論文の公開元へMolecular and pathologic characterization of avian adenovirus isolated from the oviducts of laying hens in eastern Japan
概要
The family Adenoviridae is comprised of middle-sized (70-90 nm), non-enveloped, icosahedral, double stranded DNA viruses. There are five genera which can be found in different animals: genus Mastadenovirus, in mammals; genus Aviadenovirus, in birds; genus Siadenovirus, in birds and reptiles; genus Atadenovirus, in mammals, birds, and reptiles; and genus Ichtadenovirus, in fish (Harrach et al., 2011). The adenovirus (AdV) virion is hexagonal in shape and has three exposed structural proteins. The hexon forms the capsid, the penton base anchors the fiber, and the fiber interacts with the cellular receptor. Many AdVs bind with the coxsackie and adenovirus receptor (CAR) for cellular entry (Fujino et al., 2016). Some members may have a single fiber protein, such as in most mammalian AdVs, while others have two; such as in avian adenoviruses (AAVs) (Harrach et al., 2011)
The AdV genome is a single linear molecule of double-stranded DNA (26-45 kbp) with inverted terminal repeats (30-371 bp) at its termini. The central part of the genome is usually conserved throughout the family, while the terminal portions are unique to genera. The members of genus Aviadenovirus, have the longest genomes (Harrach et al., 2011).
Avian adenoviruses (AAVs) affects birds only. However, the role of AAVs as primary pathogens remains controversial, as strains from the same serotype show variable pathogenicity (Okuda et al., 2006; Marek et al., 2010). The pathologic roles of AAVs are still unclear. In a majority of experimental infections, results have been ambiguous (Harrach et al., 2011). Some AdVs that infect birds belong to different genera other than Aviadenovirus. These are Turkey siadenovirus A (TAdV-A), the cause of hemorrhagic enteritis in turkey, which is under genus Siadenovirus; and Duck atadenovirus A (DAdV-A), the cause of egg drop syndrome (EDS) in chickens, which is under genus Atadenovirus (ICTV, 2018).
The Fowl aviadenoviruses (FAdVs), also known as group-1 AAVs, are important poultry pathogens. They are classified into five species (A to E) with 12 serotypes (1-8a, 8b- 11), of which members are associated to certain diseases (Hess, 2017). Although implicated, the pathogenicity of FAdVs is not well-defined (Niczyporuk et al., 2012). These viruses may act as primary or secondary pathogens (Toro et al., 2000; Niczyporuk et al., 2013), and there are pathogenic and non-pathogenic strains. Additionally, they can be isolated from both healthy and sick birds (McFerran and Smyth, 2000).
Pathogenic FAdV strains linked to certain diseases include: FAdV-1 to adenovrial gizzard erosion (GE) (Grafl et al., 2012), FAdV-2, 8a, 8b, and 11 to inclusion-body hepatitis (IBH) (Nakamura et al., 2011), and FAdV-4 to hydropericardium-hepatitis syndrome (HPS) (Smyth and McFerran, 2000; Joubert et al., 2014). In fact, these diseases have been replicated under controlled conditions using field isolates (Okuda et al., 2006; Mase et al., 2010). At present, the genetic determinant of pathogenicity, which may distinguish the pathogenic from non-pathogenic, is still unclear. This is true for FAdV1 (Matczuk et al., 2017), FAdV-11 (Absalón et al., 2017), and FAdV-4 (Liu et al., 2016; Mo et al., 2019), and the other serotypes.
FAdVs can spread vertically, through the fecal-oral route, or horizontally, thru progeny of infected parent stock (Figure I-1). In the field, vertical transmission of FAdV-1 (Grafl et al., 2012) and FAdV-9 (Grgic et al., 2006) has been described. Experimental studies with FAdV-4 also confirmed vertical transmission (Toro et al., 2001; Mazaheri et al., 2003). Recently, the spread by aerosols was observed (Li et al., 2019). Another mode of transmission, which is quite alarming, is via contaminated vaccines (Li et al., 2017). It is suspected that tainted NDV live vaccines may have caused some IBH or HPS outbreaks (Su et al. 2018).
Generally, pathogenic FAdV infections affect the digestive system. FAdVs have tropism for the gastrointestinal tract (GIT) (Matczuk et al., 2017) but may also affect other organs (Nakamura et al., 2011). Field cases of GE can have mortalities up to 2.35%, together with decreased egg production from 94.5% to 91.5% (Matczuk et al., 2017). Necropsy lesions include hemorrhagic proventriculus, gizzard, lungs, and kidneys; ulceration, perforation, and destruction of the gizzard koilin layer; as well as hepatitis, enteritis, and salpingitis (Domanska- Blicharz et al., 2011; Matczuk et al., 2017). IBH cases in the field can lead to mortalities up 1% to5%, and even as high as 17% (Nakamura et al., 2011). Lesions include hepatomegaly, hepatic necrosis with or without hemorrhage, and friable liver (Nakamura et al., 2011; Joubert et al., 2014). HPS cases in the field can have mortalities of 6.4 to 27.1% (Mase et al., 2009), and even as high as 60% (Cheema et al., 1989; Pan et al., 2017). Lesions include hemorrhagic parenchyma organs and the characteristic hydropericardium coupled with hepatitis (Cheema et al., 1989, Mase et al., 2009). In most experimental trials, disease can be reproduced in day-old SPF chicks only. When artificially infected SPF birds develop disease, a strain is pathogenic (Cook, 1983).
