Contagious ecthyma in Egypt: Clinical, virological and molecular explorations.

Contagious Ecthyma (CE) is a highly contagious viral disease of sheep and goats with worldwide distribution. The present study aimed to provide a clinical description of contagious ecthyma in four sheep flocks and screen the possible genetic variation in the B2L gene of the detected isolates. Oral lesions were collected and inoculated into chorioallantoic membrane (CAM) of 11 days embryonated chicken eggs. Polymerase chain reaction and direct sequencing of the B2L gene was conducted. Infected sheep exhibited anorexia with a development of nodular lesions evolving in proliferative thick scabs around oral commissures. The inoculated CAM showed small-sized white pock lesions accompanied with thickening of CAM. The partial length of B2L gene (592 bp) was successfully amplified in samples collected from four flocks. The isolated strains belong to genotype I/I and I/II. Sequence and evolutionary analysis illustrate that B2L gene (ORF011) are highly conserved among Orf viruses isolated from different countries.

Assessment of heterologous lumpy skin disease vaccine-induced immunity in pregnant cattle vaccinated at different times of gestation period and their influence on maternally derived antibodies.

The present study aimed to evaluate the cell-mediated and the humoral immune response to Romanian sheep pox vaccine in pregnant cows (n = 12) vaccinated at different times of gestation period and the duration of maternal immunity in calves born to these cows. Evaluation of cellular immunity revealed an increase in lymphocytic proliferation that peaked at 10th day post vaccination (dpv) then gradually decreased. Capripoxvirus-specific antibodies were detected by SNT and ELISA in sera collected from vaccinated dams and also in calves born to these cows. In cows, the antibody titers persisted above the protective level till the seventh month post-vaccination. Passively transferred antibody titers in newly born calves started from the first week after parturition and persisted in a protective level until 2, 3 or 4 months of ages in calves born to cows vaccinated at ≤4th, 4.5:6th, or >6:8th months of pregnancy respectively. Results proved that the average neutralizing antibody titers did not differ between pregnant cows vaccinated at different times of gestation period however, the longevity of maternally derived antibodies depends on the pregnancy stage at which the dam receive vaccine.

Lumpy skin disease outbreaks in Egypt during 2017-2018 among sheeppox vaccinated cattle: Epidemiological, pathological, and molecular findings.

The General Organization of the Veterinary Services in Egypt has adopted a sheeppox vaccination policy to control lumpy skin disease (LSD) in cattle. Over the course of the last two years, recurrent outbreaks were reported, with animals showing severe clinical signs and consequentially higher fatalities than that of cases reported in previous LSD outbreaks. A total of 1050 cattle showing typical clinical signs suggestive of LSD were clinically and pathologically investigated during 2017-2018. Skin nodules were collected and lumpy skin disease virus (LSDV) was screened in collected skin samples using PCR for the RPO-30 gene. Furthermore, the entire P32 protein coding gene was sequenced. Histopathology and immunohistochemistry of the skin nodules were also conducted. The obtained results showed an overall mortality rate of 6.86%. LSDV was confirmed in all the examined nodules as evidenced by immunohistochemistry and positive PCR amplification of the RPO30 gene. Sequencing analysis of the P32 gene revealed a highly conserved nature and genetic stability of the LSDV. The results of the present study show that the current vaccination protocol was not effective for a multitude of reasons. These results also serve as evidence for a strong recommendation of an amendment of homologous vaccine use aside from a complete coverage of cattle populations in order to reduce the incidence of LSD among cattle population in Egypt.

Treatment of Brucellosis in Guinea Pigs via a Combination of Engineered Novel pH-Responsive Curcumin Niosome Hydrogel and Doxycycline-Loaded Chitosan-Sodium Alginate Nanoparticles: an In Vitro and In Vivo Study.

Brucellosis is a common zoonotic infection, particularly in the developing world. The recommended treatment regimens for brucellosis involve the use of two medications such as doxycycline and curcumin in order to avoid relapses and prolonged use of these drugs. Doxycycline has excellent activity in the acidic phagolysosomal environment, while curcumin modulates the immune system function and macrophage activity. Due to the intracellular existence of Brucellae and the different anti-immune mechanisms of Brucella, the treatment of Brucella infection faces many limitations. The design of nanosystems is a promising treatment approach for brucellosis. The objective of this study was to design and evaluate the efficacy of in situ pH-responsive curcumin-loaded niosome hydrogel and doxycycline-loaded chitosan-sodium alginate nanoparticles as chemotherapeutic agents against brucellosis. The prepared formulae showed a spherical nano shape with a slow drug release pattern and small particle size. The prepared formulae were evaluated in vivo using Guinea pigs experimentally infected with Brucella melitensis biovar3. The prepared formula combination gave a significant high reduction rate of Brucella spleen viable count compared with that of untreated controls at p < 0.05. The results showed that the treatment schemes were not fully successful in eliminating Brucella infection in Guinea pigs; however, they significantly (p < 0.05) reduced the viable Brucella count in a shorter time and sub-therapeutic doses. Collectively the novel prepared formulae could be a successful therapy for the effective treatment of brucellosis infection at the recommended therapeutic doses. Graphical abstract.

Capripoxviruses: Exploring the genetic relatedness between field and vaccine strains from Egypt.

