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Water for IVF and ART

 
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Impact of Water

Impact of water on animal Assisted Reproductive Technologies (ART)

In in vitro fertilization (IVF), water is the main component of the culture medium used for the culture of embryos. Water is also used to rinse containers and surfaces. Water quality has been shown to affect the overall rate of success of IVF procedures. Indicators of IVF quality include the percentage and quality of blastocysts obtained (which can be expressed by inner cell mass (ICM)/total cells rate) and the ability of in vitro produced embryos to survive cryopreservation, an indicator of embryo quality. These parameters are greatly affected by culture conditions.

In artificial insemination (AI), water is used to prepare the semen extender (used as a diluent, to protect the sperm and preserve its fertilizing ability). Its quality can affect the fertility and viability of the extended semen.

Various contaminants in the water may have an effect on animal assisted reproductive technologies:
  • Bacteria and their by-products
    Bacterial contamination is a main concern in ART. Various technologies are available to minimize the levels of bacteria in water (such as reverse osmosis, 0.22 µm filters, ultra-violet light). Bacterial contamination may also occur during the storage of the purified water. It is therefore advisable to use freshly purified water when performing these procedures. In addition, bacterial endotoxins (also called lipopolysaccharides, LPS) are suspected to cause embryo fragmentation, suppress embryonic development and eventually lead to low pregnancy rates. The use of an ultrafiltration cartridge (BioPak) can reliably reduce endotoxin and bacteria levels in ultrapure water. 
  • Ions, heavy metals (copper, lead, cadmium…)
    It is important to carefully control the osmolarity of the solutions when performing ART. Therefore, the ionic levels in the water used to prepare these solutions should be kept to a minimum. In addition, heavy metals, such as copper, lead and cadmium, are known to be toxic for spermatozoa and embryo. Media supplementation with fetal calf serum (FCS), bovine serum albumin (BSA), or chelating agents (EDTA) can provide a degree of protection from these toxic substances, but is often insufficient to provide a complete protection. In addition, these supplements are not always recommended, and in many cases serum-free or protein-free medium is used. In conclusion, it is best to use water with a low level of ions (resistivity of 18.2 MOhm.cm). 
     
  • Organics
    Some organics, such as pesticides, endocrine disrupters, etc. which may be present in tap water, can greatly affect in vitro fertilization and artificial insemination. The levels of organic compounds in the water used to prepare solutions used in ART procedures should therefore be as low as possible. 
In conclusion, it is recommended to use purified water for washing glassware and instruments, and final rinses especially should be done with purified water. Water purified by reverse osmosis is generally recommended; in some cases, pure (Type 2) water or even ultrapure (Type 1) water may be required. Some specific types of washers, such as washer-disinfectors for flexible endoscopes, may require a specific water quality. It is important to follow washer manufacturer guidelines and recommendations concerning feed water quality. In addition, compliance to norms or standards may be required.

Examples illustrating the impact of water quality on ART

Many researchers have studied the importance of water quality for ART techniques. Discussed below are a few of these studies, but many more are available (see “additional references”).

1. Bovine embryonic development (IVM, IVF)
A study was conducted by Nagao et al. regarding the effects of water quality on in vitro fertilization and development of bovine oocytes in protein-free medium. Bovine in vitro matured (IVM) oocytes were inseminated and cultured for 7 days in protein-free media prepared with 4 different types of water preparations:
  • Tap water
  • Deionized water (deionized using mixed bed ion exchange resins)
  • Double-distilled water
  • Ultrapure water (double-distilled water purified with a Milli-Q® system) 
The authors observed that the frequency of development to the blastocyst stage in media prepared with ultrapure water was significantly higher than with the 3 other types of water (Figure 1):
Merck:/Freestyle/LW-Lab-Water/applications/IVF-ART/LW-LC-Impact-Image1-460x350.jpg
Figure 1: Development of bovine embryos to the blastocyst stage in protein-free media prepared with 4 different types of water (means of 4 replicates).

They also observed that early development of bovine embryos is seriously affected by the length of storage of the water used for medium preparation (Figure 2):

Merck:/Freestyle/LW-Lab-Water/applications/IVF-ART/LW-LC-Impact-Image2-430x354.jpg
Figure 2: Development of bovine embryos to the blastocyst stage in protein-free media prepared with fresh Milli-Q® water or Milli-Q® water for stored for 1 or 2 weeks.

For additional information, please refer to the original published article.

Reference
2. Swine artificial insemination (AI)
Dr. W.L. Flowers, of North Carolina State University, has studied how semen quality issues can affect the rate of success of artificial insemination programs in swine operations. In particular, the quality of the water added to the powdered semen extender was shown to play an important role in semen quality issues. Bacterial contamination was the main concern, in particular when purified water was stored. A gradual decrease in the shelf life of stored semen over time was observed when poor quality water was used. Water purified by a combination of purification technologies, including reverse osmosis, deionization and treatment with an ultraviolent light yielded the best farrowing rate and number of pigs born alive after 5 days of storage.

For additional information, please refer to the original published articles.

References
In conclusion, freshly produced ultrapure water is recommended for ART procedures. The use of an ultrafiltration cartridge at the point-of-use of the water purification system will prevent any risk of contamination of the water with endotoxins.

More Information
Additional References
  • The Efficacy of the Water Purification System with an Ultra Violet Lamp and Ultrafilter for the Preparation of Bovine Embryo Culture Media. M Nagano, Y Takahashi and S Katagiri. Journal of Reproduction and Development. Vol. 45, 1999, No. 3 June pp.239-242. http://www.jstage.jst.go.jp/article/jrd/45/3/239/_pdf
  • Macromolecular source as dependent on osmotic pressure and water source: effects on bovine in vitro embryo development and quality. P. DUQUE, C.O. HIDALGO, E. GÓMEZ, B. PINTADO, N. FACAL, C. DÍEZ. Reprod. Nutr.Dev. 43, 2003, 487–496. http://www.ncbi.nlm.nih.gov/pubmed/15141434
  • Influence of Water Quality on in Vitro Fertilization and Embryo Development for the Mouse A. FUKUDA, Y. NODA, S. TSUKUI, H. MATSUMOTO, J. YANO and T. MORI. Journal of in Vitro Fertilization and Embryo Transfer, 1987, Vol. 4, No. 1, 40-45. http://www.springerlink.com/content/mg71lk1165221qx0/ 
  • The importance of water quality for media preparation. Wiemer KE, Anderson A, Stewart B. Hum Reprod 1998, 13: 166–172. 
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