Direct Susceptibility Testing Research Articles

Assessing the effectiveness of direct susceptibility testing from positive blood culture broth using the VITEK-2 system

Assessing the effectiveness of direct susceptibility testing from positive blood culture broth using the VITEK-2 system

A Comparative Study of Bacteriological Profile and Susceptibility Pattern by Rapid Direct Manual Identification and Antimicrobial Susceptibility Testing with Routine Blood Culture Method in Neonatal Septicemia.

A Comparative Study of Bacteriological Profile and Susceptibility Pattern by Rapid Direct Manual Identification and Antimicrobial Susceptibility Testing with Routine Blood Culture Method in Neonatal Septicemia.

Direct Antibiogram Susceptibility Testing vs Isolation Followed by Subculture in Gram Negative Urine Tract Infections

Direct Antibiogram Susceptibility Testing vs Isolation Followed by Subculture in Gram Negative Urine Tract Infections

Evaluation of Direct Antimicrobial Susceptibility Testing of Gram-Negative Bacilli and Staphylococcus aureus from Positive Pediatric Blood Culture Bottles Using BD Phoenix M50.

Bloodstream infections (BSIs) are life-threatening infections for which a timely initiation of appropriate antimicrobial therapy is critical. Antibiotic susceptibility testing (AST) directly performed on positive blood culture broths can help initiate targeted antibiotic therapy sooner than the standard AST performed on colonies isolated on solid media after overnight incubation. Faster antimicrobial susceptibility testing (AST) results can improve clinical outcomes, and reduce broad-spectrum antimicrobial consumption and healthcare-associated costs in sepsis. In this study, we evaluated the accuracy of a direct AST inoculation method on the BD Phoenix M50 system using serum separator tubes to harvest bacteria from positive pediatric blood culture bottles. Direct AST was performed on 132 monomicrobial pediatric blood culture bottles that were positive for Enterobacterales (65; 49.2%), Staphylococcus aureus (46; 34.8%), and non-fermenting Gram-negative bacilli (21; 16%). Overall, the categorical and essential agreements between the direct method and standard method were 99.6% and 99.8%, respectively. Very major, major, and minor error rates were 0.1%, 0.09%, and 0.20% respectively. Direct AST performed on pediatric blood culture bottles using BD Phoenix M50 can quickly provide accurate susceptibility information to guide antimicrobial therapy in patients with BSI.

Prospective evaluation of thin-layer agar colour test in routine diagnosis of multidrug-resistant TB.

<sec><title>BACKGROUND</title>This study evaluated the diagnostic performance of the thin-layer agar MDR/XDR-TB Colour Test (CT), a Mycobacterium tuberculosis complex (MTBC) detection and direct drug susceptibility testing (DST) method with routine sputum, bronchoalveolar lavage and pleural fluid specimen.</sec><sec><title>METHODS</title>In a prospective study, the time and rate of MTBC detection were compared between CT, Löwenstein-Jensen, and MGIT media. Times until DST result, sensitivities, and specificities were evaluated between CT and MGIT 960 indirect DST.</sec><sec><title>RESULTS</title>The cultivation of 177 pulmonary specimens resulted in 83 MTBC-positive cultures. The sensitivity of CT for MTBC detection was 81.3% with a median time of 20 days compared to 13 days and 93.5% for MGIT. The sensitivity of CT for DST results was 100% for isoniazid and levofloxacin and 94.7% for rifampicin. The specificities for isoniazid and rifampicin DST were 97.3% and 98.0% for levofloxacin. The median time until a DST result was significantly shorter with CT than the BACTEC MGIT 960 system, 20 and 27 days, respectively, independent of the specimen type used.</sec><sec><title>CONCLUSIONS</title>The CT is a highly accurate and fast initial diagnostic test for high-incidence settings and could also be used as a first culture and direct DST in peripheral settings.</sec>.

