The Prevalence of Acinetobacter in hospitals and its resistance to Beta-lactams

 

Danial Dalloul¹*, Lama Doya¹, Iyad Almahmoud², HaissamYazigi³

¹Postgraduate Student in Laboratory Diagnosis Department, Faculty of Medicine, Tishreen University,

Latakia-Syria

²Doctor at Laboratory Diagnosis Department, Faculty of Medicine, Tishreen University, Latakia-Syria

³Professor at Laboratory Diagnosis Department, Faculty of Medicine, Tishreen University, Latakia-Syria

 

ABSTRACT:

Background: Acinetobacter species are ubiquitous in the environment, and have emerged as important nosocomial pathogens. They are resistant to many antibiotic groups, especially beta lactams which is the most clinically-important group. However, There is a poor knowledge about Acinetobacter susceptibility profile in Syria. Aim of the study: To detect distribution of Acinetobacter in different sections of involved hospital and its susceptibility profile to beta-lactams. Material and methods: 88 Acinetobacter samples obtained through 24 months from different departments of Al-Assad University Hospital, Latakia- Syria. Species was identified using api 20E and api 20NE (bioMérieux-France) and susceptibility profile for beta-lactams was detected using E-Test (bioMérieux-France) Results: The prevalence of Acinetobacter has reduced from 3.74% in the first year (2015) to 3.06% in the second year (2016), but the resistance to antibiotics used in the study has increased( except Azetreonam). The resistance rates were 11.4% for Ampicillin-Sulbactam, 78.4% for Ticarcillin-Clavulanic acid, 90.9% for Cefotaxime, 43.2% for Cefepime, 48.9% for Imipenem, 40.9% for Meropenem, and 44.3% for Azetreonam. Conclusion: Acinetobacter develops resistance for beta-lactams rapidly. Ampicillin-Sulbactam is the drug of choice for Acinetobacter infections in Syria.

 

 

 

INTRODUCTION:

Acinetobacter species are ubiquitous in the environment, and have emerged as important nosocomial pathogens. Resistance to drying and many commonly used antimicrobial agents are the key factors that enable these organisms to survive and spread in nosocomial environments^[1]. Acinetobacter species are non-fermenting gram-negative bacilli. Swimming motility is negative but 'twitching motility' on soft agar may occur^[1],[2].

 

Morphologically, Acinetobacter usually presents in pairs or long chains of variable length^[3]. Colonies on blood agar, which display typical shape, are white to creamy coloured, smooth or mucoid (when capsule is present). Whereas colonies display bluish to bluish gray colour on eosin-methylene blue (EMB) agar^[4]. Biochemically, the genus Acinetobacter comprises species that are strictly aerobic, catalase-positive, indole- negative, oxidase-negative, and citrate-positive^[5].

 

On the other hand; Beta-Lactams group is the most important member of "Bacterial Cell Wall Inhibitors" antibiotics. Beta-Lactams can be divided into four groups: 1: Penicillins: which involves four subgroups: natural penicillins (i.e. penicillin G), antistaphylococcal penicillins (i.e. Methicillin, Oxacillin), extended-spectrum penicillin (i.e. Ampicillin), and antipseudomonal penicillins (i.e. Piperacillin, Ticarcillin). The last two subgroups are affictive against gram-negative bacteria^[6]. 2: Cephalosporins: which are classified as 1^st generation (i.e. cefazolin), 2^nd generation (i.e. cefaclor), 3^rd generation (i.e. cefotaxime), 4^th generation (i.e. Cefepime) and advanced cephalosporins (i.e. Ceftaroline). Their activity against gram-negative organisms can be seen in 3^rd, 4^th generations and advanced cephalosporins^[6],[7]. 3: Carbapenems: (i.e. Imipenem, Meropenem). 4: Monobactams: (i.e. Aztreonam). These last two groups both have a good effect on gram-negative bacilli^[7]. All beta-lactams interfere with the synthesis of bacterial cell wall; particularly, with the final step in which the terminal D-alanine chains crossly linked by transpeptidase (penicillin binding protein PBP)^[8]. Resistance to beta-Lactams is due to one of four general mechanisms: 1. Inactivation of antibiotic by producing betalactamase. 2. Modification of target PBPs. 3. Impaired penetration of antibiotic to target PBPs which only happens in gram-negative bacteria. 4. Efflux^[7]. However, the distribution of Acinetobacter in different sections of involved hospital and its susceptibility profile to beta-lactams are the aim of this study.

