Clinical Bacteriology Lecture Notes PDF

Shape, Arrangement and Gram Stain Gram positive Gram positive Gram negative Gram negative cocci bacilli cocci bacilli Staphylococcus spp Corynebacterium Neisseria E. coli diphtheriae meningitidis Streptococcus spp Lis...

Shape, Arrangement and Gram Stain Gram positive Gram positive Gram negative Gram negative cocci bacilli cocci bacilli Staphylococcus spp Corynebacterium Neisseria E. coli diphtheriae meningitidis Streptococcus spp Listeria Neisseria Salmonella monocytogenes gonorrhoeae Enterococcus spp Clostridium spp Shigella Klebsiella spp Pseudomonas aeruginosa Proteus mirabilis Campylobacter spp 1 Gram-positive cocci 3 Major Genus Staphylococcus Streptococcus Enterococcus Staphylococcus  Staphylococcus Genus: Staphylococcus - It comprises 40 species, the most important ones are: 1. S. aureus 2. S. epidermidis 3. S. saprophyticus Habitat: - They are commensal in humans and animals - S. aureus : 15-30% in human, found in the nasal cavity and on the oropharyngeal tissue and its very resistant to environmental conditions - S.epidermidis: lives on the skin of humans Morphology: - Gram positive cocci - Grape like arrangement - Non motile, non sporulating - 90% are capsulated  Staphylococcus Disease: - S.aureus is the most virulent specie of Staphylococcus and causes nosocomial infections. - It causes : 1- Skin infections 2- Generalized infections (like UTI), its the number one bacteria between other staphylococcus species that causes bacteremia. 3- Osteomyelitis 4- Food intoxication  Staphylococcus Disease: - There is an increase in the infection rate by S.aureus and this is due to:  The high resistance of it to external environmental conditions  Its resistance to antibiotics  Poor hygiene - The staphylococcus species especially S.aureus developed a multi-drug resistance due to the excessive use of broad spectrum antibiotic. - The most common is MRSA (Methicillin-resistant Staphylococcus aureus) which causes most nosocomial infections  Staphylococcus Disease: Other species cause opportunistic infections - S. epidermidis: skin infections and endocarditis - S. saprophyticus : urinary tract infections  Staphylococcus Sample collection: - From pus - Expectorations (in case of RTI) - CSF (in case of meningitis) - Stool (in case of food intoxication) - Urine (in case of UTI)  Staphylococcus Culture characteristics: - Facultative anaerobe - Needs an enriched medium - Grows at 18-40ºC (Opt: 37ºC) - On blood agar, S. aureus gives colonies of golden color while other species give non-pigmented color. - Isolation on Chapman agar  Staphylococcus Biochemical traits, All are: - Catalase positive - Oxidase negative - DNAase positive - Glucose positive - Only S. aureus is coagulase positive, mannitol positive - S. saprophyticus is resistant to novobiocin Streptococcus  Streptococcus Genus: Streptococcus - There are 2 classification systems of Streptococcus 1. Lancefield classification: distinguishes them according to an antigen present on the cell wall to groups from A to H and from K to V. The Streptococcus species with this marker are called “groupable Streptococcus” and those without it are called “non groupable Streptococcus” Streptococcus Streptococcus A Streptococcus B Streptococcus Other groups S. pyogenes S. agalactia D  Streptococcus Genus: Streptococcus - There are 2 classification systems of Streptococcus 2. Classification according to hemolysis on blood agar: here we have 3 groups - Alpha hemolytic: S. pneumonia (pneumococcus), S. viridans - Beta hemolytic: S. pyogenes, S. agalactia - Gamma hemolytic: Streptococcus D  Streptococcus Genus: Streptococcus - The most important 5 species of Streptococcus are the follow:  S. pyogenes  S. agalactia  S. pneumonia  S. mutans  Streptococcus D  Streptococcus Habitat: - They are commensal - Opportunistic pathogens - S. pyogenes: nasopharyngeal cavity - S. agalactia: genital tract (in female) - Streptococcus D: digestive tract - S. pneumonia : upper respiratory tract - S. mutans: commensal flora of the mouth Morphology: - Gram positive cocci - Arranged in chains - Non motile, non capsulated except pneumococcus (which is usually present as diplococci)  Streptococcus Disease: - Streptococcus comes after Staphylococcus in being pyogenic. - The most virulent streptococcus specie is S. pyogenes  Causes red angina - S. agalactia causes: newborn infections, UTI or vaginal infections - S. pneumonia usually causes : Community-acquired pneumonia (CAP), sinusitis, otitis and meningitis especially in immunosuppressed patients - S. viridans causes endocarditis - S. mutans, a pathogen of dental caries, is known to be associated with bacteremia and infective endocarditis.  