Antibiotic susceptibility and serotype patterns of Pseudomonas aeruginosa from clinical isolates in Abidjan, Cote d’Ivoire

Pseudomonas aeruginosa (P. aeruginosa) has been found to be a common hospital acquired pathogen, responsible for several severe infections. The objective of this study was to describe the antibiotic resistance profile of P. aeruginosa strains from human sample. This descriptive study was carried out on 168 isolated strains of P. aeruginosa collected from January 2014 to December 2015 at the Pasteur Institute of Côte d'Ivoire. The strains identification was done according to the methods of conventional bacteriology. The antibiotic sensitivity tests were performed using the disc diffusion method in agar medium according to CA-SFM (Antibiogram committee of French society of microbiology) criteria. The serotyping of the strains was carried out by using the agglutination method slide test, with the aid of 4 specific polyvalent antisera. The most prevalent P. aeruginosa serotypes were O4 (24.4); O11 (14.6); and O6 (9.5%). The rate of antibiotic resistance to ticarcillin was 32.9%, ciprofloxacin 18.4%, ceftazidime 14.9%, Imipenem 11.3%, and amikacin 11.3%. Resistance to Imipenem was above 10% in an intensive care unit and in the pneumonology unit (PPH). Strains of O6 serotypes were the most multidrugresistant followed by O11 with respective rates of 31.2 and 28% MDR (Multidrug Resistance). P. aeruginsa are microorganism capable of developing mechanisms of complex resistance which makes it difficult to manage. The attention of hygiene rules and the rational use of antibiotics are very important in other to prevent the spread of MDR P. aeruginosa.


INTRODUCTION
Pseudomonas aeruginosa has become a major hospital acquired pathogen, responsible for several severe infections (Bertrand et al., 2013). Due to its ability to use different organic compounds as energy substrates, this strictly aerobic gram-negative bacillus lives in very humid environments (Fuentefria et al., 2011).
Hospital environment is suitable for this micro-organism because it contains numerous ecological niches (Bertrou et al., 2000). The intestinal carriage of the pyocyanic is rare in healthy subjects, but frequent among hospitalized patients, particularly in intensive care unit (Bertrand et al., 2013). Due to its natural resistance to many antibiotics and its ability to build up new resistance, the hospital environment favors the growth of P. aeruginosa despite multiple antibiotic selection pressure.
The emergence of multidrug-resistant strains of P. aeruginosa in hospital is often associated with some serotypes (Thrane et al., 2015). In order to find an epidemiological link between the strains of P. aeruginosa isolated in hospital, phenotypic markers such as antibiotype and serotype are used to differentiate strains before genetic characterization (Blot et al., 2013;Wolska et al., 2012). There is paucity of data on P.aeruginosa infections in Côted' Ivoire. The aim of this study was to evaluate the antibiotic resistance profile of P. aeruginosa and to compare these profiles with serotypes of isolated strains.

MATERIALS AND METHODS
This descriptive study was carried out on strains P. aeruginosa, isolated and collected from January 2014 to December 2015 at the Clinical Bacteriology Unit (CBU) of the Department of Bacteriology-Virology of Pasteur Institute of Côte d'Ivoire. The variables taken into account were: the origin of the strains, type of services, nature of the samples, antibiotic profile and the type of the serotype. Duplicates were not considered.

Strains of P. aeruginosa
A total of 168 strains of P. aeruginosa were isolated from various biological fluids: pus (87), sputum (320), urine (16), blood (13), material (13), Cerebrospinal fluid (6) and stools (1). Strains isolated from other media were examined for growth and pigmentation on Pseudomonas isolation agar. The strains were identified based on the standard bacteriological characteristics: appearance of colonies on King A and King B medium, presence for pyocyanine and pyoverdine, growth at 42°C, morphological appearance (gram negative bacillus, polar mobile), the positive oxidase reaction and other biochemical characteristics using the API 20 NE gallery (Bio-Mérieux®, France) according to Breed et al. (2000) and Garrity et al. (2005). The strains were stored in deep agar at room temperature.

Study of antibiotic sensitivity
Antibiotic susceptibility tests were performed using Kirby-Bauer disc diffusion method in agar medium (Kirby-Bauer, 1996). The reading and interpretation were done according to the recommendations of the Antibiogram Committee of the French Society of Microbiology (CA-SFM 2015).
In Table 2, the rate of sensitivity of 168 strains of P. aeruginosa to different antibiotics tested is summarized. The proportion of resistant strains was high for ticarcillin (34.5%) and ticarcillin-clavulanic acid (31.5%). The rate of strains resistant to cetazidime was 14.9 and 18.4% for ciprofloxacin. The resistance rate to Imipenem was 11.3%. Distribution of resistant strains according to their isolation origin showed a higher proportion from hospital strains compared to isolates from non-hospitalized patients. Among hospital services, the highest rates of resistance were observed in pneumonology unit and neurology for ticarcillin (50 and 58.3%, respectively) and ticarcillin-clavulanic acid (43.3 and 50%, respectively). In Intensive care unit, 28.6% is from resistant strain to imipenem. About 23% of strains isolated from pneumonology were resistant to ciprofloxacin (Table 3).
The serotypes of P. aeruginosa constitute epidemiological markers with the rate of 24.4, 14.9 and 9.5% for serotypes O4, O11 and O6, respectively. In Tunisia, a study carried out in 2012 reported the same serotypes with variable rates predominance of serotypes O11, followed by O6 and O4 (Zoghlami et al., 2012). Serotype O4 was predominant in prolong hospitalization units (19.8%) (Adjerald et al., 1999).
Amikacin and imipenem were the most active antimicrobial agent on P. aeruginosa isolates with resistance rates of 7.1 and 10.7%, respectively. Nigeria resistance rates were 13% for imipenem and 23% for amikacin (Iregbu et al., 2015).
In Côte d'Ivoire in 2008, P. aeruginosa isolated from infections which originate from surgical operation site shown a sensitivity of 98.5% to an imipenem, that is 1.5% resistance with lower rate (Faye-Kette et al., 2008) . A study carried out in 2013 on hospital effluents showed very high rates of resistance to imipemen with 80% of resistant strains (Guessennd et al., 2013).In Teheran, higher rate were found for imipenem (41.3%) and amikacin (28%) (Peymani et al., 2017).
The highest resistances were observed in strains isolated from the department of pulmonology with 50% resistance for ticarcillin, 23.3% for ciprofloxacin and 20% for ceftazidime. This observation is confirmed by Hamze et al. (2013) in Lebanon.

Conclusion
This study showed the importance of serotyping of P. aeruginosa isolated from clinical sources. The circulation of serotypes O4, O11 and O6 of P. aeruginosa was frequent. They were predominant in suppurative and pleuro-pulmonary secretions.
In addition, serotypes O11 and O6 were the most multiresistant. The most common phenotype of MDR is Ticarcillin-Ceftazidime-Ciprofloxacin. Knowledge of serotypes can guide the choice of antibiotic therapy in 24 h before sensitivity test results. In addition, this study should alert health professionals to an increasing rate of P. aeruginosa resistant to useful carbapenems and Fluoroquinolones. Regular monitoring of the antimicrobial resistance profile is essential to guide prescribing antibiotics and controlling the emergence of MDR P. aeruginosa strains.