Page 273 - Avian Virology: Current Research and Future Trends

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analyse because the female and male birds came from two differ- RNA between ED 4–6 and ED 10–12 (Fig. 9.10) (M.M. Miller,
ent breeder flocks and differences in vertical transmission rates K.A. Stucker and K.A. Schat, unpublished data quoted by Schat,
between the two breeder flocks cannot be excluded. 2009). The transcription in the blastoderm is most likely activated
by maternally derived transcription factors. Gonadal develop-
Transmission of CAV in SPF flocks ment with concurrent production of oestrogen occurs between
Most experimental work on CAV has been done in SPF chickens ED 4 and 6. As discussed in ‘Organization of the promoter/
but in this section the natural transmission of CAV within and enhancer region’ oestrogen is a positive regulator of transcription
between subsequent SPF flocks will be discussed. Observations of CAV explaining the presence of viral RNA during this period of
from the SPF industry suggest that if breaks occur it will most embryonal development. The second wave of transcription is also
likely be at the time of sexual development or later (Schat and likely under control of steroid hormones, but these have not been
Van Santen, 2013). The Cornell University SPF flocks (no longer identified. It is not clear if viral proteins are produced during the
in existence) were considered free of CAV but experienced a CAV viral transcription periods because production of viral proteins
break probably around 1996 (Cardona et al., 2000a,b). In 1997, during this period of development would make chickens immu-
two of the three genetic lines showed a high incidence of anti- notolerant to these viral proteins. There is no evidence supporting
bodies between 6 and 16 months of age, while the third line was or refuting the induction of tolerance.
negative at 10 months of age (Cardona et al., 2000a). This finding The Cornell SPF flocks were monitored for seroconversion
was unexpected because all birds were kept in colony cages in the during an 8-year period from 1999 to 2007 when samples were
same facility with the same air-handling unit. Spleens and tissues obtained two or three times for each bird between 108 and 521
from the reproductive tracts from male and female, antibody- days of age. During this period all birds were hatched and kept
positive and -negative birds were examined for the presence of in the same facility. Two flocks in production and two replace-
viral DNA by nested PCR. Hens and roosters were positive in ment flocks representing two genetic lines were maintained at all
70% and 67%, respectively, in one or more of the tissues inde- times. In addition to the breeder flocks, chickens were kept for
pendent of the antibody status (Table 9.4). In situ PCR showed a experimental work up to eight weeks of age. The breeding flocks
few positive cells in the white pulp of spleen, theca externa of the and young birds did not experience signs of any disease. The
ovary and infundibulum sections. Two 9-day-old embryos were percentage positive birds typically started increasing after the first
also positive for CAV by in situ hybridization. The latter observa- sampling. Some flocks had very low seroconversion rates up to
tion suggested that CAV DNA or virus could be transmitted to 464 days of age (e.g. Table 9.5: flock ID 03-3) (Miller et al., 2001;
embryos independently of the presence of antibodies. Miller et al. Schat, 2003; Schat and Schukken, 2010).
(2003) examined semen and blastodisks from antibody-negative These results have important consequences for the SPF
and positive roosters and hens by nested PCR. Positive samples industry. Flocks may have very low conversion rates that are
were found in both types of samples independently of the anti- only detected when all birds are tested for seroconversion.
body status. Spleen, bursa of Fabricius, kidney, gonad, thymus, Moreover, additional testing of all birds may be needed after
bone marrow and blood samples were collected from embryos sexual maturity to detect low levels of seroconversion once egg
between ED16 and 20. Various percentages of the tissues from production starts. The possibility of transmitting viral DNA
these embryos, including samples from antibody-negative, from sero-negative birds adds another complication. There are
PCR-positive hens were positive for CAV DNA. However, not at least two explanations for the increase in seroconversion
all organs from an individual embryo were positive suggesting rates over time and the extreme variation in seroconversion
that transmission did not occur through the germline. Egg-shell rates. The first one is that a low level of continuous horizontal
membranes harvested directly after hatching were also positive infection from seropositive to seronegative birds is responsible
but sometimes only intermittently for embryos from the same for the increase in the per cent seropositive birds. However,
hen (Miller et al., 2003). if this is the case it would be expected that all flocks would
The detection of CAV DNA in many embryonal organs and experience a fairly similar pattern in seroconversion because the
egg-shell membranes with low levels in the blastodisks suggested chickens were maintained during this period in a completely
that viral DNA needs to replicate in the embryo. Fertile eggs closed system. Moreover, horizontal transmission requires a
from CAV positive hens were tested for CAV DNA and RNA fairly high dose of virus (van Santen et al., 2004a; Tan and
from ED 0 to 12. Blastoderm and embryos were positive for viral Tannock, 2005), which is supported by the low transmission

Table 9.4 Distribution of PCR positive tissues from five different flocks. Adapted from Cardona et al., (2000b)
% Positive in
No. of % Total
Sex samples Spleen (S) Ovary (O) Infundibulum (I) S + O O + I S + O + I positive
Female 161 a 11 27 12 16 9 7 70
Spleen (S) Testis (T) Vas Deferens (V) S + VD T + V S + T + V
Male 44 7 0 27 20 2 11 67
a Only 108 infundibulum samples were tested.