Fluoroquinolone resistance in Streptococcus pneumoniae isolates in Germany from 2004-2005 to 2014-2015.

Streptococcus pneumoniae is a major cause of bacterial pneumonia, sepsis and meningitis worldwide. Prevalence of levofloxacin-resistant S. pneumoniae isolates in Germany and associated mutations in the quinolone resistance determining regions (QRDRs), as well as serotype distribution and multi locus sequence types (MLST) are shown. 21,764 invasive S. pneumoniae isolates from Germany, isolated in the epidemiological seasons from 2004/05 to 2014/15 were analyzed at the German National Reference Centre for Streptococci (GNRCS) for their levofloxacin resistance by micro broth dilution method. All resistant (minimal inhibitory concentration (MIC) ≥8µg/ml) and intermediate (MIC >2µg/ml and <8µg/ml) isolates were selected for the present study. Additionally, 29 susceptible isolates were randomly selected. A total of ninety isolates were tested for their levofloxacin-MIC by Etest, their serotype and sequence type, as well as for point-mutations at the QRDRs in the genes parC, parE, gyrA and gyrB. Twenty-five isolates exhibited levofloxacin MICs <2µg/ml (Etest) and no mutations in the QRDRs. Four isolates with MICs=2µg/ml had one mutation in parC; isolates with MICs >2µg/ml all had one or more mutations in the QRDRs. Four of nine intermediate isolates had a mutation in either parC or gyrA, and four isolates had mutations in both parC and gyrB. One isolate had mutations in both parC and gyrA. All isolates with MICs ≥8µg/ml (52) had mutations in both topoisomerase IV and gyrase. Serotypes associated with levofloxacin resistance shifted from a majority of PCV13 serotypes before the introduction of the PCV13 vaccine towards non-PCV serotypes. Resistant isolates were almost exclusively found among adults (98.1%).

Epidemiology of Streptococcus pneumoniae Serotypes in Jordan Amongst Children Younger than the Age of 5: A National Cross-Sectional Study.

INTRODUCTION: Streptococcus pneumoniae infections are a major cause of mortality and morbidity worldwide. In Jordan, pneumococcal conjugate vaccines (PCVs) are not included in the national vaccination program. Due to the current availability of several PCVs, including PCV-10, PCV-13, and PCV-15, along with PCV-20, currently undergoing pediatric approvals globally, the decision to introduce PCVs and their selection should be based on valid local data on the common serotypes of Streptococcus pneumoniae. METHODS: This cross-sectional study aimed to identify the frequency of serotypes of Streptococcus pneumoniae in children aged below 5 years hospitalized with invasive pneumococcal diseases (IPDs), including pneumonia, septicemia, and meningitis, during the study's duration in representative areas of Jordan. Serotyping for culture-positive cases was based on the capsular reaction test, known as the Quellung reaction. qPCR was conducted on the blood samples of patients with lobar pneumonia identified via X-ray or on cerebrospinal fluid for those with a positive latex agglutination test for Streptococcus pneumoniae. RESULTS: This study was based on the analysis of the serotypes of 1015 Streptococcus pneumoniae cases among children younger than the age of 5: 1006 cases with pneumonia, 6 cases with meningitis, and 3 cases with septicemia. Only 23 culture-positive cases were identified in comparison to 992 lobar pneumonia cases, which were PCR-positive but culture-negative, with a PCR positivity rate of 92%. Serotypes 6B, 6A, 14, and 19F were the most common serotypes identified in this study, with prevalence rates of 16.45%, 13.60%, 12.12%, and 8.18%, respectively. PCV-10, PCV-13, PCV-15, and PCV-20 coverage rates were 45.32%, 61.87%, 64.14%, and 68.47%, respectively. DISCUSSION: To the best of our knowledge, this is the largest prospective study from the Middle East and one of the largest studies worldwide showing the serotypes of Streptococcus pneumoniae. It reveals the urgency for the introduction of a PCV vaccination in Jordan, utilizing recently developed vaccines with a broader serotype coverage.

Superantigen genes are more important than the emm type for the invasiveness of group A Streptococcus infection.

