胞内劳森菌及其所致的增殖性肠病

本综述介绍的胞内劳森菌 ,是一种弯曲状杆菌 ,无鞭毛 ,抗酸染色阳性 ,能为Warthin Starry或Lavaditti镀银法着色。该菌存在于病畜肠细胞原生质内 ,至今不能人工培养 ,但可在一些肠细胞培养中生长 ,引起猪增殖性肠病 ,还见于仓鼠、马、鹿、雪貂、大鼠、兔、狐、鸵鸟、鸸鹋等动物。可用病理学、血清学、细菌学、PCR等方法诊断 ,防治可用抗生素 ,如金霉素、tylosin、dimetridiazole、carbadax、timulin等治疗 ,至今无疫苗。     

Lawsonia intracellularis: getting inside the pathogenesis of proliferative enteropathy

Although proliferative enteropathy (PE) has been recognised for several decades, Lawsonia intracellularis, the aetiological agent, was identified formally in only 1995. This organism is both highly fastidious and obligately intracellular bacterium, characteristics which have inevitably restricted investigations in all aspects of its biology. Despite these limitations, advances have been made in characterising and understanding L. intracellularis-host interaction both in vivo and in vitro. Based upon evidence provided by mainly pathological and histological investigations conducted to date, we review salient features of our current understanding of processes involved throughout the course of infection by this unique pathogen.     

Reproductive performance of gilts following an outbreak of acute proliferative enteropathy due to Lawsonia intracellularis

After a sudden outbreak of clinical porcine proliferative enteropathy (PPE) in a Croatian indoor breeding unit, the farm experienced decreased reproductive performance of Lawsonia intracellularis positive gilts. Conception-, farrowing-, and adjusted farrowing rates were lower (P<0.001) in gilts with positive L. intracellularis status. The number of live born and the total born litter size were significantly lower (P<0.001) in L. intracellularis positive gilts compared to seronegative ones. No differences were observed in numbers of stillborn and mummified pigs between the L. intracellularis positive and negative gilts.     

Enzyme-linked immunosorbent assay to detect Lawsonia intracellularis in rabbits with proliferative enteropathy

Lawsonia intracellularis is an obligate intracellular pathogenic bacterium that causes proliferative enteropathy in domestic and experimental animals. Antiserum against synthetic peptides of the Lawsonia surface antigen (LsaA) well recognized L. intracellularis in infected ileum by immunohistochemistry. The synthetic peptides in LsaA showed strong reaction with serum from rabbits infected with L. intracellularis by enzyme-linked immunosorbent assay. These results suggest that ELISA used synthetic peptides in LsaA and anti-LsaA serum might be useful to diagnose for proliferative enteropathy.     

Detection of Lawsonia intracellularis in the tonsils of pigs with proliferative enteropathy

The presence of Lawsonia intracellularis, the obligate intracellular bacterium causing proliferative enteropathy (PE), in the tonsils of pigs as a locus for infection or extraintestinal occurrence of the bacterium was investigated by PCR and immunohistochemistry. Tonsillar occurrence of L. intracellularis could be part of the pathogenesis of PE and an important risk factor in the spread of the disease. L. intracellularis was detected by only PCR in the tonsils of 2/32 pigs without PE at necropsy but with a clinical history of diarrhoea and detection of the bacterium in faeces 1 to 3 weeks prior to necropsy but not in four pigs with moderate PE lesions. However, L. intracellularis was detected in the tonsils of 4/9 pigs with PE complicated with necroses and in 4/4 pigs with proliferative haemorrhagic enteropathy in which L. intracellularis antigen also was demonstrated in tonsillar macrophages and as intact bacteria in the lumen of the crypts. The results show that L. intracellularis is detectable in the tonsils of pigs and that the tonsillar presence of L. intracellularis appears to be correlated to the severity of the intestinal lesions possibly as a result of local retention and not as part of the pathogenesis of PE.     

Proliferative enteropathy involving Lawsonia intracellularis infection in rabbits (Oryctlagus cuniculus)

Five rabbits suffering from diarrhea were diagnosed with proliferative enteropathy (PE). Histopathology revealed a thickened mucosa consisting of hyperplastic intestinal epithelium and infiltration of inflammatory cells mainly consisted of macrophages. In the affected epithelial cytoplasm, numerous curved bacillus-like organisms were observed in the Warthin-Starry silver stain and electron microscopy observation. In polymerase chain reactions, Lawsonia intracellularis-specific DNA fragment were amplified from affected ileal tissue extracted DNA in each case and present 5 cases were confirmed to be L. intracellularis infection. Serum collected from the affected rabbit was immunohistochemically reactive with L. intracellularis in tissue sections from pigs with porcine proliferative enteropathy, as well as with tissue sections from the five affected rabbits. Thus, serum obtained from the affected rabbit may be applicable to immunohistochemical detection for L. intracellularis infection in other species.     

