scholarly journals Blood Culture Testing Outcomes among Non-Malarial Febrile Children at Antimicrobial Resistance Surveillance Sites in Uganda, 2017–2018

2021 ◽  
Vol 6 (2) ◽  
pp. 71
Author(s):  
Rogers Kisame ◽  
Robinah Najjemba ◽  
Johan van Griensven ◽  
Freddy Eric Kitutu ◽  
Kudakwashe Takarinda ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Blood Culture ◽  
Patient Outcomes ◽  
Diagnostic Yield ◽  
Diagnostic Testing ◽  
Process Outcomes ◽  
Testing Data ◽  
Process Documentation ◽  
Pathogen Recovery ◽  
Infants And Young Children

Blood culture (BC) processes are critical to the utility of diagnostic testing, bloodstream infection (BSI) management, and antimicrobial resistance (AMR) surveillance. While Uganda has established BC guidelines, often laboratory practice does not meet the desired standards. This compromises pathogen recovery, reliability of antimicrobial susceptibility testing, and diagnostic test utility. This study assessed laboratory BC process outcomes among non-malarial febrile children below five years of age at five AMR surveillance sites in Uganda between 2017 and 2018. Secondary BC testing data was reviewed against established standards. Overall, 959 BC specimens were processed. Of these, 91% were from female patients, neonates, infants, and young children (1–48 months). A total of 37 AMR priority pathogens were identified; Staphylococcus aureus was predominant (54%), followed by Escherichia coli (19%). The diagnostic yield was low (4.9%). Only 6.3% of isolates were identified. AST was performed on 70% (18/26) of identified AMR priority isolates, and only 40% of these tests adhered to recommended standards. Interventions are needed to improve laboratory BC practices for effective patient management through targeted antimicrobial therapy and AMR surveillance in Uganda. Further research on process documentation, diagnostic yield, and a review of patient outcomes for all hospitalized febrile patients is needed.

scholarly journals 847. The Effect of Antimicrobial Administration on Blood Culture Positivity in Patients with Severe Manifestations of Sepsis

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S15-S16
Author(s):  
Matthew P Cheng ◽  
Robert Stenstrom ◽  
Katryn Paquette ◽  
Sarah Stabler ◽  
Murtaza Akhter ◽  
...  
Keyword(s):  
Blood Culture ◽  
Antimicrobial Therapy ◽  
Diagnostic Yield ◽  
The United States ◽  
Blood Cultures ◽  
Microbial Pathogens ◽  
Absolute Difference ◽  
Recovery Rates ◽  
Pathogen Recovery ◽  
Antimicrobial Administration

Abstract Background Current guidelines recommend obtaining blood cultures prior to antimicrobial therapy in patients with sepsis. Administering antimicrobials immediately without waiting for blood cultures could potentially decrease time to treatment and improve outcomes, but it is unclear the degree to which this strategy impacts diagnostic yield. Methods We performed a patient-level, single-arm, diagnostic trial. Seven urban emergency departments affiliated with academic medical centers across Canada and the United States participated in the study. Adults ≥18 years of age presenting to the emergency department with evidence of severe manifestations of sepsis, including a systolic blood pressure <90 mmHg and/or a serum lactate ≥4 mmol/L were included. Study participants had 2 sets of blood cultures drawn prior to and immediately following antimicrobial administration. The primary outcome was the difference in blood culture pathogen recovery rates before and after administration of antimicrobial therapy. Results Of the 3,164 participants screened, 325 were included in the study (mean age, 65.6 years; 63.0% men) and had repeat blood cultures drawn after the initiation of antimicrobial therapy (median time of 70 minutes, IQR 50 to 110 minutes). Pre-antimicrobial blood cultures were positive for one or more microbial pathogens in 102/325 (31.4%) patients. Fifty-four participants (52.9%) had matching blood culture results after initiation of antimicrobial treatment. The absolute difference in pathogen recovery rates was 14.5% ([95% CI 8.0 to 21.0%]; P < 0.0001) between pre- and post-antimicrobial blood cultures. Results were consistent in an analysis of the per-protocol population (absolute difference, 13.3% [95% CI 6.1 to 20.4%]; P < 0.0001). Including the results of other microbiological cultures done as part of routine care, microbial pathogens were recovered in 69 of 102 (67.7%) participants (absolute difference, 10.2% [95% CI 3.4 to 16.8%]; P < 0.0001). Conclusion Among patients with severe manifestations of sepsis, the administration of empiric antimicrobial therapy significantly reduces the yield of pathogen recovery when blood cultures are drawn shortly after treatment initiation. Disclosures All Authors: No reported Disclosures.

