A novel Enterococcus faecium phage EF-M80: unveiling the effects of hydrogel-encapsulated phage on wound infection healing.

Background: Enterococcus faecium is one of the members of ESKAPE pathogens. Due to its resistance to antimicrobial agents, treating this bacterium has become challenging. The development of innovative approaches to combat antibiotic resistance is necessary. Phage therapy has emerged as a promising method for curing antibiotic-resistant bacteria. Methods: In this study, E. faecium phages were isolated from wastewater. Phage properties were characterized through in vitro assays (e.g. morphological studies, and physicochemical properties). In addition, whole genome sequencing was performed. A hydrogel-based encapsulated phage was obtained and its structure characteristics were evaluated. Wound healing activity of the hydrogel-based phage was assessed in a wound mice model. Results: The purified phage showed remarkable properties including broad host range, tolerance to high temperature and pH and biofilm degradation feature as a stable and reliable therapeutic agent. Whole genome sequencing revealed that the genome of the EF-M80 phage had a length of 40,434 bp and harbored 65 open reading frames (ORFs) with a GC content of 34.9% (GenBank accession number is OR767211). Hydrogel-based encapsulated phage represented an optimized structure. Phage-loaded hydrogel-treated mice showed that the counting of neutrophils, fibroblasts, blood vessels, hair follicles and percentage of collagen growth were in favor of the wound healing process in the mice model. Conclusion: These findings collectively suggest the promising capability of this phage-based therapeutic strategy for the treatment of infections associated with the antibiotic-resistant E. faecium. In the near future, we hope to expect the presence of bacteriophages in the list of antibacterial compounds used in the clinical settings.

A comparative evaluation of five phenotypic methods for identification of carbapenemase-producing Enterobacteriaceae: a modified carbapenemase detection test.

Rapid detection of carbapenemase-producing Enterobacteriaceae (CPE) is urgently needed to prevent their spread in healthcare settings. Here, we have evaluated the performance of the phenotypic methods for detection of carbapenemase production directly from bacterial cultures. A total of 99 clinical and rectal Enterobacteriaceae isolates were included (81 carrying known carbapenemase-encoding genes and 18 without carbapenemase production). All isolates were subjected to the five phenotypic tests including in-house Carba NP (iCarba NP), modified-Carba NP, E-Test MBL, modified Hodge test (MHT), and commercial combination disk test. Test results were read at different time points for iCarba NP and modified-Carba (1 min, 5 min, 15 min, 1 h and 2 h). The sensitivity and specificity of the iCarba NP were 78.87% and 100%, respectively, whereas those of the modified-Carba NP test were 95.06% and 94.44%, respectively. False-negative results were detected in four OXA-48 isolates with the use of modified-Carba NP, whereas one non-carbapenemase isolate had false-positive results. The sensitivity/specificity was 91.30%/100% and 80.25%/83.33% for the E-Test MBL and MHT, respectively. The sensitivity and specificity of the aminophenylboronic acid synergy test were 100% and 97.94%, respectively, whereas those of the dipicolinic acid synergy test were 82.61% and 96.23%, respectively. Rapid, simple, and reliable methods are needed for laboratory detection of CPE isolates to improve the detection and surveillance of these clinically relevant pathogens in an epidemiological context. We conclude that the modified-Carba NP test can be one of the reliable tests for the prediction of carbapenemase-producing bacteria.IMPORTANCEThe emergence of carbapenem resistance among Gram-negative bacteria is a serious global health threat. Here, we investigate the performance of the five phenotypic assays against carbapenemase-producing and carbapenemase-non-producing Enterobacteriaceae. Accurate and rapid detection of CPE isolates is critically required for clinical management and treatment of infections caused by these organisms. Among the five evaluated phenotypic tests, the mCNP test presented the highest sensitivity (95.06%) and, therefore, can be considered the best test to be used as a screening phenotypic methodology.

First case of New Delhi metallo ß-lactamase-1 (NDM-1)-producing Burkholderia cepacia complex in Iran.

