Ecological aspects of antimicrobial susceptibility of anaerobic bacteria in Nicaragua

Abstract

Anaerobic bacteria are the predominant constituents of the normal flora on the skin and the mucous membranes of the human body. They may also act as potent pathogens in a variety of endogenous infections. Antimicrobial resistance is an increasing problem with anaerobic and aerobic bacteria. Different mechanisms for such resistance are known. However, [beta]-lactamase production is the most common mechanism of resistance. Emergence of resistant organisms in the normal flora during the administration of antimicrobial agents has a tremendous impact on the selection of antimicrobial agents for empirical therapy and is an important factor in the dissemination of resistant organisms to other patients. Anaerobic infections are common and frequently associated with severe morbidity and mortality, so empirical therapy has to be instituted as early as possible because of delays in obtaining the results of culture and susceptibility studies. The appropriate therapy should be based on local susceptibility patterns, as the prevalence of microorganisms causing infections and the antimicrobial susceptibility patterns may vary within geographic regions and even within hospitals.

The aims of this thesis are to determine the antimicrobial susceptibility patterns and [beta]-lactamase production of anaerobic bacteria from the oral and gastrointestinal microflora of (i) hospitalized antimicrobial treated adults from Nicaragua and Sweden; (ii) antimicrobial treated and healthy Nicaraguan children under 2 years of age; (iii) aerobic and anaerobic bacteria isolated from Nicaraguan patients with mixed infections; and (iv) to estimate the class-specific antibody response against Bacteroides fragilis antigens in Nicaraguan and Swedish patients with intraabdominal infections; and (v) to determine the prevalence of enterotoxigenic B. fragilis in children with and without diarrhea in order to provide the physicians with data generated in Nicaragua to facilitate the selection of antimicrobial therapy.

Bacteroides, Clostridium and Fusobacterium species resistant to at least one of the following antimicrobials: ampicillin, benzylpenicillin, piperacillin, cephalotin, cefoxitin and clindamycin, were isolated from the oral and gastrointestinal microflora of all antimicrobial treated adults included in this study. The percentage of resistant strains isolated from the Nicaraguan patients was higher than that found in the Swedish patients. Bacteroides strains resistant to ampicillin or cefoxitin were also isolated from the gastrointestinal normal flora of the children. The percentage of resistance increased with the age of the children (I month to 2 years of age). The resistant strains were mainly isolated from antimicrobial treated children. No resistance to cefoxitin was observed in the strains isolated from the healthy children until they were 18 months of age.

The antimicrobial susceptibility pattern of anaerobic and aerobic bacteria isolated from 219 Nicaraguan patients with different infectious diseases was determined against the most common antimicrobial agents used in Nicaragua. The B. fragilis group were the most commonly isolated anaerobes and the most resistant to ampicillin, cefoxitin, and clindamycin. Only five B. fragilis group strains were resistant to metronidazole. Fusobacterium strains resistant to ampicillin and cefoxitin were also isolated. However, no resistance to imipenem and chloramphenicol was found in these anaerobes. Clostridium, Peptostreptococcus, Propionibacterium and non-spore forming Gram-positive rods were sensitive to all the antimicrobial agents tested. Mixed infections were found in 66% of the patients, mainly from intraabdominal infections. Together with B. fragilis group strains, Escherichia coli strains were the most common bacterial association. Ampicillin, benzylpenicillin, cefoxitin and chloramphenicol were the antimicrobial agents less active against the E. coli, P. aeruginosa and Klebsiella strains. Gentamicin, one of the most important antimicrobial agents used, showed good activity. No more than 18% of the aerobic strains were resistant. Methicillin resistant Staphylococcus aureus was also common. No vancomycin resistance was found. Enterotoxigenic B. fragilis strains were isolated only from children with diarrhea (8.4%) and mainly when they were less than 2 years of age. All strains were resistant to ampicillin and one strain was also resistant to clindamycin.

The strongest [beta]-lactamase producers among the anaerobic bacteria isolated from normal flora and from infected sites were Bacteroides species followed by Fusobacterium species. Almost all the Gram-negative aerobic strains and S. aureus strains were [beta]-lactamase producers. A [beta]-lactamase from B. distasonis strains S:10:7 isolated from the saliva of a Nicaraguan patient was purified and characterized. The enzyme had a broad substrate profile, capable of hydrolyzing benzylpenicillin, ampicillin, piperacillin and cephalotin. No capacity to hydrolyze the [beta]-lactamase stable compounds cefoxitin and imipenem was observed. However, the B. distasonis strain S: 10:7 was resistant to cefoxitin. The enzyme was inhibited by clavulanic acid, sulbactam and tazobactam. The isoelectric point was 4.6 and the molecular weight 160,000 Daltons.

Elevated IgG titers were mainly seen against the B. fragilis capsular polysaccharide (CPS) antigens in both groups of patients with intraabdominal infections, compared with the titers observed against lipopolysaccharide (LPS) antigens. However, when the titers in Swedish and Nicaraguan IgG patients were compared, the Swedish titers were higher than the Nicaraguan titers. Patients with intraabdominal abscess and perforated appendicitis had the highest antibody responses against the B. fragilis CPS.

In conclusion, the results of the present investigation show a high level of resistance and B-lactamase production in Gram-negative anaerobic bacteria. This may be a reflection of the extensive antibiotic use in Nicaragua. In spite of these patterns, we can positively state that therapeutic alternatives are still available for the treatment of anaerobic bacterial infections because of the good results achieved by metronidazole, imipenem and chloramphenicol; less active, but still useful, were cefoxitin and clindamycin.