Gregory C' Gray, MD, MPH,1 Troy McCarthy,1 Mark LeBeck,1 Dean D' Erdman, DrPH,2

1
Gregory C. Gray, MD, MPH,1 Troy McCarthy,1 Mark
LeBeck,1 Dean D. Erdman, DrPH,2 David P.
Schnurr, PhD,3 Kevin L. Russell, MD, MTMH,4
Adriana E. Kajon, PhD,5 Jeffrey D. Dawson,
ScD,6 Gary V. Doern, PhD,7 Leta K.
Crawford-Miksza, PhD, MPH8 for the US National
Adenovirus Surveillance Team
Background - The epidemiology of human adenovirus
infections is changing. New, possibly more
virulent, strains have emerged and caused
epidemics. We sought to epidemiologically study
US clinical adenovirus isolates and to determine
risk factors for adenovirus disease. Methods -
From July 2004 through March 2005, adenovirus
viral specimens and corresponding clinical data
were collected from 23 US sites (8 military).
Adenoviruses were typed using PCR (hexon gene)
and sequencing protocols developed by the Centers
for Disease Control and Prevention. Sequences
were compared to those available from GenBank and
to in-house sequence data from the 51 recognized
human prototype strains. Identification of
adenovirus field isolates was based on achieving
identification scores 90 (both datasets) for a
given prototypic strain. A blinded
systematic-derived sample of 101 adenovirus field
isolates was studied by a collaborating
laboratory using established serotyping
techniques. Results - Of 1259 adenovirus
isolates examined (411 military and 848
civilian), 99.7 were successfully typed using
the PCR and sequencing method. Among military
specimens, adenovirus serotypes 4 (96.6),
21(1.2), and 3 (2.2) were the most prevalent.
Adenoviruses obtained from civilians were more
diverse adenovirus serotype 1 (15.9), 2
(22.8), 3 (39.9), 4 (5.1), 5 (6.1), 6 (0.6),
7 (1.1), 11 (0.5), 12 (0.8), 14 (0.4), 19
(0.2), 21 (1.8), 22 (0.2), 25 (0.1), 31
(0.9), 34 (0.7), 35 (0.7), 41 (2.0) and 45
(0.1). Adenoviruses from civilian subjects were
often associated with hospitalization (43) or
intensive care unit stay (6). The blinded
serotyping validation study demonstrated 100
agreement among the 98 isolates that were typable
by serum neutralization. Conclusion - The hexon
gene sequencing approach appears to be a very
useful method in typing human adenovirus and it
may eventually supplant traditional serotyping
methods. However, alternative sequence typing
gene targets are needed to supplement the hexon
gene approach in order to identify novel
recombinant viruses. (Revised)

• Of 694 adenovirus isolates examined thus far, 93
  were successfully typed using gene sequencing.
• Type 4 was the most prevalent among military
  populations.
• Type 3 was the most prevalent among civilian
  populations.
• Adenoviruses from civilian subjects were often
  associated with hospitalization (43) or
  intensive care unit stay (6).
• Gene typing may be useful in investigating
  nosocomial infections and in determining
  antiviral therapy (Antivir Ther 200510225-9).
• The hexon gene sequencing approach appears to be
  a very useful method in typing human adenovirus
  and it may eventually supplant traditional
  serotyping methods.

1Center for Emerging Infectious Diseases, Dept.
of Epidemiology, University of Iowa College of
Public Health, Iowa City, IA 2Respiratory Virus
Diagnostics Program, CDC/NCID/DVRD/REVB, Atlanta,
GA 3Viral and Rickettsial Disease Lab., Div. of
Communicable Disease Control, California Dept. of
Health Services, Richmond, CA 4Respiratory
Disease Laboratory, DoD Center for Deployment
Health Research, Naval Health Research Center,
San Diego, CA
5Lovelace Respiratory Research Institute,
Albuquerque, NM 6Department of Biostatistics,
University of Iowa College of Public Health, Iowa
City, IA 7Department of Pathology, University of
Iowa College of Medicine, Iowa City, IA 8Food and
Drug Laboratory Branch, California Department of
Health Services, Richmond, CA
Military The military samples came from 8
different bases in the United States. Samples
were from military personnel ranging from age 17
to 34 years old with a mean age of 20.1 2.4
years old and 88 were from males. Neither age
(p0.672) nor gender (p0.594) are predictors of
adenovirus infection in military personnel. Of
the 291 samples sequenced so far, Ad4 (n283) is
the most prevalent. Ad4, Ad3, and Ad21 were the
only other types found. There was no association
of specific adenovirus types with specific
military bases.

