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					I. GRAM POSITIVE COCCI – MICROCOCCACEA
    A. General Characteristics

    B. Clinical Significance

    C. Staph aureus

           1. Toxins

                  a) Cytolytic Exfoliative Toxin

                  b) Enterotoxin

           2. Enzymes

                  a) Lipase

                  b) Hyaluronidase

                  c) Catalase

                  d) Coagulase

           3. Culture

                  a) Beta-hemolytic

                  b) Golden or yellow

                  c) Ferments mannitol

           4. Drugs

    D. Coagulase Negative Staph

           1. Staph saprophyticus

           2. Staph epidermidis

    E. Micrococcus

II. AEROBIC GRAM POSITIVE COCCI - STREPTOCOCCUS AND
ENTEROCOCCUS
    A. General Characteristics - Streptococcus


                                          1
B. Beta Hemolytic

      1. Group A - Strep pyogenes -

            a) Diseases

            b) Classic ID

            c) Treatment

      2. Group B - Strep agalactiae

            a) Identification

            b) Diseases

            c) Treatment

C. Alpha Hemolytic

      1. Strep pneumo

            a) Characteristics

            b) Pathogenesis

            c) Treatment

      2. Other Alpha Strep

D. Gamma Hemolytic

      1. Enterococcus (Group D Entercoccus)

            a) General Characteristics

            b) Identification

            c) Disease Association

            d) Treatment

      2. Group D non-enterococci

            a) Identification

            b) Diseases



                                      2
                 c) Treatment

    E. Streptococcus - Like Organisms

                 a) Leuconostoc

                 b) Aerococcus

                 c) Pediococcus

III. GRAM NEGATIVE COCCI – NEISSERIA & MORAXELLA
(BRANHAMELLA)
    A. General Characteristics

    B. N. gonorrhoeae - gonococcus - G.C.

          1. Sources

          2. Culture

                 a) Media

                 b) Biochems

          3. Antibiotics

          4. Direct Gram Stain:

                 a) Male

                 b) Female

    C. N. meningitidis

          1. Sources

          2. Culture -

          3. Identification

          4. Antibiotics

    D. Other species

          1. Normal flora



                                        3
          2. N. lactamica, sica, mucosa

    E. Moraxella(Branhamella) catarrhalis

          1. Morphologically and biochemically resemble Neisseria.

          2. Source

          3. Identification

          4. Antibiotics

    F. Other Gram Neg Cocci Identification Methods

          1. CATSCREEN

          2. Phadebact Monoclonal – GC

IV. AEROBIC GRAM POSITIVE BACILLI
    A. Organisms

          1. Corynebacterium diphtheriae

          2. Corynebacterium (diphtheroids) sp.

          3. Arcanobacterium haemolyticum

          4. Listeria monocytogenes

          5. Erysiplothrix rhusiopathiae

          6. Lactobacillus sp.

          7. Bacillus sp.

    B. Corynbacteria

          1. General Characteristics

          2. C. diphtheriae

                 a) Culture

                 b) Diagnosis

                 c) Pathogenesis


                                           4
              d) Antibiotics

       3. Diphtheroids

              a) C. species xerosis, ulcerans, equi, pseudotuberculosis, Group JK

                       (1) Characteristics

              b) Corynebacterium jeikeium (JK- Group)

C. Arcanobacterium haemolyticum

       1. Characteristics

       2. Clinical Infection

D. Listeria monocytogenes

       1. General Characteristics

       2. Clinical Infections

       3. Treatment

E. Erysiplothrix rhusiopathiae

       1. Characteristics

       2. Clinical Aspects

       3. Treatment

F. Lactobacillus sp.

       1. General

       2. Identification

G. Bacillus sp.

       1. Characteristics

       2. Clinical Characteristics.

              a) Bacillus ceres

              b) Bacillus anthracis



                                        5
                           (1) Disease

                           (2) Characteristics

                           (3) Toxin

                           (4) Treatment

V. GRAM NEGATIVE RODS - ENTERICS
    A. General

    B. Primary Intestinal Pathogens

           1. Salmonella

                  a) General

                  b) Species

                           (1) S. typhi

                           (2) S. paratyphi

                           (3) S. cholera-suis (and other strains)

                  c) Carrier state

           2. Shigella -

                  a) General

           3. Yersinia

                  a) Yersinia pestis

                  b) Yersinia enterocolitica

    C. Identification of Salmonella,Shigella, Yersinia - see Stool Culture Protocol
    Outline Handouts+ Figure in Mahon.

           1. E. coli -

                  a) Toxins - Five main categories based on virulence factors, clinical
                  manifestations produced, epidemiology, and different O:H serotypes.
                  These include:



