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Contagious ecthyma (CE) is an infectious disease of small ruminants caused by a parapoxvirus of family Poxviridae subfamily Chordopoxvirinae. The disease is obviously distinguished by an establishment of scabby lesions and ulcerative formation on less hairy areas including muzzle, ears, nostril, and sometimes on genitalia. The disease is endemic in sheep and goats. The virus is transmissible to other ruminants and is a public health concern in humans. Although the disease is known as self-limiting, it may cause a significant economic threat and financial losses due to lower productivity in livestock production. Information with regard to the risk of the disease and epidemiology in most parts of the world is underreported. This paper aims to provide relevant information about the epidemiology of CE in selected regions of Europe, South America, North America, Asia, Africa, and Australia. An in-depth comprehension of virus infection, diagnoses, and management of the disease will enable farmers, researchers, veterinarians, abattoir workers, health personnel, and border controllers to improve their measures, skills, and effectiveness toward disease prevention and control, toward reducing unnecessary economic loss among farmers. A herd health program for significant improvement in management and productivity of livestock demands a well planned extension program that ought to encourage farmers to equip themselves with adequate skills for animal healthcare.

There is a scarcity of information for a reliable and safe protocol for semen collection in R. timorensis. The induction of general anesthesia by Ketamine-Xylazine in both intravenous and intramuscular routes provides valuable information for the practicality of the procedure. Five healthy and matured Rusa timorensisstags, aged more than 3 years, weighing an average of 68.5 ± 2.4 kg were selected for this experiment. General anaesthesia was conducted by using a combination of Ketamine and Xylazine at the dosage of 2mg/kg, and 1mg/kg respectively in both intramuscular and intravenous routes. The procedure was performed from April to November 2012 resulting in 30 collections. The physiological parameters included were respiratory rate, heart rate and temperature before, during and post-procedure. The onset, recovery and total time are significantly different between IV and IM route at 5.13 ± 0.88 min, 7.47 ± 1.61 min, 18.67 ± 1.47 min, and 11.67 ± 1.44 min, 11.73 ± 1.03 min and, 29.07 ± 4.01 min respectively. Meanwhile, there is no significant difference in the duration time of anesthesia between IV and IM route at 15.27 ± 3.03 min and 17.40 ± 3.55 min respectively. Both routes provide the same success rate and yield the same semen quality, however, the intravenous route is desirable due to fast induction and recovery time. General anaesthesia is integral for the restraining of wild cervidae, intravenous route of drug administration should be considered for semen collection in Rusa deer.

Caseous lymphadenitis (CLA) is a chronic bacterial disease that causes economic losses in small ruminant throughout the world. Corynebacterium pseudotuberculosis is the causative agent of CLA characterized by large abscesses internally/externally, abortions, weight loss, and decreased wool production in sheep and goat farming. There is paucity of information on Corynebacterium pseudotuberculosis and its immunogen mycolic acid responses in male goat during the course of CLA. As a result, this study was designed to investigate the clinical responses and pathological changes of the reproductive tract and its associated lymph nodes in male goats following inoculation with Corynebacterium pseudotuberculosis and its immunogen mycolic acids. A total of 12 healthy crossbred Boer male goats aged 16–20 months were divided into three groups, 1, 2, and 3, of four in each group. Group 1 (control group) was inoculated with 2 ml of sterile phosphate-buffered solution (PBS) via intradermal route, group 2 was inoculated with 2 ml 1 × 109 colony forming unit (cfu) of C. pseudotuberculosis intradermally and group 3 was inoculated with 2 ml of mycolic acid (1 g/ml) intradermally. The bucks were observed and assessed for changes in clinical responses daily for 60 days post challenged. At the end of the experiment, animals were slaughtered and organs of the reproductive tract and lymph nodes were harvested for histopathological investigations. In this study, all experimental bucks survived throughout the study period with some evidence of clinical manifestation such as pyrexia, weight loss, and enlargement or rupture of the lymph nodes. Microscopically, C. pseudotuberculosis and mycolic acid challenged groups showed significant cellular changes in the reproductive system and lymph nodes. Group 2 showed abscessation of the lymph nodes while group 3 did not indicate any abscess formation in the lymph nodes. For mycolic acid extract, the result revealed only mild to moderate clinic-pathological responses in bucks with no abscess formation in the lymph nodes. The result obtained added knowledge in the field of CLA research in small ruminants. In conclusion, this research revealed that C. pseudotuberculosis has a negative effect on reproductive performances of bucks and affects the clinical responses and abscess formation.

Corynebacterium pseudotuberculosis is the causative agent of caseous lymphadenitis, a debilitating chronic disease of sheep and goats. Little is known about the buck's reproductive pathophysiology with respect to inoculation with Corynebacterium pseudotuberculois and its immunogen mycolic acid extract. Therefore, this present study was designed to determine the concentration of testosterone hormone, pro-inflammatory cytokines, and semen quality of the experimental animals. A total of 12 bucks, divided into groups 1, 2, and 3 (Negative control group, Positive control group and Mycolic acid group respectively), were enrolled in this study. Following inoculation, all goats were observed for clinical responses and monitored for 60 days post-challenge and were then sacrificed. Blood samples were collected via the jugular once before inoculation and on a weekly basis post-challenge. Semen samples were collected 2 weeks post-challenge and prior to the sacrifice of the experimental animals. During the post inoculation period of 60 days, the concentration of testosterone hormone for group 2 was increased significantly (p 0.05) in weeks 5, 6, and 9 but decreased in weeks 2 and 7 post inoculation. In group 3, the mean concentration of testosterone was increased significantly (p 0.05) in weeks 5, 6, 7, and 9 post inoculation but decreased in week 2. The concentration of interleukin 6 (IL 6) in treated group 2 did not show any significant difference (p 0.05) but increased significantly (p 0.05) in week 2 post inoculation in group 3. For concentration of interleukin 1β (IL1β) in both treated groups 2 and 3 showed significant difference (p 0.05) in weeks 2 and 3 post inoculation. The tumor necrosis factor-alpha (TNF-α) concentration in both treated groups 2 and 3 did not show any significant difference (p 0.05) as compared to group 1. The concentration of interferon-γ (IFNγ) significantly increased (p 0.05) for group 2 for weeks 2, 3, 4, and 5 where else for group 3 was not in significant difference (p 0.05) compared to group 1. Both group 2 and group 3 showed a reduction in semen qualities as compared to group 1, but the severity was more intense in group 2 if compared to group 3. In conclusion, therefore, the present study concluded that the mycolic acid group revealed significant responses of testosterone hormone concentration, semen quality, and its related pro-inflammatory cytokines in bucks following infection but the severity lesser compared to Corynebacterium pseudotuberculosis group.

