Journal of General Virology (2012), 93, 2457–2465
Short
Communication
DOI 10.1099/vir.0.042028-0
Genome sequence of a waterfowl aviadenovirus,
goose adenovirus 4
Győző L. Kaján,1 Andrew J. Davison,2 Vilmos Palya,3 Balázs Harrach1
and Mária Benkő1
1
Correspondence
Institute for Veterinary Medical Research, Centre for Agricultural Research, Hungarian Academy of
Sciences, PO Box 18, H-1581 Budapest, Hungary
Győző L. Kaján
[email protected]
2
MRC – University of Glasgow Centre for Virus Research, 8 Church Street, Glasgow, G11 5JR, UK
3
CEVA-Phylaxia Inc., Szállás u. 5, H-1107 Budapest, Hungary
Received 11 May 2012
Accepted 13 August 2012
We present, to our knowledge, the first complete genome sequence of a waterfowl aviadenovirus,
goose adenovirus (GoAdV) strain P29, and an analysis of its genetic content in comparison with
five published aviadenovirus genome sequences. Of the 35 genes predicted to encode functional
proteins, the central region of the genome contains 19 (IVa2 to fiber-2) that were inherited from
the ancestor of all known adenoviruses. Of the remaining genes, nine have orthologues only in
aviadenoviruses and seven lack orthologues in any adenovirus. We also obtained limited
sequence data for a pathogenic GoAdV strain D1036/08. Phylogenetic analyses placed the two
GoAdV strains monophyletically in the genus Aviadenovirus. We propose designating strains P29
and D1036/08 as GoAdV-4 and GoAdV-5, respectively.
Adenoviruses are non-enveloped viruses with linear
dsDNA genomes that infect members of every class of
vertebrates from fish to mammals. The family Adenoviridae
contains the genera Atadenovirus, Aviadenovirus, Ichtadenovirus, Mastadenovirus and Siadenovirus (Harrach
et al., 2011). Members of the genus Aviadenovirus (known
vernacularly as aviadenoviruses) infect only birds, and
birds also host some members of the genera Atadenovirus
and Siadenovirus. The genus Aviadenovirus consists of eight
officially accepted species (Harrach & Kaján, 2011; www.
ictvonline.org), each containing viral types that are denoted
numerically (e.g. fowl adenovirus 1 in the species Fowl
adenovirus A). Names of viral types may be abbreviated to a
prefix (falcon adenovirus, FaAdV; fowl adenovirus, FAdV;
goose adenovirus, GoAdV; and turkey adenovirus, TAdV)
followed by a number (e.g. FAdV-1). Grouping of types into
species is as follows: Falcon adenovirus A (FaAdV-1), Fowl
adenovirus A (FAdV-1), Fowl adenovirus B (FAdV-5), Fowl
adenovirus C (FAdV-4 and FAdV-10), Fowl adenovirus D
(FAdV-2, FAdV-3, FAdV-9 and FAdV-11), Fowl adenovirus
E (FAdV-6, FAdV-7, FAdV-8a and FAdV-8b), Goose
adenovirus A (GoAdV-1, GoAdV-2 and GoAdV-3) and
Turkey adenovirus B (TAdV-1).
The GenBank/EMBL/DDBJ accession numbers for the genome
sequence of GoAdV strain P29 and for the partial DNA polymerase
and the hexon gene sequences of GoAdV strain D1036/08 are
JF510462, JQ178216 and JQ178217, respectively.
One supplementary table is available with the online version of this
paper.
042028 G 2012 SGM
Several adenoviruses have been identified in waterfowl,
specifically ducks and geese. Duck adenovirus 1 (egg drop
syndrome virus) is also common in geese (Bartha et al.,
1982; Ivanics et al., 2001) and belongs to the genus
Atadenovirus (Harrach et al., 2011). In contrast, serological
comparisons and preliminary molecular typing have placed
GoAdVs in the genus Aviadenovirus (Csontos, 1967; Papp
et al., 2003). The first strains from faecal samples and
unhatched eggs were described in Hungary (Csontos, 1967)
and were serologically distinguishable from FAdVs. Zsák &
Kisary (1984) isolated and studied seven GoAdV strains
from the liver and gut of young goslings and grouped them
into three types (GoAdV-1, GoAdV-2 and GoAdV-3) based
on the restriction endonuclease cleavage pattern. These
types possess a common complement-binding antigen with
FAdVs, and were apathogenic in animal infection studies.