BACKGROUND: In this study, we examined the role of superantigen genes and emm genotypes of clinical Streptococcus pyogenes isolates collected in Germany between 1997 and 2003. METHODS: Multiplex polymerase chain reaction for all 11 currently known superantigen genes and sequencing for emm types were used. RESULTS: Using a 2-step explorative data analysis procedure, we found that after combined analysis of superantigen genes and emm types, only the superantigen genes spea1-spea3, spem, and spea4 have a predictive value for invasiveness, with odds ratios of 7.992, 3.209, and 2.323, respectively. The predictive value for invasiveness of emm1 was lost after combined analysis because of the association between emm type and the highly predictive superantigen genes. On the other hand, presence of the superantigen gene ssa and of emm77 are predictors of noninvasiveness, with odds ratios of 0.370 and 0.271, respectively. CONCLUSIONS: These results indicate that the presence of superantigen genes is more important for the invasiveness of group A Streptococcus infection than emm type and may be the connection between the high-risk HLA type of the host and the pathogen. Furthermore, we found a very clear correlation between the presence of the genes spea1-spea3 and the presence of the gene emm1, which indicates that the relationship between emm1 and invasiveness is based on the superantigen gene profile. Our data suggest that the superantigen gene profile is of high importance for the clinical outcome of group A Streptococcus infections.

Prevalence of Pneumococcal Carriage among Jordanian Infants in the First 6 Months of Age, 2008-2016.

BACKGROUND: Streptococcus pneumoniae is an opportunistic human-adapted pathogen driven by nasopharyngeal carriage. AIMS: To find the pneumococcal carriage rate, resistance, serotypes, and coverage of pneumococcal conjugate vaccines (PCVs) among infants in the first six months of age in the period from March 2008 to April 2016. METHODS: Nasopharyngeal swabs (NP) were taken from healthy infants from the northern part of Jordan. Swabs were processed for cultivation, identification, resistance testing and serotyping according to standard methods. RESULTS: During the surveillance period, 484 infants of this age group were tested, with a total carriage rate of 56.2%. 96.2% of infants one to two months of age got one PCV7 injection and were 58% carriers at the time of the first injection. At age three to four months, 84.9% had received two injections, with a carriage rate of 54.9% at the time of the second injection. At ages five to six months, 12.5% had received one to three injections, with a carriage rate of 43.8%. Predominant serotypes in all age groups were 19F (12.5%), 6A (11.4%), 11A (8.4%), 19A (7.0%), 6B (6.6%), 23F (5.9%), 15B (5.1%), 15A and 23A (4.0% each). Coverage of PCV7, PCV13 and the future PCV20 among all cases were 30.5%, 50.7% and 70.6%, respectively. The highest coverage rate of 78.6% was noticed in the age group at five to six months with the future PCV20. Antibiotic resistance was the highest in the first age group. CONCLUSIONS: Pneumococcal carriage starts from the first month of the infant's life. The highest coverage was noticed for PCV20, which implies the necessity for inoculation with future vaccines.

Urban and Rural Disparities in Pneumococcal Carriage and Resistance in Jordanian Children, 2015-2019.

BACKGROUND: A pneumococcal carriage surveillance study took place examining Jordanian children in urban and rural areas in the period 2015-2019. OBJECTIVES: To determine urban and rural differences in pneumococcal carriage rate, resistance, and serotypes among healthy Jordanian children from Amman (urban) and eastern Madaba (rural). METHODS: Nasopharyngeal swabs (NP) were taken from 682 children aged 1 to 163 months. Pneumococcal identification, serotyping, and resistance were performed according to standard method. RESULTS: The number of cases tested for Amman was 267 and there were 415 cases tested for eastern Madaba. Carriage rate for eastern Madaba was 39.5% and 31.1% for Amman. Predominant serotypes for eastern Madaba and Amman were 19F (21.3%; 15.7%), 23F (12.2%; 9.6%), 14 (6.7%; 2.4%), 19A (4.9%; 2.4%), and 6A (5.5%; 3.6%). Resistance rates for eastern Madaba and Amman were as follows: penicillin (95.8%; 81.9%), clarithromycin (68.9%; 59.0%), clindamycin (40.8%; 31.3%), and trimethoprim-sulfamethoxazole (73.2%; 61.4%). Coverage of PCV7, PCV13, and the future PCV20 for Amman was 42.2%, 48.2%, and 60.2%; for eastern Madaba, coverage was 50.0%, 62.2%, and 73.2%, respectively. In Amman 25.8% of children received 1-3 PCV7 injections compared to 1.9% of children in eastern Madaba. CONCLUSIONS: There were significant differences in carriage, resistance, and coverage between both regions. The potential inclusion of a PCV vaccination program for rural areas is essential.