                                              6
                       (1) Enteropathogenic (EPEC)

                       (2) Enterotoxigenic (ETEC)

                       (3) Enteroinvasive (EIEC)

                       (4) Enterohemorrhagic (EHEC) serotype 0157:H7

                       (5) Enteroadherent (EAEC)

              b) Identification

D. Opportunistic Pathogens

       1. UTI

       2. Septicemia

       3. Pneumonia

              a) Klebsiella

       4. Meningitis

              a) E. coli

       5. Blood Culture

       6. Surgical Would Infections

       7. Catheter Related Infections

       8. Hospital Acquired Infections

E. Laboratory - Identification

       1. General

F. Characteristics

       1. Gram Stain Morphology

       2. Colony Morphology

       3. Ferments Glucose - Some ferment lactose;some don’t




                                        7
                 a) Mac

                 b) HE

                 c) EMB

          4. Oxidase negative

          5. Facultative anaerobe

          6. Nitrate +

          7. Key Rx - (Described in Lab Test Handouts + Appendix A)

                 a) ONPG – O-Nitrophenyl-beta D-galactosidase

                 b) TSI - Triple Sugar Iron

                 c) Urease

                 d) Decarboxylate - Lysine, Ornithine, Arginine

                 e) IMVC

                         (1) Indole .

                         (2) Methyl Red/VP.

