Effect of Mentha longifolia essential oil on oqxA efflux pump gene expression and biofilm formation in ciprofloxacin-resistant Klebsiella pneumoniae strains.

Background and Objectives: Today, medicinal plants and their derivatives are considered to reduce the prevalence of antibiotic resistance. The aim of this study was to investigate the effect of Mentha longifolia essential oil on oqxA efflux pump gene expression and biofilm formation in ciprofloxacin-resistant Klebsiella pneumoniae strains. Materials and Methods: A total of 50 clinical strains of K. pneumoniae resistant to ciprofloxacin were studied. The minimum inhibitory concentration (MIC) of M. longifolia essential oil and its synergistic effect with ciprofloxacin were determined using the microbroth dilution method and the fractional inhibitory concentration (FIC) method. Minimum biofilm inhibition concentration (MBIC) of M. longifolia essential oil was detected. The effect of essential oils on the expression level of the oqxA gene was detected by Real-time PCR. Results: M. longifolia essential oil showed inhibitory activity against ciprofloxacin-resistant strains of K. pneumoniae. When M. longifolia essential oil was combined with ciprofloxacin, the MIC was reduced 2-4 times. In 28% of the strains, M. longifolia with ciprofloxacin showed a synergistic effect. M. longifolia essential oil reduces the strength of biofilm formation and alters the biofilm phenotype. A significant decrease in oqxA gene expression was observed in all isolates after treatment with M. longifolia essential oil. Conclusion: Based on the results of this study, it was observed that supplementing M. longifolia essential oil can help reduce ciprofloxacin resistance and inhibit biofilm formation in fluoroquinolone-resistant K. pneumoniae strains.

Enhancing antimicrobial efficacy against Pseudomonas aeruginosa biofilm through carbon dot-mediated photodynamic inactivation.

Pseudomonas aeruginosa biofilms shield the bacteria from antibiotics and the body's defenses, often leading to chronic infections that are challenging to treat. This study aimed to assess the impact of sub-lethal doses of antimicrobial photodynamic inactivation (sAPDI) utilizing carbon dots (CDs) derived from gentamicin and imipenem on biofilm formation and the expression of genes (pelA and pslA) associated with P. aeruginosa biofilm formation.The anti-biofilm effects of sAPDI were evaluated by exposing P. aeruginosa to sub-minimum biofilm inhibitory concentrations (sub-MBIC) of CDsGEN-NH2, CDsIMP-NH2, CDsGEN-IMP, and CDsIMP-GEN, combined with sub-lethal UVA light irradiation. Biofilm formation ability was assessed by crystal violet (CV) assay and enumeration method. Additionally, the impact of sAPDI on the expression of pelF and pslA genes was evaluated using real-time quantitative polymerase chain reaction (RT-qPCR).Compared to the control group, the sAPDI treatment with CDsGEN-NH2, CDsIMP-NH2, CDsGEN-IMP, and CDsIMP-GEN resulted in a significant reduction in biofilm activity of P. aeruginosa ATCC 27853 (P < 0.0001). The CV assay method demonstrated reductions in optical density of 83.70%, 81.08%, 89.33%, and 75.71%, while the CFU counting method showed reductions of 4.03, 3.76, 4.39, and 3.21 Log10 CFU/mL. qRT-PCR analysis revealed decreased expression of the pelA and pslA genes in P. aeruginosa ATCC 27853 following sAPDI treatment compared to the control group (P < 0.05).The results indicate that sAPDI using CDs derived from gentamicin and imipenem can decrease the biofilm formation of P. aeruginosa and the expression of the pelA and pslA genes associated with its biofilm formation.

Comparison of toxin gene expression levels and molecular typing of Clostridioides difficile strains isolated from patients with diarrhea.

