Accelerated microbial identification "directly" from positive blood cultures using MALDI-TOF MS: Local clinical laboratory challenges.

Rapid identification of microbial pathogens "directly" from positive blood cultures (PBCs) is critical for prompt initiation of empirical antibiotic therapy and clinical outcomes. Towards higher microbial identification rates, we modified a published initial serum separator tubes-based MALDI-TOF-MS protocol, for blood culture specimens received at a non-hospital based standalone diagnostic laboratory, Bangalore, India: (a) "Initial" protocol #1: From 28 PBCs, identification= 39% (Gram-negative= 43%: Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa; Gram-positive: 36%: Enterococcus faecalis, Staphylococcus aureus, Staphylococcus haemolyticus); mis-identification= 14%; non-identification= 47%. (b) "Modified" protocol #2: Quality controls (ATCC colonies spiked in negative blood cultures) From 7 analysis, identification= 100% (Escherichia coli, Klebsiella pneumonia, Klebsiella oxytoca, Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus aureus); From 7 PBCs, identification= 57%; mis-identification= 14%; non-identification= 29%. Microbial preparations of highest quality and quantity for proteomic analysis and separate spectra matching reference databases for colonies and PBCs are needed for best clinical utility.

Performance evaluation of the FAST™ System and the FAST-PBC Prep™ cartridges for speeded-up positive blood culture testing.

Objectives: As time to appropriate antimicrobial therapy is major to reduce sepsis mortality, there is great interest in the development of tools for direct identification (ID) and antimicrobial susceptibility testing (AST) of positive blood cultures (PBC). Very recently, the FAST™ System (Qvella) has been developed to isolate and concentrate microorganisms directly from PBCs, resulting in the recovery of a Liquid Colony™ (LC) within 30 min. The LC can be used as equivalent of an overnight subcultured colony for downstream testing. We aimed to evaluate the performances of the FAST™ System and FAST-PBC Prep™ cartridges by testing the resulting LC for direct ID, AST and rapid resistance detection. Materials and methods: Prospectively, FAST™ System testing was carried out on each patient's first PBC with a monomicrobial Gram-stain result. In the second arm of the study, FAST™ System testing was carried out on blood cultures spiked with multidrug-resistant bacteria. Downstream testing using the LC included MALDI-TOF MS ID with the Bruker Biotyper® smart system, rapid resistance detection testing including the Abbott Diagnostics Clearview™ PBP2a SA Culture Colony Test (PBP2a) and the Bio-Rad ßLACTA™ Test (ßLT). AST was performed using the Becton Dickinson Phoenix™ System or by Bio-Rad disk diffusion using filter paper disk following EUCAST 2020 breakpoint criteria. Results: FAST™ System testing was completed on 198 prospective PBCs and 80 spiked blood cultures. After exclusion of polymicrobial blood cultures, performance evaluation compared with standard of care results was carried out on 266 PBCs. Concordant, erroneous and no ID results included 238/266 (89.5%), 1/266 (0.4%), 27/266 (10.2%) PBCs, respectively. Sensitivity and specificity for PBP2a were 100% (10/10) and 75% (15/20), respectively. Sensitivity and specificity for ßLT were 95.8% (23/24) and 100% (42/42), respectively. Categorical agreement for all 160 tested strains was 98% (2299/2346) with 1.2% (8/657) very major errors and 0.7% (10/1347) major errors. Conclusion: FAST™ System testing is a reliable approach for direct downstream testing of PBCs including MALDI-TOF MS ID, BD Phoenix™ and Bio-Rad disk diffusion AST as well as rapid resistance testing assays. Next steps include optimal integration of the FAST™ System in the PBC workflow with a view toward clinical studies.