                         (3) Citrate

                         (4) IMVIC RX

                                 (a) E. coli + + - -

                                 (b) KES        --++

                 f) Gelatin

                 g) DNAse

                 h) PA

VI. GRAM NEGATIVE RODS - NONFERMENTERS
    A. General Characteristics

          1. Microscopic


                                            8
       2. Metabolism

       3. Epidemiology – Distribution & Infections

B. Pseudomonas (or Previously ) General Information

       1. Pseudomonas aeruginosa

       2. Burkholderia cepacia

       3. Burkholderia mallei

       4. Burkholderia pseudomallei

       5. Stenotrophomonas maltophilia

       6. Pseudomonas putida & fluorescense

       7. Pseudomonas stutzeri

       8. Brevundimonas diminuta & vesicularis

       9. Shewanella putrifaciens

C. Other Non-fermenters- General Information

       1. Acinetobacter baumani.

       2. Alcalingenes faecalis.

       3. Chryseobacterium (Flavobacterium) meningosepticum

       4. Myroides (Flavobacterium) odoratus

       5. Sphingobacterium multivorum + spiritivorum

       6. Moraxella spp.

D. Identification -Clues that you have a non-fermenter

       1. Sometimes will take a little longer to grow.

       2. Colonies are smaller.

       3. Can also use antibiogram - Resistance

       4. Lactose negative



                                       9
       5. Oxidase

       6. Do not grow on MacConkeys or grow poorly

       7. OF (Oxidation-Fermentation)

       8. Decarboxylase (Lysine)

       9. May use expections to ID

       10. Pigmentation

       11. Flagella

       12. API – NFT or NE

E. Pseudomonas (or previously) species Identification

       1. Pseudomonas aeruginosa

       2. Burkholderia cepacia

       3. Burkholderia mallei

       4. Burkholderia pseudomallei

       5. Stenotrophomonas maltophilia

       6. Pseudomonas putida & fluorescense

       7. Pseudomonas stutzeri

       8. Brevundimonas diminuta & vesicularis

       9. Shewanella putrifaciens

F. Other Nonfermenters

       1. Acinetobacter baumanni.

       2. Alcaligenes fecalis.

       3. Cryseobacterium (Flavobacterium) meningosepticum

       4. Myroides (Flavobacterium) odoratum

       5. Sphingobacterium multivorum & spiritivorum



                                      10
   G. Treatment -

         1. Pseudomonas aeruginosa - Only one with valid sensitivity protocols.

         2. Follow infection control guidelines in hospitals.

VII. CAMPYLOBACTER –VIBRIO-AEROMONAS-PLEISOMONAS-
CHROMOBACTERIUM
   A. Campylobacter sp.

         1. Epidemiology

         2. Mode of Transmission

         3. Clinical Manifestations

         4. Culture

                a) Stool

                b) Atmosphere -

                c) Temperature

                d) Media

                e) Colony Morphology -

                f) Microscopic Morphology

                g) Biochems

         5. Antibiotics -

   B. Helicobacter pylori

         1. Habitat

         2. Clinical Aspects

         3. Identification

         4. Treatment

   C. Vibrio cholera/parahemolyticus/vulnificus



                                         11
          1. Habitat

          2. Mode of Transmission

          3. Virulence Factors

                 a) Enterotoxin

          4. Spectrum of Disease

          5. Culture

          6. Treatment

    D. Aeromonas

          1. Habitat

          2. Mode of Transmission

          3. Clinical Infection

          4. Culture

          5. Antimicrobiols

    E. Plesiomonas

          1. Habitat

          2. Mode of Transmission

          3. Clinical Infection

          4. Culture

    F. Chromobacterium violaceum

          1. Habitat

          2. Culture

VIII. HEMOPHILUS & OTHER FASTIDIOUS GRAM NEGATIVE RODS
    A. Hemophilus

          1. General Characteristics


                                       12
      a) Microscopic

      b) Colony

      c) Biochems

2. Pathogenesis

      a) Hemophilus influenza

             (1) Virulence

             (2) Spectrum of Disease

             (3) Habitat

             (4) Mode of Transmission

      b) Hemophilus aegypticus

             (1) Virulence

             (2) Spectrum of Disease

             (3) Habitat

             (4) Mode of Transmission

      c) Hemophilus ducreyi

             (1) Virulence

             (2) Disease Spectrum

             (3) Habitat

             (4) Mode of Transmission

      d) Other Hemophilus species. (parainfluenza, aphrophilus,
      parahemolyticus, and paraphrophilus)

             (1) Virulence

             (2) Disease Association

             (3) Habitat




                              13
                     (4) Mode of Transmission

      3. Specimen Processing

              a) H. flu

              b) H ducreyi

              c) Hemophilus sp. - growth requirements

                     (1) X - hemin

                     (2) V - NAD

                     (3) "satellitism"

                     (4) H. ducreyi and H. aegyptius - chocolate agar supplemented
                     with 1% IsoVitale X.

                     (5) CO 2 and humidity

      4. Identification

              a) Colony Morphology

              b) Microscopic morphology

              c) Biochems

      5. Treatment -

B. Brucella

      1. Pathogenesis & Epidemiology - Zoonosis.

      2. Causes "undulant fever" – Brucellosis

      3. 4 species

              a) B. suis - swine

              b) B. abortus - cattle

              c) B. melintensis - goat & sheep

              d) B. canis - dogs



                                         14
      4. Identification

                a) Microscopic

                b) Culture

                c) Biochems

                d) Special Precautions

                e) Serology

      5. Prevention

      6. Treatment

C. Bordetella

      1. Epidemiology & Pathogenicity

                a) Bordetella pertussis causes pertussis or "whooping cough".

                b) Cilia

                c) Virulence factors:

                       (1) Capsule

                       (2) Endotoxin

                       (3) Tracheal cytotoxin

                       (4) Exotoxin

                       (5) Numerous extra cellular enzymes

                d) Other species -parapertussis and bronchiseptica

      2. Types of cultures

                a) Nasopharyngeal washings and swabs -

                b) Cough plate

                c) Media

                       (1) Bordet-Gengou



                                         15
                              (2) Regan-Lowe

               3. Culture

               4. Gram Stain

               5. Differentiation
                        B. pertussis            B. parapertussis   B. bronchiseptica
Grows on Mac            -                       +                  +
Urease                  -                       + (18 hrs.)        +(4 hrs)
Nitrate                 -                       -                  +

