Description of the study area
The study was conducted in Arero district of the Borana zone, Oromia Regional State of Ethiopia. Arero district is geographically located 4°45′0′N and 38°49′0′E at a distance of 650 km south of Addis Ababa. The area is bordered on the southwest by Dire, on the west by Yabelo, on the north by Bule Hora, on the northeast by the Guji Zone, on the east by the Somali Region, and on the south by Moyale (Olani et al. 2016). The annual average temperature and rainfall are 19 °C and 716 mm, respectively. Animal husbandry in the area is characterized by an extensive pastoral production system with seasonal migration. Camels and cattle are the key livestock species in the area (Faye 2015; Mirkena et al. 2018). As aridity gradually increases and drought is a recurrent phenomenon in the area, the principal stock is shifting from cattle to camels (Dawo 2010).
Study methods and sample size determination
The study employed a cross-sectional study design (November 2013–April 2014). Arero district was selected because of its camel production potential and easy access to a major road. Three pastoral associations (PAs) were randomly included from the sampled district (i.e., Haro-Dimtu, Kaarra-Gumaata and Silala PAs). A total of 129 volunteer households were participated in this study. The sample size for respondents in the house-to-house interviews was determined using the formula (n = 0.25/SE2), at the standard error (SE) of 0.044 and 95% confidence level (Arsham et al. 2007).
A standardized, structured questionnaire was used to collect information relevant to the study objectives, such as age structure of the respondents, herd size and herding experience, CCE incidences in the past year (a year preceding the start of the study), age-wise morbidity and mortality rates attributed to CCE, seasonality of the outbreak, and opinion of camel owners on plant browse that are potentially associated with CCE occurrence. Herders’ ability to identify CCE infection from other diseases with similar clinical signs and symptoms was cross-checked by enquiring about the clinical signs of the diseases. For those who mentioned clinical signs shared by the diseases easily confused with CCE such as, Warts, the interviewers reviewed the clinical signs of CCE with camel owners to verify that the respondent had understood the disease correctly. One animal health assistant and a traditional healer were interviewed regarding the local name of CCE in each of the selected PAs. Some of the information collected during interviews was supported by field observation.
Sample collection and sampling procedures
The protocol for field studies and collection of animal samples was carried out in accordance with the ethics guideline of Jimma University College of Agriculture and Veterinary Medicine.
Fourteen (14) skin scrapings were collected from camels showing suspected clinical signs of PPV infection. Samples were immediately transferred into a cold box and transported to the National Veterinary Institute (NVI) of Ethiopia under the cold-chain system. The samples were then kept at − 20 °C until laboratory analysis.
Viral isolation on cell culture
Skin scraping samples were washed three times with sterile PBS containing antibiotics and antifungal and ground using a sterile pestle and mortar. The supernatant (0.5 mL) was inoculated onto a confluent monolayer of Vero cells grown in a 25 cm2 tissue culture flask containing 10 mL Glasgow Minimum Essential Medium (Sigma-Aldrich) supplemented with 2% fetal calf serum (Gibco). The inoculated cultures were incubated at 37 °C, 5% CO2 and observed daily for the appearance of virus-induced cytopathic effects (CPEs). Samples were considered negative when no CPE was observed following three blind passages (Gelaye et al. 2016a, b; Khalafalla et al. 2015).
Polymerase chain reaction (PCR)
After isolating genomic DNA from the virus, B2L gene was amplified using forward primers (5′-TGA GCT GGT TGG CGC TGT CCT-3′) and reverse primers (5′-CGC AGA CGT GGC TCA GTA CGT-3′). The reaction set up was prepared as follows: 5× standard reaction buffer (5 μl), 2 mM dNTPs (0.5 μl), 500 nM forward primer (1.25 μl), 500 nM reverse primer (1.25 μl), template DNA (5 μl), 2.5 U Taq DNA Polymerase (0.25 μl), nuclease-free water (to 25 μl). The thermal profile was set as follows: Initial denaturation (94 °C, 5 min, 1× cycle), Denaturation (94 °C, 1 min), Annealing (55 °C, 60 s, 35×), Extension (68 °C, 70 s), Final extension (68 °C, 5 min, 1×) (Khalafalla et al. 2020; Tedla et al. 2018).
Data collection and analysis
A database was constructed in a Microsoft Excel® to store the data. Analysis was performed using Statistical Package for Social Science (SPSS 2007 version 20) software. Descriptive (proportion) and inferential (logistic regression model) statistics were used to analyze survey findings. Potential risk factors associated with CCE occurrence were assessed by using a logistic regression model and odds ratio (OR) estimate was used to determine the strength of association between the risk factors (independent variables) and disease (dependent variable). In all the analyses, confidence levels at 95% and a p < 0.05 were used for statistical significance test.
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