Introduction to Infectious Disease 11/1/10
NOTE: This question should have gone in the endocrine study guide – N. meningitidis is a cause of Waterhouse-Friderichsen syndrome (adrenal insufficiency resulting from bilateral adrenal hemorrhage):
Test q: A 23F, previously healthy, died suddenly after complaining of a mild sore throat the previous day. At autopsy, her adrenal glands are enlarged, and there are extensive bilateral cortical hemorrhages. Infection w/which of the following organisms best accounts for these findings? Neisseria meningitidis. REPEATED x4 (2009 #46, answer is incorrectly keyed as “Histoplasma capsulatum”)
Sample Question:
An increase in the number of cases of tuberculosis in New York in the 1980s was due to:
A. multiply drug-resistant M. tuberculosis B. resistance to INH
C. genes for capsule production D. virulent strains from Tailand
E. more homeless and AIDS patients
Old test q – repeated x2
Sample question 2:
A special stain that will demonstrate bacteria, fungi, parasites and viral inclusions is:
A. Gram stain B. Acid-fast stain
C. Modified acid-fast stain
D. Giemsa stain Old test q – repeated x3 E. Silver (GMS) stain
ID Outbreaks in the U.S. Polio
Legionnaire’s Disease Toxic Shock Syndrome Lyme Disease
Human papillomavirus Tuberculosis
Hantavirus
SARS and Anthrax *MRSA
*Clostridium difficile * often hospital-acquired
“Emerging” Infectious Diseases: Microorganisms do not change much Human behavior changes a lot PARALYTIC POLIO
Poor Sanitation (U.S. in 1800) childhood diarrhea
enterovirus infection and immunity paralysis rare
Good Sanitation (U.S. in 1900)
children not infected and do not become immune young adults infected
receptors on anterior horn cells- paralysis Red areas exhibit the highest rates of paralytic polio:
Legionnaire’s Disease: Legionella DFA:
• Legionella is present in fresh water ponds, plumbing systems and aerosols • Sporadic cases occur
• American Legion convention with many elderly attendees with chronic lung disease
• New tools developed to identify Legionella- DFA, urine antigen test and DNA probes
Toxic Shock Syndrome:
• Staphylococcus aureus with TSST-1 toxin
• 1980s- Use of hyperabsorbent tampons (RELY brand) caused an epidemic of cases • Toxic shock still seen sporadically caused by both staphylococci and streptococci Rash of Toxic Shock Syndrome: Gram positive cocci on blood smear:
Lyme Disease:
• Borrelia burgdorferi (spirochete) - rash, arthritis, (meningitis and myocarditis) • deer tick and white-tail deer reservoir
• rat tick and woodrat reservoir in California • more deer-more ticks-more people • wildlife protection-housing development
Rash of Lyme disease: Spirochetes:
Looks like bulls-eye/target.
National Lyme disease risk map w/four categories of risk
Test q: A 30M comes to the physician because he has had joint pain in the right hip and left elbow and a headache for the past week. One month ago, he had similar pain in the left hip and knee, which slowly resolved. He remembers having a ring-like skin rash on his left thigh several months ago after a tick bite. On phys exam, there is joint tenderness but no swelling or deformity of the right hip and left elbow. His heart rate is slightly irregular. Which of the following infectious agents is most likely to produce these findings? Borrelia burgdorferi.
Test q: The incidence of Lyme disease in the US has increased partially due to an increase in the population of: White-tailed deer.
Human Papillomavirus:
• Causes condyloma, dysplasia and cervical carcinoma • There is a more sexually active population
• The incidence of CIS is increasing in the U.S. (but mortality is decreasing)
• ** Most new head/neck squamous carcinoma cases in the US are HPV-associated HPV DNA Types 16 and 18:
• HPV Infection produces proteins E6 and E7 • p53 protein, RB and p21 protein are inactivated
• Apoptosis does not occur and the infected cells are not killed • Dysplasia and cervical cancer may result
Condyloma caused by HPV infection- not malignant ↑
Review HPV questions that popped up on 2nd test:
Test q: A 35F is diagnosed w/squamous cell carcinoma in situ of the cervix. The five year survival rate for this patient if properly treated is: 100%. REPEATED x2
Test q: A 29F presents for pelvic exam and pap smear. What anatomic site is most likely to yield dysplastic or malignant change?
Squamocolumnar junction.
Test q: A pap smear test has the LOWEST sensitivity for detection of: Endometrial adenocarcinoma (Other choices: LGSIL, HGSIL, CIS, Invasive squamous cell carcinoma)
Test q: HPV infections of the cervix have been increasing every year in the US. The number of cases of CIS is also increasing but the number of cases of invasive cervical cancer is decreasing. What explains this paradox? Screening with pap smear.
A 35F patient has a pap smear. The diagnosis is ASC-H. What is the next step? Refer to colposcopy for biopsy
Test q: A 45F presents to a family practice doctor w/oral and vaginal thrush. A gynecological exam reveals an ulcerated and bleeding cervix. A biopsy reveals invasive squamous cell carcinoma of the cervix. Which of the following best explains the development of cancer (oncogenesis) in this patient? Protein E6 and E7 production by HPV type 16 or 18.
.Tuberculosis:
• Mycobacterium tuberculosis
• In the U.S. the people at risk are: homeless, elderly in nursing homes, prisoners and AIDS patients • If people are poor, malnourished, and have crowded living conditions, tuberculosis will flourish
Test q: An increase in the number of cases of tuberculosis in Indianapolis in 2009 is due to: More nursing home patients and immigrants (Other choices: Multiple drug-resistant M. tuberculosis and M. bovis; Resistance to INH; Genes for capsule production; Virulent strains from Thailand)
Above (left): Reported TB cases US, 1982-2004. Decreased when streptomycin was introduced. Dropped til the 1980s – then we had bad recession and introduction of HIV into the population (went back up).
Above (right): Reported TB cases by origin and race/ethnicity, US. U.S. Born: Whites and Blacks most common. Foreign Born: Asians and Hispanics most common.
