Pasteur Institute of Iran

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Introduction: Acinetobacter baumannii, a known causative agent of nosocomial infections, is one of the highly antibiotic-resistant gram-negative bacilli. Carbapenem-resistant Acinetobacter isolates are increasingly reported worldwide. Carbapenems such as imipenem and meropenem are efficient antimicrobial agents commonly used for the treatment of infections caused by multi- resistant A. baumannii strains. Some reports indicate treatment failure due to antibiotic resistant A. baumannii strains. The aim of this study was to determine antibiotic resistance pattern and prevalence carbapenemase production in A. baumannii isolates. Method: A total of 100 A. baumannii isolates were identified from clinical specimens by standard chemical tests. The samples were collected from the patients hospitalized in two teaching hospitals of Ahvaz, southwestern of Iran. The susceptibility of isolates to different antibiotics was determined by the disk diffusion method based on Clinical Laboratory Standards Institute (CLSI) direction. The Modified Hodge Test (MHT) was performed for detection of carbapenemase - producing A. baumannii isolates. Results: The isolates showed the highest resistance to ciprofloxacin (98%). The resistance rate to cefotaxime, ceftazidime, and piperacillin was 97%, gentamicin, amikacin, and meropenem 96%, imipenem 95%, cefepime 93%, and tetracycline 60%. Most of the isolates (99%) were sensitive to colistin. Among the100 A. baumannii isolates, 53 (53%) were positive for carbapenemase production by MHT. Conclusion: This study emphasizes dissemination of carbapenem resistant A. baumannii strains. Our study also showed that the MHT has an excellent sensitivity for detecting carbapenemase - producing A. baumannii isolates.

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Introduction: Acinetobacter baumannii is one of the major causes of nosocomial infections. We investigated the distribution of A. baumannii among patients and the environment in special care units of a hospital in Tehran, Iran. Methods: Sixty-nine non-duplicate clinical and environmental samples were collected from ICU, Post-ICU, and CCU in EbneSina Hospital (Tehran, Iran) from June 2015 to April 2016. The isolates were identified using standard biochemical tests, and their identity was confirmed by detection of bla_oxa51 gene. Susceptibility to 14 antibiotics was determined by disc the diffusion method, and genetic fingerprinting of the isolates was performed by random amplified polymorphic DNA (RAPD-PCR). Results: We recovered 66 A. baumannii isolates, 41 from patients, and 25 from the environment. All isolates from patients were resistant to all tested antibiotics except colistin. Environmental isolates were resistant to piperacillin (100%), tetracycline, piperacillin/tazobactam, beta-lactams and quinolones (86%), and amikacin (81%) but sensitive to colistin. The RAPD-PCR results revealed 35 clusters with 80% similarity. Despite the heterogeneity among the RAPD-PCR profiles, similar patterns were observed among 11 clusters comprising both clinical and environmental isolates. Conclusion: The results of this research suggest that the presence of A. baumannii on environmental surfaces could have played an essential role in the colonization of the hospitalized patients.

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Introduction: Diffusion of antibiotic resistance genes by horizontal gene transfer has led to the fast emergence of multidrug resistance (MDR) among bacteria. Multiple classes of integrons are effective genetic elements which play a significant role in the acquisition and nosocomial dissemination of resistance factors in strains of Gram-negative bacteria, Pseudomonas aeruginosa, and Acinetobacter baumannii. Methods: In this study, 110 sputum samples were collected from hospitalized patients with tract infections. Identification of the isolates was performed by standard biochemical tests. The most frequent Gram-negative isolates were 25 Enterobacteriaceae (62.5%), (9 Enterobacter spp, 11 Citrobacter spp, and 5 Escherichia coli), 6 P. aeruginosa (15%) and 9 Acinetobacter spp (22.5%). Susceptibility of the isolates to antibiotics was carried out by Kirby-Bauer disk diffusion method according to CLSI guidelines, and finally, the class 1 integrons were detected by PCR. Results: Maximum resistance rate among Gram-negative isolates was observed to ceftazidime, co-trimoxazole, and cefotaxime with 89%, 87%, and 82%, respectively. A low-level resistance was recognized for imipenem 32% and gentamicin 34%, while an intermediate level resistance was found against the norfloxacin 40% and ciprofloxacin 44%. Out of 6 P. aeruginosa and 9 A.  baumannii isolates, 2 (33.3%) and 3 isolates (33.3%) were positive for class 1 Integrons, respectively, while all Enterobacteriaceae isolates (100%) were negative for class 1 Integrons. Class 1 integrons were detected among of MDR isolates. Conclusion: Our results showed that monitoring MDR isolates and detection of class 1 integrons in these isolates is necessary for promotion of antibacterial stewardship.

