Comparison of two methods of seminal plasma removal on buffalo (Bubalus bubalis) sperm cryopreservation

Cryopreservation causes damage to spermatozoa, and methods minimizing this damage are therefore needed. Although much discussed, seminal plasma removal has become an alternative to improve sperm quality and viability after freezing and has been applied to different species in attempt to obtain good results. The objective of this study was to evaluate semen quality in buffaloes submitted to two methods for seminal plasma removal (filtration and centrifugation). Semen samples were collected from seven Murrah buffalo bulls (Bubalus bubalis) once a week for 8 weeks. Each ejaculate was divided into three groups: control (presence of seminal plasma), centrifugation and filtration. Sperm kinetics was evaluated with the computer‐assisted sperm analysis (CASA) system. Plasmalemma and acrosomal membrane integrity, mitochondrial membrane potential and reactive oxygen species (ROS) were measured by flow cytometry, and lipid peroxidation was evaluated by the thiobarbituric acid reactive substances (TBARS) assay. Seminal plasma removal did not improve sperm kinetics compared to the control group. Centrifugation increased the number of cells with damaged acrosomal membranes (0.77 ± 0.05) and filtration caused greater plasmalemma and acrosomal membrane damage (22.18 ± 1.07). No difference in the mitochondrial membrane potential was observed between groups. In contrast, ROS production was higher in the centrifugation group compared to the control and filtration groups, although no differences in TBARS formation were detected. In conclusion, seminal plasma removal did not improve the quality of thawed buffalo semen compared to control in terms of sperm kinetics, membrane integrity, mitochondrial membrane potential or lipid peroxidation.     

Chemical,Physicochemical, and Nutritional Evaluation of Plantago (Plantago ovata Forsk)

Plantago ovata F. are small tan‐colored seeds with ≈30% weight husk. Plantago's husk high content of soluble fiber makes it a good lubricant of the intestinal track with demonstrated effects in lowering plasma cholesterol levels in humans and experimental animals. Plantago seeds grown in Northern Mexico were analyzed for proximate composition, combustion heat, soluble and insoluble dietary fiber, fatty acids, amino acids, and protein fractionation. In vitro digestibility and digestibility of dry matter, apparent and true digestibility, and net protein ratio (in vivo) were also analyzed. Plantago seeds had 17.4% protein, 6.7% fat, 24.6% total dietary fiber, 19.6% insoluble fiber, 5.0% soluble fiber, and a combustion heat of 4.75 kcal/g. Osborne fractionation (based on solubility) yielded albumin 35.8%, globulin 23.9%, and prolamin 11.7%. The oil from plantago seeds had a high percentage of linoleic acid (40.6%) and oleic acid (39.1%) and a minor proportion of linolenic acid (6.9%). In vitro protein digestibility of the plantago seed was 77.5%, suggesting a highly digestible protein. Lysine content was 6.82 g/100 g of protein, higher than wheat and oats (2.46 and 4.20 g/100 g of protein, respectively). Rat bioassays showed values of 89.6% digestibility of dry matter, 86.0% apparent digestibility, 88.1% true digestibility, and 4.40 net protein ratio corrected (NPRc). The importance of these findings is that plantago whole grain shows favorable nutritional quality when compared with cereals and legumes.     

A novel putative member of the family Benyviridae is associated with soilborne wheat mosaic disease in Brazil

Soilborne wheat mosaic disease (SBWMD), originally attributed to infections by Soilborne wheat mosaic virus (SBWMV) and Wheat spindle streak mosaic virus (WSSMV), is one of the most frequent virus diseases and causes economic losses in wheat in southern Brazil. This study aimed to characterize molecularly the viral species associated with wheat plants showing mosaic symptoms in Brazil. Wheat leaves and stems displaying mosaic symptoms were collected from different wheat cultivars in Passo Fundo municipality, Rio Grande do Sul State, southern Brazil. Double-stranded RNA was extracted and submitted to cDNA library synthesis and next-generation sequencing. No sequences of SBWMV and WSSMV were detected but the complete genome sequence of a putative new member of the family Benyviridae was determined, for which the name wheat stripe mosaic virus (WhSMV) is proposed. WhSMV has a bipartite genome with RNA 1 and RNA 2 organization similar to that of viruses belonging to Benyviridae. WhSMV RNA 1 has a single open reading frame (ORF) encoding a polyprotein with putative viral replicase function. WhSMV RNA 2 has six ORFs encoding the coat protein, the major protein (read-through), triple gene block movement proteins (TGB 1, 2 and 3) and ORF 6 (hypothetical protein). In addition to the genomic organization and nucleotide and amino acid sequence identities, phylogenetic analyses also corroborated that WhSMV is a virus species of the Benyviridae. However, isolates of WhSMV formed a clade distinct from members of the genus Benyvirus. It was also demonstrated that the plasmodiophorid Polymyxa graminis is associated with wheat roots showing SBWMD symptoms and infected by WhSMV.     

