Portuguese Version

Year:  2003  Vol. 69   Ed. 6 - (13º)

Artigo Original

Pages: 814 to 818

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Isolation of Streptococcus pyogenes in individuals with pharyngotonsillitis and antimicrobial susceptibility testing

Author(s): Rozana Scalabrin1,
Gisele D. Buss2,
Kelly Cristina S. Iamaguchi3,
Celso Luiz Cardoso4,
Lourdes B. Garcia5

Keywords: Streptococcus pyogenes, pharyngotonsillitis, streptococcal infection

Abstract:
Aim: We investigate the occurrence of Streptococcus pyogenes in individuals with pharyngotonsillitis that spontaneously sought attendance in drugstores or in health units. Study design: Longitudinal cohorte. Material and Method: Samples from oropharynx of 58 individuals were collected with swab and tongue depressor and inoculated on sheep blood agar plates. At the moment the samples were collected, none of the individuals was under treatment with antibiotics. The presumptive identification of S. pyogenes was made by the susceptibility test to bacitracin and confirmed by the serological grouping through the extraction of the group-specific carbohydrate "C" antigen. Results: From the 58 samples of oropharynx analyzed, 32 (55.2%) were from individuals assisted in drugstores and 26 (44.8%) were obtained from those that sought health units. A total of 15 (25.9%) samples presented positive culture for S. pyogenes, being most of the strains (9/15, 60%) coming from individuals assisted in drugstores. Streptococcus pyogenes was isolated in 33.3% (11/33) of the individuals from zero to 15 years of age and in 16% (4/25) of those over 15. The strains were susceptible to all the antimicrobial agents tested. Conclusion: The results of the present study emphasize the importance of the bacteriological diagnosis in the proper treatment for the streptococcal pharyngotonsillitis that determine the prevention of the supurative or non supurative complications and the eradication of the microorganism of the oropharynx.

INTRODUCTION

Streptococcus pyogenes or group A beta-hemolytic streptococcus of Lancefield is the bacterial species more frequently associated with the etiology of primary pharyngeal and tonsil infections 1. These infections may occur at any age range, but they are common in children and adolescents aged 5 to 15 years 2. The period of usual incubation of streptococcus is 12 to 24 hours. The patient can present high fever and pain upon swallowing, with general malaise, anorexia and asthenia, symptoms that are commonly followed by nausea, vomiting and abdominal pain in children 3.

Acute streptococcal pharyngitis is more frequent in the coldest months of the year and it is contagious, since the bacteria are easily transmitted by direct contact by means of secretions of the respiratory tract, especially in environments in which there is agglomeration of people, such as schools and military facilities 4. Disseminating the primary focus of the infection, especially of pharynx and tonsil, S. pyogenes can infect different organs and tissues and cause suppurative complications. Moreover, streptococcal infections of the oropharynx can be followed by a severe sequel - rheumatic fever 1, 5, 6.

Many times it is not possible to clinically differentiate pharyngotonsillitis caused by group A streptococcus from an infection caused by other infectious agents, such as virus, since there are few specific clinical signs and they frequently fail 2. Oropharynx viral infections are normally benign and do not require treatment with specific antimicrobial agents. However, streptococcal pharyngitis should be treated with antibiotics to prevent acute rheumatic fever and suppurative complications, in addition to reducing the risk of contagious disease and minimizing symptoms 2, 5, 7.

International recommendations suggest that most cases of streptococcal pharyngitis should rely not only on clinical follow-up, but also make use of laboratory tests to confirm the presence of the bacteria in the oropharynx 2, 8. Different antibiotics can be effective in eradicating group A streptococcus in the oropharynx, including penicillin and its derivates, cephalosporins, macrolides and clindamycin. However, penicillin remains as the drug of choice when compared to other antimicrobial agents 2, 4, 5, 8.

