Objective: This research is designed to explore the methods of isolation and culture for endothelial progenitor cells from bovine bone marrow, characteristic, induced differentiative capacity in vitro.Material and methods: Main experimental reagents contain DMEM/F12, fetal bovine serum, percoll lymphocyte separating, Trypsin 1: 250, VEGF, bFGF, GF-1, EDTA and so on. Cultivation system is DMEM/F12 with 10% FBS and VEGF 10 ng/mL, cultured under 37°C, 5% CO2, saturated humidity. Cell viability is measured by trypan blue solution exclusion test. Immunofluorescent detection is used to detected cell surface markers and double swallows, while bovine chromosome is analyzed by karyotyping.  Results: We find that the majority of bovine endothelial progenitor cells (EPCs) are fibrous shaped. Frozen survival of bovine EPCs before and after cryopreservation is 95.2±0.14% and 80.9±0.30% respectively; cryopreservation affects little on the viability of bovine EPCs. Immunofluorescent detection of the cell surface markers CD34, CD133 and flk present positive, which can confirm that the cell cultured in vitro are EPCs. Then Dil-ac-LDL and FITC-UAE-1 uptake assays are carried out. Eventually, bovine EPCs are induced to differentiate into endothelial cells and smooth muscle cells respectively, demonstrating the multi-lineage differentiation potential of bovine EPCs in vitro.Conclusion: EPCs can be got with proper culture system. The little cell cryopreservation effect and stronger induced differentiation potential in vitro imply that EPCs can be applied in genetic resources conservation and reuse.http://doi.org/10.5455/javar.2017.d196

Objective: This research is designed to explore the methods of isolation and culture for endothelial progenitor cells from bovine bone marrow, characteristic, induced differentiative capacity in vitro. Material and methods: Main experimental reagents contain DMEM/F12, fetal bovine serum, percoll lymphocyte separating, Trypsin 1: 250, VEGF, bFGF, GF-1, EDTA and so on. Cultivation system is DMEM/F12 with 10% FBS and VEGF 10 ng/mL, cultured under 37°C, 5% CO2, saturated humidity. Cell viability is measured by trypan blue solution exclusion test. Immunofluorescent detection is used to detected cell surface markers and double swallows, while bovine chromosome is analyzed by karyotyping. Results: We find that the majority of bovine endothelial progenitor cells (EPCs) are fibrous shaped. Frozen survival of bovine EPCs before and after cryopreservation is 95.2±0.14% and 80.9±0.30% respectively; cryopreservation affects little on the viability of bovine EPCs. Immunofluorescent detection of the cell surface markers CD34, CD133 and flk present positive, which can confirm that the cell cultured in vitro are EPCs. Then Dil-ac-LDL and FITC-UAE-1 uptake assays are carried out. Eventually, bovine EPCs are induced to differentiate into endothelial cells and smooth muscle cells respectively, demonstrating the multi-lineage differentiation potential of bovine EPCs in vitro. Conclusion: EPCs can be got with proper culture system. The little cell cryopreservation effect and stronger induced differentiation potential in vitro imply that EPCs can be applied in genetic resources conservation and reuse. [J Adv Vet Anim Res 2017; 4(1.000): 88-96]

Objective: This study aimed at assessing the prevalence of Helicobacter spp. in ice cream sold at Qena city (Egypt) with special reference to H. pylori, and to study the antimicrobial effect of mastic gum essential oil on H. pylori.Materials and methods: A total of 60 small and large scale ice cream samples (30 each) were investigated for the presence of Helicobacter spp. Then, the essential oil of mastic gum was added to lab prepared ice cream inoculated with H. pylori isolate in a count of Log6 using 2 concentrations of 0.8 and 1.6% and one group was kept as a control group. Each group was subdivided into 2 sub-groups; one was kept at -5°C and another one was kept at -20°C.Results: Helicobacter spp. could be isolated from 11.7%, while, H. pylori could be found in 3.3% of the total examined ice cream samples. Regarding the anti-Helicobacter pylori effect of mastic gum essential oil added to lab prepared ice cream, in samples kept at -5°C it was found that after hardening the count fall into Log5 in both control and 0.8% groups and log4 in 1.6% essential mastic oil concentration. Reduction in H. pylori at the first week and third day of storage reached 100% where H. pylori couldn’t be counted or isolated from both 0.8 and 1.6% concentrations, respectively. While in ice cream samples stored at -20°C, the reduction in H. pylori load was rapid at first, it was log4 and log3 in groups of 0.8% and 1.6% concentrations, respectively versus log5 in control samples. At the third day H. pylori was not able to be counted or isolated at third day of storage for both concentrations.Conclusion: All isolates recovered from small scale ice cream samples reflexing the hygienic conditions under which samples were produced. Mastic gum essential oil exhibited a powerful anti-H. pylori effect recommending its addition to food matrix for therapeutic purposes or as a functional food.http://doi.org/10.5455/javar.2017.d199     XML     PubReader

