United Poultry Concerns February 23, 2006

Sources of Avian Influenza

Information Compiled by Mary Finelli
Assistant to the President
United Poultry Concerns
....on what was the index farm in the catastrophic outbreaks in Pennsylvania in 1983/4 the farmer had created an artificial pond to keep ducks and attract wild waterfowl (Webster and Kawaoka, 1988).

Secondary spread
Secondary spread of AI viruses is mainly by mechanical transfer of infective faeces, in which virus may bepresent at high concentrations and may survive for considerable periods (Utterback, 1984). Birds or other animals that are not themselves susceptible to infection may become contaminated and spread the virus. Shared water or food may also become contaminated. However, for domestic poultry the main source of secondary spread is man. In several specific accounts of HPAI infections strong evidence has implicated the movements of caretakers, farm owners and staff, trucks and drivers moving birds or delivering food, and artificial inseminators in the spread of virus both on to and through a farm (Wells 1963; Homme et al., 1970; Halvorson et al., 1980; Alexander and Spackman, 1981; Glass et al., 1981).


Part I: The Role of Wild and Domestic Waterfowl in Avian Influenza Outbreaks in Domestic Poultry
Written by Todd Weaver DVM, National Surveillance Unit
NAHSS Outlook, February 2005

Wildlife surveys associated with an outbreak of AI subtype H5N2 during the 1983-84 epizootic in domestic poultry in Pennsylvania, New Jersey, Maryland, and Virginia failed to isolate the responsible virus from wildlife species within the quarantine area.12  In another study, isolation attempts were made on 4,466 birds and small rodents within the quarantine area and 1,511 nearby waterfowl.13  Wild animals were not shown to be responsible for introducing the virus to domestic poultry during the Pennsylvania outbreak. A separate wildlife survey was conducted from late June through November 1984 within the Pennsylvania and Virginia quarantine zones. Of the 13 AI subtype viruses isolated from waterfowl within the quarantine zones, only one H5N2 subtype was found in a hunter-killed duck Genetic analysis determined that the HA gene could be clearly distinguished from the H5N2 family of viruses that caused the Pennsylvania epidemic. Virus isolates from this study were passaged in chickens; the isolates were found to replicate but did not produce disease. Seroprevalence studies in healthy captive waterfowl at the Baltimore zoo indicated exposure to AI.14  A survey of Louisiana waterfowl showed that AI virus is transmitted to resident mottled ducks and circulates within duck populations throughout the winter.9  In all published waterfowl AI surveys, subtype diversity has been extensive. Subtypes of H5 (0.4 percent), H7 (0.7 percent), and H9 (0.4 percent), which are associated with highly pathogenic influenza outbreaks in poultry or recent human infections, are reported to be isolated less commonly (over 3,100 isolates reported from surveys) than H3, H4, or H6 subtypes (63.8 percent of isolates).1One recent study was an exception, where H5, H7, and H9 isolates accounted for 21.5 percent of isolates.1 Spatial and temporal differences in AI isolate subtypes exist. In the next edition of NAHSS Outlook, the usefulness of waterfowl surveys as part of an ongoing AI surveillance program will be discussed.
C.W. Beard, D.V.M., USDA, ARS. Southeast Poultry Research Laboratory, Athens, GA.

Avian influenza has produced losses of variable severity, primarily in turkeys in the United States, since the mid-1960's. The disease outbreaks in turkeys in the United States have been caused by AI viruses with many of the HA designations. It was in the fall of 1983 that a highly virulent H5 virus produced severe clinical disease and high mortality in chickens, turkeys, and guinea fowl in Pennsylvania. This severe disease, clinically indistinguishable from classical fowl plague, occurred after a serologically identical but apparently mild virus had been circulating in poultry in the area for 6 months.
Outbreaks of less virulent AI have frequently been described in domestic ducks in many areas of the world. The AI viruses are often recovered from apparently healthy migratory waterfowl, shore birds, and sea birds worldwide. The epidemiologic significance of these isolations relative to outbreaks in domestic poultry has led to the generally accepted belief that waterfowl serve as the reservoir of influenza viruses.

