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Papers by Eugene Bwalya

Journal of Veterinary Medical Science

Bupivacaine, levobupivacaine and ropivacaine are potent, long acting, amide-type local anesthetic... more Bupivacaine, levobupivacaine and ropivacaine are potent, long acting, amide-type local anesthetics that have several clinical applications including intra-articular administration. The objectives of this study were to evaluate their in vitro effects on cell viability and caspase activity to elucidate whether they activate the extrinsic or intrinsic pathways of apoptosis in canine articular chondrocytes. Chondrocytes in monolayer culture were treated with culture medium as the control, or with 0.062% (0.62 mg/mL) bupivacaine, 0.062% levobupivacaine, and 0.062% ropivacaine for 24 hr. Cell viability was evaluated using the live/dead, 3-(4,5-dimehylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), and Cell Counting Kit-8 (CCK-8) assays. Evaluation of caspase-3, caspase-8, and caspase-9 activity was performed using colorimetric assays. The MTT and CCK-8 assays were used to evaluate the effect of caspase inhibitors on local anesthetic chondrotoxicity. All three local anesthetics decreased chondrocyte viability after 24 hr (P ˂ 0.001). Apoptosis was induced through both the extrinsic and intrinsic pathways. Bupivacaine increased caspase-3, caspase-8, and caspase-9 activity (P ˂ 0.001). Levobupivacaine increased caspase-3 (P = 0.03) while ropivacaine did not significantly upregulate activity for all three caspases. Caspase inhibition did not suppress bupivacaine chondrotoxicity whereas inhibition of caspase-8 and caspase-9 decreased ropivacaine chondrotoxicity and mildly attenuated levobupivacaine chondrotoxicity. In summary, the level of chondrotoxicity, the type of caspase activated, the level of caspase activation, and the response to caspase inhibitors was dependent on the type of local anesthetic. Therefore, ropivacaine may be a safer choice for intra-articular administration compared to levobupivacaine and bupivacaine.

University of Zambia Journal of Agricultural and Biomedical Sciences, 2021

Developing a price policy that ensures financial viability in order to continue providing a servi... more Developing a price policy that ensures financial viability in order to continue providing a service. The same is true for veterinary services whose economic nature has to be understood for practitioners to adjust prices for smooth income flows to the business. However, veterinary practitioners’ prices often vary across practices without a clear understanding of the economic nature of animal healthcare or elasticity of demand for the services they provide. This study, therefore, aimed to assess the price elasticity of demand for veterinary services using the University of Zambia Veterinary Clinic (UNZAVET) as a case study. To meet this objective, historical five-year data (2014-2018) was collected from the clinics’ medical and financial records. The data was then entered and analysed using both Microsoft Excel and IBM SPSS Version 20. The price elasticity of demand was calculated using an end-point method. The association between inflation rate and revenue was estimated using Simple ...

Research in Veterinary Science, 2018

Pentosan polysulfate (PPS) is currently under investigation as a potential disease-modifying anti... more Pentosan polysulfate (PPS) is currently under investigation as a potential disease-modifying antiarthritic agent. In the present study the effects of PPS on arthritic profiles based on clinical score, ankle size, histological changes, and activity of inflammatory mediators using collagen-induced arthritic rat are reported. Model of arthritis was developed in Sprague Dawley rats by intradermal injection of bovine type II collagen emulsified with incomplete Freund's adjuvant. The rats were randomly divided into four groups: normal control, arthritic control, arthritic rats treated with PPS (at dose level 20 μg/g) and arthritic rats treated with meloxicam (2 μg/g). The treatment was continued daily until the day 30. Arthritic biomarkers (cartilage oligomeric matrix protein and tartrate-resistant acid phosphatase 5b) in synovial fluid, expression of inflammatory mediators (interleukin-1β, and tumor necrosis factor-α) and osteoclast marker genes (cathepsin K, tartrate-resistant acid phosphatase) in synovial membrane were measured. Daily administration of PPS to the arthritic rats significantly decreased the severity of arthritis by effectively suppressing the symptoms of arthritis and improving the functional recovery based on clinical score and histopathological evidence. Intriguingly, identical downregulation pattern of arthritis profiles, biological markers as well as relative mRNA levels of osteoclast markers and cytokines were monitored in arthritic rats treated with PPS. In conclusion, PPS exerted protective effects against collageninduced arthritis in rats. The results suggest that PPS acts as an anti-inflammatory and anti-arthritic agent in decreasing the arthritic effects in collagen-induced arthritic rats.

Journal of Veterinary Medical Science, 2018

Although chondroinductive growth factors are considered necessary for chondrogenesis of bone marr... more Although chondroinductive growth factors are considered necessary for chondrogenesis of bone marrow-derived mesenchymal stem cells (BMSC), independent and spontaneous chondrogenesis has been previously demonstrated in adult horses, bovine calves and adult human BMSC. Surprisingly, adult canine BMSC under similar culture conditions previously failed to demonstrate chondrogenesis. The present study evaluated independent chondrogenic potential of BMSC sourced from three young dogs in the absence of known chondroinductive factors. BMSC were culture expanded in 10% DMEM up to third passage (P3). At each passage, the phenotype of BMSC was evaluated by RT-PCR gel electrophoresis and qPCR. BMSC exhibited a chondrogenic phenotype in the absence of dexamethasone and TGF-β1 as verified by the expression of Sox-9, type II collagen and aggrecan. Sox-9 was significantly downregulated (P<0.05) from P1−P3 compared to P0 while type II and X collagen, and aggrecan were significantly downregulated at P3 compared to P0. There was a significant (P<0.01) negative correlation between passaging and Sox-9, type II collagen and aggrecan gene expression. These results indicate that independent chondrogenic potential and phenotype retention of BMSC decreases in a passage-dependent pattern. Therefore, caution should be exercised for future experiments evaluating the chondrogenic potential of BMSC after extensive expansion cultures in 10% DMEM.

