First Report on Ovine Paratuberculosis in the Sudan: Diagnosis Using Different Techniques.

Paratuberculosis (PTB) has been reported in the Sudan in cattle and goats for more than 50 years but has never been reported in sheep. However, suspicion of the disease in a breeding flock of sheep in Khartoum North locality was made due to a history of unknown cause of loss of weight. Blood and faecal samples were collected from all animals (N = 59): harvested sera were tested for anti-Mycobacterium avium subsp. paratuberculosis (MAP) antibodies by Enzyme Linked Immunosorbent Assay (ELISA); faeces were screened for acid-fast bacilli by Ziehl-Neelsen staining, tested for MAP DNA by recombinase polymerase amplification (RPA) and some faecal samples were cultured for MAP isolation. Typical MAP acid-fast bacilli were seen in 10.2% (6/59) of the faecal smears, 37.5% of the tested faecal samples (12/32) were positive for MAP DNA and only 3 (5.1%) animals were seropositive for MAP. MAP positive cultures were obtained from 2 out the 6 samples showing typical MAP acid-fast bacilli; the isolates were confirmed by real-time PCR and sequencing. As sheep are animals of utmost economic importance as the main export animals for the country, this first report of ovine PTB warrants special considerations and more investigations for planning control programmes of the disease.

Molecular and serological survey of paratuberculosis in cattle in selected districts of Western Uganda.

Knowledge of Mycobacterium avium subsp. paratuberculosis (MAP) herd infection status is important to plan appropriate control and prevention strategies for Paratuberculosis (PTB); however, in Uganda MAP infection status of most herds is unknown. This study aimed at determining the MAP infection status of cattle herds and the associated risk factors for MAP infection in six western districts of Uganda. The survey covered a total of 93 herds where faecal and blood samples were collected from 1814 cattle. A Recombinase Polymerase Amplification (RPA) and an antibody-based (ELISA) assays were used to test for the presence of MAP DNA in faeces and MAP antibodies in serum, respectively. The apparent cow-level prevalence of MAP infection was 3.2 and 2.7% using ELISA and RPA respectively and the true cow-level prevalence using ELISA and RPA was 4.9 and 3% respectively. A herd-level prevalence of 43% (ELISA) and 40.8% (RPA) and a within-herd prevalence of 3.8 ± 2.1% based on ELISA were obtained. Among the risk factors investigated, long dry spells were significantly associated with high MAP infection (p < 0.05). These results indicate that MAP is actively present in most areas where surveillance was carried out. This poses a serious threat to the livestock industry and potentially to public health as MAP is highly suspected to play a role in the pathogenesis of several diseases in humans. Other areas of the country are to be surveyed as well in order to establish full data on MAP infection status to enable interventions for the control and prevention of the disease.

Metagenomic analysis of bacterial communities of Wadi Namar Lake, Riyadh, Saudi Arabia.

Wadi Namar lake is a new touristic attraction area in the south of Riyadh. Human activities around the lake may lead to changes in water quality with subsequent changes in microenvironment components including microbial diversity. The current study was designed to assess possible changes in bacterial communities of the water at Wadi Namar Lake. Therefore, water samples were collected from three different locations along the lake: L1 (no human activities, no plants), L2 (no human activity, some plants) and L3 (human activities, municipal wastes and some plants). The total DNA of the samples was extracted and subjected to 16S rDNA sequencing and metagenomic analysis; water pH, electrical conductivity (EC), total dissolved solids (TDS) as well as the concentration of Na+1, K+1, Cl-1 and total N were analysed. Metagenomic analysis showed variations in relative abundance of 17 phyla, 31 families, 43 genera and 19 species of bacteria between the locations. Proteobacteria was the most abundant phylum in all locations; however, its highest abundance was in L1. Planctomycete phylum was highly abundant in L1 and L3, while its abundance in L2 was low. The phyla Acidobacteria, Candidatus Saccharibacteria, Nitrospirae and Chloroflexi were associated with high TDS, EC, K+1 and Cl-1 concentrations in L3; various human activities around this location had possibly affected microbial diversity. Current study results help in recognising the structure of bacterial communities at Wadi Namar Lake in relation to their surroundings for planning to environment protection and future restoration of affected ecosystems.

Mycobacterium avium subsp. paratuberculosis and microbiome profile of patients in a referral gastrointestinal diseases centre in the Sudan.

