Emerging pathogens 2007
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Peter H. Gilligan PhD
Clinical Microbiology-Immunology Labs
UNC Hospitals
How I became a clinical microbiologist
• Obtained doctoral degree in microbiology at the University
of Kansas
• Did post-doctoral training (2 years) in medical and public
health microbiology at UNC Hospitals
• Director of Microbiology Labs at St Christopher’s Hospital
for Children (Philadelphia) for 4 years
• Past 20+ years, Associate Director then Director of the
Clinical Microbiology-Immunology Labs at UNC Hospitals
• Have served on medical school admission committee for
approximately 15 years and the MD/PhD advisory
(admissions) committee for the past 10 years
What do clinical microbiologists do?
• We serve:
» our patients
» our health care-providing colleagues, physicians,
nurses, physician assistants, pharmacy colleagues
» hospital administrators
• We make money for the institution
» general public by insuring the public health
• Involved in studying outbreaks of several emerging
infectious diseases
How do we serve?
• central role in the diagnosis and management of
infectious diseases
• central role in infection control and antimicrobial use
• recognize emerging disease threats and outbreaks
including bioterrorism events
• we educate & train health care providers
• we create new knowledge (research) to deal with
practical problems
Best things about my job
• Direct impact on patient care and public health of the
community
• Intellectually challenging job requiring a broad fund of
knowledge-need to know a little about a lot of things –I am
never bored!!!!!!!
• Work with highly motivated and intelligent individuals
• Get to be at the cutting edge of infectious disease
diagnosis
Worst things about my job
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Incredible amounts of governmental oversight
Increasing emphasis on financial aspects of the job
Declining talent pool of technologists
Need to be responsible for an organization that run
24/7/365-we never close. Personally have worked through
ice storms, blizzards, and hurricanes.
How you can become a clinical
microbiologist
• CLS programs available here, ECU, WCU, WSSU, Wake
Forest, UNC-CH
» Education is also available on line
• 2 more years of school to get a BS in CLS
• Take ASCP certification exam to become certified as a MT.
» Starting salary is 38,000 and up
» Career options are amazingly diverse; many former UNC
students work in leadership positions in the pharmaceutical
and biotech industries
Emerging/Re-emerging Infectious Diseases in the
past 25 years
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HIV*
Avian influenza
SARS*
Cryptosporidium*
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E. coli O157:H7*
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Nipah virus
nv Creutzfeldt-Jakob disease
Sin Nombre Virus
West Nile Virus
Clostridium difficle*
Bacillus anthracis (BT agent)
Cyclospora
CA-MRSA*
Rapidly growing mycobacterium*
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Rotavirus*
BK virus*
Chlamydia pneumoniae
Pencillinum marneffei
Legionella*
MDR- TB and pneumococcus*
Burkholderia cepacia complex*
VRE/VRSA*
Helicobacter pylori*
Invasive Group A streptococcal
disease*
HHV-6*
HPV*
HCV*
How do new pathogens emerge
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Organisms that jump species barriers
Changing ecosystems
Changes in food production techniques
Evolution of medical devices and care
» Long term survival of immunosuppressed
• Pathogens that are detected because of new
technology
• Misuse of micro-organisms
» Biocrime/bioterrorism
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Organism evolution as a result of human intervention
» Antibiotic pressure
How do microbes change?
• Bacteria, because they evolve very quickly, can readily
adapt to hostile environments
» Assume a generation time for a bacteria of 50 minutes
» 30 generations/day; or 220,000 bacterial generations for
each human generation (assume generation is 20 years)
» Bacteria have a huge evolutionary advantage over humans
How emerging pathogens develop?
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Mutation drives evolution
» constantly occurring
» usually silent or lethal
» environmental pressure such as antibiotics may select
“resistance” mutation
• Key feature of success of antibiotic resistant strains is their
genetic fitness I.e. their ability to compete in a complex microbial
environment
» Recognition that certain bacteria may be hypermutators
because of mutation in DNA repair genes
• These strains may not be as “fit” as wild-types but may
predominant in certain chronic infections such as
P.
aeruginosa causing chronic pulmonary infections in CF patients
How do emerging pathogens develop?
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Recombination
» Resistance genes from antibiotic producing
organisms
» genetic exchange of resistant genes can occur
among organisms which are genetically
diverse
• Think Cholera toxin genes to E. coli
» transfer of resistance/virulence genes can be
mediated by plasmids/phage/transposons/
integrons
Organisms that jump species barriers
• HIV, SARS, Avian flu
» HIV likely jumped from primates to humans
» SARS from pigs(?)
» Avian flu-hasn’t yet made the jump from birds to humans
because human to human spread is rare, if it occurs at all.
However mutation may result in that occurring.
