First molecular detection of hepatitis E virus genome in camel and pig faecal samples in Ethiopia.

BACKGROUND: Hepatitis E is an enteric and zoonotic disease caused by hepatitis E virus (HEV) that is mainly transmitted via the faecal-oral route through contaminated food or the environment. The virus is an emerging infectious agent causing acute human infection worldwide. A high seroprevalence of the disease was reported in pregnant women in Addis Ababa, Ethiopia, raising significant public health concern. The presence of HEV specific antibodies were also reported in dromedary camels in the country; however, the infectious virus and/or the viral genome have not been demonstrated to date in animal samples. METHODS: To address this gap, a total of 95 faecal samples collected from both apparently healthy pigs of uncharacterised types (50 samples) in Burayu and Addis Ababa areas and camels (Camelus dromedarius, 45 samples) in west Hararghe were screened for the presence of HEV genome using universal primers in a fully nested reverse transcription polymerase chain reaction (nRT-PCR). The protocol is capable of detecting HEV in faecal samples from both pigs and camels. RESULTS: The nRT-PCR detected HEV genes in six (12%) pig faecal samples and one camel sample (2.2%). Therefore, the results indicate that HEV is circulating in both pigs and camels in Ethiopia and these animals and their products could serve as a potential source of infection for humans. CONCLUSION: The detection of HEV in both animals could raise another concern regarding its public health importance as both animals' meat and camel milk are consumed in the country. Further studies to determine the prevalence and distribution of the virus in different animals and their products, water bodies, food chain, and vegetables are warranted, along with viral gene sequencing for detailed genetic characterisation of the isolates circulating in the country. This information is critically important to design and institute appropriate control and/or preventive measures.

Prediction and characterization of novel epitopes of serotype A foot-and-mouth disease viruses circulating in East Africa using site-directed mutagenesis.

Epitopes on the surface of the foot-and-mouth disease virus (FMDV) capsid have been identified by monoclonal antibody (mAb) escape mutant studies leading to the designation of four antigenic sites in serotype A FMDV. Previous work focused on viruses isolated mainly from Asia, Europe and Latin America. In this study we report on the prediction of epitopes in African serotype A FMDVs and testing of selected epitopes using reverse genetics. Twenty-four capsid amino acid residues were predicted to be of antigenic significance by analysing the capsid sequences (n = 56) using in silico methods, and six residues by correlating capsid sequence with serum-virus neutralization data. The predicted residues were distributed on the surface-exposed capsid regions, VP1-VP3. The significance of residue changes at eight of the predicted epitopes was tested by site-directed mutagenesis using a cDNA clone resulting in the generation of 12 mutant viruses involving seven sites. The effect of the amino acid substitutions on the antigenic nature of the virus was assessed by virus neutralization (VN) test. Mutations at four different positions, namely VP1-43, VP1-45, VP2-191 and VP3-132, led to significant reduction in VN titre (P value = 0.05, 0.05, 0.001 and 0.05, respectively). This is the first time, to our knowledge, that the antigenic regions encompassing amino acids VP1-43 to -45 (equivalent to antigenic site 3 in serotype O), VP2-191 and VP3-132 have been predicted as epitopes and evaluated serologically for serotype A FMDVs. This identifies novel capsid epitopes of recently circulating serotype A FMDVs in East Africa.

Assessment of Humoral Immune Response in Pre- and Post-Vaccinated Cattle Against Lumpy Skin Disease.

Introduction: Lumpy skin disease (LSD) is viral disease affecting cattle production and productivity in Ethiopia. As a prevention method, vaccinations have been used for a long period with a questionable output due to the existence of LSD outbreaks in vaccinated herds in different parts of Ethiopia. Methods: A longitudinal study was performed from October 2019 to April 2020 with the objective of assessing the humoral immune response of cattle with a serum neutralization test (SNT) from different management systems in central Ethiopia. In this study, theserum was collected from 113 cattle (extensive (60/113) and intensive (53/113) management systems) before and after vaccination. Results and Discussion: From collected sera, a limited number of cattle had seroconversion before vaccination (7.08%). On the other hand, it is obvious the seroconversion rises post vaccination. Accordingly, seroconversion starts to increase after a week (8.85% at 7 dpv) post-vaccination which proceeds to significantly increase at 30 days post vaccination (dpv) (41.65% (25/60)). Furthermore, the risk factor study before and after vaccination showed intensively managed cattle with significantly higher levels of antibody titer at 7 dpv (OR = 1.17; 95% CI = 0.22, 6.2; p = 0.016) and 30 dpv (OR = 3.67; 95% CI = 1.1, 12.29; p = 0.035) compared with that of extensively managed cattle. The other animal-related risk factor that showed a significant difference was breeds and a specific age group ([4½, 7] years) at 15 dpv (OR = 6.69; 95% CI = 2.02, 22.08; p = 0.002) and 30 dpv (OR = 4.24; 95% CI = 1.22, 14.71; p = 0.023); respectively. Conclusion: This study showed an overall lower antibody detection across the study, posing a question on the current LSD-vaccine efficacy. Therefore, a circulating strain of LSDV should be cross-checked with the vaccine strain and adaptations should be made from it.