There is a considerable body of circumstantial evidence to support the hypothesis that migratory waterfowl, sea birds, or shore birds are generally responsible for introducing the virus into poultry. Once introduced into a flock, the virus is spread from flock to flock by the usual methods involving the movement of infected birds, contaminated equipment, egg flats, feed trucks, and service crews, to mention a few. Preliminary trapping evidence indicates that garbage flies in the Pennsylvania outbreak were sources of virus on the premises of the diseased flocks. Virus may readily be isolated in large quantities from the feces and respiratory secretions of infected birds. It is logical to assume, therefore, that because virus is present in body secretions, transmission of the disease can take place through shared and contaminated drinking water. Airborne transmission may occur if birds are in close proximity and with appropriate air movement. Birds are readily infected via instillation of virus into the conjunctival sac, nares, or the trachea. Preliminary field and laboratory evidence indicates that virus can be recovered from the yolk and albumen of eggs laid by hens at the height of the disease. The possibility of vertical transmission is unresolved; however, it is unlikely infected embryos could survive and hatch. Attempts to hatch eggs in disease isolation cabinets from a broiler breeder flock at the height of disease failed to result in any AI-infected chickens. This does not mean that broken contaminated eggs could not be the source of virus to infect chicks after they hatch in the same incubator. The hatching of eggs from a diseased flock would likely be associated with considerable risk.
Infectious Diseases Society of America, last updated February 17, 2006
Routes of bird-to-bird transmission include:
Airborne transmission if birds are in close proximity
Direct contact with contaminated respiratory secretions or fecal material
Vertical transmission is not known to occur
Other factors that contribute to spread within and between flocks include the following:
Broken contaminated eggs in incubators infecting healthy chicks (see References: OIE 2002)
Movement of infected birds between flocks
Movement of fomites such as contaminated equipment, egg flats, feed trucks, and clothing and shoes of employees and service crews (see References: APHIS, Beard 1998)
Contact with infected wild birds and waterfowl
Fecal contamination of drinking water
Garbage flies (suspected of transmitting the virus during the 1983-1984 epidemic in Pennsylvania) (see References: Beard 1998)
The disease is highly contagious. One gram of contaminated manure can contain enough HPAI virus to infect 1 million birds (see References: APHIS).
University of Maryland, College Park Campus

Transmission and Control of HPAI
Epidemiological surveillance is the second line of defense against AI. Surveillance efforts during the 1983-1984 outbreaks in Pennsylvania were able to track the origin of the virus to live bird markets.
Clinical Microbiology Reviews, Taisuke Horimoto1 and Yoshihiro Kawaoka, January 2001, p. 129-149, Vol. 14, No. 1

