Gram Positive Bacteria - Microbiology - Lecture Slides, Slides for Microbiology. Aliah University
jojy30 January 2013

Gram Positive Bacteria - Microbiology - Lecture Slides, Slides for Microbiology. Aliah University

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Gram Positive Bacteria, Medical Importance, Endospores, Clostridium, Perfringens, Bacillus Anthracis, Central Endospores, Clostridium Tetani, Bacillus Anthracis, Bacillus Cereus are some points from these slides of Micro...
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The Gram-Positive Bacteria

Chapter 19

The Gram-Positive Bacilli of Medical Importance


Bacillus anthracis With central endospores

Clostridium perfringens With subterminal endospores

Clostridium tetani With terminal endospores

A reminder: Some Gram- positive soil bacteria are capable of producing endospores.


• gram-positive, endospore-forming, motile rods • mostly saprobic • aerobic & catalase positive • versatile in degrading complex macromolecules • source of antibiotics • primary habitat is soil • 2 species of medical importance

Bacillus anthracis Bacillus cereus


Bacillus anthracis • large, block shaped rods • central spores that develop under all conditions

except in the living body • virulence factors – capsule & exotoxins • 3 types of anthrax

Cutaneous – spores enter through skin • black sore- eschar; least dangerous

Pulmonary –inhalation of spores – Gastrointestinal – ingested spores

• treated with penicillin or tetracycline • vaccine – toxoid 6X over 1.5 years; annual boosters • dead livestock burned/chemically decontaminated

before burial to prevent soil contamination 5

Bacillus anthracis

• Biological warfare threat. • Letter attacks of 2001

– Resulted in only 22 cases of anthrax and five deaths

– Great disruption and concern – More than 32,000 people who

may have come into contact with the letters were given prophylactic antibiotics.


Anthrax- Domestic Bioterrorism • Starting one week after the 9/11/2001 attack letters

containing weaponized anhtrax were sent to media offices and to Democratic Senators Tom Daschle and Patrick Leahy -- the two individuals expected to resist passage of the USA PATRIOT ACT.


•The attacks came in two waves of letters with Trenton, New Jersey postmarks:

•the first, with postmarks of Sepmtember 18, were sent to ABC, NBC, CBS, the New York Post, and the National Equiror at American Media Inc. •the second, with postmarks of October 9, were sent to Senators Daschle and Leahy.


Cutaneous anthrax showing the eschar

Bacillus cereus • common airborne & dustborne • grows in foods, spores survive cooking &

reheating • ingestion of toxin-containing food causes

nausea, vomiting, abdominal cramps & diarrhea; 24 hour duration

• no treatment • spores abundant in the environment



• gram-positive, spore-forming rods • anaerobic & catalase negative • over 120 species • oval or spherical spores produced only under

anaerobic conditions • synthesize organic acids & alcohols (useful for some

biotechnology application) & exotoxins • cause wound & tissue infections & food intoxications


Clostridium perfringens

• causes gas gangrene in damaged or dead tissues

• 2nd most common cause of food poisoning, worldwide

• virulence factors – toxins – alpha toxin – causes RBC rupture, edema & tissue

destruction – collagenase – hyaluronidase – DNase


12 C. perfringens


C. perfringens wound infection

Clostridium perfringens • treatment of gangrene – debridement of diseased tissue large doses of cephalosporin or penicillin hyperbaric oxygen


Hyperbaric treatment chamber

Exposure to increased oxygen levels inhibits anaerobes and speeds healing

Clostridium difficile

• normal resident of colon, in low numbers • causes antibiotic-associated colitis

– treatment with broad-spectrum antibiotics kills the other bacteria, allowing C. difficile to overgrow

• produces entertoxins that damage intestine

• major cause of diarrhea in hospitals



Mild Severe pseudomembranous colitis

C. difficile infection. Lining of the colon as seen via a sigmoidoscope

Clostridium tetani

• common resident of soil & GI tracts of animals • causes tetanus or lockjaw, a neuromuscular

disease • spores usually enter through accidental

puncture wounds, burns, umbilical stumps, frostbite, & crushed body parts

• tetanospasmin – neurotoxin causes paralysis • vaccine booster needed every 10 years



Tetanospasmin (a potent neurotoxin). Toxin inhibits release of neurotransmitter from neurons controlling inhibition of skeletal muscle contraction

Patient with Tetanus

Figure 19.17

20 Neonatal tetanus

Clostridium botulinum

Causes 3 diseases 1. food poisoning – (an intoxication) spores are in soil,

may contaminate vegetables; improper canning does not kill spores & they germinate in the can producing botulinum toxin – toxin causes paralysis by preventing release of


2. infant botulism – caused by ingested spores that germinate in the body & release toxin

3. wound botulism – spores enter wound & cause food poisoning symptoms (although GI tract is not colonized) 21



Clostridium botulinum Botox

• In 2007, almost 4.6 million procedures were performed with FDA-approved BOTOX® Cosmetic.

