ZoonosesZoonoses LegionellosisLegionellosis PseudomonasPseudomonas GroupGroup andandA Acinetobactercinetobacter

Dr.Dr. BerekBerek ZsuzsannaZsuzsanna 0808 NovemberNovember 20112011 GRAM NEGATIVE RODS

AEROB FACULTATIV ANAEROB ANAEROB Bordetella Haemophilus Bacteroides Brucella (Z) Pasteurella (Z) Prevotella Francisella (Z) Porphyromonas Family: Fusobacterium Pseudomonas Enterobacteriaceae Acinetobacter Vibrionaceae Legionella Cardiobacterium MICROAEROPHIL Eikenella Campylobacter Kingella Helicobacter Actinobacillus ZoonosesZoonoses:: Brucella,Brucella, Francisella,Francisella, PasteurellaPasteurella andand PlaguePlague

1.1. BrucellaBrucella

Sir David Bruce (1855-1931) www.lshtm.ac.uk/.../archives/images/bruce1.jpg Brucellae 2 Morphology: Gramnegative Coccobacilli Cultivation: Agar – nutrient rich (Serum, Glycerine) CO 21 Days Resistant! staff.vbi.vt.edu/pathport/pathinfo_images/Bru...

Description: Brucella spp. Colony Characteristics: - A. Fastidious, usually not visible at 24h. - B. Grows slowly on most standard laboratory media (e.g. sheep blood, chocolate and trypticase soy agars). Pinpoint, smooth, entire translucent, non-hemolytic at 48h Morbus Bang swine Brucellosis Maltese fever All is Brucellosis „Febris undulans” RES! (undulating fever) Swine through skin lesions or conjunctiva or GI tract direct contact or contaminated food

Brucellae sick animals milk) (meat, mucosa Invasion from through - Pathogenesis, Infection, Clinical findings B. melitensisB. abortusB. suis Goat Anthropozoonosis Cattle - - Brucella – source of infection

Medmicro Brucella – portals of entry y for Brucella species.

Medmicro Figure 28-1 Portals entr of Brucella – spreading

Medmicro Figure. Acute unilateral scrotal swelling in a 27-year-old man with brucellosis.

www.medscape.com/.../art-iim441224.fig.jpg Fig.13.36 Brucellosis. Arthritis of the left knee. This was accompanied by fever, malaise, generalized myalgia and depression. Fig. 13.37 Orchitis – B. abortus Brucellosis - diagnosis Brucellosis Diagnosis Culture: min. 5 days Serology Antibody detection Tube-agglutination (Wright) IgM Chromatography ELISA Brucella IgM Therapy: Doxycyclin, Rifampicin, Streptomycin www.kit.nl Prophylaxis: exposition Treatment or anihilation of sick animals WHO – Bioterror Category B!!! ZoonosesZoonoses:: Brucella,Brucella, Francisella,Francisella, PasteurellaPasteurella andand PlaguePlague

2.2. FrancisellaFrancisella tularensistularensis

Tulare Lake; California, USA or per os or via Ectoparasites direct contact sick animals inhalation From Francisella tularensis through or NO HUMAN TO TRANSMISSION Morphology: Gramnegative rods Resistant to COLD! Cultivation is prohibited! Only special –BioterrorcategoryA!!! Laboratories WHO Pathogenesis, Infection - - Serology Diagnosis: Prophylaxiy: Expositionsprophylaxis Therapy: Streptomycin, Doxycyclin, Ciprofloxacin – Granulomatous lesions!

Francisella tularensis Clinical findings: TULARAEMIA Lymphnodes, small Granulomas + ulceration + necrosis Primar complex cutano-, oculo-, tonsilloglandular, (visible!) thoracal, abdominal - (invisible!) forms Generalisation A reported case of exposure of a patient to a wild rabbit, which subsequently died, suggested that tularemia was the likely etiology

staff.vbi.vt.edu/.../Ftularensis staff.vbi.vt.edu/.../Ftularensis

Description: Cervical Lymphadenitis in a Patient With Pharyngeal Tularemia; Patient has marked swelling and fluctuant suppuration of several anterior cervical nodes. Infection was acquired by ingestion of contaminated food or water. Source: World Health Organization Description: These Francisella tularensis colonies show characteristic opalescence on cysteine heart agar with sheep blood (cultured at 37 C for 72 hours). Note: On cysteine heart agar, F tularensis colonies are characteristically opalescent and do not discolor the medium

Description: Chest Radiograph of a Patient With Pulmonary Tularemia

staff.vbi.vt.edu/.../Ftularensis ZoonosesZoonoses:: Brucella,Brucella, Francisella,Francisella, PasteurellaPasteurella andand PlaguePlague

3.3. PasteurellaPasteurella

Louis Pasteur (1822-1895)

upload.wikimedia.org/.../4/42/Louis_Pasteur.jpg Pasteurella multocida

Morphology: Gramnegative, small rods

Cultivation: Blood- and chocolate agar

Pathogenesis: Through dog and/or cat bites

Immunocompromised: Sepsis!