Objective: This study aimed at assessing the prevalence of Helicobacter spp. in ice cream sold at Qena city (Egypt) with special reference to H. pylori, and to study the antimicrobial effect of mastic gum essential oil on H. pylori. Materials and methods: A total of 60 small and large scale ice cream samples (30 each) were investigated for the presence of Helicobacter spp. Then, the essential oil of mastic gum was added to lab prepared ice cream inoculated with H. pylori isolate in a count of Log6 using 2 concentrations of 0.8 and 1.6% and one group was kept as a control group. Each group was subdivided into 2 sub-groups; one was kept at -5°C and another one was kept at -20°C. Results: Helicobacter spp. could be isolated from 11.7%, while, H. pylori could be found in 3.3% of the total examined ice cream samples. Regarding the anti-Helicobacter pylori effect of mastic gum essential oil added to lab prepared ice cream, in samples kept at -5°C it was found that after hardening the count fall into Log5 in both control and 0.8% groups and log4 in 1.6% essential mastic oil concentration. Reduction in H. pylori at the first week and third day of storage reached 100% where H. pylori couldn’t be counted or isolated from both 0.8 and 1.6% concentrations, respectively. While in ice cream samples stored at -20°C, the reduction in H. pylori load was rapid at first, it was log4 and log3 in groups of 0.8% and 1.6% concentrations, respectively versus log5 in control samples. At the third day H. pylori was not able to be counted or isolated at third day of storage for both concentrations. Conclusion: All isolates recovered from small scale ice cream samples reflexing the hygienic conditions under which samples were produced. Mastic gum essential oil exhibited a powerful anti-H. pylori effect recommending its addition to food matrix for therapeutic purposes or as a functional food. [J Adv Vet Anim Res 2017; 4(2.000): 132-139]

Objective: Cattle are regarded as the principle reservoir of O157 and non-O157 shigatoxigenic Escherichia coli (STEC). Spreading of the STEC to human is primarily happens through contaminated meat, milk and their byproducts. The present study was aimed to explore the occurrence of STEC in the rectal swab of apparently healthy cattle.Materials and methods: A total of 60 E. coli isolates that were previously isolated from the rectal swab of cattle were used in this study. DNA were extracted from the isolates and screened by PCR to detect E. coli stx (stx1, stx2), ehxA and rfbO157 genes. Representative amplicons of the PCR products were sequenced. The prevalence of the STEC was determined based on the detection of STEC specific stx genes. The prevalence data were further analyzed by SPSS to elucidate any difference among different demographic groups of the study population.Results: Overall, 43.33% (n=26/60) of the isolates were found carrying stx genes. Based on the presence of stx and ehxA genes, 6 different types of STEC were identified, of which 20% (n=12/26) were carrying both stx1 and stx2 genes. None of the isolates was positive for rfbO157. The PCR amplicons were sequenced, and the nucleotide sequences were deposited in GenBank (accession: KM596779-KM596784).Conclusion: In this study, non-O157 STEC were found highly prevalent in the local cattle. This study suggests that the apparently healthy cattle may act as a potential source of STEC infection for humans.http://doi.org/10.5455/javar.2017.d213                 XML     PubReader