Evaluation of the resistance of Coffea canephora to Oligonychus ilicis (Acari: Tetranychidae) and the preimaginal conditioning effect on resistance using a biological life table

Oligonychus ilicis (McGregor) (Acari: Tetranychidae), is one of the most important mite pests found in coffee plants. One way to reduce intensive pest attacks is management via resistant plants. The purpose of this study was to evaluate the resistance of 14 C. canephora genotypes to O. ilicis by antixenosis and antibiosis mechanisms and to determine the preimaginal conditioning effect on resistant genotypes using O. ilicis reared on C. canephora. Resistance by antixenosis was not found; however, nine genotypes showed resistance to O. ilicis via an antibiosis mechanism. Two of these resistant genotypes showed a more persistent resistance than the O. ilicis reared on C. canephora, which indicated that a previous host can affect the selection of a genotype by O. ilicis. The study may provide a model to evaluate the preimaginal conditioning effect on plant resistance, which can be useful for plant breeders.     

Testis Morphometry and Stages of the Seminiferous Epithelium Cycle in an Epididymal Sperm‐storing Neotropical Vespertilionid,Myotis levis (Chiroptera)

Yellowish myotis, Myotis levis, is a seasonal, epididymal sperm‐storing Neotropical vespertilionid. In the dry season, males show simultaneous testis regression and sperm storage in cauda epididymis, enabling them to mate during this season. In this study, we investigated seasonal variations in body mass, diameter and height of seminiferous tubules and nuclei of Leydig cells in a population of southeastern Brazil. We also determined the frequencies of the stages of the seminiferous epithelium cycle (SEC) of mature individuals of this population. Body mass and diameter of Leydig cell nuclei showed no significant differences between dry and rainy seasons and stages of annual reproductive cycle; however, all other morphometric parameters varied significantly. The relative cumulative frequency of pre‐meiotic stages of the SEC (1–3) was 51%, of meiotic stage (4) was 2% and of post‐meiotic stages (5–8) was 47%. We confirmed that the yellowish myotis presents seasonal sperm production as revealed by testis regression and epididymal sperm storage during the dry season.     

Organic solvent-free reversed-phase ion-pairing liquid chromatography coupled to atomic fluorescence spectrometry for organoarsenic species determination in several matrices

A novel method has been developed to determine As-containing animal feed additives including roxarsone (ROX), p-arsanilic acid (p-ASA) and nitarsone (NIT), as well as other organic As species (dimethylarsonic acid (DMAA) and monomethylarsonic acid (MMAA)) by ion-pairing high-performance liquid chromatography coupled to hydride generation atomic fluorescence spectrometry (IP-HPLC-HG-AFS). A simple isocratic reversed-phase (RP) HPLC method with a mobile phase containing citric acid and sodium hexanesulfonate (pH 2.0) was developed using a C(18) column. The use of an organic solvent free mobile phase turns this methodology into an environmentally friendly alternative. Several ion pair forming agents, such as sodium hexanesulfonate, tetrabutylammonium bisulfate and perfluoroheptanoic acid, were studied. The limits of detection for As species were calculated in standard solution and resulted to be 0.2, 0.5, 0.6, 1.6, and 1.6 μg As L(-1) for MMAA, DMAA, p-ASA, ROX and NIT, respectively. This method exhibited convenient operation, high sensitivity and good repeatability. It was applied to As speciation in different samples including arugula, dog food, dog urine and chicken liver.     

Predominance of Burkholderia cenocepacia lineages causing onion sour skin in the semi-arid region of north-east Brazil

Burkholderia cepacia was originally described as the causal agent of onion sour skin. However, this bacterium is now recognized as a complex of 22 closely related species, commonly referred to as the B. cepacia complex (BCC). Only a few taxonomic studies have been undertaken with the aim of understanding the diversity of species associated with onion sour skin. In this study, we used a polyphasic analysis including rep-PCR assay, biochemical and pathological profiles, and multilocus sequence analysis (MLSA) to characterize the BCC species associated with onion sour skin in the semi-arid region of north-east Brazil. Phylogenetic analysis of the recA of strains representing the genetic variability, as determined by rep-PCR, grouped the strains into four clades. Clades I and II represent B. cenocepacia lineages IIB and IIIA, respectively, while the strains in Clades III and IV clustered more closely to Clades I and II than to strains of any other BCC species. MLSA confirmed the existence of the four clades with a 1.00 posterior probability. A distance matrix indicated a low level of divergence among the strains of the four clades found in the MLSA, proving that B. cenocepacia lineages IIIA, IIIB, and a new lineage of B. cenocepacia are associated with onion sour skin in the semi-arid region of north-east Brazil. Also, surprisingly, no strain was identified as B. cepacia, indicating that this species probably does not play a role in this disease in this region.