Gram-positive bacilli and rods

Katedra i Zaklad Mikrobiologii

UM Wrocław

lysogenic bacteria anatoxine -toxoid active immunization passive immunization zoonosis

Important terms

Important terms - lysogenic bacteria

Lysogenic bacteria - bacteria that is infected with a temporate bacteriophage and bacteriophage’s DNA is integrated into the bacteria's chromosome (viral genes are present in the bacterium without causing disruption of the bacterial cell)

TOXIN

Toxicity Antigenicity

TOXOID

Toxicity Antigenicity

Convertion : - heat - chemicals

Stimule active immunity

(toxoid vacines)

Important terms - anatoxine

anatoxine = toxoid - bacterial toxin whose toxicity has been weakened or suppressed; can be used for safe active immunization

Bacterial proteins or polysaccharides (Subunit/conjugate)

toxoid

non-virulent or attenuated pathogen

inactivated/killed pathogen active immunization - stimulation with a specific antigen to promote antibody formation in the body. Protection against disease may last several years, in some cases for life

Important terms – active immunization

Important terms - passive immunization

natural acquired

Passive immunity - providing IgG antibodies to protect against infections (immediate, but short-lived protection) :

a) natural passive immunity - maternal antibody (mainly IgG) protect the newborn baby b) acquired passive immunity - process of obtaining serum from immune individuals, and then injecting it to protect a susceptible person.

zoonosis is any disease or infection that is naturally transmissible from vertebrate animals to humans as a result of direct or indirect contactwith animal population

Important terms - zoonosis

Gram-positive Rods

Corynebacteria Listeria Erysipelothrix do not produce spores

Bacillus & Clostridium (spore forming bacteria)

Not closely related

No cause similar clinical conditions

Family: Bacillaceae

Genus: Bacillus

Genus: Clostridium

- product endoepores - facultative or obligate aerobes - many produce antibiotics

- Product endoepores - strict anaerobes - diversity of anaerobic fermentation - Wide variety of extracellular enzymes (biodegradation & invasive infection)

- B. anthracis - B. cereus - B. cereus

- Cl. tetani - Cl. botulinum - Cl. perfringens - Cl. difficile

Bacillus anthracis

• Protein capsule (poly-D-glutamic acid)

• Form spores

• Three toxin protein components (plasmid-coded )

Bioterrorism agent !!!

individulal proteins important in pathogenesis of antrax

• Protective antigen (PA) mediates cell entry of EF &LF

• Edema factor (EF) severe edema (IL-4;disable immune response )

• Lethal factor (LF) tissue necrosis

(TNF-α, IL1-β; kill the cell)

Lethal toxin Subunit A = rapid cell deth by cleaves critical intracellular signaling module

Edema toxin Subunit A = catalyzes production of cyclic AMP within host cell -> dramatic tissue edema

Protective antigen Subunit B = binds toxin to host cells and mediates it internalization)

Bacillus anthracis A-B toxines 2 A-B toxines, which share the same B component

Anthrax

• zoonosis disease

• Is not spread from person to person

How is anthrax spread?

Getting spores in a cut or broken skin

Breathing in spores

Eating contaminated meat

CUTANEOUS ANTHRAX

INHALATION ANTHRAX

INTESTINAL ANTHRAX

Cutaneous Anthrax Symptoms

• Develope 1-12 days

• Begins like an insect bite

• Becomes a fluid- filled blister

• Center turns black and becomes sunken

Inhalation Anthrax Symptoms

• Incubation time: 1-5 days

• Flu-like symptoms

• Fever/ chills

• Muscle aches

• Malaise

• Shortness of breath

Flu symptoms vs Anthrax symptoms

SYMPTOM

FLU

ANTHRAX

Sore throat Common Occasional

Runny nose Y N

Fever and chills Y Y

Cough, productive Y N

Cough, dry, unproductive

Y

Y

Schortness of breath

Rare

Common

Chest pain rare common

Intestinal Anthrax Symptoms

• Nusea, loss of appetite

• Vomiting

• Fever

• diarrhea

Anthrax Therapy:

– penicillin, tetracyclines, quinolones

– administered before the onset of lymphatic spread!

