Gram Negative Cocci and Bacilli Franzielle Jowe D. Cabrera
GRAM NEGA NEGATIVE TIVE COCCI
GRAM NEGA NEGATIVE TIVE COCCI
Neisseria • Gram negative diplococci resembling __________________ • Obligate aerobes • Oxidase and Catalase (+) • Capnophilic
Neisseria • Culture media – Thayer Martin – VCN – Modified TM – VCNT – Martin Lewis – VCAnT – New York City Agar – VCAmT
ORGANISM
ASSOCIATED DISEASE/S
N. gonorrhoeae
STD – Gonorrhea Pharyngitis, Proctitis Ophthalmia Neonatorum Disseminated Gonococcal Infection Fitz Hugh Curtis Syndrome
N. meningitidis
Meningococcemia Wa t e r h o u s e - F r i e d r i c h s e n Syndrome Localized infections in the respiratory tract
Moraxella catarrhalis
Neisseria gonorrhoeae
Neisseria meningitidis
Neisseria • Diagnostic procedures – Superoxyl Test – Carbohydrate Fermentation Test GLUCOSE N. gonorrhoeae N. meningitidis N. lactamica N. sicca M. catarrhalis
MALTOSE
SUCROSE
LACTOSE
GRAM NEGATIVE BACILLI
Enterobacteriaceae • Gram negative rods • Motile via peritrichous flagella except _____ • Facultative anaerobes • Catalase (+) • Oxidase__________ • Nitrate reduction (+) • ________ fermenter
Enterobacteriaceae • Antigenic Structure – K antigen • Capsular antigen • Heat _________ • Vi antigen _____________
– O antigen • Somatic antigen / cell wall • Heat ____________ • Mainly IgM antibodies
– H antigen • Located in the flagella • Mainly IgG antibodies
MacConkey Agar • Selects gram negative bacteria and also differentiates lactose fermenters (pink-red) from non-lactose fermenters (colorless) Bile Salts, Neutral red and crystal violet – inhibit gram (+) bacteria Lactose – Carbon source Neutral red – brown in pH 6.8 to 8.0 and pink-red at pH less than 6.8
Eosin Methylene Blue • Selects gram negative bacteria and also differentiates lactose fermenters (purple color to “green metallic sheen) from non-lactose fermenters (colorless). Eosin and Methylene Blue – inhibit gram (+) bacteria Lactose – Carbon source
Hektoen Enteric Agar • Selects for stool pathogens by inhibiting normal flora coliforms and also differentiates lactose fermenters (orange) from non-lactose fermenters (green) Bile salts (high amounts) – inhibit gram (+) bacteria and gram (-) coliforms. Lactose, sucrose and salicin – Carbon source. Bromthymol blue – indicator Sodium thiosulfate – sulfur source for H2S production. Ferric ammonium Citrate - to
Hektoen Enteric (HE) Agar • Selects for stool pathogens by inhibiting normal flora coliforms and also differentiates lactose fermenters (yellow or orange) from non-lactose fermenters (green)
E. coli
Salmonella
Shigella
XLD Agar • Selects for stool pathogens by inhibiting normal flora coliforms and also differentiates xylose fermenters (yellow) from non-xylose fermenters (red) Sodium deoxycholate – inhibit gram (+) bacteria, gram (-) coliforms and swarming of Proteus Xylose – Carbon source. Phenol red – indicator Sodium thiosulfate – sulfur source for H2S production. Ferric ammonium Citrate - to
XLD Agar • Selects for stool pathogens by inhibiting normal flora coliforms and also differentiates xylose fermenters (yellow) from non-xylose fermenters (red) Citrobacter & Proteus – yellow colonies with black centers Shigella red colonies
Xylose Fermenters Non Xylose Fermenter and H 2S producer (yellow colonies) (red colonies with black centers)
–
Salmonella-Shigella Agar • Selects for stool pathogens by inhibiting normal flora coliforms and also differentiates lactose fermenters (pink-red) from non-lactose fermenters (clear) Bile Salts and Briliant green agar – inhibit gram (+) bacteria gram (-) coliforms Lactose – Carbon source. Neutral red – indicator Sodium thiosulfate – sulfur source for H2S production. Ferric ammonium Citrate - to detect H2S production.