Objective This study investigated the suitability of siRNA targeting specific genes that cause inhibition of virus replication in vitro especially for the virus that capable of crossing placenta and we employed a novel transplacental rat cytomegalovirus that mimics infection in human. Methods Six unique siRNAs with three each targeting different regions of IE2 (ie2a, ie2b and ie2c) and DNA polymerase (dpa, dpb and dpc) were prepared and tested for antiviral activities. The efficacy as an antiviral was determined in in-vitro by measuring TCID50 virus titer, severity of virus-induced cytopathic effect (CPE), intracellular viral genome loads by droplet digital PCR, the degree of apoptosis in siRNA-treated cells and relative expression of viral mRNA in infected Rat Embryo Fibroblast (REF) cells. Findings Remarkably, the siRNAs: dpa, dpb and IE2b, significantly reduced virus yield (approximately >90%) compared to control group at day 18 post infection (p.i). Changes in CPE indicated that DNA polymerase siRNAs were capable of protecting cells against CMV infection at day 14 p.i with higher efficiency than GCV (at the concentration of 300 pmol). Gene expression analysis revealed a marked down regulation of the targeted DNA polymerase gene (73.9%, 96.0% and 90.7% for dpa, dpb and dpc siRNA, respectively) and IE2 gene (50.8%, 49.9% and 15.8% for ie2a, ie2b and ie2c siRNA, respectively) when measured by RT-qPCR. Intracellular viral DNA loads showed a significant reduction for all the DNA polymerase siRNAs (dpa: 96%, dpb: 98% and dpc:92) compared to control group (P Conclusion In conclusion, this study clearly highlighted the feasibility of RNAi as an alternative antiviral therapy that could lead to controlling the CMV infection.

Background Intramammary Infections are costly endemic diseases in dairy cows and it highly affect both the quality and quantity of milk production in addition to the animal wellbeing. It a major cause of considerable economic losses to the dairy farmers. This study was aimed to determine the prevalence of mastitis pathogens and antimicrobial resistance pattern of selected most prevalent pathogens. Results The overall culture prevalence was 67.6% (n = 503/744, CI = 64.1–70.9). The respective states’ culture prevalence was 66.4% (n = 71/107, CI = 56.5–75.0), 60.3% (184/305, CI = 54.6–65.8), 58% (n = 94/162, CI = 50-65.6), 100% (n = 75/75, CI = 93.9–100), and 83.3% (n = 79/95, CI = 73.8–89.8) for Pahang, Perak, Selangor, Negeri Sembilan and Johor respectively. The most prevalent isolates identified were non-aureus staphylococci (NAS) 39.5%, S. aureus 13.1%, K. pneumoniae 6.5%, S. agalactiae 4.8%, and S. uberis 4.3%. Resistance profile of Actinobacter spp against chloramphenicol and streptomycin were 71.4% and 35.7% respectively. For S. uberis isolates, 66.7% and 73.3% have shown resistance to tetracycline and streptomycin respectively. Conclusions This study provides knowledge of the epidemiology of bacterial isolates associated with subclinical mastitis in Malaysia and their resistance profile. The high prevalence of these pathogens in milk and multidrug resistance observed should not be over looked.

Background Cytomegalovirus (CMV) is an opportunistic pathogen that causes severe complications in congenitally infected newborns and non-immunocompetent individuals. Developing an effective vaccine is a major public health priority and current drugs are fronting resistance and side effects on recipients. In the present study, with the aim of exploring new strategies to counteract CMV replication, several anti-CMV siRNAs targeting IE2 and DNA polymerase gene regions were characterized and used as in combinations for antiviral therapy. Methods The rat embryo fibroblast (REF) cells were transfected with multi siRNA before infecting with CMV strain ALL-03. Viral growth inhibition was measured by tissue culture infectious dose (TCID50), cytopathic effect (CPE) and droplet digital PCR (ddPCR) while IE2 and DNA polymerase gene knockdown was determined by real-time PCR. Ganciclovir was deployed as a control to benchmark the efficacy of antiviral activities of respective individual siRNAs. Results There was no significant cytotoxicity encountered for all the combinations of siRNAs on REF cells analyzed by MTT colorimetric assay (P > 0.05). Cytopathic effects (CPE) in cells infected by RCMV ALL-03 had developed significantly less and at much slower rate compared to control group. The expression of targeted genes was downregulated successfully resulted in significant reduction (P Conclusions In conclusion. The study demonstrated a tremendous promise of innovative approach with the deployment of combined siRNAs targeting at several genes simultaneously with the aim to control CMV replication in host cells.