Prevention:

– possible vaccination for „risk” groups

– vaccination of animals (Pasteur 1881)

– burning of burial of animals that died of anthrax

Bacillus cereus infections

GASTROENTERITIS

Emetic form Diarrheal form

Heat-stable Proteolysis resistant enterotoxin (Rice)

Heat-labile enterotoxine (meat, vegetables)

Occular infections (necrotic toxine, cereolysine, phospholipase C)

Intravenus Catheter related sepsis

intoxication infection caused by toxin producing in vivo

Clostridium botulinum

the bacillus of

botulism

• Found worldwide in soil and aquatic sediments

• Botulus (Latin) = sausage

• 15 min, 100°C – destroying of C.botulinum and its toxin

• 121°C, 15 min (moist heat) – spores destruction

CLASSIC BOTULISM

Vegetables, fish contaminated

with spores

Improper process of

food

Spores germination

Bacill multiplying

Excrete toxin into the food

Food is ingested

Toxin absorbtion

Toxine signs appear within 24h – 48h

Appropriate conditions: - Strictly anaerobic

environment - Neutral or alkaline pH

Botulism

failure of neurotransmission flaccid paralysis

Botulinum toxin acts at the neuromuscular junction to prevent release

of acetylcholine, a neurotransmitter that stimulates muscle contraction

Manifestations

of

food-borne botulism

• Very slight gastrointestinal symptoms

• No fever, no sign of sepsis

• Normal mental status

• Descending flaccid paralysis with bulbar signs

„4 Ds” –signs of botulinism

1. Diplopia (double vision)

2. Dysarthria (difficulty forming words)

3. Dysphonia (difficulty intoning words)

4. Dysphagia (difficulty swallowing)

WOUND BOTULISM

• Rare form of botulism

• Wound becomes contamined with the organism, and toxin is absorbed from that site

INFANT BOTULISM „ floppy baby syndrome”

Early signs:

- Constipation

- Feeding problems

- lethargy

- Poor muscle tone

Not only C.botulinum….. C. baratti, C.butyricum

Botulism

CLASSIC BOTULISM (intoxication)

INFANT BOTULISM (infection caused by toxin producing in vivo)

WOUND BOTULISM -toxin producing in vivo in the wound; - symptoms similar to those of foodborne infection - incubation period is longer_

Treatment

• Antitoxin – trivalent horse antiserum (A, B, E)

• Antibiotics ( infant and wound botulism)

• Supportive measures (mechanical ventilation)

Botulinum toxin

and bioterrorism

Lethal dose of toxin for a 70-kg human:

- 0,09 to 0,15 µg intravenously

- 0,70 to 0, 90 µg inhaled

- 70 µg orally

By contaminating food (unlikely)

By inhalation (presence in aerosols)

Medical uses of botulinal toxin

• Dystonia

• Strabismus

• Hyperhydrosis

Other: migraine headaches, achalasia, cerebral palsy

Inhalational botulism does not occur naturally but would result from breathing in toxin used as a biological weapon

Iatrogenic botulism - occur due to accidental injection into the blood stream of commercial botulinum toxin

Clostridium tetani

• Normal inhabitants of the herbivores

• Produces a round terminal spore („drumstik”)

• Do not invade but remain at the side of infection

Clostridium tetani

• Tetanolysin – hemolytic toxin

• Tetanospasmin – neurotropic A-B toxin

A – responsible for toxic effect

B – binding subunit

Tetanospasmin

- Heat-labile, plasmid encoded

– Produced during stationary phase of growth

– Released when the cell is lysed

– It is destroyed by:

- heating (65°C, 5min)

- action of acid

- proteolytic enzymes

Tetanospasmin

- Being internalized & moves from the peripheral nerve

terminals to the central nervous system by retrograde

axonal transport

– Being released from the postsynaptic dendrites

– Localized within vesicles in the presynaptic nerve terminals

– Acts by blocking the release of inhibitory neurotransmitters (GABA)

– Causing exitatory synaptic activity to be unregulated

Tetanus

Predisposing factor for tetanus:

- area of low oxidation-reduction potential

(necrosis, large splinter)

- coinfection with aerobic organisms

- others:unskilled abortion, scarification rituals,

female circumcision

Tetanus

- generalized - localized - cephalic - neonatal

Tetanus

Incubation period: 4 days- several weeks

• Tingling sensation in the vicinity of the wound

• Muscular spasms

• „ lockjaw”

• Risus sardonicus -„sardonic grin”

• opisthotonus

Treatment

• Surgical management

• Tetanus toxoid – active immunisation

• Passive immunisation with antitoxin

• Critical care unit

•Laryngospasms •Fractures •Hypertension •Nosocomial infections •Pulmonary embolism •Aspiration pneumonia •Death

Complications of Tetanus

Clostridium perfringens

• Part of normal flora (vagina, GI)