               6. Treatment

               7. Prevention

      D. Pasteurella multocida

               1. Epidemiology & Pathogenesis

               2. Identification

                      a) Culture

                      b) Biochems

               3. Treatment

      E. Francisella tularensis

               1. Epidemiology & Pathogenesis

               2. Identification

                      a) Culture

                      b) Biochems

                      c) Other – Fluorescent or Serology

                      d) Precautions

               3. Treatment

      F. Legionella pneumophilia

               1. Epidemiology and Pathogenesis


                                               16
            2. Identification

                     a) Microscopic

                     b) Culture

                     c) Biochems

                     d) Identification

                     e) Precautions

            3. Therapy

     G. HACEK Group - All need CO 2 and have predeliction for cardiac tissue.

            1. Hemophilus

            2. Actinobacillus

            3. Cardiobacterium hominis

            4. Eikenella corrodens

            5. Kingella sp.

            6. Capnocytophaga sp.

     H. Bartonella

            1. Epidemiology & Pathogenesis

            2. Identification

                     a) Microscopic

                     b) Culture

                     c) Biochems

IX. Anaerobes
     A. Defined - relationship to oxygen.

            1. Do not require oxygen for life and reproduction.

                     a) Moderate (aetotolerant)


                                            17
               b) Strict (obligate)

B. Specimens

      1. Proper specimen

      2. Unacceptable specimens

C. Media

      1. PRAS

      2. Brucella Agar

      3. BBE/LKV

               a) BBE

               b) LKV

      4. PEA

      5. Supplemental Enriched Thioglycollate

      6. Transport Media

D. Anaerobic Culture Conditions

      1. Anaerobic jars

               a) Gas packs + catalyst (palladium).

               b) Add 10 cc of water and produces N 2 + H2 + CO2

               Step 1: Sodium bicarb + citric acid = production of CO 2

               C6 H8O7 + 3NaHCO 3  Na3(C6H5O7) + 3H2O + 3CO 2

               Step 2: Sodium Borohydrate + Water = Production of Hydrogen

               NaBH4 + 2H2O  NaBO 2 + 4H2

               Step 3: Removal of O 2
                      catalyst
               2H2 + O2  2H2O



                                         18
              c) Redox IndicatorsAre indicators to see if gas is at right level. These
              are redox indicators. Show reduced O.

                     (1) . Methylene blue. Blue + oxidized (O 2) present, colorless no
                     O 2.

                     (2) Resazurin - pink to colorless. Pink is oxidized (O 2) present.
                     Colorless no O 2.

       2. Anaerobic biobags

       3. Anaerobic Chamber - Glove box

E. Incubation Conditions

       1. Temperature

       2. Time

F. Identification – Handout

       1. For each colony type on Brucella Agar

              a) Gram stain .

              b) Sub to Choc and incubate aerobically

              c) Sub to Brucella and incubate anaerobically – Apply ID disks

G. Are four(five) basic groups

       1. GNR - Nonsporeforming

              a) Bacteroides fragilis group - BBE

              b) Porphyromonas sp

              c) Prevotella sp.

              d) Fusobacterium sp.

       2. GPC

              a) Peptostreptococcus sp.

       3. GPR - Nonsporeformer



                                      19
             a) Actinomyces israelii

             b) Propionibacterium sp.

             c) Lactobacillus sp.

             d) Bifidobacterium sp.

             e) Eubacterium sp.

             f) Mobiluncus sp.

