Factors associated with penicillin-nonsusceptible pneumococcal infections in Brazil

Levin, A.S.; Sessegolo, J.F.; Teixeira, L.M.; Barone, A.A.

doi:10.1590/S0100-879X2003000600017

Abstract

Resistance of Streptococcus pneumoniae is a worldwide, growing problem. Studies of factors associated with resistance to penicillin have not been conducted in Brazil. The objective of the present study was to evaluate factors associated with infection by S. pneumoniae not susceptible to penicillin. A prevalence study was conducted including all patients with a positive culture for S. pneumoniae in a hospital from July 1991 to December 1992 and the year 1994. Of 165 patients identified, 139 were considered to have clinically relevant infections and 88% of them had invasive infections. All infections were community acquired and consisted of pneumonia (44%) and of central nervous system (19%), pelvic or abdominal (12%), upper airway or ocular (12%), primary bloodstream (9%) and skin and soft tissue (5%) infections. Mortality was 25%. Susceptibility to penicillin was present in 77.6% of the isolates; 21.8% were relatively resistant, and one isolate was resistant (minimal inhibitory concentration = 4 µg/ml). Multivariate analysis showed that age below 4 years (odds ratio (OR): 3.53, 95% confidence interval (95%CI): 1.39-8.96) and renal failure (OR: 5.50, 95%CI: 1.07-28.36) were associated with lack of susceptibility to penicillin. Bacteremia occurred significantly less frequently in penicillin-nonsusceptible infections (OR: 0.34, 95%CI: 0.14-0.84), possibly suggesting that lack of penicillin susceptibility is associated with lower virulence in S. pneumoniae.

Streptococcus pneumoniae; Penicillin susceptibility; Risk factors; Prevalence study

Braz J Med Biol Res, June 2003, Volume 36(6) 807-813

Factors associated with penicillin-nonsusceptible pneumococcal infections in Brazil

A.S. Levin¹, J.F. Sessegolo², L.M. Teixeira²and A.A. Barone¹

¹Departamento de Moléstias Infecciosas, Hospital das Clínicas, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP, Brasil

²Laboratório de Microbiologia, Departamento de Microbiologia Clínica, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brasil

References

Correspondence and Footnotes ^{Correspondence and Footnotes} Correspondence and Footnotes

Abstract

Resistance of Streptococcus pneumoniae is a worldwide, growing problem. Studies of factors associated with resistance to penicillin have not been conducted in Brazil. The objective of the present study was to evaluate factors associated with infection by S. pneumoniae not susceptible to penicillin. A prevalence study was conducted including all patients with a positive culture for S. pneumoniae in a hospital from July 1991 to December 1992 and the year 1994. Of 165 patients identified, 139 were considered to have clinically relevant infections and 88% of them had invasive infections. All infections were community acquired and consisted of pneumonia (44%) and of central nervous system (19%), pelvic or abdominal (12%), upper airway or ocular (12%), primary bloodstream (9%) and skin and soft tissue (5%) infections. Mortality was 25%. Susceptibility to penicillin was present in 77.6% of the isolates; 21.8% were relatively resistant, and one isolate was resistant (minimal inhibitory concentration = 4 µg/ml). Multivariate analysis showed that age below 4 years (odds ratio (OR): 3.53, 95% confidence interval (95%CI): 1.39-8.96) and renal failure (OR: 5.50, 95%CI: 1.07-28.36) were associated with lack of susceptibility to penicillin. Bacteremia occurred significantly less frequently in penicillin-nonsusceptible infections (OR: 0.34, 95%CI: 0.14-0.84), possibly suggesting that lack of penicillin susceptibility is associated with lower virulence in S. pneumoniae.

Key words:Streptococcus pneumoniae, Penicillin susceptibility, Risk factors, Prevalence study

Introduction

Pneumococcal infections are important worldwide because of their frequency and severity. Streptococcus pneumoniae is an important cause of pneumonia, meningitis and otitis media and is a frequent cause of bacteremia. Originally, susceptibility to penicillin was universal (1) but since the first report in 1967 (2), resistance has been a growing problem worldwide (3-11). Resistance to other antimicrobial drugs also occurs and multiresistance is progressively more frequent (12-18). Epidemiological studies of the risk factors for infection or colonization by penicillin-resistant pneumococci have been done but results are few and sometimes conflicting.