Mycoplasma genitalium: test positivity in syndromic presentations and clinicians’ adherence with treatment guidelines at Sydney Sexual Health Centre

2021 ◽  
pp. 095646242110225
Author(s):  
Calvin Cheung ◽  
Christopher Bourne ◽  
Rick Varma
Keyword(s):  
Decision Making ◽  
Antimicrobial Resistance ◽  
Sexual Health ◽  
Treatment Guidelines ◽  
Health Centre ◽  
Diagnostic Testing ◽  
Mycoplasma Genitalium ◽  
Macrolide Resistance ◽  
Adherence To Treatment ◽  
Real World Data

Mycoplasma genitalium ( M.genitalium) is associated with urethritis, cervicitis, pelvic inflammatory disease, proctitis and epididymitis. Its treatment is complicated by antimicrobial resistance. To assess clinicians’ adherence to M.genitalium diagnostic testing recommendations for syndromic presentations, as well as resistance-guided management of M.genitalium at Sydney Sexual Health Centre, we reviewed patients presenting between August and December 2018. 349/372 (94%) syndromic presentations were tested for M.genitalium with 16% M.genitalium test positivity and 81% macrolide resistance. 16/27 (59%) macrolide-sensitive infections and 65/77 (84%) macrolide-resistant infections received resistance-guided treatment. Tests of cure (TOCs) were unnecessarily ordered for 82% macrolide-sensitive cases, while 88% macrolide-resistant cases were correctly ordered TOCs. Co-existing STIs at the time of macrolide-sensitive ( p = 0.30) or macrolide-resistant M.genitalium ( p = 0.94) diagnosis did not significantly affect adherence to treatment guidelines. This study confirms the expected prevalence of M.genitalium and macrolide resistance in syndromic presentations while our real-world data highlight the decision-making challenges involved with managing M.genitalium, offering insights for further research.

Diversity and Prevailing Antimicrobial Resistance in Blood Stream Pathogens at NICU of a Tertiary Care Institute in Kolkata

2021 ◽  
Vol 8 (14) ◽  
pp. 854-860
Author(s):  
Nandita Pal ◽  
Sanat Kumar Dolui ◽  
Bhuban Majhi ◽  
Manisha Das
Keyword(s):  
Antimicrobial Resistance ◽  
Blood Culture ◽  
Klebsiella Pneumoniae ◽  
Late Onset ◽  
Tertiary Care ◽  
Burkholderia Cepacia Complex ◽  
Multi Drug Resistance ◽  
Coagulase Negative Staphylococcus ◽  
Neonatal Septicaemia ◽  
Early Onset Sepsis

BACKGROUND Septicaemia is a leading cause of morbidity and mortality in neonates. Antimicrobial prescription for neonatal septicaemia (NS) should be wisely used depending on the regional spectra of infecting microbes and their antimicrobial resistance (AMR) patterns which vary over time, place and host factors. METHODS This was a descriptive cross-sectional study conducted from January 2017 to June 2017 among 102 cases of NS admitted in neonatal intensive care units (NICU). Demographic data of the concerned neonates was collected through a predesigned checklist. Blood collected from neonates was processed for culture and sepsis screen. Antimicrobial susceptibility testing was performed for the cultured isolates and phenotypic AMR patterns were observed. RESULTS Most of the studied neonates had subnormal birth weight (86). Blood culture showed predominance of gram-negative bacilli (GNB) among which Klebsiella pneumoniae was the most common followed by Burkholderia cepacia complex. Coagulase negative staphylococcus species (CoNS) was most numerous among the cultured gram-positive cocci (GPC). Candidemia of late onset was confirmed in 25 neonates. Early onset sepsis was predominantly caused by GNB (29 / 31) predominated by Klebsiella pneumoniae. Multi drug resistant (MDR) pathogens viz., extended spectrum beta-lactamase (ESBL) producing GNB, carbapenem resistant-GNB (CR-GNB) and methicillin resistant staphylococcus (MRS) phenotypes were notably prevalent. CONCLUSIONS Multidrug resistant microorganisms are notably prevalent in neonatal septicaemia. In clinically suspected very sick septicaemic neonates not showing much improvement with guideline-led empirical antimicrobial therapy, an early antibiogram guided change over to meropenem and amikacin or even to colistin in case of reported carbapenem resistance, is highly recommended. KEYWORDS Antimicrobial Resistance, Blood Culture, Multi Drug Resistance, Neonatal Septicaemia

