Development of a genetic manipulation system for Haemophilus parasuis

Haemophilus parasuis is a member of the family Pasteurellaceae and an important respiratory-tract pathogen of swine, which is the etiological agent of Glasser's disease. Because no genetic manipulation system is available for H. parasuis so far, in vivo studies about the role of its genes involved in virulence are unfeasible. Here we demonstrate that H. parasuis has a cyclic AMP (cAMP)-dependent natural transformation system that enables the uptake of DNA in which the ACCGAACTC sequence signal must be present. After improving DNA transformation parameters, such as cAMP and DNA concentration and exposition time of the exogenous DNA, a knockout mutant of H. parasuis defective in the thy gene, encoding the thymidylate synthase enzyme, has been constructed. Data presented in this work open the possibility for the functional analysis of genes involved in the infectious process of this animal pathogen.     

The prevalence of Toxocara cati in domestic cats in Mexico City

The faeces of 520 domestic cats, resident in the 16 municipal authorities of Mexico City, were analysed using the method of centrifuging with zinc sulphate (at 33%). Three hundred and ninety-nine animals lived in houses, 121 in apartments. For the purpose of this study they were divided into seven age groups, of 6 months each. Toxocara cati eggs were found in the faeces of 42.5% of the animals. 20.7% of apartment cats and 49.1% of house cats were infected. This difference in the rate of infection is statistically significant (P<0.002.) for the house cats. T. cati was found in all the age groups although most infection was found in animals less than 1 year old.The fact that infection occurred in the whole age range of the cats and in all the municipal authorities means that the risk of being contaminated with this helminth in Mexico City is high. As a consequence, the possibility of developing illness as a result of becoming a host to larva migrans is also high.     

Genetic Structure of Field Populations of Begomoviruses and of Their Vector Bemisia tabaci in Pakistan

ABSTRACT The genetic structure of field populations of begomoviruses and their whitefly vector Bemisia tabaci in Pakistan was analyzed. Begomoviruses and B. tabaci populations were sampled from different crops and weeds in different locations in Punjab and Sindh provinces, in areas where cotton leaf curl disease (CLCuD) occurs or does not occur. Phylogenetic analysis based on nucleotide sequences of the intergenic region in the viral DNA-A provided evidence of two clusters of isolates: viruses isolated from species in the family Malvaceae, and viruses isolated from other dicotyledon families. Analysis of the capsid protein (CP) open reading frame grouped isolates into three geographical clusters, corresponding to isolates collected in Punjab, Sindh, or both provinces. Random amplified polymorphic DNA analyses of the B. tabaci population showed that intrapopulation diversity was high at both the local and regional scales. Sequence analysis of the mitocondrial cytochrome oxydase I (mt COI) gene showed that the B. tabaci population was structured into at least three genetic lineages corresponding to the previously described Indian, Southeast Asian, and Mediterranean-African clades. The Indian clade was present only in Punjab, the Mediterranean-African only in Sindh, and the Southeast Asian in both provinces. B. tabaci haplotypes of the Indian clade were found only in the Punjab, where CLCuD occurs. Hence, the geographical distribution of virus and vector genotypes may be correlated, because similar phylogenetic relationships were detected for the viral CP and the vector mt COI genes.     

Histological and sex steroid hormone receptor changes in testes of immature, mature, and aged chickens

Sex steroid hormone receptors play a central role in the regulation of reproduction in male chickens. In this work, we evaluated by histomorphometric methods and Western blot analysis changes in the number of the different cell populations and in the content of sex steroid hormone receptors in testes from immature (1.5-month-old), mature (12-month-old), and aged (48-month-old) chickens. The number of Sertoli cells, germ cells, and Leydig cells per area of testicular tissue markedly changed according to chicken age. The highest number of Sertoli and Leydig cells was found in testes of immature chickens, with a dramatic decrease in those of mature chickens; however, the number of germ cells was the highest in mature chickens in comparison with other ages. The content of androgen receptor diminished in testes of mature and aged animals in comparison with that of immature chickens. In contrast, the content of estrogen receptor alpha and progesterone receptor was higher in testes of mature animals than in other ages. Both progesterone receptor isoforms were expressed in a similar proportion in testes of immature and mature animals. Interestingly, progesterone receptor isoform A was the predominant isoform in aged animals. These results suggest that there are marked age-dependent changes in chicken testes histology and in sex steroid hormone receptors content that should contribute to sex steroid hormone actions, in this tissue throughout the lifespan of chickens.     

