The high homology of RNA sequences of the HA genes derived from viruses that caused outbreaks at 3 different holdings in the German Federal State of Brandenburg and virus isolated from deep-frozen duck carcasses (A/duck meat/Bavaria/2048/2007) clearly points to a close epidemiologic link between these outbreaks. According to theoretical consideration regarding the mutation rate of influenza A viruses (14) and to practical experiences during outbreaks (4), if HPAIV had gone through a transmission chain consisting of several hosts, more extensive sequence differences would likely have resulted. This likelihood is further emphasized by the comparison of 3 subtype H5N1 virus isolates derived from a single barn of farm A (Figure 2); these viruses exhibit greater variability among each other than do viruses from the 3 different affected backyard holdings in Brandenburg. Also, distinct sequences of viruses were detected from poultry in Poland, where outbreaks occurred at the same time as the Brandenburg cases, which renders an incursion from this source highly unlikely. Among the Brandenburg cases, no epidemiologic links could be detected except that, as proven in 2 cases and assumed to have occurred in the third case, backyard chickens had access to uncooked offal from duck meat purchased separately in different supermarkets of the same national chain.

Oral uptake of virus is an efficient way of transmitting HPAIV among poultry and mammals (15,16). Infectious virus in titers of up to 10^7.2 50% egg infectious doses per gram in muscles of infected chickens, ducks, and quails has been repeatedly demonstrated (17,18). Although feeding of poultry offal to poultry or livestock is legally prohibited in Germany, unintended access of backyard poultry to poultry meat and organs is sometimes possible. If such offal is contaminated with HPAIV, transmission becomes possible, and isolated outbreaks like those reported from Brandenburg may ensue. However, this circumstance would require the presence of HPAIV in meat destined for human consumption. This possibility had previously been estimated to be low (19).

Unfortunately, no experimental evidence could be produced that unequivocally links the Bavarian farm B and Brandenburg backyard outbreaks because no material was left for virologic examination from the suspected deep-frozen duck carcasses. Therefore, we tried to collect further epidemiologic data by fully retracing the origin of the frozen ducks purchased by the owners of the backyard chickens in Brandenburg. This attempt included authorities in the involved Federal States and the management of the supermarket chain that had sold the duck meat.

The duck meat sold in autumn 2007 by the supermarket chain in Brandenburg and in the neighboring Federal States of Berlin and Mecklenburg–Western Pomerania had been purchased from a slaughterhouse in the Federal State of Lower Saxony (lots 724, 725; Figure 3). Direct links between Bavarian farm B and the slaughterhouse in Lower Saxony existed because slaughtering lots 724 and 725 included ducks from 3 fattening flocks from farm B. These ducks were slaughtered on June 14, 19, and 22, 2007, in the abattoir in Lower Saxony, because the most frequently used abattoir in Bavaria at farm A was closed for holidays (Figure 3). Among the 3 flocks was the one from barn B/3–5, which showed a suspiciously elevated cumulative proportion of deaths of 12.6% (Table; Figure 1, panel A).

This circumstantial evidence points toward a transient and limited incursion of HPAIV (H5N1) into duck meat destined for human consumption. Therefore, enhanced virologic screening of fattening ducks has been initiated in Germany. Also, reporting obligations related to deaths in duck flocks were adopted. Since the end of the Brandenburg outbreaks, a single, unrelated recurrence of HPAIV (H5N1) in Germany was detected in poultry in 2008, but none has been detected in wild birds. To date, no clinical cases of human infection with subtype H5N1 have been reported in Germany.

In avian hosts, the clinical picture of an HPAIV infection depends, among other factors, on the species affected (20). In particular, domestic waterfowl showed substantial variations in clinical features resulting from infection with strains of HPAIV (H5N1) of Asian origin (21). Factors influencing the clinical course relate to species, age of animals, and the virus strain (22,23). The cited experimental data as well as reported evidence from the field (24) show that an introduction and subsequent spread of HPAIV (H5N1) in duck flocks is likely clinically silent. Also in the cases reported here, hardly any clinical symptoms suggestive of an HPAIV infection, in particular, neurologic manifestations, were evident. Increased proportion of deaths, as seen in some of the described duck-fattening flocks, might have been fueled by bacterial co-infections, e.g., by those caused by Riemerella spp. Through silently but productively infected ducks, an endemic status of HPAIV (H5N1) infection can be established and perpetuated (25). Strains isolated from such endemic infections induce no clinical symptoms in ducks but retain high pathogenicity for chickens and turkeys. No sign of even widespread infection would be clinically apparent until the virus has become more established in flocks of highly vulnerable (gallinaceous) species (26). Such mechanisms are obviously not restricted to subtropical Southeast Asia.

In conclusion, our data show that incursions of recent strains of descendants of Qinghai lineage HPAIV (H5N1) strains into industrial duck-fattening holdings in Europe may be clinically silent, even in young ducklings. If daily mortality rates remain low, an increase in cumulative mortality may still be evident and should prompt specific virologic investigations. Undetected HPAIV (H5N1) infections of domestic waterfowl destined for human consumption raises the risk for human infection when infected birds or contaminated meat products are handled. Thus, measures must be strengthened that can prevent this zooanthroponotic virus from entering the food chain through contaminated duck meat products and spreading further. Intensified monitoring of duck herds for HPAIV infection that does not rely on syndromic surveillance would be required first. In the outbreaks reported here, the power of serologic assays to detect virus incursions is notable, compared with results by rRT-PCR. Species-independent cEIA assays detecting antibodies specific for the nucleocapsid protein of influenza A viruses were more sensitive than HI assays that used 2 different H5 antigens. Despite the intrinsically higher sensitivity of most ELISAs, the discrepancy in this study might have been aggravated by not using antigen of the outbreak viruses in HI assays. Nevertheless, cEIAs would be suitable for high-throughput analysis in extensive monitoring programs while HI techniques would not.