Also in Hungary, Ivanics et al. (2010) found pathogenic
GoAdV strains that caused hepatitis and hydropericardium
syndrome in young goslings. In Canada, inclusion body
hepatitis and mortality among young goslings were found
to be caused by an adenovirus (Riddell, 1984). In Germany,
antibodies against adenoviruses were identified in various
goose species (Hlinak et al., 1998; Kaleta et al., 1998).
Complete genome sequences are available for five galliform
poultry aviadenoviruses, each representing a different viral
species: FAdV-1 (GenBank accession no. U46933; Chiocca
et al., 1996), FAdV-4 (GU188428; Griffin & Nagy, 2011),
FAdV-8 (GU734104; Grgić et al., 2011), FAdV-9 (AF083975;
Ojkić & Nagy, 2000) and TAdV-1 (GU936707; Kaján et al.,
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2457
G. L. Kaján and others
2010). We now present the first full genome sequence of a
waterfowl aviadenovirus, GoAdV strain P29.
during passage in cell culture. The frameshift in TAdV-1
fiber-2 was previously authenticated (Kaján et al., 2010), and
that in TAdV-1 ORF19 was confirmed by inspection of the
sequence database. The genome annotations of the other
aviadenoviruses in Table 1 amount to substantial updates of
previously published versions.
GoAdV strain P29 was isolated by one of the authors (V.
Palya) in Hungary during the 1970s. Although it originated
from goslings that died young, no information is available
on its pathogenic properties. It was isolated and later
cultured afresh on goose embryo hepatocyte cells. When
the maximal cytopathic effect was observed, the cultures
were frozen and thawed three times. After low-speed
clarification, virions were concentrated from the supernatant by ultracentrifugation, and viral DNA was isolated by
phenol/chloroform extraction. Random inserts in a total of
750 plasmid clones of GoAdV DNA were sequenced on
both strands by capillary sequencing and assembled by
standard methods. Gaps and the genome ends were
amplified by PCR and sequenced by primer walking.
Genes encoding functional proteins were predicted by
standard bioinformatic approaches, which included considerations of genetic conservation.
The central region of the GoAdV genome contains 16
consecutive genes (IVa2 to pVIII) that are present in all
sequenced adenoviruses. This region also contains two
fiber genes (fiber-1 and fiber-2), of which one (the
paralogous arrangement makes it impossible to state
which) is conserved in all adenoviruses. FAdV-1, FAdV-4,
FAdV-10 and TAdV-1 also have two fiber genes (Chiocca
et al., 1996; Griffin & Nagy, 2011; Kaján et al., 2010; Marek
et al., 2012), whereas FAdV-8 and FAdV-9 have one (Grgić
et al., 2011; Ojkić & Nagy, 2000). Regardless of the number
of fiber genes, electron microscopic investigations have
revealed two fibers on each FAdV capsid vertex, these
being different in size in FAdV-1 but similar in members of
the other four species (Gelderblom & Maichle-Lauppe,
1982). The central region also contains the major late
promoter and the bipartite leader in positions equivalent to
those in other aviadenoviruses (Fig. 1; Kaján et al., 2010;
Payet et al., 1998; Sheppard et al., 1998). The U exon is
located between pVIII and fiber-1 on the complementary
strand. This exon is present in almost all adenoviruses,
presumably having been lost from some (Davison et al.,
2003). In members of the species Human adenovirus C
(genus Mastadenovirus), the U exon is spliced to two
consecutive exons located in one of the mRNA leaders of
the DNA-binding protein (DBP) gene and the DBPencoding region, so that the C-terminal region of the
protein is encoded in a different reading frame from the
overlapping DBP-coding region (Tollefson et al., 2007).
However, these downstream exons are poorly conserved in
the amino acid sequence even among human adenoviruses,
making bioinformatic identification of the GoAdV counterparts practically impossible.