The objective of the present study was to evaluate factors associated with infection by S. pneumoniae not susceptible to penicillin.

Material and Methods

This was a prevalence study involving all patients under medical attention at the Hospital das Clínicas, University of São Paulo, São Paulo, SP, Brazil, with at least one positive culture for S. pneumoniae from any clinical specimen during the period from July 1991 to December 1992 and the year of 1994. Hospital das Clínicas is a tertiary care teaching hospital associated with the University of São Paulo. It has approximately 2000 beds divided among 5 buildings. The microbiology laboratory is central and serves all buildings. The hospital is a reference center for the city of São Paulo and outskirts (approximately 15 million inhabitants).

The cases in which the isolate did not remain viable for confirmation of penicillin susceptibility were excluded. Infections were diagnosed using the criteria described by Garner et al. (19). Colonization or carrier status was defined when the isolate was obtained from a usually nonsterile site. Only cases of infection were included in the analysis of factors associated with nonsusceptibility. The following infections were considered invasive pneumococcal disease: pneumonia, meningitis and other central nervous system (CNS) infections, pelvic and abdominal infections and any bacteremic infection documented by a positive blood culture.

Identification was performed at the Microbiology Laboratory, Hospital das Clínicas, and was based on morphology, optochin susceptibility and bile-solubility tests. Susceptibility testing for penicillin, tetracycline, chloramphenicol, erythromycin, sulfamethoxazole-trimethoprim, rifampin, ampicillin, vancomycin, cephalothin and 3rd generation cephalosporins, and serotyping were done at the Microbiology Laboratory of the Clinical Microbiology Department of the Federal University of Rio de Janeiro, Brazil. These results have been published elsewhere (8,10,11). Susceptibility to penicillin was evaluated by obtaining the minimal inhibitory concentration (MIC) using the method of agar dilution. An isolate was considered susceptible if the MIC was below 0.1 µg/ml; an MIC from 0.1 to 1.0 µg/ml defined relative resistance, and an MIC above 1.0 µg/ml defined resistance. Relatively resistant and resistant isolates were considered penicillin nonsusceptible.

Patients were evaluated during hospitalization. When this was not possible data were obtained from their records. Autopsy data were available for 13 patients and were evaluated. The following data were collected: age, gender, diagnosis of infection, underlying conditions based on evaluation by the attending physician or on a clear indication in the patient's record (HIV infection, diabetes mellitus, chronic liver disease, chronic pneumopathy, cardiopathy, kidney failure, solid organ transplant, cancer, use of chemotherapy or steroids within the previous month), previous use of antimicrobial drugs, and present residence.

HIV infection was defined as a positive serologic test for HIV antibodies. Liver, kidney, heart and lung diseases were considered if there was a record of a previous medical follow-up for these conditions. The patients' present residences were divided into six zones: central (within a 5-km radius from the central point of the city), north, south, east, west, and outside the city perimeter. For the multivariate analysis the residence was considered central (within a 10-km radius from the central point) or peripheral. Information about current or previous use of antimicrobial agents was obtained from the patients or their relatives or from unmistakable evidence in the patients' chart. If not, antimicrobial use was considered unknown. Age was divided into three categories based on previously suggested high-risk age groups: younger than 4 years, from 4 to 64 years, and older than 64 years.

The data were organized using the EpiInfo software (version 6.02, Centers for Disease Control and Prevention, Atlanta, GA, USA). The group of patients infected with penicillin-nonsusceptible S. pneumoniae was compared with the penicillin-susceptible infection group. For dichotomous variables the results were expressed as odds ratios and 95% confidence intervals and the chi-square test or Fisher exact test was applied. When there were more than two categories only the chi-square test was used. For continuous variables, the mean data were compared by the Kruskal-Wallis test. Significance was set at 0.05.