BMC Veterinary Research, 2018

Background: Sodium pentosan polysulfate (NaPPS) was testified as a chondroprotective drug in with... more Background: Sodium pentosan polysulfate (NaPPS) was testified as a chondroprotective drug in with a detailed rationale of the disease-modifying activity. This study was undertaken to determine whether anti-osteoarthritis drug, NaPPS inhibited osteoclasts (OC) differentiation and function. Canine bone marrow mononuclear cells (n = 6) were differentiated to OC by maintaining with receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) for up to 7 days with the treatment of NaPPS at concentration of 0, 0.2, 1 and 5 μg/mL. Differentiation and function of OC were accessed using tartrate-resistant acid phosphate (TRAP) staining and bone resorption assay, while monitoring actin ring formation. Invasion and colocalization patterns of fluorescence-labeled NaPPS with transcribed gene in OC were monitored. Gene expression of OC for cathepsin K (CTK), matrix metallopeptidase-9 (MMP-9), nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), c-Fos, activator protein-1(AP-1) and carbonic anhydrase II was examined using real-time PCR. Results: Significant inhibition of OC differentiation was evident at NaPPS concentration of 1 and 5 μg/mL (p < 0.05). In the presence of 0.2 to 5 μg/mL NaPPS, bone resorption was attenuated (p < 0.05), while 1 and 5 μg/mL NaPPS achieved significant reduction of actin ring formation. Intriguingly, fluorescence-labeled NaPPS invaded in to cytoplasm and nucleus while colocalizing with actively transcribed gene. Gene expression of CTK, MMP-9 and NFATc1 were significantly inhibited at 1 and 5 μg/mL (p < 0.05) of NaPPS whereas inhibition of c-Fos and AP-1 was identified only at concentration of 5 μg/mL (p < 0.05). Conclusions: Taken together, all the results suggest that NaPPS is a novel inhibitor of RANKL and M-CSF-induced CTK, MMP-9, NFATc1, c-Fos, AP-1 upregulation, OC differentiation and bone resorption which might be a beneficial for treatment of inflammatory joint diseases and other bone diseases associated with excessive bone resorption.

The Journal of veterinary medical science, Jan 3, 2017

The aim of this study was to investigate osteoclastogenic properties of inflammatory cytokines at... more The aim of this study was to investigate osteoclastogenic properties of inflammatory cytokines at different time-points of osteoclastogenesis. Bone marrow-derived macrophages from five healthy dogs were stimulated with the macrophage colony-stimulating factor, receptor activator of nuclear factor-κB ligand and inflammatory cytokines such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α and IL-17. Osteoclasts (OC) formation and function were enhanced with TNF-α regardless of temporal differences. But in contrast, IL-1β suppressed the osteoclastogenesis at early phase of the process while upregulating at the late phase. Furthermore, differentiation of OC precursors into OC was suppressed at high concentrations of IL-17. Collectively, the results revealed that suppressing TNF-α would be a promising strategy to inhibit inflammation-associated bone destruction in dogs.

Research in Veterinary Science, 2017

The objective of this study was to assess the differentiation capability of synoviocytes derived ... more The objective of this study was to assess the differentiation capability of synoviocytes derived from dogs with inflammatory joint conditions. Cranial cruciate ligament ruptured (CCLr) (n = 12) and medial patella luxated (MPL) (n = 10) knee joints of the dogs were used to collect the synovial membrane (SM). Synoviocytes were enzymatically released from the SM and analyzed by flow cytometry for specific cellular markers (CD44 and CD90) of mesenchymal stem cells (MSCs), while doing histopathology from another part of SM sections. Under specific culture conditions, synoviocytes were forced to differentiate into chondrogenesis, adipogenesis, osteogenesis and osteoclastogenesis to investigate the multipotency. Upon treatments phenotypes of cell cultures were analyzed by histopathology and by semi-quantitative reverse transcriptase polymerase chain reaction for the expression of each differentiation marker genes. Although flow cytometry showing similar MSCs populations in CCLr and MPL synovium, synovial cells derived from CCLr showed higher multipotency compared to MPLderived samples. Further, synovial changes such as vascularity, mononuclear cell infiltration and cellular hypertrophy were more pronounced in CCLr-derived synovial tissue than in MPL. Taken together, these findings suggested that the differentiation capability of SM-derived multipotent stem cells varies with inflammatory severity occurring in different joint conditions.

Journal of Parasitology Research, 2015

A retrospective and prospective analysis of clinical records of dogs diagnosed withBabesiainfecti... more A retrospective and prospective analysis of clinical records of dogs diagnosed withBabesiainfections was carried out for the years 2000 to 2013 from practices in Lusaka, Zambia. Records of 363 dogs with confirmedBabesiainfections were analysed using demographic factors including sex, breed, age, and clinical signs in relation to haematological findings andBabesiaspecies. The clinical and laboratory findings observed are described as well asBabesiaspecies identification. The study included 18 breeds and the highest proportion were mongrels (32.2%), males representing 64.5% of the population. The most common presenting problems were anorexia (65.3%) and lethargy/weakness (65.3%). The most common clinical signs were fever (87.3%), pallor (52.3%), lymphadenopathy (47.4%), and presence of ticks (44.9%). Anaemia (96.4%) and nucleated erythrocytes (42.2%) were the most common laboratory findings. A mixed infection ofBabesia rossiandBabesia gibsoniwas present in 59.7% of dogs, whilst 8% and...