In developed countries, culture and quick testing for detection of streptococcal antigen in material collected from the oropharynx are advocated as routine, which is not seen in our country, in which most of the cases of pharyngotonsillitis are not submitted to laboratory follow-up. Normally, we check clinical signs and symptoms of the patient, which in many occasions leads to unnecessary prescriptions of antimicrobial agents. In view of that, our study aimed at checking the presence of S. pyogenes in cultures of the oropharynx of subjects from different age ranges with symptoms of pharyngotonsillitis that spontaneously sought for support in drugstores or in healthcare centers. Susceptibility assessment to antimicrobial agents was conducted for all isolated strains.

MATERIAL AND METHODS

Collection of samples from the oropharynx.
In the period between March 1999 and January 2000, three commercial drugstores and three healthcare centers in the city of Maringa, state of Parana, were selected to conduct the study. The oropharynx samples were collected using swabs and tongue depressors; we had 58 patients with symptoms characteristic of pharyngotonsillitis that looked for medical support in the drugstores and the healthcare centers.

Isolation and identification of Streptococcus pyogenes
After the collection, each specimen was inoculated using a technique with Petri plates containing Tryptose blood-agar base (Difco Laboratories, Detroit, MI, USA), added by 5% defibrinated sheep blood. The plates were incubated at 35-37o C at microaerophilia atmosphere for 24-48 hours. The beta-hemolytic colonies present in the agar blood were transferred to the broth Brain heart infusion (Difco) and after 18-24 hours of incubation they were stained using gram technique. The beta-hemolytic colonies formed by gram-positive cocci were initially tested concerning the production of catalase enzyme, to characterize the microorganism as belonging to Streptococcus9. The differentiation of S. pyogenes from other beta-hemolytic streptococcus was made by sensitivity tests to bacitracin and serological grouping 10.

Antibiogram
The assessment of the sensitivity of S. pyogenes strains to antimicrobials penicillin G, ceftaxime, cefpime, ofloxacin, levofloxacin, clindamycin, erythromycin, chloramphenicol and vancomycin was conducted in vitro by the method of infusion in agar, using as a control the standard strain Streptococcus pneumoniae by American Type Culture Collection (ATCC 49619)11.

RESULTS

The study of S. pyogenes was made in samples of oropharynx collected from 58 subjects. Out of the total, 15 (25.9%) presented positive culture to this microorganism (Table 1). As to site of care, 32 (55.2%) of the subjects included in the study were seen in drugstores and 26 (44.8%) were seen in the healthcare center (Table 1).

The positive result of isolation of S. pyogenes according to age range of subjects is shown in Table 2. The bacterium was isolated in the oropharynx of in 11 (33.3%) of the 33 patients aged zero to 15 years. Among the 25 patients aged over 15 years, 4 (16%) had positive culture for S. pyogenes.

The report of sore throat and hyperemia was observed in 98% of the patients and high fever was detected in 47 patients (81%).

The 15 strains of S. pyogenes isolated in our study were sensitive to all tested antimicrobial agents.

Table 1. Isolation of S. pyogenes in patients with pharyngotonsillitis seen in drugstores and healthcare centers in the city of Maringá, PR, between March 1999 and January 2000.



Table 2. Positive result of isolation of S. pyogenes in 58 patients with pharyngotonsillitis, distributed according to age range.

DISCUSSION

Acute pharyngitis is among the infectious diseases that cause the highest number of visits to physicians 8. A large number of infectious agents, more frequently virus, cause acute pharyngitis. Among bacteria, group A streptococcus is undoubtedly the most common cause of pharyngitis, being responsible for 15 to 30% of the cases in children and 5 to 15% of the cases in adults 7, 8.

Streptococcal pharyngitis presents different signs and symptoms, however, none of the parameters is specific and there may be cases of pharyngitis caused by other infectious agents of the upper respiratory tract. Thus, the definite diagnosis of acute streptococcal pharyngitis in children and adolescents should be evidenced through clinical and epidemiological aspects and supported by laboratory tests 8.