Objective: Cattle are regarded as the principle reservoir of O157 and non-O157 shigatoxigenic Escherichia coli (STEC). Spreading of the STEC to human is primarily happens through contaminated meat, milk and their byproducts. The present study was aimed to explore the occurrence of STEC in the rectal swab of apparently healthy cattle. Materials and methods: A total of 60 E. coli isolates that were previously isolated from the rectal swab of cattle were used in this study. DNA were extracted from the isolates and screened by PCR to detect E. coli stx (stx1, stx2), ehxA and rfbO157 genes. Representative amplicons of the PCR products were sequenced. The prevalence of the STEC was determined based on the detection of STEC specific stx genes. The prevalence data were further analyzed by SPSS to elucidate any difference among different demographic groups of the study population. Results: Overall, 43.33% (n=26/60) of the isolates were found carrying stx genes. Based on the presence of stx and ehxA genes, 6 different types of STEC were identified, of which 20% (n=12/26) were carrying both stx1 and stx2 genes. None of the isolates was positive for rfbO157. The PCR amplicons were sequenced, and the nucleotide sequences were deposited in GenBank (accession: KM596779-KM596784). Conclusion: In this study, non-O157 STEC were found highly prevalent in the local cattle. This study suggests that the apparently healthy cattle may act as a potential source of STEC infection for humans. [J Adv Vet Anim Res 2017; 4(2.000): 194-199]

Lespedeza cuneata (poorman’s lucerne; sericea lespedeza), a tannin-rich perennial legume, was offered as hay to dry Merino ewes in a confined feeding experiment to evaluate the effect on the level of gastrointestinal parasite infection in sheep. Medicago sativa (a low tannin containing perennial legume) was used as the control treatment. Parameters faecal egg count (FEC), FAMACHA© scores and rectal temperatures were used. FECs were substantially lower (p = 0.05) in the Lespedeza group after 35 days, together with a trend of higher rectal temperatures, compared with the Medicago group. Although non-significant (p 0.05), the higher rectal temperatures suggested a lower level of anaemia in the sheep on the Lespedeza ration and, therefore, a lower parasite-worm burden. However, FAMACHA© scores showed no significant (p 0.05) differences between treatments despite the differences in FEC that were recorded, indicating that host homeostasis was possibly mediated by improved nutrition as a result of the high protein content of both experimental diets.

Lespedeza cuneata (poorman’s lucerne; sericea lespedeza), a tannin-rich perennial legume, was offered as hay to dry Merino ewes in a confined feeding experiment to evaluate the effect on the level of gastrointestinal parasite infection in sheep. Medicago sativa (a low tannin containing perennial legume) was used as the control treatment. Parameters faecal egg count (FEC), FAMACHA© scores and rectal temperatures were used. FECs were substantially lower (p = 0.05) in the Lespedeza group after 35 days, together with a trend of higher rectal temperatures, compared with the Medicago group. Although non-significant (p > 0.05), the higher rectal temperatures suggested a lower level of anaemia in the sheep on the Lespedeza ration and, therefore, a lower parasite-worm burden. However, FAMACHA© scores showed no significant (p > 0.05) differences between treatments despite the differences in FEC that were recorded, indicating that host homeostasis was possibly mediated by improved nutrition as a result of the high protein content of both experimental diets.

The objectives of the study were to determine the species composition of ticks infesting white and black rhinoceroses in southern Africa as well as the conservation status of those tick species that prefer rhinos as hosts. Ticks were collected opportunistically from rhinos that had been immobilised for management purposes, and 447 white rhinoceroses (Ceratotherium simum) and 164 black rhinoceroses (Diceros bicornis) were sampled in South Africa, 61 black rhinos in Namibia, 18 white and 12 black rhinos in Zimbabwe, and 24 black rhinos in Zambia. Nineteen tick species were recovered, of which two species, Amblyomma rhinocerotis and Dermacentor rhinocerinus, prefer rhinos as hosts. A. rhinocerotis was collected only in the northeastern KwaZulu-Natal reserves of South Africa and is endangered, while D. rhinocerinus is present in these reserves as well as in the Kruger National Park and surrounding conservancies. Eight of the tick species collected from the rhinos are ornate, and seven species are regularly collected from cattle. The species present on rhinos in the eastern, moister reserves of South Africa were amongst others Amblyomma hebraeum, A. rhinocerotis, D. rhinocerinus, Rhipicephalus maculatus, Rhipicephalus simus and Rhipicephalus zumpti, while those on rhinos in the Karoo and the drier western regions, including Namibia, were the drought-tolerant species, Hyalomma glabrum, Hyalomma rufipes, Hyalomma truncatum and Rhipicephalus gertrudae. The species composition of ticks on rhinoceroses in Zambia differed markedly from those of the other southern African countries in that Amblyomma sparsum, Amblyomma tholloni and Amblyomma variegatum accounted for the majority of infestations.