BACKGROUND AND AIM: Lumpy skin disease (LSD) and sheep pox are economically important Capripoxvirus-induced diseases of cattle and sheep, respectively. Despite the extensive vaccination program adopted by Egyptian veterinary authorities, LSD and sheep pox are still prevalent and spread throughout the whole country. The current study was designed for molecular characterization and phylogenetic analysis of LSD virus (LSDV) and Sheep pox virus (SPPV) recovered from field cases in Egypt along with vaccinal strains to assess their genetic relatedness. MATERIALS AND METHODS: Skin biopsies were collected from naturally infected cases of LSD in Ismailia (n=3 farms) and Beni-Suef (n=2 farms) Governorates and sheep pox in Beni-Suef (n=1 flock). Virus isolation was carried out on primary ovine fetal kidney and heart cell cultures. DNA was extracted from infected materials (skin lesions, infected cell cultures) as well as LSDV Neethling vaccine strain and Romanian SPPV vaccine strain. Polymerase chain reaction was performed using oligonucleotide primers targeting the entire open reading frame of G protein-coupled receptors (GPCR) gene and gene sequences were analyzed. RESULTS: Virus isolation on primary ovine fetal kidney and heart cell culture revealed a cytopathic effect at the third passage characterized by rounding of infected cells and margination of nuclear chromatin. Comparative sequence analysis of GPCR gene revealed that Egyptian LSDV isolated from Ismailia and Beni-Suef shared 99:100% nucleotide and amino acid (AA) identities with each other. In comparison to the vaccinal strains, Egyptian LSDV isolates shared 98:99 nucleotide and AA identities with LSDV Neethling vaccine strain and 93:94% with SPPV Romanian vaccine strain. No differences at the nucleotide or AAs were observed between the SPPV vaccine and virulent strains (100% identity). Phylogenetic analyses revealed that LSDV Neethling vaccine strain is more related to field Egyptian LSDV and clustered within the LSDV group while Romanian SPPV vaccine strain clustered in a separate clade with SPPV field isolates. CONCLUSION: Comparative sequencing and phylogenetic analyses of the GPCR gene reveal a minimal genetic variation between LSDV field isolates from different locations and a close relationship between virulent field strains and homologous vaccines.

Evaluation of the General Organization of Veterinary Services control program of animal brucellosis in Egypt: An outbreak investigation of brucellosis in buffalo.

BACKGROUND AND AIM: Brucellosis is a major constraint to livestock production in Egypt as well as many developing countries worldwide. Bovine brucellosis is an economically important disease with reproductive failure as a principal manifestation resulting in abortion, premature birth and decreased milk production in females, and orchitis and epididymitis in males. In spite of the efforts of Egyptian veterinary services to overcome brucellosis, the disease is still prevalent in both animals and humans and represents one of the most important public health hazards in Egypt. The aim of the present work was to investigate the efficacy of the control program implemented by the General Organization of Veterinary Services in Brucella infected buffalo farm on serological, molecular, cultural, and histopathological basis. Brucella melitensis biovar 3 was recovered from 6 buffalo-cows. MATERIALS AND METHODS: Blood samples were collected from a total of 750 non-vaccinated lactating buffalo-cows. These animals were proved positive for Brucella by the Egyptian brucellosis national program. Sera were tested using buffered acidified plate antigen test and rose Bengal test as screening tests and complement fixation test as a confirmatory test. Positive animals were separated for slaughtering under the supervision of the Egyptian veterinary authorities. Remaining animals were tested every 3 weeks with slaughtering of positive cases and this continued until the remaining animals revealed three successive negative serological tests. Different lymph nodes (prescapular, prefemoral, mediastinal, retropharyngeal, and supramammary) were collected from 11 Brucella seropositive buffalo-cows slaughtered after being confirmed serologically as Brucella infected cases. Samples were collected and processed for bacterial isolation and nucleic acid detection using polymerase chain reaction (PCR). Parts of these specimens were fixed in 10% neutral buffered formalin for 48 h then processed by paraffin embedding technique. RESULTS: "Test and slaughter" policy was applied on Brucella infected dairy buffalo farm. The program continued for 6 months with slaughtering of positive cases until the herd was proved Brucella free. B. melitensis biovar 3 could be recovered from six buffalo-cows. Universal PCR confirmed Brucella on genus level and Bruce-ladder multiplex, PCR confirmed the presence of B. melitensis on the species level. Histopathological examination of Brucella-infected lymph nodes revealed massive rarified and depleted lymphoid areas of both sub-capsular and deep cortical lymphoid follicles, macrophage cells granulomatous reaction, as well as fat, infiltrates, and chronic vasculitis. The chronic nature of Brucella lesions has been confirmed in this study as indicated by the chronic vasculitis and collagen deposition. CONCLUSION: Freedom status from brucellosis in this study required 6 months which are considered long time allowing the spread of infection to other localities especially under unhygienic conditions, husbandry system favoring mixed populations of different ages, sex, aborted and pregnant, and lack of controlled movement of animals. Therefore, effective control of animal brucellosis requires surveillance to identify infected animal herds, elimination of the reservoirs, and vaccination of young heifers. B. melitensis biovar 3 is the cause of the Brucella outbreak in buffalo which still remains the prevalent type of Brucella in Egypt. The disease runs a chronic course allowing further spread of infection.

Evidence of intrauterine transmission of lumpy skin disease virus.

The current study describes the clinical, histopathological, molecular and serological diagnosis of lumpy skin disease (LSD) in a premature 1-day old calf that has been delivered from a cow that exhibited signs of LSD during the seventh month of pregnancy. The calf showed generalized skin lesions accompanied with signs of immaturity and died 36 h after birth. Postmortem and histopathological examinations revealed the involvement of multiple tissues. The presence of Neethling virus DNA in tissues was confirmed by polymerase chain reaction (PCR) and gene sequencing. Results of ELISA and serum neutralization test (SNT) confirmed that the calf had developed precolostral serum antibodies to LSD virus indicating in utero virus transmission. All tested sera collected from animals located in the same area were serologically positive, indicating exposure to LSD virus.