As to laboratory diagnosis of streptococcal pharyngitis, the culture is still recognized as the standard technique to investigate the presence of S. pyogenes in the oropharynx. A simple swab correctly collected from the surface of the tonsils and the posterior pharyngeal wall and cultured in agar-blood presents sensitivity of 90 to 95% 8. False negative results occur in patients that present smaller number of microorganisms in the oropharynx and false positive results can occur in carriers of S. pyogenes with acute pharyngitis of non-streptococcal origin 4, 8.

Traditional culture of streptococcus in blood agar, including the definite identification of S. pyogenes, can take 24 to 48 hours 5. For this reason, the use of quick tests to check the presence of streptococcus antigen became common. These tests detect in few seconds the presence of bacterial antigens in oropharynx secretion. The tests are available in kits of different commercial brands and can be easily used in the office by the clinician 1.

Many quick tests present excellent specificity (³95%) when compared to agar blood cultures and thus, antimicrobial therapy can be indicated based on positive results to good quality quick tests 12, 13. However, sensitivity to most quick tests is inferior to that of culture (80 to 90%), and for this reason negative quick tests in children and adolescents should be confirmed by conventional oropharynx culture 8.

The need to have bacteriological diagnosis of streptococcal pharyngitis is based on the fact that this infection should be treated with antimicrobial agents. The objective of antibiotic therapy in streptococcal pharyngitis in children and adolescents is to prevent the development of non-suppurative sequelae, such as rheumatic fever, to prevent suppurative complications (peritonsillar or retropharyngeal abscess, cervical lymphadenoiditis, mastoiditis, sinusitis, otitis media) and reduce infection levels so that patients can go back to work or school within a short period of time 4.

Owing to low incidence of pharyngitis and minimum risk of development of rheumatic fever in subjects aged 20 or over, Cooper et al.7 (2001) suggested that diagnosis in normal adults be conducted only through high sensitivity quick tests associated with clinical diagnosis or the use of clinical criteria only. To these authors, oropharynx culture is not recommended as a routine, since the results can not be assessed right in the first visit of the patient and late decision about use of antimicrobial agents eliminates the main benefit of antibiotic use in adults, that is, to ameliorate symptoms. Conversely, oropharynx cultures in adult patients would be indicated in special situations, such as the epidemiological investigation of episodes or monitoring of development and dissemination of S. pyogenes antibiotic-resistant strains 7.

When the diagnosis of streptococcal pharyngitis is confirmed, the clinician should select the most appropriate antimicrobial agent. The treatment regimen should be analyzed concerning specificity, safety and cost of drug 2.

In Brazil, many patients look for medical care through public or private healthcare system. However, a large proportion of the population makes use of self-medication or goes directly to the drugstores to ask for advice. It is important to emphasize that in our study, none of the 26 patients seen in the healthcare center were ordered laboratory analysis and the treatment regimen was prescribed based on clinical aspects only. Considering the positive results of oropharynx cultures obtained in our study, only 9 of the 32 patients that came to the drugstore should have been treated with antibiotics. Unfortunately, owing to complaints of fever, malaise and sore throat, over 80% of the subjects purchased antibiotics to treat pharyngitis.

Since 1940, penicillin has been the antimicrobial of choice for the treatment of streptococcal infections, owing to its limited action spectrum, few adverse reactions and low cost 6. All strains of S. pyogenes isolated to present are sensitive to penicillin, however, some strains present tolerance to the antibiotics, that is, to these strains the minimal bactericidal concentration of penicillin is 32 times higher than the minimal inhibitory concentration 14.

Treatment failure has been associated with inappropriate dosages of penicillin during the treatment and with the presence of penicillin-tolerant samples of S. pyogenes, capable of invading the epithelial cells in which they remain protected from the action of the antimicrobial agent 4, 15. In addition to these factors, the coexistence of beta-lactamase producing bacteria in the pharynx and tonsils can be caused by degradation of penicillin and allows survival of S. pyogenes in the infected area. In these situations, the bacteria can be eradicated in the respiratory tract by the use of amoxycillin associated with clavulanic acid 1.