Salmonella-Shigella Agar A. K. pneumoniae B. E. coli C. Salmonella D. Proteus E. Pseudomonas aeroginusa
Cefsulodin-irgasan-novobiocin • Also known as Yersinia Selective Agar • Selective media for Yersinia species
Enterobacteriaceae • TSI – Sugars: – pH indicator: – H2S indicator: • LIA – pH indicator: – H2S indicator: • Indole – Enzyme detected: – Medium: – Reagent: – Positive result:
TSI Results
Aerobic slant portion
Anaerobic deep portion
Alcaligenes
freundii
LIA Results
Klebsiella
• MRVP – Methyl Red • Mixed acid fermentation: • Reagent: • Positive result:
– Voges Proskauer • Detects: • Reagents: • Positive Result:
• Citrate – Enzyme present: – Culture media: • pH indicator:
– Positive result:
• Malonate – Differentiates Salmonella from Shigella by using malonate as sole source of carbon
• Urease – Culture media: • pH indicator
– Positive result: – Rapid urease producers: – Slow urease producers:
Urease Test results
• ONPG – Enzyme detected: – Substrate: – Positive Result:
• Decarboxylase – Enzyme removes _________ producing alkaline products – Culture media: • pH indicator:
• Deaminase – Enzyme that removes _______ resulting in an acidic product – Reagent: – Positive result for Phenylalanine:
LACTOSE FERMENTERS
Enterobacteriaceae • ESCHERICHIA – UTI – 90% – Neonatal meningitis – Nosocomial, bacteremia and Diarrheal diseases – Green metallic sheen on EMB IMViC: _________ – TSI: A/A with Gas – MUG (+) except E. coli O157:H7
• Pathogenic E. coli – ETEC ______________ • LT and ST • LT similar to _________ toxin • Montezuma’s revenge, traveller’s diarrhea, watery stool
– EIEC ______________ • Shigella like infection (dysentery) • Stool with RBC, Neutrophils and mucus
• EPEC _____________ – Non invasive, no toxin – Watery diarrhea with mucus, no blood
• EHEC _____________ – Verotoxic E. coli (E. coli O157:H7) – Most severe manifestation: ________ – SMAC: ___________
• EAEC ______________ – Adheres to host cells – Watery diarrhea
E. coli - Microscopic Exam
E. coli on EMB
Flagellated E. coli
Enterobacteriaceae • KLEBSIELLA – Mucoid colonies, large polysaccharide capsule TSI: A/A with gas – IMViC: - - + + – _______________– Friedlander’s bacillus, encapsulated and tends to string – K. oxytoca – indole positive
• ENTEROBACTER – UTI, Respiratory Tract and wound infections – ______________ - Most predominant isolate
Encapsulated
Klebsiella pneumoniae “mucoid colonies MacConkey Agar
E. Coli and Enterobacter aerogenes MacConkey Agar
E. Coli and Klebsiella pneumoniae MacConkey Agar
Eosin Methylene Blue
Greenish Metallic Sheen of E. coli in EMB
LATE LACTOSE FERMENTER
Enterobacteriaceae • ARIZONA – Related to Salmonella – ____________ lactose fermenter
• CITROBACTER – Resembles Salmonella except _________ positive and ________ negative – __________ - causes neonatal meningitis – __________ - UTI, pneumonia, endocarditis
• SERRATIA – ______________ - produces red pigment (____________) – Related to nosocomial infections
Enterobacteriaceae • YERSINIA – Y. pestis • Bipolar stain on ___________ • _____________ appearance • _____________ pattern on broth • Causative agent of ____________ – Vector _____________ – 3 types
– Y. enterocolitica • Motile at _______ but not at ________ • __________ colony on CIN • Causes _______________
Safety pin appearance
Yersinia pestis
In Europe, black rats and Norway rats. Widely blamed for the European plague epidemics in the Middle Ages.
Humans can contract bubonic plague when bitten by a flea that is carrying the plague bacterium In the American Southwest (prairie dogs)
• Bubonic plague occurs within a week of being bitten by an infected flea. Multiplication of the bacteria produces the characteristic "bubo" (swollen, painful lymph node). Bacteremia follows, causing death in about 75% of those affected.
Swollen lymph node
!Pneumonic plague occurs under crowded
conditions when contaminated respiratory droplets expelled by infected persons are directly inhaled by another person. This form is characterized by a shorter incubation period and greater mortality (90%).
