A sample preparation method which facilitates detection of bacteria in blood cultures by the polymerase chain reaction

Background: Proper identification of the causative organism in pediatric sepsis is crucial for early diagnosis and prevention of septic shock and organ failure. Objectives: The present study aimed to evaluate the multiplex Polymerase Chain Reaction (PCR) to detect Acinetobacter baumannii, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia from positive blood cultures for these pathogens isolated from children, with hospital-acquired sepsis compared to the conventional biochemical reactions for identification of these organisms. Methods: This study was a cross-sectional study performed on 100 isolates from pediatric blood cultures, including Acinetobacter baumannii, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia. The study also included 100 isolates of Escherichia coli as a negative control. All isolates were identified by API 20NE and the multiplex PCR, with primers specific to the 3 tested bacteria. Results: Multiplex PCR was positive in 96% of isolates, and 4 isolates had negative results. False positive results were reported with three E. coli strains. Multiplex PCR identified all the isolates of Acinetobacter baumannii, 29 isolates of Pseudomonas aeruginosa, and 27 isolates of Stenotrophomonas maltophilia. Compared to the biochemical identification, the diagnostic value of the multiplex PCR revealed 96.04% sensitivity, 96.9% specificity, 97.00%, positive predictive value, 96.00% negative predictive value, and 96.50% accuracy. Conclusion: The present study highlights the diagnostic value of multiplex PCR to identify Acinetobacter baumannii, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia from positive blood cultures. Multiplex PCR was sensitive, specific, and accurate. The accuracy differs according to the organisms, with 100% accuracy for Acinetobacter baumannii.

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

Frontiers in Microbiology ◽

10.3389/fmicb.2018.00481 ◽

2018 ◽

Vol 9 ◽

Cited By ~ 8

Author(s):

Hong-wei Pan ◽

Wei Li ◽

Rong-guo Li ◽

Yong Li ◽

Yi Zhang ◽

...

Keyword(s):

Sample Preparation ◽

Susceptibility Testing ◽

Preparation Method ◽

Blood Cultures ◽

Sample Preparation Method ◽

Microbial Identification ◽

Simple Sample Preparation

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Whole blood sample preparation for polymerase chain reaction using ammonium chloride and a carboxylic acid or metal carboxylate for selective red blood cell lysis (TE Ekeze and JH Kerschner, US)

Biofutur ◽

10.1016/s0294-3506(97)82269-7 ◽

1997 ◽

Vol 1997 (173) ◽

pp. 43

Keyword(s):

Polymerase Chain Reaction ◽

Carboxylic Acid ◽

Sample Preparation ◽

Blood Cell ◽

Red Blood Cell ◽

Whole Blood ◽

Metal Carboxylate ◽

Chain Reaction ◽

Polymerase Chain ◽

Whole Blood Sample

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The comparison of cultures, widal agglutination test and polymerase chain reaction as a diagnostic tool in typhoid fever

Open Medicine ◽

10.2478/s11536-008-0052-8 ◽

2008 ◽

Vol 3 (4) ◽

pp. 470-474 ◽

Cited By ~ 1

Author(s):

Ilker Devrim ◽

Koray Ergünay ◽

Ates Kara ◽

Hasan Tezer ◽

Inci Yiğitkanl ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Typhoid Fever ◽

Enteric Fever ◽

Antimicrobial Agents ◽

Blood Cultures ◽

Chain Reaction ◽

Specific Test ◽

Salmonella Infections ◽

Group I ◽

Polymerase Chain

AbstractTyphoid fever caused by Salmonella typhi, paratyphi A and B, is an important cause of morbidity and mortality in many developing countries. A rapid and sensitive method for the detection of S. typhi is essential for early diagnosis of typhoid fever and effective therapy. In this study 45 febrile patients who were suspected to have enteric fever were enrolled, and the results of blood cultures, widal agglutination tests and Polymerase Chain Reaction in these cases were evaluated. Group I consisted of 11 patients with diseases other than salmonella infections, group II represented 6 patients with positive cultures, and group III represented 28 patients with negative blood cultures negative but who were clinically suspected cases that had a medical history of using variable antimicrobial agents. Two positive PCR results were present; one of them was in culture positive group (16,6%) and the other was in culture negative group (3,5%). In our study widal agglutination tests and cultures were found not to be helpful in differential dignosis. Although PCR based detection of S. typhi is reported to be a sensitive and specific test for the diagnosis of enteric fever, in our study the benefit of this method in the diagnosis of especially patients who were treated with antimicrobial therapy was not clearly determined. Other methods to increase sensitiviy and specificity to levels such as those of real time PCR should be developed and large-scaled studies should be done in endemic and non-epidemic regions.

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Polymerase Chain Reaction-And Rna Hybridization-Based Method for the Investigation of Deep-Seated Candidiasis

Canadian Journal of Infectious Diseases ◽

10.1155/1997/520178 ◽

1997 ◽

Vol 8 (6) ◽

pp. 329-334 ◽

Cited By ~ 1

Author(s):

Peter R Couroux ◽

Zafar Hussain ◽

Frank Rutledge ◽

Robert Lannigan ◽

Edward D Ralph ◽

...

Keyword(s):

Risk Factors ◽

Polymerase Chain Reaction ◽

Invasive Candidiasis ◽

Gel Electrophoresis ◽

Tertiary Care ◽

Immunosuppressive Drugs ◽

Blood Cultures ◽

Hybridization Method ◽

Chain Reaction ◽

Polymerase Chain

OBJECTIVE: To determine the usefulness of a polymerase chain reaction (PCR) and RNA hybridization method for the diagnosis of invasive candidiasis and to compare its sensitivity with blood cultures.DESIGN: Blood cultures and a blood sample for PCR were taken from patients with suspected invasive candidiasis. A 105 base pair conserved segment within the rDNA ofCandidaspecies was amplified. The amplicon was detected by hybridization and gel electrophoresis.SETTING: Intensive care units of two tertiary care hospitals.PATIENTS: One hundred and eighteen patients 16 years of age or older with four more risk factors for invasive candidiasis were enrolled. Present or recent past treatment with broad spectrum antibiotics, cancer chemotherapy, immunosuppressive drugs, granulocytopenia or granulocytosis, intravascular catheterization, tracheal intubation, recent abdominal surgery and parenteral nutrition were considered risk factors.RESULTS: Forty-three patients had invasive candidiasis. PCR detected infections in 28 and 26 patients (sensitivity 65.1% and 60.4%) by hybridization and gel electrophoresis, respectively. The sensitivity of blood cultures was 58.1%. Of 25 patients with positive blood cultures, 17 were positive by PCR with the hybridization method. Eleven patients with invasive candidiasis had negative blood cultures but were positive by PCR.CONCLUSION: PCR, especially with a hybridization detection method, is more sensitive than blood culture for invasive candidiasis and may facilitate the diagnosis of nonfungemic disease.

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