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an in situ hybridization (ISH) technique using a digoxigenin Therefore, infections with avian orthoreovirus are best controlled
(DIG)-labelled complementary DNA (cDNA) probe (Liu by proper management procedures, primarily biosecurity and
and Giambrone, 1997), dot blot hybridization assay using a vaccination. It is unrealistic to eradicate this virus from most
radio-labelled cDNA probe (Yin and Lee, 1998b), conventional poultry operations, but a cost benefit analysis by Dobson and
RT-PCR (Xie et al., 1997; Lee et al., 1998), and RT-PCR com- Glisson (1992) determined the financial benefit of vaccinating a
bined with restriction fragment length polymorphism (RFLP) flock of broiler breeders against avian orthoreovirus during egg
(Lee et al., 1998). Liu and Giambrone (1997) and Liu et al. production was approximately 52 times higher than the cost of
(1999) developed an in situ hybridization (ISH) technique and a virulent reovirus infection. Therefore, in North America and
an in situ RT-PCR to detect avian orthoreovirus in formalin-fixed, Europe, vaccination is practised not only for the control of avian
paraffin-embedded chicken tissues and demonstrated that the orthoreovirus disease but also to obtain the vaccination associ-
latter was more sensitive. ated increase in productivity (Kibenge and Wilcox, 1983).
Recently, the application of a real-time PCR technology that Due to the age-linked susceptibility, vaccination programs to
has become more widespread in both human and veterinary control avian orthoreovirus infection have concentrated on the
diagnostics detects the PCR product in real time with a sequence- immunization of breeders with inactivated vaccines to prevent
specific probe labelled with fluorescent dyes (Spackman et al., the disease in this cohort with the added benefit of giving passive
2005b). Real-time PCR-based methods are inherently quantita- immunity to their progeny at hatch when the chicks are most sus-
tive and have high analytic sensitivity and specificity providing ceptible to reovirus disease (van der Heide et al., 1976; Wood et
high consistency in nucleic acid detection of viruses in clinical al., 1986). Meanger et al. (1997) showed that breeding hens can be
samples (Spackman et al., 2005b). Other advantages of real-time immunized with Australian RAM-1 strain of avian orthoreovirus
PCR over conventional PCR include a reduced possibility of to passively transfer neutralizing antibodies via the yolk to their
contamination due to carryover, a decreased risk of exposure progeny. However, the immunization of breeder flocks only pro-
to ethidium bromide, and a less cumbersome procedure requir- tects the first generation of their progeny and not the second (van
ing no gel electrophoresis. Relating RT-PCR specifically to der Heide et al., 1976). Rau et al. (1980) vaccinated parent birds
study this, Ke et al. (2006) developed a Light Cycler SYBR with the S1133 strain and detected progeny immunity in chicks
Green-based real-time RT-PCR to detect S2 segment of avian that hatched 2 weeks later. In addition, the vaccination of breed-
orthoreoviruses. ers blocked the egg-transmission of the virus (van Loon et al.,
Multiplexing is a modification of PCR in which multiple agents 2001). Live attenuated and inactivated oil emulsion vaccines have
can be targeted for by a rapid, cost-effective, sensitive, specific and been used successfully for decades for breeder flock vaccination.
simultaneous detection in a single reaction. Although the cost Wood and Thornton (1981) reported that killed vaccines failed
of real time RT-PCR is close to that of conventional RT-PCR, to provide protection against challenge with virulent orthoreo-
multiplexing reduces cost and time per-sample by simultane- virus strains. However, administration of attenuated vaccines to
ously testing for several agents (Spackman et al., 2005b). Several breeder birds in production can result in trans-ovarian transmis-
multiplex RT-PCR assays have been developed for detection of sion of vaccine virus leading to decreased hatchability, increased
different combinations of avian pathogens, including turkey astro- young chick mortality, and increased incidence of viral arthritis
virus type 2, turkey coronavirus, chicken origin orthoreoviruses, or tenosynovitis in chicks at 7 to 14 days of age. Therefore, when
and turkey origin orthoreoviruses (Spackman et al., 2005b); avian using live vaccine, it should be administered prior to the onset of
orthoreovirus, avian adenovirus-1, IBDV, and CAV (Caterina et egg production (Giambrone et al., 1991). Stott (1999) used an
al., 2004); avian orthoreovirus and Mycoplasma synoviae (Huang attenuated virus successfully to immunize young chicks with no
et al., 2015); and turkey rotavirus, turkey astrovirus-2, and turkey evidence of interference by maternal antibodies. van Loon et al.,
orthoreovirus (Jindal et al., 2012). (2003) reported maternal immunity in commercial broilers does
Recently developed isothermal methods for avian orthoreo- not preclude the efficacy of early age vaccination with an attenu-
virus detection include reverse transcription loop-mediated ated live avian orthoreovirus vaccine. Orthoreovirus vaccination
isothermal amplification (RT-LAMP) (Xie et al., 2012) and of breeders is usually accomplished with a live vaccine by paren-
cross-priming amplification (Woźniakowski et al., 2015). These teral, oral or coarse spray administration (Giambrone et al., 1992;
isothermal methods do not need expensive laboratory equipment, Mukiibi-Muka and Jones, 1999) at an early age followed by an
except a water bath. The main drawback of both conventional inactivated booster vaccine (Chen et al., 2004). According to Ruff
and real-time RT-PCR assays developed for avian orthoreovirus and Rosenberger (1985b), one of the most successful approaches
detection was the absolute need for the expensive thermal cyclers. for vaccinating replacement pullets for broiler breeder opera-
Moreover, the detection limit of the RT-LAMP assay was 10 fg tions is to give one or more immunizing doses of low-virulence
total RNA, which was 100-fold lower than that of RT-PCR (Xie orthoreoviruses early in life followed by inactive vaccine at 18
et al., 2012). to 20 weeks of age. Inactivated vaccines provide high titres of
humoral antibodies to the broiler parents during the production
period and approximately 50% of these antibodies are passed on
Prevention and control to the progeny as maternal antibodies resulting in a significant
Supportive care and control of secondary bacterial infections is correlation between the geometric mean antibody titres of the
the only clinical treatment available for orthoreovirus disease. broiler parents and their progeny (De Herdt et al., 1999). Guo et