• BOTOX® Cosmetic works by temporarily reducing the contractions of the muscles that cause the persistent frown lines that have develop over time.

• The benefits may last up to 4 months

• Over 11 million people have used botox

Listeria monocytogenes • non-spore-forming gram-positive • primary reservoir is soil & water • can contaminate foods & grow during refrigeration • Most cases are from contaminated dairy products,

poultry and meat • Disease is often mild/subclinical …may have fever,

diarrhea • Listerosis in immunocompromised patients, fetuses &

neonates affects brain & meninges – 20% death rate

• ampicillin & trimethoprim-sulfamethoxazole • Prevention – pasteurization & cooking


Listeria monocytogenes

• Pathogen isolated from: – 12 % of ground beef – 25-30% of

chicken/turkey carcasses – 6% of luncheon meats – also in dairy products


Erysipelothrix rhusiopathiae

• gram-positive rod widely distributed in animals & the environment

• primary reservoir – tonsils of healthy pigs • enters through skin abrasion, multiplies to

produce erysipeloid, dark red lesions • penicillin or erythromycin • vaccine for pigs



Erysipeloid lesion on hand of animal handler

Corynbacterium diptheriae

• gram-positive irregular bacilli (pleiomorphic) • produce catalase • possess mycolic acids & a unique type of

peptidoglycan • 2 stages of disease

– local infection –upper respiratory tract – diptherotoxin production & toxemia

• pseudomembrane formation can cause asphyxiation



United States



Diptheria What can happen if you don’t vaccinate!

• 1990s epidemic in former Soviet Union countries. In 1993 and 1994, more than 50,000 cases were reported during a serious outbreak of diphtheria in countries of the former Soviet Union. (Contrast US had only 15 total cases between 1994-2009)

• Reduced vaccination programs resulted in greatly reduced “herd immunity”

• Multitude of susceptible persons allowed spread in the community.

• Most cases are in children aged 1-10 in crowded settings.


Propionibacterium acnes • gram-positive rods • aerotolerant or anaerobic • nontoxigenic • common resident of

sebaceous glands • Causes 85% of acne cases


Mycobacteria • gram-positive irregular bacilli • acid-fast staining • strict aerobes • produce catalase • possess mycolic acids & a unique type of

peptidoglycan • do not form capsules, flagella or spores • grow slowly

Mycobacterium tuberculosis Mycobacterium leprae


Mycobacterium tuberculosis • produces no exotoxins or enzymes that

contribute to infectiousness • contain complex waxes & other substances that

prevent destruction by lysosomes of macrophages

• transmitted by airborne respiratory droplets • Estimated that 1/3 of world’s population is

infected • only 5-10% infected people develop clinical

disease. (still…about 2 million people die every year) 34

Primary TB

• infectious dose 10 cells • phagocytosed by alveolar

macrophages & multiply intracellularly

• after 3-4 weeks immune system attacks, forming tubercles, granulomas consisting of a central core containing bacilli surrounded by WBCs


Primary Tuberculosis

Figure 19.22a

Secondary TB • reactivation of bacilli • tubercles expand & drain into the bronchial tubes

& upper respiratory tract • gradually patient experiences more severe

symptoms – violent coughing, greenish or bloody sputum, fever,

anorexia, weight loss, fatigue • untreated 60% mortality rate

Extrapulmonary (disseminated) TB • during secondary TB, bacilli disseminate to

regional lymph nodes, kidneys, long bones, genital tract, brain, meninges = Bad News


Secondary Tuberculosis

Figure 19.22b


1. tuberculin testing (Mantoux test)

2. X rays 3. direct identification of

acid-fast bacilli in specimen

4. cultural isolation and biochemical testing



Tuberculin skin test. Tests for exposure. Tests for hypersensitivity(type IV)

Vaccination causes a positive test result…so test is only useful in countries that do employ widespread vaccination

Treatment of TB

• Current US treatment: combination of 4 antibiotics for 2 months.

• Multi-drug resistant TB treatment: much more expensive and can go on for 2 years.