Preventive Veterinary Medicine, 2011

Faecal samples were collected from January 2010 through September 2010 to determine the prevalenc... more Faecal samples were collected from January 2010 through September 2010 to determine the prevalence of gastrointestinal (GI) helminths infestation in dogs in urban Lusaka and rural Katete Districts of Zambia. A total of 452 faecal samples (n = 160 Katete, n = 292 Lusaka) were examined by faecal flotation for the presence of helminth eggs and 82.5% of dogs were positive for GI helminths in Katete compared to 76% for Lusaka. Positive results with the presence of at least one parasite corresponded to 72.9% Ancylostoma caninum, 11% Toxocara canis, 4.8% Toxascaris leonina, 2.4% Dipylidium caninum, 0.7% Taeniidae and 0.3% T. vulpis, species for Lusaka while Katete recorded 70.6% A. caninum, 18.1% T. vulpis, 11.1% T. canis, 13.1% D. caninum, 3.8% T. leonina, and 0.6% Taeniidae. Except for T. vulpis and D. caninum (p < 0.05) the results indicated no significant difference in the prevalence of the identified GI helminth between Lusaka and Katete. There was no significant difference in the prevalence between genders of GI helminth infestation demonstrated in this study and only A. caninum showed significant difference in prevalence by age category. The study also showed the presence of zoonotic intestinal helminths A. caninum, T. canis and D. caninum. The study highlights that there was no significant difference in spectrum and prevalence of GI helminths between urban and rural areas in Zambia. It further brings to light the importance of educating owners of dogs on the importance of regular deworming of dogs and control of ectoparasites in order to minimise the risk that these dogs pose to them and the public.

Pentosan polysulfate (PPS) is widely used as therapeutic intervention for joint diseases in human... more Pentosan polysulfate (PPS) is widely used as therapeutic intervention for joint diseases in humans and animals, while objective confirmation has not been established yet. The purpose of this study was to provide the objective measure of the efficacy of PPS. Twenty-five healthy Mongolian horses were randomly assigned in three groups. Three different doses of PPS, 0/1.2/3.0 mg/kg, were injected intramuscularly one a week for consecutive 4 weeks. On 14 and 28 days after the initial administration, relative ratios of serum COMP/CPII were 97.9/87.6/61.8 and 94.2/104.3/88.1 in 0/1.2/3.0 mg/kg PPS, respectively. The results revealed that balance of cartilage metabolism could be significantly brought to an anabolism dominant state by PPS injections in dose dependent manor in field fed horses in Mongolia

University of Zambia Journal of Agricultural and Biomedical Sciences

ARTICLE21ABSTRACTStray dogs destined for euthanasia were collected from January 2010 through Dece... more ARTICLE21ABSTRACTStray dogs destined for euthanasia were collected from January 2010 through December 2010 to determine the presence of zoonotic gastrointestinal (GI) helminths in Lusaka district. Atotal of 33 stray dogs (male=18, female=15) were collected, euthanizedandnecropsyinformationobtainedviahelminthological approach. 100% (33/33) were infected with one or more helminth parasites. Multiple infections involving nematodes and cestodes were the most prevalent (63.6%) followed by nematodes only (33.3%) with cestodes only being the least prevalent (3.0%). The most prevalent GI helminth parasite was Ancylostomacaninum(93.9%) with the least prevalent being Toxocaracanis(6.1%). No positive case of Trichinella spiraliswas reported from direct trichinoscopy examination of striated muscles. The mean count per dog of A. caninumwas 44.45 (SD ±58.0) with a range of 0 to 223. The presence of three important zoonotic gastrointestinal helminths (A. caninum,T. canisand Dipylidiumcaninum) repo...

Copyright © 2015 King Shimumbo Nalubamba et al. This is an open access article distributed under ... more Copyright © 2015 King Shimumbo Nalubamba et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. A retrospective and prospective analysis of clinical records of dogs diagnosed with Babesia infections was carried out for the years 2000 to 2013 from practices in Lusaka, Zambia. Records of 363 dogs with confirmed Babesia infections were analysed using demographic factors including sex, breed, age, and clinical signs in relation to haematological findings and Babesia species. The clinical and laboratory findings observed are described as well as Babesia species identification. The study included 18 breeds and the highest proportion were mongrels (32.2%), males representing 64.5 % of the population. The most common presenting problemswere anorexia (65.3%) and lethargy/weakness (65.3%).Themost common clinical signs were fever...

The present study attempted to bridge the paucity of information on the prevalence of canine endo... more The present study attempted to bridge the paucity of information on the prevalence of canine endoparasites in Zambia and to establish some of the risk factors that may be associated with endoparasitism. Faecal, blood and necropsy data on endoparasites were collected between January and December 2010 to determine the prevalence of canine endoparasites in two demographically and socio-economically diverse regions of Zambia. A total of 486 dogs were sampled (n = 160 Katete,predominantlyrural; n = 326 Lusaka,predominantlyurban).Faecal samples were examined by simple and centrifugal faecal flotation methods for the presence of helminth eggs. In Katete, 82.5 percent of dogs were positive for gastrointestinal (GI) helminths compared to 76 percent for Lusaka. The overall prevalence of single helminth infections were as follows: 72.1 percent Ancylostoma caninum, 11.3 percent Toxocara canis, 6.6 percent Trichuris vulpis, 6.2 percent Dipylidium caninum, 4.4 percent Toxascaris leonina and 0.7 p...

Transboundary and Emerging Diseases

CARTILAGE

Objective To investigate the role and characterize the molecular mechanisms regulating apoptosis ... more Objective To investigate the role and characterize the molecular mechanisms regulating apoptosis and autophagy in nitric oxide (NO)–induced chondrocyte cell death. Design Cell apoptosis and autophagy were evaluated in chondrocytes treated with sodium nitroprusside (SNP) combined with the presence or absence of interleukin-1 beta (IL-1β) and nutrient-deprived conditions. The concentration of nitrite was determined by Griess reaction. Activation of apoptosis and autophagy were determined by immunocytochemistry, Western blot, and quantitative real-time polymerase chain reaction (qPCR) analysis. Flow cytometry and MTT assay were used to assess cell viability. Results Cotreatment of chondrocytes with SNP and IL-1β under nutrient-deprived condition potentially enhanced the effect of NO-induced cell death. Immunocytochemistry, Western blot, and qPCR analysis indicated that treatment of chondrocytes with SNP significantly reduced autophagic activity, autophagic flux, and multiple autophagy-...