      4. GPR – Sporeforming – Three types: central,terminal, subterminal

             a) Clostridium group - Tetani, histolyticum, perfringens, botulinum

      5. GNC

             a) Veillonella

H. Key Characteristics of Each Group

      1. GNR - Nonsporeformer

             a) Most common is B. fragilis

                    (1) Habitat

                    (2) Culture

                    (3) Gram Stain

                    (4) Colony morphology- Bile resistant, Non-pigmented

                    (5) Biochems

                              (a) Cat+, Indole - , Esculin +

                    (6) Treatment- Resistant. May need to surgically debride.
                    Metronidazole - Flagyl Also chloramphenicol is used.

             b) Bacteroides ureolyticus

                    (1) Culture

                    (2) Colony Morphology – Bile Sensitive – Non-pigmented




                                        20
             (3) Biochems - Sensitive to K, urease -, nitrate +, indole –

      c) Prevotella sp.- (Porphyromonas)

             (1) Culture

             (2) Colony Morphology – Bile Sensitive - Pigmented

             (3) Species - Common Prevotella are intermedium – Indole +,
             lipase + and melaninogenica – indole – and lipase –

             (4) Porphyromonas asaccharolyticus, gingivalis, endodontalis
             are indole +. Not easy to ID Require GLC. ID pattern on disks
             is V=S, C=R.

      d) Fusobacterium sp.

             (1) Habitat

             (2) Gram Stain

             (3) Colony + Biochems

             (4) Species

                    (a) F. nucleatum - breadcrumb like-

                           (i) Indole +; Lipase –

                    (b) F. mortiferum - fried egg –

                           (i) Indole -; Esulin -

                    (c) F. necrophorum - nothing special-

                           (i) Indole +; Lipase +

                    (d) All Fuso are R to Vancomycin S to Colistin and
                    Kanamycin

2. GPC - Gram positive cocci

      a) Species - Peptostreptococcus magnus, micros, anaerobius

      b) Habitat

      c) Gram Stain


                               21
      d) Biochem - SPS - P. anaerobius and is sensitive

      e) Culture

      f) Treatment

3. Gram positive Nonsporeforming Rods

      a) Actinomyces israelii

             (1) Habitat

             (2) Gram Stain

             (3) Colony

             (4) Sulfur granules

             (5) Catalase negative

      b) Bifidobacterium dentium

             (1) Habitat

             (2) Gram Stain.

             (3) Biochems - Catalase -, indole -, bile esculin +

      c) Propionibacterium acnes

             (1) Habitat

             (2) Gram Stain

             (3) ID - GS , anaerobic, and catalase +. Also indole - and
             nitrate +

             (4) Treatment

      d) Eubacterium sp.

             (1) Habitat

             (2) Gram stain

             (3) All nitrate +



                                 22
      e) Lactobacillus sp.

             (1) Habitat

             (2) Biochems. - Nitrate -; indole -

      f) Mobiluncus sp.

             (1) Habitat

             (2) Gram Stain

             (3) Biochems - Are V=S; C=R

4. Gram Positive Spore Forming Rods

      a) General Info

      b) Habitat

      c) Four main pathogens

             (1) C. perfringes

             (2) C. tetani

             (3) C. botulinum

             (4) C. difficle

                     (a) Toxins –

                               (i) Entertoxin A

                               (ii) Cytotoxin B

                     (b) Culture - CCFA

      d) Biochems Used for Identifying species of clostridium

             (1) Gram Stain

             (2) Hemolytic Pattern

             (3) Biochems

                     (a) Lecithinase



                                  23
                                  (b) Lipase

                                  (c) AnIdent

X. Mycobacteria
    A. General

    B. Characteristics

          1. Acid fast- AFB

                 a)   Ziehl Neelson

                 b)   Kinyon - Cold

          2. Gram Stain

          3. Growth

    C. Mycobacterium tuberculosis complex.

          1. General Info

                 a) Includes: M. tb, M. bovis BCG, and M. africanum.