The Burkholderia cepacia complex (Bcc) is a gram-negative bacillus, which is intrinsically resistant to several used antibiotics, and is now recognized as a group of opportunistic pathogens in Cystic Fibrosis patients. Here, for the first time, we report the case of a patient with New Delhi metallo ß-lactamase (NDM)-positive Bcc lower respiratory tract infection in Iran. The patient was a 57-year-old male admitted to our hospital due to breathlessness, with a history of pulmonary thromboembolism and hypertension. On day 14, the patient underwent bronchoscopy and a bronchoalveolar lavage (BAL) specimen was taken. BAL culture grew Bcc. The drug resistance analysis showed positive NDM resistance, with susceptibility to only quinolones, therefore, levofloxacin was prescribed to the patient. He was discharged from the hospital on the 20th day, 4 days after the initiation of levofloxacin therapy, and died at home on the fifth day after discharge. This is the first report of a lung infection caused by an extensively drug-resistant NDM-positive Bcc strain in Iran.

Outbreak of colistin and carbapenem-resistant Klebsiella pneumoniae ST16 co-producing NDM-1 and OXA-48 isolates in an Iranian hospital.

BACKGROUND: Colistin and carbapenem-resistant Klebsiella pneumoniae (Col-CRKP) represent a significant and constantly growing threat to global public health. We report here an outbreak of Col-CRKP infections during the fifth wave of COVID-19 pandemic. METHODS: The outbreak occurred in an intensive care unit with 22 beds at a teaching university hospital, Isfahan, Iran. We collected eight Col-CRKP strains from seven patients and characterized these strains for their antimicrobial susceptibility, determination of hypermucoviscous phenotype, capsular serotyping, molecular detection of virulence and resistance genes. Clonal relatedness of the isolates was performed using MLST. RESULTS: The COVID-19 patients were aged 24-75 years with at least 50% pulmonary involvement and were admitted to the intensive care unit. They all had superinfection caused by Col-CRKP, and poor responses to antibiotic treatment and died. With the exception of one isolate that belonged to the ST11, all seven representative Col-CRKP strains belonged to the ST16. Of these eight isolates, one ST16 isolate carried the iucA and ybtS genes was identified as serotype K20 hypervirulent Col-CRKP. The blaSHV and blaNDM-1 genes were the most prevalent resistance genes, followed by blaOXA-48 and blaCTX-M-15 and blaTEM genes. Mobilized colistin-resistance genes were not detected in the isolates. CONCLUSIONS: The continual emergence of ST16 Col-CRKP strains is a major threat to public health worldwide due to multidrug-resistant and highly transmissible characteristics. It seems that the potential dissemination of these clones highlights the importance of appropriate monitoring and strict infection control measures to prevent the spread of resistant bacteria in hospitals.

Carbapenemase genes distribution in clonal lineages of Acinetobacter baumannii: a comprehensive study on plasmids and chromosomes.

