Brief Report

Evaluation of Antimicrobial Resistance and Frequency of bap Gene Among Acinetobacter baumannii Strains Isolated from Hospitalized Patients

Please cite this article as follows: Mousavi T, Shushtari A, Amiri E, Moshiri M, Gholami M. Evaluation of antimicrobial resistance and frequency of bap gene among acinetobacter baumannii strains isolated from hospitalized patients. Avicenna J Clin Microbiol Infect. 2025;12(4):231-234. doi:10.34172/ajcmi.3718

Introduction

Acinetobacter baumannii is a Gram-negative, aerobic bacterium with multidrug resistance (MDR) and multiple virulence factors. It possesses various antibiotic resistance mechanisms and causes serious problems in immunocompromised patients (1,2). Clinical isolates of A. baumannii can form biofilms on diverse surfaces. Notably, the emergence of biofilm-producing, antibiotic-resistant Gram-negative bacteria has become a growing concern, particularly among pathogens commonly encountered in healthcare environments (3).

A large 854-kDa protein known as biofilm-associated protein (BAP) is a key factor in biofilm production and adhesion to host cells (4). In recent years, only a few studies have examined the prevalence of the bap gene and its association with antibiotic-resistant A. baumannii strains.

Given the limited data on biofilm-forming ability and the prevalence of the biofilm-associated bap gene in A. baumannii, as well as its connection to antibiotic resistance in northern Iran, this study aimed to investigate the antibiotic susceptibility profiles and conduct a molecular evaluation of the bap gene in A. baumannii strains isolated from hospitalized patients.

Materials and Methods

Sample Collection

This cross-sectional study (February 2022 to June 2023), approved under the ethics code IR.MAZUMS.REC.1401.240, included 100 clinical samples collected from teaching hospitals in Sari County. The samples were obtained from pulmonary secretions, blood, wounds, urine, broncho-alveolar lavage (BAL), abscess, urinary catheters, pleural fluid, and cerebrospinal fluid (CSF).

Isolation and Identification of Acinetobacter baumannii Isolates

All samples were transferred to the microbiology laboratory and cultured on blood agar and MacConkey agar media. The inoculated plates were subsequently incubated at 37 °C for 24 hours to allow optimal bacterial growth. Preliminary identification of the isolates was performed using standard biochemical tests (5). Final confirmation of the A. baumannii strain was achieved through a standard polymerase chain reaction (PCR) assay targeting the bla-OXA-51 gene (bla-OXA-51-F: 5′-TAA TGC TTT GAT CGG CCT TG-3′, bla-OXA₅₁-R: 5′-TGG ATT GCA CTT CAT CTT GG -3′) (6).

Antibiotic Susceptibility

Antibiotic susceptibility of all isolates was evaluated using the Kirby-Bauer disk diffusion method on Mueller-Hinton agar, following the 2021 Clinical and Laboratory Standards Institute (CLSI) guidelines. The susceptibility of the isolates was assessed using the following antimicrobial disks: gentamicin (30 µg), tetracycline (5 µg), imipenem (10 µg), tobramycin (10 µg), trimethoprim-sulfamethoxazole (30 µg), ceftazidime (30 µg), doripenem (30 µg), piperacillin-tazobactam (10 µg), ciprofloxacin (5 µg), cefepime (30 µg), piperacillin (30 µg), cefazolin (30 µg), amikacin (30 µg), and ampicillin (30 µg) (MAST, UK).

The minimum inhibitory concentration (MIC) for colistin against A. baumannii (0.25–128 μg/mL) was determined using the CLSI broth microdilution (BMD) method. MIC values were evaluated based on CLSI guidelines (7). Escherichia coli ATCC 25922 was employed as the standard reference strain for quality verification.

Identification of bla-OXA-51 Gene in Acinetobacter baumannii Isolates

In this study, the genome of 100 isolates was obtained using the boiling extraction method. PCR amplification was conducted under the following conditions: initial denaturation at 95 °C for five minutes, 30 cycles of 95 °C for 60 seconds, 50 °C for 50 seconds, 72°C for 60 seconds, followed by a final extension at 68°C for two minutes. DNA presence, concentration, and quality were assessed using agarose gel electrophoresis.

Polymerase Chain Reaction for Detection of the bap Gene

In the present study, the primers used for bap gene detection were: bap-F: 5′-TAGGGAGGGTACCAATGCAG-3′ and bap-R: 5′-TCATGATTTGATGCTGCAGCG-3′, applied in the PCR method (8).

Results

Descriptive Epidemiology

In the current study, 100 A. baumannii isolateswere collected from different clinical specimens, including pulmonary secretions (46%), blood (30%), wound (10%), urine (5%), BAL (3%), abscess (2%), urinary catheter (2%), pleural fluid (1%), and CSF (1%). Of the isolates identified as Acinetobacter spp. phenotypically, all were confirmed as A. baumannii using bla-OXA-51 primers, which yielded the expected 353 bp bands on agarose gel electrophoresis.