Streptococcus Sample collection: - Pus - Expectorations (RTI) - Blood (bacteremia or septicemia) - CSF (meningitis) - Urine (UTI) Culture traits: - Facultative anaerobe - Mesophillic ( lives at 37 °C except S. thermophilus that tolerates higher degrees) - Neutrophillic (neutral pH) - Exigent, need enriched or selective medium and 5% CO2  Streptococcus Culture traits: on the blood agar: - They appear as colonies with a halo around them - Alpha hemolytic Streptococcus: green color around the colonies (partial hemolysis were hemoglobin turns to beliverdin which is green) - Beta hemolytic Streptococcus give a clear zone around the colonies (total hemolysis) S. pyogenes S. pneumonia (beta-hemolytic) (alpha-hemolytic) Biochemical traits: - All are catalase negative Enterococcus  Enterococcus Genus: Enterococcus - Species: E. faecalis and E. faecium Habitat: - Commensal flora in the digestive tract and genital tract - An opportunistic bacteria Morphology: - Gram positive cocci - Arranged in pairs or chains - Non motile, non capsulated  Enterococcus Disease: - UTI - Peritonitis - Endocarditis - Septicemia - Bacteremia Culture traits: - Facultative anaerobe - Non exigent Biochemical traits: - Catalase negative - Bile esculin positive (gives black) - PCR is the only way to differentiate between E. faecalis and E. faecium  Enterococcus Enterococcus have the antigenic marker D making it similar to Streptococcus D, so to differentiate between them we must know: - All Enterococcus can grow between 10-40 °C while Streptococcus at 37 °C only - Unlike Streptococcus species, Enterococcus species can grow in culture medium containing high NaCl concentration.  Enterococcus Genus Strep A Pneumococcus Strep D Enterococcus Character Hemolysis on blood Beta Alpha Gamma Gamma agar Resistance to - - - + 65g/L NaCl Resistance to - + + + bactracin Resistance to + - + + optochin Specific Capsular Cell wall A D antigen antigen antigen  Gram-positive cocci How can I differentiate between them ? Gram-positive bacilli Gram-positive bacilli 3 Major Genus Corynebacterium Listeria Clostridium Corynebacterium C. diphtheria  Corynebacterium Genus: Corynebacterium - It includes 16 species - The most important one in humans is C. diphtheria or Klebs-loeffler bacilli Habitat: - C. diphtheria is a strict parasite for humans and transmitted between them through the respiratory tract - The reservoir is seropositive patients  Corynebacterium Morphology: - Gram positive bacilli - Under microscope it appears V-shaped or like Chinese letters - Non motile, non capsulated, non sporulating  Corynebacterium Disease: - C. diphtheria causes diphtheria, a toxi-infection associated with severe intoxication signs - It causes pseudomembranous angina or white angina - Incubation period 2-6 days - It causes bull neck , fever and fatigue - In severe cases it can be deadly. - After the infection it can cause septicemia, travel to the CNS and cause paralysis or affect the heart and cause death  Corynebacterium Culture characteristics: - Aero-anaerobic - Exigent and grows on Loeffler agar or on Blood Tellurite agar. - Colonies are obtained after 24 hours incubation under 37ºC. - Hemolytic C. Diphtheria on C. Diphtheria on Blood Loeffler agar Tellurite agar  Corynebacterium Biochemical traits: - Catalase positive - Urease negative - Indole negative - Glucose positive - Nitrate reductase positive Listeria  Listeria Genus: Listeria - Contains 10 species. - The only pathogenic Listeria is Listeria monocytogenes Habitat: - In soil , plants, water | saprophytic | lives on and gets its nourishment from dead organisms or decaying organic material - Can be present in humans and animals - Found in dairy products (Cheese, Milk)  Listeria Morphology: - Gram positive bacilli - Arranged in either groups or chains (made from 3 to 4 bacteria) - Slightly similar in shape to Corynebacterium - Motile from 20-25º by peritrichous flagella, but non motile at 37ºC - Non capsulated, non sporulating  Listeria Disease: - Listeria monocytogenes causes Listeriosis, a septicemia originating from the digestive system - In adults and infants: can cause encephalitis or meningitis - In pregnant women: the infection can be lethal to the fetus  Listeria Disease:  Listeria Culture characteristics: - Facultative anaerobe - Grows between 4ºC and 45ºC - Grows on blood agar - Colonies are transparent and β-hemolytic Listeria monocytogenes colonies on blood agar  Listeria Biochemical traits: - Catalase positive - Bile-esculin positive - Glucose positive - Lecithinase positive - H2S negative  Listeria Listeria in food: how to limit its presence - Pasteurization (30 minutes at 60ºC) can destroy Listeria in dairy products - It can be killed at acidic pH below 4 - It is sensitive to aldehydes, chlorides and iodides  Listeria It is important to know that it is psychrophilic: i.e. grows at temperature 4⁰C which imposes storage problems o Raw food carries highest risk of transmission  Listeria Avoid These Foods: Ready-to-eat