Journal of General Virology

Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wid... more Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wide host range allows individual MRV to transmit into multiple species. Although several interspecies transmission and genetic reassortment events of MRVs among humans, livestock and wildlife have been reported, the genetic diversity and geographic distribution of MRVs in Africa are poorly understood. In this study, we report the first isolation and characterization of MRVs circulating in a pig population in Zambia. In our screening, MRV genomes were detected in 19.7 % (29/147) of faecal samples collected from pigs by reverse transcription PCR. Three infectious MRV strains (MRV-85, MRV-96 and MRV-117) were successfully isolated, and their complete genomes were sequenced. Recombination analyses based on the complete genome sequences of the isolated MRVs demonstrated that MRV-96 shared the S3 segment with a different MRV isolated from bats, and that the L1 and M3 segments of MRV-117 originat...

Journal of Veterinary Medical Science

This study investigated the effects of culture time on phenotype stability of canine articular ch... more This study investigated the effects of culture time on phenotype stability of canine articular chondrocytes (CACs) in non-passaged long-term monolayer culture. Third passage (P3) CACs isolated from four cartilage samples were seeded at three different initial seeding densities (0.2 × 10 4 , 1.0 × 10 4 and 5.0 × 10 4 cells/cm 2) and maintained in monolayer condition up to 8 weeks without undergoing subculture after confluence. The characteristic changes of chondrocytes during the culture period were evaluated based on the cell morphology, cell proliferation, glycosaminoglycans (GAGs) content, DNA quantification, mRNA expression and ultrastructure of chondrocytes. Chondrocytes maintained under post-confluence condition exhibited a capability to grow and proliferate up to 4 weeks. Alcian blue staining and Dimethylmethylene blue (DMMB) assay revealed that the extracellular matrix (ECM) synthesis was increased in a time-dependent manner from 2 to 8 weeks. The chondrocyte mRNA expression profile was dramatically affected by prolonged culture time, with a significant downregulation of collagen type I, whereas the expression of collagen type II, aggrecan, Sox9 and matrix metalloproteinase 13 (MMP-13) were significantly upregulated. In addition, transmission electron microscopy (TEM) result indicated dilation of rough endoplasmic reticulum (RER) in these long-term monolayer cultured chondrocytes. These findings demonstrate that the chondrocytes phenotype could be partially redifferentiated through the spontaneous redifferentiation process in long-term cultures using standard culture medium without the addition of chondrogenic supplements or tissue-culture scaffolds.

Journal of Veterinary Medical Science

Journal of Tissue Science & Engineering

Autologous chondrocyte transplantation is a promising option for the repair of isolated osteoarth... more Autologous chondrocyte transplantation is a promising option for the repair of isolated osteoarthritic cartilage lesions that requires isolation and expansion of chondrocytes from a small cartilage biopsy prior to implantation. However, when cultured in vitro, chondrocytes lose their stable phenotype and dedifferentiate to fibroblastic-like cells. The study investigated the potential of pentosan polysulfate (PPS) sodium to restore the phenotype of dedifferentiated monolayer articular chondrocytes. Canine articular chondrocytes isolated from four cartilage samples were culture expanded to establish primary culture. First passage chondrocytes were cultured as alginate beads for 18 days under normoxia in PPS concentrations of 0, 1, 5, 15 and 40 μg/mL in 20% DMEM. Effect of PPS on type I, II and X collagen, aggrecan and Runx2 gene expression were evaluated by real-time PCR. Runx2, HIF-1α and HIF-2α protein expression were evaluated by Western blot and proteoglycan deposition was determined by Alcian blue stain. Dedifferentiated chondrocytes fully retained their phenotype as evidenced by increased synthesis of cartilage-specific genes, type II collagen and aggrecan mRNA with complete suppression of type I and X collagen at PPS concentrations of 15 and 40 μg/mL. Compared to the control, type II collagen and aggrecan mRNA were significantly upregulated (P<0.05) at 5, 15 and 40 μg/mL and 5 and 15 μg/mL PPS, respectively. PPS significantly enhanced proteoglycan with peak deposition at 5 μg/mL compared to control. HIF-1α and HIF-2α proteins were detectable at protein level for the first time under normoxia condition in alginate culture. The study demonstrates for the first time the restoration of dedifferentiated canine articular chondrocytes phenotype by combining alginate encapsulation with culture in PPS without the addition of known chondrocytic growth factors. The study confirms PPS as novel chondroinductive factor with potential to offer a solution to the major challenges that exist in cartilage tissue engineering.

Viruses

Porcine sapelovirus (PSV) has been detected worldwide in pig populations. Although PSV causes var... more Porcine sapelovirus (PSV) has been detected worldwide in pig populations. Although PSV causes various symptoms such as encephalomyelitis, diarrhea, and pneumonia in pigs, the economic impact of PSV infection remains to be determined. However, information on the distribution and genetic diversity of PSV is quite limited, particularly in Africa. In this study, we investigated the prevalence of PSV infection in Zambia and characterized the isolated PSVs genetically and biologically. We screened 147 fecal samples collected in 2018 and found that the prevalences of PSV infection in suckling pigs and fattening pigs were high (36.2% and 94.0%, respectively). Phylogenetic analyses revealed that the Zambian PSVs were divided into three different lineages (Lineages 1–3) in the clade consisting of Chinese strains. The Zambian PSVs belonging to Lineages 2 and 3 replicated more efficiently than those belonging to Lineage 1 in Vero E6 and BHK cells. Bioinformatic analyses revealed that genetic re...

