Phenotypic and Genotypic Characterisation of Clinical Isolates of Nosocomial Infections

The study focuses on identification and characterization of clinical isolates of nosocomial infections with an aim to create a Bank of model microorganisms for further study of mechanisms and prospects of clinical use of novel medicines causing a reversion susceptibility to antibiotics in drug resistant pathogens. Clinical samples of nosocomial infections were collected from phthisiological hospitals in Almaty. Clinical isolates were characterized by morpho-cultural, tinctorial, physiological and biochemical properties and also by susceptibility to antibiotics. Our studies showed that the isolates are characterized by an increased ability to form biofilms that significantly complicates the therapy and prevention of these outbreaks. Moreover, isolates were characterized by varying degrees of susceptibility to antimicrobial drugs. The strains of Citrobacter were resistant to azithromycin, which is considered as a reserve drug. This fact raises a concern about the circulation and the spread in hospitals of microorganisms resistant to the latest generation of antibiotics. Obtained genome-scale contigs were used for taxonomic affiliation of the isolates and for identification of genetic determinants of antibiotic resistance. The search for genetic determinants of drug resistance in the obtained genome sequences confirmed the resistance to some antibiotics obtained by phenotypic methods. Whole genome sequences were obtained for 4 clinical isolates identified as Citrobacter koseri SCAID URN1-2019, Citrobacter freundii SCAID PHRX1-2019, Escherichia coli SCAID URN1-2019 and Streptococcus pneumoniae SCAID PHRX1-2019. The genomes were deposited in the NCBI database under accession numbers CP052059, CP052058, CP052057, and CP052060.

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Prevalence of Quinolones Resistance Proteins Encoding Genes (qnr genes) and Co-Resistance with β-lactams among Klebsiella pneumoniae Isolates from Iraqi Patients

Baghdad Science Journal ◽

10.21123/bsj.2020.17.2.0406 ◽

2020 ◽

Vol 17 (2) ◽

pp. 0406

Author(s):

Mustafa Mustafa et al.

Keyword(s):

Klebsiella Pneumoniae ◽

Biochemical Properties ◽

The Other ◽

Clinical Isolates ◽

Clinical Samples ◽

Missense Mutations ◽

High Association ◽

Resistance Proteins ◽

Major Threat ◽

Encoding Genes

This study investigated the prevalence of quinolones resistance proteins encoding genes (qnr genes) and co-resistance for fluoroquinolones and β-lactams among clinical isolates of Klebsiella pneumoniae. Out of 150 clinical samples, 50 isolates of K. pneumoniae were identified according to morphological and biochemical properties. These isolates were collected from different clinical samples, including 15 (30%) urine, 12 (24%) blood, 9 (18%) sputum, 9 (18%) wound, and 5 (10%) burn. The minimum inhibitory concentrations (MICs) assay revealed that 15 (30%) of isolates were resistant to ciprofloxacin (≥4µg/ml), 11 (22%) of isolates were resistant to levofloxacin (≥8 µg/ml), 21 (42%) of isolates were resistant to ertapenem (≥8 µg/ml), 18 (36%) of isolates were resistant to imipenem (4- ≥16µg/ml), 43 (86%) of isolates were resistant to ceftriaxone (4- ≥64 µg/ml), 42 (84%) of isolates were resistant to ceftazidime (16-64 µg/ml), and 40 (80%) of isolates were resistant to cefepime (4- ≥16µg/ml). The results revealed that all fluoroquinolone resistant K. pneumoniae isolates were resistant for β-lactams that used in this study. Genotypic detection of qnr genes revealed that qnrS and qnrB were found in 38 (76%) and 18 (36%) of K. pneumoniae isolates, respectively. On the other hand, qnrA, qnrC, and qnrD were not found among K. pneumoniae isolates. DNA sequencing of qnrB gene revealed that the presence of silent and missense mutations that may have led to increase the resistance values of MICs for ciprofloxacin and levofloxacin. These variants were registered in NCBI at the accession numbers LC373260 and LC381730. The phylogenetic tree of qnrB variants showed a significant deviation of these variants from K. pneumoniae species. The spread of qnr genes among clinical isolates of K. pneumoniae and high association observed between resistance to fluoroquinolones and β-lactams have led to a major threat to public health through development of MDR K. pneumoniae.

