NEISSERIA Basmah almaarik Lab # 6

NEISSERIA SPECIES Contains many species; 2 are human pathogens Neisseria gonorrhoea (gonococcus) Neisseria meningitidis (meningococcus) Other species are normal flora that inhabit the upper respiratory tract of humans.

Process specimen as soon as possible Neisseria gonorrhoea  Urethral discharge  Genital swabs (servical or rectal) Neisseria meningitidis  CSF Fastidious organism very sensitive to environmental factors Process specimen as soon as possible

GRAM STAIN Direct Gram Stain: Neisseria are intracellular organisms GNDC inside and outside pus cells (PMN) (kidney or bean shaped)

CULTURE Inoculate into Chocolate Agar Basal media  at 40C (no solidification) add sheep blood Keep in water bath at 70C to hemolyse blood This releases and denatures many RBCs and serum proteins, making them more readily digestible to the bacteria Colonial morphology: Grayish white, convex (raised), colony type varies, even in one strain, due to antigenic variation morphology:Grayish white, convex (raised), colony type varies, even in one strain, due to anColony tigenic variation

Vancomycin, Colistin, Nystatin, Trimethoprim. CULTURE Thayer Martin (selective medium) Chocolate agar contain antibiotics to inhibit other bacteria and fungi VCNT Vancomycin, Colistin, Nystatin, Trimethoprim.

Non-pathogenic Neisseria grow at room temperature INCUBATION Temp 35-37 ˚C ONLY Need 5-10% CO2 to grow: CO2 envelop in anaerobic jar or Candle jar For 48 hours Non-pathogenic Neisseria grow at room temperature

IDENTIFICATION Gram stain  GNDC Oxidase  +ve Carbohydrate utilization test

IDENTIFICATION Gram stain from plate  GNDC (Kidney shaped)

IDENTIFICATION All Neisseria species are Oxidase +ve Preparation of Oxidase Test: Add filter paper on a clean slide Add 2-3 drops of oxidase reagent on filter paper Use cover slip to transfer colony onto filter paper + ve  Purple color

Neisseria meningitides Neisseria gonorrhoeae IDENTIFICATION HISS medium. Broth (CTA: Cystine trypticase agar)+ serum + sugar + indicator (phenol red) Neisseria meningitides Neisseria gonorrhoeae

YOUR WORK FOR TODAY Prepare gram stain from culture. Do oxidase test. Subculture organism to chocolate agar. Incubate in the CO2 jar. Read after 48 hours. See sugar utilization tube (demonstration) See intracellular GNDC slide (demonstration slide)

http://student.ccbcmd.edu/~gkaiser/goshp. htm New diagnosis techniques: Immunoassays DNA hybridization PCR Doc Kaiser’s microbiology website http://student.ccbcmd.edu/~gkaiser/goshp. htm

Numbers in Saudi Population 13 cases of meningococcal meningitis in 2007

Morexilla Gram stain GNDC culture 35-37c Gorw on BA, CA and nutrient A. Biochemical test Oxidase +ve No Acid production in Hiss medium test.

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