Dao Huyen Tran, Nguyen Khanh Thuan, Nguyen Phuc Khanh, Nguyen Thanh Lam* 

1. Introduction

Chicken infectious anemia has emerged as an economically important disease-causing considerable health problems and economic losses to the poultry industry worldwide (Schat et al., 2009; Oluwayelu et al., 2010), especially to the broiler industry and the producers of specific pathogen-free (SPF) eggs. The disease is caused by the chicken anemia virus (CAV), the only member of the genus Gyrovirus of the family Circoviridae. (Yuasa et al., 1979), but Circoviridae taxonomy ratified by the International Committee on the Taxonomy of Viruses in 2016, which reassigns the genus Gyrovirus from the family Circoviridae to the family Anelloviridae (Rosario et al., 2017). The prime targets of this virus are the hemocytoblast of the bone marrow and precursor lymphocytes of the thymus (Noteborn., 2004).

Chicken infectious anemia is characterized by aplastic anemia and generalized lymphoid atrophy with concomitant immunosuppression and frequent association with secondary viral, bacterial, parasitic, or fungal infections (Schat et al., 2003). Mortalities and morbidities due to CAV infection may reach 55% and 80%, respectively (Lai et al., 2013). The clinical signs are mainly noticed in young chicks of 10-14 days of age, which acquire the infection vertically. Chickens older than 2-3 weeks of age are also susceptible to infection but only develop a subclinical disease evidenced by poor vaccine response (Schat et al., 2003). The virus is transmitted either vertically from hens infected for the first time during lay or horizontally in chicks devoid of maternal antibodies (Von Bulow and Schat., 1997). The severe damages caused by this virus such as weight loss, anemia, intramuscular hemorrhage, lymphoid atrophy and bone marrow aplasia are mostly seen among the young chickens less than 2 weeks of age and void of maternally-derived antibodies (Miller and Schat., 2004).

This article reviews the current state of CAV in poultry by discussing the chicken anemia virus, pathogenesis of CAV, serological evaluation of host's antibodies to CAV, an association of Marek's disease (MD) and infectious bursa disease with CAV infection, genetic diversity, and phylogenetics of CAV strains and current vaccine strategy in the control of CAV. In addition, recommendation on vaccine strategy that could be of help in the control of this virus is also highlighted.

2. Aetiology 

2.1 Virus characteristics

The CAV, the only member of the genus Gyrovirus of the family Anelloviridae (Rosario et al., 2017). It is one of the smallest, non enveloped virus; it is 23 – 25 nm in size, icosahedral, having a 2.3 Kbp circular single-stranded DNA genome (Pringle., 1999). The genome codes for three distinct viral proteins (VP1, VP2 and VP3) (Figure 1) from single major transcript three overlapping reading frames (ORF1, 2 and 3) (Noteborn et al.a 1991; Noteborn et al., 1998; Phenix et al., 1994). VP1 is the major capsid protein and has a hypervariable region spanning 13 amino acids (139 to 151) (Renshaw et al., 1996). According to a previous report, the amino acid at position 394 in VP1 could be a major genetic determinant of virulence (Yamaguchi et al., 2001). VP2 is a non-structural scaffold protein. VP1 and VP2 are the protective proteins inducing neutralizing antibodies. VP3 is an apoptin, whose ability to induce tumor-specific apoptosis makes it a promising candidate for gene therapy of various tumors.

2.2 Virus properties

Chicken anemia virus is a naked virus that has a spherical or hexagonal shape and is very small, approximately 23-25 nm in diameter. It has remarkable chemical stability and thermostability. It is able to resist exposure to pH 3, ether or chloroform, treatment for two hours at 37°C with 5% solutions of many commercial disinfectants and is able to withstand temperatures of up to 80°C for 15 minutes. These characteristics indicate that CAV is a remarkably hardy virus and, as a result, it is difficult to eradicate from the environment. The infectivity of CAV can be reduced by exposure to iodophor and formalin, and completely destroyed by hypochlorite (McNulty., 1991).

Chicken anemia virus can be propagated in chickens, embryonated eggs and in cell culture. The virus propagates in the embryo but does not reach a titre high enough to cause it to die. However, after hatching chicks develop the characteristic disease signs (McNulty., 1991). CAV does not grow in standard cell cultures such as cultured monolayer cells derived from a variety of chicken and chicken embryo tissues, nor in a variety of commonly used mammalian cell lines (Yuasa., 1983; Goryo et al., 1987), but it does grow in some lymphoblastoid cell lines established from Marek’s disease virus (MDV) and lymphoid leucosis lymphomas. The most commonly used of these is the continuous Marek’ disease chicken cell (MDCC) MSB1 cell line that consists of MDV transformed chicken lymphocytes derived from a Marek’s disease lymphoma (Yuasa., 1983; Goryo et al., the ). Unlike most cell lines, MSBl cells grow in suspension at 40°C and need to be subcultured every 2-3 days (McNulty., 1991). Infected cell cultures show a cytopathic effect. Cells enlarge and appear swollen and the nuclei contain small vacuoles and assemblies of chromatin, cell destruction occurs, the growth medium becomes alkaline and ultimately there is inability to subculture (McNulty., 1991).

2.3 Morphology

Chicken anemia virions consist of non‐enveloped, icosahedral particles with an average diameter of 25-26.5 nm, as visualized in preparations negatively stained with 1% uranyl acetate (Gelderblom et al., 1989; McNulty et al.,1990). In such preparations, two types of virus particles differing in their orientation on the grid are commonly detected. Type I particles exhibit three‐fold rotational symmetry and show a pattern of one central hollow surrounded by six neighboring hollows with a center‐to‐center distance of 7.5 nm, forming a regular surface network (Figure 2B). Type II particles exhibit five‐fold rotational symmetry and are characterized by 10 evenly spaced surface protrusions giving the impression of a “cog‐wheel” structure (Figure 2A). The appearance of these particles suggested a regular T=3 icosahedron with 32 morphologic subunits. However, modeling of unstained cryopreserved CAV particles indicated a T=1 lattice with 60 copies of VP1 in a capsid consisting of 12 pentagonal, trumpet‐shaped capsomers. These protruding capsomers distinguish CAV from Circoviridae, which have a smoother capsid surface (Crowther et al., 2003). Virions have a buoyant density in cesium chloride gradients variously reported as 1.33–1.34 g/mL (Allan et al., 1994; Todd et al., 1990) or between 1.35 and 1.37 g/mL (De Wit et al., 2004; Eltahir et al., 2011). The sedimentation coefficient of CAV has an estimated value of 91 S in isokinetic sucrose gradients(Allan et al., 1994).

3. History of disease

The first isolation of CAV was the result of an investigation in disease problems in commercial chickens vaccinated with HVT in Japan, material from one of the vaccine batches was shown to induce anemia (Yuasa et al., 1976). In 1979, Yuasa isolated a highly anemia-inducing agent by chick inoculation and was named CAV Gifu‐1 strain (Yuasa et al., 1979). However, the virus was present in chickens at least as early as 1970, when W.C. Wellenstein isolated the ConnB strain of CAV from an ampoule of MD tumor cells that had been stored in liquid nitrogen at least since 1969. The tumor material was obtained from chickens experimentally infected with MDV that had unexpectedly experienced severe anemia (Jakowski et al., 1970).

A major breakthrough was achieved in 1983 when Yuasa et al (1893) reported that virus could be propagated in certain chicken lymphoblastoid cell lines, for example, MDCC MSB‐1, causing cytopathic effects (CPE). This enabled the development of the  vitro serological assays such as indirect immunofluorescence (IIF) assays (v Bülow et al., 1985, Yuasa et al., 1985) and virus‐neutralization tests (Yuasa et al., 1983).

