In vitro activity of pristinamycin against Streptococcus pneumoniae

The activity of the pristinamycin was investigated using disk diffusion agar or ATB PNEUMO system and MIC determination using reference liquid medium method (NCCLS) on 749 S. pneumoniae strains isolated in Aquitaine in 1999. We have realized the killing curves against 10 isolates selected from erythromycin-susceptible and resistant S. pneumoniae strains. All the strains tested by ATB PNEUMO system were susceptible to pristinamycin, using disk diffusion agar, 6.8% of strains were intermediate or resistant. However the MIC's of pristinamycin determined by liquid dilution method against these strains were < 1 mg/L. These data suggest that the zone of inhibition around the disk was not correlated with MIC for erythromycin pneumococci and MIC testing must be performed. The results of killing curves showed a very good and rapid bactericidal activity of pristinamycin within two hours for concentration equal to 4 x MIC and four hours for 2 x MIC.     

In vitro activity of telithromycin (HMR 3647) against Greek Streptococcus pyogenes and Streptococcus pneumoniae clinical isolates with different macrolide susceptibilities

The susceptibilities to macrolides and telithromycin of 161 Streptococcus pyogenes and 145 Streptococcus pyogenes strains, consecutively isolated from five Greek hospitals, were determined by Etest. Moreover, mechanisms of resistance to macrolides were phenotypically and genetically determined by double disk induction test and PCR, respectively. Of the S. pneumoniae and S. pyogenes isolates, 42.8% and 30.8%, respectively, were found to be resistant to erythromycin. Of the erythromycin-resistant S. pneumoniae and S. pyogenes isolates, 57.5% and 59.5%, respectively, displayed the M phenotype and harbored the mefA/E gene. Telithromycin was found to be more active than both erythromycin and clarithromycin against both species, with considerably lower MIC₅₀ and MIC₉₀ values.     

Vancomycin-tolerant Streptococcus pneumoniae in Korea

A nationwide surveillance study was undertaken to monitor antimicrobial resistance among clinical isolates of Streptococcus pneumoniae in Korea, with a special focus on vancomycin tolerance. For the 6-month period from March to August 2002, clinical isolates of S. pneumoniae were collected from 11 university hospitals and 1 reference laboratory. One-hundred eighty-eight isolates were measured for lysis rates after exposure to vancomycin for 4 h. Two vancomycin-tolerant S. pneumoniae (VTSP) strains, S3 and H8, were isolated from sputum cultures of two patients, who had stayed in intensive-care units of different hospitals with long-term antibiotic therapy and were not treated for pneumococcal pneumonia. The penicillin, cefotaxime, and vancomycin MICs for S3 were 8 microg/ml, >16 microg/ml, and 0.5 microg/ml, and those for H8 were 2 microg/ml, 2 microg/ml, and 0.5 microg/ml, respectively. While S3 belonged to serotype 23F and was autolysin defective, H8 belonged to serotype 13F and had intact autolysin. These strains were not clonally related as determined by pulsed-field gel electrophoresis of chromosomal DNA. In agreement with previous reports, both isolates showed pairing of TIGR4 vex2 with R6 pep27 and had two identical amino acid substitutions, Q441K in vncS and N25D in vex2. These findings indicate that two VTSP strains have emerged independently in Korea, suggesting a prevalence rate of 1.1%. The emergence of VTSP would be a serious threat in Korea, where there are significant rates of penicillin resistance in S. pneumoniae. Monitoring of the prevalence of VTSP and further investigation of the clinical relevance of VTSP are warranted.     

In vitro antibacterial activity of beta-lactams and non-beta-lactams against Streptococcus pneumoniae isolates from Sydney, Australia

