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Microbes can shorten shelf-life of non-incubated eggs in the wild, finds new study

– Research led by University of California, Berkeley, biologists has implicated microbial infection as a culprit for why some birds start incubating eggs in a nest before the clutch is complete, resulting in eggs hatching at different times and putting younger chicks at a significant disadvantage that often leads to death.

Incubation seems to protect eggs from bacterial and fungal infection, according to the study, to be published in the Nov. 22 issue of the Proceedings of the Royal Society B.

 Checking chicken egg for microbes
A chicken egg is candled to determine if field experiments that exposed it to microbes and ambient temperatures have killed the embryo. (Photo by Steve Beissinger)
 

"We found that freshly laid eggs that aren't incubated have a shorter shelf life," said Steven Beissinger, principal investigator of the study and professor and chair of the Department of Environmental Science, Policy and Management at UC Berkeley's College of Natural Resources. "Without incubation, eggs are more vulnerable to infection from micro-organisms. This paper is the first to present evidence that microbes may be connected to egg viability and incubation patterns in wild birds."

Beissinger explained that birds generally produce no more than one egg per day, so there may be a one-to-two-week lag between the laying of the first and last eggs in a nest. Ornithologists have puzzled over why many birds start to incubate their eggs before all are laid, which causes them to hatch at different times. The pattern leads to a high mortality rate for the clutch's younger hatchlings because they cannot compete with their bigger siblings that got a head start in life.

"The dominant theory is that birds intentionally do this because they would not be able to adequately feed all the young if they hatched at once," said Beissinger. "By staggering the hatching order, they can control the food demand from the number of young they have to care for at any one time."
However, a number of recent studies, including one conducted by Beissinger, have caused some to question that theory. By moving eggs that were laid around the same time into one clutch - a procedure they nicknamed "egg bingo" - researchers found that bird parents were able to successfully raise more young if they hatched together rather than weeks apart.

So, the hunt began for an alternate explanation of birds' hatching patterns. Beissinger worked with Mark Cook, a UC Berkeley post-doctoral researcher in ecosystem sciences and lead author of the paper; Gary Toranzos and Roberto Rodriguez from the University of Puerto Rico's Department of Biology; and Wayne Arendt from the USDA Forest Service International Institute of Tropical Forestry.

The researchers conducted the study at two research sites in Puerto Rico. One site was at the top of the Luquillo Experimental Forest, a cool, very humid cloud forest with an average daily temperature of about 70 degrees Fahrenheit. The other site was a drier lowland forest at the base of the mountain with daily temperatures averaging 81 degrees.

The researchers used freshly laid, free range chicken eggs provided by a local supplier. They sampled the shell surface of 164 eggs for baseline bacteria levels, then randomly assigned the eggs to one of the two sites and exposed them for periods of one, three, five or seven days. After the exposure time, the eggs' yolk and albumen, or egg white, were separately tested for the presence of microbes.

They found that microbes were rapidly able to enter the egg through its pores and overwhelm the embryo within three to five days. The longer the exposure time, the greater the rate of infection for eggs at both sites. Microbes appeared on the inner membrane after just one day of exposure. Microbes reached the egg white after three days, and the yolk after five days.

Eggs in the cloud forest were more susceptible to infection. Sixty percent of the eggs were infected by microbes after five days in the cloud forest compared with 25 percent of eggs infected in the lowland forest.

"It's damper in the cloud forest, which makes it easier for the microbes to get through the egg shell," said Cook. "Water tends to help bacteria and fungi penetrate the pores of the shell. Also, once fungi take hold on the shell, it quickly starts deteriorating the thin protective cuticle that covers the shell."

The next step was to determine whether microbial infection could impact hatching success. The researchers began another round of experiments with 152 freshly laid chicken eggs. They randomly assigned the eggs to either the cloud forest or lowland forest sites, and exposed them for five days. In addition, half the eggs were wiped twice daily with alcohol to kill germs on the shell surface. After five days, the eggs were then incubated to test hatching success.

Several dozen baby chickens later, the researchers found that of the eggs in the cloud forest, 76.5 percent of the cleaned eggs hatched compared with 22.7 percent of the eggs that had been left uncleaned. In the lowland forest, only 40 percent of the eggs hatched regardless of whether they had been cleaned or not, suggesting that temperature played an important role along with microbes in controlling embryo viability.

Incubation warms eggs up to about 95 F, the optimal temperature for development. When temperatures drop to about 77 F or below, cell division stops, but the embryo can remain alive in a state of limbo.

But with temperatures in the danger zone of 77 to 95 F, tissues are more likely to develop incorrectly, leading to higher embryo mortality.
Eggs in the cloud forest that were cleaned had the highest rate of viability because they were protected from both microbial infection and dangerous temperatures.

"For the cleaned eggs that didn't hatch in the lowland forest, we're primarily seeing the impact of temperature," said Cook. "The two factors - microbes and temperature - can act independently or together to reduce the shelf life of the non-incubated eggs."

The results also shed light on why some birds partially incubate their eggs. That is, they may only incubate their eggs for two or three hours in a day.
"Some eggs are filled in the albumen with anti-microbial agents," said Beissinger. "For those agents to react with the enzymes in the albumen and work well, they need to be warmed. Warming the eggs up a bit may serve to clean out the bacteria and fungi that may have gotten in through the shell."

"Our research shows the tradeoff that birds are making in their incubating behavior," said Beissinger. "Birds may be beginning incubation early in part to maintain the viability of the embryos inside the eggs, even though they are risking a disadvantage to their latter-hatching offspring."

This research was supported by the National Science Foundation and the USDA Forest Service.