The GoAdV strain P29 genome is 43 376 bp in size and
has a nucleotide composition of 44.7 mol% G+C, which
is the lowest among the known aviadenoviruses (FAdV-1,
54.3 mol%; FAdV-4, 54.6 mol%; FAdV-8, 57.9 mol%;
FAdV-9, 53.8 mol%; TAdV-1, 66.9 mol%). Partial sequences
originating from GoAdV strain D1036/08 (Ivanics et al.,
2010) have similar compositions to that of GoAdV strain
P29. The 39 bp inverted terminal repeat in the GoAdV strain
P29 genome is the smallest among the known aviadenoviruses. Fig. 1 shows the arrangement of predicted genes and
Table 1 summarizes a comparison with the five other fully
sequenced aviadenoviruses. Thirty-five GoAdV genes were
predicted, seven of which (ORF51–ORF56 and ORF19B)
lack detectable orthologues in other aviadenoviruses. The
analysis was complicated by the observation that various
protein-coding regions in all of the published sequences are
disrupted by apparent mutations. Some of these mutations
are located in essential genes and probably represent
sequencing errors, but others are not and may have occurred
1 1C
L
pX
pVII pVI
L
51 52 2
52K pIIIa
14 12 IVa2
pol
pTP
III
Protease
Hexon
33K
22K
100K
DBP
Fiber-1
pVIII
U
Fiber-2
22 20A 20 56 55 19 54
R1
19B
R2, R3
53
?
Fig. 1. Organization of predicted functional protein-encoding genes in GoAdV strain P29. The central grey line represents the
genome, with 5 kb scale markers in white and tandem reiterations (R1–R3) in black. Protein-encoding regions are shown as
open arrows. Those shaded dark grey were apparently inherited from the common ancestor of all known adenoviruses, with
fiber-1 of the two fiber paralogues assigned to this group. Those shaded light grey have orthologues only in other
aviadenoviruses, and those shaded white are unique to GoAdV. Gene nomenclature is shown, with the prefix omitted from
names commencing with ORF. Splicing between protein-encoding regions is indicated by diagonal lines. The two exons of the
bipartite late leader (L) are also indicated. DBP, DNA-binding protein; pol, DNA polymerase; L, exon of the bipartite late leader;
pTP, terminal protein precursor; R, reiteration; U, U exon.
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Journal of General Virology 93
http://vir.sgmjournals.org
Table 1. Locations of predicted functional protein-encoding regions in aviadenovirus genomes
Features were assigned using standard approaches to analyse ATG-initiated ORFs of ¢40 codons, taking into account amino acid sequence conservation, transcriptional orientation and the
presence of potential splice sites (Davison et al., 2003; Kaján et al., 2010).
ORF
FAdV-1
FAdV-4
ORF0
ORF51
R
R
519–743
ORF1
R
794–1330
790–1314
847–1338
837–1298
886–1449
ORF52
ORF1A
R
R
,1334–1480
,1315–1482
,1339–1497
,1325–1490D
,1450–1620
ORF1B
ORF1C
R
R
1483–1668
1816–1947
1572–1808
1500–1730
1735–1893
1493–1705
1713–1871
1625–1840
1803–1955
1791–1931
Protein ORF1B
Protein ORF1C
ORF2
R
1999–2829
1850–2665
1950–2753
1922–2728
2007–2816
2012–2854
Protein ORF2
ORF14C
L
ORF14B
L
ORF14A
L
ORF14
L
ORF13
L
ORF12
L
IVa2
