Neisseria

• Gram-Negative Cocci

  • Neisseria - non-motile, non-sporeformer, aerobic organisms requiring 5-10% CO2 for growth
  • Moraxella (Moraxella catarrhalis) - formerly called Branhamella catarrhalis, Neisseria catarrhalis, separated from Neisseria because of differences in genetic composition, part of the normal flora of the oral cavity and nasopharynx, glucose non-fermenter, non-pigmented, non-hemolytic and produces gray-opaque colonies on BAP
  • Veillonella (Veillonella parvula) - small, strict anaerobe and part of the normal mouth flora

  
• Opportunistic Neisseria Pathogens

  • N. cinerea
  • N. flavescens
  • N. lactamica
  • N. mucosa
  • N. sicca
  • N. elongata
  • N. weaveri

  
• Primary Neisseria Pathogens

  • N. gonorrhoeae
  • N. meningitidis

  
• Neisseria

  • Gram-negative, kidney/coffee-bean shape with adjacent sides flattened
  • Obligate aerobes, non-motile, non-sporeformer
  • Highly susceptible to drying and exposure to unfavorable pH or sunlight
  • Most species grow on BAP except Neisseria gonorrhoeae which requires added nutrients of CAP or special enrichments
  • Catalase and oxidase positive

  
• Neisseria gonorrhoeae


  Typical Neisseria is a gram-negative, nonmotile diplococcus, approximately 0.8 um in diameter, individual cocci are kidney-shaped, when the organisms occur in pairs, the flat or concave sides are adjacent

  
• Neisseria gonorrhoeae Antigenic Structure

  • Pili (Fimbriae) - hair-like appendages that extend up to several micrometers from the gonococcal surface, enhance attachment to host cells and resistance to phagocytosis, made up of stacked pilin proteins (MW 17–21 kDa)
  • Por Protein (Outer Membrane Porin Protein) - extends through the gonococcal cell membrane, provides protection from the host's immune response, including serum complement–mediated cell death, MW 32-36 kDa
  • Opa Proteins (Opacity-associated proteins) - function in adhesion of gonococci within colonies and in attachment of gonococci to host cells, especially cells that express carcinoembryonic antigens (CD66), MW 20-28 kDa
  • Lipooligosaccharide (LOS) - in contrast to the enteric gram-negative rods, gonococcal LPS does not have long O-antigen side chains, MW 3-7 kDa, toxicity in gonococcal infections is largely due to the endotoxic effects of LOS
  • RMP (Protein III) - antigenically conserved in all gonococci, reduction-modifiable protein (Rmp) that changes its apparent molecular weight when in a reduced state, associates with Por in the formation of pores in the cell surface, MW about 30-31 kDa

  
• Gonococci elaborate an IgA1 protease that splits and inactivates IgA1, a major mucosal immunoglobulin of humans
• N. meningitidis, H. influenzae, and Streptococcus pneumoniae elaborate similar IgA1 proteases

  
• Culture Media for Neisseria

  For sterile specimen: CAP with 5-10% CO2 at 37C (Aerobic and Facultative anaerobic growth requirement)

  Selective Medium: Thayer Martin medium, Modified Thayer Martin, Transgrow Medium, Martin Lewis, New York City Medium

  
• Gonorrhea


  First described by Neisser in 1879, the most common sexually transmitted disease (STD) seen in adult, children and infants may also be affected

  
• Gonorrhea in Men


  Usually presents as acute urethritis, dysuria, purulent urethral discharge (often more severe in the morning), painful urination, may cause sterility if left untreated, the organism penetrates the mucous membranes of the urethra causing urethritis
• Gonorrhea in Men (Asymptomatic)


  Although some men will remain asymptomatic, most will complain of painful urination along with a purulent urethral discharge (pus can be expressed from the tip of the penis), possible complications include epididymitis, prostatitis, and urethral strictures
• Gonorrhea in Women

  Usually presents as vaginal discharges which are mucopurulent, dysuria, menstrual abnormalities and lower abdominal pain, may cause PID (pelvic inflammatory disease), a collective term for the inflammation of the uterus, fallopian tubes and ovary, and in some cases, may cause sterility

  
• Gonorrhea in Neonate

  Causes Ophthalmia Neonatorum, an infection in the eye of the newborn acquired from passing through an infected birth canal of the mother with gonorrhea, can be prevented by Crede's Prophylaxis

  
• Laboratory Diagnosis of Gonorrhea

  Specimen: Men - urethral discharge, oropharyngeal, rectal and prostatic discharges, Women - cervical, vaginal, urethral, oropharyngeal and rectal discharges, Note: rectal and oropharyngeal should be taken from homosexuals

  Stained smear shows a typical gram-negative diplococci, intracellular (within PMN's) which is indicative of active infection

  
• Gonorrhea Culture

  Immediately after collection, pus or mucus is streaked on enriched selective medium and incubated in an atmosphere containing 5% CO2 (candle extinction jar) at 37 °C, to avoid overgrowth by contaminants, the selective medium contains antimicrobial drugs, 48 hours after culture, the organisms can be quickly identified by their appearance on a Gram-stained smear, oxidase positivity, coagglutination, immunofluorescence staining, or other laboratory tests, the species of subcultured bacteria may be determined by fermentation reactions