Orf disease is known to be enzootic among small ruminants in Asia, Africa, and some other parts of the world. The disease caused by orf virus is highly contagious among small ruminant species. Unfortunately, it has been neglected for decades because of the general belief that it only causes a self-limiting disease. On the other hand, in the past it has been reported to cause huge cumulative financial losses in livestock farming. Orf disease is characterized by localized proliferative and persistent skin nodule lesions that can be classified into three forms: generalized, labial and mammary or genitals. It can manifest as benign or malignant types. The later type of orf can remain persistent, often fatal and usually causes a serious outbreak among small ruminant population. Morbidity and mortality rates of orf are higher especially in newly infected kids and lambs. Application of antibiotics together with antipyretic and/or analgesic is highly recommended as a supportive disease management strategy for prevention of subsequent secondary microbial invasion. The presence of various exotic orf virus strains of different origin has been reported in many countries mostly due to poorly controlled cross-border virus transmission. There have been several efforts to develop orf virus vaccines and it was with variable success. The use of conventional vaccines to control orf is a debatable topic due to the concern of short term immunity development. Following re-infection in previously vaccinated animals, it is uncommon to observe the farms involved to experience rapid virus spread and disease outbreak. Meanwhile, cases of zoonosis from infected animals to animal handler are not uncommon. Despite failures to contain the spread of orf virus by the use of conventional vaccines, vaccination of animals with live orf virus is still considered as one of the best choice. The review herein described pertinent issues with regard to the development and use of potential effective vaccines as a control measure against orf virus infection.

Polycyclic aromatic hydrocarbons are pollutants which are persistent in nature. The aryl hydrocarbon receptor is a ligand-activated cytosolic transcription factor activated by xenobiotics. The objective was to isolate and identify AHR mRNA transcript in immune organs of developing chicks and to interpret the correlation between AHR induction and dose of PAHs. Specific pathogen free embryonated eggs on day nine were inoculated with solutions of pyrene, phenanthrene, and fluoranthene dissolved in tricaprylin (vehicle) through the allantoic route at three dose levels: 0.2 mg/kg, 2 mg/kg, and 20 mg/kg. A 650 base pair product was observed by RNA extraction and reverse transcription PCR from thymus, bursa of Fabricius and spleen on 21st day. When AHR concentration was analyzed by ELISA in these organs, pyrene showed maximum potency in inducing AHR in thymus. Fluoranthene made highest concentration of AHR in bursa of Fabricius. None of these chemicals caused an increase in AHR concentration in spleen.

Haptoglobin (Hp) and Serum Amyloid A (SAA) are a group of blood proteins whose concentrations in animals can be influenced by infection, inflammation, surgical trauma or stress. Corynebacterium pseudotuberculosis is the causative agent of caseous lymphadenitis (CLA), and Mycolic acid is a virulent factor extracted from C. pseudotuberculosis. There is a dearth of sufficient evidence on the clinical implication of MAs on the responses of Hp and SAA in goats. Therefore, this study was conducted to evaluate the potential effects of Mycolic acid (MAs) and C. pseudotuberculosis on the responses of Hp and SAA in female goats. A total of 12 healthy female goats was divided into three groups; A, B and C each comprising of 4 goats and managed for a period of three months. Group (A) was inoculated with 2 mL of sterile phosphate buffered saline (as a negative control group) intradermally, while group (B) and (C) were inoculated intradermally with 2 ml each of mycolic acid and 1‏ × 109 cfu of active C. pseudotuberculosis respectively. The result of the study showed that the Hp concentration in goats inoculated with C. pseudotuberculosis was significantly increased up to 7-fold (1.17 ± 0.17 ng/L) while MAs showed a 3-fold increased (0.83 ± 0.01 ng/L) compared with the control. Whereas SAA concentration in C. pseudotuberculosis and MAs groups showed a significant 3-fold (17.85 ± 0.91 pg/mL) and 2-fold (10.97 ± 0.71 pg/mL) increased compared with the control. This study concludes that inoculation of C. pseudotuberculosis and MAs have significant effects on Hp and SAA levels, which indicates that MAs could have a role in the pathogenesis of caseous lymphadenitis.

Innumerable Escherichia coli of animal origin are identified, which are of economic significance, likewise, cattle, sheep and goats are the carrier of enterohaemorrhagic E. coli, which are less pathogenic, and can spread to people by way of direct contact and through the contamination of foodstuff or portable drinking water, causing serious illness. The immunization of ruminants has been carried out for ages and is largely acknowledged as the most economical and maintainable process of monitoring E. coli infection in ruminants. Yet, only a limited number of E. coli vaccines are obtainable. Mucosal surfaces are the most important ingress for E. coli and thus mucosal immune responses function as the primary means of fortification. Largely contemporary vaccination processes are done by parenteral administration and merely limited number of E. coli vaccines are inoculated via mucosal itinerary, due to its decreased efficacy. Nevertheless, aiming at maximal mucosal partitions to stimulate defensive immunity at both mucosal compartments and systemic site epitomises a prodigious task. Enormous determinations are involved in order to improve on novel mucosal E. coli vaccines candidate by choosing apposite antigens with potent immunogenicity, manipulating novel mucosal itineraries of inoculation and choosing immune-inducing adjuvants. The target of E. coli mucosal vaccines is to stimulate a comprehensive, effective and defensive immunity by specifically counteracting the antibodies at mucosal linings and by the stimulation of cellular immunity. Furthermore, effective E. coli mucosal vaccine would make vaccination measures stress-free and appropriate for large number of inoculation. On account of contemporary advancement in proteomics, metagenomics, metabolomics and transcriptomics research, a comprehensive appraisal of the immeasurable genes and proteins that were divulged by a bacterium is now in easy reach. Moreover, there exist marvellous prospects in this bourgeoning technologies in comprehending the host bacteria affiliation. Accordingly, the flourishing knowledge could massively guarantee to the progression of immunogenic vaccines against E. coli infections in both humans and animals. This review highlight and expounds on the current prominence of mucosal and systemic immunogenic vaccines for the prevention of E. coli infections in ruminants.