Therapy: surgery, Penicillines

medecinepharmacie.univ-fcomte.fr Pasteurella multocida

Fig. 10.55 Animal bite. Infected wound of finger following bite of domestic cat. P. multocida was isolated from the wound.

aapredbook.aappublications.org Pasteurella multocida cellulitis secondary to multiple cat bites about the face of a one- year-old child

aapredbook.aappublications.org ? ZoonosesZoonoses:: Brucella,Brucella, Francisella,Francisella, PasteurellaPasteurella andand PlaguePlague

4.4. PestPest –– YersiniaYersinia pestispestis

1863 - 1943 www.mja.com.au Genus: Enterobacteriaceae! Giemsa staining Gramnegative rods – bipolar staining

Yersinia pestis Morphology: www.lonlygunmen.de

Yersinia pestis NO CULTURE! FORBIDDEN! Only in special Laboratories WHO – Bioterrorcategory A!!!

www.idph.state.il.us, www2.cnrs.fr, ww.knowledgenews.net, Pest - XIV. Jh. spreading, fibrinolytic VIRULENCE FACTORS Capsule – Protein! V Antigen (Protein) Antiphagocytic W Antigen = Endotoxin Extracellular Substances - Plasminogen – Activator – Protein (Pla) -Toxin (kills mice)

Yersinia pestis Elimination of Rats!!! skin

Yersinia pestis Pathogenesis, Infection: Source of infection: Rats (and other rodents) → Transmission: direct contact, Rat-flea-bites Penetration: Figure 29-4 Pathogenesis of Y. pestis in plague patients. Medmicro

Yersinia pestis Yersinia pestis

Clinical findings: 1)Bubonic plague (swollen lymph nodes)

2) Septic form → haemorrhagic inflammation

3) Lung plague = Pneumonia ← direct aerogen transmission from human to human (drop- infection → primary lungplague)! Bubonic form

Fig. 13.55 Plague. Enlarged tender inguinal lymphnodes in a Vietnamese child with bubonic plague. Fig. 13.56 Advanced stage of inguinal lymphadenitis in bubonoc plague. The nodes have undergone suppuration and the lesion has drained spontaneously. By courtesy of Dr. J.R. Cantey Necrosis of finger tips of septicemic plague.

Septic form

Cutaneous Hemorrhages in Plague. Source www.cdc.gov

www.imcworldwide.org Lung plague www.imcworldwide.org Diagnosis Clinical picture Direct detection – (bipolar!) microscopic Serology – IF tube-agglutination, Therapy: Doxycyclin, Streptomycin Yersinia pestis Biological Weapons – Bioterrorism

Biological Weapon: Organism, Toxin, Virus Aim: • to kill and/or anihilate individuals and/or whole Population • economic damage

Biological war (military conflicts) Bioterrorisms (ideology!) Biological crime (personal) Biological Weapons – Bioterrorism

Categories: A, B, C Most dangerous: A B. anthracis, C. botulinum, F. tularensis, Y. pestis

Easy to culture Easy to spread/transmit – droplets, airways High letality Therapy? (too late) „Out of the blue” High number of cases ZoonosesZoonoses LegionellosisLegionellosis PseudomonasPseudomonas GroupGroup andand AAcinetobactercinetobacter

Dr.Dr. BerekBerek ZsuzsannaZsuzsanna 0808 NovemberNovember 20112011 LegionellosisLegionellosis LegionellaLegionella pneumophilapneumophila

Morphology: Gram negative rods

Aerob

www.lf3.cuni.cz Legionella Legionella

www.waterscan.co.yu Legionella pneumophila Flagellum, Fimbriae Legionella

mcb.berkeley.edu c harcoal) C; 3 days) BCYE ° xtract, Activ e ! east y ( BCYE (pH=9; Temperature 35 BCYE Cultivation: Special Media