Deli Unpasteurized or Refrigerated Meats, Hot Dogs Raw Sprouts Soft Cheeses Raw Milk Smoked Seafood and Sausages Only eat if Only eat if you Only eat after you Only eat if Only eat after you Pasteurized for 30 reheat cook thoroughly Pasteurized cook thoroughly minutes at 60⁰C Clostridium  Clostridium Genus: Clostridium - This genus contains more than 200 species. - Four of them are the most important and pathogenic to humans: 1. C. tetani 2. C. botulinum 3. C. perfringes 4. C. difficile  Clostridium Genus: Clostridium - All Clostridium species are big gram positive bacilli that produce endospores - The growth of this bacteria requires anaerobic conditions. - Most are motile by peritrichous flagella and are hemolytic.  Clostridium Clostridium tetani (C. tetani)  Clostridium (C. tetani) Habitat: its saprophytic in soli , commensal in the digestive tract of some animals and when released it is either sporulated or non sporulated in soil. Morphology: Gram positive bacilli, has terminal bulging spores (one at the extreme), motile bacterium (Peritrichous Flagella), non capsulated  Clostridium (C. tetani) Culture traits: - Strict anaerobe, - Cultural growth is very slow on blood agar plates. - It produces a fine film of growth - Swarming due to its motility  Clostridium (C. tetani) Biochemical traits: - Catalase Negative - Gas production Positive - Gelatin Hydrolysis Positive - H2S Positive - Hemolysis Positive - Nitrate Reduction Negative - DNase Positive  Clostridium (C. tetani) Disease: tetanus - C. tetani is not an invasive bacteria, the infection is limited to the site infected. - The germination and multiplication of the spore will give the vegetative form of the bacteria - It secretes tetanus toxins which includes - Tetanolysin, responsible for hemolysis, necrosis and cardiotoxicity - Tetanospasmine, the essential cause of the neurological toxicity where it interferes with the motor neurons leading to the aggressive muscle spasms and rigidity including the muscles of the jaw causing the characteristic symptom (lock jaw)  Clostridium Clostridium botulinum (C. botulinum)  Clostridium (C. botulinum) Habitat: in the intestines of animals and in canned food. Morphology: Gram positive bacilli, has sub-terminal bulging spores, motile bacterium (Peritrichous Flagella), non capsulated  Clostridium (C. botulinum) Culture traits: - Strict anaerobe, - Culture may be applied on blood agar plates. - It produces large semi-transparent colonies with a wavy outline. - Most strains are beta hemolytic  Clostridium (C. botulinum) Biochemical traits: - Catalase Negative - 20% Bile Negative - Gelatin Hydrolysis Positive - H2S Positive - Hemolysis Positive (beta-hemolytic) - Indole Negative - Oxidase Negative - Glucose Positive - Galactose Negative  Clostridium (C. botulinum) Disease: - The agent that causes botulism, - It causes food intoxication - Botulinum toxin is the most powerful bacterial toxin, 0.1 microgram of it can cause the death an adult human, - Microinjections of diluted amount of it is used as botox (2-3 months after extraction)  Clostridium Clostridium perfringens (C. perfringens)  Clostridium (C. perfringens) Habitat: - It is found in the soil or the digestive tract of humans and animals. - 5% can be found in the female genital tract.  Clostridium (C. perfringens) Morphology: - An anaerobic Gram Positive bacilli, the spores in this specie are rare. - They lack flagella (Atrichous).  Clostridium (C. perfringens) Culture traits: - Strict anaerobe - Hemolytic - Culture on a TSC agar containing egg yolk gives yellowish gray colonies with 2-4 mm opaque zones caused by lecithinase activity. - Culture of sheep blood agar shows the presence of hemolytic zones (double or single) around small gray to gray-yellow colonies with a glossy indicated production of hemolysin.  Clostridium (C. perfringens) Biochemical traits: - Catalase Negative - 20% Bile Positive - Gas Positive - Gelatin Hydrolysis Positive - 2% NaCl Positive - 6.5% NaCl Negative - Hemolysis Positive (double hemolysis zone) - Indole Negative - Oxidase Negative - Glucose Positive - Galactose Positive  Clostridium (C. perfringens) Disease: - Gas gangrene - Mild Gastroenteritis (C. perfringens type A)  Clostridium (C. perfringens) Disease: - Gas gangrene: Clostridium perfringens is found nearly everywhere. As the bacteria grow inside the body, it makes gas and harmful substances (toxins) that can damage body tissues, cells, and blood vessels. Gas gangrene develops suddenly. Gas gangrene is a highly lethal infection of soft tissue. This is synonymous with myonecrosis and is characterized by rapidly progressive gangrene of the injured tissue along with the production of foul-smelling gas. The clostridial organisms produce alpha and theta toxins that cause extensive tissue damage.  