Identification and Susceptibility Testing of Enterobacteriaceae and Pseudomonas aeruginosa by Direct Inoculation from Positive BACTEC Blood Culture Bottles into Vitek 2

PubMed Central

Bruins, Marjan J.; Bloembergen, Peter; Ruijs, Gijs J. H. M.; Wolfhagen, Maurice J. H. M.

2004-01-01

Inoculation of an automated system for rapid identification (ID) and antimicrobial susceptibility testing (AST) directly from positive blood culture bottles will reduce the turnaround time of laboratory diagnosis of septicemic patients, which benefits clinical outcome and decreases patient costs. Direct test results, however, must always be confirmed by testing a pure overnight culture, which is the “gold standard.” We studied the accuracy of direct testing versus repeat testing in order to investigate the possibility of refraining from repeat testing. We also assessed the clinical risk of reporting results based on direct testing only. We inoculated Vitek 2 (bioMérieux) directly from 410 positive BACTEC 9240 (BD) blood culture bottles containing gram-negative rods and studied the ID and AST results. In a comparison of direct inoculation with the standard method, a total of 344 isolates of Enterobacteriaceae and Pseudomonas aeruginosa were tested, and 93.0% were correctly identified. Of the 39 (10.2%) samples that contained bacilli not identifiable by Vitek 2, only 1 gave a conclusive, correct result. The overall MIC agreement among 312 isolates was 99.2%, with 0.8% very major and 0.02% major error rates. Of only three (polymicrobial) samples, the direct susceptibility pattern would be reported to the clinician as too sensitive. Vitek 2 results obtained from direct inoculation of blood culture bottles containing gram-negative bacilli are safe enough for immediate reporting, provided that ID and AST are consistent. Repeat testing is not necessary, unless Gram stain or overnight subculture results raise doubt about the purity of the culture. PMID:14715724

Diagnosis of brucellosis by using blood cultures.

PubMed Central

Ruiz, J; Lorente, I; Pérez, J; Simarro, E; Martínez-Campos, L

1997-01-01

The performances of three blood culture systems, Hemoline performance diphasic medium (bioMérieux, Marcy l'Etoile, France), Bactec Plus Aerobic/F* (Becton Dickinson, Paramus, N.J.), and Vital Aer (bioMérieux), were compared for the diagnosis of 17 cases of brucellosis. By using a 5-day incubation protocol, positive results were 52.9, 82.4, and 11.8%, respectively. When the protocol was extended to 7 days, the results were 76.5, 94.1, and 47.1%, respectively. Bactec was the fastest system (P < 0.05). PMID:9276429

[Blood stream infection and blood culture--"progress" and "blind" in blood culture testing].

PubMed

Kobayashi, Intetsu

2005-04-01

We have investigated various types of blood culture bottles which are mainly used at present and posed problems present in the blood culture bottles. First, there are differences between resin and ecosorb in the ability to adsorb and inactivate antibiotics in the blood. Second, the delay in placing the bottle (into which blood was inoculated) to the automatic instrument (delay in the start of incubation) greatly affects the automatic detection by BACTEC system and shows false negatives. Third, when the same blood is incubated in plural bottles (aerobic and anaerobic bottles), the differences among the detected organisms in the number are comparatively high, i.e., about 40%. In addition, there are differences among the organisms in the number of days required for the detection of the organisms. In this case, the detected organisms are clearly different in many cases. The technology of blood culture has been progressed remarkably. However, the efficiency of utilization of automatic instruments for diagnosis of infection depends greatly on the ability of laboratory technicians.

Evaluation of the BD BACTEC FX blood volume monitoring system as a continuous quality improvement measure.

PubMed

Coorevits, L; Van den Abeele, A-M

2015-07-01

The yield of blood cultures is proportional to the volume of blood cultured. We evaluated an automatic blood volume monitoring system, recently developed by Becton Dickinson within its BACTEC EpiCenter module, that calculates mean volumes of negative aerobic bottles and generates boxplots and histograms. First, we evaluated the filling degree of 339 aerobic glass blood cultures by calculating the weight-based volume for each bottle. A substantial amount of the bottles (48.3%) were inadequately filled. Evaluation of the accuracy of the monitoring system showed a mean bias of -1.4 mL (-15.4%). Additional evaluation, using the amended software on 287 aerobic blood culture bottles, resulted in an acceptable mean deviation of -0.3 mL (-3.3%). The new software version was also tested on 200 of the recently introduced plastic bottles, which will replace the glass bottles in the near future, showing a mean deviation of +2.8 mL (+26.7%). In conclusion, the mean calculated volumes can be used for the training of a single phlebotomist. However, filling problems appear to be masked when using them for phlebotomist groups or on wards. Here, visual interpretation of boxplots and histograms can serve as a useful tool to observe the spread of the filling degrees and to develop a continuous improvement program. Re-adjustment of the software has proven to be necessary for use with plastic bottles. Due to our findings, BD has developed further adjustments to the software for validated use with plastic bottles, which will be released soon.

Efficacy of BACTEC TB in the rapid confirmatory diagnosis of mycobacterial infections. A Lebanese tertiary care center experience.

PubMed

Itani, Lina Y; Cherry, Mohamad A; Araj, George F

2005-01-01

Rapid detection of Mycobacterium tuberculosis (MTB), especially multidrug-resistant strains, is of importance for prompt clinical management and initiation of public health control measures. Culture remains the "gold" standard in the confirmatory laboratory diagnosis of mycobacterial infections. The reliability of the automated radiometric BACTEC 460 TB (BACTEC) system for the rapid detection of mycobacteria in clinical specimens was evaluated and compared to the conventional culture on Lowenstein-Jensen (LJ) medium. All clinical specimens submitted for mycobacterial culture were processed and simultaneously cultured on both BACTEC broth medium and LJ solid medium. Acid-fast bacilli (AFB) smears were also performed on the sediments. Differentiation of mycobacterial isolates as MTB or Mycobacterium sp. other than tuberculosis (MOTT) was based on the BACTEC NAP test. All positive culture findings recovered between January 1997 and December 2003 were analyzed in this study. A total of 3300 specimens were tested of which 355 (10.7%) yielded positive cultures consisting of 233 (65.6%) MTB and 122 (34.4%) MOTT. The percentages of AFB smear-positive were 45% and 49% in clinical specimens yielding MTB & MOTT, respectively. Though several types of specimens were cultured, most isolates (72% of MTB & 91% of MOTT) were recovered from respiratory specimens. Overall, the BACTEC showed significantly higher mycobacteria recovery rate (91%) than LJ (77%). In terms of times to detection, BACTEC showed significantly shorter detection time of isolates than LJ for the overall (mean 9.6 days for BACTEC vs. 22.8 days for LJ) and for each category of AFB smear finding. The detection time is shortened for BACTEC with the increasing grade of smear positivity. BACTEC is substantially more sensitive, efficient and rapid than LJ in the laboratory diagnosis of mycobacterial infections. This system also provides rapid differentiation of MTB from MOTT and susceptibility test results on MTB

Comparison of Susceptibility Testing of Mycobacterium tuberculosis Using the ESP Culture System II with That Using the BACTEC Method

PubMed Central

Ruiz, P.; Zerolo, F. J.; Casal, M. J.

2000-01-01

The ESP Culture System II was evaluated for its capacity to test the susceptibility of 389 cultures of Mycobacterium tuberculosis to streptomycin, rifampin, ethambutol, and isoniazid. Good agreement with results with the BACTEC TB 460 was found. ESP II is a reliable, rapid, and automated method for performing susceptibility testing. PMID:11101619

Detection of Salmonella spp. with the BACTEC 9240 Automated Blood Culture System in 2008 - 2014 in Southern Iran (Shiraz): Biogrouping, MIC, and Antimicrobial Susceptibility Profiles of Isolates.

PubMed

Anvarinejad, Mojtaba; Pouladfar, Gholam Reza; Pourabbas, Bahman; Amin Shahidi, Maneli; Rafaatpour, Noroddin; Dehyadegari, Mohammad Ali; Abbasi, Pejman; Mardaneh, Jalal

2016-04-01

Human salmonellosis continues to be a major international problem, in terms of both morbidity and economic losses. The antibiotic resistance of Salmonella is an increasing public health emergency, since infections from resistant bacteria are more difficult and costly to treat. The aims of the present study were to investigate the isolation of Salmonella spp. with the BACTEC automated system from blood samples during 2008 - 2014 in southern Iran (Shiraz). Detection of subspecies, biogrouping, and antimicrobial susceptibility testing by the disc diffusion and agar dilution methods were performed. A total of 19 Salmonella spp. were consecutively isolated using BACTEC from blood samples of patients between 2008 and 2014 in Shiraz, Iran. The isolates were identified as Salmonella, based on biochemical tests embedded in the API-20E system. In order to characterize the biogroups and subspecies, biochemical testing was performed. Susceptibility testing (disc diffusion and agar dilution) and extended-spectrum β-lactamase (ESBL) detection were performed according to the clinical and laboratory standards institute (CLSI) guidelines. Of the total 19 Salmonella spp. isolates recovered by the BACTEC automated system, all belonged to the Salmonella enterica subsp. houtenae. Five isolates (26.5%) were resistant to azithromycin. Six (31.5%) isolates with the disc diffusion method and five (26.3%) with the agar dilution method displayed resistance to nalidixic acid (minimum inhibitory concentration [MIC] > 32 μg/mL). All nalidixic acid-resistant isolates were also ciprofloxacin-sensitive. All isolates were ESBL-negative. Twenty-one percent of isolates were found to be resistant to chloramphenicol (MIC ≥ 32 μg/mL), and 16% were resistant to ampicillin (MIC ≥ 32 μg/mL). The results indicate that multidrug-resistant (MDR) strains of Salmonella are increasing in number, and fewer antibiotics may be useful for treating S. enterica infections. Routine investigation and reporting

Direct identification by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) from positive blood culture bottles: An opportunity to customize growth conditions for fastidious organisms causing bloodstream infections.

PubMed

Sharma, Megha; Gautam, Vikas; Mahajan, Monika; Rana, Sudesh; Majumdar, Manasi; Ray, Pallab

2017-10-01

Culture-negative bacteraemia has been an enigmatic entity with respect to its aetiological agents. In an attempt to actively identify those positive blood cultures that escape isolation and detection on routine workflow, an additional step of MALDI-TOF MS (matrix-assisted laser desorption ionization-time of flight mass spectrometry) based detection was carried out directly from the flagged blood culture bottles. Blood samples from 200 blood culture bottles that beeped positive with automated (BACTEC) system and showed no growth of organism on routine culture media, were subjected to analysis by MALDI-TOF MS. Forty seven of the 200 (23.5%) bacterial aetiology could be established by bottle-based method. Based on these results, growth on culture medium could be achieved for the isolates by providing special growth conditions to the fastidious organisms. Direct identification by MALDI-TOF MS from BACTEC-positive bottles provided an opportunity to isolate those fastidious organisms that failed to grow on routine culture medium by providing them with necessary alterations in growth environment.

Development and Validation of a Liquid Medium (M7H9C) for Routine Culture of Mycobacterium avium subsp. paratuberculosis To Replace Modified Bactec 12B Medium

PubMed Central

Whittington, Ann-Michele; Waldron, Anna; Begg, Douglas J.; de Silva, Kumi; Purdie, Auriol C.; Plain, Karren M.

2013-01-01

Liquid culture of Mycobacterium avium subsp. paratuberculosis from clinical samples, such as feces, is the most sensitive antemortem test for the diagnosis of Johne's disease in ruminants. In Australia, New Zealand, the United States, and some other countries, the Bactec 460 system with modified Bactec 12B medium (Becton, Dickinson) has been the most commonly used liquid culture system, but it was discontinued in 2012. In this study, a new liquid culture medium, M7H9C, was developed. It consists of a Middlebrook 7H9 medium base with added Casitone, albumin, dextrose, catalase, egg yolk, mycobactin J, and a cocktail of antibiotics. We found that polyoxyethylene stearate (POES) was not essential for the cultivation of M. avium subsp. paratuberculosis in either the Bactec 12B or the M7H9C medium. The limit of detection determined using pure cultures of the C and S strains of M. avium subsp. paratuberculosis was 7 bacilli per 50 μl inoculum in the two media. The new medium was validated using 784 fecal and tissue samples from sheep and cattle, >25% of which contained viable M. avium subsp. paratuberculosis. Discrepant results for the clinical samples between the two media were mostly associated with samples that contained <10 viable bacilli per gram, but these results were relatively uncommon, and the performances of the two media were not significantly different. M7H9C medium was less than half the cost of the Bactec 12B medium and did not require regular examination during incubation, but a confirmatory IS900 PCR test had to be performed on every culture after the predetermined incubation period. PMID:24048541

Evaluation of the BD Max StaphSR Assay for Rapid Identification of Staphylococcus aureus and Methicillin-Resistant S. aureus in Positive Blood Culture Broths

PubMed Central

Hofko, Marjeta; Hamilton, Fiona; Mackenzie, Laura; Zimmermann, Stefan; Templeton, Kate

2015-01-01

We evaluated the performance of the BD Max StaphSR assay for the direct detection of Staphylococcus aureus from blood culture medium. In a two-center trial, 155 blood cultures from the BD Bactec FX system and 212 from the bioMérieux BacT/Alert system were tested; 170 bottles yielded S. aureus, and all were identified correctly by the BD Max StaphSR assay. The assay required approximately 2.5 h, thus allowing rapid identification of blood cultures flagged positive. PMID:26292311

Rapid Detection and Identification of Candidemia by Direct Blood Culturing on Solid Medium by Use of Lysis-Centrifugation Method Combined with Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS).

PubMed

Idelevich, Evgeny A; Grünastel, Barbara; Becker, Karsten

2017-01-01

Candida sepsis is a life-threatening condition with increasing prevalence. In this study, direct blood culturing on solid medium using a lysis-centrifugation procedure enabled successful Candida species identification by matrix-assisted laser desorption-ionization time of flight mass spectrometry on average 3.8 h (Sabouraud agar) or 7.4 h (chocolate agar) before the positivity signal for control samples in Bactec mycosis-IC/F or Bactec Plus aerobic/F bottles, respectively. Direct culturing on solid medium accelerated candidemia diagnostics compared to that with automated broth-based systems. Copyright © 2016 American Society for Microbiology.

A dedicated fungal culture medium is useful in the diagnosis of fungemia: a retrospective cross-sectional study.

PubMed

Zheng, Shuwei; Ng, Tong Yong; Li, Huihua; Tan, Ai Ling; Tan, Thuan Tong; Tan, Ban Hock

2016-01-01

Mortality for candidemia ranges from 15% to 35%. Current guidelines recommend inoculating blood into three aerobic and three anaerobic blood culture bottles when candidemia is suspected, without mention of a fungal blood culture bottle. To determine the value of the BACTEC Myco/F Lytic blood culture media in the diagnosis of fungemia. A two-year retrospective cross-sectional study was performed for patients who had fungemia with submitted BACTEC Plus Aerobic/F (Aer), BACTEC Plus Anaerobic/F (Anaer) or Myco/F Lytic (Myco) blood culture bottles. The detection rate of fungemia was 77.4% in 93 patients with contemporaneously submitted blood culture bottles when limited to only Aer/Anaer culture results. The detection rate improved significantly with the addition of the Myco culture bottle results (p<0.0001). A logistic regression model showed that Myco culture bottle submissions were less useful for patients with appropriate anti-fungal therapy administered within 48 hours [OR = 0.18, 95% CI = (0.06, 0.49), p = 0.001] and those with fungal growth detected within 48 hours [OR = 0.33, 95% CI = (0.12, 0.89), p = 0.001]. Among a subset of patients with concordant blood culture results, those with Myco culture bottles submission allowed earlier fungal detection and speciation by at least one day in 27.5% and 25.0% of the cases respectively. Our study highlights the importance of a dedicated fungal blood culture when fungemia is clinically suspected. Nearly a quarter of fungemias may be missed if a fungal blood culture is not performed.

Performance of two blood culture systems to detect anaerobic bacteria. Is there any difference?

PubMed

Mueller-Premru, Manica; Jeverica, Samo; Papst, Lea; Nagy, Elisabeth

2017-06-01

We studied the performance characteristics of two blood culture (BC) bottles/systems, (i) BacT/ALERT-FN Plus/3D (bioMérieux, Marcy l'Étoile, France) and (ii) BACTEC-Lytic/9000 (Becton Dickinson, Sparks, USA) for detection of growth and time-to-positivity (TTP) against a balanced and diverse collection of anaerobic bacterial strains (n = 48) that included reference strains (n = 19) and clinical isolates (n = 29) of 32 species (15 Gram-negative and 17 Gram-positive). Standard suspension of bacteria was inoculated to each bottle in duplicates and incubated in the corresponding system. Overall, 62.5% (n = 30) of strains were detected by both BC bottle types. Comparing the two, 70.8% (n = 34) and 79.2% (n = 38) of strains were detected by BacT/ALERT-FN Plus and BACTEC-Lytic bottles, respectively (p = 0.38). Among Gram-negative anaerobes (n = 25) the detection rate was 76.0% (n = 19) vs. 92.0% (n = 23) (p = 0.22), respectively. Among Gram-positive anaerobes (n = 23) the detection rate was 65.2% (n = 15) in both bottles (p = 1). The average TTP per bottle was calculated only for the strains detected by both systems (n = 30) and was 40.85 h and 28.08 h for BacT/ALERT-FN Plus and BACTEC-Lytic, respectively (p < 0.001). The mean difference was 12.76 h (95% CI: 6.21-19-31 h). Six anaerobic strains were not detected by any system, including Gram-negative Porphyromonas gingivalis, and five Gram-positive strains: Finegoldia magna, Peptostreptococcus anaerobius, Propionibacterium acnes, Clostridium novyi and Clostridium clostridioforme. Furthermore, Eggerthella lenta and Prevotella bivia were detected only by BacT/ALERT-FN Plus, while Prevotella disiens and Prevotella intermedia were detected only by BACTEC-Lytic bottles. There were no major differences in detection rate among clinical and reference strains. Anaerobic bacteria represent a minority of BC isolates, however, far from ideal detection rate was observed in this study for both

Direct detection of methicillin resistance in Staphylococcus aureus in blood culture broth by use of a penicillin binding protein 2a latex agglutination test.

PubMed

Qian, Qinfang; Venkataraman, Lata; Kirby, James E; Gold, Howard S; Yamazumi, Toshiaki

2010-04-01

We studied the utility of performing a penicillin binding protein 2a latex agglutination (PBP-LA) assay directly on Bactec blood culture broth samples containing Staphylococcus aureus to rapidly detect methicillin resistance. The sensitivity, specificity, positive predictive value, and negative predictive value of this method were 94.1%, 97.5%, 98%, and 92.9%, respectively.

Use of Positive Blood Cultures for Direct Identification and Susceptibility Testing with the Vitek 2 System

PubMed Central

de Cueto, Marina; Ceballos, Esther; Martinez-Martinez, Luis; Perea, Evelio J.; Pascual, Alvaro

2004-01-01

In order to further decrease the time lapse between initial inoculation of blood culture media and the reporting of results of identification and antimicrobial susceptibility tests for microorganisms causing bacteremia, we performed a prospective study in which specially processed fluid from positive blood culture bottles from Bactec 9240 (Becton Dickinson, Cockeysville, Md.) containing aerobic media were directly inoculated into Vitek 2 system cards (bio-Mérieux, France). Organism identification and susceptibility results were compared with those obtained from cards inoculated with a standardized bacterial suspension obtained following subculture to agar; 100 consecutive positive monomicrobic blood cultures, consisting of 50 gram-negative rods and 50 gram-positive cocci, were included in the study. For gram-negative organisms, 31 of the 50 (62%) showed complete agreement with the standard method for species identification, while none of the 50 gram-positive cocci were correctly identified by the direct method. For gram-negative rods, there were 50% categorical agreements between the direct and standard methods for all drugs tested. The very major error rate was 2.4%, and the major error rate was 0.6%. The overall error rate for gram-negatives was 6.6%. Complete agreement in clinical categories of all antimicrobial agents evaluated was obtained for 19 of 50 (38%) gram-positive cocci evaluated; the overall error rate was 8.4%, with 2.8% minor errors, 2.4% major errors, and 3.2% very major errors. These findings suggest that the Vitek 2 cards inoculated directly from positive Bactec 9240 bottles do not provide acceptable bacterial identification or susceptibility testing in comparison with corresponding cards tested by a standard method. PMID:15297523

Comparison of the recovery of Mycobacterium bovis isolates using the BACTEC MGIT 960 system, BACTEC 460 system, and Middlebrook 7H10 and 7H11 solid media.

PubMed

Hines, Nichole; Payeur, Janet B; Hoffman, Lorraine J

2006-05-01

The BACTEC Microbacteria Growth Indicator Tube (MGIT) 960 system was evaluated to determine how it compares with the BACTEC 460 radiometric system and solid media for recovery of Mycobacterium bovis from tissue samples. A total of 506 bovine lymph node samples were collected from abattoirs in the United States and Mexico between November 2003 and September 2004. Processed samples were inoculated into an MGIT 960 tube, BACTEC 460 vial, and Middlebrook 7H10 and Middlebrook 7H11 solid media. Ziehl-Neelsen slides were prepared to check for contaminants and confirm the presence of acid-fast positive bacilli. Samples containing acid-fast bacilli were confirmed as members of the Mycobacterium tuberculosis complex by a nucleic acid assay. Niacin and nitrate biochemical tests were used to distinguish M. bovis from M. tuberculosis isolates. Statistical analyses were performed to compare recovery rate, mean time to detection, contamination rates, as well as pair-wise comparisons in each category. The results showed that the MGIT 960 system had a higher recovery rate of M. bovis (122/129) than did the BACTEC 460 (102/129) and solid media system (96/129). The average time to detection was 15.8 days for the MGIT 960 system, 28.2 days for the BACTEC 460 system, and 43.4 days for solid media. Contamination rates were 6.9% for the MGIT 960 system, 3.4% for the BACTEC 460 system, and 21.7% for solid media. These results indicate the MGIT 960 system can be used as an alternative to the BACTEC 460 system for recovering M. bovis from tissue samples.

Multicenter evaluation of the Sepsityper™ extraction kit and MALDI-TOF MS for direct identification of positive blood culture isolates using the BD BACTEC™ FX and VersaTREK(®) diagnostic blood culture systems.

PubMed

Schieffer, K M; Tan, K E; Stamper, P D; Somogyi, A; Andrea, S B; Wakefield, T; Romagnoli, M; Chapin, K C; Wolk, D M; Carroll, K C

2014-04-01

(i) Evaluation of delayed time to blood culture extraction by the Sepsityper kit and impact of shipping pellets off-site for MALDI-TOF MS analysis. (ii) Comparison of Sepsityper and laboratory-developed extraction methods from a literature review. Using two blood culture systems (BD BACTEC and VersaTREK), we extracted 411 positive blood cultures using the Sepsityper kit to mimic a potential protocol for institutions without a MALDI-TOF MS. Extracted pellets were shipped and analysed on the Bruker UltraflexIII. Successful extraction of 358 (87·1%) samples was determined by the presence of detectable proteins. MALDI-TOF MS correctly identified 332 (80·8%) samples. Delayed time to extraction did not affect Sepsityper extraction or MALDI-TOF MS accuracy. The extracted pellets remain stable and provide accurate results by MALDI-TOF MS when shipped at room temperature to off-site reference laboratories. This is the first study to show that institutions without a MALDI-TOF MS can take advantage of this innovative technology by shipping a volume of blood to an off-site laboratory for extraction and MALDI-TOF MS analysis. We also performed a literature review to compare various extraction methods. © 2014 The Society for Applied Microbiology.

The sensitivity of direct identification from positive BacT/ALERT™ (bioMérieux) blood culture bottles by matrix-assisted laser desorption ionization time-of-flight mass spectrometry is low.

PubMed

Szabados, F; Michels, M; Kaase, M; Gatermann, S

2011-02-01

Recently, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been presented as a novel method for the direct identification of bacteria from positive blood culture bottles. The rate of the MALDI TOF MS-based identification in the present study from positive BacT/ALERT (bioMérieux, Marcy l'Etoile, France) blood culture bottles was 30%, which is far below the previously reported sensitivities using the BACTEC (Becton Dickinson, Franklin Lakes, NJ, USA) system. We also found evidence that the Biotyper algorithm did not identify a second pathogen in cases of positive BacT/ALERT blood culture bottles containing two different species. © 2010 The Authors. Journal Compilation © 2010 European Society of Clinical Microbiology and Infectious Diseases.

Comparison of Mycobacterium tuberculosis culture using liquid culture medium and Lowenstein Jensen medium in abdominal tuberculosis.

PubMed

Shah, Sudeep R; Shenai, Shubhada; Desai, Devendra C; Joshi, Anand; Abraham, Philip; Rodrigues, Camilla

2010-11-01

Traditionally, the Lowenstein Jensen (LJ) medium has been used for culturing Mycobacterium tuberculosis. In abdominal tuberculosis (TB), the reported yield from tissue culture is between 20% and 60%. Liquid cultures are reported to give a higher yield but there is little data available in abdominal TB. To compare the yield of TB culture with BACTEC 460TB liquid medium and LJ medium for patients with suspected abdominal TB and determine cost effectiveness. This prospective study was done in consecutive cases with clinical, radiological, endoscopic/surgical, and histological suspicion of abdominal TB. Tissue biopsies obtained at colonoscopy or surgery were processed and plated on LJ medium as well as the BACTEC 460TB system. NAP (ρ-nitro-α-acetylamino-β-hydroxy-propiophenone) differentiation was carried out to determine species. The cost of each method and cost per yield were calculated. Of the 29 cases, 22 cases (76%) were positive on BACTEC 460TB culture while 14 (48%) were positive on LJ medium giving a 64% increment in yield. However, the culture of one patient grew on LJ medium, where the BACTEC 460TB was negative. The additional cost of BACTEC 460TB is Rs. 460 and LJ is Rs. 40. Samples from patients with abdominal TB should be processed on both liquid and LJ medium. For high yield, the use of a liquid culture medium system is essential.

Comparison of the detection of periodontal pathogens in bacteraemia after tooth brushing by culture and molecular techniques.

PubMed

Marín, M-J; Figuero, E; González, I; O'Connor, A; Diz, P; Álvarez, M; Herrera, D; Sanz, M

2016-05-01

The prevalence and amounts of periodontal pathogens detected in bacteraemia samples after tooth brushing-induced by means of four diagnostic technique, three based on culture and one in a molecular-based technique, have been compared in this study. Blood samples were collected from thirty-six subjects with different periodontal status (17 were healthy, 10 with gingivitis and 9 with periodontitis) at baseline and 2 minutes after tooth brushing. Each sample was analyzed by three culture-based methods [direct anaerobic culturing (DAC), hemo-culture (BACTEC), and lysis-centrifugation (LC)] and one molecular-based technique [quantitative polymerase chain reaction (qPCR)]. With culture any bacterial isolate was detected and quantified, while with qPCR only Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans were detected and quantified. Descriptive analyses, ANOVA and Chi-squared tests, were performed. Neither BACTEC nor qPCR detected any type of bacteria in the blood samples. Only LC (2.7%) and DAC (8.3%) detected bacteraemia, although not in the same patients. Fusobacterium nucleatum was the most frequently detected bacterial species. The disparity in the results when the same samples were analyzed with four different microbiological detection methods highlights the need for a proper validation of the methodology to detect periodontal pathogens in bacteraemia samples, mainly when the presence of periodontal pathogens in blood samples after tooth brushing was very seldom.

Rapid identification of bacteria from positive blood culture bottles by use of matrix-assisted laser desorption-ionization time of flight mass spectrometry fingerprinting.

PubMed

Christner, Martin; Rohde, Holger; Wolters, Manuel; Sobottka, Ingo; Wegscheider, Karl; Aepfelbacher, Martin

2010-05-01

Early and adequate antimicrobial therapy has been shown to improve the clinical outcome in bloodstream infections (BSI). To provide rapid pathogen identification for targeted treatment, we applied matrix-assisted laser desorption-ionization time of flight (MALDI-TOF) mass spectrometry fingerprinting to bacteria directly recovered from blood culture bottles. A total of 304 aerobic and anaerobic blood cultures, reported positive by a Bactec 9240 system, were subjected in parallel to differential centrifugation with subsequent mass spectrometry fingerprinting and reference identification using established microbiological methods. A representative spectrum of bloodstream pathogens was recovered from 277 samples that grew a single bacterial isolate. Species identification by direct mass spectrometry fingerprinting matched reference identification in 95% of these samples and worked equally well for aerobic and anaerobic culture bottles. Application of commonly used score cutoffs to classify the fingerprinting results led to an identification rate of 87%. Mismatching mostly resulted from insufficient bacterial numbers and preferentially occurred with Gram-positive samples. The respective spectra showed low concordance to database references and were effectively rejected by score thresholds. Spiking experiments and examination of the respective study samples even suggested applicability of the method to mixed cultures. With turnaround times around 100 min, the approach allowed for reliable pathogen identification at the day of blood culture positivity, providing treatment-relevant information within the critical phase of septic illness.

Sample preparation method influences direct identification of anaerobic bacteria from positive blood culture bottles using MALDI-TOF MS.

PubMed

Jeverica, Samo; Nagy, Elisabeth; Mueller-Premru, Manica; Papst, Lea

2018-05-15

Rapid detection and identification of anaerobic bacteria from blood is important to adjust antimicrobial therapy by including antibiotics with activity against anaerobic bacteria. Limited data is available about direct identification of anaerobes from positive blood culture bottles using MALDI-TOF mass spectrometry (MS). In this study, we evaluated the performance of two sample preparation protocols for direct identification of anaerobes from positive blood culture bottles, the MALDI Sepsityper kit (Sepsityper) and the in-house saponin (saponin) method. Additionally, we compared two blood culture bottle types designed to support the growth of anaerobic bacteria, the BacT/ALERT-FN Plus (FN Plus) and the BACTEC-Lytic (Lytic), and their influence on direct identification. A selection of 30 anaerobe strains belonging to 22 different anaerobic species (11 reference strains and 19 clinical isolates) were inoculated to 2 blood culture bottle types in duplicate. In total, 120 bottles were inoculated and 99.2% (n = 119) signalled growth within 5 days of incubation. The Sepsityper method correctly identified 56.3% (n = 67) of anaerobes, while the saponin method correctly identified 84.9% (n = 101) of anaerobes with at least log(score) ≥1.6 (low confidence correct identification), (p < 0.001). Gram negative anaerobes were better identified with the saponin method (100% vs. 46.5%; p < 0.001), while Gram positive anaerobes were better identified with the Sepsityper method (70.8% vs. 62.5%; p = 0.454). Average log(score) values among only those isolates that were correctly identified simultaneously by both sample preparation methods were 2.119 and 2.029 in favour of the Sepsityper method, (p = 0.019). The inoculated bottle type didn't influence the performance of the two sample preparation methods. We confirmed that direct identification from positive blood culture bottles with MALDI-TOF MS is reliable for anaerobic bacteria. However, the results

CHROMagar Candida Medium for Direct Susceptibility Testing of Yeast from Blood Cultures

PubMed Central

Tan, Grace L.; Peterson, Ellena M.

2005-01-01

An evaluation was performed on 95 blood cultures positive for Candida spp. to determine the correlation of direct susceptibility testing of fluconazole versus both standardized disk diffusion and MIC methods. For direct testing, an aliquot taken from BD BACTEC Plus and/or BD BACTEC Lytic/10 bottles (Becton Dickinson [BD], Sparks, MD) positive by gram stain for yeast was subcultured to CHROMagar Candida (BD), and a 25-μg fluconazole disk (BD) was placed on the plate. The area of growth inhibition surrounding the disk was measured at 24 and 48 h. In addition, a subculture of the isolate was tested by a microdilution MIC using YeastOne (TREK Diagnostics Systems Inc., OH) and disk diffusion (NCCLS M44-A) using a standardized inoculum plated onto CHROMagar Candida as well as Mueller-Hinton agar to which 2% glucose and 0.5 μg/ml methylene blue dye was added (MH-GMB). The categorical interpretation derived from the MIC was used as the reference to which the disk diffusion results were compared. There were a total of 41 Candida albicans, 23 Candida glabrata, 20 Candida parapsilosis, 9 Candida tropicalis, and 1 each of Candida krusei and Candida lusitaniae tested. At 24 h there was full agreement among the methods for all C. albicans, C. tropicalis, C. lusitaniae, and C. krusei isolates. For the C. parapsilosis isolates at 24 h there was one very major discrepancy using the direct CHROMagar and one major error with the standardized MH-GMB. The majority of the errors were seen at 24 h with the C. glabrata isolates. Of the 23 C. glabrata isolates at 24 h by direct CHROMagar, there were 10 minor and 1 very major error; by MH-GMB there were 12 minor and 2 very major errors; and by standardized CHROMagar Candida there were 13 minor and 2 major errors. There were no very major errors with C. glabrata when all plates were read at 48 h. At 24 h by the direct and standardized CHROMagar the majority of C. glabrata isolates were more resistant, whereas by MH-GMB they were more

Differential fluorescent staining method for detection of bacteria in blood cultures, cerebrospinal fluid and other clinical specimens.

PubMed

Fazii, P; Ciancaglini, E; Riario Sforza, G

2002-05-01

The aim of this study was to evaluate a differential staining method to distinguish gram-positive from gram-negative bacteria in fluorescence. The method is based on two fluorochromes, one acting in the wavelength of red, i.e. the acridine orange, and another acting in the wavelength of green, i.e. the fluorescein. With this method, gram-positive bacteria appear yellow and gram-negative bacteria appear green. In view of the importance of a rapid aetiological diagnosis in cases of septicaemia, the differential staining method in fluorescence was compared with Gram stain for the detection of bacteria in blood. Of 5,820 blood cultures entered into the study and identified by the Bactec 9120 fluorescent series instrument (Becton Dickinson Europe, France), 774 were positive. Of the 774 positive cultures, 689 yielded only a single organism. The differential staining method in fluorescence detected 626 of the 689 cultures, while Gram stain detected 468. On the basis of these results, the sensitivity of the differential staining method in fluorescence was 90.9%, while that of Gram stain was 67.9%. The difference between the two methods was statistically significant ( P<0.001). The differential fluorescent staining method was more sensitive than Gram stain in the detection of bacteria in blood cultures during the incubation period. This technique provides a rapid, simple and highly sensitive staining method that can be used in conjunction with subculture methods. Whereas subculture requires an incubation period of 18-24 h, the fluorescent staining technique can detect bacteria on the same day that smears are prepared and examined. The differential fluorescent staining method was also evaluated for its ability to detect microorganisms in cerebrospinal fluid and other clinical specimens. The microorganisms were easily detected, even when bacterial counts in the specimens were low.

Evaluation of the BACTEC MGIT 960 SL DST Kit and the GenoType MTBDRsl Test for Detecting Extensively Drug-resistant Tuberculosis Cases.

PubMed

Tekin, Kemal; Albay, Ali; Simsek, Hulya; Sig, Ali Korhan; Guney, Mustafa

2017-10-01

The present study aimed to evaluate the performances of the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test for detecting second-line antituberculosis drug resistance in Multidrug-resistant TB (MDR-TB) cases. Forty-six MDR-TB strains were studied. Second-line antituberculosis drug resistances were detected using the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test. The Middlebrook 7H10 agar proportion method was used as the reference test. The sensitivity and specificity values for the BACTEC MGIT 960 SL DST kit were both 100% for amikacin, kanamycin, capreomycin (4 µg/mL), and ofloxacin; 100% and 95.3%, respectively, for capreomycin (10 µg/mL); and 85.7% and 100%, respectively, for moxifloxacin (0.5 µg/mL). The sensitivity and specificity values for the GenoType MTBDRsl test to detect fluoroquinolone and aminoglycoside/cyclic peptide resistance were 88.9% and 100%, respectively, for ofloxacin and 85.7% and 94.9%, respectively, for moxifloxacin (0.5 µg/mL). The accuracy of the GenoType MTBDRsl assay for kanamycin, capreomycin, ofloxacin, and moxifloxacin was lower than that of the BACTEC MGIT 960 SL DST. The BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl were successful in detecting second-line antituberculosis drug resistance. Preliminary results of the GenoType MTBDRsl are very valuable for early treatment decisions, but we still recommend additional BACTEC MGIT 960 SL DST kit usage in the routine evaluation of drug-resistant tuberculosis.

Evaluation of the BACTEC MGIT 960 SL DST Kit and the GenoType MTBDRsl Test for Detecting Extensively Drug-resistant Tuberculosis Cases

PubMed Central

Tekin, Kemal; Albay, Ali; Simsek, Hulya; Sig, Ali Korhan; Guney, Mustafa

2017-01-01

Objective: The present study aimed to evaluate the performances of the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test for detecting second-line antituberculosis drug resistance in Multidrug-resistant TB (MDR-TB) cases. Materials and Methods: Forty-six MDR-TB strains were studied. Second-line antituberculosis drug resistances were detected using the BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl test. The Middlebrook 7H10 agar proportion method was used as the reference test. Results: The sensitivity and specificity values for the BACTEC MGIT 960 SL DST kit were both 100% for amikacin, kanamycin, capreomycin (4 µg/mL), and ofloxacin; 100% and 95.3%, respectively, for capreomycin (10 µg/mL); and 85.7% and 100%, respectively, for moxifloxacin (0.5 µg/mL). The sensitivity and specificity values for the GenoType MTBDRsl test to detect fluoroquinolone and aminoglycoside/cyclic peptide resistance were 88.9% and 100%, respectively, for ofloxacin and 85.7% and 94.9%, respectively, for moxifloxacin (0.5 µg/mL). The accuracy of the GenoType MTBDRsl assay for kanamycin, capreomycin, ofloxacin, and moxifloxacin was lower than that of the BACTEC MGIT 960 SL DST. Conclusion: The BACTEC MGIT 960 SL DST kit and the GenoType MTBDRsl were successful in detecting second-line antituberculosis drug resistance. Preliminary results of the GenoType MTBDRsl are very valuable for early treatment decisions, but we still recommend additional BACTEC MGIT 960 SL DST kit usage in the routine evaluation of drug-resistant tuberculosis. PMID:29123441

Production of UC-labeled gas in BACTEC Neisseria Differentiation kits by Neisseria cinerea

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boyce, J.M.; Mitchell, E.B. Jr.; Knapp, J.S.

1985-09-01

Six strains of Neisseria cinerea were tested in BACTEC Neisseria Differentiation kits (Johnston Laboratories, Inc., Towson, Md.), and all yielded positive glucose growth indices and negative maltose and fructose growth indices. These results were similar to those achieved with Neisseria gonorrhoeae. However, most of the N. cinerea isolates tested yielded 3-h glucose growth indices that were lower than those obtained with gonococci. UC-labeled gas was produced significantly faster by N. gonorrhoeae than by N. cinerea. Additional studies suggested that the UC-labeled gas produced by N. cinerea was carbon dioxide. N. cinerea strains were similar to Branhamella catarrhalis strains because bothmore » species failed to produce detectable acid from glucose, maltose, sucrose, fructose, and lactose in cysteine-tryptic agar media. However, in contrast to N. cinerea strains, B. catarrhalis strains did not metabolize glucose in BACTEC Neisseria Differentiation kits.« less

Performance of Mycobacterium Growth Indicator Tube BACTEC 960 with Lowenstein-Jensen method for diagnosis of Mycobacterium tuberculosis at Ethiopian National Tuberculosis Reference Laboratory, Addis Ababa, Ethiopia.

PubMed

Diriba, Getu; Kebede, Abebaw; Yaregal, Zelalem; Getahun, Muluwork; Tadesse, Mengistu; Meaza, Abyot; Dagne, Zekarias; Moga, Shewki; Dilebo, Jibril; Gudena, Kebebe; Hassen, Mulu; Desta, Kassu

2017-05-10

Bacteriological confirmed active case detection remains the corner stone for diagnosing tuberculosis. Non-radiometric liquid culture system Mycobacterium Growth Indicator Tube with automated interface had been recommended by expert groups in addition to conventional solid culture media such as Lowenstein-Jensen. However in high burden resource limited countries advanced non-radiometric based tuberculosis diagnostic methods such as MGIT 960 is limited. Therefore we have evaluated the performance of MGIT 960 system compared to LJ for recovery of Mycobacterium complex (MTBC) from clinical specimens. A cross sectional study was conducted from a total of 908 samples between January 1st, 2013 to December 31st, 2014. Clinical specimens were processed following standard procedures and the final suspension was inoculated to MGIT tubes and LJ slant. Identification and confirmation of MTBC was done by ZN staining and SD Bioline test. Data was analyzed by SPSS version 20. The sensitivity, specificity, recovery rate and the average turnaround time to recover the organism was computed. From a total of 908 clinical specimens processed using both LJ and BACTEC MGIT liquid culture methods the recovery rate for LJ and MGIT, for smear positive samples was 66.7% (74/111) and 87.4% (97/ 111) respectively while for smear negative samples was 13.4% (108/797) and 17.4% (139/797) for LJ and MGIT methods respectively. The overall recovery rate for MGIT is significantly higher than LJ methods [26% (236/908; vs. 20%, 182/908, P = 0.002)]. The average turnaround time for smear positive samples was 16 and 31 days for MGIT and LJ respectively. Turnaround time for smear negative samples was 20 and 36 days for MGIT and LJ respectively. The overall agreement between MGIT and LJ was fairly good with Kappa value of 0.59 (P < 0.001). In the present study the contamination rate for MGIT is higher than the LJ methods, 15 and 9.3% respectively. The BACTEC MGIT liquid culture system has better MTBC

Comparison of MGIT and Myco/F lytic liquid-based blood culture systems for recovery of Mycobacterium tuberculosis from pleural fluid.

PubMed

Harausz, Elizabeth; Lusiba, John Kafuluma; Nsereko, Mary; Johnson, John L; Toossi, Zahra; Ogwang, Sam; Boom, W Henry; Joloba, Moses L

2015-04-01

The specificities and sensitivities of the Bactec mycobacterial growth indicator tube (MGIT) system for the recovery of Mycobacterium tuberculosis from pleural fluid are not statistically different than those of the Myco/F lytic liquid culture system. The time to positivity is shorter in the MGIT system (12.7 versus 20.7 days, respectively; P=0.007). The Myco/F lytic culture system may be an alternative to the MGIT system for diagnosing pleural tuberculosis. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Comparative performance of PCR-based assay versus microscopy and culture for the direct detection of Mycobacterium tuberculosis in clinical respiratory specimens in Lebanon.

PubMed

Araj, G F; Talhouk, R S; Itani, L Y; Jaber, W; Jamaleddine, G W

2000-09-01

American University of Beirut Medical Center, Lebanon. To assess the performance of a polymerase chain reaction (PCR) using primers that flank 542 bp within IS6110 in Mycobacterium tuberculosis (TB) vs. microscopy and BACTEC culture, in the diagnosis of tuberculosis. A total of 82 clinical respiratory pulmonary specimens and 73 samples from BACTEC vials were tested by the three methods. Of 24 smear-positive culture-positive (SP-CP) and 11 smear-negative culture-positive (SN-CP) TB specimens, PCR detected 83% and 64%, respectively. Among 17 specimens yielding mycobacteria other than tuberculosis (MOTT), the PCR was positive in 33% SP-CP and 14% SN-CP specimens. Among the 73 BACTEC vials, PCR was positive in 36 of 38 (95%) yielding culture-positive TB, and in one of 20 (5%) yielding culture positive MOTT. None of the 30 smear-negative culture-negative (SN-CN) clinical specimens and 15 of the CN vials were positive by PCR. The overall sensitivity of PCR was 77% and 95% for TB detection in respiratory specimens and BACTEC vials, respectively, and the specificity was 94% in both. Because a substantial number of TB cases are missed, especially in SN-CP specimens, a PCR-based assay utilizing these primers cannot be used reliably, alone, in clinical laboratory diagnosis of mycobacterial respiratory infections.

Novel, improved sample preparation for rapid, direct identification from positive blood cultures using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry.

PubMed

Schubert, Sören; Weinert, Kirsten; Wagner, Chris; Gunzl, Beatrix; Wieser, Andreas; Maier, Thomas; Kostrzewa, Markus

2011-11-01

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is widely used for rapid and reliable identification of bacteria and yeast grown on agar plates. Moreover, MALDI-TOF MS also holds promise for bacterial identification from blood culture (BC) broths in hospital laboratories. The most important technical step for the identification of bacteria from positive BCs by MALDI-TOF MS is sample preparation to remove blood cells and host proteins. We present a method for novel, rapid sample preparation using differential lysis of blood cells. We demonstrate the efficacy and ease of use of this sample preparation and subsequent MALDI-TOF MS identification, applying it to a total of 500 aerobic and anaerobic BCs reported to be positive by a Bactec 9240 system. In 86.5% of all BCs, the microorganism species were correctly identified. Moreover, in 18/27 mixed cultures at least one isolate was correctly identified. A novel method that adjusts the score value for MALDI-TOF MS results is proposed, further improving the proportion of correctly identified samples. The results of the present study show that the MALDI-TOF MS-based method allows rapid (<20 minutes) bacterial identification directly from positive BCs and with high accuracy. Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Blood culture bottles are superior to conventional media for vitreous culture.

PubMed

Thariya, Patsuda; Yospaiboon, Yosanan; Sinawat, Suthasinee; Sanguansak, Thuss; Bhoomibunchoo, Chavakij; Laovirojjanakul, Wipada

2016-08-01

To compare blood culture bottles and conventional media for the vitreous culture in patients with clinically suspected infectious endophthalmitis. Retrospective comparative study at KKU Eye Center, Khon Kaen University. There were 342 patients with clinically suspected infectious endophthalmitis participated in the study. The vitreous specimens were inoculated in both blood culture bottles and on conventional culture media (blood agar, MacConkey agar, chocolate agar, Sabouraud dextrose agar and thioglycolate broth). The number of positive culture yields in both blood culture bottles and conventional media. Positive culture yields in both methods were found in 151 eyes (49.5%). There were 136 of 151 eyes (90.1%) with positive culture in blood culture bottles, whereas 99 of 151 eyes (65.6%) yielded positive cultures in conventional media. These findings were different with a statistical significance (P < 0.00001) and an odds ratio of 3.47 (95% confidence interval 1.92, 6.63). A combination of blood culture bottles and conventional media improved the yield. Blood culture bottles are superior to conventional media for vitreous culture in clinically suspected infectious endophthalmitis. Vitreous culture using blood culture bottles should be recommended as the primary method for microbiological diagnosis. A combination of both methods further improves the positive culture yield. © 2016 Royal Australian and New Zealand College of Ophthalmologists.

A whole blood bactericidal assay for tuberculosis.

PubMed

Wallis, R S; Palaci, M; Vinhas, S; Hise, A G; Ribeiro, F C; Landen, K; Cheon, S H; Song, H Y; Phillips, M; Dietze, R; Ellner, J J

2001-04-15

The bactericidal activity of orally administered antituberculosis (anti-TB) drugs was determined in a whole blood culture model of intracellular infection in which microbial killing reflects the combined effects of drug and immune mechanisms. Rifampin (Rif) was the most active compound studied and reduced the number of viable bacilli by >4 logs. Isoniazid (INH), 2 quinolones, and pyrazinamide (PZA) showed intermediate levels of activity. Ethambutol exerted only a bacteristatic effect; amoxicillin/clavulanate was inactive. The combination of INH-Rif-PZA showed strong activity against 11 drug-sensitive isolates (mean, -3.8 log) but no activity against 12 multidrug-resistant (MDR) strains. The combination of levofloxacin-PZA-ethambutol had intermediate bactericidal activity against MDR isolates (mean, -1.2 log) but failed to equal that of INH-Rif-PZA against sensitive isolates (P<.001). The whole blood BACTEC method (Becton Dickinson) may be useful for the early clinical evaluation of new anti-TB drugs and in the management of individual patients.

Acid-fast bacilli culture positivity and drug resistance in abdominal tuberculosis in Mumbai, India.

PubMed

Samant, Hrishikesh; Desai, Devendra; Abraham, Philip; Joshi, Anand; Gupta, Tarun; Rodrigues, Camilla; George, Siji

2014-09-01

Culture positivity for Mycobacterium tuberculosis complex (MTB) in abdominal tuberculosis (TB) using Lowenstein Jensen medium and Bactec system varies from 25 % to 36 %. Data on the prevalence of drug resistance in primary abdominal TB is scant. Our aim was to study the acid-fast bacilli (AFB) culture positivity rate in primary abdominal TB using Bactec Mycobacterial Growth Indicator Tubes (MGIT) system and the prevalence of drug resistance in these patients. Records of patients with abdominal TB (diagnosed on clinical features, endoscopy, histology, microbiology) seen during the period 2008 to 2013 were retrieved from the Gastroenterology and Microbiology departments. Patients with extra-abdominal TB (five pulmonary, two nodal), adnexal (one), and HIV (one) were excluded from analysis. Of 61 patients, 31 (50.8 %) had a positive AFB culture. In the 30 culture-negative patients, histology showed non-caseating granulomas in 25 patients. Drug sensitivity pattern was analyzed in 18 patients; resistance was detected in eight (14.3 % of all patients and 44.4 % of patients in whom drug sensitivity was done) including three (5.4 % of all subjects and 16.6 % in whom drug sensitivity was available) who were multidrug-resistant. The rate of AFB culture positivity in primary abdominal TB was 50.8 % using Bactec MGIT. Likelihood of drug resistance was seen in 14.3 %, of whom 5.4 % were multidrug-resistant.

Stability of amoxicillin-clavulanate in BACTEC medium determined by high-performance liquid chromatography and bioassay.

PubMed Central

Moore, T D; Horton, R; Utrup, L J; Miller, L A; Poupard, J A

1996-01-01

The stabilities of amoxicillin (16 micrograms/ml) and clavulanate (8 micrograms/ml), alone and in combination in BACTEC medium (Middlebrook 7H12B medium), were determined by high-performance liquid chromatography (HPLC) and bioassay. By HPLC, the half-life of amoxicillin (trihydrate and sodium) in combination with clavulanate in nonradiolabelled 7H12B medium was 6.7 days, whereas the half-life of clavulanate in combination with amoxicillin was 2.0 days. By bioassay, the half-lives of amoxicillin trihydrate and clavulanate in radiolabelled 7H12B medium were comparable (7 and 2 days, respectively) to those determined by HPLC. When clavulanate was tested alone, the half-life was determined to be 1.88 days by HPLC and 1.87 days by bioassay. The relatively short half-life of clavulanate can be adjusted by a procedure of "topping up," or adding one-half the concentration of clavulanate every second day, in order to allow accurate amoxicillin-clavulanate MIC testing with the BACTEC mycobacterial susceptibility system. PMID:8727931

Blood culture

MedlinePlus

... into (contaminating) the blood sample and causing a false-positive result (see below). The sample is sent to a laboratory. There, it is placed in a special dish (culture). It is then watched to see if bacteria ...

Laboratory Workflow Analysis of Culture of Periprosthetic Tissues in Blood Culture Bottles.

PubMed

Peel, Trisha N; Sedarski, John A; Dylla, Brenda L; Shannon, Samantha K; Amirahmadi, Fazlollaah; Hughes, John G; Cheng, Allen C; Patel, Robin

2017-09-01

Culture of periprosthetic tissue specimens in blood culture bottles is more sensitive than conventional techniques, but the impact on laboratory workflow has yet to be addressed. Herein, we examined the impact of culture of periprosthetic tissues in blood culture bottles on laboratory workflow and cost. The workflow was process mapped, decision tree models were constructed using probabilities of positive and negative cultures drawn from our published study (T. N. Peel, B. L. Dylla, J. G. Hughes, D. T. Lynch, K. E. Greenwood-Quaintance, A. C. Cheng, J. N. Mandrekar, and R. Patel, mBio 7:e01776-15, 2016, https://doi.org/10.1128/mBio.01776-15), and the processing times and resource costs from the laboratory staff time viewpoint were used to compare periprosthetic tissues culture processes using conventional techniques with culture in blood culture bottles. Sensitivity analysis was performed using various rates of positive cultures. Annualized labor savings were estimated based on salary costs from the U.S. Labor Bureau for Laboratory staff. The model demonstrated a 60.1% reduction in mean total staff time with the adoption of tissue inoculation into blood culture bottles compared to conventional techniques (mean ± standard deviation, 30.7 ± 27.6 versus 77.0 ± 35.3 h per month, respectively; P < 0.001). The estimated annualized labor cost savings of culture using blood culture bottles was $10,876.83 (±$337.16). Sensitivity analysis was performed using various rates of culture positivity (5 to 50%). Culture in blood culture bottles was cost-effective, based on the estimated labor cost savings of $2,132.71 for each percent increase in test accuracy. In conclusion, culture of periprosthetic tissue in blood culture bottles is not only more accurate than but is also cost-saving compared to conventional culture methods. Copyright © 2017 American Society for Microbiology.

Prospective study of the clinical performance of three BACTEC media in a modern emergency department: Plus Aerobic/F, Plus Anaerobic/F, and Anaerobic Lytic/F.

PubMed

Rocchetti, Andrea; Di Matteo, Luigi; Bottino, Paolo; Foret, Benjamin; Gamalero, Elisa; Calabresi, Alessandra; Guido, Gianluca; Casagranda, Ivo

2016-11-01

The performance of 3 blood culture bottles (BACTEC Plus Aerobic/F, Plus Anaerobic/F, and Anaerobic Lytic/F) were analyzed with clinical specimens collected from 688 Emergency Department patients. A total of 270 strains belonging to 33 species were identified, with E. coli and S. aureus as the most frequently detected. Overall recovery rate (RR) of bacteria and yeast was equivalent in the Plus Aerobic/F vials (208 of 270 isolates; 77.0%) and Anaerobic Lytic/F vials (206 isolates; 76.3%) and significantly better than in the Plus Anaerobic/F vials (189 isolates; 70.0%). Median time to detection (TTD) was earliest with the Anaerobic Lytic/F vials (12.0h) compared with the Plus Aerobic/F (14.6h) and Plus Anaerobic/F vials (15.4h). Positivity rate (PR) was similar for Anaerobic Lytic/F vials (76.9%) and Plus Aerobic/F vials (76.5%), but better if compared with Plus Anaerobic/F vials (69.4%). The PR and TTD for the combination of Plus Aerobic/F with Anaerobic Lytic/F (94.5% and 12.3h, respectively) was significantly better than with Plus Aerobic/F with Plus Anaerobic/F (87.8% and 14.1h). Copyright © 2016 Elsevier B.V. All rights reserved.

Factors affecting isolation and identification of Mycobacterium avium subsp. paratuberculosis from fecal and tissue samples in a liquid culture system.

PubMed

Whittington, Richard J

2009-03-01

Culture of Mycobacterium avium subsp. paratuberculosis is the definitive diagnostic test for Johne's disease, a chronic granulomatous enteropathy of animals. Compared to solid media, the identification of all strains of the organism in liquid media can be more difficult because the appearance of colonies and mycobactin dependence are not observable, and the growth of other organisms needs to be distinguished, commonly by PCR. Factors affecting the isolation rate of S strains and the contamination rate in modified Middlebrook 7H9 broth (Bactec 12B) and 7H10 agar were studied using 11,598 fecal samples and 2,577 tissue samples from sheep from 1,421 farms over 10 years. Minimization of contamination in Bactec cultures required the avoidance of the carryover of fecal particles from the first sedimentation step in the double-incubation centrifugation method, and contamination was reduced significantly by incubating the sample in a solution containing vancomycin, amphotericin B, and nalidixic acid for 3 days compared to 2 days. The growth of irrelevant microorganisms confounded the identification of M. avium subsp. paratuberculosis in liquid culture by inhibiting IS900 PCR and in solid medium culture by inhibiting the growth of M. avium subsp. paratuberculosis or obscuring colonies. The contamination of samples was clustered in certain laboratory submissions and was reduced by including ampicillin in Bactec medium without affecting the odds of isolation of M. avium subsp. paratuberculosis. The long-term contamination rate for fecal cultures was about 7%, and that for tissue cultures was <0.2%. Liquid medium was more sensitive than solid medium culture for M. avium subsp. paratuberculosis. The applicability of these findings for C strains is discussed.

False no-growth blood cultures in pneumococcal pneumonia.

PubMed Central

Adeniyi-Jones, C C; Stevens, D L; Rasquinha, E S

1980-01-01

The growth of Streptococcus pneumoniae in commercial media containing 14C-labeled substrates was studied experimentally; the results of blood cultures that were positive for S. pneumoniae over a 14-month period were analyzed to explain no-growth but radiometrically positive blood cultures from four patients with clinically diagnosed pneumococcal pneumonia. The growth of S. pneumonoiae in aerobic blood culture vials resulted in a chocolate color in the medium. S. pneumoniae grew rapidly in both aerobic and anaerobic media, but 14CO2 evolved from the metabolism of the labeled substrates was detected only in the aerobic culture vials. Radiometric detection lagged behind growth of the organisms and was accompanied by visual changes in the media. By 24 h, the viability of the culture was on the decline; radiometric readings remained positive even when the culture had died. PMID:7419708

Neonatal blood cultures: effect of delayed entry into the blood culture machine and bacterial concentration on the time to positive growth in a simulated model.

PubMed

Jardine, Luke Anthony; Sturgess, Barbara Ruth; Inglis, Garry Donald Trevor; Davies, Mark William

2009-04-01

To determine if: time from blood culture inoculation to positive growth (total time to positive) and time from blood culture machine entry to positive growth (machine time to positive) is altered by delayed entry into the automated blood culture machine, and if the total time to positive differs by the concentration of organisms inoculated into blood culture bottles. Staphylococcus epidermidis, Escherichia coli and group B beta-haemolytic streptococci were chosen as clinically significant representative organisms. Two concentrations (> or =10 colony-forming units per millilitre and <1 colony-forming units per millilitre) were inoculated into PEDS BacT/Alert blood culture bottles and randomly allocated to one of three delayed automated blood culture machine entry times (30 min/8.5 h/15.5 h). For all organisms at all concentrations, except the Staphylococcus epidermidis, the machine time to positive was significantly decreased by delayed entry. For all organisms at all concentrations, the mean total time to positive significantly increased with increasing delayed entry into the blood culture machine. Higher concentrations of group B beta-haemolytic streptococci and Escherichia coli grew significantly faster than lower concentrations. Bacterial growth in inoculated bottles, stored at room temperature, continues although at a slower rate than in those blood culture bottles immediately entered into the machine. If a blood culture specimen has been stored at room temperature for greater than 15.5 h, the currently allowed safety margin of 36 h (before declaring a result negative) may be insufficient.

Blood culture results from healthy captive and free-ranging elasmobranchs.

PubMed

Mylniczenko, Natalie D; Harris, Brigita; Wilborn, Rachel E; Young, Forrest A

2007-09-01

Blood culture is a diagnostic tool used in confirming bacterial disease in teleostean and elasmobranch fishes. Unlike teleosts, elasmobranchs have a normal microflora in multiple organs, but their blood has generally been considered to be sterile. In regular exams of elasmobranchs conducted at a public aquarium, occasional blood samples have tested positive on culture. This finding prompted a blood culture survey of healthy captive and wild elasmobranchs (sharks and stingrays), which showed that 26.7% of all animals were positive. Stingrays alone showed a 50% occurrence of positive blood cultures, although the total number of animals was low and freshwater species were included in this number. When elasmobranchs other than stingrays were evaluated according to metabolic category, pelagic animals had a higher percentage of positive cultures than nonpelagic animals (38.7% versus 13.9%). These results indicate that a single positive blood culture without other corroborating diagnostics is not sufficient to confirm septicemia in elasmobranchs.

Validation of shortened 2-day sterility testing of mesenchymal stem cell-based therapeutic preparation on an automated culture system.

PubMed

Lysák, Daniel; Holubová, Monika; Bergerová, Tamara; Vávrová, Monika; Cangemi, Giuseppina Cristina; Ciccocioppo, Rachele; Kruzliak, Peter; Jindra, Pavel

2016-03-01

Cell therapy products represent a new trend of treatment in the field of immunotherapy and regenerative medicine. Their biological nature and multistep preparation procedure require the application of complex release criteria and quality control. Microbial contamination of cell therapy products is a potential source of morbidity in recipients. The automated blood culture systems are widely used for the detection of microorganisms in cell therapy products. However the standard 2-week cultivation period is too long for some cell-based treatments and alternative methods have to be devised. We tried to verify whether a shortened cultivation of the supernatant from the mesenchymal stem cell (MSC) culture obtained 2 days before the cell harvest could sufficiently detect microbial growth and allow the release of MSC for clinical application. We compared the standard Ph. Eur. cultivation method and the automated blood culture system BACTEC (Becton Dickinson). The time to detection (TTD) and the detection limit were analyzed for three bacterial and two fungal strains. The Staphylococcus aureus and Pseudomonas aeruginosa were recognized within 24 h with both methods (detection limit ~10 CFU). The time required for the detection of Bacillus subtilis was shorter with the automated method (TTD 10.3 vs. 60 h for 10-100 CFU). The BACTEC system reached significantly shorter times to the detection of Candida albicans and Aspergillus brasiliensis growth compared to the classical method (15.5 vs. 48 and 31.5 vs. 48 h, respectively; 10-100 CFU). The positivity was demonstrated within 48 h in all bottles, regardless of the size of the inoculum. This study validated the automated cultivation system as a method able to detect all tested microorganisms within a 48-h period with a detection limit of ~10 CFU. Only in case of B. subtilis, the lowest inoculum (~10 CFU) was not recognized. The 2-day cultivation technique is then capable of confirming the microbiological safety of MSC and

[Utility of the direct culture examination (Ziehl-Neelsen) using the Bactec system for the presumptive identification of Mycobacterium tuberculosis complex, Mycobacterium avium complex, Mycobacterium xenopi, and Mycobacterium kansasii].

