MGEM2013 Resp Disease Resp Pathogens 22-23 Part III.pptx
Transcript
MGEM2013 Respiratory pathogens lecture PART III Laboratory diagnosis and treatment Learning objectives Describe the processes involved in the laboratory diagnosis and identification of respiratory pathogens Outline the treatment regimens available for respiratory infections Diagnosis / Treatment...
MGEM2013 Respiratory pathogens lecture PART III Laboratory diagnosis and treatment Learning objectives Describe the processes involved in the laboratory diagnosis and identification of respiratory pathogens Outline the treatment regimens available for respiratory infections Diagnosis / Treatment strategies on National Institute for Health and Care Excellence (NICE) guideline / pathways osis primarily on clinical examination and symptoms presented ory investigation for C Reactive Protein (CRP – acute phase protein) – useful for pneumonia diag ology laboratory investigation needed if irresponsive to treatment / persistent infections ical testing also useful for some viral / atypical bacteria infections especially for epidemiology pu ment tics / antivirals not prescribed for self-limiting infections escribe for severe illnesses / systemic unwell Microbiology laboratory testing request Microscopy Culture & Sensitivity (MC&S) Samples for URTI – throat / laryngeal swabs , ear swabs, rarely for sinusitis but if needed – sinus drainage Samples for pneumonia – sputum Incubation duration – mostly overnight More fastidious - Neisserria spp, Haemophilius spp – may take 48 to 96 hours Mycobacterium tuberculosis (TB) – takes up to 7 or 8 weeks Microscopy Culture & Sensitivity (MC&S) Cultural medium – agar in a petri dish with different types of supplements e.g. Blood agar – solid agar with essential nutrients (proteins, ions, pH..) supplemented with blood Microscopy Culture & Sensitivity (MC&S) Some bacteria produce toxins degrading blood – haemolysins Degraded growth areas show ‘clearance zones’ - haemolysis Alpha haemolysis – incomplete degradation showing green colourisation Beta haemolysis – complete degradation showing clearance zones without blood Gamma haemolysis – no observable clearance Common RTI pathogens are beta and alpha haemolytic Microscopy Culture & Sensitivity (MC&S) Some bacteria require specific types of nutrients Chocolate Agar Blood agar heated to high temperature Releasing intracellular coenzyme Nicotinamide adenine dinucleotide (NAD) and haemin Brown colour due to breakdown of blood cells – chocolate colour For cultivation of fastidious bacteria such as Haemophilus and Neisseria Often used in conjunction with prolonged (48 to 96 hours) Microscopy Culture & Sensitivity (MC&S) Biochemical testing – metabolic profile Base on preferential utilisation of metabolites Carbohydrates (sugar) Peptides Organic compouds Positive reactions reveal by pH indicator colour change catabolite colour complex formation Series of tests form a profile – check again database for similarities Please refer the information on this slide to the video files of carbohydrate fermentation and citrate utilisation Microscopy Culture & Sensitivity (MC&S) Recognition of unique structures / antigens by specific antibodies (serotyping) Upon recognition of unique antigens by specific antibodies, ‘clumping’ occurs and can be made visible by additional reagents added to the system e.g. Lancefield grouping for beta haemolytic Streptococci Since antibodies from serum / sera of laboratory animals – this method is called serotyping Most frequently isolated bacterial pathogens Streptococcus pyogenes Aka Group A Strep / BHS (beta haemolytic Strep) Gp A Group A – Lancefield grouping (serological) for Streptococci classification Gram-positive mostly in chains but sometimes pairs Produce complete haemolysis (beta) on blood agar Common human pathogens. Pyogenic exotoxin – streptococcal toxic shock syndrome and scarlet fever Streptococcus pneumoniae Gram positive cocci, often arrange in pairs therefore appearing as diplococci Produces alpha haemolysis on blood agar Encapsulated Causing infections at all levels in the respiratory tract, including sinusitis, otitis media, bronchitis and pneumonia. Up to 60% of people carry Streptococcus pneumoniae as a commensal in the nasopharynx and infection is transmitted in airborne droplets Haemophilus influenzae: Gram-negative coccobacillus (oval in shape) Normal flora in URT Requires additional supplements – chocolate agar Pseudomonas aeruginosa: Gram-negative bacillus Encapsulated, oxidase positive Klebsiella pneumoniae : Gram negative bacillus Encapsulated, oxidase negative Bacterial capsule Varying in compositions between spp but primarily polysaccharides Render resistance to host immune response Endocytosed but resistant to phagolysosomal degradation Resistant to antibody / complement opsonisation Impenetrable by complement Many encapsulated bacterial strains form biofilm Insulated from host immune surveillance / attack Impermeable to antibiotics Atypical pneumonia caused by Legionella pneumophila Gram-negative bacillus (may exist as long filaments) widely distributed naturally in water First