The present study is aimed at finding the mutagenicity and cytotoxicity of dense form of synthetic hydroxyapatite (Source: School of Materials and Mineral Resources Engineering, Universiti Sains Malaysia) in the blood of sheep. The biomaterial was implanted in the tibia of Malin, an indigenous sheep breed of Malaysia. Blood was collected from the sheep before implantation of the biomaterial, cultured and a karyological study was made. Six weeks after implantation, blood was collected from the same animal, cultured and screened for chromosome aberrations. The mitotic indices and karyological analysis indicated that the implantation of synthetic hydroxyapatite (dense form) did not produce any cytotoxicity or chromosome aberrations in the blood of sheep.
The preterm diaphragm is functionally immature compared with its term counterpart. In utero inflammation further exacerbates preterm diaphragm dysfunction. We hypothesized that preterm lambs are more vulnerable to in utero inflammation-induced diaphragm dysfunction compared with term lambs. Pregnant ewes received intra-amniotic (IA) injections of saline or 10 mg lipopolysaccharide (LPS) 2 or 7 days before delivery at 121 days (preterm) or ∼145 days (term) of gestation. Diaphragm contractile function was assessed in vitro. Plasma cytokines, diaphragm myosin heavy chain (MHC) isoforms, and oxidative stress were evaluated. Maximum diaphragm force in preterm control lambs was significantly lower (22%) than in term control lambs ( P < 0.001). Despite similar inflammatory cytokine responses to in utero LPS exposure, diaphragm function in preterm and term lambs was affected differentially. In term lambs, maximum force after a 2-day LPS exposure was significantly lower than in controls (by ~20%, P < 0.05). In preterm lambs, maximum forces after 2-day and 7-day LPS exposures were significantly lower than in controls (by ~30%, P < 0.05). Peak twitch force after LPS exposure was significantly lower in preterm than in controls, but not in term lambs. In term lambs, LPS exposure increased the proportion of MHC-I fibers, increased twitch contraction times, and increased fatigue resistance relative to controls. In preterm diaphragm, the cross-sectional area of embryonic MHC fibers was significantly lower after 7-day versus 2-day LPS exposures. We conclude that preterm lambs are more vulnerable to IA LPS-induced diaphragm dysfunction than term lambs. In utero inflammation exacerbates diaphragm dysfunction and may increase susceptibility to postnatal respiratory failure.
This paper reports total nematode anthelmintic resistance towards albendazole, fenbendazole, levamisole and ivermectin in a commercial sheep farm located in Terengganu, Malaysia. Faecal Egg Count Reduction Test (FECRT) was conducted on 25 sheep, where five sheep in each group were treated with the respective four anthelmintics based on live bodyweight. The balance of five sheep placed in the control group were not treated with any anthelmintics. At day 13 post-treatment, faecal egg count was conducted and nematode worm egg count reduction percentage was calculated to determine the resistance status towards the respective anthelmintics tested. Results showed that nematodes were resistant to all the anthelmintics tested, namely albendazole, fenbendazole, levamisole and ivermectin with reduction percentage of 87%, 46%, 94% and 68%, respectively. Subsequently, the third stage larvae of Haemonchus contortus and Trichostrongylus colubriformis recovered from post-treatment faecal cultures were subjected to allele-specific polymerase chain reaction (AS-PCR) assay to determine the presence of the benzimidazole resistance gene. This study reports the occurrence of the classical F200Y mutation in the isotype 1 βtubulin gene, for the first time in Malaysia.
The effect of Brachiaria decumbens (signal grass) on drug-metabolizing enzymes was studied in sheep. After 14 d of grazing a pure signal grass pasture, significant declines were observed in hepatic aminopyrine N-demethylase and aniline 4-hydroxylase (phase I biotransformation) and in conjugative enzymes UDP-glucuronyltransferase and glutathione S-transferase. Kidney enzymes were significantly decreased except for UDP-glucuronyltransferase. Enzyme activities were also compared for normal sheep and cattle livers and kidneys. Lower activities were found in cattle, indicating that factors other than biotransformation are responsible for the clincial tolerance of cattle to B. decumbens toxicity.