PubMed

Moreno, C; Garrigó, M; Sánchez, F; Coll, P

1994-05-01

The usefulness of the microscopic examination of Bactec 12B and 13A growth medium as a method for the possible identification of M. tuberculosis complex, M. avium complex, M. xenopi, and M. kansasii was performed out to guide the selection of different genetic identification probes and, in the case of M. xenopi, the selection of the temperature of subcultures incubation. Upon detection of an index of growth greater than 100 in Bactec tubes, staining was performed by the Ziehl-Neelsen technique. On the basis of the morphology observed, the possible identification was performed by genetic probes. Subcultures were used for definitive identification. Three hundred forty-four positive samples were studied by radiometric technique. A total of 190 strains were identified as M. tuberculosis, 88 strains as M. avium-intracellulare (MAI), 33 strains as M. xenopi, 14 strains as M. kansasii and 19 strains were identified as: M. gordonae (10), unpigmented rapid growth microbacteria (7), and M. simiae (2). Sensitivity, specificity, positive predictive value, and negative predictive value were 97.9%, 95.4%, 96.4%, and 97.3%, respectively for M. tuberculosis complex, 84.0%, 99.2%, 97.3% 94.7% for M. avium complex; 63.6%, 98.3%, 80.7%, 96.2% for M. xenopi; 35.7%, 98.1%, 45.5% 97.2% for M. kansasii. The morphology of M. tuberculosis complex examined in the radiometric system in useful to differentiate this species from other microbacteria (MOTT), allowing the selection of specific probe used. Within the MOTT, M. avium complex also has morphological characteristics which are useful for its differentiation, the morphology usually described for the remaining species was frequently not observed.

Doing it right the first time: quality improvement and the contaminant blood culture.

PubMed

Weinbaum, F I; Lavie, S; Danek, M; Sixsmith, D; Heinrich, G F; Mills, S S

1997-03-01

The aim of the project was to determine whether the rate of contaminant blood cultures could be reduced by using a team of dedicated phlebotomists. Comparisons were made between adult patients requiring blood cultures for suspected bacteremia on medical and surgical units before and after the introduction and withdrawal of a dedicated blood culture team. The results showed that a significant reduction in the contaminant blood culture rate was achieved by the blood culture team (P < 0.001; chi(2) test). Therefore, in our experience, the rate of contaminant blood cultures can be reduced in a teaching hospital by using a team of dedicated phlebotomists. Calculations made with our data and those published by others suggest that cost savings from reducing false-positive blood cultures are greater than the cost of the blood culture team.

Blood cultures in emergency medical admissions: a key patient cohort.

PubMed

Chotirmall, Sanjay H; Callaly, Elizabeth; Lyons, Judith; O'Connell, Brian; Kelleher, Mary; Byrne, Declan; O'Riordan, Deirdre; Silke, Bernard

2016-02-01

Blood cultures are performed in the emergency room when sepsis is suspected, and a cohort of patients is thereby identified. The present study investigated the outcomes (mortality and length of hospital stay) in this group following an emergency medical admission. Prospective assessment of all emergency medical admissions presenting to the emergency department at St James's Hospital, Dublin, over an 11-year period (2002-2012) was carried out. Outcomes including 30-day in-hospital mortality and length of stay were explored in the context of an admission blood culture. Generalized estimating equations, logistic or zero-truncated Poisson multivariate models were used, with adjustment for confounding variables including illness severity, comorbidity, and chronic disabling disease, to assess the effect of an urgent blood culture on mortality and length of stay. A total of 60 864 episodes were recorded in 35 168 patients admitted over the time period assessed. Patients more likely to undergo blood cultures in the emergency department were male, younger, and had more comorbidity. Univariate and multivariate analyses showed that those who had a blood culture, irrespective of result, had increased mortality and a longer in-hospital stay. This was highest for those with a positive culture, irrespective of the organism isolated. A clinical decision to request a blood culture identified a subset of emergency admissions with markedly worse outcomes. This patient cohort warrants close monitoring in the emergency setting.

Potential Impact of Rapid Blood Culture Testing for Gram-Positive Bacteremia in Japan with the Verigene Gram-Positive Blood Culture Test

PubMed Central

Matsuda, Mari; Iguchi, Shigekazu; Mizutani, Tomonori; Hiramatsu, Keiichi; Tega-Ishii, Michiru; Sansaka, Kaori; Negishi, Kenta; Shimada, Kimie; Umemura, Jun; Notake, Shigeyuki; Yanagisawa, Hideji; Yabusaki, Reiko; Araoka, Hideki; Yoneyama, Akiko

2017-01-01

Background. Early detection of Gram-positive bacteremia and timely appropriate antimicrobial therapy are required for decreasing patient mortality. The purpose of our study was to evaluate the performance of the Verigene Gram-positive blood culture assay (BC-GP) in two special healthcare settings and determine the potential impact of rapid blood culture testing for Gram-positive bacteremia within the Japanese healthcare delivery system. Furthermore, the study included simulated blood cultures, which included a library of well-characterized methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE) isolates reflecting different geographical regions in Japan. Methods. A total 347 BC-GP assays were performed on clinical and simulated blood cultures. BC-GP results were compared to results obtained by reference methods for genus/species identification and detection of resistance genes using molecular and MALDI-TOF MS methodologies. Results. For identification and detection of resistance genes at two clinical sites and simulated blood cultures, overall concordance of BC-GP with reference methods was 327/347 (94%). The time for identification and antimicrobial resistance detection by BC-GP was significantly shorter compared to routine testing especially at the cardiology hospital, which does not offer clinical microbiology services on weekends and holidays. Conclusion. BC-GP generated accurate identification and detection of resistance markers compared with routine laboratory methods for Gram-positive organisms in specialized clinical settings providing more rapid results than current routine testing. PMID:28316631

The Importance of Mean Neutrophil Volume (MNV) Values in Differential Diagnosis of Bacterial Sepsis.

PubMed

Şafak, Birol; Baykan, Özgür; Kılınç, Osman; Özer Yıldırım, Diğdem

2017-01-01

Bloodstream infections are a significant cause of morbidity and mortality in hospitalized patients. Blood cultures and other laboratory tests are used for diagnosis. Among these tests, the mean neutrophil volume (MNV) value is reported as a potential indicator that supports the diagnosis of sepsis. Our study identified the MNV values of patients via microorganisms cultivated from blood cultures and examined the role of these MNV values in the early diagnosis of bloodstream infections. Our study surveyed retrospectively 148 adult patient blood culture samples that had been sent to our laboratory. BACTEC 9050 (Becton Dickinson, USA) and BACTEC FX 40 (Becton Dickinson, USA) devices were used in the blood culture isolation procedures. The average MNV value was found to be 159.0 (+11.3) in patients whose sepsis originated from Gram-negative bacteria, and the average MNV value was measured as 152.4 (+14.5) among patients whose sepsis originated from Gram-positive bacteria. When comparing groups of patients having Gram-negative bacteria and patients having Gram-positive bacteria, a statistically significant difference (p = 0.041) in the MNV values was observed. The MNV value was found to be statistically significant in discrimination of Gram-negative and Gram-positive sepsis. Considering these findings, measuring the MNV values can help initiate proper antibiotic treatment more quickly, and we think that this will help lower the mortality rate. However, these findings should be supported with further studies. Copyright © 2017 National Medical Association. Published by Elsevier Inc. All rights reserved.

Effect of postmortem sampling technique on the clinical significance of autopsy blood cultures.

PubMed

Hove, M; Pencil, S D

1998-02-01

Our objective was to investigate the value of postmortem autopsy blood cultures performed with an iodine-subclavian technique relative to the classical method of atrial heat searing and antemortem blood cultures. The study consisted of a prospective autopsy series with each case serving as its own control relative to subsequent testing, and a retrospective survey of patients coming to autopsy who had both autopsy blood cultures and premortem blood cultures. A busy academic autopsy service (600 cases per year) at University of Texas Medical Branch Hospitals, Galveston, Texas, served as the setting for this work. The incidence of non-clinically relevant (false-positive) culture results were compared using different methods for collecting blood samples in a prospective series of 38 adult autopsy specimens. One hundred eleven adult autopsy specimens in which both postmortem and antemortem blood cultures were obtained were studied retrospectively. For both studies, positive culture results were scored as either clinically relevant or false positives based on analysis of the autopsy findings and the clinical summary. The rate of false-positive culture results obtained by an iodine-subclavian technique from blood drawn soon after death were statistically significantly lower (13%) than using the classical method of obtaining blood through the atrium after heat searing at the time of the autopsy (34%) in the same set of autopsy subjects. When autopsy results were compared with subjects' antemortem blood culture results, there was no significant difference in the rate of non-clinically relevant culture results in a paired retrospective series of antemortem blood cultures and postmortem blood cultures using the iodine-subclavian postmortem method (11.7% v 13.5%). The results indicate that autopsy blood cultures obtained using the iodine-subclavian technique have reliability equivalent to that of antemortem blood cultures.

Comparison between MALDI-TOF MS and FilmArray Blood Culture Identification panel for rapid identification of yeast from positive blood culture.

PubMed

Paolucci, M; Foschi, C; Tamburini, M V; Ambretti, S; Lazzarotto, T; Landini, M P

2014-09-01

In this study we evaluated MALDI-TOF MS and FilmArray methods for the rapid identification of yeast from positive blood cultures. FilmArray correctly identified 20/22 of yeast species, while MALDI-TOF MS identified 9/22. FilmArray is a reliable and rapid identification system for the direct identification of yeasts from positive blood cultures. Copyright © 2014 Elsevier B.V. All rights reserved.

Blood culture-negative endocarditis

PubMed Central

Fournier, Pierre-Edouard; Gouriet, Frédérique; Casalta, Jean-Paul; Lepidi, Hubert; Chaudet, Hervé; Thuny, Franck; Collart, Frédéric; Habib, Gilbert; Raoult, Didier

2017-01-01

Abstract Blood culture-negative endocarditis (BCNE) may represent up to 70% of all endocarditis cases, depending on series. From 2001 to 2009, we implemented in our laboratory a multimodal diagnostic strategy for BCNE that included systematized testing of blood, and when available, valvular biopsy specimens using serological, broad range molecular, and histopathological assays. A causative microorganism was identified in 62.7% of patients. In this study from January 2010 to December 2015, in an effort to increase the number of identified causative microorganisms, we prospectively added to our diagnostic protocol specific real-time (RT) polymerase chain reaction (PCR) assays targeting various endocarditis agents, and applied them to all patients with BCNE admitted to the 4 public hospitals in Marseille, France. A total of 283 patients with BCNE were included in the study. Of these, 177 were classified as having definite endocarditis. Using our new multimodal diagnostic strategy, we identified an etiology in 138 patients (78.0% of cases). Of these, 3 were not infective (2.2%) and 1 was diagnosed as having Mycobacterium bovis BCG endocarditis. By adding specific PCR assays from blood and valvular biopsies, which exhibited a significantly greater sensitivity (P < 10−2) than other methods, causative agents, mostly enterococci, streptococci, and zoonotic microorganisms, were identified in an additional 27 patients (14 from valves only, 11 from blood only, and 2 from both). Finally, in another 107 patients, a pathogen was detected using serology in 37, valve culture in 8, broad spectrum PCR from valvular biopsies and blood in 19 and 2, respectively, immunohistochemistry from valves in 3, and a combination of several assays in 38. By adding specific RT-PCR assays to our systematic PCR testing of patients with BCNE, we increased the diagnostic efficiency by 24.3%, mostly by detecting enterococci and streptococci that had not been detected by other diagnostic methods

Diagnostic performance of blood culture bottles for vitreous culture compared to conventional microbiological cultures in patients with suspected endophthalmitis.

PubMed

Kehrmann, Jan; Chapot, Valerie; Buer, Jan; Rating, Philipp; Bornfeld, Norbert; Steinmann, Joerg

2018-05-01

The purpose of this investigation was to evaluate the performance of blood culture bottles in comparison to conventional microbiological culture techniques in detecting causative microorganisms of endophthalmitis and to determine their anti-infective susceptibility profiles. All consecutive cases with clinically suspected endophthalmitis in a university-based ophthalmology department between January 2009 and December 2016 were analysed in this retrospective comparative case series. Samples from 247 patients with suspected endophthalmitis underwent microbiological diagnostic work-up. All three culture methods were performed from 140 vitreous specimens. Vitreous fluid specimens were inoculated in blood culture bottles, aerobic and anaerobic broth solutions, and on solid media. Anti-infective susceptibility profiles were evaluated by semi-automated methods and/or gradient diffusion methods. Microorganisms were grown in 82 of 140 specimens for which all methods were performed (59%). Microorganisms were more frequently grown from blood culture bottles (55%) compared to broth solution (45%, p = 0.007) and solid media (33%, p < 0.0001). Considerable differences in the performance among culture media were detected for fungal pathogens. All grown fungi were detected by blood culture bottles (11 of 11, 100%). Broth solution recovered 64% and solid media 46% of grown fungi. No Gram-positive bacterium was resistant to vancomycin and all Gram-negative pathogens except for one isolate were susceptible to third-generation cephalosporins. In suspected endophthalmitis patients, blood culture bottles have a higher overall pathogen detection rate from vitreous fluid compared to conventional microbiological media, especially for fungi. The initial intravitreal antibiotic therapy with vancomycin plus third-generation cephalosporins appears to be an appropriate treatment approach for bacterial endophthalmitis.

Reduction in Blood Culture Contamination Through Use of Initial Specimen Diversion Device.

PubMed

Rupp, Mark E; Cavalieri, R Jennifer; Marolf, Cole; Lyden, Elizabeth

2017-07-15

Blood culture contamination is a clinically significant problem that results in patient harm and excess cost. In a prospective, controlled trial at an academic center Emergency Department, a device that diverts and sequesters the initial 1.5-2 mL portion of blood (which presumably carries contaminating skin cells and microbes) was tested against standard phlebotomy procedures in patients requiring blood cultures due to clinical suspicion of serious infection. In sum, 971 subjects granted informed consent and were enrolled resulting in 904 nonduplicative subjects with 1808 blood cultures. Blood culture contamination was significantly reduced through use of the initial specimen diversion device™ (ISDD) compared to standard procedure: (2/904 [0.22%] ISDD vs 16/904 [1.78%] standard practice, P = .001). Sensitivity was not compromised: true bacteremia was noted in 65/904 (7.2%) ISDD vs 69/904 (7.6%) standard procedure, P = .41. No needlestick injuries or potential bloodborne pathogen exposures were reported. The monthly rate of blood culture contamination for all nurse-drawn and phlebotomist-drawn blood cultures was modeled using Poisson regression to compare the 12-month intervention period to the 6 month before and after periods. Phlebotomists (used the ISDD) experienced a significant decrease in blood culture contamination while the nurses (did not use the ISDD) did not. In sum, 73% of phlebotomists completed a post-study anonymous survey and widespread user satisfaction was noted. Use of the ISDD was associated with a significant decrease in blood culture contamination in patients undergoing blood cultures in an Emergency Department setting. NCT02102087. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Reduction in Blood Culture Contamination Through Use of Initial Specimen Diversion Device

PubMed Central

Cavalieri, R. Jennifer; Marolf, Cole; Lyden, Elizabeth

2017-01-01

Abstract Background. Blood culture contamination is a clinically significant problem that results in patient harm and excess cost. Methods. In a prospective, controlled trial at an academic center Emergency Department, a device that diverts and sequesters the initial 1.5–2 mL portion of blood (which presumably carries contaminating skin cells and microbes) was tested against standard phlebotomy procedures in patients requiring blood cultures due to clinical suspicion of serious infection. Results. In sum, 971 subjects granted informed consent and were enrolled resulting in 904 nonduplicative subjects with 1808 blood cultures. Blood culture contamination was significantly reduced through use of the initial specimen diversion device™ (ISDD) compared to standard procedure: (2/904 [0.22%] ISDD vs 16/904 [1.78%] standard practice, P = .001). Sensitivity was not compromised: true bacteremia was noted in 65/904 (7.2%) ISDD vs 69/904 (7.6%) standard procedure, P = .41. No needlestick injuries or potential bloodborne pathogen exposures were reported. The monthly rate of blood culture contamination for all nurse-drawn and phlebotomist-drawn blood cultures was modeled using Poisson regression to compare the 12-month intervention period to the 6 month before and after periods. Phlebotomists (used the ISDD) experienced a significant decrease in blood culture contamination while the nurses (did not use the ISDD) did not. In sum, 73% of phlebotomists completed a post-study anonymous survey and widespread user satisfaction was noted. Conclusions. Use of the ISDD was associated with a significant decrease in blood culture contamination in patients undergoing blood cultures in an Emergency Department setting. Clinical Trials Registration. NCT02102087. PMID:28379370

An affordable method to obtain cultured endothelial cells from peripheral blood

PubMed Central

Bueno-Betí, Carlos; Novella, Susana; Lázaro-Franco, Macarena; Pérez-Cremades, Daniel; Heras, Magda; Sanchís, Juan; Hermenegildo, Carlos

2013-01-01

The culture of endothelial progenitor cells (EPC) provides an excellent tool to research on EPC biology and vascular regeneration and vasculogenesis. The use of different protocols to obtain EPC cultures makes it difficult to obtain comparable results in different groups. This work offers a systematic comparison of the main variables of most commonly used protocols for EPC isolation, culture and functional evaluation. Peripheral blood samples from healthy individuals were recovered and mononuclear cells were cultured. Different recovery and culture conditions were tested: blood volume, blood anticoagulant, coating matrix and percentage of foetal bovine serum (FBS) in culture media. The success of culture procedure, first colonies of endothelial cells appearance time, correlation with number of circulating EPC (cEPC) and functional comparison with human umbilical vein endothelial cells (HUVEC) were studied. The use of heparin, a minimum blood volume of 30 ml, fibronectin as a coating matrix and endothelial growing media-2 supplemented with 20% FBS increased the success of obtaining EPC cultures up to 80% of the processed samples while reducing EPC colony appearance mean time to a minimum of 13 days. Blood samples exhibiting higher cEPC numbers resulted in reduced EPC colony appearance mean time. Cells isolated by using this combination were endothelial cell-like EPCs morphological and phenotypically. Functionally, cultured EPC showed decreased growing and vasculogenic capacity when compared to HUVEC. Thus, above-mentioned conditions allow the isolation and culture of EPC with smaller blood volumes and shorter times than currently used protocols. PMID:24118735

Assessment of anaerobic blood cultures in pediatric oncology patients.

PubMed

Monsonís Cabedo, Manuel; Rives Solá, Susana; Noguera-Julian, Antoni; Urrea Ayala, Mireia; Cruz Martinez, Ofelia; Gené Giralt, Amadeu

2017-01-01

The routine use of a single aerobic bottle for blood culture in pediatric patients has become commonplace, as anaerobic bacteria are not frequently involved in clinically significant infections. The aim of this study was to assess the usefulness of routinely performing anaerobic blood cultures in pediatric oncology patients. Prospective study was conducted on pediatric (<18 years) patients affected with febrile syndrome after receiving chemotherapy for hematological or solid malignancies. Samples were inoculated into pediatric aerobic and standard anaerobic bottles (BacT/Alert automatic system). Strains were considered clinically significant, or deemed as contaminants, depending on isolation circumstances and clinical criteria. A total of 876 blood cultures from 228 patients were processed during the 21-month study period (January 2014 to September 2015). Baseline diagnosis included 143 solid tumors and 67/18 cases of leukemia/lymphoma. Bacterial growth was detected in 90 (10.2%) blood cultures for 95 different isolates, of which 62 (7.1%)/63 isolates were considered clinically significant. Among the latter, 38 (60.3%) microorganisms grew in both aerobic and anaerobic bottles, 18 (28.6%) only in aerobic bottles, and 7 (11.1%) only in anaerobic bottles. Gram-negative bacilli (33; 52.4%), mainly from the Enterobacteriaceae family, were the most frequently isolated microorganisms. Overall, only 3 out of 90 isolates (3.3%) were strict anaerobes (Propionibacterium acnes), and all of them were deemed contaminants. Strict anaerobes did not cause significant infections in febrile pediatric oncology patients, and anaerobic blood culture bottles offered no additional advantages over aerobic media. Our results suggest that routine blood cultures should be solely processed in aerobic media in this group of patients. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Does this adult patient with suspected bacteremia require blood cultures?

PubMed

Coburn, Bryan; Morris, Andrew M; Tomlinson, George; Detsky, Allan S

2012-08-01

Clinicians order blood cultures liberally among patients in whom bacteremia is suspected, though a small proportion of blood cultures yield true-positive results. Ordering blood cultures inappropriately may be both wasteful and harmful. To review the accuracy of easily obtained clinical and laboratory findings to inform the decision to obtain blood cultures in suspected bacteremia. A MEDLINE and EMBASE search (inception to April 2012) yielded 35 studies that met inclusion criteria for evaluating the accuracy of clinical variables for bacteremia in adult immunocompetent patients, representing 4566 bacteremia and 25,946 negative blood culture episodes. Data were extracted to determine the prevalence and likelihood ratios (LRs) of findings for bacteremia. The pretest probability of bacteremia varies depending on the clinical context, from low (eg, cellulitis: 2%) to high (eg, septic shock: 69%). Elevated temperatures alone do not accurately predict bacteremia (for ≥38°C [>100.3°F], LR, 1.9 [95% CI, 1.4-2.4]; for ≥38.5°C [>101.2°F], LR, 1.4 [95% CI, 1.1-2.0]), nor does isolated leukocytosis (LR, <1.7). The severity of chills graded on an ordinal scale (shaking chills, LR, 4.7; 95% CI, 3.0-7.2) may be more useful. Both the systemic inflammatory response syndrome (SIRS) and a multivariable decision rule with major and minor criteria are sensitive (but not specific) predictors of bacteremia (SIRS, negative LR, 0.09 [95% CI, 0.03-0.26]; decision rule, negative LR, 0.08 [95% CI, 0.04-0.17]). Blood cultures should not be ordered for adult patients with isolated fever or leukocytosis without considering the pretest probability. SIRS and the decision rule may be helpful in identifying patients who do not need blood cultures. These conclusions do not apply to immunocompromised patients or when endocarditis is suspected.

Utility of blood cultures in children admitted to hospital with community-acquired pneumonia.

PubMed

Davis, Tessa R; Evans, Hannah R; Murtas, Jennifer; Weisman, Aimee; Francis, J Lynn; Khan, Ahmed

2017-03-01

The aim of the study was to assess the utility of blood cultures in children admitted to hospital with community-acquired pneumonia. The primary outcome was the number of positive blood culture results, and secondary outcomes included the effect of positive blood culture results on management, and the identification of other clinical/biochemical variables that could predict blood culture results or the course of illness. A retrospective data analysis was carried out on all children admitted to Gosford Hospital during the 2-year period from July 2013 to June 2015. Included were patients under 16 years old who had a diagnosis-related group code of pneumonia. A review of blood culture results, chest X-ray, serology, C-reactive protein and white cell count and clinical outcomes were analysed. There were 215 paediatric admissions with a diagnosis of pneumonia during the 2-year study period. A blood culture was collected in 82.3% (177/215). Although seven had a positive blood culture, only two of these were finally reported as true positives and both were Streptococcus pneumoniae. Both patients were treated with a cephalosporin and demonstrated clinical improvement. No changes were made to their treatment based on the blood culture results. Blood cultures have a low yield and do not appear to be helpful when collected in all patients admitted to hospital with community-acquired pneumonia. © 2016 Paediatrics and Child Health Division (The Royal Australasian College of Physicians).

What are the critical steps in processing blood cultures? A prospective audit evaluating current practice of reporting blood cultures in a centralised laboratory serving secondary care hospitals.

PubMed

Meda, Manjula; Clayton, James; Varghese, Reela; Rangaiah, Jayakeerthi; Grundy, Clive; Dashti, Farnaz; Garner, David; Groves, Katherine; Fitzmaurice, Karen; Hutley, E

2017-04-01

To assess current procedures of processing positive blood cultures against national standards with an aim to evaluate its clinical impact and to determine the utility of currently available rapid identification and susceptibility tests in processing of blood cultures. Blood cultures from three secondary care hospitals, processed at a centralised laboratory, were prospectively audited. Data regarding processing times, communication with prescribers, changes to patient management and mortality within 30 days of a significant blood culture were collected in a preplanned pro forma for a 4-week period. Of 2206 blood cultures, 211 positive blood cultures flagged positive. Sixty-nine (3.1%) of all cultures were considered to be contaminated. Fifty per cent of blood cultures that flagged positive had a Gram stain reported within 2 hours. Two (0.99%) patients with a significant bacteraemia had escalation of antimicrobial treatment at the point of reporting the Gram stain that was subsequently deemed necessary once sensitivity results were known. Most common intervention was de-escalation of therapy for Gram-positive organisms at the point of availability of pathogen identification (25.6% in Gram positive vs 10% in Gram negative; p=0.012). For Gram-negative organisms, the most common intervention was de-escalation of therapy at the point of availability of sensitivity results (43% in Gram negatives vs 17.9% in Gram positive; p=0.0097). Overall mortality within 30 days of a positive blood culture was 10.9% (23/211). Antibiotic resistance may have contributed to mortality in four of these patients (three Gram negative and one Gram positive). Gram stain result had the least impact on antibiotic treatment interventions (escalation or de-escalation). Tests that improve identification time for Gram-positive pathogens and sensitivity time for Gram-negative pathogens had the greatest impact in making significant changes to antimicrobial treatment. Published by the BMJ

Pathogens causing blood stream infections and their drug susceptibility profile in immunocompromised patients.

PubMed

Fayyaz, Muhammad; Mirza, Irfan Ali; Ikram, Aamer; Hussain, Aamir; Ghafoor, Tahir; Shujat, Umer

2013-12-01

To determine the types of pathogens causing blood stream infections and their drug susceptibility profile in immunocompromised patients. Cross-sectional, observational study. Department of Microbiology, Armed Forces Institute of Pathology, Rawalpindi, from January to September 2012. Blood culture bottles received from immunocompromised patients were dealt by two methods, brain heart infusion (BHI) broth based manual method and automated BACTEC system. The samples yielding positive growth from either of two methods were further analyzed. The identification of isolates was done with the help of biochemical reactions and rapid tests. Antimicrobial susceptibility of the isolates was carried out as per recommendations of Clinical and Laboratory Standards Institute (CLSI). Out of the 938 blood culture specimens received from immunocompromised patients, 188 (20%) yielded positive growth. Out of these, 89 (47.3%) isolates were Gram positive and Gram negative each, while 10 (5.3%) isolates were fungi (Candida spp.). In case of Gram positive isolates, 75 (84.3%) were Staphylococcus spp. and 51 (67%) were Methicillin resistant. Amongst Gram negative group 49 (55.1%) isolates were of enterobacteriaceae family, while 40 (44.9%) were non-lactose fermenters (NLF). In vitro antimicrobial susceptibility of Staphylococci revealed 100% susceptibility to vancomycin and linezolid. The enterobacteriaceae isolates had better susceptibility against amikacin 85.7% compared to tigecycline 61.2% and imipenem 59.2%. For NLF, the in vitro efficacy of aminoglycosides was 72.5%. The frequency of Gram positive and Gram negative organisms causing blood stream infections in immunocompromised patients was equal. Vancomycin in case of Gram positive and amikacin for Gram negative organisms revealed better in vitro efficacy as compared to other antibiotics.

Routine blood cultures in the management of pyelonephritis in pregnancy for improving outcomes.

PubMed

Gomi, Harumi; Goto, Yoshihito; Laopaiboon, Malinee; Usui, Rie; Mori, Rintaro

2015-02-13

Pyelonephritis is a type of urinary tract infection (UTI) that affects the upper urinary tract and kidneys, and is one of the most common conditions for hospitalisation among pregnant women, aside from delivery. Samples of urine and blood are obtained and used for cultures as part of the diagnosis and management of the condition. Acute pyelonephritis requires hospitalisation with intravenous administration of antimicrobial agents. Several studies have questioned the necessity of obtaining blood cultures in addition to urine cultures, citing cost and questioning whether blood testing is superfluous. Pregnant women with bacteraemia require a change in the initial empirical treatment based on the blood culture. However, these cases are not common, and represent approximately 15% to 20% of cases. It is unclear whether blood cultures are essential for the effective management of the condition. To assess the effectiveness of routine blood cultures to improve health outcomes in the management of pyelonephritis in pregnant women. We searched the Cochrane Pregnancy and Childbirth Group's Trials Register without language or date restrictions (31 December 2014). Randomised controlled trials and quasi-randomised trials comparing outcomes among pregnant women with pyelonephritis who received initial management with or without blood cultures. Cluster-randomised trials were eligible for inclusion in this review but none were identified. Clinical trials using a cross-over design were not eligible for inclusion. Two review authors independently assessed one trial report for inclusion. We identified one trial report but this was excluded. No clinical trials met the inclusion criteria for this review. There are no large-scale randomised controlled trials to assess outcomes in the management of pyelonephritis in pregnancy with or without blood cultures. Randomised controlled trials are needed to evaluate the effectiveness of managing pyelonephritis in pregnant women with or without

Cultured blood versus donated blood: long-run perspectives of the economy of blood.

PubMed

Mercier Ythier, Jean

2015-01-01

Recent advances of fundamental research on the in vitro generation of red blood cells (RBCs) from hematopoietic stem cells in the laboratory open new possibilities of the utilization of cultured RBCs in transfusion medicine. We study the economic challenge of the setup and development of the mass industrial production of RBCs in mature transfusion organizations. We argue that: (i) RBC manufacturing could be set up and developed in the short-medium run for the treatment of the small proportion of transfused patients who have a rare blood type or are alloimmunized against blood antigens; (ii) manufactured RBCs could substitute for donated RBCs in the long run if the physical productivity of RBC engineering technology approaches that of bone marrow.

Factors affecting Brucella spp. blood cultures positivity in children.

PubMed

Apa, Hurşit; Devrim, Ilker; Memur, Seyma; Günay, Ilker; Gülfidan, Gamze; Celegen, Mehmet; Bayram, Nuri; Karaarslan, Utku; Bağ, Ozlem; Işgüder, Rana; Oztürk, Aysel; Inan, Seyhan; Unal, Nurrettin

2013-03-01

Brucella infections have a wide spectrum of symptoms especially in children, making the diagnosis a complicated process. The gold standard for the final diagnosis for brucellosis is to identify the Brucella spp. isolated from blood or bone marrow cultures. The main purpose of this work was to evaluate the factors affecting the isolation of Brucella spp. from blood cultures. In our study, the ratio of fever, presence of hepatomegaly, and splenomegaly were found to be higher in the bacteremic group. In addition, C-reactive protein levels and liver function enzymes were found to be higher in the bacteremic group. In our opinion, while evaluating the febrile child with suspected Brucella infection, we highly recommend sampling blood cultures regardless of the history of previous antimicrobial therapy and duration of the symptoms.

Rapid identification of fungal pathogens in BacT/ALERT, BACTEC, and BBL MGIT media using polymerase chain reaction and DNA sequencing of the internal transcribed spacer regions.

PubMed

Pryce, Todd M; Palladino, Silvano; Price, Diane M; Gardam, Dianne J; Campbell, Peter B; Christiansen, Keryn J; Murray, Ronan J

2006-04-01

We report a direct polymerase chain reaction/sequence (d-PCRS)-based method for the rapid identification of clinically significant fungi from 5 different types of commercial broth enrichment media inoculated with clinical specimens. Media including BacT/ALERT FA (BioMérieux, Marcy l'Etoile, France) (n = 87), BACTEC Plus Aerobic/F (Becton Dickinson, Microbiology Systems, Sparks, MD) (n = 16), BACTEC Peds Plus/F (Becton Dickinson) (n = 15), BACTEC Lytic/10 Anaerobic/F (Becton Dickinson) (n = 11) bottles, and BBL MGIT (Becton Dickinson) (n = 11) were inoculated with specimens from 138 patients. A universal DNA extraction method was used combining a novel pretreatment step to remove PCR inhibitors with a column-based DNA extraction kit. Target sequences in the noncoding internal transcribed spacer regions of the rRNA gene were amplified by PCR and sequenced using a rapid (24 h) automated capillary electrophoresis system. Using sequence alignment software, fungi were identified by sequence similarity with sequences derived from isolates identified by upper-level reference laboratories or isolates defined as ex-type strains. We identified Candida albicans (n = 14), Candida parapsilosis (n = 8), Candida glabrata (n = 7), Candida krusei (n = 2), Scedosporium prolificans (n = 4), and 1 each of Candida orthopsilosis, Candida dubliniensis, Candida kefyr, Candida tropicalis, Candida guilliermondii, Saccharomyces cerevisiae, Cryptococcus neoformans, Aspergillus fumigatus, Histoplasma capsulatum, and Malassezia pachydermatis by d-PCRS analysis. All d-PCRS identifications from positive broths were in agreement with the final species identification of the isolates grown from subculture. Earlier identification of fungi using d-PCRS may facilitate prompt and more appropriate antifungal therapy.

Evaluation of the HB&L System for the Microbiological Screening of Storage Medium for Organ-Cultured Corneas.

PubMed

Camposampiero, D; Grandesso, S; Zanetti, E; Mazzucato, S; Solinas, M; Parekh, M; Frigo, A C; Gion, M; Ponzin, D

2013-01-01

Aims. To compare HB&L and BACTEC systems for detecting the microorganisms contaminating the corneal storage liquid preserved at 31°C. Methods. Human donor corneas were stored at 4°C followed by preservation at 31°C. Samples of the storage medium were inoculated in BACTEC Peds Plus/F (aerobic microorganisms), BACTEC Plus Anaerobic/F (anaerobic microorganisms), and HB&L bottles. The tests were performed (a) after six days of storage, (b) end of storage, and (c) after 24 hours of preservation in deturgescent liquid sequentially. 10,655 storage and deturgescent media samples were subjected to microbiological control using BACTEC (6-day incubation) and HB&L (24-hour incubation) systems simultaneously. BACTEC positive/negative refers to both/either aerobic and anaerobic positives/negatives, whereas HB&L can only detect the aerobic microbes, and therefore the positives/negatives depend on the presence/absence of aerobic microorganisms. Results. 147 (1.38%) samples were identified positive with at least one of the two methods. 127 samples (134 identified microorganisms) were positive with both HB&L and BACTEC. 14 HB&L+/BACTEC- and 6 BACTEC+/HB&L- were identified. Sensitivity (95.5%), specificity (99.8%), and positive (90.1%) and negative predictive values (99.9%) were high with HB&L considering a 3.5% annual contamination rate. Conclusion. HB&L is a rapid system for detecting microorganisms in corneal storage medium in addition to the existing methods.

Comparative evaluation of paired blood culture (aerobic/aerobic) and single blood culture, along with clinical importance in catheter versus peripheral line at a tertiary care hospital.

PubMed

Tarai, B; Das, P; Kumar, D; Budhiraja, S

2012-01-01

Paired blood culture (PBC) is uncommon practice in hospitals in India, leading to delayed and inadequate diagnosis. Also contamination remains a critical determinant in hampering the definitive diagnosis. To establish the need of PBC over single blood culture (SBC) along with the degree of contamination, this comparative retrospective study was initiated. We processed 2553 PBC and 4350 SBC in BacT/ALERT 3D (bioMerieux) between October 2010 and June 2011. The positive cultures were identified in VITEK 2 Compact (bioMerieux). True positivity and contaminants were also analyzed in 486 samples received from catheter and peripheral line. Out of 2553 PBC samples, positivity was seen in 350 (13.70%). In 4350 SBC samples, positivity was seen in 200 samples (4.59%). In PBC true pathogens were 267 (10.45%) and contaminants were 83 (3.25%), whereas in SBC 153 (3.51%) were true positives and contaminants were 47 (1.08%). Most of the blood cultures (99.27 %) grew within 72 h and 95.8% were isolated within 48 h. In 486 PBCs received from catheter/periphery (one each), catheter positivity was found in 85 (true positives were 48, false positives 37). In peripheral samples true positives were 50 and false positives were 8. Significantly higher positive rates were seen in PBCs compared with SBCs. Automated blood culture and identification methods significantly reduced the time required for processing of samples and also facilitated yield of diverse/rare organisms. Blood culture from catheter line had higher false positives than peripheral blood culture. Thus every positive result from a catheter must be correlated with clinical findings and requires further confirmation.

Proposing an Empirically Justified Reference Threshold for Blood Culture Sampling Rates in Intensive Care Units

PubMed Central

Castell, Stefanie; Schwab, Frank; Geffers, Christine; Bongartz, Hannah; Brunkhorst, Frank M.; Gastmeier, Petra; Mikolajczyk, Rafael T.

2014-01-01

Early and appropriate blood culture sampling is recommended as a standard of care for patients with suspected bloodstream infections (BSI) but is rarely taken into account when quality indicators for BSI are evaluated. To date, sampling of about 100 to 200 blood culture sets per 1,000 patient-days is recommended as the target range for blood culture rates. However, the empirical basis of this recommendation is not clear. The aim of the current study was to analyze the association between blood culture rates and observed BSI rates and to derive a reference threshold for blood culture rates in intensive care units (ICUs). This study is based on data from 223 ICUs taking part in the German hospital infection surveillance system. We applied locally weighted regression and segmented Poisson regression to assess the association between blood culture rates and BSI rates. Below 80 to 90 blood culture sets per 1,000 patient-days, observed BSI rates increased with increasing blood culture rates, while there was no further increase above this threshold. Segmented Poisson regression located the threshold at 87 (95% confidence interval, 54 to 120) blood culture sets per 1,000 patient-days. Only one-third of the investigated ICUs displayed blood culture rates above this threshold. We provided empirical justification for a blood culture target threshold in ICUs. In the majority of the studied ICUs, blood culture sampling rates were below this threshold. This suggests that a substantial fraction of BSI cases might remain undetected; reporting observed BSI rates as a quality indicator without sufficiently high blood culture rates might be misleading. PMID:25520442

Effectiveness of introducing blood culture collection packs to reduce contamination rates.

PubMed

Bamber, A I; Cunniffe, J G; Nayar, D; Ganguly, R; Falconer, E

2009-01-01

Contaminated blood cultures result in a significant waste of healthcare resources and can lead to inappropriate antibiotic therapy. Practitioners have taken measures to reduce contamination rates. These include thorough skin disinfection, effective hand decontamination, introduction of a standardised approach to collection, and the introduction of blood culture collection packs (BCCP). This study aims to assess the impact of introducing BCCP and staff training on the rate of contamination. The study demonstrated that contamination rates are greatest in high patient throughput units where practitioners are under most pressure. The introduction of blood culture packs and staff training has reduced contamination rate significantly from 43% to 25% of the total number of positives, equating to an overall reduction of 42%. Thus, there is a demonstrable benefit in the purchase of commercially produced blood culture packs and the investment in staff training.

What proportion of Salmonella Typhi cases are detected by blood culture? A systematic literature review.

PubMed

Mogasale, Vittal; Ramani, Enusa; Mogasale, Vijayalaxmi V; Park, JuYeon

2016-05-17

Blood culture is often used in definitive diagnosis of typhoid fever while, bone marrow culture has a greater sensitivity and considered reference standard. The sensitivity of blood culture measured against bone marrow culture results in measurement bias because both tests are not fully sensitive. Here we propose a combination of the two cultures as a reference to define true positive S. Typhi cases. Based on a systematic literature review, we identified ten papers that had performed blood and bone marrow culture for S. Typhi in same subjects. We estimated the weighted mean of proportion of cases detected by culture measured against true S. Typhi positive cases using a random effects model. Of 529 true positive S. Typhi cases, 61 % (95 % CI 52-70 %) and 96 % (95 % CI 93-99 %) were detected by blood and bone marrow cultures respectively. Blood culture sensitivity was 66 % (95 % CI 56-75 %) when compared with bone marrow culture results. The use of blood culture sensitivity as a proxy measure to estimate the proportion of typhoid fever cases detected by blood culture is likely to be an underestimate. As blood culture sensitivity is used as a correction factor in estimating typhoid disease burden, epidemiologists and policy makers should account for the underestimation.

Blood cultures for women hospitalized with acute pelvic inflammatory disease. Are they necessary?

PubMed

Apuzzio, J J; Hessami, S; Rodriguez, P

2001-09-01

To determine the incidence of positive blood cultures and if the results affect the clinical management or the duration of hospital stay in patients with acute pelvic inflammatory disease (PID). Retrospective study of all patients hospitalized with a diagnosis of acute PID from January 1, 1996, to December 31, 1997. Of 93 patients in the study, 3 had significant bacterial growth from blood culture specimens. The results of blood culture specimens did not affect clinical management. Routine specimens for blood culture may not be needed from patients hospitalized with acute PID.

Simple method for culture of peripheral blood lymphocytes of Testudinidae.

PubMed

Silva, T L; Silva, M I A; Venancio, L P R; Zago, C E S; Moscheta, V A G; Lima, A V B; Vizotto, L D; Santos, J R; Bonini-Domingos, C R; Azeredo-Oliveira, M T V

2011-12-06

We developed and optimized a simple, efficient and inexpensive method for in vitro culture of peripheral blood lymphocytes from the Brazilian tortoise Chelonoidis carbonaria (Testudinidae), testing various parameters, including culture medium, mitogen concentration, mitotic index, culture volume, incubation time, and mitotic arrest. Peripheral blood samples were obtained from the costal vein of four couples. The conditions that gave a good mitotic index were lymphocytes cultured at 37°C in minimum essential medium (7.5 mL), with phytohemagglutinin as a mitogen (0.375 mL), plus streptomycin/penicillin (0.1 mL), and an incubation period of 72 h. Mitotic arrest was induced by 2-h exposure to colchicine (0.1 mL), 70 h after establishing the culture. After mitotic arrest, the cells were hypotonized with 0.075 M KCl for 2 h and fixed with methanol/acetic acid (3:1). The non-banded mitotic chromosomes were visualized by Giemsa staining. The diploid chromosome number of C. carbonaria was found to be 52 in females and males, and sex chromosomes were not observed. We were able to culture peripheral blood lymphocytes of a Brazilian tortoise in vitro, for the preparation of mitotic chromosomes.

Clinical impact of reducing routine blood culture incubation time from 7 to 5 days.

PubMed

Marginson, Michael J; Daveson, Kathryn L; Kennedy, Karina J

2014-12-01

The aim of this study was to determine the clinical impact of reducing the blood culture incubation protocol from 7 to 5 days.A laboratory data extraction identified positive blood cultures occurring after 5 or more days of incubation at Canberra Hospital, Australia between 1 January 2001 and 31 August 2011. Isolates were identified as clinically significant using a pre-existing prospective bacteraemia database. Medical records review determined whether the positive result affected clinical management.Positive blood cultures after 5 or more days of incubation accounted for 2.65% (423/15979) of all positive blood cultures, although the majority were false positives or contaminants. Eighty-five were significant/indeterminate, representing an average of eight cases per year or 0.47% (85/15979) of all positive blood cultures sets. Forty-three were isolated for the first time, representing 1.1% of all significant/indeterminate blood culture episodes. Fungi and anaerobic bacteria compromised over 50% of isolates. In 26 cases (2.4 cases per year), the culture result led to a change in patient management.A 7 day incubation protocol is preferable due to late isolation of organisms; however, if space is required to accommodate increasing blood culture numbers, reducing to a 5 or 6 day protocol would miss only a small percentage of clinically significant isolates.

Blood culture contamination in hospitalized pediatric patients: a single institution experience

PubMed Central

Min, Hyewon; Park, Cheong Soo; Kim, Dong Soo

2014-01-01

Purpose Blood culture is the most important tool for detecting bacteremia in children with fever. However, blood culture contamination rates range from 0.6% to 6.0% in adults; rates for young children have been considered higher than these, although data are limited, especially in Korea. This study determined the contamination rate and risk factors in pediatric patients visiting the emergency room (ER) or being admitted to the ward. Methods We conducted a retrospective chart review of blood cultures obtained from children who visited Yonsei Severance Hospital, Korea between 2006 and 2010. Positive blood cultures were labeled as true bacteremia or contamination according to Centers for Disease Control and Prevention/National Healthcare Safety Network definitions for laboratory-confirmed bloodstream infection, after exclusion of cultures drawn from preexisting central lines only. Results Among 40,542 blood cultures, 610 were positive, of which 479 were contaminations and 131 were true bacteremia (overall contamination rate, 1.18%). The contamination rate in the ER was significantly higher than in the ward (1.32% vs. 0.66%, P<0.001). The rate was higher in younger children (2.07%, 0.94%, and 0.61% in children aged <1 year, 1-6 years, and >6 years, respectively). Conclusion Overall, contamination rates were higher in younger children than in older children, given the difficulty of performing blood sampling in younger children. The contamination rates from the ER were higher than those from the ward, not accounted for only by overcrowding and lack of experience among personnel collecting samples. Further study to investigate other factors affecting contamination should be required. PMID:24868215

[EXPRESS IDENTIFICATION OF POSITIVE BLOOD CULTURES USING DIRECT MALDI-TOF MASS SPECTROMETRY].

PubMed

Popov, D A; Ovseenko, S T; Vostrikova, T Yu

2015-01-01

To evaluate the effectiveness of direct identification of pathogens of bacteremia by direct matrix assisted laser desorption ionization time-flight mass spectrometry (mALDI-TOF) compared to routine method. A prospective study included 211 positive blood cultures obtained from 116 patients (106 adults and 10 children, aged from 2 weeks to 77 years old in the ICU after open heart surgery. Incubation was carried out under aerobic vials with a sorbent for antibiotics Analyzer BacT/ALERT 3D 120 (bioMerieux, France) in parallel with the primary sieving blood cultures on solid nutrient media with subsequent identification of pure cultures using MALDI-TOF mass spectrometry analyzer Vitek MS, bioMerieux, France routine method), after appropriate sample preparation we carried out a direct (without screening) MALDI-TOF mass spectrometric study of monocomponental blood cultures (n = 201). using a routine method in 211 positive blood cultures we identified 23 types of microorganisms (Staphylococcus (n = 87), Enterobacteria- ceae (n = 71), Enterococci (n = 20), non-fermentative Gram-negative bacteria (n = 18), others (n = 5). The average time of incubation of samples to obtain a signal of a blood culture growth was 16.2 ± 7.4 h (from 3.75 to 51 hours.) During the first 12 hours of incubation, growth was obtained in 32.4% of the samples, and on the first day in 92.2%. In the direct mass spectrometric analysis mnonocomponental blood cultures (n = 201) is well defined up to 153 species of the sample (76.1%), while the share of successful identification of Gram-negative bacteria was higher than that of Gram-positive (85.4 and 69, 1%, respectively p = 0.01). The high degree of consistency in the results of standard and direct method of identifying blood cultures using MALDI-TOF mass spectrometry (κ = 0.96, p < 0.001; the samples included in the calculation for which both option given result). Duration of the direct mass spectrometric analysis, including sample preparation, was no

Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles.

PubMed

Peel, Trisha N; Dylla, Brenda L; Hughes, John G; Lynch, David T; Greenwood-Quaintance, Kerryl E; Cheng, Allen C; Mandrekar, Jayawant N; Patel, Robin

2016-01-05

Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs) is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM) in addition to applying the Infectious Diseases Society of America (IDSA) criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014) at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32%) met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively); this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003). The time to microorganism detection was shorter with BCBs than with standard media (P < 0.0001), with aerobic and anaerobic BCBs yielding positive results within a median of 21 and 23 h, respectively. Results of our study demonstrate that the semiautomated method of periprosthetic tissue culture in blood culture bottles is more sensitive than and as specific as agar and thioglycolate broth cultures and yields results faster. Prosthetic joint infections are a devastating complication of arthroplasty surgery. Despite this, current microbiological techniques to detect and diagnose infections are imperfect. This study examined a new approach to diagnosing infections, through the inoculation of

Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles

PubMed Central

Peel, Trisha N.; Dylla, Brenda L.; Hughes, John G.; Lynch, David T.; Greenwood-Quaintance, Kerryl E.; Cheng, Allen C.; Mandrekar, Jayawant N.

2016-01-01

ABSTRACT Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs) is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM) in addition to applying the Infectious Diseases Society of America (IDSA) criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014) at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32%) met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively); this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003). The time to microorganism detection was shorter with BCBs than with standard media (P < 0.0001), with aerobic and anaerobic BCBs yielding positive results within a median of 21 and 23 h, respectively. Results of our study demonstrate that the semiautomated method of periprosthetic tissue culture in blood culture bottles is more sensitive than and as specific as agar and thioglycolate broth cultures and yields results faster. PMID:26733067

Pseudoepidemic of Nocardia asteroides associated with a mycobacterial culture system.

PubMed Central

Patterson, J E; Chapin-Robertson, K; Waycott, S; Farrel, P; McGeer, A; McNeil, M M; Edberg, S C

1992-01-01

Nocardia isolations increased from 0.7 to 11.7/1,000 acid-fast bacillus and mycological cultures (P less than 0.000001). Only three isolations from one patient represented infection. Pseudoepidemic strain identity was confirmed by DNA fingerprinting; the isolate causing infection was distinct. The end of the pseudoepidemic was associated with changing the needle sterilizer and prolonging needle sterilization time on the BACTEC 460 machine. To our knowledge, this is the first reported Nocardia asteroides pseudoepidemic. Images PMID:1583150

Effect of a training programme on blood culture contamination rate in critical care.

PubMed

Sánchez-Sánchez, M M; Arias-Rivera, S; Fraile-Gamo, P; Jareño-Collado, R; López-Román, S; Vadillo-Obesso, P; García-González, S; Pulido-Martos, M T; Sánchez-Muñoz, E I; Cacho-Calvo, J; Martín-Pellicer, A; Panadero-Del Olmo, L; Frutos-Vivar, F

2018-03-30

Blood culture contamination can occur from extraction to processing; its rate should not exceed 3%. To evaluate the impact of a training programme on the rate of contaminated blood cultures after the implementation of sample extraction recommendations based on the best evidence. Prospective before-after study in a polyvalent intensive care unit with 18 beds. Two phases were established (January-June 2012, October 2012-October 2015) with a training period between them. Main recommendations: sterile technique, surgical mask, double skin disinfection (70° alcohol and 2% alcoholic chlorhexidine), 70° alcohol disinfection of culture flasks and injection of samples without changing needles. Including all blood cultures of patients with extraction request. demographic, severity, pathology, reason for admission, stay and results of blood cultures (negative, positive and contaminated). Basic descriptive statistics: mean (standard deviation), median (interquartile range) and percentage (95% confidence interval). Calculated contamination rates per 100 blood cultures extracted. Bivariate analysis between periods. Four hundred and eight patients were included. Eight hundred and forty-one blood cultures were taken, 33 of which were contaminated. In the demographic variables, severity, diagnosis and stay of patients with contaminated samples, no differences were observed from those with uncontaminated samples. Pre-training vs post-training contamination rates: 14 vs 5.6 per 100 blood cultures extracted (P=.00003). An evidence-based training programme reduced the contamination of samples. It is necessary to continue working on the planning of activities and care to improve the detection of pollutants and prevent contamination of samples. Copyright © 2018 Sociedad Española de Enfermería Intensiva y Unidades Coronarias (SEEIUC). Publicado por Elsevier España, S.L.U. All rights reserved.

Rapid Identification of Pathogens from Positive Blood Cultures by Multiplex PCR using the FilmArray System

PubMed Central

Blaschke, Anne J.; Heyrend, Caroline; Byington, Carrie L.; Fisher, Mark A.; Barker, Elizabeth; Garrone, Nicholas F.; Thatcher, Stephanie A.; Pavia, Andrew T.; Barney, Trenda; Alger, Garrison D.; Daly, Judy A.; Ririe, Kirk M.; Ota, Irene; Poritz, Mark A.

2012-01-01

Sepsis is a leading cause of death. Rapid and accurate identification of pathogens and antimicrobial resistance directly from blood culture could improve patient outcomes. The FilmArray® (FA; Idaho Technology, Inc., Salt Lake City, UT) Blood Culture (BC) panel can identify > 25 pathogens and 4 antibiotic resistance genes from positive blood cultures in 1 hour. We compared a development version of the panel to conventional culture and susceptibility testing on 102 archived blood cultures from adults and children with bacteremia. Of 109 pathogens identified by culture, 95% were identified by FA. Among 111 prospectively collected blood cultures, the FA identified 84 of 92 pathogens (91%) covered by the panel. Among 25 Staphylococcus aureus and 21 Enterococcus species detected, FA identified all culture-proven MRSA and VRE. The FA BC panel is an accurate method for the rapid identification of pathogens and resistance genes from blood culture. PMID:22999332

Evaluating the use of blood cultures in the management of children hospitalized for community-acquired pneumonia.

PubMed

McCulloh, Russell J; Koster, Michael P; Yin, Dwight E; Milner, Tiffany L; Ralston, Shawn L; Hill, Vanessa L; Alverson, Brian K; Biondi, Eric A

2015-01-01

Blood cultures are often recommended for the evaluation of community-acquired pneumonia (CAP). However, institutions vary in their use of blood cultures, and blood cultures have unclear utility in CAP management in hospitalized children. To identify clinical factors associated with obtaining blood cultures in children hospitalized with CAP, and to estimate the association between blood culture obtainment and hospital length of stay (LOS). We performed a multicenter retrospective cohort study of children admitted with a diagnosis of CAP to any of four pediatric hospitals in the United States from January 1, 2011-December 31, 2012. Demographics, medical history, diagnostic testing, and clinical outcomes were abstracted via manual chart review. Multivariable logistic regression evaluated patient and clinical factors for associations with obtaining blood cultures. Propensity score-matched Kaplan-Meier analysis compared patients with and without blood cultures for hospital LOS. Six hundred fourteen charts met inclusion criteria; 390 children had blood cultures obtained. Of children with blood cultures, six (1.5%) were positive for a pathogen and nine (2.3%) grew a contaminant. Factors associated with blood culture obtainment included presenting with symptoms of systemic inflammatory response syndrome (OR 1.78, 95% CI 1.10-2.89), receiving intravenous hydration (OR 3.94, 95% CI 3.22-4.83), receiving antibiotics before admission (OR 1.49, 95% CI 1.17-1.89), hospital admission from the ED (OR 1.65, 95% CI 1.05-2.60), and having health insurance (OR 0.42, 95% CI 0.30-0.60). In propensity score-matched analysis, patients with blood cultures had median 0.8 days longer LOS (2.0 vs 1.2 days, P < .0001) without increased odds of readmission (OR 0.94, 95% CI 0.45-1.97) or death (P = .25). Obtaining blood cultures in children hospitalized with CAP rarely identifies a causative pathogen and is associated with increased LOS. Our results highlight the need to refine the role of

Diagnosis of blood culture-negative endocarditis and clinical comparison between blood culture-negative and blood culture-positive cases.

PubMed

Lamas, Cristiane C; Fournier, Pierre-Edouard; Zappa, Monica; Brandão, Tatiana J D; Januário-da-Silva, Carolina A; Correia, Marcelo G; Barbosa, Giovanna Ianini F; Golebiovski, Wilma F; Weksler, Clara; Lepidi, Hubert; Raoult, Didier

2016-08-01

To analyze the clinical characteristics of blood culture-negative endocarditis (BCNE) and how it compares to those of blood culture-positive endocarditis (BCPE) cases and show how molecular tools helped establish the etiology in BCNE. Adult patients with definite infective endocarditis (IE) and having valve surgery were included. Valves were studied by polymerase chain reaction (PCR). Statistical analysis compared BCNE and BCPE. One hundred and thirty-one patients were included; 53 (40 %) had BCNE. The mean age was 45 ± 16 years; 33 (62 %) were male. BCNE was community-acquired in 41 (79 %). Most patients were referred from other hospitals (38, 73 %). Presentation was subacute in 34 (65 %), with fever in 47/53 (90 %) and a new regurgitant murmur in 34/42 (81 %). Native valves were affected in 74 %, mostly left-sided. All echocardiograms showed major criteria for IE. Antibiotics were used prior to BC collection in 31/42 (74 %). Definite histological diagnosis was established for 35/50 (70 %) valves. PCR showed oralis group streptococci in 21 (54 %), S. aureus in 3 (7.7 %), gallolyticus group streptococci in 2 (5.1 %), Coxiella burnetii in 1 (2.5 %) and Rhizobium sp. in 1 (2.5 %). In-hospital mortality was 9/53 (17 %). Fever (p = 0.06, OR 4.7, CI 0.91-24.38) and embolic complications (p = 0.003, OR 3.3, CI 1.55-6.82) were more frequent in BCPE cases, while new acute regurgitation (p = 0.05, OR 0.3, CI 0.098-0.996) and heart failure (p = 0.02, OR 0.3, CI 0.13-0.79) were less so. BCNE resulted mostly from prior antibiotics and was associated with severe hemodynamic compromise. Valve histopathology and PCR were useful in confirming the diagnosis and pointing to the etiology of BCNE.

Trypanosoma cruzi. Surface antigens of blood and culture forms

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nogueira, N.; Chaplan, S.; Tydings, J.D.

1981-03-01

The surface polypeptides of both cultured and blood forms of Trypanosoma cruzi were iodinated by the glucose oxidase-lactoperoxidase technique. Blood-form trypomastigotes (BFT) isolated form infected mice displayed a major 90,000-Mr component. In contrast, both epimastigotes and trypomastigotes obtained form acellular cultures expressed a smaller 75,000-Mr peptide. Both major surface components were presumably glycoproteins in terms of their binding to concanavalin A-Sepharose 4B. Within a 3-h period, both blood and culture forms synthesized their respective surface glycoproteins (90,000 Mr and 75,000 Mr, respectively in vitro. (/sub 35/S)methionine-labeled surface peptides were immunoprecipitated with immune sera of both human and murine origin. Amore » panel of sera form patients with chronic Chagas' disease and hyperimmunized mice recognized similar surface peptides. These immunogens were the same components as the major iodinated species. The major BFT surface peptide was readily removed by trypsin treatment of the parasites, although the procedure did not affect the 75,000-Mr peptide from the culture forms. Two-dimensional polyacrylamide gel electrophoresis revealed that the 90,000-Mr peptide found on BFT was an acidic protein of isoelectric point (pI) 5.0, whereas, the 75,000-Mr peptide form culture-form trypomastigotes has a pI of 7.2. The 90,000-Mr component is thought to be responsible for the anti-phagocytic properties of the BFT (1).« less

Tumor necrosis factor-alpha (TNF-alpha) concentrations from whole blood cultures correlate with isolated peripheral blood mononuclear cell cultures

USDA-ARS?s Scientific Manuscript database

Many cellular immune assays are impractical because they require labor-intensive isolation of cells from their natural environment. The objectives of this study were to determine the relationship between cell culture supernatant TNF-alpha from isolated peripheral blood mononuclear cells (PBMC) and w...

Minimizing the Workup of Blood Culture Contaminants: Implementation and Evaluation of a Laboratory-Based Algorithm

PubMed Central

Richter, S. S.; Beekmann, S. E.; Croco, J. L.; Diekema, D. J.; Koontz, F. P.; Pfaller, M. A.; Doern, G. V.

2002-01-01

An algorithm was implemented in the clinical microbiology laboratory to assess the clinical significance of organisms that are often considered contaminants (coagulase-negative staphylococci, aerobic and anaerobic diphtheroids, Micrococcus spp., Bacillus spp., and viridans group streptococci) when isolated from blood cultures. From 25 August 1999 through 30 April 2000, 12,374 blood cultures were submitted to the University of Iowa Clinical Microbiology Laboratory. Potential contaminants were recovered from 495 of 1,040 positive blood cultures. If one or more additional blood cultures were obtained within ±48 h and all were negative, the isolate was considered a contaminant. Antimicrobial susceptibility testing (AST) of these probable contaminants was not performed unless requested. If no additional blood cultures were submitted or there were additional positive blood cultures (within ±48 h), a pathology resident gathered patient clinical information and made a judgment regarding the isolate's significance. To evaluate the accuracy of these algorithm-based assignments, a nurse epidemiologist in approximately 60% of the cases performed a retrospective chart review. Agreement between the findings of the retrospective chart review and the automatic classification of the isolates with additional negative blood cultures as probable contaminants occurred among 85.8% of 225 isolates. In response to physician requests, AST had been performed on 15 of the 32 isolates with additional negative cultures considered significant by retrospective chart review. Agreement of pathology resident assignment with the retrospective chart review occurred among 74.6% of 71 isolates. The laboratory-based algorithm provided an acceptably accurate means for assessing the clinical significance of potential contaminants recovered from blood cultures. PMID:12089259

All in the Blood: A Review of Aboriginal Australians' Cultural Beliefs About Blood and Implications for Biospecimen Research.

PubMed

Kowal, Emma; Greenwood, Ashley; McWhirter, Rebekah E

2015-10-01

Public participation in medical research and biobanking is considered key to advances in scientific discovery and translation to improved health care. Cultural concerns relating to blood have been found to affect the participation of indigenous peoples and minorities in research, but such concerns are rarely specified in the literature. This article presents a review of the role of blood in Australian Aboriginal cultures. We discuss the range of meanings and uses of blood in traditional culture, including their use in ceremonies, healing, and sorcery. We draw on more recent literature on Aboriginal Australians and biomedicine to consider how traditional beliefs may be changing over time. These findings provide an empirical basis for researchers and bioethicists to develop culturally grounded strategies to boost the participation of Aboriginal Australians in biomedical research. They also serve as a model for integrating anthropological literature with bioethical concerns that could be applied to other indigenous and minority groups. © The Author(s) 2015.

Contamination of blood cultures during venepuncture: fact or myth?

PubMed Central

Shahar, E.; Wohl-Gottesman, B. S.; Shenkman, L.

1990-01-01

Contamination of blood cultures is believed to occur mainly during the venepuncture procedure. Consequently, meticulous preparation of the venepuncture site is widely recommended. To determine whether the contamination rate is indeed affected by the quality of the antiseptic procedure at the venepuncture site, 181 paired cultures were collected from 176 patients during a 6-month period after either strict antiseptic cleansing of skin with alcohol followed by povidone-iodine, or after brief disinfection with alcohol alone. The contamination rate was not influenced by the antiseptic procedure, and corresponded to the accepted percentage reported in most other studies. Eight false positive cultures (4.4%) were obtained after strict antisepsis of the skin and 6 (3.3%) after short simple cleansing with alcohol (P = 0.39). Our results suggest that contamination of blood cultures may not be related to the venepuncture procedure--regardless of the antiseptic technique used--but may be due to later stages of laboratory handling and processing of the specimens. Review of the literature has provided further indirect evidence to support this conclusion. PMID:2084653

Accuracy of blood culture for early diagnosis of mediastinitis in febrile patients after cardiac surgery.