Pennsylvanian outbreaks. In April 1983, an avirulent H5N2 influenza A virus (A/Chicken/Pennsylvania/1/83) appeared in chickens in Pennsylvania. Later, in October 1983, virulent influenza viruses (e.g., A/Chicken/Pennsylvania/1370/83) were isolated from chickens after they had decimated numerous flocks (>80% mortality). The virus was eventually eradicated by the destruction of over 17 million birds at a cost of over 61 million dollars 44.
Sequencing studies and RNA-RNA hybridization analyses revealed that the genes of these viruses were derived not from a virulent H5 influenza virus responsible for previous influenza outbreaks but from an avirulent virus maintained in North America 10, 89. In 1985 to 1986, H5N2 avirulent viruses were again isolated from poultry in Pennsylvania and subsequently in other states. Extensive epizoological studies by the U.S. Department of Agriculture finally traced the virus to birds in live-poultry markets in New York City, New Jersey, and Miami 90. Genetic and antigenic analyses demonstrated their close relationship to the H5N2 viruses isolated in Pennsylvania in 1983 to 1984, suggesting that live-poultry markets serve as an important reservoir for avian influenza viruses. This concern was substantiated by the Hong Kong outbreak in 1997.
Oligonucleotide mapping of RNAs from the virulent and avirulent Pennsylvania avian viruses indicated that point mutations were responsible for the acquisition of virulence 10, while a study of reassortant viruses identified the HA gene as a critical determinant of virulence 206. Subsequent comparison of the entire HA amino acid sequences of avirulent, virulent, and revertant viruses demonstrated a single amino acid substitution at residue 13 (Thr to Lys), which is thought to be responsible for the acquisition of virulence 41, 87. This substitution is remarkable in that it coincided with the loss of an oligosaccharide side chain. On the three-dimensional structure of HA, this change is located in the middle of the stalk in the vicinity of the cleavage site between HA1 and HA2, suggesting that the deleted carbohydrate side chain normally blocks the access of cellular proteases to the cleavage site 87. A unique aspect of this outbreak was that the HAs of the avirulent viruses initially isolated from chickens already possessed multiple basic amino acids (K-K-K-R) at the cleavage site, a primary requirement for the virulence phenotype; however, cleavage was blocked by a nearby carbohydrate side chain. Thus, once the carbohydrate side chain of the avirulent virus was lost, the HA became highly cleavable, leading to the acquisition of virulence.
In 1993, avirulent H5N2 viruses were again isolated in live-bird markets in Pennsylvania, as well as Florida. Phylogenetic analysis of the HA genes of these isolates indicated that they shared a common ancestor with an H5N2 virus isolated from shorebirds (ruddy turnstones) in 1991 and did not originate from the Pennsylvania virus that caused the 1983 to 1984 outbreak, suggesting that the virus had been introduced from wild waterfowl into chickens 162. The HA of these avirulent viruses contained an atypical cleavage site sequence (R-K-T-R), requiring only a single mutation for conversion to high HA cleavability 77. This isolate and the avirulent 1983 Pennsylvania virus caution us that potentially hazardous influenza viruses are more prevalent in nature than one would suspect from documented outbreaks.
Emerging Infectious Diseases, V.4 N.3
Robert G. Webster
St. Jude Children's Research Hospital, Memphis, Tennessee, USA
Most importantly, we may influence the appearance of pandemics by changing the methods of live bird marketing by separating chickens from other species, especially from aquatic birds.
Avian Influenza Response Plan 1-20-06 (doc)
Pennsylvania Dept. of Health
Live bird markets are also considered to be an important source of AI.  When AI has been detected through testing, it is important to avoid contact with these markets to avoid bringing disease back onto your premises.  Although closure of the PA, New Jersey, and New York markets for cleaning and disinfection was planned last year, it has been postponed until further notice. 
Research Structure
College of Agriculture and Natural Resources, University of Maryland, College Park.
Agricultural practices contributing to infection with influenza.