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne's disease in animals with zoonotic potential; it has been linked to many chronic diseases in humans, especially gastrointestinal diseases (GID). MAP has been extensively studied in Europe and America, but little reports were published from Africa. Sudan is a unique country with close contact between humans and livestock. Despite such interaction, the one health concept is neglected in dealing with cases of humans with GID. In this study, patients admitted to the reference GID hospital in the Sudan over a period of 8 months were screened for presence of MAP in their faeces or colonic biopsies. A total of 86 patients were recruited for this study, but only 67 were screened for MAP, as 19 did not provide the necessary samples for analysis. Both real-time PCR and culture were used to detect MAP in the collected samples and the microbial diversity in patients´ faecal samples was investigated using 16S rDNA nanopore sequencing. In total, 27 (40.3%) patients were MAP positive: they were 15 males and 12 females, of ages between 21 and 80 years. Logistic regression analysis revealed no statistical significance for all tested variables in MAP positive patients (occupation, gender, contact with animal, milk consumption, chronic disease, etc.). A unique microbiome profile of MAP-positive patients in comparison to MAP-negative was found. These findings suggest that a considerable proportion of the population could be MAP infected or carriers. Therefore, increase awareness at community level is urgently needed to decrease the risk of MAP at human/animal interface. This study represents the first report of MAP in humans in the Sudan; nevertheless, a better view of the situation of MAP in humans in the country requires a larger study including patients with other conditions.

Mycobacterium avium subsp. paratuberculosis Virulence: A Review.

To propose a solution for control of Mycobacterium avium subsp. paratuberculosis (MAP) infections in animals as well as in humans, and develop effective prevention, diagnostic and treatment strategies, it is essential to understand the molecular mechanisms of MAP pathogenesis. In the present review, we discuss the mechanisms utilised by MAP to overcome the host defense system to achieve the virulence status. Putative MAP virulence genes are mentioned and their probable roles in view of other mycobacteria are discussed. This review provides information on MAP strain diversity, putative MAP virulence factors and highlights the knowledge gaps regarding MAP virulence mechanisms that may be important in control and prevention of paratuberculosis.

Differentiation of Capripox Viruses by Nanopore Sequencing.

The genus capripoxvirus (CaPV), family Poxviridae, includes three virus species: goatpox virus (GPV), sheeppox virus (SPV) and lumpy skin disease virus (LSDV). CaPV causes disease outbreaks with consequent economic losses in Africa and the Middle East. LSDV has recently spread to Southeast Europe. As CaPVs share 96-97% genetic similarity along the length of the entire genome and are difficult to distinguish using serological assays, simple, reliable and fast methods for diagnosis and species differentiation are crucial in cases of disease outbreak. The present study aimed to develop a field-applicable CaPV differentiation method. Nanopore technology was used for whole genome sequencing. A local database of complete CaPV genomes and partial sequences of three genes (RPO30, P32 and GPCR) was established for offline Basic Local Alignment Search Tool (BLAST). Specificities of 98.04% in whole genome and 97.86% in RPO30 gene runs were obtained among the three virus species, while other databases were less specific. The total run time was shortened to approximately 2 h. Functionality of the developed procedure was proved by samples with high host background sequences. Reliable differentiation options for the quality and capacity of hardware, and sample quality of suspected cases, were derived from these findings. The whole workflow can be performed rapidly with a mobile suitcase laboratory and mini-computer, allowing application at the point-of-need with limited resource settings.

Paratuberculosis: The Hidden Killer of Small Ruminants.

Paratuberculosis (PTB) is a contagious and chronic enteric disease of ruminants and many non-ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP), and is characterised by diarrhoea and progressive emaciation with consequent serious economic losses due to death, early culling, and reduced productivity. In addition, indirect economic losses may arise from trade restrictions. Besides being a production limiting disease, PTB is a potential zoonosis; MAP has been isolated from Crohn's disease patients and was associated with other human diseases, such as rheumatoid arthritis, Hashimoto's thyroiditis, Type 1 diabetes, and multiple sclerosis. Paratuberculosis in sheep and goats may be globally distributed though information on the prevalence and economic impact in many developing countries seem to be scanty. Goats are more susceptible to infection than sheep and both species are likely to develop the clinical disease. Ingestion of feed and water contaminated with faeces of MAP-positive animals is the common route of infection, which then spreads horizontally and vertically. In African countries, PTB has been described as a "neglected disease", and in small ruminants, which support the livelihood of people in rural areas and poor communities, the disease was rarely reported. Prevention and control of small ruminants' PTB is difficult because diagnostic assays demonstrate poor sensitivity early in the disease process, in addition to the difficulties in identifying subclinically infected animals. Further studies are needed to provide more insight on molecular epidemiology, transmission, and impact on other animals or humans, socio-economic aspects, prevention and control of small ruminant PTB.