AIMS: This study was undertaken to determine the antimicrobial resistance patterns of strains of Streptococcus pneumoniae from Sydney, Australia, comparing penicillin-susceptible, -intermediate and -resistant isolates. METHODS: Non-duplicate cultures of S. pneumoniae were collected from 1 January to 31 December 2002 in the three penicillin-susceptibility categories. Minimum inhibitory concentrations (MICs) of 19 antibacterial agents were determined by agar dilution based on the National Committee for Clinical Laboratory Standards (NCCLS) methodology. Overall for 2002, 687 non-duplicate isolates were obtained, of which 190 (28%) were intermediate or resistant to penicillin. From this set, 183 isolates were selected for study: 88 (48%) in the penicillin-susceptible group (MIC or= 2.0 mg/L). RESULTS: Resistance to non-beta-lactams was more common in penicillin-intermediate or -resistant strains. Multidrug resistance (resistance to >or= 2 non-beta-lactams) was found in 3% of penicillin-susceptible, 52% of penicillin-intermediate and 87% of penicillin-resistant isolates. Erythromycin resistance was seen in 22% of the penicillin-susceptible strains but increased significantly to 60% and 89% in the penicillin-intermediate and resistant strains, respectively. Clindamycin, tetracycline and trimethoprim/sulfamethoxazole showed similar diminished activity in penicillin-intermediate and -resistant strains; 64, 84 and 91% of the penicillin-resistant isolates were resistant to clindamycin, tetracycline and to trimethoprim/sulfamethoxazole, respectively. Chloramphenicol resistance was comparatively low level except 19% of the penicillin-resistant strains were resistant. Ciprofloxacin MICs for 14 strains were raised (MICs 4-16 mg/L); three of these were penicillin-susceptible, one penicillin-intermediate and 10 penicillin-resistant. Only one isolate was resistant to moxifloxacin and to gatifloxacin. Resistance to rifampicin, vancomycin, oritavancin, or linezolid was not detected. Twenty-three isolates were intermediate and one resistant to quinupristin/dalfopristin - 22 of these were penicillin resistant. CONCLUSIONS: Streptococcus pneumoniae isolates from Sydney are commonly resistant to beta-lactams and available non-beta-lactam agents, especially if they are penicillin non-susceptible. Resistance to moxifloxacin and gatifloxacin is still rare, but some isolates were non-susceptible to quinupristin/dalfopristin. It is important to continue to survey resistance patterns to recognise emerging resistances which affect the selection of empirical antimicrobials to treat infections with S. pneumoniae.     

Comparative in vitro activity of penicillin G, levofloxacin, moxifloxacin, telithromycin, pristinamycin, quinupristin–dalfopristin and linezolid against ofloxacin-intermediate and -resistant Streptococcus pneumoniae

Screening by ofloxacin disk was carried out on 1158 strains of Streptococcus pneumoniae in order to investigate the in vitro bacteriostatic activity of penicillin G, levofloxacin, moxifloxacin, telithromycin, linezolid, pristinamycin and quinupristin–dalfopristin against ofloxacin-intermediate and -resistant S. pneumoniae strains. It was concluded that these new antimicrobial agents could be useful for the treatment of pneumococcal infections caused by penicillin-sensitive and -resistant S. pneumoniae , and would represent a valid therapeutic option for patients allergic to β -lactams, should they prove to be potent in vivo.     

A common clone of erythromycin-resistant Streptococcus pneumoniae in Greece and the UK

Objective  To investigate the possible genetic relationship among erythromycin-resistant Streptococcus pneumoniae strains isolated in Greece and the UK.
Methods  During 1995–97, 140 S. pneumoniae strains were isolated from clinical specimens submitted to the microbiology departments of the two main children's hospital in Athens. All erythromycin-resistant strains were further studied with respect to the presence of genes encoding for the two major mechanisms of macrolide resistance, their serotypes, and pulsed-field gel electrophoresis (PFGE) types, in comparison to a previously characterized UK erythromycin-resistant clone.
Results  Eleven of the 140 isolates (7.9%) were resistant to erythromycin; nine of these were susceptible to penicillin. Serotyping allocated seven, three and one isolates to serotypes 14, 19F and serogroup 6, respectively. The mef A gene was detected in seven isolates (five serotype 14 and two serotype 19F), erm B in two (one serotype 19F and the serogroup 6 isolate), whilst in the remaining two isolates no resistance gene could be detected by polymerase chain reaction (PCR). Pulsed-field gel electrophoresis of genomic DNA showed that five Greek serotype 14 isolates belonged to the same chromosomal type as the serotype 14 erythromycin-resistant UK clone.
Conclusions  The present study showed that erythromycin resistance among the S. pneumoniae isolates was mostly owing to the efflux mechanism and suggested a possible clonal spread of serotype 14 erythromycin-resistant S. pneumoniae strains between Greece and the UK.     

In Vitro Plaque-Forming Cell Responses Induced by Streptococcus pneumoniae in Humans

When peripheral blood lymphocytes (PBL) were stimulated in vitro with the rough form of type 2 Streptococcus pneumoniae R36a, the resulting plaque-forming cells (PFC) did not produce antibodies directed against phosphorylcholine, a major antigenic determinant of the cell wall C-polysaccharide. Instead, R36a stimulated polyclonal PFC in PBL and splenic lymphocytes. We compared the polyclonal responses stimulated by R36a with those induced by two well-characterized polyclonal activators (PA), Staphylococcus aureus Cowan I and pokeweed mitogen (PWM). We found that R36a was a poor mitogen for PBL, whereas the other two PA were potent mitogens; that the predominant isotype produced in response to all three PA was IgM; that adherent cells strongly inhibited the polyclonal PFC response to both R36a and Staph. aureus but not PWM; and that T cells were necessary for induction of polyclonal antibody-secreting cells by all three stimuli.     