PubMed

San Juan, R; Aguado, J M; López, M J; Lumbreras, C; Enriquez, F; Sanz, F; Chaves, F; López-Medrano, F; Lizasoain, M; Rufilanchas, J J

2005-03-01

Postsurgical mediastinitis (PSM) remains a major cause of morbidity and mortality in patients undergoing cardiac surgery procedures. Although prompt diagnosis is crucial in these patients, neither clinical data nor imaging techniques have shown enough sensitivity or specificity for early diagnosis of PSM. The aim of the present study was to assess the validity of blood cultures as a diagnostic test for the early detection of PSM in patients who become febrile after cardiac surgery procedures. During a 4-year period (1999-2002), patients who developed fever (>37.8 degrees C) in the first 60 days after a cardiac surgery procedure were evaluated. Blood cultures were drawn from these patients. PSM was defined as deep infection involving retrosternal tissue and/or the sternal bone directly observed by the surgeon and confirmed microbiologically. Three criteria for positivity of blood cultures were applied: bacteremia, staphylococcal bacteremia, or Staphylococcus aureus bacteremia. For purposes of the analysis, a positive blood culture in patients with PSM was considered a true-positive test and a negative blood culture a false-negative test. Otherwise, in febrile patients without PSM in the postsurgery period, a positive blood culture was considered a false-positive test and a negative blood culture a true-negative test. Blood cultures were drawn from 266 febrile patients in the postsurgery period. PSM occurred in 38 patients (26 cases due to S. aureus, 8 to Staphylococcus epidermidis, 3 to gram-negative enteric bacteria, and one to Pseudomonas aeruginosa). Within the 60-day postsurgical period, blood culture as a diagnostic test was most accurate in patients with S. aureus bacteremia, providing 68% sensitivity, 98% specificity, a positive predictive value of 87%, and a negative predictive value of 95%. If the analysis was limited to the period during which patients are at maximum risk for PSM (day 7-20), the values in patients with S. aureus bacteremia were as follows

Trends in blood culture contamination: a College of American Pathologists Q-Tracks study of 356 institutions.

PubMed

Bekeris, Leonas G; Tworek, Joseph A; Walsh, Molly K; Valenstein, Paul N

2005-10-01

Blood culture contamination extends hospital stays and increases the cost of care. To measure blood culture contamination rates in a large number of institutions over time and to elucidate practice patterns and demographic factors associated with sustained reduction in contamination rates. Longitudinal cohort study of 356 clinical laboratories that provided quarterly data about blood culture results, using a uniform definition of contamination. Mixed linear model analysis of the 1999 through 2003 data set. Blood culture contamination was significantly higher in institutions that used nonlaboratory personnel to collect blood (P = .03) and significantly lower in facilities that used a dedicated phlebotomy team (P < .001). Higher volume of blood collection was significantly associated with lower contamination rates (P < .001). Continued participation in the Q-Tracks monitoring program was associated with significant and progressive reduction in contamination rates. By the fifth year of participation, the median institution had reduced its blood culture contamination rate by 0.67% (P < .001). Institutions that use decentralized patient-centered personnel rather than dedicated phlebotomy teams to collect blood cultures experience significantly higher contamination rates. Long-term monitoring of contamination is associated with sustained improvement in performance.

Blood culture accuracy: discards from central venous catheters in pediatric oncology patients in the emergency department.

PubMed

Winokur, Elizabeth J; Pai, Debra; Rutledge, Dana N; Vogel, Kate; Al-Majid, Sadeeka; Marshall, Christine; Sheikewitz, Paul

2014-07-01

Lack of specific guidelines regarding collection of blood for culture from central venous catheters (CVCs) has led to inconsistencies in policies among hospitals. Currently, no specific professional or regulatory recommendations exist in relation to using, reinfusing, or discarding blood drawn from CVCs before drawing blood for a culture. Repeated wasting of blood may harm immunocompromised pediatric oncology patients. The purpose of this comparative study was to determine whether differences exist between blood cultures obtained from the first 5 mL of blood drawn from a CVC line when compared with the second 5 mL drawn. During 2009-2011, 62 pediatric oncology patients with CVCs and orders for blood cultures to determine potential sepsis were enrolled during ED visits. Trained study nurses aseptically drew blood and injected the normally discarded first 5 mL and the second specimen (usual care) into separate culture bottles. Specimens were processed in the microbiology laboratory per hospital policy. Positive cultures were evaluated to assess agreement between specimen results and to determine that the identified pathogen was not a contaminant. Out of 186 blood culture pairs, 4.8% demonstrated positive results. In all positive-positive matches, the normal discard specimen contained the same organism as the usual care specimen. In 4 matches, the normally discarded specimen demonstrated notably earlier time to positivity (4 to 31 hours) compared with the usual care specimen, which resulted in earlier initiation of definitive antibiotics. These findings support the accuracy of the specimen that is normally discarded and suggest the need to reconsider its use for blood culture testing. Copyright © 2014 Emergency Nurses Association. Published by Mosby, Inc. All rights reserved.

Hub qualitative blood culture is useful for diagnosis of catheter-related infections in critically ill patients.

PubMed

Tanguy, Michèle; Seguin, Philippe; Laviolle, Bruno; Desbordes, Laurent; Mallédant, Yannick

2005-05-01

To assess clinical safety and accuracy of qualitative blood culture drawn through the hub for ruling out catheter-related infection (CRI). Prospective observational study in a surgical intensive care unit. All patients with sepsis of unknown origin and possibly due to a CRI. Blood culture drawn through a central venous catheter (CVC) just before the catheter was cultured. In 126 patients we investigated 135 cases of sepsis of unknown origin. Using a clinical and bacteriological approach as the reference, the performance of the CVC blood culture was evaluated by the calculation of sensitivity, specificity, and positive and negative predictive values. When CVC blood culture was positive, the time to positivity was considered. Using standard definitions, seven CRIs were diagnosed. CVC blood culture identified five CRIs including the three episodes of catheter-related bacteremia. The method missed two coagulase-negative staphylococcus CRIs without bacteremia. Thirteen false-positive results occurred, including seven bacteremias from a distant source. The CVC blood culture had a sensitivity of 71% (CI 30-95%), specificity of 90% (CI 83-94%), negative predictive value of 98% (CI 93-100%), and positive predictive value of 28% (CI 11-54%). In cases of catheter-related bacteremia the time to positivity of CVC blood culture was 24 h or less. Negative CVC blood culture at 24 h seems useful for management of CVC in selected critically ill surgical patients developing a clinical sepsis. The subsequent risk of catheter-related bacteremia cannot be excluded advocating for an uninterrupted clinical and bacteriological survey.

Direct Isolation of Candida spp. from Blood Cultures on the Chromogenic Medium CHROMagar Candida

PubMed Central

Horvath, Lynn L.; Hospenthal, Duane R.; Murray, Clinton K.; Dooley, David P.

2003-01-01

CHROMagar Candida is a selective and differential chromogenic medium that has been shown to be useful for identification of Candida albicans, Candida krusei, Candida tropicalis, and perhaps Candida glabrata. Colony morphology and color have been well defined when CHROMagar Candida has been used to isolate yeast directly from clinical specimens, including stool, urine, respiratory, vaginal, oropharyngeal, and esophageal sources. Direct isolation of yeast on CHROMagar Candida from blood cultures has not been evaluated. We evaluated whether the color and colony characteristics produced by Candida spp. on CHROMagar Candida were altered when yeasts were isolated directly from blood cultures. Fifty clinical isolates of Candida were inoculated into aerobic and anaerobic blood culture bottles and incubated at 35°C in an automated blood culture system. When growth was detected, an aliquot was removed and plated onto CHROMagar Candida. As a control, CHROMagar Candida plates were inoculated with the same isolate of yeast grown on Sabouraud dextrose agar simultaneously. No significant difference was detected in color or colony morphology between the blood and control isolates in any of the tested organisms. All C. albicans (n = 12), C. tropicalis (n = 12), C. glabrata (n = 9), and C. krusei (n = 5) isolates exhibited the expected species-specific colony characteristics and color, whether isolated directly from blood or from control cultures. CHROMagar Candida can be reliably used for direct isolation of yeast from blood cultures. Direct isolation could allow mycology laboratories to more rapidly identify Candida spp., enable clinicians to more quickly make antifungal agent selections, and potentially decrease patient morbidity and mortality. PMID:12791890

Study of Umbilical Cord Blood Culture in Diagnosis of Early-onset Sepsis Among Newborns with High-risk Factors.

PubMed

Kalathia, Mitul Babubhai; Shingala, Prakash Ashokbhai; Parmar, Parin Niranjanbhai; Parikh, Yogesh Narenedrabhai; Kalathia, Ila Mitulkumar

2013-10-01

Blood culture is gold standard for diagnosis of neonatal sepsis. Low sensitivity of blood culture is usually due to small volume of blood sample, intrapartum antibiotics, and antibiotics given to newborn before sampling. We evaluated use of Umbilical cord blood culture (UCBC) in diagnosis of neonatal sepsis as compared to peripheral venous blood culture. This study was done in tertiary care teaching hospital during May-June 2012. A total of 45 newborns with presence of two or more risk factors of sepsis were included. Blood sample from placental end of umbilical cord was collected and cultured. Primary outcome was diagnosis of neonatal sepsis by use of umbilical cord blood sample as compared with venous blood sample. Secondary outcome was to compare organisms identified by UCBC and venous blood culture. Sensitivity, specificity, positive and negative predictive values of UCBC were calculated. A total of 24.44% (11 out of 45) high-risk newborns had positive UCBC. A total of 17.8% (8 out of 45) newborns had positive blood culture report. Organisms grown in UCBC were Pseudomonas (45%, 5 out of 11), Acinetobacter (27.27%, 3 out of 11), Escherichia coli (18.18%, 2 out of 11), and Klebsiella (9%, 1 out of 11). UCBC is a good method for diagnosis of neonatal sepsis among high-risk newborns as compared to venous blood culture with a sensitivity of 80% and specificity of 91.43%. Organisms grown are comparable to blood culture samples.

Blood culture contamination in Tanzania, Malawi, and the United States: a microbiological tale of three cities.

PubMed

Archibald, Lennox K; Pallangyo, Kisali; Kazembe, Peter; Reller, L Barth

2006-12-01

We conducted retrospective, comparative analyses of contamination rates for cultures of blood obtained in the emergency rooms of Muhimbili National Hospital (MNH) in Dar es Salaam, Tanzania; Lilongwe Central Hospital (LCH) in central Malawi; and the Duke University Medical Center (DUMC) in the United States. None of the emergency room patients had indwelling intravascular devices at the time that the blood samples for cultures were obtained. In addition, we reviewed the contamination rates for a cohort of patients already hospitalized in the DUMC inpatient medical service, most of whom had indwelling intravascular devices. The bloodstream infection rates among the patients at MNH (n=513) and LCH (n=486) were similar (approximately 28%); the contamination rates at the two hospitals were 1.3% (7/513) and 0.8% (4/486), respectively. Of 54 microorganisms isolated from cultures of blood collected in the DUMC emergency room, 26 (48%) were identified as skin contaminants. Cultures of blood collected in the DUMC emergency room were significantly more likely to yield growth of contaminants than the cultures of blood collected in the emergency rooms at MNH and LCH combined (26/332 versus 11/1,003; P<0.0001) or collected in the DUMC inpatient medical service (26/332 versus 7/283; P<0.01). For the MNH and LCH blood cultures, lower contamination rates were observed when skin was disinfected with isopropyl alcohol plus tincture of iodine rather than isopropyl alcohol plus povidone-iodine. In conclusion, blood culture contamination was minimized in sub-Saharan African hospitals with substantially limited resources through scrupulous attention to aseptic skin cleansing and improved venipuncture techniques. Application of these principles when blood samples for culture are obtained in U.S. hospital emergency rooms should help mitigate blood culture contamination rates and the unnecessary microbiology workup of skin contaminants.

[Prospective evaluation of blood cultures in Medical Faculty Hospital of Blacksea Technical University].

PubMed

Sucu, Nurgün; Caylan, Rahmet; Aydin, Kemalettin; Yilmaz, Gürdal; Aktoz Boz, Gönülden; Köksal, Iftihar

2005-10-01

In this study, the rate of blood culture contamination, bacterial pathogens and their antimicrobial susceptibilities causing nosocomial and community acquired bloodstream infections were investigated prospectively during the period February 2003 to February 2004. In the study period, among the 5994 blood culture samples obtained from 3114 patients, 1091 (18%) yielded positive results. Seventy-four of them (1.2%) were evaluated as false positive, 514 (9%) were pseudobacteremia or contamination. According to patients' clinical features, 503 (8%) blood culture samples were associated with blood stream infections, and 358 of them (71%) were primary episodes. Twohundred and ninetyfour of primary episodes (82%) were hospital acquired and 64 were community acquired bloodstream infections. Staphylococcus aureus was the most frequently isolated agent in the hospital and community acquired bloodstream infections at the rates of 16% and 20%, respectively. In hospital acquired blood stream infections, Escherichia coil (9%) and Pseudomonas aeruginosa (8%); in community acquired bloodstream infections Streptococcus spp (17%) and E. coli (15%) were the other most frequently isolated bacterial agents. Methicillin resistance of S. aureus isolates was determined as 54% in hospital acquired blood stream infections and 25% in community acquired blood stream infections.

[Blood culture negative endocarditis: a diagnostic challenge].

PubMed

Wälli, F; Chuard, C; Regamey, C

2005-10-12

Blood culture negative endocarditis (BCNE) account for about 5% of all cases of endocarditis. Diagnosis and initiation of antimicrobial therapy may be delayed, with a negative impact on clinical outcome. The most common cause of BCNE is antimicrobial drug therapy before blood sampling. Other common causes include slow growing and non cultivable organisms. Identification of the etiologic agent is critical in the management of BCNE and molecular tools such as broad range 16SrRNA PCR technique followed by direct automated sequencing and microorganism-specific PCR are promising. Some authors have proposed to include these techniques among major Duke's criteria for the diagnosis of BCNE.

Reducing Blood Culture Contamination in the Emergency Department: An Interrupted Time Series Quality Improvement Study

PubMed Central

Self, Wesley H.; Speroff, Theodore; Grijalva, Carlos G.; McNaughton, Candace D.; Ashburn, Jacki; Liu, Dandan; Arbogast, Patrick G.; Russ, Stephan; Storrow, Alan B.; Talbot, Thomas R.

2012-01-01

Objectives Blood culture contamination is a common problem in the emergency department (ED) that leads to unnecessary patient morbidity and health care costs. The study objective was to develop and evaluate the effectiveness of a quality improvement (QI) intervention for reducing blood culture contamination in an ED. Methods The authors developed a QI intervention to reduce blood culture contamination in the ED and then evaluated its effectiveness in a prospective interrupted times series study. The QI intervention involved changing the technique of blood culture specimen collection from the traditional clean procedure, to a new sterile procedure, with standardized use of sterile gloves and a new materials kit containing a 2% chlorhexidine skin antisepsis device, a sterile fenestrated drape, a sterile needle, and a procedural checklist. The intervention was implemented in a university-affiliated ED and its effect on blood culture contamination evaluated by comparing the biweekly percentages of blood cultures contaminated during a 48-week baseline period (clean technique), and 48-week intervention period (sterile technique), using segmented regression analysis with adjustment for secular trends and first-order autocorrelation. The goal was to achieve and maintain a contamination rate below 3%. Results During the baseline period, 321 out of 7,389 (4.3%) cultures were contaminated, compared to 111 of 6,590 (1.7%) during the intervention period (p < 0.001). In the segmented regression model, the intervention was associated with an immediate 2.9% (95% CI = 2.2% to 3.2%) absolute reduction in contamination. The contamination rate was maintained below 3% during each biweekly interval throughout the intervention period. Conclusions A QI assessment of ED blood culture contamination led to development of a targeted intervention to convert the process of blood culture collection from a clean to a fully sterile procedure. Implementation of this intervention led to an immediate

Significance of coagulase negative Staphylococcus from blood cultures: persisting problems and partial progress in resource constrained settings.

PubMed

Sidhu, Shailpreet K; Malhotra, Sita; Devi, Pushpa; Tuli, Arpandeep K

2016-12-01

Coagulase negative Staphylococcus (CoNS) is frequently isolated from blood cultures but their significance is difficult to interpret. CoNS bacteria which are often previously dismissed as culture contaminants are attracting greater importance as true pathogens in the past decades. Clinical evaluation of these isolates suggests that although there is a relative increase of CoNS associated bloodstream infections in recent years, the microorganisms still remain the most common contaminants in blood cultures. The objective of this study was to determine the significance of CoNS isolated from blood cultures. A retrospective study was conducted to evaluate the rate of contamination in blood cultures in a tertiary care hospital. The paired specimens of blood were cultured using conventional culture methods and the isolates of coagulase negative staphylococci were identified by standard methodology. Clinical data, laboratory indices, microbiological parameters and patient characteristics were analyzed. Of 3503 blood samples, CoNS were isolated from blood culture of 307 patients (8.76%). The isolates were reported as true pathogens of bloodstream infections in only 74 out of 307 cases (24.1%). In the vast majority, 212 of 307 (69.0%), they were mere blood culture contaminants and reported as insignificant/contaminant. Determining whether a growth in the blood culture is a pathogen or a contaminant is a critical issue and multiple parameters have to be considered before arriving at a conclusion. Ideally, the molecular approach is for the most part a consistent method in determining the significant isolates of CoNS. However, in countries with inadequate resources, species identification and antibiogram tests are recommended when determining significance of these isolates.

Chemical decontamination with N-acetyl-L-cysteine-sodium hydroxide improves recovery of viable Mycobacterium avium subsp. paratuberculosis organisms from cultured milk.

PubMed

Bradner, L; Robbe-Austerman, S; Beitz, D C; Stabel, J R

2013-07-01

Mycobacterium avium subsp. paratuberculosis is shed into the milk and feces of cows with advanced Johne's disease, allowing the transmission of M. avium subsp. paratuberculosis between animals. The objective of this study was to formulate an optimized protocol for the isolation of M. avium subsp. paratuberculosis in milk. The parameters investigated included chemical decontamination with N-acetyl-l-cysteine-sodium hydroxide (NALC-NaOH), alone and in combination with antibiotics (vancomycin, amphotericin B, and nalidixic acid), and the efficacy of solid (Herrold's egg yolk medium [HEY]) and liquid (Bactec 12B and para-JEM) culture media. For each experiment, raw milk samples from a known noninfected cow were inoculated with 10(2) to 10(8) CFU/ml of live M. avium subsp. paratuberculosis organisms. The results indicate that an increased length of exposure to NALC-NaOH from 5 to 30 min and an increased concentration of NaOH from 0.5 to 2.0% did not affect the viability of M. avium subsp. paratuberculosis. Additional treatment of milk samples with the antibiotics following NALC-NaOH treatment decreased the recovery of viable M. avium subsp. paratuberculosis cells more than treatment with NALC-NaOH alone. The Bactec 12B medium was the superior medium of the three evaluated for the isolation of M. avium subsp. paratuberculosis from milk, as it achieved the lowest threshold of detection. The optimal conditions for NALC-NaOH decontamination were determined to be exposure to 1.50% NaOH for 15 min followed by culture in Bactec 12B medium. This study demonstrates that chemical decontamination with NALC-NaOH resulted in a greater recovery of viable M. avium subsp. paratuberculosis cells from milk than from samples treated with hexadecylpyridinium chloride (HPC). Therefore, it is important to optimize milk decontamination protocols to ensure that low concentrations of M. avium subsp. paratuberculosis can be detected.

Blood Culture Testing via a Mobile App That Uses a Mobile Phone Camera: A Feasibility Study

PubMed Central

Chong, Yong Pil; Jang, Seongsoo; Kim, Mi Na; Kim, Jeong Hoon; Kim, Woo Sung

2016-01-01

Background To evaluate patients with fever of unknown origin or those with suspected bacteremia, the precision of blood culture tests is critical. An inappropriate step in the test process or error in a parameter could lead to a false-positive result, which could then affect the direction of treatment in critical conditions. Mobile health apps can be used to resolve problems with blood culture tests, and such apps can hence ensure that point-of-care guidelines are followed and processes are monitored for blood culture tests. Objective In this pilot project, we aimed to investigate the feasibility of using a mobile blood culture app to manage blood culture test quality. We implemented the app at a university hospital in South Korea to assess the potential for its utilization in a clinical environment by reviewing the usage data among a small group of users and by assessing their feedback and the data related to blood culture sampling. Methods We used an iOS-based blood culture app that uses an embedded camera to scan the patient identification and sample number bar codes. A total of 4 medical interns working at 2 medical intensive care units (MICUs) participated in this project, which spanned 3 weeks. App usage and blood culture sampling parameters (including sampler, sampling site, sampling time, and sample volume) were analyzed. The compliance of sampling parameter entry was also measured. In addition, the participants’ opinions regarding patient safety, timeliness, efficiency, and usability were recorded. Results In total, 356/644 (55.3%) of all blood culture samples obtained at the MICUs were examined using the app, including 254/356 (71.3%) with blood collection volumes of 5-7 mL and 256/356 (71.9%) with blood collection from the peripheral veins. The sampling volume differed among the participants. Sampling parameters were completely entered in 354/356 cases (99.4%). All the participants agreed that the app ensured good patient safety, disagreed on its

Blood Culture Testing via a Mobile App That Uses a Mobile Phone Camera: A Feasibility Study.

PubMed

Lee, Guna; Lee, Yura; Chong, Yong Pil; Jang, Seongsoo; Kim, Mi Na; Kim, Jeong Hoon; Kim, Woo Sung; Lee, Jae-Ho

2016-10-26

To evaluate patients with fever of unknown origin or those with suspected bacteremia, the precision of blood culture tests is critical. An inappropriate step in the test process or error in a parameter could lead to a false-positive result, which could then affect the direction of treatment in critical conditions. Mobile health apps can be used to resolve problems with blood culture tests, and such apps can hence ensure that point-of-care guidelines are followed and processes are monitored for blood culture tests. In this pilot project, we aimed to investigate the feasibility of using a mobile blood culture app to manage blood culture test quality. We implemented the app at a university hospital in South Korea to assess the potential for its utilization in a clinical environment by reviewing the usage data among a small group of users and by assessing their feedback and the data related to blood culture sampling. We used an iOS-based blood culture app that uses an embedded camera to scan the patient identification and sample number bar codes. A total of 4 medical interns working at 2 medical intensive care units (MICUs) participated in this project, which spanned 3 weeks. App usage and blood culture sampling parameters (including sampler, sampling site, sampling time, and sample volume) were analyzed. The compliance of sampling parameter entry was also measured. In addition, the participants' opinions regarding patient safety, timeliness, efficiency, and usability were recorded. In total, 356/644 (55.3%) of all blood culture samples obtained at the MICUs were examined using the app, including 254/356 (71.3%) with blood collection volumes of 5-7 mL and 256/356 (71.9%) with blood collection from the peripheral veins. The sampling volume differed among the participants. Sampling parameters were completely entered in 354/356 cases (99.4%). All the participants agreed that the app ensured good patient safety, disagreed on its timeliness, and did not believe that it was

The preanalytical optimization of blood cultures: a review and the clinical importance of benchmarking in 5 Belgian hospitals.

PubMed

Willems, Elise; Smismans, Annick; Cartuyvels, Reinoud; Coppens, Guy; Van Vaerenbergh, Kristien; Van den Abeele, Anne-Marie; Frans, Johan

2012-05-01

Bloodstream infections remain a major challenge in medicine. Optimal detection of pathogens is only possible if the quality of preanalytical factors is thoroughly controlled. Since the laboratory is responsible for this preanalytical phase, the quality control of critical factors should be integrated in its quality control program. The numerous recommendations regarding blood culture collection contain controversies. Only an unambiguous guideline permits standardization and interlaboratory quality control. We present an evidence-based concise guideline of critical preanalytical determinants for blood culture collection and summarize key performance indicators with their concomitant target values. In an attempt to benchmark, we compared the true-positive rate, contamination rate, and collected blood volume of blood culture bottles in 5 Belgian hospital laboratories. The true-positive blood culture rate fell within previously defined acceptation criteria by Baron et al. (2005) in all 5 hospitals, whereas the contamination rate exceeded the target value in 4 locations. Most unexpected, in each of the 5 laboratories, more than one third of the blood culture bottles were incorrectly filled, irrespective of the manufacturer of the blood culture vials. As a consequence of this shortcoming, one manufacturer recently developed an automatic blood volume monitoring system. In conclusion, clear recommendations for standardized blood culture collection combined with quality control of critical factors of the preanalytical phase are essential for diagnostic blood culture improvement. Copyright © 2012 Elsevier Inc. All rights reserved.

Decreased mortality associated with prompt Gram staining of blood cultures.

PubMed

Barenfanger, Joan; Graham, Donald R; Kolluri, Lavanya; Sangwan, Gaurav; Lawhorn, Jerry; Drake, Cheryl A; Verhulst, Steven J; Peterson, Ryan; Moja, Lauren B; Ertmoed, Matthew M; Moja, Ashley B; Shevlin, Douglas W; Vautrain, Robert; Callahan, Charles D

2008-12-01

Gram stains of positive blood cultures are the most important factor influencing appropriate therapy. The sooner appropriate therapy is initiated, the better. Therefore, it is reasonable to expect that the sooner Gram stains are performed, the better. To determine the value of timely Gram stains and whether improvement in Gram stain turnaround time (TAT) is feasible, we compared data for matched pairs of patients with cultures processed promptly (<1 hour TAT) with data for patients with cultures not processed promptly (> or =1 hour TAT) and then monitored TAT by control charting.In 99 matched pairs, average difference in time to detection of positive blood cultures within a pair of patients was less than 0.1 hour. For the less than 1 hour TAT group, the average TAT and crude mortality were 0.1 hour and 10.1%, respectively; for the 1 hour or longer TAT group, they were 3.3 hours and 19.2%, respectively (P < .0001 and P = .0389, respectively). After multifaceted efforts, we achieved significant improvement in the TAT for Gram stains.

Comparison of methods for identifying causative bacterial microorganisms in presumed acute endophthalmitis: conventional culture, blood culture, and PCR.

PubMed

Pongsachareonnont, Pear; Honglertnapakul, Worawalun; Chatsuwan, Tanittha

2017-02-21

Identification of bacterial pathogens in endophthalmitis is important to inform antibiotic selection and treatment decisions. Hemoculture bottles and polymerase chain reaction (PCR) analysis have been proposed to offer good detection sensitivity. This study compared the sensitivity and accuracy of a blood culture system, a PCR approach, and conventional culture methods for identification of causative bacteria in cases of acute endophthalmitis. Twenty-nine patients with a diagnosis of presumed acute bacterial endophthalmitis who underwent vitreous specimen collection at King Chulalongkorn Memorial Hospital were enrolled in this study. Forty-one specimens were collected. Each specimen was divided into three parts, and each part was analyzed using one of three microbial identification techniques: conventional plate culture, blood culture, and polymerase chain reaction and sequencing. The results of the three methods were then compared. Bacteria were identified in 15 of the 41 specimens (36.5%). Five (12.2%) specimens were positive by conventional culture methods, 11 (26.8%) were positive by hemoculture, and 11 (26.8%) were positive by PCR. Cohen's kappa analysis revealed p-values for conventional methods vs. hemoculture, conventional methods vs. PCR, and hemoculture vs. PCR of 0.057, 0.33, and 0.009, respectively. Higher detection rates of Enterococcus faecalis were observed for hemoculture and PCR than for conventional methods. Blood culture bottles and PCR detection may facilitate bacterial identification in cases of presumed acute endophthalmitis. These techniques should be used in addition to conventional plate culture methods because they provide a greater degree of sensitivity than conventional plate culture alone for the detection of specific microorganisms such as E. faecalis. Thai Clinical Trial Register No. TCTR20110000024 .

[Problems found in the isolation of mycelial fungi (Fusarium solani) from blood culture (Bacter NR-860)].

PubMed

Sanz, F; Soledad Cuétara, M; del Palacio, A; Campbell, C K; Johnson, L; Urruzunu, P; Pía Roiz, M

1993-11-01

F. solani fungemia is unusual. Patients at risk are immunosuppressed, have underlying malignancy or severe debilitating diseases. We report two cases of F. solani fungemia in two non neutropenic patients who had been treated with wide-spectrum antibiotics an/or systemic corticosteroids, parenteral nutrition and intravenous lines. Bactec NR-860 (Becton-Dickinson) system was used, and growth was detected in aerobic conditions (between 3-7 days of incubation). Removal of the catheters with or without i.v. amphotericin B were used successfully. The spectrum of Fusarium sp. fungemia is discussed. Current available antifungal therapy is also reviewed.

Rapid culture-based detection of living mycobacteria using microchannel electrical impedance spectroscopy (m-EIS).

PubMed

Kargupta, Roli; Puttaswamy, Sachidevi; Lee, Aiden J; Butler, Timothy E; Li, Zhongyu; Chakraborty, Sounak; Sengupta, Shramik

2017-06-10

Multiple techniques exist for detecting Mycobacteria, each having its own advantages and drawbacks. Among them, automated culture-based systems like the BACTEC-MGIT™ are popular because they are inexpensive, reliable and highly accurate. However, they have a relatively long "time-to-detection" (TTD). Hence, a method that retains the reliability and low-cost of the MGIT system, while reducing TTD would be highly desirable. Living bacterial cells possess a membrane potential, on account of which they store charge when subjected to an AC-field. This charge storage (bulk capacitance) can be estimated using impedance measurements at multiple frequencies. An increase in the number of living cells during culture is reflected in an increase in bulk capacitance, and this forms the basis of our detection. M. bovis BCG and M. smegmatis suspensions with differing initial loads are cultured in MGIT media supplemented with OADC and Middlebrook 7H9 media respectively, electrical "scans" taken at regular intervals and the bulk capacitance estimated from the scans. Bulk capacitance estimates at later time-points are statistically compared to the suspension's baseline value. A statistically significant increase is assumed to indicate the presence of proliferating mycobacteria. Our TTDs were 60 and 36 h for M. bovis BCG and 20 and 9 h for M. smegmatis with initial loads of 1000 CFU/ml and 100,000 CFU/ml respectively. The corresponding TTDs for the commercial BACTEC MGIT 960 system were 131 and 84.6 h for M. bovis BCG and 41.7 and 12 h for M smegmatis, respectively. Our culture-based detection method using multi-frequency impedance measurements is capable of detecting mycobacteria faster than current commercial systems.

Clinical significance of repeat blood cultures during febrile neutropenia in adult acute myeloid leukaemia patients undergoing intensive chemotherapy.

PubMed

Kimura, Shun-Ichi; Gomyo, Ayumi; Hayakawa, Jin; Tamaki, Masaharu; Akahoshi, Yu; Harada, Naonori; Ugai, Tomotaka; Kusuda, Machiko; Kameda, Kazuaki; Wada, Hidenori; Ishihara, Yuko; Kawamura, Koji; Sakamoto, Kana; Sato, Miki; Terasako-Saito, Kiriko; Kikuchi, Misato; Nakasone, Hideki; Kako, Shinichi; Tanihara, Aki; Kanda, Yoshinobu

2017-10-01

We evaluated the clinical significance of repeat blood cultures in persistent and recurrent fever during neutropenia in adult acute myeloid leukaemia (AML) and myelodysplastic syndrome (MDS) patients undergoing intensive chemotherapy. We retrospectively reviewed the chemotherapy cycles at our centre between January 2007 and December 2015. Blood cultures obtained within three days after initial febrile neutropenia (FN) were defined as initial blood cultures and those obtained on or after day 4 were defined as repeat blood cultures. Overall, 321 chemotherapy cycles in 89 patients were subjected to review. FN was identified in 276 (86.0%) chemotherapy cycles. In persistent FN (134 episodes), the causative pathogens were detected by repeat blood cultures in seven episodes (5.2%), including only three episodes (2.2%) of new infection. Shaking chills and high body temperature were identified as significant predictors for bloodstream infection (BSI). In recurrent FN (85 episodes), the causative pathogens were detected in seven episodes (8.2%), and all of these were new organisms. The frequency of detecting new pathogens by repeat blood cultures in recurrent FN (7/85) was higher than that in persistent FN (3/134) (p = .0491). A history of recent BSI was identified as a significant predictor for BSI in recurrent FN. The diagnostic yield of repeat blood cultures for persistent FN was low in intensive chemotherapy for AML and MDS. The frequency of repeat blood cultures for persistent FN could be reduced based on predictors. On the other hand, blood cultures were considered to be essential in cases with recurrent FN.

Use of amplified Mycobacterium tuberculosis direct test in respiratory samples from HIV-infected patients in Brazil.

PubMed

Barreto, Leonardo Bruno Paz Ferreira; Lourenço, Maria Cristina da Silva; Rolla, Valéria Cavalcanti; Veloso, Valdiléia Gonçalves; Huf, Gisele

2014-01-01

To compare the accuracy of the amplified Mycobacterium tuberculosis direct (AMTD) test with reference methods for the laboratory diagnosis of tuberculosis in HIV-infected patients. This was a study of diagnostic accuracy comparing AMTD test results with those obtained by culture on Löwenstein-Jensen (LJ) medium and by the BACTEC Mycobacteria Growth Indicator Tube 960 (BACTEC MGIT 960) system in respiratory samples analyzed at the Bioassay and Bacteriology Laboratory of the Oswaldo Cruz Foundation Evandro Chagas Clinical Research Institute in the city of Rio de Janeiro, Brazil. We analyzed respiratory samples collected from 118 patients, of whom 88 (74.4%) were male. The mean age was 36.6 ± 10.6 years. Using the AMTD test, the BACTEC MGIT 960 system, and LJ culture, we identified M. tuberculosis complex in 31.0%, 29.7%, and 27.1% of the samples, respectively. In comparison with LJ culture, the AMTD test had a sensitivity, specificity, positive predictive value, and negative predictive value of 87.5%, 89.4%, 75.7%, and 95.0%, respectively, for LJ culture, whereas, in comparison with the BACTEC MGIT 960 system, it showed values of 88.6%, 92.4%, 83.8%, and 94.8%, respectively. The AMTD test showed good sensitivity and specificity in the population studied, enabling the laboratory detection of M. tuberculosis complex in paucibacillary respiratory specimens.

Blood Cultures Drawn From Arterial Catheters Are Reliable for the Detection of Bloodstream Infection in Critically Ill Children.

PubMed

Berger, Itay; Gil Margolis, Merav; Nahum, Elhanan; Dagan, Ovdi; Levy, Itzhak; Kaplan, Eytan; Shostak, Eran; Shmuelov, Esther; Schiller, Ofer; Kadmon, Gili

2018-05-01

Arterial catheters may serve as an additional source for blood cultures in children when peripheral venipuncture is challenging. The aim of the study was to evaluate the accuracy of cultures obtained through indwelling arterial catheters for the diagnosis of bloodstream infections in critically ill pediatric patients. Observational and comparative. General and cardiac ICUs of a tertiary, university-affiliated pediatric medical center. The study group consisted of 138 patients admitted to the general or cardiac PICU in 2014-2015 who met the following criteria: presence of an indwelling arterial catheter and indication for blood culture. Blood was drawn by peripheral venipuncture and through the arterial catheter for each patient and sent for culture (total 276 culture pairs). Two specialists blinded to the blood source evaluated each positive culture to determine if the result represented true bloodstream infection or contamination. The sensitivity, specificity, and positive and negative predictive values of the arterial catheter and peripheral cultures for the diagnosis of bloodstream infection were calculated. Of the 56 positive cultures, 41 (15% of total samples) were considered diagnostic of true bloodstream infection. In the other 15 (5%), the results were attributed to contamination. The rate of false-positive results was higher for arterial catheter than for peripheral venipuncture cultures (4% vs 1.5%) but did not lead to prolonged unnecessary antibiotic treatment. On statistical analysis, arterial catheter blood cultures had high sensitivity (85%) and specificity (95%) for the diagnosis of true bloodstream infection, with comparable performance to peripheral blood cultures. Cultures of arterial catheter-drawn blood are reliable for the detection of bloodstream infection in PICUs.

Evaluation of negative results of BacT/Alert 3D automated blood culture system.

PubMed

Kocoglu, M Esra; Bayram, Aysen; Balci, Iclal

2005-06-01

Although automated continuous-monitoring blood culture systems are both rapid and sensitive, false-positive and false-negative results still occur. The objective of this study, then, was to evaluate negative results occurring with BacT/Alert 3D blood culture systems. A total of 1032 samples were cultured with the BacT/Alert 3D automated blood culture system, using both aerobic (FA) and anaerobic (FN) [corrected] media, and 128 of these samples yielded positive results. A total of 904 negative blood samples were then subcultured in 5% sheep blood agar, eosin methylene blue, chocolate agar, and sabouraud-dextrose agar. Organisms growing on these subcultures were subsequently identified using both Vitek32 (bioMerieux, Durham, NC) and conventional methods. Twenty four (2.6%) of the 904 subcultures grew on the subculture media. The majority (83.3%) of these were determined to be gram-positive microorganisms. Fourteen (58.3%) were coagulase-negative staphylococci, two (8.3%) were Bacillus spp., one (4.2%) was Staphylococcus aureus, and one (4.2%) was identified as Enterococcus faecium. Streptococcus pneumoniae and Neisseria spp. were isolated together in two (8.3%) vials. Gram-negative microorganisms comprised 12.5% of the subcultures, of which two (8.3%) were found to be Pseudomonas aeruginosa, and one (4.2%) was Pseudomonas fluorescens. The other isolate (4.2%) was identified as Candida albicans. We conclude that the subculture of negative results is valuable in the BacT/Alert 3D system, especially in situations in which only one set of blood cultures is taken.

Performance Evaluation of the Verigene Gram-Positive and Gram-Negative Blood Culture Test for Direct Identification of Bacteria and Their Resistance Determinants from Positive Blood Cultures in Hong Kong

PubMed Central

Siu, Gilman K. H.; Chen, Jonathan H. K.; Ng, T. K.; Lee, Rodney A.; Fung, Kitty S. C.; To, Sabrina W. C.; Wong, Barry K. C.; Cheung, Sherman; Wong, Ivan W. F.; Tam, Marble M. P.; Lee, Swing S. W.; Yam, W. C.

2015-01-01

Background A multicenter study was conducted to evaluate the diagnostic performance and the time to identifcation of the Verigene Blood Culture Test, the BC-GP and BC-GN assays, to identify both Gram-positive and Gram-negative bacteria and their drug resistance determinants directly from positive blood cultures collected in Hong Kong. Methods and Results A total of 364 blood cultures were prospectively collected from four public hospitals, in which 114 and 250 cultures yielded Gram-positive and Gram-negative bacteria, and were tested with the BC-GP and BC-GN assay respectively. The overall identification agreement for Gram-positive and Gram-negative bacteria were 89.6% and 90.5% in monomicrobial cultures and 62.5% and 53.6% in polymicrobial cultures, respectively. The sensitivities for most genus/species achieved at least 80% except Enterococcus spp. (60%), K.oxytoca (0%), K.pneumoniae (69.2%), whereas the specificities for all targets ranged from 98.9% to 100%. Of note, 50% (7/14) cultures containing K.pneumoniae that were missed by the BC-GN assay were subsequently identified as K.variicola. Approximately 5.5% (20/364) cultures contained non-target organisms, of which Aeromonas spp. accounted for 25% and are of particular concern. For drug resistance determination, the Verigene test showed 100% sensitivity for identification of MRSA, VRE and carbapenem resistant Acinetobacter, and 84.4% for ESBL-producing Enterobacteriaceae based on the positive detection of mecA, vanA, bla OXA and bla CTXM respectively. Conclusion Overall, the Verigene test provided acceptable accuracy for identification of bacteria and resistance markers with a range of turnaround time 40.5 to 99.2 h faster than conventional methods in our region. PMID:26431434

A Retrospective Evaluation of Critical Care Blood Culture Yield - Do Support Services Contribute to the "Weekend Effect"?

PubMed

Morton, Ben; Nagaraja, Shankara; Collins, Andrea; Pennington, Shaun H; Blakey, John D

2015-01-01

The "weekend effect" describes an increase in adverse outcomes for patients admitted at the weekend. Critical care units have moved to higher intensity working patterns to address this with some improved outcomes. However, support services have persisted with traditional working patterns. Blood cultures are an essential diagnostic tool for patients with sepsis but yield is dependent on sampling technique and processing. We therefore used blood culture yield as a surrogate for the quality of support service provision. We hypothesized that blood culture yields would be lower over the weekend as a consequence of reduced support services. We performed a retrospective observational study examining 1575 blood culture samples in a university hospital critical care unit over a one-year period. Patients with positive cultures had, on average, higher APACHE II scores (p = 0.015), longer durations of stay (p = 0.03), required more renal replacement therapy (p<0.001) and had higher mortality (p = 0.024). Blood culture yield decreased with repeated sampling with an increased proportion of contaminants. Blood cultures were 26.7% less likely to be positive if taken at the weekend (p = 0.0402). This effect size is the equivalent to the impact of sampling before and after antibiotic administration. Our study demonstrates that blood culture yield is lower at the weekend. This is likely caused by delays or errors in incubation and processing, reflecting the reduced provision of support services at the weekend. Reorganization of services to address the "weekend effect" should acknowledge the interdependent nature of healthcare service delivery.

Diet and Blood Pressure Control in Chinese Canadians: Cultural Considerations.

PubMed

Zou, Ping

2017-04-01

Hypertension is highly prevalent in Chinese Canadians and diet has been identified as an important modifiable risk factor for hypertension. The current anti-hypertensive dietary recommendations in hypertension care guidelines lack examination of cultural factors, are not culturally sensitive to ethnic populations, and cannot be translated to Chinese Canadian populations without cultural considerations. Guided by Leininger's Sunrise Model of culture care theory, this paper investigates how cultural factors impact Chinese Canadians' dietary practice. It is proposed that English language proficiency, health literacy, traditional Chinese diet, migration and acculturation, and Traditional Chinese Medicine influence Chinese Canadians' dietary practices. A culturally congruent nursing intervention should be established and tailored according to related cultural factors to facilitate Chinese Canadians' blood pressure control. In addition, further study is needed to test the model adapted from Sunrise Model and understand its mechanism.

Impact of antibiotic administration on blood culture positivity at the beginning of sepsis: a prospective clinical cohort study.

PubMed

Scheer, Christian S; Fuchs, Christian; Gründling, Matthias; Vollmer, Marcus; Bast, Juliane; Bohnert, Jürgen A; Zimmermann, Kathrin; Hahnenkamp, Klaus; Rehberg, Sebastian; Kuhn, Sven-Olaf

2018-06-04

Sepsis guidelines recommend obtaining blood cultures before starting anti-infective therapy in patients with sepsis. However, little is known how antibiotic treatment prior to sampling affects bacterial growth. The aim of this study was to compare the results of blood cultures drawn prior to and under antibiotic therapy. Prospective clinical cohort study of septic patients. Adult ICU patients with 2 or 3 blood culture (BC) sets at the beginning of sepsis between 2010 and 2017 were included. Patients with blood culture samplings obtained prior to antibiotic therapy were compared to patients with samplings under antibiotic therapy. Blood culture positivity, defined as microbiological pathogen finding, was compared between the groups. Logistic regression was performed to adjust the impact of different factors with respect to blood culture positivity. In total, 559 patients with 1364 blood culture sets at the beginning of sepsis were analyzed. BC positivity was 50.6% (78/154) among septic patients who did not receive antibiotics and only 27.7% (112/405) in those who were already under antibiotics (P<0.001). Logistic regression revealed antibiotic therapy as an independent factor for less pathogen identification (Odds ratio 0.4; 95%CI 0.3-0.6). Gram-positive pathogens (28.3%(111/392) vs. 11.9%(116/972);P<0.001) and also gram-negative pathogens (16.3%(64/392) vs. 9.3%(90/972);P<0.001) were more frequent in BC sets drawn prior to antibiotic therapy compared to sets under antibiotics. Obtaining blood cultures under antibiotic therapy is associated with a significant loss of pathogen detection. This strongly emphasizes the current recommendation to obtain blood cultures prior to antibiotic administration in patients with sepsis. Copyright © 2018. Published by Elsevier Ltd.

Prospective identification of erythroid elements in cultured peripheral blood.

PubMed

Miller, J L; Njoroge, J M; Gubin, A N; Rodgers, G P

1999-04-01

We have developed a prospective approach to identify the generation of erythroid cells derived from cultured peripheral blood mononuclear cells (PBMC) by monitoring the expression of the cell surface protein CD48. Unpurified populations of PBMC obtained from the buffy coats of normal volunteers were grown in suspension culture in the absence or presence of erythropoietin. A profile of surface CD48 expression permitted a flow cytometric identification of erythropoietin responsive populations at various stages of their maturation. In the absence of erythropoietin (EPO) supplemented media, the CD48- cells represented <5% of the total population of PBMC remaining in culture. In cultures supplemented with 1 U/mL EPO, the mean percentage of CD48- cells increased to 34.7 + 14.9% (p < 0.01) after 14 days in culture. Coordinated CD34 and CD71 (transferrin receptor) expression, morphology, gamma-globin transcription, and colony formation in methylcellulose were observed during the 14-day culture period. Flow cytometric monitoring of bulk cultured PBMC provides a simple and reliable means for the prospective or real-time study of human erythropoiesis.

Microbiology of liver abscesses and the predictive value of abscess gram stain and associated blood cultures.

PubMed

Chemaly, Roy F; Hall, Gerri S; Keys, Thomas F; Procop, Gary W

2003-08-01

Although rare, pyogenic liver abscesses are potentially fatal. We evaluated the predictive value of Gram stain of liver abscess aspirates and temporally associated blood cultures. Gram stains detected bacteria in 79% of the liver abscesses tested. The sensitivity and specificity of Gram stain of the liver abscesses were 90% and 100% for Gram-positive cocci (GPC) and 52% and 94% for Gram-negative bacilli (GNB). The sensitivities of the blood cultures for any GPC and GNB present in the liver abscess were 30% and 39%, respectively. Although, Gram stains and blood cultures offer incomplete detection of the microbial contents of pyogenic liver abscesses, both tests should always accompany liver abscess cultures.

Educational intervention as an effective step for reducing blood culture contamination: a prospective cohort study.

PubMed

Park, W B; Myung, S J; Oh, M-d; Lee, J; Kim, N-J; Kim, E-C; Park, J S

2015-10-01

Contaminated blood cultures lead to diagnostic challenges and place a burden on healthcare services. To determine the impact of introducing a clinical skills test (CST) as part of the medical licensing examination and an institutional education programme on the contamination rates of blood cultures. A prospective cohort study was conducted from 2009 through 2013 in all wards of a tertiary-care teaching hospital. We evaluated the effects of the CST, which was added to the National Medical Licensing Examination in Korea (KMLE) in 2010 and our institutional education programme, which began in 2013. The medical interns in charge of collection of blood for culture were divided in three groups with presence or absence of CST and the institutional education programme. The primary outcome was the percentage of blood cultures contaminated in each group, which were compared using the Poisson regression model. Participants' self-rated scores for the blood draw procedure were also analysed. Although introduction of the CST in the KMLE failed to reduce blood culture contamination rate (1.36% vs 1.35%; P = 0.734), the institutional education programme significantly reduced the contamination rate (1.35% vs 1.00%; P < 0.0001). Most participants answered that they always followed each step correctly except for waiting the recommended contact time after applying the antiseptic. The educational intervention, not the introduction of CST in the KMLE, was effective in reducing overall contamination rates. Copyright © 2015 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

Towards cultural materialism in the medical humanities: the case of blood rejuvenation

PubMed Central

2018-01-01

This paper argues for an approach within the medical humanities that draws on the theoretical legacy of cultural materialism as a framework for reading cultural practices and their relationship to the social and economic order. It revisits the origins and development of cultural materialism in cultural studies and literary studies between the 1970s and 1990s and considers how, with adaptation, this methodology might facilitate ideological criticism focused on material formations of health, disease and the human body. I outline three key characteristics of a medicocultural materialist approach along these lines: (a) interdisciplinary work on a broad range of medical and cultural sources, including those drawn from ‘popular’ forms of culture; (b) the combination of historicist analysis with scrutiny of present-day contexts; (c) analyses that engage with political economy perspectives and/or the work of medical sociology in this area. The subsequent sections of the paper employ a medicocultural materialist approach to examine conjectural understandings of, and empirical investigations into, the capacity of transfused human blood to rejuvenate the ageing body. I trace textual faultlines that expose the structures of power which inform the movement of blood between bodies in ‘medical gothic’ fictions from the 19th-century fin de siècle, including Mary Elizabeth Braddon's ‘Good Lady Ducayne’ (1896) and Bram Stoker's Dracula (1897). I conclude with a critique of biomedical innovations in blood rejuvenation in the era of medical neoliberalism, before considering the potential applications of medicocultural materialism to other topics within the field of the medical humanities. PMID:28495908

A randomized trial of povidone-iodine compared with iodine tincture for venipuncture site disinfection: effects on rates of blood culture contamination.

PubMed

Little, J R; Murray, P R; Traynor, P S; Spitznagel, E

1999-08-01

Contamination of blood cultures creates problems in their interpretation and unneeded resource utilization. Because skin flora comprise the major group of contaminant species, more effective skin disinfection at the venipuncture site could reduce contamination. We performed a randomized trial in adult inpatients at a tertiary care teaching hospital. Antecubital venipuncture sites were randomly disinfected with povidone-iodine or iodine tincture, and blood cultures (two bottles, 10 mL of blood) were drawn by professional phlebotomists. Scoring of contaminant species was restricted to skin flora. Hospital resource utilization was compared among patients with contaminated blood cultures and those with sterile blood cultures. Of the 3,851 blood cultures collected during the study, 120 (3.1%) were contaminated with skin flora. The contamination rate for blood cultures collected after povidone-iodine was 3.8% (74 of 1,947), compared with a rate of 2.4% (46 of 1,904, P = 0.01) after iodine tincture. The difference in mean total hospital costs for patients with contaminated blood cultures and those with sterile blood cultures was $4,100 (95% confidence interval: $740 to $7,400, P = 0.02). Iodine tincture is superior to povidone-iodine for venipuncture site antisepsis before blood culture sampling. Because of the high costs associated with contaminated blood cultures, hospitals should consider switching from povidone-iodine to iodine tincture. Reduction of the contamination rate may improve the quality of patient care and reduce hospital costs.

The Clinical Impact of Rapid, Direct MALDI-ToF Identification of Bacteria from Positive Blood Cultures.

PubMed

French, Kathryn; Evans, Jason; Tanner, Hannah; Gossain, Savita; Hussain, Abid

2016-01-01

Faster identification of bacterial isolates from blood cultures can enable earlier clinical intervention for patients with sepsis. We evaluated the clinical impact of direct identification of micro-organisms from positive blood cultures using MALDI-ToF. Positive blood cultures with organisms seen on Gram stain were included over a four week period. For each patient case, comparison was made between the clinical advice given on day one with only a Gram stain result, and the follow up advice given on day two with the benefit of organism identification. Culture results were then compared with direct MALDI-ToF identification. For 73 of 115 cases (63.5%), direct organism identification was obtained by MALDI-ToF. Of those 73, 70 (95.5%) had a result concordant with that of the plate culture. In 28 of the 115 cases (24.3%) direct MALDI-ToF identification on day one would have had a clear clinical benefit. In 11 cases it would have helped to identify the potential source of bacteraemia. In 11 cases it would have indicated a different antibiotic regimen on day one, with five patients receiving appropriate antibiotics 24 hours earlier. For 14 cases the blood culture isolate could have been designated as unlikely to be clinically significant. We have demonstrated that organism identification on day one of blood culture positivity can have a direct clinical impact. Faster identification using MALDI-ToF assists the clinician in assessing the significance of a blood culture isolate on day one. It can allow earlier appropriate choice of antimicrobial agent, even in the absence of susceptibility testing, and help narrow down the potential source of infection providing a focus for further investigation in a more timely way than conventional techniques alone.

Time to Positivity and Detection of Growth in Anaerobic Blood Culture Vials Predict the Presence of Candida glabrata in Candidemia: a Two-Center European Cohort Study

PubMed Central

Kaasch, Achim J.; Soriano, Alex; Torres, Jorge-Luis; Vergara, Andrea; Morata, Laura; Zboromyrska, Yuliya; De La Calle, Cristina; Alejo, Izaskun; Hernández, Cristina; Cardozo, Celia; Marco, Franscesc; Del Río, Ana; Almela, Manel; Mensa, Josep; Martínez, José Antonio

2014-01-01

This study shows the accuracy of exclusive or earlier growth in anaerobic vials to predict Candida glabrata in a large series of candidemic patients from two European hospitals using the Bactec 9240 system. Alternatively, C. glabrata can be predicted by a time to positivity cutoff value, which should be determined for each setting. PMID:24899027

Use of BBL CHROMagar MRSA Medium for Identification of Methicillin-Resistant Staphylococcus aureus Directly from Blood Cultures

PubMed Central

Pape, John; Wadlin, Jill; Nachamkin, Irving

2006-01-01

We evaluated the ability of BBL CHROMagar MRSA medium (Becton Dickinson, Sparks, MD) to identify methicillin-resistant Staphylococcus aureus (MRSA) directly upon subculture from positive blood culture bottles. There were 124 MRSA isolates recovered from blood cultures in the study. BBL CHROMagar MRSA medium was highly sensitive (97.6% [121/124] at 18 to 24 h of incubation and 100% [124/124] at 48 h) and 99.9% specific for identifying MRSA from positive blood cultures. PMID:16825383

Detection of bacterial growth by gas absorption.

PubMed

Waters, J R

1992-05-01

When 24 different aerobic organisms were grown in a shaken culture, all were found to first absorb gas from the headspace. In a rudimentary medium, such as tryptic soy broth, 16 of the 24 organisms did not produce gas following the initial gas absorption. We have developed a simple, noninvasive method for detecting both gas absorption and production in multiple culture vials. The time to positivity was compared with that obtained by the BACTEC 460 blood culture system. For nearly all of these organisms, there was no difference. For some of those organisms that did not produce gas, e.g. Staphylococcus epidermidis, Moraxella osloensis, and Neisseria meningitidis, detection by gas absorption was a few hours faster. Gas absorption appears to be a promising technique for a new automated blood culture system because of its simplicity and because medium without special additives can be used to detect organisms that do not produce gas.

Reducing time to identification of positive blood cultures with MALDI-TOF MS analysis after a 5-h subculture.

PubMed

Verroken, A; Defourny, L; Lechgar, L; Magnette, A; Delmée, M; Glupczynski, Y

2015-02-01

Speeding up the turn-around time of positive blood culture identifications is essential in order to optimize the treatment of septic patients. Several sample preparation techniques have been developed allowing direct matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) identification of positive blood cultures. Yet, the hands-on time restrains their routine workflow. In this study, we evaluated an approach whereby MALDI-TOF MS identification without any additional steps was carried out on short subcultured colonies from positive blood bottles with the objective of allowing results reporting on the day of positivity detection. Over a 7-month period in 2012, positive blood cultures detected by 9 am with an automated system were inoculated onto a Columbia blood agar and processed after a 5-h incubation on a MALDI-TOF MicroFlex platform (Bruker Daltonik GmbH). Single-spotted colonies were covered with 1 μl formic acid and 1 μl matrix solution. The results were compared to the validated identification techniques. A total of 925 positive blood culture bottles (representing 470 bacteremic episodes) were included. Concordant identification was obtained in 727 (81.1 %) of the 896 monomicrobial blood cultures, with failure being mostly observed with anaerobes and yeasts. In 17 episodes of polymicrobic bacteremia, the identification of one of the two isolates was achieved in 24/29 (82.7 %) positive cultures. Routine implementation of MALDI-TOF MS identification on young positive blood subcultures provides correct results to the clinician in more than 80 % of the bacteremic episodes and allows access to identification results on the day of blood culture positivity detection, potentially accelerating the implementation of targeted clinical treatments.

Blood culture-PCR to optimise typhoid fever diagnosis after controlled human infection identifies frequent asymptomatic cases and evidence of primary bacteraemia.

PubMed

Darton, Thomas C; Zhou, Liqing; Blohmke, Christoph J; Jones, Claire; Waddington, Claire S; Baker, Stephen; Pollard, Andrew J

2017-04-01

Improved diagnostics for typhoid are needed; a typhoid controlled human infection model may accelerate their development and translation. Here, we evaluated a blood culture-PCR assay for detecting infection after controlled human infection with S. Typhi and compared test performance with optimally performed blood cultures. Culture-PCR amplification of blood samples was performed alongside daily blood culture in 41 participants undergoing typhoid challenge. Study endpoints for typhoid diagnosis (TD) were fever and/or bacteraemia. Overall, 24/41 (59%) participants reached TD, of whom 21/24 (86%) had ≥1 positive blood culture (53/674, 7.9% of all cultures) or 18/24 (75%) had ≥1 positive culture-PCR assay result (57/684, 8.3%). A further five non-bacteraemic participants produced culture-PCR amplicons indicating infection; overall sensitivity/specificity of the assay compared to the study endpoints were 70%/65%. We found no significant difference between blood culture and culture-PCR methods in ability to identify cases (12 mismatching pairs, p = 0.77, binomial test). Clinical and stool culture metadata demonstrated that additional culture-PCR amplification positive individuals likely represented true cases missed by blood culture, suggesting the overall attack rate may be 30/41 (73%) rather than 24/41 (59%). Several participants had positive culture-PCR results soon after ingesting challenge providing new evidence for occurrence of an early primary bacteraemia. Overall the culture-PCR assay performed well, identifying extra typhoid cases compared with routine blood culture alone. Despite limitations to widespread field-use, the benefits of increased diagnostic yield, reduced blood volume and faster turn-around-time, suggest that this assay could enhance laboratory typhoid diagnostics in research applications and high-incidence settings. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Identification of blood culture isolates directly from positive blood cultures by use of matrix-assisted laser desorption ionization-time of flight mass spectrometry and a commercial extraction system: analysis of performance, cost, and turnaround time.

PubMed

Lagacé-Wiens, Philippe R S; Adam, Heather J; Karlowsky, James A; Nichol, Kimberly A; Pang, Paulette F; Guenther, Jodi; Webb, Amanda A; Miller, Crystal; Alfa, Michelle J

2012-10-01

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry represents a revolution in the rapid identification of bacterial and fungal pathogens in the clinical microbiology laboratory. Recently, MALDI-TOF has been applied directly to positive blood culture bottles for the rapid identification of pathogens, leading to reductions in turnaround time and potentially beneficial patient impacts. The development of a commercially available extraction kit (Bruker Sepsityper) for use with the Bruker MALDI BioTyper has facilitated the processing required for identification of pathogens directly from positive from blood cultures. We report the results of an evaluation of the accuracy, cost, and turnaround time of this method for 61 positive monomicrobial and 2 polymicrobial cultures representing 26 species. The Bruker MALDI BioTyper with the Sepsityper gave a valid (score, >1.7) identification for 85.2% of positive blood cultures with no misidentifications. The mean reduction in turnaround time to identification was 34.3 h (P < 0.0001) in the ideal situation where MALDI-TOF was used for all blood cultures and 26.5 h in a more practical setting where conventional identification or identification from subcultures was required for isolates that could not be directly identified by MALDI-TOF. Implementation of a MALDI-TOF-based identification system for direct identification of pathogens from blood cultures is expected to be associated with a marginal increase in operating costs for most laboratories. However, the use of MALDI-TOF for direct identification is accurate and should result in reduced turnaround time to identification.

Comparative usefulness of inflammatory markers to indicate bacterial infection-analyzed according to blood culture results and related clinical factors.

PubMed

Nishikawa, Hirokazu; Shirano, Michinori; Kasamatsu, Yu; Morimura, Ayumi; Iida, Ko; Kishi, Tomomi; Goto, Tetsushi; Okamoto, Saki; Ehara, Eiji

2016-01-01

To assess relationships of inflammatory markers and 2 related clinical factors with blood culture results, we retrospectively investigated inpatients' blood culture and blood chemistry findings that were recorded from January to December 2014 using electronic medical records and analyzed the data of 852 subjects (426 culture-positive and 426 culture-negative). Results suggested that the risk of positive blood culture statistically increased as inflammatory marker levels and the number of related factors increased. Concerning the effectiveness of inflammatory markers, when the outcome definition was also changed for C-reactive protein (CRP), the odds ratio had a similar value, whereas when the outcome definition of blood culture positivity was used for procalcitonin (PCT), the greatest effectiveness of that was detected. Therefore, the current results suggest that PCT is more useful than CRP as an auxiliary indication of bacterial infection. Copyright © 2016 Elsevier Inc. All rights reserved.

Using electronic data to predict the probability of true bacteremia from positive blood cultures.

PubMed

Wang, S J; Kuperman, G J; Ohno-Machado, L; Onderdonk, A; Sandige, H; Bates, D W

2000-01-01

As part of a project to help physicians make more appropriate treatment decisions, we implemented a clinical prediction rule that computes the probability of true bacteremia for positive blood cultures and displays this information when culture results are viewed online. Prior to implementing the rule, we performed a revalidation study to verify the accuracy of the previously published logistic regression model. We randomly selected 114 cases of positive blood cultures from a recent one-year period and performed a paper chart review with the help of infectious disease experts to determine whether the cultures were true positives or contaminants. Based on the results of this revalidation study, we updated the probabilities reported by the model and made additional enhancements to improve the accuracy of the rule. Next, we implemented the rule into our hospital's laboratory computer system so that the probability information was displayed with all positive blood culture results. We displayed the prediction rule information on approximately half of the 2184 positive blood cultures at our hospital that were randomly selected during a 6-month period. During the study, we surveyed 54 housestaff to obtain their opinions about the usefulness of this intervention. Fifty percent (27/54) indicated that the information had influenced their belief of the probability of bacteremia in their patients, and in 28% (15/54) of cases it changed their treatment decision. Almost all (98% (53/54)) indicated that they wanted to continue receiving this information. We conclude that the probability information provided by this clinical prediction rule is considered useful to physicians when making treatment decisions.

Development of a rapid and simplified protocol for direct bacterial identification from positive blood cultures by using matrix assisted laser desorption ionization time-of- flight mass spectrometry.