Journal of Veterinary Medical Science

Bupivacaine, levobupivacaine and ropivacaine are potent, long acting, amide-type local anesthetic... more Bupivacaine, levobupivacaine and ropivacaine are potent, long acting, amide-type local anesthetics that have several clinical applications including intra-articular administration. The objectives of this study were to evaluate their in vitro effects on cell viability and caspase activity to elucidate whether they activate the extrinsic or intrinsic pathways of apoptosis in canine articular chondrocytes. Chondrocytes in monolayer culture were treated with culture medium as the control, or with 0.062% (0.62 mg/mL) bupivacaine, 0.062% levobupivacaine, and 0.062% ropivacaine for 24 hr. Cell viability was evaluated using the live/dead, 3-(4,5-dimehylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), and Cell Counting Kit-8 (CCK-8) assays. Evaluation of caspase-3, caspase-8, and caspase-9 activity was performed using colorimetric assays. The MTT and CCK-8 assays were used to evaluate the effect of caspase inhibitors on local anesthetic chondrotoxicity. All three local anesthetics decreased chondrocyte viability after 24 hr (P ˂ 0.001). Apoptosis was induced through both the extrinsic and intrinsic pathways. Bupivacaine increased caspase-3, caspase-8, and caspase-9 activity (P ˂ 0.001). Levobupivacaine increased caspase-3 (P = 0.03) while ropivacaine did not significantly upregulate activity for all three caspases. Caspase inhibition did not suppress bupivacaine chondrotoxicity whereas inhibition of caspase-8 and caspase-9 decreased ropivacaine chondrotoxicity and mildly attenuated levobupivacaine chondrotoxicity. In summary, the level of chondrotoxicity, the type of caspase activated, the level of caspase activation, and the response to caspase inhibitors was dependent on the type of local anesthetic. Therefore, ropivacaine may be a safer choice for intra-articular administration compared to levobupivacaine and bupivacaine.

University of Zambia Journal of Agricultural and Biomedical Sciences, 2021

Developing a price policy that ensures financial viability in order to continue providing a servi... more Developing a price policy that ensures financial viability in order to continue providing a service. The same is true for veterinary services whose economic nature has to be understood for practitioners to adjust prices for smooth income flows to the business. However, veterinary practitioners’ prices often vary across practices without a clear understanding of the economic nature of animal healthcare or elasticity of demand for the services they provide. This study, therefore, aimed to assess the price elasticity of demand for veterinary services using the University of Zambia Veterinary Clinic (UNZAVET) as a case study. To meet this objective, historical five-year data (2014-2018) was collected from the clinics’ medical and financial records. The data was then entered and analysed using both Microsoft Excel and IBM SPSS Version 20. The price elasticity of demand was calculated using an end-point method. The association between inflation rate and revenue was estimated using Simple ...

Research in Veterinary Science, 2018

Pentosan polysulfate (PPS) is currently under investigation as a potential disease-modifying anti... more Pentosan polysulfate (PPS) is currently under investigation as a potential disease-modifying antiarthritic agent. In the present study the effects of PPS on arthritic profiles based on clinical score, ankle size, histological changes, and activity of inflammatory mediators using collagen-induced arthritic rat are reported. Model of arthritis was developed in Sprague Dawley rats by intradermal injection of bovine type II collagen emulsified with incomplete Freund's adjuvant. The rats were randomly divided into four groups: normal control, arthritic control, arthritic rats treated with PPS (at dose level 20 μg/g) and arthritic rats treated with meloxicam (2 μg/g). The treatment was continued daily until the day 30. Arthritic biomarkers (cartilage oligomeric matrix protein and tartrate-resistant acid phosphatase 5b) in synovial fluid, expression of inflammatory mediators (interleukin-1β, and tumor necrosis factor-α) and osteoclast marker genes (cathepsin K, tartrate-resistant acid phosphatase) in synovial membrane were measured. Daily administration of PPS to the arthritic rats significantly decreased the severity of arthritis by effectively suppressing the symptoms of arthritis and improving the functional recovery based on clinical score and histopathological evidence. Intriguingly, identical downregulation pattern of arthritis profiles, biological markers as well as relative mRNA levels of osteoclast markers and cytokines were monitored in arthritic rats treated with PPS. In conclusion, PPS exerted protective effects against collageninduced arthritis in rats. The results suggest that PPS acts as an anti-inflammatory and anti-arthritic agent in decreasing the arthritic effects in collagen-induced arthritic rats.

Journal of Veterinary Medical Science, 2018

Although chondroinductive growth factors are considered necessary for chondrogenesis of bone marr... more Although chondroinductive growth factors are considered necessary for chondrogenesis of bone marrow-derived mesenchymal stem cells (BMSC), independent and spontaneous chondrogenesis has been previously demonstrated in adult horses, bovine calves and adult human BMSC. Surprisingly, adult canine BMSC under similar culture conditions previously failed to demonstrate chondrogenesis. The present study evaluated independent chondrogenic potential of BMSC sourced from three young dogs in the absence of known chondroinductive factors. BMSC were culture expanded in 10% DMEM up to third passage (P3). At each passage, the phenotype of BMSC was evaluated by RT-PCR gel electrophoresis and qPCR. BMSC exhibited a chondrogenic phenotype in the absence of dexamethasone and TGF-β1 as verified by the expression of Sox-9, type II collagen and aggrecan. Sox-9 was significantly downregulated (P<0.05) from P1−P3 compared to P0 while type II and X collagen, and aggrecan were significantly downregulated at P3 compared to P0. There was a significant (P<0.01) negative correlation between passaging and Sox-9, type II collagen and aggrecan gene expression. These results indicate that independent chondrogenic potential and phenotype retention of BMSC decreases in a passage-dependent pattern. Therefore, caution should be exercised for future experiments evaluating the chondrogenic potential of BMSC after extensive expansion cultures in 10% DMEM.