Soiné et al (1993, 1994) and Renshaw et al (1996) confirmed by polymerase chain reaction (PCR) and sequencing that the ConnB strain was indeed CAV. Toro et al (2006) confirmed that CAV infections were present in the United States as early as 1959 by analyzing banked sera from chickens used for the production of all antisera.  In Europe, CAV antibodies were found in 75% and 89.5% of the flocks in The Netherlands and Switzerland, respectively (Wunderwald and Hoop., 2002; De Wit et al., 2004). The combined evidence of these studies suggests that CAV is not a new virus spread by MD vaccination, but that it is an “old” virus that has been present for a long time without causing overt problems. 

In the early 1990s, remarkable progress was made in research on the molecular biology of CAV (Schat., 2009). This resulted in the development of refined diagnostic methods and the potential for the development of new types of vaccines (Koch et al., 1995; Noteborn  et al., 1998). In 2004, based on the model proposed for the control of virus replication showed that CAV has evolved as a successful pathogen that can be maintained in successive generations of chickens without causing disease (Miller and Schat., 2004). However, in the modern high-stress environment of commercial poultry production, it is a major pathogen that can cause significant economic (Schat and van Santen., 2008).

4.  Epidemiology

4.1 Geographic distribution

Chicken anemia virus appears to have a worldwide distribution based on virus isolation and serological evaluations (Vielitz., 1989). There is ample evidence that CAV is not a new pathogen, but a pathogenic virus that had gone unrecognized for many years until its isolation by (Yuasa et al., 1979). It has been found in both commercial and SPF flocks. SPF flock infections have the potential to lead to vaccine contamination since CAV can be vertically (egg) transmitted (Yuasa and Yoshida., 1983).

In the U.S.A., CAV was initially isolated from a farm on the Delmarva peninsula in 1987 (Rosenberger and Cloud., 1988). Serologic evidence of CAV infection exists from the Delmarva peninsula, Texas and Alabama (McNulty et al., 1989), North Carolina, Delmarva and Virginia (Rosenberger and Cloud., 1989), and Arkansas, Connecticut, Georgia, Kansas, Maine, Michigan, New York and Pennsylvania (Lucio et al., 1990). In Egypt, El-Lethi et al (1990) reported the suspicion of CAV in dressed poultry and serological investigation has been proved the intensive exposure of commercial chicken to CA. Hailemariam et al (2008) detected chicken anemia virus from commercial broiler breeder chickens in Malaysia. In India, CIA has been reported from poultry flocks of some states and included in the list the of emerging and important viruses that are a severe threat to the Indian poultry industry (Wani et al., 2013). A number of breeders and commercial flocks in China were reported to suffer from problems suggestive of CAV and the complication described (Cui et al., 1992). Tantaswasdi et al (1996) based on virological and serological studies to demonstrate the presence of CAV in commercial chickens in Thailand. Virological and serological evidence has demonstrated the presence of CAV in the world many years ago. Today, CAV is a concern for the poultry industry in most countries around the world.

4.2 Susceptible hosts

Chicken anemia virus first reported by (Yuasa et al., 1979) from contaminated vaccines in Japan. The chicken is the only natural host for the virus, which is ubiquitous not only in commercial poultry but also in SPF stocks (Cardona et al., 2000). 

Though chickens of all ages are susceptible to CAV infection, but susceptibility to anemia rapidly decreases in immunologically intact chicks during the first one to three weeks of life (Yuasa et al., 1979; Rosenberger and Cloud., 1989), the clinical disease is mainly noticed in young chicks of up to 3-4 weeks of age, which usually acquire the infection vertically (Pope ., 1991; Dhama et al., 2008), in these flocks growth was retarded and mortality was generally between 10 and 20%, but occasionally it reached 60% (Gelderblom et al., 1989), but after 3 weeks of age susceptibility to clinical disease decreases.

CAV-infected birds develop a profound immunosuppression in the presence of concurrent infection with other viruses such as MDV (McNeilly et al., 1991), fowl adenovirus (FAV) (Toro et al., 2001), reoviruses (McNeilly et al., 1995) and Newcastle disease virus (NDV) (De Boer et al., 1994) leading to synergistic effects of both agents (Pope ., 1991) and also causes decreased immune response against several vaccine viruses, resulting in vaccination failures or aggravation of the residual pathogenicity of attenuated vaccine viruses (Schat., 2003; Toro et al., 2006).

Antibodies to CAV have been detected in Japanese quail (Farkas et al., 1998), Netherlands fancy chicken breeds (De Wit et al., 2004), jackdaws, rooks, and rare avian breeds in Ireland (Campbell., 2001), failed to detect antibodies in turkey and duck, pigeons, pheasant, duck, turkey (Campbell ., 2001; Kaffashi et al., 2006).

4.3 Transmission

Chicken anemia virus spreads both horizontally and vertically (Smyth and Schat., 2013). The natural route of chicken anemia virus transmission involves transmission via feather, oral contamination and feces from infected chickens are the main source of virus for horizontal transmission among chickens (Davidson et al., 2008). However, infection via the respiratory route, as shown in chicks after intratracheal inoculation (Rosenberger and Cloud., 1989), also may be possible in the field. Virus is shed in feces and feather follicle epithelium (Davidson et al., 2008). Transmission occurs readily via contaminated litter (Islam et al., 2013). CAV spreads easily among chickens in a group only if they are immunosuppressed (Yuasa et al., 1980). In naturally exposed flocks, it commonly takes two to four weeks until most birds have seroconverted (McNulty et al., 1988; Sommer and Cardona., 2003).

Chicken anemia virus can also be transmitted vertically through hatching eggs. Breeder flocks may become infected before they begin to lay fertile eggs and virus subsequently is transmitted vertically for as long as the hen is viremic. The virus attacks young chicks without maternal antibodies within the first two weeks of age and causes severe damage to tissues and organs (Miller and Schat., 2004). Among the chicks that have maternally inherited antibodies, vertical transmission of the virus is not possible but horizontal transmission with subclinical symptoms is possible as the antibodies wane (Davidson et al., 2008). This causes poor growth and makes chickens susceptible to some secondary infection as MD, infectious bursa disease and other adenoviral infections (Natesan et al., 2006).

Vertical transmission occurs for a period of 3 to 9 weeks after chicken anemia virus infection. When 1-day-old susceptible chicks are inoculated with chicken anemia virus, viremia occurs within 24 hours and virus can be recovered from most organs and rectal contents for up to 35 days. 

5. Pathogenesis

The histological lesions in CAV infected birds are the hemocytoblast of the bone marrow and precursor lymphocytes of the thymus (Noteborn., 2004), or dividing T cells in response to antigenic stimulation. Hemocytoblasts in the bone marrow and lymphoblasts in the thymus cortex are primarily involved in early cytolytic infection at six to eight days post infection (PI) leading to a rapid depletion by apoptosis of these cells. Depletion of lymphoid cells and occasional necrosis in the cloacal bursa, spleen, and lymphoid foci of other tissues have not been detected before 10 to 12 days PI (v Bülow et al., 1986; Goryo et al., 1989; Smyth et al., 1993). Repopulation of the thymus with lymphocytes, repopulation of the bone marrow with proerythroblasts and promyelocytes, and recovery of hematopoietic activity, beginning 16 days PI all appear to coincide with the beginning of antibody formation. These events result in complete recovery by 32-36 days. Large numbers of cortical thymic lymphoblasts become virus positive within four to six days PI. In addition, intrasinusoidal and extrasinusoidal hemocytoblasts, reticular cells in the bone marrow, and mature T cells in the spleen can be virus antigen positive. Infected cells in the thymus and bone marrow are most abundant at six to seven days PI and can be detected until 10 to 12 days or even later. 