Legionella pneumophila Bloodagar Legionella Culture

BCYE

www.lf3.cuni.cz Culture Legionella

www.contractlaboratory.com medecinepharmacie.univ-fcomte.fr

Legionella Source of infection, habitat: ubiquitous (air-conditions, water-showers, Humidity (floor), Biofilms) Legionella pneumophila Pathogenesis-1: aerogen - dropinfection! transmission: www.ratsteachmicro.com www.eco-khlamx.com www.lifespan.org

Legionella pneumophila Facultative intracellular! Pathogenesis-2: In water: Protozoon In humans: Leukocytes, Inflammatory reaction, Proteases, Phospholipases Legionella and amoeba

FIGURE 40-7 Electron micrograph showing L. pneumophila serogroup 1 in the process of dividing (arrows) within a vesicle of an amoeba (Hartmanella veriformis) cell. Pathogenic cycle of Legionella

microbewiki.kenyon.edu textbookofbacteriology.net/Legionella.jpeg

BAL: direct IF Legionair’s disease → avoiding of Legionella containing Aerosole Biopsy! 2) Pontiac-Fever (non-pneumonia) Diagnosis: Clinical findings: Legionellosis 1) Pneumonia detection – Erythromycin, Tetracyclin, Rifampicin, FQ Cultivation Direct antigen detection – Urine! Antibody detection – Serology; ELISA Prophylaxis: Therapy: Legionella pneumophila Fig. 2.29 Legionnaires’ disease. Chest radiograph showing extensive consolidation affecting parts of all lobes of the lungs. Diagnosis

Fig. 2.28 Legionella pneumophila. Specimen from bronchial biopsy taken through fibreoptic bronchoscope in a patient with fulminant Legionnaires’s disease. The organism can be isolated on selective culture media or by guinea pig inoculation. By courtesy of Dr. S. Fischer-Hoch Fig. 2.30 Legionnaires’ disease. Autospy specimen showing consolidation of upper and lower lobes of right lung. ZoonosesZoonoses LegionellosisLegionellosis PseudomonasPseudomonas GroupGroup andand AAcinetobactercinetobacter

Dr.Dr. BerekBerek ZsuzsannaZsuzsanna 0808 NovemberNovember 20112011 PseudomonasPseudomonas GroupGroup

Pseudomonas P. aeruginosa P

Burkholderia B. mallei P B. pseudomallei P B. cepacia

Stenotrophomonas S. maltophilia (Xanthomonas)

P = obligate pathogen Pseudomonas aeruginosa

Morphology and culture: Gramnegative rods 1-2 μm, no demands; Biofilmbuilding! Pseudomonas aeruginosa

Pigments: 1. Pyocyanin 2. Fluorescein

Haemolysis (β) Pseudomonas aeruginosa

VERY RESISTANT !

-to Heat Light Dry

Desinfectants

Antibiotics

www.fiu.edu Pseudomonas aeruginosa

ANTIGENS AND VIRULENCE FACTORS: Adhaesion and Colonisation - "0", "H", Pili/Fimbriae - Capsule = Glycocalyx - Alginate slime = mucoid Exopolysaccharid Î Biofilmbuilding Invasion, Penetration - Extracellular Proteases, Exoenzymes (many!) - Cytotoxin = Leukocidin and Haemolysins -Pigments Dissemination - Exotoxin A – inhibit Proteinsynthesis (EF2ÎDiphtheria) - Exotoxin/Exoenzym S – at burns; detecting in blood - Endotoxin

LD50 – at burnt wounds 30 Normal skin 108 Pseudomonas aeruginosa – Structure

Medmicro No single factor is decisive for virulence.

www.ratsteachmicro.com Pseudomonas aeruginosa Toxin-A – mode of action

Pseudomonas aeruginosa – elmi Pseudomonas aeruginosa

Pathogenesis, Infection:

Habitat (normally): Soil, water (swimming-pools, Pools!), sewege water Human GI tract Respiratory tract: animals

Source of infection: sick, carriers contaminated enviroment Solutions (humid Milieu!) Kunststoffe transmission: direct, indirect Contact Pseudomonas aeruginosa

Clinical findings: Nosokomial infections - Meningitis, Pneumonia (Respirator!) -Sepsis - Burns! Æ skin, Wounds - Urogenital Infections (Katheters), - GI tract (!), newborn/babies - Otitis media, externa - EYE + Contact lenses

