Molecular characterization of endemic salmonella infections in cattle

Salmonellosis is one of the most significant zoonoses worldwide and also in Finland. The major serovars causing infections in humans are Salmonella Enteritidis and Salmonella Typhimurium. Salmonella Typhimurium definitive phage type (DT) 1 and Salmonella Infantis are considered endemic in Finland. These serovars have frequently caused outbreaks among humans, the source of which is often detected. For the sporadic cases they usually remain unknown. Salmonella Agona was not frequently encountered in Finland until a small outbreak among cattle farms occurred in 1994-1995. S. Agona became the third most common Salmonella serovar in cattle in Finland in 1995. The two more common serovars were S. Infantis and S. Typhimurium DT1. Bacterial typing methods are used for outbreak investigations and for surveillance, where the data can be used for risk assessment calculations in addition to the future prevention of outbreaks. In particular the identification of factors that contribute to the persistence and spread of infection in endemic situations, estimations of the effect of animal reservoirs on human cases, and the identification of other risk factors for human infections are among the important reasons for typing. Salmonella Infantis became more common in cattle in the 1980s, after it spread in the broiler chicken production in Finland in 1971. Subsequently it caused outbreaks among broilers and humans in the 1970s and 1980s. In the 1990s, S. Infantis became the predominant serovar among cattle in Finland. In 1995, a feedborne outbreak of S. Infantis in cattle occurred. We were able to identify the feedstuff-related genotype by XbaI-PFGE methodology. It belonged to the major endemic type, pf1, but differed from it by having a plasmid visible as an intensive band of 60 kb in XbaI-PFGE (plasmid subtype pf1/39). Farms infected with the feedstuff-related genotype pf1/39 or the related genotypes pf1/43, pf1/44, pf1/45, or pf1/46 containing the same 60 kb plasmid were also identified. The stability of the feedstuff-related genotype was followed on selected farms. The plasmid was stable on the farms during the follow-up period. The feedstuff-related genotype did not persist in the cattle population. Moreover, there was a general decline in bovine salmonella infections from 1997 onwards. The genetic diversity of the S. Infantis isolates taken from Finnish cattle was also assessed: the S. Infantis infection in cattle was highly clonal as 99 per cent of the isolates had XbaI-PFGE profiles clonally related to each other. The major genotype pf1 was predominant both at the starting year of our analysis in 1985 and as the infection seemed to fade out in 2003. Traditionally, only one isolate per farm is stored in the national collection. However, an infection may have existed subclinically for a long time in a herd until the first Salmonella isolates were obtained. In our analysis of successive isolates from the same herds, we frequently detected minor changes in banding patterns during long-lasting infections in individual herds. The sampling and testing of several isolates from a herd in outbreak investigations is therefore advisable. There is a trend towards less genetic diversity of the S. Infantis infection among domestic isolates from humans and poultry. Up to eight different ribo/IS200-types were detected in the 1980s isolates, whereas in the 1990s only two different ribo/IS200-types (1A, 1B) were seen. In cattle, 89 per cent of the analysed isolates possessed the ribo/IS200-type 1A, although four different ribo/IS200-types were recorded in the 1980s isolates, and two ribo/IS200-types in the 2000s isolates. The ribo/IS200-types and the most common XbaI-PFGE profiles determined amongst the analysed cattle isolates could also be detected among domestic isolates from poultry and humans. After the outbreak in cattle in the years 1994-1995, isolates of S. Agona taken from the 1984 to 1999 period were characterized by PFGE using XbaI, BlnI, SpeI, and NotI enzymes. Two outbreakrelated genotypes, which were not detected in the earlier isolates of S. Agona, were identified. Another small possibly genetically related outbreak among cattle farms occurred in 1997. In 1999, a large outbreak of S. Agona of domestic origin involving more than 50 human cases occurred. Despite epidemiological investigations carried out by the local authorities, the source of the outbreak remained unknown. Based on our typing data, this outbreak was unrelated to the cattle farm outbreaks, though it did occur in the same region of Finland. The outbreak profile for the 1999 outbreak could not be found in any of the other isolates. As no recent foreign isolates were available, a foreign source of the human infection cannot be disregarded. Salmonella Typhimurium DT1 has become the most common S. Typhimurium phage type among cattle farms in Finland, and has been detected annually since 1980. Domestic and foreign isolates of S. Typhimurium DT1 from 1981 to 1999 were characterised. Furthermore, two clusters formed by the effects of the combination of the XbaI-, BlnI-, and SpeI-PFGE profiles, IS200-profiles and possession of the serovar-specific virulence plasmid were analysed. The major cluster had no virulence plasmid and included the most common XbaI-PFGE profile 10 and IS200-profile D,typical of our endemic infection. The results of XbaI-, BlnI-, and SpeI-PFGE gave 54 different combination profiles, which can be applied in the analysis of outbreaks. In contrast, molecular subtyping by XbaI-PFGE alone is not discriminatory enough in analysing our endemic infection. The source of sporadic human infections is unknown in most cases, and molecular typing did not reveal any clear infection source. The most common XbaI-PFGE profile 10 was also seen in hedgehogs and wild birds. They might act as important reservoirs, maintaining a minimum baseline level of S. Typhimurium DT1 in the environment and consequently be possible sources of human infections.