The virus infects hemocytoblasts, causing pancytopenia evident as anemia, leukocytopenia and thrombocytopenia (Figure 4). Packed cell volumes are low, and blood smears often reveal anemia and leukopenia. Blood may be watery and clot slowly as a consequence of thrombocytopenia. The loss of thrombocytes and granulocytes is important because two cell types are both important effector cells during bacterial infections and, as a consequence, secondary bacterial infections (e.g., “blue-wing disease”) are frequently associated with CAV-induced immunosuppression. Besides, the hematopoietic cells in the bone marrow become damaged, thereby reducing drastically the number of erythrocyte and myeloid cells, which contributes to the level of anemia in the host (Santen  et al., 2004).

The T lymphocytes are also a major target of CAV with effect on the downstream adaptive immunity (Figure 5). The B cells are not susceptible to CAV directly but indirect impact on B cells has been associated with damage to cytokines and other molecules (Adair., 2000). Different studies have shown the reduction of cytokines such as interleukin 2 (IL-2) with downstream effect on macrophages, neutrophils and the phagocytic activities of the immune system, which is the main cause of the immunosuppressive action of CAV (Natesan et al., 2006; Oluwayelu et al., 2010). In addition to the suppression of the immune molecules, interferon gamma (IFN-γ) has been reported to increase in first few days of infection, followed by gradual reduction (Natesan et al., 2006). Drastic reduction of cytokines (IL-2, IL-1, IL-12) at all doses with a 3–15-fold initial increase of IFN- γ at the early stage of infection was also established. 

Besides, quantitative (q)RT-PCR assays have been used to investigate the effects of CAV infection on cytokines in relation to virus replication. Unfortunately, the few published results have not included the effect of virus replication prior to 7 days PI, when high levels of virus replication occur (Markowski‐Grimsrud and Schat., 2003; Santen et al., 2004). At that time, immunosuppressive effects are already evident with impairment of macrophages (McConnell et al., 1993) and cytotoxic T lymphocytes (CTL), but IFN-γ, IL-2 and IL-1β mRNA levels were not affected (Markowski‐Grimsrud and Schat., 2003). 

Clearly, additional studies using (q)RT-PCR assays or enzyme-linked immunoassays (ELISA) are needed to determine the impact of CAV infection on cytokines starting at 2 to 3 days PI, because viral antigens can be detected in lymphoid tissues and bone marrow as early as 3 to 4 days PI (Smyth et al., 1993). 

The reduction of CD4+ and CD8+ in CAV infected chicks has also been reported (Adair., 2000; Wani et al., 2016). Transient severe depletion of CD4+ and CD8+ lymphocytes, or a selective decrease in CTL, may play an important role in the mechanism of CAV induced immunosuppression (Jeurissen et al., 1989a; Hu et al., 1993). Recently, Haridy et al (2011) reported that infection in 4-week-old chickens resulted in a moderate loss of CD41 and CD81 cells in the spleen and thymus. Infection in 1-day-old chicks causes a more severe depletion of CD41 and CD81 cells (Hu et al., 1993). The effect of virus replication in these cells is especially important when replication of CAV occurs at the same time that CTL are generated in response to vaccination or infection with a second pathogen.

Markowski‐Grimsrud and Schat (2003) reported the absence of REV-specific CTL 7 days after birds were co-infected with REV and CAV, at which time CAV was actively replicating, based on RT-PCR analysis. Because there was no effect of CAV infection on transcription of IL-2 or IFN-γ at 7 days PI, it was suggested that the lack of pathogen-specific CTL was caused by CAV-induced apoptosis of CD81 cells during the generation of CTL. 

In contrast to the effect on CTL, natural killer (NK) cells were not affected by CAV infection (Markowski-Grimsrud and Schat., 2001). Based on their studies of CAV infection in MSB-1 cells, (Peters et al., 2006) suggested that VP2 may also play a role in immunosuppression through down-regulation of major histocompatibility complex (MHC) class I antigens. The importance of this observation for immunosuppression is difficult to evaluate because the assumption is that CAV-infected cells will become apoptotic.

CAV-induced immunosuppression has been causally linked to increased incidence of other diseases (Schat and van Santen., 2008). For example, infection with CAV can aggravate infectious bronchitis virus (IBV) -induced disease (Toro et al., 2006), likely by affecting both CMI and antibody responses. Van Ginkel et al., 2008 demonstrated reduced local antibody responses to IBV in the Harderian gland and lacrimal fluids in CAV-infected chickens. This effect was most likely caused by a decrease in CD41 Th cells as a consequence of CAV infection.

6. Clinical signs of disease and pathology

6.1 Clinical signs

Naturally occurring disease

Typical symptoms observed at the onset of clinical disease include weakness, depression, anorexia and stunting and an increased daily mortality ensues (Figure 6A, B). Anaemia is noticeable on the non-feathered areas such as the comb and wattles, eyelids and legs and the carcass appears quite pale. The clinical disease is also characterized by skin lesions, most commonly appearing on the wings (Figure 7), which are prone to secondary bacterial infections (Yuasa et al., 1979; McNulty., 1991). The disease is acute. Peak mortality occurs within 5 to 6 days of onset of disease signs, and mortality has often declined to normal levels after a further 5 to 6 days (Engström and Luthman., 1984; Yuasa et al., 1987). The severity or intensity of the symptoms may be related to the viral dose, age of the infection, the maternal antibody status and the route of infection (Rosenberger and Cloud., 1989).

Affected birds often have focal skin lesions. These occur most commonly on the wings, but may also be present on the head, around the rump, on the sides of the thorax and abdomen, on the thighs and legs and feet. These lesions appear to be due to ecchymotic skin hemorrhages. The skin turns blue and breaks, releasing a serosanguinous exudate. These lesions are prone to secondary bacterial infection, leading to gangrenous dermatitis. These signs and lesions have given rise to 

The specific sign of CAV infection is anemia, with a peak at 14 to 16 days PI. Anemia is characterized by hematocrit values ranging from 6-27% (Figure 8). Affected birds may die between 12 and 28 days PI. If mortality occurs, it generally does not exceed 30%. Surviving chicks completely recover from depression and anemia by 20-28 days PI (Yuasa et al., 1979; Goryo et al., 1985; v Bülow et al., 1986; Rosenberger and Cloud.,1989), although retarded recovery and increased mortality may be associated with secondary bacterial or viral infections. Secondary infections, causing more severe clinical signs, are frequently seen in field cases, but they may also occur inadvertently in experimental chicks (Goryo et al., 1987; Engström et al., 1988; Vielitz and Landgraf., 1988).

Experimental disease

The proportion of chicks that show clinical signs following experimental parenteral inoculation with CAV at 1-day-old is variable. In a comparison of 11 Japanese isolates, (Yuasa and Imai., 1986) recorded mortality rates varying from 20% to 70%. However, morbidity rates, as assessed by the development of anemia, were 100% for each isolate. In contrast, (Engström et al., 1988) observed no mortality and only slight decreases in hematocrit values in chicks inoculated with their Swedish isolate. As mortality rates in different experiments using the same isolate may be highly variable (McNulty et al., 1990), it is difficult to draw firm conclusions regarding the relative pathogenicity of different isolates. 

Following intramuscular inoculation at one day of age, mortality and disease due to CAV usually occurred in chicks between 10 and 24 days of age. Disease signs were similar to those in the naturally occurring disease, except that skin lesions were not a feature of the experimental disease. The weight gains of inoculated chicks were decreased relative to uninoculated controls. Dead or moribund chicks had yellow bone marrow, severe atrophy of the thymus and bursa of Fabricius, and discoloration and swelling of the liver, kidneys and spleen (Figure 9A, B, C). Subcutaneous and intramuscular hemorrhages, and hemorrhage in the mucosa of the proventriculus, were present in some birds (v Bülow et al., 1986; Goryo et al., 1987; Otaki et al., 1987; Engström et al., 1988).