Combined Adjuvant Formulations Enhanced an Immune Response of Trivalent Foot and Mouth Disease Vaccine in Cattle.

Introduction: Foot-and-mouth disease is globally one of the most economically important viral diseases of cloven-hoofed animals that can be controlled by different strategies, where vaccination plays an important role. Selection of adjuvant added to vaccine preparation is crucial in ensuring the protective effect of the vaccine. Aluminum hydroxide gel mixed with saponin (AS) is widely used adjuvant, with its suboptimal immune response in FMD vaccine. The present study was undertaken to evaluate different ingredients of adjuvants for inactivated trivalent (A, O and SAT 2) FMD vaccine and to demonstrate the effect of booster dose in cattle. Methods: Cattle were grouped into five; four experimental and one control, with six animals in each group and immunized with trivalent vaccine with various formulations of adjuvants. Immune response was measured using Solid Phase Competitive Enzyme Linked Immune Sorbent Assay (SPCE). Results: The antibody level in cattle immunised with a vaccine formulation containing a mixture of aluminum hydroxide gel and saponin (AS) were significantly lower than AS boosted group for the three serotypes (p<0.05, t-test), which directs the need for booster dose. Whereas the antibody response in the AS + oil group was higher followed by oil alone. The AS preparation with a booster dose has shown better immune response compared to the group without. Conclusion: The findings of this study could suggest that oil based and AS with oil could replace the conventional aluminum hydroxide gel and saponin adjuvants in FMD vaccine preparations. Challenge test was not successful indicating the need for further research on the virus infectivity.

Genogrouping of Infectious Bursal Disease Viruses Circulating in Ethiopian Chickens: Proposal for Assigning Very Virulent Strains in the Country into New Sub Genogroup 3d.

INTRODUCTION: In 2017 infectious bursal disease viruses (IBDVs) were reclassified into genogroups based on nature of clustering on a phylogenetic tree constructed using VP2 gene sequence data rather than according to their pathotype and/or antigenic types. Ethiopian IBD viruses were not reclassified according to the proposed genogrouping. METHODS: In order to genogroup the Ethiopian IBDVs, available VP2 gene sequences data together with reference strain sequences were retrieved from GenBank and genogrouped as recently recommended based on evolutionary tree reconstruction and determination of their clustering on the phylogenetic tree. RESULTS: The Ethiopian IBDVs were grouped into genogroups 1 and 3 that antigenically represent classically virulent and very virulent IBDVs, respectively. The genogroup 1 IBDVs were clustered with the vaccine strain while the genogroup 3 viruses were clustered with four known viruses belonging to sub-genogroup 3a and sub-genogroup 3b. Almost half of the Ethiopian IBDVs reported did not cluster with the specific sub-groups of genogroup 3; rather, the isolates were clustered differently suggesting they deserve a different sub-genogroup tentatively proposed as 3d. The two genogroups observed based on clustering on a phylogenetic tree were supported by corresponding deduced amino acid changes in similar positions in VP2 sequences. In addition, virulence marker amino acid genes coupled with second major hydrophilic region (amino acid positions 314-325) were predicted in these sequences that could be responsible for the occurrence of IBD outbreaks. CONCLUSION: A new sub-genogroup of IBDVs, 3d, were observed in the sequences that could be one of the reasons for the frequent occurrence of IBD outbreaks and questions the protective potential of the existing vaccine. To institute disease control in the country, the effectiveness of the vaccine in use needs to be assessed in vivo against both genogroups 1 and 3 viruses and all three sub-genogroup 3 viruses circulating in the country.

Rotavirus in Calves and Its Zoonotic Importance.

Rotavirus is a major pathogen responsible for diarrheal disease in calves, resulting in loss of productivity and economy of farmers. However, various facets of diarrheal disease caused by rotavirus in calves in the world are inadequately understood, considering that diarrheal disease caused by rotavirus is a vital health problem in calves that interrupts production benefits with reduced weight gain and increased mortality, and its potential for zoonotic spread. The pathological changes made by rotavirus are almost exclusively limited to the small intestine that leads to diarrhea. It is environmentally distributed worldwide and was extensively studied. Reassortment is one of the important mechanisms for generating genetic diversity of rotaviruses and eventually for viral evolution. So, the primary strategy is to reduce the burden of rotavirus infections by practicing early colostrum's feeding in newborn calves, using vaccine, and improving livestock management. Rotaviruses have a wide host range, infecting many animal species as well as humans. As it was found that certain animal rotavirus strains had antigenic similarities to some human strains, this may be an indication for an animal to play a role as a source of rotavirus infection in humans. Groups A to C have been shown to infect both humans and animals. The most commonly detected strains in both human and animals are G2, G3, G4, and G9, P [6]. Therefore, this review was made to get overview epidemiology status and zoonotic importance of bovine rotavirus.

