Spermatic and oxidative profile of domestic cat (Felis catus) epididymal sperm subjected to different cooling times (24, 48 and 72 hours)

Cooling stored epididymal samples for several days allows facilities to transport and process genetic material post‐mortem. Improvements to this practice allow the preservation of sperm from domestic cats, which are the ideal study model for wild felids. However, the modifications in spermatic features and the oxidative profile are not fully understood in cats. This information is necessary for the development of biotechniques, such as new extenders for cryopreservation. Therefore, the purpose of this study was to evaluate the spermatic and oxidative profile in samples from the epididymal cauda of domestic cats cooled at 5°C for 24, 48 and 72 hr. Spermatozoa were collected from the epididymis cauda. Evaluations consisted of computer‐assisted sperm analysis (CASA), plasma membrane integrity (eosin/nigrosin), acrosome integrity (fast green/rose bengal), sperm morphology, sperm DNA integrity (toluidine blue), mitochondrial activity (3′3 diaminobenzidine), activity of the antioxidant enzymes glutathione peroxidase (GPx) and superoxide dismutase (SOD), measurement of lipid peroxidation (TBARS) and protein oxidation. A decrease in sperm motility parameters was observed after 72 hr of cooling (i.e. total and progressive) with a higher percentage of minor (37.7 ± 6.3%) and total defects (53.4 ± 6.3%). Additionally, a decrease in high mitochondrial activity (Class I: 16.6 ± 2.2%) occurred after 72 hr. The decrease in motility rates after a long cooling time probably was caused by the increase in sperm abnormalities. A long cooling time causes cold shock and mitochondrial exhaustion, but there was no observed change with the oxidative stress condition. Therefore, cat epididymal sperm stored at 5°C appear to maintain a high quality for up to 48 hr of cooling time.     

A fast,low‐cost and efficient method for the diagnosis of sperm DNA fragmentation in several species

Sperm DNA fragmentation is a condition that interferes directly in the reproductive efficiency. Currently, there are several methods for assessing the sperm DNA integrity, such as Alkaline Comet, TUNEL and Sperm Chromatin Structure Assay. However, many of these techniques are laborious and require high‐precision equipment. Thus, the development of new techniques can optimize the evaluation of sperm DNA damage. Therefore, the aim of this study was to standardize the toluidine blue (TB) stain technique for the analysis of DNA fragmentation of dog, cat, bull, stallion and ram spermatozoa. For this purpose, we used six animals of each specie (n = 30), in reproductive age. Sperm was collected by different methods according to the particularities of each species, and such samples were divided into two aliquots: a sperm sample was kept at 5°C (considered as intact sperm DNA), and the remaining samples were submitted to the induction of DNA fragmentation by exposure to ultraviolet light for 4 hr. Samples were then mixed with the intact sample to obtain known and progressive proportions of sperm with fragmented DNA (0%, 25%, 50%, 75% and 100%). Semen smears were performed and subjected to staining with TB. Blue‐stained spermatozoa were considered to have DNA fragmentation. We observed high linear regression coefficients between the expected proportion of damaged DNA and the results of TB for dog, cat, ram, bull and stallion samples. In conclusion, TB stain was considered a fast and effective technique for the study of spermatozoa DNA in several species.     

Greenhouse and field screening of wild<Emphasis Type="Italic">Lycopersicon</Emphasis> germplasm for resistance to the whitefly<Emphasis Type="Italic">Bemisia argentifolii</Emphasis>

Thirty-two accessions of wild tomato (Lycopersicon spp.) germplasm were evaluated for resistance to the whiteflyBemisia argentifolii Bellows ⇐p; Perring in a greenhouse choice bioassay. Density data were recorded for the adaxial and abaxial leaf surfaces for (i) all life stages ofB. argentifolii and (ii) types I, IV, V and VI trichomes. Individual plant selections (33 from 22 wild tomato accessions) with high resistance were subsequently tested in the field to verify the resistance found in the greenhouse screening. Resistance was defined by the density of all life stages of the whitefly observed on the eight leaflets sampled at nodes 5 and 7. Only type IV trichomes had a consistent (but low) and significant negative correlation between trichome density and whitefly density for various life stages. The highest whitefly resistance was observed inLycopersicon pennellii accessions LA 716, LA 1340 and LA 2560. The most resistantL. hirsutum f.typicum accessions were LA 1777 and LA 1353. http://www.phytoparasitica.org posting Dec. 9, 2002.     

Kisspeptin,c‐Fos and CRFR Type 2 Co‐expression in the Hypothalamus after Insulin‐Induced Hypoglycaemia