• vaccine based on attenuated bacilli Calmet- Guerin (BCG) strain of M. bovis used in other countries (not U.S.)

• Success rate of vaccination ??? ~80% in children and 20-50% in adults.


Directly Observed Therapy (DOT) …to assure that medication is actually taken


Mycobacterium leprae

• Hansen’s bacillus • strict parasite – has not been grown on artificial

media or tissue culture • slowest growing of all species • multiplies within host cells in large packets called

globi • causes leprosy, a chronic disease that begins in the

skin & mucous membranes & progresses into nerves



• endemic regions throughout the world • spread through direct inoculation from leprotics • 2 forms

– tuberculoid – superficial infection without skin disfigurement which damages nerves and causes loss of pain perception

– lepromatous – a deeply nodular infection that causes severe disfigurement of the face & extremities

• treatment by long-term combined therapy


46 Leprosy




Feather test for leprosy.

Tickling to determine sensitivity to touch


• Genera Actinomyces & Nocardia are nonmotile filamentous bacteria related to mycobacteria

• may cause chronic infection of skin and soft tissues • Actinomyces sp – responsible for diseases of the oral

cavity & intestines • Nocardia brasiliensis causes pulmonary disease

similar to TB



Periodontal infection. Infection works its way to the surface of the face to rupture and drain




Pulmonary infection that has disseminated to the skin


• Ferment carbohydrates to lactic acid. The acidity inhibits many competing bacteria and creates an ecological niche where they have the advantage.

• Major inhabitant of the vaginal tract. Not a pathogen.

• Used in production of many foods

L. delbrueckii used in yogurt production – L. acidophilus used in acidophilus milk production

Gram Positive Cocci

Oral streptococci on agar plate

General characteristics of the Staphylococci

• Spherical cells arranged in irregular clusters • Gram positive • Common inhabitant of the skin & mucous

membranes • Lack spores and flagella • May have capsules • numerous species


Staphylococcus aureus

• grows in large, round, opaque colonies • optimum temperature of 37oC • facultative anaerobe • withstands high salt, extremes in pH, & high

temperatures • produces many virulence factors


S. aureus


SEM showing “grape-like” clusters

Enzymes of S. aureus coagulase – coagulates plasma and blood;

produced by 97% of human isolates; diagnostic

• hyaluronidase • staphylokinase • DNase • lipases • penicillinase


S. aureus


Catalase Test

Coagulase Test

Toxins of S. aureus

• hemolysins – lyse RBCs; α, β, γ • leukocidin • enterotoxins • exfoliative toxin • toxic shock syndrome toxin


S. aureus

• Present in most environments frequented by humans

• Readily isolated from fomites • Carriage rate for healthy adults is 20-60% • Carriage is mostly in anterior nares, skin,

nasopharynx, intestine


S. aureus diseases

• Ranges from localized to systemic • localized -abscess, folliculitis, furuncle,

carbuncle, impetigo • systemic – osteomyelitis, bacteremia • toxigenic disease – food intoxication, scalded

skin syndrome, toxic shock syndrome


S. aureus


Skin Lesions Boil or Furuncle


S. aureus



S. aureus


Staphylococcal scalded skin syndrome (SSSS)

Epidermal shedding/separation

Exfoliative toxin

Staphylococcus aureus Food Poisoning  Food contaminated by handling with bare

hands. Bacteria grow and produce toxin.  Refrigerate food to prevent growth. Heating

doesn’t always destroy toxin.  Symptoms usually start within 1-6 hours after

eating contaminated food (usually gone in a day)

 Symptoms include: diarrhea, vomiting,

nausea, cramps

Staphylococcus aureus

Toxic Shock Syndrome (TSS) • First linked to use of ultra-absorbent

tampons that bound Mg ions and created a favorable environment for increased colonization and growth of vaginal S. aureus and increased TSS toxin production.

• Toxin enters bloodstream and causes fever, vomiting organ damage and potential fatality.

• Height of problem 1980-1984…15,000 affected/yr with 15% mortality.


SuperBug ?

• MRSA: Methicillin Resistant Staphylococcus aureus – A strain of staph that is highly resistant to broad-spectrum

antibiotics. – Once found almost exclusively in hospital settings the

bacteria is now showing up in the wider community – People can be colonized by the bacteria (skin and nose

primarily) and be healthy but act as carriers to spread the microbe.

– It is thought that perhaps 25-30% of people may harbor S.aureus.