Erythromycin should be the drug of choice for the treatment of patients allergic to penicillin. First and second generation cephalosporins are also indicated for the treatment of allergic patients that do not manifest immediate hypersensitivity to beta-lactam agents 5.

S. pyogenes samples resistant to macrolides are prevalent in some areas of the world and this characteristics had resulted from treatment failure. Less than 5% of the strains of S. pyogenes isolated in different countries such as Canada, United States, Sweden and Turkey, are resistant to erythromycin, however, the highest levels are found in Finland (20%), Spain (26.6%) and Italy (31%) 16-18.

In addition to erythromycin, another macrolide, azythromycin, has been used with advantages in penicillin allergic patients, since it is better tolerated than erythromycin and can be administered in a single dose by oral route for only 5 days, contrarily to penicillin, erythromycin and cephalosporins that require a 10-day period of treatment. However, the average price of a 5-day treatment course with the recommended dose is seventy reais, compared to six reais spent in treating with a single dose of intramuscular penicillin G (600,000 U) or twenty-two reais spent with the use of oral amoxycillin (250mg, TID). In addition, resistance to macrolides is developed quickly with the intensive use of drugs, which does not occur with penicillin. Thus, the use of new macrolides such as azythromycin as first line therapy should be carefully considered 2, 8.

It is important to emphasize that short period of treatment, associated with high price and lack of knowledge of selection of resistant strains caused by indiscriminate use of antimicrobial agents have stimulated the sales of azythromycin by drugstore sales clerks, as opposed to more effective and cheaper antimicrobial agents.

Some antimicrobial agents such as sulphoniamide and tetracyclines are not recommended for the treatment of streptococcal pharyngitis owing to the high rates of microbial resistance and the frequent failure in eradicating even sensitive strains 2.

Primary prevention of rheumatic fever is obtained by appropriate treatment of streptococcal pharyngitis by eradicating oropharynx group A streptococcus. Rheumatic fever is a world health problem since it is the main cause of acquired heart disease in children in the world, more frequently seen in countries in which medical assistance is deficient and in many children that live under poor sanitary conditions 1, 6. As to inappropriate prevention measures, high rates of rheumatic fever can be found if compared to communities in the developed world who have access to appropriate medical care 6.
Another aspect that should be highlighted is that the oropharynx culture and the diagnostic quick test cannot differentiate asymptomatic carriers from pharyngitis of non-streptococcal etiology. Thus, the importance of these tests is that they demonstrate that in most patients with acute pharyngitis there is no need to take antibiotics, which is confirmed by the negative test. In summary, the association of clinical and laboratory diagnoses allows better definition of treatment of streptococcal pharyngotonsillitis by rationally using the effective antibiotics that determine the appropriate form to cure infections, prevent suppurative and non-suppurative complications and eradicate oropharynx microorganism.

CONCLUSION

The results of the present study emphasized the importance of the bacteriological diagnosis in treating streptococcal pharyngotonsillitis to allow the prevention of suppurative or non-suppurative complications and the eradication of oropharynx microorganism.

ACKNOWLEDGEMENT

To undergraduate Solange A. Braziel, for the collection of clinical specimen in patients seen by the healthcare unit.

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1 Specialization Course on Health Sciences under course, State University of Maringá (UEM).
2 Undergraduate, School of Pharmacy, UEM.
3 Undergraduate, School of Nursing, UEM.
4 Faculty Professor, Discipline of Microbiology, UEM.
5 Joint Professor, Discipline of Microbiology, UEM.
Address correspondence to: Dra. Lourdes Botelho Garcia. Avenida Brasil, 475 Bairro Aeroporto Maringá PR 87050-000.
Tel (55 44) 261-4429 - E-mail: lbgarcia@uem.br
Article submitted on September 01, 2003. Article accepted on September 25, 2003.

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