Multicenter study of pneumococcal carriage in children 2 to 4 years of age in the winter seasons of 2017-2019 in Irbid and Madaba governorates of Jordan.

Streptococcus pneumoniae is one of the leading causes of death worldwide. It disseminates through colonizers and causes serious infections. Aims of this study are to determine pneumococcal carriage rate, resistance, serotype distribution, and coverage of pneumococcal conjugate vaccines from children attending day care centers from Irbid and Madaba in Jordan. Nasopharyngeal swabs were collected from day care centers (DCCs) of healthy Jordanian children 2-4 years of age from four regions of Madaba (n = 596), and from eastern Irbid (n = 423). Swabs were cultivated on Columbia blood agar base supplemented with 5% sheep blood and incubated for 18-24 hours at 37°C with 5% CO2. Alpha-hemolytic isolates were tested for optochin sensitivity and bile solubility for identification. Isolates were analyzed for antimicrobial susceptibility by the Vitek2 system and E-test (BioMérieux). Serotyping was performed using the Neufeld Quellung method. A total of 341 pneumococcal strains were isolated from 1019 nasopharyngeal (NP) samples of healthy children attending DCCs for two winter seasons from 2017-2019. Carriage rate in eastern Irbid for both seasons was 29.6% and for Madaba 37.9%. Resistance rates for Irbid and Madaba, respectively, were as follows: Penicillin (86.3%; 94.4%), erythromycin (57.0%; 78.2%), clindamycin (30.8%; 47.2%), trimethoprim-sulfamethoxazole (68.6%; 86.6%), and tetracycline (45.7%; 51.9%). Predominant serotypes for Irbid were 19F (20.8%), 23F (12.0%), 6A (10.4%), and 6B (9.6%); whereas for Madaba were 19F (24.5%), 14 (7.4%), 6A (6.9%) and 23F (6.5%). Serotype coverage of the thirteen valent pneumococcal conjugate vaccine (PCV13) was about 65% for both regions. Over 96% of isolates with PCV13 serotypes in this study were resistant to penicillin with the exception of serotypes 3 and 5. As a conclusion resistance and carriage rates among the age group 2 to 4 years reached an alarming rate especially among vaccine types, which can be controlled by pneumococcal conjugate vaccination strategies.

Mechanisms of macrolide resistance among Streptococcus pneumoniae isolates from Russia.

Among 76 macrolide-nonsusceptible Streptococcus pneumoniae isolates collected between 2003 and 2005 from Central Russia, the resistance mechanisms detected in the isolates included erm(B) alone (50%), mef alone [mef(E), mef(I), or a different mef subclass; 19.7%], or both erm(B) and mef(E) (30.3%). Isolates with dual resistance genes [erm(B) and mef(E)] belonged to clonal complex CC81 or CC271.

Time-kill kinetics of Streptococcus pneumoniae with reduced susceptibility to telithromycin.