Background: The global spread of plasmids carrying carbapenemase genes within carbapenem resistant Acinetobacter baumannii (CRAB) strains poses a worldwide public health issue. In this study, we conducted a comprehensive genetic analysis of plasmids and chromosomes harboring the major carbapenemase genes (bla NDM, bla KPC, bla VIM, bla IMP, bla GES, bla OXA-58-like, bla OXA-24/40-like, bla OXA-143-like, and bla OXA-23-like) in CRAB strains using bioinformatic tools. Methods: We retrieved plasmids and chromosomes carrying the major carbapenemase genes from GenBank. The size, replicon type, and conjugal apparatus of the plasmids were also determined. Furthermore, allele types, co-existence of other antimicrobial resistance genes alongside carbapenemases in plasmids or chromosomes, co-occurrence of carbapenemase genes, gene repetition, and sequence types (ST) of whole genomes were characterized. Results: The database contained 113 plasmids and 38 chromosomes harboring carbapenemase genes. This investigation revealed that bla NDM and bla OXA-58-like were the predominant allele types in both the plasmids and chromosomes. Nine (7.96%) plasmids with bla NDM-1 were potentially conjugative. The most common replicon types of the plasmids were R3-T1, R3-T8, R3-T2, R3-T23, and RP-T1. The analysis revealed that bla NDM-1 and bla OXA-58-like genes possessed the highest variety of co-existence with other antibiotic resistance genes. The co-occurrence of dual carbapenemases was identified in 12 plasmids and 19 chromosomes. Carbapenemase gene repetitions were identified in 10 plasmids and one chromosome. Circular alignment revealed that the plasmids carrying the co-occurrence of bla NDM-1 and bla OXA-58 were more homogeneous. However, there was heterogeneity in certain regions of these plasmids. According to the minimum spanning tree (MST) results, the majority of the plasmids belonged to the genomes of ST2Pas, ST1Pas, ST422Pas, ST622Pas, and ST85Pas. Conclusion: A. baumannii appears to have a strong ability for genome plasticity to incorporate carbapenemase genes on its plasmids and chromosomes to develop resistance against carbapenems. Mobilizable plasmids harboring carbapenemases significantly contribute to the dissemination of these genes. The genetic structure of the plasmids revealed a strong associations of class I integrons, ISAba-like structures, Tn4401 elements, and aac (6')-Ib with carbapenemases. Furthermore, gene repetition may also be associated with carbapenem heteroresistance.

Secondary bacterial infection caused by ST16 NDM-1 and OXA-48-producing colistin and carbapenem-resistant Klebsiella pneumoniae treated with tigecycline in a pregnant woman with COVID-19.

During the COVID-19 pandemic, the rapid emergence of carbapenem and colistin-resistant Klebsiella pneumoniae has resulted in an alarming situation worldwide. We aimed to describe secondary infections and antimicrobial use, in a pregnant woman admitted to hospital with COVID-19. A 28-year-old pregnant woman was admitted to the hospital due to COVID-19. According to the clinical conditions, the patient was transferred to the ICU on the second day. She was empirically treated with ampicillin and clindamycin. Mechanical ventilation through an endotracheal tube was started on the 10th day. During her hospitalization in the ICU, she was infected with ESBL-producing K. pneumonia, Enterobacter spp and carbapenemase-producing colistin-resistant Klebsiella pneumoniae isolates. Finally, the patient was treated with tigecycline monotherapy that was associated with ventilator-associated pneumonia clearance. Bacterial co-infection is relatively infrequent in hospitalized patients with COVID-19. Treatment of infections caused by carbapenemase-producing colistin-resistant K. pneumoniae isolates is challenging, with limited antimicrobials available in Iran. In order to prevent the spread of extensively drug-resistant bacteria, infection control programs must be implemented more seriously.

Assessing the knowledge, attitudes and practices of physicians on antibiotic use and antimicrobial resistance in Iran: a cross-sectional survey.

BACKGROUND: Antimicrobial resistance (AMR) is a global public health issue. Physicians should play a key role to fight AMR, and medical education is a fundamental issue to combat it. Understanding the knowledge, attitudes and practices of physicians regarding antibiotic prescription and antibiotic resistance is fundamental for controlling the irrational antibiotic use. This study was conducted to assess the knowledge, attitudes and the practices of physicians in Iran with respect to antibiotic resistance and usage. METHODS: A cross-sectional study was performed from June to October 2021 among physicians at primary care centers and academic hospitals in the region of Isfahan, Iran. A total of 182 physicians were surveyed. Participants were invited to complete a self-reported questionnaire (paper based or online questionnaire). The questions were based on knowledge, attitude, and practice toward antibiotic usage and AMR. Data were analyzed using SPSS version 18 software following the objective of the study. RESULTS: Out of 182 study participants, 100, 50 and 32 responders were medical doctors (MD), internist and other specialists, respectively. Regarding the knowledge section of the questionnaire, almost less than 10% of participants declared to know the antibiotics of Iran's antimicrobial stewardship program. Also, the percentage of participants who correctly responded to clinical quizzes was 23% for treatment of extended-spectrum beta-lactamase (ESBL) producers, 59.3% about the treatment of severe sepsis, 22% about the intrinsic resistance of Proteus mirabilis and 43.4% for experimental treatment with vancomycin in community-acquired pneumonia. Regarding attitude, most participants (97.2%) were aware of the antimicrobial resistance problem in Iran, and 95.6% agreed that prescribing antimicrobials was not the appropriate in our country. Regarding practice, only 65.9% of participants said that before prescribing antibiotics they use of local and international antimicrobial therapy guidelines and less than 50% of physicians were in contact with a microbiology laboratory. CONCLUSION: This data revealed that our physicians' level of knowledge about AMR and antimicrobial stewardship is poor, so there is the need to increase training on antibiotic resistance and antimicrobial stewardship.

