the detection of antibodies to H. influenzae in 23.7% of children with acute pneumonia and in 46.9% of children with chronic pneumonia. In the CFT antibodies to H. influenzae are also more often detected in children with chronic pneumonia (48%) than in those with acute respiratory infections (12.2%). In the PHA test high titers of antibodies to type b H. influenzae capsular polysaccharide occur in 11.9% of children with acute pneumonia and in 8.2% of children with chronic pneumonia.     

Prevalence and chemosusceptibility of Neisseria meningitidis and Haemophilus influenzae in a population of central Italy

The purpose of this study was to determine the prevalence of nasopharyngeal carriers of Neisseria meningitidis and Haemophilus influenzae, and to evaluate their chemoresistance. N. meningitidis was more frequently isolated in adolescents (10-15 years). All of meningococci were susceptible to ceftriaxone and rifampin, only one isolate showed reduced susceptibility to chloramphenicol and four strains showed reduced penicillin susceptibility. The results show that these drugs are still effective for prophylaxis and treatment in our area. All strains of H. influenzae were susceptible to penicillin, ceftriaxone, chloramphenicol, rifampin, azithromicin and gentamicin. 6 nontypeable strains were resistant to sulfamethoxazole-trimethoprim, 7 strains of type a and c-f, and 3 non-typeable strains showed reduced susceptibility to tetracycline. In contrast with the current trend in the world, in our area the susceptibility of H. influenzae to betalactams was 100%, therefore these antibiotics are still the drugs of choice for treatment of invasive diseases.     

Haemophilus influenzae meningitis in Manitoba and the Keewatin District,NWT: potential for mass vaccination.

A community-based surveillance study of all central nervous system infections was carried out in Manitoba and the Keewatin District, NWT, between Apr. 1, 1981, and Mar. 31, 1984. There were 201 cases of bacterial meningitis in Manitoba over the study period, 81 (40%) caused by Haemophilus influenzae; all but one isolate tested were type b (Hib). There were nine cases of H. influenzae meningitis in the Keewatin District. The overall annual incidence rate of H. influenzae meningitis in Manitoba was 2.5/100,000; for children under 5 years the rate was 32.1/100,000. For the Keewatin District the corresponding rates were 69.6/100,000 and 530/100,000. A total of 85% and 100% of the cases of H. influenzae meningitis occurred by 24 months of age in Manitoba and the Keewatin District respectively. The age at onset was earlier in native Indian children (22 cases) and Inuit children (9 cases) than in non-native children (59 cases) (p less than 0.005); thus, vaccine prevention of Hib meningitis will likely be more difficult in native Indian and Métis children. Without evaluating the increased potential of H. influenzae vaccines to prevent nonmeningitic forms of disease, we concluded that mass childhood vaccination with polyribosylribitolphosphate (PRP) vaccine is not warranted in Manitoba or the Keewatin District. Immunogenicity studies suggest that administration of conjugated Hib vaccines such as PRP-D in infancy may prevent approximately one-third to two-thirds of cases of H. influenzae meningitis; these vaccines warrant consideration for use in mass childhood vaccination programs.     

Intracellular Production of Brucella L Forms I. Recovery of L Forms from Tissue Culture Cells Infected with Brucella abortus