Nigeria is naturally blessed with wide diversity of native animal genetic resources. Indigenous ruminant livestock such as cattle, camel, donkey, sheep and goat contributes largely in both protein supply, revenue generation and national economy. In Nigeria, these animal resources are mismanaged and undermined through the indiscriminate slaughter of pregnant animals and foetal losses in abattoirs. This unethical practice resulted in the loss of genetic diversity, preferred traits and superior females ruminant animals. The current research focus on reported incidences across abattoirs, which is a centre where such practice is highly occurs within the country. Lack of modern facilities, law enforcement, poor management and animal welfare in abattoirs to protect pregnant animals are among few factors responsible for an increase in incidences. It is unprofitable to continue the tradition of pregnant animal slaughter that causes foetal losses. This is a condition that significantly threatens the animal genetic resources and general livestock industry in Nigeria. This practice must be discard with a proper conservation and documentation of these valuable animal genetic resources. Both long and short terms conservation programs must aim for substantial benefits of these resources. Laws must be enforced with strict penalties to those involved in pregnant animal slaughter. Genetic resources of these species and meat industry future could be safe with proper implementation of these laws and conservation measures.
Bartonellosis is an emerging zoonotic infection responsible for a variety of clinical syndromes in humans and animals. Members of the genus Bartonella exhibit high degrees of genetic diversity and ecologic plasticity. The infection is usually transmitted to animals and humans through blood-feeding arthropod vectors such as fleas, lice, ticks and sandflies. This study was conducted to investigate the prevalence of Bartonella species in 184 beef cattle, 40 dairy cattle, 40 sheep and 40 goats in eight animal farms across Peninsular Malaysia. Bartonella-specific PCR assays and sequence analysis of partial fragments of the citrate synthase gene were used for detection and identification of B. bovis. Isolation of B. bovis was attempted from PCR-positive blood samples. Molecular heterogeneity of the isolates was investigated based on sequence analysis of gltA, ITS, rpoB genes, ERIC-PCR, as well as using an established multilocus sequence typing (MLST) method. The carriage rate of B. bovis in ticks was also determined in this study.
This study reports the molecular detection of Theileria spp. from six cattle farms, a sheep farm and a goat farm located at different states in Peninsular Malaysia. Animal blood samples were screened for the presence of Theileria DNA using a conventional polymerase chain reaction (PCR) assay. A total of 155 (69.2%) of 224 cattle investigated were PCR-positive for Theileria DNA. The occurrences of Theileria spp. ranged from 17.5% to 100.0% across six cattle farms. Theileria DNA was detected from 90.0% of 40 sheep but none of 40 goats examined in this study. Sequence analyses of amplified 18S rRNA partial fragments (335-338bp) confirmed the identification of Theileria buffeli, Theileria sergenti, and Theileria sinensis in representative samples of cattle and ticks. T. luwenshuni was identified in the infected sheep. The high occurrences of Theileria spp. in our farm animals highlight the needs for appropriate control and preventive measures for theileriosis.
Interaction of bromophenol blue (BPB) with serum albumins from different mammalian species, namely, human (HSA), bovine (BSA), goat (GSA), sheep (SSA), rabbit (RbSA), porcine (PSA) and dog (DSA) was studied using absorption and absorption difference spectroscopy. BPB-albumin complexes showed significant differences in the spectral characteristics, i.e., extent of bathochromic shift and hypochromism relative to the spectral features of free BPB. Absorption difference spectra of these complexes also showed variations in the position of maxima and absorption difference (deltaAbs.) values. Absorption difference spectra of different bilirubin (BR)-albumin complexes showed a significant blue shift accompanied by decrease in deltaAbs. values in presence of BPB which were indicative of the displacement of bound BR from its binding site in BR-albumin complexes. These changes in the difference spectral characteristics of BR-albumin complexes were more marked at higher BPB concentration. However, the extent of these changes was different for different BR-albumin complexes. Taken together, all these results suggest that BPB partially shares BR binding site on albumin and different mammalian albumins show differences in the microenvironment of the BR/BPB binding site.
