Lecturer name:Dr. Ali Somily &Dr.FawziaAl-otaibiDepartment of Pathology, Microbiology Unit
Lecture Title:Anaerobes of clinical Importance
(Foundation Block, Microbiology)
By the end of this lecture the student should be able to:Describe anaerobic bacteria including their sensitivity to oxygen and where they may be found in the environment and the human body.Differentiate the various types of anaerobes with regard to atmospheric requirement (i.e. obligate anaerobes,Faculativeanaerobes andaerotolerentanaerobes.Describe how anaerobes, as part of endogenousmicrobiota, initiate and establish infection.Name the endogenous anaerobes commonly involved in human infection.
Recognize specimens that are acceptable and unacceptable for anaerobic culture.Give the clues(sign and manifestations) to anaerobic infection, name the most probable etiologic agents of the following(Wound botulism, gasgangrene,tetanus,Actinomycosis,Pseudomembranouscolitis and bacterialvaginosis)Describe the microscopic and colony morphology and the results of differentiating anaerobic isolates.Discuss antimicrobial susceptibility testing of anaerobes including methods and antimicrobial agents to be tested.Describe the major approaches to treat anaerobic-associated diseases either medical or surgical.
Anaerobicspore forming bacilli (Clostridia)Gram negative bacilli non-sporingforming (Bacteroides)Anaerobic streptococci (Peptostreptococcus)Anaerobic staphylococcus (Peptococcus)Gram negativediplococci(Veillonella)Gram positive bacilli (Actinomyces)
Lackcytochrome-cannot use oxygen as hydrogen acceptorMost LackCatalase&PeroxidaseContainflavoproteinso in the presence of oxygen produce H2O2 which is toxicSome lack enzymesuperoxide dismutaseso many killed , peroxide and toxicradiclesenzyme likefumaratereductasemust be in reduced form to work
HABITAT I :
Theseorganism are normal flora in:A. Oropharynxeg. 1.Provetellamelaninogenicus2.Fusobacteria3.VeillonellaB. Gastrointestinal tractFound mainly in the large colon in large numbersTotal number of anaerobes = 1011While all aerobes (including E.coli) =1014examples are(1) Bacteroidesfragilis(2)BifidobacteriumspeciesC. Female genital tract (mainly in the vagina)
FEATURES OF ANAEROBIC INFECTIONS
Infectionsare always near to the site of the body which are habitat.Infection from animal bites.Deep abscessesThe infections are alsopolymicrobialGas formation, foul smellDetection of "Sulphurgranules"' due toactinomycosisFailure to grow organism from pus if not cultureanaerobically.Failure to respond to usual antibiotics.
HOW DOES THE INFECTION BEGIN ?
DISRUPTIONOF BARRIERSTRAUMAOPERATIONSCANCEROUS INVASION OF TISSUESDISRUPTION OF BLOOD SUPPLYDROPS OXYGEN CONTENT OF TISSUEDECREASE IN Eh POTENTIALTISSUE NECROSIS
WHAT ARE THE INFECTION CAUSED BY THESE ANAEROBIC ORGANISMS I
Post operative wound infectionBrain, dental, lungabscessIntra abdominal abscess, appendicitis,diverculitisInfectionof the female genitaltract: Septic abortion, puerperalinfection andendometritis, pelvic abscess or breast abscessDiabeticfootinfections andpilonidalsinus
Whenanaerobic infection is suspected;a) Specimens have to be collected from the site containing necrotic tissue.b) Pus is better than swabs.c) Specimens has to be send to the laboratory within 1/2 hour why?d) Fluid media like cooked meat broth are the best culture media.e) Specimens have to incubatedanaerobicallyfor 48 hours.
Bacteroidesfragilisis always resistant to penicillin.But penicillin can he used for other anaerobesFlagyl(metronidazole) is the drug of choice.Clindamycincan also be used.