Aim: This study aimed to evaluate the expression of tcdA, tcdB, and binary toxin genes (cdtA and cdtB) by Real-Time PCR and molecular typing of Clostridioides difficile isolated from patient diarrhea samples from Hamadan Hospitals, west of Iran. Background: The concentration of C. difficile toxins (CDTs) is associated with the severity of the disease and the mortality rate. Measuring CDT levels could provide a reliable and objective means of determining the severity of C. difficile infection (CDI). Methods: From November 2018 to September 2019, 130 diarrhea samples were collected from hospitalized patients in three hospitals in Hamadan. C. difficle isolates were detected by culture and PCR. The presence of the genes encoding the toxin was identified by PCR, whereas the measurement of toxin expression was conducted using a relative Real-Time PCR technique. Genetic linkage of the isolates was also assessed by Ribotyping and Repetitive Extragenic Palindromic (rep-PCR) methods. Results: Among 130 diarrhea samples, 16 (12.3%) were positive for C. difficile. Genes encoding cdtA and tcdB were detected in all isolates, and 8 (50%) and 6 (37.5%) isolates were positive for the cdtA and cdtB genes. Real-time PCR results showed different expression levels of the toxin genes. A significant increase in the expression of the tcdA gene was observed compared with the control strain (P<0.05). Besides, more expression of cdtA gene was observed in the strains compared with cdtB gene. Ribotyping and rep-PCR results showed high genetic diversity of C. difficile among hospitals investigated. Conclusion: We encountered toxigenic C. difficile strains with various toxin expression levels, ribotypes, and rep types based on the findings of this study. This indicated that various clones from various sources circulate in the hospitals and among patients.

Vancomycin and nisin-modified magnetic Fe3O4@SiO2 nanostructures coated with chitosan to enhance antibacterial efficiency against methicillin resistant Staphylococcus aureus (MRSA) infection in a murine superficial wound model.

BACKGROUND: The objective of this research was to prepare some Fe3O4@SiO2@Chitosan (CS) magnetic nanocomposites coupled with nisin, and vancomycin to evaluate their antibacterial efficacy under both in vitro and in vivo against the methicillin-resistant Staphylococcus. aureus (MRSA). METHODS: In this survey, the Fe3O4@SiO2 magnetic nanoparticles (MNPs) were constructed as a core and covered the surface of MNPs via crosslinking CS by glutaraldehyde as a shell, then functionalized with vancomycin and nisin to enhance the inhibitory effects of nanoparticles (NPs). X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FT-IR), field emission scanning electron microscope (FE-SEM), vibrating sample magnetometer (VSM), and dynamic light scattering (DLS) techniques were then used to describe the nanostructures. RESULTS: Based on the XRD, and FE-SEM findings, the average size of the modified magnetic nanomaterials were estimated to be around 22-35 nm, and 34-47 nm, respectively. The vancomycin was conjugated in three polymer-drug ratios; 1:1, 2:1 and 3:1, with the percentages of 45.52%, 35.68%, and 24.4%, respectively. The polymer/drug ratio of 1:1 exhibited the slowest release rate of vancomycin from the Fe3O4@SiO2@CS-VANCO nanocomposites during 24 h, which was selected to examine their antimicrobial effects under in vivo conditions. The nisin was grafted onto the nanocomposites at around 73.2-87.2%. All the compounds resulted in a marked reduction in the bacterial burden (P-value < 0.05). CONCLUSION: The vancomycin-functionalized nanocomposites exhibited to be more efficient in eradicating the bacterial cells both in vitro and in vivo. These findings introduce a novel bacteriocin-metallic nanocomposite that can suppress the normal bacterial function on demand for the treatment of MRSA skin infections.

Evaluation of antibiotic resistance, toxin-antitoxin systems, virulence factors, biofilm-forming strength and genetic linkage of Escherichia coli strains isolated from bloodstream infections of leukemia patients.

BACKGROUND: One of the most common complications in patients with febrile neutropenia, lymphoma, leukemia, and multiple myeloma is a bloodstream infection (BSI). OBJECTIVE: This study aimed to evaluate the antibiotic resistance patterns, virulence factors, biofilm-forming strength, and genetic linkage of Escherichia coli strains isolated from bloodstream infections (BSIs) of leukemia patients. METHODS: The study conducted in Iran from June 2021 to December 2022, isolated 67 E. coli strains from leukemia patients' bloodstream infections in hospitals in two different areas. Several techniques including disk diffusion and broth microdilution were used to identify patterns of antibiotic resistance, microtiter plate assay to measure biofilm formation, and PCR to evaluate the prevalence of different genes such as virulence factors, toxin-antitoxin systems, resistance to ß-lactams and fluoroquinolone antibiotics of E. coli strains. Additionally, the genetic linkage of the isolates was analyzed using the Enterobacterial Repeat Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) method. RESULTS: The results showed that higher frequency of BSI caused by E. coli in man than female patients, and patients with acute leukemia had a higher frequency of BSI. Ampicillin and Amoxicillin-clavulanic acid showed the highest resistance, while Imipenem was identified as a suitable antibiotic for treating BSIs by E. coli. Multidrug-resistant (MDR) phenotypes were present in 22% of the isolates, while 53% of the isolates were ESBL-producing with the blaCTX-M gene as the most frequent ß-lactamase gene. The fluoroquinolone resistance genes qnrB and qnrS were present in 50% and 28% of the isolates, respectively. More than 80% of the isolates showed the ability to form biofilms. The traT gene was more frequent than other virulence genes. The toxin-antitoxin system genes (mazF, ccdAB, and relB) showed a comparable frequency. The genetic diversity was detected in E. coli isolates. CONCLUSION: Our results demonstrate that highly diverse, resistant and pathogenic E. coli clones are circulating among leukemia patients in Iranian hospitals. More attention should be paid to the treatment and management of E. coli bloodstream infections in patients with leukemia.

