Prevalence of Drug-Resistant Strains of Streptococcus Pneumoniae in Abakaliki

Streptococcus pneumoniae is a major cause of illness such as pneumonia, meningitis, bacteremia and otitis media in children and the elderly. The emergence of drug-resistant strains threatens to complicate the management of these diseases. We conducted a hospital-based surveillance for drug-resistant Strep. Pneumoniae in outpatients with pneumococcal infection in Abakaliki Ebonyi State Nigeria. Between 20032005 from January through December, a cross-sectional study was conducted in Ebonyi State University Teaching Hospital (EBSUTH) Abakaliki to assess the prevalence of drug resistant strains of Streptococcus pneumoniae isolated from sputum samples of patients with pneumococcal infections attending the outpatient clinics. A total of 305 sputum samples of patients with clinically diagnosed pneumonia were collected and inoculated on 5% sheep-blood agar, incubated at 35°C for 24 h in 5-10% CO2. Colonies were Gram-stained; alpha-hemolysis colonies were tested with a 6mm optochin disk followed by bile solubility test. Susceptibility testing panels of the following antibiotics: penicillin, ciprofloxacin, clavulanic acid/amoxicillin, septrin, erythromycin, gentamycin, clarithromycin, cefotaxime and cefuroxime were tested against isolated strains of Streptococcus pneumoniae using disc diffusion method. Of the 305 sputum samples collected 30 (9.8%) of S. pneumoniae was isolated from patients within the age range of 20-40 years, 140 (45.9%) from patients within 41-60, 135(44.2%) from patients within 60 years and above. Susceptibility studies showed that the highest resistance was 182 (59.6%), for penicillin followed by septrin 156 (51.2%), erythromycin 120(39.3%), clavulanic/amoxicillin 118(38.7%) cefotaxime 114(37.4%) clarithromycin 100(32.7%), ciprofloxacin 94(30.8%), gentamycin 75(24.6%), cefuroxime 70(22.9%) and ceftriaxone 69(22.6%). The prevalence of S. pneumoniae resistance was relatively high and we suggest that proper antibiotics use should be adopted to avert development of multi-drug resistant strains in future.


INTRODUCTION
Streptococcus pneumoniae (pneumococcus) is the most common cause of community acquired pneumonia, meningitis Invasive pneumococcal disease is defined as isolation of S. pneumoniae from a normally sterile site which most frequently affects children less than 2 years old, adult at least 65 years AJID Science Publications old and immunocompromised individuals (Lynch and Zhanei, 2009a). Invasive pneumococcal disease is defined as isolation of S. pneumoniae from a normally sterile site which most frequently affects children less than 2 years old, adult at least 65 years old and immunocompromised individuals (children or adults) (Naheed et al., 2009). Globally, an estimated 1.6 million people including 1 million children less than 5 years old die of invasive pneumonia disease annually (Lynch and Zhanei, 2009b).
In the past, approximately 80 percent of patients hospitalized with bacteremic pneumococcal infections died of their illness (Peter et al., 2008). With effective antimicrobial agents, mortality has decreased but remains at nearly 20% for bacteremic diseases in elderly adults (Lynch and Zhanei, 2009a;Ramdani-Bouguessa and Rahal, 2003). In the United States of America over $4 billion is spent in treatment of pneumonia and other related diseases and emergency of drug resistant strains of S. pneumoniae will make these common infection more difficult to treat (Spika et al., 1991;Pallares et al., 1987;Viladrich et al., 1991;Michael, 1994). Following the initial detection of penicillin non-susceptible S. pneumoniae in a few geographic regions (e.g., South Africa, India, Australia, Spain) in the 1970s, resistance to penicillin and other antibiotics classes has spread worldwide (Kiran et al., 2010;Lynch and Zhanel, 2009b). However, the incidence of S. pneumoniae resistance varies markedly among countries or regions (e.g., Spain, France, Asia) but remains low (<5%) in some countries (e.g., Finland, Sweden, Germany) (Kiran et al., 2010). Rates of S. pneumoniae resistance in the United States and Canada are in the moderate range (Wierzbowski et al., 2007;. Prior antibiotic use is the dominant risk factor associated with drug resistant pneumococci (Kiran et al., 2010). Here in Nigeria, information on epidemiology of S. pneumoniae resistance is scare; hence the present study was designed to determine the prevalence of drug resistance strains of S. pneumoniae isolated from sputum samples of adult visiting outpatients clinics of EBSUTH Abakaliki, Ebonyi State Nigeria.

Collection of Clinical Specimen (Sputum)
Between 2003-2005, a total of 305 sputum samples of adults attending the outpatient clinic of Ebonyi State University teaching hospital Abakaliki were collected. The sputum samples were collected from patients with symptoms of lower respiratory tract infection especially cough and who was clinically diagnose of having respiratory tract infection. A case of pneumococcal infection was defined by the isolation of S. pneumoniae from sputum samples. Sputum samples of patients devoid of S. pneumoniae but was diagnosed of upper respiratory infection were not included in the study. Patients were also provided with questionnaire to identify risk factors for drug resistance S. pneumoniae acquisition which includes previous hospitalization, respiratory diseases, intravenous drug use, use of antibiotics for any care within the past years, chronic illness and tuberculosis infections. Consent of patients was obtained before samples were collected.

Isolation of S. Pneumoniae from Sputum Sample
Patient's sputum samples were collected and inoculated on 5% sheep-blood agar and incubated at 35°C for 24 h in 5-10% CO 2 . Colonies were Gramstained; alpha-hemolysis colonies were tested with a 6 mm optochin disk in sheep-blood agar dish followed by bile solubility test. Further identification and characterization was carried out using standard microbiology technique (Cheesbrough, 2006).