• 23 viral labs linked across the United States
  submitting all adenoviral isolates x 3 years

• All specimens were typed using a DNA hexon gene
  sequencing procedure that highly correlates with
  the more tedious classical serological typing
  systems.

Coinvestigators - Dean D. Erdman, DrPH David P.
Schnurr, PhD Kevin L. Russell, MD, MTMH
Adriana E. Kajon, PhD Jeffrey D. Dawson, ScD
Gary V. Doern, PhD Leta K. Crawford-Miksza,
PhD, MPH Site principal investigators - James
Chappell, MD, PhD, Nashville, TN Gail J.
Demmler, MD, Houston, TX Christine C. Ginocchio,
PhD, Manhasset, NY Diane C. Halstead, PhD,
DABMM, FAAM, Jacksonville, FL Nancy G. Henshaw,
PhD, MPH, Durham, NC Sue C. Kehl, PhD,
Milwaukee, WI Deanna L. Kiska, PhD, DABMM,
Syracuse, NY Marie L. Landry, MD, New Haven, CT
Diane S. Leland, PhD, Indianapolis, IN Melissa
Miller, MD, Chapel Hill, NC Christine Robinson,
PhD, Denver, CO Michael A. Saubolle, PhD, Tempe,
AZ Rangaraj Selvarangan, BVSc, PhD, Kansas City,
MO Gregory A. Storch, MD, St. Louis, MO and
Danielle Zerr, MD, MPH, Seattle, WA Collaborators
- Howard Lehmkuhl, PhD, Kevin Knudson, PhDCEID
Staff - Whitney Baker, Ana Capuano, Mark Lebeck,
Ghazi Kayali, Troy McCarthy, Sharon
Setterquist Funding NIH/NIAID R01 AI053034
Civilian Civilian isolates came from 15 sites.
Civilian adenovirus patients ranged in age from
8 days to 78 years, with a mean age of 6.9 12.1
yrs. 59 were from males. Age and gender were
not predictors of specific adenovirus types.
43.4 of the civilian samples were adenovirus
types 3, 4, 7 or 21 with Ad3 being most
prevalent. Other types observed were Ad1, Ad11,
Ad12, Ad14, Ad19, Ad22, Ad 31, Ad41, Ad5, and
Ad6. No specific strains were strongly
associated with hospitalization or ICU admission.

• The wild type sequences were compared to the CEID
  adenovirus sequence library that we created by
  sequencing 51 prototypic adenovirus isolates.

• Sites sent adenovirus positive specimens with
  associated data on supplied forms to be typed.

Validations A panel of 101 blinded specimens was
sent to CAs Viral and Rickettsial Disease
Laboratory. 98 were typed by classical
serotyping method with 100 agreement with UI
hexon gene sequencing method The other 3
specimens were sent to the Navy Respiratory
Disease Laboratory for a PCR typing algorithm
comparison where they found 100 agreement with
UI hexon gene sequence method.

• Why Study Human Adenovirus?
• Recognition of emergent more virulent adenovirus
  strains
• Recognition of adenovirus associations with
  numerous chronic diseases
• Recognition of adenovirus as a major cause of
  morbidity among transplant patients
• Recognition that some species of adenovirus may
  be responsive to antiviral therapy

• Clinical data collected from populations at high
  risk of adv infection
• children lt7 years of age
• allogeneic stem cell or solid organ
• transplant patients