Co-delivery of streptomycin and hydroxychloroquine by labeled solid lipid nanoparticles to treat brucellosis: an animal study.

Can brucellosis-related biochemical and immunological parameters be used as diagnostic and treatment indicators? The goal of this project was to look at biochemical parameters, trace elements, and inflammatory factors in the acute and chronic stages of brucellosis after treatment with streptomycin and hydroxychloroquine-loaded solid lipid nanoparticles (STR-HCQ-SLN). The double emulsion method was used for the synthesis of nanoparticles. Serum levels of trace elements, vitamin D, CRP, and biochemical parameters were measured in rats involved in brucellosis. The therapeutic effect of STR-HCQ-SLN was compared with that of free drugs. In both healthy and infected rats, serum concentrations of copper, zinc, iron, magnesium, potassium, and biochemical parameters of the liver were significantly different. By altering the serum levels of the aforementioned factors, treatment with STR-HCQ-SLN had a positive therapeutic effect on chronic brucellosis. Vitamin D levels declined (46.4%) and CRP levels rose (from 7.5 mg to less than 1 mg) throughout the acute and chronic stages of brucellosis. This study showed that by comparing the biochemical parameters and the levels of trace elements in the serum of healthy and diseased mice in the acute and chronic stages of brucellosis, it is possible to get help from other routine methods for diagnosis.

Molecular Epidemiology of Stenotrophomonas maltophilia Strains Isolated from Bacteremia in Hospitalized Children.

Objectives: The aim of this study was to determine the phenotypic and genotypic characteristics of Stenotrophomonas maltophilia isolates obtained from blood culture samples of pediatric patients hospitalized in Borujerd and Hamadan hospitals in western Iran. Methods: Oxidase-negative isolates were collected from the blood cultures of pediatric patients. S. maltophilia isolates were identified and confirmed by routine microbiological and molecular testing. Antibiotic susceptibility of the isolates was determined. The phenotypic and genotypic biofilm-forming ability of the isolates were investigated. Molecular typing of all isolates was performed by repetitive element sequence-based polymerase chain reaction. Results: Out of 450 oxidase-negative bacilli, 72 (16.0%) were identified as S. maltophilia isolates. Biofilm assay results showed strong biofilm formation in 19 (26.4%) isolates, moderate in 38 (52.8%), weak in 10 (13.9%), and no biofilm formation in five (6.9%) isolates. Biofilm-associated genes rmlA, rpfF, and spgM were detected respectively in 59 (81.9%), 54 (75.0%), and 72 (100%) of isolates. Antimicrobial susceptibility testing showed that 67 (93.1%) isolates were sensitive to trimethoprim-sulfamethoxazole. All isolates were sensitive to levofloxacin and resistant to ceftazidime. The S. maltophilia isolates were grouped into 14 different types of repetitive sequence by repetitive element sequence-based polymerase chain reaction analysis. Conclusions: The results of this study indicate that S. maltophilia should be considered an important opportunistic pathogen in pediatric units. Different genotypes of S. maltophilia with the ability to form a biofilm (an important virulence factor) were circulating in the hospitals investigated. Levofloxacin and trimethoprim-sulfamethoxazole are recommended to treat S. maltophilia infections.

Association of qnr Genes and OqxAB Efflux Pump in Fluoroquinolone-Resistant Klebsiella pneumoniae Strains.