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Presence of β-lactamase encoding genes in clinical isolates of Y. enterocolitica bioserotype 1B/O8mm isolated in Poland in 2009

Medycyna Doświadczalna i Mikrobiologia ◽

10.32394/mdm.70.2.4 ◽

2018 ◽

pp. 149-157

Author(s):

Katarzyna Zacharczuk

Keyword(s):

Sensu Stricto ◽

Clinical Isolates ◽

Clinical Samples ◽

Broth Microdilution ◽

Genetic Determinants ◽

Intrinsic Resistance ◽

Variable Activity ◽

Genes Encoding ◽

Encoding Genes ◽

Mic Value

Introduction: The intrinsic resistance to β-lactam antibiotics of Y. enterocolitica is mainly due to activity of two type β - lactamases, BlaA and BlaB, which are encoding by genes located on chromosome, blaA and ampC, respectively. High-pathogenicity bioserotype 1B/O8 of Y. enterocolitica has been isolated from clinical samples since 2004 year in Poland. The study shown that clinical isolates of Y. enterocolitica 1B/O8 collected from 2004 to 2009 in Poland constituted the same sensu stricto strain. The aim of present study was to determine the occurrence of genes encoding β-lactamases BlaA and BlaB with the MIC value characterization for selected antibiotics in clinical isolates of Yersinia enterocolitica bioserotype 1B/O isolated in Poland in 2009. Methods: Ampicilin and ticarcillin minimum inhibitory concentrations (MICs) for 64 clinical Y. enterocolitica bioserotype 1B/O8 isolates were determined by Etest and broth microdilution respectively. The blaA i blaB genes were detected by PCR. Results: All investigated isolates were resistant to ticarcillin (MIC 16 – 128 mg/ml), whereas 9,5% to ampicilin (MIC 12 mg/l) according to EUCAST interpretive criteria. All isolates yielded the expected amplicons of blaA and ampC genes. Conclusions: Observed differences in the resistance to ampicillin among analyzed isolates belonging to the epidemic sensu stricto strain can indicate the variable activity of genetic determinants of resistance to β-lactam antibiotics of Y. enterocolitica bioserotype 1B/O8.

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Analysis of the monitoring study of the antibiotic-resistance of the agents of purulent-inflammatory processes of soft tissue

Reports of Vinnytsia National Medical University ◽

10.31393/reports-vnmedical-2018-22(2)-10 ◽

2018 ◽

Vol 22 (2) ◽

pp. 285-288

Author(s):

A.P. Prevar ◽

A.V. Kryzshanovskaya ◽

V.A. Radionov ◽

V.M. Mrug

Keyword(s):

Soft Tissues ◽

High Sensitivity ◽

Biochemical Properties ◽

Antimicrobial Drugs ◽

Strict Adherence ◽

E Coli ◽

Monitoring Study ◽

Treatment Measures ◽

Inflammatory Processes ◽

Main Factor

The main factor in the treatment of suppurative and inflammatory processes is the timely optimization of treatment measures taking into account the nature of the microflora and its susceptibility to antimicrobial drugs. The purpose of the study is to monitor the spectrum of microorganisms – pathogens of purulent-inflammatory processes of soft tissues in surgical patients; study of the sensitivity of isolated strains to antibiotics. The material was collected in accordance with aseptic rules. The identification of a pure culture of bacteria was carried out according to morphological, culture, biochemical properties, and the presence of virulence enzymes. Sensitivity of bacteria to antibiotics was determined by the standard disks method (by Kirby-Bauer’s). 255 patients with purulent-inflammatory processes of soft tissues were examined for the period from 2014 to 2017. 229 strains of isolated bacteria were included to Escherichia coli, Citrobacer freundii, Enterobacter cloacae, E.aerogenes, Proteus vulgaris, Staphylococcus aureus, S.epidermidis, Streptococcus pyogenes, S.viridians, S.agalactiae, Pseudomonas aeruginosa. The main cause of purulent-inflammatory processes of soft tissues is Staphylococci (67,2%). Compared to previous studies, the number of P.aeruginosa isolated cultures increased (7.9%). In monoculture and in association with other microorganisms, E. coli (9.6% of cases), E.cloacae et aerogenes (3.9% of cases), P.vulgaris (3.9% of cases), C.freundi (2.5% of cases), S.agalactiae, S.pyogenes, S.viridans (3.5%). The number of associated sows reaches 12%. Clinical strains of microorganisms remain most sensitive to fluoroquinolones, cephalosporins, and also retains high sensitivity to gentamicin, lincomycin, rifampicin, which is important for empirical antibiotic therapy. To increase the effectiveness of antibacterial therapy, strict adherence to the mode of appointment of antibiotics, justification of indications, a combination of antibiotics of different spectrum of action, mandatory correction after determining the sensitivity of the pathogen.