• Ubiquitous in nature

• Cl. perfringens strains are grouped A through E type

Pathogenesis

Exogenic infection: penetrating wound, contaminated with spores

Endogenic infection: after gut rupture, septic abortion (rare)

Clostridium perfringens

SOFT TISSUE INFECTIONS

GASTROENTERITIS

CELLULITIS

FASCIITIS

MYONECROSIS FOOD POISONING

NECROTIZING ENTERITIS

SEPSIS

Remember, sometimes just colonization

After: - gut rupture - septic abortion

SOFT TISSUE INFECTIONS

CELLULITIS FASCIITIS = suppurative myositis results from cellulitis Accumulation of pus in the muscle planes no necrosis no sistemic symptoms

MYONECROSIS = gas gangrene systemic spraed necrosiss of muscle painful gas formation

edema erythema gas formation in tissue generally nonpeinful

LOCALIZED ! SYSTEMIC ! -> shock, renal failure

Clostridial cellulitis

• No systemic toxemia

• Infected tissue looks similar to gas gangrene due to gas bubbles fermentation

Myonecrosis (gas gangrene)

Is marked by:

high fever

intense pain

brownish pus

gas bubbles

(CO2, H2S, CH4)

skin decoloration

foul odor

Clostridium perfringens soft tissue infections

- Typically polymicrobic: Cl. perfringens (10-48h) Cl. septicum (2-3 days) Cl. histolyticum (3-6 days) Cl. novyi (5-6 days) - Incubation period in gas gangrene: 1-7 days

Clostridium perfringens pathogenesis

EXOTOXINS: Known 12 exotoxins (α,β,γ,δ,ε etc.)

toxin α = lecithinase A) MASIVE LYSIS OF: : endothelial cells, erythrocytes Leukocytes,platelets B) INCREASED VASCULAR PERMEABILITY C) HEPATIC TOXITI, MYOCARDIAL DYSFUNCTION

ENTEROTOXIN:

- Heat-labile protein - leading tp loss of fluid and

proteins

Role:

Pathogenesis IN

FOOD POISONING

DEGRADATIVE ENZYMES

Proteases

Dnases

Hyaluronidase

collagenases

a) LIQUEFY TISSUE b) PROMOTE THE SPREAD OF INFECTION

Role:

Role:

attention !

Microscopic examination of necrotic tissue reveals: gram-positive

rods in the, which is the result of lysis by clostridial toxins

Cl. perfringens type : A B C D E

All produces α exotoxine

Produces ENTEROTOXIN (food poisoning)

Produces the bigest amount of α TOXINE

Produces Beta toxine responsible for: - intestinal stasis - loss of mucosa with formation of necrotic lesions

Clostridium perfringens human infections

SOFT TISSU INFECTIONS FOOD POISONING PRIMARY SEPTICEMIA

Cl. perfringens type A

NECROTIZING ENTERITIS

Cl. perfringens type C

C.Perfringens Food-Borne Disease

-Intoxication - abdominal cramps, watery diarrhea

- no vomiting, no fever -occurs 8-18h after eating contaminated food (meet products)

- lasts 24-48h

Cl.perfringens Necrotizing enteritis (Pig-bel disease)

- β toxin produced by Cl. perfringens type C

- Toxine can be inactivated by trypsin BUT: Groups of risk: Papua New Guinea inhabitants (sweet patatoes) population with malnutration - presentation: acute abdominal pain, ulceration of small intestine, bloody diarrhea perforation of the intestinal wall septic shock

Treatment – Surgical debriment

– Very high dose penicillin

– Supportive therapy (HBOT)

Prevention: Cleaning and watching wounds for signs of infection

Clostridium difficile

• AAD (Antibiotic- Associated Diarrheas)

• PMC (Pseudomembranous Colitis)

Most common antimicrobial drugs associated with C.difficile diarrhea

• Cephalosporins

• Ampicillin

• Clindamycin

Clostridium difficile

Endogenous source:

Broad spectrum antibiotics alters the normal enteric flora

Exogenous source:

The hospitalized patients are more susceptible to the exogenous aquisition of C. difficile – nosocomial infection

Cl. difficile pathogenesis

Toxin A

enterotoxin that causes fluid accumulation

stimulates an inflammantory response

weak cytotoxin

Toxin B

potent cytotoxin (distrupts protein synthesis)

Resistant gene casettes (ermB MLS)

Diagnosis AAD

Presence of diarrhea associated with antibiotic therapy in the preceding 4-6 weeks