Test q: In the US in 2004, the population at greatest risk for acquiring active tuberculosis is: Foreign-born Asians. (Other choices: US-born whites, Foreign-born whites, US-born Asians, Foreign-born blacks)
Hantavirus:
• 4 corners area (SW USA – Utah, Colorado, Arizona, New Mexico)
• flu-like illness to hemorrhagic fever with sudden death (carried by deer mice) – similar to Ebola virus
• excess rain leads to bumper crop of pinon nuts • deer mice eat nuts and population increases • Navajos harvest pinon nuts
Test q: HIV has spread world-wide but Ebola Virus has primarily remained in a single, small geographic area. What is the explanation of this inability to create a pandemic like HIV? Ebola Virus is rapidly symptomatic and usually fatal.
MRSA and C. difficile: MRSA
Wide use beta-lactams
Poor handwashing- more patients per nurse/doctor
Clostridium difficile Use of Clindamycin
Use of proton pump inhibitors
Use of quinolones (gattifloxacin, moxifloxacin) Plus not hand-washing
Emerging Diseases ??
The human population changes behavior Microorganisms do not change behavior Koch’s Postulates:
• Microrganism is found in lesions of the disease • Organism is isolated on solid media cultures
• Organisms from culture causes lesions in experimental animals
• Organisms can be recovered on solid media cultures from lesions in the animals Problems with Koch’s Postulates:
• Viruses, Rickettsiae, Chlamydia etc. do not grow on solid media
• DNA Probes and Amplified DNA Probes are more sensitive and more specific than culture (on solid media or tissue culture)
• Therefore, the “Gold Standard” is often NOT culture Categories of Infectious Agents:
• Prions- kuru, C-J Disease, mad cow disease • Viruses
• Bacteriophage/Plasmid • Bacteria
• Chlamidiae/Rickettsiae/Mycoplasma • Fungi
• Parasites (protozoa, worms, ectoparasites)
TB in the upper lobe- caseous granulomas. Immune system breaks down, then TB reactivates in the upper lobe. Ghon Complex or Primary Complex: (LN and
peripheral lesion – macrophages migrate) Tuberculosis is a
communicable disease.
The deermouse and family:
Microorganisms have to overcome a lot of resistance in order to establish an infection. At every point, they are attacked by different defense systems.
Release and Transmission:
• Contact- wound, body fluids or mucosal surfaces (as in STDs) • Cough with aerosol as in TB
• Insect vectors as in malaria, Lyme disease
• Diseases spread from person to person are said to be contagious or COMMUNICABLE
• Diseases acquired in the hospital are NOSOCOMIAL
Test q: A nosocomial infection is best described as one that is: acquired in the hospital.
How Do Microorganisms Cause Disease?
• Direct contact with cell death (Streptococcus produces cellulase) • Release of toxins that enter the blood and kill target tissue (C.
diphtheriae)
• The host response, such as abscess (staphylococci) or granuloma (TB), that destroys host tissue
Viral Surface Protein Receptors:
• EBV- **CD21 (CR2) receptor on B-cells and perhaps
macrophages; atypical lymphocytes in blood are mainly CD8+** • Rabies- acetylcholine receptor on neurons
• Rhinovirus- ICAM-1 on mucosal cells • HIV- CD4 , CXCR4 or CCR-5
Figure: HIV mechanism of attachment HIV Kills CD4 (Helper) T- Lymphocytes: • Only 1/100,000 CD4 lymphocytes are
infected
• Infected cells induce non-infected cells to commit suicide by APOPTOSIS and other mechanisms
How viruses work: take over host machinery.
Bacterial Injury: • Virulence genes
• Salmonella and E. coli have similar virulence genes but E. coli lacks genes for attachment and invasion
• Helicobacter pylori and Corynebacterium diphtheria strains that do NOT produce toxins are not pathogenic (most diphtheria strains do not make a toxin)
Bacterial Adhesins:
• Lipoteichoic acids on Streptococci bind tightly to blood cells and oral epithelial cells • Pili on GN rods and cocci- proteins at the tips of the pili bind sugars
• type I- mannose (UTI)
• type P- galactose (pyelonephritis) • type S- sialic acid (meningitis)
Bacterial Targets:
• Unlike viruses that invade many cell types bacteria primarily infect epithelial cells (Shigella, E. coli), macrophages (TB) or both (Salmonella, Listeria)
• Macrophages- receptors for Ab or C’
• Epithelial Cells- many bacteria bind to integrins such as CR3 (complement iC3b) Bacterial Toxins: Endotoxin, Exotoxin, Enterotoxin
Figure: Diphtheria Exotoxin Fragment B (COOH)- attachment Fragment A (NH2)- enzymatically active Connected by a S-S bridge
Toxin produced in upper airway but travels through blood and causes heart damage. Figure (below): Immune Evasion:
1. Inaccessibility- C. difficile in the intestinal lumen – sends its toxin to do the dirty work 2. Block phagocytosis- S. pneumoniae’s mucopolysaccharide capsule
3. Antigenic Variation- influenza virus and rhinovirus 4. Immunosuppression- HIV and EBV
Diagnosing Infectious Diseases: • Special Stains
• Laboratory Culture
• Nucleic acid probes (DNA probes) • Amplified nucleic acid probes • Clinical History
Special Stains for Microorganisms:
• Gram Stain- G+ and G- bacteria; Candida
• Giemsa Stain- bacteria, fungi (all), viral inclusions, parasites - EVERYTHING
• Silver Stain (GMS)- fungi and a few bacteria
GRAM STAIN: • Any fluid or tissue
• Screen for or identify bacteria • 30 minute turnaround
• GPC, GPR, GNC, GNR and yeast
• Poor sensitivity for filamentous fungi, mycobacteria, parasites and viral inclusions
Gram-negative bacilli: Gram stain of Clostridium perfringens Gram Stain of Candida albicans
Gram-positive rods. Most fungi do not stain w/Gram stain at all.
Giemsa Stain or Diff-Quick (rapid Giemsa) Stain: • Any tissue or fluid
• Works best on smears
• Bacteria, fungi, parasites, viral inclusions • Rapid preparation (1 minute)
Giemsa stain of perispinal abscess: Giemsa stain of H. capsulatum: Giemsa stain of CMV:
Shows many intracellular cocci Histoplasma capsulatum. RBCs, Cytomegallovirus – inclusions (dark areas) See neutrophils, bands. Staphylococcus macrophage w/yeast inside.