Aim: The objective of this research is to report parameters for breeding soundness evaluation, semen extension, and cryopreservation in Rusa timorensis. Materials and Methods: Seven healthy stags were chosen for semen collection using an electroejaculator. The collections were performed twice in a breeding season between February and June 2016. Samples were collected between 2 and 3 weeks interval, collected twice for each animal. Semen was evaluated, extended, and cryopreserved using four different extenders; Andromed®, BioXcell®, Triladyl®, and a modified Tris-egg yolk combined with Eurycoma longifolia Jack. Results: R. timorensis semen characteristics according to volume (ml), color, sperm concentration (106/ml), general motility (%), progressive motility (%), and % morphology of normal spermatozoa are 0.86±0.18 ml, thin milky to milky, 1194.2±346.1 106/ml, 82.9±2.8%, 76.1±4.8%, and 83.9±4.8%, respectively. Conclusion: Semen characteristics of R. timorensis collected by electroejaculation is good allowing for cryopreservation and future artificial insemination work. The most suitable extender for Rusa deer semen is Andromed®.

Limit of detection (LOD), limit of quantification, and the dynamic range of detection of hepatitis B surface antigen antibody (anti-HBs) using a surface plasmon resonance (SPR) chip-based approach with Pichia pastoris-derived recombinant hepatitis B surface antigen (HBsAg) as recognition element were established through the scouting for optimal conditions for the improvement of immobilization efficiency and in the use of optimal regeneration buffer. Recombinant HBsAg was immobilized onto the sensor surface of a CM5 chip at a concentration of 150 mg/L in sodium acetate buffer at pH 4 with added 0.6% Triton X-100. A regeneration solution of 20 mM HCl was optimally found to effectively unbind analytes from the ligand, thus allowing for multiple screening cycles. A dynamic range of detection of ∼0.00098-0.25 mg/L was obtained, and a sevenfold higher LOD, as well as a twofold increase in coefficient of variance of the replicated results, was shown as compared with enzyme-linked immunosorbent assay (ELISA). Evaluation of the assay for specificity showed no cross-reactivity with other antibodies tested. The ability of SPR chip-based assay and ELISA to detect anti-HBs in human serum was comparable, indicating that the SPR chip-based assay with its multiple screening capacity has greater advantage over ELISA.

Intramammary infections are costly endemic disease in dairy cows and it highly affects both the quality and quantity of milk production in addition to the animal well-being. It a major cause of considerable economic losses to the dairy farmers. This study was aimed to determine the prevalence of mastitis pathogens and antimicrobial resistance pattern of selected most prevalent pathogens. A total of 1945 quarter samples from 517 cows across 33 dairy herds were used. Isolation and identification of mastitis pathogens was carried out based on standard bacteriological procedures. Antimicrobial resistance profile was conducted by disk diffusion technique. The overall culture prevalence was 67.6% (n=503/744, CI= 64.1-70.9). The respective states’ culture prevalence were 66.4% (n=71/107, CI= 56.5-75.0), 60.3% (184/305, CI= 54.6-65.8), 58% (n=94/162, CI= 50-65.6), 100% (n=75/75, CI= 93.9-100), and 83.3% (n=79/95, CI= 73.8-89.8) for Pahang, Perak, Selangor, Negeri Sembilan and Johor respectively. The most prevalent isolates identified were non-aureus staphylococci (NAS) 39.5%, S. aureus 13.1%, K. pneumoniae 6.5%, S. agalactiae 4.8%, and S. uberis 4.3%. Resistance profile of K. pneumoniae isolates against ampicillin and Penicillin G were 70.4% and 88.9% respectively while that of Actinobacter spp against chloramphenicol and streptomycin were 71.4% and 35.7% respectively. For S. uberis isolates, 66.7% and 73.3% have shown resistance to tetracycline and streptomycin respectively. This study provides knowledge of the epidemiology of bacterial isolates associated with subclinical mastitis in Malaysia and their resistance profile. The high prevalence of these pathogens in milk and multidrug resistance observed should not be over looked.

We report the whole-genome sequence of Escherichia coli sequence type 127 (ST127) strain 1538RHQ, recovered from a mastitic cow in a dairy herd in Selangor, Malaysia. The objective of this study was to identify the antigenic and virulence properties that can be used as suitable targets for vaccine development against bovine mastitis.

There is a little information on the characterization of Orf virus strains that are endemic in Malaysia. The relationship between the severity of disease and the molecular genetic profile of Orf virus strains has not been fully elucidated. This study documented the first confirmed report of contagious ecthyma causing by Orf virus in goats from a selected state of eastern peninsular Malaysia. The disease causes significant debilitation due to the inability of affected animals to suckle which brings a great economic loss to the farmers. A total of 504 animals were examined individually to recognize the affected animals with Orf lesion. Skin scrapping was used to collect the scab material from the infected animals. The presence of Orf virus was confirmed by combination of methods including virus isolation on vero cells, identification by Transmission Electron Microscopy (TEM) and molecular technique using PCR and Sanger sequencing. The results showed the successful isolation of four Orf virus strains with a typical cytopathic effects on the cultured vero cells line. The morphology was confirmed to be Orf virus with a distinctive ovoid and criss cross structure. The phylogenetic analysis revealed that these isolated strains were closely related to each other and to other previously isolated Malaysian orf viruses. In addition these Orf virus strains were closely related to Orf viruses from China and India. This study provides more valuable insight in terms of genotype of Orf virus circulating in Malaysia.

Volume 8, no. 6, e01618-18, 2019, [https://doi.org/10.1128/MRA.01618-18][1]. Page 1: The author byline should appear as given in this correction. [1]: /lookup/doi/10.1128/MRA.01618-18

Streptococcus agalactiae, commonly known as group B streptococcus (GBS), is among the most implicated pathogens in bovine mastitis worldwide. Proper control measures can curb both economic and public health effects it may cause., Streptococcus agalactiae, commonly known as group B streptococcus (GBS), is among the most implicated pathogens in bovine mastitis worldwide. Proper control measures can curb both economic and public health effects it may cause. Here, we report the sequenced genome of S. agalactiae sequence type 167 (ST167) strain 3966RFQB obtained from a bovine mastitis case at a dairy herd in Banting, Selangor, Malaysia (longitude 2.8121°N, latitude 101.5026°E).

