First report on molecular characterization and phylogenetic analysis of Reticuloendotheliosis virus in Sudan.

The reticuloendotheliosis virus (REV) group of retroviruses infects a wide range of avian species, including chickens, turkeys, ducks, geese, quail, and prairie chickens. The infection can result in immunosuppression, runting syndrome, high mortality, acute reticular cell neoplasia, or T- and/or B-cell lymphoma. One PCR positive chicken spleen sample obtained in a previous study in addition to one Marek's disease and three fowl pox (FP) vaccine samples were investigated in this study. A PCR assay was performed to detect the presence of REV provirus DNA in these samples. The results indicated the contamination of fowl pox virus and Marek's disease vaccines with REV. In addition, detection of integration of REV inside the genome of fowl pox vaccine was confirmed using primers corresponding to the FPV DNA regions flanking the REV integration site. Alignments of two sequences, one from the spleen tissue and the other from contaminated FP vaccine with REV, with other REV (env) gene sequences obtained from GenBank indicated their high similarity. Furthermore, phylogenetic analysis indicated that the partial part of (env) gene of our two isolates was closely related to variants from India, USA, Taiwan, and China. These results confirmed the contamination of commercial fowl pox and Marek's disease vaccines used in Sudan with REV. Phylogenetic analysis indicated that the partial part of (env) gene sequences from Sudan was closely related to variants from India, USA, Taiwan, and China.

Serological and molecular identification of Reticuloendotheliosis virus (REV) in chickens in Sudan.

BACKGROUND: Reticuloendotheliosis virus (REV) is a gammaretrovirus that belongs to the family of Retroviridae. The infection can result in immunosuppression, runting syndrome, high mortality, acute reticular cell neoplasia or T- and/ or B-cell lymphoma, in a variety of domestic and wild birds. The disease is widespread around the world. No related data have been reported in Sudan about the disease. The present study was conducted to determine the prevalence of REV antibodies and DNA in local and commercial breeds of chickens older than 20 weeks from June 2014 to February, 2017. METHODS: A total of 460 sera samples and 150 (50 liver and 100 spleen) tissue samples were collected from local and commercial breeds of chickens older than 20 weeks and screened for anti-REV antibodies in four states of Sudan using a commercial REV antibody ELISA test kit (IDEXX). Polymerase chain reaction (PCR) was performed to detect REV DNA in tissue samples in Khartoum State. RESULTS: The results revealed that the overall seroprevalence of REV was 74.6% among local and commercial chicken breeds, but in commercial it was 79.5% (190/239) and 69.2% in local breeds (153/221). One hundred and fifty tissue samples of chickens (50 liver, 100 spleen) were tested using PCR for detection of REV using primer sets of the conserved region in envelope glycoprotein (env) gene with a band length of 850 bp. Five out of 50 (10%) liver samples were RE provirus DNA positive detected by PCR, whereas 15 out of 100 (15%) spleen samples were PCR positive. Univariate analysis revealed there was a difference (p ≤ 0.05) between locality and breed of chickens and seropositivity to REV. CONCLUSIONS: The prevalence of the disease was high in Sudan and more studies are needed to evaluate the epidemiology and pathogenesis of the virus.

Theileria lestoquardi in Sudan is highly diverse and genetically distinct from that in Oman.

Malignant ovine theileriosis is a severe tick-borne protozoan disease of sheep and other small ruminants which is widespread in sub-Saharan Africa and the Middle East. The disease is of considerable economic importance in Sudan as the export of livestock provides a major contribution to the gross domestic product of this country. Molecular surveys have demonstrated a high prevalence of sub-clinical infections of Theileria lestoquardi, the causative agent, among small ruminants. No information is currently available on the extent of genetic diversity and genetic exchange among parasites in different areas of the country. The present study used a panel of T. lestoquardi specific micro- and mini-satellite genetic markers to assess diversity of parasites in Sudan (Africa) and compared it to that of the parasite population in Oman (Asia). A moderate level of genetic diversity was observed among parasites in Sudan, similar to that previously documented among parasites in Oman. However, a higher level of mixed-genotype infection was identified in Sudanese animals compared to Omani animals, consistent with a higher rate of tick transmission. In addition, the T. lestoquardi genotypes detected in these two countries form genetically distinct groups. The results of this work highlight the need for analysis of T. lestoquardi populations in other endemic areas in the region to inform on novel approaches for controlling malignant theileriosis.

Survey of causative agents for acute respiratory infections among patients in Khartoum-State, Sudan, 2010-2011.

BACKGROUND: This study was carried out to determine causative agents of acute respiratory illness of patients in Khartoum State, Sudan. METHODS: Four hundred patients experiencing respiratory infections within January-March 2010 and January-March 2011 were admitted at Khartoum Hospital and had their throat swab samples subjected to multiplex real-time RT-PCR to detect influenza viruses (including subtypes) and other viral agents. Isolation, nucleotide sequence and phylogenetic analysis on some influenza viruses based on the HA gene were done. RESULTS: Out of 400 patients, 66 were found to have influenza viruses (35, 27, 2, and 2 with types A, B, C, and A and B co-infections, respectively). Influenza viruses were detected in 28, 33 and 5 patients in the age groups <1, 1-10, and 11-30 years old, respectively but none in the 31-50 years old group. Out of 334 patients negative for influenza viruses, 27, 14, and 2 were positive for human respiratory syncytial virus, rhinovirus and adenovirus, respectively. Phylogenetic tree on influenza A (H1N1) pdm09 subtype shows that Sudan strains belong to the same clade and are related to those strains from several countries such as USA, Japan, Italy, United Kingdom, Germany, Russia, Greece, Denmark, Taiwan, Turkey and Kenya. Seasonal A H3 subtypes have close similarity to strains from Singapore, Brazil, Canada, Denmark, USA and Nicaragua. For influenza B, Sudan strains belong to two different clades, and just like influenza A (H1N1) pdm09 and A H3 subtypes, seem to be part of worldwide endemic population (Kenya, USA, Brazil, Russia, Taiwan and Singapore). CONCLUSIONS: In Sudan, the existence of respiratory viruses in patients with acute respiratory infection was confirmed and characterized for the first time by using molecular techniques.

Attenuation of Theileria lestoquardi infected cells and immunization of sheep against malignant ovine theileriosis.

Malignant ovine theileriosis caused by Theileria lestoquardi is an economically important disease infecting small ruminants in the Sudan. The disease causes massive losses among sheep in many regions of Northern Sudan. The present studies were done to isolate lymphoblastoid cells infected with malignant ovine theileriosis and attenuate them by passage using culture media to develop and produce schizonts candidate vaccine, then test its efficacy and safety by exposing immunized lambs to field challenge in an area endemic with T. lestoquardi. In the present experiments we isolated and established an in vitro culture of T. lestoquardi infected lymphoblast cell line. Long-term culture of T. lestoquardi infected lymphoplastoid cells was shown to result in attenuation of their virulence and lambs inoculated with different doses of such cells at passage 105 exhibited very mild reactions with fever that lasted for 1-5 days and parasitaemia of <0.2%. The experimental lambs immunized with this candidate vaccine were immune and protected when exposed to field challenge in an area endemic of ovine theileriosis, while morbidity and mortality among non-immunized animals reached 76.9% and 46.15%, respectively, and they exhibited the clinical signs of malignant ovine theileriosis that included, high fever, loss of appetite, enlargement of lymph nodes, jaundice, loss of weight and death. The present study demonstrates the efficacy and the safety of this attenuated cell line as a live attenuated candidate vaccine.