Bacterial Detection, Identification, and Antimicrobial Susceptibility Testing

A special issue of Antibiotics (ISSN 2079-6382).

Deadline for manuscript submissions: closed (31 March 2024) | Viewed by 8640

Special Issue Editor


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Guest Editor
Microbiology and Virology Unit, Turin University, 10124 Turin, Italy
Interests: clinical microbiology; rapid diagnostic; antibiotic resistance; MALDI-TOF MS; lateral flow immunoassay; Real Time PCR; DNA sequencing; β-lactamases; antimicrobial susceptibility testing; synergy testing; time kill assay; new β-lactam/β-lactamase
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Special Issue Information

Dear Colleagues,

Microbiological diagnostics plays a key role in the determination of the appropriate treatment for infectious diseases. Recent technological advances have enabled accurate microbiological diagnoses with shorter turnaround times. In addition, several antimicrobial molecules have been introduced to the market for the treatment of infections due to multidrug-resistant bacteria.

Therefore, for this Special Issue, we invite authors to contribute original research and review papers describing the application of innovative diagnostic techniques for the detection or identification of bacterial agents and the application of rapid or conventional methods for the determination of in vitro antimicrobial susceptibility.

Dr. Gabriele Bianco
Guest Editor

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Antibiotics is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2900 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • identification methods
  • rapid diagnostics
  • MALDI-TOF MS
  • molecular testing
  • rapid antimicrobial susceptibility testing
  • antimicrobial susceptibility
  • synergy testing
  • β-lactam/β-lactamase inhibitor
  • fosfomycin
  • colistin

Published Papers (6 papers)