Background: The aim of this study was to investigate the frequency and relationship between plasmid-mediated quinolone resistance genes and OqxAB pump genes, as well as the genetic linkage in K. pneumoniae strains isolated from Hamadan hospitals in the west of Iran. Materials and Methods: In this study, 100 K. pneumoniae clinical strains were isolated from clinical samples of inpatients at Hamadan Hospital in 2021. The antimicrobial susceptibility testing was performed using the disk diffusion method. The frequencies of genes encoding OqxAB efflux pumps and qnr were investigated by PCR. Molecular typing of qnr-positive K. pneumoniae isolates was assessed by ERIC-PCR. Results: Antibiotic susceptibility testing showed high resistance (>80%) to fluoroquinolones. The gene encoding the OqxAB efflux pump was detected in more than 90% of K. pneumomiae strains. All K. pneumoniae isolates were negative for qnrA, and 20% and 9% of the isolates were positive for qnrB and qnrS, respectively. The genes encoding oqxA and oqxB were detected in 96% of qnr-positive strains. A qnrB + /qnrS + profile was observed in 16% of qnr-positive K. pneumoniae strains. Ciprofloxacin MIC ≥ 256 µg/ml was detected in 20% of qnr-positive strains. Genetic association analysis by ERIC-PCR revealed genetic diversity among 25 different qnr-positive strains of K. pneumonia. Conclusion: However, no significant correlation was found between the qnr and the OqxAB efflux pump genes in this study. The high rate of fluoroquinolone resistance and determinants of antibiotic resistance among diverse K. pneumoniae strains increase the risk of fluoroquinolone-resistance transmission by K. pneumoniae strains in hospitals.

Relationship of OqxAB efflux pump to antibiotic resistance, mainly fluoroquinolones in Klebsiella pneumoniae, isolated from hospitalized patients.

Objectives: This research was designed to study the prevalence of OqxAB efflux pump genes and also to investigate the relationship between efflux pump and resistance to antibiotics, especially to fluoroquinolones, evaluate the expression levels of OqxAB genes, and molecular typing of Klebsiella pneumoniae isolated from hospitalized patients in Hamadan hospitals, west of Iran. Materials and Methods: In a cross-sectional study, 100 clinical strains of K. pneumoniae were isolated from hospitalized patients in three major teaching hospitals from January to June 2021. The antibiotic susceptibility of isolates was evaluated by the disk-diffusion agar method. The frequency of genes encoding oqxA and oqxB of efflux pump genes was investigated by PCR, and the expression of the oqxA efflux pump gene was investigated by the Real-time PCR method. The genetic relationship of K. pneumoniae isolates was analyzed by the Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR technique. Results: According to our results, the multi-drug resistance phenotype (MDR) in 65% and high prevalence resistance to ciprofloxacin in 89% of K. pneumoniae isolates was detected. The higher prevalence of oqxA (95%) and oqxB (98%) was also detected. There was a significant relationship between ciprofloxacin resistance and the oqxB gene as well as between ceftriaxone and chloramphenicol resistance and the oqxA gene. The expression of the oqxA gene was higher in ciprofloxacin-resistant isolates. Conclusion: The results of this study suggest a potential reservoir for the spread of OqxAB genes among hospital-acquired bacteria. Infection control strategies should be used prudently to reduce the spread of resistant strains of K. pneumoniae in hospitals.

Use of the quantum dot-labeled solid lipid nanoparticles for delivery of streptomycin and hydroxychloroquine: A new therapeutic approach for treatment of intracellular Brucella abortus infection.

Brucellosis is considered one of the most important infectious diseases affecting any tissue and organ in the human body. Due to the intracellular pathogenesis of Brucella species, the use of conventional antibiotics for managing chronic brucellosis has several limitations. Therefore, the study focused on the use of solid lipid nanoparticles (SLN) to deliver streptomycin (STR) for intracellular infection, with or without the combination of hydroxychloroquine (HCQ) to evaluate if there might be a boost in the antibiotic effect when using the STR or STR-NPs alone. We used the double emulsion technique to synthesize Nano drug carriers; afterward, the physicochemical characteristics of synthesized Nano drug carriers were determined. The in vitro antibacterial activity of free drugs and Nano drug carriers were evaluated using well diffusion, broth microdilution assays (BMD), and murine macrophage-like cells cell line J774A.1. Additionally, acute and chronic phases of brucellosis were inducted into Wistar rats, and healing capacity of Nano drug carriers on liver and spleen tissues was compared with free drugs. The zeta potential of nanoparticles, means of size, Polydispersity Index (PDI), drugs loading, and encapsulation efficiency were 15.2 mV, 312.5 ± 26 nm, 0.433 ± 0.09, 16.6% and 89.5%, respectively. Well diffusion and BMD methods did not show a significantly differ between free drugs and nano drug carriers. However, the Nano drug carriers remarkably decreased the number of bacteria in the cell line compared to the free drugs. STR/HCQ-SLN enhanced the healing processes of the liver and spleen after brucellosis induction. STR/HCQ-SLN showed better inhibitory effects against the chronic phase of B. abortus infection in comparison to the STR-SLN, but this difference was not statistically significant. Using nanoplatforms to enhance conventional anti-brucellosis agents is promising, green and safe. Due to the continuous release of drugs, drugs increase their accumulation at the site of infection, causing a more significant effect on the chronic and acute phases of brucellosis.

