Gram-negativeCocci Gram-negative...

Gram-negative Cocci Gram-negative bacilli

Miklos Fuzi

Neisseria

Pyogen ic Cocci GRAM -

Aerobic: Oxidase +Neisseria N. gonorrhoeae

N. meningitidisothers (N. sicca, N. subflava, N. flavescens and apathogenic spp.)

Moraxella M. catarrhalis

Anaerobic: Veillonella spp.

Veillonellae vietsciences.free.fr

N. gonorrhoeae and N. meningitidis

Morpholog yGram-negativeDiplococci

www.waterscan.co.yu/images

path.upmc.edu

Gram-negative Diplococci

N. gonorrhoeae and N. meningitidis

Oxidase +

path.upmc.edu

Culture:Special demandsNutrient rich medium(Chocolate agar and5-10% CO2)

Resistance:Sensitive bacteria:Dry, Heat, Disinfectants, Antibiotics

http://www.mfi.ku.dk/ppaulev/chapter33/images/33-3.jpg

Antigens and Virulence factors:Pili/Fimbriae (Antigenic variations!)IgA-Proteases!outer membrane proteins (OMP)

(Antigenic variations!)LOS (Mimicry!- sialiation in serum prevents

immune response)Cell-wall Peptidoglycan (Toxic)Inhibits apoptosis in macrophages (prevention

of immune response)

N. gonorrhoeae = Gonococcus

N. gonorrhoeae = Gonococcuste

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Pili

neisseria.org/images/ng-lym2.jpg

Gonococcus-Lymphocyte Interaction



Source of infectionSick humans

Transmission- Direct (sexual) Contact

Clinical findingsGonorrhea = TripperOphthalmoblenorrhea neonatorum

NO IMMUNITY!(Antigenic variations!)

Medmicro

Pathogenesis

Gonorrhea – acute Urethritis

www.stdservices.on.net ww

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Gonorrhea – acute Urethritis

www.boltonlgb.co.uk

Gonorrhea – acute Cervicitis

Gonorrhea – acute Cervicitis

Gonorrhea – acute ConjuctivitisBlenorrhea neonatorum

www.mc3.edu

www.slackbooks.com

Corneal ulcers due to gonococcus are very destructive and have a tendency to perforate the cornea.

Gonorrhea – Chronic and disseminated Form

Endometritis, Salpingitis, Prostatitispurulent Arthritis, Vasculitis

Important! anorectal GO and Pharyngitis(„alternative Genitals”)

Fig. 8.33 Gonococcal arthritis. Dactylitis secondary to gonococcal bacteriaemia. By courtesy of Dr. S.E. Thompson

Fig. 8.33 Gonococcal septic arthritis. Arthritis due to N. gonorrhoeae in a 24-year-old woman, showing marked erythema and swelling of the right ankle and leg. By courtesy of Dr. T.F. Sellers Jr.

Gonorrhea – Diagnosis – acute Disease

MicroscopicDirect detection – phagocytosed diplococciGram stainingMethylenblue staining, Direct Immunofluorescent (DIF)

www2.mf.uni-lj.si,www.uni-ulm.de,

pathmicro.med.sc.edu

GO – Gram staining – presumptive Diagnosis only!

GO – Gram staining – only presumptive Diagnosis!

www.med.uni-giessen.de

Gonorrhea – Diagnosis

Culture :„bedside” Thayer-Martin medium and chocolate agar, 5% CO2

Identification: ox+, glu+, mal-

Antigen detection (direct):Latex-agglutination

Detection of bact. DNA: PCR

www2.mf.uni-lj.si,www.uni-ulm.de,

pathmicro.med.sc.edu

Therapy:3. Generation Cephalosporin (Ceftriaxone) or Spectinomycin (Aminoglycoside)

Prophylaxis:GO- Exposition (safe sex)- Source of infection: find and treat! - Early Diagnosis and treatment

Ophthalmia neonatorum : Application of 1% silver-nitrate in conjunctival sack

NO VACCINE! (Antigenic variants!)