A study on direct antimicrobial susceptibility testing of positive blood culture broth using BacT/Alert 3D microbial identification system in a tertiary care hospital

A study on direct antimicrobial susceptibility testing of positive blood culture broth using BacT/Alert 3D microbial identification system in a tertiary care hospital

Using the CLSI rAST breakpoints of Enterobacterales in positive blood cultures

ObjectivesThe objective of this study was to provide the clinic with rapid and accurate results of antimicrobial susceptibility testing for the treatment of patients with bloodstream infections. To achieve this, we applied the Clinical and Laboratory Standards Institute (CLSI) blood culture direct rapid antimicrobial susceptibility test (rAST) to assess the susceptibility of the most common Enterobacterales found in blood cultures. MethodsIn this study, we utilized the CLSI blood culture direct rapid antimicrobial susceptibility test to assess the susceptibility (rAST) of the most common Enterobacterales present in blood cultures. We chose this method for its simplicity in analysis, and our aim was to predict minimum inhibitory concentrations (MICs) using the rAST. As a benchmark, we assumed that Broth Macrodilution method (BMD) results were 100% accurate. For data evaluation, we employed the terms categorical agreement (CA), very major errors (VME), and major errors (ME). ResultsOur findings demonstrate that the CLSI rAST method is reliable for rapidly determining the in vitro susceptibility of Enterobacterales to common antimicrobial drugs in bloodstream infections. We achieved a concordance rate of 90% in classification within a 10-hour timeframe. We identified a total of 112 carbapenem-resistant Enterobacterales (CRE) strains, and there was no significant difference in the detection rate of CRE at 6, 10, and 16 hours. This suggests that CRE can be identified as early as 6 hours. ConclusionThe CLSI rAST is a valuable tool that can be utilized in clinical practice to quickly determine the susceptibility of Enterobacterales to antimicrobial drugs within 10 hours. This capability can greatly assist in the clinical management of patients with bloodstream infections.

Evaluation of three protocols for direct susceptibility testing for Gram-negative rods from flagged positive blood culture bottles.

Bloodstream infections are associated with high mortality that can be reduced by targeted antibiotic therapy in the early stages of infection. Direct antibiotic susceptibility testing (AST) from flagged positive blood cultures may facilitate the administration of early effective antimicrobials much before the routine AST. Clinical and Laboratory Standards Institute-recommended direct AST can be performed with a limited number of antibiotic disks only. On the other hand, using an automated system for direct AST will not only allow effective laboratory workflow with reduced turnaround time but also provide the minimum inhibitory concentration values of tested antibiotics. However, using expensive automated systems for direct AST may not be feasible for resource-limited laboratories. Therefore, in this study, we aimed to evaluate the CLSI-recommended method and two other direct AST protocols (one with an automated system and the other with disk diffusion) for Gram-negative rods from flagged positive blood cultures.

Evaluation of Direct Antimicrobial Susceptibility Testing from Positive Flagged Blood Cultures in Sepsis Patients.

Presently, many laboratories are equipped with automated system for antimicrobial susceptibility testing (AST) for minimum inhibitory concentration-based reporting which enables the clinician to choose the right antimicrobial for timely treatment of sepsis. The study aimed to assess performance of direct AST from blood culture positive broth using automated AST system for accuracy and time taken to release the report. The present study conducted in a 25-bedded ICU in North India for 12 months. Single morphotype of bacteria on gram stain from positively flagged blood culture bottles were included, which was directly identified (using an in-house protocol) with MALDI-TOF-MS from positive blood culture broths. DAST was carried out from 200 such blood culture broths and results were compared with reference AST (RAST) which was also done using VITEK-2 using overnight grown bacterial colonies as per standard protocol. Among 60 isolates of Enterobacterales, 99% categorical agreement for both E. coli and K. pneumoniae observed by two methods were tested for AST. Among non-fermenters, Pseudomonas aeruginosa showed a categorical agreement of 99.6%, as compared with Acinetobacter spp. and exotic GNBs, which showed 95-96% agreement. A significant difference of 18-24 hours was noted in time to release the report between DAST and RAST, for GNB and GPC both. Direct AST from positive flagged blood culture bottles can significantly reduce the time to release the bacterial susceptibility report by up to 24 hours, at the same time maintaining the accuracy. Singh V, Agarwal J, Nath SS, Sharma A. Evaluation of Direct Antimicrobial Susceptibility Testing from Positive Flagged Blood Cultures in Sepsis Patients. Indian J Crit Care Med 2024;28(4):387-392.

