Diphtheria and Neisseria



Genus Neisseria:


This genus contains several more or less fastidious species of which two, N. gonorrhea and N. meningitides are important pathogens.




Characteristics: Non-motile Gram-negative diplococci with fastidious growth requirements: capnophilics; capsulate.


Diseases:      1)   Meningitis

2)   Septicemia in absence of meningitis

3)   Chronic meningococcal arthritis


Transmission: Human pathogens; no animal reservoir. This carried in pharynx. Carriage rate in population increases during epidemics. Droplet spread N. meningitidis has several immunologically distinct capsular types (A, B, C).


Laboratory Diagnosis


Sample collection:


  1. Cerebrospinal fluid
  2. Blood for culture in septicaemia
  3. Aspirate from skin leasion or pus from infected joint
  4. Swab from nasopharynx and throat to detect carriers

Transportation of sample: Transport sample as early as possible. Never refrigerate the sample. It should be kept at 35º-37ºc


Processing of the sample:


  1. C.S.F. sample is collected in two sterile test tubes- sample no. 1 for culture and sample no. 2 for cell count , microscopy and biochemistry
  2. The smears are prepared directly from the sample if it is turbid.if slight sloudy,centrifuge the sample and sediment is used for preparation of smears and culture


Methods for identification of organism:


 Direct methods:

  1. Gram staining: Gram-negative diplococcic with adjacent sides flattened cells often in pairs and chains and some are seen intracellular in polymorphoneuclear cells.
  2. Culture: Culture media: chocolate agar

Grow well at 37°C in moist atmosphere containing 5-10% CO2

  1. Biochemical reactions: a) Oxidase test positive
  2. b) Sugar fermentation test:

Glucose   –   Acid no gas                      Maltose  –      Acid no gas


  1. Antibiotic sensitivity test


Indirect methods: Slide agglutination test, coaglutination test and latex agglutination test




Characteristics: Non-motile Gram-negative diplococci with fastidious growth requirements: capnophilics; Noncapsulated.


Diseases:     N. gonorrhea:       1)   Gonorrhea,

2)  Pelvic inflammatory disease

3)  Salpingitis in females

4)  Ophthalmia neonatorum in infants


Transmission: Human pathogens; no animal reservoir. This may be carried in genital tract, nasopharynx and anus. Spread by sexual or intimate contact.


Laboratory D1agnosis:


Sample Collection:


  1. Urethral discharge collected by calcium alginate swab
  2. Cervical swab from non-puerperal women


Transportation of sample: Samples are transported in Amies transport medium.


Methods for identification of organism:


 Direct methods:


  1. Gram staining: Gram-negative diplococcic with concave adjacent sides (kidney shape), seen intracellularly  in polymorphoneuclear cells.


  1. Culture: Culture media: Chocolate agar, Modified Thayer martin madium

Grow well at 37 c in moist atmosphere containing 5-10% co2


  1. Biochemical reactions: a) Oxidase test positive
  2.  b)   Sugar fermentation test:

Glucose   –   Acid no gas             Maltose  –      No reaction


  1. Antibiotic sensitivity test


Indirect methods: Direct fluroscent antibody test and Slide agglutination test





This genus contains many species, is widely distributed in nature, and is part of a spectrum, with Mycobacterium and Nocardia, of similar cell wall structure containing mycolic acids. The species of major importance is C. diphtheriae. This and other pathogens within the genus need to be distinguished from commensal corynebacteria.


Corynebacterium diphtheriae


Characteristic: Gram-positive, non-capsulate, non-sporing, non-motile rods, 2-6 µm in length. In Gram-stained films cells arranged as “Chinese letters” or palisades and showing irregular staining or granule formation are characteristic.  Non-fastidious, but growth enhanced by inspissated serum (Loeffler medium). Capable of aerobic and anaerobic respiration.


Transmission: Normal habitat: usually nasopharynx, occasionally skin of humans. Infection is usually spread by aerosol. Patients may carry toxigenic organisms for up to 2-3 months after infection.



Pathogenesis: Diseases is due to production of diphtheria toxin controlled by the tox gene, which is integrated into the bacterial chromosome on a lysogenic phage. When concentration of exogenous inorganic iron (Fe3+) is very low, toxin production is maximal; the selective advantage to the organism is unknown. The mode of action of the toxin is to block protein synthesis of the host cells by inactivating an elongation factor.


Diseases: Diphtheria caused by toxigenic strains of C. diphtheriae. Focus of infection may be the throat or, increasingly commonly the skin.





Commensal Corynebacteria are normally present in the throat, skin, conjunctiva and other area. These may sometimes be mistaken for C.diphtheriae and are called diphtheroids. In general, diphtheroids possess few or no metachromatic granuals and are arranged in palisade (Parallel rows) rather than in Chinese letter pattern. Some diphtheroids are indistinguishable from diphtheria bacilli microscopically and required to be differentiated by biochemical test and more reliably by virulence test.




Sample collection:

  1. Throat and nasopharyngeal swab
  2. skin swab from for diagnosis of cutaneous diphtheria


Methods for identification of orgainism:

Gram-positive, non-capsulate, non-sporing, non-motile rods


 Direct methods:


    1. Gram staining : Gram-positive rods are seen arranged as Chinese letters” or palisade( commansal corynebacteria)


  1. Albert staining: diphtherae stains green and beaded due to the presence of dark staining granules in rods. These granules,known as volutin granules or metachromatic granules, are energy storing inorgainic poly-phosphate units


  1. Methylene blue staining: diphtherae stains unevenly with dark blue staining granules in rods


  1. Culture: Culture media: Loffler’serum medium, Tellurite blood agar  (Black colony)

Grow well aerobically. Temperature range for growth is 20-40ºC


  1. Biochemical reactions:

Sugar fermentation test: ferment Glucose and maltose with production of acid

Catalase positive


  1. Animal inoculation:

        Intracutaneous injection in to guineapig leads to death of the animal.


Indirect methods:

Toxigenicity testing of C.diphtheria by  Elek gel precipitation test and   Schick test

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