Gram-negative diplococci failed to grow at 48 hours in 35 degree Celsius.

An urethral swab was obtained from a man with an urethral exudate was plated directly on chocolate agar and modified Thayer-Martin agar and a gram stain was made. The culture plates were incubated at 35 degrees celsius, but showed no growth at 48 hours. The gram stain showed gram-negative diplococci.

What is a Thayer-Martin agar? A Thayer-Martin agar is a Mueller-Hinton agar with 5% chocolate sheep blood and a combination of antibiotics. The antibiotics include vancomycin, colistin, nystatin, and trimethoprim. Vancomycin kills most gram-positive organisms, except Lactobacillus and Pediococcus species are inherently resistant. Colistin is added to kill most gram-negative organisms except Neisseria. Nystatin is added to kill most fungi. Trimethoprim inhibits gram-negative organisms especially Proteus species.

A Thayer-Martin agar is used to isolate suspected pathogenic Neisseria gonorrhoeae and meningitidis.

A chocolate blood agar (CBA, CHOC) is a nonselective enriched growth media used for isolation of pathogenic bacteria. The chocolate agar is used for growing fastidious respiratory bacteria such as Haemophilus influenzae or Neisseria meningitidis. Some of the fastidious bacteria need NAD and hemin which are inside RBCs, which are lysed in the chocolate agar.

Typically when a pathogenic Haemophilus or Neisseria is suspected, the swab will first be plated onto a chocolate agar, and then a more selective media, either being the chocolate agar with the addition of bacitracin for selection of Haemophilus, or the Thayer-Martin agar for the selection of Neisseria species.

Because of the source of the swab, being from a urethral exudate, this makes Neisseria more likely so sample was restreaked onto a Thayer-Martin agar.

Neisseria gonorrhoeae is a gram-negative diplococci (which was observed on upon gram stain). It is oxidase positive, aerobic, and non-mobile. It survives within PMNs within the host and is the causative agent of gonorrhea.

The reason that it failed to grow at 35 degrees celsius after 48 hours is because Neisseria gonorrhoeae requires an environment enriched with 3-7% carbon dioxide.  This species is capnophilic meaning that it strives in higher levels of carbon dioxide.

A way to differentiate Neisseria gonorrhoeae from Neisseria meningitidis is from a carbohydrate utilization test. N. gonorrhoeae ONLY oxidizes glucose while N. meningitidis is able to ferment glucose and maltose.

 

BOC Online Study Guide; Microbiology

 

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