PubMed

Jakovljev, Aleksandra; Bergh, Kåre

2015-11-06

Bloodstream infections represent serious conditions carrying a high mortality and morbidity rate. Rapid identification of microorganisms and prompt institution of adequate antimicrobial therapy is of utmost importance for a successful outcome. Aiming at the development of a rapid, simplified and efficient protocol, we developed and compared two in-house preparatory methods for the direct identification of bacteria from positive blood culture flasks (BD BACTEC FX system) by using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF MS). Both methods employed saponin and distilled water for erythrocyte lysis. In method A the cellular pellet was overlaid with formic acid on the MALDI TOF target plate for protein extraction, whereas in method B the pellet was exposed to formic acid followed by acetonitrile prior to placing on the target plate. Best results were obtained by method A. Direct identification was achieved for 81.9 % and 65.8 % (50.3 % and 26.2 % with scores >2.0) of organisms by method A and method B, respectively. Overall concordance with final identification was 100 % to genus and 97.9 % to species level. By applying a lower cut-off score value, the levels of identification obtained by method A and method B increased to 89.3 % and 77.8 % of organisms (81.9 % and 65.8 % identified with scores >1.7), respectively. Using the lowered score criteria, concordance with final results was obtained for 99.3 % of genus and 96.6 % of species identifications. The reliability of results, rapid performance (approximately 25 min) and applicability of in-house method A have contributed to implementation of this robust and cost-effective method in our laboratory.

Comparative evaluation of two rapid Salmonella-IgM tests and blood culture in the diagnosis of enteric fever.

PubMed

Prasad, K J; Oberoi, J K; Goel, N; Wattal, C

2015-01-01

Enteric fever is a major public health problem in developing countries like India. An early and accurate diagnosis is necessary for a prompt and effective treatment. We have evaluated the diagnostic accuracy of two Rapid Salmonella-IgM tests (Typhidot-IgM and Enteroscreen-IgM) as compared to blood culture in rapid and early diagnosis of enteric fever. A total of 2,699 patients' serum samples were tested by Rapid Salmonella-IgM tests and blood culture. Patients were divided into two groups. Test group - patients with enteric fever and blood culture positives for Salmonella Typhi; and three types of Controls, i.e. patients with non-enteric fever illnesses, normal healthy controls and patients positive for S. Paratyphi- A. In addition to this we have also evaluated the significance of positive Salmonella-IgM tests among blood culture-negative cases. The overall sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of the Typhidot-IgM test and Enteroscreen-IgM test considering blood culture as gold standard were 97.29% and 88.13%, 97.40% and 87.83%, 98.18% and 92.03%, 96.15% and 82.27%, respectively. Typhidot-IgM test was found to be significantly more sensitive and specific as compared to Enteroscreen-IgM. Among blood culture-negative patients, Rapid Salmonella-IgM tests detected 72.25% additional cases of enteric fever. Although the Rapid Salmonella-IgM tests are meant to diagnose S. Typhi only, but these tests detect S. Paratyphi- A also. Thirty-eight patients who were blood culture-positive for S. Paratyphi- A were also positive by Rapid Salmonella-IgM tests. Rapid Salmonella-IgM tests offer an advantage of increased sensitivity, rapidity, early diagnosis and simplicity over blood culture.

Measurement of Blood Coagulation Factor Synthesis in Cultures of Human Hepatocytes.

PubMed

Heinz, Stefan; Braspenning, Joris

2015-01-01

An important function of the liver is the synthesis and secretion of blood coagulation factors. Within the liver, hepatocytes are involved in the synthesis of most blood coagulation factors, such as fibrinogen, prothrombin, factor V, VII, IX, X, XI, XII, as well as protein C and S, and antithrombin, whereas liver sinusoidal endothelial cells produce factor VIII and von Willebrand factor. Here, we describe methods for the detection and quantification of most blood coagulation factors in hepatocytes in vitro. Hepatocyte cultures indeed provide a valuable tool to study blood coagulation factors. In addition, the generation and expansion of hepatocytes or hepatocyte-like cells may be used in future for cell-based therapies of liver diseases, including blood coagulation factor deficiencies.

Predicting bacteremia based on nurse-assessed food consumption at the time of blood culture.

PubMed

Komatsu, Takayuki; Onda, Toshihito; Murayama, Go; Yamanouchi, Masashi; Inukai, Minori; Sakai, Ai; Kikuta, Masumi; Branch, Joel; Aoki, Makoto; Tierney, Lawrence M; Inoue, Kenji

2012-01-01

Bacteremia and its complications are important causes of morbidity and mortality in hospitalized patients. However, the yield of blood cultures is relatively low, with many false-positive results from bacterial contamination. We investigated the relationship between patient food consumption and the presence of bacteremia. This was an observational analysis of a cohort of 1179 patients who underwent blood culture analysis between January 2005 and December 2009. Patients with anorexia-inducing conditions, such as gastrointestinal illness and malignant disease treated with chemotherapy, were excluded. Food consumption was rated by nurses as the percentage of food consumed during the meal preceding the blood culture. Groupings were as follows: low consumption (<50%), moderate (>50% to <80%), and high (>80%). Low consumption was observed in 39.8% of patients, moderate in 17.8%, and high in 41.6%. The average body temperature was 38.1 ± 1.1°C. Bacteremia was present in 18.5%, 3.9%, and 1.4% of patients in the low, moderate, and high food consumption groups, respectively. The negative predictive value was 98.3%, suggesting that bacteremia is very unlikely in the setting of good food intake. Bacteremia is an unlikely occurrence in hospitalized patients who maintain adequate food consumption at the time of blood culture. Copyright © 2012 Society of Hospital Medicine.

PCR identification of bacteria in blood culture does not fit the daily workflow of a routine microbiology laboratory.

PubMed

Karumaa, Santra; Kärpänoja, Pauliina; Sarkkinen, Hannu

2012-03-01

We have evaluated the GenoType blood culture assay (Hain Lifescience, Nehren, Germany) for the identification of bacteria in 233 positive blood cultures and assessed its suitability in the workflow of a routine microbiology laboratory. In 68/233 (29.2%) samples, the culture result could not be confirmed by the GenoType assay due to a lack of primers in the test, multiple organisms in the sample, or inconsistency with respect to the identification by culture. Although the GenoType blood culture assay gives satisfactory results for bacteria for which primers are available, there are difficulties in applying the test in the routine microbiology laboratory.

Clinical and Laboratory Potential Predictors of Blood Culture Positivity in Under Five Children with Clinically Severe Pneumonia - Khartoum -Sudan.

PubMed

Salih, Karimeldin Mohamed Ali; El-Samani, El-Fatih; Bilal, Jalal Ali; Eldouch, Widad; Ibrahim, Salah Ahmed

2015-08-01

Blood culture is necessary for appropriate management of clinically severe pneumonia in children under five years of age. However, in limited resource countries it might be unduly costly and waste of valuable time because of the high negative culture rate. This study aims to identify clinical and laboratory parameters that potentially predict a positive blood culture in cases of severe pneumonia. A hospital based study, enrolled 189 cases satisfying the WHO definition of severe pneumonia. Age, gender, clinical history, physical examination, temperature, complete blood count, C-reactive protein, blood culture and Chest X Ray for all the patients were recorded. Forty one patients had positive blood culture giving a prevalence of 21.7%. All variables were used in a dichotomous manner. White Blood Count (WBC) more than 20 000, very high C-reactive protein (C-RP ≥8mg/L) and Temperature more than 40(o)C, had a positive predictive value of 46.1%, 44.3% and 40.0% respectively for a positive culture as well as a Negative Predictive Value of 91.1%, 91.6% and 91.7% respectively. The WBC more than 20 000 and temperature above 40(o)C had a significant association with a positive blood culture. Their adjusted Odds Ratios were 3.9 (95% CI: 1.4-10.90) and 3.1 (95% CI: 1.2-8.4) respectively. This was not the case for C-RP (Odds Ratio=2.2, 95% CI: 0.7-2.2) or positive Chest X Ray (Odds Ratio=1.5, 95% CI: 0.6-3.6). Temperature of more than 40(o)C, Very high C-RP and WBC of more than 20 000 are good indicators of a potential positive blood culture. It is therefore recommended that further research be undertaken to refine these predictors as screening tools before resorting to blood culture. It is also recommended that antibiotic treatment may be initiated on the basis of the high temperature and WBC, while waiting for the culture results.

Identification of bacteria in blood culture broths using matrix-assisted laser desorption-ionization Sepsityper™ and time of flight mass spectrometry.

PubMed

Kok, Jen; Thomas, Lee C; Olma, Thomas; Chen, Sharon C A; Iredell, Jonathan R

2011-01-01

Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) is a novel method for the direct identification of bacteria from blood culture broths. We evaluate for the first time, the performance of the MALDI Sepsityper™ Kit and MS for the identification of bacteria compared to standard phenotypic methods using the manufacturer's specified bacterial identification criteria (spectral scores ≥1.700-1.999 and ≥2.000 indicated identification to genus and species level, respectively). Five hundred and seven positive blood culture broths were prospectively examined, of which 379 (74.8%; 358 monomicrobial, 21 polymicrobial) were identified by MALDI-TOF MS; 195 (100%) and 132 (67.7%) of 195 gram-positive; and 163 (100%) and 149 (91.4%) of 163 gram-negative organisms from monomicrobial blood cultures were correctly identified to genus and species level, respectively. Spectral scores <1.700 (no identification) were obtained in 128/507 (25.2%) positive blood culture broths, including 31.6% and 32.3% of gram-positive and polymicrobial blood cultures, respectively. Significantly more gram-negative organisms were identified compared to gram-positive organisms at species level (p<0.0001). Five blood cultures were misidentified, but at species level only; including four monomicrobial blood cultures with Streptococcus oralis/mitis that were misidentified as Streptococcus pneumoniae. Positive predictive values for the direct identification of both gram-positive and gram-negative bacteria from monomicrobial blood culture broths to genus level were 100%. A diagnostic algorithm for positive blood culture broths that incorporates gram staining and MALDI-TOF MS should identify the majority of pathogens, particularly to genus level.

Time to Detection with BacT/Alert FA Plus Compared to BacT/Alert FA Blood Culture Media.

PubMed

Nutman, A; Fisher Even-Tsur, S; Shapiro, G; Braun, T; Schwartz, D; Carmeli, Y

2016-09-01

Rapid identification of the causative pathogen in patients with bacteremia allows adjustment of antibiotic therapy and improves patient outcomes. We compared in vitro and real-life time to detection (TTD) of two blood culture media, BacT/Alert FA (FA) and BacT/Alert FA Plus (FA Plus), for the nine most common species of bacterial pathogens recovered from blood samples. Experimental data from simulated cultures was compared with microbiology records of TTD for both culture media with growth of the species of interest in clinical blood cultures. In the experimental conditions, median TTD was 3.8 hours (23.9 %) shorter using FA Plus media. The magnitude of reduction differed between species. Similarly, in real life data, FA Plus had shorter TTD than FA media; however, the difference between culture media was smaller, and median TTD was only 1 hour (8.5 %) less. We found shorter TTD with BacT/Alert FA Plus culture media, both experimentally and in real-life conditions and unrelated to antibiotic neutralization, highlighting the importance of appropriate blood culture media selection.

Comparison of the lysis centrifugation method with the conventional blood culture method in cases of sepsis in a tertiary care hospital.

PubMed

Parikh, Harshal R; De, Anuradha S; Baveja, Sujata M

2012-07-01

Physicians and microbiologists have long recognized that the presence of living microorganisms in the blood of a patient carries with it considerable morbidity and mortality. Hence, blood cultures have become critically important and frequently performed test in clinical microbiology laboratories for diagnosis of sepsis. To compare the conventional blood culture method with the lysis centrifugation method in cases of sepsis. Two hundred nonduplicate blood cultures from cases of sepsis were analyzed using two blood culture methods concurrently for recovery of bacteria from patients diagnosed clinically with sepsis - the conventional blood culture method using trypticase soy broth and the lysis centrifugation method using saponin by centrifuging at 3000 g for 30 minutes. Overall bacteria recovered from 200 blood cultures were 17.5%. The conventional blood culture method had a higher yield of organisms, especially Gram positive cocci. The lysis centrifugation method was comparable with the former method with respect to Gram negative bacilli. The sensitivity of lysis centrifugation method in comparison to conventional blood culture method was 49.75% in this study, specificity was 98.21% and diagnostic accuracy was 89.5%. In almost every instance, the time required for detection of the growth was earlier by lysis centrifugation method, which was statistically significant. Contamination by lysis centrifugation was minimal, while that by conventional method was high. Time to growth by the lysis centrifugation method was highly significant (P value 0.000) as compared to time to growth by the conventional blood culture method. For the diagnosis of sepsis, combination of the lysis centrifugation method and the conventional blood culture method with trypticase soy broth or biphasic media is advocable, in order to achieve faster recovery and a better yield of microorganisms.

Towards cultural materialism in the medical humanities: the case of blood rejuvenation.

PubMed

Oakley, Catherine

2018-03-01

This paper argues for an approach within the medical humanities that draws on the theoretical legacy of cultural materialism as a framework for reading cultural practices and their relationship to the social and economic order. It revisits the origins and development of cultural materialism in cultural studies and literary studies between the 1970s and 1990s and considers how, with adaptation, this methodology might facilitate ideological criticism focused on material formations of health, disease and the human body. I outline three key characteristics of a medicocultural materialist approach along these lines: (a) interdisciplinary work on a broad range of medical and cultural sources, including those drawn from 'popular' forms of culture; (b) the combination of historicist analysis with scrutiny of present-day contexts; (c) analyses that engage with political economy perspectives and/or the work of medical sociology in this area. The subsequent sections of the paper employ a medicocultural materialist approach to examine conjectural understandings of, and empirical investigations into, the capacity of transfused human blood to rejuvenate the ageing body. I trace textual faultlines that expose the structures of power which inform the movement of blood between bodies in 'medical gothic' fictions from the 19th-century fin de siècle, including Mary Elizabeth Braddon's 'Good Lady Ducayne' (1896) and Bram Stoker's Dracula (1897). I conclude with a critique of biomedical innovations in blood rejuvenation in the era of medical neoliberalism, before considering the potential applications of medicocultural materialism to other topics within the field of the medical humanities. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

Time to positivity and detection of growth in anaerobic blood culture vials predict the presence of Candida glabrata in candidemia: a two-center European cohort study.

PubMed

Cobos-Trigueros, Nazaret; Kaasch, Achim J; Soriano, Alex; Torres, Jorge-Luis; Vergara, Andrea; Morata, Laura; Zboromyrska, Yuliya; De La Calle, Cristina; Alejo, Izaskun; Hernández, Cristina; Cardozo, Celia; Marco, Franscesc; Del Río, Ana; Almela, Manel; Mensa, Josep; Martínez, José Antonio

2014-08-01

This study shows the accuracy of exclusive or earlier growth in anaerobic vials to predict Candida glabrata in a large series of candidemic patients from two European hospitals using the Bactec 9240 system. Alternatively, C. glabrata can be predicted by a time to positivity cutoff value, which should be determined for each setting. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Polymerase chain reaction detection of Bartonella spp. in dogs from Spain with blood culture-negative infectious endocarditis.

PubMed

Roura, X; Santamarina, G; Tabar, M-D; Francino, O; Altet, L

2018-05-25

The presence of Bartonella spp. was detected by polymerase chain reaction (PCR) in dogs from Spain with blood culture-negative endocarditis. The aim of this study is to add information about canine infectious endocarditis in Europe. Thirty dogs with naturally occurring blood culture-negative endocarditis were examined from 2010 to 2017 at three veterinary referral hospitals, located in northwest, northeast, and southeast of Spain. It is a retrospective study. Medical records were reviewed to extract relevant data. Frozen or paraffin-embedded cardiac valve tissue and/or ethylenediamine tetraacetic acid blood samples were evaluated by PCR for the presence of Bartonella DNA. Positive results were sequenced to confirm the species. Polymerase chain reaction was positive for eight out of 30 dogs included (26.6%). Bartonella rochalimae, Bartonella vinsonii subsp. berkhoffii, and Bartonella koehlerae were detected in valve tissue or blood. Bartonella could be an important cause of blood culture-negative infectious endocarditis in dogs from Spain. The outcome for those dogs affected with Bartonella spp. was grave. Prompt empirical treatment with amoxicillin-clavulanate plus fluoroquinolones could be of value in cases of blood culture-negative endocarditis. Copyright © 2018 Elsevier B.V. All rights reserved.

Influence of postmortem time on the outcome of blood cultures among cadaveric tissue donors.

PubMed

Saegeman, V; Verhaegen, J; Lismont, D; Verduyckt, B; De Rijdt, T; Ectors, N

2009-02-01

Tissue banks provide tissues of human cadaver donors for transplantation. The maximal time limit for tissue retrieval has been set at 24 h postmortem. This study aimed at evaluating the evidence for this limit from a microbiological point of view. The delay of growth in postmortem blood cultures, the identification of the species isolated and clinical/environmental factors were investigated among 100 potential tissue donors. No significant difference was found in the rate of donors with grown blood cultures within (25/65=38%) compared with after (24/65=37%) 24 h of death. Coagulase-negative staphylococci and gastro-intestinal microorganisms were isolated within and after 24 h of death. Two factors--antimicrobial therapy and "delay before body cooling"--were significantly inversely related with donors' blood culture results. From a microbiological point of view, there is no evidence for avoiding tissue retrieval among donors after 24 h of death.

Studies that report unexpected positive blood cultures for Lyme borrelia - are they valid?

PubMed

Wormser, Gary P; Shapiro, Eugene D; Strle, Franc

2017-11-01

Positive blood cultures for Lyme borrelia have been well documented in untreated patients with early Lyme disease. In this report we review the validity of three studies that reported the recovery of Borrelia burgdorferi sensu lato from the blood of a high proportion of patients for whom no evidence was presented, and no claim was made, that the patients had untreated early Lyme disease. In two of the studies the patients had been treated extensively with antibiotics for Lyme disease before the cultures were obtained. Critical evaluation of the three reports suggests that they are invalid. Indeed, two subsequently published studies could not reproduce the results of one of the reports. In a published analysis of another of the reports, investigators from the Centers for Disease Control and Prevention concluded that the cultures were likely to have been contaminated. When the biologic plausibility of recovering borrelia from blood is extremely low, the level of scientific rigor required of a study that claims a positive result should be particularly high. Copyright © 2017 Elsevier Inc. All rights reserved.

Understanding Bacterial Isolates in Blood Culture and Approaches Used to Define Bacteria as Contaminants: A Literature Review.

PubMed

Hossain, Belal; Islam, Mohammad Shahidul; Rahman, Atiqur; Marzan, Mahfuza; Rafiqullah, Iftekhar; Connor, Nicholas E; Hasanuzzaman, Mohammad; Islam, Maksuda; Hamer, Davidson H; Hibberd, Patricia L; Saha, Samir K

2016-05-01

Interpretation of blood culture isolates is challenging due to a lack of standard methodologies for identifying contaminants. This problem becomes more complex when the specimens are from sick young infants, as a wide range of bacteria can cause illness among this group. We used 43 key words to find articles published between 1970 and 2011 on blood culture isolates and possible contaminants in the PubMed database. Experts were also consulted to obtain other relevant articles. Selection of articles followed systematic methods considering opinions from more than 1 reviewer. After reviewing the titles of 3869 articles extracted from the database, we found 307 relevant to our objective. Based on the abstracts, 42 articles were selected for the literature review. In addition, we included 7 more articles based on cross-references and expert advice. The most common methods for differentiating blood culture isolates were multiple blood cultures from the same subject, antibiograms and molecular testing. Streptococcus pneumoniae, Hemophilus influenzae, Neisseria meningitidis and group A and B streptococcus were always considered as pathogens, whereas Bacillus sp., Diphtheroids, Propionibacterium and Micrococcus were commonly regarded as contaminants. Coagulase-negative staphylococci were the most frequent isolates and usually reported as contaminants unless the patient had a specific condition, such as long-term hospitalization or use of invasive devices (catheters). Inaccurate interpretation of blood culture may falsely guide treatment and also has long-term policy implications. The combination of clinical and microbiological knowledge, patient's clinical history and laboratory findings are essential for appropriate interpretation of blood culture.

[Application value of Xpert MTB/RIF in diagnosis of spinal tuberculosis and detection of rifampin resistance].

PubMed

Jin, Yang-Hui; Shi, Shi-Yuan; Zheng, Qi; Shen, Jian; Ying, Xiao-Zhang; Wang, Yi-Fan

2017-09-25

To investigate the application value of Xpert MTB/RIF in diagnosis of spinal tuberculosis and detection of rifampin resistance. The 109 pus specimens were obtained from patients who were primaryly diagnosed as spinal tuberculosis. All of the pus specimens were detected by acid-fast stain, liquid fast culturing by BACTEC MGIT 960 and Xpert MTB/RIF assay to definite the differences in sensitivity and specificity of mycobacterium tuberculosis among detecting methods. Pus specimens obtained by different methods were deteceded by MTB/RIF test to analyze the self-influence on Xpert MTB/RIF test. The result of liquid fast culturing by BACTEC MGIT 960 was used as the gold standard; and the value of Xpert MTB/RIF assay in detecting rifampin resistance was analyzed. The sensitivity of acid-fast stain, liquid fast culturing by BACTEC MGIT 960 and Xpert MTB/RIF assay were 25.92%, 48.15%, 77.78%, respectively. The sensitivity of pus specimens obtained from open surgery, ultrasound positioning puncture and biopsy the sensitivity were 83.78%, 76.47%, 44.68% respectively deteceded by MTB/RIF test. According to the gold standard of the results of liquid fast culturing by BACTEC MGIT 960 assay, the sensitivity and specificity of Xpert MTB/RIF assay in detecting rifampin resistance were 80%(4/5) and 90.70%(39/43), respectively. Xpert MTB/RIF assay has higher value in diagnosis of spinal tuberculosi, and also can detect rifampin resistance. The number of mycobacterium tuberculosis in pus specimens has a great influence in the sensitivity of Xpert MTB/RIF assay.

[Rapid identification of microorganisms by mass spectrometry in a blood culture system. Comparison of two procedures].

PubMed

Cattani, María E; Posse, Tamara; Hermes, Ricardo L; Kaufman, Sara C

2015-01-01

Rapid identification of microorganisms is critical in hospitalized infected patients. Blood culture is currently the gold standard for detecting and identifying microorganisms causing bacteremia or sepsis. The introduction of mass spectrometry by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF MS) in microbiology laboratories, especially in microorganisms growing in blood culture bottles, provides rapid identification. This study evaluates the performance of the Maldi Sepsityper Biotyper procedure (hereinafter, MS) compared to that of an in-home method (hereinafter, HF). Eight hundred and forty (840) positive blood culture bottles were processed using the HF procedure, 542 of which were also processed using MS. The organisms were identified in 670 (79.76%) and 391 (72.14%) bottles respectively (p = 0,0013). This study demonstrates the effectiveness of both procedures for identifying microorganisms directly from positive blood culture bottles. However, the HF procedure proved to be more effective than MS, especially in the presence of Gram positive organisms. Copyright © 2015 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Efficacy of the FilmArray blood culture identification panel for direct molecular diagnosis of infectious diseases from samples other than blood.

PubMed

Micó, Miquel; Navarro, Ferran; de Miniac, Daniela; González, Yésica; Brell, Albert; López, Cristina; Sánchez-Reus, Ferran; Mirelis, Beatriz; Coll, Pere

2015-12-01

Molecular-based techniques reduce the delay in diagnosing infectious diseases and therefore contribute to better patient outcomes. We assessed the FilmArray blood culture identification (BCID) panel (Biofire Diagnostics/bioMérieux) directly on clinical specimens other than blood: cerebrospinal, joint, pleural and ascitic fluids, bronchoscopy samples and abscesses. We compared the results from 88 samples obtained by culture-based techniques. The percentage of agreement between the two methods was 75 % with a Cohen κ value of 0.51. Global sensitivity and specificity using the FilmArray BCID panel were 71 and 97 %, respectively. Sensitivity was poorer in samples with a low bacterial load, such as ascitic and pleural fluids (25 %), whereas the sensitivity for abscess samples was high (89 %). These findings suggest that the FilmArray BCID panel could be useful to perform microbiological diagnosis directly from samples other than positive blood cultures, as it offers acceptable sensitivity and moderate agreement with conventional microbiological methods. Nevertheless, cost-benefit studies should be performed before introducing this method into algorithms for microbiological diagnostics.

[Stimulation of cell cultures recovery after cryopreservation by the cattle cord blood FRACTION (below 5 kDa) or Actovegin].

PubMed

Gulevskiĭ, A K; Trifonova, A V; Lavrik, A A

2013-01-01

The capacities of the cattle cord blood low-molecular fraction (below 5 kDa) and Actovegin (the vealer blood fraction (below 5 kDa)) for recovering functions of cell cultures after cryopreservation compared. Their influence proliferation of the flozen-thawed cell cultures, certain stages of their growth, cell attachment, rate of cell spreading, and mitotic regiment has been studied. Both the cord blood low-molecular fraction and Actovegin were shown to stimulate growth of the cell cultures after cryopreservation more efficiently at the concentration of 224 μg/ml. However, despite the stimulating effect discovered, their application did not bring proliferative indices on the 1st passage after cryopreservation to the values of the native culture. The effects of the cord blood low-molecular fraction and Actovegin on the human fibroblast culture were identical by the following parameters: cell attachment, rates of cell spreading and proliferation. In culture BHK-21 clone 13/04 the efficiency of Actovegin was low, while the cord blood low-molecular fraction has a conspicuous stimulating effect on its adhesion and proliferation. The investigations carried out can serve as a basis for the development of regenerative media containing the cattle cord blood low-molecular fraction (below 5 kDa) or Actovegin as active components at the concentration of 224 μg/ml with the purpose of fast recovery of culture prolifetative properties after cryopreservation.

Clinical and Diagnostic Aspects of Brucellosis and Antimicrobial Susceptibility of Brucella Isolates in Hamedan, Iran.

PubMed

Torkaman Asadi, Fatemeh; Hashemi, Seyyed Hamid; Alikhani, Mohammad Yousef; Moghimbeigi, Abbas; Naseri, Zahra

2017-05-24

Current drug regimens for brucellosis are associated with relatively high rates of therapeutic failure or relapse. Reduced antimicrobial susceptibility of Brucella spp. has been proposed recently as a potential cause of therapeutic failure. The aim of this study was to evaluate the antibiotic resistance pattern of Brucella melitensis clinical isolates by E-test method in Hamadan, west of Iran. In a 15-month period, all patients with suspected brucellosis were enrolled. Blood specimens were collected for diagnosis of brucellosis by BACTEC system and serological tests. Antimicrobial susceptibility of clinical isolates to 7 antibiotics was assessed by the E-test method. One hundred forty-nine patients with brucellosis were evaluated. 38.3% of cultures of clinical samples were positive for BACTEC system, of which 91.2% were associated with a positive serological test result. No significant associations were found between serology and the culture method. All Brucella isolates were susceptible to doxycycline, streptomycin, gentamicin, ciprofloxacin, and moxifloxacin. However, decreased sensitivity to rifampin and trimethoprim-sulfamethoxazole was found in 35.1% and 3.5% of isolates, respectively. Because of the high rates of intermediate sensitivity to rifampin among Brucella isolates, this drug should be prescribed with caution. We recommend restricting the use of rifampin for treatment of brucellosis except as an alternative drug for special situations.

Evaluation of the FilmArray Blood Culture Identification Panel: Results of a Multicenter Controlled Trial

PubMed Central

Salimnia, Hossein; Lephart, Paul R.; Schreckenberger, Paul; DesJarlais, Sharon M.; Johnson, J. Kristie; Robinson, Gwen; Carroll, Karen C.; Greer, Amy; Morgan, Margie; Chan, Raymond; Loeffelholz, Michael; Valencia-Shelton, Frances; Jenkins, Stephen; Schuetz, Audrey N.; Daly, Judy A.; Barney, Trenda; Hemmert, Andrew; Kanack, Kristen J.

2016-01-01

Sepsis is a major cause of morbidity, mortality, and increased medical expense. Rapid diagnosis improves outcomes and reduces costs. The FilmArray blood culture identification panel (BioFire Diagnostics LLC, Salt Lake City, UT), a highly multiplexed PCR assay, can identify 24 etiologic agents of sepsis (8 Gram-positive, 11 Gram-negative, and 5 yeast species) and three antimicrobial resistance genes (mecA, vanA/B, and blaKPC) from positive blood culture bottles. It provides results in about 1 h with 2 min for assay setup. We present the results of an eight-center trial comparing the sensitivity and specificity of the panel with those of the laboratories' standard phenotypic identification techniques, as well as with molecular methods used to distinguish Acinetobacter baumannii from other members of the A. calcoaceticus-A. baumannii complex and to detect antimicrobial resistance genes. Testing included 2,207 positive aerobic blood culture samples, 1,568 clinical and 639 seeded. Samples were tested fresh or were frozen for later testing within 8 h after the bottles were flagged as positive by an automated blood culture system. At least one organism was detected by the panel in 1,382 (88.1%) of the positive clinical specimens. The others contained primarily off-panel organisms. The panel reported multiple organisms in 81 (5.86%) positive clinical specimens. The unresolved blood culture identification sensitivity for all target detections exceeded 96%, except for Klebsiella oxytoca (92.2%), which achieved 98.3% sensitivity after resolution of an unavoidable phenotypic error. The sensitivity and specificity for vanA/B and blaKPC were 100%; those for mecA were 98.4 and 98.3%, respectively. PMID:26739158

Optimal turnaround time for direct identification of microorganisms by mass spectrometry in blood culture.

PubMed

Randazzo, Adrien; Simon, Marc; Goffinet, Pierre; Classen, Jean-François; Hougardy, Nicolas; Pierre, Pascal; Kinzinger, Philippe; Mauel, Etienne; Goffinet, Jean-Sébastien

2016-11-01

During the past few years, several studies describing direct identification of bacteria from blood culture using mass spectrometry have been published. These methods cannot, however, be easily integrated into a common laboratory workflow because of the high hands-on time they require. In this paper, we propose a new method of identification with a short hands-on time and a turnaround time shorter than 15min. Positive blood bottles were homogenised and 600μL of blood were transferred to an Eppendorf tube where 600μL of lysis buffer were added. After homogenisation, a centrifugation step of 4min at 10,500g was performed and the supernatant was discarded. The pellet was then washed and loaded in quadruplicate into wells of a Vitek® MS-DS plate. Each well was covered with a saturated matrix solution and a MALDI-TOF mass spectrometry analysis was performed. Species were identified using the software Myla 3.2.0-2. We analysed 266 positive blood culture bottles. A microorganism grew in 261 cultures, while five bottles remained sterile after 48h of incubation in subculture. Our method reaches a probability of detection at the species level of 77.8% (203/261) with a positive predictive value of 99.5% (202/203). We developed a new method for the identification of microorganisms using mass spectrometry, directly performed from a positive blood culture. This method has short hands-on time and turnaround time and can easily take place in the workflow of a laboratory, with comparable results in performance with other methods reported in the literature. Copyright © 2016 Elsevier B.V. All rights reserved.

Gram-negative and -positive bacteria differentiation in blood culture samples by headspace volatile compound analysis.

PubMed

Dolch, Michael E; Janitza, Silke; Boulesteix, Anne-Laure; Graßmann-Lichtenauer, Carola; Praun, Siegfried; Denzer, Wolfgang; Schelling, Gustav; Schubert, Sören

2016-12-01

Identification of microorganisms in positive blood cultures still relies on standard techniques such as Gram staining followed by culturing with definite microorganism identification. Alternatively, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry or the analysis of headspace volatile compound (VC) composition produced by cultures can help to differentiate between microorganisms under experimental conditions. This study assessed the efficacy of volatile compound based microorganism differentiation into Gram-negatives and -positives in unselected positive blood culture samples from patients. Headspace gas samples of positive blood culture samples were transferred to sterilized, sealed, and evacuated 20 ml glass vials and stored at -30 °C until batch analysis. Headspace gas VC content analysis was carried out via an auto sampler connected to an ion-molecule reaction mass spectrometer (IMR-MS). Measurements covered a mass range from 16 to 135 u including CO2, H2, N2, and O2. Prediction rules for microorganism identification based on VC composition were derived using a training data set and evaluated using a validation data set within a random split validation procedure. One-hundred-fifty-two aerobic samples growing 27 Gram-negatives, 106 Gram-positives, and 19 fungi and 130 anaerobic samples growing 37 Gram-negatives, 91 Gram-positives, and two fungi were analysed. In anaerobic samples, ten discriminators were identified by the random forest method allowing for bacteria differentiation into Gram-negative and -positive (error rate: 16.7 % in validation data set). For aerobic samples the error rate was not better than random. In anaerobic blood culture samples of patients IMR-MS based headspace VC composition analysis facilitates bacteria differentiation into Gram-negative and -positive.

Rapid identification of bacteria in positive blood culture by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

PubMed

Schmidt, V; Jarosch, A; März, P; Sander, C; Vacata, V; Kalka-Moll, W

2012-03-01

Blood culture is probably the most significant specimen used for the diagnosis of bacterial infections, especially for bloodstream infections. In the present study, we compared the resin-containing BD BACTEC™ Plus-Aerobic (Becton Dickinson), non-charcoal-containing BacT/Alert(®) SA (bioMérieux), and charcoal-containing BacT/Alert(®) FA (bioMérieux) blood culture bottles with direct identification by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). A total of 103 bacterial isolates, from clinical blood cultures, representing the most frequent 13 genera and 24 species were examined. Bacteria were extracted from positive blood culture broth by density centrifugation and then subjected to identification by MALDI-TOF MS using two different volumes and chemical treatments. Overall, correct identification by MALDI-TOF MS was obtained for the BD BACTEC™ Plus-Aerobic, BacT/Alert(®) SA, and BacT/Alert(®) FA blood culture bottles in 72%, 45.6%, and 23%, respectively, for gram-negative bacteria in 86.6%, 69.2%, and 47.1%, respectively, and for gram-positive bacteria in 60.0%, 28.8%, and 5.4%, respectively. The lack of identification was observed mainly with viridans streptococci. Depending on the blood culture bottles used in routine diagnostic procedures and the protocol for bacterial preparation, the applied MALDI-TOF MS represents an efficient and rapid method for direct bacterial identification.

Does maternal intrapartum antibiotic treatment prolong the incubation time required for blood cultures to become positive for infants with early-onset sepsis?

PubMed

Sarkar, Siddhartha Sean; Bhagat, Indira; Bhatt-Mehta, Varsha; Sarkar, Subrata

2015-03-01

We hypothesized that maternal intrapartum antibiotic treatment delays the growth of the organism in the blood culture obtained during the work-up for infants with suspected early-onset sepsis (EOS). Single center, retrospective review of infants with blood culture-proven EOS over 13.5 years period. EOS was defined by isolation of a pathogen from blood culture obtained within 72 hours of birth and antibiotic treatment for  ≥ 5 days. Among 81 infants with positive blood cultures, 38 were deemed to have EOS and 43 were deemed contaminants. The organisms grown were as follows: Escherichia coli in 17 infants, Group B streptococcus in 10 infants, and others in 11 infants. Overall, 17 infants with EOS did not receive intrapartum antibiotics and had blood cultures drawn for being symptomatic after birth. The other 21 infants who received intrapartum antibiotics had blood culture drawn primarily for maternal chorioamnionitis. The median (interquartile range [IQR]) incubation time to blood culture positivity was not different in infants who received intrapartum antibiotics compared with infants who did not (19.6 hours, IQR 16-28 hours vs. 19.5 hours, IQR 17.2-21.6 hours, p = 0.7489). Maternal intrapartum antibiotic treatment did not delay the time to blood culture positivity in infants with EOS. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Blood cultures taken from patients attending emergency departments in South Africa are an important antibiotic stewardship tool, which directly influences patient management.

PubMed

Boyles, Tom H; Davis, Kelly; Crede, Thomas; Malan, Jacques; Mendelson, Marc; Lesosky, Maia

2015-10-06

Febrile illness with suspected blood stream infection (BSI) is a common reason for admission to hospital in Africa and blood cultures are therefore an important investigation. Data on the prevalence and causes of community acquired BSI in Africa are scarce and there are no studies from South Africa. There are no validated clinical prediction rules for use of blood cultures in Africa. A prospective observational cohort study of patients attending 2 urban emergency departments in Cape Town, South Africa. The decision to take a blood culture was made by the attending clinician and information available at the time of blood draw was collected. Bottles were weighed to measure volume of blood inoculated. 500 blood culture sets were obtained from 489 patients. 39 (7.8 %) were positive for pathogens and 13 (2.6 %) for contaminants. Significant independent predictors of positive cultures were diastolic blood pressure <60 mmHg, pulse >120 bpm, diabetes and a suspected biliary source of infection, but not HIV infection. Positive results influenced patient management in 36 of 38 (95 %) cases with the organism being resistant to the chosen empiric antibiotic in 9 of 38 (24 %). Taking <8 ml of blood was predictive of a negative culture. The best clinical prediction rule had a negative predictive value (NPV) of 92 % which is unlikely to be high enough to be clinically useful. Blood cultures taken from patients attending emergency departments in a high HIV prevalent city in South Africa are frequently positive and almost always influence patient management. At least 8 ml of blood should be inoculated into each bottle. Blood cultures should be taken from all patients attending EDs in South Africa suspected of having BSI particularly if diabetic, with hypotension, tachycardia or if biliary sepsis is suspected.

Direct Antimicrobial Susceptibility Testing of Gram-Negative Bacilli in Blood Cultures by an Electrochemical Method

PubMed Central

Huang, Ay Huey; Wu, Jiunn Jong; Weng, Yu Mei; Ding, Hwia Cheng; Chang, Tsung Chain

1998-01-01

Nonfastidious aerobic gram-negative bacilli (GNB) are commonly isolated from blood cultures. The feasibility of using an electrochemical method for direct antimicrobial susceptibility testing of GNB in positive blood cultures was evaluated. An aliquot (10 μl) of 1:10-diluted positive blood cultures containing GNB was inoculated into the Bactometer module well (bioMérieux Vitek, Hazelwood, Mo.) containing 1 ml of Mueller-Hinton broth supplemented with an antibiotic. Susceptibility tests were performed in a breakpoint broth dilution format, with the results being categorized as resistant, intermediate, or susceptible. Seven antibiotics (ampicillin, cephalothin, gentamicin, amikacin, cefamandole, cefotaxime, and ciprofloxacin) were used in this study, with each agent being tested at the two interpretive breakpoint concentrations. The inoculated modules were incubated at 35°C, and the change in impedance in each well was continuously monitored for 24 h by the Bactometer. The MICs of the seven antibiotics for each blood isolate were also determined by the standardized broth microdilution method. Of 146 positive blood cultures (1,022 microorganism-antibiotic combinations) containing GNB tested by the direct method, the rates of very major, major, and minor errors were 0, 1.1, and 2.5%, respectively. The impedance method was simple; no centrifugation, preincubation, or standardization of the inocula was required, and the susceptibility results were normally available within 3 to 6 h after inoculation. The rapid method may allow proper antimicrobial treatment almost 30 to 40 h before the results of the standard methods are available. PMID:9738038

A model of psychosocial and cultural antecedents of blood pressure control.

PubMed Central

Bosworth, Hayden B.; Oddone, Eugene Z.

2002-01-01

Hypertension is a major modifiable risk factor for stroke, congestive heart failure, and end-stage renal disease. Hypertension is particularly prevalent and deadly among African Americans. Effective treatment for hypertension has been available for decades, yet only one fourth of all individuals have their blood pressure under control. Despite the potential impact of hypertension, interventions to improve control have had limited success. We present a model of how to understand antecedents of blood pressure control according to three interrelated categories: patient characteristics, social and cultural environment, and medical environment. This theoretical paper was conducted using a literature review and a model to explain psychosocial antecedents of blood pressure control is presented. We conclude that improved understanding of important antecedents of blood pressure control coupled with technological advances, such as tailored interventions, provide clinicians with a tool that may lead to improved blood pressure control. These interventions will require the involvement of clinicians and consideration of sociocultural factors to be successful. PMID:11991336

Mycobacterial blood culture as the only means of diagnosis of disseminated tuberculosis in advanced HIV infection.

PubMed

Mathuram, Alice Joan; Michael, Joy Sarojini; Turaka, Vijay Prakash; Jasmine, Sudha; Carey, Ronald; Ramya, I

2018-04-01

The diagnosis of disseminated tuberculosis (TB) in advanced HIV infection is often delayed because of difficulty in obtaining suitable specimens for culture. A total of 32 such patients from South India with positive mycobacterial blood cultures were studied over ten years. Almost all (90%) had a febrile illness and the majority (68.7%) had clinical lung involvement, but only 27.3% had positive sputum smears. Liver biopsy yielded a positive diagnosis in only 1/7. Cytopenia was almost universal (96.9%). Bone marrow cultures were, however, positive in 54.8%, of whom one-quarter grew atypical mycobacteria. Mycobacterial blood culture is therefore a useful adjunct test to diagnose TB in advanced HIV.

Blood cell oxidative stress precedes hemolysis in whole blood-liver slice co-cultures of rat, dog, and human tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vickers, Alison E.M., E-mail: vickers_alison@allergan.co; Sinclair, John R.; Fisher, Robyn L.

A novel in vitro model to investigate time-dependent and concentration-dependent responses in blood cells and hemolytic events is studied for rat, dog, and human tissues. Whole blood is co-cultured with a precision-cut liver slice. Methimazole (MMI) was selected as a reference compound, since metabolism of its imidazole thione moiety is linked with hematologic disorders and hepatotoxicity. An oxidative stress response occurred in all three species, marked by a decline in blood GSH levels by 24 h that progressed, and preceded hemolysis, which occurred at high MMI concentrations in the presence of a liver slice with rat (>= 1000 muM atmore » 48 h) and human tissues (>= 1000 muM at 48 h, >= 750 muM at 72 h) but not dog. Human blood-only cultures exhibited a decline of GSH levels but minimal to no hemolysis. The up-regulation of liver genes for heme degradation (Hmox1 and Prdx1), iron cellular transport (Slc40a1), and GSH synthesis and utilization (mGST1 and Gclc) were early markers of the oxidative stress response. The up-regulation of the Kupffer cell lectin Lgals3 gene expression indicated a response to damaged red blood cells, and Hp (haptoglobin) up-regulation is indicative of increased hemoglobin uptake. Up-regulation of liver IL-6 and IL-8 gene expression suggested an activation of an inflammatory response by liver endothelial cells. In summary, MMI exposure led to an oxidative stress response in blood cells, and an up-regulation of liver genes involved with oxidative stress and heme homeostasis, which was clearly separate and preceded frank hemolysis.« less

Evaluation of the Verigene Gram-Positive Blood Culture Nucleic Acid Test for Rapid Detection of Bacteria and Resistance Determinants

PubMed Central

Wojewoda, Christina M.; Sercia, Linda; Navas, Maria; Tuohy, Marion; Wilson, Deborah; Hall, Geraldine S.; Procop, Gary W.

2013-01-01

Rapid identification of pathogens from blood cultures can decrease lengths of stay and improve patient outcomes. We evaluated the accuracy of the Verigene Gram-positive blood culture (BC-GP) nucleic acid test for investigational use only (Nanosphere, Inc., Northbrook, IL) for the identification of Gram-positive bacteria from blood cultures. The detection of resistance genes (mecA in Staphylococcus aureus and Staphylococcus epidermidis and vanA or vanB in Enterococcus faecium and Enterococcus faecalis) by the BC-GP assay also was assessed. A total of 186 positive blood cultures (in BacT/Alert FA bottles) with Gram-positive cocci observed with Gram staining were analyzed using the BC-GP assay. The BC-GP results were compared with the identification and susceptibility profiles obtained with routine methods in the clinical laboratory. Discordant results were arbitrated with additional biochemical, cefoxitin disk, and repeat BC-GP testing. The initial BC-GP organism identification was concordant with routine method results for 94.6% of the blood cultures. Only 40% of the Streptococcus pneumoniae identifications were correct. The detection of the mecA gene for 69 blood cultures with only S. aureus or S. epidermidis was concordant with susceptibility testing results. For 3 of 6 cultures with multiple Staphylococcus spp., mecA detection was reported but was correlated with oxacillin resistance in a species other than S. aureus or S. epidermidis. The detection of vanA agreed with susceptibility testing results for 45 of 46 cultures with E. faecalis or E. faecium. Comparison of the mean times to results for each organism group showed that BC-GP results were available 31 to 42 h earlier than phenotypic identifications and 41 to 50 h earlier than susceptibility results. PMID:23596240

Evaluation of the Verigene Gram-positive blood culture nucleic acid test for rapid detection of bacteria and resistance determinants.

PubMed

Wojewoda, Christina M; Sercia, Linda; Navas, Maria; Tuohy, Marion; Wilson, Deborah; Hall, Geraldine S; Procop, Gary W; Richter, Sandra S

2013-07-01

Rapid identification of pathogens from blood cultures can decrease lengths of stay and improve patient outcomes. We evaluated the accuracy of the Verigene Gram-positive blood culture (BC-GP) nucleic acid test for investigational use only (Nanosphere, Inc., Northbrook, IL) for the identification of Gram-positive bacteria from blood cultures. The detection of resistance genes (mecA in Staphylococcus aureus and Staphylococcus epidermidis and vanA or vanB in Enterococcus faecium and Enterococcus faecalis) by the BC-GP assay also was assessed. A total of 186 positive blood cultures (in BacT/Alert FA bottles) with Gram-positive cocci observed with Gram staining were analyzed using the BC-GP assay. The BC-GP results were compared with the identification and susceptibility profiles obtained with routine methods in the clinical laboratory. Discordant results were arbitrated with additional biochemical, cefoxitin disk, and repeat BC-GP testing. The initial BC-GP organism identification was concordant with routine method results for 94.6% of the blood cultures. Only 40% of the Streptococcus pneumoniae identifications were correct. The detection of the mecA gene for 69 blood cultures with only S. aureus or S. epidermidis was concordant with susceptibility testing results. For 3 of 6 cultures with multiple Staphylococcus spp., mecA detection was reported but was correlated with oxacillin resistance in a species other than S. aureus or S. epidermidis. The detection of vanA agreed with susceptibility testing results for 45 of 46 cultures with E. faecalis or E. faecium. Comparison of the mean times to results for each organism group showed that BC-GP results were available 31 to 42 h earlier than phenotypic identifications and 41 to 50 h earlier than susceptibility results.

Blood culture positive Nocardia asteroides infection: a case report.

PubMed

Höpler, Wolfgang; Laferl, Hermann; Szell, Marton; Pongratz, Peter; Brandl, Irmgard; Tucek, Gerhard; Wenisch, Christoph

2013-01-01

We report a case of nocardiosis in a patient with several risk factors for this rare infection. Radiologically, the patient's multiple lung abscesses were misinterpreted as pulmonary metastases. Diagnosis was finally reached by the growth of Nocardia asteroides in two different blood culture sets. Nocardia bacteraemia is a rare clinical event. Despite initiation of an effective antibiotic therapy, the patient died. Autopsy revealed disseminated nocardial abscesses in the lungs, the kidneys and the brain.

CULTURES FROM THE BLOOD In SEPTICÆMIA, PNEUMONIA, MENINGITIS AND CHRONIC DISEASES

PubMed Central

White, Franklin Warren

1899-01-01

Our conclusions from the literature and our own experiments may be summarized as follows: I. Blood for bacteriological examination during life should be taken directly from the veins and in considerable quantity. II. Resorption of toxines is the most important feature in cases of sepsis; pyogenie bacteria invade the general circulation in a rather small proportion even of severe eases, and, as a rule, late in the course of the disease. III. A general infection by the pnenmococcus can be demonstrated occasionally in the late stages of acute lobar pneumonia. IV. The value of blood cultures as a means of diagnosis in obscure cases of sepsis is limited by the fact that invasion of the blood by the specific organism cannot be demonstrated during life in the majority of cases. Positive cultures are very valuable; negative cultures do not exclude local septic infections. V. The detection of specific bacteria in the blood of cases of sepsis and of pneumonia gives a very unfavorable prognosis in most cases. VI. General terminal infections with pyogenic cocci occasionally occur as an immediate cause of death in chronic disease. Local infections processes play this part more frequently. VII. As far as our experiments have shown, invasion of the blood by bacteria during the death agony, with subsequent distribution of the genus to the organs by the circulation, is a rather uncommon occurrence. VIII. Owing to the relative infrequency of agonal invasion, we believe that in the majority of cases where the autopsy is performed promptly after death, the bacteria which are found in the organs succeeded in reaching these organs previously to the death agony, and are associated with the course of the disease. IX. The presence of bacteria in the organs of late autopsies is due in many cases to post-mortem extension from one organ to another, and in some cases to the post-mortem growth of small numbers of genus which were distributed to the organs by means of the circulation. PMID

The efficacy of 2% chlorhexidine gluconate in 70% alcohol compared with 10% povidone iodine in reducing blood culture contamination in pediatric patients.

PubMed

Tangsathapompong, Auchara; Banjongmanee, Pornumpa; Unrit, Kengkaj; Sritipsukho, Paskorn; Mungkornkaew, Narisara; Sajak, Sirinporn

2014-08-01

Blood culture is the gold standard for diagnosis of septicemia. However; false-positive blood cultures are associated with increased health care costs due to unnecessary treatment. To evaluate the efficacy of 2% chlorhexidine gluconate in 70% alcohol compared with 10% povidone iodine in reducing blood culture contamination inpediatricpatients. This is a prospective study of pediatric patients who were admitted at a tertiary-care hospital. Pediatric patients who neededpercutaneous blood cultures were recruitedfrom two general pediatric wards and the pediatric intensive care unit. The authors used 10% povidone iodine as an antiseptic in odd months and 2% chlorhexidine gluconate in 70% alcohol as an antiseptic in even months in obtaining the blood culture samples. There were 1,269 blood culture specimens taken from 821 patients. 654 specimens used 10%povidone iodine as an antiseptic and 619 specimens used 2% chlorhexidine gluconate in 70% alcohol as an antiseptic. The 10% povidone iodine group and the 2% chlorhexidine gluconate in 70% alcohol group had the risk of blood culture contamination of 3.21% (95%CI: 2.00%-4.87%) and 2.28% (95% CI: 1.25%-3.79%) respectively. The risk difference ofblood culture contamination was 0.93% (95% confidence interval: 0. 86-2.72%) with p = 0.31. The most common contamination organism was Coagulase negative staphylococci (68.57%). No adverse skin reactions were observed in both antiseptic solutions groups. Use of2% chlorhexidine gluconate in 70% alcohol as an antiseptic seems to reduce the risk of blood culture contamination compared to use of 10% povidone iodine. In addition, neither of the antiseptic solutions resulted in adverse skin reactions.

Simple Sample Preparation Method for Direct Microbial Identification and Susceptibility Testing From Positive Blood Cultures.

PubMed

Pan, Hong-Wei; Li, Wei; Li, Rong-Guo; Li, Yong; Zhang, Yi; Sun, En-Hua

2018-01-01

Rapid identification and determination of the antibiotic susceptibility profiles of the infectious agents in patients with bloodstream infections are critical steps in choosing an effective targeted antibiotic for treatment. However, there has been minimal effort focused on developing combined methods for the simultaneous direct identification and antibiotic susceptibility determination of bacteria in positive blood cultures. In this study, we constructed a lysis-centrifugation-wash procedure to prepare a bacterial pellet from positive blood cultures, which can be used directly for identification by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and antibiotic susceptibility testing by the Vitek 2 system. The method was evaluated using a total of 129 clinical bacteria-positive blood cultures. The whole sample preparation process could be completed in <15 min. The correct rate of direct MALDI-TOF MS identification was 96.49% for gram-negative bacteria and 97.22% for gram-positive bacteria. Vitek 2 antimicrobial susceptibility testing of gram-negative bacteria showed an agreement rate of antimicrobial categories of 96.89% with a minor error, major error, and very major error rate of 2.63, 0.24, and 0.24%, respectively. Category agreement of antimicrobials against gram-positive bacteria was 92.81%, with a minor error, major error, and very major error rate of 4.51, 1.22, and 1.46%, respectively. These results indicated that our direct antibiotic susceptibility analysis method worked well compared to the conventional culture-dependent laboratory method. Overall, this fast, easy, and accurate method can facilitate the direct identification and antibiotic susceptibility testing of bacteria in positive blood cultures.

Evaluation of the Nanosphere Verigene Gram-Positive Blood Culture Assay with the VersaTREK Blood Culture System and Assessment of Possible Impact on Selected Patients

PubMed Central

Beal, Stacy G.; Ciurca, Jane; Smith, Geremy; John, Jeffrey; Lee, Francesca; Doern, Christopher D.

2013-01-01

The Verigene Gram-positive blood culture (BC-GP) assay (Nanosphere, Northbrook, IL) is a molecular method for the rapid identification of Gram-positive organisms and resistance markers directly from blood culture bottles. A total of 148 VersaTREK REDOX 1 40-ml aerobic bottles demonstrating Gram-positive bacteria were tested. Results were compared with those from conventional biochemical and matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) identifications. We obtained isolates of methicillin-resistant Staphylococcus aureus (MRSA) (24), methicillin-susceptible Staphylococcus aureus (MSSA) (14), methicillin-resistant Staphylococcus epidermidis (MRSE) (17), methicillin-susceptible Staphylococcus epidermidis (MSSE) (9), other coagulase-negative staphylococci (19), Streptococcus salivarius (5), Streptococcus parasanguinis (2), Streptococcus sanguinis (1), Streptococcus cristatus (1), the Streptococcus bovis group (5), Streptococcus agalactiae (9), the Streptococcus anginosus group (1), Streptococcus pneumoniae (6), vancomycin-resistant Enterococcus faecium (VRE FCM) (16), vancomycin-susceptible Enterococcus faecalis (3), Aerococcus viridans (2), Bacillus (6), Corynebacterium (8), Lactobacillus (2), Micrococcus (2), Neisseria mucosa (1), Escherichia coli (3), Candida tropicalis (1), Propionibacterium (1), and Rothia (1). Overall agreement with the culture results was 95%. A total of 137 of 138 (99%) monomicrobial cultures were concordant. We tested 9 polymicrobial samples and found 33% agreement. A chart review of 31 patients with MRSA, MSSA, or VRE demonstrated that the Nanosphere BC-GP assay might have led to more appropriate antibiotic selection for these patients an average of 42 h earlier. Additionally, contact isolation could have been initiated an average of 37 h earlier for patients with MRSA or VRE. The BC-GP assay may have a positive impact on patient care, health care costs, and antibiotic stewardship. PMID:24048531

Is the Presence of Actinomyces spp. in Blood Culture Always Significant?

PubMed Central

Nic-Fhogartaigh, Caoimhe; Millar, Michael

2016-01-01

The isolation of Actinomyces spp. from sterile clinical samples is traditionally regarded as significant. We reviewed the demographic characteristics, clinical risk factors, and outcomes of patients with Actinomyces spp. isolated from blood cultures in our NHS Trust and found that this is not necessarily the case. PMID:26818678

Western Culture in Japanese Film: Kurosawa's "Throne of Blood" and "Ran."

ERIC Educational Resources Information Center

Kane, Peter E.

Akira Kurosawa, the most popular Asian film maker with audiences in the United States, has found in William Shakespeare's plays themes and plots that resonate within Japanese culture. While the translations of "Macbeth" into "Throne of Blood" and "King Lear" into "Ran" are quite direct and literal with only…

Prevalence of ESBL-producing Enterobacteriaceae isolated from blood cultures in Mali.

PubMed

Sangare, Samba Adama; Maiga, Almoustapha Issiaka; Guindo, Ibrehima; Maiga, Aminata; Camara, Namory; Dicko, Oumar Agaly; Diallo, Souleymane; Bougoudogo, Flabou; Armand-Lefevre, Laurence; Andremont, Antoine; Maiga, Ibrahim Izetiegouma

2016-10-31

The increasing frequency of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae is becoming a serious public health concern. This study sought to determine ESBL frequency in Enterobacteriaceae isolated from patients' blood cultures in two university teaching hospitals of Bamako, Mali. During a three-month period, the presence of Enterobacteriaceae from blood cultures of patients admitted to the university teaching hospitals of Bamako was evaluated. The microbial identifications were initially performed with an API 20E gallery and VITEK2 locally in Mali, and then confirmation in France was performed with a mass spectrometry MALDI-TOF in the bacteriology laboratory of the university teaching hospital of Bichat. Antibiotic susceptibility profiles were determined by the diffusion method as recommended by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). The isolated species were K. pneumoniae (14/40; 35.0%), E. coli (11/40; 27.5%), and E. cloacae (9/40; 22.5%). Of the strains isolated, 21/34 (61.8%) had an ESBL phenotype, including 10/14 (71.4%) K. pneumoniae, 8/11 (72.7%) E. coli, and 3/9 (33.3%) E. cloacae. Resistances associated with ESBL strains of K. pneumoniae, E. coli, and E. cloacae were as follows: gentamicin (10/10, 100%; 6/8, 75%; 2/3, 67%, respectively), amikacin (2/10, 20%; 0/8, 0%; 0/3, 0%, respectively), ofloxacin (8/10, 80%; 7/8, 87%; 3/3, 100%, respectively), and cotrimoxazole (10/10, 100%; 6/8, 75%; 3/3, 100%, respectively). Almost two-thirds (61.8%) of Enterobacteriaceae isolated from our blood cultures were ESBL producers. Only susceptibilities to carbapenems and to amikacin were fully conserved within the strains.

Acceleration of Antimicrobial Susceptibility Testing of Positive Blood Cultures by Inoculation of Vitek 2 Cards with Briefly Incubated Solid Medium Cultures

PubMed Central

Idelevich, Evgeny A.; Schüle, Isabel; Grünastel, Barbara; Wüllenweber, Jörg; Peters, Georg

2014-01-01

Briefly incubated agar cultures from positive blood cultures were used for antimicrobial susceptibility testing (AST) by Vitek 2. The cultivation time until inoculation was 3.8 h for Gram-positive cocci and 2.4 h for Gram-negative rods. The error rates were low, providing early and reliable AST without additional time or cost expenditure. PMID:25165084

Knowledge of Good Blood Culture Sampling Practice among Healthcare Staffs in An Emergency Department - Are We Getting It Right?

PubMed

Chew, K S; Mohd Hashairi, F; Jusoh, A F; Aziz, A A; Nik Hisamuddin, N A R; Siti Asma, H

2013-08-01

Although a vital test, blood culture is often plagued with the problem of contamination and false results, especially in a chaotic emergency department setting. The objectives of this pilot study is to find out the level of understanding among healthcare staffs in emergency department, Hospital Universiti Sains Malaysia (HUSM) regarding good blood culture sampling practice. All healthcare staffs in emergency department, HUSM who consented to this study were given a set of selfadministered anonymous questionnaire to fill. More than half (53.1%) of the 64 participants are emergency medicine residents. Majority of them (75%) have been working in the emergency medicine, HUSM for more than 2 years. More than half of them were able to answer correctly the amount of blood volume needed for culture in adult and pediatric patients. When asked what are the factors required to improve the true yield as well as to reduce the risk of culture contamination, the four commonest answers given were observing proper aseptic technique during blood sampling, donning sterile glove, proper hand scrubbing as well as ensuring the sterility of the equipments. This study suggests that there is a lack of proper knowledge of good blood culture sampling practice among our healthcare staffs in emergency department.

Molecular Detection of Streptococcus pneumoniae on Dried Blood Spots from Febrile Nigerian Children Compared to Culture

PubMed Central

Iroh Tam, Pui-Ying; Hernandez-Alvarado, Nelmary; Schleiss, Mark R.; Hassan-Hanga, Fatimah; Onuchukwu, Chuma; Umoru, Dominic; Obaro, Stephen K.

2016-01-01

Background Nigeria has one of the highest burdens of pneumococcal disease in the world, but accurate surveillance is lacking. Molecular detection of infectious pathogens in dried blood spots (DBS) is an ideal method for surveillance of infections in resource-limited settings because of its low cost, minimal blood volumes involved, and ease of storage at ambient temperature. Our study aim was to evaluate a Streptococcus pneumoniae real-time polymerase chain reaction (rt-PCR) assay on DBS from febrile Nigerian children on Whatman 903 and FTA filter papers, compared to the gold standard of culture. Methods Between September 2011 to May 2015, blood was collected from children 5 years of age or under who presented to six hospital study sites throughout northern and central Nigeria with febrile illness, and inoculated into blood culture bottles or spotted onto Whatman 903 or FTA filter paper. Culture and rt-PCR were performed on all samples. Results A total of 537 DBS specimens from 535 children were included in the study, of which 15 were culture-positive for S. pneumoniae. The rt-PCR assay detected S. pneumoniae in 12 DBS specimens (2.2%). One positive rt-PCR result was identified in a culture-negative specimen from a high-risk subject, and two positive rt-PCR results were negative on repeat testing. Six culture-confirmed cases of S. pneumoniae bacteremia were missed. Compared to culture, the overall sensitivities of Whatman 903 and FTA DBS for detection of S. pneumoniae were 57.1% (95% CI 18.4–90.1%) and 62.5% (95% CI 24.5–91.5%), respectively. Nonspecific amplification was noted in an additional 22 DBS (4.1%). Among these, six were positive for a non-S. pneumoniae pathogen on culture. Conclusions Rt-PCR was able to detect S. pneumoniae from clinical DBS specimens, including from a culture-negative specimen. Our findings show promise of this approach as a surveillance diagnostic, but also raise important cautionary questions. Several DBS specimens were detected as

Molecular Detection of Streptococcus pneumoniae on Dried Blood Spots from Febrile Nigerian Children Compared to Culture.