The correlation between viral shedding duration and blood biomarkers in COVID-19-infected patients.

Background: Since December 2019, the world is struggling with an outbreak of coronavirus disease-2019 (COVID-19) infection mostly represented as an acute respiratory distress syndrome and has turned into the most critical health issue worldwide. Limited information is available about the association between dynamic changes in the naso/oropharyngeal viral shedding in infected patients and biomarkers, aiming to be assessed in the current study. Materials and Methods: This quasi-cohort study was conducted on 31 patients with moderate severity of COVID-19 manifestations, whose real-time polymerase chain reaction (RT-PCR) test was positive for severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) RNA at baseline. RT-PCR was rechecked for patients every 3-4 days until achieving two negative ones. In parallel, biomarkers, including lymphocyte count, lactate dehydrogenase (LDH), and C-reactive protein (CRP), were assessed every other day, as well. Viral shedding also was assessed. Results: Spearman's correlation test revealed a significant direct correlation between the viral shedding from the symptom onset and the time, in which CRP (P = 0.0015, r = 0.54) and LDH (P = 0.001, r = 0.6207) return to normal levels after symptom onset, but not for lymphocyte count (P = 0.068, r = 0.34). Conclusion: Based on the current study's findings, the duration of SARS-CoV-2 RNA shedding was directly correlated with the required time for LDH and CRP return to normal levels. Therefore, these factors can be considered the determinants for patients' discharge, isolation, and return to social activities; however, further investigations are required to generalize the outcomes.

Barriers to rational antibiotic prescription in Iran: a descriptive qualitative study.

INTRODUCTION: Rational antibiotic prescription (RAP) refers to the purposeful and appropriate antibiotic prescription with correct dose and course to produce the most possible benefits and less possible side effects. Identification and management of the barriers to RAP can help promote RAP. The aim of the study was to explore the barriers to RAP in Iran. METHODS: This descriptive qualitative study was conducted in 2021 on 46 physicians (including general physicians, specialists, and subspecialists), pharmacologists, microbiologists, and nurses. Participants were purposefully selected from five specialty and subspecialty hospitals in Isfahan, Iran, and the Treatment Administration of Isfahan University of Medical Sciences, Isfahan, Iran. Data were collected via semi-structured interviews and were analyzed via conventional content analysis. RESULTS: The barriers to RAP in Iran came into sixteen subcategories and four main categories, namely physicians' limited professional competence (with six subcategories), poor informational and functional resources (with four subcategories), ineffective supervision of RAP (with three subcategories), and inappropriate context for RAP (with three subcategories). The subcategories of these categories were physicians' limited professional knowledge, physicians' poor attitude towards RAP, physicians' routine-based practice instead of evidence-based practice, physicians' limited accountability, physicians' fear over the legal consequences of not prescribing antibiotics, physicians' financial motives, limited access to quality educational materials, poor in-service training for physicians, lack of culturally appropriate guidelines, inefficiency of the stewardship committee, limited supervision of physicians' performance, ineffective managerial supervision, limited supervision of sampling for antimicrobial susceptibility testing, sociocultural factors contributing to irrational antibiotic prescription, poor adherence of insurance companies to their financial commitments, and financial incentives of pharmaceutical companies for physicians. CONCLUSION: The barriers to RAP are different and complex and include physician-related, resource-related, supervision-related, and contextual factors. Physicians with limited professional competence, limited access to resources, and limited supervision will have problems in RAP. Effective management of the barriers to RAP can promote RAP and minimize irrational antibiotic prescription and its consequences, chiefly antimicrobial resistance.