Seroprevalence of Mycobacterium avium subsp. paratuberculosis in Dairy Cattle in Khartoum State, Sudan.

Paratuberculosis, caused by Mycobacterium avium subspecies paratuberculosis (MAP), is a chronic wasting disease mainly of domestic and wild ruminants. It occurs worldwide, causing significant economic losses through decreased productivity, low fertility, increased cull rates and mortality. It is listed by the OIE (World Organization for Animal Health) as a disease of concern to trade in animals. Prevalence of this disease can be studied by detecting anti-MAP antibodies by Enzyme linked immunosorbent Assay (ELISA). The aim of this study was to investigate the current prevalence of MAP infection in cattle in Khartoum State. The overall apparent prevalence of MAP infection was found to be 6.3% and 18.9% at animal and herd levels, respectively. All seropositive animals were cross-bred females of good body condition; most of them (>90%) were >3 years old and >50% were from medium-sized herds in Omdurman. No significant association (p > 0.05) was found between seropositivity and animal herd size. The prevalence of MAP infection in Khartoum State is still low to medium compared to other parts of the world, but it is comparable to those reported from other African countries. Further studies with the view of designing nationwide surveys in domestic ruminants and camels in other states of the country are needed for establishing control programmes.

Buffalopox Virus: An Emerging Virus in Livestock and Humans.

Buffalopox virus (BPXV) is the cause of buffalopox, which was recognized by the FAO/WHO Joint Expert Committee on Zoonosis as an important zoonotic disease. Buffalopox was first described in India, later in other countries, and has become an emerging contagious viral zoonotic disease infecting milkers with high morbidity among affected domestic buffalo and cattle. BPXV is a member of the genus Orthopoxvirus and a close variant of the vaccinia virus (VACV). Recent genome data show that BPXV shares a most recent common ancestor of VACV Lister strain, which had been used for inoculating buffalo calves to produce a Smallpox vaccine. Over time, VACV evolved into BPXV by establishing itself in buffaloes to be increasingly pathogenic to this host and to make infections in cattle and humans. Together with the current pandemic of SARS-COV2/COVID 19, BPXV infections illustrate how vulnerable the human population is to the emergence and re-emergence of viral pathogens from unsuspected sources. In view that majority of the world population are not vaccinated against smallpox and are most vulnerable in the event of its re-emergence, reviewing and understanding the biology of vaccinia-like viruses are necessary for developing a new generation of safer smallpox vaccines in the smallpox-free world.

Paratuberculosis: A Potential Zoonosis and a Neglected Disease in Africa.

The Mycobacterium avium subspecies paratuberculosis (MAP) is the causative agent of paratuberculosis, which is an economically important disease of ruminants. The zoonotic role of MAP in Crohn's disease and, to a lesser extent, in ulcerative colitis, the two major forms of idiopathic inflammatory bowel disease (IIBD), has been debated for decades and evidence continues to mount in support of that hypothesis. The aim of this paper is to present a review of the current information on paratuberculosis in animals and the two major forms of IIBD in Africa. The occurrence, epidemiology, economic significance and "control of MAP and its involvement IIBD in Africa" are discussed. Although the occurrence of MAP is worldwide and has been documented in several African countries, the epidemiology and socioeconomic impacts remain undetermined and limited research information is available from the continent. At present, there are still significant knowledge gaps in all these areas as far as Africa is concerned. Due to the limited research on paratuberculosis in Africa, in spite of growing global concerns, it may rightfully be considered a neglected tropical disease with a potentially zoonotic role.

Evaluation of antibiotic-induced behavioral changes in mice.