Gonorrhea

www.tiscali.co.uk

N. meningitidis = Meningococcus

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Antigens and Virulence factors:Capsule – Polysaccharide, antiphagocytic

Multiple serotypes (A, B, C, W135, Y!)Capsule change

Pili/FimbriaeIgA-Proteases!Outer Membrane Protein s (OMP)LOS (Mimicry, sialisation: Serum resistant,

very toxic)

zdsys.chgb.org.cn

Meningococcus

Source of infectionhuman – carriers (sick, healthy)

Transmission, Portal of entry- Direct, drop-infection- Nose, throat

Clinical findingPharyngitisMeningitis cerebrospinalis epidemicaSepsis = Waterhouse-Friderichsen Syndrome


Fig. 10.56 Acute meningococcaemia. Note the variabl e size of the lesions and their peripheral distribution. Some of the lesions are obviously purpuric, others macular or papular.

Fig. 10.60 Acute meningococcaemia. Petechia on bulb ar conjunctiva.

Fig. 10.62 Acute meningococcaemia. Gangrene of the extremities following a near-fatal illness with hypotension.

Fig. 10.63 Acute meningococcaemia. Gangrene of both legs in a black man with acute meningococcal infection. Bilateral below knee amputations were later required.

The characteristic skin rash of meningococcal septi caemia, caused by Neisseria meningitidis . (Courtesy of Wellcome Trust Photographic Library) srs.dl.ac.uk

Waterhouse- Friderichsen Syndrome: schwere nekrotisierende Hautläsionen bei Meningokokkensepsis mit Verbrauchskoagulopathie (R. E. Rieger, Univ.-Kinderklinik Marburg).

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www.gesundheit.de

The patient with Waterhouse-Friderichsen syndrome h as sepsis with DIC and marked purpura. medlib.med.utah.edu

Purulent meningitis with hemorrhage in the frontal lobe (gross findings).

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Acute hemorrhage in bilateral adrenals caused acute adrenal insufficiency (Waterhouse-Friderichsen syndrome).

pathy.fujita-hu.ac.jp

Meningitis Diagnosis

Samples, specimen:Liquor (cerebrospinal fluid) ! – Lumbar punctionBloodcarriers: throat

DetectionMicroscopic examination(Liquor, blood culture)CultureLiquor, Blood, ThroatDirect detection of antigen(Liquor) – Latex-agglutinationDirect detection of bact. DNAReal-time PCR from blood, CSF

Meningitis Diagnosis

Culture: Bloodagar, Chocolate agar

Identification:glu+, mal+

MIC (E-test)

DiagnosisN. meningitidis

Meningococcus meningitis

Therapy:Penicillin and/orCeftriaxone; cefotaximeNO Beta-lactamase production

Prophylaxis:Active ImmunisationVaccine for:- Risk groups- Traveler(Meningitis belt!)

Chemoprophylaxis:Ciprofloxacin; rifampicin(Contacts)

Meningitis belt

Neisseria meningitidis - B

Europe!

NO VACCINE!

Rifampicin only

www.versapharm.com

Haemophilus

GRAM-NEGATIVE COCCOBACILLI

Genus SpeciesHaemophilus H. influenzae

H. parainfluenzaeH. aegyptius H. ducreyi

Bordetella B. pertussis B. parapertussis

P: Pathogen

Haemophilus influenzaeHaemophilus influenzaeHaemophilus influenzaeHaemophilus influenzae

Morphology:Gram - Coccobacillus, ca. 1 µm

Cultivation:Growth factors !(chocolate, X= Haem, V= NAD, Satellite-phenomenon; Satellitism


phil.cdc.gov Blood agar plate culture showing Haemophilus influe nzae satelliting around Staphylococcus aureus.


Antigens and Virulence factors:Capsule – Polysaccharide

Types: a, b, c, d, e, f (HiB!)

IgA-Protease!Surface antigens:

Outer Membrane Proteine (OMP)LPS

Haemophilus influenzae Type b (Hib)

www.soundmedicine.iu.edu


Clinical findings:Meningitis , Sepsis

Cellulitis

Upper respiratory tract:Epiglottitis !, Nasopharyngitis, Sinusitis, Otitis media

Lower respiratory tract:Bronchitis, Pneumonia,

An infant with severe vasculitis with disseminated intravascular coagulation (DIC) with gangrene of the hand secondary to Haemophilus influenzae type b septicemia - prior to the availability of the Hib vaccine. -Image provided by: Visual Red Book on CD-ROM--(2000 Red Book: 25th Edition, Report of the Committee on Infectious Diseases)

www.ecbt.org


Sepsis

Periorbital cellulitis. © Neal Halsy, MD www.cispimmunize.org



Clinical findings:Meningitis , Sepsis

Cellulitis

Upper respiratory tract:Epiglottitis! , Nasopharyngitis, Sinusitis, Otitis media

Lower respiratory tract:Bronchitis, Pneumonia,

HiB-epiglottitis

Haemophilus influenzaeHaemophilus influenzaeHaemophilus influenzaeHaemophilus influenzaeDiagnosis:Samples, specimen

�LIQUOR! (CSF)� Site of infection (Nose, throat, Sputum etc.)

