Ferret Study Reinforces Role of Aerosols in SARS-CoV-2 Spread

To study virus transmission, Herfst and his colleagues stacked two ferret containers on top of one another, connected only by a 15 cm–wide duct made of PVC pipe with four 90-degree turns. They infected each of three groups of four ferrets, called “donor ferrets,” with a different virus. One set received SARS-CoV-2; another group got SARS-CoV, the virus responsible for the 2003 SARS outbreak; and a third was infected with H1N1 influenza, the virus behind the 2009 flu pandemic, as a positive control. “The proof for airborne transmission in humans is super limited, and you can’t do those experiments in humans for obvious reasons,” says Erasmus Medical Center virologist Sander Herfst, the senior author of the new study.

The researchers placed each donor ferret by itself in the bottom of the two-cage apparatus and a single healthy ferret, called the “indirect recipient ferret,” in the top cage, with a steel grate closing both ends of the duct to trap any food, feces, or other large particles. A fan carried air from the bottom cage to the top cage at 100 liters per minute. If COVID-19 transmission were only possible via large droplets, the researchers hypothesized that no indirect recipient ferrets should become infected, as the steel mesh, the duct’s right-angle turns, the one-meter distance between animals, and gravity should all work against large droplets rising to the top cage.

University of California, Davis, engineer William Ristenpart, who studies the dispersion of airborne pathogens and was not involved with the study, says the experimental setup mimics classic experiments from the 1960s investigating the transmission of influenza. “The little serpentine ductwork they have really prevents ballistic droplets to be carried upward in the airflow,” he tells The Scientist. 

ABOVE: In the experiment, an infected donor ferret was housed in the bottom cage, and a healthy indirect recipient ferret was housed in the top cage. Air from the donor ferret’s cage traveled upward to the indirect recipient ferret through a duct with multiple right angles meant to capture large particles.

JASMIN KUTTER, ERASMUS MEDICAL CENTER

The researchers swabbed the ferrets’ noses and throats every other day to measure the amounts of viral RNA and infectious virus present. By the end of the 13-day experiment, all indirect recipient ferrets in the influenza and SARS-CoV groups had caught the viruses from the donor ferrets, while two of the four SARS-CoV-2 recipients were infected. The four donor and four recipient ferrets infected with SARS-CoV became seriously ill, exhibiting breathing difficulties and less activity than usual, while those infected with SARS-CoV-2 were fully asymptomatic.

“It’s another brick in the edifice of evidence for long-range aerosol transmission,” says Ristenpart.

The efficiency of long-range SARS-CoV-2 transmission in ferrets may also point to why the virus has spread so aggressively through minks, which also belong to the Mustelidae family, Herfst says. Millions of minks have been culled in the Netherlands and Denmark due to virus outbreaks.

Historically, scientists have considered aerosols to be particles smaller than five microns across, although Marr says she does not know how that number was derived. In a letter published in Science earlier this month (October 16), Marr called for the size threshold to be raised to 100 microns, noting that particles larger than five microns can behave as aerosols—lingering in the air, rather than dropping to the ground shortly after being emitted.

The authors of the new study note that they found particles larger than 10 microns at the entrance of both cages, but they could not specify their maximum size or their composition, a limitation of the aerodynamic particle sizer they used. They suspect that the particles came from the lower cage because of the direction and high speed of the airflow.

Ferrets groom themselves by licking their fur, which could send a cloud of virus-laden fur particulates into the air. The study authors found tiny amounts of viral RNA but no infectious virus on the fur of the SARS-CoV donor ferrets. Ristenpart, who helped find that guinea pigs could transmit influenza through aerosolized fur particles in an August study published in Nature Communications, says he appreciates that the authors discussed the possibility that the virus could be transmitted via fur particles. 

Although the authors acknowledge that their experiments do not “discriminate between transmission via small aerosols, large droplets, and fomites,” Marr says she thinks it’s highly unlikely that particles larger than 50 microns traveled to the upper cage based on the cross-sectional area of the duct, the speed of the air, and the 90-degree turns.

The study comes weeks after the US Centers for Disease Control and Prevention acknowledged that the novel coronavirus can spread via aerosols. The agency updated its guidelines to include airborne transmission in mid-September, but removed the language the next day. If the virus were found to spread among people predominantly through aerosols, Marr says, that would carry practical implications for recommendations of types of masks and ventilation systems best suited to filter out airborne particles. 

Now that Herfst has found that the virus transmits effectively through the air, he says he hopes to next tackle how long infectious virus can be detected in the air around infected ferrets, which would help answer how long individuals remain contagious.

J.S. Kutter et al., “SARS-CoV and SARS-CoV-2 are transmitted through the air between ferrets over more than one meter distance,” bioRxiv, doi:10.1101/2020.10.19.345363, 2020.