Evaluation of three protocols for direct susceptibility testing for gram negative-Enterobacteriaceae from patient samples in Uganda with SMS reporting

In Uganda, the challenge of generating and timely reporting essential antimicrobial resistance (AMR) data has led to overreliance on empirical antibiotic therapy, exacerbating the AMR crisis. To address this issue, this study aimed to adapt a one-step AMR testing protocol alongside an SMS (Short Message Service) result relay system (SRRS), with the potential to reduce the turnaround time for AMR testing and result communication from 4 days or more to 1 day in Ugandan clinical microbiology laboratories. Out of the 377 samples examined, 54 isolates were obtained. Notably, E. coli (61%) and K. pneumoniae (33%) were the most frequently identified, majority testing positive for ESBL. Evaluation of three AMR testing protocols revealed varying sensitivity and specificity, with Protocol A (ChromID ESBL-based) demonstrating high sensitivity (100%) but no calculable specificity, Protocol B (ceftazidime-based) showing high sensitivity (100%) and relatively low specificity (7.1%), and Protocol C (cefotaxime-based) exhibiting high sensitivity (97.8%) but no calculable specificity. ESBL positivity strongly correlated with resistance to specific antibiotics, including cefotaxime, ampicillin, and aztreonam (100%), cefuroxime (96%), ceftriaxone (93%), and trimethoprim sulfamethoxazole (87%). The potential of integrating an SRRS underscored the crucial role this could have in enabling efficient healthcare communication in AMR management. This study underscores the substantial potential of the tested protocols for accurately detecting ESBL production in clinical samples, potentially, providing a critical foundation for predicting and reporting AMR patterns. Although considerations related to specificity warrant careful assessment before widespread clinical adoption.

Evaluation of a Modified Protocol for the SepsiPrep Kit for Direct Identification and Antimicrobial Susceptibility Testing From Positive Blood Culture Using BACTEC Plus and BacT/Alert Blood Culture Bottles

Evaluation of a Modified Protocol for the SepsiPrep Kit for Direct Identification and Antimicrobial Susceptibility Testing From Positive Blood Culture Using BACTEC Plus and BacT/Alert Blood Culture Bottles

Evaluation of the VITEK®2 Cards for Rapid Direct Identification and Antimicrobial Susceptibility Testing of Human Pathogenic Bacteria

Evaluation of the VITEK®2 Cards for Rapid Direct Identification and Antimicrobial Susceptibility Testing of Human Pathogenic Bacteria

A prospective study evaluating the effect of a ‘Diagnostic Stewardship Care-Bundle’ for automated blood culture diagnostics

We prospectively implemented a diagnostic stewardship care-bundle checklist, 'Sepsis-48 DSB', with the aim of reducing intervening duration of key steps of automated blood culture diagnostics (aBCD). Sepsis-48 DSB was implemented for automated blood culture bottles (BCBs) received from adult intensive care units (AICUs) during the intervention period (P2; July 2020-June 2021) and intervening durations were compared with those during the retrospective, pre-intervention period (P1; March-June 2020). During both periods, provisional blood culture reports (pBCR) were issued wherein direct microbial identification (dID) was performed in BCBs with Gram-negatives by directly inoculating conventional biochemical tests and direct antimicrobial susceptibility testing (dAST) using EUCAST RAST method. The results were compared with the standard of care (SoC) method (i.e. full incubation followed by identification and AST by VITEKⓇ-2 Compact). During P2, significant reductions in loading time (LT) [median: 63.5 vs. 32 minutes, P < 0.001], time to dID+dAST performance (TTD) [186 vs. 115 minutes, P=0.0018] and an increase in compliance to bundle targets [LT ≤45: 44% vs. 66%, P=0.006 and TTD ≤120: 34% vs. 51.7%, P=0.03] were observed. Using dID+dAST method, results were read 694 minutes earlier than SoC method. Of 176 pBCR, 165 (94%) were concordant with SoC in microbial identification of species. Categorical agreement for any drug-bug combination was 92.7% (1079/1164) and corresponding major, very major, and minor error rates were 8.8% (19/216), 4.9% (45/921), and 1.8% (21/1164), respectively. The 'diagnostic stewardship care-bundle' strategy was successfully implemented with considerable diagnostic accuracy leading to significant reductions in duration of targeted steps of aBCD.