BACKGROUND: Telithromycin is a new ketolide increasingly used in Europe and the United States. Only very few telithromycin-resistant isolates have been described to date. METHODS: The anti-pneumococcal activity of telithromycin was determined against four clinical isolates of Streptococcus pneumoniae with reduced susceptibility to telithromycin by time-kill methodology. RESULTS: All four telithromycin non-susceptible strains had the constitutive macrolide-lincosamide-streptogramin B phenotype and the ermB genotype. Pneumococcal strains had telithromycin minimum inhibitory concentrations (MICs) ranging between 2 and 8 microg/ml. Mulitlocus sequence typing and serotyping showed three isolates to harbour the identical serotype (serotype 14) and sequence type (sequence type 143) indicating a genetic relatedness of strains. Telithromycin was only bactericidal against the isolates with telithromycin resistance, with 4-8 times the MIC after 24 h. CONCLUSION: The killing by telithromycin of S. pneumoniae isolates having an ermB resistance determinant and a telithromycin MIC of > or =2 microg/ml is slow. Achievable concentrations in serum, alveolar macrophages and epithelial lining fluid are below the concentrations which are necessary for bactericidal killing of highly telithromycin-resistant strains.

Regional differences in the epidemiology of invasive pneumococcal disease in toddlers in Germany.

In a population-based study, regional differences in incidence, serotype distribution and resistance rates in invasive pneumococcal disease in 1-2-year-old children were related to different day care attendance rates. Day-care attendance appears to be a relevant risk factor in some German states and should be considered for inclusion in the recommendations for pneumococcal vaccination of children at risk.

Prediction of the potential benefit of different pneumococcal conjugate vaccines on invasive pneumococcal disease in German children.

BACKGROUND: In the US a pneumococcal conjugate vaccination program with a 7-valent conjugate vaccine was successfully implemented in 2000. How much invasive pneumococcal disease can potentially be prevented by the 7-valent (or 11-valent) vaccine in Europe? METHODS: Prospective, active surveillance of invasive pneumococcal disease in German children age <16 years was performed between 1997 and 2000. Age- and disease-specific coverage and incidence rates were assessed in children old enough to benefit from complete vaccination to estimate the annual number of cases potentially preventable. RESULTS: A total of 1,743 cases were reported; 667 isolates were serotyped. Coverage of 7-valent (11-valent) conjugate vaccines in children age 6 months and older was age- and diagnosis-dependent, ranging from 10.5% (15.8%) to 78.3% (82.6%) for meningitis and from 13.6% (68.2%) to 75.0% (89.3%) for nonmeningitis invasive pneumococcal disease cases. Of an estimated annual number of 176 children with pneumococcal meningitis age 6 months or older, 112 (122) cases had serotypes included in the 7-valent (11-valent) conjugate vaccine compared with 181 (254) of 324 nonmeningitis invasive pneumococcal disease cases, with 37 of the 73 cases covered by the 11-valent vaccine only in children older than 5 years. Regarding meningitis in this age group the potential benefit was equally poor for both the 7-valent (12 of 37 cases) and the 11-valent vaccine (15 of 37 cases). CONCLUSION: Coverage of the 7- and 11-valent conjugate vaccines depends markedly on age and disease. The additional potential benefit of the 11-valent compared with the 7-valent vaccine for pneumococcal meningitis was marginal.

Characterization of German penicillin non-susceptible serotype 23F pneumococci using multilocus sequence typing.

Three nationwide multicentre studies (n = 5071) showed an increase in antibiotic resistance in pneumococci in Germany. Serotype 23F was the predominant serotype (n = 45, 22.4 %), followed by 6B (n = 30, 14.9 %) and 9V (n = 19, 9.5 %). Multilocus sequence typing was used to characterize 45 serotype 23F strains with reduced penicillin susceptibility. The Spanish(23F)-1 clone [profile 4-4-2-4-4-1-1, sequence type (ST) 81] contributes significantly to the emergence of penicillin resistance in Germany (n = 21, 46.7 % of all penicillin non-susceptible serotype 23F isolates). Isolates of ST 277 (profile 7-13-8-6-6-12-8), which has been found previously in the Netherlands, are also observed, particularly in western Germany (n = 8, 17.8 %). A high proportion of strains (n = 11, 24.4 %) have sequence types that have not been reported to date from other countries (STs 353-362). The major penicillin-resistant clones are present in Germany, a country with relatively low levels of beta-lactam resistance.

Molecular characterisation of macrolide resistance mechanisms of Streptococcus pneumoniae and Streptococcus pyogenes isolated in Germany, 2002-2003.