 

MATERIAL AND METHODS:

This study was carried out in Al-Assad University Hospital, Lattakia, Syria from Jan. 2015 to Dec. 2016. A pre-total of 94 of suspected isolates were collected from different sections of mentioned hospital, depending on microscopic morphology, cultures' characteristics and api 20E (bioMérieux-France) results - as a routine method for bacterial identification used in the hospital's laboratory-. All the isolates were suspended in 20% glycerol stock and kept at -70° C for long-term storage. The isolates were sub-cultured twice on to nutrient agar plates (blood agar and EMB agar) and incubated at 37°C overnight before they were used. The accurate detection for species was accomplished using api 20NE (bioMérieux-France). A total of 88 isolates (n=47 in 2015, n=41 in 2016) were considered as Acinetobacter species according to api 20NE results. Susceptibility to beta-lactams were determined using the E-Test (bioMérieux-France) method. Depending on their activity against gram-negative bacteria, a collection of seven beta-lactams were chosen from the four groups: Ampicillin-Sulbactam and Ticarcillin-Clavulanic acid from penicillins' group, Cefotaxime and Cefepime from cephalosporins' group, Imipenem and Meropenem from Carbapenems' group, and Azetreonam from Monobactams' group. We also use a disk of Teicoplanin (glycopeptide antibiotic which have no effect on gram-negative bacteria) as a negative control.

 

Image (1): E-Test method used in our study:

 

Api 20NE: is a standardized system for the identification of non-fastidious, non-enteric Gram-negative rods (e.g. Pseudomonas, Acinetobacter, etc), combining 8 conventional tests, 12 assimilation tests and a database. The API 20 NE strip consists of 20 micro-tubes containing dehydrated substrates. The conventional tests are inoculated with a saline bacterial suspension which reconstitutes the media. During incubation, metabolism produces color changes that are either spontaneous or revealed by the addition of reagents. The assimilation tests are inoculated with a minimal medium and the bacteria grow if they are capable of utilizing the corresponding substrate. The reactions are read according to the Reading Table and the identification is obtained by referring to the Analytical Profile Index or using the identification software. Api 20NE is considered as good bacterial identification method on species level^[9].

 

E-Test: The E-Test is a dilution test based on the diffusion of a continuous concentration gradient of an antimicrobial agent from a plastic strip into an agar medium. The plastic strip, which has a predefined concentration of dried and stabilized drug on one side and a continuous MIC interpretive scale on the other, is placed on the surface of an agar medium inoculated with the organism to be tested. The plate is incubated according to the atmosphere and time required for the specific organism. After incubation, an ellipse of growth inhibition is formed around the strip, and the MIC is read at the point on the scale where the ellipse intersects the strip^[10] . Depending on MIC values, strains are divided into three groups: Sensitive (S), Intermediate (I), Resistant (R). E-Test yields excellent category agreement results when compared with the approved methods of detecting antimicrobial susceptibility ^{[11] , [12]}.

 

Data analysis:

Data management and statistical analysis were performed using IBM SPSS version 20.

 

 

 

 

RESULT AND DISCUSSION:

The 88 study isolates were distributed as shown in Table (1). There was a lack of Acinetobacter rates in approximately all sections and all types of clinical samples between the two years of the study. Acinetobacter involved in 3.74% of nosocomial infections in 2015, compared to 3.04% in 2016. That may due to the elevation of other bacterial infections. That probably related to the increasing in other bacteria involved in nosocomial infections.