Characterization of hemin binding activity of Streptococcus pneumoniae.

Streptococcus pneumoniae is a causative agent of bacterial pneumonia, otitis media, meningitis, and bacteremia. It causes considerable morbidity and mortality throughout the world, especially among children, the elderly, and immunocompromised individuals. We have demonstrated previously that the growth of S. pneumoniae is limited under iron-depleted conditions and can be restored by the addition of either hemin or hemoglobin. In the present study, we showed that S. pneumoniae had the ability to bind hemin and that the level of hemin binding activity was not affected by supplementation of the growth medium with iron. Approximately 70 to 80% of the hemin binding activity was mediated by proteinase-resistant components, and the remainder was mediated by proteins. Hemin binding proteins were located in both soluble extract and envelope fractions of pneumococcal cells. By batch affinity chromatography, a major hemin binding polypeptide with an apparent molecular mass of 43 kDa was identified in the cell lysate of S. pneumoniae. Polyclonal antibodies against this polypeptide were raised. By immunoblot analysis, this hemin binding polypeptide was localized in the envelope and did not exhibit any variation in molecular weight among all serotypes tested. The subcellular distribution of hemin binding activity may have functional implications.     

In vitro activity of ceftobiprole and seven other antimicrobial agents against invasive Streptococcus pneumoniae isolates in Spain

The in vitro activity of ceftobiprole was compared with that of seven antimicrobial agents against invasive Streptococcus pneumoniae isolated from adult patients (>15 years old). Characterization of erythromycin-resistant strains and serotype distribution of all pneumococci were also evaluated. Seventy invasive S. pneumoniae strains were isolated from December 2007 to January 2009. Serotyping was carried out by Quellung reaction. Antibiotic susceptibility was tested by broth microdilution (CLSI guidelines). The comparator agents were penicillin, cefotaxime, erythromycin, clindamycin, telithromycin, tetracycline and moxifloxacin. Phenotypic characterization of macrolide resistance was performed by the double disk method. Macrolide resistance genes [erm(B) and mef(A/E)] and the promoter of erm(B) were detected by PCR. Twenty-five different serotypes were detected of which 87% were non-PCV7 types. The percentages of resistance to erythromycin, clindamycin and tetracycline were 20%, 8.6% and 16%, respectively. A penicillin MIC ≥0.12 mg/L was observed in 14 of the 70 invasive pneumococci strains. The cefotaxime and ceftobiprole MIC₅₀/MIC₉₀ of these 14 strains were 1/4 and 0.03/1 mg/L, respectively. Ceftobiprole showed higher in vitro activity than penicillin and cefotaxime with all isolates being inhibited by ≤1 mg/L. Its high in vitro activity should make ceftobiprole a very promising drug for the treatment of pneumococcal infections.     

Anti-adhesive activity of human casein against Streptococcus pneumoniae and Haemophilus influenzae

The casein fraction of human milk was found to inhibit the attachment of Streptococcus pneumoniae and Haemophilus influenzae human respiratory tract epithelial cells. The inhibitory activity for S. pneumoniae remained after heat and trypsin treatment of the casein and was found in oligosaccharides released from casein. kappa-Casein, which is the most highly glycosylated casein component, inhibited pneumococcal attachment at concentrations similar to the whole casein fraction. The results are consistent with the known recognition of GlcNAc beta 1-3Gal by S. pneumoniae, since human milk and bovine colostrum, which contain GlcNAc, inhibited attachment, but mature bovine milk lacking GlcNAc did not. The effect on H. influenzae was similar to that on S. pneumoniae in that the attachment was inhibited by human casein and bovine colostrum, but not by either mature bovine milk or by the bovine casein fraction. The kappa-casein component of human milk was a less efficient inhibitor of H. influenzae attachment than the whole casein fraction and the free oligosaccharides were inactive. This anti-microbial effect of human casein represents a new mechanism for the protection by breast-milk against respiratory tract infection.     

