Development and evaluation of loop-mediated isothermal amplification assay for rapid detection of Capripoxvirus

Research (Published online: 05-11-2015)

3. Development and evaluation of loop-mediated isothermal amplification assay for rapid detection of Capripoxvirus - Kanisht Batra, Aman Kumar, Vinay Kumar, Trilok Nanda, Narender S. Maan and Sushila Maan

Veterinary World, 8(11): 1286-1292

   doi: 10.14202/vetworld.2015.1286-1292

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Abstract

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Aim: The present study was undertaken to develop a nucleic acid-based diagnostic assay loop-mediated isothermal amplification assay (LAMP) targeting highly conserved genomic regions of Capripoxvirus (CaPVs) and its comparative evaluation with real-time polymerase chain reaction (PCR).

Material and Methods: Lyophilized vaccine strain of sheeppox virus (SPPV) was used for optimization of LAMP assay. The LAMP assay was designed using envelope immunogenic protein (P32) coding gene targeting highly conserved genomic regions of CaPV responsible for causing sheep pox, goat pox, and lumpy skin disease in sheep, goat and cattle respectively. Serial tenfold dilution of SPPV recombinant plasmid DNA was used for a calculating limit of detection. Analytical sensitivity and specificity were performed.

Results: The test described is quick (30 min), sensitive and specific for detection of CaPVs. The described assay did not show any cross-reactivity to other related viruses that cause apparently similar clinical signs. It was found to be ten times more sensitive than conventional PCR however, 100 times less sensitive than quantitative PCR (qPCR). LAMP assay results were monitored by color change method using picogreen dye and agarose gel electrophoresis.

Conclusion: LAMP assay can be a very good alternative for CaPV detection to other molecular techniques requiring sophisticated equipments.

Keywords: Capripoxvirus, loop-mediated isothermal amplification assay, real-time polymerase chain reaction, sensitivity, specificity.

Cross antigenicity of immunodominant polypeptides of somatic antigen of Oesophagostomum columbianum with other helminths by western blotting

Research (Published online: 05-11-2015)

2. Cross antigenicity of immunodominant polypeptides of somatic antigen of Oesophagostomum columbianum with other helminths by western blotting - Sunita Dalal, Arvind Prasad, Abdul Nasir and Vijesh Kumar Saini

Veterinary World, 8(11): 1279-1285

   doi: 10.14202/vetworld.2015.1279-1285

Abstract

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Aim: Oesophagostomum columbianum in small ruminants in India is found as mixed infection commonly in sheep and goat. Haemonchus contortus, an abomasal nematode is found as concurrent infection with it. Eggs of Haemonchus and O. columbianum cannot be easily distinguished. Diagnosis of O. columbianum may only be possible if a non-cross antigenic polypeptide was available for immunodiagnosis.

Materials and Methods: Somatic antigen (SoAg) of O. columbianum was fractionated by sodium dodecyl sulfatepolyacrylamide gel electrophoresis and immunodominant polypeptides were identified by western blotting with homologous hyperimmune serum (HIS) and experimental sera of sheep or goat infected with other helminths.

Results: SoAg of O. columbianum was immunoaffinity purified. Sharp polypeptide bands of 130, 72 and 68 KDa were observed along with several faint bands of lower molecular weight. Western blot of purified SoAg of O. columbianum with homologous HIS showed reaction with all the protein bands of 17, 28, 30, 32, 35, 38, 50, 68, 100, 130, 150, and 170 kDa. For identification of non-cross antigenic polypeptide, immunoaffinity purified SoAg of O. columbianum was reacted to heterologous HIS against H. contortus, Paramphistomum epiclitum, and Fasciola gigantica in western blotting utilizing completely dry method (i-blot). Among high molecular weight polypeptides 100 and 150 kDa were non-cross antigenic and among low molecular weight except 50 kDa polypeptide, 17, 30, 32, 35, and 38 kDa of O. columbianum were not cross antigenic with other helminths.

Conclusions: Hence, polypeptides of 17, 30, 32, 35 and 38 kDa as well as 100 and 150 kDa polypeptides of O. columbianum may be exploited for immunodiagnosis of the infection in sheep and goat with extensive studies on cross antigenicity.

Keywords: cross antigenicity, hyperimmune sera, immunodiagnostic polypeptides, Oesophagostomum columbianum. 

