Zoology Unit 4 PDF

. In Vitro Fertilization and Embryo Transfer ~. \ It involves retrieving In vitro fertilizat ion (IVF) is a type of assistive reproductive technology (ART)....

. In Vitro Fertilization and Embryo Transfer ~. \ It involves retrieving In vitro fertilizat ion (IVF) is a type of assistive reproductive technology (ART). This fertilized egg is known as an eggs from a woman's ovaries and fertilizing them with sperm. embryo which is then transferred in the-~om an's uterus. genetic problems and IVF-ET is a complex series of procedures used to help with fertility or prevent endometriosis, cervical assist with the conception of a child. It is advisable in case of tubal disease, y and failed inductio n hostility , unexplained infertility , male factor infertilit follicular growth, oocyte In vitro fertilizat ion involves induction of super ovulation, monitor ing of refrieva l and in vitro fertilization. This is followed by embryo transfer. The IVF comprises of basic 5 steps. They are enlisted as follows: a. Stimulation b. Egg retrieval c. Insemination and fertilization d. Embryo culture e. Embryo selection and embryo transfer Step I: Stimulation It is also termed as super ovulation. cycle. In a normal condition, a woman produce one egg during each menstrual eggs increase the However, IVF has requirem ent of multiple eggs because use of multiple probabi lity of developing a viable embryo. boosts up. Fertility drugs are prescribed to the woman, so that the egg production substanc es, that causes The fertility drugs consist of exogenous gonadotropins and similar of eggs per cycle. hormon al stimulat ion of the ovary causing the production of large number followin g the use On the third day of menstruation, treatme nt cycles are generally started, ment of multiple follicles of the ovaries. of fertility medicines to trigger the develop inal ultrasou nds and blood During this step, females are required to undergo regular transvag tests to check hormon e levels. The stimulat ion of ovary can be performed by 2 major protocols: of the pituitary ovarian axis 1. Lengthy protocol is the one where the suppression (down regulation) e (GnRH) agonist. is perform ed by the prolonged use of a gonadotropin-releasing hormon usually after 10-14 days, subsequent ovary Once the process of down regulation is accomplished, hypersti mulation generally using follicle stimulating hormone (FSH) starts. consist of prescription of 2. Short protocol is the one where down regulation part is neglected and development of multiple injectable gonadotropins under regular monitor ing in order to trigger the follicles of the ovaries. r growth is checked The frequen t monitor ing checks the level of oestradiol, and the follicula via gynaecologic ultrasonography. Usually, ten days of injection is required. prevents the During the last days of stimulation, the use of GnRH antagonists usually spontaneous ovulatio n during the cycle. ~f the ovulation It blocks the natural surge of luteinizing hormones (LH) facilitating the start process by use of injectable human chorionic gonadotropins. Step II: Egg retrieva: It is also termed a·s follicular aspiration. s body. It is a minor surgery perform ed for the removal of eggs from the woman' final maturat ion is After the ovarian follicles reach a certain level of degree of development, induced by an injection of human chorioni c gonadot ropin (hCG). hCG hormone plays a role as that of luteinizing hormon e (LH). 40 hours. After a single hCG injection, ovulation would take place between 38 and , which is, just However, the eggs are retrieved between 14 and 36 hours after hCG injection before the rupture of foilicles.  This assists for scheduling the process of egg retrieval at a time when the eggs are completely matured. A technique called transvaginal oocyte retrieval is used to retrieve eggs In this process, the woman is given anaesthesia, prior to surgery. The health care provider by using ultrasound images as a guide, inserts a thin needle through the vagina into the ovary and sacs (follicles). Then, the needle is connected to a suction device, that pulls the egg and fluid out of each follicle, one at a time. The same process is repeated for another ovary. Generally, 10-30 eggs are removed. Step Ill: Insemination and fertilization The best quality of embryos that are potent for successful pregnancy are selected. It is also termed as oocyte selection. Along with it, the process called as sperm washing is also conducted. In this process, the inactive cells and seminal fluids are removed from semen in order to prepare it for fertilization. In the case where semen is supplied by a sperm donor, the preparation for treatment takes place before being frozen and quarantined, then it will be thawed ready for use. For about 18hrs, the incubation of sperms and egg (at the ratio of about 75000:1) is done i,n the culture