MICROWAVE GMS (Silver) STAIN: • Any tissue or fluid- 30 minutes • All fungi including Pneumocystis
• Histoplasma, Cryptococcus, Mucor, Aspergillus • Most sensitive stain for fungi
• Poor for bacteria and mycobacteria • O.K. for Nocardia and Actinomyces
ACID-FAST STAINS (Kinyoun, Ziel-Neelsen, etc.): • Carbolfucsin or fluorescent(more sensitive) • “modified” AFB for Nocardia- weak acid wash
• Mycobacteria, Nocardia, Rhodococcus, Cryptosporidium
• Sputum must be decontaminated and concentrated for high sensitivity
Fluorescent Acid-Fast Stain: Acid-Fast Stain (Mycobacterium avium): Acid-fast stain:
M. tuberculosis Lymph node biopsy – traditional acid-fast stain (red) M. tuberculosis - “cording”
Cording – wrap around each other like cords in a rope. BAD sign – high virulent strength.
Nocardia Filamentous Nocardia: Nocardia (modified-acid-fast stain):
Partially acid-fast and may grow on “TB” media Gram Stain (Gram positive filaments)
If a question says “WEAKLY ACID FAST” = NOCARDIA. SPECIAL SILVER STAINS:
Warthin Starry, Dieterle – spirochetes Treponema pallidum
Legionella Bartonella
DIRECT FLUORESCENT ANTIBODY (DFA) STAIN: Sputum or respiratory washings
Legionella, Bordetella persussis Herpes virus and others **Currently we are identifying Influenza A by DFA and most are H1N1 (by PCR)
Dieterle/Warthin-Starry Stain: Fluorescent antibody stain: Herpes-infected cells
Laboratory Culture:
• Solid Media- wounds, throat, urine, sputum • Broth- blood and sterile body fluids
• Tissue Culture- virus, Chlamydia (rare) • Now, most chlamydia testing is
molecular.
DNA Probes:
• GC and Chlamydia from urogenital specimens *(including Pap Smears)
• ID of bacterial, fungal and mycobacterial growth in broth and on solid media
• Candida, Gardnerella and Trichomonas from vaginal discharges
DNA Probe:
If we use a direct DNA probe, we target ribosomal RNA because there are 10,000 copies/cell, while there are very few DNA copies per cell. We see if it hybridizes (HPA, above).
Amplified DNA Probes: :
• PCR, LCR, TMA, SDA
• GC and Chlamydia- urogenital swabs or urines • M. tuberculosis in respiratory specimens
(TMA – transcription-mediated amplification) (SDA – strand displacement amplification)
SDA: TMA: Linear SDA:
Real-time PCR for MRSA, GBS and C. difficile: • Results in 1-2 hours (short as 45 minutes)
• Methicillin-resistant Staphylococcus aureus (MRSA) and Group-B streptococcus/ S. agalactiae (GBS) are FDA-approved tests
• C. difficile
• Other tests are ASRs (analyte specific reagents) that must be validated in your own laboratory; NO ready-to-use kits
Test q: The chief advantage for choosing real-time PCR over direct DNA probe or amplified DNA probes is: decreased turnaround time for results
Common DNA – blue. Unique DNA to organism – pink. Yellow is what we test for by PCR.
MRSA: MecA gene IDI-MRSA™ Provides a Definitive Result
Methylicillin-resistant staph – standard beta-lactam antibiotics don’t work because of altered penicillin- binding protein.
Test q: A 50F is admitted for a right knee replacement. The surgical procedure was a success, but 3 days (72hr) post-operation, the surgical site is red, warm, and swollen w/drainage of pus. Real-time PCR is performed on the pus and is (+) for Staphylococcus aureus w/MecA gene present. This history is consistent with: Nosocominal MRSA. (Other choices: Community-acquired MRSA; S. aureus, methicillin susceptible; Coagulase-negative
staphylococcus; Viridans streptococci)
Target Specific Fluorogenic Probe – Molecular Beacon:
Rapid IDI-MRSA/MSSA Assay* - under development
• Assay under development, allowing the direct detection of MRSA and MSSA in under 2 hours of laboratory time • Plan to validate four different specimen types:
• Positive blood bottles, wound swabs, nasal swabs, and rectal swabs
• DNA detection of the SCCmec-orfX junction found only in MRSA provides definitive identification of MRSA
• Only molecular method to definitively identify MRSA in specimens which contain methicillin-resistant coagulase negative
Staphylococci and mecA negative S. aureus. Normally such a
combination would yield false-positives in other PCR methods • Detects both HA-MRSA and CA-MRSA strains (i.e. USA300)
Fluorescence is monitored for every cycle:
The secret to real-time PCR is that the reaction doesn’t have to go to Positives and negatives. completion – can read during the beginning of the reaction.
Inflammatory Response to Infectious Diseases: • Host Dependent- eg. Greatly reduced in AIDS
• PMNs- pyogenic response to many bacteria: staphylococci, streptococci, GNRs • Granulomatous- tuberculosis, histoplasmosis
• Lymphocytic – viruses
• Bacteria – usually neutrophils (PMNs); liquefactive necrosis; abscess • Fungi – usually macrophages/lymphocytes; granulomas
• Virus – usually lymphocytes; may show cytopathic effect (CPE)
• Immune Deficient Host- there may be absence of the normal inflammatory response and high numbers of microorganisms (eg. M. avium in AIDS) “All bets are off”
Bacterial abscess in the brain:
Abscess: TB Granuloma:
Pus pocket Destroyed tissue, lots of neutrophils Kidney filled w/caseating
granulomatous inflammation
Lung slice, TB granuloma:
TB granuloma – spontaneously healed. Most important mediator is IFN-γ
Primary and Secondary TB: 1° – peripheral lesion and lymph node involvement, mid lung fields.
