Structural and ultrastructural characteristics of the yolk syncytial layer in Prochilodus lineatus (Valenciennes, 1836) (Teleostei; Prochilodontidae)

The yolk syncytial layer (YSL) has been regarded as one of the main obstacles for a successful cryopreservation of fish embryos. The purpose of this study was to identify and characterize the YSL in Prochilodus lineatus, a fish species found in southeastern Brazil and considered a very important fishery resource. Embryos were obtained through artificial breeding by hormonal induction. After fertilization, the eggs were incubated in vertical incubators with a controlled temperature (28 degrees C). Embryos were collected in several periods of development up to hatching and then fixed with 2% glutaralclehyde and 4% paraformaldehyde in 0.1 M sodium phosphate buffer (pH 7.3). Morphological analyses were carried out under either light, transmission or scanning electron microscopy. The formation of the YSL in P. lineatus embryos starts at the end of the cleavage stage (morula), mainly at the margin of the blastoderm, and develops along the embryo finally covering the entire yolk mass (late gastrula) and producing a distinct intermediate zone between the yolk and the endodermal cells. The YSL was characterized by the presence of microvilli on the contact region with the yolk endoderm. A cytoplasmic mass, full of mitochondria, vacuoles, ribosomes, endomembrane nets and euchromatic nuclei, indicated a high metabolic activity. This layer is shown as an interface between the yolk and the embryo cells that, besides sustaining and separating the yolk, acts as a structure that makes it available for the embryo. The structural analyses identified no possible barriers to cryoprotectant penetration.

Structural analysis of the Pimelodus maculatus (Lacépède, 1803) embryogenesis (Siluriformes: Pimelodidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Cryogenic preservation of embryos of Prochilodus lineatus (Valenciennes, 1836) (Characiforme; Prochilodontidae)

While the freezing techniques of mammal embryos have been providing promising results, the cryopreservation of teleostean eggs and embryos have remained unsuccessful up to now. Therefore, this work aimed to develop a procedure of cryogenic preservation of embryos of Prochilodus lineatus and to observe, at both structural and ultrastructural levels, the morphological alterations that took place after the application of freezing/thawing techniques. The embryos at the morula stage could not tolerate exposure to the cryoprotectants ethylene glycol monomethyl ether, propylene glycol monomethyl ether, methanol, dimethyl sulphoxide and propylene glycol, presenting 100% of mortality. Embryos at the 4- to 6-somites stage tolerated exposure to propylene glycol and dimethyl sulphoxide, and the results revealed no significant differences (alpha = 0.05) regarding survival from both treatments. None of the freezing, thawing and hydration protocols was effective on preserving embryo viability. The ultrastructural analyses of frozen and thawed embryos showed that cells from ectoderm, somites, notochord and endoderm were structurally intact, with well preserved nuclei and mitochondria. The yolk globules were able to tolerate the freezing process, but the yolk syncytial layer was unorganized, displaying an electron-dense and compacted appearance, collapsed reticules, nuclei with modified chromatin and ruptures on the plasmatic membrane at the contact zone with endoderm. It might be concluded that the procedures tested for freezing were unable to avoid the formation of intracellular ice crystals, leading to drastic morphological modifications and making P. lineatus embryos unviable.

Ground annatto seeds (Bixa orellana L.) in sorghum-based commercial layer diets and their effects on performance, egg quality, and yolk pigmentation

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

The traceability of animal meals in layer diets as detected by stable carbon and nitrogen isotope analyses of eggs

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Feed type for induced molting of commercial layer hens

An experiment employing three hundred and twenty 81-week-old Lohmann LSL commercial-breed hens was conducted to compare alternative induced-molting methods with the conventional method (fasting). Induced molting lasted 28 days at most, production and quality being monitored for four periods of 28 days thereafter. A completely randomized experimental design with five treatments, eight replicates of eight birds each per plot was adopted. The following experimental treatments were applied until a loss of 26% of body weight was reached: T1 - fasting, T2 - wheat bran ad libitum, T3 - rice bran ad libitum, T4 - cracked rice ad libitum, T5 - ground alfalfa ad libitum. Birds were then fed production diet ad libitum, except for those on treatment T1 (fasting) which received 30, 60 and 100 g/bird/day and then feed ad libitum. During induced molting the birds were exposed to a natural photoperiod and at day 28 that period was increased by 30 minutes/week until reaching 16 hours of light/day. The characteristics evaluated during induced molting were: feed intake, body weight changes and laying percentage. In the post-molt period, performance (feed intake, laying percentage, egg weight, egg mass, feed conversion ratio per dozen and per egg mass and percentage of broken eggs) and egg quality (specific gravity, eggshell breaking strength, percentages of eggshell, yolk, and albumen, eggshell thickness, yolk color and Haugh unit) were evaluated. Every 28 days one egg was collected from each repetition for three consecutive days for quality assessment. The use of rice bran and wheat bran is viable as molting inducers since the birds given those treatments display performance and egg quality similar to those fasted during the induced molting and also because these ingredients promote easier handling, eliminates the need for grinding and feed-mixing equipment and, being less aggressive, provide greater bird welfare.