Evaluation of a lateral flow immunoassay to detectCTX-M extended-spectrum B-lactamases (ESBL)directly from positive blood cultures for its potentialuse in Antimicrobial Stewardship programs / Evaluación de una inmunocromatografía paradetectar beta-lactamasas de espectro extendidoCTX-M directamente de hemocultivos positivospara su potencial uso en programas deoptimización de antibioterapia

Background. Bloodstream infections (BSI) caused by extended-spectrum beta-lactamases Enterobacteriaceae (ESBL-E) are associated with high rates of treatment failure andincreased mortality, especially when appropriate antimicrobialtherapy is delayed. Our aim was to evaluate the anticipationof ESBLs detection and the potential improvement of the timeresponse of the Vitek 2 System (BioMérieux; France).Methods. We compared this lateral flow immunoassaywhen used directly on fluid from positive blood cultures to theVITEK2 AST system. We evaluated 80 isolates, 61 tested directlyon fluid from positive blood cultures and 19 tested on fluidfrom blood cultures spiked with known ESBL positive and negative organisms.Results. The concordance between the CTX-LFIA and thereference method (Vitek 2) had a Cohen´s Kappa coefficient of0.97, indicating a particularly good correlation between bothcompared methods.Conclusion. This lateral flow immunoassay can be morerapid than the Vitek 2 for earlier presumptive identification ofCTX- M ESBLs and can be useful to anticipate results and theadjustment of antimicrobial therapy. (AU)

Antecedentes. Las bacteriemias causadas por Enterobacteriaceae productoras beta-lactamasas de espectro extendido(BLEE) están asociadas con altas tasas de fallo de tratamientoy mortalidad, especialmente cuando se retrasa el tratamientoapropiado. Nuestro objetivo ha sido evaluar la anticipación dela detección de estas BLEE y la potencial mejora en el tiempode respuesta respecto al VITEK2 System (Biomerieux; Francia).Métodos. Se comparó una inmunocromatografía para sudetección con el VITEK2 AST system directamente del hemocultivo. Se evaluaton 80 aislados, 61 evaluados directamentede hemocultivos positivos y 19 de la misma manera pero inoculados con microorganismos productores y no productores deBLEE.Resultados. La concordancia entre la inmunocromatografía y el VITEK2 AST mostró un coeficiente Kappa de 0,97,indicando una buena correlación entre ambas técnicas.Conclusión. Esta inmunocromatografía puede ser másrápida que el VITEK2 para una identificación de BLEE tipo CTXM y puede ser útil para anticipar resultados y ajustar la terapiaantimicrobiana. (AU)

Evaluation of a lateral flow immunoassay to detect CTX-M extended-spectrum ß-lactamases (ESBL) directly from positive blood cultures for its potential use in Antimicrobial Stewardship programs.

OBJECTIVE: Bloodstream infections (BSI) caused by extended-spectrum beta-lactamases Enterobacteriaceae (ESBL-E) are associated with high rates of treatment failure and increased mortality, especially when appropriate antimicrobial therapy is delayed. Our aim was to evaluate the anticipation of ESBLs detection and the potential improvement of the time response of the Vitek 2 System (BioMérieux; France). METHODS: We compared this lateral flow immunoassay when used directly on fluid from positive blood cultures to the VITEK2 AST system. We evaluated 80 isolates, 61 tested directly on fluid from positive blood cultures and 19 tested on fluid from blood cultures spiked with known ESBL positive and negative organisms. RESULTS: The concordance between the CTX-LFIA and the reference method (Vitek 2) had a Cohen´s Kappa coefficient of 0.97, indicating a particularly good correlation between both compared methods. CONCLUSIONS: This lateral flow immunoassay can be more rapid than the Vitek 2 for earlier presumptive identification of CTX-M ESBLs and can be useful to anticipate results and the adjustment of antimicrobial therapy.

Bacteraemia and antibiotic sensitivity in a tertiary neonatal intensive care unit.