Clostridium Clostridium difficile (C. difficile)  Clostridium (C. difficile) Habitat: - Found in soil and water and also detected in hospitals. - The active form is present in human intestine.  Clostridium (C. difficile) Morphology: - The germ is a Gram positive anaerobic bacilli. - The germ is a spore forming. - The germ is capsulated and motile by peritrichous flagella.  Clostridium (C. difficile) Culture traits: - Strict anaerobic, grows at 37°C - Needs a selective medium TCCA (Taurocholate Cycloserine Cefoxitin Agar); A selective and differential agar medium that contains cycloserine, cefoxitin, fructose, and egg yolk was developed to facilitate the isolation of C. difficile from fecal specimens. - Under UV radiations colonies present a green fluorescence  Clostridium (C. difficile) Biochemical traits: - Catalase Negative - Gelatin Hydrolysis Positive - H2S Positive - Urease Negative - VP Negative - Nitrate Reduction Negative - Hemolysis Negative - Indole Negative - Oxidase Negative - Glucose Positive - Galactose Negative  Clostridium (C. difficile) Disease: - C. difficile is a spore-forming, Gram-positive anaerobic bacillus that produces two exotoxins: toxin A and toxin B. - It is a common cause of antibiotic-associated diarrhea (AAD) and accounts for 15 to 25% of all episodes of AAD. Prolonged use of antibiotics  Clostridium (C. difficile) Disease: - It causes opportunistic infections - In case of using a broad spectrum antibiotic for a long time, - It secretes 2 toxins A and B Toxin A: an enterotoxin that causes cytokine release and hemorrhagic necrosis. Toxin B: is a cytotoxin that causes tissue damage - Symptoms are diarrhea, fever, abdominal pain - The diarrhea becomes strong leading to mega-colon and then pseudomembranous colitis. Clostridium (Endospores) Gram-negative cocci Gram-negative cocci 1 Major Genus Neisseria Neisseria meningitidis Neisseria gonorrhoeae  Neisseria Genus: Neisseria - Most of the bacteria in this family are commensal except for two species that are pathogenic:  N. meningitidis (causes meningitis)  N. gonorrhoeae (causes genital tract infection).  Neisseria meningitides (N. meningitidis) - Also called meningococcus - It is a strict parasite for humans and cannot survive outside the human body  Neisseria meningitides (N. meningitidis) - Morphology: coffee bean shape, diplococci, non motile and capsulated.  Neisseria meningitides (N. meningitides) Culture traits: - Strict aerobic, need specific temperature (35-37⁰) at 5% CO2. - Grows on chocolate agar, incubated for 24 – 48 hours and gives mucoidal colonies - Needs humidity to grow.  Neisseria meningitides (N. meningitides) Biochemical traits: - Catalase Positive - Oxidase Positive - Nitrate Reductase Negative - Maltose Positive (difference with N. gonorrhoeae) - Glucose Positive - DNAse Negative - Colistin Resistance  Neisseria meningitides (N. meningitides) Disease: - It is transmitted by inhalation or prolonged contact between humans - The incubation period of it is 2 – 4 days - Causes rhino pharyngitis, but in some cases it can reach the blood and then the CNS leading to meningitis. - Meningitis by N. meningitidis may not be severe but it can be fatal if not treated. - It can lead to the Water house friderichsen syndrome  Neisseria meningitides (N. meningitides) Disease: - Meningococcemia is a life-threatening infection that occurs when the bacteria Neisseria meningitidis invades the blood stream. - Waterhouse-Friderichsen syndrome (WFS) is a group of symptoms resulting from the failure of the adrenal glands to function normally as a result of bleeding into the gland. - Adrenal glands, also known as suprarenal glands, are small, triangular-shaped glands located on top of both kidneys. - Adrenal glands produce hormones that help regulate your metabolism, immune system, blood pressure, response to stress and other essential functions.  Neisseria meningitides (N. meningitides) Sample collection: - CSF - Rhino pharyngeal sample - Blood  Neisseria meningitides (N. gonorrheae) - It is a strict parasite for humans. - Very sensitive to environmental conditions - In the past it used to be the number one cause of STDs (Sexual Transmitted Diseases), now it comes in second after Chlamydia.  Neisseria meningitides (N. gonorrheae) Morphology: - Coffee bean shape, diplococci and non- capsulated. - Motile: gonorrhoeae cells possess a form of motility that enables them to easily explore surfaces. This type of motility is called “twitching”. Twitching motility is powered by cycles of elongations and retractions of pili. - In severe infections it can be intracellular and in clusters  Neisseria meningitides (N. gonorrheae) Culture traits: - This organism is typically cultured using an agar medium such as chocolate agar plate. - N. gonorrhoeae requires enriched media in a CO2 – (carbon dioxide- enriched (5%) atmosphere) atmosphere at 35 – 37 ºC for growth.  