Antimicrobial Susceptibility Testing

The MIC results indicated that out of the 100 A. baumannii strains, 97 were susceptible and 3 were resistant to colistin. Susceptibility to other antibiotics was assessed using the standard disk diffusion assay. The detailed susceptibility profiles of other tested antibiotics are presented in Figure 1.

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Figure 1.

Antibiotic Resistance Patterns of A. baumannii Isolates. Note: AK: Amikacine; AMS: Ampicillin-sulbactam; CAZ: Cefazolin; CFZ: Ceftazidime; CIP: Ciprofloxacine; COL: Colistin; CPM: Cefepime; DOR: Doripenem; GEN: Gentamicin; IPM: Imipenem; PRL: Piperacillin; SXT: Trimethoprim-sulfamethoxazole; TE: Tetracycline; TOB: Tobramycin; TPZ: Piperacillin-tazobactam

Figure 1.

Antibiotic Resistance Patterns of A. baumannii Isolates. Note: AK: Amikacine; AMS: Ampicillin-sulbactam; CAZ: Cefazolin; CFZ: Ceftazidime; CIP: Ciprofloxacine; COL: Colistin; CPM: Cefepime; DOR: Doripenem; GEN: Gentamicin; IPM: Imipenem; PRL: Piperacillin; SXT: Trimethoprim-sulfamethoxazole; TE: Tetracycline; TOB: Tobramycin; TPZ: Piperacillin-tazobactam

Presence of bap

All isolates involved in this study carried the A. baumannii bap gene (Figure 2).

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Figure 2.

PCR Image of Three A. baumannii Isolates (1,2,3) Showing 405 bp DNA Bands Representing the bap Gene. Note. M: DNA ladder; N: Negative control; P: Positive control

Figure 2.

PCR Image of Three A. baumannii Isolates (1,2,3) Showing 405 bp DNA Bands Representing the bap Gene. Note. M: DNA ladder; N: Negative control; P: Positive control

Discussion

Over the past decade, A. baumannii has attracted increasing research interest as a major multidrug-resistant pathogen (9-11). Among its virulence-related genes, the bap gene has received considerable attention due to its critical role in biofilm formation, which enhances the persistence of A. baumannii in hospital environments where exposure to disinfectants is frequent (12,13).

In Iran, several studies have reported that the frequency of the bap gene among A. baumannii isolates ranges from 43% (14) to 100% (15). These high frequencies highlight the worldwide distribution and dissemination of this important virulence factor in A. baumannii strains. Similarly, in a study conducted on 53 A. baumannii clinical isolates collected from inpatients in Iraq, the frequency of the bap gene was reported to be 79% (16). The lowest frequency (43%) was reported by Zeighami et al, who examined 100 A. baumannii strains recovered from immunocompromised patients hospitalized in the intensive care units (ICUs) (14). The low frequencies observed in some studies may be due to geographical variations and differences in the source of isolates. For example, one of our reference hospitals (Shahid Zare) is a burn-specialized hospital where A. baumannii strains are exposed to extreme amounts of disinfectants and prophylactic antibiotics. The capacity of A. baumannii to form biofilms has been closely linked to the development of an MDR phenotype. Consistent with our observations, previous studies have reported a strong association between the presence of the bap gene and biofilm formation, as well as increased biofilm production in MDR isolates, highlighting the critical role of bap-related biofilms in enhancing antimicrobial resistance mechanisms in A. baumannii (17)

However, carbapenems (imipenem and doripenem) and cephalosporins (ceftazidime, cefepime, and cefazoline) were found to be completely ineffective against A. baumannii isolates, as none of the isolates exhibited susceptibility to these drugs. Interestingly, all isolates were resistant to imipenem and doripenem, as these drugs (Carbapenem family) are the first-line treatment for A. baumannii infections in many areas. Consistent with our findings, multiple studies have reported similarly high resistance rates of A. baumannii isolates to carbapenems (16,18-21).

These results suggest that carbapenems should be decommissioned as the drugs of choice for A. baumannii infections. In contrast, colistin and aminoglycosides (tobramycin and gentamicin) showed excellent activity against A. baumannii isolates. Several other studies have similarly reported a high susceptibility profile of A. baumannii isolates to colistin and aminoglycosides (16,18), supporting their use as potential therapeutic options for A. baumannii infections in Mazandaran province and northern Iran.

Conclusion

In conclusion, our study highlights the significant burden that A. baumannii imposes on healthcare systems and patients, primarily due to its MDR nature. We identified colistin, tobramycin, and gentamicin as effective treatment options, underscoring the need to reconsider reliance on traditional antibiotics such as carbapenems.

Acknowledgments

We thank the hospital staff for their assistance in collecting clinical isolates and preparing patient data.

Competing Interests

The authors declare no conflict of interests.

Ethical Approval

This study was approved by the Research Ethics Committees of Mazandaran University of Medical Sciences (MAZUMS) (Approval ID. IR.MAZUMS.REC.1401.240).

Funding

This work was funded by Mazandaran University of Medical Sciences, Sari, Iran (Grant No. 13945).

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