L
2892–3527;
15 081–15 086
3549–4448;
15 081–15 086
4462–5385;
15 081–15 086
5366–6538
2667–3311;
16 952–16 957
3373–3981;
16 952–16 957
3997–4656;
16 952–16 957
4667–5314;
16 952–16 957
5330–6217;
16 952–16 957
6207–7118;
16 952–16 957
7090–8274
2837–3517;
16 276–16 281
3536–4222;
16 276–16 281
4261–5247;
16 276–16 281
5243–6160;
16 276–16 281
6129–7337
2901–3575;
16 758–16 763
3600–4278D;
16 758–16 763
4315–5259d;
16 758–16 763
5255–6184;
16 758–16 763
6208–7350
2857–3522;
16 474–16 482
3519–4397;
16 474–16 482
4466–5248;
16 474–16 482
5373–6314;
16 474–16 482
6295–7467
pol
pTP
L
L
52K
R
6501–10 268
10 269–12 155;
15 081–15 086
12 193–13 329
8258–12 016
12 021–13 913;
16 952–16 957
13 959–15 158
7334–11247D
11 244–13 184;
16 276–16 281
13 241–14 449
7347–11262D
11 259–13 562;
16 758–16 763
13 655–14 884
7469–11431
11 382–13 376;
16 474–16 482
13 445–14 671
pIIIa
R
13 316–15 043
15 145–16 917
14 436–16 211
14 871–16 636D
14 658–16 373
III
R
15 110–16 657
16 989–18 566
16 292–17 929
16 796–18 454
16 498–18 072
478–717
FAdV-9
FAdV-8
TAdV-1
575–808
566–799
548–622; 696–872
GoAdV
407–538;
602–724
795–1229
1226–1831
Protein
Protein ORF0
Protein ORF51
Deoxyuridine
triphosphatase
Protein ORF52
Protein ORF1A
Assigned tentatively
Possibly ORF2 family
Exon encoding N
terminus unidentified
Protein ORF14C
Contains hydrophobic
domain; assigned
tentatively to GoAdV
Contains parvovirus
NS1 (Rep) domain;
ORF2 family
ORF14 family
Protein ORF14B
ORF14 family
Protein ORF14A
ORF14 family
2866–3591;
Protein ORF14
14 110–14 115
Protein ORF13
3603–4514;
Protein ORF12
14 110–14 115
4471–5667
Encapsidation
protein IVa2
5633–9397
DNA polymerase
9400–11 205;
Terminal protein
14 110–14 115 precursor pTP
11 232–12 383
Encapsidation
protein 52K
12 367–14 094
Capsid protein
precursor pIIIa
14 131–15 717
Capsid protein III
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Comments
ORF14 family
ORF2 family
ORF2 family
Penton base
Goose adenovirus 4 genome
2459
Or.*
ORF
Or.*
FAdV-1
pVII
R
16 679–16 897
pX
R
pVI
Journal of General Virology 93
FAdV-9
FAdV-8
TAdV-1
GoAdV
Protein
18 575–18 808
17 971–18 202D
18 492–18 728
18 081–18 326
15 722–15 976
16 965–17 495
18 980–19 519
18 436–19 035
18 952–19 542
18 576–19 139
16 023–16 580
R
17 559–18 230
19 611–20 294
19 165–19 851
19 718–20 405D
19 290–19 970
16 649–17 329
hexon
protease
DBP
R
R
L
18 289–21 117
21 134–21 754
21 899–23 340;
23 455–23 662
20 340–23 153
23 171–23 800
23 921–25 359;
25 464–25 743
19 964–22 810
22 827–23 444
23 557–24 989;
25 087–25 324
20 531–23 374
23 405–24 034
24 142–25 655;
25 796–25 964
20 044–22 866
22 882–23 514
23 676–25 501;
25 668–26 112
17 371–20 172
20 186–20 806
20 877–22 273;
22 392–22 602
100K
R
23 680–26 634
25 775–28 940D
25 388–28 556D
26 022–29 262D
26 201–29 452
22 660–25 602
22K
R
26 324–26 878
28 558–29 145
28 237–28 781D
28 910–29 484D
29 079–29 723
25 205–25 798
33K
R
pVIII
R
26 324–26 654;
26 836–27 119
27 149–27 886
28 558–28 936;
29 124–29 434
29 460–30 203
28 237–28 603;
28 811–29 145
29 185–29 910
28 910–29 305D;