Secondary Bacterial Infection and Clinical Characteristics in Patients With COVID-19 Admitted to Two Intensive Care Units of an Academic Hospital in Iran During the First Wave of the Pandemic.

Data on the prevalence of bacterial co-infections and secondary infection among adults with COVID-19 admitted to the intensive care unit (ICU) are rare. We aimed to determine the frequency of secondary bacterial infection, antibiotic use, and clinical characteristics in patients admitted to the ICU with severe SARS-CoV-2 pneumonia. This was a retrospective cohort study of adults with severe COVID-19 admitted to two ICUs from March 6 to September 7, 2020 in an academic medical center in Isfahan, Iran. To detect COVID-19, reverse transcription real-time polymerase chain reaction was performed and also typical pattern of CT scan was used for the diagnosis of COVID-19. Data collection included the age, gender, main symptoms, history of underlying disease, demographics, hospital stay, outcomes, and antibiotic regimen of the patient. Antimicrobial susceptibility testing was carried out according to the CLSI guidelines. During the study period, 553 patients were referred to the both ICUs for COVID-19 with severe pneumonia. Secondary bacterial infection was detected in 65 (11.9%) patients. The median age was 69.4 (range 21-95) years; 42 (63.6%) were men. Notably, 100% (n = 65) of the patients with superinfection were prescribed empirical antibiotics before first positive culture, predominantly meropenem (86.2%) with a median duration of 12 (range 2-32) days and levofloxacin (73.8%) with a median duration of nine (range 2-24) days. Most prevalent causative agents for secondary bacterial infection were Klebsiella pneumoniae (n = 44) and Acinetobacter baumannii (n = 33). Most patients with secondary bacterial infection showed extensive drug-resistance. The mortality among patients who acquired superinfections was 83% against an overall mortality of 38.1% in total admitted COVID-19 patients. We found a high prevalence of carbapenem-resistant Gram-negative bacilli in COVID-19 patients admitted to our ICUs, with a high proportion of K. pneumoniae followed by A. baumannii. These findings emphasize the importance of implementation of strict infection control measures and highlight the role of antimicrobial stewardship during a pandemic.

Comparative resistome and virulome analysis of clinical NDM-1-producing carbapenem-resistant Enterobacter cloacae complex.

OBJECTIVES: The Enterobacter cloacae complex (ECC) are causatives of hospital-acquired infections. The antimicrobial resistance (AMR) and virulence profiling of ECC promotes our knowledge for their elimination in clinical settings. METHODS: We assembled the whole genome of four clinical carbapenem-resistant ECCs and characterized their AMR and virulence profiles using whole genome sequencing. RESULTS: The chromosomes length scaled from minimum 3 949 952 bp (for P2) to maximum 4 976 575 bp (for P3). Strains P1 and P2 belonged to sequence type (ST)182. P3 and P4 belonged to ST477 and ST134, respectively. The blaCTX-M-15 gene was detected in P1 plasmid. P1 and P4 harboured the blaTEM-1 and blaOXA-1 genes. blaNDM-1 was found in P1, P3, and P4. No blaOXA-48, blaKPC, blaVIM, or blaIMP were identified. The plasmids were nontransferable and had IncFIB, IncFII, Col, and IncC incompatibility (Inc) groups . Class 1 integron was detected in all strains. Virulence genes related to biofilms, adhesins, siderophores (aerobactin, enterobactin, and salmochelin), intrinsic antimicrobial efflux pumps, secretory systems type I to VI, environmental and antibiotic stress response , outer membrane proteins, and heavy metal (copper, tellurite, arsenic, and zinc) resistance were found. The number of positive virulence factors was higher for P1 compared with other strains. CONCLUSION: The accumulation of AMR genes in Enterobacter spp. and their high endurance in hostile environments is a serious health problem. More genomic investigations are required to determine their AMR and virulence genetic reservoirs at the global level.

