BACT211

CLINICAL BACTERIOLOGY
BATCH
2021

COLLEGE OF MEDICAL LABORATORY SCIENCE-OUR LADY OF FATIMA UNIVERSITY (LAGUNA CAMPUS)

Neisseria and Moraxella

Neisseria spp. Neisseria gonorrhoeae

• Family Neisseriaceae Characteristics:
• Kingella, Eikenella, Simonsiella, Alysiella • Only host: human
• aerobic, nonmotile, non–spore-forming, capnophilic • Gonorrhoea “flow of seed” “clap” meaning “brothel”
• can grow anaerobically if nitrite is present (alternative electron acceptor) - acute pyogenic infection of nonciliated columnar and transitional epithelium
• Gram (-) diplococci - urethra, endocervix, anal canal, pharynx, and conjunctiva
N. elongata
- STI
N. weaveri bacili
N. bacilliformis - primary reservoir: asymptomatic carrier
- Incubation period: 2-7 days
• cytochrome oxidase (+) and catalase (+) N. elongata • Men: Urethra
• normal flora: mucous membranes of the RT and UGT N. bacilliformis - 3-5% asymptomatic: AHU strain arginine, hypoxanthine, uracil
• Women: Endocervix
Primary human pathogen - 50% asymptomatic
Neisseria gonorrhoeae - complications: sterility, ectopic pregnancy, or perihepatitis Fitz-Hugh–Curtis syndrome
• gonococci • Inhibited by SPS (Sodium polyanethole sulfonate)
• not part of normal flora - not recovered in blood
• always pathogenic •Fastidious organisms - to neutralize the effect: gelatin
•Requires iron for growth • Newborns: ophthalmia neonatorum
Neisseria meningitides • gonococcal eye infection
•meningococci • Connatal infection
• result in blindness
•commensal inhabitant of URT of carriers
•invasive pathogen
WORKFLOW
Specimen Collection
• Specimen of choice:
- Men: urethra
- Women: endocervix
• Avoid use of disinfectants
• Calcium alginate and cotton swabs: inhibitory
- Dacron or rayon swabs are preferred
• extremely susceptible to drying and temperature changes direct plating

Direct Microscopic Examination

• Urogenital specimen: Gram stain
• Pharyngeal specimens: Gram stain NOT recommended commensal Neisseria spp.
• Gram (-) intracellular diplococci in pairs with adjacent sides flattened
'' Kidney bean shape”
• Gram stain with > 5 PMN/ field but no bacteria:
nongonococcal urethritis C. trachomatis or Ureaplasma urealyticum

Culture
• Medium of choice: CAP
- no growth in BAP
• Plated ASAP
Virulence factors: • Susceptible to cold temperature
1. Receptors for human transferrin - Medium should be warmed at RT before inoculation
2. Capsule Nesseria meningitides • Acinetobacter spp., Capnocytophaga spp., & Kingella denitrificans
3. Pili (fimbriae) - (+) growth on gonococcal media
• N. gonorrhoeae - oxidase and catalase tests
• Colony types: Type 1-5 •Inhibitors:
• Basis: Presence or absence of pili •Vancomycin, lincomycin: Gram (+)
• Virulent forms: Type 1 & 2 •Colistin: Gram (-)
•Nystatin, Anisomycin, Amphotericin B: yeast
4. Proteins in CM •Trimethoprim: Swarming Proteus spp.
• Antigenic variation
• Protein I: major outer membrane porin protein (Por)
• Thayer-Martin
- porA & porB: N. meningitides - Vancomycin, colistin, nystatin
- porB: N. gonorrhoeae • Modified Thayer-Martin
•Protein II (Opa)
- Vancomycin, colistin, nystatin, trimethoprim
- adherence to phagocytic and epithelial cells •Martin-Lewis
•Protein III (reduction modified protein [Rmp])
- Vancomycin, colistin, anisomycin, trimethprim
- blocks host serum bactericidal (IgG) action •New York City
- Vancomycin, colistin, amphotericin B, trimethoprim
5. Core lipooligosaccharide (LOS) or endotoxin •GC-LECT
• Major in vivo virulence factor - Vancomycin, lincomycin, colistin, amphotericin B, trimethoprim
• mediates damage to body tissues and elicits an inflammatory response
• “blebs”: outer membrane fragments released during rapid growth containing the •Incubation:
LOS • 35° C in a 3% to 5% CO2 atmosphere
CO2 incubator CO2 generating pouch candle jar
6. Lipid moiety
• differentiation from the lipopolysaccharide found in most gram (-) bacilli
7. IgA protease
• cleaves IgA on mucosal surfaces

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 BACT211
CLINICAL BACTERIOLOGY
BATCH
2021

COLLEGE OF MEDICAL LABORATORY SCIENCE-OUR LADY OF FATIMA UNIVERSITY (LAGUNA CAMPUS)