The objectives of the study were to determine the species composition of ticks infesting white and black rhinoceroses in southern Africa as well as the conservation status of those tick species that prefer rhinos as hosts. Ticks were collected opportunistically from rhinos that had been immobilised for management purposes, and 447 white rhinoceroses (Ceratotherium simum) and 164 black rhinoceroses (Diceros bicornis) were sampled in South Africa, 61 black rhinos in Namibia, 18 white and 12 black rhinos in Zimbabwe, and 24 black rhinos in Zambia. Nineteen tick species were recovered, of which two species, Amblyomma rhinocerotis and Dermacentor rhinocerinus, prefer rhinos as hosts. A. rhinocerotis was collected only in the northeastern KwaZulu-Natal reserves of South Africa and is endangered, while D. rhinocerinus is present in these reserves as well as in the Kruger National Park and surrounding conservancies. Eight of the tick species collected from the rhinos are ornate, and seven species are regularly collected from cattle. The species present on rhinos in the eastern, moister reserves of South Africa were amongst others Amblyomma hebraeum, A. rhinocerotis, D. rhinocerinus, Rhipicephalus maculatus, Rhipicephalus simus and Rhipicephalus zumpti, while those on rhinos in the Karoo and the drier western regions, including Namibia, were the drought-tolerant species, Hyalomma glabrum, Hyalomma rufipes, Hyalomma truncatum and Rhipicephalus gertrudae. The species composition of ticks on rhinoceroses in Zambia differed markedly from those of the other southern African countries in that Amblyomma sparsum, Amblyomma tholloni and Amblyomma variegatum accounted for the majority of infestations.

One hundred milk samples were collected from camel’s milk for the isolation of Staphylococcus aureus. Thirty-one isolates were S. aureus, 45 were other forms of staphylococci and 24 represented other bacteria. Five isolates from S. aureus were methicillin resistant S. aureus (MRSA) and 26 samples were methicillin susceptible S. aureus (MSSA). The whole genome sequence of S. aureus was annotated and visualised by rapid annotation using subsystem technology (RAST) which is a fully-automated service for annotating complete or nearly complete bacterial genomes. Four isolates from MSSA strains were subjected to multi-locus sequence typing (MLST). Three multilocus sequences types or sequence types (MLST/ST) were found, namely ST15, ST1153 and ST130. The phylogenetic analysis of the concatenated sequences of the seven genes forming the MLST profile of S. aureus classification revealed a high degree of similarity and close relationship between the ST15 and ST1153 while the third ST (ST130) was located in a different cluster.

One hundred milk samples were collected from camel’s milk for the isolation of Staphylococcus aureus. Thirty-one isolates were S. aureus, 45 were other forms of staphylococci and 24 represented other bacteria. Five isolates from S. aureus were methicillin resistant S. aureus (MRSA) and 26 samples were methicillin susceptible S. aureus (MSSA). The whole genome sequence of S. aureus was annotated and visualised by rapid annotation using subsystem technology (RAST) which is a fully-automated service for annotating complete or nearly complete bacterial genomes. Four isolates from MSSA strains were subjected to multi-locus sequence typing (MLST). Three multilocus sequences types or sequence types (MLST/ST) were found, namely ST15, ST1153 and ST130. The phylogenetic analysis of the concatenated sequences of the seven genes forming the MLST profile of S. aureus classification revealed a high degree of similarity and close relationship between the ST15 and ST1153 while the third ST (ST130) was located in a different cluster.

