Frequency of isolation of Salmonella in wetland wild ducks in the province of Entre Ríos, Argentina. Preliminary studies

Argentina is the second largest country in the Neotropics and stands out for its great biodiversity, encompassing five Phyto-geographic Areas and the Tropical, Magellan and Antarctic Oceans (Cabrera, 1976). It has a wide variety of wetlands, such as marine coasts, estuaries, Andean-Patagonian Lakes, pampean lagoons, marshes and estuaries, rivers and streams of plains and saline altitude lakes, distributed in six large areas of wetlands: Cuenca del Plata, Chaco, Pampas, Patagonia, Puna and Patagonian Coastal Zone (Canevari et al., 1998). It also has a wide altitudinal variation and latitudinal distribution of its wetlands, from the Tropic of Capricorn to 55 ° S, and from sea level up to the lagoons of the Puna (4500 m above sea level). These features allow the development of a wide variety of aquatic environments containing about 250 species of resident and migratory water birds, which represents about 25% of the species of waterfowl in the world (Delany & Scott, 2006), and approximately the same percentage of the total number of bird species of Argentina (Narosky & Izurieta, 2003).

Entre Ríos is a province in the northeast of Argentina, located in the so-called Mesopotamian region. With an area of 78,781 square kilometers (territorially spread over 66,976 km2 of land and 11,805 km² of Islands and land prone to flooding); its limits are set by rivers or streams: to the West and South, the Paraná River; to the North the Guayquiraró River, the Mocoretá River and the Basualdo and Tunas streams, and to the East, the Uruguay River. This somehow turns the province into a sort of island. Annual rainfall amounts to an average of 1,150 mm and the climate characteristic are subtropical, without dry season to the North and the pampas temperate to the South (De Chemin et al., 1992). These conditions allow for many aquatic birds to find a proper place for temporary or permanent stay. Furthermore, this environment belongs to wetland corridor of the Argentinian river coast and del Plata Basin, which fulfills irreplaceable functions in flood mitigation, ensuring the quality of water and maintaining the large natural areas used in fish farming (Head of the Cabinet of Ministers, Ministry of environment and sustainable development of Argentina and aquatic resources work group, 2006). On the other hand, Entre Ríos has around 2,490 poultry farms, which correspond to 47 per cent of the farms in the country. Of the total of the existing broilers, the Uruguay River coast stands out, concentrating around 70% of farms in comparison to the Paraná River shore, which concentrates 70% of farms producing eggs for consumption (Schell et al., 2010).

For its part, Salmonella, a bacterium belonging to the Enterobacteriaceae family, is a pathogen of humans and domestic and wild animals (Bell & Kyriakides, 2002). Fecal excretion of animals containing this bacterium contributes to spread Salmonella in the environment. Therefore, carrier animals play an important role in the spread of Salmonellosis. Different serotypes of Salmonella are considered to be pathogenic to commercial birds (Gast, 2003). However, little is known concerning the salmonella that affect wild aquatic birds. Bueno et al. (2010) reported a lower prevalence of Salmonella in a survey of 432 aquatic birds in the West of Entre Ríos and in a  department of Corrientes, performed through individual cloacal swab samples of these birds. This type of birds can spread Salmonella near the local poultry farm areas and have a negative impact on them. Therefore, the aim of this work was to study the frequency of isolation of Salmonella in wild ducks in the wetlands of Entre Ríos, using different types of samples for analysis.

Materials & Methods

Sampling and survey sites
Samples were collected throughout 8 capture actions, from June 2010 to February 2011, in the wetlands of the departments of Gualeguay (Santa Rosa), Paraná (Paraná River, lower arroyo Antonio Thomas and Hernandarias islands), Uruguay (San Ramón Colony, and Gená stream) and Victoria (Rincón del Doll, El Timbocito-laguna La Larga, laguna stream, Grande lagoon-Espinillo stream) in the province of Entre Ríos, Argentina (Table 1). The birds were captured using special networks for this purpose or firearms, during the hunting period or for scientific hunting, and named as proposed by Narosky & Izurieta (2003). 85 wild aquatic birds of the Anatidae family (Anseriforms), belonging to 6 species. The samples collected were: a cloacal swab, portion of liver and cecum of each bird blind; the samples were individually placed in a sterile tube. These samples were transported at 4 ° C in ice boxes from the field to the Laboratory of Bird Health of the EEA INTA Conception del Uruguay (Entre Ríos, Argentina) and maintained at this temperature until processing.

Isolation of Salmonella sp.
At the laboratory, samples were cultured individually, in 5 ml of base tetrathionate broth (Acumedia, United States) with the addition of iodine solution, bright green and novobiocin for 5 days at 35 ± 2 º C. On the 1st and 5th day of incubation, the culture was planted in stretch marks by exhaustion, in Hektoen enteric agar (Britania, Argentina) and Salmonella-shigella Agar (SS, Merck, Germany) and incubated at 35 ± 2 º C for 24 hours. Colonies that were compatible with Salmonella spp. Were characterized for biochemical tests and serum typification, using commercial somatic antisera.

Only one sample of a Cutirí duck cecum (Amazonetta brasiliensis) tested positive for Salmonella sp. Based on its biochemical characteristics, it corresponded to motile Salmonella, isolated only from Hektoen agar, planted on the 1st day of incubation of the tetrathionate broth. This sample was taken in the Gená stream, Pronunciamiento, belonging to the Department of Uruguay, in the East of Entre Ríos, during the month of February, 2011.

Birds sampled in our study are considered by Narosky & Izurieta (2003) as common, or easy to see in Argentina, except for the Necklace duck, of low-frequency or little chance of observing. Thus, all these species can migrate to Argentina from other, especially neighboring countries (Narosky & Izurieta, 2003), increasing the possibility of transmission of Salmonella and other micro-organisms to wild birds' resident in Argentina and to other migratory birds. This can complicate the control of any disease in provinces like Entre Ríos, where wetlands, place of residence to wild birds, are close to poultry farms. On the other hand, the birds sampled in the Uruguay Department are considered common residents and they nest in the area (Raffo et al., 2009). This Department is characterized by a high density of broiler chickens (Schell et al., 2010)

Mitchell & Ridgwell (1971) analyzed 477 samples of wild duck excretes in London natural reserve and 4% of the samples tested positive to Salmonella, mostly Salmonella Typhymurium. On the other hand, Goope et al. (2000) studied the presence of Salmonella spp. in wild birds in captivity in a zoo in Trinidad and found a frequency of isolation of 3% for this bacterium. Furthermore, Waldrup and Kocan (1985) could not isolate Salmonella from swab cultures of wild aquatic birds in Oklahoma, United States, but some of the samples showed a serological reaction to this bacterium. The results of our study were similar to the ones described above and reported by Bueno et al. (2010), showing a very low frequency of isolation of Salmonella SP., which in our case was 0% for cloacal swab and liver samples, and 1.2% for cecum samples.

The low frequency of isolation of Salmonella in wild ducks in the wetlands of Entre Ríos indicates that these birds do not represent a high risk for commercial production of poultry in this province. However, monitoring of this bacterium should continue and expand to other wetlands in the province, in order to be able to locate possible Salmonella outbreaks, which could compromise commercial poultry production in Entre Ríos, the province with the highest density and commercial poultry population in Argentina.

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