29 518–29 831
29 885–30 610
29 079–29 475;
29 675–30 024
30 086–30 829
25 205–25 640;
25 827–26 074
26 096–26 821
Core protein
precursor pVII
Core protein
precursor pX
Capsid protein
precursor pVI
Hexon
Protease
Single-stranded
DNA-binding
protein
Hexon assembly
protein 100K
Encapsidation
protein 22K
Protein 33K
U
L
,27 894–28 115
,30 218–30 436
,29 929–30 162
,30 631–30 864
,30 902–31 120
,26 828–27 073
fiber-1
R
28 114–30 495
30 435–31 733
30 161–31 876
30 863–32 434
31 135–32 424
27 097–28 377
Fiber-1
fiber-2
R
30 536–31 768
31 717–33 141
32 431–33 769D
28 367–29 749
Fiber-2
ORF22
ORF20A
L
L
L
33 194–33 781
33 784–34 200;
35 196–35 216
34 201–35 123D;
35 196–35 216
33 814–34 386
34 415–35 080;
36 239–36 259
35 162–36 118;
36 239–36 259
ORF56
L
29 741–30 352
30 355–31 131;
32 172–32 192
31 116–32 069;
32 172–32 192
32 263–32 745;
39 233–39 277
Protein ORF22
Protein ORF20A
ORF20
31 812–32 429
32 431–32 871;
33 886–33 906
32 892–33 809;
33 886–33 906
ORF55
L
Protein ORF55
ORF19
L
32 697–33 101;
39 233–39 277
33 092–35 005;
39 233–39 277
ORF54
L
34 238–36 201D;
36 349–36 396
FAdV-4
35 330–37 288;
37 356–37 373
31 930–32 502
32 506–32 985;
33 998–34 018
32 986–33 903;
33 998–34 018
34 220–36 346;
36 426–36 443
32 501–33 080D
33 084–33 644;
34 644–34 664
33 648–34 562;
34 644–34 664
34 726–37 095;
37 180–37 194
36 430–39 002D;
39 190–39 252
34 999–35 388;
39 233–39 277
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Capsid protein
precursor pVIII
Protein U
Comments
Capsid protein II
Exon encoding C terminus
unidentified
Capsid protein IV-1; fiber
family
Capsid protein IV-2; fiber
family
Contains hydrophobic
domains
Protein ORF20
Protein ORF56
Protein ORF19
Protein ORF54
Type I membrane protein;
contains Ig domain;
possibly ORF11 family
Type I membrane protein
Contains lipase domain;
contains signal peptide;
type I membrane protein
in most; ORF19 family
Type 1 membrane protein
G. L. Kaján and others
2460
Table 1. cont.
http://vir.sgmjournals.org
Table 1. cont.
ORF
Or.*
FAdV-1
FAdV-4
L
ORF43
ORF8
ORF17
ORF16
ORF9
R
R
L
L
R
37 391–38 239
38 717–39 256
39 286–39 705
40 082–41 002
ORF10
R
41 113–41 853
ORF11
R
41 958–42 315;
42 385–42 645;
42 720–42 973
ORF23
L
ORF26
R
ORF19A
R
42 029–42 034;
42 117–44 678
ORF4
ORF25
R
R
44 799–45 308
ORF53
L
37 716–38 381
39 245–40 060
40 680–41 132
41 149–41 556
37 859–38 668
40 596–41 066
FAdV-8
38 652–39 486D
39 682–40 152
TAdV-1
GoAdV
Protein
35 395–39 162;
39 233–39 277
Protein ORF19B
Protein ORF43
GAM-1
Protein ORF17
Protein ORF16
Protein ORF9
40 468–41 328
41 621–42 220
42 556–43 494
Protein ORF10
41 461–41 821;
41 899–42 398
40 513–41 194;
4179–41 445
42 660–43 595
41 782–42 715D
43 073–43 255;
43 327–43 436;
43 531–43 663
43 699–44 048;
44 118–44 619
Protein ORF11
Protein ORF23
44 791–44 832;
44 902–45 011;
45 078–45 216
Protein ORF26
Protein ORF19A
44 050–44 058;
44 137–44 637
Protein ORF4
Protein ORF25
43 147–43 152;
43 234–43 773
40 451–41 080
*Or., Orientation; L, leftward; R, rightward.
DDisrupted by a frameshift.
dDisrupted by an in-frame stop codon.