Hatten, Betty A. (The University of Texas Southwestern Medical School, Dallas), and S. Edward Sulkin. Intracellular production of Brucella L forms. I. Recovery of L forms from tissue culture cells infected with Brucella abortus. J. Bacteriol. 91:285-296. 1966.-Infectivity of virulent Brucella abortus strain 3183 was less for hamster macrophages after a 2-hr adsorption period than for an attenuated strain (S19) and its tissue culture variant (30). Both strains S19 and 30 were very toxic for the cells, but 3183 was not toxic. Two types of L forms were recovered from a large percentage of hamster kidney cell cultures when disintegration of infected cells was accelerated by tissue culture medium of high pH. One type grew in finely granular microcolonies, was isolated from cells infected for short periods of time, and often reverted to the bacterial form. The other type occurred in small irregularly shaped forms which later developed into round bodies. Both stained specifically with fluorescein-conjugated B. abortus antiserum. Semisolid media containing 0.7% agar provided optimal subsurface L-form growth. L forms also grew well in Thioglycollate Medium but grew poorly in other liquid media. Surface L-form growth was supported by several agar media, but CO(2) was required for optimal growth. Monolayers infected with strain 3183 and examined immediately after adsorption contained occasional small, round bodies. Bizarre forms increased in number with time and, after 24 to 72 hr, large pink-staining inclusions were often present which persisted for several days. Also appearing at about the same time were smaller, dark-staining forms which were first seen in clusters but later dispersed and finally occurred in chainlike configurations. Direct fluorescent-antibody stains of infected cells established that the intracellular forms were related to the infecting strain of B. abortus.     

Occurrence of the Fimbria Gene hifA in clinical isolates of nonencapsulated Haemophilus influenzae

The adherence of Haemophilus influenzae to epithelial cells plays a crucial role in infections. However, little is known about the occurrence of fimbriae. In this study, we examined the distribution of the fimbria gene (hifA) by PCR among 167 H. influenzae strains isolated from patients with respiratory infections. Almost all (163; 98%) of the isolates were nonencapsulated strains. The carriage rate of hifA by the nonencapsulated strains was 18.4%. Electron microscopy showed that fimbriae were abundantly present on the cell surface of hifA-positive strains tested. Only four (2.4%) isolates were encapsulated, all of which were type b and did not possess hifA. The present work suggests that fimbriae may play a considerable role as adhesins in nonencapsulated H. influenzae strains.     

Effect of Penicillin on the Multiplication of Meningopneumonitis Organisms (Chlamydia psittaci)

Although formation of infectious particles of meningopneumonitis organism in L cells was completely inhibited by 1 or more units of penicillin per ml, multiplication of reticulate bodies was observed, by light microscopy, in the presence of 200 units of penicillin per ml in stained smears of infected cells. When reticulate bodies were purified from cultures containing penicillin after 18, 30, and 45 hr of incubation, continuously increasing yields were obtained. When penicillin was added to infected cultures 0 to 15 hr after infection, no increase in infectivity was observed at 40 hr, but when antibiotic was added between 20 and 35 hr, partial synthesis of infectious particles was observed at 40 hr. On the other hand, removal of penicillin from an infected culture before 15 hr after infection did not affect the final yields of infectivity when assayed at 40 hr, but elimination of penicillin after 20 hr resulted in a decrease in infectivity. In suspensions of (32)P-labeled purified reticulate bodies grown in cultures containing penicillin and harvested 18 and 40 hr after infection, the (32)P distributions obtained by acid fractionation were similar to those of reticulate bodies from penicillin-free cultures. Cell membranes of reticulate bodies were also prepared from 40-hr cultures with penicillin. The size and shape of purified membranes, as seen by electron microscopy, and their amino acid compositions were similar to membranes prepared from reticulate bodies grown without penicillin, except that very small structures were observed in membranes from cultures containing penicillin. These results indicated that penicillin does not inhibit reproduction of reticulate bodies and formation of their cell membranes, but does inhibit the formation of elementary body cell envelopes.     

Human lactoferrin receptor activity in non-encapsulated Haemophilus influenzae

Since the ability of bacteria to compete with lactoferrin for iron contributes to the pathogenesis of mucosal infections, the presence of lactoferrin receptor activity in non-encapsulated Haemophilus influenzae was investigated. The growth of 18 H. influenzae isolates from the sputum samples of chronic bronchitis patients and of six of seven H. influenzae throat isolates from healthy adults was stimulated by iron saturated human lactoferrin. Apo-lactoferrin did not stimulate the growth of H. influenzae. Human lactoferrin binding to iron limited bacteria was detected for 16 H. influenzae strains from chronic bronchitis patients and for five of seven isolates from healthy adults. We conclude that the majority of H. influenzae isolates tested bind human lactoferrin and that the iron from lactoferrin is used for growth.     