Variables with P£0.20 in the univariate analysis were included in the multivariate analysis, which was performed by multiple logistic regression using the Stata program (version 7, StataCorp, College Station, TX, USA).

Results

The study involved 165 patients, 56% of whom were males. There were 107 adults (above 14 years of age) and 58 children. Mean (± SD) age was 43 ± 18 years for adults and 30 ± 32 months for children.

Most patients (76%) presented underlying conditions such as cancer (N = 20), liver disease (N = 18), HIV infection (N = 17), chronic lung disease (N = 13), steroid use within the previous month (N = 10), chronic heart disease (N = 8), renal failure (N = 8), and chemotherapy in the previous month (N = 8).

There were 128 (77.6%) isolates susceptible to penicillin, 21.8% relatively resistant, and one resistant isolate (MIC: 4 µg/ml). MIC50 for penicillin was 0.03 µg/ml and MIC90 was 0.25 µg/ml. Data concerning susceptibility to penicillin and other drugs and serotypes have been described elsewhere (8,10,11).

Twenty-six patients were considered to be noninfected carriers and 139 (84%) presented infection which was invasive in 122 (88%). The infections were: pneumonia (44%), CNS infections (19%), pelvic or abdominal infections (12%), upper airway or ocular infections (12%), primary bloodstream infections or endocarditis (9%), and skin or soft tissue infections (5%). Overall mortality was 25% for patients with pneumococcal infection, 33% in adults and 14% in children.

The results of univariate analysis can be seen in Table 1. Mean (± SD) age was 41.6 ± 17.4 years for patients with penicillin-susceptible infections and 44.5 ± 22.2 years for patients with nonsusceptible infection (P = 0.91). Among children, mean age was 18.1 ± 14.0 months for patients with penicillin-susceptible infections and 29.2 ± 28.9 months for patients with nonsusceptible infection (P = 0.15). Adequate information on previous antimicrobial use was only available for 37 cases of infection. The following variables were included in the multivariate analysis: age category, presence of bacteremia, cancer, renal failure, and present residence. Table 2 presents the final model of multiple logistic regression.

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Factors associated with penicillin-nonsusceptible pneumococcal infections in Brazil. A.S. Levin, J.F. Sessegolo, L.M. Teixeira and A.A. Barone. Brazilian Journal of Medical and Biological Research, 36 (6): 807, 2003.

Thumbnail

Factors associated with penicillin-nonsusceptible pneumococcal infections in Brazil. A.S. Levin, J.F. Sessegolo, L.M. Teixeira and A.A. Barone. Brazilian Journal of Medical and Biological Research, 36 (6): 807, 2003.

Discussion

There are relatively few studies involving factors associated with penicillin-nonsusceptible pneumococcal acquisition. Table 3 presents the most important studies in this area. Many involve a small number of patients or analyze only subsets of patients. In our study the factors associated with penicillin nonsusceptibility were: younger age, the absence of bacteremia and renal failure.

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Factors associated with penicillin-nonsusceptible pneumococcal infections in Brazil. A.S. Levin, J.F. Sessegolo, L.M. Teixeira and A.A. Barone. Brazilian Journal of Medical and Biological Research, 36 (6): 807, 2003.

In many studies young age is considered to be a risk factor for penicillin-resistant infection (20-24) or colonization (25) for reasons that are not clear. This may reflect the high prevalence of antimicrobial use among young children, especially those in day care or in other situations of extended contact with other children (26). On the other hand, age has been considered to be an independent risk factor in multivariate analyses (20,27). The previous use of antimicrobial drugs has been the factor most frequently associated with nonsusceptibility (18,20-22,25,27-32). In our study reliable data on previous antimicrobial use were not available for most patients and could not be properly evaluated. However, one of the problems met when analyzing the use of antimicrobials as a factor associated with resistance is using a design such as the one used in the present study. If patients with susceptible and nonsusceptible pneumococcus are compared this may lead to a selection bias because patients with previous antimicrobial use probably will not have susceptible isolates or may not have pneumococci at all. When these groups are compared the non-use of antimicrobials may be associated with having a susceptible isolate and not an association of resistance with antimicrobial use, as suggested in the literature. It has been suggested that to evaluate the role of antimicrobial use in resistance, cases with resistant isolates should be compared with controls from the same population not chosen for presenting susceptible strains (33). We feel that the design used in our study may not be suited to evaluating the impact of antimicrobial use on resistance.