media. In majority of the cases, the egg will be fertilized by that time and the fertilized egg shows two pronuclei. In specific cases such as low sperm count or motility, intracytoplasmic sperm injection (ISCI) can be used to inject a single sperm directly into the egg. Now, the fertilized egg is transferred to a special growth medium and left for about 48hrs until the egg reaches the 6-8 cell stage. Step IV: Embryo culture After, the fertilized egg reaches 6-8 celled stage, embryos are cultured usually 3 days after retrieval. Embryo culture can be performed either in an artificial culture medium or in an autologous endometrial co-culture. Embryo culture in artificial culture medium: In this type of culture, there can be e~ther the same culture medium throughout the process or embryo can be sequentially placed in different media by use of sequential system. Ex. One medium can be used for cul_ture to day 3, and second medium is employed for culture after it, when culturing to blastocyst stage. For the culture of human embryos to the blastocyst stage, both the single and sequential medium are equally effective. The media for artificial culture usually contain glucose, pyruvate, and energy supplying components. However, the addition of the nucleotides, amino acids, vitamins, and cholesterol enhances the performance of embryonic growth and development. The techniques that allow dynamic embryo culture along with fluid flow and embryo movement are also present. A new techniqu2 in development where the embryos are encapsulated in permeable intrauterine vessel. Step V: Embryo selection and embryo transfer 1. Embryo selection: On the basis of the number of cells, evenness of growth and degree of fragmentation, embryos are graded by embryologists. For the selection of embryos, morphological scoring system is considered as best strategy that optimizes pregnancy rates as well. If it is to be choose between embryos of morphologically equal quality, presence of soluble human leukocyte antigen-G (HLA-G) is regarded as a second parameter. Embryos that have reached 6-8 celled stage are then transferred 3 days after retrieval. It has been s~en that blastocyst stage transfer results in higher pregnancy rates. 2. Embryo transfer: The number of embryos that are to be transferred depends on the number available, the age of the woman and other health and diagnostic factors. Most of the clinics and country regulatory bodies tend to reduce the risk of pregnancies ~rrying multiples. \) &; The best embryos are transferred to the patient's uterus by means of a thin plastic catheter that goes through the cervix. Summaries of steps of IVF: ► The follicle maturation along with ovulation is promoted by ovarian hormonal stimulation. ► To achieve fertilization by assisted reproductive technology (ART) several fertilization methods are used. ► Under the cultured conditions, the re-implantation embryo spends certain time that will influence its further development. ► Pre implantation embryo biopsies can be used during this period of time. ► Finally, the embryo is transferred to a recipient female. Risks of In vitro fertilization (IVF): ! Multiple births: There are chances of multiple births if more than one embryo is transferred to the uterus. A multi- fetus pregnancy carries a higher risk of early labour and low birth weight than a single-fetus pregnancy. ! Premature birth and low body weight: It is suggested that IVF causes the birth prior to the normal delivery time with low weight of baby. Ovarian hypersti~ulation syndrome: The use of fertility ·drugs like human chorionic gonadotropins (hCG) to induce ovulation can result in ovarian hyperstimulation syndrome. In this condition, the ovaries get swollen and becomes painful. ! M·iscarriage: For women who conceive using IVF with fresh embryos, the incidence of miscarriage is close to that of women who conceive naturally, i.e. around 15% to25%, however the rate rises with maternal age. ! Complications of egg-retrieval process: Bleeding, inflammation or damage to the intestines, bladder or blood vessels could be caused by the use of an aspirating needle to collect eggs. Sedation and general anesthesia, if used, are also associated with risks. ! Ectopic pregnancy: Aho11t 2% to 5% of women who 11c:;e IVF may have an Pr.torir rrpgnancy. [ctopic pregnancy is the condition in which the fertilized egg implants outside the uterus, generally in a-fallopian tube. Outside the uterus, the fertilized egg can't survive, and there's no way to continue the pregnancy. ! Birth defects: The mother's age is the main risk factor, no matter how the baby is conceived, in the development of birth defects. To decide if babies conceived using IVF might be at increased risk of certain birth defects, further research is required.  