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Research

17 pages, 2143 KiB  
Article
Rapid Minimum Inhibitory Concentration (MIC) Analysis Using Lyophilized Reagent Beads in a Novel Multiphase, Single-Vessel Assay
by Tejas Suresh Khire, Wei Gao, Brian Bales, Kuangwen Hsieh, Greg Grossmann, Dong Jin M. Park, Christine O’Keefe, Arnyah Brown-Countess, Sara Peterson, Fan-En Chen, Ralf Lenigk, Alex Trick, Tza-Huei Wang and Christopher Puleo
Antibiotics 2023, 12(11), 1641; https://doi.org/10.3390/antibiotics12111641 - 19 Nov 2023
Viewed by 1583
Abstract
Antimicrobial resistance (AMR) is a global threat fueled by incorrect (and overuse) of antibiotic drugs, giving rise to the evolution of multi- and extreme drug-resistant bacterial strains. The longer time to antibiotic administration (TTA) associated with the gold standard bacterial culture method has [...] Read more.
Antimicrobial resistance (AMR) is a global threat fueled by incorrect (and overuse) of antibiotic drugs, giving rise to the evolution of multi- and extreme drug-resistant bacterial strains. The longer time to antibiotic administration (TTA) associated with the gold standard bacterial culture method has been responsible for the empirical usage of antibiotics and is a key factor in the rise of AMR. While polymerase chain reaction (PCR) and other nucleic acid amplification methods are rapidly replacing traditional culture methods, their scope has been restricted mainly to detect genotypic determinants of resistance and provide little to no information on phenotypic susceptibility to antibiotics. The work presented here aims to provide phenotypic antimicrobial susceptibility testing (AST) information by pairing short growth periods (~3–4 h) with downstream PCR assays to ultimately predict minimum inhibitory concentration (MIC) values of antibiotic treatment. To further simplify the dual workflows of the AST and PCR assays, these reactions are carried out in a single-vessel format (PCR tube) using novel lyophilized reagent beads (LRBs), which store dried PCR reagents along with primers and enzymes, and antibiotic drugs separately. The two reactions are separated in space and time using a melting paraffin wax seal, thus eliminating the need to transfer reagents across different consumables and minimizing user interactions. Finally, these two-step single-vessel reactions are multiplexed by using a microfluidic manifold that allows simultaneous testing of an unknown bacterial sample against different antibiotics at varying concentrations. The LRBs used in the microfluidic system showed no interference with the bacterial growth and PCR assays and provided an innovative platform for rapid point-of-care diagnostics (POC-Dx). Full article
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12 pages, 573 KiB  
Article
Polyclonal Multidrug ESBL-Producing Klebsiella pneumoniae and Emergence of Susceptible Hypervirulent Klebsiella pneumoniae ST23 Isolates in Mozambique
by José João Sumbana, Antonella Santona, Nader Abdelmalek, Maura Fiamma, Massimo Deligios, Alice Manjate, Jahit Sacarlal, Salvatore Rubino and Bianca Paglietti
Antibiotics 2023, 12(9), 1439; https://doi.org/10.3390/antibiotics12091439 - 12 Sep 2023
Viewed by 943
Abstract
Globally, antibiotic-resistant Klebsiella spp. cause healthcare-associated infections with high mortality rates, and the rise of hypervirulent Klebsiella pneumoniae (hvKp) poses a significant threat to human health linked to community-acquired infections and increasing non-susceptibility. We investigated the phenotypic and genetic features of [...] Read more.
Globally, antibiotic-resistant Klebsiella spp. cause healthcare-associated infections with high mortality rates, and the rise of hypervirulent Klebsiella pneumoniae (hvKp) poses a significant threat to human health linked to community-acquired infections and increasing non-susceptibility. We investigated the phenotypic and genetic features of 36 Klebsiella isolates recovered from invasive infections at Hospital Central of Maputo in Mozambique during one year. The majority of the isolates displayed multidrug resistance (MDR) (29/36) to cephalosporins, gentamicin, ciprofloxacin, and trimethoprim–sulfamethoxazole but retained susceptibility to amikacin, carbapenems, and colistin. Most isolates were ESBLs-producing (28/36), predominantly carrying the blaCTX-M-15 and other beta-lactamase genes (blaSHV, blaTEM-1, and blaOXA-1). Among the 16 genomes sequenced, multiple resistance genes from different antibiotic classes were identified, with blaCTX-M-15, mostly in the ISEcp1-blaCTX-M-15-orf477 genetic environment, co-existing with blaTEM-1 and aac(3)-IIa in five isolates. Our results highlight the presence of polyclonal MDR ESBL-producing K. pneumoniae from eight sequence types (ST), mostly harbouring distinct yersiniabactin within the conjugative integrative element (ICE). Further, we identified susceptible hvKp ST23, O1-K1-type isolates carrying yersiniabactin (ybt1/ICEKp10), colibactin, salmochelin, aerobactin, and hypermucoid locus (rmpADC), associated with severe infections in humans. These findings are worrying and underline the importance of implementing surveillance strategies to avoid the risk of the emergence of the most threatening MDR hvKp. Full article