“Biosecurity is the first line of defense against all avian influenza viruses. Preventing the introduction of avian influenza by eliminating all contact between commercial poultry and wild birds, swine farms, and  LBMs is a common, routine and successful practice” (USAHA 2002 Resolution Nº 28.) Unfortunately, implementation of strict biosecurity measures in the LBM [live bird markets] systems in the US are difficult because of environmental risks factors and cultural practices that contribute to the perpetuation of viruses in this marketing systems. There are many LBMs in the US, largely concentrated in densely populated areas such as New York (NY), Boston (MA), Chicago (IL), Los Angeles (CA), San Francisco (CA), San Antonio (TX), just to name a few. These LBMs could have a major impact on public health, because HPAI can be a zoonotic disease (18, 85). Intensive education programs are necessary to make owners, suppliers, wholesalers, dealers of commercial poultry and the general public aware of the threat of avian influenza not only for poultry but also potentially to humans. The recent outbreaks of H7 AI in Virginia in 2002 was caused by the introduction of a virus first identified via surveillance in the NY/NJ area and traced to an auction in PA (USAHA and Virginia Dept. of Ag and Cons. Serv.) Likewise, the recent outbreak of H7 viruses in Delaware and Maryland in 2004 can be traced back to viruses circulating in the LBMs. H7 AI viruses were first isolated through sampling conducted in the LBM system of the East Coast in 1994 and efforts to eradicate them have been unsuccessful. A LPAI virus characterized the outbreak in Virginia; where more than 4,000,000 birds were sacrificed to prevent the spread and possible emergence of a HPAI virus. The outbreak caused the state of Virginia huge economic losses. Not controlling the spread of low-pathogenic avian influenza viruses can have devastating consequences as exemplified by the outbreak of H5 in Pennsylvania in 1983-1984 that resulted in the emergence of a HPAI (1, 14, 48, 117). This outbreak led to the declaration of a state of emergency and the implementation of a vigorous eradication campaign. The campaign cost over $63 million in Federal funds and an additional $350 million in increased consumer costs (24). Over 17 million birds died or were slaughtered. The epidemic disrupted the market supply of eggs and poultry meat and impacted also in the cost of pork and beef, which had small increases. Nowadays, a new outbreak of a HPAI such as the 1983 outbreak in Pennsylvania would have an economic impact of up to $120 million to the poultry industry. The costs of not controlling an outbreak would be even more astonishing to the consumers, in excess of $10 billion. It is clearly of interest to devise mechanisms to best contain future epidemics of highly pathogenic AI.
Most efforts at controlling avian influenza in poultry have been focused on the H5 and H7 subtypes; however, control measures should arguably be in place to diagnose and control influenza viruses regardless of the HA subtype. LPAI H9N2 and H6N1 strains circulating in LBMs served as gene donors in the reassortments leading to the emergence of the 1997 Hong Kong H5N1 in China (35, 42, 86). This virus infected at least 18 people; killing six (19, 20, 65, 99). The continuous circulation of non-pathogenic non-H5, non-H7 LPAI in poultry markets of China has contributed to the subsequent outbreaks of highly pathogenic H5N1 viruses in 2001, 2002, 2003, amd 2004, including two more human infections, one of them fatal. Fortunately, these H5 viruses have not spread from person-to-person, although major surveillance efforts in LBMs in China continue to monitor closely the emergence of such a virus. Poultry markets around the world must be considered convenient sampling sites to detect the presence of influenza viruses that can be devastating not only for poultry but also to humans. In the US, Suarez et al. have shown that the LPAI H7 viruses circulating in the LBM system of the Northeastern States have undergone repeated reassortment with viruses of other subtypes (95). It is possible that these reassortments may have contributed to successfully maintaining H7 viruses in the markets. Interestingly, LPAI H7 viruses have circulated in the NY/NJ markets for many years without - to our knowledge- acquiring highly virulent properties. These considerations underscore the importance of interspecies transmission of waterfowl AI to poultry, and alert to the large gap of knowledge regarding selection forces and molecular mechanisms that lead to the emergence of highly pathogenic viruses.
The New York Times, October 19, 2004
To the Editor:
Re ''A War and a Mystery: Confronting Avian Flu'' (Oct. 12): It is important to point out the role of factory farming in the spread of avian influenza.
According to the United Nations Food and Agriculture Organization, the recent outbreak may have been facilitated by huge poultry and pig operations that have sprung up near urban centers in Thailand, Vietnam and China. Avian flu is just one of many animal diseases that have been perpetuated by the squalid conditions on factory farms.
Protecting people from these infections will require more than vaccines. The best defenses are sanitary methods of meat production and clean animals.
The writer is a food and agriculture specialist at the Worldwatch Institute.
And see: AVIAN FLU IN THE UNITED STATES (p.3 of document)
CRS Report for Congress
Updated May 14, 2004
Avian Influenza: Multiple Strains Cause Different Effects Worldwide
Jim Monke, Analyst in Agricultural Policy Resources, Science, and Industry Division


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