The finding that bacteremia was significantly more frequent among penicillin-susceptible S. pneumoniae infections is interesting and not easily explained. There have been studies with similar results in which invasive or severe infections have been associated with susceptibility to penicillin (20,25,34), suggesting that nonsusceptibility leads to less virulent strains. On the other hand, there has been an association between nonsusceptibility to penicillin and immune suppression (21,22), HIV infection (20), severe underlying disease or conditions (21), and, in our study, renal failure. The significance of this is unknown but may also be related to the possible inability of resistant strains to cause severe infections in immunocompetent patients. The mechanisms for this remain obscure and deserve further investigation.

Nosocomial infection and colonization by penicillin-resistant S. pneumoniae have been reported (35). Previous hospitalization is considered to be a risk factor in some studies (18,24,34). In our study the infections were community acquired. Renal failure was significantly associated with resistance even though it was a relatively rare event occurring in only 8 patients. We did not find evidence that these patients had previous hospitalizations which might explain resistance. Renal failure is a known risk factor for pneumococcal disease and is considered to be an independent prognostic factor for mortality (36) but, to our knowledge, an association of renal failure with resistance has not been described before. Further studies will be needed to confirm this finding.

Address for correspondence: A.S. Levin, Rua Harmonia, 564/52, 05435-000 São Paulo, SP, Brasil. Fax: +55-11-3088-7095. E-mail: gcih@hcnet.usp.br

Publication supported by FAPESP. Received April 5, 2002. Accepted January 31, 2003.