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Protein ORF53
Comments
Contains lipase domain;
type 1 membrane protein;
ORF19 family
Type 1 membrane protein;
contains Ig domain;
ORF11 family
Type 1 membrane protein;
contains Ig domain;
ORF11 family
Type 1 membrane protein;
contains Ig domain;
ORF11 family
Type 1 membrane protein;
contains Ig domain;
ORF11 family
Contains C-type lectin
domain; annotation may
be incomplete as
similarity extends
upstream
Contains lipase domain;
type 1 membrane protein;
ORF19 family
Contains US22 domain
Type 1 membrane protein
Goose adenovirus 4 genome
ORF19B
FAdV-9
G. L. Kaján and others
(a) Complete
hexon
human 5
human 12
bovine 1
957
Mast
bat 3
1000
1000
canine 1
murine 1
1000
snake 1
1000
At
bovine 4
998
duck 1
turkey 3
1000
Si
raptor 1
goose P29
1000
falcon 1
fowl 10
1000
fowl 4 Fowl C
Avi
997 fowl 9
1000
fowl 8
812
turkey 1
fowl 1
sturgeon 1 Icht
1000
(b) Partial pol
fowl 6
994 fowl 7
fowl 8a
fowl 8b
961
fowl 2
fowl 3
Fowl D
1000 fowl 9
fowl 11
fowl 5
1000 fowl 10
Fowl C
fowl 4
fowl 1
turkey 1
goose P29
goose D1036/08
Meyer’s parrot 1
turkey 3
raptor 1
959
818
fowl 9
876
970
fowl 3
1000
Si
bovine 4
duck 1
snake 1
murine 1
826
1000
Avi
1000
995
0.2
(c) Partial
hexon
Fowl E
fowl 11 Fowl D
1000
778
fowl 2
At
bat 3
canine 1
bovine 1
human 5
human 12
Mast
turkey 2
973
0.1
fowl 8a
1000
fowl 6
888
976
Fowl E
fowl 8b
991
fowl 7
Fowl B
fowl 5
Fowl A
fowl 1
turkey 1
Turkey B
1000
fowl 10
Fowl C
866
fowl 4
fowl 8
(d) Complete
penton base
1000
fowl 9
fowl 10
Fowl C
1000
fowl 4
984
psittacine 1
757
pigeon 1
falcon 1
falcon 1 Falcon A
goose P29
goose P29
1000
raptor 1
goose D1036/08
0.1
turkey 1
fowl 1
0.1
Fig. 2. Phylogenetic analyses of GoAdV sequences. Virus types are represented by host name and type or strain number.
Genus names are abbreviated (Avi, Aviadenovirus; At, Atadenovirus; Icht, Ichtadenovirus; Mast, Mastadenovirus; Si,
Siadenovirus), as are species names (e.g. Fowl A, Fowl adenovirus A). Tree topology was tested by bootstrapping (1000
datasets) and bootstrap values .750 are shown. The GenBank accession nos used are listed in Table S1 (available in JGV
Online). (a) Amino acid sequence of the complete hexon. The tree was rooted using white sturgeon adenovirus 1. (b) Partial
amino acid sequence of pol. The tree was rooted at the midpoint. (c) Partial DNA sequence of hexon. The tree was rooted at the
midpoint. (d) Amino acid sequence of the complete penton base. The tree was rooted using raptor adenovirus 1. The edited
alignment lengths used in (a), (b) and (d) were 833, 91 and 446 aa, respectively, and that used in (c) was 483 nt. Bars, amino
acid (a, b, d) or nucleotide (c) substitutions per site.
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Journal of General Virology 93
Goose adenovirus 4 genome
The regions towards the ends of the GoAdV genome are
more divergent than the central region, and contain genes
that are specific to aviadenoviruses or unique to GoAdV.
Genes in the latter category are inherently the most difficult
to predict. Towards the left genome end, all sequenced
aviadenoviruses have ORF1 and ORF2. ORF12 and ORF14
are predicted to be expressed by splicing from a short
protein-coding exon that is also part of the terminal
protein precursor (pTP) gene. Among aviadenoviruses,
ORF12 and ORF13 are related to ORF2 (Table 1; Corredor
et al., 2006; Davison et al., 2003; Washietl & Eisenhaber,
2003), and only the former is present in GoAdV. GoAdV
has a single member of the ORF14 gene family, and other
aviadenoviruses have up to four, which we have designated
the series ORF14, ORF14A, ORF14B and ORF14C in Table
1. The leftmost gene in most aviadenoviruses, ORF0, is
absent from GoAdV, and the unrelated ORF51 has been
assigned tentatively to this location. Between ORF1 and
ORF2, other aviadenoviruses have ORF1A (for which an
upstream protein-coding exon remains unidentified),
ORF1B and, except for FAdV-4, ORF1C. GoAdV lacks
ORF1A and ORF1B and instead has the unrelated ORF52,
which may encode an ORF2 family member (Fig. 1).
