Pneumonia and Influenza

8/12/2019 Pneumonia and Influenza

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Diagnostic Testing for

Community-Acquired

Pneumonia (CAP) and

Influenza

Norman Moore, Ph.D.

Director of Clinical Affairs

[email protected]


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Objectives

Discuss the etiological agents for pneumonia and which

age groups are most prone to the infection.

Describe what clinical samples should be taken and

how they should be transported to the laboratory for

analysis

State the diagnostic testing methods recommended for

community-acquired pneumoniaand influenza

Show how influenza can lead to pneumonia


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Infectious Disease in the US

1970: William Stewart, the Surgeon General of the United Statesdeclared the U.S. was ready to close the book on infectiousdisease as a major health threat; modern antibiotics, vaccination,and sanitation methods had done the job.

1995: Infectious disease had again become the third leading causeof death, and its incidence is still growing!

Pneumonia is the sixth leading cause of death in the US and

the major cause of death from infectious disease in the US.


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Current Number of Pneumonia Cases (US)

37 million ambulatory care visits per year for acute respiratoryinfections (physician and ER visits combined)

Community-Acquired Pneumonia (CAP)

Each year 2- 3 million cases of CAP result in ~ 10 million physicianvisits & 500,000 hospitalizations in the US

Average mortality is 10-25% in hospitalized patients with CAP

Nosocomial Pneumonia

Standard definition: onset of symptoms occurs approx 3 days afteradmission

250,000 - 350,000 cases of nosocomial pneumonia per year

25 - 50% mortality rate


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Etiological Agents

Newborns (0 to 30 days)

Group B Streptococcus,Lysteria monocytogenes,

or Gram negative rods are commonRSV in premature babies

Infants and toddlers

90% of lower respiratory tract infections are viral

with the most common being RSV, Influenza

A&B, and parainfluenza. Bacterial infections are

rare, but could be S. pneumoniae, Hib, or S.

aureus.


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Etiological Agents

Outpatient

S. pneumoniae,H. influenzae,M. pneumoniae, C.

pneumoniae, and respiratory viruses Inpatient (non-ICU)

With the above agents, addL. pneumophila

Inpatient (ICU)

S. pneumoniae, S. aureus,L. pneumophila, Gram-

negative bacteria, andH. influenzae


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Streptococcus pneumoniae

TypesOver 90 serotypes exist, with 88% of disease

covered in the 23-valent vaccine

Incidence100,000 to 135,000 cases of pneumonia

requiring hospitalization up to the year 2000

Around 80% of CAP

Cases are dropping due to the S. pneumoniaevaccine

TransmissionPerson to person

Risk groupsThe young and elderly

Most common identificationBlood culture and sputum

culture


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Haemophilus influenzae

TypesThe original risk was H. influenzaeType B

(Hib), but vaccine has dramatically reduced pneumonia

due to Hib, but other types and non-typeable strains still

cause disease

IncidenceVariable


Risk groupsThe young and elderly

Most common identificationBlood culture and sputum

culture


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Chlamydia pneumoniae

IncidenceOverall is unknown, but in the literature, itseems to go in cycles so high incidence in some yearsand low in others.

Can be considered 3rd

most common etiological agent in respiratorytract infections of young adults behindMycoplasma pneumoniaeandinfluenza


Risk groupsAll age groups, but more common in

school-age children. Most common identificationSerology

Personal contact with Barry FieldsChief of Respiratoryinfections from CDCrates of C. pneumoniaehave

been extremely low for years and he currently doesntview this as a significant infection.


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Mycoplasma pneumoniae

IncidenceEstimated 2 million cases and 100,000

pneumonia related hospitalizations in US

TransmissionPerson to person by respiratory

secretions, usually close contact

Outbreaks in crowded conditions like military and college which can

last several months

Risk groupsAll age groups, but more common in

school-age children and young adults. Most common etiological agent for adults younger than 30

Most common identification - Serology


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Legionella pneumophila

IncidenceEstimates vary greatly from 15,000 per year

to 100,000 per year in US

TransmissionContaminated water

Outbreaks in hospitals, ships, hotels, etc.