1. Allen KD (1991). Penicillin-resistant pneumococci. Journal of Hospital Infection, 17: 3-13.
2. Hansman D & Bullen MM (1967). A resistant pneumococcus. Lancet, 2: 264-265.
3. Bermejo J, Pallares R, Podzamczer D & Gudiol F (1988). Infecciones por neumococos resistentes a penicilina: a propósito de tres observaciones. Medicina, 48: 285-289.
4. Chowdhury MNH (1983). A penicillin-resistant Streptococcus pneumoniae; case report. Tropical and Geographical Medicine, 35: 309-311.
5. Di Nubile MJ (1990). Meningitis due to pneumococcus. Reviews of Infectious Diseases, 12: 717-718.
6. Dronda F, Montilla P, Moreno S & Martinéz-Luengas F (1989). Endocarditis por Streptococcus pneumoniae resistente a penicilina. Revista Clinica Española, 185: 383-384.
7. Gellert G, Bock BV, Meyers H, Robertson C & Ehling LR (1991). Penicillin-resistant pneumococcal meningitis in an HIV-infected man. New England Journal of Medicine, 325: 1047-1048.
8. Levin AS, Teixeira LM, Sessegolo JF & Barone AA (1996). Resistance of Streptococcus pneumoniae to antimicrobials in São Paulo, Brazil: clinical features and serotypes. Revista do Instituto de Medicina Tropical de São Paulo, 38: 187-192.
9. Paton JH & Reeves DS (1987). First multiresistant pneumococcus in Britain. British Medical Journal, 295: 810-811.
10. Sessegolo JF, Levin AS, Levy CE, Asensi M, Facklam RR & Teixeira LM (1994). Distribution of serotypes and antimicrobial resistance of Streptococcus pneumoniae strains isolated in Brazil from 1988 to 1992. Journal of Clinical Microbiology, 32: 906-911.
11. Teixeira LM, Carvalho MG, Castineiras TM, Fracalanzza SA, Levin AS & Facklam RR (1997). Serotyping distribution and antimicrobial resistance of Streptococcus pneumoniae isolated in Brazil (1992-1996). Advances in Experimental Medicine and Biology, 418: 269-271.
12. Centers for Disease Control (1985). Isolation of multiple antibiotic-resistant pneumococci. Journal of the American Medical Association, 254: 1885-1886.
13. Chalkley LJ & Koornhof HJ (1988). Penicillin-binding proteins of Streptococcus pneumoniae Journal of Antimicrobial Chemotherapy, 22: 791-800.
14. Collingnon PJ, Bell J, Hufton IW & Mitchell D (1988). Meningitis caused by a penicillin- and chloramphenicol-resistant Streptococcus pneumoniae. Medical Journal of Australia, 149: 497-498.
15. Feldman C, Kallenbach JM, Miller SD, Thorburn JR & Koornhof HJ (1985). Community-acquired pneumonia due to penicillin-resistant pneumococci. New England Journal of Medicine, 313: 615-617.
16. Hebert JP & Caillet R (1980). Pneumocoques résistants aux antibiotiques - une observation. Nouvelle Presse Médicale, 9: 1098-1099.
17. Myint J & Panigrahi H (1991). Penicillin-resistant Streptococcus pneumoniae Journal of Hospital Infection, 17: 317-319.
18. Pallares R, Gudiol F, Liñares J, Ariza J, Rufi G, Murgui L, Dorca J & Viladrich PF (1987). Risk factors and response to antibiotic therapy in adults with bacteremic pneumonia caused by penicillin-resistant pneumococci. New England Journal of Medicine, 317: 18-22.
19. Garner JS, Jarvis WR, Emori TG, Horan TC & Hughes JM (1988). CDC definitions for nosocomial infections, 1988. American Journal of Infection Control, 16: 128-140.
20. Bedos JP, Chevret S, Chastang C, Geslin P & Regnier B (1996). Epidemiological features of and risk factors for infection by Streptococcus pneumoniae strains with diminished susceptibility to penicillin: findings of a French survey. Clinical Infectious Diseases, 22: 63-72.
21. Garcia-Leoni ME, Cercenado E, Rodeño P, Quirás JCLB, Martínez-Hernández D & Bouza E (1992). Susceptibility of Streptococcus pneumoniae to penicillin: a prospective microbiological and clinical study. Clinical Infectious Diseases, 14: 427-435.
22. Nava JM, Bella F, Garau J et al. (1994). Predictive factors for invasive disease due to penicillin-resistant Streptococcus pneumoniae: a population-based study. Clinical Infectious Diseases, 19: 884-890.
23. Reichler MR, Allphin AA, Breiman RF et al. (1992). The spread of multiply resistant Streptococcus pneumoniae at a day care center in Ohio. Journal of Infectious Diseases, 166: 1346-1353.
24. Reichler MR, Rakovsky J, Sobotová A, Sláciková M, Hlavácová B, Hill B, Krajciková L, Tarina P, Facklam RR & Breiman RF (1995). Multiple antimicrobial resistance of pneumococci in children with otitis media, bacteremia, and meningitis in Slovakia. Journal of Infectious Diseases, 171: 1491-1496.
25. Melander E, Molstad S, Persson K, Hansson HB, Soderstrom M & Ekdahl K (1998). Previous antibiotic consumption and other risk factors for carriage of penicillin-resistant Streptococcus pneumoniae in children. European Journal of Clinical Microbiology and Infectious Diseases, 17: 834-838.
26. Kronenberger CB, Hoffman RE, Lezotte DC & Marine WM (1996). Invasive penicillin-resistant pneumococcal infections: a prevalence and historical cohort study. Emerging Infectious Diseases, 2: 121-124.
27. Deeks SL, Palacio R, Ruvinsky R, Kertesz DA, Hortal M, Rossi A, Spika JS & DiFabio JL (1999). Risk factors and course of illness among children with invasive penicillin-resistant Streptococcus pneumoniae The Streptococcus pneumoniae Working Group. Pediatrics, 103: 409-413.
28. Dejthevapor C, Vibhagool A, Thakkinstian A, Sirinavi S & Varachit M (2000). Risk factors for penicillin-resistant Streptococcus pneumoniae acquisition in patients in Bangkok. Southeast Asian Journal of Tropical Medicine and Public Health, 31: 679-683.
29. Diekema DJ, Brueggemann AB & Doern GV (2000). Antimicrobial-drug use and changes in resistance in Streptococcus pneumoniae Emerging Infectious Diseases, 6: 552-556.
30. Einarsson S, Kristjansson M, Kristinsson KG, Kjartansson G & Jonsson S (1998). Pneumonia caused by penicillin-non-susceptible and penicillin-susceptible pneumococci in adults: a case-control study. Scandinavian Journal of Infectious Diseases, 30: 253-256.
31. Nasrin D, Collingnon PJ, Roberts L, Wilson EJ, Pilotto LS & Douglas RM (2002). Effect of ß lactam antibiotic use in children on pneumococcal resistance to penicillin: prospective cohort study. British Medical Journal, 324: 1-4.
32. Tan TQ, Mason Jr EO & Kaplan SL (1993). Penicillin-resistant systemic pneumococcal infections in children: a retrospective case-control study. Pediatrics, 92: 761-767.
33. Harris AD, Karchmer TB, Carmeli Y & Samore MH (2001). Methodological principals of case-control studies that analyzed risk factors for antibiotic resistance: a systematic review. Clinical Infectious Diseases, 32: 1055-1061.
34. Winston LG, Perlman JL, Rose DA & Gerberding JL (1999). Penicillin-nonsusceptible Streptococcus pneumoniae at San Francisco General Hospital. Clinical Infectious Diseases, 29: 580-585.
35. Jacobs MR, Koornhof HJ, Robins-Browne RM et al. (1978). Emergence of multiply resistant pneumococci. New England Journal of Medicine, 299: 735-740.
36. Kim BN, Bae LG, Kim MN, Park SJ, Woo JH, Ryu J & Kim YS (2002). Risk factors for penicillin resistance and mortality in Korean adults with Streptococcus pneumoniae bacteremia. European Journal of Clinical Microbiology and Infectious Diseases, 21: 35-42.
37. Saah AJ, Mallonee JP, Tarpay M, Thornsberry CT, Roberts MA & Rhoades ER (1980). Relative resistance to penicillin in the pneumococcus. A prevalence and case-control study. Journal of the American Medical Association, 243: 1824-1827.
38. Mannheimer SB, Riley LW & Roberts RB (1996). Association of penicillin-resistant pneumococci with residence in a pediatric chronic facility. Journal of Infectious Diseases, 174: 513-519.
39. Roberts RB, Tomasz A, Corso A, Hargrave J, Severina E & PRP Collaborative Study Group (2001). Penicillin-resistant Streptococcus pneumoniae in metropolitan New York hospitals: case control study and molecular typing of resistant strains. Microbial Drug Resistance, 7: 137-152.