In the present study, a real-time PCR protocol was developed for the detection of macrolide resistance determinants and was validated in a nationwide study in Germany covering a total of 236 Streptococcus pyogenes and 241 Streptococcus pneumoniae strains collected from children < or = 16 years of age with community-acquired infections. Macrolide resistance was observed in 19.9% of pneumococcal strains and 14% of S. pyogenes isolates. Of the erythromycin A-resistant S. pyogenes strains, 93.9% showed the efflux type mef(A); 62.5% of the S. pneumoniae strains were mef(A)- and 37.5% erm(B)-positive. The correlation of the results of real-time PCR assay genotyping in the present study compared with those of conventional PCR genotyping and resistance phenotyping was 100%. Macrolide resistance is of growing concern in Germany. This highly sensitive and specific PCR assay to detect macrolide resistance has the potential to provide sufficiently rapid results to improve antibiotic treatment of streptococcal infections.

Seven-year surveillance of emm types of pediatric Group A streptococcal pharyngitis isolates in Western Greece.

BACKGROUND: An experimental 26-valent M protein Group A streptococcal (GAS) vaccine has entered clinical studies. Pharyngeal GAS emm type surveillances in different areas and time-periods enhance the understanding of the epidemiology of GAS pharyngitis. Moreover, these surveillances, combined with the data on GAS invasive disease, can play a significant role in the formulation of multivalent type-specific vaccines. METHODS: During a 7-year period (1999-2005), 2408 GAS isolates were recovered from consecutive children with pharyngitis in Western Greece. The overall macrolide resistance rate was 22.8%. Along the study period we noted a tendency towards significantly decreased rates of resistance, with the lowest rates occurring in 2002 (15.3%), 2003 (15%) and 2004 (16.7%). A random sample of isolates from each year, 338 (61.7%) of the 548 macrolide-resistant and 205 (11%) of the macrolide-susceptible, underwent molecular analysis, including emm typing. RESULTS: The 543 typed isolates had 28 different emm types. A statistically significant association was found between macrolide resistance and emm4, emm22 and emm77, whereas emm1, emm3, emm6, emm12, emm87 and emm89 were associated with macrolide susceptibility. A significant yearly fluctuation was observed in emm4, emm28 and emm77. The most common macrolide-resistant GAS were emm77 isolates harboring erm(A), either alone or in combination with mef(A), emm4 carrying mef(A), emm28 possessing erm(B), emm75 carrying mef(A), emm12 harboring mef(A) and emm22 carrying erm(A). We estimated that 82.8% of the isolates belonged to emm types included in the novel 26-valent M protein vaccine. The vaccine coverage rate was determined mainly by the increased frequency of nonvaccine emm4 isolates. CONCLUSIONS: A limited number of emm types dominated among macrolide-susceptible and macrolide-resistant GAS isolates. We observed seasonal fluctuations, which were significant for emm4, emm28 and emm77. This type of data can serve as baseline information if the novel 26-valent M protein GAS vaccine is introduced into practice.

Susceptibility to tuberculosis is associated with TLR1 polymorphisms resulting in a lack of TLR1 cell surface expression.

Human TLR1 plays an important role in host defense against Mycobacterium tuberculosis. Our aim was to analyze the association of the loss of TLR1 surface expression and TLR1 SNPs with susceptibility to TB. TLR1neg and TLR1pos cells from healthy individuals were identified by flow cytometry and compared by sequencing. TLR1 expression was measured using quantitative real-time PCR and immunoblotting. TLR1 SNP analyses of healthy individuals and TB patients from EU-C and Ghana were performed, and association of the TLR1 genotypes with increased risk of developing TB was statistically evaluated. Lack of TLR1 surface expression accompanied by impaired function was strongly associated with TLR1 SNP G743A. Genotyping of EU-C controls and TB patients revealed an association of TLR1 743A/1805G alleles [OR 2.37 (95% CI 1.13, 4.93), P=0.0219; OR 2.74 (95% CI 1.26, 6.05), P=0.0059] as well as TLR1neg 743AA/1805GG versus TLR1pos genotypes 743AG/1805TG [OR 4.98 (95% CI 1.64, 15.15), P=0.0034; OR 5.70 (95% CI 1.69, 20.35), P=0.0015] and 743AG + GG/1805TG + TT [OR 3.54 (95% CI 1.29, 9.90), P=0.0086; OR 4.17 (95% CI 1.52, 11.67), P=0.0025] with increased susceptibility to TB. No association of G743A with TB was found in Ghana as a result of a low frequency of genotype 743AA. Our data gain new insights in the role of TLR1 in M. tuberculosis defense and provide the first evidence that TLR1 variants are associated with susceptibility to TB in a low-incidence country.

