Introduction
Chicken infectious anemia (CIA) or chicken anemia (CA) is an emerging immunosuppressive disease of great importance to the poultry industry [1,2]. The immunosuppression associated with CIA results in decreased response to subsequent vaccination and increased susceptibility to secondary infections [1]. The disease is caused by the chicken infectious anemia virus (CIAV), which is the only member of the genus gyrovirus belonging to the family Anelloviridae [3,4]. Chicken infectious anemia virus (CAV) was initially identified during the investigation of contaminated Marek’s disease (MD) vaccine [4,5]. Toro et al. [6] found antibodies to CIAV in chicken sera stored since 1959, suggesting that the CIA virus had been in circulation before its initial isolation. The control of CIA involved strict biosecurity measures and vaccination of birds against immunosuppressive pathogens such as Marek’s disease virus, infectious bursal disease virus, and avian adenovirus [7]. Also, proper vaccination of breeder stocks against CIAV to confer maternal immunity to their progenies was suggested [8,9,10].
The city of Jos, located in Plateau State, Nigeria, boasts a vibrant poultry industry characterized by numerous live bird markets (LBMs). These markets serve as crucial hubs for the trade and exchange of poultry, facilitating the movement of birds across various regions. The dynamics of LBMs, with their diverse poultry populations and frequent bird interactions, create an environment conducive to the transmission of infectious agents, including CIAV. Despite the potential threat posed by CIAV, there is a noticeable gap in our understanding of its prevalence and distribution within LBMs in Jos, Plateau State. This research aimed to address this gap by conducting a seroprevalence study of chicken infectious anemia virus antibodies in chickens within the LBMs of Jos and by exploring the prevalence and distribution patterns of CIAV antibodies.
Materials and Methods
Ethics statement
The use of animals in this study was in accordance with the ethics of the Ahmadu Bello University Committee on Animal Use and Care.
Study area
This study was conducted in Jos, the capital of Plateau state, Nigeria, which is composed of two Local Government Areas: Jos-North and Jos-South with a total population of approximately 900,000 residents, as indicated by the 2006 census. Situated between latitudes 9° 45’ 3.4’’N and 10° 0 ’ 2.3’’N and longitudes 8° 45’E and 9° 0 ’ 1.2’’E, the city covers an approximate area of 496.1 km2. Positioned at a high altitude on the Jos Plateau, it boasts an elevation of 4,062 ft (1,217 m) above sea level [11,12]. The avian species commonly encountered in the study area include local and commercial chickens, turkeys, guinea fowls, ducks, geese, and pigeons. Two LBMs, namely Yan Kaji (located at Sarkin Mangu Street off Amingo Junction in Jos-North LGA) and Kugiya LBM (located at Bukuru in Jos-South LGA), were identified for blood sample collection for serological studies.
Sample Size Determination
The sample size was determined using the formula by Thrusfield [13]:
n=Z2 P (1-P)/d2
where n=required sample size,
P=prevalence,
Z=Z value (at 95 %confidence level=1.96),
d=precision,
P=89% [14],
d=4.6%
Sample size (n)=176
A total of 176 chickens were sampled for the determination of seroprevalence of CIAV antibodies.
Sample collection
Blood samples (5 ml each) for antibody detection were collected from chickens (88 local and 88 exotic) in LBMs in Jos-North (87 chickens) and Jos-South (89 chickens) from February to May 2023 (every 2 weeks) at slaughter point into labeled plain tubes and allowed to clot. Thereafter, the blood was centrifuged, serum harvested into labeled tubes, and stored at −20°C in a deep freezer at the Laboratory until tested for CIAV antibodies using enzyme-linked immunosorbent assay (ELISA).
Detection of Chicken Infectious Anaemia Virus Antibodies using Enzyme Linked Immunosorbent Assay
Indirect ELISA was used to detect CIAV antibodies in the sera collected from the chickens. The test was performed using a CAV ELISA kit (AsurDxTM Chicken Anemia Virus Antibody Test Kit- Biostone Animal Health LLC, Texas, USA) by following the manufacturers’ instructions. The serum samples to be tested, CAV antigen-coated plates, and the ELISA kit reagents were adjusted to room temperature prior to the test. Also, the serum samples were diluted in 50 folds (1:50) by adding 5 µl of test serum to 245 µl of avian sample diluent provided prior to the assay. To each antigen-coated plate, 90 µl of CAV Assay diluents were added, followed by 10 µl of the positive and negative controls in duplicates and each diluted serum was then dispensed into the subsequent wells of the plate. The solution in the plates was mixed by gentle rocking of the plates for 1 minute, then the plates were covered with foil and were incubated for 30 minutes at room temperature. This was followed by washing each well 5 times using 250 µl of wash solution. After the final wash, the plates were tapped on paper towels to remove residual wash fluid. 100 µl of HRP- conjugated antibody solution reagent was then added to each well and the plates were incubated at room temperature for 30 minutes. Each well was washed again 5 times with 350 µl of wash solution and after the final wash, the plates were tapped on paper towels to remove residual wash fluid. 100 µl of TMB substrate was dispensed into each well, the plate was covered with foil and incubated at room temperature for 15 minutes. Finally, 100 µl of stop solution was dispensed into each well to stop the reaction. The optical density (OD) of the wells was measured and recorded at 620 nm wavelength using ELISA microtitre plate reader.