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Introduction: Acinetobacter baumannii (AB) is a Gram-negative bacteria associated with various hospital infections. The present study deals with in silico analysis of the biofilm-associated protein (Bap) in this pathogen. Method: Sixty-eight multi-drug resistant (MDR) AB were isolated from two hospitals in Kerman, Iran. Biofilm-formation was investigated using the microtiter method and PCR followed by sequencing to detect the bap gene in the strongest biofilm-forming isolate. The physicochemical parameters of Bap protein were determined by the ProtParam tool using the ExPasy program. The 3D models from the primary amino acid sequence were constructed using the I-TASSER modeling platform based on multiple-threading alignments by LOMETS. Nevertheless, to ensure the correct initial structure, the protein was minimized in energy through the 3DRefine software of the deep learning system. For the accuracy of predicted models, calculation of the orientation of dihedral angles, including the phi (φ) and psi (ψ) and backbone conformation using the PROCHECK module of the PDB Sum server was performed. The domains and key amino acids involved in protein structure were studied by the Pfam and Interpro softwares. Results: Analysis of the amino acid content of the Bap protein revealed the absence of Arg and Cys in the protein structure. Our Bap protein exhibited ~99.6% identity with other Bap sequences in the GenBank database. Stereochemical simulation identified 19 antiparallel β-sheets with two small α-helices. The N-terminal of Bap protein formed oligomers that mediate cellular adhesion. Conclusion: This study adds considerable information about Bap protein 3D structure, its conformation, domain analysis, and amino acids involved in cellular attachment.

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Introduction: In the past decade, multidrug-resistant Acinetobacter baumannii has become one of the most critical challenges in treating infected patients. The AdeABC efflux pump is the most important among the various resistance mechanisms. This pump can force various antibiotics and ethidium bromide out of the bacterial cell to the surrounding environment. Methods: In this study, nine A. baumannii clinical isolates were isolated and identified using different biochemicals (catalase, oxidase, TSI, hemolysis, growth at 44°C, and indole) and molecular (bla_Oxa51 gene) tests. Following the antibiogram test, the antibiotic resistance changes in the isolates in the presence and absence of efflux pump inhibitor (CCCP) were determined for tetracycline, and ciprofloxacin AdeABC efflux pump genes, including adeA, adeB, and adeC, were amplified by PCR. Finally, the presence of the AdeABC efflux pump was investigated using the agar ethidium-bromide cartwheel method (AEBCM). Results: According to the antibiogram test, all isolates were MDR. In the presence of efflux pump inhibitor, a reduced resistance for tetracycline was observed, but not for ciprofloxacin. The AdeABC efflux pump genes were detected in all isolates. An increase in the AdeABC pump activity in four isolates was confirmed using AEBCM. Conclusion: AEBCM, a fast and convenient tool for assessing the ethidium bromide secretion in various bacteria, provides a quick diagnosis and treatment of multidrug-resistant bacteria.

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Introduction: Acinetobacter baumannii is one of the most important causes of nosocomial infections. In this bacteria, several mechanisms contribute to resistance against antimicrobial agents. The present study investigated the prevalence of adeS and adeH genes and the role of efflux pumps in imipenem and colistin-resistant A. baumannii clinical isolates. Methods: This study included 60 A. baumannii isolates collected from medical centers affiliated with the Shahid Beheshti University of Medical Science, Tehran, Iran. The antibiotic susceptibility pattern was examined using the broth microdilution MIC method according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Also, the adeS and adeH genes were amplified by PCR. Results: The isolates were 100% imipenem-resistant and 86.7% colistin-resistant. All isolates were positive for the 51-blaOXA gene. The adeH and adeS genes were detected in 95% and 80% of the isolates. Conclusion: The high frequency of adeS and adeH efflux pump genes and the high drug resistance in A. baumannii clinical isolates indicated that adeS and adeH efflux pump genes contribute to antibiotic resistance in this species. Therefore, our results provide essential information about high drug resistance in A. baumannii clinical isolates that can help limit the horizontal and vertical transmission of efflux pump genes in antibiotic-resistant A. baumannii isolates that causes nosocomial infections in susceptible strains.