PubMed

Iroh Tam, Pui-Ying; Hernandez-Alvarado, Nelmary; Schleiss, Mark R; Hassan-Hanga, Fatimah; Onuchukwu, Chuma; Umoru, Dominic; Obaro, Stephen K

2016-01-01

Nigeria has one of the highest burdens of pneumococcal disease in the world, but accurate surveillance is lacking. Molecular detection of infectious pathogens in dried blood spots (DBS) is an ideal method for surveillance of infections in resource-limited settings because of its low cost, minimal blood volumes involved, and ease of storage at ambient temperature. Our study aim was to evaluate a Streptococcus pneumoniae real-time polymerase chain reaction (rt-PCR) assay on DBS from febrile Nigerian children on Whatman 903 and FTA filter papers, compared to the gold standard of culture. Between September 2011 to May 2015, blood was collected from children 5 years of age or under who presented to six hospital study sites throughout northern and central Nigeria with febrile illness, and inoculated into blood culture bottles or spotted onto Whatman 903 or FTA filter paper. Culture and rt-PCR were performed on all samples. A total of 537 DBS specimens from 535 children were included in the study, of which 15 were culture-positive for S. pneumoniae. The rt-PCR assay detected S. pneumoniae in 12 DBS specimens (2.2%). One positive rt-PCR result was identified in a culture-negative specimen from a high-risk subject, and two positive rt-PCR results were negative on repeat testing. Six culture-confirmed cases of S. pneumoniae bacteremia were missed. Compared to culture, the overall sensitivities of Whatman 903 and FTA DBS for detection of S. pneumoniae were 57.1% (95% CI 18.4-90.1%) and 62.5% (95% CI 24.5-91.5%), respectively. Nonspecific amplification was noted in an additional 22 DBS (4.1%). Among these, six were positive for a non-S. pneumoniae pathogen on culture. Rt-PCR was able to detect S. pneumoniae from clinical DBS specimens, including from a culture-negative specimen. Our findings show promise of this approach as a surveillance diagnostic, but also raise important cautionary questions. Several DBS specimens were detected as S. pneumoniae by rt-PCR despite growth of

Impact of Implementation of an Automated Liquid Culture System on Diagnosis of Tuberculous Pleurisy.

PubMed

Lee, Byung Hee; Yoon, Seong Hoon; Yeo, Hye Ju; Kim, Dong Wan; Lee, Seung Eun; Cho, Woo Hyun; Lee, Su Jin; Kim, Yun Seong; Jeon, Doosoo

2015-07-01

This study was conducted to evaluate the impact of implementation of an automated liquid culture system on the diagnosis of tuberculous pleurisy in an HIV-uninfected patient population. We retrospectively compared the culture yield, time to positivity, and contamination rate of pleural effusion samples in the BACTEC Mycobacteria Growth Indicator Tube 960 (MGIT) and Ogawa media among patients with tuberculous pleurisy. Out of 104 effusion samples, 43 (41.3%) were culture positive on either the MGIT or the Ogawa media. The culture yield of MGIT was higher (40.4%, 42/104) than that of Ogawa media (18.3%, 19/104) (P<0.001). One of the samples was positive only on the Ogawa medium. The median time to positivity was faster in the MGIT (18 days, range 8-32 days) than in the Ogawa media (37 days, range 20-59 days) (P<0.001). No contamination or growth of nontuberculous mycobacterium was observed on either of the culture media. In conclusion, the automated liquid culture system could provide approximately twice as high yields and fast results in effusion culture, compared to solid media. Supplemental solid media may have a limited impact on maximizing sensitivity in effusion culture; however, further studies are required.

Cost analysis of strategies to reduce blood culture contamination in the emergency department: sterile collection kits and phlebotomy teams.

PubMed

Self, Wesley H; Talbot, Thomas R; Paul, Barbara R; Collins, Sean P; Ward, Michael J

2014-08-01

Blood culture collection practices that reduce contamination, such as sterile blood culture collection kits and phlebotomy teams, increase up-front costs for collecting cultures but may lead to net savings by eliminating downstream costs associated with contamination. The study objective was to compare overall hospital costs associated with 3 collection strategies: usual care, sterile kits, and phlebotomy teams. Cost analysis. This analysis was conducted from the perspective of a hospital leadership team selecting a blood culture collection strategy for an adult emergency department (ED) with 8,000 cultures drawn annually. Total hospital costs associated with 3 strategies were compared: (1) usual care, with nurses collecting cultures without a standardized protocol; (2) sterile kits, with nurses using a dedicated sterile collection kit; and (3) phlebotomy teams, with cultures collected by laboratory-based phlebotomists. In the base case, contamination rates associated with usual care, sterile kits, and phlebotomy teams were assumed to be 4.34%, 1.68%, and 1.10%, respectively. Total hospital costs included costs of collecting cultures and hospitalization costs according to culture results (negative, true positive, and contaminated). Compared with usual care, annual net savings using the sterile kit and phlebotomy team strategies were $483,219 and $288,980, respectively. Both strategies remained less costly than usual care across a broad range of sensitivity analyses. EDs with high blood culture contamination rates should strongly consider evidence-based strategies to reduce contamination. In addition to improving quality, implementing a sterile collection kit or phlebotomy team strategy is likely to result in net cost savings.

Direct identification of bacteria in positive blood culture bottles by matrix-assisted laser desorption ionisation time-of-flight mass spectrometry.

PubMed

La Scola, Bernard; Raoult, Didier

2009-11-25

With long delays observed between sampling and availability of results, the usefulness of blood cultures in the context of emergency infectious diseases has recently been questioned. Among methods that allow quicker bacterial identification from growing colonies, matrix-assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry was demonstrated to accurately identify bacteria routinely isolated in a clinical biology laboratory. In order to speed up the identification process, in the present work we attempted bacterial identification directly from blood culture bottles detected positive by the automate. We prospectively analysed routine MALDI-TOF identification of bacteria detected in blood culture by two different protocols involving successive centrifugations and then lysis by trifluoroacetic acid or formic acid. Of the 562 blood culture broths detected as positive by the automate and containing one bacterial species, 370 (66%) were correctly identified. Changing the protocol from trifluoroacetic acid to formic acid improved identification of Staphylococci, and overall correct identification increased from 59% to 76%. Lack of identification was observed mostly with viridans streptococci, and only one false positive was observed. In the 22 positive blood culture broths that contained two or more different species, only one of the species was identified in 18 samples, no species were identified in two samples and false species identifications were obtained in two cases. The positive predictive value of bacterial identification using this procedure was 99.2%. MALDI-TOF MS is an efficient method for direct routine identification of bacterial isolates in blood culture, with the exception of polymicrobial samples and viridans streptococci. It may replace routine identification performed on colonies, provided improvement for the specificity of blood culture broths growing viridans streptococci is obtained in the near future.

Unique blood culture for diagnosis of bloodstream infections in emergency departments: a prospective multicentre study.

PubMed

Dargère, S; Parienti, J-J; Roupie, E; Gancel, P-E; Wiel, E; Smaiti, N; Loiez, C; Joly, L-M; Lemée, L; Pestel-Caron, M; du Cheyron, D; Verdon, R; Leclercq, R; Cattoir, V

2014-11-01

Detection of microorganisms by blood cultures (BCs) is essential in managing patients with bacteraemia. Rather than the number of punctures, the volume of blood drawn is considered paramount in efficient and reliable detection of microorganisms. We performed a 1-year prospective multicentre study in adult emergency departments of three French university hospitals comparing two methods for BCs: a unique blood culture (UBC) collecting a large volume of blood (40 mL) and the standard method of multiple blood cultures (MBC). The performances of both methods for bacterial contamination and efficient microbial detection were compared, each patient serving as his own control. Amongst the 2314 patients included, three hundred were positive for pathogens (n=245) or contaminants (n=55). Out of the 245 patients, 11 were positive for pathogens by UBC but negative by MBC and seven negative by UBC but positive by MBC (p 0.480). In the subgroup of 137 patients with only two BCs, UBC was superior to MBC (p 0.044). Seven and 17 patients had contaminated BCs by UBC and MBC only, respectively (p 0.062). Considering the sums of pathogens missed and contaminants, UBC significantly outperformed MBC (p 0.045). Considering the complete picture of cost savings, efficient detection of microorganisms and decrease in contaminations, UBC offers an interesting alternative to MBC. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Is the Presence of Actinomyces spp. in Blood Culture Always Significant?

PubMed

Jeffery-Smith, Anna; Nic-Fhogartaigh, Caoimhe; Millar, Michael

2016-04-01

The isolation ofActinomycesspp. from sterile clinical samples is traditionally regarded as significant. We reviewed the demographic characteristics, clinical risk factors, and outcomes of patients withActinomycesspp. isolated from blood cultures in our NHS Trust and found that this is not necessarily the case. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Activity of Nitazoxanide and Tizoxanide against Mycobacterium tuberculosis in vitro and in whole blood culture

PubMed Central

Harausz, Elizabeth P.; Chervenak, Keith A.; Good, Caryn E.; Jacobs, Michael R.; Wallis, Robert S.; Sanchez-Felix, Manuel; Boom, W. Henry

2016-01-01

Nitazoxanide (NTZ) and its metabolite tizoxanide (TIZ) were studied as antimycobacterial agents in vitro (in mycobacterial growth indicator tube [MGIT] cultures) and in a whole blood bactericidal assay. Both NTZ and TIZ show high protein binding. In MGIT cultures (albumin concentration=78 µM), inhibition of Mycobacterium tuberculosis growth occurred at total drug concentrations of ≥16 µg/ml, whereas in whole blood cultures (albumin concentration=350 µM), ≥128 µg/ml was required. Free drug fractions at these two conditions were estimated to be 69% and 2%, respectively. Co-incubation of NTZ and TIZ in human plasma for 72 hours nearly completely eliminated their ability to inhibit mycobacterial growth in MGIT. Interactions with plasma proteins may limit the potential of NTZ and TIZ as drugs for human tuberculosis. PMID:27156623

MALDI-TOF mass spectrometry for early identification of bacteria grown in blood culture bottles.

PubMed

Zabbe, Jean-Benoît; Zanardo, Laura; Mégraud, Francis; Bessède, Emilie

2015-08-01

This note reports an interesting way to rapidly identify bacteria grown from blood culture bottles. Chocolate agar plates were inoculated with 1 drop of the positive blood bottle medium. After a 3-hour incubation, the growth veil was submitted to MALDI-TOF mass spectrometry: 77% of the bacteria present have been correctly identified. Copyright © 2015 Elsevier B.V. All rights reserved.

Potential substitution of cord blood for infant blood in the neonatal sepsis evaluation.

PubMed

Hansen, Anne; Forbes, Peter; Buck, Rosanne

2005-01-01

Evaluation of sepsis accounts for one third of all nursery triage admissions. If umbilical cord blood could be accurately substituted for infant blood, it would spare infants the discomfort of an invasive procedure and save both time and resources. While awaiting 48-hour blood culture results, we decide on clinical management based on whether the white blood cell (WBC) immature to total (I:T) granulocyte ratio is >or=0.2. Our goal was to assess the correlation of complete blood count (CBC), I:T ratio and blood culture results between umbilical cord and infant blood. We conducted a prospective cohort study comparing CBC/differential and blood culture results of paired samples of umbilical cord and infant blood from term newborns. We sent 113 paired samples of cord and infant venous blood for CBC/differential and blood culture. All 113 umbilical cord and infant blood cultures were negative, yielding a false-positive blood culture rate of zero. For 92% of babies, both the cord and infant blood I:T ratio were <0.2 or both were >or=0.2. Cord and infant WBC, hematocrit and platelet counts were moderately to highly correlated. We conclude that cord blood can be safely substituted for infant blood in routine sepsis evaluations of asymptomatic, term infants based on both the low false-positive cord blood culture rate and the significant association between high I:T ratios in cord and infant blood. Copyright (c) 2005 S. Karger AG, Basel.

Misinterpretation of Gram Stain from the Stationary Growth Phase of Positive Blood Cultures for Brucella and Acinetobacter Species.

PubMed

Bazzi, Ali M; Al-Tawfiq, Jaffar A; Rabaan, Ali A

2017-01-01

Acinetobacter baumannii and Brucella species are Gram-negative organisms that are vulnerable to misinterpretation as Gram-positive or Gram-variable in blood cultures. We assess the random errors in gram stain interpretation to reduce the likelihood of such errors and therefore patient harm. Aerobic and anaerobic blood cultures from two patients in an acute care facility in Saudi Arabia were subjected to preliminary Gram-staining. In case 1, VITEK-2 Anaerobe Identification, repeat Gram staining from a blood agar plate, Remel BactiDrop™ Oxidase test, Urea Agar urease test and real-time PCR were used to confirm presence of Brucella and absence of Coryneform species. In case 2, repeat Gram- staining from the plate and the vials, VITEK-2 Gram-Negative Identification, real-time PCR and subculture on to Columbia agar, blood agar, and MacConkey agar were carried out to identify A. baumannii . In case 1, initially pleomorphic Gram-positive bacteria were identified. Coryneform species were suspected. Tiny growth was observed after 24 h on blood agar plates, and good growth by 48 h. Presence of Brucella species was ultimately confirmed. In case 2, preliminary Gram-stain results suggested giant Gram-positive oval cocci. Further testing over 18-24 h identified A. baumannii . Oxidase test from the plate and urease test from the culture vial is recommended after apparent identification of pleomorphic Gram-positive bacilli from blood culture, once tiny growth is observed, to distinguish Brucella from Corynebacterium species. If giant Gram-positive oval cocci are indicated by preliminary Gram-staining, it is recommended that the Gram stain be repeated from the plate after 4-6 h, or culture should be tested in Triple Sugar Iron (TSI) medium and the Gram stain repeated after 2-4 h incubation.

FACTORS ASSOCIATED WITH ODONTOGENIC BACTERAEMIA IN ORTHODONTIC PATIENTS.

PubMed

Umeh, O D; Sanu, O O; Utomi, I L; Nwaokorie, F O

2016-01-01

Various researches have investigated factors associated with the prevalence and intensity of bacteraemia following oral procedures including orthodontic procedures. The aim of this study was to determine the effect of age, gender, plaque and gingival indices on the occurrence of odontogenic bacteraemia following orthodontic treatment procedures. Orthodontic Clinic, Lagos University Teaching Hospital (LUTH), Lagos , Nigeria. Using the consecutive, convenience sampling method, a total of 100 subjects who met the inclusion criteria were recruited for the study and peripheral blood was collected before and again within 2 minutes of completion of orthodontic procedures for microbiologic analysis using the BACTEC automated blood culture system and the lysis filtration methods of blood culturing. The subjects were randomly placed in one of four orthodontic procedures investigated: alginate impression making (Group I), separator placement (Group II), band cementation (Group III) and arch wire change (Group IV). Plaque and gingival indices were assessed using the plaque component of the Simplified Oral Hygiene Index (OHI-S) (Greene & Vermillion) and Modified gingival index (Lobene) respectively before blood collection. Spearman Point bi-serial correlations and logistic regression statistics were used for statistical evaluations at p < 0.05 level. An overall baseline prevalence of bacteraemia of 3% and 17% were observed using the BACCTEC and lysis filtration methods respectively. Similarly, overall prevalence of bacteraemia following orthodontic treatment procedures of 16% and 28% were observed respectively using the BACTEC and lysis filtration methods. A statistically significant increase in the prevalence of bateraemia was observed following separator placement (p=0.016). An increase in age, plaque index scores and modified gingival index scores of the subjects were found to be associated with an increase in the prevalence of bacteraemia following orthodontic treatment

An Automated Sample Preparation Instrument to Accelerate Positive Blood Cultures Microbial Identification by MALDI-TOF Mass Spectrometry (Vitek®MS).

PubMed

Broyer, Patrick; Perrot, Nadine; Rostaing, Hervé; Blaze, Jérome; Pinston, Frederic; Gervasi, Gaspard; Charles, Marie-Hélène; Dachaud, Fabien; Dachaud, Jacques; Moulin, Frederic; Cordier, Sylvain; Dauwalder, Olivier; Meugnier, Hélène; Vandenesch, Francois

2018-01-01

Sepsis is the leading cause of death among patients in intensive care units (ICUs) requiring an early diagnosis to introduce efficient therapeutic intervention. Rapid identification (ID) of a causative pathogen is key to guide directed antimicrobial selection and was recently shown to reduce hospitalization length in ICUs. Direct processing of positive blood cultures by MALDI-TOF MS technology is one of the several currently available tools used to generate rapid microbial ID. However, all recently published protocols are still manual and time consuming, requiring dedicated technician availability and specific strategies for batch processing. We present here a new prototype instrument for automated preparation of Vitek ® MS slides directly from positive blood culture broth based on an "all-in-one" extraction strip. This bench top instrument was evaluated on 111 and 22 organisms processed using artificially inoculated blood culture bottles in the BacT/ALERT ® 3D (SA/SN blood culture bottles) or the BacT/ALERT Virtuo TM system (FA/FN Plus bottles), respectively. Overall, this new preparation station provided reliable and accurate Vitek MS species-level identification of 87% (Gram-negative bacteria = 85%, Gram-positive bacteria = 88%, and yeast = 100%) when used with BacT/ALERT ® 3D and of 84% (Gram-negative bacteria = 86%, Gram-positive bacteria = 86%, and yeast = 75%) with Virtuo ® instruments, respectively. The prototype was then evaluated in a clinical microbiology laboratory on 102 clinical blood culture bottles and compared to routine laboratory ID procedures. Overall, the correlation of ID on monomicrobial bottles was 83% (Gram-negative bacteria = 89%, Gram-positive bacteria = 79%, and yeast = 78%), demonstrating roughly equivalent performance between manual and automatized extraction methods. This prototype instrument exhibited a high level of performance regardless of bottle type or BacT/ALERT system. Furthermore, blood culture workflow could potentially

Prospective comparison of eubacterial PCR and measurement of procalcitonin levels with blood culture for diagnosing septicemia in intensive care unit patients.

PubMed

Rowther, Farjana B; Rodrigues, Camilla S; Deshmukh, Minal S; Kapadia, Farhad N; Hegde, Ashit; Mehta, Ajita P; Joshi, Vinay R

2009-09-01

Rapid identification of infection has a major impact on the clinical course, management, and outcome of critically ill intensive care unit (ICU) patients. We compared the results of PCR and procalcitonin with blood culture for ICU patients suspected of having septicemia. Ninety patients (60 patients meeting the criteria for sepsis and 30 patients not meeting the criteria for sepsis) were evaluated. Compared with blood culture as the gold standard, the sensitivity, specificity, and positive and negative predictive values for PCR were 100%, 43.33%, 46.87%, and 100%, respectively, and for procalcitonin were 100%, 61.66%, 56.6%, and 100%, respectively. The average times required to produce a final result were as follows: PCR, 10 h; blood culture, 33 h; procalcitonin, 45 min. Both PCR and procalcitonin may be useful as rapid tests for detecting septicemia but compared with blood cultures lacked specificity.

Utility of Acridine Orange staining for detection of bacteria from positive blood cultures.

PubMed

Neeraja, M; Lakshmi, V; Padmasri, C; Padmaja, K

2017-08-01

The diagnostic performance of AO stain was evaluated for the detection of bacteria and or fungi from positive blood cultures. The sensitivity of Gram stain (GS) was 98.26% while Acridine Orange (AO) stain proved to be more sensitive (100%) with a Positive and Negative Predictive Value of 100% each. The specificity of both the stains was 100%. Overall agreement between the two stains was 98.23% (688/700). The organisms that were missed by GS and positive by AO were Candida species (Sutton, 2006) and Gram negative bacilli (GNB) (Sutton, 2006). Sensitivity of GS was 82.35% and AO was 100% among mixed cultures. Immediate reporting of the results of AO stain would have a significant impact on clinical management of patients with serious blood stream infections. Copyright © 2017 Elsevier B.V. All rights reserved.

Modeling the Blood-Brain Barrier in a 3D triple co-culture microfluidic system.

PubMed

Adriani, G; Ma, D; Pavesi, A; Goh, E L K; Kamm, R D

2015-01-01

The need for a blood-brain barrier (BBB) model that accurately mimics the physiological characteristics of the in-vivo situation is well-recognized by researchers in academia and industry. However, there is currently no in-vitro model allowing studies of neuronal growth and/or function influenced by factors from the blood that cross through the BBB. Therefore, we established a 3D triple co-culture microfluidic system using human umbilical vein endothelial cells (HUVEC) together with primary rat astrocytes and neurons. Immunostaining confirmed the successful triple co-culture system consisting of an intact BBB with tight intercellular junctions in the endothelial monolayer. The BBB selective permeability was determined by a fluorescent-based assay using dextrans of different molecular weights. Finally, neuron functionality was demonstrated by calcium imaging.

Deficits in knowledge, attitude, and practice towards blood culture sampling: results of a nationwide mixed-methods study among inpatient care physicians in Germany.

PubMed

Raupach-Rosin, Heike; Duddeck, Arne; Gehrlich, Maike; Helmke, Charlotte; Huebner, Johannes; Pletz, Mathias W; Mikolajczyk, Rafael; Karch, André

2017-08-01

Blood culture (BC) sampling rates in Germany are considerably lower than recommended. Aim of our study was to assess knowledge, attitudes, and practice of physicians in Germany regarding BC diagnostics. We conducted a cross-sectional mixed-methods study among physicians working in inpatient care in Germany. Based on the results of qualitative focus groups, a questionnaire-based quantitative study was conducted in 2015-2016. In total, 706 medical doctors and final-year medical students from 11 out of 16 federal states in Germany participated. BC sampling was considered an important diagnostic tool by 95% of the participants. However, only 23% of them would collect BCs in three scenarios for which BC ordering is recommended by present guidelines in Germany; almost one out of ten physicians would not have taken blood cultures in any of the three scenarios. The majority of participants (74%) reported not to adhere to the guideline recommendation that blood culture sampling should include at least two blood culture sets from two different injection sites. High routine in blood culture sampling, perceived importance of blood culture diagnostics, the availability of an in-house microbiological lab, and the department the physician worked in were identified as predictors for good blood culture practice. Our study suggests that there are substantial deficits in BC ordering and the application of guidelines for good BC practice in Germany. Based on these findings, multimodal interventions appear necessary for improving BC diagnostics.

[Pooled Umbilical Cord Blood Plasma for Culturing UCMSC and Ex Vivo Expanding Umbilical Cord Blood CD34⁺ Cells].

PubMed

Wu, Jie-Ying; Lu, Yan; Chen, Jin-Song; Wu, Shao-Qing; Tang, Xue-Wei; Li, Yan

2015-08-01

To investigate the feasibility of umbilical cord blood plasma (UCP) as a replacement for fetal bovine serum (FBS) for culturing mesenchymal stem cells (MSC) derived from umbilical cord, and to observe the supporting effects of these cells (served as a feeder layer) on ex vivo expanding of human umbilical cord blood CD34(+) cells. Umbilical cord blood (UCB) units were suitable if the Guangzhou cord blood bank donor selection criteria strictly were fulfilled. UCP were ready to use after the collection from the plasma depletion/reduction during the processing and pooling of suitable UCB units (at least 30 units were screened for pathogens and microorganisms, and qualified). Umbilical cord mesenchymal stem cells (UCMSC) were harvested from the umbilical cord tissue of health full-term newborns after delivery by enzyme digestion and divided into 3 groups: group 1 and 2 were cultured in the presence of DMEM/F12 containing either FBS or UCP; and group 3 was cultured in serum-free medium (StemPro® MSC SFM CTS™). Morphology, proliferation and surface marker expression were examined by flow cytometry, and the differentiation toward adipogenic and osteogenic lineages was used for investigating the effect of media on UCMSC after 3-5 passages. Next, the cells cultured in the three different media were cryopreserved and thawed, then prepared as feeder layers with the name of UCMSC(FBS), UCMSC(UCP), and UCMSC(SFM), respectively. The CD34⁺ cells were separated from UCB by magnetic activated cell sorting (MACS) and divided into 4 groups cultured in StemPro(-34) SFM medium added with hematopoietic cytokine combination (StemSpan® CC100). The control group included only CD34⁺ cells as group A (blank control) and experimental groups included UCMSC(FBS) + CD34⁺ cells as group B, UCMSC(UCP) + CD34⁺ cells as group C, UCMSC(SFM) + CD34⁺ cells as group D, and cells in all groups were cultured ex vivo for 7 days. The nucleated cell (NC) number was counted by cell counter, CD34�

Lack of Utility of the Lysis-Centrifugation Blood Culture Method for Detection of Fungemia in Immunocompromised Cancer Patients

PubMed Central

Creger, Richard J.; Weeman, Kisa E.; Jacobs, Michael R.; Morrissey, Anne; Parker, Pamela; Fox, Robert M.; Lazarus, Hillard M.

1998-01-01

We retrospectively compared the utility of a fungal isolation device (Isolator) versus conventional techniques for recovering fungal organisms from blood cultures obtained from neutropenic cancer patients. Positive cultures were deemed true pathogens, possible pathogens, or contaminants according to laboratory and clinical criteria. Fifty-three patients had 66 positive blood cultures for fungi, nine on multiple occasions. In 20 episodes true pathogens were recovered, 6 from broth medium alone, 4 from the Isolator system alone, and 10 from both systems. False-negative cultures were noted in 4 of 20 (20%) cases in which broth medium was used and in 6 of 20 (30%) cases in which the Isolator system was used. Possible pathogens were detected in 4 of 66 blood culture-positive cases. Forty-two positive cultures were considered contaminants, 1 collected from standard medium and 41 of 42 (98%) which grew only in Isolators. Eleven of 18 patients with true fungal infections expired as a result of infection, while 4 of 33 patients with a contaminant expired, none from a fungal cause. We do not advocate the routine use of Isolator tubes in the evaluation of the febrile, neutropenic patient due to the high rates of false positives and of contamination. PMID:9431970

Induction of vascular endothelial phenotype and cellular proliferation from human cord blood stem cells cultured in simulated microgravity

NASA Astrophysics Data System (ADS)

Chiu, Brian; Z-M Wan, Jim; Abley, Doris; Akabutu, John

2005-05-01

Recent studies have demonstrated that stem cells derived from adult hematopoietic tissues are capable of trans-differentiation into non-hematopoietic cells, and that the culture in microgravity ( μg) may modulate the proliferation and differentiation. We investigated the application of μg to human umbilical cord blood stem cells (CBSC) in the induction of vascular endothelial phenotype expression and cellular proliferation. CD34+ mononuclear cells were isolated from waste human umbilical cord blood samples and cultured in simulated μg for 14 days. The cells were seeded in rotary wall vessels (RWV) with or without microcarrier beads (MCB) and vascular endothelial growth factor was added during culture. Controls consisted of culture in 1 G. The cell cultures in RWV were examined by inverted microscopy. Cell counts, endothelial cell and leukocyte markers performed by flow-cytometry and FACS scan were assayed at days 1, 4, 7 and at the termination of the experiments. Culture in RWV revealed significantly increased cellular proliferation with three-dimensional (3D) tissue-like aggregates. At day 4, CD34+ cells cultured in RWV bioreactor without MCB developed vascular tubular assemblies and exhibited endothelial phenotypic markers. These data suggest that CD34+ human umbilical cord blood progenitors are capable of trans-differentiation into vascular endothelial cell phenotype and assemble into 3D tissue structures. Culture of CBSC in simulated μg may be potentially beneficial in the fields of stem cell biology and somatic cell therapy.

Assessment of clinical significance of positive blood cultures of relatively low-virulence isolates.

PubMed

Hirakata, Y; Furuya, N; Iwata, M; Kashitani, F; Ishikawa, M; Yumoto, S; Yasui, K; Endoh, H; Marui, A; Kaku, M; Tateda, K; Yamaguchi, K

1996-03-01

In Omori Hospital, Toho University School of Medicine, relatively low-virulence blood isolates, including coagulase-negative staphylococci (CNS), enterococci and nonfermentative gram-negative rods other than Pseudomonas aeruginosa comprised c. 60% of total blood isolates. A retrospective study was conducted to assess their clinical significance by reviewing a total of 91 hospital charts. The physicians' assessments of these positive blood cultures as recorded in the charts were classified into four categories--sepsis, possible sepsis, contamination and no comment. The episodes classified as sepsis accounted for 5.0-19.6%. These episodes were also evaluated by a graded clinical significance score based on multiple factors, including number of positive cultures and clinical signs. The scores for the 91 episodes covered a wide range from 1 to 9, indicating that both contaminants and causative organisms may have been involved. The episodes judged as sepsis or possible sepsis tended to have higher scores. The scores for the episodes associated with enterococci were also higher than those involving CNS or non-fermentative gram-negative rods. The scores for episodes associated with intravenous hyperalimentation catheters were higher than those not associated with the catheters.

Multiplex Identification of Gram-Positive Bacteria and Resistance Determinants Directly from Positive Blood Culture Broths: Evaluation of an Automated Microarray-Based Nucleic Acid Test

PubMed Central

Buchan, Blake W.; Ginocchio, Christine C.; Manii, Ryhana; Cavagnolo, Robert; Pancholi, Preeti; Swyers, Lettie; Thomson, Richard B.; Anderson, Christopher; Kaul, Karen; Ledeboer, Nathan A.

2013-01-01

Background A multicenter study was conducted to evaluate the diagnostic accuracy (sensitivity and specificity) of the Verigene Gram-Positive Blood Culture Test (BC-GP) test to identify 12 Gram-positive bacterial gene targets and three genetic resistance determinants directly from positive blood culture broths containing Gram-positive bacteria. Methods and Findings 1,252 blood cultures containing Gram-positive bacteria were prospectively collected and tested at five clinical centers between April, 2011 and January, 2012. An additional 387 contrived blood cultures containing uncommon targets (e.g., Listeria spp., S. lugdunensis, vanB-positive Enterococci) were included to fully evaluate the performance of the BC-GP test. Sensitivity and specificity for the 12 specific genus or species targets identified by the BC-GP test ranged from 92.6%–100% and 95.4%–100%, respectively. Identification of the mecA gene in 599 cultures containing S. aureus or S. epidermidis was 98.6% sensitive and 94.3% specific compared to cefoxitin disk method. Identification of the vanA gene in 81 cultures containing Enterococcus faecium or E. faecalis was 100% sensitive and specific. Approximately 7.5% (87/1,157) of single-organism cultures contained Gram-positive bacteria not present on the BC-GP test panel. In 95 cultures containing multiple organisms the BC-GP test was in 71.6% (68/95) agreement with culture results. Retrospective analysis of 107 separate blood cultures demonstrated that identification of methicillin resistant S. aureus and vancomycin resistant Enterococcus spp. was completed an average of 41.8 to 42.4 h earlier using the BC-GP test compared to routine culture methods. The BC-GP test was unable to assign mecA to a specific organism in cultures containing more than one Staphylococcus isolate and does not identify common blood culture contaminants such as Micrococcus, Corynebacterium, and Bacillus. Conclusions The BC-GP test is a multiplex test capable of detecting most

Revisiting the Diego Blood Group System in Amerindians: Evidence for Gene-Culture Comigration.

PubMed

Bégat, Christophe; Bailly, Pascal; Chiaroni, Jacques; Mazières, Stéphane

2015-01-01

Six decades ago the DI*A allele of the Diego blood group system was instrumental in proving Native American populations originated from Siberia. Since then, it has received scant attention. The present study was undertaken to reappraise distribution of the DI*A allele in 144 Native American populations based on current knowledge. Using analysis of variance tests, frequency distribution was studied according to geographical, environmental, and cultural parameters. Frequencies were highest in Amazonian populations. In contrast, DI*A was undetectable in subarctic, Fuegian, Panamanian, Chaco and Yanomama populations. Closer study revealed a correlation that this unequal distribution was correlated with language, suggesting that linguistic divergence was a driving force in the expansion of DI*A among Native Americans. The absence of DI*A in circumpolar Eskimo-Aleut and Na-Dene speakers was consistent with a late migratory event confined to North America. Distribution of DI*A in subtropical areas indicated that gene and culture exchanges were more intense within than between ecozones. Bolstering the utility of classical genetic markers in biological anthropology, the present study of the expansion of Diego blood group genetic polymorphism in Native Americans shows strong evidence of gene-culture comigration.

[Infective endocarditis in intensive cardiac care unit - clinical and biochemical differences of blood-culture negative infective endocarditis].

PubMed

Kaziród-Wolski, Karol; Sielski, Janusz; Ciuraszkiewicz, Katarzyna

2017-01-23

Diagnosis and treatment of infective endocarditis (IE) is still a challenge for physicians. Group of patients with the worst prognosis is treated in Intensive Cardiac Care Unit (ICCU). Etiologic agent can not be identified in a substantial number of patients. The aim of study is to find differences between patients with blood culture negative infective endocarditis (BCNIE) and blood culture positive infective endocarditis (BCPIE) treated in ICCU by comparing their clinical course and laboratory parameters. Retrospective analysis of 30 patients with IE hospitalized in ICCU Swietokrzyskie Cardiac Centre between 2010 and 2016. This group consist of 26 men (86,67%) and 4 women (13,3%). Mean age was 58 years ±13. Most of the cases were new disease, recurrence of the disease was observed in 2 cases (6,7%). 8 patients (26,7%) required artificial ventilation, 11 (36,7%) received inotropes and 6 (20%) vasopresors. In 14 (46,7%) cases blood cultures was negative (BCNIE), the rest of patients (16, 53,3%) was blood cultures - positive infective endocarditis (BCIE). Both of the groups were clinically similar. There were no statistically significant differences in incidence of cardiac implants, localization of bacterial vegetations, administered catecholamines, antibiotic therapy, artificial ventilation, surgical treatment, complication and in-hospital mortality. Incidence of cardiac complications in all of BCNIE cases and in 81,3% cases of BCPIE draws attention, but it is not statistically significant difference (p=0,08). There was statistically significant difference in mean BNP blood concentration (3005,17 ng/ml ±2045,2 vs 1013,42 ng/ml ±1087,6; p=0,01), but there were no statistically significant differences in rest of laboratory parameters. BCNIE group has got higher mean BNP blood concentration than BCPIE group. There were no statistically significant differences between these groups in others laboratory parameters, clinical course and administered antibiotic therapy

Blood in ancient Jewish culture.

PubMed

Kottek, Samuel S

2005-01-01

The article analyzes the Jewish attitude towards blood, conceived both as the vehicle of life, and as a polluting product of feminine bodies. The author analyzes numerous Biblical sources concerning the 'unapproachable' blood of menstruation, the role of blood in the generation of the fetus, the blood as source of illness, the practice of bloodletting, and finally the idea that male menstruation exists as a peculiarity of the Jews.

Comparison of EBV DNA viral load in whole blood, plasma, B-cells and B-cell culture supernatant.

PubMed

Ouedraogo, David Eric; Bollore, Karine; Viljoen, Johannes; Foulongne, Vincent; Reynes, Jacques; Cartron, Guillaume; Vendrell, Jean-Pierre; Van de Perre, Philippe; Tuaillon, Edouard

2014-05-01

Epstein-Barr virus (EBV) genome quantitation in whole blood is used widely for therapeutic monitoring of EBV-associated disorders in immunosuppressed individuals and in patients with EBV-associated lymphoma. However, the most appropriate biological material to be used for EBV DNA quantitation remains a subject of debate. This study compare the detection rate and levels of EBV DNA from whole blood, plasma, enriched B-cells, and B-cell short-term culture supernatant using quantitative real-time PCR. Samples were collected from 33 subjects with either HIV infection or B-cell lymphoma. Overall, EBV DNA was detected in 100% of enriched B-cell samples, in 82% of B-cell culture supernatants, in 57% of plasma, and 42% of whole blood samples. A significant correlation for EBV viral load was found between enriched B-cell and B-cell culture supernatant material (ρ = 0.92; P < 0.0001), but no significant correlation existed between EBV DNA levels in whole blood and enriched B-cells (ρ = -0.02; P = 0.89), whole blood and plasma (ρ = 0.24; P = 0.24), or enriched B-cells and plasma (ρ = 0.08; P = 0.77). Testing of enriched B-cells appeared to be the most sensitive method for detection of EBV DNA as well as for exploration of the cellular reservoir. Quantitation of EBV DNA in plasma and B-cell culture supernatant may be of interest to assess EBV reactivation dynamics and response to treatment as well as to decipher EBV host-pathogen interactions in various clinical scenarios. © 2013 Wiley Periodicals, Inc.

Skin antiseptics in venous puncture site disinfection for preventing blood culture contamination: A Bayesian network meta-analysis of randomized controlled trials.

PubMed

Liu, Wenjie; Duan, Yuchen; Cui, Wenyao; Li, Li; Wang, Xia; Dai, Heling; You, Chao; Chen, Maojun

2016-07-01

To compare the efficacy of several antiseptics in decreasing the blood culture contamination rate. Network meta-analysis. Electronic searches of PubMed and Embase were conducted up to November 2015. Only randomized controlled trials or quasi-randomized controlled trials were eligible. We applied no language restriction. A comprehensive review of articles in the reference lists was also accomplished for possible relevant studies. Relevant studies evaluating efficacy of different antiseptics in venous puncture site for decreasing the blood culture contamination rate were included. The data were extracted from the included randomized controlled trials by two authors independently. The risk of bias was evaluated using Detsky scale by two authors independently. We used WinBUGS1.43 software and statistic model described by Chaimani to perform this network meta-analysis. Then graphs of statistical results of WinBUGS1.43 software were generated using 'networkplot', 'ifplot', 'netfunnel' and 'sucra' procedure by STATA13.0. Odds ratio and 95% confidence intervals were assessed for dichotomous data. A probability of p less than 0.05 was considered to be statistically significant. Compared with ordinary meta-analyses, this network meta-analysis offered hierarchies for the efficacy of different antiseptics in decreasing the blood culture contamination rate. Seven randomized controlled trials involving 34,408 blood samples were eligible for the meta-analysis. No significant difference was found in blood culture contamination rate among different antiseptics. No significant difference was found between non-alcoholic antiseptics and alcoholic antiseptics, alcoholic chlorhexidine and povidone iodine, chlorhexidine and iodine compounds, povidone iodine and iodine tincture in this aspect, respectively. Different antiseptics may not affect the blood culture contamination rate. Different intervals between the skin disinfection and the venous puncture, the different settings (emergency

Integration of DPC and clinical microbiological data in Japan reveals importance of confirming a negative follow-up blood culture in patients with MRSA bacteremia.

PubMed

Miyamoto, Naoki; Yahara, Koji; Horita, Rie; Yano, Tomomi; Tashiro, Naotaka; Morii, Daiichi; Tsutsui, Atsuko; Yaita, Kenichiro; Shibayama, Keigo; Watanabe, Hiroshi

2017-10-01

Methicillin-resistant Staphylococcus aureus (MRSA) bacteremia is one of the commonest and most life-threatening of all infectious diseases. The morbidity and mortality rates associated with MRSA bacteremia are higher than those associated with bacteremia caused by other pathogens. A common guideline in MRSA bacteremia treatment is to confirm bacteremia clearance through additional blood cultures 2-4 days after initial positive cultures and as needed thereafter. However, no study has presented statistical evidence of how and to what extent confirming a negative follow-up blood culture impacts clinical outcome. We present this evidence for the first time, by combining clinical microbiological data of blood cultures and the DPC administrative claims database; both had been systematically accumulated through routine medical care in hospitals. We used electronic medical records to investigate the clinical background and infection source in detail. By analyzing data from a university hospital, we revealed how survival curves change when a negative follow-up blood culture is confirmed. We also demonstrated confirmation of a negative culture is significantly associated with clinical outcomes: there was a more than three-fold increase in mortality risk (after adjusting for clinical background) if a negative blood culture was not confirmed within 14 days of the initial positive blood culture. Although we used data from only one university hospital, our novel approach and results will be a basis for future studies in several hospitals in Japan to provide statistical evidence of the clinical importance of confirming a negative follow-up blood culture in bacteremia patients, including those with MRSA infections. Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Time-to-positivity, type of culture media and oxidase test performed on positive blood culture vials to predict Pseudomonas aeruginosa in patients with Gram-negative bacilli bacteraemia.

PubMed

Cobos-Triguero, N; Zboromyrska, Y; Morata, L; Alejo, I; De La Calle, C; Vergara, A; Cardozo, C; Arcas, M P; Soriano, A; Marco, F; Mensa, J; Almela, M; Martínez, J A

2017-02-01

The aim of this study was to determine the usefulness of oxidase test and time-to-positivity (TTP) in aerobic and anaerobic blood culture vials to detect the presence of Pseudomonas aeruginosa in patients with Gram-negative bacilli (GNB) bacteraemia. TTP was recorded for each aerobic and anaerobic blood culture vial of monomicrobial bacteraemia due to GNB. Oxidase test was performed in a pellet of the centrifuged content of the positive blood culture. An algorithm was developed in order to perform the oxidase test efficiently taking into account TTP and type of vial. A total of 341 episodes of GNB bacteraemia were analysed. Sensitivity, specificity, positive predictive value and negative predictive value of the oxidase test performed on positive vials with GNB to predict P. aeruginosa were 95%, 99%, 91%, and 99%, respectively. When growth was first or exclusively detected in anaerobic vials, P. aeruginosa was never identified hence the performance of the oxidase test could be avoided. When growth was only or first detected in aerobic vials, a TTP≥8h predicted P. aeruginosa in 37% or cases (63 of 169), therefore oxidase test is highly recommended. Oxidase test performed onto positive blood culture vials previously selected by TTP and type of vials is an easy and inexpensive way to predict P. aeruginosa. In most cases, this can lead to optimization of treatment in less than 24 hours.

Highly Sensitive Bacteriophage-Based Detection of Brucella abortus in Mixed Culture and Spiked Blood

PubMed Central

Sergueev, Kirill V.; Filippov, Andrey A.; Nikolich, Mikeljon P.

2017-01-01

For decades, bacteriophages (phages) have been used for Brucella species identification in the diagnosis and epidemiology of brucellosis. Traditional Brucella phage typing is a multi-day procedure including the isolation of a pure culture, a step that can take up to three weeks. In this study, we focused on the use of brucellaphages for sensitive detection of the pathogen in clinical and other complex samples, and developed an indirect method of Brucella detection using real-time quantitative PCR monitoring of brucellaphage DNA amplification via replication on live Brucella cells. This assay allowed the detection of single bacteria (down to 1 colony-forming unit per milliliter) within 72 h without DNA extraction and purification steps. The technique was equally efficient with Brucella abortus pure culture and with mixed cultures of B. abortus and α-proteobacterial near neighbors that can be misidentified as Brucella spp., Ochrobactrum anthropi and Afipia felis. The addition of a simple short sample preparation step enabled the indirect phage-based detection of B. abortus in spiked blood, with the same high sensitivity. This indirect phage-based detection assay enables the rapid and sensitive detection of live B. abortus in mixed cultures and in blood samples, and can potentially be applied for detection in other clinical samples and other complex sample types. PMID:28604602

Rapid identification of bacteria from bioMérieux BacT/ALERT blood culture bottles by MALDI-TOF MS.

PubMed

Haigh, J D; Green, I M; Ball, D; Eydmann, M; Millar, M; Wilks, M

2013-01-01

Several studies have reported poor results when trying to identify microorganisms directly from the bioMérieux BacT/ALERT blood culture system using matrix-assisted laser desorption/ionisation-time of flight (MALDI-TOF) mass spectrometry. The aim of this study is to evaluate two new methods, Sepsityper and an enrichment method for direct identification of microorganisms from this system. For both methods the samples were processed using the Bruker Microflex LT mass spectrometer (Biotyper) using the Microflex Control software to obtain spectra. The results from direct analysis were compared with those obtained by subculture and subsequent identification. A total of 350 positive blood cultures were processed simultaneously by the two methods. Fifty-three cultures were polymocrobial or failed to grow any organism on subculture, and these results were not included as there was either no subculture result, or for polymicrobial cultures it was known that the Biotyper would not be able to distinguish the constituent organisms correctly. Overall, the results showed that, contrary to previous reports, it is possible to identify bacteria directly from bioMérieux blood culture bottles, as 219/297 (74%) correct identifications were obtained using the Bruker Sepsityper method and 228/297 (77%) were obtained for the enrichment method when there is only one organism was present. Although the enrichment method was simpler, the reagent costs for the Sepsityper method were approximately pound 4.00 per sample compared to pound 0.50. An even simpler and cheaper method, which was less labour-intensive and did not require further reagents, was investigated. Seventy-seven specimens from positive signalled blood cultures were analysed by inoculating prewarmed blood agar plates and analysing any growth after 1-, 2- and 4-h periods of incubation at 37 degrees C, by either direct transfer or alcohol extraction. This method gave the highest number of correct identifications, 66/77 (86

Revisiting the IFN-γ release assay: Whole blood or PBMC cultures? - And other factors of influence.

PubMed

Hartmann, Sofie Bruun; Emnéus, Jenny; Wolff, Anders; Jungersen, Gregers

2016-07-01

The interferon-γ release assay (IGRA) is a widely used test for the presence of a cell-mediated immune (CMI) response in vitro. This measure is used to test for infection with intracellular pathogens or for validating vaccine efficacy, and it is a widely used test for both human as well as cattle. However, there is no consensus whether to use whole blood cultures or purified PBMCs for the assay, and both cell populations are being used and results compared. Therefore the aim of this study was to compare different culture settings using immune cells from previously vaccinated calves, and to shed light on external factors that could influence the read out in terms of IFN-γ levels. It was found that optimal culture conditions varied between individual animals; when polyclonal activated, cells from whole blood cultures were most responsive, but when activated specifically, the optimal cell concentration/population varied with whole blood, 10×10(6)cells/ml PBMC and 5×10(6)cells/ml PBMC being the highest performing conditions. A further investigation of the distribution of cell populations in PBMCs compared to whole blood was conducted, and a significant (p<0.001) decrease in the percentage of CD3(+) T lymphocytes within the PBMCs was found. More specifically, this reduction was due to a significant (p<0.01) decrease in the percentage of γδ(+) T lymphocytes. Thus measuring immune responses on purified PBMCs might not give a physiologically relevant output. Additionally, it was tested if the choice of incubation plate would interfere with the level of secreted IFN-γ in whole blood cultures from five calves. Six plates (a-f) were tested and no significant difference in absolute levels of IFN-γ was detected in the six plates when cells were polyclonal and specifically activated. However, we observed a significant (p<0.05) higher background level in a flat-bottom plate from Corning® (cat# 3595) (plate d) compared to two different flat-bottom plates from Corning

Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes for Rapid Identification of Candida albicans Directly from Blood Culture Bottles

PubMed Central

Rigby, Susan; Procop, Gary W.; Haase, Gerhard; Wilson, Deborah; Hall, Geraldine; Kurtzman, Cletus; Oliveira, Kenneth; Von Oy, Sabina; Hyldig-Nielsen, Jens J.; Coull, James; Stender, Henrik

2002-01-01

A new fluorescence in situ hybridization (FISH) method that uses peptide nucleic acid (PNA) probes for identification of Candida albicans directly from positive-blood-culture bottles in which yeast was observed by Gram staining (herein referred to as yeast-positive blood culture bottles) is described. The test (the C. albicans PNA FISH method) is based on a fluorescein-labeled PNA probe that targets C. albicans 26S rRNA. The PNA probe is added to smears made directly from the contents of the blood culture bottle and hybridized for 90 min at 55°C. Unhybridized PNA probe is removed by washing of the mixture (30 min), and the smears are examined by fluorescence microscopy. The specificity of the method was confirmed with 23 reference strains representing phylogenetically related yeast species and 148 clinical isolates covering the clinically most significant yeast species, including C. albicans (n = 72), C. dubliniensis (n = 58), C. glabrata (n = 5), C. krusei (n = 2), C. parapsilosis (n = 4), and C. tropicalis (n = 3). The performance of the C. albicans PNA FISH method as a diagnostic test was evaluated with 33 routine and 25 simulated yeast-positive blood culture bottles and showed 100% sensitivity and 100% specificity. It is concluded that this 2.5-h method for the definitive identification of C. albicans directly from yeast-positive blood culture bottles provides important information for optimal antifungal therapy and patient management. PMID:12037084

Reducing time to identification of aerobic bacteria and fastidious micro-organisms in positive blood cultures.

PubMed

Intra, J; Sala, M R; Falbo, R; Cappellini, F; Brambilla, P

2016-12-01

Rapid and early identification of micro-organisms in blood has a key role in the diagnosis of a febrile patient, in particular, in guiding the clinician to define the correct antibiotic therapy. This study presents a simple and very fast method with high performances for identifying bacteria by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) after only 4 h of incubation. We used early bacterial growth on PolyViteX chocolate agar plates inoculated with five drops of blood-broth medium deposited in the same point and spread with a sterile loop, followed by a direct transfer procedure on MALDI-TOF MS target slides without additional modification. Ninety-nine percentage of aerobic bacteria were correctly identified from 600 monomicrobial-positive blood cultures. This procedure allowed obtaining the correct identification of fastidious pathogens, such as Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae that need complex nutritional and environmental requirements in order to grow. Compared to the traditional pathogen identification from blood cultures that takes over 24 h, the reliability of results, rapid performance and suitability of this protocol allowed a more rapid administration of optimal antimicrobial treatment in the patients. Bloodstream infections are serious conditions with a high mortality and morbidity rate. Rapid identification of pathogens and appropriate antimicrobial therapy have a key role for successful patient outcome. In this work, we developed a rapid, simplified, accurate, and efficient method, reaching 99 % identification of aerobic bacteria from monomicrobial-positive blood cultures by using early growth on enriched medium, direct transfer to target plate without additional procedures, matrix-assisted laser desorption ionization-time of flight mass spectrometry and SARAMIS database. The application of this protocol allows to anticipate appropriate antibiotic therapy. �

IDENTIFICATION OF BACTERIA IN BLOOD CULTURES FROM CLINICALLY ILL CAPTIVE ANTILLEAN MANATEES (TRICHECHUS MANATUS MANATUS).

PubMed

Silva, Mariana C O; Attademo, Fernanda F L; Freire, Augusto C B; Sousa, Glaucia P; Luna, Fábia O; Lima, Débora C V; Mota, Rinaldo A; Mendes, Emiko S; Silva, Jean C R

2017-03-01

Between September 2001 and March 2013, 62 bacterial cultures (37 aerobic and 25 anaerobic) were performed on 37 blood samples from 23 Antillean manatees ( Trichechus manatus manatus) that were kept in captivity at the Brazilian National Center for Research and Conservation of Aquatic Mammals (CMA) in Pernambuco (CMA-PE) and Alagoas (CMA-AL), Brazil. All of the animals sampled exhibited clinical signs at the time of sampling including abscesses (n = 8), debilitation and anorexia (n = 22), and profound lethargy-moribundity (n = 7). The 4 animals with profound lethargy-moribundity died shortly after sampling of unknown causes. Bacteria were isolated from 15/37 (40.5%) and aerobic blood cultures from 13/23 animals (56.5%). None of the anaerobic cultures were positive. Aeromonas caviae , Aeromonas hydrophila , Aeromonas sp., Escherichia coli , Leclercia adecarboxylata , Pantoea agglomerans , Pseudomonas aeruginosa , Pseudomonas stutzeri , Pseudomonas sp., Sphingomonas paucimobilis , coagulase-negative Staphylococcus, and Staphylococcus epidermidis were each found in only one animal; Staphylococcus spp. was found in two; and Vibrio fluvialis in four. Thirteen samples had only one bacteria isolated, one sample had two bacteria, and one sample had three bacteria isolated. Regarding sex, age group, and origin among the manatees examined, 54.5% (6/11) of the females, 58.3% (7/12) of the males, 40% (2/5) of the calves, 66.7% (8/12) of the juveniles, 50% (3/6) of the adults, 55.5% (10/18) at CMA-PE, and 60% (3/5) at CMA-AL were found to be positive for bacterial growth during at least one sampling time. All Antillean manatees were clinically ill. Regarding clinical signs, bacteria were found in 50% (11/22) of blood samples of the animals showing debilitation and anorexia, 1 of 8 (12.5%) of blood samples of the animals showing abscesses, and 3 of 7 (42.9%) of blood samples of the animals showing profound lethargy-moribundity.

The combined rapid detection and species-level identification of yeasts in simulated blood culture using a colorimetric sensor array.

PubMed

Shrestha, Nabin K; Lim, Sung H; Wilson, Deborah A; SalasVargas, Ana Victoria; Churi, Yair S; Rhodes, Paul A; Mazzone, Peter J; Procop, Gary W

2017-01-01

A colorimetric sensor array (CSA) has been demonstrated to rapidly detect and identify bacteria growing in blood cultures by obtaining a species-specific "fingerprint" of the volatile organic compounds (VOCs) produced during growth. This capability has been demonstrated in prokaryotes, but has not been reported for eukaryotic cells growing in culture. The purpose of this study was to explore if a disposable CSA could differentially identify 7 species of pathogenic yeasts growing in blood culture. Culture trials of whole blood inoculated with a panel of clinically important pathogenic yeasts at four different microorganism loads were performed. Cultures were done in both standard BacT/Alert and CSA-embedded bottles, after adding 10 mL of spiked blood to each bottle. Color changes in the CSA were captured as images by an optical scanner at defined time intervals. The captured images were analyzed to identify the yeast species. Time to detection by the CSA was compared to that in the BacT/Alert system. One hundred sixty-two yeast culture trials were performed, including strains of several species of Candida (Ca. albicans, Ca. glabrata, Ca. parapsilosis, and Ca. tropicalis), Clavispora (synonym Candida) lusitaniae, Pichia kudriavzevii (synonym Candida krusei) and Cryptococcus neoformans, at loads of 8.2 × 105, 8.3 × 103, 8.5 × 101, and 1.7 CFU/mL. In addition, 8 negative trials (no yeast) were conducted. All negative trials were correctly identified as negative, and all positive trials were detected. Colorimetric responses were species-specific and did not vary by inoculum load over the 500000-fold range of loads tested, allowing for accurate species-level identification. The mean sensitivity for species-level identification by CSA was 74% at detection, and increased with time, reaching almost 95% at 4 hours after detection. At an inoculum load of 1.7 CFU/mL, mean time to detection with the CSA was 6.8 hours (17%) less than with the BacT/Alert platform. The CSA

Value of blood culture time to positivity in identifying complicated nontyphoidal Salmonella bacteremia.

PubMed

Chen, Shang-Yu; Weng, Tzu-Hua; Tseng, Wen-Pin; Fu, Chia-Ming; Lin, Hui-Wen; Liao, Chun-Hsing; Lee, Tai-Fen; Hsueh, Po-Ren; Fang, Cheng-Chung; Chen, Shey-Ying

2018-02-13

Few studies analyzed the association between blood culture time to positivity (TTP) and risk of complicated nontyphoidal Salmonella (NTS) bacteremia. We conducted a retrospective study of 206 patients (aged 60.4 ± 17.4 years) with NTS bacteremia during a 30-month period. Complicated NTS bacteremia was defined as the presence of 30-day mortality, complicated infection requiring surgery or abscess drainage, or requirement of intensive care unit admission. Serogroup D (75.7%) was the predominant isolates. Malignancy (44.7%) was the most prevalent comorbidity. Patients with rapid TTP (<10 h) were more likely to have thrombocytopenia, septic shock, persistent bacteremia, complicated infection, and a higher intensive care unit admission rate. In multivariate logistic regression model, a TTP <10 h was an independent predictor for complicated NTS bacteremia (adjusted odd ratio, 5.683, 95% confidence interval, 2.396-13.482). Our study showed that blood culture TTP provides important diagnostic and prognostic information in the treatment of NTS bacteremia patients. Copyright © 2018. Published by Elsevier Inc.

Comparative efficacies of amoxicillin, clindamycin, and moxifloxacin in prevention of bacteremia following dental extractions.

PubMed

Diz Dios, P; Tomás Carmona, I; Limeres Posse, J; Medina Henríquez, J; Fernández Feijoo, J; Alvarez Fernández, M

2006-09-01

We evaluated the efficacies of oral prophylactic treatment with amoxicillin (AMX), clindamycin (CLI), and moxifloxacin (MXF) in the prevention of bacteremia following dental extractions (BDE). Two hundred twenty-one adults who required dental extractions under general anesthesia were randomly assigned to a control group, an AMX group, a CLI group, and an MXF group (the individuals in the drug treatment groups received 2 g, 600 mg, and 400 mg, respectively, 1 to 2 h before anesthesia induction). Venous blood samples were collected from each patient at the baseline and 30 s, 15 min, and 1 h after the dental extractions. The samples were inoculated into BACTEC Plus aerobic and anaerobic blood culture bottles and were processed in a BACTEC 9240 instrument. Subculture and the further identification of the isolated bacteria were performed by conventional microbiological techniques. The prevalences of BDE in the control group, AMX group, CLI group, and MXF group were 96, 46, 85, and 57%, respectively, at 30 s; 64, 11, 70, and 24%, respectively, at 15 min; and 20, 4, 22, and 7%, respectively, at 1 h. Streptococcus spp. were the most frequently identified bacteria in all groups (44 to 68%), with the lowest percentage being detected in the AMX group (44%). AMX and MXF prophylaxis showed high efficacies in reducing the prevalence and duration of BDE, but CLI prophylaxis was noneffective. As a consequence, MXF prophylaxis is a promising antibiotic alternative for the prevention of BDE when beta-lactams are not indicated.

Microbial identification and automated antibiotic susceptibility testing directly from positive blood cultures using MALDI-TOF MS and VITEK 2.

PubMed

Wattal, C; Oberoi, J K

2016-01-01

The study addresses the utility of Matrix Assisted Laser Desorption/Ionisation Time-Of-Flight mass spectrometry (MALDI-TOF MS) using VITEK MS and the VITEK 2 antimicrobial susceptibility testing (AST) system for direct identification (ID) and timely AST from positive blood culture bottles using a lysis-filtration method (LFM). Between July and December 2014, a total of 140 non-duplicate mono-microbial blood cultures were processed. An aliquot of positive blood culture broth was incubated with lysis buffer before the bacteria were filtered and washed. Micro-organisms recovered from the filter were first identified using VITEK MS and its suspension was used for direct AST by VITEK 2 once the ID was known. Direct ID and AST results were compared with classical methods using solid growth. Out of the 140 bottles tested, VITEK MS resulted in 70.7 % correct identification to the genus and/ or species level. For the 103 bottles where identification was possible, there was agreement in 97 samples (94.17 %) with classical culture. Compared to the routine method, the direct AST resulted in category agreement in 860 (96.5 %) of 891 bacteria-antimicrobial agent combinations tested. The results of direct ID and AST were available 16.1 hours before those of the standard approach on average. The combined use of VITEK MS and VITEK 2 directly on samples from positive blood culture bottles using a LFM technique can result in rapid and reliable ID and AST results in blood stream infections to result in early institution of targeted treatment. The combination of LFM and AST using VITEK 2 was found to expedite AST more reliably.

Blood culture-negative endocarditis: Improving the diagnostic yield using new diagnostic tools.

PubMed

Fournier, Pierre-Edouard; Gouriet, Frédérique; Casalta, Jean-Paul; Lepidi, Hubert; Chaudet, Hervé; Thuny, Franck; Collart, Frédéric; Habib, Gilbert; Raoult, Didier

2017-11-01

Blood culture-negative endocarditis (BCNE) may represent up to 70% of all endocarditis cases, depending on series. From 2001 to 2009, we implemented in our laboratory a multimodal diagnostic strategy for BCNE that included systematized testing of blood, and when available, valvular biopsy specimens using serological, broad range molecular, and histopathological assays. A causative microorganism was identified in 62.7% of patients.In this study from January 2010 to December 2015, in an effort to increase the number of identified causative microorganisms, we prospectively added to our diagnostic protocol specific real-time (RT) polymerase chain reaction (PCR) assays targeting various endocarditis agents, and applied them to all patients with BCNE admitted to the 4 public hospitals in Marseille, France.A total of 283 patients with BCNE were included in the study. Of these, 177 were classified as having definite endocarditis. Using our new multimodal diagnostic strategy, we identified an etiology in 138 patients (78.0% of cases). Of these, 3 were not infective (2.2%) and 1 was diagnosed as having Mycobacterium bovis BCG endocarditis. By adding specific PCR assays from blood and valvular biopsies, which exhibited a significantly greater sensitivity (P < 10) than other methods, causative agents, mostly enterococci, streptococci, and zoonotic microorganisms, were identified in an additional 27 patients (14 from valves only, 11 from blood only, and 2 from both). Finally, in another 107 patients, a pathogen was detected using serology in 37, valve culture in 8, broad spectrum PCR from valvular biopsies and blood in 19 and 2, respectively, immunohistochemistry from valves in 3, and a combination of several assays in 38.By adding specific RT-PCR assays to our systematic PCR testing of patients with BCNE, we increased the diagnostic efficiency by 24.3%, mostly by detecting enterococci and streptococci that had not been detected by other diagnostic methods, but also agents

Prevalence of multi-drug resistant organisms in stool of paediatric patients with acute leukaemia and correlation with blood culture positivity: A single institution experience.

PubMed

Shankar, Krupa; Radhakrishnan, Venkatraman; Vijayakumar, Varalakskmi; Ramamoorthy, Jaikumar; Ganesan, Prasanth; Dhanushkodi, Manikandan; Ganesan, T S; Sagar, T G

2018-01-01

Multi-drug resistant (MDR) bacteria are associated with increased morbidity and mortality in children with acute leukaemia. The present study was conducted to assess the prevalence of MDR bacteria in stool cultures of patients with acute leukaemia at presentation to the hospital. The results were then correlated with blood cultures when patients developed septicaemia. The study involved analysis of case records of patients with newly diagnosed acute leukaemia less than 18 years of age treated at our centre from January 2015 to December 2015. Stool cultures were sent within 72 hr of hospital admission and blood cultures were sent when clinically indicated. MDR was defined as resistance to at least one antibiotic in three or more following antimicrobial groups: cephalosporins, β-lactam/β-lactamase inhibitor, carbapenems, fluoroquinolones and aminoglycosides. The analysis included 85 patients with acute leukaemia, among whom 48 of 85 (56%) patients had positive stool cultures and 42 of 85 (50%) patients were positive for MDR bacteria. Blood cultures were positive in 13 of 48 patients (27%, seven MDR and six non-MDR) with positive stool cultures and three of 37 patients (8%, one MDR and two non-MDR) with negative stool cultures (P = 0.01). The concordance between stool and blood culture for similar organism was 61%. There were seven deaths in 48 stool culture positive patients and two deaths in 37 stool culture negative patients. This study shows the high prevalence of MDR bacteria in newly diagnosed children with acute leukaemia. Colonisation with MDR bacteria in stools is associated with increased positivity of blood cultures and mortality. © 2017 Wiley Periodicals, Inc.

Preparation of positive blood cultures for direct MALDI-ToF MS identification.

PubMed

Robinson, Andrew M; Ussher, James E

2016-08-01

MALDI-ToF MS can be used to identify microorganisms directly from blood cultures. This study compared two methods of sample preparation. Similar levels of genus- (91% vs 90%) and species-level identifications (79% vs 74%) were obtained with differential centrifugation and SDS methods. The SDS method is faster and requires minimal handling. Copyright © 2016 Elsevier B.V. All rights reserved.