Neisseria meningitides (N. gonorrheae) Biochemical traits: - Catalase Positive - Gas Production Negative - H2S Negative - Hemolysis Negative - Nitrate Reductase Negative - Oxidase Positive - Glucose Positive - Lactose Positive - Maltose Negative  Neisseria meningitides (N. gonorrheae) Sample collection: - Blood - Conjunctiva - Reproductive tract  Neisseria meningitides (N. gonorrheae) Disease: it causes genito-urinary infections  In males : it can cause urethritis  In females: it can cause vulvo-vaginitis, dysuria, endo-cervical infection  In newborns: Conjonctivitis  Neisseria meningitides (N. gonorrheae) - Disease : it causes genito-urinary infections  Neisseria meningitides (N. gonorrheae) Transmission: 1- Sexual transmission 2- During delivery: the infection is transmitted from the mother to her newborn causing conjunctivitis Gram-negative Bacilli  Enterobacteria Family: Enterobacteriaceae It is a large and heterogeneous group of gram negative bacilli that comprises more than 30 genera. The clinically significant species fall into less than 25 species. Despite being pathological , they are widely used in bio industry (fermentation, pharmaceutical industry, production of antivirals)  Enterobacteria Habitat 1. Some species are saprophytic and live in the environment (soil and water) like: Proteus, enterobacter Serratia. 2. Most of the species are commensal and isolated from humans and animals particularly from the intestine. 3.They are referred to as the fermentative flora of the intestine - These flora have a role in digestion and gas production. E.Coli presents 80% of the aerobic flora, in addition to Klebsiella, Proteus and Enterobacter that are less frequent. - These commensal flora can also be transitory flora and inhabit the moist regions in the buccal cavity, nasal region and the female genital organs  Enterobacteria Pathogenesis - Opportunistic Pathogens Cause infection in immunosupressed Example: Klebsiella spp - Opportunistic in a different location Example: E.coli causes UTI when it leaves from the intestine to the urinary tract Klebsiella causes RTI and UTI if it reaches these regions - Strict pathogens Mostly digestive tract infections Other: Salmonella typhi causes Typhoid fever Yersinia Pestis Causes Plague  Enterobacteria Seven criteria apply to the bacteria (with exceptions) 1. All Enterobacteria are gram negative bacilli with average size 2. Non exigent, grow on usual culture media at 37 degrees (except for Yersenia enterolitica) 3. They are oxidase negative 4. They are nitrate reductase positive 5. Facultative anaerobes 6. Ferment glucose ( W or W/O gas production) 7. Most are non motile, except some species that are motile by peritrichous flagella.  Enterobacteria Culture traits: - 2 culture media are used: Drigalski agar and EMB agar - Drigalski: selective for gram negative species and enterobacteria. It contains inhibitors for gram positive bacteria which are Crystal violet and sodium deoxycholate The PH indicator in it is bromothymol blue which gives a yellow color when PH is acidic (upon lactose fermentation) - EMB: specific for enterobacteria contains eosine and methylene blue which are inhibitors of Drigalski agar the growth of gram positive bacteria  Enterobacteria: Salmonella Habitat - It is a strict parasite for humans, mammals and birds. Culture traits - Facultative anaerobic - On EMB agar : it gives transparent colonies (lactose negative) - On SS agar: it gives black colonies. - On DCL and Hekteon agar : gives also black colonies since they detect H2S Biochemical traits - Catalase positive - Oxidase negative - Gelatinase negative - Nitrate reductase positive - Glucose positive with gas production - Lactose negative - Succrose negative - Manitol positive - H2S positive - Indole negative - Urea megative  Enterobacteria: Salmonella Pathogenesis - After entry through the oral route it leads to several infections termed (Salmonellosis): - Typhoid and Para-typhoid fever - Gastroenteritis - Collective food toxi-infection - Mode of contamination: ingestion of contaminated food like dairy products, eggs, meat or contaminated water  Enterobacteria: Salmonella Typhoid and Para-typhoid fever - They are severe cases ( also called severe or major salmonellosis) and cause death if not treated - The clinical symptoms are: high fever, intestinal irritation (severe and bloody diarrhea) or septicemia - Diagnosis is through a stool or blood sample ( in case of septicemia) Salmonella gastroenteritis - It is minor salmonellosis that is non invasive and does not usually cause septicemia only diarrhea - Detected by copro-culture and treatment is basically rehydration. Collective food toxi-infection (CFTI) - The simultaneous consumption of contaminated food by many people - Incubation period is 10 -18 hours and trouble for 2-5 days - Diagnosed by copro-culture