MRSA has become a subject for some intensive public education efforts


• A CDC survey completed recently suggests that in 2005 there were ~ 94,000 cases of MRSA in the US and that perhaps 19,000 people died. (A higher rate than AIDS in the US)

• 85% of the cases were in hospitals, nursing homes etc. But many are concerned about the large number of infections acquired in normal community environments.

• Spread may be by hands (direct contact) or from contaminated surfaces.

• Infections range from an inconsequential skin lesion to a fatal septicemia or pneumonia

Other Staphylococci

S. epidermidis – lives on skin & mucous membranes; endocarditis, bacteremia, UTI

S. hominis – lives around apocrine sweat glands

S. capitis – live on scalp, face, external ear • All 3 may cause wound infections • S. saprophyticus – infrequently lives on skin,

intestine, vagina; UTI



• Gram-positive spherical/ovoid cocci arranged in long chains

• Non-spore-forming, nonmotile • Can form capsules & slime layers • Facultative anaerobes • Do not form catalase !!! • Most parasitic forms are fastidious & require enriched

media • Small, nonpigmented colonies • numerous species





• Lancefield classification system based on cell wall Ag – 14 groups (A,B,C,….)

• Another classification system is based on hemolysis reactions

• β-hemolysis – A,B,C,G & some D strains • α –hemolysis – S. pneumoniae & others

collectively called viridans



Human streptococcal pathogens

S. pyogenes S. agalactiae • viridans streptococci • S. pneumoniae Enterococcus faecalis


β-hemolytic S. pyogenes Group A • Most serious streptococcal

pathogen • Strict parasite…Inhabits

throat, nasopharynx, occasionally skin

• Produces multiple virulence factors


S. pyogenes • Humans only reservoir • Transmission – contact, droplets, food, fomites • Skin infections –pyoderma, impetigo, erysipelas • Systemic infections – strep throat, pharyngitis,

scarlet fever • Sequelae -rheumatic fever, glomerulonephritis



Pharyngitis and Tonsilitis Strep Throat

Note white pus patches on tonsils

Toxic sequelae to Strep throat

82 /tessman.strep.throat.kare


Impetigo lesions


Streptococcal skin infections

FLESH-EATING Bacteria Necrotizing fasciitis

84 Streptococcus pyogenes


Rheumatic fever is a possible sequelae to S. pyogenes infection.

Antibodies against the bacteria may cross-react with host tissue leading to damage of heart valves

Group B: S. agalactiae

• Regularly resides in human vagina, pharynx & large intestine

• can be transferred to infant during delivery & cause severe infection – Most prevalent cause of neonatal pneumonia, sepsis, &

meningitis – 15,000 infections & 5,000 deaths in US – Pregnant women often screened & treated

• wound and skin infections & endocarditis in debilitated people


Enterococcus faecalis, & E. faecium • Normal colonists of human large intestine • Cause opportunistic urinary, wound, and skin

infections, particularly in debilitated persons • Enterococcus …Genus related to streptococci

and so studied together


Viridans group • α-hemolytic • Large complex group • Most numerous &

widespread residents of the oral cavity & also found in nasopharynx, genital tract, skin

• Not very invasive but…dental or surgical procedures facilitate entrance


Viridans group

• Bacteremia, meningitis, abdominal infection, tooth abscesses

• Most serious infection – subacute endocarditis – blood-borne bacteria settle & grow on heart lining or valves

• Persons with preexisting heart disease are at high risk & may receive prophylactic antibiotics before surgery or dental procedures



Subacute bacterial endocarditis.

Colonization of heart valve surfaces lead to layers of fibrin and bacteria called vegetations

Viridans group • S. mutans produces slime layers that adhere

to teeth, basis for plaque • involved in dental caries


SEM of Plaque

S. pneumoniae

• Causes 60-70% of all bacterial pneumonias • arranged in pairs and short chains • All pathogenic strains form large capsules –

major virulence factor • Causes pneumonia & otitis media • Vaccine available for high risk people


93 Gram stain of sputum from pneumonia patient showing diplococci

S. pneumoniae

• 5-50% of all people carry it as normal flora in pharynx

• Very delicate, does not survive long outside of its habitat

• Pneumonia occurs when cells are aspirated into the lungs of susceptible individuals

• Pneumococci multiply & induce an overwhelming inflammatory response

• Treated with penicillin 94

S. pneumoniae


Middle ear infections (otitis media)


S. pneumoniae


Normal eardrum Buldging eardrum (otitis media)


Tubes are usually left in for 8 to 18 months and most often fall out on their own

Healthy eardrum Ruptured/torn eardrum


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