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Introduction: Acinetobacter baumannii is the cause of nosocomial infections, primarily in intensive care units. The pgaA gene plays an essential role in biofilm formation, making it a promising target for developing new strategies to tackle A. baumannii infections. This study investigated the meropenem effect on pgaA gene expression and biofilm formation in A. baumannii. Methods: Over five months, 50 urine samples were taken from patients receiving medical care in the intensive care unit, of which 20 A. baumannii isolates were detected. Antibiotic susceptibility was determined with meropenem, imipenem, trimethoprim/sulfamethoxazole, ceftazidime, ciprofloxacin, tetracycline, amikacin, as well as gentamicin disks by the Kirby-Bauer method. The minimum inhibitory concentration (MIC) of meropenem was determined using the microdilution method. Biofilm formation was investigated through the tissue culture plate (TCP) technique and imaged using an atomic force microscope (AFM). Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) determined the expression level of the pgaA gene. Results: Antibiotic susceptibility testing revealed that all A. baumannii isolates were resistant to meropenem, imipenem, ciprofloxacin, and amikacin, and 71.42% were resistant to tetracycline. The MIC for meropenem could not be determined for isolates. Meropenem prevented biofilm formation in more than 70% of the isolates, and AFM imaging revealed thin biofilms. The RT-PCR showed that exposure to meropenem significantly decreased the pgaA expression gene in over 95% of the isolates (P < 0.0001). Conclusion: Meropenem inhibited biofilm formation in most A. baumannii isolates by downregulating the pgaA expression, suggesting a potential role in preventing A. baumannii infections by reducing biofilm formation

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Introduction: Increased multidrug-resistant (MDR) Acinetobacter baumannii infections pose a significant challenge in hospital settings. Enhanced resistance to antibiotics like fluoroquinolones and β-lactams necessitates adopting alternative treatment strategies such as metal oxide nanoparticles. This study investigated the synergistic effect of zinc oxide nanoparticles (ZnO-NPs) on ciprofloxacin and ceftazidime activity against MDR A. baumannii. Methods: We examined 30 MDR A. baumannii isolates from intensive care unit (ICU) patients in Iran. ZnO-NPs were synthesized via the solvothermal method and characterized using X-ray diffraction (XRD) and field emission scanning electron microscopy (FESEM) to ascertain their crystalline structure and morphology. Antibacterial activity was evaluated by determining minimum inhibitory concentrations (MICs) and inhibition zones through broth microdilution and disk diffusion methods, using concentrations of ciprofloxacin and ceftazidime in combination with ZnO-NPs. Results: ZnO-NPs combined with ciprofloxacin 8 μg/mL and ceftazidime 32 μg/mL exhibited inhibition growth percentage (GI%) increases of 44.9% and 31.65%.  Conclusion: The enhanced in vitro antibacterial effects of combined ZnO-NPs and antibiotics against MDR A. baumannii indicate a synergy. Considering the limited number of isolates, comprehensive research incorporating in vivo models and clinical trials is warranted to evaluate the practicality of this approach in overcoming antibiotic resistance.

 

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Introduction: Secondary bacterial and fungal infections are a significant concern in COVID-19 patients, particularly those critically ill and requiring intensive care. This retrospective study investigated the prevalence and spectrum of secondary infections among COVID-19 patients admitted to the intensive care unit (ICU) at a tertiary care hospital in Navi Mumbai. Additionally, we explored the association between secondary infections and patient comorbidities. Methods: We performed a single-center, retrospective cohort study of 3234 COVID-19 patients admitted to a tertiary care hospital in Navi Mumbai, India, between August 2020 and August 2021. Microbiological data from various clinical specimens, including blood, sputum, bronchoalveolar lavage (BAL) fluid, urine, and tissue cultures, were retrospectively analyzed. Patient demographics and comorbidities were extracted from medical records. We employed descriptive statistics and Pearson's Chi-square test for data analysis to identify associations between secondary infections and patient characteristics. Results: Among the 3234 COVID-19 patients, 195 (6.02%) presented with clinical features suggestive of secondary infections. Microbiological analysis confirmed secondary infections in 98 patients (3.03%), with a culture positivity rate of 50.3%. Among bacterial isolates, Klebsiella pneumoniae was the most prevalent (43.28%), followed by Acinetobacter baumannii (25.37%). Aspergillus spp. emerged as the dominant fungal pathogen. Notably, Escherichia coli isolation was significantly associated with various specimen types (P < 0.001). However, no significant correlation was found between secondary infection rates and patient comorbidities. Conclusion: Gram-negative bacteria, specifically K. pneumoniae and A. baumannii, were the primary pathogens responsible for secondary infections in our cohort of critically ill COVID-19 patients admitted to the ICU. These findings underscore the importance of ongoing surveillance and monitoring of secondary infection trends, including fungal pathogens, to inform and optimize management strategies in this high-risk population.