and treatment by rehydration  Enterobacteria: Shigella Habitat - Only parasitic to humans Morphology - Gram negative bacilli - Non motile - Non exigent on ordinary agar - Grows at 37 degrees within 24 hours - Colonies are average in size, round, regular and brilliant - Unlike Salmonella there is no specific enriched medium for Shigella Biochemical traits - Urease negative - Deaminase and lysine decarboxylase negative - H2S negative - Glucose positive W/O gas production - Simon citrate negative - Acetoin production negative - Very close to E.coli but lactose fermenter  Enterobacteria: Shigella Pathogenesis - Causes bacillary dysentery or Shigellosis - Transmission: by fecal- oral route - The most virulent is Shigella dysentery or shiga bacilli - It’s the only one that secretes the exotoxin (Shiga toxin) - Symptoms: fever, diarrhea, paroxysmal colitis, abdominal pain - Stool is mucoidal, bloody and with pus - Other Shigella species also cause infections but they are often milder.  Enterobacteria: Yersinia Habitat - They are environmental bacteria present in soil, water and vegetables - Can be present in animals but rarely in humans Morphology - Gram negative bacilli - Non motile, non sporulating, non capsulated Pathogenesis - Many species cause Yersiniosis - Y. Pestis: plague - Y. pseudotuberculosis - Y. entercolitic: intestinal infections  Enterobacteria: Yersinia Culture traits - Grows on ordinary media - Thermodepandent and can’t tolerate any temperature change - Optimal growth : 25⁰- 30⁰c - Slow growth at 37⁰c - Some are psychrophillic and grow at 4⁰c - Specific media: CIN agar, incubation for 24 hours at 30⁰- 32⁰c where it gives colonies transparent at the edges and red in the center - SSDC agar: will give big transparent colonies Biochemical traits: - Oxidase and lactose negative - Catalase positive - Nitrate reductase positive - Glucose positive W/O gas production - ONPG positive - Urease positive - Sucrose and sorbitol positive  Enterobacteria: E.coli Habitat - It’s a commensal bacteria found in the digestive tract of humans and animals - It constitutes 80% of the aerobic flora Morphology - Gram negative bacilli - Mostly motile Culture traits - Grows on ordinary culture media with lactose - On solid agar selective for enterobacteria they appear after 18-24 hours as rounded regular edged soft colonies - On EMB agar they form metallic green colonies - On Macconkey it gives pinkish colonies  Enterobacteria: E.coli E.Coli on EMB agar E.Coli on Macconkey agar  Enterobacteria: E.coli Biochemical traits - Indole positive - Lactose positive - Citrate negative - Urease negative - MR+/VP- - Glucose positive with gas production - Deaminase negstive  Enterobacteria: E.coli Pathogenesis - It is an opportunistic pathogen - Some species are related to human and animal diseases such as diarrhea, UTI, septicemia… - E.coli species that cause diarrhea are divided according to the symptoms they cause to: - Enterotoxigenic E.coli - Enteroinvasive E.coli - Enterohemorrhagic E.coli - Enteroaggregative E.coli - Other species are responsible for extra-intestinal infections like UTI, meningitis and septicemia  Enterobacteria: Proteus mirabilis Habitat - Commensal in the digestive tract of humans - It is the second agent causing UTI after E.coli Morphology - Gram negative bacilli - Motile Biochemical traits - Urease positive - Tryptophane deaminase positive  Enterobacteria: Klebsiella Habitat - Comprises many species ( K. pneumonia, K. oxytoca…) - Commensal in the digestive tract and the upper respiratory tract - It is the third agent causing UTI after E.coli and Proteus mirabilis - In some cases it can cause lower respiratory tract infections Biochemical traits - Urease negative - MR-/VP+ - Lactose positive Morphology - It has a polysaccharide capsule that protects it from antibiotics and phagocytosis, and this makes Klebsiella pneumonia on the colonies appear mucoidal Macconkey agar  Haemophilus inflenzae Genus: Haemophilus - Family: Pasteurellacae - Comprises 16 species, strict pathogenic in humans is H. inflenzae Habitat - Commensal in the upper respiratory tract Morphology - cocco-bacilli - Non motile, non sporulating - Virulent is capsulated, non virulent is non capsulated ( commensal opportunistic pathogen) - Its difficult to visualize the capsule  Haemophilus inflenzae Cultural traits - Facultative anaerobe - Grows on chocolate agar - Exigent, needs a specific medium with the presence of 2 factors : Factor X ( hemin extracted from blood) and Factor V ( NAD extracted from yeast) - Grows after 24-48 hours of incubation at 37 degrees - Colonies appear clear  Haemophilus inflenzae Pathogenesis - There are 2 types of Haemophilus inflenzae 1. Non capsulated: its commensal in the RTI, responsible for local complications in immunocomprimised ( rhino pharyngitis that can complicate to sinusitis