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Isolation of Stenotrophomonas maltophilia from Clinical Samples in Some Hospitals in Shiraz, Iran

Clinical and Experimental Investigations ◽

10.31487/j.cei.2020.03.01 ◽

2020 ◽

pp. 1-3

Author(s):

Majid Baserisalehi ◽

Samira Zarezadeh ◽

Majid Baserisalehi ◽

Saeed Shoa

Keyword(s):

Stenotrophomonas Maltophilia ◽

Enterobacter Aerogenes ◽

Late Summer ◽

Early Spring ◽

Clinical Samples ◽

Bacillus Species ◽

Microbiological Analysis ◽

Blood Samples ◽

Healthcare Facilities ◽

Phenotypic Methods

Stenotrophomonas maltophilia is an emerging pathogenic non-fermentative Gram-negative Bacillus species. It has caused many nosocomial infections and can be isolated from various hospital wards and healthcare facilities. Research has shown that most of its strains are inherently resistant to many antibiotics and have multidrug resistance. This research intended to determine its occurrence frequency at some Hospitals in shiraz, Iran. The present study was conducted in six months (from early spring to late summer 2019). Clinical samples (Blood, Urine and cerebrospinal fluid (CSF)) collected from 120 patients afflicted with various infections. The samples were transferred to the Laboratory and subjected to microbiological analysis. Identification of the isolates was carried out by phenotypic methods and Stenotrophomonas maltophilia isolates verified using molecular methods. In total, various bacteria were isolated from 84 clinical samples. The isolates were Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Stenotrophomonas maltophilia, Staphylococcus aureus and Pseudomonas aeruginosa. Stenotrophomonas maltophilia was isolated from 17 (20.2%) positive samples and most of them were isolated from blood samples. Our finding indicated that Stenotrophomonas maltophilia isolated more from blood samples follow by CSF sample. In addition, our finding illustrated that Stenotrophomonas maltophilia can be considered as the common nosocomial agent at hospitals in Shiraz, Iran.

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1615. Isolation of Lytic Bacteriophages with Broad Host Range Activity Against Pseudomonas aeruginosa Strains Isolated from Respiratory Samples from Cystic Fibrosis Patients Intended for Therapeutic Application

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa439.1795 ◽

2020 ◽

Vol 7 (Supplement_1) ◽

pp. S801-S801

Author(s):

Jose Alexander ◽

Daniel Navas ◽

Marly Flowers ◽

Angela Charles ◽

Amy Carr

Keyword(s):