Towards the right genome end, all GoAdV genes are oriented
leftwards. This region also contains tandem reiterations R1–
R3. Sequence variations were noted just to the left of R2, with
variations at 6 nt that result in two different sequences. The
only convincing gene assigned to the region between R1 and
R2 is ORF53. The other genes fall into two groups (ORF19B,
ORF54, ORF19, ORF55 and ORF56; and ORF20 and
ORF20A), each expressed by splicing from a short proteinencoding exon, plus ORF22, which appears not to be spliced.
ORF19, ORF20, ORF20A and ORF22 are conserved in all
sequenced aviadenoviruses (Table 1), ORF19 encoding a
lipase-related protein (Davison et al., 2003; Ojkić & Nagy,
2000). GoAdV ORF19B and ORF19 are related, both
proteins containing a GXSXG consensus motif, which is
characteristic of enzymically active lipases (Corredor et al.,
2008; Schrag & Cygler, 1997). FAdV-4 also has a second gene
(ORF19A) encoding a lipase-like protein, although it is
oriented rightwards (Table 1). The remaining three ORFs in
this region of the GoAdV genome (ORF54, ORF55 and
ORF56) encode type I membrane proteins. The ORF56
protein contains an immunoglobulin-like domain and is
possibly a member of the ORF11 family, which is located
further to the right in other aviadenoviruses. ORF11 family
proteins may have immunomodulatory effects in the host
(Corredor et al., 2008; Davison et al., 2003; Le Goff et al.,
2005).
Several adenovirus proteins are processed by the viral
protease. In GoAdV, type I cleavage signals [(M/L/I/V/
F)XGG’X)] are present in pTP (numbering from the first
residue of the signal, at 301 and 306), pIIIa (the variant
NXGG’X at 184, as in every other aviadenovirus), pX (6 and
126), pVI (25) and pVIII (35, 110, 128 and 139). Type II
signals [(M/L/I/V/N/Q)X(A/G)X’G] are present in pTP (167
and 184), pIIIa (524, 561 and the variant FTGT’G at 506),
http://vir.sgmjournals.org
pVII (32 and 55), pX (101), pVI (209) and pVIII (160). A
variant (NRGW’G) of the type IIb signal (NTGW’G) is
present in pVII (10), as in most other aviadenoviruses. Type
III signals [(M/L/I)XAX’G] are present in pTP (the variant
SGAS’G at 187) and pIIIa (the variant VDAD’G at 262).
Only three of these 23 signals are not conserved in any other
aviadenovirus: in pTP (301) and pIIIa (524 and 561). The
pVIc cofactor, which is responsible for activation of the viral
protease (Mangel et al., 2003; Webster et al., 1993), is present
at the C terminus of the pVI protein. Its consensus in
aviadenoviruses is GV(A/T/Q)XXX(R/K)R(M/V)CY.
Phylogenetic analyses also incorporated data for GoAdV
strain D1036/08, which was isolated in Hungary and causes
hepatitis and hydropericardium syndrome in young goslings
(Ivanics et al., 2010). The partial sequence of the DNA
polymerase gene (pol) of this strain has been published
(Ivanics et al., 2010), and in addition, we determined the
partial hexon gene sequence via a PCR-based approach
(Meulemans et al., 2001). Phylogenetic calculations were
performed by using PHYLIP (Felsenstein, 1989). For amino
acid sequences, PROTDIST was used with the Categories
model, and for DNA sequences, DNADIST with the Kimura
two-parameters model. For tree reconstruction, FITCH was
used with the Fitch–Margoliash method with global
rearrangements, and trees were visualized by using MEGA5
(Tamura et al., 2011). Calculations were based on partial pol
(Kaján et al., 2011; Wellehan et al., 2004) and hexon
(Meulemans et al., 2001, 2004) sequences, which are more
suitable for analysis within genera, and on complete
sequences of hexon and penton base, which are more
suitable for analysis among genera. The trees showed that
GoAdV strain P29 groups with aviadenoviruses (Fig. 2). The
two GoAdV strains are monophyletic (Fig. 2b, c), having
identical partial pol sequences (Fig. 2b) but different hexon
sequences (Fig. 2c). Branching of the GoAdV strains from
other aviadenoviruses lies deep within the genus for the
complete (Fig. 2a, d) and partial (Fig. 2b, c) genes.
