Misinterpretation of Gram Stain from the Stationary Growth Phase of Positive Blood Cultures for Brucella and Acinetobacter Species



Ali M. Bazzi1, *, Jaffar A. Al-Tawfiq3, 4, Ali A. Rabaan2
1 Microbiology Lab, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia
2 Molecular Diagnostic Lab, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia
3 Specialty Internal Medicine, Johns Hopkins Aramco Healthcare, Dhahran, Saudi Arabia
4 Department of Medicine Indiana University School of Medicine, Indianapolis, IN, USA


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© 2017 Bazzi et al.

open-access license: This is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International Public License (CC-BY 4.0), a copy of which is available at: https://creativecommons.org/licenses/by/4.0/legalcode. This license permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

* Address correspondence to this author at the John Hopkins Aramco Health care Saudi Aramco -Dhahran, zip code 31311, Dhahran health center, bdg 62, room 273, Saudi Arabia; Tel: +966138776636; Fax: 966138776629; E-mails: bazziamh@gmail.com; ali.bazzi@JHAH.com


Abstract

Introduction:

Acinetobacter baumannii and Brucella species are Gram-negative organisms that are vulnerable to misinterpretation as Gram-positive or Gram-variable in blood cultures.

Objective:

We assess the random errors in gram stain interpretation to reduce the likelihood of such errors and therefore patient harm.

Methodology:

Aerobic and anaerobic blood cultures from two patients in an acute care facility in Saudi Arabia were subjected to preliminary Gram-staining. In case 1, VITEK-2 Anaerobe Identification, repeat Gram staining from a blood agar plate, Remel BactiDrop™ Oxidase test, Urea Agar urease test and real-time PCR were used to confirm presence of Brucella and absence of Coryneform species. In case 2, repeat Gram- staining from the plate and the vials, VITEK-2 Gram-Negative Identification, real-time PCR and subculture on to Columbia agar, blood agar, and MacConkey agar were carried out to identify A. baumannii.

Results:

In case 1, initially pleomorphic Gram-positive bacteria were identified. Coryneform species were suspected. Tiny growth was observed after 24 h on blood agar plates, and good growth by 48 h. Presence of Brucella species was ultimately confirmed. In case 2, preliminary Gram-stain results suggested giant Gram-positive oval cocci. Further testing over 18-24 h identified A. baumannii.

Conclusions:

Oxidase test from the plate and urease test from the culture vial is recommended after apparent identification of pleomorphic Gram-positive bacilli from blood culture, once tiny growth is observed, to distinguish Brucella from Corynebacterium species. If giant Gram-positive oval cocci are indicated by preliminary Gram-staining, it is recommended that the Gram stain be repeated from the plate after 4-6 h, or culture should be tested in Triple Sugar Iron (TSI) medium and the Gram stain repeated after 2-4 h incubation.

Keywords: Gram-stain, Blood culture, Brucella, Acinetobacter, Coryneform.