Cystic Fibrosis ◽

Host Range ◽

Phage Therapy ◽

Clinical Isolates ◽

Clinical Samples ◽

Plaque Morphology ◽

Broad Host Range ◽

Chronic Infections ◽

Clinically Significant ◽

Host Target

Abstract Background With the rise of the antimicrobial resistance between different genera and species of bacteria, Phage Therapy is becoming a more realistic and accessible option for patients with limited or no antimicrobial options. Being able to have rapid access to a collection of clinical active phages is key for rapid implementation of phage therapy. The Microbiology Department at AdventHealth Orlando is performing routine screening of environmental and patient samples for isolation of phages against non-fermenting Gram negative bacteria to develop a Phage Bank. Methods Protocols for phage isolation from environmental sources such as lakes, rivers and sewers and clinical samples were developed. A series of respiratory, throat, stool and urine samples were processed following an internal protocol that includes centrifugation, filtration and enrichment. Clinical samples were centrifugated for 10 minutes, filtered using 0.45µm centrifugation filters, seeded with targeted host bacteria (clinical isolates) and incubated at 35°C for 24 hours. The enriched samples were centrifugated and filtered for a final phage enriched solution. Screening and isolation were performed using the Gracia method over trypticase soybean agar (TSA) for plaque morphology and quantification. Host range screening of other clinical isolates of P. aeruginosa was performed using the new isolated and purified phages. Results 4 lytic phages against clinical strains of P. aeruginosa from patient with diagnosis of cystic fibrosis (CF), were isolated and purified from 4 different respiratory samples, including sputum and bronchial alveolar lavage. All phages showed phenotypical characteristics of lytic activity. 1 phage was active against 4 strains of P. aeruginosa, 1 phage was active against 2 strains of P. aeruginosa and the remaining 2 phages were active only against the initial host target strain. Conclusion With this study we demonstrated the potential use of clinical samples as source for isolating active bacteriophages against clinically significant bacteria strains. Clinical samples from vulnerable population of patients with chronic infections are part of our routine “phage-hunting” process to stock and grow our Phage Bank project for future clinical use. Disclosures All Authors: No reported disclosures

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1280. In-Vitro Activity of Cefiderocol, Imipenem/Relebactam, & Ceftazidime/Avibactam in Ceftolozane/Tazobactam Resistant Strains of Multidrug Resistant Pseudomonas aeruginosa

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa439.1463 ◽

2020 ◽

Vol 7 (Supplement_1) ◽

pp. S655-S655

Author(s):

Daniel Navas ◽

Angela Charles ◽

Amy Carr ◽

Jose Alexander

Keyword(s):

Multidrug Resistant ◽

Resistance Mechanisms ◽

Vitro Activity ◽

Clinical Isolates ◽

Resistance Testing ◽

Clinical Samples ◽

In Vitro Activity ◽

Carbapenemase Gene ◽

Resistant Strains

Abstract Background The activity of imipenem/relebactam (I/R), ceftazidime/avibactam (CZA) and cefiderocol (FDC) were evaluated against clinical isolates of multidrug resistant (MDR) strains of P. aeruginosa which was resistant to ceftolozane/tazobactam (C/T). The recent increase of MDR P. aeruginosa strains isolated from clinical samples has prompted research and development of new antimicrobials that can withstand its multiple resistance mechanisms. C/T is an effective option for treatment of MDR P. aeruginosa in our facility with only 10% of resistance in MDR strains, but the emergence of resistance may occur due to the presence of a carbapenemase gene or an ampC mutation. Methods Antimicrobial susceptibility testing for C/T Etest® (bioMérieux, Inc.) were performed on all MDR strains initially screened by the VITEK2® (bioMérieux, Inc.). 10% (n=20) of all MDR isolates were resistant to C/T by the CLSI 2019 breakpoints. These resistant isolates were tested for presence of a carbapenemase gene using the GeneXpert CARBA-R (Cepheid®) PCR and against CZA Etest® (bioMérieux, Inc.) I/R gradient strips (Liofilchem®) and FDC broth microdilution (Thermo Scientific™ Sensititre™). Results A total of 20 clinical isolates of MDR P. aeruginosa resistant to C/T were tested following standardized CLSI protocols and techniques. All 20 isolates were screened for the presence of a carbapenemase gene (blaVIM, blaNDM, blaKPC, blaOXA-48, blaIMP). A blaVIM gene was detected in 6 (30%) out of 20 isolates. FDC demonstrated the greatest activity with 85% (n=17) of susceptible isolates (CLSI MIC <4µg/dL). CZA (CLSI MIC <8µg/dL) and I/R (FDA MIC <2µg/dL) showed 15% (n=3) and 10% (n=2) of susceptible isolates respectively. FDC was active against all 6 blaVIM isolates, where all 6 strains were resistant to CZA and I/R as expected. 3 isolates tested non-susceptible against FDC; additional characterization was not performed at this time. Conclusion Based on these results, FDC demonstrated the greatest in-vitro activity against C/T resistant strains of MDR P. aeruginosa. FDC also demonstrated activity against all 6 MDR P. aeruginosa carrying blaVIM gene. FDC is a strong option to consider on MDR P. aeruginosa strains based on a resistance testing algorithm and a cost/effective protocol. Disclosures All Authors: No reported disclosures

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Strain Typing of Trichosporon asahii Clinical Isolates by Random Amplification of Polymorphic DNA (RAPD) Analysis

Journal of Laboratory Physicians ◽

10.1055/s-0041-1731111 ◽

2021 ◽

Author(s):

Thayanidhi Premamalini ◽

Vijayaraman Rajyoganandh ◽

Ramaraj Vijayakumar ◽

Hemanth Veena ◽

Anupma Jyoti Kindo ◽

...