APPLICATION OF STEM CELL TECHNOLOGY~ ) develop into many differe nt or specialized Stem cells: Stem cells are the cells that have the potential to ive unspecialized cells that are able to divide and cell types. These stem cells can be thoug ht as primit cells, muscle cells, blood cells and other cells of become specialized cells of the body. Such as liver ic cell type is known as differe ntiatio n. specific functions. The process of changing into specif renew themselves by producing daughter Stem cells are the cells in the body that continually e and become cells that are specific to differe nt cells. Some of these daughter cells go on to chang which all other cells in our body come from their tissues in the body, stem cells are the source cells from cells (lung, heart, skin, retina etc.) is what makes ability to give rise to a variety of differe nt types of is a variety of stem cells seen in our body. These them so valuable to biomedic~I world today. There stem cells while other type of stem cells seen in stem cells present in our bodies are known as adult as embryonic stem cells. Embryonic stem cells developing embryos of humans and animals are known they have the ability to develop in~o a wide variety are easier to isolate days after fertilization of egg and tly. ''---- of cell types which is the topic of great interest curren I y of new cell types. The idea is used to Stem cells are very important: As they give rise to a variet to get new undamaged tissue ex. In brain, heart I replace the damaged tissue by the addition of stem cells ed, stem cell therap y could be applied. etc. In any situation where tissue damage was occurr leukemia and other blood diseases have Stem cells have therapeutic uses: as for years people with treatm ent of their disease. The bone marro w is the beery gettin g bone marro w transplants for successful to replace the diseased cells. Now new uses of stem source of stem cells that give rise to new blood cells cells are being tested in animals. born babies are also found to be rich source of In addition cord blood cells from umbilical cords of new blood diseases. blood stem cells and have been used to treat various disease, diabetes, arthritis, brain disease and eye Stem cel'ls have application in the areas of heart applications. d by high blood sugar (glucose) levels Diabetes:- Diabetes mellitus is a metabolic disease characterize or both. Diabetes was first identi fied as.disease that result from defect in insulin secretion or action glucose (hyperglycemia) leads spillage of glucose associated with sweet urine. Increased levels of blood into urine hence called sweet urine. a hormone produced by pancreatic cells. Insulin Normally blood glucose levels are controlled by insulin elevates (ex after eating food) insulin is released lowers the blood glucose levels when blood glucose ts with diabetes the absence or in sufficient from pancreas to normalize the glucose level. In patien is a chronic medical condition, it lasts lifelong. production of insulin causes hyperglycemia. Diabetes Diabetes can lead to blindness kidney failure, nerve damage. These are caused because of result of damage to small blood vessels called micro vascular disease. Diabetes can also cause strokes, coronary diseases and other large blood vessel diseases called macro- vascular disease. In other application of stem cell therapy for a disease state, diabetic rats have been injected with embryonic stem cells that express insulin. Improvement in glucose control was observed. Transplantation of bone marrow derived cells were found to normalize glucose anctirrsalin levels in diabetic mice and their survival rate was better. Diabetes patients lose the function of insulin producing beta- cells within pancreas. Human embryonic stem cells may be grown in cell culture and stimulated to form insulin producing cells that can be transplanted into the patient. Treated patients may have the chance to return to normal activities. Parkinson's disease: It is a chronic nervous disorder characterized by a fine, slowly spreading tremor, muscular weakness and rigidity and peculiar gait (posture). Muscles become rigid and limbs tremble uncontrollably. Parkinson's disease results from loss of a particular type of brain cell called dopaminergic (DA) neurons in the part of brain. Stem cells are taken and grown into new brain cells and transplant these cells into patient stem cells are induced to become neurons. Such cells could be ideal for developing and testing new drugs to treat brain diseases. Embryonic stem cells are responsible for development of the whole body. Some stem cells will be left in the body even after the removal of umbilical cord after birth. These cells have the capacity to repair damaged cells. However they are limited in numbers.  