DIRECT ANTIMICROBIAL SUSCEPTIBILITY TESTING OF GRAM NEGATIVE BACILLI FROM POSITIVE BLOOD CULTURES

Conventional blood culture and susceptibility pattern takes around 48 hours after blood culture bottles turns positive on Bact/ALERT system. Rapid initiation of antimicrobial therapy is required in management of sepsis patients to reduce its adverse effects on mortality and morbidity. Objective of this study was to execute rapid and accurate direct antimicrobial susceptibility testing (DST) of gram negative bacilli on positive blood culture bottles by the disk diffusion method and compare the results with those of standard antimicrobial susceptibility testing (AST). This prospective study was performed at the Saveetha Institute of Medical and Technical Science, Sriperumbudur, with mono-bacterial gram negative bacilli. 100-gram negative bacilli (GNB) samples were agged from positive blood cultures with duration of 3 months. Both DST and AST results were interpreted, and the isolates were identied by vitek2. On 100 GNB isolates, the most common organism was Escherichia coli, followed by Klebsiella pneumoniae (27%), Pseudomonas aeruginosa (11%), Acinetobacter baumanii (8%), Salmonella typhi (7%), Enterobacter cloacae (3%), Acinetobacter lwof(3%) and Serratia marsacens. The DST and AST correlations were found to be in 99% categorical agreement, with 1% minor error, 0% major error, and 0% very major error. The concentration of bacterial inoculums impacts the results between DST and AST. DST reduces the crucial time from 24 to 48 hours for patients with bloodstream infections treated with empirical antibiotic therapy, which may or may not be resistant. DST initiates early antibiotic therapy that is susceptible to treating a bloodstream infection or sepsis. Aims and Objective: To evaluate categorical agreement of antimicrobials between DST and AST from positive blood culture bottles by Kirby-Bauer disk diffusion method. Materials and methods: A prospective study was conducted in tertiary care hospital, with sample size of 100 gram negative bacilli from positive blood culture bottles in patients suspected with blood stream infections during the period from October 2022 to December 2022. RESULTS: Out of 100 gram negative bacilli , Escherichia coli (37%) was the most common isolate, followed by Klebsiella pneumonia (27%), Pseudomonass aeroginosa (11%), Acinetobacter baumanii (8%), Salmonella typhi (7%), Enterobacter clocae (3%), Acinetobacter lw of(3%) and Serratia marsacens (2%). Conclusion: During this study, there was 99% categorical agreement between DST and AST. Earlier treatment with antimicrobials can reduce the mortality and morbidity in bacteraemia patients

A COMPARATIVE STUDY TO DETERMINE THE ACCURACY OF DIRECT SUSCEPTIBILITY TESTING WITH CONVENTIONAL ANTIMICROBIAL SUSCEPTIBILITY TESTING