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Introduction: Non-fermenting Gram-negative bacilli (NFGNB), particularly the Acinetobacter baumannii complex (ABC) and Pseudomonas aeruginosa, are common causes of infections in both hospitalized patients and outpatients, posing significant clinical and therapeutic challenges. The primary objective of this study was to conduct a comprehensive analysis of NFGNB, specifically ABC and P. aeruginosa, isolated from pus specimens obtained from both hospitalized patients and outpatients. Methods: This study investigated the antibiotic resistance patterns of NFGNB, focusing on ABC and P. aeruginosa, isolated from pus samples collected from both hospitalized patients and outpatients. The isolates were tested for multidrug resistance (MDR) and extensive drug resistance (XDR) using standardized microbiological protocols. The data were analyzed using descriptive statistics to summarize the findings. Results: Out of 1234 pus samples received, 117 (9.5%) NFGNB were isolated, accounting for 30% of the total Gram-negative bacilli (GNB) isolates. The majority of NFGNB (82.9%, n = 97/117) were isolated from inpatients, with surgical site infections being the most common clinical condition (33.3%, n = 39/117). Among the NFGNB isolates, P. aeruginosa was the predominant species (76.9%, n = 90/117), followed by A. baumannii (22.2%, n = 26/117). Antimicrobial susceptibility testing revealed that 37.7% (n = 34/90) of P. aeruginosa isolates were MDR and 13% (n = 12/90) were XDR, while 65% (n = 17/26) of A. baumannii isolates were MDR and 26.9% (n = 7/26) were XDR. Conclusion: This study highlights the emergence of NFGNB as significant nosocomial pathogens, exhibiting a high degree of resistance to commonly used antibiotics. The findings underscore the urgent need to enhance and strictly implement effective antibiotic stewardship policies, including the development of new antibiotic regimens and antimicrobial resistance surveillance programs, to combat the growing resistance of nosocomial pathogens and ultimately improve patient outcomes.

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Introduction: Acinetobacter baumannii is notorious for its high resistance levels, and the development of clinically effective antimicrobial agents is an urgent medical challenge. Single-chain variable fragments (scFvs) that exhibit antibacterial properties against challenging pathogens, such as Pseudomonas aeruginosa and Staphylococcus aureus, have the potential to improve therapeutic strategies significantly. Their unique ability to function independently of the host immune system makes scFvs a highly promising option for effective treatment. In our previous studies, we identified two human scFvs (EB211 and EB279) that showed direct growth inhibition activity against A. baumannii strains in vitro and therapeutic effectiveness in immunocompromised mice with pneumonia caused by an extensively drug-resistant A. baumannii strain. In the present study, we endeavored to demonstrate how EB211 and EB279 could inhibit the growth of A. baumannii. Methods: A. baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa strains were individually incubated with the scFv in the presence of a high concentration of magnesium (MgSO₄; 20 mM). Epitope mapping and immunoblotting were conducted to identify A. baumannii proteins likely bound by EB211 and EB279. Results: It was found that EB211 and EB279, similar to colistin sulfate, lost their activity in the presence of magnesium. Moreover, immunoblotting revealed that EB211 and EB279 might bind the OprD family outer membrane porin and TonB family C-terminal domain protein, respectively. Conclusion: EB211 and EB279 elicit direct growth inhibitory activity against A. baumannii without needing immune cells or complements, which could be helpful for immunocompromised patients.