CHICAGO — The mysterious deaths this week of seven puppies shortly after landing on a plane in Chicago has rattled the nerves of animal lovers. But there is much that airlines and pet owners can do to ensure the well-being of the 2 million dogs, cats, livestock and other creatures transported by air every year, experts said.

On Wednesday, investigators were looking at factors ranging from heat stroke to carbon monoxide poisoning to pre-existing health problems in connection with a shipment of 14 puppies inside the cargo hold of an American Airlines MD-80 plane from Tulsa, Okla., to Chicago on Tuesday.

The dogs, which were sent by an unidentified commercial dog-breeding operation in Oklahoma, were to make connections at O'Hare to flights that would take them to pet stores in other states. Only seven of the 14 dogs were deemed "airworthy," and they were placed on the second flight, American spokeswoman Mary Frances Fagan said.

Five puppies died at O'Hare, and two expired either while being taken to veterinarians or after arriving at clinics, Fagan said.

"The animals were alive when they came off the plane," she said. But airline baggage handlers taking the puppies to a holding area for animals observed that some of the dogs "looked lethargic," Fagan said. "People were trying to cool them off."

A variety of breeds were in the shipment, she said, adding that necropsy reports on the dead animals won't be available for a week.

Each airline has formulated its own rules for animal transport, according to the U.S. Department of Agriculture. Passengers who are planning to carry their small pets on board with them should note that some airlines restrict the maximum number of animals allowed in the passenger cabin.

The carriers must abide by federal Animal Welfare Act regulations governing the transport of dogs and cats in aircraft cargo bays, USDA spokesman David Sacks said. The regulations include keeping cargo areas heated or cooled as necessary to maintain a temperature and humidity that ensures the well-being of the animals. The cargo areas also must be pressurized above 8,000 feet.

Pet owners must have a health certificate from a veterinarian stating that the animal is fit to fly. In addition, owners shipping their animals should make sure they use containers that have adequate openings for ventilation. They also must have portable carriers large enough to give the animal enough space to turn around while standing as well as to sit upright, according to the International Airline Transportation Association. Food and water containers accessible from outside the portable carrier are also required.

Officials said pet owners also should:

—Book animals during moderate weather, if possible, and book them on flights that have the least number of connections (none is preferable.)

—Avoid holidays and weekends when airline baggage-handling staffing is typically at lower levels.

—Have someone waiting at the destination airport to pick up the animal as soon as possible.

—In addition, the USDA stipulates that no more than two puppies or kittens, eight weeks to six months of age, may be transported in the same enclosure aboard air carriers.

Despite concerns about the stress animals experience while being transported as live cargo, statistics show that air travel is the safest form of transportation for animals as well as people.

Some 144 animals, including 122 dogs, died while being transported via air in the past five years, according to data reported by the U.S. Department of Transportation. Those are only the cases involving pets reported by the airlines to the government. The tally does not include deaths involving animals shipped by commercial breeders or livestock.

The department last month released data showing that "short-faced" dog breeds, such as pugs and bulldogs, represent about half of the canines that die while being transported on planes.

Continental Airlines reported 40 animal deaths during transport in 2009, the highest number among U.S. carriers. American was second, with 28 deaths; followed in the top five by Delta Air Lines, 17 deaths; Alaska Airlines, 16; and United Airlines, 13 deaths, according to the U.S. DOT.