Keyword(s):

Genetic Relatedness ◽

Rapd Analysis ◽

Clinical Isolates ◽

Clinical Samples ◽

Strain Typing ◽

Trichosporon Asahii ◽

Pcr Fingerprinting ◽

Specific Pcr ◽

Rapd Primer ◽

Random Amplification

Abstract Objective The aim of this study was to identify and isolate Trichosporon asahii (T. asahii) from clinical samples and to assess the genetic relatedness of the most frequently isolated strains of T. asahii using random amplification of polymorphic DNA (RAPD) primers GAC-1 and M13. Methods All the clinical samples that grew Trichosporon species, identified and confirmed by polymerase chain reaction (PCR) using Trichosporon genus-specific primers, were considered for the study. Confirmation of the species T. asahii was carried out by T. asahii-specific PCR. Fingerprinting of the most frequently isolated T. asahii isolates was carried out by RAPD using random primers GAC-1 and M13. Results Among the 72 clinical isolates of Trichosporon sp. confirmed by Trichosporon-specific PCR, 65 were found to be T. asahii as identified by T. asahii-specific PCR. Fingerprinting of the 65 isolates confirmed as T. asahii using GAC-1 RAPD primer yielded 11 different patterns, whereas that of M13 primer produced only 5 patterns. The pattern I was found to be the most predominant type (29.2%) followed by pattern III (16.9%) by GAC-1 primer. Conclusions This study being the first of its kind in India on strain typing of T. asahii isolates by adopting RAPD analysis throws light on genetic diversity among the T. asahii isolates from clinical samples. Fingerprinting by RAPD primer GAC-1 identified more heterogeneity among the T. asahii isolates than M13.

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Complete Genome Sequences of Four Extensively Drug-Resistant Acinetobacter baumannii Isolates from Thailand

Microbiology Resource Announcements ◽

10.1128/mra.00949-20 ◽

2020 ◽

Vol 9 (40) ◽

Author(s):

Peechanika Chopjitt ◽

Thidathip Wongsurawat ◽

Piroon Jenjaroenpun ◽

Parichart Boueroy ◽

Rujirat Hatrongjit ◽

...

Keyword(s):

Acinetobacter Baumannii ◽

Complete Genome ◽

Sequence Type ◽

Clinical Isolates ◽

Drug Resistant ◽

Genome Sequences ◽

Content Type ◽

Type Isolate ◽

Resistant Genes ◽

Extensively Drug Resistant

ABSTRACT Here, we report the complete genome sequences of four clinical isolates of extensively drug-resistant Acinetobacter baumannii (XDRAB), isolated in Thailand. These results revealed multiple antimicrobial-resistant genes, each involving two sequence type 16 (ST16) isolates, ST2, and a novel sequence type isolate, ST1479.

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First Draft Genome Sequences of Malaysian Clinical Isolates of Corynebacterium diphtheriae

Genome Announcements ◽

10.1128/genomea.01670-16 ◽

2017 ◽

Vol 5 (9) ◽

Cited By ~ 2

Author(s):

Norazah Ahmad ◽

Shirley Yi Fen Hii ◽

Mohd Khairul Nizam Mohd Khalid ◽

Muhammad Adib Abd Wahab ◽

Rohaidah Hashim ◽

...

Keyword(s):

Draft Genome ◽

Corynebacterium Diphtheriae ◽

Clinical Isolates ◽

Genome Sequences ◽

Content Type

ABSTRACT Corynebacterium diphtheriae has caused multiple isolated diphtheria cases in Malaysia over the years. Here, we report the first draft genome sequences of 15 Malaysia C. diphtheriae clinical isolates collected from the years 1981 to 2016.

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