Enterotoxaemia (pulpy kidney) is a common bacterial disease of sheep caused by Clostridium perfringens type D epsilon toxin. It has mortality rates of up to 30% in non-vaccinated animals. Current vaccines from whole cell cultures are expensive to manufacture and can induce local inflammatory responses in sheep. They usually have reduced immunogenicity because of the difficulty of standardising the inactivation step in vaccine manufacturing. In the current study, we evaluated the safety and potency of a recombinant plant-made epsilon toxoid protein (r-Etox) as an affordable and safer alternative vaccine for developing countries. Results of injection site reactions, rectal temperature and toxin neutralisation test in single and prime– boost inoculations of mice, guinea pigs and sheep suggest that the product is not toxic to animals and could protect sheep against enterotoxaemia.

Enterotoxaemia (pulpy kidney) is a common bacterial disease of sheep caused by Clostridium perfringens type D epsilon toxin. It has mortality rates of up to 30% in non-vaccinated animals. Current vaccines from whole cell cultures are expensive to manufacture and can induce local inflammatory responses in sheep. They usually have reduced immunogenicity because of the difficulty of standardising the inactivation step in vaccine manufacturing. In the current study, we evaluated the safety and potency of a recombinant plant-made epsilon toxoid protein (r-Etox) as an affordable and safer alternative vaccine for developing countries. Results of injection site reactions, rectal temperature and toxin neutralisation test in single and prime– boost inoculations of mice, guinea pigs and sheep suggest that the product is not toxic to animals and could protect sheep against enterotoxaemia.

There is paucity of information on the prevalence of leptospirosis in wildlife in Nigeria. This study investigated the prevalence and renal pathology of leptospirosis in wild animals in Southwest Nigeria. One hundred and five kidney samples were examined from 10 different wildlife species (antelope) greater cane rat (GCR), hare, African giant rat (AGR), tree hyrax, civet cat, monitor lizard, python, bushbuck and partridge) using a combination of Ellinghausen McCullough Johnson Harris (EMJH) medium, microscopic agglutination test (MAT), Warthin– Starry silver stain (WSss) and immunohistochemistry. Chi-square test was used with confidence level set at 0.05 to ascertain associations between positive cases and sex and species. Eightytwo (78.1%) samples were culturally positive, while 67.7% (63/93), 57.0% (16/28) and 66.7% (8/12) were WSss, MAT and immunohistochemically positive, respectively. Interstitial nephritis (41.0%) and tubular nephrosis (81.0%) were the most prominent histopathological changes. Pathogenic Leptospira organisms were highest in GCR (32.1%) and antelope (14.3%). Serovars hardjo (11.54%), bratislava (3.9%), canicola (3.9%), icterohaemorrhagiae (15.4%), pomona (7.14%) gripptotyphosa (19.2%) and undetermined isolates were also detected in other animals. The result showed high prevalence of Leptospira infection in the wild and the possibility of domestic animals and humans contracting the disease. This study is the first documentation of evidence of pathogenic Leptospira species in wildlife in Nigeria.

There is paucity of information on the prevalence of leptospirosis in wildlife in Nigeria. This study investigated the prevalence and renal pathology of leptospirosis in wild animals in Southwest Nigeria. One hundred and five kidney samples were examined from 10 different wildlife species (antelope) greater cane rat (GCR), hare, African giant rat (AGR), tree hyrax, civet cat, monitor lizard, python, bushbuck and partridge) using a combination of Ellinghausen McCullough Johnson Harris (EMJH) medium, microscopic agglutination test (MAT), Warthin– Starry silver stain (WSss) and immunohistochemistry. Chi-square test was used with confidence level set at 0.05 to ascertain associations between positive cases and sex and species. Eightytwo (78.1%) samples were culturally positive, while 67.7% (63/93), 57.0% (16/28) and 66.7% (8/12) were WSss, MAT and immunohistochemically positive, respectively. Interstitial nephritis (41.0%) and tubular nephrosis (81.0%) were the most prominent histopathological changes. Pathogenic Leptospira organisms were highest in GCR (32.1%) and antelope (14.3%). Serovars hardjo (11.54%), bratislava (3.9%), canicola (3.9%), icterohaemorrhagiae (15.4%), pomona (7.14%) gripptotyphosa (19.2%) and undetermined isolates were also detected in other animals. The result showed high prevalence of Leptospira infection in the wild and the possibility of domestic animals and humans contracting the disease. This study is the first documentation of evidence of pathogenic Leptospira species in wildlife in Nigeria.