isolated in war verterans contracted infection from contaminated central humidifier system (Legionnaires’ disease) Non human-to-human infections - contaminated aerosol Severe pneumonia Confusion, diarrhoea, abdominal pain and respiratory failure Associated with high mortality Culture requires special media containing cysteine and iron Fungal pneumonia Cryptococcus neoformans Found in soil and in bird droppings Affect immunocompromised individuals Thick polysaccharide capsule render resistance to phagolysosomal degradation and other host immune response Pulmonary infections spread to brain causing meningitis Aspergillus Mainly in immuno-compromised patients Conidia small enough to reach the alveoli After attachment extracellular proteases and phospholipases produced Colonisation leads to invasion of pulmonary tissues and penetration of blood vessels by septate hyphae Mortality for invasive aspergillosis is 100% For non-culturable microorganisms Serological testing for antibodies (ELISA, latex / haemagglutination) Most commonly used Convenient Easy to perform Cost effective (mostly) e.g. Mycoplasma, Chlamydia, Infectious mononucleosis Molecular diagnostic techniques (PCR / sequencing) Not conventional unless under specific circumstances such as pandemic Require specific instrumentation and training Highly specific results Non-culturable atypical bacteria Resistant to penicillin type antibiotics due to atypical bacterial cell wall structure Mainly by cell culture (not carried out routinely) Diagnosis often made retrospectively by antibody testing Chlamydia Obligate intracellular parasites - cannot reproduce outside host cell Mycoplasma The smallest bacteria – unlike Rickettsia and Chlamydia, survive outside host cells Rickettsiae Gram negative pleomorphic bacteria – cocci, rods or filamentous Human as ‘accidental hosts’ Transmitted by ticks, fleas and lice e.g. Rocky Mountain fever, typhus and scrub typhus Treatment for RTI NICE guidelines for bacterial RTI Only provide immediate antibiotics prescription if: Systematically unwell Symptoms suggestive of serious illness or complications Primarily Penicillin V / Amoxicillin For penicillin allergy – Clarithromycin / erythromycin If no improvement – amoxicillin coupled with beta lactamase inhibitor ? Which one For reference / interest only: Treatment pathway for URTI https://pathways.nice.org.uk/pathways/self-limiting-respiratory-tract-and-ear-infections-antibiotic-prescribing Treatment pathway for LRTI https://pathways.nice.org.uk/pathways/pneumonia#content=view-node%3Anodes-pharmacological-treatment Treatment of viral infections Primarily supportive Rehydration Anti-inflammatory (why) Rest For immunocompromised or severely affected individuals Aciclovir Aciclovir converted to aciclovir triphosphate by viral enzyme Structurally similar to guanosine triphosphate but blocks further elongation of new strand Inhibiting viral DNA synthesis by DNA polymerase Summary Diagnosis / Treatment strategy ed on National Institute for Health and Care Institute (NICE) guidance / pathways gnosis based on clinical examination and symptoms oratory investigation for C Reactive Protein (CRP) – useful for pneumonia diagnosis microbials not prescribed for self-limiting infections y prescribe for severe illnesses / systemic unwell robiology laboratory investigation needed if irresponsive to treatment / persistent infections – MC ological testing also useful for some viral / atypical bacteria infections especially for epidemiolog Microbiology laboratory testing request Microscopy Culture & Sensitivity (MC&S) Samples for URTI – throat / laryngeal swabs , ear swabs, rarely for sinusitis but if needed – sinus drainage Samples for pneumonia – sputum TB culture takes 7 to 8 weeks as opposed to non-TB bacterial respiratory infection samples investigation which takes typically an overnight incubation or for some cases, within a week For non-culturable (routine) microorganisms Serological testing for antibodies (ELISA, latex / haemagglutination) Most commonly used Convenient Easy to perform Cost effective (mostly) Molecular diagnostic techniques (PCR / sequencing) Not conventional unless under specific circumstances such as pandemic Require specific instrumentation and training Highly specific results NICE guidance for RTI Only provide immediate antibiotics prescription if: Systematically unwell Symptoms suggestive of serious illness or complications Primarily Penicillin V / Amoxicillin For penicillin allergy – Clarithromycin / erythromycin If no improvement – beta lactamase inhibitor ? Which one Treatment of viral infections Primarily supportive Rehydration Anti-inflammatory (why) Rest For immunocompromised or severely affected individuals Aciclovir Aciclovir converted to aciclovir triphosphate Inhibiting viral DNA synthesis by DNA polymerase Learning objectives Describe the processes involved in the laboratory diagnosis and identification of respiratory pathogens Outline the treatment regimes available for respiratory infections nk you for having completed the whole set of three mini-lect Please raise questions in the contents support forum