Gut microbiota (GM) plays a critical role in health maintenance. Previous reports connected GM with metabolic, immunologic and neurologic pathways. The main purpose of the current investigation was to study whether antibiotic-induced disturbances of GM affects psychological or behavioral conditions on mice as animal model. Mice were exposed to clindamycin or amoxicillin, and their behaviors were evaluated. Antibiotic-treated groups displayed reduced recognition memory and increased depression. No significant changes in the locomotor activity and anxiety were observed. Our data suggested that changes in GM composition by antibiotics may lead to the cognitive and behavioral deficit.

Species-Specific Conservation of Linear Antigenic Sites on Vaccinia Virus A27 Protein Homologs of Orthopoxviruses.

The vaccinia virus (VACV) A27 protein and its homologs, which are found in a large number of members of the genus Orthopoxvirus (OPXV), are targets of viral neutralization by host antibodies. We have mapped six binding sites (epitopes #1A: aa 32-39, #1B: aa 28-33, #1C: aa 26-31, #1D: 28-34, #4: aa 9-14, and #5: aa 68-71) of A27 specific monoclonal antibodies (mAbs) using peptide arrays. MAbs recognizing epitopes #1A-D and #4 neutralized VACV Elstree in a complement dependent way (50% plaque-reduction: 12.5-200 µg/mL). Fusion of VACV at low pH was blocked through inhibition of epitope #1A. To determine the sequence variability of the six antigenic sites, 391 sequences of A27 protein homologs available were compared. Epitopes #4 and #5 were conserved among most of the OPXVs, while the sequential epitope complex #1A-D was more variable and, therefore, responsible for species-specific epitope characteristics. The accurate and reliable mapping of defined epitopes on immuno-protective proteins such as the A27 of VACV enables phylogenetic studies and insights into OPXV evolution as well as to pave the way to the development of safer vaccines and chemical or biological antivirals.

Prevalence of bluetongue virus antibodies and associated risk factors among cattle in East Darfur State, Western Sudan.

BACKGROUND: Bluetongue virus (BTV) is an insect-transmitted virus, which causes bluetongue disease (BT) in sheep and a fatal hemorrhagic infection in North American white-tailed deer. However, in cattle the disease is typically asymptomatic and no overt clinical signs of disease appear to be associated with BTV infection. Serological evidence and isolation of different BTV serotypes have been reported in Sudan, however, no information is currently available in regard to previous exposure of Sudanese livestock to BTV infection in East Darfur State, Sudan. AIMS: To determine the prevalence of BTV antibodies and to identify the potential risk factors associated with BTV infection among cattle in East Darfur State, Sudan. METHODS: A total of 224 blood samples were collected randomly from five localities in East Darfur State, Sudan. The serum samples were screened for detection of BTV-specific immunoglobulin G (IgG) antibodies using a competitive enzyme-linked immunosorbent assay (c-ELISA). RESULTS: Serological evidence of BTV infection was observed in 150 out of 224 animals accounting for a 67% prevalence rate among cattle in East Darfur State. Older cattle (>2 years of age) were six times more likely to be infected with BTV (OR = 6.62, CI = 2.87-15.26, p-value = 0.01). Regarding animal source (contact with other herds) as a risk factor, it was shown that cattle purchased from market or introduced from other herds were 3 times at higher risk of being infected with BTV (OR = 3.87, CI = 1.07-13.87, p value = 0.03). Exposure of cattle to the insect vector increased the risk of contracting BTV infection by six times compared to non-exposed cattle (OR = 6.44, CI = 1.53-27.08, p value = 0.01). CONCLUSION: The present study indicated that age, animal source and the intensity of the insect vector are influential risk factors for BTV infection in cattle in the Darfur region. Surveillance for BTV infection should be extended to include other susceptible ruminants and to study the distribution of the insect vectors to better predict and respond to a possible BTV outbreak in the State of East Darfur, Sudan.

First report on circulation of Echinococcus ortleppi in the one humped camel (Camelus dromedaries), Sudan.