6.2 Pathology

Lesions associated with CIA may vary dependent on the route of infection, age of exposure, viral dose, and immune status of the host. Moreover, CAV infection may often be involved in diseases caused by other pathogens and can be complicated by other pathogens. The pathology will be described for uncomplicated infections mostly based on experimental infections, as part of the hemorrhagic‐aplastic anemia syndrome, and as a complicating factor in other diseases.

Gross lesion

Thymic atrophy (Figure 10), sometimes resulting in an almost complete absence of thymic lobes, is the most consistent lesion especially when chicks develop age resistance to anemia (Goryo et al., 1985; Smyth et al., 2006). The thymic remnants may have a dark reddish color. Bone marrow atrophy is the most characteristic lesion seen and is best evaluated in the femur (Goryo et al., 1989). The affected bone marrow becomes fatty and yellowish or pink (Figure 11). Print some instances, its color appears dark red, although distinct lesions can be detected by histologic examination. Bursal atrophy is less commonly associated with CAV infection. In a small proportion of birds, the size of the cloacal bursa (bursa of Fabricius) may be reduced. In many cases, the outer bursal wall appears translucent, so plicae become visible. Hemorrhages in the proventricular mucosa and subcutaneous and muscular hemorrhages are sometimes associated with severe anemia. More pronounced hemorrhages or bursal atrophy, and lesions in other tissues, for example, swollen and mottled livers have also been reported but may be caused by secondary infections with other agents (Goryo et al., 1989; Rosenberger and Cloud., 1989).

Hemorrhagic‐aplastic anemia syndrome

Outbreaks of infectious anemia in field flocks are mostly associated with the so‐called hemorrhagic syndrome, with or without concurrent (gangrenous) dermatitis (Figure 12) (Dorn et al., 1981; Bisgaard., 1983; Engström and Luthman., 1984; Yuasa et al., 1987; Landgraf., 1988; Chettle et al., 1989). CAV is also involved in the etiology of aplastic anemia associated with inclusion body hepatitis (IBH) (v Bülow et al., 1986) and with the IBH/hydropericardium syndrome or infectious bursal disease (IBD) (Pope ., 1991). Hemorrhages seen in chickens with IBD may, in most instances, be a sequel of CAV rather than infectious bursal disease virus (IBDV) infection.

Characteristic lesions of the so‐called hemorrhagic syndrome are intracutaneous, subcutaneous, and intramuscular hemorrhages (Figures 13 and 14). Punctuate hemorrhages may be present even more frequently in the mucosa of the distal part of the proventriculus (Figure 15). Intracutaneous or subcutaneous hemorrhages of the wings are often complicated by severe edema and subsequent dermatitis, which may become gangrenous due to bacterial infection (Engström and Luthman., 1984). Subcutaneous hemorrhage of shanks and feet may result in the formation of ulcers. Affected chicks also sometimes appear to be predisposed to develop pododermatitis. 

Hemorrhages are not consistently seen in anemic chicks, although their occurrence is mostly correlated with the severity of anemia. Increased clotting time associated with thrombocytopenia, therefore, does not completely explain hemorrhages. Endothelial lesions and impaired liver functions, partly caused by a viral infection and enhanced by secondary bacterial infection, are likely to be more important in the pathogenesis of hemorrhagic diathesis.

7. Diagnosis

7.1 Differential diagnosis

Infection criteria have only limited value in the diagnosis of CAV‐induced disease because CAV is virtually ubiquitous among chickens. Demonstration of the virus, viral antigens, or viral DNA may be considered etiologically significant if detected at sufficiently high levels in a high proportion of affected birds. In chickens under six weeks of age, a typical combination of signs, hematologic changes, gross and microscopic lesions, and flock history are suggestive of CIA. However, no particular lesions can be considered pathognomonic.

Aplastic anemia, but not ced by erythroblastosis virus can be distinguished from CAV‐induced anemia by microscopic examination of blood smears. MDV can cause severe atrophy of the thymus and cloacal bursa, especially after infection with very virulent viruses (Miles et al., 2001). IBDV induces atrophy of lymphoid tissues with typical histologic lesions but normally does not affect the thymus. MDV and IBDV normally do not cause anemia, although anemia has been described with some strains of MDV (Gilka and Spencer., 1995). Aplastic anemia that may be associated with acute IBDV occurs and disappears much earlier than CAV‐induced anemia (Nunoya et al., 1992). Adenovirus is a major cause of an inclusion body hepatitis‐aplastic anemia syndrome that occurs most frequently between 5 and 10 weeks of age (Cowen., 1992). It does not, however, induce aplastic anemia after a single infection of experimental chickens.

Intoxication with high doses of sulfonamides, or mycotoxins such as aflatoxin, can result in aplastic anemia and “hemorrhagic syndrome”. Aflatoxin also may impair the immune system. In the field, however, chickens are rarely exposed to doses of aflatoxin or sulfonamides that are sufficient to cause acute disease. On the other hand, subclinical intoxication of chickens might add to the pathogenicity of CAV or vice versa.

7.2 Laboratory diagnosis

A definitive diagnosis of CIA can be made by virus isolation, demonstration of virus antigen in impression smears, and cryostat tissue sections (Oluwayelu., 2010). Also, the detection of serum antibodies to the virus and detection of CAV nucleic acid in tissues from diseased birds using Marek’s disease virus-transformed chicken lymphoblastoid (MDCC-MSB1) cell line (McNulty., 1998) and PCR (Adedeji et al., 2016) have been demonstrated. The serological diagnosis of CIA includes ELISA (Shettima et al., 2017), indirect IIF, immunoperoxidase tests, and VN test (Oluwayelu., 2010).

8. Treatment

No specific treatment for chickens affected by CAV infection is available. Treatment with broad‐spectrum antibiotics to control bacterial infections usually associated with CIA might be indicated. The potential for herbal immunomodulatory and hematinic supplements to ameliorate the immunosuppressive, anemia, and growth-suppressive effects of CAV infection has been investigated (Gopal et al., 2015; Latheef et al., 2017).

VEMEDIM PRODUCTS FOR ENHANCING IMMUNE RESPONSE TO RESIST THE PATHOGENS

- Vime -Booster: 1 mL/liter of water. 

- B complex C: 100 g/100 liters of water or 100 g/50-80 kg of feed

- Vita C 250: 1g per 5 liters of drinking water or 1g per 2.5kg of feed for 3-5 days.

- Vimix plus: One 100 g pack to be dissolved in 60 liters of drinking water

- Vimeperos: 1g/5 liters of drinking water

- AD₃E: 15g/1 liter of water or 15g/0.5kg of feed

- Aminovit: 1g/ 20liters of water 

9. Control and prevention

The control measures for CIA include vaccination and good poultry health and management practices (Oluwayelu., 2010). Immunity to the chicken anemia virus is complex. Neutralizing antibodies are protective against disease but do not completely protect chickens against infection or result in virus clearance. The presence of antibodies in breeders greatly reduces vertical as well as horizontal transmission. Several commercial vaccines are available and are mainly used in broiler breeders. Maternal antibodies and controlled exposure are primary methods for control in broilers.

The immunization of breeder flocks against CAV has been reported to ensure more protective levels of passive immunity for the progeny chicks during the first few weeks of life (Vielitz et al., 1987). This has been reported to help minimize vertical transmission of the CAV (Oluwayelu., 2010). The introduction of CAV vaccination program requires consideration to the nature and immunopathogenesis of CAV infection in relation to other agents (Oluwayelu., 2010). Because severe disease results from coinfection with immunosuppressive viruses such as MDV, control of these other pathogens also is important. Besides, co-infection with IBDV enhances the pathogenicity of CAV (Owoade et al., 2013), IBD control has been suggested to be integrated into the CAV control program. 