Detection:Microscopic, Culture, Capsule Ag detection (Latex-agglutination)Real-time PCR from blood, CSF

Therapy:1. Ampicillin + III. gen. Cephalosporins2. Ampicillin + Aminoglycosides

Prophylaxis:Active Immunisation - HiB Conjugate-Vaccine

(Polysaccharide + Protein)

Lipopolysaccharid Extract - Vaccine

ibs-isb.nrc-cnrc.gc.ca

www.kmhk.kmu.edu.tw

Haemophilus ducreyiHaemophilus ducreyiHaemophilus ducreyiHaemophilus ducreyi

Causing: Ulcus molle = Chancroid == soft Chancre

Haemophilus aegyptiusHaemophilus aegyptiusHaemophilus aegyptiusHaemophilus aegyptius

Causing: Brasilian Purpuric Fever

Haemophilus parainfluenzaeHaemophilus parainfluenzaeHaemophilus parainfluenzaeHaemophilus parainfluenzae

Pharyngitis, Endocarditis, Conjunctivitis

Ulcus molle

medinfo.ufl.edu

www.smu.eduChancroid in female

Bordetella

Bordetella pertussis

Morpholog y:Gram-negative Coccobacillus, ca. 1 µm



Culture:Special MediumBordet – Gengou

www.szu.cznobelprize.org

Antigens and Virulence factors:CapsuleFimbriae, filamentous HaemagglutininOuter Membrane Proteine (OMP)LPSPertactin

Extracellular Toxins:Pertussis ToxinAdenylate-cyclase ToxinTracheal cytotoxinDermatonecrotic Toxin


FIGURE 31-2 Virulence factors of B pertussis .

Medmicro

Pertussis toxin

www.med.sc.edu:85

Pathogenesis, Infection:Source: sick – in prodromal and catarrhal Stadium

Portal of entry: Respiratory tract

Transmission: drop-infection → sensitive! 55°C; 30’


FIGURE 31-1 Pathogenesis of whooping cough.

Medmicro

www.my-pharm.ac.jp

FIGURE 31-3 Binding of pertussis toxin to cell memb ranes.

Medmicro

FIGURE 31-4 Synergy between pertussis toxin and the filamentous hemagglutinin in binding to ciliated respiratory epithelial cells.

Medmicro


Clinical finding:Whooping caugh / Pertussis(Peribronchial inflammation, Intersticiale Pneumonia)

4-Phases:Prodromal, Catarrhal, Paroxysmal, Convalescent

Colonization of tracheal epithelial cells by B. pertussis web.umr.edu/~microbio

www.gesundes-kind.de,www.vaccineinformation.org

Pertussis –paroxysmal Phase

www.thecrookstoncollection.com aapredbook.aappublications.org

www.med.sc.edu

Pertussis - Diagnosis

Lymphocytosis


Diagnosis

Cultivation:Bordet – GengouDirect caugh!Charcoal MediumSerology:IgM, IgA, IgG Detection of DNAPCR

medinfo.ufl.edu


Therapy:Macrolides

Prophylaxis:Active Immunisation – acellular Vaccine DaPT

ToxoidFH/PilusPertactin

DPT = DiPerTe – killed B. pertussis

Pertussis in the USA – 2012 (CDC)

< 1 year 4516 1-6 years 7312 7-10 years 8349 11-19 12484 20+ years 8890 Unknown 329 Total 41880

Brucella

BrucellaeBrucellaeBrucellaeBrucellae

Morpholog y:Gram-negative coccobacilli

Cultivation :Agar – nutrient rich(Serum, Glycerine)Atmosphere: CO2Incubation: days-weeks

Description: Brucella spp. Colony Characteristics: - A. Fastidious, usuall y not visible at 24h. - B. Grows slowly on most standard l aboratory media (e.g. sheep blood, chocolate and trypticase soy agars). Pinpoin t, smooth, entire translucent, non-hemolytic at 48h

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BrucellaeBrucellaeBrucellaeBrucellae

Pathogenesis, Infection, Clinical findings

B. melitensis Goat Maltese feverB. abortus Cattle Morbus BangB. suis Pig Swine Brucellosis

Anthropozoonosis All is Brucellosis„Febris undulans” RES!(undulating fever: „wavelike”)

- from sick animals (meat, milk)- through direct contact or contaminated food- invasion through skin lesions or conjunctiva or GI tract

mucosa

Brucella – source of infection

Medmicro

Brucella –

portals of entry

Figure 28-1 Portals of entry for Brucella species.