Characterization of Staphylococcus aureus isolates from pastry samples by rep-PCR and phage typing.

Background and Objectives: Staphylococcus aureus is one of the most common causes of food poisoning. This study aimed to identify S. aureus isolated from pastries, the virulence factors, antimicrobial resistance patterns, biofilm formation, and then classification based on SCCmec types, phage types, and also Rep types. Materials and Methods: In this study, 370 creamy and dried pastry samples have been randomly collected from different confectioneries in Hamadan city. The S. aureus isolates were identified by conventional microbiological methods and nuc gene amplification. The virulence factors and prophage genes were detected. After that, the biofilm production and antibiotic susceptibility assay of S. aureus isolates were examined. Finally, the isolates were classified by rep-PCR typing. Results: Among 370 samples, 97 creamy (34.64%) and 3 dried (3.33%) pastry samples were contaminated with S. aureus. Antibiotic sensitivity results showed the highest resistance to penicillin (90%) but none of them were MRSA. According to biofilm formation assay, 14 strains (45%) were strongly adhesive. The dominant phage among isolates was SGF, especially SGFa subgroup. About half of the isolates carried SCCmec Types I and III. Analysis of the genetic linkage between isolates by rep-PCR showed ≥80% genetic similarity and also different rep-types of S. aureus isolates. Conclusion: The presence of different prophage encoded virulence factors and antibiotic resistance enable S. aureus strains to produce a broad range of diseases. Thus, consumption of creamy pastries increases the risk of infection with S. aureus and it is a serious warning to the health system.

Genotyping of Chlamydia abortus using multiple loci variable number of tandem repeats analysis technique.

BACKGROUND: The correlation between various factors (geographical region, clinical incidence, and host type) and the genomic heterogeneity has been shown in several bacterial strains including Chlamydia abortus. METHODS: The aim of this study was to survey the predominant types of C. abortus strains isolated from ruminants in Iran by the multiple loci variable number of tandem repeats (VNTR) analysis (MLVA) method. C. abortus infection was evaluated in a total of 117 aborted fetuses by real-time PCR. The isolation was done via the inoculation of the positive samples in chicken embryo and the L929 cell line. Genotyping was carried out by MLVA typing technique. RESULTS: Forty samples (34.2%) were detected with C. abortus infection; however, chlamydial infection in ruminants of Charmahal/Bakhtiari (3 bovines and 35 sheep) was higher than that of Khuzestan (2 sheep). All MLVA types (MT1-MT8) were detected in the collected samples from Charmahal/Bakhtiari but only 2 types (MT1 and MT3) were reported in samples from Khuzestan. The main MT type was MT1 (32% of aborted fetuses). Although in this study only 9 cow samples were investigated, they possessed similar clusters to those obtained from sheep (MT1 and MT6). Variation of type in sheep samples (MT1 to MT8) was more than that of bovine samples (MT1, and MT6). CONCLUSION: By this research revealed that C.abortus was responsible for a significant percentage of ruminant abortion in two studied regions. The main MT type was MT1 (32% of aborted fetuses) and also 7 different genotypes were involved in infections. So it is concluded that diversity in C.abortus genotyping is high in two regions.

Prevalence of Shiga Toxin-Producing Escherichia coli O157 and Non-O157 Serogroups Isolated from Fresh Raw Beef Meat Samples in an Industrial Slaughterhouse.

