Comparison of three enzyme-linked immunosorbant assays with the indirect immunofluorescent antibody test for the diagnosis of canine infection with Ehrlichia canis

The aim of this study was to compare three different enzyme-linked immunosorbant assays (recombinant major antigenic protein 2 (rMAP2)-ELISA, the Immunocomb (Biogal, Israel) and the Snap 3Dx assay (IDEXX Laboratories Inc., USA)) with the indirect immunofluorescent antibody test in detecting anti-Ehrlichia canis immunoglobulin-G (IgG) antibodies. Samples tested were collected from dogs suspected to be naturally infected with E. canis and from experimentally infected dogs.When qualitative results (positive/negative) were compared, there was an overall agreement of 81% (54/67) between the indirect immunofluorescence antibody (IFA) test and the rMAP2-ELISA. An overall agreement of 94% (63/67) was found between the IFA test and the Immunocomb, and an overall agreement of 91% (61/67) was found between the IFA test and the Snap 3Dx assay. In 50 of 67 (74.6%) samples tested, complete agreement in the qualitative results was found in all four tests. Sixteen of 17 samples with disagreement in the qualitative results were found to have IFA titers of 1:320 or less. The sensitivities and specificities of the tests were found to be 0.71 and 0.85 for the rMAP2-ELISA, 0.86 and 0.98 for the Immunocomb, and 0.71 and 1.00 for the Snap 3Dx assay.The tests performed in this study were found to be highly specific in detecting E. canis antibodies. Their sensitivity was found to be low with sera having IFA titers of < or =1:320, while high with sera having titers greater than 1:320. Repeating the serological tests 1-2 weeks after the first antibody assay may overcome the sensitivity problem with titers of < or =1:320.     

Novel and simple approach using synthesized nickel nanoparticles to control blood-sucking parasites

The present study was on assessment of the anti-parasitic activities of nickel nanoparticles (Ni NPs) against the larvae of cattle ticks Rhipicephalus (Boophilus) microplus and Hyalomma anatolicum (a.) anatolicum (Acari: Ixodidae), fourth instar larvae of Anopheles subpictus, Culex quinquefasciatus and Culex gelidus (Diptera: Culicidae). The metallic Ni NPs were synthesized by polyol process from Ni-hydrazine as precursor and Tween 80 as both the medium and the stabilizing reagent. The synthesized Ni NPs were characterized by Fourier transform infrared (FTIR) spectroscopy analysis which indicated the presence of Ni NPs. Synthesized Ni NPs showed the X-ray diffraction (XRD) peaks at 42.76°, 53.40°, and 76.44°, identified as 1 1 1, 2 2 0, and 2 0 0 reflections, respectively. Scanning electron microscopy (SEM) analysis of the synthesized Ni NPs clearly showed that the Ni NPs were spherical in shape with an average size of 150 nm. The Ni NPs showed maximum activity against the larvae of R. (B.) microplus, H. a. anatolicum, A. subpictus, C. quinquefasciatus and C. gelidus with LC₅₀ values of 10.17, 10.81, 4.93, 5.56 and 4.94 mg/L; r² values of 0.990, 0.993, 0.992, 0.950 and 0.988 and the efficacy of Ni-hydrazine complexes showed the LC₅₀ values of 20.35, 22.72, 8.29, 9.69 and 7.83 mg/L; r² values of 0.988, 0.986, 0.989, 0.944 and 0.978, respectively. The findings revealed that synthesized Ni NPs possess excellent larvicidal parasitic activity. To the best of our knowledge, this is the first report on larvicidal activity of blood feeding parasites using synthesized Ni NPs.     

Study of genetic diversity in Indian and exotic sesame (Sesamum indicum L.) germplasm using random amplified polymorphic DNA (RAPD) markers

Fifty-eight accessions of sesame (Sesamum indicum L.), an important oil seed crop of the tropics and subtropics were analysed using random amplified polymorphic DNA (RAPD) technique. The material analysed comprised 36 collections from 18 different states of India and four adjoining countries of the Indian subcontinent, and 22 exotic accessions from 21 sesame growing countries around the world. The results from PCR amplifications with the selected 24 random 10-mer primers were statistically analysed. The value of Jaccard’s similarity coefficients ranged from 0.19 to 0.89. The results indicated the presence of high level of genetic diversity. However, the extent of genetic diversity was greater in the collections from Indian subcontinent as compared to the exotics. Among the Indian accessions, the collections from Rajasthan and North-eastern states were highly diverse. The phenetic analysis grouped 48 out of 58 accessions in six clusters and the remaining highly diverse accessions were placed outside these close-knit clusters. The Bootstrap estimates obtained by Wagner parsimony analysis were significant for seven out of 49 nodes in the majority-rule consensus tree (<95% occurrence). The results of both the analyses were, however, broadly comparable when the constitution of the individual clusters were considered. The principal components analysis indicated that the first two components accounted for only 21% of the total variations and in order to explain <75% of variations 18 components were required. The high level of genetic diversity prevalent among the Indian collections is probably indicative of the nativity of this crop species. Similarly, the relatively lower level of polymorphism in exotic germplasm could be ascribed to the comparatively recent introductions of limited germplasm of this crop into some of the non-traditional sesame growing countries. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effect of Nitrogen and Sulphur Application on Yield and Fatty acid Composition of Mustard (Brassica juncea L.) Oil