Cystic Fibrosis (mucoid strains) Fig. 10.2 Pseudomonas folliculitis. Papulopustuler rash over the buttocks and thights following use of a spa pool. Fig. 13-6 Ecthyma gangrenosum. Necrotic round lesion on the buttock of a child with Pseudomonas septicaemia associated with immunodeficiency. Fig. 12.46 Bacterial keratitis. Contact lens-associated keratitis due to Pseudomonas aeruginosa. By courtesy of Dr. A.N. Carlson Diffuse infiltrate Thinned corneal stoma

Edge of epithelium

Fig. 12.47 Bacterial keratitis, in this case due to P. aeruginosa. An infiltrate is seen with corneal thinning. By courtesy of Mr. P.A. Hunter Fig. 12.48 Bacterial keratitis. A massive inflammatory response in anterior uveitis leads to precipitation of the cells as pus in the anterior chamber. This is called hypopyon. By courtesy of Mr. S. Harding Fig. 12.49 Bacterial keratitis. P. aeruginosa eye infection showing corneal ulceration and hypopyon formation in this rapidly progressive eye infection. Pseudomonas aeruginosa

Clinical findings: Nosokomial infections - Meningitis, Pneumonia (Respirator!) -Sepsis - Burns! Æ skin, Wounds - Urogenital Infections (Katheters), - GI tract (!), newborn/babies - Otitis media, externa - EYE + Contactlenses

Cystic Fibrosis (mucoid strains)

Pseudomonas aeruginosa Pseudomonas aeruginosa

Diagnosis: Detecting the bacterium, identification, Oxidase +

Therapy and Prophylaxis: Antibiogram!!! Aminoglycosides, Carbenicillin, antipseudomonas Cephalosporines, Fluoroquinolones

Expositionsprophylaxis: CLEAN! Disinfection! Active Immunisation – in cystic fibrosis Vaccine

A conjugate vaccine in the final clinical phase is Aerugen®, the first and only vaccine for the prophylaxis of fatal Pseudomonas aeruginosa infections in cystic fibrosis patients. The polyvalent conjugate vaccine combines 8 prevalent P. aeruginosa serotypes and the bacterial exotoxin A. It is the first conjugate vaccine based on a lipopolysaccharide component. www.bernabiotech.com Burkholderia mallei

Clinical finding: Malleus (glanders) – horse, donkey Humans: often fatal! Occupational disease Bioterror category B!

A horse with glanders and with positive mallein test

microbewiki.kenyon.edu/images/3/39/Horse.jpg www.vef.hr/vetarhiv/papers/68-5/alani2.jpg Burkholderia pseudomallei

Clinical finding: Melioidosis (pseudoglanders)

– Pneumonia, Sepsis subtropical and tropical area B. pseudomallei

www.co.collin.tx.us www.asm.org

B. pseudomallei

B. pseudomallei on Ashdown's agar after incubation at 37°C in air for 3 days Burkholderia cepacia, Stenotrophomonas maltophilia

Nosokomial infections

www.cdc.gov/.../web%20images/01-0535-8t.jpg

Figure 1. Electron micrographs showing expression of flagella by SMDP92. Stenotrophomonas maltophilia strains can have one (A) to several flagella (B,C). The flagella on these bacteria show a polar disposition. Bars, 0.5 µm Figure 8. Ultrastructural analysis of Stenotrophomonas maltophilia adhering to plastic. (A) Scanning electron micrographs showing the tight adhesion of SMDP92 to the plastic surface. (B) Structures resembling flagella seem to be protruding and interconnecting bacteria (arrowheads) or connecting bacteria to the plastic (arrows). (C) In addition to the flagellalike filaments (arrowheads), high-power magnification shows the presence of thin fibrillar structures connecting bacteria to the abiotic surface. Bars: A 10 mm, B 1 mm, C 2 mm.

www.cdc.gov/.../web%20images/01-0535-8t.jpg Stenotrophomonas maltophilia www.uni-ulm.de Burkholderia sp. www.oregon.gov B. cepacia www.uni-ulm.de AcinetobacterAcinetobacter spp.spp.

Morphology: Gramnegative dimorphic rods, coccobacilli

www.acinetobacter.org Acinetobacter spp. www.acinetobacter.org

Culture: Agar, Bloodagar important: Temperature 40-44°C Non-motile – „akinetic” Pathogenesis and clinical findings: like Pseudomonas aeruginosa

Nosokomial infections! Multiresistant! THE END Istanbul, 2006