Reactivation (2°) – primarily infection in upper lung because TB likes oxygen (and most oxygen is in lung apices) Inflammatory Response:
• Cytopathic- viral inclusions in Herpes and CMV or dysplastic changes in HPV • Necrosis- Clostridium perfringens in gas gangrene
Koilocytosis of HPV:
Example in HPV – infects cervical epithelial cells koilocytosis
ID Lab Preview 11/2/10
Special Stains for Microorganisms:
• Gram Stain- G+ and G- bacteria; Candida
• Giemsa Stain- bacteria, fungi (all), viral inclusions, parasites • Silver Stain (GMS)- fungi and a few bacteria
• (Modified)-Acid-Fast Stain- mycobacteria and Nocardia
GRAM STAIN: Gram negative bacilli:
• Any fluid or tissue
• Screen for or identify bacteria • 30 minute turnaround
• GPC, GPR, GNC, GNR and yeast
• GPC = Gram+ cocci; GPR = Gram+ rods; GNC = Gram neg. cocci; GNR = Gram neg. rods
• Poor sensitivity for filamentous fungi, mycobacteria, parasites and viral inclusions
Gram (+) Clostridium perfringens: Gram Stain of Candida albicans:
Ocassionally, yeast can be seen w/a Gram stain. Giemsa Stain or Diff-Quick (rapid Giemsa) Stain:
• Any tissue or fluid
• Works best on peripheral blood smears • Bacteria, fungi, parasites, viral inclusions • Rapid preparation (1 minute)
Giemsa stain of a perispinal abscess: Giemsa stain: Giemsa stain of CMV:
Staphylococcus: see many intracellular cocci. Histoplasma capsulatum Cytomegallovirus: viral inclusions in the
nuclei. Also have cytoplasmic inclusions.
MICROWAVE GMS (Silver) STAIN: • Any tissue or fluid- 30 minutes
• All fungi including Pneumocystis, Histoplasma, Cryptococcus, Mucor, Aspergillus • Most sensitive stain for fungi
• Poor for bacteria and mycobacteria • O.K. for Nocardia and Actinomyces
Ribbony appearance of Mucor: Silver Stain of H. capsulatum: Cysts of Pneumocystis jiroveci:
Above: Can look like a tea cup from the
side – can also have groove in it (top left) or dot in it (middle). Pneumocystis is actually a fungus, but see no budding. Has characteristic forms on Silver stain.
ACID-FAST STAINS (Kinyoun, Ziel-Neelsen, etc.):
• Carbolfucsin or fluorescent (more sensitive for Mycobacterium tuberculosis) • “modified” AFB for Nocardia- weak acid wash
• Mycobacterium, Nocardia, Rhodococcus, Cryptosporidium
• Sputum must be decontaminated and concentrated for high sensitivity
Acid fast stain: some organisms can be stained w/certain stains, and if you bleach them w/acid, the stain goes away (this is the way most bacteria are – non-acid fast). If you stain an organism and unsuccessfully try to remove the stain w/acid, the organism is acid-fast. MODIFIED Acid fast used for NOCARDIA.
M. tuberculosis in a gastric biopsy: Fluorescent Acid-Fast Stain of M. tuberculosis
TB shows “cording” (Mycobacteria wrapping More sensitive stain.
around each other); blue organisms are Candida
SPECIAL SILVER STAINS: • Warthin Starry, Dieterle • Treponema pallidum • Legionella
• Bartonella
Silver stains used on spirochetes and other small rods.
DIRECT FLUORESCENT ANTIBODY STAIN • Sputum or respiratory washings
• Legionella, Bordetella persussis, Herpes virus and others
• High specificity/Low sensitivity
Dieterle/Warthin-Starry Stain: Fluorescent antibody stain: Herpes-infected cells
Shows Treponema pallidum (syphilis)
Bronchopneumonia:
• Patchy or “hit and miss”
• Staphylococcus, GNRs, anaerobes • Aspiration with spread through the airways
Bronchopneumonia: Patchy bronchopneumonia:
* = Pneumonia. Abnormal part is paler – lots of WBCs. Can also see patchy involvement on CXR. Lobar pneumonia:
• Involvement of the entire lobe (most of it)
• Streptococcus pneumoniae, Klebsiella pneumoniae, Haemophilus influenzae • Encapsulated bacteria
• Aspiration then spread through alveolar walls (pores of Kohn)
Test q: Bacteria noted to cause lobar pneumonia include: Streptococcus pneumoniae and Klebsiella pneumonia.
Above (left): In lobar pneumonia, have pores that the organisms can squeeze through because they have a capsule. Most bacteria
and other organisms cannot. Get involvement of one alveolus spreads to the next one, to the next one, and so on.
Above (middle): Grossly, have entire lobe involved (bottom) and entire lobe uninvolved (top)
Above (right): Lobe is completely whited out. No breath sounds heard in this area. Percussion – dullness over area.
Bronchopneumonia – would hear crackles.
Paragonimus westermani:
• Human lung fluke – one of the few parasites that migrates through the lungs
• Granulomatous reaction to the eggs
Tremendous granulomatous inflammatory response Within granulomas,
have multi-nucleated histiocytes, rim of lymphocytes/fibrosis.
Can see organism’s eggs, too.
Aspergillus fumigatus (A. flavus):
• Opportunistic infection in transplant and hematology/oncology patients
• Neutropenia
• Narrow, septate hyphae that branch at acute angles (45 degrees or less)
Figure: High power – Silver stain (best for these fungi). See cross-walls/septae. Branches at 45* angles or less.
Aspergillus – hyphae in blood vessel walls: Vascular invasion is common (w/infarction):
Aspergillus is noted to invade blood vessles – Lung w/multiple thrombi in vessels. this is a bv that has thrombosed.
Mucormycosis (Rhizipomycosis):: • Mucor and Rhizopus
• Diabetic ketoacidosis (DKA), transplant/heme-onc • Nasal sinuses, lung, GI tract, brain
• Rhinocerebral mucormycosis – most disastrous
complication – grows through bone, sinuses, into brain. • Morphologic identification or culture
• Broad irregular ribbon-like hyphae, no septae (aseptate), right-angle branching
• Aseptate fungal hyphae (no crosswalls) • Branch at 90 degrees (right angles)
• Diabetics (poorly controlled) and transplantation patients • Northern USA – Mucor. Southern USA - Rhizopus • 90° branch coming off. Very folded, ribbony
Above (left): Involves blood vessles. Looks like jigsaw puzzle – irregular. Donut shapes = hyphae cut in cross section (like a pipe). Above (right): All the donuts are hyphae cut in cross-section. Broad hyphae, no cross-walls. Can see branching over 45°.