Molecular Characterization of Lumpy Skin Disease Virus Isolates from Outbreak Cases in Cattle from Sawena District of Bale Zone, Oromia, Ethiopia.

Lumpy skin disease (LSD) is a viral disease caused by LSD virus and is one of the most economically significant transboundary and emerging diseases of cattle. LSD causes considerable economic losses due to emaciation, damage to hides, infertility, and loss of milk production. In Ethiopia, the disease is distributed almost in all regions and is regarded as one of the most economically important livestock diseases in the country. An outbreak investigation of the disease was monitored from October 2016 to April 2017 in southern pastoral areas of Bale Zone, Oromia, Ethiopia. In December 2016, LSD outbreak occurred in Sawena district of Bale Zone, from which necessary biopsy samples were collected from actively infected animals for the purpose of virus isolation, and characterization using different molecular techniques at National Animal Health and Diagnostic Investigation Center (NAHDIC) of Sebeta, Ethiopia. In addition, clinical examination of infected and in-contact animals was carried out together with a questionnaire survey. Based on the clinical manifestations, LSD was recorded in 18% (94/522) of examined cattle, whereas biopsy samples from 20 clinically positive animals were collected for further laboratory process. The morbidity rate was higher in animals less than two years 28.97% (31/107) than other ages and showed a statistically significant difference with P < 0.05. Female animals showed higher morbidity rate of 20.59% (76/369) than male animals (11.76%) (18/153) with a significant difference at P ≤ 0.003. Mortality rate and case fatality were also significantly higher in young animals than other age groups. Viruses were isolated from both skin biopsies and nasal swabs on Vero cell line. From both skin biopsies and nasal swabs, the virus DNA was identified by amplifying the 172 bp DNA fragment using real-time and conventional PCR. Providing adequate diagnostic facilities, establishing strategic policies for effective control and eradication and awareness creations for communities for early identification or reporting were recommendations made to minimize economic losses of the disease.

Assessment of Major Animal Health Problems and Their Impact on Beef Cattle Production in Doba District of West Harerghe Zone, Ethiopia.

The aim of the current study was to assess the major animal health problems and their impact on beef cattle production in Doba district of West Harerghe Zone, Ethiopia. The study area was purposively selected, and a simple random sampling method was used to selected households' fatteners from each kebele and interviewed using structured questionnaires. The present study showed that the overall prevalence of the diseases was internal and external parasite 93.3%, bloat 53.3%, black leg 71.1%, pasteurolosis 71.8%, wound 71.8%, FMD 22.2%, and anthrax 13.33% which affect fattening cattle, respectively, in the study area. All the respondents (100%) involved in the study were experienced with deworming of their animals to protect from parasites. But, only 46.7% and 42.2% of the respondents have accessed veterinary services with limited regularity and vaccination program, respectively, in the study area. Hence, the beef cattle fatting sector should be supported through considering alleviating the major disease affecting this sector and encouraging the farmers' indigenous knowledge practice with technology.

Immunogenicity and Efficacy Evaluation of Vero Cell-Adapted Infectious Bursal Disease Virus LC-75 Vaccine Strain.

INTRODUCTION: Infectious bursal disease virus (IBDV) is an avian viral pathogen that causes infectious bursal disease (IBD) of chickens. The disease has been endemic in Ethiopia since 2002, and vaccination has been practiced as the major means of disease prevention and control. An IBD vaccine is produced in Ethiopia using primary chicken embryo fibroblast (CEF) cell, which is time-consuming, laborious, and uneconomical. The present study was carried out to develop cell-based IBDV LC-75 vaccine using Vero cells and to evaluate the safety, immunogenicity and protection level. METHODS: Identity of the vaccine seed was confirmed with gene-specific primers using reverse transcription polymerase chain reaction (RT-PCR). Confluent monolayer of Vero cells was infected with vaccine virus and serial passage continued till passage 10. A characteristic virus-induced cytopathic effect (CPE) was observed starting from passage 2 on the third day post-infection. The infectious titer of adapted virus showed a linear increment along the passage level. The virus-induced specific antibody was determined using indirect ELISA after vaccination of chicks through ocular route. RESULTS: The antibody titer measured from Vero cells vaccinated chicks revealed similar level with the currently available CEF cell-based vaccine, hence no significant difference. Chicks vaccinated with Vero cell adapted virus showed complete protection against very virulent IBDV, while unvaccinated group had 60% morbidity and 25% mortality. CONCLUSION: It is concluded that the IBDV vaccine strain well adapted on Vero cells and found to be immunogenic induces antibody development and successfully protects chicks against challenge with the circulating field IBDV isolate. Hence, it is recommended to produce IBD vaccine using Vero cell culture at the industrial scale to conquer the limitations caused by using CEF cells and thus to vaccinate chicks population to protect against the circulating IBDV infection.