Introduction: Clinically significant bacteria are identified in the laboratory and the accurate information isprovided with antimicrobial susceptibility testing (AST) which is essential for an accuratemanagementofpatientssufferingwith abacterial infection orbacterialdisease[1].As the process of AST istime consuming and Results of AST are provided with in a timespan of 48–72 h after sampling.Disk diffusion has many benefits, which includes low expenditure, time duration reduction ofresults and few more basic benefits. One of the benefits is the probability or chances ofexecuting direct susceptibility testing (DST).when is potentially very useful in the management of critically used selectively and interpreted carefully, DST on clinical samples ill patients, as the time to results is shortened by approximately 24 h. Aim:To do a comparative study for determining the accuracy of direct susceptibility testing with conventional antimicrobial susceptibility testing. Material And Method:This cross- sectional study was conducted in the Department of Microbiology, peoples College of MedicalSciences and Research Centre, over a period of 1 year-from February 2021 to February 2022. Result:A total of 311 samples of urine, pus and body fluid were collected and positive sample processed from 124males and 187 females from age groups 1 to 90 years.From total of 311 samples Out of all the samples 132 were of pus, 175 were of urine specimen 4 were offluidssamples.Comparison of the bacterial response to 14 antimicrobial agents using directantimicrobial sensitivity testing (DST) versus standard antimicrobial sensitivity testing(AST) for Gram positive bacteriaOut of 91 gram positive bacteria we found that on performing AST and DST on norfloxacin probability valuewas 1, which shows that there is no significant difference in results by these two methods. Similarly,Ciprofloxacin, Cefoxitin, Doxycycline, Amoxy-clavulanic acid, Erythromycin, Clindamycin, Linezolid,Chloramphenicol, Cefuroxime all have a p-value as 1 on performing both the different types of antimicrobialsusceptibility testing method which indicates a similarity in their results. Conclusion:After 1 year of systematic study even though we found that AST and DST both gives almost similar result, but keeping in mind major and minor differences in antibiotic sensitivity test . We should consider it only foremergencypurposes.In emergency it is observed that DST proved to be more successful because DST provides results with in 24to 32 hours, this providing results faster but it should always be followed by AST for confirmation of results.With DST, we have to apply more no. of antibiotics disk to coues all bacteria increasingcost of ABST.

Evaluation of a sterile, filter-based, in-house method for rapid direct bacterial identification and antimicrobial susceptibility testing using positive blood culture.

This study aimed to assess the performance of our in-house method for rapid direct bacterial identification (ID) and antimicrobial susceptibility testing (AST) using a positive blood culture (BC) broth. For Gram-negative bacteria, 4mL of BC broth was aspirated and passed through a Sartorius Minisart syringe filter with a pore size of 5µm. The filtrate was then centrifuged and washed. A small volume of the pellet was used for ID, using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and for AST, using automated broth microdilution. For Gram-positive cocci, 4mL of BC broth was passed through the Minisart syringe filter. Then, 4mL of sterile distilled water was injected in the direction opposite to that of the filtration to collect the bacterial residue trapped in the filter. Compared with the conventional method performed with pure colonies on agar plates, 94.0% (234/249) were correctly identified using the in-house method, with rates of 91.4% (127/139) and 97.3% (107/110) for Gram-positive and Gram-negative isolates, respectively. Of 234 correctly identified isolates, 230 were assessed by AST. Categorical agreement and essential agreement were 93.3% and 94.5%, respectively, with a minor error rate of 3.8%, a major error rate of 3.4%, and a very major error rate of 1.6%. Our in-house preparation method showed good performance in rapid direct ID and AST using positive BC broths compared to the conventional method. This simple method can shorten the conventional turnaround time for ID and AST by at least 1day, potentially contributing to better patient management.

EUCAST rapid antimicrobial susceptibility testing (RAST) compared to conventional susceptibility testing: implementation and potential added value in a tertiary hospital in Belgium

ABSTRACT Objectives EUCAST breakpoints for short incubation disk diffusion allow rapid antimicrobial susceptibility testing (RAST) directly from positive blood cultures. We evaluate the RAST methodology and assess its potential added value in a setting of low prevalence of multidrug-resistant (MDR) organisms. Methods In our two-part study, we performed RAST on 127 clinical blood cultures at 6 and 8 h and determined categorical agreement with direct susceptibility testing. We also measure the impact of susceptibility results on antimicrobial therapy compared to empirical treatment. Results Categorical agreement was 96.2% at 6 h (575/598 isolate-drug combinations) and 96.6% at 8 h (568/588 combinations). Major errors involved piperacillin/tazobactam in 16 of 31 cases. The second part of our study shows that AST reporting proved essential in correcting ineffective empirical therapy in 6.3% of the patients (8/126). Conclusion EUCAST RAST is an inexpensive and reliable method of susceptibility testing, although care must be taken with reporting piperacillin/tazobactam. In support of RAST implementation, we show that AST remains of great importance in providing effective therapy, even in a setting of low MDR prevalence and elaborate antibiotic guidelines.

