Pseudomonas & and other Non-fermenters
Are they pathogens or are they not?
• a complex mixture of opportunistic
pathogens widely in nature on organic substrates and colonizing plants,
animals and human.
• are ubiquitous organisms found in soil,
decaying organic matter, vegetation, and
water. Unfortunately, they are also found
throughout the hospital environment.
The group characterization
• small, gram-negative rods typically arranged in pairs, simple nutritional needs and quickly aquired
resistance to antibiotics they are common in hospital environment causing colonisation and, hence,
infections of hospitalized patient.
• obligate aerobe; glucose oxidizer (enterobacteria –
glucose fermenters); simple nutritional needs, ussually
oxidase positive (enterobacteria – negative)
Virulence (factors)
• they are not very well characterized
• virulence factors – structural
components (e.g. capsule – suppresses neutrophil and lymphocyte activity, pili –
adhesin, LPS – endotoxin activity, ), toxin
& enzymes (e.g. exotoxins – tissue
destruction, hemolysins), ATB and
desinfectant resistance
Colonization and/or infection
• transiently colonize the respiratory and gastrointestinal tracts of hospitalized
patients, particularly those treated with broad-spectrum antibiotics
• most of the infections in humans are caused Pseudomonas aeruginosa,
Burkholderia cepacia, Stenotrophomonas
maltophilia, Acinetobacter baumannii
Treatment, prevention & control
• combined use of effective antibiotics (e.g., aminoglycoside and β-lactam
antibiotics); monotherapy is generally ineffective
preventing contamination of sterile
medical equipment and nosocomial
transmission; unnecessary use of broad- spectrum antibiotics can select for
resistant organisms
Species and infection
• Pseudomonas aeruginosa
• pulmonary infections
• bacteremia
• primary skin infections
• urinary tract infections
• ear and eye infections
• many virulence factors Structural components
* adhesins – pili, nonpilus (binding to epith.cells)
* polysacharide capsule – alginate (anchor
bacteria to epit.cells, protect from phagocytosis and complement and antibiotics , biofilm in vivo)
* endotoxin (next slides)
* pyocyanin – blue pigment, toxic forms of oxygen (superoxide, hydrogen peroxide)– tissue
damage
Virulence factors of P. aeruginosa
Pseudomonas aeruginosa - the most significat pathogen are well known comparing to other non-fermenter species.
STRUCTURAL COMPONENTS: Pilli (fimbriae) and nonpilus adhesins bind to epithelial cells, similar in structure to the pilli found in N.
gonorrhoeae; capsule contain mucoid polysaccharide, adhesin,
inhibits antibiotic killing (e.g. aminoglycoside); suppresses neutrophil and lymphocyte activity; lipopolysaccharide – endotoxic activity;
pyocyanin – inhibit ciliary function; increases release of cytokines to mediate inflammation; mediates tissue damage through production of toxic oxygen radicals (e.g. hydrogen peroxide).
TOXINS & ENZYMES: exotoxin A - inhibits protein synthesis, produces tissue damage (e.g. skin, cornea); immunosuppressive; exotoxin S - inhibits protein synthesis; immunosuppression, cytotoxin (leukocidin) - cytotoxic for eukaryotic membranes (e.g., disrupts leukocyte
function, pulmonary microvascular injury), elastase - destruction of elastin-containing tissues (e.g., blood vessels, lung tissue, skin), collagen, immunoglobulins, and complement factors, phospholipase C - hemolysin; mediates tissue damage; stimulates inflammatory response, alkaline phosphatase - destruction of tissue
Virulence factors
Any tissue or organ may be affected with P. aeruginosa infections Localized infections: the infection follow trauma including also
medical instrumentation or stay in a risk area (e.g. swimming pools).