Association of fluoroquinolone resistance and ESBL production in hypervirulent Klebsiella pneumoniae ST11 and ST893 in Iran.

The spread of multidrug resistance in Klebsiella pneumoniae is a serious threat to the public health. In this study, the prevalence of fluoroquinolone resistance and virulence determinants among ESBL-producing K. pneumoniae isolates was investigated. A total of 50 third-generation cephalosporin resistant K. pneumoniae strains were collected from patients' clinical cultures between September 1st, 2019 and February 30th, 2020. Clonal relatedness of clinical isolates was determined by multilocus sequence typing. All 50 isolates were multidrug-resistant (MDR) and carried at least one of the ESBL resistance determinants. The bla CTX-M-15 gene was the major ESBL determinant found in K. pneumoniae (88%), followed by bla SHV (86%) and bla TEM (78%). PMQR was detected in 96% of the isolates and aac(6')-Ib-cr was the most common (78%) as well as multiple mutations in gyrA (S83I, D87G) and parC (S80I) were found. Selected isolates were assigned to seven sequence types (STs) (ST11, ST893, ST147, ST16, ST377, ST13, and ST392). Overall, hypervirulent phenotypes were identified in 26 (52%) of the isolates. Among the 50 isolates, 28 (56%) were positive for ybt, 23 (46%) for rmpA, 17 (34%) for iroB, 15 (30%) for magA, 4 (8%) for alls and 3 (6%) for iucA genes. The K1 capsular type was the most prevalent (11/50; 22%) among isolates. The emergence of hypervirulent K. pneumoniae (hvKp) ST11 and ST893, which co-carried ESBL, PMQR determinants and different virulence genes has become a threat to the treatment of inpatients in the clinical setting.

Comprehensive pan-genomic, resistome and virulome analysis of clinical OXA-48 producing carbapenem-resistant Serratia marcescens strains.

BACKGROUND: Carbapenem-resistant Enterobacteriaceae (CRE) have been thoroughly studied as the pathogens associated with hospital acquired infections. However, data on Serratia marcescens are not enough. S. marcescens is now becoming a propensity for its highly antimicrobial-resistant clinical infections. METHODS: Four carbapenem-resistant S. marcescens (CR-SM) isolates were obtained from hospitalized patients through routine microbiological experiments. We assembled the isolates genomes using whole genome sequencing (WGS) and compared their resistome and virulome patterns. RESULTS: The average length and CG content of chromosomes was 5.33 Mbp and 59.8%, respectively. The number of coding sequences (CDSs) ranged from 4,959 to 4,989. All strains had one single putative conjugative plasmid with IncL incompatibility (Inc) group. The strains harbored blaCTX-M-15, blaTEM-1 and blaSHV-134. All plamsids were positive for blaOXA-48. No blaNDM-1, blaKPC, blaVIM and blaIMP were identified. The blaSRT-2 and aac(6')-Ic genes were chromosomally-encoded. Class 1 integron was detected in strains P8, P11 and P14. The Escher_RCS47 and Salmon_SJ46 prophages played major role in plasmid-mediated carraige of extended spectrum ß-lactamases (ESBLs). The CR-SM strains were equipt with typical virulence factors of oppotunistic pathogens including biofilm formation, adhesins, secretory systems and siderophores. The strains did not have ability to produce prodigiosin but were positive for chitinase and EstA. CONCLUSION: The presence of conjugative plasmids harboring major ß-lactamases within prophage and class 1 integron structures highlights the role of different mobile genetic elements (MGEs) in distribution of AMR factors and more specifically carbapenemases. More molecular studies are required to determine the status of carbapenem resistance in clinical starins. However, appropriate strategies to control the global dissemination of CR-SM are urgent.