Normal reproductive function is dependent upon availability of glucose and insulin‐induced hypoglycaemia is a metabolic stressor known to disrupt the ovine oestrous cycle. We have recently shown that IIH has the ability to delay the LH surge of intact ewes. In the present study, we examined brain tissue to determine: (i) which hypothalamic regions are activated with respect to IIH and (ii) the effect of IIH on kisspeptin cell activation and CRFR type 2 immunoreactivity, all of which may be involved in disruptive mechanisms. Follicular phases were synchronized with progesterone vaginal pessaries and at 28 h after progesterone withdrawal (PW), animals received saline (n = 6) or insulin (4 IU/kg; n = 5) and were subsequently killed at 31 h after PW (i.e., 3 h after insulin administration). Peripheral hormone concentrations were evaluated, and hypothalamic sections were immunostained for either kisspeptin and c‐Fos (a marker of neuronal activation) or CRFR type 2. Within 3 h of treatment, cortisol concentrations had increased whereas plasma oestradiol concentrations decreased in peripheral plasma (p < 0.05 for both). In the arcuate nucleus (ARC), insulin‐treated ewes had an increased expression of c‐Fos. Furthermore, the percentage of kisspeptin cells co‐expressing c‐Fos increased in the ARC (from 11 to 51%; p < 0.05), but there was no change in the medial pre‐optic area (mPOA; 14 vs 19%). CRFR type 2 expression in the lower part of the ARC and the median eminence was not altered by insulin treatment. Thus, disruption of the LH surge after IIH in the follicular phase is not associated with decreased kisspeptin cell activation or an increase in CRFR type 2 in the ARC but may involve other cell types located in the ARC nucleus which are activated in response to IIH.     

A procedure for combining a quantitative inheritance study with the first cycle of a breeding program

Summary The number of effective factors controlling the inheritance of a quantitative character can be estimated by analysis of F₃ lines where the parental lines are homozygous. The procedure is illustrated with data from a study of pod length inheritance in common bean (Phaseolus vulgaris L.). The mean pod length of Sprite, the parent with longer pods, was compared statistically with the means of F₃ progenies derived from F₂ plants obtained from the appropriate tail of the F₂ frequency distribution for pod length. The data indicate that analysis of F₃ progenies avoids much of the underestimation of the effective factor number which is characteristic of other procedures. The number of effective factors was estimated to be a minimum of 4 and the correlation between random F₂ parents and F₃ progeny means was r=0.82.Florida Agricultural Experiment Station Journal Series No. 406.     

Efficient detection of asparagus monoploids for the production of colchiploid inbreds

Summary An experiment was conducted to develop a method for monoploid seedling detection which is more efficient than making cytological examinations of twin seedlings. A recessive seedling marker gene (basal stem color) was used to detect parthenogenesis in female plants. Out of 10 seedlings showing the marker character, 9 were monoploid as expected. The frequency of parthenogenesis among single seedlings was approximately 1 seedling in 2,400. The marker gene technique increases the efficiency of cytological examination from approximately 1 or 2% to 90%.Florida Agricultural Experiment Station Journal Series No 3681     

Inheritance of the virgata pattern of partly colored seed coats in ‘Early Giant’ common bean

The inheritance of the virgata pattern of partly colored seed coats found in common bean (Phaseolus vulgaris L.) Early Giant (EG) was studied by a series of test crosses with line 5-593 and genetic stocks developed by backcrossing selected genes into the recurrent parent 5-593, a Florida dry bean breeding line with a self-colored, black seed coat with genotype T Z Bip P [C r] J G B V Rk. Analysis of the F₂ from the cross EG × 5-593 led to the hypothesis that the virgata pattern of EG has genotype t z bip^vgt, where vgt stands for virgata. The test cross EG × t z virgarcus BC₃ 5-593 confirmed the hypothesis that EG carries t z from data recorded in F₁, F₂, and 27 F₃ progenies from randomly selected F₂ plants. The F₃ segregation was also consistent with the hypothesis that a single recessive gene converts virgarcus into virgata. The test cross EG × t z bip bipunctata BC₃ 5-593 failed to show genetic complementation in F₁ progeny, and the F₂ segregated 3:1 for the parental phenotypes virgata and bipunctata, respectively. Including previously published data, all possible crosses were made among bipunctata, virgata, and virgarcus parents, supporting a multiple allelic series at Bip. We propose the gene symbol bip^vgt for the new allele at Bip, where the allelic series has the order of gene dominance Bip > bip^vgt > bip. Based on test crosses, the complete seed-coat color and pattern genotype of EG is tz bip^vgt P [C r] J G B v^lae rk^d.     

Ice sheet and solid Earth influences on far-field sea-level histories

Previous predictions of sea-level change subsequent to the last glacial maximum show significant, systematic discrepancies between observations at Tahiti, Huon Peninsula, and Sunda Shelf during Lateglacial time (approximately 14,000 to 9000 calibrated years before the present). We demonstrate that a model of glacial isostatic adjustment characterized by both a high-viscosity lower mantle (4 x 10(22) Pa s) and a large contribution from the Antarctic ice sheet to meltwater pulse IA (approximately 15-meters eustatic equivalent) resolves these discrepancies. This result supports arguments that an early and rapid Antarctic deglaciation contributed to a sequence of climatic events that ended the most recent glacial period of the current ice age.     

Widmanstaetten patterns in josephinite, a metal-bearing terrestrial rock

Widmanstaetten patterns have been found in several specimens of josephinite, a complex, metamorphosed, metal-bearing rock from placers on serpentinized peridotite in southwest Oregon. The patterns, in interior less-altered regions of the specimens, are typical of exsolution textures produced during slow cooling of a homogeneous metal. The bulk composition of the metal phases indicates that the homogeneous metal must have existed at temperatures above 500 degrees C. Josephinite Widmanstaetten patterns are the first known in terrestrial rock. We interpret them as further evidence that josephinite was derived from the mantle.     

Augmentation of bone repair by inductively coupled electromagnetic fields

Pulsing electromagnetic fields of low frequency and strength have been inductively coupled across skin, directly to bone, to enhance the repair of canine osteotomies. The induced voltage field in bone appears to increase the organization and strength of the repair process at 28 days after "fracture."