Medm

icro

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

DiagnosisCulture: min. 5 daysSerologyAntibody detection

Tube-agglutination (Wright)IgM ChromatographyELISA

Direct detection of DNA : PCRTherapy:Doxycyclin, Rifampicin, Streptomycin

Prophylaxis:Avoid expositionTreatment or annihilation of sick animals

WHO – Bioterror Category B!!!

Brucella IgM

www.kit.nl

BrucellosisBrucellosisBrucellosisBrucellosis

Francisella

Francisella tularensisFrancisella tularensisFrancisella tularensisFrancisella tularensis

Morphology:Gram-negative rods

Survives in wet and cold environment.

Cultivation is prohibited!Only in special LaboratoriesWHO – Bioterror category A!!!

Pathogenesis, Infection- From sick animals- through direct contact or inhalation or per os or via ectoparasites

NO HUMAN-TO-HUMAN TRANSMISSION

Francisella tularensisFrancisella tularensisFrancisella tularensisFrancisella tularensis

Clinical findings:TULAREMIALymphnodes, small granulomas+ ulceration+ necrosisAssociated with diverse symptomscutano-, oculo-, tonsilloglandular, (visible!)thoracal, abdominal - (invisible!) formsGeneralisation – Granulomatous lesions!

Diagnosis: SerologyDetection of DNA: PCRTherapy:Streptomycin, Doxycyclin,Ciprofloxacin

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

Description: Cervical Lymphadenitis in a Patient With Pharyngeal Tularemia ; Patient has marked swelling and fluctuant suppurati on of several anterior cervical nodes. Infection was acquired by ingestion of contaminated food or water. Source: World Health Organization


Description: Chest Radiograph of a Patient With Pulmonary Tularemia

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


Yersinia

Yersinia pestisYersinia pestisYersinia pestisYersinia pestis Genus: Enterobacteriaceae!

Morphology: Gram-negative rods – bipolar staining

www.lonlygunmen.deGiemsa staining www.mja.com.au

www.idph.state.il.us, www2.cnrs.fr, ww.knowledgenews.net,

Yersinia pestisYersinia pestisYersinia pestisYersinia pestis

NO CULTURE! FORBIDDEN!Only in special LaboratoriesWHO – Bioterrorcategory A!!!

Plague: 14th century

Plague in medieval Europe


VIRULENCE FACTORS

Capsule – Protein!V Antigen (Protein) AntiphagocyticW Antigen = Endotoxin

Extracellular Substances- Plasminogen – Activator – Protein (Pla)

spreading, fibrinolytic-Toxin (kills mice)


Pathogenesis, Infection:Source of infection:Rats (and other rodents)→ Elimination of Rats!!!

Transmission:direct contact,Rat-flea-bites

Penetration: skin


Figure 29-4 Pathogenesis of Y. pestis in plague patients.

Medmicro

Clinical findings :1)Bubon ic plague(swollen lymph nodes)

2) Septic form → haemorrhagic inflammation

3) Pulmonary form = Pneumonia ← direct aerogen transmission from human to human (airborne infection → primary pulmonary plague)!


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

Bubonic form

Necrosis of finger tips of septicemic plague.

www.imcworldwide.org

Cutaneous Hemorrhages in Plague. Source www.cdc.gov

Septic form

www.imcworldwide.orgPulmonary plague


DiagnosisClinical pictureDirect detection – microscopic (bipolar!)Real-time PCRSerology – tube-agglutination, IF

Therapy:Doxycyclin, Streptomycin

Biological Weapons – Bioterrorism

Biological Weapon: Microbe, ToxinAim:• to kill individuals and/or whole population • economic damage

Biological war (military conflicts)Bioterrorisms (ideology!)Biological crime (personal)

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

Easy to cultureEasy to spread/transmit – airborneHigh rate of mortalityTherapy? (too late)High number of cases

Biological Weapons – Bioterrorism

THE ENDKorfu, 2006

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