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12 pages, 1587 KiB  
Article
QUIRMIA—A Phenotype-Based Algorithm for the Inference of Quinolone Resistance Mechanisms in Escherichia coli
by Frank Imkamp, Elias Bodendoerfer and Stefano Mancini
Antibiotics 2023, 12(7), 1119; https://doi.org/10.3390/antibiotics12071119 - 28 Jun 2023
Cited by 3 | Viewed by 1017
Abstract
Objectives: Quinolone resistance in Escherichia coli occurs mainly as a result of mutations in the quinolone-resistance-determining regions of gyrA and parC, which encode the drugs’ primary targets. Mutational alterations affecting drug permeability or efflux as well as plasmid-based resistance mechanisms can also [...] Read more.
Objectives: Quinolone resistance in Escherichia coli occurs mainly as a result of mutations in the quinolone-resistance-determining regions of gyrA and parC, which encode the drugs’ primary targets. Mutational alterations affecting drug permeability or efflux as well as plasmid-based resistance mechanisms can also contribute to resistance, albeit to a lesser extent. Simplifying and generalizing complex evolutionary trajectories, low-level resistance towards fluoroquinolones arises from a single mutation in gyrA, while clinical high-level resistance is associated with two mutations in gyrA plus one mutation in parC. Both low- and high-level resistance can be detected phenotypically using nalidixic acid and fluoroquinolones such as ciprofloxacin, respectively. The aim of this study was to develop a decision tree based on disc diffusion data and to define epidemiological cut-offs to infer resistance mechanisms and to predict clinical resistance in E. coli. This diagnostic algorithm should provide a coherent genotype/phenotype classification, which separates the wildtype from any non-wildtype and further differentiates within the non-wildtype. Methods: Phenotypic susceptibility of 553 clinical E. coli isolates towards nalidixic acid, ciprofloxacin, norfloxacin and levofloxacin was determined by disc diffusion, and the genomes were sequenced. Based on epidemiological cut-offs, we developed a QUInolone Resistance Mechanisms Inference Algorithm (QUIRMIA) to infer the underlying resistance mechanisms responsible for the corresponding phenotypes, resulting in the categorization as “susceptible” (wildtype), “low-level resistance” (non-wildtype) and “high-level resistance” (non-wildtype). The congruence of phenotypes and whole genome sequencing (WGS)-derived genotypes was then assigned using QUIRMIA- and EUCAST-based AST interpretation. Results: QUIRMIA-based inference of resistance mechanisms and sequencing data were highly congruent (542/553, 98%). In contrast, EUCAST-based classification with its binary classification into “susceptible” and “resistant” isolates failed to recognize and properly categorize low-level resistant isolates. Conclusions: QUIRMIA provides a coherent genotype/phenotype categorization and may be integrated in the EUCAST expert rule set, thereby enabling reliable detection of low-level resistant isolates, which may help to better predict outcome and to prevent the emergence of clinical resistance. Full article
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11 pages, 736 KiB  
Article
Influence of Multiplex PCR in the Management of Antibiotic Treatment in Patients with Bacteremia
by Alina-Ioana Andrei, Daniela Tălăpan, Alexandru Rafila and Gabriel Adrian Popescu
Antibiotics 2023, 12(6), 1038; https://doi.org/10.3390/antibiotics12061038 - 10 Jun 2023
Viewed by 1240
Abstract
The multiplex PCR assay can be a helpful diagnostic tool for patients with bacteremia. Herein, we assessed the impact of a Blood Culture Identification Panel (BCID) on both the diagnosis and treatment of patients with bacteremia. We performed a retrospective study using laboratory [...] Read more.
The multiplex PCR assay can be a helpful diagnostic tool for patients with bacteremia. Herein, we assessed the impact of a Blood Culture Identification Panel (BCID) on both the diagnosis and treatment of patients with bacteremia. We performed a retrospective study using laboratory and clinical data to evaluate the impact of syndromic testing using a multiplex PCR testing system (BioFire® FilmArray) for the management of patients with bloodstream infections. BCID detected the pathogen in 102 (87.9%) samples out of the 116 positive blood cultures tested. The average time from the blood culture collection to the communication of the molecular test result was 23.93 h (range: 10.67–69.27 h). The main pathogen detected was Klebsiella pneumoniae (17.6%). The antimicrobial therapy was changed in accordance with the BCID results in 28 (40.6%) out of the 69 cases, wherein the treatment could have been theoretically adjusted. This allowed the adjustment of the therapy to be performed 1305.1 h faster than it would have been possible if conventional diagnostic methods had been used; this was the case for only 35.1% of the time gained if treatment was adjusted for all patients with positive BCID. Thus, although molecular tests can make a difference in the management of bloodstream infections, there is room for improvement in the clinical application of BCID results. Full article