and otitis) 2. Capsulated: especially type B that causes more than 95% of invasive manifestations, mostly meningitis and epiglottitis. It can also cause septicemia and less likely arthritis. - Sample : non capsulated form can be collected from bronchial secretions, pus and expectorations. The sample of the capsulated form can be collected from blood and CSF  Haemophilus inflenzae Diagnosis - It is usually direct - They are grown on 2 petri-dish, one with the 2 factors and the other without - The colonies that grow in the presence of 2 factors : H. influenzae - The colonies that grow without the factors: Para-influenza Para H. influenzae influenzae Factor X Factor V Factor X and V  Campylobacter Genus : Campylobacter - Comprises 17 species, the most important are: C. jejuni, C. coli, C. fetus Habitat - Commensal in many birds and mammals Morphology - Spiral - Motile by polar flagella - Non sporulating  Campylobacter Culture traits - Microaerophillic - Needs specific atmospheric conditions: 5%[O2], 10%[CO2], 85%[NO2] - Exigent bacteria - Needs either Colombia agar or specific medium CAMP - Optimal growth at 42ºc Biochemical traits - Catalase positive - Oxidase positive - Nitrate reductase positive - Glucide negative ( can’t degrade lipids) - Direct diagnosis is based on detection of bacteria in stool or blood  Campylobacter Pathogenesis - Transmitted by ingestion of contaminated poultry, food or water - C.coli and C. jejuni are mainly responsible for intestinal infection and diarrhea - The are the main cause of diarrhea, followed by E.coli - The symptoms are: bloody, mucoidal and watery diarrhea in addition to fever, nausea and vomiting - Symptoms last 1-10 days - C. fetus is responsible for extra intestinal manifestations  Bordetella pertussis Genus : Bordetella - The main species identified are B. pertussis, B. parapertussis, B. bronchiseptica, B. avium and B. hinzii Morphology - Cocco-bacilli - Motile with pilli - Capsulated Culture traits - Strict aerobic with optimal growth at 35-37ºc with incubation 2-3 days - Exigent, needs medium with nicotinamide ( Bordet-gengou agar) - Colonies are small with hemolysis Biochemical traits - Catalase and oxidase positive - Glucide negative  Bordetella pertussis Pathogenesis - It causes respiratory tract infection in enfants (whooping cough) - The RTI leads to severe complications like pneumonia, encephalitis which can be deadly in weak patients - Transmission is by direct contact or inhalation - Vaccination dramatically decreased the incidence of infection - Incubation period lasts from few days to several weeks Sample - Mucus taken from patients with bronchitis  Helicobacter pylori Genus : Helicobacter - The principle identified bacteria is H. pylori Habitat - Exclusively present in the digestive tract of humans ( stomach and duodenum) - It is present in : 90% of patients with duodenal ulcer and 80%of patients with gastric ulcer - It is the only bacteria known to live in acidic conditions Morphology - Gram negative bacilli - Helicoidal form - Non capsulated and non sporulating - Motile with peritrichous flagella  Helicobacter pylori Culture traits - Microaerophillic - Grows on chocolate agar or PYL agar ( selective for H. pylori) or colombia agar with blood added to it - Grows at 37ºc it gives small colonies that appear after 4 days of incubation Biochemical traits - Urease + + + - Catalase and oxidase positive  Helicobacter pylori Pathogenesis - H. pylori is the most common type of chronic infection in humans that can persist for decades or even the whole life - Its a strict pathogen and the main etiology of duodenal and gastric ulcer - In most individuals the infection is asymptomatic - 10% of people affected have ulcers and 1% have cancer  Helicobacter pylori Diagnosis: 1. Urea breath test:  Helicobacter pylori Diagnosis: 2. Serodiagnosis : search for antibodies against H. pylori IgG by ELISA 3. Invasive method: take a biopsy and analyze it under microscope 4. Direct diagnosis: searching for the bacteria in a stool sample Treatment - The triple therapy is used as a first line treatment and it consists of givine a combination of 2 antibiotics ( Clarithromycin with either metronidazole or amoxicillin BID plus a PPI for 14 days)  Vibrio cholera Genus Vibrio - Family: vibrionaceae - Most important specie V. cholera Habitat - Found mostly in water - Has high resistance to environmental condition - Found in the stool of seropositive patients Morphology - Gram negative bacilli - Comma shaped - Motile with monotrichous flagella  Vibrio cholera Culture traits - Facultative anaerobe - Grows on medium containing 3% [NaCl] (specific for V.cholera and PH 8.5 (basic) - The culture media used are: TCBS (thiosulfate, citrate, bile, succhrose) in which colonies appear yellow; other media NAA (nutritive alkaline Agar) Biochemical traits - Oxidase