Viability of Sarcocystis neurona sporocysts after long-term storage

The effect of long-term storage on the viability and infectivity of Sarcocystis neurona sporocysts was investigated. S. neurona sporocysts were harvested from the small intestine of Virginia opossums from 1996 to 2002 and stored at 4 degrees C. Viability of sporocysts was assessed by propidium iodide (PI) exclusion assay, in vitro excystation and development in tissue cultures, and bioassay in gamma-interferon gene knockout (gamma-IFN-KO) mice. The rate of excystation was apparently unaffected by long-term storage; sporocysts retained their ability to excyst after 7 years of storage at 4 degrees C. However, the ability of sporocysts to exclude PI stain, to invade and proliferate in cells in vitro, and to cause disease and lesions in gamma-IFN-KO mice appeared to decline as sporocysts age. The results demonstrated that sporocysts of S. neurona were able to survive and maintain moderate to high viability for up to 7 years when stored in phosphate buffered saline and Hank's balanced salt solution containing antibiotic-antimycotic mixture at 4 degrees C.     

Prevalence of and risk factors associated with the presence of Sarcocystis neurona sporocysts in opossum (Didelphis virginiana) from Michigan: a retrospective study

From April 1996 to December 2002 the prevalence of Sarcocystis neurona sporocysts in North American opossum (Didelphis virginiana) in Southern Michigan was estimated. Sporocysts of S. neurona were found in intestinal scrapings from 31 (15%) of 206 examined opossum. The frequency of infection was higher in adult animals (26/206; 12.6%) and females (19/206; 9.2%) than in juveniles (5/206; 2.4%) and males (12/206; 5.8%). Also, prevalence of S. neurona sporocysts in opossums in relation to factors such as age, sex, season, body condition, presence of concomitant infection, and presence of young in the pouch of females was studied in detail over the course of the year, 2002. Univariate analyses identified the following factors as being associated with the presence of S. neurona sporocysts in opossums: (i) for age, adult (odd ratio [OR] = 2.074, P = 0.0005); (ii) for sex, female (OR = 7.016, P = 0.0119); (iii) for season, summer (OR = 7.917, P = 0.0032) and spring (OR = 4.071, P = 0.1063); (iv) for body condition, poor (OR = 3.50, P = 0.1200) and good (OR = 1.167, P = 0.8637); (v) for the presence of concomitant infection (OR = 23.056, P = 0001), and (vi) for the presence of young in the pouch of females (OR = 40.083, P = 0.0001). Multivariate logistic-regression analyses selected the following factors as being significantly associated with presence of S. neurona sporocysts in opossums: (i) for the presence of concomitant infection (OR = 8.722, P = 0.0160) and (ii) for the presence of young in the pouch of females (OR = 31.915, P = 0.0065). The prevalence of S. neurona sporocysts in D. virginiana suggests that this opossum may constitute an ample reservoir of infection to other animals in the northern United States.     

Molecular typing of Sarcocystis neurona: current status and future trends

Sarcocystis neurona is an important protozoal pathogen because it causes the serious neurological disease equine protozoal myeloencephalitis (EPM). The capacity of this organism to cause a wide spectrum of neurological signs in horses and the broad geographic distribution of observed cases in the Americas drive the need for sensitive, reliable and rapid typing methods to characterize strains. Various molecular methods have been developed and used to diagnose EPM due to S. neurona, to identify S. neurona isolates and to determine the heterogeneity and evolutionary relatedness within this species and related Sarcocystis spp. These methods included sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), immuno-fluorescent assay (IFA), slide agglutination test (SAT), SnSAG-specific ELISA, random amplified polymorphic DNA (RAPD), PCR-based restriction fragment length polymorphism (RFLP), amplified fragment length polymorphism (AFLP) fingerprinting, and sequence analysis of surface protein genes, ribosomal genes, microsatellite alleles and other molecular markers. Here, the utility of these molecular methods is reviewed and evaluated with respect to the need for molecular approaches that utilize well-characterized polymorphic, simple, independent, and stable genetic markers. These tools have the potential to add to knowledge of the genetic population structure of S. neurona and to provide new insights into the pathogenesis of EPM and S. neurona epidemiology. In particular, these methods provide new tools to address the hypothesis that particular genetic variants are associated with adverse clinical outcomes (severe pathotypes). The ultimate goal is to utilize them in future studies to improve treatment and prevention strategies.     

Comparison of nutritional and antinutritional factors in soybean and fababean seeds with or without cortex

The composition and contents of nutritional factors such as proteins, lipids, carbohydrates, fibers, amino acids, and antinutritional factors such as trypsin inhibitors, phytic acid, and tannins were compared in soybean and fababean seeds with emphasis placed on the nutritional improvement of the seeds by cortex removal. Protein hydrolysis analysis for both whole seeds and seed with cortex removed revealed the presence of a large amount of lysine, arginine, aspartic acid, glutamic acid, glycine, and leucine while these seeds contained a low level of tryptophan, cystine, and methionine. Some antinutritional factors such as trypsin inhibitors, phytic acid, and tannins were detected in soybean and fababean seeds: phytic acid content and trypsin inhibitor activity were higher in soybean seeds than in fababean seeds while the difference in the tannin content was less pronounced. It was found that most of the tannins occurred in the cortex of the soybean and fababean seeds. Tannins are polyphenolic compounds that readily form indigestible complexes with proteins and other macromolecules under specific environmental conditions. By removal of the cortex, tannins were almost completely eliminated without changes in the protein composition and amino acids. From these results, it is assumed that since soybean and fababean seeds contained a high concentration of antinutritional factors in the cortex such as tannins, the utilization of the legume seeds after removal of all of the cortex is suitable for human diet or industrial products.     