scholarly journals Emergence of high rates of antimicrobial resistance among viridans group streptococci in the United States.

1996 ◽  
Vol 40 (4) ◽  
pp. 891-894 ◽  
Author(s):  
G V Doern ◽  
M J Ferraro ◽  
A B Brueggemann ◽  
K L Ruoff
Keyword(s):  
United States ◽  
Antimicrobial Resistance ◽  
Blood Culture ◽  
Rank Order ◽  
The United States ◽  
Medical Centers ◽  
Streptococcus Milleri ◽  
Intermediate Resistance ◽  
High Level ◽  
Viridans Group Streptococci

Three hundred fifty-two blood culture isolates of viridans group streptococci obtained from 43 U.S. medical centers during 1993 and 1994 were characterized. Included were 48 isolates of "Streptococcus milleri," 219 S. mitis isolates, 29 S. salivarius isolates, and 56 S. sanguis isolates. High-level penicillin resistance (MIC, > or = 4.0 micrograms/ml) was noted among 13.4% of the strains; for 42.9% of the strains, penicillin MICs were 0.25 to 2.0 micrograms/ml (i.e., intermediate resistance). In general, amoxicillin was slightly more active than penicillin. The rank order of activity for five cephalosporins versus viridans group streptococci was cefpodoxime = ceftriaxone > cefprozil = cefuroxime > cephalexin. The percentages of isolates resistant (MIC, > or = 2 micrograms/ml) to these agents were 15, 17, 18, 20, and 96, respectively. The rates of resistance to erythromycin, tetracycline, and trimethoprim-sulfamethoxazole were 12 to 38%. Resistance to either chloramphenicol or ofloxacin was uncommon (i.e., < 1%). In general, among the four species, S. mitis was the most resistant and "S. milleri" was the most susceptible.

scholarly journals 1439. High Rates of Drug Resistance in Escherichia coli from a Pilot Antimicrobial Resistance Surveillance System in Cambodia

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S723-S724
Author(s):  
Vandarith Nov ◽  
Darapheak Chau ◽  
Chhorvann Chhea
Keyword(s):  
Public Health ◽  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antimicrobial Resistance ◽  
Blood Culture ◽  
Klebsiella Pneumoniae ◽  
Surveillance System ◽  
Methicillin Resistance ◽  
Salmonella Typhi ◽  
Blood Samples

Abstract Background Antimicrobial resistance (AMR) is a major and growing global public health problem. The Cambodia Ministry of Health established a pilot laboratory-based AMR surveillance system for blood specimens in 2017. The objective of this study is to characterize AMR among pathogenic isolates from blood samples. Methods A retrospective analysis was performed using one year of data from a pilot AMR Surveillance system in Cambodia. Four blood culture isolate pathogens were included: Escherichia coli, Klebsiella pneumoniae, Salmonella Typhi /Salmonella Paratyphi A and Staphylococcus aureus. Blood culture isolates that were referred from eleven sentinel sites were analyzed at the National Public Health Laboratory for identification. Antibiotic susceptibility testing (AST) was done using disk diffusion, minimum inhibitory concentration method following Clinical Laboratory Standard Institute (CLSI) guidelines. Results Among 214 pathogenic isolates from blood samples, E.coli was the most common (56.1%), followed by Salmonella Typhi/Salmonella Paratyphi A (18.7%), Staphylococcus aureus (13.5%), and Klebsiella pneumoniae (11.7%). Methicillin Resistance Staphylococcus aureus (MRSA) was detected in half of the isolates. E.coli was resistant to ampicillin (94.4%), trimethoprim-sulfamethoxazole (84.5%), and ceftriaxone (79.2%). Salmonella Typhi was resistant to ampicillin (73.3%) and trimethoprim-sulfamethoxazole (60.0%) and Salmonella Paratyphi A were resistant to fluoroquinolones (91.7%). For last resort antibiotics, E.coli was resistant to carbapenem groups (3.2% for imipenem, 4.9% for meropenem, and 5.0% for ertapenem). Klebsiella pneumoniae was not resistant to any groups. Conclusion E.coli was found at high rates in blood samples and was resistant to common antibiotics used in Cambodia. These pilot data show the importance of establishing a national AMR surveillance system in Cambodia to monitor AMR trends following GLASS guidelines. Disclosures All Authors: No reported disclosures