BACKGROUND: Neonatal sepsis is an important cause of mortality and morbidity in neonatal intensive care populations worldwide. Data on rates of bacteraemia and antibiotic resistance patterns are limited, particularly in the developing world. METHODS: We retrospectively reviewed positive blood cultures obtained in the neonatal intensive care unit between 01 January 2015 and 31 December 2015. All neonates, either born at the tertiary hospital or transferred from referral units, regardless of diagnosis, who had a positive blood culture were included. RESULTS: There were 702 admissions during the study period and 437 positive cultures. Male patients made up 55.1% (65/118), and the gender was unknown for 11.0% (13/118). Late onset sepsis accounted for 85.7% (102/119) and early onset sepsis, 14.3% (17/119). Of the 119 organisms cultured, 76 (63.8%) were Gram-negative, 35 (29.4%) were Gram-positive and 8 (6.7%) were Candida species. Klebsiella was the most common genus at 42% (50/119). Of the clinically relevant organisms recovered, 37.0% (44/119) were susceptible to the empiric first-line regimen of penicillin and gentamycin. Furthermore, 69.7% (53/76) of the Gram-negative organisms produced extended-spectrum beta-lactamases. CONCLUSION: The majority of organisms cultured were considered contaminants and were not clinically relevant. Improvements in culture collection processes are needed. The majority of organisms considered clinically relevant were resistant to the first-line antibiotic regimen. To improve the likelihood of clinical success, empiric antibiotic regimens should be based on local data, if possible.

False-positive blood cultures: The need for follow-up.

The diagnosis of blood-borne infections in immunocompromised patients is a major challenge for the clinical microbiology laboratory. Isolation of bloodborne pathogens in these patients has profound clinical implications, yet is fraught with technical problems, including the presence of unusual and difficult to isolate pathogens. Coupled with this is the problem of false-positive blood culture signals from automated blood culture systems which further delays the definitive diagnosis. Here, we present a case of an 8-year-old boy with Ph +ve acute lymphoblastic leukaemia who has repeated 'false positive' blood cultures and later grew an uncommon organism.

Prediction of blood culture outcome using hybrid neural network model based on electronic health records.

BACKGROUND: Blood cultures are often performed to detect patients who has a serious illness without infections and patients with bloodstream infections. Early positive blood culture prediction is important, as bloodstream infections may cause inflammation of the body, even organ failure or death. However, existing work mainly adopts statistical models with laboratory indicators, and fails to make full use of textual description information from EHRs. METHODS: We study the problem of positive blood culture prediction by using neural network model. Specifically, we first construct dataset from raw EHRs. Then we propose a hybrid neural network which incorporates attention based Bi-directional Long Short-Term Memory and Autoencoder networks to fully capture the information in EHRs. RESULTS: In order to evaluate the proposed method, we constructe a dataset which consists of totally 5963 patients who had one or more blood cultures tests during hospitalization. Experimental results show that the proposed neural model gets 91.23% F-measure for this task. CONCLUSIONS: The comparison results of different models demonstrated the effectiveness of our model. The proposed model outperformed traditional statistical models.

Performance Evaluation of the MBT STAR®-Carba IVD Assay for the Detection of Carbapenemases With MALDI-TOF MS.

Objectives: The increasing rate of carbapenem resistance in Gram-negative bacteria is a major public health problem and rapid detection is essential for infection management. We evaluated the performances of the MBT STAR®-Carba IVD assay (Bruker Daltonics) to detect carbapenemase-producing organisms (CPO) from bacterial colonies and directly from positive blood culture bottles with MALDI-TOF MS. Methods: We analyzed 130 strains with a reduced susceptibility to at least one carbapenem including 109 CPO (6 KPC, 27 NDM, 21 VIM, 1 IMP, 41 OXA-48-like, 8 OXA-23, 2 OXA-24/-40, and 2 OXA-58) and 21 non-CPO. The assay on colonies was performed with all 130 strains while the assay on spiked blood cultures was performed with 45 strains. Samples were prepared with the MBT STAR®-CARBA IVD kit and imipenem hydrolysis by the potential carbapenemase was analyzed with the MBT STAR®-BL module (Bruker Daltonics) on MALDI-TOF MS. Results: Performed on colonies, the assay detected all carbapenemase-producing Enterobacteriaceae (n = 78), Pseudomonas spp. (n = 19) and Acinetobacter spp. (n = 12). All 21 tested non-CPO remained negative resulting in sensitivity and specificity of 100%. Performed on positive blood cultures, the assay detected all carbapenemase-producing Enterobacteriaceae (n = 23) and Pseudomonas spp. (n = 4) but missed 9/12 carbapenemase-producing Acinetobacter spp. However, a prolonged imipenem-incubation time of the strain pellet improved carbapenemase detection. Non-CPO from positive blood culture bottles remained negative (n = 5) with the assay with the exception of one Klebsiella pneumoniae isolate. Conclusion: The MBT STAR®-Carba IVD assay is a highly reliable method for the detection of carbapenemase activity in Gram-negative bacteria. However, time-consuming sample preparation steps and reagent costs need to be considered before implementation in a routine clinical microbiology laboratory.