BACKGROUND: The aims of the current study are the identification of O157 and non-O157 Shiga Toxin-Producing Escherichia coli (STEC) serogroups isolated from fresh raw beef meat samples in an industrial slaughterhouse, determination of antimicrobial resistance patterns, and genetic linkage of STEC isolates. MATERIALS AND METHODS: A total of 110 beef samples were collected from the depth of the rump of cattle slaughtered at Hamadan industrial slaughterhouse. After detection of E. coli isolates, STEC strains were identified according to PCR for stx1, stx2, eaeA, and hlyA virulence genes, and STEC serogroups (O157 and non-O157) were identified by PCR. The genetic linkage of STEC isolates was analyzed by the ERIC- (Enterobacterial Repetitive Intergenic Consensus-) PCR method. The antimicrobial susceptibility of STEC isolates was detected by the disk diffusion method according to CLSI guidelines. RESULTS: Among 110 collected beef samples, 77 (70%) were positive for E. coli. The prevalence of STEC in E. coli isolates was 8 (10.4%). The overall prevalence of O157 and non-O157 STEC isolates was 12.5% (one isolate) and 87.5% (7 isolates), respectively. The hemolysin gene was detected in 25% (2 isolates) of STEC strains. Evaluation of antibiotic resistance indicated that 100% of STEC isolates were resistant to ampicillin, ampicillin-sulbactam, amoxicillin-clavulanic acid, and cefazolin. Resistance to tetracycline and ciprofloxacin was detected in 62.5% and 12.5% of isolates, respectively. The analysis of the ERIC-PCR results showed five different ERIC types among the STEC isolates. CONCLUSION: The isolation of different clones STECs from beef and the presence of antibiotic-resistant isolates indicate that more attention should be paid to the hygiene of slaughterhouses.

Investigation of Antibiotic Resistance and Biofilm Formation in Clinical Isolates of Klebsiella pneumoniae.

AIM: Klebsiella pneumoniae (K. pneumoniae) is an encapsulated Gram-negative bacterium that can lead to 14-20% of nosocomial infections. The ability of biofilm formation in this bacterium decreases the host immune response and antibiotic efficacy. This may impose a huge impact on patients and healthcare settings. This study aimed to evaluate the antibiotic resistance pattern and biofilm formation in K. pneumoniae strains isolated from two major Hamadan hospitals, west of Iran. METHODS: A total of 83 K. pneumoniae strains were isolated from clinical samples of patients in different wards of Hamadan hospitals from September 2018 to March 2019. Determination of antimicrobial susceptibility was performed using the disk diffusion method. Biofilm formation was evaluated by the crystal violet method. Data were analyzed by the SPSS software and chi-square test. RESULTS: The results showed that clinical samples included 18 urinary tract samples (22%), 6 wound samples (7%), 6 blood samples (7%), 17 tracheal tube aspiration samples (20%), 32 throat cultures (38%), 2 sputum samples (2.5%), and 2 abscess drain cultures (2.5%). High-level resistance to cefotaxime was detected in 92%, and all of isolates were susceptible to colistin. Biofilm formation was seen in 62 (75%) isolates. Strong biofilm formation was observed in 17 (20%) strains. A significant correlation was seen between biofilm formation and antibiotic resistance (P value <0.05). CONCLUSION: Our findings emphasize the need for proper diagnosis, control, and treatment of infections caused by K. pneumoniae especially in respiratory tract infections due to the strong biofilm formation and high antibiotic resistance in these strains.

Molecular typing, biofilm production, and detection of carbapenemase genes in multidrug-resistant Acinetobacter baumannii isolated from different infection sites using ERIC-PCR in Hamadan, west of Iran.

BACKGROUND: Acinetobacter baumannii is an opportunistic pathogen that can cause several kinds of nosocomial infections. Increasing antibiotic resistance as well as identifying genetic diversity and factors associated with pathogenicity and prevalence of this bacterium is important. The aim of this study was the investigation of molecular typing, biofilm production, and detection of carbapenemase genes in multidrug-resistant Acinetobacter baumannii isolated from different infection sites using ERIC-PCR in Iran. METHODS: Forty isolates of A. baumannii were obtained from various wards of the central hospital, in the west of Iran. Phenotypic identification and genetic diversity, biofilm production assay, and detection of Carbapenemase genes carried out. RESULTS: Tracheal samples 26 (61.9 %) are the most frequent isolates, and 95 % of isolates were identified as MDR. 32.5 % of all A. baumannii strains were capable to form a strong biofilm. It was founded that antimicrobial resistance patterns had a significant relationship with strong biofilm formation (P = 0.001). Most frequencies of the studied genes were in the order of VIM (81 %), SPM (45.2 %), and IMP (35.7 %) genes. The VIM gene was the most frequent in all isolates which were significant (P = 0.006). 14 different ERIC-types were observed including 7 common types and 7 unique or single types. F type is the largest common type consisting of nine isolates and B, D, and E types contain two isolates separately. CONCLUSIONS: ERIC-PCR technique was used to genetically classify A. baumannii isolates as one of the most common microorganisms in nosocomial infections.