ANAEROBIC NON SPORE FORMING BACILLI
ACTINOMYCES SPPACTINOMYCOSISPROPIONIBACTERIUM SPPACNEMOBILUNCUS SPPBACTERIAL VAGINOSISLACTOBACILLUS SPPENDOCARDITISEUBACTERIUM SPPBIFIDOBACTERIUM SPP
Actinomycesare branching anaerobic ormicroaerophilicGram positive bacilliSource of the infection is normal flora and the host usually normal hostPrimary site of the infection is mouth, lung, appendix, uterus with IUD (chronic infection)Infection can spread to the brain, liver, bone and bloodDiagnosis by Gram stain with sulfur granules and growth of molar tooth coloniesTreatment penicillin,clindamycinor tetracycline
STRICTANAEROBEPLEOMORPHICGRAM NEGATIVE BACILLI (COCCO BACILLI)NORMAL FLORA INOROPHARYNXGASTROINTESTINALTRACTVAGINA
GROUP =B. FRAGILIS, B. VULGARIS, B.THETAIOTAMICRON, B. UNIFORMISACCOUNT FOR 1/3 OF ALL ISOLATESRESISTANT TO 20% BILERESISTANT TO MANY ANTIBIOTICSPENICILLIN, KANAMYCIN, VANCOMYCIN, COLISTIN – AND MANY MORENO PIGMENTATION OF COLONIES OR FLUORESCENCE
BACTEROIDES OTHER SP
BACTEROIDESSPECIES OTHER THAN B. FRAGILIS GROUPBILE SENSITIVERESISTANT TO KANAMYCIN ONLYSOME PIGMENTED
OTHER GRAM NEGATIVE RODS
FUSOBACTERIUM NECROPHORUMGRAM NEGATIVE BACILLIPERITONISILLARINTRNAL JUGULAR VEIN THROMBOSISEMBOLI TO THE LUNGPEPTOCOCCUSGRAM POSITIVE COCCI IN CLUSTERSPEPTOSTREPTOCOCCUSGRAM POSITIVE COCCI IN CHAINSBRAIN ABSCESSVEILLONELLA PARVULAGRAM NEGATIVE COCCI
LARGEGRAM POSITIVE RODSSPORE FORMATIONCausative Agents For1.Gas gangrene :Cl.perfringensand othere.gsepticum2.Tetanus :Cl.tetani3.Botulism :Cl.botulinum4.Toxicenterocolitis:Cl.difficile(Pseudomembernouscolitis)
Morphologylarge rods gram +vewith bulgingendosporesLaboratory diagnosisSmear Gram stain Large Gram positive bacilli with few or no WBCsCultureBlood agar withhaemolyticcolonies (double zone ofhaemolysis)Cooked meat mediumGives the NAGLAR'S Reaction & toxin neutralization on Egg yolk medium & toxin is aphospholipase
Can leads to the following diseases1) Wound Contamination2) Wound infection3)Gas Gangrene- most important disease4) Gas Gangrene of the uterus in criminal abortion5) Food Poisoning : Spores are swallowedGerminate in gut after 18 hours(Toxin production)abdominal pain anddiarrhoea
Pathogenesis:Traumatic open wounds or compound fractures lead to muscle damages and contamination with dirt etc,Mainly in war wounds, old age, low blood supply and amputation of thigh (required prophylaxis with penicillinPrevention and TreatmentRemove dead tissue , debris and foreign bodies .Penicillin and hyperbaric oxygen in some cases
Morphologygram +veanaerobic with terminalspore.Drum Stick appearanceLives in soil and animalfeaces.e,ghorse and any wound can infected if contaminated by sporesFace & neck wounds are more dangerous
Clinical FeaturesIncubation period 1-3 weeks (time from infection to the appearance of symptoms)Symptoms:local (not common), cephalic(rare), generalized (most common)Painful muscle spasm around infected wound and Contraction of muscles in the face calledTrismus(Lockjaw) ,RisusSardonicus(facial muscle)Arachingof Back-strychnineOpisthotonusin children.Opisthomeaning "behind" andtonosmeaning "tension",duetoextrapyramidaleffect and is caused by spasm of the axial along the spinal column .
PathogenesisMainly due totetanospasminwhich is powerfulexotoxin(protein) .This organism does not lead to invasion orBacteraemia. Its function to inhibits transmission of normal inhibitory messages from central nervous system at anterior horn cells of cord.DiagnosisMainly by clinical and it is strict anaerobe very motile , spread on agar.
Preventionby vaccinationTreatmentCleaning of wound and removal of Foreign bodySpecific by antitoxin form horse serum but it can lead to anaphylaxis & shock must be tested first or human immunoglobulin. Antibiotics .like penicillin. Supportive treatment by keeping the patient in dark pace, fluids and sedative valium
Foundin soil ponds and lakesToxin isexotoxin(protein) heat labile at 100OC and resist gastrointestinal enzymesItis the most powerful toxin known Lethal dose 1 µg human and 3 kg kill all population of the world .It dictated for bylysogenicphageBotulismFrom canned food., sea foode_g. salmon when it is not well cooked (Spores resist heat at 100oC)then multiply and producetoxin
SymptomsAbnormal eye movement as if cranial nerve affected when bulbar area of the brain affected. Finally the patient might develop respiratory and circulatory collapseInfantileBotulismIngestion ofSporesgermination in thegutBotulism.Child present with week child, cranial nerve and constipationBotulismPatogenesisAttacks neuromuscular junctions and prevents release of acetylcholine that can leads to paralysis
Laboratory diagnosisSuspected food from the patientfaecesculture or serum toxin detection by mice inoculation after weeksparalysis and deathTreatmentMainlysupportive and horse antitoxin in sever casesPreventionAdequate pressure cooking autoclaving and heating of food for 10 minutes at 100OC
Normalflora ingastroentestinaltract after exposure to antibiotics and killing of other normal flora, this organism will multiply witch then produce toxin that has two componentsA–Subunitenterotoxin(cause diarrhea)B-SubunitCytotoxic( kill the cellsienecrosis)PSEUDOMEMBRANE COLITISis the clinical manifestation of this disease which composed of bacteria , fibrin , WBCs and dead tissue cellsSever dehydration , intestinal obstruction and perforation are some of complication of this syndrome
Laboratory diagnosis: this organism hard to grow in the laboratory required special media and growth of the organism in solid media required cell line culture to illustratecytotoxicityof the organism. The simplest method for diagnosis by detection of the toxin in the stool by immunological testing (ELISA)
Treatment:Metronidazoleor and oralvancomycinin sever casesPrevention: This organism form spores and hard to control in the hospital because they are resistant to alcohol decontamination ( use Nahypochlorideinstead).Patient need to be isolated and contact need to be screened to find out if they carrying the toxic strain of the bacteria.
Reference book and the relevant page numbers..
Sherries Medical Microbiology, an introduction to Infectious Diseases.Latest edition, Kenneth Ryan and George Ray. Publisher: McGrawHill.
Dr. Ali Somily &Dr.FawziaAl-otaibi
(Foundation Block, Microbiology)