What is a Gram stain?

From the Guidelines

Gram staining morphology is a useful tool in predicting Staphylococcus aureus in respiratory samples of patients with ventilator-associated pneumonia (VAP), with a pooled sensitivity and specificity of 68% and 95%, respectively, as reported in a systematic review and meta-analysis published in 2020 1. The diagnostic accuracy of Gram staining in predicting VAP caused by S. aureus is supported by a systematic review and meta-analysis that found a positive likelihood ratio of 12.7 and a negative likelihood ratio of 0.34 1.

Key Findings

• The study found that Gram staining morphology suggestive of S. aureus in respiratory samples has a high negative predictive value, which can help clinicians rule out VAP caused by S. aureus in ICUs with low to moderate prevalence of S. aureus 1.
• The presence of Gram-positive cocci in clusters on Gram staining supports the introduction of an anti-staphylococcal antibiotic for ICUs with low prevalence of S. aureus 1.
• The use of Gram staining morphology in clinical decision-making processes can provide rapid and useful information for empirical antibiotic coverage for VAP, although randomized clinical trials with patient-centered outcomes are needed for strong recommendations in clinical practice 1.

Clinical Implications

• Gram staining morphology should be used to support clinical decision-making processes for empirical antibiotic coverage in VAP, particularly in ICUs with low to moderate prevalence of S. aureus.
• Clinicians should consider the results of Gram staining morphology in conjunction with other risk factors and severity of illness when making decisions about empirical antibiotic therapy.
• Further studies are needed to evaluate the impact of Gram staining morphology on patient outcomes, costs, and empirical antibiotic choices.

From the Research

Gram Stain Overview

• The Gram stain is a method used to distinguish between Gram-positive and Gram-negative bacteria based on differential staining with a crystal violet-iodine complex and a safranin counterstain 2, 3.
• The cell walls of Gram-positive organisms retain the crystal violet-iodine complex after treatment with alcohol and appear purple, whereas Gram-negative organisms decolorize following such treatment and appear pink 2, 3.

Gram Stain Mechanism

• The Gram stain differentiates bacteria into two fundamental varieties of cells based on the chemical and structural makeup of their cell walls 4.
• Gram-positive bacteria have a thick, relatively impermeable wall that resists decolorization and is composed of peptidoglycan and secondary polymers, while Gram-negative bacteria have a thin peptidoglycan layer plus an overlying lipid-protein bilayer known as the outer membrane 4.

Applications of Gram Stain

• The Gram stain is useful for assessing bacterial contamination of tissue culture samples or for examining the Gram stain status and morphological features of bacteria isolated from mixed or isolated bacterial cultures 2, 3.
• The Gram stain can be used in combination with other methods, such as the Accelerate Pheno™ system, for rapid identification and antimicrobial susceptibility testing of Gram-positive and Gram-negative bacteria 5.

Limitations of Gram Stain

• The Gram stain is not a useful differentiating tool for some bacteria, such as those in the Archaea domain, which have variability of wall structure 4.
• The Gram stain has limitations in identifying certain types of bacteria, such as streptococci and coagulase-negative staphylococci 5.