Ileal and jejunal Peyer’s patches in buffalo calves: Histomorphological comparison

Research (Published online: 05-11-2015)

1. Ileal and jejunal Peyer’s patches in buffalo calves: Histomorphological comparison - Kritima Kapoor and Opinder Singh

Veterinary World, 8(11): 1273-1278

   doi: 10.14202/vetworld.2015.1273-1278

Abstract

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Aim: The present study was aimed to elucidate the histomorphology of ileal and jejunal Peyer’s patches in the small intestine of buffalo calves and their structural comparison.
Materials and Methods: The study was conducted on neonatal (n=10) and pre-pubertal (n=10) buffalo calves. The age of the postnatal buffalo calves was estimated by their temporary and permanent dentition.
Results: The study revealed that several layers of oval to elongate elliptical lymphoid follicles were observed in submucosa on the anti-mesenteric side in the ileum of early neonatal calves. However, the follicles at this age, in jejunum were of all shapes present within one layer. The interfollicular space was occupied by the interfollicular tissue, which was diffuse and wider around jejunal lymphoid follicles as compared to ileal lymphoid follicles. However, toward the pubertal stage, the number of layers of lymphoid follicles was reduced in ileum due to involution while it remained similar in number in jejunum at this stage.
Conclusion: The ileal Peyer’s patches were found to have started involution more or less around reaching puberty, whereas the jejunal Peyer’s patches appear to be functional throughout the lifespan of the animal.
Keywords: histomorphology, ileum, jejunum, lymphoid follicle, Peyer’s patch.

Assessment of semen quality in pure and crossbred Jersey bulls

Research (Published online: 31-10-2015)

18. Assessment of semen quality in pure and crossbred Jersey bulls - Umesh Kumar, Ajay P. Gawande, Sunil K. Sahatpure, Manoj S. Patil, Chetan K. Lakde, Sachin W. Bonde, Pradnyankur L. Borkar, Ajay J. Poharkar and Baldeo R. Ramteke

Veterinary World, 8(10): 1266-1272

   doi: 10.14202/vetworld.2015.1266-1272

Abstract

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Aim: To compare the seminal attributes of neat, pre-freeze (at equilibration), and post-freeze (24 h after freezing) semen in pure and crossbred Jersey bulls.

Materials and Methods: Total 36 ejaculates (3 ejaculates from each bull) were collected from 6 pure Jersey and 6 crossbred Jersey bulls and evaluated for various seminal attributes during neat, pre-freeze, and post-freeze semen.

Results: The mean (±standard error [SE]) values of neat semen characteristics in pure and crossbred Jersey bulls were recorded such as volume (ml), color, consistency, mass activity (scale: 0-5), and sperm concentration (millions/ml). The extended semen was further investigated at pre-freeze and post-freeze stages and the mean (±SE) values recorded at neat, pre-freeze, and post-freeze semen were compared between pure and crossbred Jersey bulls; sperm motility (80.55±1.70%, 62.77±1.35%, 46.11±1.43% vs. 80.00±1.80%, 65.00±1.66%, 47.22±1.08%), live sperm count (83.63±1.08%, 71.72±1.09%, 58.67±1.02% vs. 80.00±1.08%, 67.91±1.20%, 51.63±0.97%), total abnormal sperm count (8.38±0.32%, 12.30±0.39%, 16.75±0.42% vs. 9.00±0.45%, 12.19±0.48%, 18.11±0.64%), hypo-osmotic swelling (HOS) reacted spermatozoa (71.88±0.77%, 62.05±0.80%, 47.27±1.05% vs. 72.77±1.02%, 62.11±0.89%, 45.94±1.33%), acrosome integrity (89.05±0.83%, 81.33±0.71%, 71.94±0.86% vs. 86.55±0.57%, 78.66±0.42%, 69.38±0.53%), and DNA integrity (99.88±0.07%, 100, 99.66±0.11% vs. 99.94±0.05%, 100, 99.44±0.18%,). The volume, color, consistency, sperm concentration, and initial motility in pure and crossbred Jersey bulls did not differ significantly (p>0.05). The mass activity was significantly (p<0.05) higher in pure Jersey as compare to crossbred Jersey bulls. Live sperm percentage and acrosome integrity was significantly (p<0.01) higher in pure Jersey bulls as compared to crossbred Jersey bulls. However, no statistical difference (p>0.05) was observed in abnormal sperm; HOS reacted spermatozoa and DNA integrity percentage among breeds.

Conclusion: It may be concluded that the quality of pure Jersey bull semen was comparatively better than the crossbred Jersey bulls.

Keywords: acridine orange, acrosomal integrity, crossbred Jersey, DNA integrity, Jersey, semen.

Evaluation of Spirulina platensis extract as natural antivirus against foot and mouth disease virus strains (A, O, SAT2)

Research (Published online: 31-10-2015)

17. Evaluation of Spirulina platensis extract as natural antivirus against foot and mouth disease virus strains (A, O, SAT2) - Hind M. Daoud and Eman M. Soliman

Veterinary World, 8(10): 1260-1265

   doi: 10.14202/vetworld.2015.1260-1265

Abstract

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Aim: This work was aimed to document the antiviral activates of Spirulina platensis extract against foot and mouth disease virus (FMDV) different types to evaluate its replication in Baby Hamster Kidney (BHK) cell culture and in baby mice.

Materials and Methods: Cytotoxicity assay studied for S. platensis extract on BHK cells to determine the non-toxic dose. The non-toxic dose of Spirulina extract was mixed with each type of FMDV (A, O, SAT2). Then 10-fold dilutions from each mixture were done. FMDV titer for each type of treated FMDV was calculated to evaluate the antiviral activity of the Spirulina extract against FMDV. Furthermore, old baby Swiss mice were inoculated with 0.1 ml intraperitonially from the mixture of FMDV different types and different concentration of Spirulina extracts. After 48 h post inoculation, all the baby mice examined to evaluate the antiviral action of Spirulina extract.