Rapid Identification of Staphylococcus aureus Directly from Blood Cultures by Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes

PubMed Central

Oliveira, Kenneth; Procop, Gary W.; Wilson, Deborah; Coull, James; Stender, Henrik

2002-01-01

A new fluorescence in situ hybridization (FISH) method with peptide nucleic acid (PNA) probes for identification of Staphylococcus aureus directly from positive blood culture bottles that contain gram-positive cocci in clusters (GPCC) is described. The test (the S. aureus PNA FISH assay) is based on a fluorescein-labeled PNA probe that targets a species-specific sequence of the 16S rRNA of S. aureus. Evaluations with 17 reference strains and 48 clinical isolates, including methicillin-resistant and methicillin-susceptible S. aureus species, coagulase-negative Staphylococcus species, and other clinically relevant and phylogenetically related bacteria and yeast species, showed that the assay had 100% sensitivity and 96% specificity. Clinical trials with 87 blood cultures positive for GPCC correctly identified 36 of 37 (97%) of the S. aureus-positive cultures identified by standard microbiological methods. The positive and negative predictive values were 100 and 98%, respectively. It is concluded that this rapid method (2.5 h) for identification of S. aureus directly from blood culture bottles that contain GPCC offers important information for optimal antibiotic therapy. PMID:11773123

Multiplex Real-Time PCR Assay for Rapid Detection of Methicillin-Resistant Staphylococci Directly from Positive Blood Cultures

PubMed Central

Wang, Hye-young; Kim, Sunghyun; Kim, Jungho; Park, Soon-Deok

2014-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) is the most prevalent cause of bloodstream infections (BSIs) and is recognized as a major nosocomial pathogen. This study aimed to evaluate a newly designed multiplex real-time PCR assay capable of the simultaneous detection of mecA, S. aureus, and coagulase-negative staphylococci (CoNS) in blood culture specimens. The Real-MRSA and Real-MRCoNS multiplex real-time PCR assays (M&D, Republic of Korea) use the TaqMan probes 16S rRNA for Staphylococcus spp., the nuc gene for S. aureus, and the mecA gene for methicillin resistance. The detection limit of the multiplex real-time PCR assay was 103 CFU/ml per PCR for each gene target. The multiplex real-time PCR assay was evaluated using 118 clinical isolates from various specimen types and a total of 350 positive blood cultures from a continuous monitoring blood culture system. The results obtained with the multiplex real-time PCR assay for the three targets were in agreement with those of conventional identification and susceptibility testing methods except for one organism. Of 350 positive bottle cultures, the sensitivities of the multiplex real-time PCR kit were 100% (166/166 cultures), 97.2% (35/36 cultures), and 99.2% (117/118 cultures) for the 16S rRNA, nuc, and mecA genes, respectively, and the specificities for all three targets were 100%. The Real-MRSA and Real-MRCoNS multiplex real-time PCR assays are very useful for the rapid accurate diagnosis of staphylococcal BSIs. In addition, the Real-MRSA and Real-MRCoNS multiplex real-time PCR assays could have an important impact on the choice of appropriate antimicrobial therapy, based on detection of the mecA gene. PMID:24648566

A new rapid method for direct antimicrobial susceptibility testing of bacteria from positive blood cultures.

PubMed

Barnini, Simona; Brucculeri, Veronica; Morici, Paola; Ghelardi, Emilia; Florio, Walter; Lupetti, Antonella

2016-08-12

Rapid identification and antimicrobial susceptibility testing (AST) of the causative agent(s) of bloodstream infections can lead to prompt appropriate antimicrobial therapy. To shorten species identification, in this study bacteria were recovered from monomicrobial blood cultures by serum separator tubes and spotted onto the target plate for direct MALDI-TOF MS identification. Proper antibiotics were selected for direct AST based on species identification. In order to obtain rapid AST results, bacteria were recovered from positive blood cultures by two different protocols: by serum separator tubes (further referred to as PR1), or after a short-term subculture in liquid medium (further referred to as PR2). The results were compared with those obtained by the method currently used in our laboratory consisting in identification by MALDI-TOF and AST by Vitek 2 or Sensititre on isolated colonies. The direct MALDI-TOF method concordantly identified with the current method 97.5 % of the Gram-negative bacteria and 96.1 % of the Gram-positive cocci contained in monomicrobial blood cultures. The direct AST by PR1 and PR2 for all isolate/antimicrobial agent combinations was concordant/correct with the current method for 87.8 and 90.5 % of Gram-negative bacteria and for 93.1 and 93.8 % of Gram-positive cocci, respectively. In particular, 100 % categorical agreement was found with levofloxacin for Enterobacteriaceae by both PR1 and PR2, and 99.0 and 100 % categorical agreement was observed with linezolid for Gram-positive cocci by PR1 and PR2, respectively. There was no significant difference in accuracy between PR1 and PR2 for Gram-negative bacteria and Gram-positive cocci. This newly described method seems promising for providing accurate AST results. Most importantly, these results would be available in a few hours from blood culture positivity, which would help clinicians to promptly confirm or streamline an effective antibiotic therapy in patients with bloodstream

Intravenous amoxicillin/clavulanate for the prevention of bacteraemia following dental procedures: a randomized clinical trial.

PubMed

Limeres Posse, J; Álvarez Fernández, M; Fernández Feijoo, J; Medina Henríquez, J; Lockhart, P B; Chu, V H; Diz Dios, P

2016-07-01

Although controversy exists regarding the efficacy of antibiotic prophylaxis for patients at risk of infective endocarditis, expert committees continue to publish recommendations for antibiotic prophylaxis regimens. This study aimed to evaluate the efficacy of four antimicrobial regimens for the prevention of bacteraemia following dental extractions. The study population included 266 adults requiring dental extractions who were randomly assigned to the following five groups: control (no prophylaxis); 1000/200 mg of amoxicillin/clavulanate intravenously; 2 g of amoxicillin by mouth; 600 mg of clindamycin by mouth; and 600 mg of azithromycin by mouth. Venous blood samples were collected from each patient at baseline and at 30 s, 15 min and 1 h after dental extractions. Samples were inoculated into BACTEC Plus culture bottles and processed in the BACTEC 9240. Conventional microbiological techniques were used for subcultures and further identification of the isolated bacteria. The trial was registered at ClinicalTrials.gov with ID number NCT02115776. The incidence of bacteraemia in the control, amoxicillin/clavulanate, amoxicillin, clindamycin and azithromycin groups was: 96%, 0%, 50%, 87% and 81%, respectively, at 30 s; 65%, 0%, 10%, 65% and 49% at 15 min; and 18%, 0%, 4%, 19% and 18% at 1 h. Streptococci were the most frequently identified bacteria. The percentage of positive blood cultures at 30 s post-extraction was lower in the amoxicillin/clavulanate group than in the amoxicillin group (P < 0.001). The incidence of bacteraemia in the clindamycin group was similar to that in the control group. Bacteraemia following dental extractions was undetectable with amoxicillin/clavulanate prophylaxis. Alternative antimicrobial regimens should be sought for patients allergic to the β-lactams. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e

Evaluation of a direct blood culture disk diffusion antimicrobial susceptibility test.

PubMed Central

Doern, G V; Scott, D R; Rashad, A L; Kim, K S

1981-01-01

A total of 556 unique blood culture isolates of nonfastidious aerobic and facultatively anaerobic bacteria were examined by direct and standardized disk susceptibility test methods (4,234 antibiotic-organism comparisons). When discrepancies which could be accounted for by the variability inherent in disk diffusion susceptibility tests were excluded, the direct method demonstrated 96.8% overall agreement with the standardized method. A total of 1.6% minor, 1.5% major, and 0.1% very major discrepancies were noted. PMID:7325634

The combined rapid detection and species-level identification of yeasts in simulated blood culture using a colorimetric sensor array

PubMed Central

Lim, Sung H.; Wilson, Deborah A.; SalasVargas, Ana Victoria; Churi, Yair S.; Rhodes, Paul A.; Mazzone, Peter J.; Procop, Gary W.

2017-01-01

Background A colorimetric sensor array (CSA) has been demonstrated to rapidly detect and identify bacteria growing in blood cultures by obtaining a species-specific “fingerprint” of the volatile organic compounds (VOCs) produced during growth. This capability has been demonstrated in prokaryotes, but has not been reported for eukaryotic cells growing in culture. The purpose of this study was to explore if a disposable CSA could differentially identify 7 species of pathogenic yeasts growing in blood culture. Methods Culture trials of whole blood inoculated with a panel of clinically important pathogenic yeasts at four different microorganism loads were performed. Cultures were done in both standard BacT/Alert and CSA-embedded bottles, after adding 10 mL of spiked blood to each bottle. Color changes in the CSA were captured as images by an optical scanner at defined time intervals. The captured images were analyzed to identify the yeast species. Time to detection by the CSA was compared to that in the BacT/Alert system. Results One hundred sixty-two yeast culture trials were performed, including strains of several species of Candida (Ca. albicans, Ca. glabrata, Ca. parapsilosis, and Ca. tropicalis), Clavispora (synonym Candida) lusitaniae, Pichia kudriavzevii (synonym Candida krusei) and Cryptococcus neoformans, at loads of 8.2 × 105, 8.3 × 103, 8.5 × 101, and 1.7 CFU/mL. In addition, 8 negative trials (no yeast) were conducted. All negative trials were correctly identified as negative, and all positive trials were detected. Colorimetric responses were species-specific and did not vary by inoculum load over the 500000-fold range of loads tested, allowing for accurate species-level identification. The mean sensitivity for species-level identification by CSA was 74% at detection, and increased with time, reaching almost 95% at 4 hours after detection. At an inoculum load of 1.7 CFU/mL, mean time to detection with the CSA was 6.8 hours (17%) less than with the

Rapid Identification of Bacteria in Positive Blood Culture Broths by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry▿

PubMed Central

Stevenson, Lindsay G.; Drake, Steven K.; Murray, Patrick R.

2010-01-01

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry is a rapid, accurate method for identifying bacteria and fungi recovered on agar culture media. We report herein a method for the direct identification of bacteria in positive blood culture broths by MALDI-TOF mass spectrometry. A total of 212 positive cultures were examined, representing 32 genera and 60 species or groups. The identification of bacterial isolates by MALDI-TOF mass spectrometry was compared with biochemical testing, and discrepancies were resolved by gene sequencing. No identification (spectral score of <1.7) was obtained for 42 (19.8%) of the isolates, due most commonly to insufficient numbers of bacteria in the blood culture broth. Of the bacteria with a spectral score of ≥1.7, 162 (95.3%) of 170 isolates were correctly identified. All 8 isolates of Streptococcus mitis were misidentified as being Streptococcus pneumoniae isolates. This method provides a rapid, accurate, definitive identification of bacteria within 1 h of detection in positive blood cultures with the caveat that the identification of S. pneumoniae would have to be confirmed by an alternative test. PMID:19955282

Rapid identification of bacteria in positive blood culture broths by matrix-assisted laser desorption ionization-time of flight mass spectrometry.

PubMed

Stevenson, Lindsay G; Drake, Steven K; Murray, Patrick R

2010-02-01

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry is a rapid, accurate method for identifying bacteria and fungi recovered on agar culture media. We report herein a method for the direct identification of bacteria in positive blood culture broths by MALDI-TOF mass spectrometry. A total of 212 positive cultures were examined, representing 32 genera and 60 species or groups. The identification of bacterial isolates by MALDI-TOF mass spectrometry was compared with biochemical testing, and discrepancies were resolved by gene sequencing. No identification (spectral score of < 1.7) was obtained for 42 (19.8%) of the isolates, due most commonly to insufficient numbers of bacteria in the blood culture broth. Of the bacteria with a spectral score of > or = 1.7, 162 (95.3%) of 170 isolates were correctly identified. All 8 isolates of Streptococcus mitis were misidentified as being Streptococcus pneumoniae isolates. This method provides a rapid, accurate, definitive identification of bacteria within 1 h of detection in positive blood cultures with the caveat that the identification of S. pneumoniae would have to be confirmed by an alternative test.

Use of an automated blood culture system (BD BACTEC™) for diagnosis of prosthetic joint infections: easy and fast.

PubMed

Minassian, Angela M; Newnham, Robert; Kalimeris, Elizabeth; Bejon, Philip; Atkins, Bridget L; Bowler, Ian C J W

2014-05-04

For the diagnosis of prosthetic joint infection (PJI) automated BACTEC™ blood culture bottle methods have comparable sensitivity, specificity and a shorter time to positivity than traditional cooked meat enrichment broth methods. We evaluate the culture incubation period required to maximise sensitivity and specificity of microbiological diagnosis, and the ability of BACTEC™ to detect slow growing Propionibacteria spp. Multiple periprosthetic tissue samples taken by a standardised method from 332 patients undergoing prosthetic joint revision arthroplasty were cultured for 14 days, using a BD BACTEC™ instrumented blood culture system, in a prospective study from 1st January to 31st August 2012. The "gold standard" definition for PJI was the presence of at least one histological criterion, the presence of a sinus tract or purulence around the device. Cases where > =2 samples yielded indistinguishable isolates were considered culture-positive. 1000 BACTEC™ bottle cultures which were negative after 14 days incubation were sub-cultured for Propionibacteria spp. 79 patients fulfilled the definition for PJI, and 66 of these were culture-positive. All but 1 of these 66 culture-positive cases of PJI were detected within 3 days of incubation. Only one additional (clinically-insignificant) Propionibacterium spp. was identified on terminal subculture of 1000 bottles. Prolonged microbiological culture for 2 weeks is unnecessary when using BACTEC™ culture methods. The majority of clinically significant organisms grow within 3 days, and Propionibacteria spp. are identified without the need for terminal subculture. These findings should facilitate earlier decisions on final antimicrobial prescribing.

Bactec™ blood culture bottles allied to MALDI-TOF mass spectrometry: rapid etiologic diagnosis of bacterial endophthalmitis.

PubMed

Tanaka, Tatiana; Oliveira, Luiza Manhezi de Freitas; Ferreira, Bruno Fortaleza de Aquino; Kato, Juliana Mika; Rossi, Flavia; Correa, Karoline de Lemes Giuntini; Pimentel, Sergio Luis Gianotti; Yamamoto, Joyce Hisae; Almeida Junior, João Nóbrega

2017-07-01

Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) has been used for direct identification of pathogens from blood-inoculated blood culture bottles (BCBs). We showed that MALDI-TOF MS is an useful technique for rapid identification of the causative agents of endophthalmitis from vitreous humor-inoculated BCBs with a simple protocol. Copyright © 2017 Elsevier Inc. All rights reserved.

Mortality following blood culture in premature infants: increased with Gram-negative bacteremia and candidemia, but not Gram-positive bacteremia.

PubMed

Benjamin, Daniel K; DeLong, Elizabeth; Cotten, Charles M; Garges, Harmony P; Steinbach, William J; Clark, Reese H

2004-03-01

To describe survival following nosocomial bloodstream infections and quantify excess mortality associated with positive blood culture. Multicenter cohort study of premature infants. First blood culture was negative for 4648/5497 (78%) of the neonates--390/4648 (8%) died prior to discharge. Mortality prior to discharge was 19% in the 161 infants with Gram-negative rod (GNR) bacteremia, 8% in the 854 neonates with coagulase negative staphylococcus (CONS), 6% in the 169 infants infected with other Gram-positive bacteria (GP-o), and 26% in the 115 neonates with candidemia. The excess 7-day mortality was 0% for Gram-positive organisms and 83% for GNR bacteremia and candidemia. Using negative blood culture as referent, GNR [hazard ratio (HR)=2.61] and candidemia (HR=2.27) were associated with increased mortality; CONS (HR=1.08) and GP-o (HR=0.97) were not. Nosocomial GNR bacteremia and candidemia were associated with increased mortality but Gram-positive bacteremia was not.

Multicenter Clinical Evaluation of BacT/Alert Virtuo Blood Culture System.

PubMed

Jacobs, Michael R; Mazzulli, Tony; Hazen, Kevin C; Good, Caryn E; Abdelhamed, Ayman M; Lo, Pauline; Shum, Bianche; Roman, Katharine P; Robinson, Danielle C

2017-08-01

BacT/Alert Virtuo is an advanced, automated blood culture system incorporating improved automation and an enhanced detection algorithm to shorten time to detection. A multicenter study of the investigational Virtuo system (bioMérieux, Inc., Durham, NC) compared to BacT/Alert 3D (BTA3D) for detection of bacteremia/fungemia in four bottle types, SA and FA Plus (aerobic) and SN and FN Plus (anaerobic), was performed in a clinical setting with patient samples in a matched system design clinical trial. Blood was added to paired aerobic or anaerobic bottles, with the volume in each bottle in each pair required to be ≤10 ml and with the volumes required to be within 30% of each other. Of 5,709 bottle sets (52.5% aerobic pairs and 47.5% anaerobic pairs), 430 (7.5%) were positive for bacterial or fungal growth, with 342 (6.0%) clinically significant and 83 (1.5%) contaminated. A total of 3,539 sets (62.0%) were volume compliant, with 203 sets (5.7%) clinically significant. The positivity rates for volume-compliant bottle pairs determined by the two systems were comparable, with 68.7% of clinically significant isolates detected by both instruments, 15.7% by Virtuo only, and 15.7% by BTA3D only. Virtuo detected microbial growth nearly 2 h sooner overall than BTA3D (mean, 15.9 h versus 17.7 h). Shorter time to detection by Virtuo was related to organism group, with the time to detection being significantly shorter for enteric Gram-negative bacilli and enterococci (means, 3.6 h and 2.3 h shorter, respectively). This large clinical study demonstrated that the Virtuo blood culture system produced results comparable to those seen with the long-established BTA3D system, with significantly shorter time to detection. Copyright © 2017 Jacobs et al.

Sequential production of leukaemia inhibitory factor by blood cell culture in patients with ARDS.

PubMed

Gruson, D; Hilbert, G; Juzan, M; Taupin, J L; Coulon, V; Moreau, J F; Gualde, N; Gbikpi-Benissan, G

1998-04-01

Leukaemia inhibitory factor (LIF) is a polyfunctional cytokine integrated in cytokine networks and its concentration has been shown to be elevated in bronchoalveolar lavage fluid of patients with the acute respiratory distress syndrome (ARDS). The aim of our study was to evaluate the production of LIF by culturing blood cells from patients with ARDS. 8 patients with ARDS, 8 patients with pneumonia and 5 healthy subjects. The blood samples were taken on day 1 after onset of ARDS. LIF was measured, in the cell-free supernatant, with an enzyme-linked immunosorbent assay after 24 h, 48 h and 72 h of blood cell culture. LIF was detectable in some patients in the ARDS group: at i) at 24 h and 48 h: in 2 patients ii) at 72 h in 4/5 patients (140 +/- 231 pg/ml). Only in the 4 patients in whom LIF was measured at 72 h was ARDS associated with the multiple organ dysfunction syndrome. Furthermore, among the 5 patients with ARDS who subsequently died, 4 had a detectable LIF. We have observed that LIF was produced only in ARDS, but not in all patients. The production of LIF seems to be a good indicator of the severity of ARDS. These preliminary results must be confirmed by a larger study.

Loss of the ability to generate large burst-forming unit-like megakaryocytic colonies from thawed cord blood in semisolid cultures after short term suspension culture.

PubMed

Eskola, M; Bäckman, S; Möttönen, S; Kekomäki, R

2015-04-01

Total colony-forming cells from thawed cord blood units (CBUs) include megakaryocytic colony-forming units (CFU-Mks), which survive the freezing process. The aim of this study was to evaluate whether different megakaryocytic progenitors from unseparated CBUs survive the freezing process and a short-term liquid culture. Thawed samples of CBUs were cultured in liquid medium. During the cultures, serial samples were drawn to assess the growth of different megakaryocytic progenitors in a semisolid collagen medium with identical cytokines as in the liquid medium. Megakaryocytic cells were detected using immunohistochemistry and flow cytometry. In suspension culture, the megakaryocytic progenitors almost completely lost the ability to generate large (burst-forming unit-like, BFU-like) megakaryocytic colonies in semisolid cultures (large colonies, median count per chamber d0: 7.25 vs. d7: 1.5; P < 0.0001), whereas the number of small colonies (median count per chamber d0: 7.25 vs. d7: 16.0; P = 0.0505) peaked at day seven. Further 7-day culture in suspension resulted in the decline of small colonies as well (d7: 16.0 vs. d14: 5.75; P = 0.0088). Total CFU-Mk count declined from 23.3 (range 12.5-34.0) at d0 to 7.25 (range 1.0-13.5) at d14 (P < 0.0001). Immediately post-thaw, CBUs possess an ability to generate large BFU-like megakaryocytic colonies, whereas the colonies were not detectable in most CBUs in semisolid culture after a short suspension culture. Small CFU-Mks were observed throughout the cultures. It may be that the BFU-Mk colonies matured and acquired CFU-Mk behaviour. © 2014 International Society of Blood Transfusion.

In vitro evaluation of BacT/Alert FA blood culture bottles and T2Candida assay for the detection of Candida in the presence of antifungals.

PubMed

Beyda, Nicholas D; Amadio, Jonathan; Rodriguez, Jose R; Malinowski, Karen; Garey, Kevin W; Wanger, Audrey; Ostrosky-Zeichner, Luis

2018-06-13

The T2Candida assay is a novel, non-culture based assay for the diagnosis of candidemia directly from whole blood. The impact of antifungals on the performance of the T2Candida assay and blood culture bottles have not been well described. In this study, the performance of the T2Candida assay was compared to that of blood culture in detecting Candida spp. in spiked blood cultures with or without the presence of antifungals. Clinical bloodstream isolates of Candida spp. were inoculated into human whole blood at low (1 - 5 cells/mL) and high (10-50 cells/mL) concentrations with or without presence of caspofungin and fluconazole. Time to detection (TTD) was assessed for prepared samples using BacTAlert FA aerobic blood culture bottles or the T2Candida assay. In the absence of antifungals, T2Candida assay sensitivity was comparable to that of blood culture at both the low and high inoculum (95% vs. 97.5% and 100% vs. 100%, respectively) and had an average TTD that was significantly faster (5.1 hrs vs 27.2 - 30 hrs, respectively). Neither caspofungin nor fluconazole was observed to impact the sensitivity or TTD of the T2Candida assay, while fluconazole reduced overall blood culture sensitivity by 7.5% - 12.5% (at low and high inoculum, respectively) and significantly prolonged the TTD of C. albicans, C. tropicalis , and C. parapsilosis by 14.8 - 67 hrs. Neither caspofungin nor fluconazole impacted the performance of the T2Candida assay in-vitro and may be useful for the diagnosis of candidemia in patients receiving antifungal therapy. Copyright © 2018 American Society for Microbiology.

[Assessment of two DNA extraction methods to amplify the pneumolysin gene (PLY) from blood culture samples of Streptococcus pneumoniae].

PubMed

Hernández, Carolina; Durán, Claudia; Ulloa, María Teresa; Prado, Valeria

2004-05-01

Streptococcus pneumoniae is a common etiologic agent of invasive respiratory infections among children under 5 years of age and older adults. Isolation rates of S. pneumoniae by traditional culture techniques are low. To study the sensitivity and specificity of two different DNA extraction methods to amplify the ply gene, applied to three different types of blood culture broths, experimentally inoculated with S. pneumoniae. DNA was extracted from the cultures using an organic method or a technique that consists in dilution, washing with NaOH and concentration of the sample. This was followed by PCR amplification of a 355 pb fragment of the pneumolysin gene (ply). The organic DNA extraction method inhibited the PCR reaction at all concentrations studied (0.6 to 10(6) colony forming units/mL). Using the NaOH extraction, ply gene amplification was positive in all three blood culture broths, but only at concentrations of 10(3) colony forming units/mL, or higher. Using the same DNA extraction method, PCR was negative when the broths were inoculated with seven other related bacterial species, which results in a 100% specificity. Detection of S. pneumoniae by amplification of ply gene from blood cultures using the protocol of NaOH for DNA extraction is specific and provides results in a short lapse. However, the diagnostic sensitivity is not optimal, which limits its clinical use.

Blood culture gram stain, acridine orange stain and direct sensitivity-based antimicrobial therapy of bloodstream infection in patients with trauma.

PubMed

Behera, B; Mathur, P; Gupta, B

2010-01-01

The purpose of this study was to ascertain if the simple practice of Gram stain, acridine orange stain and direct sensitivity determination of positive blood culture bottles could be used to guide early and appropriate treatment in trauma patients with clinical suspicion of sepsis. The study also aimed to evaluate the error in interpreting antimicrobial sensitivity by direct method when compared to standard method and find out if specific antibiotic-organism combination had more discrepancies. Findings from consecutive episodes of blood stream infection at an Apex Trauma centre over a 12-month period are summarized. A total of 509 consecutive positive blood cultures were subjected to Gram staining. AO staining was done in BacT/ALERT-positive Gram-stain negative blood cultures. Direct sensitivity was performed from 369 blood culture broths, showing single type of growth in Gram and acridine orange staining. Results of direct sensitivity were compared to conventional sensitivity for errors. No 'very major' discrepancy was found in this study. About 5.2 and 1.8% minor error rates were noted in gram-positive and gram-negative bacteria, respectively, while comparing the two methods. Most of the discrepancies in gram-negative bacteria were noted in beta lactam - beta lactamase inhibitor combinations. Direct sensitivity testing was not reliable for reporting of methicillin and vancomycin resistance in Staphylococci. Gram stain result together with direct sensitivity testing is required for optimizing initial antimicrobial therapy in trauma patients with clinical suspicion of sepsis. Gram staining and AO staining proved particularly helpful in the early detection of candidaemia.

Evaluation of three sample preparation methods for the direct identification of bacteria in positive blood cultures by MALDI-TOF.

PubMed

Tanner, Hannah; Evans, Jason T; Gossain, Savita; Hussain, Abid

2017-01-18

Patient mortality is significantly reduced by rapid identification of bacteria from sterile sites. MALDI-TOF can identify bacteria directly from positive blood cultures and multiple sample preparation methods are available. We evaluated three sample preparation methods and two MALDI-TOF score cut-off values. Positive blood culture bottles with organisms present in Gram stains were prospectively analysed by MALDI-TOF. Three lysis reagents (Saponin, SDS, and SepsiTyper lysis bufer) were applied to each positive culture followed by centrifugation, washing and protein extraction steps. Methods were compared using the McNemar test and 16S rDNA sequencing was used to assess discordant results. In 144 monomicrobial cultures, using ≥2.000 as the cut-off value, species level identifications were obtained from 69/144 (48%) samples using Saponin, 86/144 (60%) using SDS, and 91/144 (63%) using SepsiTyper. The difference between SDS and SepsiTyper was not statistically significant (P = 0.228). Differences between Saponin and the other two reagents were significant (P < 0.01). Using ≥1.700 plus top three results matching as the cut-off value, species level identifications were obtained from 100/144 (69%) samples using Saponin, 103/144 (72%) using SDS, and 106/144 (74%) using SepsiTyper and there was no statistical difference between the methods. No true discordances between culture and direct MALDI-TOF identification were observed in monomicrobial cultures. In 32 polymicrobial cultures, MALDI-TOF identified one organism in 34-75% of samples depending on the method. This study demonstrates two inexpensive in-house detergent lysis methods are non-inferior to a commercial kit for analysis of positive blood cultures by direct MALDI-TOF in a clinical diagnostic microbiology laboratory.

Direct identification of microorganisms from positive blood cultures by MALDI-TOF MS using an in-house saponin method.

PubMed

Yonetani, Shota; Ohnishi, Hiroaki; Ohkusu, Kiyofumi; Matsumoto, Tetsuya; Watanabe, Takashi

2016-11-01

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a fast and reliable method for the identification of bacteria. A MALDI Sepsityper kit is generally used to prepare samples obtained directly from culture bottles. However, the relatively high cost of this kit is a major obstacle to introducing this method into routine clinical use. In this study, the accuracies of three different preparation methods for rapid direct identification of bacteria from positive blood culture bottles by MALDI-TOF MS analysis were compared. In total, 195 positive bottles were included in this study. Overall, 78.5%, 68.7%, and 76.4% of bacteria were correctly identified to the genus level (score ≥1.7) directly from positive blood cultures using the Sepsityper, centrifugation, and saponin methods, respectively. The identification rates using the Sepsityper and saponin methods were significantly higher than that using the centrifugation method (Sepsityper vs. centrifugation, p<0.001; saponin vs. centrifugation, p=0.003). These results suggest that the saponin method is superior to the centrifugation method and comparable to the Sepsityper method in the accuracy of rapid bacterial identification directly from blood culture bottles, and could be a less expensive alternative to the Sepsityper method. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Development of a Multiplexed Microsphere PCR for Culture-Free Detection and Gram-Typing of Bacteria in Human Blood Samples.

PubMed

Liang, Fang; Browne, Daniel J; Gray, Megan J; Gartlan, Kate H; Smith, David D; Barnard, Ross T; Hill, Geoffrey R; Corrie, Simon R; Markey, Kate A

2018-05-11

Bloodstream infection is a significant clinical problem, particularly in vulnerable patient groups such as those undergoing chemotherapy and bone marrow transplantation. Clinical diagnostics for suspected bloodstream infection remain centered around blood culture (highly variable timing, in the order of hours to days to become positive), and empiric use of broad-spectrum antibiotics is therefore employed for patients presenting with febrile neutropenia. Gram-typing provides the first opportunity to target therapy (e.g., combinations containing vancomycin or teicoplanin for Gram-positives; piperacillin-tazobactam or a carbapenem for Gram-negatives); however, current approaches require blood culture. In this study, we describe a multiplexed microsphere-PCR assay with flow cytometry readout, which can distinguish Gram-positive from Gram-negative bacterial DNA in a 3.5 h time period. The combination of a simple assay design (amplicon-dependent release of Gram-type specific Cy3-labeled oligonucleotides) and the Luminex-based readout (for quantifying each specific Cy3-labeled sequence) opens opportunities for further multiplexing. We demonstrate the feasibility of detecting common Gram-positive and Gram-negative organisms after spiking whole bacteria into healthy human blood prior to DNA extraction. Further development of DNA extraction methods is required to reach detection limits comparable to blood culture.

Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Use with Positive Blood Cultures: Methodology, Performance, and Optimization.

PubMed

Faron, Matthew L; Buchan, Blake W; Ledeboer, Nathan A

2017-12-01

Early initiation of effective antibiotics for septic patients is essential for patient survival. Matrix-assisted desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has revolutionized clinical microbiology for isolate identification and has the possibility to impact how blood culture testing is performed. This review discusses the various uses of MALDI-TOF MS for the identification and susceptibility testing of positive blood cultures, the performance of these methods, and the outcomes involved with its implementation. Copyright © 2017 American Society for Microbiology.

In vitro studies of the blood-brain barrier using isolated brain capillaries and cultured endothelial cells.

PubMed

Goldstein, G W; Betz, A L; Bowman, P D; Dorovini-Zis, K

1986-01-01

The endothelial cells in brain capillaries are the anatomic site of the blood-brain barrier. To learn more about the biology of these specialized cells, we developed methods to prepare suspensions of purified brain microvessels as well as primary cultures of endothelial cells in monolayer. These two preparations allow for direct investigation of the metabolism, transport properties, and receptor content of the brain capillary. We used isolated brain microvessels to study distribution of membrane carriers between the luminal and the abluminal plasma membrane of endothelial cells. We found that Na+K+-ATPase and the A-system amino-acid transport system are located predominantly on the abluminal surface of brain capillary endothelial cells. This distribution of transport carriers is consistent with the low permeability of potassium and small neutral amino acids in the blood-to-brain direction. It suggests, however, that both solutes can be actively transported across brain capillaries from the brain interstitial fluid to the blood. In tissue culture, the endothelial cells form continuous tight junctions with their neighbors. This results in a cellular layer impermeable to protein tracers. When exposed to hyperosmolar solutions, in an attempt to mimic the conditions that open the blood-brain barrier in vivo, we found a reversible separation of the tight junctions between contiguous endothelial cells. No indication of activation of pinocytosis was observed. In vitro systems provide a novel approach for studying the function of the blood-brain barrier and allow for observations not possible with intact animals.

Work System Assessment to Facilitate the Dissemination of a Quality Improvement Program for Optimizing Blood Culture Use: A Case Study Using a Human Factors Engineering Approach.

PubMed

Xie, Anping; Woods-Hill, Charlotte Z; King, Anne F; Enos-Graves, Heather; Ascenzi, Judy; Gurses, Ayse P; Klaus, Sybil A; Fackler, James C; Milstone, Aaron M

2017-11-20

Work system assessments can facilitate successful implementation of quality improvement programs. Using a human factors engineering approach, we conducted a work system assessment to facilitate the dissemination of a quality improvement program for optimizing blood culture use in pediatric intensive care units at 2 hospitals. Semistructured face-to-face interviews were conducted with clinicians from Johns Hopkins All Children's Hospital and University of Virginia Medical Center. Interview data were analyzed using qualitative content analysis. Blood culture-ordering practices are influenced by various work system factors, including people, tasks, tools and technologies, the physical environment, organizational conditions, and the external environment. A clinical decision-support tool could facilitate implementation by (1) standardizing blood culture-ordering practices, (2) ensuring that prescribing clinicians review the patient's condition before ordering a blood culture, (3) facilitating critical thinking, and (4) empowering nurses to communicate with physicians and advocate for adherence to blood culture-ordering guidelines. The success of interventions for optimizing blood culture use relies heavily on the local context. A work system analysis using a human factors engineering approach can identify key areas to be addressed for the successful dissemination of quality improvement interventions. © The Author 2017. Published by Oxford University Press on behalf of The Journal of the Pediatric Infectious Diseases Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Comparison of 'time to detection' values between BacT/ALERT VIRTUO and BacT/ALERT 3D instruments for clinical blood culture samples.

PubMed

Congestrì, Francesco; Pedna, Maria Federica; Fantini, Michela; Samuelli, Michela; Schiavone, Pasqua; Torri, Arianna; Bertini, Stefania; Sambri, Vittorio

2017-09-01

The early detection of bacteraemia and fungemia is of paramount importance to guide antimicrobial therapy in septic patients. In this study the 'time to detection' (TTD) value for the new blood culture system BacT/ALERT VIRTUO (VIRTUO) was evaluated in 1462 positive clinical bottles and compared with the TTD for 1601 positive clinical bottles incubated in the BacT/ALERT 3D system (BTA-3D). The most representative microorganisms isolated from bottles incubated in both blood culture systems were divided into eight categories (in order of frequency): coagulase-negative staphylococci (CoNS), Escherichia coli, Enterobacteriaceae (other than E. coli), Staphylococcus aureus, Enterococcus spp, viridans group streptococci, Pseudomonas aeruginosa, and Candida spp. The comparison of TTD values for the two blood culture systems strongly indicated that growth of the first five groups listed above was detected earlier with VIRTUO than with BTA-3D (p < 0.05). The new VIRTUO blood culture system can reduce the TTD for more than 75% of isolated microorganisms. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus: methicillin-resistant isolates are detected directly in blood cultures by multiplex PCR.

PubMed

Pereira, Eliezer M; Schuenck, Ricardo P; Malvar, Karoline L; Iorio, Natalia L P; Matos, Pricilla D M; Olendzki, André N; Oelemann, Walter M R; dos Santos, Kátia R N

2010-03-31

In this study, we standardized and evaluated a multiplex-PCR methodology using specific primers to identify Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus and their methicillin-resistance directly from blood cultures. Staphylococci clinical isolates (149) and control strains (16) previously identified by conventional methods were used to establish the multiplex PCR protocol. Subsequently, this methodology was evaluated using a fast and cheap DNA extraction protocol from 25 staphylococci positive blood cultures. A wash step of the pellet with 0.1% bovine serum albumin (BSA) solution was performed to reduce PCR inhibitors. Amplicons of 154bp (mecA gene), 271bp (S. haemolyticus mvaA gene) and 108 and 124bp (S. aureus and S. epidermidis species-specific fragments, respectively) were observed. Reliable results were obtained for 100% of the evaluated strains, suggesting that this new multiplex-PCR combined with an appropriate DNA-extraction method could be useful in the laboratory for fast and accurate identification of three staphylococci species and simultaneously their methicillin resistance directly in blood cultures.

Comparative Efficacies of Amoxicillin, Clindamycin, and Moxifloxacin in Prevention of Bacteremia following Dental Extractions

PubMed Central

Diz Dios, P.; Tomás Carmona, I.; Limeres Posse, J.; Medina Henríquez, J.; Fernández Feijoo, J.; Álvarez Fernández, M.

2006-01-01

We evaluated the efficacies of oral prophylactic treatment with amoxicillin (AMX), clindamycin (CLI), and moxifloxacin (MXF) in the prevention of bacteremia following dental extractions (BDE). Two hundred twenty-one adults who required dental extractions under general anesthesia were randomly assigned to a control group, an AMX group, a CLI group, and an MXF group (the individuals in the drug treatment groups received 2 g, 600 mg, and 400 mg, respectively, 1 to 2 h before anesthesia induction). Venous blood samples were collected from each patient at the baseline and 30 s, 15 min, and 1 h after the dental extractions. The samples were inoculated into BACTEC Plus aerobic and anaerobic blood culture bottles and were processed in a BACTEC 9240 instrument. Subculture and the further identification of the isolated bacteria were performed by conventional microbiological techniques. The prevalences of BDE in the control group, AMX group, CLI group, and MXF group were 96, 46, 85, and 57%, respectively, at 30 s; 64, 11, 70, and 24%, respectively, at 15 min; and 20, 4, 22, and 7%, respectively, at 1 h. Streptococcus spp. were the most frequently identified bacteria in all groups (44 to 68%), with the lowest percentage being detected in the AMX group (44%). AMX and MXF prophylaxis showed high efficacies in reducing the prevalence and duration of BDE, but CLI prophylaxis was noneffective. As a consequence, MXF prophylaxis is a promising antibiotic alternative for the prevention of BDE when beta-lactams are not indicated. PMID:16940094

3D cultured immortalized human hepatocytes useful to develop drugs for blood-borne HCV

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aly, Hussein Hassan; Shimotohno, Kunitada; Hijikata, Makoto

2009-02-06

Due to the high polymorphism of natural hepatitis C virus (HCV) variants, existing recombinant HCV replication models have failed to be effective in developing effective anti-HCV agents. In the current study, we describe an in vitro system that supports the infection and replication of natural HCV from patient blood using an immortalized primary human hepatocyte cell line cultured in a three-dimensional (3D) culture system. Comparison of the gene expression profile of cells cultured in the 3D system to those cultured in the existing 2D system demonstrated an up-regulation of several genes activated by peroxisome proliferator-activated receptor alpha (PPAR{alpha}) signaling. Furthermore,more » using PPAR{alpha} agonists and antagonists, we also analyzed the effect of PPAR{alpha} signaling on the modulation of HCV replication using this system. The 3D in vitro system described in this study provides significant insight into the search for novel anti-HCV strategies that are specific to various strains of HCV.« less

Procalcitonin levels in patients with positive blood culture, positive body fluid culture, sepsis, and severe sepsis: a cross-sectional study.

PubMed

Yu, Ying; Li, Xia-Xi; Jiang, Ling-Xiao; Du, Meng; Liu, Zhan-Guo; Cen, Zhong-Ran; Wang, Hua; Guo, Zhen-Hui; Chang, Ping

2016-01-01

Numerous investigations on procalcitonin (PCT) have been carried out, although few with large sample size. To deal with the complexity of sepsis, an understanding of PCT in heterogeneous clinical conditions is required. Hospitalized patients aged 10-79 years were included in this retrospective and cross-sectional study. PCT tests were assayed within 2 days of blood culture. A total of 2952 cases (from 2538 patients) were enrolled in this study, including 440 cases in the 'positive BC' group, 123 cases in the 'positive body fluid culture' group, and 2389 cases in the 'negative all culture' group. Median PCT values were 4.53 ng/ml, 2.95 ng/ml, and 0.49 ng/ml, respectively. Median PCT values in the gram-negative BC group and gram-positive BC group, respectively, were 6.99 ng/ml and 2.96 ng/ml. Median PCT values in the 'positive hydrothorax culture' group, 'positive ascites culture' group, 'positive bile culture' group, and 'positive cerebrospinal fluid culture' group, respectively, were 1.39 ng/ml, 8.32 ng/ml, 5.98 ng/ml, and 0.46 ng/ml. In all, 357 cases were classified into the 'sepsis' group, 150 of them were classified into the 'severe sepsis' group. Median PCT values were 5.63 ng/ml and 11.06 ng/ml, respectively. PCT could be used in clinical algorithms to diagnose positive infections and sepsis. Different PCT levels could be related to different kinds of microbemia, different infection sites, and differing severity of sepsis.

Comparison of methods for the identification of microorganisms isolated from blood cultures.

PubMed

Monteiro, Aydir Cecília Marinho; Fortaleza, Carlos Magno Castelo Branco; Ferreira, Adriano Martison; Cavalcante, Ricardo de Souza; Mondelli, Alessandro Lia; Bagagli, Eduardo; da Cunha, Maria de Lourdes Ribeiro de Souza

2016-08-05

Bloodstream infections are responsible for thousands of deaths each year. The rapid identification of the microorganisms causing these infections permits correct therapeutic management that will improve the prognosis of the patient. In an attempt to reduce the time spent on this step, microorganism identification devices have been developed, including the VITEK(®) 2 system, which is currently used in routine clinical microbiology laboratories. This study evaluated the accuracy of the VITEK(®) 2 system in the identification of 400 microorganisms isolated from blood cultures and compared the results to those obtained with conventional phenotypic and genotypic methods. In parallel to the phenotypic identification methods, the DNA of these microorganisms was extracted directly from the blood culture bottles for genotypic identification by the polymerase chain reaction (PCR) and DNA sequencing. The automated VITEK(®) 2 system correctly identified 94.7 % (379/400) of the isolates. The YST and GN cards resulted in 100 % correct identifications of yeasts (15/15) and Gram-negative bacilli (165/165), respectively. The GP card correctly identified 92.6 % (199/215) of Gram-positive cocci, while the ANC card was unable to correctly identify any Gram-positive bacilli (0/5). The performance of the VITEK(®) 2 system was considered acceptable and statistical analysis showed that the system is a suitable option for routine clinical microbiology laboratories to identify different microorganisms.

Re-using blood products as an alternative supplement in the optimisation of clinical-grade adipose-derived mesenchymal stem cell culture

PubMed Central

Phetfong, J.; Tawonsawatruk, T.; Seenprachawong, K.; Srisarin, A.; Isarankura-Na-Ayudhya, C.

2017-01-01

Objectives Adipose-derived mesenchymal stem cells (ADMSCs) are a promising strategy for orthopaedic applications, particularly in bone repair. Ex vivo expansion of ADMSCs is required to obtain sufficient cell numbers. Xenogenic supplements should be avoided in order to minimise the risk of infections and immunological reactions. Human platelet lysate and human plasma may be an excellent material source for ADMSC expansion. In the present study, use of blood products after their recommended transfusion date to prepare human platelet lysate (HPL) and human plasma (Hplasma) was evaluated for in vitro culture expansion and osteogenesis of ADMSCs. Methods Human ADMSCs were cultured in medium supplemented with HPL, Hplasma and a combination of HPL and Hplasma (HPL+Hplasma). Characteristics of these ADMSCs, including osteogenesis, were evaluated in comparison with those cultured in fetal bovine serum (FBS). Results HPL and HPL+Hplasma had a significantly greater growth-promoting effect than FBS, while Hplasma exhibited a similar growth-promoting effect to that of FBS. ADMSCs cultured in HPL and/or Hplasma generated more colony-forming unit fibroblasts (CFU-F) than those cultured in FBS. After long-term culture, ADMSCs cultured in HPL and/or Hplasma showed reduced cellular senescence, retained typical cell phenotypes, and retained differentiation capacities into osteogenic and adipogenic lineages. Conclusion HPL and Hplasma prepared from blood products after their recommended transfusion date can be used as an alternative and effective source for large-scale ex vivo expansion of ADMSCs. Cite this article: J. Phetfong, T. Tawonsawatruk, K. Seenprachawong, A. Srisarin, C. Isarankura-Na-Ayudhya, A. Supokawej. Re-using blood products as an alternative supplement in the optimisation of clinical-grade adipose-derived mesenchymal stem cell culture. Bone Joint Res 2017;6:414–422. DOI: 10.1302/2046-3758.67.BJR-2016-0342.R1. PMID:28720606

Isolation of Francisella tularensis and Yersinia pestis from Blood Cultures by Plasma Purification and Immunomagnetic Separation Accelerates Antibiotic Susceptibility Determination

PubMed Central

Aloni-Grinstein, Ronit; Schuster, Ofir; Yitzhaki, Shmuel; Aftalion, Moshe; Maoz, Sharon; Steinberger-Levy, Ida; Ber, Raphael

2017-01-01

The early symptoms of tularemia and plague, which are caused by Francisella tularensis and Yersinia pestis infection, respectively, are common to other illnesses, resulting in a low index of suspicion among clinicians. Moreover, because these diseases can be treated only with antibiotics, rapid isolation of the bacteria and antibiotic susceptibility testing (AST) are preferable. Blood cultures of patients may serve as a source for bacteria isolation. However, due to the slow growth rates of F. tularensis and Y. pestis on solid media, isolation by plating blood culture samples on proper agar plates may require several days. Thus, improving the isolation procedure prior to antibiotic susceptibility determination is a major clinically relevant need. In this study, we developed a rapid, selective procedure for the isolation of F. tularensis and Y. pestis from blood cultures. We examined drop-plating and plasma purification followed by immunomagnetic separation (IMS) as alternative isolation methods. We determined that replacing the classical isolation method with drop-plating is advantageous with respect to time at the expense of specificity. Hence, we also examined isolation by IMS. Sub-localization of F. tularensis within blood cultures of infected mice has revealed that the majority of the bacteria are located within the extracellular fraction, in the plasma. Y. pestis also resides within the plasma. Therefore, the plasma fraction was isolated from blood cultures and subjected to an IMS procedure using polyclonal anti-F. tularensis live vaccine strain (LVS) or anti-Y. pestis antibodies conjugated to 50-nm nano-beads. The time required to reach an inoculum of sufficient bacteria for AST was shortest when using the plasma and IMSs for both bacteria, saving up to 2 days of incubation for F. tularensis and 1 day for Y. pestis. Our isolation procedure provides a proof of concept for the clinical relevance of rapid isolation for AST from F. tularensis- and Y. pestis

Rapid identification of pathogens directly from blood culture bottles by Bruker matrix-assisted laser desorption laser ionization-time of flight mass spectrometry versus routine methods.

PubMed

Jamal, Wafaa; Saleem, Rola; Rotimi, Vincent O

2013-08-01

The use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for identification of microorganisms directly from blood culture is an exciting dimension to the microbiologists. We evaluated the performance of Bruker SepsiTyper kit™ (STK) for direct identification of bacteria from positive blood culture. This was done in parallel with conventional methods. Nonrepetitive positive blood cultures from 160 consecutive patients were prospectively evaluated by both methods. Of 160 positive blood cultures, the STK identified 114 (75.6%) isolates and routine conventional method 150 (93%). Thirty-six isolates were misidentified or not identified by the kit. Of these, 5 had score of >2.000 and 31 had an unreliable low score of <1.7. Four of 8 yeasts were identified correctly. The average turnaround time using the STK was 35 min, including extraction steps and 30:12 to 36:12 h with routine method. The STK holds promise for timely management of bacteremic patients. Copyright © 2013 Elsevier Inc. All rights reserved.

Identification and susceptibility testing of microorganism by direct inoculation from positive blood culture bottles by combining MALDI-TOF and Vitek-2 Compact is rapid and effective.

PubMed

Romero-Gómez, María-Pilar; Gómez-Gil, Rosa; Paño-Pardo, Jose Ramón; Mingorance, Jesús

2012-12-01

The objective of this study was to evaluate the reliability and accuracy of the combined use of MALDI-TOF MS bacterial identification and the Vitek-2 Compact antimicrobial susceptibility testing (AST) directly from positive blood cultures. Direct identification by MALDI-TOF MS and AST were performed in parallel to the standard methods in all positively flagged blood cultures bottles during the study period. Three hundred and twenty four monomicrobial positive blood cultures were included in the present study, with 257 Gram-negative and 67 Gram-positive isolates. MALDI-TOF MS identification directly from blood bottles reported the correct identification for Enterobacteriaceae in 97.7%, non-fermentative Gram-negative bacilli 75.0%, Staphylococcus aureus 75.8%, coagulase negative staphylococci 63.3% and enterococci 63.3%. A total 6156 isolate/antimicrobial agent combinations were tested. Enterobacteriaceae group and non-fermentative Gram-negative Bacilli showed an agreement of 96.67% and 92.30%, respectively, for the Gram-positive cocci the overall agreement found was 97.84%. We conclude that direct identification by MALDI-TOF and inoculation of Vitek-2 Compact AST with positive blood culture bottles yielded very good results and decreased time between initial inoculation of blood culture media and determination of the antibiotic susceptibility for Gram-negative rods and Gram-positive cocci causing bacteremia. Copyright © 2012 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

A rapid, highly sensitive and culture-free detection of pathogens from blood by positive enrichment.

PubMed

Vutukuru, Manjula Ramya; Sharma, Divya Khandige; Ragavendar, M S; Schmolke, Susanne; Huang, Yiwei; Gumbrecht, Walter; Mitra, Nivedita

2016-12-01

Molecular diagnostics is a promising alternative to culture based methods for the detection of bloodstream infections, notably due to its overall lower turnaround time when starting directly from patient samples. Whole blood is usually the starting diagnostic sample in suspected bloodstream infections. The detection of low concentrations of pathogens in blood using a molecular assay necessitates a fairly high starting volume of blood sample in the range of 5-10mL. This large volume of blood sample has a substantial accompanying human genomic content that interferes with pathogen detection. In this study, we have established a workflow using magnetic beads coated with Apolipoprotein H that makes it possible to concentrate pathogens from a 5.0mL whole blood sample, thereby enriching pathogens from whole blood background and also reducing the sample volume to ~200μL or less. We have also demonstrated that this method of enrichment allows detection of 1CFU/mL of Escherichia coli, Enterococcus gallinarum and Candida tropicalis from 5mL blood using quantitative PCR; a detection limit that is not possible in unenriched samples. The enrichment method demonstrated here took 30min to complete and can be easily integrated with various downstream molecular and microbiological techniques. Copyright © 2016 Elsevier B.V. All rights reserved.

An eight-year review of blood culture and susceptibility among sepsis cases in an emergency department in Northeastern Malaysia.

PubMed

Hashairi, F; Hasan, H; Azlan, K; Deris, Z Z

2011-12-01

An understanding of common pathogens and their antibiotic sensitivity patterns is critical for proper management of sepsis in Emergency Department (ED). The goal of the study was to identify common organisms isolated from blood cultures of patients attended to ED and their antimicrobial susceptibility. Beginning from 2002, all cases of positive blood culture collected by the ED, Hospital Universiti Sains Malaysia (HUSM) were recorded and analysed. Over the period of eight years, we documented 995 cases of positive blood cultures. Of these samples, 549 (55.2%) were Gram-negative bacteria; 419 (42.1%) were Gram-positive bacteria; 10 (1.0%) were anaerobic organisms; 10 (1.0%) were fungus; and 7 (0.7%) cases were mixed organisms. Gram-negative bacteria were observed to develop more resistance to antimicrobial agents, especially those commonly used in an outpatient setting with less than 80% sensitivity to ampicillin, cotrimoxazole and ciprofloxacin. By contrast, there has been no marked change in the sensitivity trends of Gram-positive bacteria over the same period. In conclusion, ED physicians are more equipped to initiate empirical antimicrobial therapy especially when dealing with possibility of Gram-negative sepsis.

Rapid Identification of Microorganisms from Positive Blood Culture by MALDI-TOF MS After Short-Term Incubation on Solid Medium.

PubMed

Curtoni, Antonio; Cipriani, Raffaella; Marra, Elisa Simona; Barbui, Anna Maria; Cavallo, Rossana; Costa, Cristina

2017-01-01

Matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a useful tool for rapid identification of microorganisms. Unfortunately, its direct application to positive blood culture is still lacking standardized procedures. In this study, we evaluated an easy- and rapid-to-perform protocol for MALDI-TOF MS direct identification of microorganisms from positive blood culture after a short-term incubation on solid medium. This protocol was used to evaluate direct identification of microorganisms from 162 positive monomicrobial blood cultures; at different incubation times (3, 5, 24 h), MALDI-TOF MS assay was performed from the growing microorganism patina. Overall, MALDI-TOF MS concordance with conventional methods at species level was 60.5, 80.2, and 93.8% at 3, 5, and 24 h, respectively. Considering only bacteria, the identification performances at species level were 64.1, 85.0, and 94.1% at 3, 5, and 24 h, respectively. This protocol applied to a commercially available MS typing system may represent, a fast and powerful diagnostic tool for pathogen direct identification and for a promptly and pathogen-driven antimicrobial therapy in selected cases.

Isolation of Corynebacterium tuscaniae sp. nov. from Blood Cultures of a Patient with Endocarditis

PubMed Central

Riegel, Philippe; Creti, Roberta; Mattei, Romano; Nieri, Alfredo; von Hunolstein, Christina

2006-01-01

A strain of an unknown coryneform bacterium was repeatedly isolated in pure culture from the blood of a patient affected by endocarditis. Comparative 16S rRNA gene sequence analysis revealed that this isolate represented a new subline within the genus Corynebacterium. This new taxon can be identified by the presence of corynomycolic acids and its enzymatic activities and fermentation of sugars. Acid production from glucose and maltose, pyrazinamidase and alkaline phoshatase activities, and hippurate hydrolysis were the most characteristic phenotypic features of the bacterium. On the basis of both phenotypic and phylogenetic evidence, it is proposed that this isolate be classified as a novel species, Corynebacterium tuscaniae sp. nov. The type strain, ISS-5309, has been deposited in the American Type Culture Collection (ATCC BAA-1141) and in the Culture Collection of the University of Göteborg (CCUG 51321). PMID:16455875

Rapid Detection of Staphylococcus aureus and Methicillin-Resistant S. aureus (MRSA) in Wound Specimens and Blood Cultures: Multicenter Preclinical Evaluation of the Cepheid Xpert MRSA/SA Skin and Soft Tissue and Blood Culture Assays▿

PubMed Central

Wolk, D. M.; Struelens, M. J.; Pancholi, P.; Davis, T.; Della-Latta, P.; Fuller, D.; Picton, E.; Dickenson, R.; Denis, O.; Johnson, D.; Chapin, K.

2009-01-01

A multicenter preclinical evaluation was conducted to evaluate the performance of two Cepheid Xpert assays for detection of methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus. Sensitivity was 97.1% and 98.3% for MRSA in wound and blood culture specimens, respectively. Sensitivity was 100% for S. aureus from both specimen types. PMID:19144803

Nanomechanical sensor applied to blood culture pellets: a fast approach to determine the antibiotic susceptibility against agents of bloodstream infections.

PubMed

Stupar, P; Opota, O; Longo, G; Prod'hom, G; Dietler, G; Greub, G; Kasas, S

2017-06-01

The management of bloodstream infection, a life-threatening disease, largely relies on early detection of infecting microorganisms and accurate determination of their antibiotic susceptibility to reduce both mortality and morbidity. Recently we developed a new technique based on atomic force microscopy capable of detecting movements of biologic samples at the nanoscale. Such sensor is able to monitor the response of bacteria to antibiotic's pressure, allowing a fast and versatile susceptibility test. Furthermore, rapid preparation of a bacterial pellet from a positive blood culture can improve downstream characterization of the recovered pathogen as a result of the increased bacterial concentration obtained. Using artificially inoculated blood cultures, we combined these two innovative procedures and validated them in double-blind experiments to determine the susceptibility and resistance of Escherichia coli strains (ATCC 25933 as susceptible and a characterized clinical isolate as resistant strain) towards a selection of antibiotics commonly used in clinical settings. On the basis of the variance of the sensor movements, we were able to positively discriminate the resistant from the susceptible E. coli strains in 16 of 17 blindly investigated cases. Furthermore, we defined a variance change threshold of 60% that discriminates susceptible from resistant strains. By combining the nanomotion sensor with the rapid preparation method of blood culture pellets, we obtained an innovative, rapid and relatively accurate method for antibiotic susceptibility test directly from positive blood culture bottles, without the need for bacterial subculture. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Bacteremia following dental implant surgery: preliminary results.

PubMed

Bölükbaşı, Nilüfer; Özdemir, Tayfun; Öksüz, Lütfiye; Gürler, Nezahat

2012-01-01

The aims of this study were to investigate the incidence of bacteremia, bacteriology and antibiotic susceptibility against to causative bacteria associated with dental implant installation. 30 generally healthy patients were enrolled in this study. Blood samples were collected at baseline and at 30 minutes after dental implant installation and 24 hours after dental implant surgery. Blood samples were cultured in a BACTEC system. The isolated bacteria were identified using conventional methods. Antimicrobial sensitivity tests were performed by disc diffusion. No bacteria were isolated at the baseline and 24 hours after surgery, whereas the prevalence of bacteremia at 30 minutes after dental implant installation was 23%. The isolated bacteria species were Staphylococcus epidermidis, Eubacterium spp., Corynebacterium spp. and Streptococcus viridans. The Staphylococcus epidermidis, which was isolated in three patients, was found to be resistant to penicillin which is first choice of many clinicians. Our findings suggest that installation of dental implants can produce bacteremia. Within the limitations of this study, it can be speculated that the resistance of antibiotics may compromise the routine prophylaxis against infective endocarditis. Therefore use of blood cultures and antibiograms may be suggested in risky patients. The outcome of the present study should be verified using a larger patient group with varying conditions.

Evaluation of growth based rapid microbiological methods for sterility testing of vaccines and other biological products.

PubMed

Parveen, Seema; Kaur, Simleen; David, Selwyn A Wilson; Kenney, James L; McCormick, William M; Gupta, Rajesh K

2011-10-19

Most biological products, including vaccines, administered by the parenteral route are required to be tested for sterility at the final container and also at various stages during manufacture. The sterility testing method described in the Code of Federal Regulations (21 CFR 610.12) and the United States Pharmacopoeia (USP, Chapter <71>) is based on the observation of turbidity in liquid culture media due to growth of potential contaminants. We evaluated rapid microbiological methods (RMM) based on detection of growth 1) by adenosine triphosphate (ATP) bioluminescence technology (Rapid Milliflex(®) Detection System [RMDS]), and 2) by CO(2) monitoring technologies (BacT/Alert and the BACTEC systems), as alternate sterility methods. Microorganisms representing Gram negative, Gram positive, aerobic, anaerobic, spore forming, slow growing bacteria, yeast, and fungi were prepared in aliquots of Fluid A or a biological matrix (including inactivated influenza vaccines) to contain approximately 0.1, 1, 10 and 100 colony forming units (CFU) in an inoculum of 10 ml. These preparations were inoculated to the specific media required for the various methods: 1) fluid thioglycollate medium (FTM) and tryptic soy broth (TSB) of the compendial sterility method (both membrane filtration and direct inoculation); 2) tryptic soy agar (TSA), Sabouraud dextrose agar (SDA) and Schaedler blood agar (SBA) of the RMDS; 3) iAST and iNST media of the BacT/Alert system and 4) Standard 10 Aerobic/F and Standard Anaerobic/F media of the BACTEC system. RMDS was significantly more sensitive in detecting various microorganisms at 0.1CFU than the compendial methods (p<0.05), whereas the compendial membrane filtration method was significantly more sensitive than the BACTEC and BacT/Alert methods (p<0.05). RMDS detected all microorganisms significantly faster than the compendial method (p<0.05). BacT/Alert and BACTEC methods detected most microorganisms significantly faster than the compendial method

Nanobacteria from blood: the smallest culturable autonomously replicating agent on Earth

NASA Astrophysics Data System (ADS)

Kajander, E. Olavi; Kuronen, Ilpo; Akerman, Kari K.; Pelttari, Alpo; Ciftcioglu, Neva

1997-07-01

Nanobacteria are the first mineral forming bacteria isolated from blood and blood products. They are coccoid cell-walled organisms with a size of 0.08 - 0.5 micrometers in EM, occure in clusters, produce a biofilm containing carbonate or hydroxyl apatite, and are highly resistant to heat, gamma-irradiation and antibiotics. Their growth rate is about one hundredth that of ordinary bacteria and they divide via several mechanisms. Taq polymerase was able to use their nontraditional nucleic acid as a template. 16S rRNA gene sequence results positioned them into the alpha-2 subgroup of Proteobacteria. Nanobacteria are smallest cell-walled bacteria since they can pass through 0.07 micrometers pores. In low-serum cultures, they form even smaller elementary particles or tubular units. How can blood be infected with such slow growing, heat and radio-resistant bacteria? The answer may lie in their phylogeny: alpha-2 subgroup has organisms from soil exposed to radiation and heat, that can penetrate into eukaryotic cells. Nanobacteria grow so slowly that they require a niche `cleaned' with heat, radiation or immunodefence. For survival they cloak themselves in apatite, a normal constituent of mammalian body. This may link nanobacteria to nannobacteria discovered from sedimentary rocks by Dr. Folk. Both have similar size, size variation, clustering and mineral deposits. They may resemble the probable ancient bacterial fossils in the Martian meteorite ALH84001.

An in-house assay is superior to Sepsityper for direct matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry identification of yeast species in blood cultures.

PubMed

Bidart, Marie; Bonnet, Isabelle; Hennebique, Aurélie; Kherraf, Zine Eddine; Pelloux, Hervé; Berger, François; Cornet, Muriel; Bailly, Sébastien; Maubon, Danièle

2015-05-01

We developed an in-house assay for the direct identification, by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, of yeasts in blood culture. Sixty-one representative strains from 12 species were analyzed in spiked blood cultures. Our assay accurately identified 95 of 107 (88.8%) positive blood cultures and outperformed the commercial Sepsityper kit (81.7% identification). Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Shortened Time to Identify Staphylococcus Species from Blood Cultures and Methicillin Resistance Testing Using CHROMAgar

PubMed Central

Chihara, Shingo; Hayden, Mary K.; Minogue-Corbett, Eileen; Singh, Kamaljit

2009-01-01

The ability to rapidly differentiate coagulase-negative staphylococcus (CoNS) from Staphylococcus aureus and to determine methicillin resistance is important as it affects the decision to treat empiric antibiotic selection. The objective of this study was to evaluate CHROMagar S. aureus and CHROMagar MRSA (Becton Dickinson) for rapid identification of Staphylococcus spp. directly from blood cultures. Consecutive blood culture bottles (BacT Alert 3D SA and SN, bioMérieux) growing gram-positive cocci in clusters were evaluated. An aliquot was plated onto CHROMagar MRSA (C-MRSA) and CHROMagar S. aureus (C-SA) plates, which were read at 12 to 16 hours. C-SA correctly identified 147/147 S. aureus (100% sensitivity); 2 CoNS were misidentified as S. aureus (98% specificity). C-MRSA correctly identified 74/77 MRSA (96% sensitivity). None of the MSSA isolates grew on C-MRSA (100% specificity). In conclusion, CHROMagar is a rapid and sensitive method to distinguish MRSA, MSSA, and coagulase-negative Staphylococcus and may decrease time of reporting positive results. PMID:20016679

Detection of Legionella pneumophila serogroup 1 in blood cultures from a patient treated with tumor necrosis factor-alpha inhibitor.