Vineyard yield estimation by combining remote sensing,computer vision and artificial neural network techniques

In viticulture, it is critical to predict productivity levels of the different vineyard zones to undertake appropriate cropping practices. To overcome this challenge, the final yield was predicted by combining vegetation indices (VIs) to sense the health status of the crop and by computer vision to obtain the vegetated fraction cover (F_c) as a measure of plant vigour. Multispectral imagery obtained from an unmanned aerial vehicle (UAV) is used to obtain VIs and F_c, which are used together with artificial neural networks (ANN) to model the relationship between VIs, F_c and yield. The proposed methodology was applied in a vineyard, where different irrigation and fertilisation doses were applied. The results showed that using computer vision techniques to differentiate between canopy and soil is necessary in precision viticulture to obtain accurate results. In addition, the combination of VIs (reflectance approach) and F_c (geometric approach) to predict vineyard yield results in higher accuracy (root mean square error (RMSE)?=?0.9 kg vine^?1 and relative error (RE)?=?21.8% for the image when close to harvest) compared to the simple use of VIs (RMSE?=?1.2 kg vine^?1 and RE?=?28.7%). The implementation of machine learning techniques resulted in much more accurate results than linear models (RMSE?=?0.5 kg vine^?1 and RE?=?12.1%). More precise yield predictions were obtained when images were taken close to the harvest date, although promising results were obtained at earlier stages. Given the perennial nature of grapevines and the multiple environmental and endogenous factors determining yield, seasonal calibration for yield prediction is required.

     

Osmoregulatory changes in wedge sole (Dicologoglossa cuneata Moreau, 1881) after acclimation to different environmental salinities

The osmoregulatory responses of 20 days of acclimation to environmental salinities of 5‰, 15‰, 25‰, 35‰ and 55‰ were assessed in juveniles of wedge sole ( Dicologoglossa cuneata Moreau, 1881). This sole shows a good capacity to adapt to this range of environmental salinities. A direct linear relationship between environmental salinity and plasma osmolality was observed, with a calculated isosmotic point of 10.4‰ (284 mOsm kg⁻¹). Na⁺, K⁺-ATPase activity in the gills followed a 'U-shaped' relationship with environmental salinity, and a direct linear relationship in kidney tissue. Plasma cortisol levels were elevated in fish held in extreme salinities, and glucose levels were higher only in the group maintained at the highest environmental salinity. In the liver, a decrease in glycogen, lactate and amino acid contents was observed in specimens acclimated to extreme salinities (5‰ and 55‰), suggesting mobilization of liver metabolites. Metabolite levels in white muscle showed a pattern similar to the liver, with lower values in specimens acclimated to extreme salinities. We conclude that wedge sole is strongly euryhaline, but acclimation to extreme salinities comes with an energetic cost.     

A highly accurate, single PCR reaction for parentage assignment in Senegal sole based on eight informative microsatellite loci

From a battery of microsatellite markers (100 loci), recently identified by our group, we have selected eight for parentage assignment in Senegal sole ( Solea senegalensis ). This tool is based on microsatellite loci obtained from four genomic DNA libraries and one cDNA library. Within the eight loci (six from anonymous genomic DNA sequences and two located in expressed sequence tags of known genes), we have found, in an analysis of a reproductive broodstock, between nine and 16 alleles. The expected heterozygosity was between 0.616 and 0.860. In addition, we have optimized the polymerase chain reaction (PCR) conditions to amplify all loci simultaneously in a single multiplex PCR reaction, and we have tested three lots of male and female (five to six individuals) and three offspring (50–60 larvae each). The use of the eight microsatellite loci, the possibility of amplifying them in a single PCR reaction and the high value of the exclusion probability (0.9992) make this multiplex PCR method a unique tool for parentage assignment.
Finally, analysing one meiotic gynogenetic progeny, we have determined the relative distance of six of these loci to the centromere, and we have also found that all of them are unlinked. All these characteristics confer this tool with a high accuracy for parentage studies and genetic population analyses of Senegal sole.     

Use of intraoperative impression smear cytology to guide successful treatment of a large renal cyst in a dog: a case report

A 13-yr-old Shih tzu was referred for surgical management of right-sided cranial abdominal mass, which corresponded to large, cavitated renal mass on ultrasonography, and was suspected to represent neoplasia. Intraoperative impression smear cytology (ISC) of the renal mass wall was consistent with benign renal cyst (RC), without evidence of neoplasia or infection. Deroofing and omentalisation were performed and histopathology was consistent with benign RC. Chronic kidney disease was diagnosed 4 mon postoperatively, however, the dog was asymptomatic, without cyst reoccurrence. Intraoperative ISC is an expedient and inexpensive diagnostic technique that can guide most appropriate treatment in dogs with large RCs.     

酶水解法制取大米蛋白的机理研究

(1郑州轻工业学院食品与生物工程学院,郑州 450002;2河南科技大学食品与生物工程学院,洛阳 471003; 3江南大学食品学院,无锡 214036)