Risk groupsUsually elderly, smokers

Most common identificationUrinary antigen


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Viral pneumonia

Adults may get viral pneumonia by influenza,

adenovirus, cytomegalovirus, parainfluenza, varicella,

rubeola, or respiratory syncytial virus, particularly during

epidemics

Viral pneumonia, especially influenza, may cause

a secondary bacterial disease, such as

pneumococcal pneumonia


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Influenza A&B

Impact of influenza in the US

Hospitalizations up from 114,000 to 226,000

36,000 deaths annually

Influenza target population: 188MM in US

5-20% of US population affected by influenza each year

Most deaths affect elderly and young children

Also affects otherwise healthy individuals


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Influenza Treatment

Antiviral drugs are available

Must be administered within 48 hr of onset of symptoms

Generally reduce duration of symptoms by one day

First generation drugs (amantidine, rimantidine) are

cheaper but only treat influenza A

Second generation drugs (Tamiflu, Relenza) are more

expensive but treat both influenza A and B

Reason to differentiate between influenza A and B


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Respiratory Syncytial Virus

Almost all children with have RVS by their second

birthday

25% to 40% will have signs or symptoms of

bronchiolitis or pneumonia

0.5% to 2% will require hospitalization

Recovery is in 1 to 2 weeks, but they can spread

virus for 1 to 3 weeks The elderly can get a usually mild RSV infection due to

a weakened immune system

Rapid tests are not recommended on this

population


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Specimen Collection


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Swab collection

Swab should remain moist and cultured within 4

hours

If longer than 4 hours to get to culturing, shoulduse transport medium

Refrigeration, not frozen


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Sputum Collection

Quality of specimen

Care should be taken in collection since a lower respiratory tract

sample can be contaminated with upper unless collected by an

invasive technique Collection

Patient is instructed to give a deep coughed specimen

Put into sterile container, trying to minimize saliva

Transport to lab immediately Patient unable to give specimen can be given an aerosol-

induced specimen


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Blood culture

Usually done with fever spike

Standard is to take two sets of blood cultures one hour

apart


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Urine

Urine can be used for Legionellaand Streptococcus

pneumoniae

Antigen test

Noninvasive sample

Does not need to be qualified like a sputum

sample


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Influenza Sample Collection

Appropriate specimens

Nasal wash/aspirate, nasopharyngeal swab, or nasal swab

Throat swabs have dramatically reduced sensitivity

Samples should be collected within first 24 to 48 hours

of symptoms since that is when viral titers are highest

and antiviral therapy is effective

Testing can be done immediately with rapids or sample

placed in transport media

Infectivity is maintained up to 5 days when stored @ 4-

8C

If the sample cannot be evaluated in this time period, the

sample should be frozen @ -70C.


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Diagnostic Methods Available


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Infectious Disease Society of America/American Thoracic

Society Consensus Guidelines on the Management ofCommunity-Acquired Pneumonia in Adults (2007)

Diagnostic Testing

Suggestive clinical features combined with a chest radiograph or other

imaging technique is required for the diagnosis of pneumonia

It is recommended that patients with CAP should be investigated for

specific pathogens that would significantly alter standard (empirical)

management decisions, when the presence of such pathogens is

suspected on the basis of clinical and epidemiologic clues.


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Infectious Disease Society of America/American Thoracic

Society CAP Guidelines 2007

When to apply diagnostic tests

Optional for outpatients with CAP

Blood culture and sputum culture for inpatientswith productive cough*

All adult patients with severe CAP, should have

blood culture, sputum culture,Legionellaurinary

antigen and S. pneumoniaeurinary antigen tests*


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Common Diagnostic Tests Gram stain

Sputum culture

Blood culture

Latex agglutination assays

DFA/IFA

PCR

Serology Urinary antigen


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Gram stain

Apply sample to microscope slide

Apply stains & view using standard microscope

Pros: Inexpensive

Rapid (~15 minutes)

Cons: Difficult to get good sample (50% are inadequate)Should have less than 10 squamous epithelial cells

per low power field (100x)

Requires trained personnel to read


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Sputum CultureBacterial Culture

Pros: Inexpensive

Standard media for mostSheep blood agar, MacConkeyagar, and chocolate agar, BCYE for Legionella

Allows for antibiotic susceptibility testing

Cons: Requires live bacteriaantibiotics can affect resultsDifficult to get good sample

Requires dedicated tech time / experienced personnel

Results take 24 hours to >1 week


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LegionellaCulture

Legionel la

Legionellaneeds specific growth conditions

Buffered charcoal yeast extract (BCYE) plate

Clinical sample may need to be acid treated to reduce generalmicroflora

May take 3 to 10 days to get result


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Cell culture for Chlamydia pneumoniae

Chlamydia cultures should be transported in 2-sucrose

phosphate or other transport medium

Use HeLa cell line rather than McCoy that is used for C.

trachomitis

May take 3 to 10 days and is labor-intensive


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Culture for Mycoplasma pneumoniae

Specialty media

May take over 3 weeks for result

Vial is inspected daily and is prone to contamination (usually

indicated by color shift in first 5 days)

Needs subculturing to agar

Highly labor intensive


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Blood Culture

Pros: Inexpensive

Allows for antibiotic susceptibility testing

High specificity

Cons: Requires live bacteriaantibiotics can affect resultsRequires dedicated tech time / experienced personnel

Results take 24 hours to >1 week

Many bacterial infections dont

progress to bacteremia


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Latex Agglutination

Detecting antigen associated w/certain serogroups

Polystyrene latex particles coated with antibodies

Pros: Relatively simple

Rapid (~15 minutes)

Cons: Does not detect all serogroups of S. pneumoniae

Procedure associated with urine is cumbersome

Interpretation of results can be subjective


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Fluorescent Antibody (DFA/IFA)