Mastitis is the inflammation of the mammary gland due to microbial infiltration causing a reduced mammary function. This study aims at developing a vaccine using Malaysian local isolate of Staphylococcus aureus and evaluating serum amyloid A, Interleukin-10, IgM and IgG responses periodically. Four bacterin concentrations (106, 107, 108 and 109 cfu/ml of the local isolate of S. aureus) were adjuvanted with aluminium potassium sulphate. Thirty cows grouped into 4 treatment groups (G-) were vaccinated (2 ml) intramuscularly, with a fifth G-A as control. The mean concentration (MC) of serum amyloid A (SAA) was significantly different (sig-d) (p ˂ 0.05) in G-D at 0 h post vaccination (PV), 3 h PV, 24 h PV, weeks 1, 2, 3 and 4 PV (6-, 15-, 5-, 12-, 11-, 4- and 11-fold increased (FI) respectively). The MC of serum amyloid A was also sig-d in G-E at 0 h PV, weeks 1, 2 and 4 PV (3, 8, 5 and 8 FI respectively). The MC of IL-10 was sig-d in G-D and C at 3 h PV and week 2 PV (5 and 2 FI respectively). The IgM MC was sig-d in G-B and C at 3 h PV (5 and 6 FI respectively), at 24 h PV (5 and 9 FI respectively), at week 3 PV(2 and 2 FI respectively) and week 4 PV (3 and 4 FI respectively). The MC of IgG was sig-d in G-E at 0 h, 3 h and week 3 PV(5, 6 and 2 FI respectively) and in G-D at weeks 1–4 (3, 3, 3 and 5 FI respectively). In conclusion, elevated levels of SAA, IgG and IL-10 in G-D(108) informed our choice of best dosage which can be used to evoke immunity in cows.

Mastitis is an inflammatory condition of the udder that occurs as a result of the release of leucocytes into the udder in a response to bacterial invasion. The major causes of mastitis are an array of gram positive and negative bacteria, however, algae, virus, fungi, mechanical or thermal injury to the gland have also been identified as possible causes. Mastitis vaccines are yet to be developed using Malaysian local isolate of bacteria. The objective of the present experimental trial was to develop a monovalent vaccine against mastitis using S. aureus of Malaysian isolate and to evaluate the clinical responses such as temperature, respiratory rates and heart rates in vaccinated cows. S. aureus is a major causative bacteria in clinical and subclinical types of mastitis in cows. Four concentrations of the bacterin (106, 107, 108 and 109 cfu/ml of the local isolate of S. aureus) were prepared using Aluminium potassium sulfate adjuvant. Thirty cows were grouped into four treatment groups (B, C, D and E) with a fifth group as control (A). These groups were vaccinated intramuscularly(IM) with the prepared monovalent vaccine and its influence on the vital signs were intermittently measured. The mean of rectal temperature was significantly different (p˂ 0.05) at 0hr Post Vaccination [1]" in groups D and E (39.5 ± 0.15 °C and 39.4 ± 0.15 °C respectively) and at 3 h PV in groups C, D and E (39.8 ± 0.14 °C, 39.9 ± 0.14 °C and 40.3 ± 0.14 °C respectively) compared to the control group. This indicated a sharp increased rectal temperatures between 0hr and 3 h PV in groups C, D and E which later declined at 24 h PV. The mean of rectal temperature of group E was significantly different (p˂ 0.05) at weeks 1 and 2 PV (39.87 ± 0.19 °C and 39.80 ± 0.18 °C respectively) compared to the control group. The mean of heart rate was significantly different (p˂ 0.05) at week 1 PV in groups D and E (83.0 ± 3.8 beats/minute and 80.0 ± 3.8 °C respectively) compared to control. A trending decrease was however observed in heart rates of group E from weeks through 4 PV and in group D from weeks 1 through 3 PV. The mean of respiratory rates was significantly different (p˂ 0.05) at week 3 PV in group B and D (31.0 ± 1.2 breaths/minute and 28.0 ± 1.2 breaths/minute) compared to control. In conclusion, this study highlights responses of these vital signs due to vaccination against S. aureus causing mastitis in cows. To the best of our knowledge the findings of this study adds value to the shallow literature on vital signs alterations in cows vaccinated against mastitis as elevated levels of temperature and heart rates of group D and E indicated obvious response.

Univariate screening on factors affecting the purification performance of recombinant hepatitis B surface antigen (HBsAg) on ion exchange chromatography (IEC) and size exclusion chromatography (SEC) and the establishment of a two-step purification strategy were performed. Amongst four IEC adsorbents examined, the use of Q Sepharose XL IEC adsorbent under optimized conditions together with optimized SEC purification was able to efficiently purify HBsAg. An established purification strategy comprising the two techniques further demonstrated adaptability for scale-up operations with a final total purification factor (PF) of 94.82 ± 16.20, HBsAg purity of 95.48% and recovery yield of 78.07%.

