Reid Robertson wasn’t quite sure what he was looking at on the screen in front of him at the University of Arizona Veterinary Diagnostic Laboratory. The test should have returned a definitive negative – the preferred outcome – or a clear positive. This one was neither, and lurking in that grey area was a potentially devastating poultry condition called virulent Newcastle Disease.

According to the United States Department of Agriculture Animal and Plant Health Inspection Service, virulent Newcastle Disease, or vND, classified as reportable in the U.S., is one of the most infectious diseases of poultry in the world. The disease is so virulent many birds die without showing any clinical signs, which include sneezing and coughing, gasping for air, nasal discharge, diarrhea, tremors, circling, drooping wings, twisting of head and neck, complete stiffness, swelling around the eyes and neck, and sudden death.

Thanks to stringent biosecurity measures, U.S. domestic poultry had enjoyed 15 years of vND-free status. But on May 18, the virus was identified in “backyard exhibition chickens” in California, where 94 cases have since been confirmed by the USDA APHIS.

California’s Cooperative Extension estimates there are 100,000 backyard bird enthusiasts in the state. In addition to raising chickens for egg and meat production, many owners of personal flocks show their chickens, often traveling across state lines to do so.

“The movement of backyard domestic poultry across state lines is more difficult to regulate than commercial poultry,” said veterinarian Sharon Dial, director of the UA Veterinary Diagnostic Laboratory, or VDL.

The UA VDL’s mission is to protect production animal agriculture and the citizens of Arizona from foreign animal diseases, zoonotic diseases and food safety issues, any of which could have serious economic consequences. In California, for example, the chicken industry was responsible for as much as $11.34 billion in total economic activity in 2016, while creating or support as many as 50,688 total jobs, according to the U.S. Poultry and Egg Association.

“In Arizona, we have had an enhanced surveillance in place for sick bird reports since the high pathogenic avian influenza outbreak of 2015 that occurred in the Midwest,” said Arizona State Veterinarian Peter Mundschenk, whose office collects the samples that are then sent to the UA VDL.

Surveillance testing is one of the main purposes of the UA VDL, which acts as the state veterinary diagnostic laboratory for Arizona and is Arizona’s only member laboratory of the National Animal Health Laboratory Network, or NAHLN.  The National Animal Health Laboratory Network is a nationally coordinated network and partnership of more than 60 federal, state and university-associated animal health laboratories managed by the National Veterinary Services Laboratories, or NVSL. NAHLN laboratories provide diagnostic testing to detect biological threats to U.S. food animals, thus protecting animal health, public health and the nation's food supply. 

When vND was reported in California, the State Veterinarian’s Office decided to investigate all reports of death loss in backyard domestic poultry for enhanced surveillance. They also distributed educational materials on the disease and biosecurity to all National Poultry Improvement Plan participants in Arizona.

One team visited a breeder who had called expressing concern about periodic deaths among his chickens. Those samples ended up on Robertson’s desk at the UA VDL.

“It was surveillance for avian paramyxovirus and avian influenza,” said Robertson, who graduated from the UA with a bachelor’s degree in microbiology and has been working at the UA VDL for two years. “We perform molecular testing using PCR, which is polymerase chain reaction. We're looking for the nucleic acid of the virus itself.”

If the virus is present, in the presence of the proper reagents, its DNA will replicate quickly in a process called amplification. When charted on a graph, the amplification curve will rise exponentially until it passes a specific threshold, and then level out as the reaction runs out of reagents. If no virus is present, the curve won’t rise as there is no DNA to replicate.

“What happened with this one is we had our initial curve, exponential growth, and then it tapered off. The trajectory became curved instead of plateauing like it should,” Robertson said. “Normally, either you have nothing or you have something. It's a pretty self-explanatory test. But for this one, I noticed it was a bit strange.”

The test wasn’t positive, but it wasn’t completely negative, either. While Robertson could have reported the results as negative – the curve hadn’t passed the threshold, so it was, technically, negative – he couldn’t ignore the strange results, especially on a test for a reportable disease.

“I informed my superior and we ended up talking with the National Veterinary Service Laboratory in Ames, Iowa,” Robertson said. “I sent them the information I had, which was the curve, and asked them, ‘What am I looking at here? What's going on?’”

Through the NAHLN system, suspect or detected samples are sent to the NVSL for test confirmation. When the NVSL examined the sample, they found DNA viral particles that were classified as a non-virulent to domestic poultry avian paramyxovirus. Avian paramyxovirus has many subspecies, one of which is virulent Newcastle Disease.

The NVSL cultured the virus, which was found to be a subspecies of paramyxovirus that is virulent to pigeons. Historically, Arizona has diagnosed pigeon paramyxovirus when there have been sporadic deaths in pigeons, though the subspecies did not affect domestic poultry. Because this subspecies was found on a swab from a chicken, there was some concern the virus might be mutating to affect domestic poultry.

Further testing by the NVSL yielded negative results.

“We all did a big happy dance,” Dial said. “vND is fairly high pathogenicity – animals start dying quickly. It's a high mortality disease. It's economically hard on production poultry; it's emotionally hard and economically hard on people who are not traditional poultry farmers.

“While this is in backyard chickens and while most of our production poultry industry is very good about biosecurity, all it takes it one small break and then it can get into the production poultry, and that can have a substantial economic impact on Arizona,” said Dial, who is serving as interim associate dean for academic and faculty affairs for the UA’s proposed College of Veterinary Medicine. “This gave us an opportunity to say, are we ready to deal with this if it is positive? Because we must be prepared.

“We are a border state, and the comment has been made many times by many people, including the state veterinarian and the FBI, that it's not if, it's when we are going to have an outbreak here. No one wants to be the director of a laboratory when that happens, even though it would bring attention to our mission.  But, if I am director when it happens, I am very pleased to know that the processes and people, like Reid, are in place and the system is working.”

As for Robertson, his emotions ran the gamut from fear — did I do something wrong? — to apprehension of identifying the outbreak of a deadly disease. When it was all said and done, he simply felt relaxed, and maybe a bit more experienced than the day before.

“I'd never seen anything like that before,” Robertson said of the sample that, ironically, came from a chicken that appeared healthy. “Surveillance is really the first line of protection. You never know when and where that next outbreak may appear.”

Which is why, Mundschenk added, “The Department of Agriculture and State Veterinarian’s Office will continue surveillance of reported sick birds and any high-risk birds to make sure we are protecting our commercial industry through early detection.”