positive - Indole positive - Glucose positive - Nitrate reductase positive Diagnosis - By copro-culture  Vibrio cholera Pathogenesis - It is a strict human disease that is non invasive - Patients mostly at risk are those having a weak immune system (enfants and elderly) - The diarrhea is watery (like rice water) and contains mucus, epithelial cells and many vibrions - Treatment is by rehydration (usually antibiotics are not used). It decreases mortality. - In absence of treatment the disease can be lethal in 50% of cases  Pseudomonas aeruginosa Genus pseudomonas - Family pseudomonadaceae - It comprises 42 species. P. aeruginosa is the most encountered in infections Habitat - Survives in environment, soil, water and on moist surfaces - Can sometimes be found in the digestive tract flora Leads to nosocomial infections Morphology - Gram negative cocco-bacilli - Motile - Non capsulated, non sporulating  Pseudomonas aeruginosa Culture traits - Strict aerobic, non exigent - Grows on blood agar within hours and colonies appear metallic - On macConkey it gives whitish-beige colonies without changing the color of the medium ( lactose negative) - On Muller Hinton agar colonies appear green - Cetrimide agar for pseudomonas gives greenish color due to the pigments Pyoverdine and pyocyanine On Muller Hinton On Macconkey On blood agar  Pseudomonas aeruginosa Biochemical traits - Oxidase positive - Lactose negative - Nitrate reductase positive - Glucose negative - Indole negative - H2S negative - Arginine dehydrogenase positive - Urease positive - Mannitol positive Diagnosis - On Cetrimide agar it gives greenish colonies - P. aeroginosa is the only specie that lives at 41 degrees - P. fluorescens is used in detecting pollutants  Pseudomonas aeruginosa Pathogenesis - It is an opportunistic pathogen - Can cause : sinusitis, otitis, ocular infections, infect burn wounds, UTI - It fabricates an endotoxin that can cause septicemia  Acinetobacter Genus Acinetobacter - It comprises almost 26 named species ( A. baumannii , A. caloaceticus) - Most clinically important is Acinetobacter baumannii Morphology - Gram negative cocco-bacilli - Pleomorphic - Non motile, non sporulating Habitat - Ubiquitous in soil and on water surfaces - Colonizes the skin, respiratory and oro-pharyngeal secretions - It is one of the ESCAPE* organisms  Acinetobacter Culture traits  Strict aerobic , non fastidious  Incubated at 37 degrees under aerobic conditions  Grows on nutrient agar, blood agar, and MacConkey agar plates  On blood agar, nonhemolytic, opaque, circular, and gray color colonies will be observed.  On Macconkey agar, non-lactose, fermenting, opaque, and circular colonies will be observed  Acinetobacter Biochemical traits - Catalase positive - Oxidase negative - Urease negative - Indole negative - H2S negative Sample collected - Depends mainly on the site of infection - Blood - Urine - CSF - Sputum  Acinetobacter Pathogenesis - It is an opportunistic pathogen - Associated with hospital acquired infections - It targets moist tissues like the mucous membranes or exposed skin areas - Leads to skin infections that have a characteristic ( peau d’orange) appearance initially, then a sand –paper like presentation that eventually leaves clear vesicles - Can also cause UTI, meningitis and respiratory tract infections - If untreated it can lead to septicemia and probably death - The antibiotic resistance of Acinetobacter buamannii has been dramatically increasing in the past decades.  Brucella melitensis Genus Brucella - Family: Brucellaceae - 10 species are identified, 4 of them are pathogenic to humans : B. melitensis, B. suis, B. abortus, B. canis Habitat - Parasite for mammals - Brucellosis is a type of zoonosis - Transmission between humans is rare, its mostly from animals to humans Morphology - Gram negative cocco-bacilli - Non motile, non sporulating, non capsulated  Brucella melitensis Culture traits - Strict aerobic - Colonies take time to appear 48hours – 4days (binary fission is slow) - Culture media must be enriched by: thiamine, niacin amide, biotin - Grow on Colombia agar or chocolate agar at 34-35ºc - Some species grow at 5-10% [CO2] Biochemical traits - Catalase positive - Oxidase positive - Urease positive - Nitrate reductase positive  Brucella melitensis Pathogenesis - Transmission can occur either directly, by contact between animals and humans, or indirectly through the digestive tract by ingestion of contaminated dairy products - Farmers and butchers are mostly at risk - The disease is called Mediterranean fever or Malta fever - It can also cause generalized infections or septicemia Diagnosis - Direct: by hemoculture in case of septicemia, growth is usually slow (2-4 days) - Indirect : based on serdiagnosis i.e searching for anti-brucella antibiodies in the serum. >1/40 is considered significant