BMC Veterinary Research, 2018

Background: Sodium pentosan polysulfate (NaPPS) was testified as a chondroprotective drug in with... more Background: Sodium pentosan polysulfate (NaPPS) was testified as a chondroprotective drug in with a detailed rationale of the disease-modifying activity. This study was undertaken to determine whether anti-osteoarthritis drug, NaPPS inhibited osteoclasts (OC) differentiation and function. Canine bone marrow mononuclear cells (n = 6) were differentiated to OC by maintaining with receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) for up to 7 days with the treatment of NaPPS at concentration of 0, 0.2, 1 and 5 μg/mL. Differentiation and function of OC were accessed using tartrate-resistant acid phosphate (TRAP) staining and bone resorption assay, while monitoring actin ring formation. Invasion and colocalization patterns of fluorescence-labeled NaPPS with transcribed gene in OC were monitored. Gene expression of OC for cathepsin K (CTK), matrix metallopeptidase-9 (MMP-9), nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), c-Fos, activator protein-1(AP-1) and carbonic anhydrase II was examined using real-time PCR. Results: Significant inhibition of OC differentiation was evident at NaPPS concentration of 1 and 5 μg/mL (p < 0.05). In the presence of 0.2 to 5 μg/mL NaPPS, bone resorption was attenuated (p < 0.05), while 1 and 5 μg/mL NaPPS achieved significant reduction of actin ring formation. Intriguingly, fluorescence-labeled NaPPS invaded in to cytoplasm and nucleus while colocalizing with actively transcribed gene. Gene expression of CTK, MMP-9 and NFATc1 were significantly inhibited at 1 and 5 μg/mL (p < 0.05) of NaPPS whereas inhibition of c-Fos and AP-1 was identified only at concentration of 5 μg/mL (p < 0.05). Conclusions: Taken together, all the results suggest that NaPPS is a novel inhibitor of RANKL and M-CSF-induced CTK, MMP-9, NFATc1, c-Fos, AP-1 upregulation, OC differentiation and bone resorption which might be a beneficial for treatment of inflammatory joint diseases and other bone diseases associated with excessive bone resorption.

The Journal of veterinary medical science, Jan 3, 2017

The aim of this study was to investigate osteoclastogenic properties of inflammatory cytokines at... more The aim of this study was to investigate osteoclastogenic properties of inflammatory cytokines at different time-points of osteoclastogenesis. Bone marrow-derived macrophages from five healthy dogs were stimulated with the macrophage colony-stimulating factor, receptor activator of nuclear factor-κB ligand and inflammatory cytokines such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α and IL-17. Osteoclasts (OC) formation and function were enhanced with TNF-α regardless of temporal differences. But in contrast, IL-1β suppressed the osteoclastogenesis at early phase of the process while upregulating at the late phase. Furthermore, differentiation of OC precursors into OC was suppressed at high concentrations of IL-17. Collectively, the results revealed that suppressing TNF-α would be a promising strategy to inhibit inflammation-associated bone destruction in dogs.

Research in Veterinary Science, 2017

The objective of this study was to assess the differentiation capability of synoviocytes derived ... more The objective of this study was to assess the differentiation capability of synoviocytes derived from dogs with inflammatory joint conditions. Cranial cruciate ligament ruptured (CCLr) (n = 12) and medial patella luxated (MPL) (n = 10) knee joints of the dogs were used to collect the synovial membrane (SM). Synoviocytes were enzymatically released from the SM and analyzed by flow cytometry for specific cellular markers (CD44 and CD90) of mesenchymal stem cells (MSCs), while doing histopathology from another part of SM sections. Under specific culture conditions, synoviocytes were forced to differentiate into chondrogenesis, adipogenesis, osteogenesis and osteoclastogenesis to investigate the multipotency. Upon treatments phenotypes of cell cultures were analyzed by histopathology and by semi-quantitative reverse transcriptase polymerase chain reaction for the expression of each differentiation marker genes. Although flow cytometry showing similar MSCs populations in CCLr and MPL synovium, synovial cells derived from CCLr showed higher multipotency compared to MPLderived samples. Further, synovial changes such as vascularity, mononuclear cell infiltration and cellular hypertrophy were more pronounced in CCLr-derived synovial tissue than in MPL. Taken together, these findings suggested that the differentiation capability of SM-derived multipotent stem cells varies with inflammatory severity occurring in different joint conditions.

Journal of Parasitology Research, 2015

A retrospective and prospective analysis of clinical records of dogs diagnosed withBabesiainfecti... more A retrospective and prospective analysis of clinical records of dogs diagnosed withBabesiainfections was carried out for the years 2000 to 2013 from practices in Lusaka, Zambia. Records of 363 dogs with confirmedBabesiainfections were analysed using demographic factors including sex, breed, age, and clinical signs in relation to haematological findings andBabesiaspecies. The clinical and laboratory findings observed are described as well asBabesiaspecies identification. The study included 18 breeds and the highest proportion were mongrels (32.2%), males representing 64.5% of the population. The most common presenting problems were anorexia (65.3%) and lethargy/weakness (65.3%). The most common clinical signs were fever (87.3%), pallor (52.3%), lymphadenopathy (47.4%), and presence of ticks (44.9%). Anaemia (96.4%) and nucleated erythrocytes (42.2%) were the most common laboratory findings. A mixed infection ofBabesia rossiandBabesia gibsoniwas present in 59.7% of dogs, whilst 8% and...