Economic health care costs of blood culture contamination: A systematic review.

BACKGROUND: Blood culture contamination with gram-positive organisms is a common occurrence in patients suspected of bloodstream infections, especially in emergency departments. Although numerous research studies have investigated the cost implications of blood culture contamination, a contemporary systematic review of the literature has not been performed. The aim of this project was to perform a systematic review of the published literature on the economic costs of blood culture contamination. METHODS: PubMed was searched (January 1, 1978, to July 15, 2018) using the search terms "blood culture contamination" or "false-positive blood cultures." Articles were title searched and abstracts were reviewed for eligible articles that reported immediate or downstream economic costs of blood culture contamination. RESULTS AND DISCUSSION: The PubMed search identified 151 relevant articles by title search, with 49 articles included after abstract review. From the studies included, overall blood culture contamination rates ranged from 0.9%-41%. Up to 59% of patients received unnecessary treatment with parenteral vancomycin as a result of blood culture contamination, resulting in increased pharmacy charges between $210 and $12,611 per patient. Increases in total laboratory charges between $2,397 and $11,152 per patient were reported. Attributable hospital length of stay increases due to blood culture contamination ranged from 1-22 days. CONCLUSIONS: This systematic review of the literature identified several areas of health care expenditure associated with blood culture contamination. Interventions to reduce the risk of blood culture contamination would avoid downstream economic costs.

Procalcitonin-guided Antibiotic Treatment in Patients With Positive Blood Cultures: A Patient-level Meta-analysis of Randomized Trials.

BACKGROUND: Whether procalcitonin (PCT)-guided antibiotic management in patients with positive blood cultures is safe remains understudied. We performed a patient-level meta-analysis to investigate effects of PCT-guided antibiotic management in patients with bacteremia. METHODS: We extracted and analyzed individual data of 523 patients with positive blood cultures included in 13 trials, in which patients were randomly assigned to receive antibiotics based on PCT levels (PCT group) or a control group. The main efficacy endpoint was duration of antibiotic treatment. The main safety endpoint was mortality within 30 days. RESULTS: Mean duration of antibiotic therapy was significantly shorter for 253 patients who received PCT-guided treatment than for 270 control patients (-2.86 days [95% confidence interval [CI], -4.88 to -.84]; P = .006). Mortality was similar in both arms (16.6% vs 20.0%; P = .263). In subgroup analyses by type of pathogen, we noted a trend of shorter mean antibiotic durations in the PCT arm for patients infected with gram-positive organisms or Escherichia coli and significantly shorter treatment for subjects with pneumococcal bacteremia. In analysis by site of infection, antibiotic exposure was shortened in PCT subjects with Streptococcus pneumoniae respiratory infection and those with E. coli urogenital infections. CONCLUSIONS: This meta-analysis of patients with bacteremia receiving PCT-guided antibiotic management demonstrates lower antibiotic exposure without an apparent increase in mortality. Few differences were demonstrated in subgroup analysis stratified by type or site of infection but notable for decreased exposure in patients with pneumococcal pneumonia and E. coli urogenital infections.

ICU intervention during induction chemotherapy for adult patients with newly diagnosed acute myeloid leukemia.

We carried out a retrospective study on newly diagnosed AML patients to identify the risk factors associated with intensive care unit (ICU) intervention. One hundred and twenty consecutive AML patients were included. The median cycle of induction therapy (IT) was 2 (range 1-4). Ten patients (8%) needed ICU intervention during IT. The median time from first IT to ICU transfer was 16days (range 2-88days). Three patients required vasopressor/s, three mechanical ventilation, and four both. The cumulative probability for ICU intervention rose progressively with increasing cycles of IT received, from 2.5% during first induction to 27.5% at fourth induction. Age, sex, presentation white cell counts and coagulation profiles, cytogenetics, pre-chemotherapy ventricular ejection fractions, and prior chemoradiation were not risk factors. Univariate analysis identified a history of inflammatory bowel disease (IBD) (p=0.004) (RR=5.7; p=0.03) and positive blood cultures (BC) (p=0.03) (RR=3; p=0.06) as significant risks. Multivariate analysis found a history of IBD as the only significant factor (p=0.03), while positive BC (p=0.1) trending towards significance. AML patients with a history of IBD and positive BC are at increased risks for ICU intervention during IT.