Actinomycosis:
• Sulfur granules (yellow on gross) • Sulfur granules (pink/blue on H&E) • Filamentous anaerobic bacteria
• Gram-positive, acid-fast- negative filamentous bacteria (Actinomyces)
• Aspiration
• Draining fistulas are common
Grows in the mouth. Poor oral hygiene can cause it. Can get “lumpy jaw” or even fistulas growing into the lung or abdomen. Nocardia is also filamentous but AEROBIC. Nocardiosis is acid-fast POSITIVE.
Gram-positive, filamentous Actinomyces
At high magnification, see filamentous bacteria that make up the colonies.
Test q: A 56M farmer presents to the ED w/cough, fever, weight loss, and a draining fistula in his neck. Grossly, the draining material contains yellow granules. Filamentous bacteria are recovered on anaerobic media. The bacteria are Gram-variable and non-acid-fast. Diagnosis? Actinomyces israelii.
Test q: A 70F has a fever and a cough productive of yellow sputum. On phys exam, there is dullness to percussion at the left lung base. A chest radiograph shows areas of consolidation in the left lower lobe. Despite antibiotic therapy, the course of the disease is complicated by abscess formation, and she dies. At autopsy, there is a bronchopleural fistula surrounded by a pronounced fibroblastic reaction. Small, yellow, 1-2mm “sulfur granules” are grossly visible within the area of abscess formation. Which of the following organisms is most likely to produce these autopsy findings? Actinomyces israelii.
Colonies of Actinomyces: Colony of Actinomyces in a bronchiole: Actinomyces colonies on H&E Stain:
Yellow sulfur granules H&E Stain Have orange-pinkish rim around them. Background cells are neutrophils. Salpingitis: Inflammation/infection of the fallopian tube.
• Chlamydia trachomatis • Neisseria gonorrhoeae • Sterility, ectopic pregnancy
Chlamydia (C) and gonorrhea (G) do the same things when they infect cervix/fallopian tubes. G is more virulent – will do more damage, more likely to be symptomatic. C – less damage, more likely to be asymptomatic (so more likely to go untreated). Can destroy/scar tubes so much that eggs can’t pass ectopic pregnancies, problems, sterility.
Above: Salpingitis. All the material inside the lumen is pus. Will see lots of pink areas – congested blood vessels.
Fimbriae – should be lined by columnar epithelium w/cilia. In lumen is the pus.
Abscess w/segmented neutrophils:
Above (left): Lung abscesses. Often see air-fluid level in an abscess (green circle) – see black = air, gray = dense, fluid. Above (middle): Abscess in brain – if chronic, can get fibrotic edge.
Above (right): See lots of neutrophils in abscesses – some have 3 lobes. Some undergoing apoptosis – live 4 hours and die by cell
suicide.
Appendicitis:
• Neutrophils are easiest to see in the muscle and fat
• Leukemoid reaction (left shift) is common – high granulocyte count. • Pyuria due to involvement of the ureter
Neutrophils:
Pyelonephritis:
• Most are ascending (post bladder infection)
• E. coli is most common cause bc it’s the most common cause of cystitis – organisms swim upstream to kidney. • High fever, chills , pain
• “Thyroidization of the kidney” seen on H&E-stained sections Thyroidization:
Look like thyroid follicles w/blue cells (~parafollicular cells). These See glomeruli, so know it’s kidney. are actually tubules in the kidney filled w/protein and lymphocytes.
Diphtheria:
• Corynebacterium diphtheriae
• C. diphtheriae can be normal flora- do toxin assay for diagnosis of diphtheria
• Exotoxin destroys myocytes myocarditis can cause a fatal arrythmia heart failure • Abscesses in the myocardium – die of either heart failure or myocarditis
Pseudomembrane:
Syphilis:
• Obliterative endarteritis seen in primary, secondary and tertiary syphilis – arteries are attacked
• Plasma cells surround vasa vasorum
• Tertiary- ascending or thoracic aorta is most commonly involved (80%) • Aneurysms in abdominal aorta usually due to atherosclerosis. • Treponema pallidum
Chancre on Chest
Aneurysm in ascending aorta: Inflammation in aorta wall: Silver stain:
Shows disorganized elastic tissue. Reticular and elastic fibers are destroyed – wall is weakened and balloons out to form aneurysm.
Tuberculosis:
• Mycobacterium tuberculosis • Communicable
• Necrotizing granulomas (microscopic) is usual but non-necrotizing granulomas are possible • Caseous necrosis (gross)
• Ghon complex in lung (primary TB)
• Apical disease (secondary/reactivation TB)
Microscopically, can see bronchus (cartilage) w/pseudomembrane – mixture of inflammatory cells, dead epithelial cells, and
Necrotizing granuloma:
* Large granulomas (hilar lymph nodes) ** Small granulomas (miliary TB)
Lymphadenopathy – esp. in hilar/central part of lung
Histoplasma capsulatum: • Not communicable
• Necrotizing granulomas – look just like TB granulomas. Until you culture or special stain, can’t distinguish. • Small (2-3 microns), budding yeast
• Intracellular
• Disease almost identical to TB but not communicable. Get it by inhaling conidia from soil.
H&E Stain with yeasts of H. capsulatum: Giemsa stain: GMS (Silver) stain:
Hard to see yeast. Sample from blood. Best seen w/silver stain.
Coccidioides immitis: “West Coast Histo” • California and Arizona
• Necrotizing granulomas
C. immitis Spherules – H&E: GMS (silver) Stain of C. immitis: PAS Stain of Coccidioides immitis:
Above: Inflammatory cells outside. Can see endospores in spherule. SPHERULES = COCCIDIOIDES!
Test q: A 50y/o resident of Phoenix, Arizona (in 2005, was a resident of Southern Cali) has had a cough that has persisted for one month. On phys exam, his temp is 38.1C. A chest radiograph shows a 3.5-cm opacity w/central cavitation in the right apical region. An open lung biopsy is performed to exclude cancer. Microscopic exam of the biopsy specimen shows caseating granulomatous inflammation containing 60-µm spherules filled w/smaller, rounded structures. Which of the following organisms is most likely to produce these findings? Coccidioides immitis. REPEATED x2
Test q: A 50M who recently moved to Indianapolis from Arizona develops fever, cough, and lymphadenopathy. A fine-needle aspiration of a cervical lymph node shows large (50-75 microns) round bodies. Budding is not seen. Diagnosis: Coccidioides immitis.