Correspondence and Footnotes

Publication Dates

Publication in this collection
03 June 2003
Date of issue
June 2003

History

Received
05 Apr 2002
Accepted
31 Jan 2003

This work is licensed under a Creative Commons Attribution 4.0 International License.

[1] 1. Allen KD (1991). Penicillin-resistant pneumococci. Journal of Hospital Infection, 17: 3-13.

[2] 2. Hansman D & Bullen MM (1967). A resistant pneumococcus. Lancet, 2: 264-265.

[3] 3. Bermejo J, Pallares R, Podzamczer D & Gudiol F (1988). Infecciones por neumococos resistentes a penicilina: a propósito de tres observaciones. Medicina, 48: 285-289.

[4] 4. Chowdhury MNH (1983). A penicillin-resistant Streptococcus pneumoniae; case report. Tropical and Geographical Medicine, 35: 309-311.

[5] 5. Di Nubile MJ (1990). Meningitis due to pneumococcus. Reviews of Infectious Diseases, 12: 717-718.

[6] 6. Dronda F, Montilla P, Moreno S & Martinéz-Luengas F (1989). Endocarditis por Streptococcus pneumoniae resistente a penicilina. Revista Clinica Española, 185: 383-384.

[7] 7. Gellert G, Bock BV, Meyers H, Robertson C & Ehling LR (1991). Penicillin-resistant pneumococcal meningitis in an HIV-infected man. New England Journal of Medicine, 325: 1047-1048.

