Embden-Meyerhof-Parnas

(EMP) pathway

O₂ requirements: none

Final acceptors: organic compound (pyruvic acid converted to lactic acid/mixed acids, produces strong acid products and usually gas)

Used by: anaerobes and facultative anaerobes, fermenters

Entner-Douderoff (ED)

pathway

O₂ requirements: oxygen used

Final acceptor: oxygen, water (no gas) is end product

Used by: aerobic and facultative anaerobes (nonfermenters)

What indicates

a nonfermenter?

TSI: K/K (slant/butt)

Oxidase + (some nf. are oxidase - such as aeromonas & vibrio so still need to do TSI)

Some nf. grow poorly on MacConkey but when they do, they appear as lactose nfs.

Low virulance: nosocomial infections/opportunists in debilitated/immuocompromised patients

Specific cond. or diseases: Malignancies, Instrumentation/surgery, Prolonged steroid use/antibiotics/anti-cancer therapies, Underlying metabolic/chronic disease, Burns/open wounds/lesions

Natural habitat:

Humidifiers, water baths, disinfectant/irrigating solutions, distilled water, hand creams, etc.

Anesthesia eqpt., forceps, thermometers

Moist parts of skin

Various domestic animals

Isolation pattern:

Pseudomonas areuginosa 75%

Acinetobacter species 20%

Other Pseudomonas 3.5%

Other nfs. 1%

Pseudomonas aeruginosa

General Characteristics

 Oxidase +, Grows well on MAC *atypical for nfs.* clear colonies, BAP colonies large, spreading, green, grape odor, Motile, Produces pigment pyocyanin (greenish color, metallic sheen)

Pseudomonas aeruginosa

Infections

Pseudomonas aeruginosa

Identification

Mac plate - colorless colony (lactose -)

TSI - K/K

Oxidase +

Grows @ 42°C (exception to 35-37°C normality)

Flourescent agar - detects pyocyanin (green) pigment

Very resistant to many antibiotics - use aminoglycosides

Acinetobacter species

General characteristics

Gram - coccobacillary (often misread on gram stain)

Small translucent colonies on BAP w/ammonia like odor

Non-motile

Grows well on Mac plate

Oxidase - (lacks cytochrome oxidase)

Acinetobacter species

Natural habitat

Widely dispersed in soil/water

Found in regions of human skin

May be normal flora of oropharynx/vajayjay

Acinetobacter species

Infections

Pneumonia - often related to endotracheal tubes

Endocarditis, meningitis, skin/wound

Peritonitis in patients receiving peritoneal dialysis

UTIs

Tend to be resistant to antibiotics

EVEN aminoglycosides

Haemophilus and growth factors

General characteristics

Tiny gram - rods

Fastidious aerobes

- require growth factors

- X factor - hemin, hematin

-Comes for RBCs

-Heat stable

-V factor -nicotinamide adenine dinucleotide (NAD - coenzyme in production of ATP)

- Heat liable

- Found in yeast extract, Staph/Neisseria species

Factors in media

BAP, Rabbit/Horse BAP, Chocolate, TSA and MacConkey

BAP - Both X and V factor but also contains V-inhibiting substance making V factor unavailable

Rabbit/Horse blood agars - Contain both X/V factors and lack V inhibiting substance, but are very expensive

Chocolate Agar - Produced by increasing temp. while producing BAP

- Lysis of RBCs

- Destroys V-inhibiting substance

- Contains both X/V factors

TSA plate - Basic nutrient agar w/no factors

-Discs that contain factors are placed

-X, V, and XV

-Examine for growth around disc(s)

May see small colonies of Haemophilus surrounding Staph/Neisseria species

Due to BAP providing X factor along with Staph/Neisseria providing V factor

Haemophlus influenzae

&

Haemophilus parainfluenzae

Infections

H. influenzae: meningitis

pneumonia

epiglottis

pink eye

 H. parainfluenzae: rarely pathogenic

H. influenzae

& H. parainfluenzae

Factors

H. influenzae requires both X and V factors

H. parainfluenzae only requires V factor and will grow wherever present

Haemophilus

Factor Test

Some strains of Haemophilus contain beta lactamase which:

has an enzyme that breaks down Penicillin (PCN)

Test: PCN is suspended in broth

Bacteria placed in suspension

If bac. contains beta lac., PCN is broken down into penicillinoic acid

Causes a drop in pH and detected by yellow color of broth (original color is pink/red)

Neisseriaceae

General Chars.