BACKGROUND: Echinococcus granulosus (EG) complex, the cause of cystic echinococcosis (CE), infects humans and several other animal species worldwide and hence the disease is of public health importance. Ten genetic variants, or genotypes designated as (G1-G10), are distributed worldwide based on genetic diversity. The objective of this study was to provide some sequence data and phylogeny of EG isolates recovered from the Sudanese one-humped camel (Camelus dromedaries). Fifty samples of hydatid cysts were collected from the one- humped camels (Camelus dromedaries) at Taboul slaughter house, central Sudan. DNAs were extracted from protoscolices and/or associated germinal layers of hydatid cysts using a commercial kit. The mitochondrial NADH dehydrogenase subunit 1 (NADH1) gene and the cytochrome C oxidase subunit 1 (cox1) gene were used as targets for polymerase chain reaction (PCR) amplification. The PCR products were purified and partial sequences were generated. Sequences were further examined by sequence analysis and subsequent phylogeny to compare these sequences to those from known strains of EG circulating globally. RESULTS: The identity of the PCR products were confirmed as NADH1 and cox1 nucleotide sequences using the Basic Local Alignment Search Tool (BLAST) of NCBI (National Center for Biotechnology Information, Bethesda, MD). The phylogenetic analysis showed that 98% (n = 49) of the isolates clustered with Echinococcus canadensis genotype 6 (G6), whereas only one isolate (2%) clustered with Echinococcus ortleppi (G5). CONCLUSIONS: This investigation expands on the existing sequence data generated from EG isolates recovered from camel in the Sudan. The circulation of the cattle genotype (G5) in the one-humped camel is reported here for the first time.

Development and evaluation of loop-mediated isothermal amplification assay for detection of Crimean Congo hemorrhagic fever virus in Sudan.

Crimean-Congo hemorrhagic fever (CCHF) virus (CCHFV) activity has been detected in Kordufan region of the Sudan in 2008 with high case-fatality rates in villages and rural hospitals in the region. Therefore, in the present study, a reverse transcription (RT) loop-mediated isothermal amplification (RT-LAMP) assay was developed and compared to nested RT-PCR for rapid detection of CCHFV targeting the small (S) RNA segment. A set of RT-LAMP primers, designed from a highly conserved region of the S segment of the viral genome, was employed to identify all the Sudanese CCHFV strains. The sensitivity studies indicated that the RT-LAMP detected 10fg of CCHFV RNA as determined by naked eye turbidity read out, which is more likely the way it would be read in a resource-poor setting. This level of sensitivity is good enough to detect most acute cases. Using agarose gel electrophoresis, the RT-LAMP assay detected as little as 0.1fg of viral RNA (equivalent to 50 viral particle). There was 100% agreement between results of the RT-LAMP and the nested PCR when testing 10-fold serial dilution of CCHFV RNA. The specificity studies indicated that there was no cross-reactivity with other related hemorrhagic fever viruses circulating in Sudan including, Rift Valley fever virus (RVFV), Dengue fever virus, and yellow fever virus. The RT-LAMP was performed under isothermal conditions at 63°C and no special apparatus was needed, which rendered the assay more economical and practical than real-time PCR in such developing countries, like Sudan. In addition, the RT-LAMP provides a valuable tool for rapid detection and differentiation of CCHFV during an outbreak of the disease in remote areas and in rural hospitals with resource-poor settings.

A nosocomial transmission of crimean-congo hemorrhagic fever to an attending physician in North Kordufan, Sudan.

BACKGROUND: Crimean-Congo hemorrhagic fever (CCHF), a tick-borne disease caused by Crimean-Congo hemorrhagic fever virus (CCHFV), is a member of the genus Nairovirus in the family Bunyaviridae. Recently, CCHFV has been reported as an important emerging infectious viral pathogen in Sudan. Sporadic cases and multiple CCHF outbreaks, associated with nosocomial chain of transmission, have been reported in the Kordufan region of Sudan. AIMS: To confirm CCHF in an index patient and attending physician in North Kordufan region, Sudan, and to provide some information on virus genetic lineages. METHODS: Antibody captured ELISA, reverse transcription PCR, partial S segment sequences of the virus and subsequent phylogenetic analysis were used to confirm the CCHFV infection and to determine the virus genetic lineages. RESULTS: CCHF was confirmed by monitoring specific IgM antibody and by detection of the viral genome using RT-PCR. Treatment with oral ribavirin, replacement with fluid therapy, blood transfusion and administration of platelets concentrate resulted in rapid improvement of the health condition of the female physician. Phylogenetic analysis of the partial S segment sequences of the 2 CCHFV indicates that both strains are identical and belong to Group III virus lineage, which includes viruses from Africa including, Sudan, Mauritania, South Africa and Nigeria. CONCLUSION: Further epidemiologic studies including, CCHFV complete genome analysis and implementation of improved surveillance are urgently needed to better predict and respond to CCHF outbreaks in the Kordufan region, Sudan.