[8] 8. Levin AS, Teixeira LM, Sessegolo JF & Barone AA (1996). Resistance of Streptococcus pneumoniae to antimicrobials in São Paulo, Brazil: clinical features and serotypes. Revista do Instituto de Medicina Tropical de São Paulo, 38: 187-192.

[9] 9. Paton JH & Reeves DS (1987). First multiresistant pneumococcus in Britain. British Medical Journal, 295: 810-811.

[10] 10. Sessegolo JF, Levin AS, Levy CE, Asensi M, Facklam RR & Teixeira LM (1994). Distribution of serotypes and antimicrobial resistance of Streptococcus pneumoniae strains isolated in Brazil from 1988 to 1992. Journal of Clinical Microbiology, 32: 906-911.

[11] 11. Teixeira LM, Carvalho MG, Castineiras TM, Fracalanzza SA, Levin AS & Facklam RR (1997). Serotyping distribution and antimicrobial resistance of Streptococcus pneumoniae isolated in Brazil (1992-1996). Advances in Experimental Medicine and Biology, 418: 269-271.

[12] 12. Centers for Disease Control (1985). Isolation of multiple antibiotic-resistant pneumococci. Journal of the American Medical Association, 254: 1885-1886.

[13] 13. Chalkley LJ & Koornhof HJ (1988). Penicillin-binding proteins of Streptococcus pneumoniae Journal of Antimicrobial Chemotherapy, 22: 791-800.

[14] 14. Collingnon PJ, Bell J, Hufton IW & Mitchell D (1988). Meningitis caused by a penicillin- and chloramphenicol-resistant Streptococcus pneumoniae. Medical Journal of Australia, 149: 497-498.

[15] 15. Feldman C, Kallenbach JM, Miller SD, Thorburn JR & Koornhof HJ (1985). Community-acquired pneumonia due to penicillin-resistant pneumococci. New England Journal of Medicine, 313: 615-617.

[16] 16. Hebert JP & Caillet R (1980). Pneumocoques résistants aux antibiotiques - une observation. Nouvelle Presse Médicale, 9: 1098-1099.

[17] 17. Myint J & Panigrahi H (1991). Penicillin-resistant Streptococcus pneumoniae Journal of Hospital Infection, 17: 317-319.

[18] 18. Pallares R, Gudiol F, Liñares J, Ariza J, Rufi G, Murgui L, Dorca J & Viladrich PF (1987). Risk factors and response to antibiotic therapy in adults with bacteremic pneumonia caused by penicillin-resistant pneumococci. New England Journal of Medicine, 317: 18-22.

[19] 19. Garner JS, Jarvis WR, Emori TG, Horan TC & Hughes JM (1988). CDC definitions for nosocomial infections, 1988. American Journal of Infection Control, 16: 128-140.

[20] 20. Bedos JP, Chevret S, Chastang C, Geslin P & Regnier B (1996). Epidemiological features of and risk factors for infection by Streptococcus pneumoniae strains with diminished susceptibility to penicillin: findings of a French survey. Clinical Infectious Diseases, 22: 63-72.

[21] 21. Garcia-Leoni ME, Cercenado E, Rodeño P, Quirás JCLB, Martínez-Hernández D & Bouza E (1992). Susceptibility of Streptococcus pneumoniae to penicillin: a prospective microbiological and clinical study. Clinical Infectious Diseases, 14: 427-435.

[22] 22. Nava JM, Bella F, Garau J et al. (1994). Predictive factors for invasive disease due to penicillin-resistant Streptococcus pneumoniae: a population-based study. Clinical Infectious Diseases, 19: 884-890.

[23] 23. Reichler MR, Allphin AA, Breiman RF et al. (1992). The spread of multiply resistant Streptococcus pneumoniae at a day care center in Ohio. Journal of Infectious Diseases, 166: 1346-1353.

[24] 24. Reichler MR, Rakovsky J, Sobotová A, Sláciková M, Hlavácová B, Hill B, Krajciková L, Tarina P, Facklam RR & Breiman RF (1995). Multiple antimicrobial resistance of pneumococci in children with otitis media, bacteremia, and meningitis in Slovakia. Journal of Infectious Diseases, 171: 1491-1496.