Macroscopic Examination Neisseria meningitides

• small, gray to tan, translucent, and raised Characteristics:
- T1 & T2: Bright, smaller and raised • Only found in human
- T3-T5: larger, flatter colonies • commensal & invasive pathogen
• autolytic enzyme • Incubation period: 1-10 days
- make the isolate nonviable on prolonged incubation • adhere to the nasopharyngeal mucosa leading to colonization
• endemic and epidemic meningitis and meningococcemia
Microscopic Examination • Complication: Waterhouse-Friderichsen syndrome
Waterhouse -Friderichsen syndrome
• Gram (-) diplococci • 12-48 hrs of onset: death (WFS) is a group of symptoms resulting
• Acinetobacter & Kingella sp. • Meningococcal pneumonia: serogroup Y from the failure of the adrenal glands to
•10 U Penicillin disk function normally as a result of bleeding
into the gland

Biochemical Tests WORKFLOW:

• Oxidase test
- done on all suspected isolates of N. gonorrhoeae
- Reagent: 1% dimethyl-p-phenylenediamine dihydrochloride
or tetramethyl-p-phenylenediamine dihydrochloride
•Result:
Specimen Collection:
• CSF, blood, nasopharyngeal swabs and aspirates, joint fluids
• Inhibited by SPS
Direct Microscopic Examination
• Gram (-) intracellular and extracellular diplococci
• CSF: centrifuged at 1000 x g for 10 mins

Culture
• BAP & CAP
• 35° C in a 3% to 5% CO2 atmosphere
(+): purple to black color after 10 mins
( -): no change in color Macroscopic Examination
• medium-sized, gray, and convex, and encapsulated strains are mucoid
• Carbohydrate Utilization • Green tinge under BAP
• Cystine Trypticase Agar (CTA)
Microscopic Examination
- 1% of the individual carbohydrate
• Gram (-) diplococci with adjacent sides flattened
- phenol red (indicator)
• Result:
(+) yellow: acid production Biochemical Test
• Oxidase (+)
(-): no change: no acid produced
• Carbohydrate utilization test: glucose and maltose
• γ-glutamyl aminopeptidase: (+) N. meningitidis ; (-) N. gonorrhoeae, N. lactamica,
& M. catarrhalis.
• N. lactamica: glucose, maltose, lactose
• ONPG (+)

Moraxella catarrhalis
Characteristics
• Family: Moraxellaceae
• Genera: Moraxella, Acinetobacter, and Psychrobacter

• M. catarrhalis
- opportunistic pathogen
- commensal of URT
•Infections: URTI
- 3rd most common cause of acute otitis media and sinusitis in children

Specimen Collection
• middle ear effusion, nasopharynx, sinus aspirates, sputum aspirates, or bronchial
aspirates

Direct Microscopic Examination

• intracellular gram (-) diplococci

Culture
•BAP & CAP
•Grow well at 28° C
•Inhibited by colistin

Macroscopic Examination
• smooth, opaque, gray-to-white colonies

Microscopic Examination
• gram (-) diplococci

Biochemical Tests
• Oxidase & Catalase (+)
• Carbohydrate Utilization Test: asaccharolytic
• Dnase & butyrate esterase (-)
- Tributyrin as substrate

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 BACT211
CLINICAL BACTERIOLOGY
BATCH
2021

COLLEGE OF MEDICAL LABORATORY SCIENCE-OUR LADY OF FATIMA UNIVERSITY (LAGUNA CAMPUS)

Neisseria lactamica
• nasopharynx of infants and children
• similar to N. polysaccharea encountered in meningococcal carrier surveys
• Similar colonial morphology to N. meningitides
• Carbohydrate utilization test:
- Glucose, maltose, lactose
•ONPG (+)

Neisseria subflava
• “less yellow”
• part of the upper respiratory microbiota

Neisseria elongata
•N. weaveri & N. bacilliformis
- Rod shaped neisseria
•commensals in the upper respiratory tract
•opportunistic pathogens

Commensal Neisseria Species

Neisseria cinerea
• Colonial morphology similar to T3 of N. gonorrhoeae in CAP
• CAP & BAP
•To differentiate from N. gonorrhoeae
- Susceptible to 10-μg colistin disk (≥10 mm zone of inhibition)
•To differentiate from M. catarrhalis
- Nitrite reduction (+)
- Dnase reaction (+)
•To differentiate from N. flavescens
- No yellow pigment production

Neisseria flavescens
• yellow-pigmented
• asaccharolytic
•To differentiate from N. cinerea
- (+) growth on BAP & Cap at 22° C
- Yellow colonies

Neisseria polysaccharea
• produces large amounts of extracellular polysaccharide when grown in media with
1% or 5% sucrose
• To differentiate from N. gonorrhoreae
- (+) growth at NA at 35° C
- production of polysaccharide from 1% or 5% sucrose

Neisseria mucosa
• Large, mucoid colonies
• Isolated in nasopharynx of children & young adults
• Islolated in airways of dolphins
• Carbohydrate utilization
- Glucose, maltose, sucrose, fructose
• (+) reduction of nitrate & nitrite

Neisseria sicca
• dry, wrinkled, adherent, and breadcrumb-like colonies

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