Blastomyces dermatitidis:
• Large yeast with Broad-Based Buds
• Pseudoepitheliomatous hyperplasia mimics (clinically and microscopically) squamous carcinoma in skin and in the larynx – epithelium proliferates, piles up, looks like cancer
• Inflammation is mixed: histiocytes with giant cells and abscesses with neutrophils
H&E shows broad-based-budding: GMS (silver) Stain:
Test q: A 55M presents to the Derm clinic w/fever, cough, and a tumor-like growth on his nose. A biopsy of the nose mass is performed and the H&E-stained tissue shows pseudo-epitheliomatous hyperplasia but no cancer. A silver (GMS) stain shows large yeasts that are at least 12 microns in diameter and show broad-based buds. A capsule stain (mucicarmine) is negative. Diagnosis? Blastomyces dermatitidis.
Test q: Granulomatous inflammation w/numerous polymorphonuclear leukocytes is typical of: Blastomycosis
Pneumocystis pneumonia:
• (Pneumocystis carinii pneumonia) or PCP • Pneumocystis jiroveci – new name • Two forms: cysts and trophozoites • Cysts on GMS stain
• Trophozoites on Giemsa stain
Pneumocystis pneumonia on H&E Stain
Giemsa Stain of BAL: GMS stain:
Cysts Trophozoites Pneumocystis cyst with 8 trophozoites. Cysts (“cups, targets, grooves”)
(BAL = bronchoalveolar lavage)
P. jiroveci has cup-saucer shapes, target middle shapes, grooves, but NO BUDDING.
Cryptococcus neoformans:
• Variably-sized budding yeast with thick capsule of mucopolysaccharides • India Ink stain (poor sensitivity and specificity)
• Latex agglutination on CSF or serum has high sensitivity and specificity • Cryptococcal meningitis 100% fatal if untreated
Test q: The optimal test for Cryptococcus neoformans in CSF is: Latex agglutination.
Test q: A 25y/o AIDS patient presents in the ED w/a severe headache. You are concerned that he might have cryptococcal meningitis which if untreated, is 100% fatal. The most sensitive and specific test for detection of Cryptococcus neoformans in spinal fluid (CSF) is: Latex agglutination. C. Neoformans in glomeruli (H&E): C. Neoformans in bone marrow: Bone marrow (higher magnification):
Above: Glomeruli filled with little round bodies – always lots of clear space around them because of capsule.
GMS Stain- C. neoformans India Ink:
Chest wall biopsy. Capsule lots of space. C. Neoformans in CSF
Latex Agglutination: See +/- tests below. Particles coated w/antibody, mixed w/CSF – if C. Neoformans antigen present, cross-link and clump.
Polio:
• Anterior/motor horns have receptors
• Paralytic polio occurs in non-vaccinated adults if infected by the virus
Polio: Anterior horns are destroyed. Best seen on low power
Entameba histolytica:
• Flask-shaped ulcers in the colon
• Erythrophagocytosis- amoebae ingest RBCs; if so, it is E.
histolytica
H&E- Flask-shaped ulcer of E. histolytica: Erythrophagocytosis:
E. histolytica – see erythrophagocytosis
Strongyloidiasis:
• Autoinfection- the entire life cycle can occur in humans if they are immune compromised • Association with HTLV-1 infections
The only worm parasite that can go through entire life cycle in the host. No intermediary/secondary host needed. Often patients come in w/pneumonia, are
coughing up the organisms. Calcified worms H&E Stain of
Strongyloides
(calcified worm)
Can find ameobae containing RBCs – erythrophagocytosis.
Adenovirus:
• Nucleus only inclusions • Cowdry B type in Adenovirus • Cowdry A type in Herpes
H&E Stain with Adenovirus inclusions in hepatocyte nuclei Big viral inclusions – see big purple blobs in the nuclei. By history, we know it’s adeno (would not necessarily know by this pic).
CMV:
• Enlarged cells
• Intranuclear (Cowdry B) and cytoplasmic inclusions are present in some cells Below: Arrows = CMV-infected cells CMV infection in lung:
Enlarged (cytomegallic) cells – see outline of nucleus, viral particles are concentrated in one area.
Varicella – in Herpes family, associated w/nerves • Nuclear inclusions
• Cowdry B
• Ganglia and adrenal medulla
• Won’t be able to tell apart from CMV histologically. Involves ganglia. Below: VZV-infected cells and hemorrhage: VZV – Adrenal Gland:
Toxoplasmosis: • Brain abscess
• “Ring-enhancing” lesions • Cysts
H&E Stain of brain biopsy with Toxoplasma: Cysts of T. gondii in brain- H&E Stain:
See cyst forms of toxoplasmosis.
Hepatitis:
• Lymphocytes in the portal zones • Loss of hepatocytes
• Cirrhosis
Chronic Hepatitis: Hepatocytes w/inflammatory cells. Chronic Hepatitis: Plasma cells, lymphocytes.
Get lots of lymphocytes (viral infection) – inflammation around the bile ducts/artery/vein.
Respiratory Infections 11/8/10
OBJECTIVES:
• Understand how anatomical barriers protect the host from developing respiratory infections • Describe the modes of transmission for specific microorganisms that cause respiratory infections • List major factors that predispose people to respiratory infections
• Discuss the pathogenesis of specific respiratory infections • Describe the pathologic features of specific respiratory infections
• Know how infectious diseases of the respiratory tract are diagnosed in the laboratory EXAMPLE QUESTION
ICAM-1 serves as the receptor for attachment of: A. Mycobacterium tuberculosis
B. Influenza viruses C. Haemophilus influenzae
D. Rhinoviruses Old test q – repeated x2 E. Histoplasma capsulatum
Topics for today’s lecture: • Influenza
• Bacterial pneumonia • Bronchopneumonia • Lobar pneumonia
• Selected pathogens and diagnostic considerations • Tuberculosis
• Histoplasmosis • Coccidioidomycosis
Intro: Respiratory Tract Infections:
• Among the most common and least preventable of all infectious diseases
• A diverse array of infections ranging from common colds in previously well persons to life-threatening pneumonia in patients with other severe problems
• Caused by a wide variety of microbial agents – will be talking about mostly acute but mycobacterium is more chronic.