Molecular characterization of pneumococcal isolates from pets and laboratory animals.

BACKGROUND: Between 1986 and 2008 Streptococcus pneumoniae was isolated from 41 pets/zoo animals (guinea pigs (n = 17), cats (n = 12), horses (n = 4), dogs (n = 3), dolphins (n = 2), rat (n = 2), gorilla (n = 1)) treated in medical veterinary laboratories and zoos, and 44 laboratory animals (mastomys (multimammate mice; n = 32), mice (n = 6), rats (n = 4), guinea pigs (n = 2)) during routine health monitoring in an animal facility. S. pneumoniae was isolated from nose, lung and respiratory tract, eye, ear and other sites. METHODOLOGY/PRINCIPAL FINDINGS: Carriage of the same isolate of S. pneumoniae over a period of up to 22 weeks was shown for four mastomys. Forty-one animals showed disease symptoms. Pneumococcal isolates were characterized by optochin sensitivity, bile solubility, DNA hybridization, pneumolysin PCR, serotyping and multilocus sequence typing. Eighteen of the 32 mastomys isolates (56%) were optochin resistant, all other isolates were optochin susceptible. All mastomys isolates were serotype 14, all guinea pig isolates serotype 19F, all horse isolates serotype 3. Rats had serotypes 14 or 19A, mice 33A or 33F. Dolphins had serotype 23F, the gorilla serotype 14. Cats and dogs had many different serotypes. Four isolates were resistant to macrolides, three isolates also to clindamycin and tetracycline. Mastomys isolates were sequence type (ST) 15 (serotype 14), an ST/serotype combination commonly found in human isolates. Cats, dogs, pet rats, gorilla and dolphins showed various human ST/serotype combinations. Lab rats and lab mice showed single locus variants (SLV) of human STs, in human ST/serotype combinations. All guinea pig isolates showed the same completely new combination of known alleles. The horse isolates showed an unknown allele combination and three new alleles. CONCLUSIONS/SIGNIFICANCE: The isolates found in mastomys, mice, rats, cats, dogs, gorilla and dolphins are most likely identical to human pneumococcal isolates. Isolates from guinea pigs and horses appear to be specialized clones for these animals. Our data redraw attention to the fact that pneumococci are not strictly human pathogens. Pet animals that live in close contact to humans, especially children, can be infected by human isolates and also carriage of even resistant isolates is a realistic possibility.

Clonal spread of mef-positive macrolide-resistant Streptococcus pneumoniae isolates causing invasive disease in adults in Germany.

Isolates (3,845) obtained from German adults with invasive pneumococcal disease between 1992 and 2004 were investigated. Of these, 430 isolates (11.2%) were erythromycin A nonsusceptible. Macrolide resistance genotypes and multilocus sequence types were determined. Among the isolates, 35.6% were erm(B) positive and 63.5% were mef positive. Over the study period, the frequency of resistance rose significantly from 2.2 to 17.0% (P < 0.001). A serotype 14, sequence type 9 clone was the most widespread.

Genetic diversity of pneumococcal surface protein A of Streptococcus pneumoniae meningitis in German children.