Frequency of toxin genes and antibiotic resistance pattern of Clostridioides difficile isolates in diarrheal samples among hospitalized patients in Hamadan, Iran.

AIM: This study was designed to investigate the prevalence of Clostridioides difficile, its toxin-producing genes, and antibiotic resistance patterns in diarrheal samples from hospitalized patients in Hamadan, Iran. BACKGROUND: Today, concerns over Clostridioides difficile infection (CDI) have significantly increased due to reduced susceptibility to antibiotics used for CDI treatment. Toxins produced by C. difficile strains are associated with disease severity and outcome. METHODS: In this cross-sectional study, a total of 130 diarrheal samples of patients admitted to different wards of three hospitals in Hamadan from November 2018 to September 2019 were collected. C. difficile isolates were identified by culture on CCFA and PCR (Polymerase chain reaction). The presence of toxin-encoding genes (tcdA and tcdB) and binary toxin genes (cdtA and cdtB) was analyzed by PCR. Resistance of the isolates to metronidazole, vancomycin and clindamycin antibiotics was determined using agar dilution method. RESULTS: Out of 130 diarrheal samples from hospitalized patients, 16 (12.3%) C. difficile isolates were obtained. PCR results were positive for two toxin-producing genes, tcdA and tcdB, in all (100%) C. difficile isolates, and the binary toxin genes cdtA and cdtB were detected in 6 (37.5%) and 8 (50%) isolates, respectively. The results of antibiotic susceptibility testing showed resistance to metronidazole, vancomycin, and clindamycin in 3 (18.7%), 3 (18.7%), and 2 (12.5%) isolates, respectively, and all isolates were resistant to rifampicin. CONCLUSION: The results of this study showed toxigenic C. difficile with tcdA + /tcdB + profile is a major cause of nosocomial diarrhea in Hamadan, and clinical laboratories should routinely perform C. difficile diagnostic testing on diarrheal specimens of hospitalized patients. Resistance to conventional antibiotic therapy against C. difficile should be considered as a warning to prevent irrational administration of antibiotics.

Genetic diversity of colistin resistance Nosocomial Acinetobacter baumannii strains from Iran.

BACKGROUND: Drug-resistant Acinetobacter baumannii is a global health problem since its ability to acquire new resistance mechanisms. Here, we aimed to determine the association of common types of A. baumannii and assess their drug resistance of A. baumannii and contribution of integrons (Ints) and oxacillinase genes in Zanjan, Iran. MATERIALS AND METHODS: Among 68 isolated Acinetobacters from patients, 48 isolates were A. baumannii strains. Antibiotic susceptibility pattern and colistin resistance were determined by disk diffusion and broth microdilution, respectively. The presence of Int I, II, III, and oxacillinase genes examined using polymerase chain reaction. The clonal relationship of clinical isolates of A. baumannii determined by Pulsed Field Gel Electrophoresis method. RESULTS: The results showed the highest antibiotic susceptibility (58%) for colistin. 96% of isolates were considered as multidrug resistant, and 46% as extensively drug resistant, and 16% as pandrug resistant. Frequencies of Int I, II, III resistance genes were 60%, 28%, and 0%, respectively, and 12% of strains had no isoform of Ints. Frequencies of Carbapenem resistance genes were 74%, 24%, 100%, and 4% for blaOXA-23, blaOXA-24, blaOXA-51, and blaOXA-58, respectively. The above samples were group into 26 pulsotypes. CONCLUSION: The studied A. baumannii strains had several resistance genes, and the colistin resistance showed an extraordinary ascending tendency that could be a severe issue in nosocomial infections, and the presence of high genetic diversity indicated a variation in A. baumannii strains and possibly a variety of sources of contamination or infection.

The Prevalence of Shiga Toxin-Producing Escherichia coli and Enteropathogenic Escherichia coli Isolated from Raw Chicken Meat Samples.