» Technology allows us to quickly develop diagnostics for new
pathogens
• Took years to develop HIV diagnostics
• Took weeks to develop SARS diagnostics
Changing ecosystems
• Lyme disease
» A perfect storm
• Farmland in New England returned to forest
• Natural predators for deer were eliminated
• Deer populations and the ticks they carried increased
because of ecosystem changes
• People built homes and spent increasing amounts of time
in the woods
• This resulted in increased exposure to deer ticks that
carried Borrelia
» Ticks were pencil point in size and often difficult to see
Changes in food production techniques
• Increased use of factory farming
• Feedlots bring together large numbers of animals who
produce large amounts of waste
» Waste can lead to run-off of EHEC that can contaminant
adjacent fields as was seen in recent spinach outbreaks
• Large meat packing operations can result in 50 ton lots of
ground meat containing 100s of animals
» Meat can be distributed throughout the US
» Contaminated lots can then lead to large scale outbreaks
Changes in medical care
• Immunosuppression either as a result of HIV or medically
therapy (ex. transplants) results in emerging infections
» Pneumocystis, MAC, toxoplasma and CMV in HIV patients
» CMV, adenovirus and HHV-6 in transplant patients
• The use of indwelling artificial materials such as catheters,
shunts, artificial joints present new ecosystems and new
organisms
» Examples-coagulase negative staphylococci growing as a
biofilm on artificial joints/catheters/shunts
» Rapidly growing mycobacteria causing keratitis following
LASIK surgery
Pathogens detected with new technology
• Prime example is HCV
» Viral genome elucidated using molecular cloning techniques
• Broad range 16S RNA primers are used to detect noncultivable bacteria
• Next big thing- application of molecular tools to
understand how mixed microbial populations cause
disease
» Likely diseases caused by mixed microbial populations are
bacterial vaginosis, peridontal disease, inflammatory bowel
disease, CF lung disease
How does bacterial resistance
develop?
• Bacterial resistance develops in response to antimicrobial
pressure
» It is estimated that 3 million lbs of antimicrobials are used
each year in the US
• Much of it is used in children to treat viral respiratory
illness
• Estimated that 3/4 of children in US younger than two
receive antimicrobials
• Children then may serve as a key role for the emergence of
antimicrobial resistance
» 10x that amount are used in animals
» End result- tremendous selective pressure that results in the
emergence of bacterial resistance
UNC-ED
• 6% of wounds from ED in 1st quarter of 2005 grew MRSA
• 45% of wounds from ED in 2nd quarter of 2005 grew
MRSA
• ? Due to proliferation of CA-MRSA?
• GOAL
» To characterize and determine the prevalence of CA-MRSA
isolates at UNC hospitals
Molecular analysis: CA- vs. HA-MRSA
Adapted from Weber, CID, 2005:41S
Virulence of CA-MRSA
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Panton-Valentine leukocidin (PVL)
» Hemolysin first reported in 1932 by Panton and Valentine
» Located on mobile phage
» 2 co-transcribed genes, lukS-PV and lukF-PV
» The two subunits form a hexameric pore-forming cytolytic toxin with
a high affinity for PMNs and macrophages
•PVL producing strains associated with skin and soft tissue
infections and necrotizing pneumonia
•Rarely associated with osteomyelitis, septicemia, or
endocarditis
•Rare HA-MRSA strains with PVL have similar clinical syndrome
•Usually only 2% of all S. aureus isolates produce PVL but
found in the majority of epidemic CA-MRSA strains
SCCmec types
(Staphylococcal chromosome cassette)
21-24
Adapted from Diederen and Kluytmans, JID, 2005
Susceptibility Patterns
tmp-smz
ery
vanc
gent
pen
clinda
ery
doxy
cefox
clinda
cefox
vanc
doxy
gent
tmp-smz
pen
HA-MRSA
CA-MRSA
93% are erythromycin resistant
16% clindamycin resistant
CA-MRSA Timeline
Prison and
jail populations
Children without
identifiable risk factors
Mid 1990s
Outbreak in Detroit
2/3 of patients were IVDU
2003
Late 1990s
1980
2000
1998 - Athletes/sports teams
1999 - Native Americans
Necrotizing pneumonia,
United States and Europe
IVDU=intravenous drug users
Naimi TS et al. JAMA. 2003;290:2976-2984.
Zetola N et al. Lancet Infect Dis. 2005;5:275-286.
Levine DP et al. Ann Intern Med. 1982;97:330-338.
CDC. Morb Mortal Wkly Rep. 2003;52:793-795.
Groom AV et al. JAMA. 2001;286:1201-1205.
Herold BC et al. JAMA. 1998;279:593-598.
CDC. Morb Mortal Wkly Rep. 2001;50:919-922.
Gillet Y et al. Lancet. 2002;359:753-759.
CDC. Morb Mortal Wkly Rep. 1999;48:707-710.