Emergence of carbapenem resistant Acinetobacter baumannii clonal complexes CC2 and CC10 among fecal carriages in an educational hospital.

Carbapenem-resistant Acinetobacter baumannii strains are increasing worldwide. In this study, samples were collected from hospital environments, extra hospital environments, and fecal carriages. 76% (89/117) of bacterial isolates were detected as A. baumannii strains. The imipenem resistance in the hospital environment, fecal carriages, extra hospital environments, and clinical isolates was 37.7% (17/45), 100% (9/9), 0% (0/45), and 92.9% (92/99), respectively. The blaVIM and blaOXA-23 were detected in 6.6% (3/45) and 2.2% (1/45) of strains isolated from hospital environments. Interestingly, strains isolated from fecal carriages had blaVIM, blaOXA-23, and blaIMP genes which resembled carbapenem resistance genes in clinical strains. The structure of clonal relatedness among all non-clinical isolates was as follows: CC2, 37% (33/89); CC1, 22.4% (20/89); CC3, 12.3% (11/89); CC25, 7.8% (7/89); CC10, 4.4% (4/89) and CC15, 2.2% (2/89). Comparison of clonal relatedness among clinical and non-clinical isolates indicated that widespread clones including CC2, CC3, and CC10 were common clonal complexes between two categories.

Comparative genome analysis of colistin-resistant OXA-48-producing Klebsiella pneumoniae clinical strains isolated from two Iranian hospitals.

BACKGROUND: Carbapenemase-producing Klebsiella pneumoniae (CP-KP) is becoming extensively disseminated in Iranian medical centers. Colistin is among the few agents that retains its activity against CP-KP. However, the administration of colistin for treatment of carbapenem-resistant infections has increased resistance against this antibiotic. Therefore, the identification of genetic background of co-carbapenem, colistin-resistance K. pneumoniae (Co-CCRKp) is urgent for implementation of serious infection control strategies. METHODS: Fourteen Co-CCRKp strains obtained from routine microbiological examinations were subjected to molecular analysis of antimicrobial resistance (AMR) using whole genome sequencing (WGS). RESULTS: Nine of 14 K. pneumoniae strains belonged to sequence type (ST)-11 and 50% of the isolates had K-locus type 15. All strains carried blaOXA-48 except for P26. blaNDM-1 was detected in only two plasmids associated with P6 and P26 strains belonging to incompatibility (Inc) groups; IncFIB, IncHI1B and IncFII. No blaKPC, blaVIM and blaIMP were identified. Multi-drug resistant (MDR) conjugative plasmids were identified in strains P6, P31, P35, P38 and P40. MICcolistin of K. pneumoniae strains ranged from 4 to 32 µg/ml. Modification of PmrA, PmrB, PhoQ, RamA and CrrB regulators as well as MgrB was identified as the mechanism of colistin resistance in our isolates. Single amino acid polymorphysims (SAPs) in PhoQ (D150G) and PmrB (R256G) were identified in all strains except for P35 and P38. CrrB was absent in P37 and modified in P7 (A200E). Insertion of ISKpn72 (P32), establishment of stop codon (Q30*) (P35 and P38), nucleotides deletion (P37), and amino acid substitution at position 28 were identified in MgrB (P33 and P42). None of the isolates were positive for plasmid-mediated colistin resistance (mcr) genes. P35 and P38 strains carried iutA, iucD, iucC, iucB and iucA genes and are considered as MDR-hypervirulent strains. P6, P7 and P43 had ICEKp4 variant and ICEKp3 was identified in 78% of the strains with specific carriage in ST11. CONCLUSION: In our study, different genetic modifications in chromosomal coding regions of some regulator genes resulted in phenotypic resistance to colistin. However, the extra-chromosomal colistin resistance through mcr genes was not detected. Continuous genomic investigations need to be conducted to accurately depict the status of colistin resistance in clinical settings.

