Diagnosis of porcine proliferative enteropathy: detection of Lawsonia intracellularis by pathological examinations, polymerase chain reaction and cell culture inoculation.
A total of 21 pigs aged 7-17 weeks with clinical symptoms suggestive for Porcine Proliferative Enteropathy were examined for Lawsonia intracellularis by analysing the following parameters: (i) intestinal gross and histological lesions, (ii) presence of comma-shaped bacteria in enterocytes by Warthin-Starry and a modified Ziehl-Neelsen stain, (iii) PCR amplification of L. intracellularis DNA from intestinal mucosa by using two oligonucleotide primer pairs targeting a 255-bp DNA fragment of the 16S rDNA-gene and a 319-bp DNA fragment of the L. intracellularis chromosome. Specificity of PCR reactions was confirmed by using DNA extracted from the L. intracellularis reference strain N343 (ATCC 55672) as well as by DNA sequence comparisons of PCR amplification products with data bank entries. Intestinal gross lesion indicative for PPE were observed in 20 pigs (95.2%). For all 21 pigs, the L. intracellularis aetiology was confirmed by histological as well as bacterioscopical examinations. Specific PCR amplification products were obtained from 20 pigs (95.2%). Taking PCR positivity as the definite criterion, L. intracellularis was diagnosed in 20 pigs from 11 herds in seven Swiss cantons (Argovia, Berne, Fribourg, Grisons, Lucerne, Schwyz, Thurgovia). To grow L. intracellularisin vitro, the cell culture method of Lawson et al. (J. Clin. Microbiol. 1993: 31, 1136-1142) was adopted. Inocula prepared from heavily infected fresh and frozen ileal mucosa of 15 pigs were cultured in rat enterocytic IEC-18 cells (ATCC CRL 1589). Six cell culture passages of 10 days each were completed. The reference strain N343 was examined for cultivability, accordingly. Except for occasional specific PCR amplifications from cell cultures up to the second passage, any indications for growth of L. intracellularis in IEC-18 cells were not found.
Dittmar
M
M
Institute of Veterinary Bacteriology, University of Zurich, Winterthurerstrasse 270, CH-8057 Zurich, Switzerland.
Hoelzle
L E
LE
Hoelzle
K
K
Sydler
T
T
Corboz
L
L
Miserez
R
R
Wittenbrink
M M
MM
D000818:Animals; D018929:Cell Culture Techniques; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis; D016905:Gram-Negative Bacterial Infections / Q000175:diagnosis / Q000662:veterinary*; D007413:Intestinal Mucosa / Q000382:microbiology*; D020639:Lawsonia Bacteria / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D016133:Polymerase Chain Reaction / Q000592:standards / Q000662:veterinary*; D011237:Predictive Value of Tests; D051381:Rats; D013047:Specific Pathogen-Free Organisms; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis* / Q000473:pathology
D017931:DNA Primers; D004269:DNA, Bacterial