Concurrent infections with other viruses, in the field, is not uncommon. This includes FAdV coinfection with: Chick anemia virus (CAV), Infectious bronchitis virus (IBV), Avian metapneumo virus (aMPV), Reovirus, and Infectious bursal disease virus (IBDV) (De Herdt et al., 2013). FAdV infections mixed with two or more viruses include: CAV, and IBDV (Choi et al., 2012); CAV, Marek disease virus (MDV), and IBDV (Krishan et al., 2015); Avian leukosis virus (ALV), reticuloendotheliosis virus (REV), and CAV (Meng et al., 2018); and CAV, IBDV, and H9N2 avian influenza virus (H9N2-AI) (Yu et al., 2019). Synergism with other viruses has been confirmed under experimental conditions. Viruses that are synergistic with pathogenic FAdV4 include NDV, CAV (Su et al., 2018), and H9-AIV (Niu et al., 2019).
Infection with FAdVs doesn’t always lead to clinical disease. Apparently, there are other variables involved. These include host-related factors, such as: type of bird, age, immune competence, and antibodies in the serum. Virus-related factors include the route of infection, genetic profile, concurrent infections, and infectious dose (Hess, 2017). The study by Grgic et al. (2006) imply not only vertical transmission in the field, but latent infection as well. FAdVs are ubiquitous in poultry, that they are reported worldwide from both sick and healthy birds (McFerran and Smyth, 2000). It is speculated that there might be a wild bird reservoir (Figure I-1). This carrier, upon contact with domesticated birds, could be a source of infection (Li et al., 2017). In fact, FAdV isolates from pigeons has induced IBH in SPF chicks (Takase, et al., 1990).
FAdV infections are diagnosed thru clinicopathological observations, histopathological examination, microscopy, serological techniques, virus isolation, and molecular methods. Virus isolation is the gold standard (Li et al., 2017). Primary cell lines suitable for use include: chick kidney cells (CKC), chick embryo liver cell (CEL), and chick embryo fibroblasts (CEF). For chicken cases, CEL and CKC are the best; virus propagation in homologous cell type is ideal (McFerran and Smyth, 2000). FAdVs produce round cell cytopathic effect (CPE). When possible, isolates must be checked for hemagglutination reaction to exclude Orthomyxoviridae, Paramyxoviridae, and other hemagglutinating AAVs (McFerran and Smyth, 2000). Transformed cells can also be used for propagating FAdVs, these include the quail fibroblast cell line QT35 (Schonewille et al., 2010) and the liver cell line LMH (Yu et al., 2018).
Because of their pathogenic potential, and economic impact, control measures against FAdVs are necessary; this includes vaccination and biosecurity. Inactivated, attenuated, and subunit vaccines against IBH or HPS are available in some countries (Shah et al., 2017). Each vaccine type has its own advantages and disadvantages (Li et al., 2017). Like other viral diseases, there is no specific treatment during outbreaks, thus, disinfection, good husbandry, and biosecurity are important (Shah et al., 2017). Ruano et al. (2001) compared the efficacy of commercial poultry disinfectants. Their results showed that glutaraldehyde (at 1:20) and ionophor (at 1:80) were effective against FAdV even at 10 minutes exposure.
The first case of IBH in Japan was reported in 1972 (Otsuki et al., 1976). The first case of HPS was reported in 1996 (Abe et al., 1998), followed by outbreaks of GE in 1998 (Ono et al., 2001). Since then, cases of FAdV-associated diseases have been reported sporadically in Japanese poultry. In IBH cases, mortalities ranged from 2% to 17% (Nakamura et al., 2011), while mortalities in HPS cases ranged from 6.4% to 26.1% (Mase et al., 2010). The GE cases in 1998 was asymptomatic, and signs were noticed only at slaughter (Ono et al., 2001). FAdV has also been isolated from pigeons with IBH; and these isolates were able to induce disease in SPF chicks (Takase et al., 1990).
Studies on Japanese FAdV isolates show that, compared to other nations, serotype-2 is the major cause of IBH. There is also indication that progeny were infected by parent stock (Nakamura et al., 2011). In other instances, serotypes 2 and 8 induced HPS in SPF chicks (Nakamura et al., 2000). Fiber2 genetic analysis of Japanese HPS strains showed that the strains formed two distinct clades: HPS and non-HPS. Included in the non-HPS clade were non- pathogenic KR5 and Indian strains (Mase et al., 2010).
The objective of this study is to examine two strains of AAV that were isolated from the oviduct of laying hens with poor egg production. Microbial tests, microscopic examination, molecular methods, virus isolation, and artificial infection were used to describe the: pathological, biochemical, and genetic properties of these AAVs. Though these data, the goal is to contribute new information regarding the nature of AAV infections.
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