Complete elimination of chicken anemia virus from chicken flocks is very challenging and, therefore, there is a need to improve hygiene and biosecurity on poultry farms (Adedeji et al., 2016). This may help to prevent cross-contamination between flocks and the introduction of viruses from indigenous to commercial chickens, or vice versa. In addition, periodic decontamination and proper timing of flocks should be practiced (Oluwayelu., 2010). Generally, good farm management and hygiene procedures are important to minimize the economic impact associated with chicken anemia virus infection.

10. Vaccination

Current vaccination strategies are based on the prevention of vertical and horizontal transmission of the virus to very young chicks by immunization of breeder flocks and have been successful in reducing the incidence of anemia in young chicks (Engström., 1999).

Artificial exposure of young breeder flocks was originally achieved by the transfer of litter from CAV‐infected flocks or by providing drinking water containing CAV‐positive tissue homogenate. This method is still used in countries where vaccines are not available or where vaccines are not applied for economic reasons. However, these procedures are very risky with regard to hygiene and level of exposure and should be discouraged (Vielitz and Landgraf., 1988).

Commercial live vaccines for pullets are available in several countries (Vielitz  et al., 1991; Vielitz and Voss., 1994). Vaccination should be performed at about 9 to 15 weeks of age, but never later than three to four weeks before the first collection of hatching eggs to avoid the hazard of vaccine virus spread through the egg. Vaccines can be applied in the drinking water or by injection.

Based on the negative effect of CAV on the generation of CTL when infection occurs after maternal antibodies have disappeared (Markowski‐Grimsrud and Schat., 2003) vaccination for broilers may also be necessary. Several strategies utilizing various combinations of natural exposure, monitoring for seroconversion, and vaccination of breeders are actually used in the broiler industry. Vaziry et al (2011) showed that inoculation of one‐day‐old SPF chickens with a vaccine licensed for pullets resulted in persistence of the vaccine virus in the thymus and spleen in some birds, altering the thymopoiesis and inducing a low antibody response to CAV. Currently, all live CAV vaccines are intended for use in parent and grandparent flocks for the vaccination of chickens older than eight weeks of age. Thus biosecurity measures must be employed to prevent infection before vaccination.

The paucity of information on the stability of current vaccines and the effects of vaccination on immune responses has led to several approaches to develop safer vaccines. Inactivated vaccines have been tested in SPF breeder hens. Vaccinated hens showed seroconversion and their offspring were protected against challenge (Pagès‐Manté et al., 1997; Zhang et al., 2015). Unfortunately, viral titers in MSB‐1 cells and embryos are generally low (McNulty., 1991) and therefore inactivated vaccines may not be cost‐effective.

Schat et al (2011) inoculated one‐day‐old SPF chickens with antigen-antibody complexes and showed that specific combinations of antigen and antibody did induce a protective response against challenge with a different strain. Based on the work by Koch et al., (1995) and Noteborn et al., (1998) indicating that co-expression of VP1 and VP2 is required for induction of neutralizing antibodies, recombinant vaccines expressing VP1 and VP2 are possible. Finally, Moeini et al., (2011) also showed that Lactobacillus acidophilus displaying purified VP1, which was synthesized in E.coli as a fusion protein with a cell wall binding domain in the presence of VP2, can be potentially used as an oral vaccine although the antibody titers were rather low.

11. Chicken anemia virus effect on vaccination

Chicken anemia virus alone or in combination with IBDV has been shown to affect the immune response to several vaccine viruses (Box et al., 1988; De Boer et al., 1989). Inoculation with CAV between 1 and 14 days of age resulted in depressed vaccinal immunity against MDV (Otaaki et al., 1988). It was suggested that failure of vaccination with turkey herpesvirus or attenuated serotype 1 MDV may be related to compromised T cell-mediated immunity resulting from enhanced pathogenicity of CAV in vaccinated chicks (Otaaki et al., 1988). (Box et al., 1988) reported that breeder flocks that seroconverted to CAV at 8 and 22 weeks of age showed impaired response to killed Newcastle disease (ND) vaccine, but no impaired response to killed infectious bronchitis or killed IBD vaccination. No impairment to killed ND vaccination was seen in flocks that were CAV-antibody negative (Box et al., 1988). De Boer et al., (1989) recovered CAV from severe outbreaks of MD in the U.S.A., Israel and The Netherlands. The MDV isolates were of normal virulence and all flocks had been vaccinated with bivalent MDV vaccines. Rosenberger and Cloud (1989) demonstrated that the administration of CAV at the day of hatch to SPF chicks reduced the response to NDV fowl pox and infectious laryngotracheitis (ILT) vaccination. IBDV exacerbated the increase in susceptibility seen in CAV inoculates with the exception of the ILT vaccinated group. Chicks that received CAV-IBDV on day of hatch were more resistant to ILT virus challenge at 4 weeks of age than vaccinated controls. SPF birds inoculated at 2 weeks of age with CAV and/the IBDV were more resistant to challenge with NDV, fowl pox and ILT virus than were the day of hatch inoculates. Rosenberger and Cloud (1989) also found that commercial broilers exposed to CAV-IBDV had lower antibody titers to IBV and NDV vaccination than birds infected with CAV or IBDV alone. Birds not infected with CAV or IBDV had the highest IBV and NDV titers.

12. References

AboElkhair, M., et al. (2014). "Molecular characterization of chicken anemia virus circulating in chicken flocks in Egypt." 2014. 

Adair, B. M. (2000). Immunopathogenesis of chicken anemia virus infection. Developmental & Comparative Immunology, 24(2-3), 247-255.

Adedeji, A., et al. (2016). "Concurrent infections of chicken infectious anemia and infectious bursal disease in 5 weeks old pullets in Jos, Plateau State, Nigeria."  2(3): 60-65.

Allan, G., et al. (1994). "Some biological and physico‐chemical properties of porcine circovirus."  41(1‐10): 17-26.

Authority, E. F. S. (2010). "Analysis of the baseline survey on the prevalence of Campylobacter in broiler batches and of Campylobacter and Salmonella on broiler carcasses in the EU, 2008 - Part A: Campylobacter and Salmonella prevalence estimates."  8(3): 1503.

Bisgaard, M. J. N. v. (1983). "An age related and breeder flock associated hemorrhagic disorder in Danish broilers." 35(11): 397-407.

Box, P., et al. (1988). "Impaired response to killed Newcastle disease vaccine in chicken possessing circulating antibody to chicken anaemia agent."  17(3): 713-723.

Campbell, G. (2001). Investigation into evidence of exposure to infectious bursal disease virus (IBDV) and chick infectious anaemia virus (CAV) in wild birds in Ireland. Proc. 2nd Int. Symp. Infectious Bursal Disease and Chicken Infectious Anaemia, Rauischholzhausen, Germany. Institut für Geflügelkrankheiten, Justus Liebig University, Giessen, Germany.

Cardona, C., et al. (2000). "Humoral immune responses to chicken infectious anemia virus in three strains of chickens in a closed flock." 661-667.

Chansiripornchai, N. J. T. T. J. o. V. M. (2016). "Field study of seroconversion of three different commercial vaccines of chicken infectious anemia virus in Thailand."  46(4): 699.

Chettle, N., et al. (1989). "An outbreak of disease due to chicken anaemia agent in broiler chickens in England." 124(9): 211-215.

Chu, D. K., et al. (2012). "Characterization of a novel gyrovirus in human stool and chicken meat."  55(3): 209-213.