NON-LACTOSE FERMENTERS
Enterobacteriaceae • SALMONELLA – Black colonies on SSA, BSA – Ingestion of milk, dairy products, shellfish, eggs, meat – Species: • __________– typhoid fever, meningitis, osteomyelitis • S. paratyphi A and B – paratyphoid fever • S. paratyphi C ( S. cholera suis ) – septicemia • S. enteritidis – gastroenteritis
– ___________ - tube dilution agglutination test for detection of Salmonella infection
Enterobacteriaceae • SHIGELLA – Non-motile, colorless colonies on SSA – Intestinal pathogen – causes ____________ O Ag A B C D
ONPG
CATALASE
MANNITOL
Shigella • Shigellosis or Bacillary Dysentery – watery diarrhea – bloody stool with WBC’s & mucus – fecal-oral route (human – only known reservoir) – self-limiting but highly communicable bec. of low infective dose (200 bacilli) – Gay bowel syndrome (S. flexneri)
Shigella • Reiter's syndrome is a late complication of S. flexneri infection. (3%) – pains in their joints, irritation of the eyes, and painful urination – can lead to chronic arthritis
Enterobacteriaceae • PROTEUS, PROVIDENCIA, MORGANELLA – LDA positive – Urease positive – R/A on TSI • Proteus – Swarming motility – P. vulgaris – indole ________ – P. mirabilis – indole ________
Proteus • Proteus mirabilis
• Swarming phenomenon
Enterobacteriaceae • EDWARDSIELLA – Similar to __________ on IMViC – TSI similar to ___________ – __________ - most common human species
PSEUDOMONAS AERUGINOSA • Motile and rod shaped, obligate aerobe • Sweet or grape-like, corn-tortilla like odor due to _____________ • Grows well at ________ deg C • Oxidase positive • TSI reaction ____________ • Produces blue-green pigment – Blue – – Yellow –
PSEUDOMONAS AERUGINOSA • Wound infections causing blue-green pus • Meningitis, pneuomonia, sepsis • Mild otitis media/externa ______________ • Skin lesions following bacteremia ________
BURKHOLDERIA • BURKHOLDERIA MALLEI – Causative agent of _____________ , disease of the horses
• BURKHOLDERIA PSEUDOMALLEI – Causative agent of _____________ , glander’s like disease – Also known as ______________ bacillus
• BURKHOLDERIA CEPACIA – Commonly isolated on patients with _______________
Vibrio • Facultative anaerobe • Curved, comma shaped, motile via _____ flagella • Oxidase _________ • 0129 ___________ • Halophilic except _________________________ • Sucrose fermenter except ___________________ • Alkaline loving: APW and TCBS
Vibrio • Species – V. cholerae • 2 strains ________________ • Rice watery stool • String test (+) – Reagent: _______________
• 3 serogroups: _____________________ • 2 biovars: ____________________
CLASSICAL RBC hemolysis VP Polymxyin B RBC agglutination
EL TOR
V. parahaemolyticus • 2nd most common sp implicated in gastroenteritis • 1 cause of summer diarrhea in Japan • Associated with consumption of improperly handled seafood (oysters, clams, crabs, lobster, scallops, sardines and shrimp) • Kanagawa phenomenon – _______________
• AEROMONAS HYDROPHILIA – Oxidase ____________ – _____________ hemolytic
• PLESIOMONAS SHIGELLOIDES – Oxidase ____________ – _____________ hemolytic
• STENOTR STENOTROPHOMONA OPHOMONAS S MAL M ALT TOPHILI OPHILIA A – Oxidizes _________ faster than glucose – Lavender green colony – Oxidase _______
• ACINETOBACTER – Oxidase _____ , non-motile – UTI, wound infection, diarrhea _____________ ______ - oxidixer oxidixer – _______ _____________ ______ - non-utilizer non-utilizer – _______
• ALCALIGENES FAECALIS – Oxidase and Catalase _________ – Asaccharolytic – Apple like odor
Haemophilus • Non motile • Facultative anaerobes • Fastidious _________ – Staph streak
Haemophilus – H. influenzae • • • •
AKA ____________________ Six serotypes; most commonly encountered __________ Meningitis among children <5 years old Causes epiglottitis
– H. aegyptius • AKA ______ • Causative agent of __________
– H. ducreyi • Causative agent of ___________ • Short bacilli in __________________ arrangement
Haemophilus D-ALA (Porphyrin test) H. influenzae H . parainfluenzae H . parahemolyticus H. aegyptius H. ducreyi
X FACTOR
V FACTOR
CAMPYLOBACTER • C. jejuni and C. coli (associated with gastritis and diarrhea) • Microaerophilic and Capnophilic • Optimum growth temperature ___________ • Motility __________ • Special culture media ___________ • Microscopic _____________
HELICOBACTER • Associated with __________ • Found in human stomach • Relationship between H. pylori and ulcers
BORDETELLA • Agent of whooping cough ______________ – 3 stages • Catarrhal • Paroxysmal • Convalescent
• Pertussis like syndrome ________________ • Only motile species _________________ • Special culture media ___________________
BRUCELLA • Causes ___________________ – Morning NO FEVER – Night WITH FEVER
• Infection acquired through ingestion of animal products and via direct contact • Special culture media __________________
GROWTH IN THIONINE B. abortus
B. melitensis
B. suis
B. canis
GROWTH IN BASIC FUCHSIN
LEGIONELLA • Legionnaire’s disease _______________ – Pontiac fever
• Found in both natural and artificial water sources – May be present inside free living amoebas
• Special culture media _______________
FRANCISELLA TULARENSIS • Faintly staining, gram negative coccobacilli • Causes _____________ • Special growth requirement _____________ • Infection via inhalation and via vectors ______ • Special culture media _______________
PASTEURELLA • ____________ agent of pasteurellosis • Present in the oral cavity of domestic animals (cat and dogs) • Bipolar staining, safety pin appearance, with capsule • ______________ are the most common manifestation