The relative level of antibody in the sample was determined by calculating the percentage positivity (PP) as shown below:
Results were considered valid when: mean OD620 positive control—mean OD620 negative control >0.1; the mean OD620 positive control ≥ 0.2. chicken serum samples with PP < 25 % were considered negative for CAV antibodies while those with PP ≥ 25 % were considered positive for CAV antibodies.
Data analyses
The prevalence of CIAV antibodies was presented as percentages. The chi-square test was used to compare between type of chickens and the location of chickens in this study and the results were presented using tables. Statistical Package for Social Science (IBM SPSS Statistics 25) was used for these analyses.
Results
Overall seroprevalence
A total of 176 chickens were sampled within Jos, out of which 87 were from Jos North and 89 from Jos South; 88 chickens each were local and commercial (Table 1).
Table 1.
Seroprevalence of CIAV in chickens sampled at LBMs within Jos, Nigeria.
| Number of chicken sampled | Number positive for CIAV antibodies | Seroprevalence (%) | X2 | p value | ||
|---|---|---|---|---|---|---|
| Location | Jos North | 87 | 45 | 51.7 | 21.48 | 0.000* |
| Jos South | 89 | 75 | 84.3 | |||
| Type of chicken | Local | 88 | 66 | 75.0 | 3.771 | 0.075 |
| Commercial | 88 | 54 | 61.4 | |||
| Overall | 176 | 120 | 68.2 |
*showed significant difference at p < 0.05.
Out of the 176 samples, 120 (68.2%) were seropositive for chicken infectious anemia virus (CIAV) antibodies, with 45 (51.7%) from Jos North and 75 (84.3%) from Jos South. Based on the type of chicken, 66 (75.0%) local and 54 (61.4%) exotic chickens were seropositive for CIAV antibodies. The CIAV antibodies seroprevalence showed significant (p < 0.05) difference between Jos North and South, but did not differ significantly (p > 0.05) between the local and exotic chickens (Table 1).
Seroprevalence based on location
Out of the 87 chickens sampled for CIAV antibodies in Jos North, 23 (52.3%) local and 22 (51.7%) exotic chickens were seropositive (Table 2). In Jos South, 43 (97.7%) local and 32 (71.1%) exotic chickens were seropositive for CIAV antibodies out of the 89 chickens sampled. The CIAV antibodies seroprevalence showed no significant (p > 0.05) difference between local and exotic chickens in Jos North but were significantly different (p < 0.05) between the local and exotic chickens in Jos South (Table 2).
Table 2.
Distribution of CIAV antibodies based on breed of chickens and location of the study area.
| Location | Type of chicken | Number of chicken sampled | Number positive for CIAV antibody | Seroprevalence (%) | X2 | p value |
|---|---|---|---|---|---|---|
| Jos North | Local | 44 | 23 | 52.3 | 0.011 | 1.000 |
| Commercial | 43 | 22 | 51.7 | |||
| Jos South | Local | 44 | 43 | 97.7 | 11.889 | 0.001* |
| Commercial | 45 | 32 | 71.1 |
*showed significant difference at p < 0.05.
Discussion
Chicken infectious anemia virus (CIAV) constitutes one of the most important ubiquitous immunosuppressive pathogens in chickens, which causes great economic losses in the poultry industry globally [10]. The overall seroprevalence of CIAV in local and exotic chickens within Jos Metropolis in this study was 68.2% with 75.0% in local and 61.4% in commercial chickens. This seroprevalence showed variations from those reported in other studies. In Nigeria, previous studies showed a range of seroprevalence: Owoade et al. [15] reported 55% in commercial poultry flocks, and Emikpe et al. [14] reported 88.9% in local chickens in Southwestern Nigeria; Okpanachi [16] found 59.97% in Zaria, and Kadiri [17] recorded 62% in commercial poultry in Kaduna, in Northeastern Nigeria; Shettima et al. [2] reported 38.5% in Maiduguri, Northeastern Nigeria. In addition, Orakpoghenor et al. [18] reported 6.7% of free-living wild birds within Zaria and its surroundings.