scholarly journals Large-scale assessment of antimicrobial resistance marker databases for genetic phenotype prediction: a systematic review

2020 ◽  
Vol 75 (11) ◽  
pp. 3099-3108
Author(s):  
Norhan Mahfouz ◽  
Inês Ferreira ◽  
Stephan Beisken ◽  
Arndt von Haeseler ◽  
Andreas E Posch
Keyword(s):  
Antimicrobial Resistance ◽  
Clinical Microbiology ◽  
Large Scale ◽  
Diagnostic Testing ◽  
Error Rates ◽  
Systematic Evaluation ◽  
Large Scale Assessment ◽  
Performance Requirements ◽  
Public Repositories ◽  
The Right

Abstract Background Antimicrobial resistance (AMR) is a rising health threat with 10 million annual casualties estimated by 2050. Appropriate treatment of infectious diseases with the right antibiotics reduces the spread of antibiotic resistance. Today, clinical practice relies on molecular and PCR techniques for pathogen identification and culture-based antibiotic susceptibility testing (AST). Recently, WGS has started to transform clinical microbiology, enabling prediction of resistance phenotypes from genotypes and allowing for more informed treatment decisions. WGS-based AST (WGS-AST) depends on the detection of AMR markers in sequenced isolates and therefore requires AMR reference databases. The completeness and quality of these databases are material to increase WGS-AST performance. Methods We present a systematic evaluation of the performance of publicly available AMR marker databases for resistance prediction on clinical isolates. We used the public databases CARD and ResFinder with a final dataset of 2587 isolates across five clinically relevant pathogens from PATRIC and NDARO, public repositories of antibiotic-resistant bacterial isolates. Results CARD and ResFinder WGS-AST performance had an overall balanced accuracy of 0.52 (±0.12) and 0.66 (±0.18), respectively. Major error rates were higher in CARD (42.68%) than ResFinder (25.06%). However, CARD showed almost no very major errors (1.17%) compared with ResFinder (4.42%). Conclusions We show that AMR databases need further expansion, improved marker annotations per antibiotic rather than per antibiotic class and validated multivariate marker panels to achieve clinical utility, e.g. in order to meet performance requirements such as provided by the FDA for clinical microbiology diagnostic testing.

scholarly journals Predictors of Time to Effective and Optimal Antimicrobial Therapy in Patients With Positive Blood Cultures Identified via Molecular Rapid Diagnostic Testing

2018 ◽  
Vol 6 (1) ◽  
Author(s):  
Maya Beganovic ◽  
Tristan T Timbrook ◽  
Sarah M Wieczorkiewicz
Keyword(s):  
Patient Outcomes ◽  
Antimicrobial Stewardship ◽  
Antimicrobial Therapy ◽  
Diagnostic Tests ◽  
Diagnostic Testing ◽  
Rapid Diagnostic Tests ◽  
Blood Cultures ◽  
Effective Therapy ◽  
Rapid Diagnostic Testing ◽  
Optimal Therapy

Abstract Antimicrobial stewardship (AMS) programs integrated with rapid diagnostic tests optimize patient outcomes and reduce time to effective therapy (TTET) and time to optimal therapy (TTOT). This study identifies predictors of TTET and TTOT among patients with positive blood cultures and identifies limitations to current TTOT definitions and outcomes.

scholarly journals 701. Rapid Detection of Antimicrobial Resistance Determinants with the BioFire® System