Gram - cocci

Aerobic/facultative anaerobic

Some species require increase in CO₂

capnophilic

Oxidase +

Catalase +

Non-motile

Some species are fastidious and require enriched media

Some non-pathogenic species make a yellow/brownish colonies xanthrophyll pigment

Normal habitat is mucous membranes

Pathogenic species

of Neisseriaceae

Neisseria gonorrhoeae (most common pathogen)

Neisseria meningitidis

Moraxella catarrhalis

Types of specimens for

pathogenic Neisseriaceae

-Swabs from mouth, anus, vajayjay, urethra of penis

-Sterile fluids such as CSF and synovial joint fluid

-Sputum specimens

-Blood cultures

Collection/Isolation

of

Neisseriaceae

Transport: Best to innoculate media immediately after collection

Specimen needs to be moist and at least room temp (Refrigeration can kill it)

Increase amt of CO₂ in specimen container (swabs with crushable chamber that produces CO₂)

Isolation media: BAP

Chocolate Agar @ 35°C w/CO₂

Modified Thayer-Martin (MTM) @ 35°C w/CO₂: Use for non-sterile specimens, medium contains antimicrobial agents that inhibit normal flora, enriched medium for isolation of N. gonorrhoeae

Heavily inoculate each tube w/org.

If bacteria contain enzymes needed to metabolize the carbohydrate, then acidic products are made

-Drop in pH = color change of phenol red indicator in medium

No color = negative

Yellow = positive

Determines if bac. has enzyme to utilize nitrate

Nitrate --> nitrite + water

2 reagents:

sulfanilic acid and dimethyle-α-naphthylamine

React with Nitrite producing

Red = positive

No color change = negative

Some Neisseria can further break down nitrite into nitrogen gas

Nitrite --> ammonia + nitrogen gas

2 reagents + a pinch of zinc powder

Zinc reacts w/nitrate producing

Red color = negative

Clear color = positive (nitrate broken down completely into nitrogen gas)

                         DEX  MAL  LAC  SUC  NO₃

N. Gonorrhoeae   +       -        -       -       -

N. Meningitis       +       +       -        -       -

N. Lactamica        +       +       +        -       -

M. catarrhalis      -       -        -        -       +

Most prevalent bacterial infection in the world

-Over 3mil new infections in US alone

STD (almost exclusively)

-Increased incidence among sexually active teens/young adults

-Increased infection of oropharynx/rectum due to change in sexual beh.

Primary infection: Anogenital (anus, urethra, cervix)

Non-genital (eye pharynx, neonates may get infection during birth)

Neisseria gonorrhoeae

Infection in Men

Visible symptoms 1-7 days following exposure

Purulent urethral discharge

Severe pain

Preliminary ID: gram stain of discharge

Not normal flora in urethra

Gram - diplococci that are intracellular (inside white blood cells)

Start Tx

Gram stain eng or unclear: ID by other means

Neisseria gonorrhoeae

Infection in Women

Symptoms

Vaginal discharge

Lower abdominal pain

Abnormal bleeding

Symptoms may go unnoticed until beginning of menstrual cycle

Vaginal discharge will contain normal flora

Gram stain is not useful in presumptive ID

Must use other means of ID

Neisseria gonorrhoeae

Complications

Disseminated gonococcal infection

results from lack of Tx

Pelvic inflammatory disease (PID)

leading cause of sterility in females

Septicemia

Disseminated arthritis

bacteria invade joints

Neisseria gonorrhoeae

Identification

Opaque, gray/white glistening colonies

Oxidase +

Catalase +

Carb. ferm., nitrate test

Molecular detection:

detects bacterial nucleic acid

more rapid than culture (hrs vs. days)

more sensitive than culture

replaces culture except for abuse or non-genital source

Neisseria meningitidis

Identification

Colonies are round, convex, glistening and blue/gray color

Aerobic

Non-motile

Oxidase +

Catalase +

Will grow on BAP whereas N. gonorrhoeae will NOT

Neisseria meningitidis

Infections

Epidemic outbreaks of meningitidis

colonizes nasopharynx initially

invades bloodstream and CNS

Sudden onset of frontal headache/stiff neck/rigidity of spine

Progression of inf. - vomiting, myalgia

Progress rapidly to shock, respiratory distress/petechial rash

Final is Disseminated Intravascular Coagulation

-Use up clotting factor so bleed out into organs

Causes Waterhouse-Frederichsen syndrome

-Bleeding in adrenal glands

-Causes rapid death

Moraxella catarrhalis

Identification

Gray/white, smooth opaque colonies

Oxidase +

Catalase +

Nitrate +

DNase +

Grows well on BAP and Choc

Moraxella catarrhalis

Infection/Tx

*normal flora of upper respiratory tract

Infections: acute bronchitis and pneumonia, maxillary sinusitis, meningitis, endocarditis

Tx: Penicillin/Ampicillin resistant

Have β lactamase enzyme

Usually treated with erythromycin, tetracycline, chloramphenicol