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Intra-catheter leukocyte stain and culture as a diagnostic tool in an experimental animal model of port-related bloodstream infection

Abstract number: P1938

Serrera A., Martinez A., Alonso M., Guillen-Grima F., Aguinaga A., Gonzalez R., Leiva J., del Pozo J.L.

Objectives: Port-related bloodstream infections (PRBI) are difficult to diagnose without port removal. Quantitative blood cultures (QBC) have become the gold standard method for conservative diagnosis. However, this method is laboring, costly and sometimes insensitive. Intra-catheter leukocytes can be isolated, stained by acridine orange (AO) and cultured. The aim of this study was to analyse the usefulness of this method to perform a rapid diagnose of PRBI in an experimental animal model. Secondary we determined the optimal threshold for intracatheter leukocyte culture (ILC) for the diagnosis of PRBI.

Methods: 32 sheep were implanted with venous access ports. Three days after, a standard Staphylococcus aureus inoculum was locked inside the port and allowed to dwell for 24 hours. Blood was drawn through port and a peripheral vein three days post-inoculation. Paired QBC, ILC and AO stain were performed. To obtain the intra-catheter leucocytes, 1mL of the blood sample was layered over 1 ml of the extraction solution in a sterile shell vial tube, and centrifuged at 3500 g for 30 minutes. A cellular monolayer was prepared from 50 microliters of the leukocyte band and slides were stained with AO. A positive result was the detection of one microorganism after observing 20 fields by fluorescence microscopy (470 nm). Fifty microliters from the leucocyte band were inoculated into 2 blood agar plates for the ILC. Any cfu observed in the ILC was considered as a positive culture.

Results: We could obtain paired samples in 28 of the 32 animals; devices were obstructed in 4 animals. QBC met criteria for PRBI in 89% of the animals. ILC culture was positive in 100% of the animals, and we could detect some microorganism in 21.4% of the AO stains. The ROC analysis showed that the best threshold for the ILC was 420 cfu/mL (77% sensitivity and 40% specificity). When the cfu in the ILC were higher than 5000 per mililiter, the AO stain improved its sensitivity to 66%.

Conclusion: The results obtained in our model suggest that ILC could be a valid conservative method to diagnose PRBI using a small amount of port blood (1 ml). The AO stain, although it is a simple and quick test, is not very useful for the diagnosis of PRBI. Limitations of this study are the small sample size and the short period of time studied after inoculation, so more studies are warranted.

This work was supported by a grant from Proy. Inst. Carlos III ref. 03/1102

Session Details

Date: 19/04/2008
Time: 00:00-00:00
Session name: 18th European Congress of Clinical Microbiology and Infectious Diseases
Subject:
Location: Barcelona, Spain
Presentation type:
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