These infections may occur in the eye (e.g. keratitis, endophtalmitis), ear (e.g. necrotizing otitis externa), skin (wound sepsis), urinary and respiratory tract infections usually in hospitalized patients after
medical instrumentation (e.g. catheters), gastrointestinal infections – from mild diarrhea in children to necrotizing enterocolitis in
neutropenic cancer patients, central nervous infections – meningitis and brain abscesses usually associated with trauma or tumors. The localized infection coud lead to disseminated infection. Respiratory tract infections in patients with cystic fibrosis(CF) - P. aeruginosa is pathogen of chronic infection of patient with CF (as single infection or coinfection with Burkholderia cepacia complex) and chronic lung
diseases. Because the strains adapt to the changing environment during the infection usualy produce large mucoid polysaccharide (alginate) and biofilm in host or mucoid colonies on culture media.
Even susceptible strains to antibiotics which produce biofilm are usualy not susceptible to many of them.
Localized infections
Systemic infections
as bacteremia, endocarditis, bone and joint infections are sequale of spread of localized infection.
Infection caused by other non-fermenters e.g. Acinetobacter
baumannii, Stenotrophomonas maltophilia are significant agents of hospitally aquired infections.
ANTIBIOTIC RESISTANCE. P. aeruginosa is inherently resistant to many antibiotics and can mutate to even more resistant strains during therapy. P. aeruginosa produces a number of different β- lactamases that can inactivate many β-lactam antibiotics (e.g., penicillins, cephalosporins, carbapenems). Although numerous resistance mechanisms have been identified, the mutation of porin proteins constitutes the major mechanism of resistance.
Penetration of antibiotics into the pseudomonad cell is primarily through pores in the outer membrane. If the proteins forming the walls of these pores are altered to restrict flow into the cell,
resistance to many classes of antibiotics can develop simultaneously.
a) Specimens: pus, sputum, urine, blood, and other clinical material depending upon localization of the
process.
b) Microscopy: gram-negative rods usually in pairs.
c) Culture: because their simple nutritional need they grow well on basic culture media (nutrient agar), but because detection of some properties (e.g. hemolysis) enriched media (blood agar) are also used (fig.1). In case competetive flora appear in the samples a selective media are used.
Laboratory diagnosis
P. aeruginosa culture (nutrient agar) produced blue pigment (pyocyanin)
are focused on detection of specific biological properties of the agent (microscopy, bacterial colonies, enzymes detection, mass spectrum
Screening tests (preliminary identification): detection of some phenotypic properties are demonstrated in fig.
Identification
A B C
D
Fig. Pseudomonads and other (glucose) non-fermenters usually produce enzyme oxidase (A – blue color indicate production of the enzyme)
comparing to enterobacteriae (B, oxidase negative reaction) which are also gram-negative bacteria and produce similar bacterial colonies on culture media. Pseudomonads and other non-fermenters never ferment glucose (C), comparing with glucose feremntation in enterobacteria. The property is tested in semisolid medium containing glucose overlaid with mineral oil.
Maldi-TOF identification
Endotoxin is a lipopolysaccharide integrated in the outer membrane of Gram-negative bacteria
http://pathmicro.med.sc.edu/fox/lps.jpg
-Lipid A embedded in outer membrane, core and O antigen portions extending from the bacterial surface
Endotoxin properties
- Lipid A is toxic portion of endotoxin (activate complement and cytokines cascade)
- Exerts its effect only when bacteria lyse (result of complement atack, ingeston and killing by phagocytes or certain antibiotics)
- Lipid A become toxic if its in too high concentration
- Consequences – local inflamation (activation of complement) and systemic response (septic
shock)
• shock - clinical term, a set of events that lead to collapse of the circulatory system and can result in multiorgan system failure
• etiology - variety of insults but bacterial infection is the most common cause
• 1. step - production of cytokines by
monocytes, macrophages and other cells
• 2. step - activation of complement cascade
• 3. step – activation of coagulation cascade
Endotoxin and septic shock
Diagram of septic shock
• exotoxin A, S (blocking elongation factor inhibit proteosynthesis)
• Elastase (damage elastin in tissue - lung parenchymal damage, hemorragic lesions proteases – figures below, phospholipase C – tissue damase mediate and disrupting of inflamatory response
Toxin and enzymes
ecthyma granulosum