Cowen, B. S. J. W. s. P. S. J. (1992). "Inclusion body hepatitis–anaemia and hydropericardium syndromes: aetiology and control."  48(3): 247-254.

Crowther, R., et al. (2003). "Comparison of the structures of three circoviruses: chicken anemia virus, porcine circovirus type 2, and beak and feather disease virus."  77(24): 13036-13041.

Cui, X., et al. (1992). "Identification of chicken infectious anemia virus."  6: 3-5.

Davidson, I., et al. (2008). "The contribution of feathers in the spread of chicken anemia virus."  132(1-2): 152-159.

De Boer, G., et al. (1989). Enhancing effects of chicken anemia agent (CAA) on Marek's disease pathogenesis. Pro-ceedings of the 38th Western Poultry Diseases Conference, Tempe, Ariz.

De Boer, G., et al. (1994). "Interaction between chicken anaemia virus and live Newcastle disease vaccine."  23(2): 263-275.

De Wit, J., et al. (2004). "A serological survey for pathogens in old fancy chicken breeds in central and eastern part of The Netherlands."  129(10): 324-327.

Del Valle, F., et al. (2020). "The diseases suspected of the involvement of chicken anemia virus infection in 11 to 14-weeks old replacement pullets from eastern Japan: a case report." 82(5): 520-526.

Dhama, K., et al. (2008). "Chicken infectious anaemia virus: an immunosuppressive pathogen of poultry—a review." 32(2): 158-167.

Dorn, P., et al. (1981). "Anaemie, Ruckbildung der lymphatischen Organe under Dermatitis. Beobachtungen zu einem neuen Krankheitsbild in der Geflugelmast."

Ducatez, M., et al. (2008). "Molecular epidemiology of chicken anemia virus (CAV) in southeastern Chinese live birds markets."  52(1): 68-73.

El-Lethi, A. (1990). Viral contamination of dressed poultry. MV Sc, thesis, Cairo University, Egypt.

Eltahir, Y. M., et al. (2011). "Analysis of chicken anemia virus genome: evidence of intersubtype recombination."  8(1): 1-7.

Engström, B., et al. (1988). "Blue wing disease of chickens: experimental infection with a Swedish isolate of chicken anaemia agent and an avian reovirus."  17(1): 33-50.

Engström, B. and M. J. A. P. Luthman (1984). "Blue wing disease of chickens: signs, pathology and natural transmission." 13(1): 1-12.

Engström, B. J. A. V. S. (1999). "Prevalence of antibody to chicken anaemia virus (CAV) in Swedish chicken breeding flocks correlated to outbreaks of blue wing disease (BWD) in their progeny." 40(2): 97-107.

Fang, L., et al. (2017). "Genetic analysis of two chicken infectious anemia virus variants-related gyrovirus in stray mice and dogs: The first report in China, 2015."  2017.

Farkas, T., et al. (1998). "A serological survey of chickens, Japanese quail, pigeons, ducks and crows for antibodies to chicken anaemia virus (CAV) in Japan."  27(3): 316-320.

Gelderblom, H., et al. (1989). "Morphological characterization of chicken anaemia agent (CAA)."  109(1-2): 115-120.

Gilka, F. and J. J. A. P. Spencer (1995). "Extravascular haemolytic anaemia in chicks infected with highly pathogenic Marek's disease viruses."  24(3): 393-410.

Gopal, K., et al. (2015). "Immunomodulatory and protective effects of a polyherbal formulation (Immon) against infectious anemia virus infection in broiler."  11(5): 470-476.

Goryo, M., et al. (1987). "Outbreak of anemia associated with chicken anemia agent in young chicks."

Goryo, M., et al. (1985). "Isolation of an agent inducing chicken anaemia."  14(4): 483-496.

Goryo, M., et al. (1987). "Serial propagation and purification of chicken anaemia agent in MDCC‐MSB1 cell line."  16(1): 149-163.

Goryo, M., et al. (1989). "Histopathology of chicks inoculated with chicken anaemia agent (MSB1‐TK5803 strain)."  18(1): 73-89.

Hailemariam, Z., et al. (2008). "Detection and characterization of chicken anemia virus from commercial broiler breeder chickens." 5(1): 1-11.

Haridy, M., et al. (2011). "Pathological and immunohistochemical studies of subclinical infection of chicken anemia virus in 4-week-old chickens." 1201210763-1201210763.

Hu, L.-b., et al. (1993). "Depletion of CD4+ and CD8+ T lymphocyte subpopulations by CIA-1, a chicken infectious anemia virus." 492-500.

Huynh, L. T. M., et al. (2020). "Chicken infectious anaemia virus infections in chickens in northern Vietnam: epidemiological features and genetic characterization of the causative agent." 49(1): 5-14.

Islam, A., et al. (2013). "Development of a chick bioassay for determination of infectivity of viral pathogens in poultry litter."  91(1-2): 65-71.

Jakowski, R. M., et al. (1970). "Hematopoietic destruction in Marek's disease." 374-385.

Jeurissen, S., et al. (1989). "Transient depletion of cortical thymocytes induced by chicken anaemia agent."  14(1-3): 115-123.

K. A. Schat, “Chicken infectious anemia,” in Diseases of Poultry, Y. M. Saif, H. J. Barnes, J. R. Glisson, A. M. Fadly, L. R. McDougald, and E. Swayne, Eds., pp. 182–202, Iowa StateUniversity Press, Ames, Iowa, USA, 11th edition, 2003

Kaffashi, A., et al. (2006). "Viral load in 1-day-old and 6-week-old chickens infected with chicken anaemia virus by the intraocular route."  35(6): 471-474.

Kim, H.-R., et al. (2010). "Molecular characterization of chicken infectious anemia viruses detected from breeder and broiler chickens in South Korea."  89(11): 2426-2431.

Koch, G., et al. (1995). "Immunogenic and protective properties of chicken anaemia virus proteins expressed by baculovirus."  13(8): 763-770.

Kye, S.-J., et al. (2013). "Phylogenetic analysis and genetic characterization of chicken anemia virus isolates from Cambodia."  92(10): 2681-2686.

Lai, G.-H., et al. (2013). "Expression and characterization of highly antigenic domains of chicken anemia virus viral VP2 and VP3 subunit proteins in a recombinant E. coli for sero-diagnostic applications."  9(1): 1-10.

Latheef, S. K., et al. (2017). "Immunomodulatory and prophylactic efficacy of herbal extracts against experimentally induced chicken infectious anaemia in chicks: assessing the viral load and cell mediated immunity."  28(1): 115-120.

Li, Y., et al. (2017). "Genomic characterization of recent chicken anemia virus isolates in China."  8: 401.

Li, Y., et al. (2021). "Molecular evolution and pathogenicity of chicken anemia virus isolates in China." 166(2): 439-449.

Lucio, B., et al. (1990). "Identification of the chicken anemia agent, reproduction of the disease, and serological survey in the United States." 146-153.

McNulty MS (1998) Chicken anemia virus. In: Swayne DE, Glisson JR, Jackwood MW, Pearson JE, Reeds WM (Eds) A Laboratory Manual for the Isolation and Identification of Avian Pathogens. 4th edition. Kenneth square: American Association of Avian Pathologists, Pp. 146-149

Markowski-Grimsrud, C. and K. Schat (2001). Impairment of cell-mediated immune responses during chicken infectious anemia virus infection. Proc 2nd Int Symp Infect Bursal Dis Chick Infect Anaemia, Rauischholzhausen, Germany.

Markowski‐Grimsrud, C. J. and K. A. J. I. Schat (2003). "Infection with chicken anaemia virus impairs the generation of pathogen‐specific cytotoxic T lymphocytes."  109(2): 283-294.