2018 ◽  
Vol 5 (suppl_1) ◽  
pp. S252-S253
Author(s):  
Stefanie Marxreiter ◽  
Eric Lo ◽  
Cody Oswald ◽  
Aubrie Hopper ◽  
Becki Barr ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Blood Culture ◽  
Clinical Samples ◽  
Intrinsic Resistance ◽  
Correct Treatment ◽  
Phenotypic Data ◽  
University Of Utah ◽  
Children's Hospital ◽  
Children’S Hospital ◽  
Research Grant

Abstract Background Antimicrobial-resistant (AMR) bacteria are a rising healthcare concern and are associated with an estimated five-fold increase in mortality for infected patients. Correct treatment requires antimicrobial susceptibility knowledge, but standard testing methods require multiple days for an accurate phenotype. Rapid identification of AMR immediately after blood culture positivity could potentially improve health outcomes, lower economic cost, prevent the spread of multidrug-resistant outbreaks and assist with antimicrobial stewardship goals. Methods The BioFire® Antimicrobial Resistance (AMR) Panel is a research use only multiplex-nested PCR system with 47 assays for 30 genes conferring resistance to cephalosporins, carbapenems, aminoglycosides, and fluoroquinolones which can be found in E. coli, K. pneumoniae, P. aeruginosa, A. baumannii, and E. cloacae complex. We tested 86 residual positive blood culture samples collected from Primary Children’s Hospital, University of Utah Hospital and Huntsman Cancer Hospital with the BioFire AMR Panel. Molecular genotypic results were compared with phenotypic susceptibility information for each blood culture specimen to confirm resistance detections. Results Of the 86 samples tested, there were 33 cultures phenotypically resistant (beyond intrinsic resistance) to at least one antibiotic class targeted by the panel. BioFire AMR Panel identified resistance to gentamicin, cefoxitin, all penicillins tested, and ciprofloxacin with 100% positive predictive value (PPV). For tobramycin, ceftazidime, and ceftriaxone, the PPV was greater than 85%. Carbapenem resistance was not detected, likely due to the low number of resistant organisms present in our patient population. Conclusion The BioFire AMR Panel provides identification of genetic AMR determinants in a rapid, easy-to-use system that accurately correlates with phenotypic data for specific antimicrobials. Studies will continue to test additional clinical samples at various geographical locations to further evaluate the relationship between genotypic and phenotypic resistance assessment. Data presented is from an assay that has not been cleared or approved by US FDA or other regulatory agencies for in vitro diagnostic use. Disclosures S. Marxreiter, NIH NIAID: Grant Investigator, Research grant. BioFire Diagnostics, LLC: Employee, Salary. E. Lo, BioFire Diagnostics, LLC: Employee, Salary. NIH NIAID: Grant Investigator, Research grant. C. Oswald, BioFire Diagnostics, LLC: Employee, Salary. NIH NIAID: Grant Investigator, Research grant. A. Hopper, Primary Children’s Hospital: Investigator, Research grant. B. Barr, Primary Children’s Hospital: Grant Investigator, Research grant. J. A. Daly, Primary Children’s Hospital: Grant Investigator, Research grant. University of Utah: Grant Investigator, Research grant. C. C. Ginocchio, Biomerieux: Employee, Salary. R. Crisp, BioFire Diagnostics, LLC: Employee, Salary. A. Hemmert, BioFire Diagnostics, LLC: Employee, Salary. NIH NIAID: Grant Investigator, Research grant.

scholarly journals A SYSTEMATIC EVALUATION OF RAPID DOT-EIA, BLOOD CULTURE AND WIDAL TEST IN THE DIAGNOSIS OF TYPHOID FEVER

2013 ◽  
Vol 03 (01) ◽  
pp. 21-24 ◽  
Author(s):  
H. Sanjeev ◽  
Sweetha Nayak ◽  
Pai Asha K. B. ◽  
Rai Rekha ◽  
Vimal Karnaker ◽  
...  
Keyword(s):  
Blood Culture ◽  
Typhoid Fever ◽  
Gold Standard ◽  
Salmonella Enterica ◽  
Diagnostic Yield ◽  
Far East ◽  
Health Care Facilities ◽  
Systematic Evaluation ◽  
Widal Test ◽  
South Central