 

 

Table(1): The distribution of study isolates in clinical samples and different departments in Al-Assad University Hospital

*N: number of total bacterial samples, **A: number of Acinetobacter isolates, ***P: percentage of Acinetobacter isolates. (I) D.: Department, (II) S.: Sample type. (1) ICU: Intensive Care Unite, (2) Sur. : Surgery department, (3) Int. : Internal medicine department, (4) Ped. : Pediatric and Premature department, (5) Gyn. : Gynecology and Obstetrics department, (6) Resp.Sam. : Respiratory System Samples (sputum, bronchial lavage, pleural fluid), (7) C.S.F : Cerebro-spinal fluid

 

 

In general, the prevalence of Acinetobacter in Al-Assad University Hospital as elucidated in Table (2) shows that intensive care unit (ICU) was the most affected section by Acinetobacter as 11.8% of isolates came from ICU were identified as Acinetobacter. That is probably related to the increasingly invasive diagnostic and therapeutic procedures used in hospital ICUs nowadays ^[13]. On the other hand, the highest rate of infections caused by Acinetobacter were bacteremia especially in pediatric and neonatal (6.6% of positive blood cultures were Acinetobacter) followed by wounds infections (5.72% of all wounds infections were caused by Acinetobacter).

 

 

Table(2): The prevalence of Acinetobacter  over the study period

*P: percentage of Acinetobacter distribution. (I) D.: Department, (II) S.: Sample type. (1) ICU: Intensive Care Unite, (2) Sur. : Surgery department, (3) Int. : Internal medicine department, (4) Ped. : Pediatric and Premature department, (5) Gyn. : Gynecology and Obstetrics department, (6) Resp.Sam. : Respiratory System Samples (sputum, bronchial lavage, pleural fluid), (7) C.S.F : Cerebro-spinal fluid

 

On contrary, the resistance rate to selected beta-lactams antibiotics has elevated through the two years. Acinetobacter strains in 2016 became more resistant to all beta-lactams used in the study (except Azetreonam) in comparison with their resistance rate in 2015.The lack

 

of Azetreonam popularity in clinical usage probably what lies behind its stable resistance rate. The resistance rates to each beta-lactams antibiotic used in our study in both 2015 and 2016 years are illustrated in Figure(1).

 

 

Figure(1): The resistance rate to selected beta-lactams  antibiotics throughout the study period

(1) AB: Ampicillin-Sulbactam. (2) TLC: Ticarcillin-Clavulanic acid. (3) CT: Cefotaxime. (4) PM: Cefepime. (5) IP: Imipenem.

(6) MP: Meropenem. (7) AT: Azetreonam

 

 

Table(3): resistance rate for beta-lactams used in the study

(1) AB: Ampicillin-Sulbactam. (2) TLC: Ticarcillin-Clavulanic acid. (3) CT: Cefotaxime. (4) PM: Cefepime. (5) IP: Imipenem.

(6) MP: Meropenem. (7) AT: Azetreonam

 

*Anti: antibiotics which used in our study

 

Figure (2): Isolates distribution depending on number of resisted antibiotics

 

However, the overall resistance rates for beta-lactams antibiotics used in the study are indicated in Table(3). The highest resistance rate was for Cefotaxime (CT) 90.9% this result was emerged in other research ^{[14], [15]}. Whereas, the lowest resistance rate was for Ampicillin-Sulbactam (AB) 11.4% which has no correspondence with some other research ^[16].

 

The resistance has not increased in its range only, but also in its spectrum. Number of isolates and number of beta-lactams antibiotics they resist are shown in Table(4). According to the table(4), there were two isolates in 2015 (4.3%) that had no resistance to any beta-lactams antibiotics, whereas in 2016 there were not isolates without resistance. On the other hand, isolates that resisted 6-7 beta-lactams antibiotics increased in 2016. Overall, most isolates were resistant to 2-5 beta-lactams antibiotics.

 

We observed that isolates tended to resist more beta-lactams antibiotics in 2016 in comparison with 2015. Figure(2) illustrates the distribution of isolates depending on number of resisted antibiotics. It is observed from Figure(2) that most isolates in the year 2015 were resistant for no more than 3 beta-lactams antibiotics, whereas most isolates in 2016 were resistant for 4 beta-lactams antibiotics at least.

 

CONCLUSION:

ICU is the most affected section by Acinetobacter. Bacteremia and wounds infections are the most common infections caused be this species. More procedures should be done in order to reduce this incidence like affective sterilization and using vaccination ^[17]. Acinetobacter develops resistance rapidly ^[18]and more research should be done to detect the resistance mechanisms. Ampicillin-Sulbactam is the drug of choice in Acinetobacter infections in Syria according to our results

 

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Received on 16.10.2017         Modified on 17.11.2017

Accepted on 28.12.2017      © RJPT All right reserved