水稻不同种植方式的比较试验与评价

为了比较水稻不同种植方式的实施效果,在中国三大稻区安排了机械插秧、机械直播、机械钵苗行栽3种种植方式与常规育秧手工插秧方式比较试验,试验的品种为当地种植面积较大的6个品种,采集了不同试验方式的各项成本,测定了水稻产量指标和作业环节消耗工时等数据.试验结果表明,南方一季稻区优选种植方式的排序为:机械插秧、机械直播和手工插秧;南方双季稻区机械插秧方式优于手工插秧;北方稻区机械钵苗行栽方式优于手工插秧.其中机械插秧方式平均增产7.53%、平均省工41.44%,平均增加效益46.78%;机械直播方式产量平均下降6.34%,平均省工31.30%,而对效益的影响因品种不同而表现出差异性;机械钵苗行栽方式平均增产8.95%、省工64.29%,增加效益22.57%.     

Effect of daily administration of pyrantel tartrate in preventing infection in horses experimentally challenged with Sarcocystis neurona

OBJECTIVE: To determine whether daily administration of pyrantel tartrate can prevent infection in horses experimentally challenged with Sarcocystis neurona. ANIMALS: 24 mixed-breed specific-pathogen-free weanling horses, 10 adult horses, 1 opossum, and 6 mice. PROCEDURE: Sarcocystis neurona-na?ve weanling horses were randomly allocated to 2 groups. Group A received pyrantel tartrate at the labeled dose, and group B received a nonmedicated pellet. Both groups were orally inoculated with 100 sporocysts/d for 28 days, 500 sporocysts/d for 28 days, and 1000 sporocysts/d for 56 days. Blood samples were collected weekly, and CSF was collected monthly. Ten seronegative adult horses were monitored as untreated, uninfected control animals. All serum and CSF samples were tested by use of western blot tests to detect antibodies against S. neurona. At the end of the study, the number of seropositive and CSF-positive horses in groups A and B were compared by use of the Fisher exact test. Time to seroconversion on the basis of treatment groups and sex of horses was compared in 2 univariable Cox proportional hazards models. RESULTS: After 134 days of sporocyst inoculation, no significant differences were found between groups A and B for results of western blot tests of serum or CSF There were no significant differences in number of days to seroconversion on the basis of treatment groups or sex of horses. The control horses remained seronegative. CONCLUSIONS AND CLINICAL RELEVANCE: Daily administration of pyrantel tartrate at the current labeled dose does not prevent S. neurona infection in horses.     

Use of next‐generation sequencing for the identification and characterization of Maize chlorotic mottle virus and Sugarcane mosaic virus causing maize lethal necrosis in Kenya

The diagnosis of novel unidentified viral plant diseases can be problematic, as the conventional methods such as real‐time PCR or ELISA may be too specific to a particular species or even strain of a virus, whilst alternatives such as electron microscopy (EM) or sap inoculation of indicator species do not usually give species level diagnosis. Next‐generation sequencing (NGS) offers an alternative solution where sequence is generated in a non‐specific fashion and identification is based on similarity searching against GenBank. The conventional and NGS techniques were applied to a damaging and apparently new disease of maize, which was first identified in Kenya in 2011. ELISA and TEM provided negative results, whilst inoculation of other cereal species identified the presence of an unidentified sap transmissible virus. RNA was purified from material showing symptoms and sequenced using a Roche 454 GS‐FLX+. Database searching of the resulting sequence identified the presence of Maize chlorotic mottle virus and Sugarcane mosaic virus, a combination previously reported to cause maize lethal necrosis disease. Over 90% of both viral genome sequences were obtained, allowing strain characterization and the development of specific real‐time PCR assays which were used to confirm the presence of the virus in material with symptoms from six different fields in two different regions of Kenya. The availability of these assays should aid the assessment of the disease and may be used for routine diagnosis. The work shows that next‐generation sequencing is a valuable investigational technique for rapidly identifying potential disease‐causing agents such as viruses.     

Providing an intelligible explanation to pet owners by using three-dimensional CT images: Use of clinical imaging for better informed consent

The purpose of the study reported here was to discuss the level of intelligibility of three-dimensional computed tomography (3D-CT) images of clinical patients for persons who are lacking veterinary knowledge. 143 undergraduate students were participated in this study. They were entirely unfamiliar with veterinary anatomy. A survey was conducted using the visual analog scale method with a focus on the level of intelligibility of the 3D-CT images compared to that of the two-dimensional CT images or radiographs. As a result, the mean value of the intelligibility of the 3D-CT image was 88.4 ± 17.8 points out of 100 points. In clinical cases, it is very difficult for pet owners lacking veterinary knowledge to understand the clinical status of their pets even with explanations using illustrations, radiographs, and transverse CT images. It is considered that better informed consent and patient satisfaction will be obtained provided veterinarians make an effort towards intelligible patient education by using 3D-CT images.