PubMed

Kaku, Norihito; Yanagihara, Katsunori; Morinaga, Yoshitomo; Sato, Tsuyoshi; Nakashima, Munetoshi; Sakai, Takahiro; Tominaga, Hiroo; Wakigawa, Fumiko; Nagashima, Seiji; Fukuda, Minoru; Hashiguchi, Kohji; Kohno, Shigeru

2013-02-01

A 65-year-old man was admitted to our hospital with a temperature of 39.3 °C, cough, sputum, and pharyngeal discomfort that had persisted for 3 days. He had been treated with methotrexate and adalimumab (a tumor necrosis factor-alpha [TNF-α] inhibitor) for rheumatoid arthritis for 2 years, and he had also been treated with S-1 (tegafur, gimeracil, and oteracil potassium) for pancreatic metastasis of gastric cancer for 2 months. Regardless of the underlying pathologies, his general condition was good and he had worked as an electrician until 2 days before admission. However, his appetite had suddenly decreased from the day before admission, and high fever and hypoxia were also evident upon admission. A chest X-ray and computed tomography scan revealed left pleural effusion and consolidation in both lungs. The pneumonia severity index score was 165 and the risk class was V. Accordingly, we started to treat the pneumonia with a combination of levofloxacin and meropenem. Thereafter, we received positive urinary antigen test findings for Legionella pneumophila. After hospitalization, hypoxia was progressed and hypotension was emerged. Despite the application of appropriate antibiotics, vasopressors, and oxygenation, the patient died 8 h after admission. Even after his death, blood cultures were continued to consider the possibility of bacterial co-infection. Although no bacteria were detected from blood cultures, Gimenez staining revealed pink bacteria in blood culture fluids. Subsequent blood fluid culture in selective medium revealed L. pneumophila serogroup 1. Recently, TNF-α inhibitors have been described as a risk factor for Legionnaires' disease. In consideration of the increased frequency of TNF-α inhibitors, we may need to recognize anew that L. pneumophila might be a pathogen of severe community-acquired pneumonia.

Impact of reporting gram stain results from blood culture bottles on the selection of antimicrobial agents.

PubMed

Uehara, Yuki; Yagoshi, Michiko; Tanimichi, Yumiko; Yamada, Hiroko; Shimoguchi, Kazuo; Yamamoto, Sachiyo; Yanai, Mitsuru; Kumasaka, Kazunari

2009-07-01

We assessed the usefulness of reporting direct blood Gram stain results compared with the results of positive blood cultures in 482 episodes and monitored impact on selection of antimicrobial treatment. We found that the reporting groups "Staphylococcus spp," "Pseudomonas spp and related organisms," and "yeasts" identified in this way matched perfectly with later culture identification. When the report indicated Staphylococcus spp or Pseudomonas spp and related organisms, physicians started or changed antimicrobials suitable for these bacteria more frequently than when "other streptococci" and "family Enterobacteriaceae" were reported (P < .05). Incorrect recognition of Acinetobacter spp as Enterobacteriaceae family is still the most challenging problem in this context. Gram stain results that definitively identify Staphylococcus spp, Pseudomonas spp and related organisms, and yeasts reliably can be rapidly provided by clinical laboratories; this information has a significant impact on early selection of effective antimicrobials. Further investigation is needed to assess the clinical impact of reporting Gram stain results in bacteremia.

Applications of copolymer for rapid identification of bacteria in blood culture broths using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

PubMed

Ashizawa, Kazuho; Murata, Syota; Terada, Takashi; Ito, Daisuke; Bunya, Masaru; Watanabe, Koji; Teruuchi, Yoko; Tsuchida, Sachio; Satoh, Mamoru; Nishimura, Motoi; Matsushita, Kazuyuki; Sugama, Yuji; Nomura, Fumio

2017-08-01

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) can be used to identify pathogens in blood culture samples. However, sample pretreatment is needed for direct identification of microbes in blood culture bottles. Conventional protocols are complex and time-consuming. Therefore, in this study, we developed a method for collecting bacteria using polyallylamine-polystyrene copolymer for application in wastewater treatment technology. Using representative bacterial species Escherichia coli and Staphylococcus capitis, we found that polyallylamine-polystyrene can form visible aggregates with bacteria, which can be identified using MALDI-TOF MS. The processing time of our protocol was as short as 15min. Hemoglobin interference in MALDI spectra analysis was significantly decreased in our method compared with the conventional method. In a preliminary experiment, we evaluated the use of our protocol to identify clinical isolates from blood culture bottles. MALDI-TOF MS-based identification of 17 strains from five bacterial species (E. coli, Klebsiella pneumoniae, Enterococcus faecalis, S. aureus, and S. capitis) collected by our protocol was satisfactory. Prospective large-scale studies are needed to further evaluate the clinical application of this novel and simple method of collecting bacteria in blood culture bottles. Copyright © 2017 Elsevier B.V. All rights reserved.

Integration of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in blood culture diagnostics: a fast and effective approach.

PubMed

Klein, Sabrina; Zimmermann, Stefan; Köhler, Christine; Mischnik, Alexander; Alle, Werner; Bode, Konrad A

2012-03-01

Sepsis is a major cause of mortality in hospitalized patients worldwide, with lethality rates ranging from 30 to 70 %. Sepsis is caused by a variety of different pathogens, and rapid diagnosis is of outstanding importance, as early and adequate antimicrobial therapy correlates with positive clinical outcome. In recent years, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) fingerprinting has become a powerful tool in microbiological diagnostics. The direct identification of micro-organisms in a positive blood culture by MALDI-TOF MS can shorten the diagnostic procedure significantly. Therefore, the aim of the present study was to evaluate whether identification rates could be improved by using the new Sepsityper kit from Bruker Daltonics for direct isolation and identification of bacteria from positive blood cultures by MALDI-TOF MS compared with the use of conventional separator gel columns, and to integrate the MALDI-TOF MS-based identification method into the routine course of blood culture diagnostics in the setting of a microbiological laboratory at a university hospital in Germany. The identification of Gram-negative bacteria by MALDI-TOF MS was significantly better using the Sepsityper kit compared with a separator gel tube-based method (99 and 68 % correct identification, respectively). For Gram-positive bacteria, only 73 % were correctly identified by MALDI-TOF with the Sepsityper kit and 59 % with the separator gel tube assay. A major problem of both methods was the poor identification of Gram-positive grape-like clustered cocci. As differentiation of Staphylococcus aureus from coagulase-negative staphylococci is of clinical importance, a PCR was additionally established that was capable of identifying S. aureus directly from positive blood cultures, thus closing this diagnostic gap. Another benefit of the PCR approach is the possibility of directly detecting the genes responsible for meticillin

An evaluation of three processing methods and the effect of reduced culture times for faster direct identification of pathogens from BacT/ALERT blood cultures by MALDI-TOF MS.

PubMed

Loonen, A J M; Jansz, A R; Stalpers, J; Wolffs, P F G; van den Brule, A J C

2012-07-01

Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) is a fast and reliable method for the identification of bacteria from agar media. Direct identification from positive blood cultures should decrease the time to obtaining the result. In this study, three different processing methods for the rapid direct identification of bacteria from positive blood culture bottles were compared. In total, 101 positive aerobe BacT/ALERT bottles were included in this study. Aliquots from all bottles were used for three bacterial processing methods, i.e. the commercially available Bruker's MALDI Sepsityper kit, the commercially available Molzym's MolYsis Basic5 kit and a centrifugation/washing method. In addition, the best method was used to evaluate the possibility of MALDI application after a reduced incubation time of 7 h of Staphylococcus aureus- and Escherichia coli-spiked (1,000, 100 and 10 colony-forming units [CFU]) aerobe BacT/ALERT blood cultures. Sixty-six (65%), 51 (50.5%) and 79 (78%) bottles were identified correctly at the species level when the centrifugation/washing method, MolYsis Basic 5 and Sepsityper were used, respectively. Incorrect identification was obtained in 35 (35%), 50 (49.5%) and 22 (22%) bottles, respectively. Gram-positive cocci were correctly identified in 33/52 (64%) of the cases. However, Gram-negative rods showed a correct identification in 45/47 (96%) of all bottles when the Sepsityper kit was used. Seven hours of pre-incubation of S. aureus- and E. coli-spiked aerobe BacT/ALERT blood cultures never resulted in reliable identification with MALDI-TOF MS. Sepsityper is superior for the direct identification of microorganisms from aerobe BacT/ALERT bottles. Gram-negative pathogens show better results compared to Gram-positive bacteria. Reduced incubation followed by MALDI-TOF MS did not result in faster reliable identification.

Cord blood testing

MedlinePlus

... Blood culture (if an infection is suspected) Blood gases (including oxygen, carbon dioxide, and pH levels) Blood ... 2018, A.D.A.M., Inc. Duplication for commercial use must be authorized in writing by ADAM ...

In the era of the 24 h laboratory, does communicating Gram stain results from blood cultures flagging positive outside of conventional working hours alter patient management?

PubMed

Moore, Jonathan S; Koerner, Roland J

2015-11-01

As laboratories move towards 24 h a day working patterns, we aim to evaluate if expediting the availability of provisional blood culture results outside of normal working hours would derive clinical benefit. 116 blood cultures flagging positive outside of conventional working hours (20:00-09:00) were studied. In each case, medical records were reviewed and cases discussed with clinicians to determine if earlier communication of results would have altered management and affected the outcome. Organisms were seen in 102/116 blood cultures. In total, 76/82 (92.7%) patients with cultures deemed to be significant were on an antibiotic. The isolate was sensitive to the prescribed antibiotic in 56/74 (76%) cases. Input from a microbiologist is likely to have altered management in 14 (13.7%) cases, but unlikely to have affected any outcomes. We found no compelling evidence that expediting the availability of Gram stain results from positive blood cultures alone improves patient outcome. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Optimisation of decontamination method and influence of culture media on the recovery of Mycobacterium avium subspecies paratuberculosis from spiked water sediments.

PubMed

Aboagye, G; Rowe, M T

2018-07-01

The recovery of Mycobacterium avium subspecies paratuberculosis (Map) from the environment can be a laborious process - owing to Map being fastidious, its low number, and also high numbers of other microbial populations in such settings. Protocols i.e. filtration, decontamination and modified elution were devised to recover Map from spiked water sediments. Three culture media: Herrold's Egg Yolk Media (HEYM), Middlebrook 7H10 (M-7H10) and Bactec 12B were then employed to grow the organism following its elution. In the sterile sediment samples the recovery of Map was significant between the time of exposure for each of HEYM and M-7H10, and insignificant between both media (P < 0.05). However, in the non-sterile sediment samples, the HEYM grew other background microflora including moulds at all the times of exposure whilst 4 h followed by M-7H10 culture yielded Map colonies without any background microflora. Using sterile samples only for the Bactec 12B, the recovery of Map decreased as time of exposure increased. Based on these findings, M-7H10 should be considered for the recovery of Map from the natural environment including water sediments where the recovery of diverse microbial species remains a challenge. Map is a robust pathogen that abides in the environment. In water treatment operations, Map associates with floccules and other particulate matter including sediments. It is also a fastidious organism, and its detection and recovery from the water environment is a laborious process and can be misleading within the abundance of other mycobacterial species owing to their close resemblance in phylogenetic traits. In the absence of a reliable recovery method, Map continues to pose public health risks through biofilm in household water tanks, hence the need for the development of a reliable recovery protocol to monitor the presence of Map in water systems in order to curtail its public health risks. Copyright © 2018 Elsevier B.V. All rights reserved.

Cytokine Production in Mixed Cultures of Mesenchymal Stromal Cells from Wharton's Jelly and Peripheral Blood Lymphocytes.

PubMed

Poltavtsev, A M; Poltavtseva, R A; Yushina, M N; Volgina, N E; Svirshchevskaya, E V

2017-05-01

We compared the production of 19 humoral factors in mixed cultures of mesenchymal stromal cells from Wharton's jelly and allogenic peripheral blood lymphocytes. For evaluation of the specificity of immunosuppressive activity of mesenchymal stromal cells, comparative analysis of the production of these humoral factors in mixed cultures of lymphocytes and epithelial BxPC-3 cells was conducted. The production of soluble factors in both mono- and mixed cultures significantly correlated (p<0.05). The maximum production was found for proinflammatory chemokine IP-10 and IFN-γ and anti-inflammatory cytokine IL-10. The major difference of mesenchymal stromal cells from epithelial BxPC-3 cells was 7-fold higher production of IL-10, which can explain the immunosuppressive effect of mesenchymal stromal cells.

Evaluation of a Semiquantitative Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry Method for Rapid Antimicrobial Susceptibility Testing of Positive Blood Cultures.

PubMed

Jung, Jette S; Hamacher, Christina; Gross, Birgit; Sparbier, Katrin; Lange, Christoph; Kostrzewa, Markus; Schubert, Sören

2016-11-01

With the increasing prevalence of multidrug-resistant Gram-negative bacteria, rapid identification of the pathogen and its individual antibiotic resistance is crucial to ensure adequate antiinfective treatment at the earliest time point. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry for the identification of bacteria directly from the blood culture bottle has been widely established; however, there is still an urgent need for new methods that permit rapid resistance testing. Recently, a semiquantitative MALDI-TOF mass spectrometry-based method for the prediction of antibiotic resistance was described. We evaluated this method for detecting nonsusceptibility against two β-lactam and two non-β-lactam antibiotics. A collection of 30 spiked blood cultures was tested for nonsusceptibility against gentamicin and ciprofloxacin. Furthermore, 99 patient-derived blood cultures were tested for nonsusceptibility against cefotaxime, piperacillin-tazobactam, and ciprofloxacin in parallel with MALDI-TOF mass spectrometry identification from the blood culture fluid. The assay correctly classified all isolates tested for nonsusceptibility against gentamicin and cefotaxime. One misclassification for ciprofloxacin nonsusceptibility and five misclassifications for piperacillin-tazobactam nonsusceptibility occurred. Identification of the bacterium and prediction of nonsusceptibility was possible within approximately 4 h. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Evaluation of the usefulness of a quantitative blood culture in the diagnosis of catheter-related bloodstream infection: Comparative analysis of two periods (2002 and 2012).

PubMed

Planes, Anna Maria; Calleja, Raquel; Bernet, Albert; Campins-Martí, Magda; Almirante, Benito; Pumarola, Tomàs; Fernández-Hidalgo, Núria

2016-10-01

A retrospective study was conducted to investigate the usefulness of systematic quantitative blood culture (QBC) in the diagnosis of catheter-related bloodstream infection (CRBSI) during two 1-year periods (2002 and 2012). The study included all QBC requests sent to the microbiology laboratory for suspected CRBSI in adults (≥18 years) with any type of intravascular catheter (IVC). Based on a ratio of ≥4:1CFU/mL of the same microorganism between IVC blood culture from any lumen and peripheral blood culture, 5 diagnostic groups were defined: confirmed or probable CRBSI, primary BSI, other focus of infection, and colonization. In total, 4521 QBCs were evaluated; 24% positive in 2002 and 16% in 2012 (P<0.0001). There were 243 episodes of suspected CRBSI (101 in 2002 and 142 in 2012). Confirmed CRBSI episodes were higher in 2002 than 2012 (56% vs 34%) (P<0.0001), whereas colonization episodes were lower (18% vs 38%) (P=0.0006). Gram-positive cocci decrease in 2012 relative to 2002 (56% vs 79.7%) (P=0.022). Almost one-third (32%) of confirmed CRBSI would have been missed if blood from all catheter lumens had not been cultured. QBC is a useful method for diagnosing CRBSI. Blood samples from all catheter lumens must be cultured to avoid missing around one-third of CRBSI diagnoses. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Rapid identification of pneumococci, enterococci, beta-haemolytic streptococci and S. aureus from positive blood cultures enabling early reports.

PubMed

Larsson, Marie C; Karlsson, Ewa; Woksepp, Hanna; Frölander, Kerstin; Mårtensson, Agneta; Rashed, Foad; Annika, Wistedt; Schön, Thomas; Serrander, Lena

2014-03-19

The aim of this study was to evaluate diagnostic tests in order to introduce a diagnostic strategy to identify the most common gram-positive bacteria (pneumococci, enterococci, β-haemolytic streptococci and S. aureus) found in blood cultures within 6 hours after signalling growth. The tube coagulase test was optimized and several latex agglutination tests were compared and evaluated before a validation period of 11 months was performed on consecutive positive blood culture patient samples from Kalmar County Hospital, Sweden. During the validation period 150 (91%) of a total of 166 gram-positive cocci (119 in clusters, 45 in chains or pairs and 2 undefined morphology) were correctly identified as S. aureus, CoNS, Pneumococci, Enterococci or group A streptococci (GAS), group B streptococci (GBS), group G streptococci (GGS) within 6 hours with a minimal increase in work-load and costs. The remaining samples (9%) were correctly identified during the next day. No samples were incorrectly grouped with this diagnostic strategy and no patient came to risk by early reporting. A simple strategy gives reliable and cost-effective reporting of >90% of the most common gram-positive cocci within 6 hours after a blood cultures become positive. The high specificity of the tests used makes preliminary reports reliable. The reports can be used to indicate the focus of infection and not the least, support faster administration of proper antimicrobial treatment for patients with serious bacterial infections.

Human brucellosis among pyrexia of unknown origin cases and occupationally exposed individuals in Goa Region, India.

PubMed

Pathak, Ajay D; Dubal, Zunjar B; Doijad, Swapnil; Raorane, Abhay; Rodrigues, Savio; Naik, Rajeshwar; Naik-Gaonkar, Shraddha; Kalorey, Dewanand R; Kurkure, Nitin V; Naik, Rajesh; Barbuddhe, Sukhadeo B

2014-01-01

Brucellosis is a widespread zoonotic infection. This disease is endemic in many parts of Asia, including India. Brucellosis is a major cause of pyrexia of unknown origin (PUO). Persons exposed to infected animals or contaminated animal products are at high risk. Seropositivity among animal handlers, veterinarians and dairy workers has been documented in India. Thus, the present study was aimed to determine prevalence of brucellosis among PUO cases and occupationally exposed individuals. In this study, serum samples (n=282) from cases of pyrexia of unknown origin (PUO) (n=243), and occupationally exposed individuals (n=39) were collected and tested for brucellosis by Rose Bengal plate test (RBPT), serum agglutination test (SAT), indirect ELISA, IgG and IgM ELISA. Blood culture for isolation of Brucella was performed for 10 serologically positive patients using BACTEC 9050 automated blood culture system. Biochemical tests and PCR techniques were used for confirmation of the isolates. Of the samples tested, 4.25%, 3.54%, 6.02% and 4.96% samples were positive by RBPT, SAT, indirect ELISA and IgG ELISA, respectively. None of the sample was positive for IgM ELISA. Of the 10 blood samples cultured bacteriologically, one Brucella isolate was recovered. The isolate was confirmed as Brucella abortus. Amplification of the bcsp31 and IS711 genes was also observed. Seropositivity for brucellosis was observed among PUO cases, animal handlers and dairy workers in Goa, India. The serological tests showed variable results. One Brucella isolate was obtained by performing blood culture. Confirmation of the case was done rapidly using molecular tools. General awareness about clinical symptoms should be increased which will improve proper diagnosis within short time frame.

Mesenchymal stem cells from the Wharton's jelly of umbilical cord segments provide stromal support for the maintenance of cord blood hematopoietic stem cells during long-term ex vivo culture.

PubMed

Bakhshi, Tiki; Zabriskie, Ryan C; Bodie, Shamanique; Kidd, Shannon; Ramin, Susan; Paganessi, Laura A; Gregory, Stephanie A; Fung, Henry C; Christopherson, Kent W

2008-12-01

Hematopoietic stem cells (HSCs) are routinely obtained from marrow, mobilized peripheral blood, and umbilical cord blood. Mesenchymal stem cells (MSCs) are traditionally isolated from marrow. Bone marrow-derived MSCs (BM-MSCs) have previously demonstrated their ability to act as a feeder layer in support of ex vivo cord blood expansion. However, the use of BM-MSCs to support the growth, differentiation, and engraftment of cord blood may not be ideal for transplant purposes. Therefore, the potential of MSCs from a novel source, the Wharton's jelly of umbilical cords, to act as stromal support for the long-term culture of cord blood HSC was evaluated. Umbilical cord-derived MSCs (UC-MSCs) were cultured from the Wharton's jelly of umbilical cord segments. The UC-MSCs were then profiled for expression of 12 cell surface receptors and tested for their ability to support cord blood HSCs in a long-term culture-initiating cell (LTC-IC) assay. Upon culture, UC-MSCs express a defined set of cell surface markers (CD29, CD44, CD73, CD90, CD105, CD166, and HLA-A) and lack other markers (CD45, CD34, CD38, CD117, and HLA-DR) similar to BM-MSCs. Like BM-MSCs, UC-MSCs effectively support the growth of CD34+ cord blood cells in LTC-IC assays. These data suggest the potential therapeutic application of Wharton's jelly-derived UC-MSCs to provide stromal support structure for the long-term culture of cord blood HSCs as well as the possibility of cotransplantation of genetically identical, HLA-matched, or unmatched cord blood HSCs and UC-MSCs in the setting of HSC transplantation.

The Influence of Acute Handling Stress on Some Blood Parameters in Cultured Sea Bream (Sparus Aurata Linnaeus, 1758)

PubMed Central

Fazio, Francesco; Ferrantelli, Vincenzo; Fortino, Gianluca; Giangrosso, Giuseppe; Faggio, Caterina

2015-01-01

The effect of acute handling stress on haematological profile, blood glucose and lactate (secondary stress markers) in cultured sea bream Sparus aurata was evaluated. Sixty six Sparus aurata were used and equally divided into two groups (A and B). Group A was not subjected to stress, Group B was subjected to acute handling stress. From each fish, biometric data and blood samples were collected to evaluate haematological profile, blood glucose and lactate. Unpaired t-test Student was applied to evaluate possible differences in parameters between the two groups. Red blood cells, haematocrit, haemoglobin, white blood cells (WBC), glucose and lactate showed an increase (P<0.05) in Group B compared with Group A, while mean corpuscular volume decreased (P<0.05) in Group B. The results highlight the role of studied parameters in monitoring the stressful conditions of aquaculture production which affect animal welfare and fish products quality. PMID:27800375

Antifungal Activity of Propolis Against Yeasts Isolated From Blood Culture: In Vitro Evaluation.

PubMed

Mutlu Sariguzel, Fatma; Berk, Elife; Koc, Ayes Nedret; Sav, Hafize; Demir, Gonca

2016-09-01

Due to the failure of available antifungal agents in the treatment of candidemia and the toxic activities of these drugs, a lot of researches are being conducted to develop new nontoxic and effective antifungal agents for optimal control of fungal pathogens. The aim of this study is to evaluate the in vitro antifungal activity of propolis against yeasts isolated from the blood cultures of intensive care unit patients. Seventy-six strains were included in this study. The in vitro antifungal activity of propolis, fluconazole (FLU), and itraconazole (ITR) was investigated by the microdilution broth methods (CLSI guidelines M27-A3 for yeast). The propolis sample was collected from Kayseri, Turkey. Of the 76 isolates, 33 were identified as Candida albicans while 37 were C. parapsilosis, three were C. tropicalis, and three were identified as C. glabrata. The geometric mean range for MIC (μg/ml) with regard to all isolates was 0.077 to 3 μg/ml for FLU and ITR, and 0.375 to 0.70 μg/ml for propolis. It was shown that propolis had significant antifungal activity against all Candida strains and the MIC range of propolis was determined as 0185 to 3 μg/ml. This study demonstrated that propolis had significant antifungal activity against yeasts isolated from blood culture compared with FLU and ITR. The propolis MIC in azole-resistant strains such as C. glabrata was found lower than the FLU MIC. © 2015 Wiley Periodicals, Inc.

Bacterial rapid identification with matrix assisted laser desorption/ionization time-of-flight mass spectrometry: development of an 'in-house method' and comparison with Bruker Sepsityper(®) kit.

PubMed

Frédéric Ric, S; Antoine, M; Bodson, A; Lissoir, B

2015-10-01

The objective of this study was to compare an in-house matrix-assisted laser desorption ionization with time of flight (MALDI-TOF) method and a commercial MALDI-TOF kit (Sepsityper(®) kit) for direct bacterial identification in positive blood cultures. We also evaluated the time saved and the cost associated with the rapid identification techniques. We used the BACTEC(®) automated system for detecting positive blood cultures. Direct identification using Sepsityper kit and the in-house method were compared with conventional identification by MALDI-TOF using pure bacterial culture on the solid phase. We also evaluated different cut-off scores for rapid bacterial identification. In total, 127 positive blood vials were selected. The rate of rapid identification with the MALDI Sepsityper kit was 25.2% with the standard cut-off and 33.9% with the enlarged cut-off, while the results for the in-house method were 44.1 and 61.4%, respectively. Error rates with the enlarged cut-off were 6.98 (n = 3) and 2.56% (n = 2) for Sepsityper and the in-house method, respectively. Identification rates were higher for gram-negative bacteria. Direct bacterial identification succeeded in supplying rapid identification of the causative organism in cases of sepsis. The time taken to obtain a result was nearly 24  hours shorter for the direct bacterial identification methods than for conventional MALDI-TOF on solid phase culture. Compared with the Sepsityper kit, the in-house method offered better results and fewer errors, was more cost-effective and easier to use.

NanoFlares for the detection, isolation, and culture of live tumor cells from human blood.

PubMed

Halo, Tiffany L; McMahon, Kaylin M; Angeloni, Nicholas L; Xu, Yilin; Wang, Wei; Chinen, Alyssa B; Malin, Dmitry; Strekalova, Elena; Cryns, Vincent L; Cheng, Chonghui; Mirkin, Chad A; Thaxton, C Shad

2014-12-02

Metastasis portends a poor prognosis for cancer patients. Primary tumor cells disseminate through the bloodstream before the appearance of detectable metastatic lesions. The analysis of cancer cells in blood—so-called circulating tumor cells (CTCs)—may provide unprecedented opportunities for metastatic risk assessment and investigation. NanoFlares are nanoconstructs that enable live-cell detection of intracellular mRNA. NanoFlares, when coupled with flow cytometry, can be used to fluorescently detect genetic markers of CTCs in the context of whole blood. They allow one to detect as few as 100 live cancer cells per mL of blood and subsequently culture those cells. This technique can also be used to detect CTCs in a murine model of metastatic breast cancer. As such, NanoFlares provide, to our knowledge, the first genetic-based approach for detecting, isolating, and characterizing live cancer cells from blood and may provide new opportunities for cancer diagnosis, prognosis, and personalized therapy.

NanoFlares for the detection, isolation, and culture of live tumor cells from human blood

PubMed Central

Halo, Tiffany L.; McMahon, Kaylin M.; Angeloni, Nicholas L.; Xu, Yilin; Wang, Wei; Chinen, Alyssa B.; Malin, Dmitry; Strekalova, Elena; Cryns, Vincent L.; Cheng, Chonghui; Mirkin, Chad A.; Thaxton, C. Shad

2014-01-01

Metastasis portends a poor prognosis for cancer patients. Primary tumor cells disseminate through the bloodstream before the appearance of detectable metastatic lesions. The analysis of cancer cells in blood—so-called circulating tumor cells (CTCs)—may provide unprecedented opportunities for metastatic risk assessment and investigation. NanoFlares are nanoconstructs that enable live-cell detection of intracellular mRNA. NanoFlares, when coupled with flow cytometry, can be used to fluorescently detect genetic markers of CTCs in the context of whole blood. They allow one to detect as few as 100 live cancer cells per mL of blood and subsequently culture those cells. This technique can also be used to detect CTCs in a murine model of metastatic breast cancer. As such, NanoFlares provide, to our knowledge, the first genetic-based approach for detecting, isolating, and characterizing live cancer cells from blood and may provide new opportunities for cancer diagnosis, prognosis, and personalized therapy. PMID:25404304

Rapid presumptive identification of the Mycobacterium tuberculosis-bovis complex by radiometric determination of heat stable urease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gandy, J.H.; Pruden, E.L.; Cox, F.R.

1983-12-01

Simple and rapid Bactec methodologies for the determination of neat (unaltered) and heat stable urease activity of mycobacteria are presented. Clinical isolates (63) and stock cultures (32)--consisting of: M. tuberculosis (19), M. bovis (5), M. kansasii (15), M. marinum (4), M. simiae (3), M. scrofulaceum (16), M. gordonae (6), M. szulgai (6), M. flavescens (1), M. gastri (1), M. intracellulare (6), M. fortuitum-chelonei complex (12), and M. smegmatis (1)--were tested for neat urease activity by Bactec radiometry. Mycobacterial isolates (50-100 mg wet weight) were incubated at 35 degrees C for 30 minutes with microCi14C-urea. Urease-positive mycobacteria gave Bactec growth indexmore » (GI) values greater than 100 units, whereas urease-negative species gave values less than 10 GI units. Eighty-three isolates possessing neat urease activity were heated at 80 degrees C for 30 minutes followed by incubation at 35 degrees C for 30 minutes with 1 microCi14C-urea. Mycobacterium tuberculosis-bovis complex demonstrated heat-stable urease activity (GI more than 130 units) and could be distinguished from mycobacteria other than tuberculosis (MOTT), which gave GI values equal to or less than 40 units.« less

Improved Clinical Outcomes in Patients with Positive Blood Culture by Proactive Intervention of Antimicrobial Use-Impact of Antimicrobial Stewardship Program Implementation on Clinical Practice.

PubMed

Yamada, Takehiro; Kagami, Keisuke; Imai, Shungo; Akizawa, Koji; Iwasaki, Sumio; Fukumoto, Tatsuya; Ishiguro, Nobuhisa; Iseki, Ken

2017-01-01

Bacteremia is one of the most serious infectious illness resulting from nosocomial infection. Therefore, appropriate antimicrobial chemotherapy should be provided as soon as possible to patients exhibiting symptoms of infectious disease and having positive blood culture results. Antimicrobial stewardship (AS) guidelines were recently released by the Infectious Diseases Society of America. The guidelines recommend "proactive intervention and feedback" as one of the core strategies for implementing optimal antimicrobial drug use to improve patient outcomes in clinical settings. We began using the AS program for optimizing antimicrobial chemotherapy in patients with positive blood culture results. The results of blood cultures and antimicrobial prescriptions for the corresponding patients were daily reviewed by a pharmacist and a physician, members of the infection control team (ICT). If the antimicrobial agents selected were inappropriate, ICT made a recommendation to the attending physicians who prescribed the antibiotics. To evaluate the outcomes of this program, we conducted a single-center, retrospective investigation for near a hundred of patients who underwent intervention by infection-control physician and pharmacist. Resolution of bacteremia (determined by blood culture results) was 96.3% in the group that accepted intervention, whereas only 16.7% of the cases resolved in the group that did not accept intervention. These results strongly suggest the importance of the infection disease-specialist team intervention. This program could become an important method for improving clinical outcomes in patients with bacteremia.

Evaluation of Antigen-Specific IgM and IgG Production during an In Vitro Peripheral Blood Mononuclear Cell Culture Assay.

PubMed

Matsuda, Yoshiko; Imamura, Ryoichi; Takahara, Shiro

2017-01-01

The recent attention given to diseases associated with memory B-cell (mBC)-produced antibodies (Abs) suggests the need for a similar in vitro assay to evaluate the functions of mBCs. Here, we cultured peripheral blood mononuclear cells (PBMCs) with the intent to collect mBC-derived Abs in vitro and maintain their cell-cell contact-dependent interactions with helper T-cells. PBMCs were cultured with interleukin (IL)-21, CpG-oligodeoxynucleotides (ODN), phorbol myristate acetate (PMA), and phytohemagglutinin/leucoagglutinin (PHA-L) in 24-well flat-bottom plates (5 × 10 5  cells/well). A culture supernatant analysis of PBMCs from healthy donors ( n  = 10) indicated that antigen-specific IgM Ab levels in a PBMC culture supernatant might be better able to demonstrate the antigen sensitization status in a smaller peripheral blood sample, compared to IgG because Epstein-Barr virus-specific IgM mBCs circulate peripherally at a significantly higher frequency once antiviral humoral immunity has stabilized. Thus, our in vitro assay demonstrated the potential significance of antigen-specific IgM Ab production in the culture supernatants. Furthermore, an analysis of cultured PBMCs from allograft kidney recipients ( n  = 16) sensitized with de novo donor-specific human leukocyte antigen (HLA)-specific Abs (DSAs) showed that IgM-type HLA-specific Abs were detected mainly from the culture supernatants from PBMCs of patients with stable graft function, whereas IgG isotype HLA Abs were detectable only from patients with biopsy-proven antibody-mediated rejection. In other words, these IgG isotype Abs also represented an activated humoral immune response in vivo . Additionally, IgM- and IgG-expressing mBCs from healthy donors ( n  = 5) were cultured with IL-21, CpG-ODN, and a supernatant produced by stimulating CD19 + B-cell-depleted PBMCs with PHA-L and PMA in 24-well flat-bottom plates (1 × 10 5  cells/well), and the resulting in vitro analysis provided

Rapid method for direct identification of bacteria in urine and blood culture samples by matrix-assisted laser desorption ionization time-of-flight mass spectrometry: intact cell vs. extraction method.

PubMed

Ferreira, L; Sánchez-Juanes, F; Muñoz-Bellido, J L; González-Buitrago, J M

2011-07-01

Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a fast and reliable technology for the identification of microorganisms with proteomics approaches. Here, we compare an intact cell method and a protein extraction method before application on the MALDI plate for the direct identification of microorganisms in both urine and blood culture samples from clinical microbiology laboratories. The results show that the intact cell method provides excellent results for urine and is a good initial method for blood cultures. The extraction method complements the intact cell method, improving microorganism identification from blood culture. Thus, we consider that MALDI-TOF MS performed directly on urine and blood culture samples, with the protocols that we propose, is a suitable technique for microorganism identification, as compared with the routine methods used in the clinical microbiology laboratory. © 2010 The Authors. Clinical Microbiology and Infection © 2010 European Society of Clinical Microbiology and Infectious Diseases.

[Tuberculous meningoencephalitis--own observations].

PubMed

Garlicki, Aleksander; Kluba-Wojewoda, Urszula; Bociaga-Jasik, Monika; Kalinowska-Nowak, Anna; Sobczyk-Krupiarz, Iwona; Mach, Tomasz

2003-01-01

Nine cases of the tuberculous meningoencephalitis in adult men and women treated in years 1993-2000 have been presented. The diagnosis was established on the basis of clinical picture and examination of the cerebrospinal fluid (CSF). Routine analysis of CSF was done, as well as the microscopic examination, biological assay, and culture on the Lowenstein-Jensena medium and using Bactec 460 TB system. Only in one case Mycobacterium tuberculosis was found in the smear of CSF stained by Ziehl-Nielsen method. In one case the biologic assay was positive and Mycobacterium tuberculosis was cultured on Lowenstein-Jensen medium. In another 4 cases the pathogen was detected by Bactec 460 TB technique. In 4 cases Mycobacterium tuberculosis was also cultured from the secretion of the respiratory tract, and in one patient urine. On chest X-ray characteristic changes for pulmonary tuberculosis were observed in 5 cases (56%). Seven patients (78%) had hydrocephalus detected on CT scan of the head. One patient died, one (11%) developed persistent complication in form of spastic paralysis of lower extremities and sphincters dysfunction. Seven patients (78%) recovered completely after the 12 months therapy with standard chemotherapeutic regimen for tuberculosis.

Design of a Tablet Computer App for Facilitation of a Molecular Blood Culture Test in Clinical Microbiology and Preliminary Usability Evaluation.

PubMed

Samson, Lasse L; Pape-Haugaard, Louise; Meltzer, Michelle C; Fuchs, Martin; Schønheyder, Henrik C; Hejlesen, Ole

2016-03-18

User mobility is an important aspect of the development of clinical information systems for health care professionals. Mobile phones and tablet computers have obtained widespread use by health care professionals, offering an opportunity for supporting the access to patient information through specialized applications (apps) while supporting the mobility of the users. The use of apps for mobile phones and tablet computers may support workflow of complex tasks, for example, molecular-based diagnostic tests in clinical microbiology. Multiplex Blood Culture Test (MuxBCT) is a molecular-based diagnostic test used for rapid identification of pathogens in positive blood cultures. To facilitate the workflow of the MuxBCT, a specialized tablet computer app was developed as an accessory to the diagnostic test. The app aims to reduce the complexity of the test by step-by-step guidance of microscopy and to assist users in reaching an exact bacterial or fungal diagnosis based on blood specimen observations and controls. Additionally, the app allows for entry of test results, and communication thereof to the laboratory information system (LIS). The objective of the study was to describe the design considerations of the MuxBCT app and the results of a preliminary usability evaluation. The MuxBCT tablet app was developed and set up for use in a clinical microbiology laboratory. A near-live simulation study was conducted in the clinical microbiology laboratory to evaluate the usability of the MuxBCT app. The study was designed to achieve a high degree of realism as participants carried out a scenario representing the context of use for the MuxBCT app. As the MuxBCT was under development, the scenario involved the use of molecular blood culture tests similar to the MuxBCT for identification of microorganisms from positive blood culture samples. The study participants were observed, and their interactions with the app were recorded. After the study, the participants were debriefed to

Design of a Tablet Computer App for Facilitation of a Molecular Blood Culture Test in Clinical Microbiology and Preliminary Usability Evaluation

PubMed Central

Pape-Haugaard, Louise; Meltzer, Michelle C; Fuchs, Martin; Schønheyder, Henrik C; Hejlesen, Ole

2016-01-01

Background User mobility is an important aspect of the development of clinical information systems for health care professionals. Mobile phones and tablet computers have obtained widespread use by health care professionals, offering an opportunity for supporting the access to patient information through specialized applications (apps) while supporting the mobility of the users. The use of apps for mobile phones and tablet computers may support workflow of complex tasks, for example, molecular-based diagnostic tests in clinical microbiology. Multiplex Blood Culture Test (MuxBCT) is a molecular-based diagnostic test used for rapid identification of pathogens in positive blood cultures. To facilitate the workflow of the MuxBCT, a specialized tablet computer app was developed as an accessory to the diagnostic test. The app aims to reduce the complexity of the test by step-by-step guidance of microscopy and to assist users in reaching an exact bacterial or fungal diagnosis based on blood specimen observations and controls. Additionally, the app allows for entry of test results, and communication thereof to the laboratory information system (LIS). Objective The objective of the study was to describe the design considerations of the MuxBCT app and the results of a preliminary usability evaluation. Methods The MuxBCT tablet app was developed and set up for use in a clinical microbiology laboratory. A near-live simulation study was conducted in the clinical microbiology laboratory to evaluate the usability of the MuxBCT app. The study was designed to achieve a high degree of realism as participants carried out a scenario representing the context of use for the MuxBCT app. As the MuxBCT was under development, the scenario involved the use of molecular blood culture tests similar to the MuxBCT for identification of microorganisms from positive blood culture samples. The study participants were observed, and their interactions with the app were recorded. After the study, the

Rapid identification of moulds and arthroconidial yeasts from positive blood cultures by MALDI-TOF mass spectrometry.

PubMed

de Almeida, João N; Sztajnbok, Jaques; da Silva, Afonso Rafael; Vieira, Vinicius Adriano; Galastri, Anne Layze; Bissoli, Leandro; Litvinov, Nadia; Del Negro, Gilda Maria Barbaro; Motta, Adriana Lopes; Rossi, Flávia; Benard, Gil

2016-11-01

Moulds and arthroconidial yeasts are potential life-threatening agents of fungemia in immunocompromised patients. Fast and accurate identification (ID) of these pathogens hastens initiation of targeted antifungal therapy, thereby improving the patients' prognosis. We describe a new strategy that enabled the identification of moulds and arthroconidial yeasts directly from positive blood cultures by MALDI-TOF mass spectrometry (MS). Positive blood cultures (BCs) with Gram staining showing hyphae and/or arthroconidia were prospectively selected and submitted to an in-house protein extraction protocol. Mass spectra were obtained by Vitek MS™ system, and identifications were carried out with in the research use only (RUO) mode with an extended database (SARAMIS™ [v.4.12] plus in-house database). Fusarium solani, Fusarium verticillioides, Exophiala dermatitidis, Saprochaete clavata, and Trichosporon asahii had correct species ID by MALDI-TOF MS analysis of positive BCs. All cases were related to critically ill patients with high mortality fungemia and direct ID from positive BCs was helpful for rapid administration of targeted antifungal therapy. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Long-term culture of chicken primordial germ cells isolated from embryonic blood and production of germline chimaeric chickens.

PubMed

Naito, Mitsuru; Harumi, Takashi; Kuwana, Takashi

2015-02-01

Production of germline chimaeric chickens by the transfer of cultured primordial germ cells (PGC) is a useful system for germline manipulation. A novel culture system was developed for chicken PGC isolated from embryonic blood. The isolated PGC were cultured on feeder cells derived from chicken embryonic fibroblast. The cultured PGC formed colonies and they proliferated about 300-times during the first 30 days. The cultured PGC retained the ability to migrate to recipient gonads and were also chicken VASA homologue (CVH)-positive. Female PGC were present in the mixed-sex PGC populations cultured for more than 90 days and gave rise to viable offspring efficiently via germline chimaeric chickens. Male cultured PGC were transferred to recipient embryos and produced putative chimaeric chickens. The DNA derived from the cultured PGC was detected in the sperm samples of male putative chimaeric chickens, but no donor derived offspring were obtained. Donor-derived offspring were also obtained from germline chimaeric chickens by the transfer of frozen-thawed cultured PGC. The culture method for PGC developed in the present study is useful for manipulation of the germline in chickens, such as preservation of genetic resources and gene transfer. Copyright © 2014 Elsevier B.V. All rights reserved.

Mesenchymal stem cells from the Wharton’s jelly of umbilical cord segments provide stromal support for the maintenance of cord blood hematopoietic stem cells during long-term ex vivo culture

PubMed Central

Bakhshi, Tiki; Zabriskie, Ryan C.; Bodie, Shamanique; Kidd, Shannon; Ramin, Susan; Paganessi, Laura A.; Gregory, Stephanie A.; Fung, Henry C.; Christopherson, Kent W.

2012-01-01

BACKGROUND Hematopoietic stem cells (HSCs) are routinely obtained from marrow, mobilized peripheral blood, and umbilical cord blood. Mesenchymal stem cells (MSCs) are traditionally isolated from marrow. Bone marrow–derived MSCs (BM-MSCs) have previously demonstrated their ability to act as a feeder layer in support of ex vivo cord blood expansion. However, the use of BM-MSCs to support the growth, differentiation, and engraftment of cord blood may not be ideal for transplant purposes. Therefore, the potential of MSCs from a novel source, the Wharton’s jelly of umbilical cords, to act as stromal support for the long-term culture of cord blood HSC was evaluated. STUDY DESIGN AND METHODS Umbilical cord–derived MSCs (UC-MSCs) were cultured from the Wharton’s jelly of umbilical cord segments. The UC-MSCs were then profiled for expression of 12 cell surface receptors and tested for their ability to support cord blood HSCs in a long-term culture-initiating cell (LTC-IC) assay. RESULTS Upon culture, UC-MSCs express a defined set of cell surface markers (CD29, CD44, CD73, CD90, CD105, CD166, and HLA-A) and lack other markers (CD45, CD34, CD38, CD117, and HLA-DR) similar to BM-MSCs. Like BM-MSCs, UC-MSCs effectively support the growth of CD34+ cord blood cells in LTC-IC assays. CONCLUSION These data suggest the potential therapeutic application of Wharton’s jelly–derived UC-MSCs to provide stromal support structure for the long-term culture of cord blood HSCs as well as the possibility of cotransplantation of genetically identical, HLA-matched, or unmatched cord blood HSCs and UC-MSCs in the setting of HSC transplantation. PMID:18798803

Acceptability and feasibility of a culturally tailored Internet-delivered intervention to promote blood donation in Blacks.

PubMed

Robbins, Mark L; Paiva, Andrea L; Amoyal, Nicole R; Brick, Leslie; Kessler, Debra A; Burditt, Caitlin; Caltabiano, Melinda; Shaz, Beth H

2015-03-01

A pilot test of a computer-tailored intervention designed to promote blood donation among Blacks was conducted. Intervention content, based on the transtheoretical model, offered participants individually and culturally tailored information on blood donation with emphasis on need specific to race (e.g., sickle-cell disease). Black adults (N = 150) with a diversity of blood donation experience were recruited from a blood center and a survey recruitment website. Posttest assessment included a 14-item evaluation and transtheoretical model questions. Participants rated the program positively (81.3% to 98.7% of participants agreeing or strongly agreeing with evaluation items). For example, 98.7% of respondents reported that the program gave sound advice and that personal feedback was easily understood, and 87.3% felt the program was designed for people like themselves. Ninety-five percent of participants reported that they would recommend the program to others. There were no significant differences in ratings based on demographics. Qualitative responses support program acceptability. Furthermore, pre- and postprogram assessments indicated an increase in intention to donate, t(149) = 3.56, p = .001, d = .29. With acceptability and feasibility confirmed, the next steps are to test efficacy and cost-effectiveness for use to increase blood donation, particularly in priority populations. © 2014 Society for Public Health Education.

Quantitative interpretations of Visible-NIR reflectance spectra of blood.

PubMed

Serebrennikova, Yulia M; Smith, Jennifer M; Huffman, Debra E; Leparc, German F; García-Rubio, Luis H

2008-10-27

This paper illustrates the implementation of a new theoretical model for rapid quantitative analysis of the Vis-NIR diffuse reflectance spectra of blood cultures. This new model is based on the photon diffusion theory and Mie scattering theory that have been formulated to account for multiple scattering populations and absorptive components. This study stresses the significance of the thorough solution of the scattering and absorption problem in order to accurately resolve for optically relevant parameters of blood culture components. With advantages of being calibration-free and computationally fast, the new model has two basic requirements. First, wavelength-dependent refractive indices of the basic chemical constituents of blood culture components are needed. Second, multi-wavelength measurements or at least the measurements of characteristic wavelengths equal to the degrees of freedom, i.e. number of optically relevant parameters, of blood culture system are required. The blood culture analysis model was tested with a large number of diffuse reflectance spectra of blood culture samples characterized by an extensive range of the relevant parameters.

Comparative analysis of Gram's stain, PNA-FISH and Sepsityper with MALDI-TOF MS for the identification of yeast direct from positive blood cultures.

PubMed

Gorton, Rebecca L; Ramnarain, P; Barker, K; Stone, N; Rattenbury, S; McHugh, T D; Kibbler, C C

2014-10-01

Fungaemia diagnosis could be improved by reducing the time to identification of yeast from blood cultures. This study aimed to evaluate three rapid methods for the identification of yeast direct from blood cultures; Gram's stain analysis, the AdvanDX Peptide Nucleic Acid in Situ Hybridisation Yeast Traffic Light system (PNA-FISH YTL) and Bruker Sepsityper alongside matrix-assisted laser desorption ionisation time of flight mass spectrometry (MALDI-TOF MS). Fifty blood cultures spiked with a known single yeast strain were analysed by blinded operators experienced in each method. Identifications were compared with MALDI-TOF MS CHROMagar Candida culture and ITS rRNA sequence-based identifications. On first attempt, success rates of 96% (48/50) and 76% (36/50) were achieved using PNA-FISH YTL and Gram's stain respectively. MALDI-TOF MS demonstrated a success rate of 56% (28/50) when applying manufacturer's species log score thresholds and 76% (38/50) using in-house parameters, including lowering the species log score threshold to >1.5. In conclusion, PNA-FISH YTL demonstrated a high success rate successfully identifying yeast commonly encountered in fungaemia. Sepsityper(™) with MALDI-TOF MS was accurate but increased sensitivity is required. Due to the misidentification of commonly encountered yeast Gram's stain analysis demonstrated limited utility in this setting. © 2014 Blackwell Verlag GmbH.

Direct identification of microorganisms from positive blood cultures using the lysis-filtration technique and matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS): a multicentre study.

PubMed

Farina, Claudio; Arena, Fabio; Casprini, Patrizia; Cichero, Paola; Clementi, Massimo; Cosentino, Marina; Degl'Innocenti, Roberto; Giani, Tommaso; Luzzaro, Francesco; Mattei, Romano; Mauri, Carola; Nardone, Maria; Rossolini, Gian Maria; Serna Ortega, Paula Andrea; Vailati, Francesca

2015-04-01

Microbial identification from blood cultures is essential to institute optimal antibiotic therapy and improve survival possibilities. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been successfully applied to identify bacteria and yeasts from positive blood cultures broths. The aim of this multicentre study was to evaluate the reliability of the lysis-filtration technique associated with MALDI-TOF MS to directly identify microorganisms from 765 positive blood cultures collected in six Italian hospitals. Overall, 675/765 (78.1%) blood isolates were correctly identified at the species level, with significant differences between Gram-negative and Gram-positive bacteria (92.6%, and 69.8%, respectively). Some difficulties arise in identifying Streptococcus pneumoniae, Staphylococcus aureus, yeasts and anaerobes. The lysis-filtration protocol is a suitable procedure in terms of performance in identifying microorganisms, but it is quite expensive and technically time-consuming since the time of filtration is not regular for all the samples. The application of the MALDI-TOF MS technique to the direct microbial identification from positive blood cultures is a very promising approach, even if more experience must be gained to minimize errors and costs.

Beyond Blood Culture and Gram Stain Analysis: A Review of Molecular Techniques for the Early Detection of Bacteremia in Surgical Patients.

PubMed

Scerbo, Michelle H; Kaplan, Heidi B; Dua, Anahita; Litwin, Douglas B; Ambrose, Catherine G; Moore, Laura J; Murray, Col Clinton K; Wade, Charles E; Holcomb, John B

2016-06-01

Sepsis from bacteremia occurs in 250,000 cases annually in the United States, has a mortality rate as high as 60%, and is associated with a poorer prognosis than localized infection. Because of these high figures, empiric antibiotic administration for patients with systemic inflammatory response syndrome (SIRS) and suspected infection is the second most common indication for antibiotic administration in intensive care units (ICU)s. However, overuse of empiric antibiotics contributes to the development of opportunistic infections, antibiotic resistance, and the increase in multi-drug-resistant bacterial strains. The current method of diagnosing and ruling out bacteremia is via blood culture (BC) and Gram stain (GS) analysis. Conventional and molecular methods for diagnosing bacteremia were reviewed and compared. The clinical implications, use, and current clinical trials of polymerase chain reaction (PCR)-based methods to detect bacterial pathogens in the blood stream were detailed. BC/GS has several disadvantages. These include: some bacteria do not grow in culture media; others do not GS appropriately; and cultures can require up to 5 d to guide or discontinue antibiotic treatment. PCR-based methods can be potentially applied to detect rapidly, accurately, and directly microbes in human blood samples. Compared with the conventional BC/GS, particular advantages to molecular methods (specifically, PCR-based methods) include faster results, leading to possible improved antibiotic stewardship when bacteremia is not present.

Novel, high-yield red blood cell production methods from CD34-positive cells derived from human embryonic stem, yolk sac, fetal liver, cord blood, and peripheral blood.

PubMed

Olivier, Emmanuel; Qiu, Caihong; Bouhassira, Eric E

2012-08-01

The current supply of red blood cells expressing rare blood groups is not sufficient to cover all the existing transfusion needs for chronically transfused patients, such as sickle cell disease homozygous carriers, because of alloimmunization. In vitro production of cultured red blood cells is slowly emerging as a possible complement to the existing collection-based red blood cell procurement system. The yield of cultured red blood cells can theoretically be maximized by amplifying the stem, progenitor, or precursor compartment. Here, we combined methods designed to expand these three compartments to optimize the yield of cultured red blood cells and found that exposing CD34(+) cells to a short pulse of cytokines favorable for erythroid differentiation prior to stem cell expansion followed by progenitor expansion produced the highest yield of erythroid cells. This novel serum-free red blood cell production protocol was efficient on CD34(+) cells derived from human embryonic stem cells, 6-8-week yolk sacs, 16-18-week fetal livers, cord blood, and peripheral blood. The yields of cells obtained with these new protocols were larger by an order of magnitude than the yields observed previously. Globin expression analysis by high-performance liquid chromatography revealed that these expansion protocols generally yielded red blood cells that expressed a globin profile similar to that expected for the developmental age of the CD34(+) cells.

Does neutralization of gastric aspirates from children with suspected intrathoracic tuberculosis affect mycobacterial yields on MGIT culture?

PubMed

Parashar, Deepak; Kabra, Sushil K; Lodha, Rakesh; Singh, Varinder; Mukherjee, Aparna; Arya, Tina; Grewal, Harleen M S; Singh, Sarman

2013-06-01

The microbiological confirmation of pulmonary tuberculosis in children relies on cultures of gastric aspirate (GA) specimens. Conventionally, GAs are neutralized to improve culture yields of mycobacteria. However, there are limited data to support this practice. To study the utility of neutralization of GAs with sodium bicarbonate in children with intrathoracic tuberculosis, a total of 116 children of either sex, aged 6 months to 14 years (median age, 120 months; interquartile range [IQR], 7 to 192 months), underwent gastric aspiration on 2 consecutive days. Gastric aspirates were divided into two aliquots, and only one aliquot was neutralized with 1% sodium bicarbonate. Both aliquots were processed for smear and culture examinations. Out of the 232 gastric aspirates, 12 (5.17%) were acid-fast bacilli (AFB) smear positive. There were no differences in smear positivity rates from samples with or without neutralization. The yield of Mycobacterium tuberculosis on a Bactec MGIT 960 culture system was significantly lower in the neutralized samples (16.3% [38/232]) than in the nonneutralized samples (21.5% [50/232]) (P = 0.023). There was no significant difference between the neutralized and the nonneutralized samples in time to detection using the MGIT 960 system (average, 24.6 days; IQR, 12 to 37 days) (P = 0.9). The contamination rates were significantly higher in the neutralized samples than in the nonneutralized samples (17.2% [40/232] versus 3.9% [9/232]) (P = 0.001). The agreement for positive mycobacterial culture between the two approaches was 66.5% (P = 0.001). Hence, we recommend that gastric aspirate samples not be neutralized with sodium bicarbonate prior to culture for M. tuberculosis.

Effectiveness of practices to reduce blood culture contamination: A Laboratory Medicine Best Practices systematic review and meta-analysis☆

PubMed Central

Snyder, Susan R.; Favoretto, Alessandra M.; Baetz, Rich Ann; Derzon, James H.; Madison, Bereneice M.; Mass, Diana; Shaw, Colleen S.; Layfield, Christopher D.; Christenson, Robert H.; Liebow, Edward B.

2015-01-01

Objectives This article is a systematic review of the effectiveness of three practices for reducing blood culture contamination rates: venipuncture, phlebotomy teams, and prepackaged preparation/collection (prep) kits. Design and methods The CDC-funded Laboratory Medicine Best Practices Initiative systematic review methods for quality improvement practices were used. Results Studies included as evidence were: 9 venipuncture (vs. versus intravenous catheter), 5 phlebotomy team; and 7 prep kit. All studies for venipuncture and phlebotomy teams favored these practices, with meta-analysis mean odds ratios for venipuncture of 2.69 and phlebotomy teams of 2.58. For prep kits 6 studies’ effect sizes were not statistically significantly different from no effect (meta-analysis mean odds ratio 1.12). Conclusions Venipuncture and the use of phlebotomy teams are effective practices for reducing blood culture contamination rates in diverse hospital settings and are recommended as evidence-based “best practices” with high overall strength of evidence and substantial effect size ratings. No recommendation is made for or against prep kits based on uncertain improvement. PMID:22709932

Effectiveness of practices to reduce blood culture contamination: a Laboratory Medicine Best Practices systematic review and meta-analysis.

PubMed

Snyder, Susan R; Favoretto, Alessandra M; Baetz, Rich Ann; Derzon, James H; Madison, Bereneice M; Mass, Diana; Shaw, Colleen S; Layfield, Christopher D; Christenson, Robert H; Liebow, Edward B

2012-09-01

This article is a systematic review of the effectiveness of three practices for reducing blood culture contamination rates: venipuncture, phlebotomy teams, and prepackaged preparation/collection (prep) kits. The CDC-funded Laboratory Medicine Best Practices Initiative systematic review methods for quality improvement practices were used. Studies included as evidence were: 9 venipuncture (vs. versus intravenous catheter), 5 phlebotomy team; and 7 prep kit. All studies for venipuncture and phlebotomy teams favored these practices, with meta-analysis mean odds ratios for venipuncture of 2.69 and phlebotomy teams of 2.58. For prep kits 6 studies' effect sizes were not statistically significantly different from no effect (meta-analysis mean odds ratio 1.12). Venipuncture and the use of phlebotomy teams are effective practices for reducing blood culture contamination rates in diverse hospital settings and are recommended as evidence-based "best practices" with high overall strength of evidence and substantial effect size ratings. No recommendation is made for or against prep kits based on uncertain improvement. Copyright © 2012 The Canadian Society of Clinical Chemists. All rights reserved.

Blood culture contamination with Enterococci and skin organisms: implications for surveillance definitions of primary bloodstream infections.

PubMed

Freeman, Joshua T; Chen, Luke Francis; Sexton, Daniel J; Anderson, Deverick J

2011-06-01

Enterococci are a common cause of bacteremia but are also common contaminants. In our institution, approximately 17% of positive blood cultures with enterococci are mixed with skin organisms. Such isolates are probable contaminants. The specificity of the current definition of primary bloodstream infection could be increased by excluding enterococci mixed with skin organisms. Copyright © 2011 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Mosby, Inc. All rights reserved.

Tooth Tissue Engineering: The Importance of Blood Products as a Supplement in Tissue Culture Medium for Human Pulp Dental Stem Cells.

PubMed

Pisciolaro, Ricardo Luiz; Duailibi, Monica Talarico; Novo, Neil Ferreira; Juliano, Yara; Pallos, Debora; Yelick, Pamela Crotty; Vacanti, Joseph Phillip; Ferreira, Lydia Masako; Duailibi, Silvio Eduardo

2015-11-01

One of the goals in using cells for tissue engineering (TE) and cell therapy consists of optimizing the medium for cell culture. The present study compares three different blood product supplements for improved cell proliferation and protection against DNA damage in cultured human dental pulp stem cells for tooth TE applications. Human cells from dental pulp were first characterized as adult stem cells (ectomesenchymal mixed origin) by flow cytometry. Next, four different cell culture conditions were tested: I, supplement-free; II, supplemented with fetal bovine serum; III, allogeneic human serum; and IV, autologous human serum. Cultured cells were then characterized for cell proliferation, mineralized nodule formation, and colony-forming units (CFU) capability. After 28 days in culture, the comet assay was performed to assess possible damage in cellular DNA. Our results revealed that Protocol IV achieved higher cell proliferation than Protocol I (p = 0.0112). Protocols II and III resulted in higher cell proliferation than Protocol I, but no statistical differences were found relative to Protocol IV. The comet assay revealed less cell damage in cells cultured using Protocol IV as compared to Protocols II and III. The damage percentage observed on Protocol II was significantly higher than all other protocols. CFUs capability was highest using Protocol IV (p = 0.0018) and III, respectively, and the highest degree of mineralization was observed using Protocol IV as compared to Protocols II and III. Protocol IV resulted in significantly improved cell proliferation, and no cell damage was observed. These results demonstrate that human blood product supplements can be used as feasible supplements for culturing adult human dental stem cells.

Organizational safety culture/climate and worker compliance with hazardous drug guidelines: lessons from the blood-borne pathogen experience.

PubMed

McDiarmid, Melissa A; Condon, Marian

2005-07-01

The health risks posed to health care workers (HCW) handling antineoplastic and other hazardous drugs (HDs) are well established. However, despite nearly 20 years of professional practice standards, compliance with safe handling procedures is poor. We present documentation of undercompliance with recommended safety procedures for HDs. Then, we examine a similar problem, HCW compliance with blood-borne pathogen universal precautions (UP) and its partial solution tied to the strength of a facility's safety culture. Lessons learned here may be applicable to the HD issue. It is proposed that analyzing a facility's safety culture may enlarge our understanding of the barriers contributing to HD under-compliance and suggest strategies to improve it. The Safety Culture paradigm offers many targets for intervention to enhance and promote worker compliance with safe HD handling practices thus mitigating internal exposure.

Mouse preantral follicle growth in 3D co-culture system using human menstrual blood mesenchymal stem cell.

PubMed

Rajabi, Zahra; Yazdekhasti, Hossein; Noori Mugahi, Seyed Mohammad Hossein; Abbasi, Mehdi; Kazemnejad, Somaieh; Shirazi, Abolfazl; Majidi, Masoumeh; Zarnani, Amir-Hassan

2018-03-01

Follicle culture provides a condition which can help investigators to evaluate various aspects of ovarian follicle growth and development and impact of different components and supplementations as well as presumably application of follicle culture approach in fertility preservation procedures. Mesenchymal Stem Cells (MSCs), particularly those isolated from menstrual blood has the potential to be used as a tool for improvement of fertility. In the current study, a 3D co-culture system with mice preantral follicles and human Menstrual Blood Mesenchymal Stem Cells (MenSCs) using either collagen or alginate beads was designed to investigate whether this system allows better preantral follicles growth and development. Results showed that MenSCs increase the indices of follicular growth including survival rate, diameter, and antrum formation as well as the rate of in vitro maturation (IVM) in both collagen and alginates beads. Although statistically not significant, alginate was found to be superior in terms of supporting survival rate and antrum formation. Hormone assay demonstrated that the amount of secreted 17 β-estradiol and progesterone in both 3D systems increased dramatically after 12 days, with the highest levels in system employing MenSCs. Data also demonstrated that relative expression of studied genes increased for Bmp15 and Gdf9 and decreased for Mater when follicles were cultured in the presence of MenSCs. Collectively, results of the present study showed that MenSCs could improve indices of follicular growth and maturation in vitro. Further studies are needed before a clinical application of MenSCs-induced IVM is considered. Copyright © 2018 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. All rights reserved.