Data on the prevalence and distribution of carbapenemase genes in Enterobacterales species isolated from clinical specimens in the center of Irans.

Carbapenem resistance in Enterobacterales is a major and persistent public health problem worldwide. In current research, we present data of 96 Enterobacterales species collected from a clinical hospital in Isfahan, Iran. The bacterial identification was performed by standard biochemical tests and API 20E methods. Agar disk diffusion assay was performed to determine the phenotypic antibiotic resistance of strains. Polymerase chain reaction (PCR) was carried out to detect carbapenemase genes. In this manuscript, multiple antimicrobial resistance phenotype such as multiple carbapenem resistance determinants were detected. The data would provide important information on distribution of carbapenemase genes of those pathogenic bacteria in Iran.

The face of hypervirulent Klebsiella pneumoniae isolated from clinical samples of two Iranian teaching hospitals.

Hypervirulent Klebsiella pneumoniae (hvKp) has emerged as a pathogen of global concern. In this study, both phenotypic and genotypic tests were used to detect hvKp. Antimicrobial resistance profiles and clonal relatedness of clinical isolates were also determined. We found that 34.2% (163/477) of the isolates were tellurite resistant, and among them 102 hvKp isolates detected with iucA or iutA or peg-344 as molecular markers. The blaSHV (80.4%), followed by blaCTX-M-15 (76.5%) and blaTEM (67.6%), blaOXA-48 (53.9%), and blaNDM-1 (32.3%) were detected, while blaKPC-1 was not present in any hvKp isolates. It was found that the majority of hvKp isolates belonged to capsular serotype K20 and ompK36 group C, which is related to clonal group (CG) 23 (e.g. ST23). A high percentage of multidrug-resistant hvKp (76.6%) and high resistance to imipenem (67%) indicated a serious problem that should be addressed in the clinical setting.

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.

Molecular characterization of carbapenem-resistant serotype K1 hypervirulent Klebsiella pneumoniae ST11 harbouring blaNDM-1 and blaOXA-48 carbapenemases in Iran.

Carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp) has been increasingly reported and is now recognized as a public health concern. The aim of this study was to investigate the molecular epidemiology of CR-hvKp strains that were isolated from an Iranian hospital. A total of 74 non-duplicated carbapenem-resistant K. pneumoniae (CR-Kp) were collected from patients' clinical or surveillance cultures. Resistance/virulence genes were identified by PCR and sequencing. String test, capsular genotyping, conjugation assays, PCR-based replicon typing, pulsed field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and were performed. All 74 CR-Kp isolates were carbapenemase producers, which co-carried multiple resistance genes such as blaCTX-M-15, blaTEM-1, blaSHV-type, qnrB1, and qnrS1. The most common carbapenemase gene was blaOXA-48 (67/74 90.5%), followed by blaNDM-1 (18/74 24.3%), and blaNDM-7 (3/74 4%). The blaOXA-48 and blaNDM-1 were found on IncL/M and IncFII conjugative plasmids, respectively. Of 74 CR-Kp isolates, 49 were positive for string test. Capsular genotyping revealed that 34 and 10 CR-Kp strains belonged to the K1 and K2 serotypes, respectively. rmpA was the most prevalent virulence gene detected in 64.8% of the isolates. Fifty two strains were identified as CR-hvKp. PFGE typing showed 5 different clusters with two major clusters B (39 isolates, 52.7%) associated with sequence type 11 (ST11), and A (21 isolates, 28.4%) associated with ST893. Furthermore, ST147, ST392, and ST15 carbapenemase producers have also been sporadically identified. One isolate belonging to ST11 was resistant to colistin and were negative for mcr-1-2-3 genes. Insertional inactivation of mgrB due to IS elements was observed in the colistin-resistant isolate. Our findings suggest that ST11 CR-hvKP strain has a clonal distribution in our hospital. Therefore, immediate implementation of infection-control measures may be the best way to prevent the spread of these clones.