Results: The result showed that the non-toxic doses of S. platensis (50 ug/ml) revealed 35.7%, 28.5%, and 31% reductions in FMDV titers Type O, A, and SAT2 on BHK cells, respectively. The same non-toxic dose gave 50% of the inhibitory concentration in baby mice without cytotoxic effect.

Conclusion: This study confirmed the biological activity of the ethanol extract of S. platensis against FMDV Types O, A, and SAT2. From the results, S. platensis could be useful as antiviral lead to limitation of infection among animals during outbreaks but further studies need to evaluate the S. platensis on experimental or natural infected farm animals to establish the effective dose side affected period of treatment of S. platensis.

Keywords: antiviral, foot and mouth disease virus, Spirulina platensis.

Application of hyaluronic acid in the healing of non-experimental open wounds: A pilot study on 12 wounds in 10 client-owned dogs

Research (Published online: 28-10-2015)

16. Application of hyaluronic acid in the healing of non-experimental open wounds: A pilot study on 12 wounds in 10 client-owned dogs -Roberta Ferrari, Patrizia Boracchi, Stefano Romussi, Giuliano Ravasio and Damiano Stefanello

Veterinary World, 8(10): 1247-1259

   doi: 10.14202/vetworld.2015.1247-1259

Abstract

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Aim: Veterinarians have frequently to deal with wounds to the skin, subcutis, and underlying muscle. The aim was to explore the application of hyaluronic acid (HA)-containing dressing on open skin wounds in dogs. The progress of healing was assessed by wound area reduction and two scoring scales applied in human medicine.

Materials and Methods: Ten client-owned dogs with 12 cutaneous open wounds healed by the second intention were included. All wounds were treated using available in commerce HA-containing wound dressing from admission to complete re-epithelialization. At every clinical examination, wound area and scale scoring assessments were performed.

Results: After debridement, an increased wound size was obtained while an improvement was determined by both grading systems. The median numbers of return to the clinic for bandage change were 5 times. The median time to complete wound healing was 34.5 days. The mean wound area at day 7, 14, 21, and 28 were, respectively, 90.4%, 47.7%, 22.4%, and 14.8% of the original size (for linear measurement) and 95.5%, 54.4%, 23.10%, and 14.8% of the original size (for software measurement). Regarding wound healing assessment tools, the agreement between two operators was considered high for both scales.

Conclusions: HA-containing dressing may be a possible wound treatment for cutaneous open wounds in dogs. The assessment of wound quality using scale scoring system could be useful especially in the 1st week and to direct clinical decision-making process.

Keywords: advance wound dressing, canine, wound assessment.

Conventional and fluorescent based semen quality assessment in Karan Fries bulls

Research (Published online: 28-10-2015)

15. Conventional and fluorescent based semen quality assessment in Karan Fries bulls - A. Panmei, A. K. Gupta, P. R. Shivahre, M. Bhakat and A. Upadhyay

Veterinary World, 8(10): 1243-1246

   doi: 10.14202/vetworld.2015.1243-1246

Abstract

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Aim: The present study was carried out on semen ejaculates of 15 Karan Fries (KF) bulls maintained at Artificial Breeding Research Centre, National Dairy Research Institute, Karnal, India with an objective to evaluate the relationship between the conventional and fluorescent based semen quality analysis of the bulls.

Materials and Methods: A total of 96 ejaculates were collected from 15 KF (Holstein Friesian [HF] crossbred) bulls. Semen were evaluated for color, volume, mass activity (MA) and percentage of individual motility (IM), sperm concentration, percent live spermatozoa, hypo-osmotic swelling test and acrosome integrity, chromatin integrity, sperm viability, and membrane integrity. Data were analyzed using SPSS software package for descriptive analysis. The correlation between rankings of sires based on conventional and fluorescent semen parameters were calculated by Spearman’s rank correlation coefficient.

Results: The average ejaculates volume (ml), sperm concentration (106/ml), MA, IM (%), live (%), morphological abnormalities (%), host (%), acrosome integrity (%), chromomycin A3 (CMA3) (%), SYBR-PI (%), and fluorescent isothiocyanate-peanut agglutinin (FITC-PNA) (%) were 4.57±0.36, 1162.98±97.93, 2.95±0.09, 60.8±1.22, 71.41±2.10, 9.31±1.15, 65.5±1.81, 86.6±1.59, 3.53±0.43, 65.39±2.23 and 74.47±2.53, respectively. Rank correlations were found to be significant for SYBR-PI and FITC-PNA with most of the parameters evaluated by conventional methods. Overall, among conventional criteria, IM revealed ranking of bulls almost similar to that of fluorescent criteria.

Conclusion: Overview of our results indicated that, among conventional criteria, MA and IM revealed ranking of bulls almost similar to that of fluorescent criteria.

Keywords: conventional method, fluorescent technique, Karan Fries, semen quality.