Performed directly from sample on microscope slide

Sputum, pleural fluid, aspirated material, or tissue

Add fluorescent-tagged antibody specific for specific bacteria Observefor fluorescence using a special microscope

Pros: Relatively quick turn around time (~1 hour)

Cons: More labor intensive than rapid tests

Requires trained technologist and special microscope

Few labs equipped to perform DFA on

2nd/3rdshifts

Sensitivity can be poor (25% to 75%

on Legionella)


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Polymerase Chain Reaction (PCR)

Molecular technique using a clinical sample

Extract and amplify nucleic acid (DNA or RNA) of specific pathogen

Pros: Extremely sensitivecan detect one microorganism

Detects both live and dead pathogens

Cons: Requires highly trained technologist, expensive equipment

More labor intensive than rapid tests

Prone to cross-contamination (false positives)


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Serology

Chlamydia pneumoniae

Measurement usually of acute and convalescent serum

A four-fold rise in titer is considered diagnostic

A single IgM titer of 16 or greater or IgG of 512 or greater isconsidered suggestive of recent infection

Mycoplasma pneumoniae

A fourfold rise from acute to convalescent serum or complement

fixation titer of 1:128 in single serum specimen


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Urinary antigen

Tests are available for S. pneumoniaeand L.

pneumophila serogroup1

With Legionella, antigen appears in the urine 1 to 3

days after infection

Noninvasive sample

Easy-to-use


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Test Procedure for Urinary Antigen

Collect urine sample (no pre-treatment i.e. concentrating, boiling, filtering,etc.)

Open device pouch and lay flat

Dip provided sampling swab into urine

Place swab in lower hole of swab well and push up Add required number of drops of Reagent A (2 drops for Legionellatest

and 3 for S. pneumoniae)

Close device

Wait 15 minutes

Interpret results


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Diagnostic Methods for Influenza

Culture

DFA

PCR

Rapid Tests


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Viral Culture

Pro

Highly sensitive as long as sample is properly

handled Con

Cant give same day result to help monitor

therapy

High level of difficult/equipment


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DFA

Pro

Usually considered to have high level of

sensitivity

Can usually test for other respiratory pathogens at

the same time

Results can be achieved in same day

Con

Labor intensive needed experienced users

Turn-around time from lab usually takes many

hours


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PCR

Pro

For respiratory specimens, high performance

Same day results Con

Turn around time from lab is extensive, especially

if batching specimens

Expensive

Requires experienced technicians, labs, dedicated

equipment, etc.


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Rapid Tests

Pro

Tests take minimal time

Some tests are so simple that they can be CLIA-waived

Can be used to triage patients

Positive results can be used to rule out other

issues like pneumonia so dont give unnecessarychest x-ray, antibiotics, etc.

Con

Performance is not as good as culture, PCR, and

DFA


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The Connection BetweenInfluenza and S. pneumoniae


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Pandemic outbreaks

In 1957 and 1968 influenza pandemic outbreaks, it was

shown that a bacterial agent was present in

approximately 70% of the serious (life-threatening or

death) cases. In contrast, in non-pandemic years, only 25% of serious

cases had a secondary bacterial infection.


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Synergy Between Influenza and S. pneumoniae

Influenza neuraminidase found to prime lung for S.

pneumoniaeinvasion.

S. pneumoniaehas its own neuraminidase that it

uses to promote binding to cells.

In a mouse model, if neuraminidase inhibitors

were added, then mortality went down.

Recombinant versions of influenza strains of past 50years were made.

1957 and 1997 pandemic strains that were related

to bacterial pneumonia had highest levels of

neuraminidase activity.


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S. pneumoniaeand Penicillin


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Penicillin Breakpoint

IV Penicillin

Less expensive than broad spectrum antibiotics

Reduce broad spectrum antibiotic resistance

Less liver/kidney resistance

Minimum Inhibitory Concentration (MIC) (mcg/mL)

Susceptible (S) Intermediate (I) Resistant (R)

Updated 2 4 8

Previous 0.06 0.12-1.0 2


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Reference

Mandell, L.A., R.G. Wunderink, A. Anzueto, J.G.Bartlett, G.D. Campbell, N.C. Dean, S.F. Dowell, T.M.File, D.M. Musher, M.S. Niederman, A. Torres, andC.G. Whitney. Infectious Diseases Society of

America/American Thoracic Society ConsensusGuidelines on the Management of Community-AcquiredPneumonia in Adults. Clinical Infectious Diseases.2007; 44:S27-72.

Murray, P.R., E.J. Baron, J.H. Jorgensen, M.A. Pfaller,and R.H. Yolken. Manual of Clinical Microbiology 8thEdition.

Forbes, B.A., D.F. Sahm, and A.S. Weissfeld. Bailey &Scotts Diagnostic Microbiology 12thEdition.

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Pneumonia and Influenza