• Antimicrobial resistance is increasing
• Diagnostic methods are inadequate but evolving • Community acquired pneumonia is common
• 5.6 million people (est.) annually in US resulting in 1.3 million hospital admissions per year • And in those over age 65, the number one cause of death from infectious diseases
• Nosocomial (hospital acquired) pneumonia is the leading cause of death from infection in US hospitals; its occurrence prolongs hospital stays about 8 days
• Cost of pneumonia care in US hospitals estimated to be over $40 billion Viral Respiratory Infections:
• Frequent, contagious, spread by droplets; range from common cold to pneumonia • May damage bronchial epithelium, obstruct airways & lead to bacterial superinfection • Upper Respiratory: rhinitis, sinusitis, otitis media, pharyngitis & tonsillitis
• Lower Respiratory: laryngotracheobronchitis, bronchiolitis, interstitial pneumonia, & pleuritis • Most important: Rhinoviruses & Influenza viruses
Rhinoviruses:
• 60% of common colds due to rhinoviruses: other causes = coronavirus 15% (also is agent of SARS - Severe
Acute Respiratory Syndrome), influenza virus, parainfluenza virus, respiratory syncytial virus (RSV), adenovirus,
& enterovirus (1-10% of colds)
• Rhinoviruses: picornavirus family (small RNA viruses with single stranded RNA genome) • >100 serotypes
• Rhinovirus binds intercellular adhesion molecule (ICAM-1) on respiratory epithelial cells & induces mucus secretion via bradykinin release
• Dx by clinical findings, but bx of nasal mucosa would show: abundant mucin/mucosal edema, lymphocytes & plasma cells (nonspecific)
Test q: A 24M has a fever and runny nose, sneezing, and coughing that have worsened over the past four days. The symptoms abate, and he has sequelae (on 2005 test, he has no sequelae). This infection is most likely to be promoted by binding of which of the following organisms to intercellular adhesion molecule-1 (ICAM-1)? Rhinovirus, REPEATED x2
Test q: Most respiratory viral infections are accompanied by an interstitial infiltrate of: Lymphocytes.
Influenza Viruses:
• Single-stranded RNA • Types A, B, or C
• Subtypes (H1 - H3: N1 or N2) determined by viral hemagglutinin & neuraminidase in lipid envelope – e.g., H3N2, H1N1 (swine flu 2009)
• Viruses are spread person to person by airborne droplets or contact with contaminated hands or surfaces • Major mode of prevention?
• Epidemics through mutations of hemagglutinin (H) & neuraminidase (N) that allow virus to escape host antibodies – Antigenic drift: minor antigenic change due to point mutations
– Antigenic shift: major antigenic change (both H and N replaced) due to genetic reassortment between animal & human influenza A viruses (but not B or C)
• Pandemics due to novel influenza viruses: 1918, 1947, 1957, 1968, 1968, 1977, and 2009 – 1918 Spanish flu killed 20-40 million world wide
– As of 11-1-09, pandemic H1N1 2009 was in > 199 countries & overseas territories, & had infected > 482,300 & killed ~ 6000
Test q: The H5N1 stain of avian influenza A in Southeast Asia led to over 85 human cases w/over 200 deaths within the past year. There are reports that the virus has the potential to jump from birds to humans, and it has been found in pigs in China, raising fears that they could serve as a “mixing bowl” making it easier for the virus to mutate and spread to humans. This “mixing” which would result in a major antigenic change (both H and N replaced) is termed: Antigenic shift.
Test q: The following is the mechanism by which influenza causes disease: Mutations in hemagglutinin and neuraminidase allow escape from host antibodies. REPEATED x2
Influenza: More Characteristics:
• Clinical: rapid onset of fever, chills, headache, myalgias, arthralgias, dyspnea, cough & anorexia
• Pathology: nonspecific mucosal hyperemia, lymphomonocytic and plasmacytic infiltrate in submucosa, mucus overproduction
• Pneumonia: persons with underlying heart & lung disease at risk
• Clearance of infection - occurs when cytotoxic T cells kill virus-infected cells • Host antibodies to H and N prevent future infection with that specific virus
• Lab dx: swab of nasopharynx, throat swab, combined nose/throat swab, nasal washes, or bronchial lavages – virus isolation/culture – gold standard but takes 3-7 d
– viral Ag detection by direct FA (fluorescent antibody); sensitivity at CPL ~ 85-90%; takes 4 h
– multiplex RT-PCR (reverse transcription PCR) also used at CPL; more sensitive; but slower than FA; takes overnight
– Rapid Ag point-of-care tests are the least sensitive/false negs; takes 15 min
• Rx: Tamiflu (oestelamivir) and Relenza (zanamivir) effective in 1st 24-48h vs influenza A (neuraminidase inhibitors of both influenza A & B)
Above: Tremedous mucin outpouring in the lumen – Mucosa is flattened, eroded, ulcerated.
Monocytic infiltrate – lymphocytes, macrophages
Routes By Which Bacteria Get Into Lungs: • Aspiration of contaminated oropharyngeal
contents (e.g. pneumococci, GNRs, anaerobes)
• Inhalation (M. tuberculosis, Legionella, plague)
• Bacteremia
• Direct extension into lungs
Factors That Predispose To Bacterial Pneumonia:
Age (extremes) Cystic fibrosis Alcohol Debility in general Anesthesia Edema; congestion Marrow transplant (CHF)
Malignancy Cerebrovascular illness Chemotherapy; Immune deficiencies immunosuppression Obstruction
COPD Viral infections Splenic dysfunction Cigarette smoking Ventilator use Cirrhosis
Autopsy findings in lungs of a patient who died (Fall 09) of H1N1/09
Very severe pneumonia, involved most lobes, was bilateral
1. Bronchus – in the lumen is mucoid material w/lots of cells. Epithelial cells have fallen apart – sloughed.
2. Marked congestion and hemorrhage. Much cellular activity – including macrophages (very active there), lymphocytes.
3. Alveoli filled w/edema fluid. Alveolar septae are widened, filled w/RBCs 4. See foci of acute inflammation, PMNs in the mix.