Susceptibility Studies
Susceptibility testing panels that includes antibiotics used in EBSUTH in treating patients with pneumonia which includes penicillin (10 µg), ciprofloxacin (5 µg), clavulanic acid/amoxicillin (20/10 µg), septrin (25 µg), erythromycin (15µg), gentamycin (10 µg), clarithromycin (25 µg), cefotaxime (30µg) and cefuroxime (30 µg) were tested against isolated strains of S. pneumoniae using disc diffusion method. Briefly, A Mueller-Hinton agar plate was prepared using manufacturers instruction, test organisms equivalent to 0.5 McFarlane equivalent standard was inoculated on the surface of sterile agar plate and was allowed for 15 min to pre-diffuse, then antibiotics disc as mentioned above was placed on the agar with sterile forceps. This was incubated for 18-24 h at 37°C, after which the inhibition zone diameter was taken with a transparent meter rule to the nearest millimeter. Susceptibility result was interpreted as resistance or susceptible or intermediate according to the definitions of the National Committee for Clinical and Laboratory Standard Institute.

DISCUSSION
The result obtained in the present study described the epidemiology period of one of the two major hospitals in Abakaliki, Ebonyi State in which low susceptibility to different classes of antibiotics was observed especially with penicillin, a common drug of choice for treatment of pneumonia. The hospital were this study was carried out represent this town population because it is the only University teaching hospital which serves as a referral hospital for patients with serious medical problems in Ebonyi state. Infections caused by Streptococcus pneumoniae such as otitis media, sinusitis, bronchitis and meningitis, which have led to a number of deaths especially in children and the elderly are a reason for worldwide concern. The overall prevalence of pneumonia infection that occurred within our study period (2003)(2004)(2005) was low. Previous antibiotic exposure has been documented as a risk factor for antibiotics resistance in many studies (Kaplan and Mason, 1998) but this could not be confirmed in our study. We could not demonstrate any relationship between the occurrence of resistant strain and the other risk factors previously described such as extreme age, hospital-acquired infections, probably due to small sample size. There are reports of high incidence of pneumonia in children and adults from different parts of the world (Hofman et al., 1995). Highest prevalence of resistant strains of S. pneumoniae was observed with penicillin in both adult and children. Such findings have been reported in and outside Nigeria (Jacobs et al., 1978;Ramdani-Bouguessa and Rahal, 2003;Simberkoff et al., 1986) and such was observed in our study. Recent studies of adults and children in a variety of communities have found high proportions of penicillin-resistant pneumoncocci. Discovery of penicillin was one of the major medical advances of the 20th century, being indicated for the treatment of community pneumonia caused by Streptococcus pneumoniae. Nevertheless, in the beginning of the 21st century it has been losing its role in the treatment of pneumonia due to the crescent increase of antimicrobial resistance (Barlett et al., 1998). Amoxicillin and clavulanic acid alone or associated macrolide can be used as an alternative drug of choice in the treatment of Streptococccus pneumoniae infections, although the optimal therapy for infections with drugresistant pneumococci is not well defined. Studies have suggested that cephalosporins or high dose penicillin may be effective in patients with non-meningeal bacteremic infections (Tan et al., 1992;Garcia-Leoni et al., 1992).
There are report that children six years of age were more likely than older children and adults to be infected with cefotaxime-resistant or multi-resistant isolates. However, we found a disturbingly high incidence of drug resistant pneumococcal infections among adults. Our data suggest that recommendations for empirical therapy are needed for pneumococcal infections in adults as well as children, particularly in communities in which the prevalence of drug resistant is high. The geographic variation in the prevalence of drug-resistant strains of pneumococci in Nigeria highlights the importance of community-based monitoring of pneumococcal susceptibility to antimicrobial agents to guide therapy. In addition, the increased prevalence of drug-resistant S. pneumoniae emphasizes the critical need for preventive strategies in populations at risk from serious pneumococcal infections.
Frequent and prophylactic use of antimicrobial drugs has been associated with a risk of drug-resistant pneumococcal infections but such was not observed in our study as all the risk factors checked could not be associated with the prevalence of multi-drug resistant in S. pneumoniae. Despite a low incidence of pneumococcal infection in Abakaliki particularly in patients between the ages of 20-40 years, all age groups was found to be associated with drug-resistant S. pneumoniae. Patients within the age range of 60 years and above were also associated with an increased rate of drug resistant S. pneumoniae. Rural residence and poverty may be responsible for poor health status and little or no access to medical care. As a result, people tend to see the roadside chemist/pharmacy dealers and procure cheap drugs without doctor's prescription because there is no restriction or guidelines to the use of antibiotics in Nigeria. A critical component for the control of drug-resistant S. pneumoniae will be community-wide educational programs for clinicians and the public on the importance of appropriate antibiotic use.
The vast majority of S. pneumoniae isolates had a relatively high resistant to all the antibiotics tested namely; penicillin, cefotaxime, septrin, erythromycin, clavulanic acid/amoxicillin, cefotaxime, clarithromycin, ciprofloxacin, gentamycin, cefuroxime and ceftriaxone. With the continuing spread of drug-resistant strains of pneumococci, treatment options will become more limited and prevention measures will become critical.

CONCLUSION
This study emphasizes the importance of antimicrobial-susceptibility testing of all S. pneumoniae infection in both children and adults in Abakaliki, Ebonyi State as well as a crucial need for communitybased programs of surveillance for drug-resistant pneumococcus to aid clinicians in their choice of therapy for pneumococcal infections. The prevention of infections with S. pneumoniae will require strategies to encourage judicious antibiotics use and recommendation for the treatment and prevention of pneumococcal infections must be addressed by health care and public health agencies. Our data suggest an urgent need for consensus guidelines to prevent development of multidrug resistant strains of Streptococcus pneumoniae in this region in future.