Rapid detection and identification of pathogens in blood cultures by fluorescence in situ hybridization and flow cytometry

Kempf, Volkhard A. J.; Mändle, Tanja; Schumacher, Ulrike; Schäfer, Andrea; Autenrieth, Ingo B.

doi:10.1016/j.ijmm.2004.12.006

Cited by 46 publications

(41 citation statements)

References 18 publications

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“…100 The technique is based on fluorescently labeled oligonucleotide probes specific for ribosomal RNA, which have been used to detect .95% of bacteria and fungi in BC. 101,102 The most commonly used target in prokaryotes is 16S rRNA.…”

Section: Hybridization: Pna-fish ® and Rebamentioning

confidence: 99%

In vitro diagnosis of sepsis: a review

Marcello

Tumolo²,

Donno³

et al. 2016

PLMI

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Sepsis, severe sepsis and septic shock, systemic inflammatory response, and other related manifestations represent a relevant medical problem with high morbidity and mortality, despite the improvements in diagnosis, treatment, and preventive measures over the last few decades. The limited knowledge of the pathophysiology in association with the lack of in vitro diagnostic methods for the certain and quick determination of the causative microbiological agents and their antibiotic resistance means the condition is still critical and of high impact in health care. The current gold standard method to detect the sepsis-causing pathogens, which is based on blood culture, is still insufficiently sensitive and slow. The new culture-independent molecular biology-based techniques can lead to the identification of a broad range of microorganisms and resistance markers within a few hours and with high sensitivity and specificity; nevertheless, limitations of, for example, the polymerase chain reaction-based methods still hamper their application in the clinical routine. This review summarizes the in vitro diagnostic methods and their approach in the clinical diagnosis of the bloodstream infections, and explores their advantages and disadvantages at the current state of the art. A quick analysis of the future prospective in multiplex technologies for microbiological diagnosis of sepsis is also provided.

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Section: Hybridization: Pna-fish ® and Rebamentioning

confidence: 99%

In vitro diagnosis of sepsis: a review

Marcello

Tumolo²,

Donno³

et al. 2016

PLMI

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“…In the clinical realm, FISH-FC using both DNA and PNA probes has been explored for rapid detection of pathogens in blood cultures, including Gram-negative rods, Staphylococcus aureus and Candida spp. (52,67). Access to rapid, robust and culture-independent methods for pathogen detection or surveillance is expected to be important to this sector, as the benefits of rapid clinical diagnostics have been clearly identified in terms of both cost savings and improved patient outcome (reduced mortality) (68,69).…”

Section: Combining Fish and Flow Cytometrymentioning

confidence: 99%

Methods for Whole Cell Detection of Microorganisms

Brehm-Stecher

2008

ACS Symposium Series

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Microbes are ubiquitous, and can be found occupying nearly every imaginable niche on Earth. These include organic and inorganic surfaces, interfacial boundaries and within macroscopically solid matrices, such as the pore space of rocks. Because phylogenetically divergent microbes may be visually indistinguishable, understanding the species distribution and ecological significance of environmental microbes requires diagnostic tools that extend beyond simple phenotypic description. Methods for microbial diagnostics can be divided into two broad categories: cellular and acellular. Acellular techniques, such as the polymerase chain reaction or certain immunoassay formats may be effective at detecting molecular, structural or biochemical targets associated with specific cell types, but this information is provided out of its "natural", and arguably most meaningful context -that of the individual microbial cell. In contrast, cellular methods have the potential to preserve an abundance of valuable information. Apart from molecular identity, this includes information regarding cell morphology and other physiological characteristics, cell number and distribution within a sample, and physical or spatial associations with other cell types. The aim of this chapter is to provide an overview of whole cell methods for microbial detection, including both existing approaches and those still in development. The tools described here are expected to find wide application for the detection of microbes on surfaces or within complex matrices across a number of parallel or allied fields, including environmental, food and clinical microbiologies.

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“…in blood cultures are critical. [2,10,11] Current conventional methods for the identification of Candida spp. from positive blood cultures are based on subculture to appropriate fungal medium, including Sabouraud dextrose agar, or CHROMagar Candida medium, and isolated yeast colonies are identified by the phenotypic (germ tube formation, carbohydrate assimilation [e.g., API 20C, ID 32C], and chlamydospore formation on cornmeal agar).…”

mentioning

confidence: 99%

“…[12] More recently, molecular techniques have found a broad application in the detection and identification of microorganisms in clinical samples without cultivation. [10,13,14] Molecular techniques such as DNA sequence analysis, real-time polymerase chain reaction (RT-PCR), and multiplex PCR have enabled the detection and rapid identification of pathogenic fungi, including Candida species in clinical samples. [14][15][16] However, most of these tests are still time-consuming and too expensive for routine use.…”

mentioning

confidence: 99%

“…[17,18] Fluorescent in situ hybridization (FISH) with rRNA-targeted oligonucleotide probes has also become a highly valuable tool for the identification of microorganisms without cultivation. [10,19] This method has been reported to be a reasonable and rapid method in the detection and identification of pathogens. Moreover, this method is cheaper and easier to perform than the other molecular methods in the microbiology laboratory.…”

mentioning

confidence: 99%

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Identification Of Candida Species From Blood Cultures With Fluorescent In Situ Hybridization (fish), Polymerase Chain Reaction-restriction Fragment Length Polymorphism (pcr-rflp) And Conventional Methods

Börekçi¹,

Ersöz²,

Otağ³

et al. 2009

TUTFD

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Objectives: Rapid and accurate identification of Candida species from blood cultures is crucial to ensure effective antifungal therapy and to reduce the morbidity and mortality associated with bloodstream fungal infections. In this study, we aimed to identify Candida spp. from blood culture samples with fluorescent in situ hybridization (FISH), polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and conventional methods. Materials and Methods:A total of 50 yeast positive samples out of 325 blood culture positive samples and 50 blood culture negative samples were examined by FISH, PCR-RFLP and conventional methods to identify Candida spp.Results: All three methods generally were compatible for identification of single-species Candida spp. (p<0.001). But, FISH and PCR-RFLP for the identification of multi-species Candida spp. were found more compatible than conventional methods (p<0.001). Besides, FISH is cheaper and quicker than the other two methods in the identification of Candida spp. from blood culture positive samples. The rates of multi-species candidemia with FISH, PCR-RFLP and conventional methods were 20%, 6% and 4%, respectively. Conclusion:Both PCR-RFLP and FISH methods might be preferred for the rapid identification of Candida spp. from blood culture positive samples. However, FISH is a more suitable method for the detection of multi-species candidemia.

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Rapid detection and identification of pathogens in blood cultures by fluorescence in situ hybridization and flow cytometry

Cited by 46 publications

References 18 publications

In vitro diagnosis of sepsis: a review

In vitro diagnosis of sepsis: a review

Methods for Whole Cell Detection of Microorganisms

Identification Of Candida Species From Blood Cultures With Fluorescent In Situ Hybridization (fish), Polymerase Chain Reaction-restriction Fragment Length Polymorphism (pcr-rflp) And Conventional Methods

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