Gonado-hypophyseal lesions and reproductive hormonal changes in Brucella melitensis-infected mice and its lipopolysaccharide (LPS) were investigated. Forty-eight mice (24 males and 24 females) were used as infection models in this study, and they were divided into three equal separate groups of eight mice based on their sex: M1, M2, and M3 and F1, F2, and F3 (M = Male, F = Female). Groups M1 and F1 were orally inoculated with sterile phosphate-buffered saline (PBS) and served as the negative control. Groups M2 and F2 were orally inoculated with 0.4 ml of 109 colony-forming units (cfu) of B. melitensis, while groups M3 and F3 were orally inoculated with 0.4 ml of LPSs from 109 cfu of B. melitensis. All the groups were observed for 10 days for changes in clinical signs or signs of mortality and then euthanized. Whole blood was collected via cardiac puncture, while tissues were fixed in 10 % formalin for histopathological processing. Histopathological lesions in the male and female reproductive tracts ranged from mild to moderate degenerative, inflammatory, and necrotic lesions. These changes were moderate in M2 than in M3 in males and in F3 than in F2 groups in females. Lesions in the pituitary gland ranged from mild to moderate inflammation and necrosis with astrogliosis. F2 and M2 groups had more lesion distribution in the pituitary gland than F3 and M3 groups. There was a 13 % decline in progesterone concentration in F2 and a 44 % decline in F3. Estradiol level declined by 33 % in F2 and slightly increased by 4.2 % in F3. Progesterone-estrogen ratio (P/E) increased by 31 % in F2 and declined by 46 % in F3. Testosterone concentration increased by 74 % in M2 and decreased by 56 % in M3. The study showed that oral administration of B. melitensis affects the pituitary gland and may alter the hypothalamic-pituitary-gonadal axis.

The gram-negative bacterium Pasteurella multocida serotype B: 2 is an important veterinary and opportunistic human pathogen that causes deplorable welfare problems and colossal economic production losses in bovine and other animal species throughout the world. Pasteurella multocida type B: 2 and its LPS are associated with Haemorrhagic Septicaemia (HS) in mice as animal model. Sixty healthy Balb c mice were placed in twelve plastic cages each one containing five mice. The mice were divided into three major groups (A, B and C). Group A is the control group (n = 10) and these were inoculated with 0.4 mL of PBS pH 7.4 orally. The treatment groups (B; n = 25 and C; n = 25) were inoculated with P. multocida type B: 2 and lipopolysaccharide respectively. The mice in group B and C were further divided into five subgroups. The subgroups were designated based on the graded doses as B101, B103, B105, B107 and B109 for Pasteurella multocida and C101, C103, C105, C107 and C109 for LPS respectively. The mice were observed for 120 h post-inoculation. The concentration of IL-1I² was significantly higher (p

A study of the Mut+ phenotype for the expression of recombinant hepatitis B surface antigen (HBsAg) in Pichia pastoris strain GS115 using the pPIC3.5K vector with a two-phase fed-batch protocol in a shake flask system is described. Expression levels of HBsAg protein of 6.74 g L−1 Dry Cell Weight (DCW) and 26.07 mg L−1 of HBsAg concentration were achieved 48 h from the induction point which added to a 120 h reduction in production period in comparison with MutS expression (168 h). The use of the pPIC3.5K-HBsAg plasmid in the Mut+ phenotype enhanced the expression of HBsAg by a nearly 13 times higher volumetric productivity in the first 24 h and 35 times higher at peak production concentration. Comparison of AOX expression cassettes relative to the HBsAg gene in the role of mRNA secondary structure during translation initiation revealed that HBsAg possesses lower folding stability with AOX1 Mut+ phenotype. The results from this study demonstrated that expression of HBsAg with Mut+ AOX1 promoter is adequate as an alternative for the production of HBsAg. In addition, the AOX promoter of the Mut+ phenotype was observed to be better suited for HBsAg expression, which correlates with the ease of translation initiation under shake flask conditions.

Polycyclic aromatic hydrocarbons (PAHs) are persistent pollutants and chemically a class of structurally similar chemical compounds characterized by the presence of fused aromatic rings. This research was undertaken to find out immunotoxic effects produced by pyrene, phenanthrene and fluoranthene. These chemicals were injected into developing chicks at three dose levels (0.2, 2 and 20 mg per kg) through allantioc route to rule out possible mechanisms involved in immunotoxicity. DNA adduct produced by PAHs in immune organs were analyzed by DNA adduct enzyme-linked immunosorbent assay (ELISA) kit and DNA damage was assessed by comet assay. A significant increase in the DNA adduct levels was found in thymus and bursa in 2 mg and 20 mg dose levels of pyrene, fluoranthene and phenanthrene treated groups, whereas those in spleen simulated the value of controls. Comet assay indicated that PAHs especially pyrene, fluoranthene and phenanthrene were capable of inducing increased level of comet parameters in thymus at all the dose levels. Bursa of Fabricius and spleen also showed a gradual rise in comet parameters corresponding to all dose levels, but the increase was more marked as in thymus. Thus, it can be concluded that DNA adducts produced by PAHs lead to single-strand breaks and reduced DNA repair, which ultimately begin a carcinogenic process. Hence, this experiment can be considered as a strong evidence of genotoxic potential of PAHs like pyrene, phenanthrene and fluoranthene in developing chicks.

Background Rat cytomegalovirus ALL-03 (Malaysian strain) which was isolated from a placenta and uterus of a house rat, Rattus rattus diardii has the ability to cross the placenta and infecting the fetus. To further elucidate the pathogenesis of the Malaysian strain of Rat Cytomegalovirus ALL-03 (RCMV ALL-03), detailed analysis on the viral genome sequence is crucial. Methods Genome sequencing of RCMV ALL-03 was carried out in order to identify the open reading frame (ORF), homology comparison of ORF with other strains of CMV, phylogenetic analysis, classifying ORF with its corresponding conserved genes, and determination of functional proteins and grouping of gene families in order to obtain fundamental knowledge of the genome. Results The present study revealed a total of 123 Coding DNA sequences (CDS) from RCMV ALL-03 with 37 conserved ORF domains as with all herpesvirus genomes. All the CDS possess similar function with RCMV-England followed by RCMV-Berlin, RCMV-Maastricht, and Human CMV. The phylogenetic analysis of RCMV ALL-03 based on conserving genes of herpes virus showed that the Malaysian RCMV isolate is closest to RCMV-English and RCMV-Berlin strains, with 99% and 97% homology, respectively. Similarly, it also demonstrated an evolutionary relationship between RCMV ALL-03 and other strains of herpesviruses from all the three subfamilies. Interestingly, betaherpesvirus subfamily, which has been shown to be more closely related with gammaherpesviruses as compared to alphaherpesviruses, shares some of the functional ORFs. In addition, the arrangement of gene blocks for RCMV ALL-03, which was conserved among herpesvirus family members was also observed in the RCMV ALL-03 genome. Conclusion Genomic analysis of RCMV ALL-03 provided an overall picture of the whole genome organization and it served as a good platform for further understanding on the divergence in the family of Herpesviridae.