Emergence of a Streptococcus pneumoniae clinical isolate highly resistant to telithromycin and fluoroquinolones

Streptococcus pneumoniae is a major pathogen causing community-acquired pneumonia and acute bronchitis. Macrolides, fluoroquinolones (FQs), and, recently, telithromycin (TEL) constitute primary therapeutic options, and rare cases of resistance have been reported. In this report, we describe the emergence of an S. pneumoniae clinical isolate with high-level TEL resistance (MIC, 256 microg/ml) and simultaneous resistance to FQs. Ongoing studies are oriented to elucidate the precise mechanism of resistance to TEL.     

in vitro activity of telithromycin against Haemophilus influenzae

The aim of this study was to evaluate the in vitro activity of telithromycin against 142 strains of Haemophilus influenzae using determination of MICs by agar dilution method and to evaluate the correlation between MICs and inhibition diameter zones obtained by disk diffusion testing. MIC50 and MIC90 of telithromycin were 1 and 2 mg/L respectively. Telithromycin activity against H. influenzae was similar to that of azithromycin, superior to erythromycin and clarithromycin and irrespective of the susceptibility to betalactams. Distribution of diameter zones showed a similar pattern to that of MICs but correlation between MICs and diameter zones was poor with correlation coefficients inferior to 0.5 whatever the agar media used.     

Activity of telithromycin and seven other agents against 1034 pediatric Streptococcus pneumoniae isolates from ten central and eastern European centers

Objective  To test the activity of telithromycin against 1034 Streptococcus pneumoniae isolates from pediatric patients in ten centers from ten central and eastern European countries during 2000–2001, and to compare it with the activities of erythromycin A, azithromycin, clarithromycin, clindamycin, and quinupristin–dalfopristin.
Methods  The minimum inhibitory concentrations (MICs) of telithromycin, erythromycin A, azithromycin, clarithromycin, clindamycin, levofloxacin, quinupristin–dalfopristin and penicillin G were tested by the agar dilution method with incubation in air, and mechanisms of resistance to macrolides and quinolones were investigated.
Results  Strains were isolated from sputum, tracheal aspirates, ear, eye, blood, and cerebrospinal fluid. Among S. pneumoniae strains tested, 36% had raised penicillin G MICs (≥ 0.12 mg/L). Susceptibilities were as follows: telithromycin, quinupristin–dalfopristin and levofloxacin, ≥ 99%; clindamycin, 83%; and erythromycin A, azithromycin and clarithromycin, 78%. Of 230 (22.3%) erythromycin A-resistant S. pneumoniae strains, 176 (79.6%) had erm(B) , 38 (16.1%) had mef(A) , and 10 (4.3%) had mutations in 23S ribosomal RNA or in ribosomal protein L4. The rates of drug-resistant S. pneumoniae are high in all centers except Kaunas, Riga, and Prague.
Conclusion  Telithromycin had low MICs against all strains, irrespective of macrolide, azalide or clindamycin resistance. Ribosomal methylation was the most prevalent resistance mechanism among all resistant strains, except in Sofia, where the prevalence of the efflux mechanism was higher.     

Emergence and spread of antimicrobial resistance of Streptococcus pneumoniae in Korea

Pneumococcal resistance has become a global issue during the past three decades. One of the major foci of pneumococcal resistance worldwide is the Asian region including Korea, Japan, and Hong Kong. Korea had not been recognized as a focus of pneumococcal resistance until 1995, when serial reports documented the alarmingly high prevalence of penicillin resistance among clinical isolates. Serial reports on penicillin resistance among pneumococcal isolates in Korea ranged from 68% to 77% as of 1995. Multidrug resistance was also noted in 34% of Korean isolates. Penicillin-binding protein profile analysis, pulsed-field gel electrophoresis, ribotyping, and fingerprinting analysis of pbp genes showed that antibiotic-resistant pneumococci isolated in Korea were genetically related. Data documented the extensive spread of a resistant clone within Korea and between different countries. Besides the injudicious use of antimicrobial agents or the high prevalence of serotypes 23 and 19, the spread of a resistant clone may play an important role in the rapid increase of penicillin resistance in Korea.     

Kinetic bactericidal activity of telithromycin, azithromycin and clarithromycin against respiratory pathogens

The present study assessed the comparative in vitro killing kinetics of telithromycin, azithromycin and clarithromycin. Minimal inhibitory concentrations (MIC) and minimal bactericidal concentrations (MBC) were determined against Streptococcus pneumoniae, beta-haemolytic streptococci, Haemophilus influenzae and Moraxella catarrhalis strains characterized by different susceptibilities to beta-lactams and macrolides. For each bacterial species, representative strains were chosen for time-kill studies. Telithromycin showed high activity against all the tested strains with MIC ranging from < or = 0.004 to 0.5 mg/L for streptococci, from 0.008 to 8 mg/L for H. influenzae, and from 0.008 to 0.5 mg/L for M. catarrhalis. In time-kill studies, telithromycin showed an overall superior bactericidal activity in respect to macrolides, particularly against resistant strains. In conclusion, telithromycin proved to possess bactericidal activity against a wide range of respiratory pathogens, including strains resistant to common macrolides.     