Permanent alterations of the L-forms of proteus and salmonella under various conditions

L-forms obtained from three strains of Proteus and from one strain of Salmonella have been kept for 15 to 20 years by weekly or monthly transfers on agar plates containing penicillin. The morphology and growth requirements of these strains have changed. They now grow abundantly on the surface of agar and in broth. The cultures consist of large bodies, small granules, and transitional forms. These organisms are more resistant to distortion and stain more deeply than organisms of the usual L-forms. In broth and to a lesser extent on agar, branching filaments develop, on the ends of which both the large, round organisms and small organisms are produced. The filaments are a transitional stage in the development of the cultures. Usual bacillary forms were not present in the culture and did not appear in successive transfers in the absence of penicillin. Bacilli reappeared on exposure of the L cultures to the influence of a spore-bearing bacillus. A similar transformation of L-forms has also been observed developing within a short time in recently isolated A and B type L cultures of one Proteus strain during the process of reversion to the bacterial form. The altered cultures are fixed in a stage of transition between the B type L-form and the regular bacteria.     

Isolation of spheroplastic forms of Haemophilus influenzae from sputum in conventionally treated chronic bronchial sepsis using selective medium supplemented with N-acetyl-D-glucosamine: possible reservoir for re-emergence of infection.

The isolation rate of Haemophilus influenzae from patients with persistent production of purulent sputum has been increased by the routine use of a selective medium. Nevertheless, some purulent sputum still fails to yield a pathogen. The selective medium was supplemented with N-acetyl-D-glucosamine to encourage primary isolation of colony forming, spheroplastic H influenzae, which reverted to normal forms on subculture. On the basis of in vitro experiments it is postulated that these spheroplastic forms of H influenzae may be induced by inadequate antimicrobial chemotherapy and may be responsible for re-emergence of symptoms in these patients during or shortly after stopping chemotherapy.     

In Silico Metabolic Model and Protein Expression of Haemophilus influenzae Strain Rd KW20 in Rich Medium

The intermediary metabolism of Haemophilus influenzae strain Rd KW20 was studied by a combination of protein expression analysis using a recently developed direct proteomics approach, mutational analysis, and mathematical modeling. Special emphasis was placed on carbon utilization, sugar fermentation, TCA cycle, and electron transport of H. influenzae cells grown microaerobically and anaerobically in a rich medium. The data indicate that several H. influenzae metabolic proteins similar to Escherichia coli proteins, known to be regulated by low concentrations of oxygen, were well expressed in both growth conditions in H. influenzae. An in silico model of the H. influenzae metabolic network was used to study the effects of selective deletion of certain enzymatic steps. This allowed us to define proteins predicted to be essential or non-essential for cell growth and to address numerous unresolved questions about intermediary metabolism of H. influenzae. Comparison of data from in vivo protein expression with the protein list associated with a genome-scale metabolic model showed significant coverage of the known metabolic proteome. This study demonstrates the significance of an integrated approach to the characterization of H. influenzae metabolism.     

Evolving concepts in biofilm infections

Several pathogens associated with chronic infections, including Pseudomonas aeruginosa in cystic fibrosis pneumonia, Haemophilus influenzae and Streptococcus pneumoniae in chronic otitis media, Staphylococcus aureus in chronic rhinosinusitis and enteropathogenic Escherichia coli in recurrent urinary tract infections, are linked to biofilm formation. Biofilms are usually defined as surface-associated microbial communities, surrounded by an extracellular polymeric substance (EPS) matrix. Biofilm formation has been demonstrated for numerous pathogens and is clearly an important microbial survival strategy. However, outside of dental plaques, fewer reports have investigated biofilm development in clinical samples. Typically biofilms are found in chronic diseases that resist host immune responses and antibiotic treatment and these characteristics are often cited for the ability of bacteria to persist in vivo . This review examines some recent attempts to examine the biofilm phenotype in vivo and discusses the challenges and implications for defining a biofilm phenotype.