Internationally, different seroprevalence rates were reported: Hegazy et al. [19] found 81.7% in Egypt, Bhatt et al. [20] found 86.9% in India, Snoeck et al. [21] reported 36.7% in the Central African Republic and 34.9% in Cameroon in 2009. Also, Sharma et al. [22] reported 92.9% in layers and 58.3% in broilers in Grenada, West Indies, while Kabir et al. [23] reported 83.6% in broiler breeder and layer farms in Bangladesh. Variations in environmental conditions, husbandry practices, sample sizes, species and types of birds, and sero-detection tests used could explain the differences in CIAV seroprevalence across studies [24,25].
Among the sampled chickens, there was a notable difference in CIAV seroprevalence between Jos North and Jos South. Specifically, a higher percentage of chickens from Jos South (84.3%) tested positive for CIAV antibodies compared to those from Jos North (51.7%). This discrepancy in seroprevalence rates between the two regions suggests spatial heterogeneity in the prevalence of CIAV, which could be influenced by factors such as environmental conditions, farming practices, the density of LBMs, and the likelihood of cross contamination in the LBM. Previous studies have demonstrated similar geographic variations in CIAV prevalence, highlighting the role of localized factors in disease transmission dynamics [2,23,25].
In addition, the study showed a significant difference in CIAV seroprevalence between local and exotic chicken populations. This suggests that the risk of CIAV infection may be significantly influenced by the type of chicken, whether local or exotic [2,15]. However, this finding was not uncommon due to the fact that most local chickens and raised under an extensive system of management (with no vaccinations) which constantly exposes them to the virus in the environment, hence they are more likely to transmit the virus to their offspring [26]. This seroprevalence might also be attributed to horizontal transmission of the CIAV resulting from the interaction between different groups of chickens raised under an extensive system of management and/or LBMs. The ability of CIAV to persist in the ovaries of laying hens might be another reason for the seroprevalence observed in the local chickens, especially if the eggs were obtained from infected breeders [26]. Similarly, these might be also the likely reasons for CIAV seroprevalence observed in the exotic chickens in this study.
The slight difference in CIAV seroprevalence between local and exotic chickens could be attributed to various factors, including genetic susceptibility, biosecurity measures, and exposure to infectious agents. In addition, management practices in LBMs, such as overcrowding and poor sanitation, could facilitate the spread of CIAV among susceptible populations. Previous studies have suggested that genetic factors, management practices, and exposure to risk factors in LBMs could contribute to differential susceptibility between local and commercial chicken populations [2,15,23].
Furthermore, the study examined seroprevalence rates within specific geographic regions and chicken types. In Jos North, there was no significant difference in CIAV seroprevalence between local and commercial chickens. However, in Jos South, a significant difference was observed, with a higher percentage of local chickens testing positive for CIAV antibodies than exotic chickens. This finding suggests that the risk of CIAV infection may vary between local and exotic chicken populations depending on the geographic region.
It is interesting to note that the CIAV seroprevalence in chickens does not necessarily relate to overt disease but might be indicative of subclinical infection and/or vaccination [24]. In the case of subclinical infection, this might lead to possible immunosuppressive effects such as increased predisposition to other viral or bacterial infections or suboptimal response to vaccination [1,27]. Thus, in this study, the CIAV seroprevalence observed might be linked to subclinical infection as CIAV vaccination is not a common practice in Nigeria [24]. Another reason for this suggestion might be associated with the fact that the ELISA kit used could not differentiate between antibodies due to infection from that due to vaccination.
Conclusion and Recommendation
The outcomes of this study underscore the importance of continued surveillance and control measures to mitigate the spread of CIAV in poultry populations within Jos and beyond. Chicken infectious anemia virus is circulating in the chicken population in Jos, Plateau State, Nigeria; therefore, poultry farmers should consider implementing strict biosecurity protocols, and vaccination programs, and promoting good farming practices to reduce the risk of CIAV transmission and minimize economic losses associated with the disease. Further research is therefore needed to explore the underlying mechanisms driving differential susceptibility to CIAV among local and commercial chicken populations and to develop targeted interventions to effectively manage the disease.
Acknowledgments
The authors wish to appreciate the members of the Live Bird Marketers Association of Jos-north and Jos-south for their cooperation during sampling as well as Mr Rabiata Shuaib for his technical assistance in the laboratory.
Conflict of interest
All authors declare no conflict of interest.
