Prevalence of Escherichia coli O157 in lambs at slaughter in Rome, Central Italy

Antonio
1* Battisti ,

Sarah Alessia Alessandra Di Stefano 1Istituto Zooprofilattico Sperimentale delle Regioni Lazio e Toscana 2Istituto Superiore di Sanit, Dipartimento di Sanit Alimentare e Animale
Introduction Materials and Methods

1 Lovari ,

1 Franco ,

1 Egidio ,

2 Morabito

Escherichia coli O157 is a human pathogen whose infection can cause life-threatening diseases such as haemorrhagic colitis and haemolytic uremic syndrome. Outbreaks involving large numbers of persons have often been linked to the consumption of undercooked minced beef or cross-contaminated cooked meats. Many studies have indicated that cattle represent the main reservoir of VTEC O157 (Armstrong et al. 1996; Hancock et al. 1998). Other ruminants and domestic animals harbour these bacteria, in particular VTEC O157 has been detected both in meat and milk from sheep and goat and sheep is now considered as a possible reservoir for human infection. In Italy 70-80 thousand tons of sheep and goat meat is consumed every year, of which 55-65% comes from national flocks (U.N.A., 2004). Lamb is one of the most popular types of sheep meat and can be divided in two categories: suckling lambs, 20-30 days old animals weighing between 7 and 10 kg and exclusively milk-fed, and weaned lambs, usually less than 6 months of age and with a polygastric digestive physiology. The aim of this study was to estimate the prevalence of faecal carriage of E. coli O157 by lambs intended for slaughter in the principal public slaughterhouse of Rome, Italy, and locally distributed for consumption.

Between March and December 2002, a total of 643 faecal samples were collected at the major abattoir of Rome, Lazio region, Central Italy. Two hundred sixty-five samples were collected from suckling lambs and 378 were from weaned lambs. For prevalence estimates, a simple random sampling technique was used. A desired sample size was calculated based on a four percent expected prevalence, a 95% level of confidence and a 1.5% desired absolute precision, stratified by age class (suckling or weaned lambs) and origin (local or foreign origin). Samples collection of weaned lambs was equally distributed in the four seasons, with samples collected in March, July, October and December, while samples from suckling lambs were obtained in March and December. Weaned lambs were from different origin: 252 animals (67%) came from Eastern Europe (Romania, Slovakia, Hungary and Poland) and 126 (33%) were from farms located in central Italy. Suckling lambs were exclusively of local origin (Central Italy). The abattoir had dimensions ranging from 120.000 to 150.000 lambs slaughtered per year. From each animal faeces were collected from the rectum immediately after slaughter. All samples were examined for the presence of VTEC O157 using an immunomagnetic separation (IMS) technique. Specimens were analysed by 1 gr of faeces each and enriched in 45 ml of BPW (buffered peptone water), prewarmed at 37C and incubated at 42 C for 24h. One plate Sorbitol MacConkey Agar supplemented with 0.05 mg/l cefixime and 2.5 mg/l tellurite (CT-SMAC) and one plate of CHROMagar O:157 were seeded with two 50 ml-aliquots of IMS-enriched material. The plates were incubated at 37C for 16-20 h. Polymerase chain reaction (PCR) was used to assess the presence of virulence genes. Primer pairs KS7-KS8 and GK3-GK4 were used for for vt1 and vt2, respectively. The intimin-coding eae gene was detected according to Oswald et al. (2000).

Results E. coli O157 agglutinating strains were isolated from the intestinal contents of five out of 643 animals examined with an overall prevalence of 0.8%(95% C. I. 0.3-1.9%). The characteristics and of the isolates are reported in Table 1. All the positive animals came from Eastern Europe. When only the VT-producing strains (virulent strains) are considered, the prevalence decreases to 0.2% (95% C. I. 0.0-1.0). E. coli O157 was isolated only from weaned lambs (Table 2).
Table 1. Origin and characteristics of Escherichia coli O157 isolates from weaned lambs Strain Lambs origin eae Vt E-hly Motility
10140 10550/16 10550/1 15830 18526 Slovakia Slovakia Slovakia Hungary Hungary +ve +ve -ve -ve -ve +ve -ve -ve -ve -ve +ve -ve -ve -ve -ve -ve +ve +ve +ve +ve

Table 2. Escherichia coli O:157 estimated prevalence in lambs from the major slaughterhouse in Rome

Suckling lambs (n=265) Weaned lambs (n=378)

Discussion

Overall Prevalence (95% C. I.) 0 (Upper limit 1.4%) 1.3% (0.5-3.2%)

Prevalence VT +ve, eae +ve 0 (Upper limit 1.4%) 0.3% (0.0-1.7%)

Prevalence eae +ve, VT ve 0 (Upper limit 1.4%) 0.3% (0.0-1.7%)

Prevalence eae-ve, VT -ve 0 (Upper limit 1.4%)

0.8 (0.2-2.5%)

The presence of E. coli O157 isolates lacking of all or part of the virulence gene set (carried by Mobile Genetic Elements, MGE), could indicate that in lamb gut they are subjected to a negative selection that forces them to lose these MGE. On the other hand, none of the E. coli O157 isolates possessed the H7 antigen, that identify the most virulent clone of EHEC O157. Therefore, they could be part of a more unstable clone which could lose spontaneously the MGE carrying the virulence genes. In conclusion, the estimated risk of E. coli VTEC O157 carriage in the lambs sampled in this study can be considered low. Therefore, the risk of human exposure to VTEC O157 due to the consumption of the lamb meat produced in the slaughterhouse in Rome, should be reasonably low, particularly when suckling lamb meat is considered. Moreover, the observations on the genetic instability of the ovine E. coli O157 may explain why the consumption of sheep meat has never been associated with E. coli O157 infections in humans and raise a question on the real impact of the ovine species in the spreading of E. coli O157 infections. Further investigations are needed to clarify this matter.
References
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