The effects of nitrogen and sulphur on the yield and fatty acid composition of mustard ( Brassica juncea L.) oil were studied in a field experiment. Significantly higher grain and oil yields were obtained with N and S application. Applications of nitrogen up to 60 kg ha⁻¹ and sulphur up to 40 kg ha⁻¹ favourably influenced the grain yield. Increasing levels of N decreased the oil content while application of sulphur improved the oil content. The contents of linoleic and linolenic acid were maximum (16.82 and 8.73%, respectively) with 60 kg N along with 40 kg S ha⁻¹. No use of fertilizers led to higher contents of undesirable fatty acids such as palmitic (hypercholesterimic) and erucic (do not have food value) acids.     

Organ culture of chicken bursa as a model to study the pathogenicity of infectious bursal disease virus isolates.

In vitro study with chicken bursal organ culture was attempted to assess the pathogenicity of locally isolated infectious bursal disease virus (IBDV) initially isolated from the bursa of naturally infected birds. In bursal organ culture, lymphoblastic transformation was noticed as early as 24 hr postinoculation and reached maximum at 72 hr postinoculation. The other microscopic changes were increased number of macrophages and formation of plasma cells. The IBDV antigen was detected 24 hr onward by coagglutination test with antibody coated Staphylococcus aureus strain Cowan I. On the basis of lesion score, the three isolates of IBDV (A, B, and C) were graded as virulent (B isolate) and moderately virulent (A and C isolates). A similar pattern of pathogenicity was also observed in the in vivo pathogenicity studies in chicken based on bursa: body weight ratio and histopathologic lesion score. The bursal organ culture thus provides a useful experimental model to differentiate the IBDV isolates on the basis of their virulence.     

Changes in colostrum of Murrah buffaloes after calving

Colostrum samples were collected from 8 Murrah buffaloes on days 1, 2, 3, 4 and 5 after calving. Levels of IgG averaged 54.0 mg/ml at calving, then decreased significantly (P < 0.01). IgA and IgM on day 1 were 3.22 mg/ml and 5.22 mg/ml, respectively; both decreased during the first five days after calving. Values of IgA and IgM were higher than those reported in cows. SCC values, which were high at calving (500 000 per ml), reduced significantly (P < 0.01) on day 2, then decreased slightly until day 5 (180 000 per ml). At calving, macrophages were the most prominent cells in buffalo colostrum, followed by lymphocytes and neutrophils. Phagocytic activity was 23% at calving and reduced significantly (P < 0.01) to 14% on day 5. Phagocytic index was highest in the first colostrum, and then decreased non-significantly.     

Effect of fasting duration on clinical pathology results in Wistar rats

Background: Fasting is an important preanalytical factor that may affect the interpretation of hematology and clinical biochemistry data in toxicology or pharmacology studies. Limited information is available on how the results may be affected by different durations of fasting. Objective: The purpose of this study was to assess the influence of fasting duration on clinical pathology results in male and female rats and to determine an optimum fasting time for preclinical studies. Methods: Male and female Wistar rats (10 each per group) were fasted for 0, 4, 8, 16, 24, and 48 hours. Changes in body weight and in the results of routine CBC and clinical chemistry analysis were evaluated by 1‐way ANOVA. Results: Body weight was significantly decreased by 4 hours of fasting in all rats, and hemoglobin concentration was significantly increased at 16 hours in male rats. Serum glucose and triglyceride concentrations in both sexes and cholesterol and high‐density lipoprotein‐C concentrations in female rats were also significantly decreased beginning at 16 hours. The creatinine concentration was increased in females after 16 hours of fasting. Serum alkaline phosphatase and alanine aminotransferase activities were significantly decreased after 8 hours in males and 16 hours in females. Conclusions: Fasting‐induced changes in clinical pathology results were consistent with hemoconcentration and altered nutrition and metabolic function. Most changes occurred at 16 hours, with minimal subsequent changes. Hence, a 16‐hour fasting duration may be recommended for preclinical studies involving clinical pathology measurements.     

Identification of a 66 kDa Haemonchus contortus excretory/secretory antigen that inhibits host monocytes

A 66 kDa adult Haemonchus contortus excretory/secretory (E/S) antigen was identified in Western blot by reaction with sera from the infected goats. The protein was purified from the adult worm extract and E/S products by anion exchange and ConA-Sepharose chromatography. The purified protein inhibited monocyte function in vitro as judged by decreased production of hydrogen peroxide and nitric oxide in the culture medium. The protein also caused proliferation of peripheral blood mononuclear cells. The absence of protein in the free living L3 larvae suggests that the expression of this protein coincides with the adaptation to the parasitic life. A correlation of antibody titre and worm burden was observed in the infected goats with higher antibody levels in high worm burdened animals. Anti-protein antibody caused loss of adult worm motility in vitro resulting in the death of the parasite. The fact that the protein is recognized by the host together with in vitro killing of adult parasites by antibodies makes this protein a promising candidate for vaccination trial.