BIOPESTICIDES Biopesticides are certain types of pesticides that are derived from natural materials like plants (Botanical origin), bacteria, fungi and virus (Microbial origin) and certain minerals. They tend to pose fewer risks than conventional chemicals. Very small quantities can be effective and they tend to break down more quickly, which means less pollution. These include for eg: fungi such as Beauveria sp., bacteria such as Bacillus sp., neem extract and pheromones. Similarly, canola oil and baking soda have pesticide applications and are considered as biopesticides. These pest management tools are based on beneficial micro-organisms and other safe biologically active ingredients. Benefits of biopesticides include effective control of insects, plants diseases and weeds, as well as human and environment safety. It also plays an important role in providing pest management, niche markets and environmental concerns limit the use of chemical pesticide product. Characteristic features of Biopesticides ► Have a narrow target and a vey specific mode of action. ► Are slow acting ► Haave relatively critical application times. ► Suppress, rather than eliminate a pest popukation. ► Have limited filed persistence and a short shelf life. ► Are safe to human and the environment that conventional pesticide. ► Present no residue problems. Types of biopesticides: Bio pesticides Microbial pesticides Plant Incorporated- 1 Protectent {PIPsJ : 1. Microbial Pesticides: It consists of a naturally occurring or genetically controlled microorgsnism (e.g., bacteria, fungus, virus or protozoa) as active ingredients. These pesticides can control many different kinds of pest, althougheach separate active ingredient is relatively specific for its target pest. Microbial suppress pest by- ► Producing a toxin specific to the pest ► Causing a disease ► Preventing establishment of other microorganisms through competition. An example ofa widely used microbial pesticide is subspecies and strains of Bacillus thuringiensis, or "Bt" 2. Plant-lncorporated-Protectant (PIPs): These pesticides are produced from genetic material that has been added to the plant. For e.g., the gene for the Bt pesticide protein is introduced into the plant's own genetic material. Then the plant, instead of the Bt bacterium, manufactures the substance that destroys the pest. - 3. Biochemical pesticides: These are naturally occurring substances that control pest by nontoxic mechanism. Biochemical pesticides include substances, such as insect sex pheromones, that interfere with mating, as well as various scented plant extracts that attracts insect pests to traps. Man - made pheromones are used to disrupt insect mating by creating confusion during the search of mates or can be used to attract male insect to traps. The most commonly used plants are neem (Azardirachta indica), pongamia and mahua. 2-5% neem or mahua seed kernel extract has been found to be effective against rice cutworm, tobacco caterpillar, rice green leafhopper and several species of aphids and mites. BACTERIAL BIOPESTICIDES Bacillus thuringiensis (Bt) is a rod shaped, gram-positive bacteria that forms a spore and is found in the soil. Bt is toxic to caterpillars, some fly larvae, and some beetle larvae but not toxic to other organisms. A few strains of Bt are available in products used are ✓ Bt var.kurstaki is toxic to lepidopteran (butterfly, skipper, and moth) larvae; ✓ Bt var. aizawai is toxic to wax moth larvae; ✓ Bt var. israelensis is toxic to mosquito, midge, fungus gnats, and blackfly larvae; ✓ Bt var. galleriae is toxic to larvae of May or June beetles {white grubs); ✓ Bt var.tenebrionis (or var. San Diego) is toxic to Colorado potato beetle, elm leaf beetle, and willow leaf beetle larvae. ✓ Bt var. tenebrionis does not kill all leaf beetles. MODE OF TOXIN ACTION: A crystalline toxin and spore is usually produced by Bt cells. The toxin is called a delta endotoxin. Bt products usually contain the toxin and spores {environmental resistant stage of the bacterium) but some products do not contain spores. Spores may become bacterial cells inside the insect. Once the insect eats the Bt the delta endotoxin is activated in the insect's gut by enzymes and alkaline (basic) conditions of the gut. A specific pH is required to activate the endotoxin. The endotoxin disrupts the cell walls of the gut. Bacterial cells enter the body of the insect. Infected insects stop feeding in a few hours and die· in a few hours to weeks (frequently 2-3 days). Different strains of Bt have different endotoxins and kill different insects. The endotoxin is not activated in the gut of humans. In summary, Bt is a microbial biopesticide that is very specific to certain insects. It causes insects to stop feeding in a few hours and usually kills insects in a few days. It must be eaten and kills larvae. It does not last long on the plant, may require frequent applications, is considered organic, and is not toxic to beneficials. Bt G.eoe is inset1ed Into crop Crop ls Infected by Europe.an com borer Pest oles when f·eedlng on any plant part