  
• Biochemical Tests for Gonorrhea

  Oxidase Test - presumptive identification test, not specific from Neisseria since Pseudomonas is also detected

  Sugar Fermentation tests - glucose is the only sugar fermented by the organism (TSI - K/A no H2S)

  Superoxol Test - production of bubbles from 30% H2O2 (Catalase)

  Immunologic test - FAT, Coagglutination

  
• Oxidase Test


  Principle: To determine the presence of bacterial cytochrome oxidase using the oxidation of the substrate tetramethyl-p-phenylenediamine dihydrochloride to indophenol, a dark purple–colored end product

  Reagent: 1% tetramethyl-p-phenylenediamine dihydrochloride

  Procedure: Add several drops of the reagent into the colony on a solid medium or add 1 drop of the reagent to a filter paper and spread the colony onto it

  Result: (+) colonies become pink, then red and finally purple black
• Neisseria Transmission, Treatment, and Prevention

  Transmission: Sexual intercourse (oral, anal, vaginal), Passing through an infected birth canal, Soiled articles/clothing (rare/less likely)

  Treatment: Strain I (Penicillin Sensitive N. gonorrhoreae) - 2.5 mega units of penicillin, Strain II (Penicillin Resistant N. gonorrhoeae) - 4.9 mega units of penicillin, Strain III (PPNG - Penicillinase Producing N. gonorrhoeae) - Bactrim, Spectrin and Spectinomycin

  Prevention: Avoid sexual intercourse especially with asymptomatic carriers, Prompt treatment after early diagnosis, Education

  
• Neisseria meningitidis


  Small, gram-negative diplococcus found inside and outside the cells, Virulence Factors: Lipopolysaccharide Capsule - inhibits phagocytosis, Lipooligosaccharide Endotoxin Complex - the major virulence factor which activates the clotting cascades producing DIC and also responsible for vascular damage

  
• Neisseria meningitidis Pathogenesis

  Humans are the only natural hosts for whom meningococci are pathogenic, the nasopharynx is the portal of entry, there, the organisms attach to epithelial cells with the aid of pili and may form part of the transient microbiota without producing symptoms, from the nasopharynx, organisms may reach the bloodstream, producing bacteremia, fulminant meningococcemia is more severe, with high fever and hemorrhagic rash, there may be disseminated intravascular coagulation and circulatory collapse (Waterhouse-Friderichsen syndrome)

  
• Waterhouse Friderichen Syndrome


  A highly fatal fulminating meningococcemia characterized by lesions of disseminated intravascular coagulation (DIC) with thrombosis and acute hemorrhage in the adrenal glands
• Neisseria meningitidis Transmission

  Meningococci enter the body through the upper respiratory tract including the conjunctiva and establish on the mucous membranes of the nasopharynx, then dissemination occurs in the bloodstream

  
• Neisseria meningitidis Meningitis

  Cerebrospinal meningitis - when the organisms are brought to the CNS through the blood stream

  
• Neisseria meningitidis Laboratory Diagnosis

  Specimen: Blood, CSF, synovial fluid, pleural fluid, aspirates of petechiae, nasopharyngeal and throat swabs (for carrier)

  Stained smear

  Culture - CAP under 10% CO2

  Biochemical tests: Oxidase Test - presumptive test, CHO Fermentation Test - Glucose and Maltose fermenter in contrast to Neisseria gonorrhoeae which ferments glucose only, Serological tests (Counter immunoelectrophoresis and Particle agglutination tests)

  
• Neisseria meningitidis Culture

  Culture media without sodium polyanethol sulfonate are helpful in culturing blood specimens, Cerebrospinal fluid specimens are plated on "chocolate" agar and incubated at 37 °C in an atmosphere of 5% CO2 (candle jar), Freshly drawn spinal fluid can be directly incubated at 37 °C if agar culture media are not immediately available, A modified Thayer-Martin medium with antibiotics (vancomycin, colistin, amphotericol) favors the growth of neisseriae, inhibits many other bacteria, and is used for nasopharyngeal cultures, Presumptive colonies of neisseriae on solid media, particularly in mixed culture, can be identified by oxidase test

  
• Neisseria meningitidis Treatment

  Penicillin - drug of choice, Chloramphenicol, Ceftriaxone for penicillin allergic patients

  Prophylaxis: Rifampin and Ciprofloxacin
• Other Neisseria Species

  Neisseria lactamica - very rarely causes disease but is important because it grows in the selective media (eg, modified Thayer-Martin medium) used for cultures of gonococci and meningococci from clinical specimens, can be cultured from the nasopharynx of 3–40% of persons and most often is found in children, unlike the other neisseriae, it ferments lactose

  Neisseria sicca, Neisseria subflava, Neisseria cinerea, Neisseria mucosa, and Neisseria flavescens - members of the normal flora of the respiratory tract, particularly the nasopharynx, and very rarely produce disease, N cinerea sometimes resembles N gonorrhoeae because of its morphology and positive hydroxyprolyl aminopeptidase reaction