BACKGROUND: Shiga toxin-producing Escherichia coli (STEC) is known as a crucial zoonotic food-borne pathogen. A total of 257 raw chicken meat samples were collected from different markets in Hamadan, west of Iran, from January 2016 to May 2017. MATERIALS AND METHODS: The samples were cultured in selective and differential culture media, and the virulence genes of E. coli isolates were analyzed by PCR assay. The antibiotic resistance patterns of E. coli isolates were determined by the disk diffusion method. The genetic relatedness of the E. coli O157 isolates was analyzed by ERIC-PCR. RESULTS: In total, 93 (36% ± 3.12) of the isolates were identified as E. coli in this study. Based on serological and microbiological tests, 36 (38.7% ± 9.9), 7 (7.5% ± 5.35), and 12 (12.9% ± 6.81) of the E. coli isolates were characterized as STEC, enteropathogenic E. coli (EPEC), and attaching and effacing E. coli (AEEC) strains, respectively. A high level of resistance to nalidixic acid (91.4% ± 5.7), tetracycline (89.2% ± 6.31), ampicillin (82.8% ± 7.67), and trimotoprime-sulfametoxazole (71% ± 9.22) was detected among the E. coli isolates. The analysis of the ERIC-PCR results showed five different ERIC types among the E. coli O157 isolates. CONCLUSIONS: Based on our findings, control and check-up of poultry meats should be considered as a crucial issue for public health.

Emerge of NDM-1-Producing Multidrug-Resistant Pseudomonas aeruginosa and Co-Harboring of Carbapenemase Genes in South of Iran.

BACKGROUND: New Delhi metallo-beta-lactamase-1 (NDM-1) is one of the most important emerging antibiotic resistance. Co-harboring three or four carbapenemases is rare and only a few reports exist in the literature. We described the characteristics of the large epidemic outbreaks and reports co-producing blaNDM-1 with the other carbapenemase genes in P. aeruginosa isolates. METHODS: This present cross-sectional research was conducted on 369 P. aeruginosa isolates obtained from burn and general hospitals within years 2013 to 2016. Beta-lactamase classes A, B and D genes were identified by PCR method. Modified hodge test (MHT), double-disk potentiation tests (DDPT) and double disk synergy test (DDST) were performed for detection carbapenemase and metallo beta-lactamase (MBL) production of blaNDM-1 positive P. aeruginos isolates. RESULTS: From 236 carbapenem-resistant P. aeruginosa (CRPA), 116 isolates have had MBL genes and twenty-nine isolates were found positive for blaNDM-1 . In CRPA isolates, blaIMP-1 , blaVIM-2 and blaOXA-10 were identified in 27.5%, 21.1% and 32.2% of isolates respectively, while co-producing blaNDM-1 , blaIMP-1 , blaOXA-10 , co-producing blaNDM-1 , blaVIM-2 , blaOXA-10 and co-producing blaIMP-1 , blaVIM-2 were determined in 11 (4.6%), 8 (3.4%) and 27 (11.4%) of isolates respectively. CONCLUSION: The finding of this co-existence of multiple carbapenemase resistance genes is threating for public health. Dipicolinic acid is a superior MBL inhibitor in DDPT antique than EDTA in DDST method for the detection of MBL-blaNDM-1 producing P. aeruginosa.

Detection of adeABC efllux pump encoding genes and antimicrobial effect of Mentha longifolia and Menthol on MICs of imipenem and ciprofloxacin in clinical isolates of Acinetobacter baumannii.

BACKGROUND: Acinetobacter baumannii is an opportunistic pathogen that causes nosocomial infections especially in patients in intensive care units (ICUs). Accordingly, the aim of our study was to detection of adeABC efllux pump encoding genes and antimicrobial effect of the essential oil of Mentha longifolia and Menthol on the minimum inhibitory concentration (MIC) of imipenem and ciprofloxacin in clinical isolates of A. baumannii. METHODS: A total of 75 clinical isolates of A. baumannii were collected. The presence of efflux pump genes was detected by polymerase chain reaction (PCR). The minimum inhibitory concentration (MIC) of the essential oil of Mentha longifolia and Menthol and their combined effect with antibiotics were measured by microbroth dilution method and fractional inhibitory concentration (FIC) index. RESULTS: The frequency of adeA, adeB, and adeC genes in clinical isolates of A. baumannii were 86.7, 90.7, and 92%, respectively. When the essential oil of Mentha longifolia was combined with ciprofloxacin and imipenem, MICs decreased 4- and 8-fold, respectively. In the combination of menthol with imipenem, the resistance to imipenem was reduced from 0- to 16-fold in 90% (63/70) of the isolates. CONCLUSION: The presence of efflux pump genes in more than 90% of A. baumannii isolates indicates its potential role in inducing imipenem- and ciprofloxacin-resistance in this bacterium. Menthol has an antimicrobial effect as an active ingredient in Mentha longifolia. In the future, the combination of medicinal plants with antibiotics can be used as a complement in treating diseases caused by drug-resistant bacteria such as A. baumannii infections.