Failure of Gram stain to detect Propionibacterium acnes in specimens from clinically significant infections

Abstract

We have read with interest the article by Johnson et al. (7) about the variability of the results of Gram staining encountered with some anaerobes. Gram stain is one of the cornerstones for bacterial identification, but it also serves as a useful technique for rapid detection of organisms in clinical samples (6). Propionibacterium acnes is a non-spore-forming, pleomorphic, anaerobic gram-positive rod (3) that is frequently considered to be a contaminant in clinical samples but may also be implicated as the agent of human infection (2, 4, 8). In our experience, Gram stain directly performed on clinical samples from patients with documented infection by P. acnes frequently failed to detect this organism. In order to evaluate the usefulness of Gram stain for direct detection of P. acnes, the laboratory records of the Medical Microbiology Department of the Fundación Jiménez Díaz (a 600-bed university hospital in Madrid, Spain) between January 1988 and August 1995 were reviewed for culture evidence of the presence of P. acnes. Those samples with heavy (3-streak) or moderate (2-streak) growth of P. acnes together with evidence of an inflammatory reaction (as detected on Gram smear by the presence of polymorphonuclear leukocytes) and yielding pure or predominant growth of P. acnes without any other more probable pathogen were selected. Critical review of patient charts was done to evaluate the clinical significance of the isolate. In 39 cases, meeting the above mentioned selection criteria, P. acnes was judged to have clinical significance. For controlling these results we later selected randomly other charts and assessed the Gram stain utility for other infections caused by various grampositive and gram-negative organisms. Table 1 shows the results of Gram stain for P. acnes infections and controls, indicating where the ability of the Gram stain to detect P. acnes was poor, especially when growth was only moderate, as compared with the ability of the Gram stain to reveal other grampositive and gram-negative organisms. Murray et al. (5) attribute misleading Gram stain results in clinical samples to cell wall deterioration due to the inflammatory response. Alterations in the P. acnes cell wall because of interaction with leukocytes or monocytes have been reported previously (9), and this fact, together with the pleomorphic nature of this bacteria (occasionally misinterpreted as a mixed infection [1, 8]) plus the presence of fibrinous debris in the samples, may explain the observations made in our study. In conclusion, Gram stain may not be a reliable technique for the rapid diagnosis of P. acnes infections, and when evidence of an abundant inflammatory response is noted in the Gram-stained smear, a more careful evaluation of cultures must be performed, particularly when nosocomial infection is of concern.

First case of Propionibacterium acnes urinary tract infection in a dog

“…In this case, P. acnes was not detected in urine via Gram staining. This was also demonstrated in a previous study [ 11 ], and should be taken into account when considering P. acnes infection as a differential.…”

A Fluorescent Gram Stain for Flow Cytometry and Epifluorescence Microscopy

“…has been associated with delayed diagnosis and inappropriate antibiotic therapy (7,20). All of the anaerobic gram-positive organisms in this study were characteristically decolorized (5,12) during the traditional Gram stain procedure. Using a modified Gram stain to study the staining reaction by electron microscopy, Beveridge (2) demonstrated that cytoplasmic voids were formed close to the cell wall and septation sites in P. acnes, causing the cells to lyse and appear gram negative.…”

Scanning Electron Microscope: A New Potential Tool to Replace Gram Staining for Microbe Identification in Blood Cultures

“…This type of strategy will lead to the creation of an image database and the development of a software using machine learning, allowing for the automatic identification of the objects observed. In the long term, this would replace the technical and microbiological knowledge necessary to be able to identify the bacteria with the commonly-used staining methods such as Gram and other dyes used for microorganisms [13,30]. A major part of the cost for our method comes from the purchase of a TM4000 Plus SEM (50 k EUR).…”

“…However, Gram staining results are not always reliable due to multiple factors, such as inter-operator variability, false staining for some species, etc. [11][12][13]. Within the last decade, insufficient identifications were switched to culture and eventually relied on other high accuracy methods for microbe identification in blood cultures such as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF/MS) [14], molecular biology [15][16][17], multiplex PCR assays, specific hybridization assays, or microarrays [18][19][20][21].…”