In Vitro Susceptibility to Gemifloxacin and Trovafloxacin of Streptococcus pneumoniae Strains Exhibiting Decreased Susceptibility to Ciprofloxacin

 The in vitro susceptibility to trovafloxacin and gemifloxacin of Streptococcus pneumoniae strains exhibiting decreased susceptibility to ciprofloxacin (MIC ≥2 μg/ml; 30 strains with intermediate resistance [MIC 2 μg/ml] and 43 strains with complete resistance [MIC ≥4 μg/ml]) was determined. Seventy-three strains collected in a surveillance study carried out from May 1996 to April 1997 in Spain (prior to commercialisation of trovafloxacin and gemifloxacin) from patients with respiratory tract infections were tested. The antibacterial activity of gemifloxacin was affected to a lesser extent than that of trovafloxacin by the increase in the MIC of ciprofloxacin, with gemifloxacin showing significantly (P≤0.001) better antibacterial activity than trovafloxacin in all ciprofloxacin MIC categories (MIC50/MIC90 values of 0.015/0.03, 0.015/0.06, 0.03/0.06 and 0.12/0.25 μg/ml for gemifloxacin vs 0.12/0.12, 0.12/1, 0.25/0.5 and 2/4 μg/ml for trovafloxacin in the 2, 4, 8 and ≥16 μg/ml ciprofloxacin MIC categories, respectively). Nine (12.3%) of these 73 strains exhibited decreased susceptibility to trovafloxacin (≥2 μg/ml), whereas all strains were inhibited by 0.25 μg/ml of gemifloxacin.     

Comparison of the in vitro activity of pristinamycin and quinupristin/dalfopristin against Streptococcus pneumoniae

The in vitro activity of quinupristin/dalfopristin, a new injectable streptogramin, and pristinamycin was evaluated against 200 recently isolated clinical Streptococcus pneumoniae strains expressing various degrees of susceptibility to penicillin G and erythromycin. MICs were determined by the agar dilution method. All strains were susceptible to pristinamycin irrespective of their susceptibility to penicillin G or erythromycin (MIC90: 0.25 mg/L for each phenotype). The activity of quinupristin/dalfopristin was slightly lower than that of pristinamycin against 42 penicillin G-susceptible/erythromycin-susceptible strains (MIC90: 0.5 mg/L), 13 penicillin G-susceptible/erythromycin-resistant strains (MIC90: 1 mg/L), 25 penicillin G-intermediate or -resistant/erythromycin-susceptible strains (MIC90: 0.5 mg/L) and 120 penicillin G-intermediate or -resistant/erythromycin-resistant strains (MIC90: 0.5 mg/L). The activity of both streptogramins was not significantly altered in case of erythromycin resistance. Thus, both streptogramins might be useful for the treatment of penumococcal infections, especially in cases of multiresistant strains.     

Multidrug-resistant Streptococcus pneumoniae serotype 6D clones in South Korea

To investigate the characteristics of main Streptococcus pneumoniae clones of serotype 6D (ST282 and ST3171) in South Korea, antimicrobial susceptibility testing was performed, and 11 genes around the cps locus were sequenced on ST282(6D), ST3171(6D), and ST81(6A) isolates. The antimicrobial susceptibility patterns were very similar between clones belonging to the same clonal complex, ST81(6A) and ST282(6D); nonsusceptibilities to penicillin and cefuroxime, high MICs of ceftriaxone, and high resistance rates to trimethoprim-sulfamethoxazole. However, ST3171(6D) isolates showed resistance to only macrolides and clindamycin. The sequences of 11 genes around the cps locus indicated the same genetic backgrounds between the ST81(6A) and ST282(6D) isolates. On the other hand, ST3171(6D) isolates showed nucleotide and amino acid differences from ST81(6A) and ST282(6D) isolates in most genes, indicating a different genetic background. The mosaic structure of dexB gene in ST282(6D) isolates indicated that recombination might occur in the dexB gene. Our results suggest that the multidrug-resistant ST282(6D) pneumococcal clone has emerged by serial genetic recombination, including capsular switch.