Limited information is available on tropical ticks and tick-borne bacteria affecting the health of humans and animals in the Southeast Asia region. Francisella tularensis is a tick-borne bacterium which causes a potentially life-threatening disease known as tularemia. This study was conducted to determine the occurrence of Francisella spp. in questing ticks collected from Malaysian forest reserve areas. A total of 106 ticks (mainly Dermacentor and Haemaphysalis spp.) were examined for Francisella DNA using a Polymerase chain reaction (PCR) assay targeting the bacterial 16S rDNA. Francisella DNA was detected from 12 Dermacentor ticks. Sequence analysis of the amplified 16S rDNA sequences (1035 bp) show >99% identity with that of Francisella endosymbiont reported in a tick from Thailand. A dendrogram constructed based on the bacterial 16S rDNA shows that the Francisella spp. were distantly related to the pathogenic strains of F. tularensis. Three Francisella-positive ticks were identified as Dermacentor atrosignatus, based on sequence analysis of the tick mitochondrial 16S rRNA gene. Further screening of cattle and sheep ticks (Haemaphysalis bispinosa and Rhipicephalus microplus) and animal samples (cattle, sheep, and goats) did not yield any positive findings. Our findings provide the first molecular data on the occurrence of a Francisella strain with unknown pathogenicity in Dermacentor questing ticks in Malaysia.
A survey of Caseous Lymphadenitis (CLA), a bacterial infection in sheep and goats was conducted on small ruminant farms in two districts in Perak, namely Kinta and Hilir Perak. The objective of this survey is to determine the status of CLA infection in small ruminants. A total of 8 farms were screened, involving a total of 579 animals. Agar Gel Precipitation Test (AGPT) and Enzyme Linked Immuno Absorbent Assay (ELISA) were conducted on serum samples obtained from the animals. Results show that 8.5% of the animals had a positive reaction for AGPT test. It was found that 36 samples (17%) were found positive using both AGPT and ELISA methods, 9 samples (4%) were found positive only using AGPT method, 14 samples (6%)were found positive only using ELISA and 157 samples (73%) were found negative using both methods. Since there is no available data on the prevalence of the disease in the country, further epidemiological studies as well as reliable diagnostic detection methods need to be assessed for aiding in control and eradication programmes for this disease.
Delivering therapeutics to the brain using conventional dosage forms is always a challenge, thus the present study was aimed to formulate mucoadhesive nanoemulsion (MNE) of aripiprazole (ARP) for intranasal delivery to transport the drug directly to the brain. Therefore, a TPGS based ARP-MNE was formulated and optimized using the Box-Behnken statistical design. The improved in vitro release profile of the formulation was in agreement to enhanced ex vivo permeation through sheep mucous membranes with a maximum rate of permeation co-efficient (62.87 cm h-1 × 103) and flux (31.43 μg cm-2.h-1). The pharmacokinetic profile following single-dose administration showed the maximum concentration of drug in the brain (Cmax) of 15.19 ± 2.51 μg mL-1 and Tmax of 1 h in animals with ARP-MNE as compared to 10.57 ± 1.88 μg mL-1 and 1 h, and 2.52 ± 0.38 μg mL-1 and 3 h upon intranasal and intravenous administration of ARP-NE, respectively. Further, higher values of % drug targeting efficiency (96.9%) and % drug targeting potential (89.73%) of ARP-MNE through intranasal administration were investigated. The studies in Wistar rats showed no existence of extrapyramidal symptoms through the catalepsy test and forelimb retraction results. No ex vivo ciliotoxicity on nasal mucosa reflects the safety of the components and delivery tool. Further, findings on locomotor activity and hind-limb retraction test in ARP-MNE treated animals established its antipsychotic efficacy. Thus, it can be inferred that the developed ARP-MNE could effectively be explored as brain delivery cargo in the effective treatment of schizophrenia without producing any toxic manifestation.