Abstract Background: Typhoid fever, caused by Salmonella enterica serotype Typhi, is endemic in the Indian sub-continent including Bangladesh, South-east and Far-east Asia, Africa and South Central America. The disease can occur in all age group with highest incidence among children. Blood culture is regarded as the gold standard for diagnosis and carry 70-75% diagnostic yield in the first week of illness. However, this requires laboratory equipment and technical training that are beyond the means of most primary health care facilities in the developing world. Typhidot is a rapid dot-enzyme immune assay (EIA), which detects IgG and IgM antibodies to a specific 50 kD outer membrane protein (OMP) antigen of Salmonella enterica serotype Typhi. Typhidot becomes positive as early as in the first week of fever. The results can be visually interpreted and is available within one hour. Materials and method: Fifty blood samples, collected aseptically from patients clinically diagnosed of Typhoid fever, were evaluated by blood culture, Widal test and Typhidot. Results: Of the 50 patients, 33 (66%) were positive by blood culture. Widal test was positive in 33(66%) patients which included 26 in blood culture positive patients and 7 in blood culture negative patients. Typhidot was positive in 37 (74%) patients. Thus, in comparison to the gold standard test i:e blood culture, Typhidot and Widal test had sensitivity and specificity of 100% & 76% and 78.78% & 58.82% respectively. Conclusion: Typhidot is found to have high sensitivity and good specificity and could be applied as a good alternate in resource poor nation. Further, it is simple to perform, reliable when compared to Widal test, and rapid, with results being available in one hour when compared to 48 hours for blood culture and 18 hours for Widal test.

scholarly journals Antimicrobial Resistance Patterns of Bacterial Isolates from Blood Culture among HIV/AIDS Patients at Felege Hiwot Referral Hospital, Northwest Ethiopia

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
Mohabaw Jemal ◽  
Teshiwal Deress ◽  
Teshome Belachew ◽  
Yesuf Adem
Keyword(s):  
Antimicrobial Resistance ◽  
Blood Culture ◽  
Agar Plate ◽  
Blood Cultures ◽  
Resistant Bacteria ◽  
Bacterial Isolates ◽  
Aids Patients ◽  
Gram Positive ◽  
Gram Negative ◽  
Hiv Aids

Background. The emergence and spread of antimicrobial resistance in bacteria is recognized as a global public health problem. Bloodstream infection with antimicrobial-resistant bacteria in HIV/AIDS patients makes the problem more challenging. So, regular and periodic diagnosis and use of the appropriate antimicrobial susceptibility pattern determination is the only option for decreasing the prevalence and development of drug-resistant bacteria. Methods. An institution-based cross-sectional study was conducted among 384 HIV/AIDS patients. Sociodemographic data of patients were recorded using structured questionnaires. Blood cultures were collected with BACTEC aerobic blood culture bottles. A pair of samples was collected from each patient aseptically and incubated at 37°. If samples are positive for bacterial agents, they were subcultured to solid media such as blood agar plate, chocolate agar plate, and MacConkey agar plates. Identification was performed using colony characteristics and standard biochemical techniques. The antimicrobial susceptibility test was determined by the Kirby–Bauer disc diffusion method. Data entry and analysis were performed while using SPSS version 20. Descriptive statistics were performed to calculate frequencies. Results. Altogether, 384 patients were included, and 123 blood cultures were positive, so that the yield was thus 32%. About 46 (37.4%) of Gram-negative and 77 (62.6%) of Gram-positive bacterial species were identified. Among Gram-negative bacterial isolates, K. pneumoniae was the leading pathogen, 19 (41.3%), whereas S. aureus, 38 (49.4%), was predominant among Gram-positive isolates. In his study, the majority of Gram-positive isolates showed high level of resistance to penicillin, 72 (95.5%), tetracycline, 55 (71.4%), and cotrimoxazole, 45 (58.4%). About 28 (73.6%) of S. aureus isolates were also methicillin-resistant. Gram-negative bacterial isolates also showed a high resistance to ampicillin (91.3%), tetracycline (91.3%), and gentamicin (47.8%). Overall, about 78% of multidrug resistance was observed. Conclusion. Several pathogens were resistant to greater than five antimicrobial agents, so that proper management of patients with bacteremia is needed, and a careful selection of effective antibiotics should be practiced.

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