[25] 25. Melander E, Molstad S, Persson K, Hansson HB, Soderstrom M & Ekdahl K (1998). Previous antibiotic consumption and other risk factors for carriage of penicillin-resistant Streptococcus pneumoniae in children. European Journal of Clinical Microbiology and Infectious Diseases, 17: 834-838.

[26] 26. Kronenberger CB, Hoffman RE, Lezotte DC & Marine WM (1996). Invasive penicillin-resistant pneumococcal infections: a prevalence and historical cohort study. Emerging Infectious Diseases, 2: 121-124.

[27] 27. Deeks SL, Palacio R, Ruvinsky R, Kertesz DA, Hortal M, Rossi A, Spika JS & DiFabio JL (1999). Risk factors and course of illness among children with invasive penicillin-resistant Streptococcus pneumoniae The Streptococcus pneumoniae Working Group. Pediatrics, 103: 409-413.

[28] 28. Dejthevapor C, Vibhagool A, Thakkinstian A, Sirinavi S & Varachit M (2000). Risk factors for penicillin-resistant Streptococcus pneumoniae acquisition in patients in Bangkok. Southeast Asian Journal of Tropical Medicine and Public Health, 31: 679-683.

[29] 29. Diekema DJ, Brueggemann AB & Doern GV (2000). Antimicrobial-drug use and changes in resistance in Streptococcus pneumoniae Emerging Infectious Diseases, 6: 552-556.

[30] 30. Einarsson S, Kristjansson M, Kristinsson KG, Kjartansson G & Jonsson S (1998). Pneumonia caused by penicillin-non-susceptible and penicillin-susceptible pneumococci in adults: a case-control study. Scandinavian Journal of Infectious Diseases, 30: 253-256.

[31] 31. Nasrin D, Collingnon PJ, Roberts L, Wilson EJ, Pilotto LS & Douglas RM (2002). Effect of ß lactam antibiotic use in children on pneumococcal resistance to penicillin: prospective cohort study. British Medical Journal, 324: 1-4.

[32] 32. Tan TQ, Mason Jr EO & Kaplan SL (1993). Penicillin-resistant systemic pneumococcal infections in children: a retrospective case-control study. Pediatrics, 92: 761-767.

[33] 33. Harris AD, Karchmer TB, Carmeli Y & Samore MH (2001). Methodological principals of case-control studies that analyzed risk factors for antibiotic resistance: a systematic review. Clinical Infectious Diseases, 32: 1055-1061.

[34] 34. Winston LG, Perlman JL, Rose DA & Gerberding JL (1999). Penicillin-nonsusceptible Streptococcus pneumoniae at San Francisco General Hospital. Clinical Infectious Diseases, 29: 580-585.

[35] 35. Jacobs MR, Koornhof HJ, Robins-Browne RM et al. (1978). Emergence of multiply resistant pneumococci. New England Journal of Medicine, 299: 735-740.

[36] 36. Kim BN, Bae LG, Kim MN, Park SJ, Woo JH, Ryu J & Kim YS (2002). Risk factors for penicillin resistance and mortality in Korean adults with Streptococcus pneumoniae bacteremia. European Journal of Clinical Microbiology and Infectious Diseases, 21: 35-42.

[37] 37. Saah AJ, Mallonee JP, Tarpay M, Thornsberry CT, Roberts MA & Rhoades ER (1980). Relative resistance to penicillin in the pneumococcus. A prevalence and case-control study. Journal of the American Medical Association, 243: 1824-1827.

[38] 38. Mannheimer SB, Riley LW & Roberts RB (1996). Association of penicillin-resistant pneumococci with residence in a pediatric chronic facility. Journal of Infectious Diseases, 174: 513-519.

[39] 39. Roberts RB, Tomasz A, Corso A, Hargrave J, Severina E & PRP Collaborative Study Group (2001). Penicillin-resistant Streptococcus pneumoniae in metropolitan New York hospitals: case control study and molecular typing of resistant strains. Microbial Drug Resistance, 7: 137-152.