The tight junctions between capillary endothelial cells of the blood-brain barrier (BBB) restricts the entry of therapeutics into the brain. Potential of the intranasal delivery tool has been explored in administering the therapeutics directly to the brain, thus bypassing BBB. The objective of this study was to develop and optimize an intranasal mucoadhesive nanoemulsion (MNE) of asenapine maleate (ASP) in order to enhance the nasomucosal adhesion and direct brain targetability for improved efficacy and safety. Box-Behnken statistical design was used to recognize the crucial formulation variables influencing droplet size, size distribution and surface charge of ASP-NE. ASP-MNE was obtained by incorporating GRAS mucoadhesive polymer, Carbopol 971 in the optimized NE. Optimized ASP-MNE displayed spherical morphology with a droplet size of 21.2 ± 0.15 nm and 0.355 polydispersity index. Improved ex-vivo permeation was observed in ASP-NE and ASP-MNE, compared to the ASP-solution. Finally, the optimized formulation was found to be safe in ex-vivo ciliotoxicity study on sheep nasal mucosa. The single-dose pharmacokinetic study in male Wistar rats revealed a significant increase in concentration of ASP in the brain upon intranasal administration of ASP-MNE, with a maximum of 284.33 ± 5.5 ng/mL. The time required to reach maximum brain concentration (1 h) was reduced compared to intravenous administration of ASP-NE (3 h). Furthermore, it has been established during the course of present study, that the brain targeting capability of ASP via intranasal administration had enhanced drug-targeting efficiency and drug-targeting potential. In the animal behavioral studies, no extrapyramidal symptoms were observed after intranasal administration of ASP-MNE, while good locomotor activity and hind-limb retraction test established its antipsychotic activity in treated animals. Thus, it can be concluded that the developed intranasal ASP-MNE could be used as an effective and safe tool for brain targeting of ASP in the treatment of psychotic disorders.
Spectroscopic examination of purified extracts of the rumen content of sheep intoxicated by Brachiaria decumbens revealed the presence of two spirostanes, identified as epi-sarsasapogenin and epi-smilagenin. Sarsasapogenone was obtained by the oxidation of sarsasapogenin. The reduction of sarsasapogenone using lithium aluminum hydride yielded isomeric products, sarsasapogenin (20%) and epi-sarsasapogenin (80%).
1. Anaesthesia caused marked decreases in the plasma concentrations of triiodothyronine (T3) and thyroxine (T4) and in the body temperature of young fowl. 2. Exogenous T4 or a thyroid hormone secretagogue (somatostatin antiserum), increased endogenous T3 and T4 concentrations and body temperature in conscious birds and prevented the body temperature decline in anaesthetized fowl. 3. These results provide further evidence for a role of T3 and T4 in temperature regulation in birds, particularly during anaesthesia.