These findings show variation – in immunocompromised host, can have chronic findings that you don’t see in others.
1
2
Community-Acquired Pneumonia: Pathogens: • Streptococcus pneumoniae • Haemophilus influenzae • Moraxella catarrhalis • Staphylococcus aureus • Legionella spp.
• Enterobacteriaceae (e.g., Klebsiella
pneumoniae)
• Pseudomonas aeruginosa
• Viruses, Mycoplasma, Chlamydophila
Hospital-Acquired Pneumonia:
• Staphylococcus aureus: MRSA > MSSA • Gram-negative rods
– Enterobacteriaceae (Klebsiella spp.,
Escherichia coli, and others)
– Pseudomonas aeruginosa,
Stenotrophomonas maltophilia, Acinetobacter spp.
• Legionella spp.
• Anaerobes (aspiration)
• Viruses (Influenza, RSV, Parainfluenza Morphology of Bacterial Pneumonia:
• 2 frequently overlapping GROSS morphologic patterns – Bronchopneumonia
– Lobar pneumonia
• Caused by a variety of Gram-positive and Gram-negative bacteria
Bronchopneumonia Comparison of Bronchopneumonia and Lobar Pneumonia:
• Patchy exudative consolidation of lung parenchyma
• Gross: dispersed, elevated, focal firm areas • Microscopic: acute neutrophilic suppurative exudate filling air spaces and airways at level of bronchi and bronchioles (with or without lots of edema fluid or RBCs)
• Aggressive disease may result in abcess • Organization may result in fibrous scarring
Lobar pneumonia:
• Involves large portion of lobe or entire lobe • Bacteria spread alveolus-to-alveolus through
pores of Köhn
• Most common cause = pneumococcus (Streptococcus pneumoniae), but occas. K.
pneumoniae, staphylococci, streptococci, H. influenzae
• “Classic” sequence of stages portray natural history of uncomplicated lobar pneumonia – but infrequently seen because of antibiotic therapy
“Classic” stages of Lobar Pneumonia: • Stage 1
– Congestion and inflammatory edema – 1st 24 hours • Stage 2
– Red hepatization (consolidation) – looks like liver tissue
– Gross: red, firm, liver-like
– Micro: confluent exudate with neutrophils and RBCs • Stage 3
– Grey hepatization – blood removed, abundant neutrophils, fibrin mixed together
– Gross: gray-brown
– Microscopic: rbcs disintegrate leaving fibrinosuppurative exudate
• Stage 4
– Resolution – enzymatic and cellular degradation – normal structure restored
Resolution is less likely in Klebsiella pneumonia.
Lobar pneumonia: Somewhere between
congestion and red hepatization. Alveolus full of neutrophils, engorgement of alveolar septal vessels w/RBCs
Red hepatization – looks like liver. Neutrophils, exudate.
Complications of Lobar Pneumonia: • Abscess
• Pleuritis and empyema – thoracic cavity pus • Organizing pneumonia (leaves residual fibrosis) • Bacteremia and sepsis
• Infarct
Laboratory Diagnosis of Pneumonia:
• Sputum specimen collected and sent to lab • Direct smears for Gram stain
• Specimen plated on various media
Sputum being prepared for Gram stain
Diagnosis of Pneumonia due to Streptococcus pneumoniae:
• Gram-stained sputum containing many neutrophils and typical Gram-positive, lancet-shaped diplococci supports diagnosis of pneumococcal pneumonia
• But remember, S. pneumonia is part of oropharyngeal microbiota in 20% of adults
• Its isolation from blood cultures is more specific but less sensitive than sputum culture; only 25-30% of patients have positive blood cultures
Test q: A 68F from a nursing home develops fever and shortness of breath. Percussion reveals dullness over the entire right lower lobe w/other lung fields normal. A sputum specimen reveals neutrophils and Gram-positive cocci. You suspect: Streptococcus pneumoniae.
Streptococcus pneumoniae: Sputum Gram Stain Streptococcus pneumoniae:
Big red objects in the background – neutrophils. Little objects stained dark – look like two little footballs w/pointed ends together = pneumococci.
Streptococcus pneumoniae: colonies
Blood agar plate, organisms more flat on the surface. Surface drops when they begin to autolyze – start to self-destruct. Show zone of partial hemolysis.
Gray hepatization (bottom portion) – consolidated, confluent appearance. Top right – congestion.
Infections with Haemophilus influenzae: • Respiratory
– Life threatening epiglottitis - submucosal inflammatory edema may obstruct airway < 24h after onset; generally children 2-4 yrs
– Otitis media, chronic bronchitis, bronchopneumonia – Pathology: dense fibrin-rich exudates of neutrophils
Acute Otitis Media: Fibrinosuppurative exudate on H&E: H. influenzae: Acute Purulent Bronchitis:
Bulging tympanic membrane – Bronchus disrupted by neutrophilic exudate -
acute OM – both Haemophilus sloughing of the mucosa
and S. Pneumoniae are common.
Diagnosis of H. influenzae Infection: H. influenzae: Gram-Negaitve Rods in Sputum: • Tiny gram-negative coccobacilli in gram-stained
smears of sputum or CSF
• Culture on chocolate agar - requires X & V factors for growth
• Rapid I.D. -- e.g., latex agglutination or DNA probe
Test q: A 28y/o immigrant from Haiti is seen in the Wishard ED for cough and fever. Sputum shows an encapsulated gram-negative rod. Hemin and NAD are required for growth. Diagnosis? Haemophilus influenzae.
Pseudomonas aeruginosa Infection: • P. aeruginosa necrotizing pneumonia • With necrotizing vasculitis
• Leads to necrosis in lungs – small cavities in lung tissue become necrotic w/lots of acute inflammation.
• Common in hospital-acquired pneumonia, especially in ICU patients and the immunocompromised. • Small, GNR. Produces complete hemolysis on blood agar – green colonies, smells fruity.
Get purple haze of bacteria on H&E –