Blood culture contamination rate in an intensive care setting: Effectiveness of an education-based intervention.

BACKGROUND: Blood culture (BC) contamination rate is an indicator of quality of care scarcely explored in intensive care units (ICUs). We analyzed the BC contamination rate in our ICU to assess the effectiveness of an education-based intervention. METHODS: We conducted an interventional study with concurrent controls. Consecutive BCs drawn during a 6-month period were included. An education-based intervention was presented to case nurses (optimal technique). The remaining nurses comprised the control group (standard technique). Two independent observers assessed clinical significance of saprophytic skin bacteria isolated in BCs. RESULTS: Six hundred fifty-six BCs were obtained: 308 (47%) via optimal technique and 348 (53%) via standard technique (47%). One hundred eighty-seven BCs were positive for saprophytic microorganisms; 127 (89%) were considered unrelated to infection. Coagulase-negative staphylococci isolation was lower in the optimal technique group (14% vs 26%; P < .001), as well as contamination due to coagulase-negative staphylococci (12% vs 21%; P = .002) or Acinetobacter baumannii (0.3% vs 2%; P = .013). BC contamination rate was 13% in the optimal technique group versus 23% in the standard group (P < .005). In the optimal technique group, BC contamination rate was higher in BCs drawn through the catheter (17% vs 7%; P = .028). CONCLUSIONS: An education-based intervention significantly reduced the BC contamination rate in our ICU. It seems necessary to design a tool to extract BCs through the catheter to minimize the risk of contamination.

Rapid detection of Gram-negative bacteria and their drug resistance genes from positive blood cultures using an automated microarray assay.

We evaluated the performance of the Verigene Gram-negative blood culture (BC-GN) assay (CE-IVD version) for identification of Gram-negative (GN) bacteria and detection of resistance genes. A total of 163 GN organisms (72 characterized strains and 91 clinical isolates from 86 patients) were tested; among the clinical isolates, 86 (94.5%) isolates were included in the BC-GN panel. For identification, the agreement was 98.6% (146/148, 95% confidence interval [CI], 92.1-100) and 70% (7/10, 95% CI, 53.5-100) for monomicrobial and polymicrobial cultures, respectively. Of the 48 resistance genes harbored by 43 characterized strains, all were correctly detected. Of the 19 clinical isolates harboring resistance genes, 1 CTX-M-producing Escherichia coli isolated in polymicrobial culture was not detected. Overall, BC-GN assay provides acceptable accuracy for rapid identification of Gram-negative bacteria and detection of resistance genes, compared with routine laboratory methods despite that it has limitations in the number of genus/species and resistance gene included in the panel and it shows lower sensitivity in polymicrobial cultures.

The Danish Collaborative Bacteraemia Network (DACOBAN) database.

The Danish Collaborative Bacteraemia Network (DACOBAN) research database includes microbiological data obtained from positive blood cultures from a geographically and demographically well-defined population serviced by three clinical microbiology departments (1.7 million residents, 32% of the Danish population). The database also includes data on comorbidity from the Danish National Patient Registry, vital status from the Danish Civil Registration System, and clinical data on 31% of nonselected records in the database. Use of the unique civil registration number given to all Danish residents enables linkage to additional registries for specific research projects. The DACOBAN database is continuously updated, and it currently comprises 39,292 patients with 49,951 bacteremic episodes from 2000 through 2011. The database is part of an international network of population-based bacteremia registries from five developed countries on three continents. The main purpose of the DACOBAN database is to study surveillance, risk, and prognosis. Sex- and age-specific data on background populations enables the computation of incidence rates. In addition, the high number of patients facilitates studies of rare microorganisms. Thus far, studies on Staphylococcus aureus, enterococci, computer algorithms for the classification of bacteremic episodes, and prognosis and risk in relation to socioeconomic factors have been published.