The complete genome sequence of the ALL-03 strain of rat cytomegalovirus (RCMV) has been determined. The RCMV genome has a length of 197,958 bp and is arranged as a single unique sequence flanked by 504-bp terminal direct repeats. This strain is closely related to the English strain of RCMV in terms of genetic arrangement but differs slightly in size.

Pasteurella multocida type B: 2 is responsible for major animal diseases of economic importance in both developed and developing countries. Haemorrhagic Septicaemia (HS) could inflict devastating effects on blood tissues and organs in the host animal. Therefore, the current study aims to investigate the haematological and clinico- pathological responses in mice following oral inoculation of graded doses of Pasteurella multocida type B: 2 and its lipopolysaccharide. Sixty healthy Balb c mice were placed in twelve plastic cages each one containing five mice. The mice were divided into three major groups (A, B and C). Group A is the control group (n=10) and these were inoculated with 0.4 ml of PBS pH 7.4 orally. The treatment groups (B; n=25 and C; n=25) were inoculated with P. multocida type B: 2 and its lipopolysaccharide respectively. The mice in group B and C were further divided into five subgroups. The subgroups were designated based on the graded doses as B101, B103, B105, B107 and B109 for Pasteurella multocida and C101, C103, C105, C107 and C109 for LPS respectively. The mice were observed for 120 hours post- inoculation. The clinical signs (Ruffled fur, Ocular discharges, Level of alertness and Laboured breathing) were significantly different (p

All the three plants were analysed in liquid chromatography-mass spectrometry-electrospray ionisation to identify plant active principles. Chromatographic separation of the compound of interest was achieved on C-18 column with detection in positive ion mode. The test showed that the most active principles found in Curcuma longa L. were curcuminoids mainly curcumin, demethoxycurcumin, bisdemethoxycurcumin, and dihydrocurcumin. In Centella asiatica L. madecassic acid was found at ion/mz 490. While the active principle in Strobilanthes crispus L. is verbascoside, which was detected at ion/mz 625. As plant active principles were excellently determined, the plant bioactivities such as anti-cancer, antiviral and/or antibacterial could be explored and developed. Key words: Centella asiatica L., Curcuma longa L., Strobilanthes crispus L., liquid chromatography–mass spectrometry-electrospray ionisation (LC-ESI- MS), curcuminoids, madecassic acid, verbascosid.

Endothelial cells have been implicated as key cells in promoting the pathogenesis and spread of cytomegalovirus (CMV) infection. This study describes the isolation and culture of rat brain endothelial cells (RBEC) and further evaluates the infectious potential of a Malaysian rat CMV (RCMV ALL-03) in these cultured cells. Brain tissues were mechanically fragmented, exposed to enzymatic digestion, purified by gradient density centrifugation, and cultured in vitro. Morphological characteristics and expression of von Willebrand factor (factor VIII-related antigen) verified the cells were of endothelial origin. RBEC were found to be permissive to the virus by cytopathic effects with detectable plaques formed within 7 d of infection. This was confirmed by electron microscopy examination which proved the existence of the viral particles in the infected cells. The susceptibility of the virus to these target cells under the experimental conditions described in this report provides a platform for developing a cell-culture-based experimental model for studies of RCMV pathogenesis and allows stimulation of further studies on host cell responses imposed by congenital viral infections.

Background The successful isolation, purification, and culture of caprine islets has recently been reported. The present study shows arange of size distribution in caprine islet diameter from 50 to 250 μm, in which 80% of the total islet yield was comprised of small islets. Methods Caprine islets were isolated and purified. Islets were handpicked and the diameter of the islets was recorded using light microscopy. Viablility of the islets was analyzed by confocal microscopy. Insulin secretion assay was carried out and analyzed by ELISA. Results When tested at 48 h after isolation, these small islets were 29.3% more viable compared to the large-sized islets. Large islets showed a high ratio (P

This study probes into the prospect of cross-reactivity of HCMV with RCMV which has not been acknowledged to date. We describe the uncovering of a protein with an estimated size of between 61-68 kDa from local RCMV strains which reacted with HCMV positive sera. Our findings are a first disclosure of a plausible immunological cross-reactivity between RCMV with its human counterpart which grounds substantial interest implying existence of conserved determinants between rat and human CMV polypeptides. The cross-reactive protein most likely represents an enveloped glycoprotein, though the precise identification and its degree of similarity needs to be evidently defined and further elucidated in forthcoming experiments.

The present study described the kinetics of Rat cytomegalovirus (RCMV) infection in newborn rats by monitoring infectious virus and viral antigens in various organs, viral DNA in the blood (DNAemia) and antibody response. These parameters were evaluated quantitatively using double-antibody sandwich ELISA (DAS-ELISA), real-time PCR, indirect ELISA and virus infectivity assay. For the first time DAS-ELISA was used for detection of RCMV antigen directly from organ samples. The relationships between the presence of viral antigens in the infected organs and antibody levels were established by the Spearman's rank test. It was found that the virus was present in the blood, spleen, liver, lungs, and kidneys earlier than in the salivary glands. Furthermore, the early immunity of the newborn rats led to a delayed seroconversion. We suggested that the prolonged presence of the virus in salivary glands could augment the antibody response that conversely might be responsible for a reduction of viremia. This study expanded our understanding of RCMV pathogenesis leading to improved therapeutic and preventive treatment regimens particularly for the neonatal Human cytomegalovirus (HCMV) infections. Additionally, the detection procedures developed in this study such as DAS-ELISA and real-time PCR could serve as alternative techniques for rapid screening of large number of samples.