A simple double differential centrifugation-wash procedure to rapidly obtain bacterial identification and direct antimicrobial susceptibility testing from positive blood cultures

IntroductionThis study proposes a simple and rapid method for both bacterial identification and direct antimicrobial susceptibility testing (AST) by using MALDI-TOF and a double differential centrifugation-wash procedure from positive blood cultures. MethodsFifty-two positive blood cultures (37 gramnegative bacilli and 15 grampositive cocci) were studied by two methods for identification and AST: a reference method, and the rapid MALDI-TOF method obtaining a purified pellet by using a double differential centrifugation procedure. ResultsA total of 1101 MIC values (mg/l) were interpreted according to EUCAST clinical breakpoints and compared using the two methods simultaneously. Discrepancies in 81 MIC values (7.35%) were detected. By analyzing standard parameters, we obtained 98.28% essential agreement and 92.65% categorical agreement considering all isolates tested. ConclusionThis method provides rapid bacterial identification and AST, offering definitive results 24–48h earlier than the conventional method (p<0.001) and improving the turnaround time in blood culture diagnostics, especially in laboratories without 24-h on-call.

Comparative analysis of conventional and direct antimicrobial susceptibility testing of blood cultures in a tertiary care cancer hospital: the way forward

ABSTRACT Bloodstream infections (BSI) are associated with high mortality rates, especially in immunocompromised patients. Identifying pathogens early and selecting appropriate antimicrobials to treat BSI is integral in reducing the mortality rate. There is a need to reduce the turnaround time (TAT) of pathogen identification as well as to accelerate the antimicrobial susceptibility testing (AST) of blood cultures, which can be achieved by following relevant identification methods and performing the direct AST (DAST) by the disk diffusion method. In this study, blood samples were collected from patients with suspected bacteremia/septicemia, and aseptic precautions were taken to prevent contamination. Samples containing gram-negative bacilli (GNB) were then analyzed by DAST and conventional AST (CAST). We tested 118 GNB-positive isolates in total to compare the results of DAST and CAST. DAST and CAST showed good categorical agreement (CA) for various groups of microorganisms: 98.9% and 99.6% for Enterobacterales and Pseudomonas spp., respectively. Early detection of pathogens in blood along with the determination of their antibiotic susceptibility patterns is a need of the hour. By performing DAST on positive blood culture broth, clinical teams can obtain the information necessary for switching from empirical therapy to definitive treatment one day faster. This rapid identification of the pathogen, along with corresponding AST results, will help clinicians to accelerate targeted antimicrobial therapy for critical patients and, thus, reduce mortality and morbidity rates in patients with bloodstream infections.

Direct single-cell antimicrobial susceptibility testing of Escherichia coli in urine using a ready-to-use 3D microwell array chip.

Empirical antibiotic therapies are prescribed for treating uncomplicated urinary tract infections (UTIs) due to the long turnaround time of conventional antimicrobial susceptibility testing (AST), leading to the prevalence of multi-drug resistant pathogens. We present a ready-to-use 3D microwell array chip to directly conduct comprehensive AST of pathogenic agents in urine at the single-cell level. The developed device features a highly integrated 3D microwell array, offering a dynamic range from 102 to 107 CFU mL-1, and a capillary valve-based flow distributor for flow equidistribution in dispensing channels and uniform sample distribution. The chip with pre-loaded reagents and negative pressure inside only requires the user to initiate AST by loading samples (∼3 s) and can work independently. We demonstrate an accessible sample-to-result workflow, including syringe filter-based bacteria separation and rapid single-cell AST on chip, which enables us to bypass the time-consuming bacteria isolation and pre-culture, speeding up the AST in ∼3 h from 2 days of conventional methods. Moreover, the bacterial concentration and AST with minimum inhibitory concentrations can be assessed simultaneously to provide comprehensive information on infections. With further development for multiple antibiotic conditions, the Dsc-AST assay could contribute to timely prescription of targeted drugs for better patient outcomes and mitigation of the threat of drug-resistant bacteria.