In a world in which sheep producers are facing increasing problems due to the rapid spread of anthelmintic resistance, the battle against gastrointestinal parasitic nematodes is a difficult one. One of the potential new tools for integrated control strategies is biological control by means of the nematode-destroying microfungus Duddingtonia flagrans. This fungus forms sticky traps that catch developing larval stages of parasitic nematodes in the fecal environment. When resting spores (chlamydospores) of this fungus are fed daily to grazing animals for a period of time, the pasture infectivity and thus, the worm burden of grazing animals are lowered, especially in young lambs. Research has been conducted throughout the world covering many different climates and management systems. An Australian parasite model showed that if the fungus performs efficiently (> or =90% reduction in worm burden) for 2 or 3 mo, it should contribute significantly to a reduction in the number of dead lambs otherwise occurring when managed only by anthelmintic treatment and grazing management. Feeding or field trials have clearly demonstrated that dosing with a few hundred thousand spores per kilogram of live BW not only reduced the number of infective larvae but also increased the BW of the lambs compared with controls not given fungus. Initial Australian work with feeding spores by means of a block formulation or a slow-release device has shown some promise, but further work is needed to fully develop these delivery systems. In tropical Malaysia, small paddock trials and field studies resulted in significant improvements, in terms of lower worm burdens and increased live BW, when feeding half a million spores daily to grazing lambs. Additional benefits have been observed when the fungus is employed in combination with a fast rotational grazing system. Research has also demonstrated that spores can be delivered in slightly moist feed block material, but only if such blocks are consumed rapidly, because of their very short shelf life. In the northern, temperate Danish climate it has been demonstrated that daily feeding of half a million spores per kilogram of live BW can lead to significant production benefits, with increased live BW gain in fungus-exposed animals. Biological control of parasitic nematodes in sheep seems to hold promise for the future, but to be able to assist producers, the optimal delivery system needs to be refined and further developed. In addition, more work will be needed to define the best use of this technology in different geographic regions.
The extract of the Psophocarpus tetragonolobus pods has been tested for antimicrobial activity in a disk diffusion assay on eight human pathogenic bacteria and two human pathogenic yeasts. The extracts of P. tetragonolobus possessed antimicrobial activity against all tested strains. The ethanolic extract of P. tetragonolobus pods was further tested for in vivo brine shrimp lethality test and in vitro sheep erythrocyte cytotoxic assay. The brine shrimp lethality test exhibited no significant toxicity (LC(50)=1.88 mg/ml) against Artemia salina, whereas sheep erythrocyte test showed significant toxicity. The reason for haemolysis of erythrocyte was discussed. The P. tetragonolobus extract with high LC(50) value signified that this plant is not toxic to human. This result also suggested that the ethanolic extract of P. tetragonolobus pods is potential source for novel antimicrobial compounds.
One thousand and forty-five tissue samples of skeletal muscles, tongue, heart, diaphragm and esophagus were collected from 209 animals (43 sheep, 89 goats and 77 cattle) from an abattoir in Selangor between February and October, 2013. Each sample was divided into three pieces with each piece measuring 2-3 mm3. Each piece was then squeezed between two glass slides and examined microscopically at x 10 magnification for the presence of sarcocystosis. Three positive samples from each animal species were then fixed in 10% formalin for histological processing. Seven positive samples collected from each animal species were preserved at -80°C or 90% ethanol for gene expression studies. Microsarcocysts were detected in 114 (54.5%) animals by light microscopy (LM). The infection rates in sheep, goat and cattle were 86, 61.8 and 28.6% respectively. The highest rate of infection was in the skeletal muscles of sheep (64.9%) and goats (63.6%) and in the heart of cattle (63.6%). The cysts were spindle to oval in shape and two stages were recognized, the peripheral metrocytes and centrally located banana-shaped bradyzoites. 18S rRNA gene expression studies confirmed the isolates from the sheep as S. ovicanis, goats as S. capracanis and cattle as S. bovicanis. This, to the best of our knowledge, is the first molecular identification of an isolate of S. ovicanis and S. capracanis in Malaysia. Further studies with electron microscopy (EM) are required in the future to compare the features of different types of Sarcocysts spp.
Corynebacterium pseudotuberculosis is the etiological agent of caseous lymphadenitis that affects sheep and goats. This study was designed to determine the presence of the causative organism in the female reproductive organs and associated lymph nodes in non-pregnant does experimentally inoculated through intradermal route in the chronic form.
Laboratories intending to adopt cycle sequencing of PCR products in their routine analysis often face a confusing range of methods and kits. Through the study of mitochondrial cytochrome b, we have shown that clean and highly reproducible sequences could be obtained by using a combination of existing simple and economical methods in the preparation of DNA templates, PCR, purification of PCR products and sequencing. Our protocol is useful in itself or as a standard in typing other PCR-amplified DNA at the population level.