A total of 157 stool samples were examined for Group A rotaviruses in diarrheic children admitted to 8 different major hospitals in Malaysia. The overall incidence rate in this study was 19.7% (31 of 157) with a variation of 9.5% to 39.1% in different locations. Majority of the infections detected were in those under 2 years of age and there were fewer admissions in the older age group. The stool samples were initially screened for rotavirus Group A by latex agglutination method and followed by RNA electrophoresis. The size and the characteristics wheel-shaped morphology of the viral preparations when examined by electron-microscopy further confirmed the presence of rotaviruses in the positive stool samples. Analysis of the RNA pattern showed that majority of the isolates, 51.6% (16 of 31) were Type IIC ('long' with comigration of RNA segments 7 and 8), 35.5% (11 of 31) with Type IIG ('long' with comigration of segments 7, 8, 9), 9.7% (3 of 31) with Type IG ('short' with comigration of RNA segments 7, 8, 9) and 3.2% (1 of 31) of mixed or atypical pattern. It appeared that over a 12 year interval, only one new or unusual rotavirus electropherotype was found. This is the first comprehensive report on the electropherotypes of rotaviruses covering eight different geographical locations in Malaysia and the data obtained is useful for understanding the geographic distribution and types of rotaviruses transmitting in Malaysia.

Corynebacterium pseudotuberculosis is the causative agent of caseous lymphadenitis (CLA), a common disease in small ruminant populations across the globe. The following report describes two cases of a 3 year old Boer cross does showing large mass beneath the jaw, on the ventral side of the mandible with strong pain reaction to touch of the affected region. The consistency of the mass was non–movable, solid and hard on deep palpation. The mass was lanced and purulent discharges were evacuated. Phenotypic and biochemical identification methods allocated the isolates in C. pseudotuberculosis biovar ovis. The vast majority of the isolates was able to produce phospholipase D and was susceptible to most of the antimicrobial compounds tested. Furthermore, the isolates were compared with C. pseudotuberculosis isolated from other states with different geographical locations and showed high similarity index, suggesting the prevalence of dominant clones and a potential dissemination across the country.

A new rat cytomegalovirus (RCMV) isolated from the placenta/uterus of a house rat (Rattus rattus diardii) was found to productively infect rat embryo fibroblast (REF) cells. The virus produced typical herpesvirus-like cytopathic effects characterized by a lytic infection. The well-known herpesvirus morphology was confirmed by electron microscopy. Its slow growth in cell culture indicated that the virus is belonging to subfamily Betaherpesvirinae. Electron microscopy techniques and immunohistochemistry confirmed the presence of herpesviral inclusion bodies and virus related particles in the cytoplasm and nucleus of infected cells. Hyperimmune serum against the Maastricht strain of RCMV revealed the virus identity in neutralization test, immunoperoxidase and immunofluorescence techniques. Despite typical characteristics of CMV, the viral genome is significantly different from that of Maastricht, English, UPM/Sg and UPM/Kn strains. The dissimilarities, which have not been reported before, had been confirmed by mean of restriction endonuclease analysis. The new RCMV strain, a virus that infects placenta and uterus of rats, has been named as ALL-03.

Both wild-type virulent and mutant strains of pseudorabies virus (PrV) were used in this study. Mutants used were derived from the plaque purified strain PrVmAIP. A total of six drug resistant mutants, three bromodeoxyuridine (BUdR) resistant and three iododeoxyuridine (IUdR) resistant, respectively, were isolated and passaged in chicken embryo fibroblast (CEF) cells. The DNA of these PrVs were compared with the wild-type isolates by means of the restriction fragment pattern (RFP) findings produced with Bam HI, Kpn I, Hind III and Bgl II restriction enzymes (RE). Compared to the wild-type PrVs (PrV-VBA1-parental strain of PrVmAIP; PrV-VBA2; PrV-VBA3), the RFP of PrVmAIP showed the presence of mutations within the RE sites studied. Both PrV-VBA1 and PrV-VBA2 appeared to be closely related but their RFPs differed from PrV-VBA3. Significant differences either in the number, size or migrations of the DNA fragments could also be detected in the BUdR resistant strains. Even though different features of cytopathic effect (GPE) were observed in the IUdR resistant PrVs, the RFP findings remained identical. The PrVs studied showed considerable differences from the reference PrV (PrV-CD).

The genomic mapping of Red Jungle Fowl (Gallus gallus), local Village Chicken, and broiler was carried out by random amplified polymorphism DNA (RAPD) technique. Two different sets of arbitrary primers were used (Operon OPAO1-20 and Genemed GM01-50). All the genomes of the three species of chickens were amplified with OPA01-20 primers. The genomes of the Red Jungle Fowl and local Village Chicken were further amplified with GM01-50 primers. Analysis of the results based on band sharing (BS) and the molecular size of individually amplified DNA fragments showed that Red Jungle Fowl and local Village Chicken shared the species similarity of 66% with Operon primers 01-20, 64% between local Village Chicken and broiler, and 63% when DNA bands between Red Jungle Fowl and broiler were compared. With GM01-50, the BS between Red Jungle Fowl and local village chicken increased to 72%. The results showed that the local village chicken is more closely related to Red Jungle Fowl than to broiler in the genetic distance. On the other hand, broiler is 1% closer in genetic distance to local village chicken than to Red Jungle Fowl. The results also indicated that primers like OPA-7, 8 and 9 can be used as species specific DNA markers for these three species of chickens.