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Physiopathology of AEEC infection in weaned pigs

The main objective of WP11 is to develop an infection model for AEEC infection in weaned pigs.

Major finding during the project:

The main objective of WP11 was to develop an infection model for AEEC infection in weaned pigs. In this line one of the deliverable (D23) was to develop an oral AEEC model. The efforts in the first 4 experiments were concentrated on 5-6 weeks old pigs, inoculated once with TSB grown 1010 CFU of the standard Canadian 1390 (O45) AEEC bacteria/pig (all from farm A). The strain was provided by CR6 (Dr. J. Fairbrother). In all subsequent experiments we used 4 weeks old pigs, from farms B, C, and D and inoculated them twice on subsequent two days. All together 159 pigs were used in 10 different experiments.

In 5 of these experiments different predisposing factors were also used (Fumonisin mycotoxin, TGE-virus, F18+ETEC bacteria and high energy or high protein diet). In none of these experiments were repeatable attaching effacing lesion in a consequent number of pigs when ilea and caeca were tested by fluorescence microscopy (IF), and HE sections, at 3-5 days post infection. The predisposing effect of the high protein diet (per se) could reproducibly proven by the concomitant use of ETEC infections. To increase the chance of selecting more virulent strains, there were also 3 pools (with 4 AEEC strains in each) formed using Canadian and Hungarian AEEC strains. Slight AE lesions were only detected in 3 of all together 12 pigs inoculated that way. Based on these results it was concluded that 4 weeks old commercial pigs are basically quite resistant to the oral infection by porcine AEEC strains.

The other objective (D24) within the above main objective was to establish an alternative virulence model in weaned pigs. For these experiments we have used all together 48 pigs that provided over 200 loops to be investigated for the pathogenicity and pathophysiology of the AEEC infection in these animals by IF and HE sections. The ligated intestinal loops were prepared in 4 weeks old pigs (as described in previous report). Loops were inoculated with approx. 5 x108 CFU of AEEC bacteria (or negative control). Pigs were kept in post surgical anaesthesia for 16-18 hours (as described before) We have used the Canadian O45 standard strain (86-1390) in every pig as a positive control strain, together with a standard negative (nonpathogenic) E. coli control strain.

The other test organisms were Hungarian 9 different AEEC strains of different O-types and intimin types (described below, and characterized in detail in WP17). These strains were isolated from the upper intestine or from the rectal flora, and a human O157:H7 EHEC, as well as a bovine AEEC (O126 strain from J.Mainil) for their adhesive pathophysiology. For all these types of AEEC bacteria the ligated loop model in 4-week-old weaned pigs proved to be a useful model and detected differences between EHEC and porcine AEEC, as well as within porcine AEEC. Using this weaned pig loop model we could also complete the objective to describe potential new porcine types of AEEC (D25). These were reported within WP17 and they are the O4/O123, O28, O76, O108, O149, O157, (most of them are of b-type of intimin but some are of g, and some are untypable).
This model was also succesfully used to study intestinal cellular immunity as a response to AEEC infection (Trizol-fixed intestinal wall and Peyer's patch samples were sent to CR7 for amplifications by RT-PCR to measure intestinal cytokine response. See WP13).

DNA vaccine test was performed on 24 pigs for 7 weeks (between 4-11 weeks of age), using the DNA fragments AE, Tir, IL-4 and IL-6 in different combinations as well as controls (See WP13).


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