7/29/2019 Gram's staining Face to Face with Microbiologists Gram Staining

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GRAM STAINING

Face to Face on FACEBOOK

Edited by Dr.T.V.Rao MD

Importance and Limitations of Gram Staining in Diagnostic Microbiology

Majority of the Microbiology Laboratories do a culture work for Bacterial identification from several

specimens arise from the Hospitals. Gram staining is a common traditional procedure and an age old

procedure since Christian Grams contribution in 18th

Century, for Bacterial studies. The differentiation

of bacteria into either the gram-positive or the gram-negative group is fundamental to most bacterial

identification systems. This task is usually accomplished through the use of Grams Staining Method.

Unfortunately, the gram stain methodology is complex and prone to error. The technique is used as a

tool for the differentiation of Gram-positive and Gram-negative bacteria, as a first step to determinethe identity of a particular bacterial sample. The Gram stain is not an infallible procedure for diagnosis,

identification, or phylogeny. However it is of extremely limited use when saprophytes and

commensals are associated with careless specimen collection AND PRECESSING IN OUR Laboratories

however it is a procedure which helps when Gram stains are performed on body fluids, CSF or biopsy

when infection is suspected. It yields results much more quickly than culture, and is especially

important when infection would make an important difference in the patient's treatment and

prognosis; examples are cerebrospinal fluid for meningitis and synovial fluid for septic arthritis. Many

Grams stains from specimens with Polymicrobial flora are fallacious as it is in Diabetic foot, Sputum,

faecal, and urinary specimens. Many juniors and even the Seniors Microbiologists believe the first

impressions and come to hasty conclusion in advising the therapeutic options. My experience proves

that when we process THE BACTERIA from the Culture plates, by Macroscopic observation many of

our conclusions may not be true, and have little determination and patience to think all you see and

define by naked eye ( Macroscopic Observations ) may not prove correct when we really do a Grams

staining, many Micrococci, Diptheroids, and Candida spp and other normal flora mimic as pathogenic

isolates and processed with confusing uncharacterised Biochemical reaction, and tested for

Antibiograms, this is an area where the Grams staining plays wonders to enlighten us. Problems with

Grams Method have led to a search for other tests that correlate with the cell wall structure of the

gram-positive and the gram-negative cells. Several improvements/alternatives to the classical gram

stain have appeared in the literature. Many conflicts in our Laboratories between members of the

staff and technicians are due to ignorance as when we have a little interest to go in a Methodical and

Scientific way. However Microbes prosper with advantage by our ignorance in Diagnosis and

Treatment.

D.S. Murthy from Osmania Medical College, Hyderabad India true sir. But, at the same time, Gram

stain may provide the clues, which we have to confirm by further processing. It is only a preliminary

test or the first step but not a conclusive evidence of the infecting agent. While the technique is error

prone, we can reduce the errors by standardizing the protocol in the laboratory with different

specimens and periodic quality assessment. Most errors are because of overconfidence (and resulting

negligence) than because of the technique per se.

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Sridhar Rao JJM Medical College, Davangere India Yes sir, Gram stain is the most fundamental

step in the laboratory diagnosis of bacterial infectious disease. The smear from the clinical specimen

not only provides useful information on the likely aetiology but also reveals the quality of the

specimen. Gram stained smear examination (in my opinion) is a highly useful preliminary examination

that MUST be followed on all exudate specimens (irrespective of requisition). This will help the

identification and interpretation of culture results. Unfortunately, it is overlooked in many

laboratories. Sir, as you have rightly pointed out, it helps in getting rid of misidentification of

organisms by their colony morphology alone. Many times organisms identified as Staphylococcus turn

out to be something else on Gram stained smear examination. This age-old staining technique

certainly has its limitations, but under an experienced microbiologist, it is still an indispensable tool.

T.V.Rao Travancore Medical College, Kollam India I request all the Postgraduate students to take

greater interest in doing and reading Gram's stains by themselves, Many Case reports published by

people who are keenly interested in Microscopic observations. As we are all aware we have no

modern facilities in Medical colleges, Good observation can definitely give many clues in diagnosing

Bacterial infections

Vijendra Kwale yes Post Graduate in MD Microbiology Osmania Medial College Hyderabad Sir. We

face this very frequently - colony morphologies very similar to Staphylococcus But it turns out to be

something else in the smear! , many times Diptheroids/micro & sometimes Candida too when we

cross check with the direct gram stain of sample, it does also help rule out contamination done by us

during plating. But as a PG, i must admit sir, that its very difficult at this stage for me to make out

organisms properly in directs especially when they are scanty in the sample still, we keep trying to

search more carefully and differentiate from things other than the organisms ..

T.V.Rao Travancore Medical College, Kollam Kerala IndiaVery Nice Kawle about your writing on Gram

staining, At least in 10 - 20 % of predictions by Macroscopic observation may not be true when we see

under Microscope, I have seen at many departments these matters are left to Technicians and Juniors

Microbiologists Many organisms which are just contaminants and normal flora are processed as

assumed pathogens giving a conflicting Biochemical reaction then inexperienced name them as rare

isolates for Academic presentation. Please remember human body contains 10 times more normal

floral bacteria than our own cells. I request the message known to many this is the reason where

Bacteriology reporting went wrong in many Microbiology Departments. I wish that you contribute

more good ideas for the benefit of many with good wishes

Email; doctortvrao@gmail.com