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Analytical Chemistry

New method casts light on a chicken and egg problem

Raman spectroscopy could make chicken breeding more humane by identifying the sex of embryos still in the egg

by Deirdre Lockwood
September 6, 2016

Photograph of 3 eggs in different phases of being analyzed by Raman spectroscopy.
Credit: Anal. Chem.
Researchers use a laser to cut away a piece of shell, revealing the blood vessels in the developing chicken embryo. After analysis by Raman spectroscopy, they seal the hole with tape and leave the embryo to develop to maturity.

For chickens bred to lay eggs, being male is a gloomy prospect. These cockerels develop too slowly to be raised for meat, so they are usually killed within days of hatching by methods including gassing and grinding. The practice culls billions of chicks each year, raising ethical concerns for consumers and animal rights advocates. As a result, both United Egg Producers, the U.S. industry group that represents most hatcheries for egg-laying hens, and the German government have pledged to end the practice in coming years, or once an alternative is available. Now researchers have developed an approach that could help speed this transition: using spectroscopy to identify the sex of a developing chicken embryo while it’s still in the egg (Anal. Chem. 2016, DOI: 10.1021/acs.analchem.6b01868). The method, which has up to 95% accuracy, could allow hatcheries to cull male chick embryos just three days into development, before they are sensitive to pain.

Currently, the sex of chicks can be determined before they hatch by sampling hormone levels or DNA from within the egg after removing a piece of shell. But hormonal tests must be done on about day nine of development, and chicks become sensitive to pain at about day seven, says Roberta Galli of Dresden University of Technology. Moreover, these testing methods require taking a sample from each egg, followed by chemical analysis, which may not be feasible on an industrial scale.

Credit: Roberta Galli
A new method uses Raman spectroscopy to detect the sex of developing chicken embryos within the egg by removing a piece of shell and illuminating the blood vessels within.
Photograph of eggs inside Raman spectroscopy instrument.
Credit: Roberta Galli
A new method uses Raman spectroscopy to detect the sex of developing chicken embryos within the egg by removing a piece of shell and illuminating the blood vessels within.

Galli and her colleagues wanted to develop a less invasive method that could be applied earlier in development. The team has used Raman spectroscopy for other sensitive biomedical applications, so they thought the approach might be able to determine sex, which imparts differences to blood biochemistry. Male blood has different protein and sugar profiles and about 2% more DNA than female blood.

The method the team developed uses a laser to cut a 15-mm-diameter circle in the end of an eggshell. When the researchers remove the shell piece on day three of development, the embryo’s blood vessels are visible. They shine near-infrared light on the vessels and detect the scattering with a Raman spectrometer; the spectrum is rapidly assigned to a sex based on algorithms the team developed. For 101 eggs whose sex was also determined by DNA test, the algorithm correctly identified embryo sex in 90% of cases. However, Galli says they have since optimized the system, nudging the accuracy to 95%—closer to the 98% accuracy of manual sex determination used in industry based on examining the feathers or genitals. After the analysis, the researchers close up the egg with surgical adhesive tape and allow development to continue. About 81% of the eggs they tracked after the test hatched and developed normally, compared to 92% of control eggs, though other control studies report hatching rates of 84–90%.

The team’s lab system can process two to three eggs per minute—much slower than expert chick sexers, who can work at five to eight times that rate. But the team is building an industrial prototype to automate the process and has partnered to test it with Lohmann Tierzucht, a major commercial producer of egg-laying hens in Germany, where demand for an alternative to chick culling is high. Right now the team does not have a cost estimate for the prototype, Galli says, but the fact that the method requires minimal consumable products may keep expenses down.

Rodrigo Gallardo, an expert in poultry biology at the University of California, Davis, calls the technique “very promising” because it can be applied so early in development and is less invasive than other methods. However, he says, it “needs further development and refinement to be used in the poultry industry,” including lowering the processing time, improving the accuracy, and ensuring that the method does not harm or contaminate developing chicks.


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