McConnell, C., et al. (1993). "Effects of chicken anemia virus on macrophage function in chickens." 358-365.

McMullin, P. F. (2020). Diseases of poultry 14th edition: David E. Swayne, Martine Boulianne, Catherine M. Logue, Larry R. McDougald, Venugopal Nair, David L. Suarez, Sjaak de Wit, Tom Grimes, Deirdre Johnson, Michelle Kromm, Teguh Yodiantara Prajitno, Ian Rubinoff & Guillermo Zavala (Eds.), Hoboken, NJ, John Wiley & Sons, 2020, 1451 pp.,£ 190 (hardcover)/£ 171.99 (e-book), ISBN 9781119371168, Taylor & Francis.

McNeilly, F., et al. (1991). "Detection of chicken anaemia agent in chickens by immunofluorescence and immunoperoxidase staining."  20(1): 125-132.

McNeilly, F., et al. (1995). "Synergism between chicken anemia virus (CAV) and avian reovirus following dual infection of 1-day-old chicks by a natural route." 532-537.

McNulty, M., et al. (1988). "A serological survey of domestic poultry in the United Kingdom for antibody to chicken anaemia agent."  17(2): 315-324.

McNulty, M., et al. (1990). "Preliminary characterisation of isolates of chicken anaemia agent from the United Kingdom." 19(1): 67-73.

McNulty, M., et al. (1989). "Chicken anemia agent in the United States: isolation of the virus and detection of antibody in broiler breeder flocks." 691-694.

McNulty, M., et al. (1990). "Chicken anemia agent: an electron microscopic study." 736-743.

McNulty, M. S. (1991). "Chicken anaemia agent: A review." Avian Pathology 20(2): 187-203.

Miles, A. M., et al. (2001). "Coinfection of specific-pathogen-free chickens with Marek's disease virus (MDV) and chicken infectious anemia virus: effect of MDV pathotype." 9-18.

Miller, M. M. and K. J. A. d. Schat (2004). "Chicken infectious anemia virus: an example of the ultimate host–parasite relationship."  48(4): 734-745.

Moeini, H., et al. (2011). "Lactobacillus acidophilus as a live vehicle for oral immunization against chicken anemia virus."  90(1): 77-88.

Natesan, S., et al. (2006). "Biological and molecular characterization of chicken anaemia virus isolates of Indian origin." 118(1-2): 78-86.

Noteborn, M., et al. (1991). "Characterization of cloned chicken anemia virus DNA that contains all elements for the infectious replication cycle."  65(6): 3131-3139.

Noteborn, M., et al. (1998). "Simultaneous expression of recombinant baculovirus-encoded chicken anaemia virus (CAV) proteins VP1 and VP2 is required for formation of the CAV-specific neutralizing epitope."  79(12): 3073-3077.

Noteborn, M. H. J. V. m. (2004). "Chicken anemia virus induced apoptosis: underlying molecular mechanisms."  98(2): 89-94.

Nunoya, T., et al. (1992). "Occurrence of acute infectious bursal disease with high mortality in Japan and pathogenicity of field isolates in specific-pathogen-free chickens." 597-609.

Oluwayelu, D., et al. (2010). "Biological characterization of Nigerian chicken anaemia virus isolates."  39: 21-26.

Oluwayelu, D., et al. (2005). "Isolation and preliminary characterization of chicken anemia virus from chickens in Nigeria." 49(3): 446-450.

Oluwayelu, D. O. J. A. J. o. B. (2010). "Diagnosis and epidemiology of chicken infectious anemia in Africa." 9(14): 2043-2049.

Otaaki, Y., et al. (1988). "Depression of vaccinal immunity to Marek's disease by infection with chicken anaemia agent." 17(2): 333-347.

Otaki, Y., et al. (1987). "Isolation of chicken anaemia agent and Marek's disease virus from chickens vaccinated with turkey herpesvirus and lesions induced in chicks by inoculating both agents." 16(2): 291-306.

Owoade, A., et al. (2013). "Report of Mixed Infection of Infectious Bursal Disease and Chicken Infectious Anaemia Viruses." 9(1): 1-5.

Pagès‐Manté, A., et al. (1997). "Experimental evaluation of an inactivated vaccine against chicken anaemia virus." 26(4): 721-729.

Peters, M. A., et al. (2006). "Site-directed mutagenesis of the VP2 gene of chicken anemia virus affects virus replication, cytopathology and host-cell MHC class I expression."  87(4): 823-831.

Phenix, K. V., et al. (1994). "Transcriptional analysis and genome expression of chicken anaemia virus."  75(4): 905-909.

Pope, C. R. (1991). Chicken anemia agent. Veterinary immunology and immunopathology, 30(1), 51-65.

Pringle, C. (1999). Chicken Anemia Virus. Proceedings of the XI International Congress of Virology on Virus Taxonomy, Sydney, Australia.

Renshaw, R. W., et al. (1996). "A hypervariable region in VP1 of chicken infectious anemia virus mediates rate of spread and cell tropism in tissue culture." 70(12): 8872-8878.

Rijsewijk, F. A., et al. (2011). "Discovery of a genome of a distant relative of chicken anemia virus reveals a new member of the genus Gyrovirus."  156(6): 1097-1100.

Rosario, K., et al. (2017). "Revisiting the taxonomy of the family Circoviridae: establishment of the genus Cyclovirus and removal of the genus Gyrovirus."  162(5): 1447-1463.

Rosenberger, J. and S. Cloud (1988). Characterization of chicken anemia agent (CAA) isolated from Delmarva chickens. 23rd National meeting on Poultry Health and Condemnations. Proceedings.

Rosenberger, J. and S. Cloud (1989). Immunosuppressive aspects of chicken anemia agent (CAA). Proc. 24th National Meeting on Poultry Health and Condemnations, Ocean City, MD. Delmarva Poultry Industry, Georgetown, DE.

Rosenberger, J. K. and S. S. J. A. d. Cloud (1989). "The effects of age, route of exposure, and coinfection with infectious bursal disease virus on the pathogenicity and transmissibility of chicken anemia agent (CAA)." 753-759.

Rosenberger, J. K. and S. S. J. A. d. Cloud (1989). "The isolation and characterization of chicken anemia agent (CAA) from broilers in the United States." 707-713.

Santen, V. L. v., et al. (2004). "Pathogenesis of chicken anemia virus: comparison of the oral and the intramuscular routes of infection."  48(3): 494-504.

Schat, K. (2003). Marek’s disease, chicken infectious anemia and immunosuppression: a nasty combination. 15th Annual Australian Poultry Science Symposium, Sydney, Australia.

Schat, K. and V. J. D. o. P. A. F. van Santen, ed. Blackwell Publ. Ames IA (2008). "Chicken anemia virus and other circovirus infections." 211-235.

Schat, K. A., et al. (2011). "Immune complex vaccines for chicken infectious anemia virus."  55(1): 90-96.

Schat, K. J. T. V. (2009). "Chicken anemia virus." 151-183.

Shettima, Y. M., et al. (2017). "Seroprevalence of chicken infectious anemia virus infection among some poultry species in Maiduguri, Nigeria."  4(4): 385-389.

Shivaprasad, H. and B. Lucio (1994). "Anemia Infecciosa Aviar." 

Smyth, J., et al. (2006). "Chicken anaemia virus inoculated by the oral route causes lymphocyte depletion in the thymus in 3-week-old and 6-week-old chickens." 35(3): 254-259.

Smyth, J., et al. (1993). "A sequential histopathologic and immunocytochemical study of chicken anemia virus infection at one day of age." 324-338.

Smyth, J. and K. J. I. d. o. p. I. G. Schat, ed. Servet, Zaragoza, Spain (2013). "Virus-induced immunosuppression: chicken infectious anemia." 91-121.