&

recovery of Cl.difficile organism and/or toxin from the stool

PMC

demonstration of pseudomembranes by colanoscopy & recovery of Cl.difficile from stool

Treatment

• Oral administration of metronidazole or vancomycin

• Avoid: antidiarrheal agents( prevent colonic stasis)

• Volume resuscitation for dehydrated patients

Coryneforms = diphtheroids

Natural human flora of:

- skin - upper respiratory tract - urogenital tract - gastrointestinal tract

Can function as

opportunistic pathogens

strict pathogens:

lysogenic Corynebacterium diphtheriae Corynebacterium ulcerans (zoonosis)

- Pleomorphic rods - Form clumps that look like Chinese characters

Diphtheria - Corynebacterium diphtheriae

•incubation period is 2–5 days but occasionally longer •humans are the only reservoir •The disease has now reached endemic status (less developed countries with unvaccinated or partially vaccinated citizens ) •Diphtheria is a disease that must be notified immediately

DIPHTHERIA

CUTANEOUS diphtheria Upper RESPIRATORY tract infection

Mortality rate : genreal population: 5%–10% children younger than age 5 years and adults older than age 40 years: up to 20%

occurs through inhalation of saliva droplets that contain the bacteria

spread through direct contact with skin lesions of an infected host

Diphtheria

Exotoxin responsible for the disease symptoms

Diphtheria should be considered: - In unvaccinated patients with pharyngitis - low grade fever - cervical adenopathy - adherent gray exudate in throat

gene for toxin production occurs on the chromosome of the prophage, but a bacterial repressor protein controls the expression of this gene. The repressor is activated by iron. Toxin is synthesized only by lysogenic bacteria under conditions of iron deficiency.

EXOTOXINE of Corynebacterium diphtheriae

A B

A-B type of toxine

A -ctive fragment of toxine (Inhibits synthesis of human proteins)

B - ind toxine to the receptor & mediates delivery A to its target

Thic, grayish, adherent excudate = Pseudomembrane

Fragment A catalyzes the transfer of ADP-ribose from NAD to the eucaryotic Elongation Factor 2 which inhibits the function of the latter in protein synthesis. Ultimately, inactivation of all of the host cell EF-2 molecules causes death of the cell.

Diphtheria

LOCAL INFECTION

(throat, skin)

BUT SYSTEMIC EFFECTS

(heart, peripheral nerves)

Treatment of Diphtheria

- neutralization of toxin (horse serum antitoxin) - eradycation of bacteria ( penicillin, erytromycin) - immunization with toxoid

PREVENTION: immunization with toxoid DTaP triple vaccine (+tetanus toxoid & pertussis antigens)

Other coryneforms

Corynebacterium ulcerans can carry diphtheria gene-tox may produce diphtheria infection (transmission can also occur through food vehicles such as milk or other dairy products- zoonosis)

Corynebacterium urealyticum- important pathogen of urinary tract Corynebacterium amycolatum- opportunistic pathogen capable of causing endocarditis and sepsis

Corynebacterium jeikeium –opportunistic pathogen of immunocompromised patients

Erysipelothrix rhusiopathiae

Pleomorphic G+ rod distributed worldwide (animals, environment) uncomon human pathogen (occupational disease in people working with animals) erysipeloid (localized or septicemic) – zoonosis complications: endocarditis, abscess formation, osteomyelitis virulence factors: - hyaluronidase - neuraminidase - capsule like structure

Listeria monocytogenes

- widespread among animals and in an environment - growth at a broad temperature range (1- 45°C) - facultative intracellular bacteria - naturally resistant to cephalosporins

Listeriosis Although the number of cases of listeriosis is small, the high rate of death associated with this infection makes it a significant public health concern sporadic disease in healthy adults ( influenzae-like illnes with or without gastroenteritis), but is serious disease in patients from risk groups: - pregnant women - newborns - immunocompromised patients - eldery

Virulence factors: -internalins - listeriolysin O - intracellular survival - ActA protein

LISTERIOSIS

GIT self-limiting febrile gastroenteritis

syndrome

CNS - meningitis - meningoencephalitis - rhomboencephalitis

bacteremia and non-CNS focal infections focal bone infection

native and prosthetic joints infection osteomyelitis liver infection

splenic abscess others…

neonatal infections - early onset: granulomatosis infantisepticam - late-onset: - pneumonia - septicemia - meningitis

pregnant women: miscarriage, premature delivery

amnionitis,

Thank you for attention