Pneumococcal surface protein A (PspA) is a possible candidate for the development of a pneumococcal vaccine that has the potential to offer a broad range of protection. PspA genes of pneumococcal meningitis isolates (n=40) isolated as part of an ongoing population-based nation-wide surveillance program on invasive pneumococcal disease in children in Germany were analyzed to expand our knowledge on the distribution of PspA families of this important vaccine candidate in Germany. The serotype distribution of the strains was as follows: serotype 4 (n=3), 6B (5), 9V (2), 14 (8), 18C (6), 19F (5), 23F (6), and 7F (5). The pspA genes of these strains could be assigned to 2 families containing 20 pneumococcal strains each. Family I could be subdivided into 2 clades with 17 strains in clade 1 and 3 strains in clade 2, and family II could be subdivided into 3 subgroups (clades 3-5) containing 16, 3, and 1 strain, respectively. Pneumococcal serotypes were distributed evenly over all clades and families. Interestingly, the distribution of the PspA gene families in Germany was seen to differ slightly that found in other countries.

A new closed-tube multiplex real-time PCR to detect eleven superantigens of Streptococcus pyogenes identifies a strain without superantigen activity.

Superantigens (SAgs) are very potent microbial toxins that are involved in severe diseases such as necrotizing fasciitis and toxic shock syndrome. There are currently 11 different SAgs that have been identified from Streptococcus pyogenes. In the present study, two sets of multiplex PCRs were developed for detection of these 11 SAg genes. The first group comprises spea1-3+5, spec, speg, spej, spek, and spel. The second group consists of spea1-4, speh, spei, spem, ssa, and smez. The presence of Streptococcus pyogenes SAg genes can be immediately identified using a real-time method with SYBR-Green, thus providing an excellent tool in clinical diagnostics. After testing more than 300 clinical isolates, we identified one strain without any SAg gene. This finding contrasts with previous reports describing SAg genes located on every Streptococcus pyogenes genome. This SAg gene-negative strain also did not show any mitogenic activity. It is hypothesized that clinical isolates from patients may overrepresent bacterial strains with pathogenic factors, such as SAgs.

Resistance to erythromycin and telithromycin in Streptococcus pyogenes isolates obtained between 1999 and 2002 from Greek children with tonsillopharyngitis: phenotypic and genotypic analysis.

Since the late 1990s, the prevalence of erythromycin-resistant Streptococcus pyogenes has significantly increased in several European countries. Between January 1999 and December 2002, 1,577 isolates of S. pyogenes were recovered from children with tonsillopharyngitis living in various areas of Western Greece. Erythromycin resistance was observed in 379 (24%) of the 1,577 isolates. All erythromycin-resistant strains along with 153 randomly selected erythromycin-susceptible S. pyogenes isolates were tested for their antimicrobial susceptibility, resistance phenotypes, and genotypes. Representative isolates underwent emm gene sequence typing. Isolates with reduced susceptibility to telithromycin (MIC, > or = 2 microg/ml) were studied for multilocus sequence type, L22, L4, and 23S rRNA mutations. Of the total 379 erythromycin-resistant isolates, 193 (50.9%) harbored the mef(A) gene, 163 (43%) erm(A), 1 (0.3%) mef(A) plus erm(A), and 22 (5.8%) the erm(B) gene. Among the erythromycin-susceptible isolates, emm 1 (25%), emm 2 (12.5%), and emm 77 (12.5%) predominated. Furthermore, among the erythromycin-resistant isolates, emm 4 (30.6%), emm 28 (22.2%), and emm 77 (12.5%) prevailed. Resistance to telithromycin was observed in 22 (5.8%) of the erythromycin-resistant isolates. Sixteen (72.7%) of the 22 isolates appeared to be clonally related, since all of them belonged to emm type 28 and multilocus sequence type 52. One of the well-known mutations (T2166C) in 23S rRNA, as well as a new one (T2136C), was detected in erythromycin- and telithromycin-resistant isolates. High incidence of macrolide resistance and clonal spread of telithromycin resistance were the characteristics of the Greek S. pyogenes isolates obtained from 1999 to 2002.

Telithromycin-nonsusceptible clinical isolates of Streptococcus pneumoniae from Europe.

Telithromycin-nonsusceptible pneumococcal clinical isolates (n = 17) were analyzed for their antimicrobial susceptibility, macrolide resistance mechanisms, and genetic relatedness. All strains showed the erm(B) genotype and showed a wide range of combinations of macrolide resistance mechanisms. The predominant clone (n = 7) was serotype 14, sequence type 143.