Soine, C., et al. (1993). "Determination of the detection limit of the polymerase chain reaction for chicken infectious anemia virus." 467-476.

Sommer, F. and C. J. A. d. Cardona (2003). "Chicken anemia virus in broilers: dynamics of the infection in two commercial broiler flocks."  47(4): 1466-1473.

Sonie, C. (1994). Sequence analysis of cell-culture and non-cell culture-adapted strains of chicken infectious anemia virus. Proceedings of the International Symposium on Infectious Bursal Disease and Chicken Infectious Anaemia. Rauischholzhausen, Germa.

Tan, C., et al. (2020). "Epidemiology, molecular characterization, and recombination analysis of chicken anemia virus in Guangdong province, China."  165(6): 1409-1417.

Taniguchi, T., et al. (1982). "Hematopathological changes in dead and moribund chicks induced by chicken anemia agent." 22(2): 61-69.

Tantaswasdi, U., et al. (1996). "Virological and serological studies of chicken anaemia virus in Thailand."  47(3/4): 45-56.

Todd, D., et al. (1990). "Purification and biochemical characterization of chicken anaemia agent."  71(4): 819-823.

Toro, H., et al. (2006). "Serological evidence of chicken infectious anemia virus in the United States at least since 1959."  50(1): 124-126.

Toro, H., et al. (2001). "Vertical induction of the inclusion body hepatitis/hydropericardium syndrome with fowl adenovirus and chicken anemia virus." 215-222.

Toro, H., et al. (2006). "Epidemiological and experimental evidence for immunodeficiency affecting avian infectious bronchitis."  35(6): 455-464.

Trinh D. Q., Ogawa, H., Bui, V. N., Nguyen, T. T. H., Gronsang, D., Baatartsogt, T., Kizito, M. K., Aboelkhair, M., Yamaguchi, S., Nguyen, V. K. & Imai, K. (2015). Development of a blocking latex agglutination test for the detection of antibodies to chicken anemia virus, Journal of Virological Methods, 221: 74-80

v Bülow, V., et al. (1985). "Untersuchungen über den Erreger der infektiösen Anämie bei Hühnerküken (CAA) in vitro: Vermehrung, Titration, Serumneutralisationstest und indirekter Immunfluoreszenztest."  32(1‐10): 679-693.

v Bülow, V., et al. (1986). "Erhöhte Pathogenität des Erregers der aviären infektiösen Anämie bei Hühnerküken (CAA) bei simultaner Infektion mit Virus der marekschen Krankheit (MDV), Bursitisvirus (IBDV) oder Reticuloendotheliosevirus (REV)."  33(1‐10): 93-116.

Van Dong, H., et al. (2019). "Chicken anemia virus in northern Vietnam: molecular characterization reveals multiple genotypes and evidence of recombination."  55(5): 643-653.

Van Ginkel, F., et al. (2008). "Infectious bronchitis virus in the chicken Harderian gland and lachrymal fluid: viral load, infectivity, immune cell responses, and effects of viral immunodeficiency."  52(4): 608-617.

Vaziry, A., et al. (2011). "Chicken infectious anaemia vaccinal strain persists in the spleen and thymus of young chicks and induces thymic lymphoid cell disorders."  40(4): 377-385.

Vielitz, E., et al. (1991). "Vaccination against infectious anemia of poultry (CAA)--results of field studies." 98(4): 144-147.

Vielitz, E. and H. J. A. P. Landgraf (1988). "Anaemia‐dermatitis of broilers: Field observations on its occurrence, transmission and prevention."  17(1): 113-120.

Vielitz, E., et al. (1987). "Anemia in broilers--development of a vaccine for breeder stock."  34(8): 553-557.

Vielitz, E. and M. Voss (1994). Experiences with a commercial CAV vaccine. Proc Int Symp Infect Bursal Dis Chick Infect Anaemia, Rauischholzhausen, Germany.

Vielitz, E. J. D.-S.-s., Hameln, West Germany (1989). "Avian infectious anemia-occurrence, transmission and prevention." 78-99.

Von Bulow, V. and K. Schat (1997). Chicken infectious anemia: In: Diseases of Poultry, Iowa: Iowa State University Press Ames.

Wanasawaeng, W., et al. (2013). "Molecular Characterization of chicken infectious anemia virus outbreaks during 2008-2011 in Thailand."  43(4): 497.

Wani, M., et al. (2013). "Molecular detection and epidemiology of chicken infectious anaemia virus in India." 3(4): 145-151.

Wani, M. Y., et al. (2016). "Impact of virus load on immunocytological and histopathological parameters during clinical chicken anemia virus (CAV) infection in poultry."  96: 42-51.

Wunderwald, C. and R. K. J. A. P. Hoop (2002). "Serological monitoring of 40 Swiss fancy breed poultry flocks."  31(2): 157-162.

Yamaguchi, S., et al. (2001). "Identification of a genetic determinant of pathogenicity in chicken anaemia virusThe GenBank accession numbers of the sequences reported in this paper are AB031296 and AB046587–AB046590."  82(5): 1233-1238.

Xie, Z., et al. (2014). "Full-genome sequence of chicken anemia virus strain GXC060821, isolated from a Guangxi Sanhuang chicken."  2(1).

Yao, S., et al. (2019). "Molecular epidemiology of chicken anaemia virus in sick chickens in China from 2014 to 2015." 14(1): e0210696.

Yuasa, N., et al. (1985). "Survey of antibody against chicken anaemia agent (CAA) by an indirect immunofluorescent antibody technique in breeder flocks in Japan."  14(4): 521-530.

Yuasa, N., et al. (1987). "Aetiological examination of an outbreak of haemorrhagic syndrome in a broiler flock in Japan."  16(3): 521-526.

Yuasa, N. and K. J. A. P. Imai (1986). "Pathogenicity and antigenicity of eleven isolates of chicken anaemia agent (CAA)." 15(4): 639-645.

Yuasa, N., et al. (1983). "Isolation of chicken anemia agent with MDCC-MSB1 cells from chickens in the field."  23(3): 75-77.

Yuasa, N., et al. (1983). "Distribution of chicken anemia agent (CAA) and detection of neutralizing antibody in chicks experimentally inoculated with CAA."  23(3): 78-81.

Yuasa, N., et al. (1980). "Effect of infectious bursal disease virus infection on incidence of anemia by chicken anemia agent." 202-209.

Yuasa, N., et al. (1979). "Isolation and some characteristics of an agent inducing anemia in chicks." 366-385.

Yuasa, N., et al. (1976). "Isolation of a reticuloendotheliosis virus from chickens inoculated with Marek's disease vaccine."  16(4): 141-151.

Yuasa, N. and I. J. N. I. o. A. H. Q. Yoshida (1983). "Experimental egg transmission of chicken anemia agent." 23(3): 99-100.

Yuasa, N. J. N. I. o. A. H. Q. (1983). "Propagation and infectivity titration of the Gifu-1 strain of chicken anemia agent in a cell line (MDCC-MSB1) derived from Marek's disease lymphoma." 23(1): 13-20.

Zhang, W., et al. (2014). "What is for dinner? Viral metagenomics of US store bought beef, pork, and chicken." 468: 303-310.

Zhang, X., et al. (2015). "Assessing the efficacy of an inactivated chicken anemia virus vaccine."  33(16): 1916-1922.

* Nguyen Thanh Lam, DVM., MSc., PhD

Department of Veterinary Medicine, College of Agriculture, Can Tho University

Address: Campus II, 3/2 street, Ninh Kieu district, Can Tho city, Viet Nam

Phone: +84 (0) 939-468-525

Email: ntlam@ctu.edu.vn