Preventive Veterinary Medicine, 2011

Faecal samples were collected from January 2010 through September 2010 to determine the prevalenc... more Faecal samples were collected from January 2010 through September 2010 to determine the prevalence of gastrointestinal (GI) helminths infestation in dogs in urban Lusaka and rural Katete Districts of Zambia. A total of 452 faecal samples (n = 160 Katete, n = 292 Lusaka) were examined by faecal flotation for the presence of helminth eggs and 82.5% of dogs were positive for GI helminths in Katete compared to 76% for Lusaka. Positive results with the presence of at least one parasite corresponded to 72.9% Ancylostoma caninum, 11% Toxocara canis, 4.8% Toxascaris leonina, 2.4% Dipylidium caninum, 0.7% Taeniidae and 0.3% T. vulpis, species for Lusaka while Katete recorded 70.6% A. caninum, 18.1% T. vulpis, 11.1% T. canis, 13.1% D. caninum, 3.8% T. leonina, and 0.6% Taeniidae. Except for T. vulpis and D. caninum (p < 0.05) the results indicated no significant difference in the prevalence of the identified GI helminth between Lusaka and Katete. There was no significant difference in the prevalence between genders of GI helminth infestation demonstrated in this study and only A. caninum showed significant difference in prevalence by age category. The study also showed the presence of zoonotic intestinal helminths A. caninum, T. canis and D. caninum. The study highlights that there was no significant difference in spectrum and prevalence of GI helminths between urban and rural areas in Zambia. It further brings to light the importance of educating owners of dogs on the importance of regular deworming of dogs and control of ectoparasites in order to minimise the risk that these dogs pose to them and the public.

Pentosan polysulfate (PPS) is widely used as therapeutic intervention for joint diseases in human... more Pentosan polysulfate (PPS) is widely used as therapeutic intervention for joint diseases in humans and animals, while objective confirmation has not been established yet. The purpose of this study was to provide the objective measure of the efficacy of PPS. Twenty-five healthy Mongolian horses were randomly assigned in three groups. Three different doses of PPS, 0/1.2/3.0 mg/kg, were injected intramuscularly one a week for consecutive 4 weeks. On 14 and 28 days after the initial administration, relative ratios of serum COMP/CPII were 97.9/87.6/61.8 and 94.2/104.3/88.1 in 0/1.2/3.0 mg/kg PPS, respectively. The results revealed that balance of cartilage metabolism could be significantly brought to an anabolism dominant state by PPS injections in dose dependent manor in field fed horses in Mongolia

University of Zambia Journal of Agricultural and Biomedical Sciences

ARTICLE21ABSTRACTStray dogs destined for euthanasia were collected from January 2010 through Dece... more ARTICLE21ABSTRACTStray dogs destined for euthanasia were collected from January 2010 through December 2010 to determine the presence of zoonotic gastrointestinal (GI) helminths in Lusaka district. Atotal of 33 stray dogs (male=18, female=15) were collected, euthanizedandnecropsyinformationobtainedviahelminthological approach. 100% (33/33) were infected with one or more helminth parasites. Multiple infections involving nematodes and cestodes were the most prevalent (63.6%) followed by nematodes only (33.3%) with cestodes only being the least prevalent (3.0%). The most prevalent GI helminth parasite was Ancylostomacaninum(93.9%) with the least prevalent being Toxocaracanis(6.1%). No positive case of Trichinella spiraliswas reported from direct trichinoscopy examination of striated muscles. The mean count per dog of A. caninumwas 44.45 (SD ±58.0) with a range of 0 to 223. The presence of three important zoonotic gastrointestinal helminths (A. caninum,T. canisand Dipylidiumcaninum) repo...

Copyright © 2015 King Shimumbo Nalubamba et al. This is an open access article distributed under ... more Copyright © 2015 King Shimumbo Nalubamba et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. A retrospective and prospective analysis of clinical records of dogs diagnosed with Babesia infections was carried out for the years 2000 to 2013 from practices in Lusaka, Zambia. Records of 363 dogs with confirmed Babesia infections were analysed using demographic factors including sex, breed, age, and clinical signs in relation to haematological findings and Babesia species. The clinical and laboratory findings observed are described as well as Babesia species identification. The study included 18 breeds and the highest proportion were mongrels (32.2%), males representing 64.5 % of the population. The most common presenting problemswere anorexia (65.3%) and lethargy/weakness (65.3%).Themost common clinical signs were fever...

The present study attempted to bridge the paucity of information on the prevalence of canine endo... more The present study attempted to bridge the paucity of information on the prevalence of canine endoparasites in Zambia and to establish some of the risk factors that may be associated with endoparasitism. Faecal, blood and necropsy data on endoparasites were collected between January and December 2010 to determine the prevalence of canine endoparasites in two demographically and socio-economically diverse regions of Zambia. A total of 486 dogs were sampled (n = 160 Katete,predominantlyrural; n = 326 Lusaka,predominantlyurban).Faecal samples were examined by simple and centrifugal faecal flotation methods for the presence of helminth eggs. In Katete, 82.5 percent of dogs were positive for gastrointestinal (GI) helminths compared to 76 percent for Lusaka. The overall prevalence of single helminth infections were as follows: 72.1 percent Ancylostoma caninum, 11.3 percent Toxocara canis, 6.6 percent Trichuris vulpis, 6.2 percent Dipylidium caninum, 4.4 percent Toxascaris leonina and 0.7 p...

Transboundary and Emerging Diseases

CARTILAGE

Objective To investigate the role and characterize the molecular mechanisms regulating apoptosis ... more Objective To investigate the role and characterize the molecular mechanisms regulating apoptosis and autophagy in nitric oxide (NO)–induced chondrocyte cell death. Design Cell apoptosis and autophagy were evaluated in chondrocytes treated with sodium nitroprusside (SNP) combined with the presence or absence of interleukin-1 beta (IL-1β) and nutrient-deprived conditions. The concentration of nitrite was determined by Griess reaction. Activation of apoptosis and autophagy were determined by immunocytochemistry, Western blot, and quantitative real-time polymerase chain reaction (qPCR) analysis. Flow cytometry and MTT assay were used to assess cell viability. Results Cotreatment of chondrocytes with SNP and IL-1β under nutrient-deprived condition potentially enhanced the effect of NO-induced cell death. Immunocytochemistry, Western blot, and qPCR analysis indicated that treatment of chondrocytes with SNP significantly reduced autophagic activity, autophagic flux, and multiple autophagy-...