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12 pages, 295 KiB  
Article
In Vitro Activities of Ceftobiprole, Dalbavancin, Tedizolid and Comparators against Clinical Isolates of Methicillin-Resistant Staphylococcus aureus Associated with Skin and Soft Tissue Infections
by Sofia Maraki, Viktoria Eirini Mavromanolaki, Dimitra Stafylaki, Evangelia Iliaki-Giannakoudaki and George Hamilos
Antibiotics 2023, 12(5), 900; https://doi.org/10.3390/antibiotics12050900 - 12 May 2023
Cited by 1 | Viewed by 1310
Abstract
Skin and soft tissue infections (SSTIs) are associated with significant morbidity and healthcare costs, especially when caused by methicillin-resistant Staphylococcus aureus (MRSA). Vancomycin is a preferred antimicrobial therapy for the management of complicated SSTIs (cSSTIs) caused by MRSA, with linezolid and daptomycin regarded [...] Read more.
Skin and soft tissue infections (SSTIs) are associated with significant morbidity and healthcare costs, especially when caused by methicillin-resistant Staphylococcus aureus (MRSA). Vancomycin is a preferred antimicrobial therapy for the management of complicated SSTIs (cSSTIs) caused by MRSA, with linezolid and daptomycin regarded as alternative therapeutic options. Due to the increased rates of antimicrobial resistance in MRSA, several new antibiotics with activity against MRSA have been recently introduced in clinical practice, including ceftobiprole, dalbavancin, and tedizolid. We evaluated the in vitro activities of the aforementioned antibiotics against 124 clinical isolates of MRSA obtained from consecutive patients with SSTIs during the study period (2020–2022). Minimum inhibitory concentrations (MICs) for vancomycin, daptomycin, ceftobiprole, dalbavancin, linezolid and tedizolid were evaluated by the MIC Test Strip using Liofilchem strips. We found that when compared to the in vitro activity of vancomycin (MIC90 = 2 μg/mL), dalbavancin possessed the lowest MIC90 (MIC90 = 0.094 μg/mL), followed by tedizolid (MIC90 = 0.38 μg/mL), linezolid, ceftobiprole, and daptomycin (MIC90 = 1 μg/mL). Dalbavancin demonstrated significantly lower MIC50 and MIC90 values compared to vancomycin (0.064 vs. 1 and 0.094 vs. 2, respectively). Tedizolid exhibited an almost threefold greater level of in vitro activity than linezolid, and also had superior in vitro activity compared to ceftobiprole, daptomycin and vancomycin. Multidrug-resistant (MDR) phenotypes were detected among 71.8% of the isolates. In conclusion, ceftobiprole, dalbavancin and tedizolid exhibited potent activity against MRSA and are promising antimicrobials in the management of SSTIs caused by MRSA. Full article
9 pages, 250 KiB  
Article
Disc Diffusion and ComASP® Cefiderocol Microdilution Panel to Overcome the Challenge of Cefiderocol Susceptibility Testing in Clinical Laboratory Routine
by Gabriele Bianco, Matteo Boattini, Sara Comini, Giuliana Banche, Rossana Cavallo and Cristina Costa
Antibiotics 2023, 12(3), 604; https://doi.org/10.3390/antibiotics12030604 - 17 Mar 2023
Cited by 5 | Viewed by 1758
Abstract
Cefiderocol susceptibility testing represents a major challenge for clinical microbiology. Although disc diffusion showed robustness to test cefiderocol susceptibility, large areas of technical uncertainty (ATU) are reported by current EUCAST breakpoints. Herein, we evaluated the in vitro activity of cefiderocol on a collection [...] Read more.
Cefiderocol susceptibility testing represents a major challenge for clinical microbiology. Although disc diffusion showed robustness to test cefiderocol susceptibility, large areas of technical uncertainty (ATU) are reported by current EUCAST breakpoints. Herein, we evaluated the in vitro activity of cefiderocol on a collection of 286 difficult-to-treat Gram-negative isolates using disc diffusion and ComASP® cefiderocol microdilution panel. Broth microdilution (BMD) in iron-depleted Mueller–Hinton broth was used as reference method. Following the EUCAST guidelines, disc diffusion allowed to determine cefiderocol susceptibility (susceptible or resistant) in 78.6%, 88.1%, 85.4% and 100% of Enterobacterales, P. aeruginosa, A. baumannii and S. maltophilia isolates tested, respectively. ComASP® cefiderocol panel showed 94% and 84% of overall categorical agreement and essential agreement. Only one very major error and two major errors were observed, for MIC values nearly close to the resistance breakpoint (2 mg/L). Overall, 20.5% of the carbapenemase-producing Enterobacterales that achieved ATU results by the disc diffusion method tested resistant by both ComASP® panel and reference BMD. Conversely, all VIM-producing P. aeruginosa showed MIC values in the susceptible range (≤2 mg/L). Lastly, only six out of seven (85.7%) A. baumannii isolates showing inhibition zones <17 mm tested resistant by both ComASP® panel and the reference BMD suggesting that inhibition zone <17 mm are not unequivocally suggestive of resistance. Our results, although obtained on a limited number of isolates, suggest that the combination of disc diffusion with a ComASP® cefiderocol microdilution panel could be a viable solution to overcome the challenge of cefiderocol susceptibility testing in routine microbiology laboratories. Full article
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