          2. Epidemiology

          3. Mode of transmission

          4. Pathogenesis

    D. Nontuberculous Mycobacteria

          1. Habitat

          2. Epidemiology

                 a) M. avium

                 b) M. gordonae

                 c) M. marinum

                 d) M kansasii

    E. Drugs: INH (Isoniazid), Rifampin, EMB (Ethambutol), and Pyrazinamide.



                                           24
F. Skin Test

G. Vaccination

      1. BCG- Bacille Calmette- Guerin

H. Types of Clinical Specimens

      1. Pulmonary

      2. Gastric lavage

      3. Urine

      4. Misc

I. Collection And Processing

      1. Prevent Contaminants.

      2. Number

      3. Decontaminate

               a) N-acetyl cysteine- liquefaction(digestion) and sodium hydroxide -
               decontamination = Gold std

               b) Zephiran-trisodium phosphate

               c) Sputolysin-Oxalic Acid

J. Media

      1. Agar based

               a) Middlebrook 7H10

      2. Egg based

               a) Lowenstien-Jensen LJ

               b) Petragnani

      3. Liquid

               a) Bactec 12 B




                                      25
              b) Middlebrook 7H9 Broth

K. Identification

       1. Grow

       2. Sub and look for

              a) Rate of growth

              b) Pigment

              c) Colony morphology

       3. Then Work up

              a) Direct Smears

                    (1) Acid fast Stains

                    (2) Fluorescent Techniques

                    (3) Molecular Techniques

              b) Rate of Growth

                    (1) Rapid growers

                    (2) Slow grower

              c) Pigmentation.

                    (1) Photochromogen

                    (2) Scotochromogens

                    (3) Nonchromogens

              d) Biochems

                    (1) M. tb complex

                             (a) Niacin, nitrate reduction +, Catalase –

                             (b) Buff colored – nonchromogenic, non photoreactive,

              e) Flow Charts



                                      26
    L. Noncultivatable - Mycobacterium Leprae

          1. Characteristics

          2. Epidemiology

          3. Pathogenesis

          4. Treatment

XI. SPIROCHETES
    A. Order: Spirochaetales; Two Families Spirochaetaceae – Two genera:Treponema &
    Borrelia and Leptospiraceae – One genus –Leptospira.

          1. Treponema

                 a) T. palladum subsp. pallidum= venereal syphilis (named after a
                 shepherd)

                 b) T. pallidum subsp. pertenue = Yaws

                 c) T. pallidum subsp -endemicus - Endemic non-venereal syphilis

                 d) T. carateum - Pinta

          2. Borrelia

                 a) B. recurrentis = Relapsing fever

                 b) B. burgdorferi = Lyme disease

          3. Leptospira - tissue spirochete

                 a) L. interrogans

    B. General Characteristics

          1. Microscopic Morphology

          2. Differences

                 a) Treponema

                 b) Borrelia

                 c) Leptospira


                                          27
C. Treponema

      1. T. pallidum subsp pallidum

                a) Venereal Syphilis

                        (1) Pathogenesis

                        (2) Diagnosis

      2. Other treponemal diseases

                a) Yaws

                b) Pinta.

                c) Non-venereal syphilis

      3. Treatment

D. Borrelia

      1. General

      2. B. recurrentis

      3. B. burgdorferi

      4. 4. Treatment

                a) Relapsing Fever

                b) Lyme disease

E. Leptospira

      1. L. interrogans

                a) Epidemiology

                b) Pathogenesis

                c) Id

                        (1) Darkfield

                        (2) Serology



                                        28
                            (3) Culture

                     d) Drugs – Penicillin, Amoxicillin, Doxycycline, Tetracycline

XII.   CHLAMYDIA - OBLIGATE INTRACELLULAR BACTERIA
       A. General Characteristics

       B. Species)

             1. C. trachomatis .

                     a) Trachoma - Endemic trachoma

                     b) Non-gonococcal urethritis & cervicitis

                     c) Conjunctivitis

                     d) LGV = Lymphogranuloma venereum

                     e) Diagnosis and recovery of C. trachomatis

                            (1) Cell culture

                            (2) Tissue section

                            (3) Cytology

                            (4) Serology

                            (5) DFA

                            (6) ELISA

             2. C. psittaci - Psittacosis

                     a) General Characteristics

             3. C. pneumoniae

XIII. MYCOPLASMA and UREAPLASMA - CELL WALL-DEFICIENT
BACTERIA
       A. Characteristics

       B. M. pneumoniae



                                               29
       C. Mycoplasma hominis and Ureaplasma urealyticum

       D. Culture

       E. Treatment – Are self-limiting and usually do not require treatment.


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