Clinical presentation CA-MRSA
• CA-MRSA
» SSTIs (abscesses, cellulitis, folliculitis, impetigo,
furunculosis*)
• Typically treated with excision and drainage; +/- oral antibiotics
• Occasionally require IV antibiotics, hospitalization and surgical
intervention
» Necrotizing pneumonia especially in young people
secondary to influenza was reported this flu season
• Mortality was over 50%- median time to death 3.5 days
• Median age was 17.5 years
• 5 isolates from Louisiana were CA-MRSA genotype of the
same PFGE type
• Both levofloxacin and inducible clindamycin resistance seen in
these isolates
Case 5
• The patient is a 16 yo who presents with shoulder and left
chest wall pain
• An MRI is ordered because of concerns about a abscess
• The patient becomes hypotensive, SOB, is intubated and
admitted to the MICU.
• Prior to admission, he denied fever, chills, cough and night
sweats
• He lives on a farm in rural central NC with exposures to
dogs cats and horses
» In the past year a horse had been put done due to
“strangles.” Strangles is a respiratory infection caused by
Streptococcus equi
Case 5
• No contributory travel or sexual history. Does not use
drugs or alcohol
• Two months previously he had a right-sided preauricular
abscess incised and drained
» Treated with Augmentin and infection resolved
• On PE, afebrile, pulse was 103 bpm, RR 30 and BP 99/62
• Skin examination was significant for a small violaceous
lesion at the site of the prior abscess
• Had several pustules on his leg and a hyperpigmented
macules on his left great toe
• LDH was highly elevated, he was anemic and had a sed
rate of 60
Gram stain of sputum
Culture from blood bottle
Study Design
I.
II.
Outpatient wound cultures (SSTIs) with MRSA (6/05 to 3/06), n=233
Nosocomial MRSA isolates (blood) (6/05 to 4/06), n=76
III. Wound cultures with MRSA regardless of location (6/06-7/06), n=100
IV. Respiratory cultures with MRSA from Cystic Fibrosis (CF) patients
(10/05 to 4/07), n=339
V. Child care centers
VI. All isolates recovered at Lilongwe Medical Center (6-06-2-07) n>100
Definition of CA-MRSA
Panton-Valentine leukocidin positive
SCCmec type IV
I. PVL and SCCmec Characterization
of outpatient wound isolates
180
168
SCCmec typing**
160
140
IV
72%
II
500
120
100
SCCmec II
80
SCCmec IV
60
SCCmec undetermined
40
26
22
20
0.5%
9%
1
0
PVL positive
(n= 191)
15
11%
6%
PVL negative
(n= 42)
** Oliveira and Lencastre (2002) Antimicrob Agents Chemother 46, 2155-61.
0.5%
1
n=233
Study Design
I.
II.
Outpatient wound cultures (SSTIs) with MRSA (6/05 to 3/06), n=233
Nosocomial MRSA isolates (blood) (6/05 to 4/06), n=76
III. Wound cultures with MRSA regardless of location (6/06-7/06), n=100
IV. Respiratory cultures with MRSA from Cystic Fibrosis (CF) patients
(10/05 to 4/07), n=339
V. Child care centers
VI. All isolates recovered at Lilongwe Medical center (in June 07)
Definition of CA-MRSA
Panton-Valentine leukocidin positive
SCCmec type IV
II. PVL and SCCmec Characterization
nosocomial blood isolates
# of isolates
45
42
40
SCCmec II
35
SCCmec IV
SCCmec typing
IV
SCCmec undetermined
II
500
30
25
55%
20
16
15
11
10
7
21%
15%
5
0
0
0
PVL positive
(n= 16)
PVL negative
(n= 60)
9%
n=76
II. Clinical and Molecular Analysis
nosocomial blood isolates
Clinical Characterization
N=76
CA
HA
I
CA
1
14
1
16
HA
0
41
1
42
I
1
17
0
18
2
72
2
Study Design
I.
II.
Outpatient wound cultures (SSTIs) with MRSA (6/05 to 3/06), n=233
Nosocomial MRSA isolates (blood) (6/05 to 4/06), n=76
III. Wound cultures with MRSA regardless of location (6/06-7/06), n=100
IV. Respiratory cultures with MRSA from Cystic Fibrosis (CF) patients
(10/05 to 4/07), n=339
V. Child care centers
VI. All isolates recovered at Lilongwe Medical center (in June 07)
Definition of CA-MRSA
Panton-Valentine leukocidin positive
SCCmec type IV
III. PVL and SCCmec Characterization
of 2006 wound isolates
SCCmec typing
80
72
IV
70
II
500
# of isolates
60
50
72%
40
SCCmec II
30
SCCmec IV
indeterminate
20
10
9
9%
0
0
PVL positive
(n= 81)
10
8
10%
8%
1
PVL negative
(n= 19)
n=100
• Thanks to:
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Melissa Miller
Jennifer Goodrich
Joel Wedd
Mwai Makoka and the UNC project Lilongwe
Tameaka Sutton-Shields
Kyle Rodino
All the CMIL technologists who identify, save and
freeze isolates so we can do this research
As Brian the scientist would say, “Any
Questions?”