On the basis of host species, genome organization, gene
content, splice sites, protease cleavage signals and phylogeny,
GoAdV strain P29 belongs to the genus Aviadenovirus.
Restriction endonuclease patterns predicted for this strain
(not shown) are sufficiently different from those reported
for GoAdV-1, GoAdV-2 and GoAdV-3 (Zsák & Kisary,
1984) to indicate that it should be designated a new type,
GoAdV-4. Since GoAdV-1, GoAdV-2 and GoAdV-3 are
apathogenic (Zsák & Kisary, 1984) and GoAdV-4 has not
been reported to be pathogenic, GoAdV strain D1036/08
might represent another new type (GoAdV-5) in the same
species as GoAdV-4, because it is pathogenic (Ivanics et al.,
2010) and distinguishable from strain P29 at the hexon locus
(Fig. 2c). Determining the species affiliation of these two
viruses will require sequence data from the three established
types in the species Goose adenovirus A.
Our earlier analyses, based on partial hexon sequences,
showed that waterfowl aviadenoviruses cluster phylogenetically on a sister branch of the FAdVs (Papp et al., 2003).
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2463
G. L. Kaján and others
We suggested that this topology may reflect coevolution of
adenoviruses with their hosts, as birds in the orders
Galliformes and Anseriformes evolved from an ancient
clade, the Galloanserae. However, our present analyses (Fig.
2) included a greater number of aviadenoviruses and resulted
in an overall topology that is not fully concordant with
coevolution. Specifically, the distance between FAdVs and
FaAdV-1 (Schrenzel et al., 2005) is similar to that between
FAdVs and GoAdVs (Fig. 2a, c, d), despite the fact that the
order Galliformes is more distantly related to the order
Falconiformes than to the order Anseriformes (Hackett et al.,
2008). Although coevolution of adenoviruses with their
hosts is generally well supported (Benkö & Harrach, 2003;
Davison et al., 2003), resolution of the situation among
aviadenoviruses will require further scrutiny.
Harrach, B., Benkő, M., Both, G. W., Brown, M., Davison, A. J.,
Echavarrı́a, M., Hess, M., Jones, M. S., Kajon, A. & other authors
(2011). Family Adenoviridae. In Virus Taxonomy: Classification and
Nomenclature of Viruses: Ninth Report of the International Committee
on Taxonomy of Viruses, pp. 125–141. Edited by A. M. Q. King, M.
J. Adams, E. B. Carstens & E. Lefkowitz. San Diego, CA: Elsevier
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Hlinak, A., Müller, T., Kramer, M., Mühle, R. U., Liebherr, H. & Ziedler, K.
(1998). Serological survey of viral pathogens in bean and white-fronted
geese from Germany. J Wildl Dis 34, 479–486.
Ivanics, É., Palya, V., Glávits, R., Dán, A., Pálfi, V., Révész, T. & Benkő, M.
(2001). The role of egg drop syndrome virus in acute respiratory disease
of goslings. Avian Pathol 30, 201–208.
Ivanics, É., Palya, V., Markos, B., Dán, A., Ursu, K., Harrach, B., Kaján, G.
& Glávits, R. (2010). Hepatitis and hydropericardium syndrome
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Acknowledgements
This work was partially supported by the Hungarian Scientific
Research Fund (grant OTKA K72484) and the UK MRC. We thank
Éva Ivanics, Róbert Glávits and Ádám Dán (National Food Chain
Safety Office, Hungary) for allowing us to study the field isolate of
GoAdV strain D1036/08.
Kaján, G. L., Sameti, S. & Benkő, M. (2011). Partial sequence of the
DNA-dependent DNA polymerase gene of fowl adenoviruses: a
reference panel for a general diagnostic PCR in poultry. Acta Vet
Hung 59, 279–285.
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serologic detection of virus-induced infections in the domestic goose
(Anser anser dom.)]. Tierarztl Prax Ausg G Grosstiere Nutztiere 26,
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