Superior survival of ex vivo cultured human reticulocytes following transfusion into mice.

PubMed

Kupzig, Sabine; Parsons, Stephen F; Curnow, Elinor; Anstee, David J; Blair, Allison

2017-03-01

The generation of cultured red blood cells from stem cell sources may fill an unmet clinical need for transfusion-dependent patients, particularly in countries that lack a sufficient and safe blood supply. Cultured red blood cells were generated from human CD34 + cells from adult peripheral blood or cord blood by ex vivo expansion, and a comprehensive in vivo survival comparison with standard red cell concentrates was undertaken. Significant amplification (>10 5 -fold) was achieved using CD34 + cells from both cord blood and peripheral blood, generating high yields of enucleated cultured red blood cells. Following transfusion, higher levels of cultured red cells could be detected in the murine circulation compared to standard adult red cells. The proportions of cultured blood cells from cord or peripheral blood sources remained high 24 hours post-transfusion (82±5% and 78±9%, respectively), while standard adult blood cells declined rapidly to only 49±9% by this time. In addition, the survival time of cultured blood cells in mice was longer than that of standard adult red cells. A paired comparison of cultured blood cells and standard adult red blood cells from the same donor confirmed the enhanced in vivo survival capacity of the cultured cells. The study herein represents the first demonstration that ex vivo generated cultured red blood cells survive longer than donor red cells using an in vivo model that more closely mimics clinical transfusion. Cultured red blood cells may offer advantages for transfusion-dependent patients by reducing the number of transfusions required. Copyright© Ferrata Storti Foundation.

Prevalence of extended-spectrum beta-lactamase-producing Enterobacteriaceae isolated from blood cultures in Africa.

PubMed

Sangare, S A; Maiga, A I; Guindo, I; Maiga, A; Camara, N; Savadogo, S; Diallo, S; Bougoudogo, F; Armand-Lefevre, L; Andremont, A; Maiga, I I

2015-09-01

Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae have been isolated from many regions of the world. Epidemiological studies are being conducted in Europe, North America, and Asia. No study has however been conducted in Africa to determine the prevalence and distribution of ESBLs on the continent. This literature review aimed at describing the prevalence of ESBL-producing Enterobacteriaceae isolated from blood cultures, as well as the ESBL genes involved at the international level. Our focus was mainly on Africa. We conducted a literature review on PubMed. Articles related to our study field and published between 1996 and 2014 were reviewed and entirely read for most of them, while we only focused on the abstracts of some other articles. Relevant articles to our study were then carefully reviewed and included in the review. The prevalence of ESBL-producing Enterobacteriaceae differs from one country to another. The results of our literature review however indicate that class A ESBLs prevail over the other types. We took into consideration articles focusing on various types of samples to assess the prevalence of ESBL-producing Enterobacteriaceae, but information on isolates from blood cultures is limited. The worldwide prevalence of ESBL-producing Enterobacteriaceae has increased over time. Evidence of ESBL-producing Enterobacteriaceae can be found in all regions of the world. Studies conducted in Africa mainly focused on the Northern and Eastern parts of the continent, while only rare studies were carried out in the rest of the continent. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Evaluation of fluorescence in situ hybridisation (FISH) for the detection of fungi directly from blood cultures and cerebrospinal fluid from patients with suspected invasive mycoses.

PubMed

Da Silva, Roberto Moreira; Da Silva Neto, João Ricardo; Santos, Carla Silvana; Frickmann, Hagen; Poppert, Sven; Cruz, Kátia Santana; Koshikene, Daniela; De Souza, João Vicente Braga

2015-01-31

The aim of this study was to evaluate the diagnostic performance of in-house FISH (fluorescence in situ hybridisation) procedures for the direct identification of invasive fungal infections in blood cultures and cerebrospinal fluid (CSF) samples and to compare these FISH results with those obtained using traditional microbiological techniques and PCR targeting of the ITS1 region of the rRNA gene. In total, 112 CSF samples and 30 positive blood cultures were investigated by microscopic examination, culture, PCR-RFLP and FISH. The sensitivity of FISH for fungal infections in CSF proved to be slightly better than that of conventional microscopy (India ink) under the experimental conditions, detecting 48 (instead of 46) infections in 112 samples. The discriminatory powers of traditional microbiology, PCR-RFLP and FISH for fungal bloodstream infections were equivalent, with the detection of 14 fungal infections in 30 samples. However, the mean times to diagnosis after the detection of microbial growth by automated blood culture systems were 5 hours, 20 hours and 6 days for FISH, PCR-RFLP and traditional microbiology, respectively. The results demonstrate that FISH is a valuable tool for the identification of invasive mycoses that can be implemented in the diagnostic routine of hospital laboratories.

Diagnostic accuracy of the InBiOS AMD rapid diagnostic test for the detection of Burkholderia pseudomallei antigen in grown blood culture broth.

PubMed

Peeters, Marjan; Chung, Panha; Lin, Hua; Mortelmans, Kristien; Phe, Chhundy; San, Chentha; Kuijpers, Laura Maria Francisca; Teav, Syna; Phe, Thong; Jacobs, Jan

2018-06-01

To assess the diagnostic and operational performance of the InBiOS AMD rapid diagnostic test (RDT) (Seattle, USA) for the detection of B. pseudomallei in grown blood culture broth. The InBiOS RDT is a lateral flow immunoassay in a strip format detecting B. pseudomallei capsular polysaccharide in culture fluids, marketed for research only. Broth of blood culture bottles (BacT/Alert, bioMérieux, Marcy L'Etoile, France) sampled in adult patients at the Sihanouk Hospital Center of HOPE, Phnom Penh, Cambodia, during 2010-2017 and stored at - 80 °C was tested. They included samples grown with B. pseudomallei (n = 114), samples with no growth (n = 12), and samples with growth of other pathogens (n = 139, among which Burkholderia cepacia (n = 5)). Diagnostic sensitivity and specificity were 96.5% [95% confidence interval (CI): 91.3-98.6%] and 100% [CI: 97.5-100%] respectively. Background clearance and line intensities were good and very good. The RDT's test strip, not housed in a cassette, caused difficulties in manipulation and biosafety. The centrifugation step prescribed by the procedure challenged biosafety, but processing of 19 B. pseudomallei samples without centrifugation showed similar results for line intensity and background clearance, compared to centrifugation. The InBiOS RDT showed excellent accuracy for detection of B. pseudomallei in grown blood culture broth. Provided operational adaptations such as cassette housing, it has the potential to reduce time to diagnosis of melioidosis.

Typhidot M and Diazo test vis-à-vis blood culture and Widal test in the early diagnosis of typhoid fever in children in a resource poor setting.

PubMed

Beig, Farzana K; Ahmad, Faraz; Ekram, Mohd; Shukla, Indu

2010-01-01

Typhoid fever is a major public health problem. A test which is simple, reliable and can be carried out in small laboratories is the need of the hour. We prospectively evaluated typhidot M and Diazo tests vis-à-vis blood culture and Widal test in children. Patients aged 6 months to 12 years, having fever of more than four days duration with clinical suspicion of typhoid fever were enrolled. Patients in whom other diagnosis was made served as control. The tests under scrutiny were validated against blood culture and then all the four tests were evaluated among patients who presented in the first week of illness. Blood culture was positive in only 27.3% of the cases. Among these culture positive cases, typhidot M test had the highest sensitivity, specificity, PPV and NPV of 90% (95% CI = 74.4-96.5), 100% (95% CI = 90.1-100), 100% (95% CI = 87.5-100), and 92.1% (95% CI = 79.2-97.3) respectively. Diazo test ranked next with sensitivity, specificity, PPV and NPV of 86.7% (95% CI = 70.3-94.7), 85.7% (95% CI = 70.6-93.7), 83.9% (95% CI = 67.4-92.9), 88.2% (95% CI = 73.4-95.3) respectively. Among clinically suspected typhoid cases, the overall sensitivity, of blood culture, Widal, typhidot M, Diazo was 27.3% (95% CI = 19.8- 36.3), 64.6% (95% CI = 55.3-72.9), 89.1% (95% CI = 81.9-93.7), 80.9% (95% CI = 72.6-87.2) respectively. In the first week of illness, typhidot M showed the best sensitivity [86.2% (95% CI = 69.4-94.5)] followed by Diazo [79% (95% CI = 61.6-90.2)], Widal [41.4% (95% CI = 25.5-59.3)] and blood culture [31% (95% CI = 17.3-49.2)]. Both Typhidot M and Diazo are good screening tests for the diagnosis of typhoid fever. Typhidot M is superior to Diazo but the latter is more suitable to resource poor settings being economic and easy to perform.

Latent cytomegalovirus infection in blood donors

PubMed Central

Diosi, Peter; Moldovan, Eva; Tomescu, Nicholas

1969-01-01

Twenty-one out of 32 apparently healthy blood donors aged 21 to 65 years yielded positive complement fixation tests with a cytomegalovirus antigen, at titres ranging from 1:8 to 1:64. Virus was present in leucocyte cultures of fresh peripheral blood of two seropositive subjects from a total of 35 donors examined. Plasma and 48-hour stored blood specimens failed to disclose virus in culture. Viruria could not be demonstrated, and there was no evidence of recent illness in the study group. These findings suggest that subclinical viraemia is not uncommon in blood donors. PMID:4311727

The use of Gram stain and matrix-assisted laser desorption ionization time-of-flight mass spectrometry on positive blood culture: synergy between new and old technology.

PubMed

Fuglsang-Damgaard, David; Nielsen, Camilla Houlberg; Mandrup, Elisabeth; Fuursted, Kurt

2011-10-01

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) is promising as an alternative to more costly and cumbersome methods for direct identifications in blood cultures. We wanted to evaluate a simplified pre-treatment method for using MALDI-TOF-MS directly on positive blood cultures using BacT/Alert blood culture system, and to test an algorithm combining the result of the initial microscopy with the result suggested by MALDI-TOF-MS. Using the recommended cut-off score of 1.7 the best results were obtained among Gram-negative rods with correct identifications in 91% of Enterobacteriaceae, 83% in aerobic/non-fermentative Gram-negative rods, whereas results were more modest among Gram-positive cocci with correct identifications in 52% of Staphylococci, 54% in Enterococci and only 20% in Streptococci. Combining the results of Gram stain with the top reports by MALDI-TOF-MS, increased the sensitivity from 91% to 93% in the score range from 1.5 to 1.7 and from 48% to 85% in the score range from 1.3 to 1.5. Thus, using this strategy and accepting a cut-off at 1.3 instead of the suggested 1.7, overall sensitivity could be increased from 88.1% to 96.3%. MALDI-TOF-MS is an efficient method for direct routine identification of bacterial isolates in blood culture, especially when combined with the result of the Gram stain. © 2011 The Authors. APMIS © 2011 APMIS.

Beyond Blood Culture and Gram Stain Analysis: A Review of Molecular Techniques for the Early Detection of Bacteremia in Surgical Patients

PubMed Central

Kaplan, Heidi B.; Dua, Anahita; Litwin, Douglas B.; Ambrose, Catherine G.; Moore, Laura J.; Murray, COL Clinton K.; Wade, Charles E.; Holcomb, John B.

2016-01-01

Abstract Background: Sepsis from bacteremia occurs in 250,000 cases annually in the United States, has a mortality rate as high as 60%, and is associated with a poorer prognosis than localized infection. Because of these high figures, empiric antibiotic administration for patients with systemic inflammatory response syndrome (SIRS) and suspected infection is the second most common indication for antibiotic administration in intensive care units (ICU)s. However, overuse of empiric antibiotics contributes to the development of opportunistic infections, antibiotic resistance, and the increase in multi-drug-resistant bacterial strains. The current method of diagnosing and ruling out bacteremia is via blood culture (BC) and Gram stain (GS) analysis. Methods: Conventional and molecular methods for diagnosing bacteremia were reviewed and compared. The clinical implications, use, and current clinical trials of polymerase chain reaction (PCR)-based methods to detect bacterial pathogens in the blood stream were detailed. Results: BC/GS has several disadvantages. These include: some bacteria do not grow in culture media; others do not GS appropriately; and cultures can require up to 5 d to guide or discontinue antibiotic treatment. PCR-based methods can be potentially applied to detect rapidly, accurately, and directly microbes in human blood samples. Conclusions: Compared with the conventional BC/GS, particular advantages to molecular methods (specifically, PCR-based methods) include faster results, leading to possible improved antibiotic stewardship when bacteremia is not present. PMID:26918696

Direct identification of bacteria from charcoal-containing blood culture bottles using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.

PubMed

Wüppenhorst, N; Consoir, C; Lörch, D; Schneider, C

2012-10-01

Several protocols for direct matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) from positive blood cultures are currently used to speed up the diagnostic process of bacteraemia. Identification rates are high and results are accurate for the BACTEC™ system and for charcoal-free bottles. Only a few studies have evaluated protocols for charcoal-containing BacT/ALERT bottles reaching substantially lower identification rates. We established a new protocol for sample preparation from aerobic and anaerobic positive charcoal-containing BacT/ALERT blood culture bottles and measured the protein profiles (n = 167). Then, we integrated this protocol in the routine workflow of our laboratory (n = 212). During the establishment of our protocol, 74.3 % of bacteria were correctly identified to the species level, in 23.4 %, no result and in 2.4 %, a false identification were obtained. Reliable criteria for correct species identification were a score value ≥1.400 and a best match on rank 1-3 of the same species. Identification rates during routine workflow were 77.8 % for correct identification, 20.8 % for not identified samples and 1.4 % for discordant identification. In conclusion, our results indicate that MALDI-TOF MS is possible, even from charcoal-containing blood cultures. Reliable criteria for correct species identification are a score value ≥1.400 and a best match on rank 1-3 of a single species.

Evaluation of whole blood IFNgamma test using PPD and recombinant antigen challenge for diagnosis of pulmonary and extra-pulmonary tuberculosis.

PubMed

Kalantri, Yatiraj; Hemvani, Nanda; Chitnis, D S

2009-06-01

Quantiferon TB gold (QFT-G) with recombinant antigen cocktail is well evaluated for diagnosis of pulmonary tuberculosis (PTB). However, diagnosis of extra-pulmonary tuberculosis (EPTB) is more difficult due to limitations of conventional techniques. This study compares recombinant antigens based QFT-G and low cost PPD based interferon test for the diagnosis of PTB and EPTB. IFNgamma release, with recombinant antigens and PPD, was assayed by ELISA from 140 cases of EPTB, 100 cases of PTB along with acid fast bacillus (AFB) detection, AFB culture on LJ and MGIT BACTEC. Sensitivity and specificity for QFT-G recombinant antigens was 84.29% and 96%, while for PPD based interferon was 70% and 84% for EPTB group. The sensitivity was far superior to AFB smear and culture for both the antigens. Nine samples were identified as non-tubercular mycobacteria (NTM) in the EPTB group and all were negative for QFT-G, but six of them were positive for PPD based test. Results of the study show that QFT-G using recombinant antigen is sensitive and specific for both PTB and EPTB diagnosis. The PPD based test is economic and offers comparable performance for PTB and EPTB diagnosis and also useful for diagnosis of NTM.

Using qualitative research methods in biomedical innovation: the case of cultured red blood cells for transfusion.

PubMed

Lyall, Catherine; King, Emma

2016-05-11

Qualitative research has a key role to play in biomedical innovation projects. This article focuses on the appropriate use of robust social science methodologies (primarily focus group studies) for identifying the public's willingness and preference for emerging medical technologies. Our study was part of the BloodPharma project (now known as the Novosang project) to deliver industrially generated red blood cells for transfusion. Previous work on blood substitutes shows that the public prefers donated human blood. However, no research has been conducted concerning attitudes to stem cell derived red blood cells. Qualitative research methods including interviews and focus groups provide the methodological context for this paper. Focus groups were used to elicit views from sub-sections of the UK population about the potential use of such cultured red blood cells. We reflect on the appropriateness of that methodology in the context of the BloodPharma project. Findings are in the form of lessons transferable to other interdisciplinary, science-led teams about what a social science dimension can bring; why qualitative research should be included; and how it can be used effectively. Qualitative data collection offers the strength of exploring ambivalence and investigating the reasons for views, but not necessarily their prevalence in wider society. The inherent value of a qualitative method, such as focus groups, therefore lies in its ability to uncover new information. This contrasts with a quantitative approach to simply 'measuring' public opinion on a topic about which participants may have little prior knowledge. We discuss a number of challenges including: appropriate roles for embedded social scientists and the intricacies of doing upstream engagement as well as some of the design issues and limitations associated with the focus group method.

[A case of sphenoid sinusitis which could be diagnosed by orbital computed tomography after detected Strepotococcus pneumoniae from blood culture].

PubMed

Kimura, Takuma; Aoki, Makoto; Aoki, Yasuko; Tonhyo, Chong

2005-03-01

We report a case of sphenoid sinusitis which could be diagnosed by orbital CT after detecting Strepotococcus pneumoniae from blood culture. A previously healthy 47 year-old Japanese male was admitted to our hospital with severe left-sided headache of 2 days duration. From 9 days before hospitalization (1st day), the patient complained of cough and sputum. On physical examination, his neck was supple and his temperature was 38.3 degrees C. The rest of the examination was normal. A chest radiograph, sinus radiograph, and head computed tomographic (CT) scan without contrast material disclosed no abnormalities. Lumbar puncture was done and cerebrospinal fluid was clear and cell counts and the levels of glucose and protein were normal. The peripheral white blood cell count was 14,400/fl, and the C-reactive protein level was 9.6 mg/dl. After blood, urine, pharyngeal mucus and cerebrospinal fluid cultures were obtained, empirical antibiotic therapy with 2 gms of piperacillin twice daily was begun. He complained sever left-sided retro-orbital headahe on the next day too. The lumbar puncture and head CT scan with contrast material was done again but gave no diagnostic clues. The examinations by the otolaryngologist, ophthalmologist and dentist found no abnormal findings. On the 3rd hospitalized day, Strepotococcus pneumoniae was detected from the blood culture taken on the 1st hospitalized day. A CT scan focused on orbita was done and revealed a low density area of the left sphenoid sinus. The dose of piperacillin was increased to 4 gms twice daily and continued for 24 days. The patient's headache improved and piperacillin was changed to oral levofloxacin 100 mg, three times daily on the 26th day. The medication was stopped on the 73th day. Isolated sphenoid sinusitis is rare, but crtitical complications such as cranial nerve involvement, brain abscess, and bacterial meningitis may happen. It is necessary to also think of sphenoid sinusitis in practices of patients with

Amplification of ST50 gene using dry-reagent-based polymerase chain reaction for the detection of Salmonella typhi.

PubMed

Aziah, Ismail; Ravichandran, Manickam; Ismail, Asma

2007-12-01

Conventional polymerase chain reaction (PCR) testing requires many pipetting steps and has to be transported and stored in cold chain. To overcome these limitations, we designed a ready-to-use PCR test for Salmonella typhi using PCR reagents, primers against the ST50 gene of S. typhi, a built-in internal amplification control (IAC), and gel loading dye mixed and freeze-dried in a single tube. The 2-step dry-reagent-based assay was used to amplify a 1238-bp target gene and an 810-bp IAC gene from 73 BACTEC blood culture broths (33 true positives for S. typhi and 40 true negatives for non-S. typhi). The sensitivity, specificity, positive predictive value, and negative predictive value of the PCR assay were 87.9%, 100%, 100%, and 90.9%, respectively. We suggest that this rapid 2-step PCR test could be used for the rapid diagnosis of typhoid fever.

Pleural fluid culture

MedlinePlus

... Risks of thoracentesis are: Collapsed lung ( pneumothorax ) Excessive loss of blood Fluid reaccumulation Infection Pulmonary edema Respiratory distress Serious complications are uncommon Alternative Names Culture - pleural fluid Images Pleural culture References Chernecky CC, ...

[Evaluation of PNA-FISH method for direct identification of Candida species in blood culture samples and its potential impact on guidance of antifungal therapy].

PubMed

Doğan, Özlem; İnkaya, Ahmet Çağkan; Gülmez, Dolunay; Uzun, Ömrüm; Akova, Murat; Arıkan Akdağlı, Sevtap

2016-10-01

Early antifungal therapy has a major influence on survival in candidemia. Rapid identification of the species has importance for the treatment, prediction of the species-specific primary resistance and variable antifungal susceptibility. Recently, molecular-based methods attempt to reduce the time between the positive signal of a blood culture and identification of the fungus. PNA-FISH (Peptide nucleic acid fluorescence in situ hybridization) assay distinguishes a number of frequently isolated Candida species in groups following the growth in blood culture. The aim of this study was to investigate the correlation of the species identified by PNA-FISH with conventional identification methods in yeast positive blood cultures and its influence on the selection of antifungal therapy. Specimens of adult patients diagnosed as yeast with Gram stain in signal-positive blood cultures between August to December 2013, were included in the study. The strains were concomitantly cultivated by subculturing from the blood culture bottles onto solid media and identified by conventional methods (germ tube test, ID32C and morphology on cornmeal Tween 80 agar). Rapid species identification was performed by Yeast Traffic Light PNA-FISH, which generates green flourescence for Candida albicans and Candida parapsilosis, yellow for Candida tropicalis, and red for Candida krusei and Candida glabrata. C.tropicalis was identified as a single species whereas the others were identified in pairs. The time points when the yeast positive blood culture bottle was received by the mycology laboratory and reporting of the species identification results by PNA-FISH and the conventional methods were recorded. Seven C.albicans, six C.glabrata, three C.parapsilosis, one C.tropicalis, one C.krusei, one Cryptococcus neoformans, one Saprochaete capitata (Blastoschizomyces capitatus), one C.albicans and Candida dubliniensis, one C.krusei and C.dubliniensis, and one C.glabrata and C.parapsilosis were

Culture Phenotypes of Genomically and Geographically Diverse Mycobacterium avium subsp. paratuberculosis Isolates from Different Hosts▿

PubMed Central

Whittington, Richard J.; Marsh, Ian B.; Saunders, Vanessa; Grant, Irene R.; Juste, Ramon; Sevilla, Iker A.; Manning, Elizabeth J. B.; Whitlock, Robert H.

2011-01-01

Mycobacterium avium subsp. paratuberculosis causes paratuberculosis (Johne's disease) in ruminants in most countries. Historical data suggest substantial differences in culturability of M. avium subsp. paratuberculosis isolates from small ruminants and cattle; however, a systematic comparison of culture media and isolates from different countries and hosts has not been undertaken. Here, 35 field isolates from the United States, Spain, Northern Ireland, and Australia were propagated in Bactec 12B medium and Middlebrook 7H10 agar, genomically characterized, and subcultured to Lowenstein-Jensen (LJ), Herrold's egg yolk (HEY), modified Middlebrook 7H10, Middlebrook 7H11, and Watson-Reid (WR) agars, all with and without mycobactin J and some with sodium pyruvate. Fourteen genotypes of M. avium subsp. paratuberculosis were represented as determined by BstEII IS900 and IS1311 restriction fragment length polymorphism analysis. There was no correlation between genotype and overall culturability, although most S strains tended to grow poorly on HEY agar. Pyruvate was inhibitory to some isolates. All strains grew on modified Middlebrook 7H10 agar but more slowly and less prolifically on LJ agar. Mycobactin J was required for growth on all media except 7H11 agar, but growth was improved by the addition of mycobactin J to 7H11 agar. WR agar supported the growth of few isolates. The differences in growth of M. avium subsp. paratuberculosis that have historically been reported in diverse settings have been strongly influenced by the type of culture medium used. When an optimal culture medium, such as modified Middlebrook 7H10 agar, is used, very little difference between the growth phenotypes of diverse strains of M. avium subsp. paratuberculosis was observed. This optimal medium is recommended to remove bias in the isolation and cultivation of M. avium subsp. paratuberculosis. PMID:21430104

MALDI-TOF mass spectrometry following short incubation on a solid medium is a valuable tool for rapid pathogen identification from positive blood cultures.

PubMed

Kohlmann, Rebekka; Hoffmann, Alexander; Geis, Gabriele; Gatermann, Sören

2015-01-01

Rapid identification of the causative microorganism is a key element in appropriate antimicrobial therapy of bloodstream infections. Whereas traditional analysis of positive blood cultures requires subculture over at least 16-24h prior to pathogen identification by, e.g. matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), sample preparation procedures enabling direct MALDI-TOF MS, i.e. without preceding subculture, are associated with additional effort and costs. Hence, we integrated an alternative MALDI-TOF MS approach in diagnostic routine using a short incubation on a solid medium. Positive blood cultures were routinely plated on chocolate agar plates and incubated for 4h (37 °C, 5% CO2). Subsequently, MALDI-TOF MS using a Microflex LT instrument (Bruker Daltonics) and direct smear method was performed once per sample. For successful identification of bacteria at species level, score cut-off values were used as proposed by the manufacturer (≥ 2.0) and in a modified form (≥ 1.5 for MALDI-TOF MS results referring to Gram-positive cocci and ≥ 1.7 for MALDI-TOF MS results referring to bacteria other than Gram-positive cocci). Further data analysis also included an assessment of the clinical impact of the MALDI-TOF MS result. Applying the modified score cut-off values, our approach led to an overall correct species identification in 69.5% with misidentification in 3.4% (original cut-offs: 49.2% and 1.8%, respectively); for Gram-positive cocci, correct identification in 68.4% (100% for Staphylococcus aureus and enterococci, 80% for beta-hemolytic streptococci), for Gram-negative bacteria, correct identification in 97.6%. In polymicrobial blood cultures, in 72.7% one of the pathogens was correctly identified. Results were not reliable for Gram-positive rods and yeasts. The approach was easy to implement in diagnostic routine. In cases with available clinical data and successful pathogen identification, in 51.1% our

Proliferation of Peripheral Blood Lymphocytes and Mesenchymal Stromal Cells Derived from Wharton's Jelly in Mixed and Membrane-Separated Cultures.

PubMed

Poltavtsev, A M; Poltavtseva, R A; Yushina, M N; Pavlovich, S V; Svirshchevskaya, E V

2017-08-01

We studied the effect of mesenchymal stromal cells on proliferation of CFSE-stained T cells in mixed and membrane-separated (Transwell) cultures and in 3D culture of mesenchymal stromal cells from Wharton's jelly. The interaction of mesenchymal stromal cells with mitogen-activated peripheral blood lymphocytes from an allogeneic donor was followed by suppression of T-cell proliferation in a wide range of cell proportions. Culturing in the Transwell system showed the absence of suppression assessed by the fraction of proliferating cells and by the cell cycle analysis. In 3D cultures, contact interaction of mesenchymal stromal cells and lymphocytes was demonstrated that led to accumulation of G2/M phase lymphocytes and G0/G1 phase mesenchymal stromal cells. The suppressive effect of mesenchymal stromal cells from Wharton's jelly is mediated by two mechanisms. The effects are realized within 6 days, which suggests that the therapeutic effects of mesenchymal stromal cells persist until their complete elimination from the body.

Evaluation of the LightCycler Staphylococcus MGRADE Kits on Positive Blood Cultures That Contained Gram-Positive Cocci in Clusters

PubMed Central

Shrestha, Nabin K.; Tuohy, Marion J.; Padmanabhan, Ravindran A.; Hall, Gerri S.; Procop, Gary W.

2005-01-01

We evaluated the Roche LightCycler Staphylococcus MGRADE kits to differentiate between Staphylococcus aureus and coagulase-negative staphylococci in blood cultures growing clusters of gram-positive cocci. Testing 100 bottles (36 containing S. aureus), the assay was 100% sensitive and 98.44% specific for S. aureus and 100% sensitive and specific for coagulase-negative staphylococci. PMID:16333115

Modelling the endothelial blood-CNS barriers: a method for the production of robust in vitro models of the rat blood-brain barrier and blood-spinal cord barrier

PubMed Central

2013-01-01

Background Modelling the blood-CNS barriers of the brain and spinal cord in vitro continues to provide a considerable challenge for research studying the passage of large and small molecules in and out of the central nervous system, both within the context of basic biology and for pharmaceutical drug discovery. Although there has been considerable success over the previous two decades in establishing useful in vitro primary endothelial cell cultures from the blood-CNS barriers, no model fully mimics the high electrical resistance, low paracellular permeability and selective influx/efflux characteristics of the in vivo situation. Furthermore, such primary-derived cultures are typically labour-intensive and generate low yields of cells, limiting scope for experimental work. We thus aimed to establish protocols for the high yield isolation and culture of endothelial cells from both rat brain and spinal cord. Our aim was to optimise in vitro conditions for inducing phenotypic characteristics in these cells that were reminiscent of the in vivo situation, such that they developed into tight endothelial barriers suitable for performing investigative biology and permeability studies. Methods Brain and spinal cord tissue was taken from the same rats and used to specifically isolate endothelial cells to reconstitute as in vitro blood-CNS barrier models. Isolated endothelial cells were cultured to expand the cellular yield and then passaged onto cell culture inserts for further investigation. Cell culture conditions were optimised using commercially available reagents and the resulting barrier-forming endothelial monolayers were characterised by functional permeability experiments and in vitro phenotyping by immunocytochemistry and western blotting. Results Using a combination of modified handling techniques and cell culture conditions, we have established and optimised a protocol for the in vitro culture of brain and, for the first time in rat, spinal cord endothelial cells

Modelling the endothelial blood-CNS barriers: a method for the production of robust in vitro models of the rat blood-brain barrier and blood-spinal cord barrier.

PubMed

Watson, P Marc D; Paterson, Judy C; Thom, George; Ginman, Ulrika; Lundquist, Stefan; Webster, Carl I

2013-06-18

Modelling the blood-CNS barriers of the brain and spinal cord in vitro continues to provide a considerable challenge for research studying the passage of large and small molecules in and out of the central nervous system, both within the context of basic biology and for pharmaceutical drug discovery. Although there has been considerable success over the previous two decades in establishing useful in vitro primary endothelial cell cultures from the blood-CNS barriers, no model fully mimics the high electrical resistance, low paracellular permeability and selective influx/efflux characteristics of the in vivo situation. Furthermore, such primary-derived cultures are typically labour-intensive and generate low yields of cells, limiting scope for experimental work. We thus aimed to establish protocols for the high yield isolation and culture of endothelial cells from both rat brain and spinal cord. Our aim was to optimise in vitro conditions for inducing phenotypic characteristics in these cells that were reminiscent of the in vivo situation, such that they developed into tight endothelial barriers suitable for performing investigative biology and permeability studies. Brain and spinal cord tissue was taken from the same rats and used to specifically isolate endothelial cells to reconstitute as in vitro blood-CNS barrier models. Isolated endothelial cells were cultured to expand the cellular yield and then passaged onto cell culture inserts for further investigation. Cell culture conditions were optimised using commercially available reagents and the resulting barrier-forming endothelial monolayers were characterised by functional permeability experiments and in vitro phenotyping by immunocytochemistry and western blotting. Using a combination of modified handling techniques and cell culture conditions, we have established and optimised a protocol for the in vitro culture of brain and, for the first time in rat, spinal cord endothelial cells. High yields of both CNS

Efficient, Validated Method for Detection of Mycobacterial Growth in Liquid Culture Media by Use of Bead Beating, Magnetic-Particle-Based Nucleic Acid Isolation, and Quantitative PCR

PubMed Central

Waldron, Anna M.; Begg, Douglas J.; de Silva, Kumudika; Purdie, Auriol C.; Whittington, Richard J.

2015-01-01

Pathogenic mycobacteria are difficult to culture, requiring specialized media and a long incubation time, and have complex and exceedingly robust cell walls. Mycobacterium avium subsp. paratuberculosis (MAP), the causative agent of Johne's disease, a chronic wasting disease of ruminants, is a typical example. Culture of MAP from the feces and intestinal tissues is a commonly used test for confirmation of infection. Liquid medium offers greater sensitivity than solid medium for detection of MAP; however, support for the BD Bactec 460 system commonly used for this purpose has been discontinued. We previously developed a new liquid culture medium, M7H9C, to replace it, with confirmation of growth reliant on PCR. Here, we report an efficient DNA isolation and quantitative PCR methodology for the specific detection and confirmation of MAP growth in liquid culture media containing egg yolk. The analytical sensitivity was at least 104-fold higher than a commonly used method involving ethanol precipitation of DNA and conventional PCR; this may be partly due to the addition of a bead-beating step to manually disrupt the cell wall of the mycobacteria. The limit of detection, determined using pure cultures of two different MAP strains, was 100 to 1,000 MAP organisms/ml. The diagnostic accuracy was confirmed using a panel of cattle fecal (n = 54) and sheep fecal and tissue (n = 90) culture samples. This technique is directly relevant for diagnostic laboratories that perform MAP cultures but may also be applicable to the detection of other species, including M. avium and M. tuberculosis. PMID:25609725

Evaluation of the Accelerate Pheno System for Fast Identification and Antimicrobial Susceptibility Testing from Positive Blood Cultures in Bloodstream Infections Caused by Gram-Negative Pathogens.

PubMed

Marschal, Matthias; Bachmaier, Johanna; Autenrieth, Ingo; Oberhettinger, Philipp; Willmann, Matthias; Peter, Silke

2017-07-01

Bloodstream infections (BSI) are an important cause of morbidity and mortality. Increasing rates of antimicrobial-resistant pathogens limit treatment options, prompting an empirical use of broad-range antibiotics. Fast and reliable diagnostic tools are needed to provide adequate therapy in a timely manner and to enable a de-escalation of treatment. The Accelerate Pheno system (Accelerate Diagnostics, USA) is a fully automated test system that performs both identification and antimicrobial susceptibility testing (AST) directly from positive blood cultures within approximately 7 h. In total, 115 episodes of BSI with Gram-negative bacteria were included in our study and compared to conventional culture-based methods. The Accelerate Pheno system correctly identified 88.7% (102 of 115) of all BSI episodes and 97.1% (102 of 105) of isolates that are covered by the system's identification panel. The Accelerate Pheno system generated an AST result for 91.3% (95 of 104) samples in which the Accelerate Pheno system identified a Gram-negative pathogen. The overall category agreement between the Accelerate Pheno system and culture-based AST was 96.4%, the rates for minor discrepancies 1.4%, major discrepancies 2.3%, and very major discrepancies 1.0%. Of note, ceftriaxone, piperacillin-tazobactam, and carbapenem resistance was correctly detected in blood culture specimens with extended-spectrum beta-lactamase-producing Escherichia coli ( n = 7) and multidrug-resistant Pseudomonas aeruginosa ( n = 3) strains. The utilization of the Accelerate Pheno system reduced the time to result for identification by 27.49 h ( P < 0.0001) and for AST by 40.39 h ( P < 0.0001) compared to culture-based methods in our laboratory setting. In conclusion, the Accelerate Pheno system provided fast, reliable results while significantly improving turnaround time in blood culture diagnostics of Gram-negative BSI. Copyright © 2017 American Society for Microbiology.

Characterization of an Enterococcus faecium small-colony variant isolated from blood culture.

PubMed

Gröbner, Sabine; Beck, Julia; Schaller, Martin; Autenrieth, Ingo B; Schulte, Berit

2012-01-01

Small-colony variants (SCVs) of bacteria are slow-growing subpopulations which can cause latent or recurrent infections due to better intracellular survival compared to their wild-type counterparts. Atypical colony morphology and altered biochemical profile may lead to failure in identification of SCV strains. We here report for the first time the isolation of an Enterococcus faecium SCV phenotype. The case of a 65-year-old woman with acute myeloid leukaemia who developed symptoms of sepsis during induction chemotherapy is presented. E. faecium with normal and SCV phenotype was isolated from blood cultures. At the same time urine culture was positive with E. faecium suggesting that bacteraemia originated from the urinary tract. The SCV phenotype was characterized by atypical growth behaviour. Electron microscopic analyses revealed perturbation of the separation of daughter cells and the accumulation of cell wall material. Accordingly, the SCV variant showed a dysfunction or lack of spontaneous autolysis whereas the normal phenotype did not. In contrast to conventional identification systems based on biochemical characteristics, the E. faecium SCV was precisely identified by MALDI-TOF MS analysis implemented in our laboratory. Hence, the increasing use of MALDI-TOF MS analysis for the identification of bacteria might be an appropriate tool for the detection of SCV variants, the diagnosis of which is of importance for the clinical outcome and the antibiotic treatment. Copyright © 2011. Published by Elsevier GmbH.

Instant screening and verification of carbapenemase activity in Bacteroides fragilis in positive blood culture, using matrix-assisted laser desorption ionization--time of flight mass spectrometry.

PubMed

Johansson, Åsa; Nagy, Elisabeth; Sóki, József

2014-08-01

Rapid identification of isolates in positive blood cultures are of great importance to secure correct treatment of septicaemic patients. As antimicrobial resistance is increasing, rapid detection of resistance is crucial. Carbapenem resistance in Bacteroides fragilis associated with cfiA-encoded class B metallo-beta-lactamase is emerging. In our study we spiked blood culture bottles with 26 B. fragilis strains with various cfiA-status and ertapenem MICs. By using main spectra specific for cfiA-positive and cfiA-negative B. fragilis strains, isolates could be screened for resistance. To verify strains that were positive in the screening, a carbapenemase assay was performed where the specific peaks of intact and hydrolysed ertapenem were analysed with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). We show here that it is possible to correctly identify B. fragilis and to screen for enzymic carbapenem resistance directly from the pellet of positive blood cultures. The carbapenemase assay to verify the presence of the enzyme was successfully performed on the pellet from the direct identification despite the presence of blood components. The result of the procedure was achieved in 3 h. Also the Bruker mass spectrometric β-lactamase assay (MSBL assay) prototype software was proven not only to be based on an algorithm that correlated with the manual inspection of the spectra, but also to improve the interpretation by showing the variation in the dataset. © 2014 The Authors.

Direct identification of bacteria from BacT/ALERT anaerobic positive blood cultures by MALDI-TOF MS: MALDI Sepsityper kit versus an in-house saponin method for bacterial extraction.

PubMed

Meex, Cécile; Neuville, Florence; Descy, Julie; Huynen, Pascale; Hayette, Marie-Pierre; De Mol, Patrick; Melin, Pierrette

2012-11-01

In cases of bacteraemia, a rapid species identification of the causal agent directly from positive blood culture broths could assist clinicians in the timely targeting of empirical antimicrobial therapy. For this purpose, we evaluated the direct identification of micro-organisms from BacT/ALERT (bioMérieux) anaerobic positive blood cultures without charcoal using the Microflex matrix-assisted laser desorption/ionization (MALDI) time of flight MS (Bruker), after bacterial extraction by using two different methods: the MALDI Sepsityper kit (Bruker) and an in-house saponin lysis method. Bruker's recommended criteria for identification were expanded in this study, with acceptance of the species identification when the first three results with the best matches with the MALDI Biotyper database were identical, whatever the scores were. In total, 107 monobacterial cultures and six polymicrobial cultures from 77 different patients were included in this study. Among monomicrobial cultures, we identified up to the species level 67 and 66 % of bacteria with the MALDI Sepsityper kit and the saponin method, respectively. There was no significant difference between the two extraction methods. The direct species identification was particularly inconclusive for Gram-positive bacteria, as only 58 and 52 % of them were identified to the species level with the MALDI Sepsityper kit and the saponin method, respectively. Results for Gram-negative bacilli were better, with 82.5 and 90 % of correct identification to the species level with the MALDI Sepsityper kit and the saponin method, respectively. No misidentifications were given by the direct procedures when compared with identifications provided by the conventional method. Concerning the six polymicrobial blood cultures, whatever the extraction method used, a correct direct identification was only provided for one of the isolated bacteria on solid medium in all cases. The analysis of the time-to-result demonstrated a reduction

Pneumococcal septic arthritis in adults: clinical analysis and review.

PubMed

Belkhir, L; Rodriguez-Villalobos, H; Vandercam, B; Marot, J C; Cornu, O; Lambert, M; Yombi, J C

2014-01-01

Septic arthritis (SA) is a rheumatological emergency that can lead to rapid joint destruction and irreversible loss of function. The most common pathogen causing SA is Staphylococcus aureus which is responsible for 37-65% of cases. Streptococcus pneumoniae is traditionally described as an uncommon cause of SA of a native joint. The objective of our study was to analyse clinical characteristics, treatment, and outcome of all cases of pneumococcal septic arthritis treated in our institution, and to compare them with other series published in the literature. We conducted a retrospective study of pneumococcal SA identified among all cases of SA diagnosed in a teaching hospital of one thousand beds between 2004 and 2009. Diagnosis was based on culture of joint liquid or by the presence of pneumococcal bacteraemia and purulent (more than 50 000/mm(3) white blood cells with more than 90% neutrophils) joint fluid aspiration. Among 266 cases of SA, nine patients (3·3%) were diagnosed as having pneumococcal SA. The median age was 75 years. The main affected joint was the knee (7/9). No patient had more than one joint involved. Four patients suffered from concomitant pneumonia. Joint culture and blood cultures were positive in 7/9 and 5/9, respectively. Median (range) length of stay was 18 days (3-47 days). One patient with associated pneumococcal bacteraemia died 19 days after admission. Seven patients recovered completely. Streptococcus pneumoniae is now being increasingly recognized as a common agent of SA. This organism is frequently associated with pneumococcal pneumonia or bacteraemia, particularly in patients with advanced age and comorbidities. Direct inoculation of joint fluid into blood culture medium BACTEC system increases the probability of microbiological diagnosis. The prognosis is usually favourable if the disease is promptly recognized and treated (antibiotic therapy combined with joint drainage).

Comparison of Diagnostic Accuracy of Periprosthetic Tissue Culture in Blood Culture Bottles to That of Prosthesis Sonication Fluid Culture for Diagnosis of Prosthetic Joint Infection (PJI) by Use of Bayesian Latent Class Modeling and IDSA PJI Criteria for Classification.

PubMed

Yan, Qun; Karau, Melissa J; Greenwood-Quaintance, Kerryl E; Mandrekar, Jayawant N; Osmon, Douglas R; Abdel, Matthew P; Patel, Robin

2018-06-01

We have previously demonstrated that culturing periprosthetic tissue in blood culture bottles (BCBs) improves sensitivity compared to conventional agar and broth culture methods for diagnosis of prosthetic joint infection (PJI). We have also shown that prosthesis sonication culture improves sensitivity compared to periprosthetic tissue culture using conventional agar and broth methods. The purpose of this study was to compare the diagnostic accuracy of tissue culture in BCBs (subsequently referred to as tissue culture) to prosthesis sonication culture (subsequently referred to as sonicate fluid culture). We studied 229 subjects who underwent arthroplasty revision or resection surgery between March 2016 and October 2017 at Mayo Clinic in Rochester, Minnesota. Using the Infectious Diseases Society of America (IDSA) PJI diagnostic criteria (omitting culture criteria) as the gold standard, the sensitivity of tissue culture was similar to that of the sonicate fluid culture (66.4% versus 73.1%, P = 0.07) but was significantly lower than that of the two tests combined (66.4% versus 76.9%, P < 0.001). Using Bayesian latent class modeling, which assumes no gold standard for PJI diagnosis, the sensitivity of tissue culture was slightly lower than that of sonicate fluid culture (86.3% versus 88.7%) and much lower than that of the two tests combined (86.3% versus 99.1%). In conclusion, tissue culture in BCBs reached sensitivity similar to that of prosthesis sonicate fluid culture for diagnosis of PJI, but the two tests combined had the highest sensitivity without compromising specificity. The combination of tissue culture in BCBs and sonicate fluid culture is recommended to achieve the highest level of microbiological diagnosis of PJI. Copyright © 2018 American Society for Microbiology.

Comparative analysis of Micrococcus luteus isolates from blood cultures of patients with pulmonary hypertension receiving epoprostenol continuous infusion.

PubMed

Hirata, Yoshinori; Sata, Makoto; Makiuchi, Yuko; Morikane, Keita; Wada, Akihito; Okabe, Nobuhiko; Tomoike, Hitonobu

2009-12-01

During the period 2002-2008, at the National Cardiovascular Center, Osaka, 28 Micrococcus luteus isolates and one Kocuria spp. isolate were obtained from blood cultures of pulmonary hypertension (PH) patients who were receiving continuous infusion therapy with epoprostenol. Pulsed-field gel electrophoresis patterns of the isolates were unrelated, suggesting that the infections had multiple origins. The preparation of epoprostenol solution by patients themselves was thought to be a risk factor.

Prevalence of bloodstream pathogens is higher in neonatal encephalopathy cases vs. controls using a novel panel of real-time PCR assays.

PubMed

Tann, Cally J; Nkurunziza, Peter; Nakakeeto, Margaret; Oweka, James; Kurinczuk, Jennifer J; Were, Jackson; Nyombi, Natasha; Hughes, Peter; Willey, Barbara A; Elliott, Alison M; Robertson, Nicola J; Klein, Nigel; Harris, Kathryn A

2014-01-01

In neonatal encephalopathy (NE), infectious co-morbidity is difficult to diagnose accurately, but may increase the vulnerability of the developing brain to hypoxia-ischemia. We developed a novel panel of species-specific real-time PCR assays to identify bloodstream pathogens amongst newborns with and without NE in Uganda. Multiplex real-time PCR assays for important neonatal bloodstream pathogens (gram positive and gram negative bacteria, cytomegalovirus (CMV), herpes simplex virus(HSV) and P. falciparum) were performed on whole blood taken from 202 encephalopathic and 101 control infants. Automated blood culture (BACTEC) was performed for all cases and unwell controls. Prevalence of pathogenic bacterial species amongst infants with NE was 3.6%, 6.9% and 8.9%, with culture, PCR and both tests in combination, respectively. More encephalopathic infants than controls had pathogenic bacterial species detected (8.9%vs2.0%, p = 0.028) using culture and PCR in combination. PCR detected bacteremia in 11 culture negative encephalopathic infants (3 Group B Streptococcus, 1 Group A Streptococcus, 1 Staphylococcus aureus and 6 Enterobacteriacae). Coagulase negative staphylococcus, frequently detected by PCR amongst case and control infants, was considered a contaminant. Prevalence of CMV, HSV and malaria amongst cases was low (1.5%, 0.5% and 0.5%, respectively). This real-time PCR panel detected more bacteremia than culture alone and provides a novel tool for detection of neonatal bloodstream pathogens that may be applied across a range of clinical situations and settings. Significantly more encephalopathic infants than controls had pathogenic bacterial species detected suggesting that infection may be an important risk factor for NE in this setting.

Development of cultured Plasmodium falciparum blood-stage malaria cell banks for early phase in vivo clinical trial assessment of anti-malaria drugs and vaccines.

PubMed

Stanisic, Danielle I; Liu, Xue Q; De, Sai Lata; Batzloff, Michael R; Forbes, Tanya; Davis, Christopher B; Sekuloski, Silvana; Chavchich, Marina; Chung, Wendy; Trenholme, Katharine; McCarthy, James S; Li, Tao; Sim, B Kim Lee; Hoffman, Stephen L; Good, Michael F

2015-04-07

The ability to undertake controlled human malaria infection (CHMI) studies for preliminary evaluation of malaria vaccine candidates and anti-malaria drug efficacy has been limited by the need for access to sporozoite infected mosquitoes, aseptic, purified, cryopreserved sporozoites or blood-stage malaria parasites derived ex vivo from malaria infected individuals. Three different strategies are described for the manufacture of clinical grade cultured malaria cell banks suitable for use in CHMI studies. Good Manufacturing Practices (GMP)-grade Plasmodium falciparum NF54, clinically isolated 3D7, and research-grade P. falciparum 7G8 blood-stage malaria parasites were cultured separately in GMP-compliant facilities using screened blood components and then cryopreserved to produce three P. falciparum blood-stage malaria cell banks. These cell banks were evaluated according to specific criteria (parasitaemia, identity, viability, sterility, presence of endotoxin, presence of mycoplasma or other viral agents and in vitro anti-malarial drug sensitivity of the cell bank malaria parasites) to ensure they met the criteria to permit product release according to GMP requirements. The P. falciparum NF54, 3D7 and 7G8 cell banks consisted of >78% ring stage parasites with a ring stage parasitaemia of >1.4%. Parasites were viable in vitro following thawing. The cell banks were free from contamination with bacteria, mycoplasma and a broad panel of viruses. The P. falciparum NF54, 3D7 and 7G8 parasites exhibited differential anti-malarial drug susceptibilities. The P. falciparum NF54 and 3D7 parasites were susceptible to all anti-malaria compounds tested, whereas the P. falciparum 7G8 parasites were resistant/had decreased susceptibility to four compounds. Following testing, all defined release criteria were met and the P. falciparum cell banks were deemed suitable for release. Ethical approval has been obtained for administration to human volunteers. The production of cultured P

Bartonella Species, an Emerging Cause of Blood-Culture-Negative Endocarditis.

PubMed

Okaro, Udoka; Addisu, Anteneh; Casanas, Beata; Anderson, Burt

2017-07-01

Since the reclassification of the genus Bartonella in 1993, the number of species has grown from 1 to 45 currently designated members. Likewise, the association of different Bartonella species with human disease continues to grow, as does the range of clinical presentations associated with these bacteria. Among these, blood-culture-negative endocarditis stands out as a common, often undiagnosed, clinical presentation of infection with several different Bartonella species. The limitations of laboratory tests resulting in this underdiagnosis of Bartonella endocarditis are discussed. The varied clinical picture of Bartonella infection and a review of clinical aspects of endocarditis caused by Bartonella are presented. We also summarize the current knowledge of the molecular basis of Bartonella pathogenesis, focusing on surface adhesins in the two Bartonella species that most commonly cause endocarditis, B. henselae and B. quintana . We discuss evidence that surface adhesins are important factors for autoaggregation and biofilm formation by Bartonella species. Finally, we propose that biofilm formation is a critical step in the formation of vegetative masses during Bartonella -mediated endocarditis and represents a potential reservoir for persistence by these bacteria. Copyright © 2017 American Society for Microbiology.

The effects of a culturally-tailored campaign to increase blood donation knowledge, attitudes and intentions among African migrants in two Australian States: Victoria and South Australia

PubMed Central

Polonsky, Michael J.; Jones, Sandra C.; Renzaho, Andre M. N.

2017-01-01

Research suggests that African migrants are often positively predisposed towards blood donation, but are under-represented in participation. A culturally-tailored intervention targeting the African migrant community in Australia was developed and implemented, to enhance knowledge about blood donation, improve attitudes towards donating, increase intentions to donate blood, and increase the number of new African donors in Australia. Four weeks after a targeted campaign, a survey evaluation process commenced, administered face-to-face by bilingual interviewers from the African community in Melbourne and Adelaide, Australia (community survey). The questionnaires covered demographics, campaign awareness, blood donation knowledge and intentions, medical mistrust and perceived discrimination, and were analysed to evaluate changes in knowledge and intention. Sixty-two percent of survey participants (n = 454) reported being aware of the campaign. With increasing campaign awareness, there was a 0.28 increase in knowledge score (p = .005); previous blood donation was also associated with an increased blood donation knowledge score. Blood donation intention scores were not associated with campaign awareness (p = 0.272), but were associated with previous blood donation behaviour and a positive blood donation attitude score. More positive scores on the blood donation attitude measure were associated with increasing blood donation intentions, self-efficacy and campaign awareness (score increases of 0.27, 0.30 and 0.04, respectively, all p<0.05). Data were collected on the ethnicity of new blood donors in six blood collection centres before and after the intervention, and independent of the intervention evaluation survey. These data were also used to assess behavioural changes and the proportions of donors from different countries before and after the survey. There was no difference in the number of new African migrant donors, before and after the intervention. The culturally

The effects of a culturally-tailored campaign to increase blood donation knowledge, attitudes and intentions among African migrants in two Australian States: Victoria and South Australia.

PubMed

Francis, Kate L; Polonsky, Michael J; Jones, Sandra C; Renzaho, Andre M N

2017-01-01

Research suggests that African migrants are often positively predisposed towards blood donation, but are under-represented in participation. A culturally-tailored intervention targeting the African migrant community in Australia was developed and implemented, to enhance knowledge about blood donation, improve attitudes towards donating, increase intentions to donate blood, and increase the number of new African donors in Australia. Four weeks after a targeted campaign, a survey evaluation process commenced, administered face-to-face by bilingual interviewers from the African community in Melbourne and Adelaide, Australia (community survey). The questionnaires covered demographics, campaign awareness, blood donation knowledge and intentions, medical mistrust and perceived discrimination, and were analysed to evaluate changes in knowledge and intention. Sixty-two percent of survey participants (n = 454) reported being aware of the campaign. With increasing campaign awareness, there was a 0.28 increase in knowledge score (p = .005); previous blood donation was also associated with an increased blood donation knowledge score. Blood donation intention scores were not associated with campaign awareness (p = 0.272), but were associated with previous blood donation behaviour and a positive blood donation attitude score. More positive scores on the blood donation attitude measure were associated with increasing blood donation intentions, self-efficacy and campaign awareness (score increases of 0.27, 0.30 and 0.04, respectively, all p<0.05). Data were collected on the ethnicity of new blood donors in six blood collection centres before and after the intervention, and independent of the intervention evaluation survey. These data were also used to assess behavioural changes and the proportions of donors from different countries before and after the survey. There was no difference in the number of new African migrant donors, before and after the intervention. The culturally

A Triple Culture Model of the Blood-Brain Barrier Using Porcine Brain Endothelial cells, Astrocytes and Pericytes.

PubMed

Thomsen, Louiza Bohn; Burkhart, Annette; Moos, Torben

2015-01-01

In vitro blood-brain barrier (BBB) models based on primary brain endothelial cells (BECs) cultured as monoculture or in co-culture with primary astrocytes and pericytes are useful for studying many properties of the BBB. The BECs retain their expression of tight junction proteins and efflux transporters leading to high trans-endothelial electric resistance (TEER) and low passive paracellular permeability. The BECs, astrocytes and pericytes are often isolated from small rodents. Larger species as cows and pigs however, reveal a higher yield, are readily available and have a closer resemblance to humans, which make them favorable high-throughput sources for cellular isolation. The aim of the present study has been to determine if the preferable combination of purely porcine cells isolated from the 6 months old domestic pigs, i.e. porcine brain endothelial cells (PBECs) in co-culture with porcine astrocytes and pericytes, would compare with PBECs co-cultured with astrocytes and pericytes isolated from newborn rats with respect to TEER value and low passive permeability. The astrocytes and pericytes were grown both as contact and non-contact co-cultures as well as in triple culture to examine their effects on the PBECs for barrier formation as revealed by TEER, passive permeability, and expression patterns of tight junction proteins, efflux transporters and the transferrin receptor. This syngenic porcine in vitro BBB model is comparable to triple cultures using PBECs, rat astrocytes and rat pericytes with respect to TEER formation, low passive permeability, and expression of hallmark proteins signifying the brain endothelium (tight junction proteins claudin 5 and occludin, the efflux transporters P-glycoprotein (PgP) and breast cancer related protein (BCRP), and the transferrin receptor).

Impact of definition and procedures used for absent blood culture data on the rate of intravascular catheter infection during parenteral nutrition.

PubMed

Austin, P D; Hand, K S; Elia, M

2016-06-01

Diagnosis of intravascular catheter infection may be affected by the definition and procedures applied in the absence of blood culture data. To examine the extent to which different definitions of catheter infection and procedures for handling absent blood culture data can affect reported catheter infection rates. Catheter infection rates were established in a cohort of hospitalized patients administered parenteral nutrition according to three clinical and four published definitions. Paired and unpaired comparisons were made using available case analyses, sensitivity analyses and intention-to-categorize analyses. Complete data were available for each clinical definition (N = 193), and there were missing data (4.1-26.9%) for the published definitions. In an available case analysis, the catheter infection rate was 13.0-36.8% for the clinical definitions and 2.1-12.4% for the published definitions. For the published definitions, the rate was 1.6-32.1% in a sensitivity analysis and 11.4-16.9% in an intention-to-categorize analysis, with suggestion of bias towards a higher catheter infection rate in those with missing data, in keeping with the analyses of the clinical definitions. For paired comparisons, the strength of agreement between definitions varied from 'poor' (Cohen's kappa <0.21) to 'very good' (Cohen's kappa ≥0.81). The use of different definitions of catheter infection and procedures applied in the absence of blood culture data produced widely different catheter infection rates, which could compromise measurements or comparisons of service quality or study outcome. As such, there is a need to establish and use a valid, consistent and practical definition. Copyright © 2016 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

[Fever during blood transfusion. A case of coincidence instead of causality].

PubMed

Strobel, E; Bender, H-U

2013-10-01

A boy aged 2 years and 5 months showed agitation, shivering and fever with a temperature of 38.9 °C during a red blood cell transfusion. Examination of the assumed adverse transfusion reaction gave no indications of erythrocyte incompatibility, hemolysis or IgA incompatibility. No antibodies against HLA class I antigens or HPA antigens were found in the recipient's blood. Sterility testing of the blood product showed no growth, but in the blood cultures taken from the patient immediately after the blood transfusion, Enterobacter cloacae was detected which also could be found in blood cultures and nose and throat swabs taken 3 days before. Therefore, the fever during blood transfusion was not a case of causality but of coincidence. This case underlines the recommendation to examine blood cultures from the recipient in all suspected cases of adverse transfusion reaction.

Evaluation of the LightCycler Staphylococcus M GRADE kits on positive blood cultures that contained gram-positive cocci in clusters.

PubMed

Shrestha, Nabin K; Tuohy, Marion J; Padmanabhan, Ravindran A; Hall, Gerri S; Procop, Gary W

2005-12-01

We evaluated the Roche LightCycler Staphylococcus M(GRADE) kits to differentiate between Staphylococcus aureus and coagulase-negative staphylococci in blood cultures growing clusters of gram-positive cocci. Testing 100 bottles (36 containing S. aureus), the assay was 100% sensitive and 98.44% specific for S. aureus and 100% sensitive and specific for coagulase-negative staphylococci.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry for direct bacterial identification from positive blood culture pellets.

PubMed

Prod'hom, Guy; Bizzini, Alain; Durussel, Christian; Bille, Jacques; Greub, Gilbert

2010-04-01

An ammonium chloride erythrocyte-lysing procedure was used to prepare a bacterial pellet from positive blood cultures for direct matrix-assisted laser desorption-ionization time of flight (MALDI-TOF) mass spectrometry analysis. Identification was obtained for 78.7% of the pellets tested. Moreover, 99% of the MALDI-TOF identifications were congruent at the species level when considering valid scores. This fast and accurate method is promising.

Time to positivity of blood cultures in patients with bloodstream infections: A useful prognostic tool.

PubMed

Martín-Gutiérrez, Guillermo; Martín-Pérez, Carlos; Gutiérrez-Pizarraya, Antonio; Lepe, José A; Cisneros, José M; Aznar, Javier

2017-12-01

The time to positivity (TTP) of blood cultures in patients with bloodstream infections (BSIs) has been considered to be a possible prognostic tool for some bacterial species. However, notable differences have been found between sampling designs and statistical methods in published studies to date, which makes it difficult to compare results or to derive reliable conclusions. Our objective was to evaluate the clinical and microbiological implications of TTP among patients with BSI caused by the most common pathogens. A total of 361 episodes of BSI were reported for 332 patients. The survival of the entire cohort was measured from the time of blood culture sampling. In order to compare our results with those of previous studies, TTP was divided in three different groups based on log rank (short TTP <12h; medium TTP ≥12h to ≤27h, and long TTP >27h). Cox proportional hazard models were used to calculate crude and adjusted hazard ratios (HR). The Cox proportional hazard model revealed that TTP is an independent predictor of mortality (HR=1.00, p=0.031) in patients with BSIs. A higher mortality was found in the group of patients with the shortest TTP (<12h) (HR=2.100, p=0.047), as well as those with longest TTP (>27h) (HR=3.277, p=0.031). It seems that TTP may provide a useful prognostic tool associated with a higher risk of mortality, not only in patients with shorter TTP, but also in those with longer TTP. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Biocompatible and label-free separation of cancer cells from cell culture lines from white blood cells in ferrofluids.

PubMed

Zhao, Wujun; Cheng, Rui; Lim, So Hyun; Miller, Joshua R; Zhang, Weizhong; Tang, Wei; Xie, Jin; Mao, Leidong

2017-06-27

This paper reports a biocompatible and label-free cell separation method using ferrofluids that can separate a variety of low-concentration cancer cells from cell culture lines (∼100 cancer cells per mL) from undiluted white blood cells, with a throughput of 1.2 mL h -1 and an average separation efficiency of 82.2%. The separation is based on the size difference of the cancer cells and white blood cells, and is conducted in a custom-made biocompatible ferrofluid that retains not only excellent short-term viabilities but also normal proliferations of 7 commonly used cancer cell lines. A microfluidic device is designed and optimized specifically to shorten the time of live cells' exposure to ferrofluids from hours to seconds, by eliminating time-consuming off-chip sample preparation and extraction steps and integrating them on-chip to achieve a one-step process. As a proof-of-concept demonstration, a ferrofluid with 0.26% volume fraction was used in this microfluidic device to separate spiked cancer cells from cell lines at a concentration of ∼100 cells per mL from white blood cells with a throughput of 1.2 mL h -1 . The separation efficiencies were 80 ± 3%, 81 ± 5%, 82 ± 5%, 82 ± 4%, and 86 ± 6% for A549 lung cancer, H1299 lung cancer, MCF-7 breast cancer, MDA-MB-231 breast cancer, and PC-3 prostate cancer cell lines, respectively. The separated cancer cells' purity was between 25.3% and 28.8%. In addition, the separated cancer cells from this strategy showed an average short-term viability of 94.4 ± 1.3%, and these separated cells were cultured and demonstrated normal proliferation to confluence even after the separation process. Owing to its excellent biocompatibility and label-free operation and its ability to recover low concentrations of cancer cells from white blood cells, this method could lead to a promising tool for rare cell separation.

Membrane culture and reduced oxygen tension enhances cartilage matrix formation from equine cord blood mesenchymal stromal cells in vitro.

PubMed