Journal of General Virology

Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wid... more Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wide host range allows individual MRV to transmit into multiple species. Although several interspecies transmission and genetic reassortment events of MRVs among humans, livestock and wildlife have been reported, the genetic diversity and geographic distribution of MRVs in Africa are poorly understood. In this study, we report the first isolation and characterization of MRVs circulating in a pig population in Zambia. In our screening, MRV genomes were detected in 19.7 % (29/147) of faecal samples collected from pigs by reverse transcription PCR. Three infectious MRV strains (MRV-85, MRV-96 and MRV-117) were successfully isolated, and their complete genomes were sequenced. Recombination analyses based on the complete genome sequences of the isolated MRVs demonstrated that MRV-96 shared the S3 segment with a different MRV isolated from bats, and that the L1 and M3 segments of MRV-117 originat...

Journal of Veterinary Medical Science

This study investigated the effects of culture time on phenotype stability of canine articular ch... more This study investigated the effects of culture time on phenotype stability of canine articular chondrocytes (CACs) in non-passaged long-term monolayer culture. Third passage (P3) CACs isolated from four cartilage samples were seeded at three different initial seeding densities (0.2 × 10 4 , 1.0 × 10 4 and 5.0 × 10 4 cells/cm 2) and maintained in monolayer condition up to 8 weeks without undergoing subculture after confluence. The characteristic changes of chondrocytes during the culture period were evaluated based on the cell morphology, cell proliferation, glycosaminoglycans (GAGs) content, DNA quantification, mRNA expression and ultrastructure of chondrocytes. Chondrocytes maintained under post-confluence condition exhibited a capability to grow and proliferate up to 4 weeks. Alcian blue staining and Dimethylmethylene blue (DMMB) assay revealed that the extracellular matrix (ECM) synthesis was increased in a time-dependent manner from 2 to 8 weeks. The chondrocyte mRNA expression profile was dramatically affected by prolonged culture time, with a significant downregulation of collagen type I, whereas the expression of collagen type II, aggrecan, Sox9 and matrix metalloproteinase 13 (MMP-13) were significantly upregulated. In addition, transmission electron microscopy (TEM) result indicated dilation of rough endoplasmic reticulum (RER) in these long-term monolayer cultured chondrocytes. These findings demonstrate that the chondrocytes phenotype could be partially redifferentiated through the spontaneous redifferentiation process in long-term cultures using standard culture medium without the addition of chondrogenic supplements or tissue-culture scaffolds.

Journal of Veterinary Medical Science

Journal of Tissue Science & Engineering

Autologous chondrocyte transplantation is a promising option for the repair of isolated osteoarth... more Autologous chondrocyte transplantation is a promising option for the repair of isolated osteoarthritic cartilage lesions that requires isolation and expansion of chondrocytes from a small cartilage biopsy prior to implantation. However, when cultured in vitro, chondrocytes lose their stable phenotype and dedifferentiate to fibroblastic-like cells. The study investigated the potential of pentosan polysulfate (PPS) sodium to restore the phenotype of dedifferentiated monolayer articular chondrocytes. Canine articular chondrocytes isolated from four cartilage samples were culture expanded to establish primary culture. First passage chondrocytes were cultured as alginate beads for 18 days under normoxia in PPS concentrations of 0, 1, 5, 15 and 40 μg/mL in 20% DMEM. Effect of PPS on type I, II and X collagen, aggrecan and Runx2 gene expression were evaluated by real-time PCR. Runx2, HIF-1α and HIF-2α protein expression were evaluated by Western blot and proteoglycan deposition was determined by Alcian blue stain. Dedifferentiated chondrocytes fully retained their phenotype as evidenced by increased synthesis of cartilage-specific genes, type II collagen and aggrecan mRNA with complete suppression of type I and X collagen at PPS concentrations of 15 and 40 μg/mL. Compared to the control, type II collagen and aggrecan mRNA were significantly upregulated (P<0.05) at 5, 15 and 40 μg/mL and 5 and 15 μg/mL PPS, respectively. PPS significantly enhanced proteoglycan with peak deposition at 5 μg/mL compared to control. HIF-1α and HIF-2α proteins were detectable at protein level for the first time under normoxia condition in alginate culture. The study demonstrates for the first time the restoration of dedifferentiated canine articular chondrocytes phenotype by combining alginate encapsulation with culture in PPS without the addition of known chondrocytic growth factors. The study confirms PPS as novel chondroinductive factor with potential to offer a solution to the major challenges that exist in cartilage tissue engineering.

Viruses

Porcine sapelovirus (PSV) has been detected worldwide in pig populations. Although PSV causes var... more Porcine sapelovirus (PSV) has been detected worldwide in pig populations. Although PSV causes various symptoms such as encephalomyelitis, diarrhea, and pneumonia in pigs, the economic impact of PSV infection remains to be determined. However, information on the distribution and genetic diversity of PSV is quite limited, particularly in Africa. In this study, we investigated the prevalence of PSV infection in Zambia and characterized the isolated PSVs genetically and biologically. We screened 147 fecal samples collected in 2018 and found that the prevalences of PSV infection in suckling pigs and fattening pigs were high (36.2% and 94.0%, respectively). Phylogenetic analyses revealed that the Zambian PSVs were divided into three different lineages (Lineages 1–3) in the clade consisting of Chinese strains. The Zambian PSVs belonging to Lineages 2 and 3 replicated more efficiently than those belonging to Lineage 1 in Vero E6 and BHK cells. Bioinformatic analyses revealed that genetic re...