Isolation of Lawsonia intracellularis in Korea and reproduction of proliferative enteropathy in pigs and hamsters

Lawsonia intracellularis (L. intracellularis) was isolated from a Korean pig suffering acute proliferative enteropathy. In vitro culture conditions of L. intracellularis were established in McCoy cells. Pigs and hamsters experimentally infected with the pure culture of L. intracellularis reproduced clinical signs and intestinal lesions of proliferative enteropathy. The presence of L. intracellularis in the intestinal lesions was confirmed by immunohistochemistry with L. intracellularis-specific monoclonal antibodies.     

Porcine proliferative enteropathy in Estonian pig herds: histopathology and detection of Lawsonia intracellularis by PCR

Lawsonia intracellularis is the causative agent of proliferative enteritis in pigs (PPE). This bacterium is difficult to culture from clinical samples and antemortem demonstration is therefore usually performed by PCR on faecal samples. The aim of this study was to elucidate the frequency of L. intracellularis infection in pig herds in Estonia using PCR, histopathological methods and electronmicroscopical studies. The frequency of demonstration of L. intracellularis was highest in 9-12 weeks old pigs (68.1%). It was more frequent in growing pigs with enteritis on small farms where the system of "all-in all-out" was not practiced and where standards of hygiene were poor. Gross and histopathological studies demonstrated that characteristic macroscopic changes associated with PPE were localised to the distal jejunum and ileum.Thickened longitudinal and circumferential folds occurred in the mucosa of the affected regions of the bowel. Samples from pigs aged 4 to 20 weeks exhibited the most intensive inflammatory changes. The distal part of the jejunum, ileum and the upper third of proximal colon and cecum wall were visibly thickened with reduced luminal diameter. Hyperplasia of lymphoid tissue and, in many cases, pseudomembranous or fibrinous inflammation was found. L. intracellularis was detected in 56 young pigs using histopathological methods. Additionally, in 8 of these pigs intracellular bacteria were demonstrated in ilial epithelial cells by transmission electronmicroscopical (TEM) investigation. On the basis of these TEM investigations it was concluded that L. intracellularis causes disturbances of normal growth, differentiation and apoptosis of the epithelial cells of ileum.     

Immunohistochemistry and polymerase chain reaction for the detection of Lawsonia intracellularis in porcine intestinal tissues with proliferative enteropathy

Detection method of Lawsonia intracellularis was studied in formalin-fixed paraffin-embedded intestinal tissues from 5 naturally infected pigs by immunohistochemistry with a monoclonal antibody against outer membrane protein of L. intracellularis. Warthin-Starry silver stain revealed clusters of argyrophilic, slightly curved rod-shaped organisms in the apical cytoplasm of enterocytes. Immunohistochemical staining with a L. intracellularis-specific monoclonal antibody confirmed the presence of the organism in the apical cytoplasm of hyperplastic enterocytes. The presence of L. intracellularis in the ileum of pig with proliferative enteropathy was confirmed by polymerase chain reaction (PCR) further on the basis of amplification of 319 base pair products specific for porcine L. intracellularis chromosomal DNA. Immunohistochemistry and PCR may be a complementary method to confirm the diagnosis of L. intracellularis infection in pigs.     

In vitro cultivation and partial characterization of Lawsonia intracellularis from a Japanese field case of porcine proliferative enteropathy

Lawsonia intracellularis isolated from a Japanese field case of porcine proliferative enteropathy (PPE) was cultivated and partially characterized. The bacterial cells isolated from the intestinal mucosa of a pig suffering from the acute form of PPE were used to inoculate rat small intestine cells (IEC-18) and human epithelial cells (HEp-2). Infected foci, which were stained with L. intracellularis-specific rabbit antiserum, were observed in the cell culture at 5 days post inoculation. The DNA sequence of several genes in the Japanese isolate had high similarity with those of the L. intracellularis type strain, suggesting the genetically close relationship of the two strains. This is the first report describing the cultivation and partial characterization of L. intracellularis originated in Japan.     

Lawsonia intracellularis proliferative enteropathy in a weanling foal,with a tentative histological diagnosis of lymphocytic plasmacytic enteritis

This Case Report describes a weanling filly with protein-losing enteropathy associated with Lawsonia intracellularis infection. This was diagnosed on the basis of a significant antibody response and a positive faecal PCR result. The histopathological lesion observed in proliferative enteropathy is mucosal hyperplasia, commonly affecting the ileum and colon in foals. Duodenal biopsies obtained from this filly revealed a lymphocytic plasmacytic infiltrate. The filly recovered completely following treatment with erythromycin and no additional medication was administered to treat the lymphocytic plasmacytic infiltrate. This Case Report suggests that lymphocytic plasmacytic infiltrates observed on duodenal biopsies may represent a nonspecific intestinal immune response.     

Comparison of intestinal mucosa homogenate and pure culture of the homologous Lawsonia intracellularis isolate in reproducing proliferative enteropathy in swine

Little information is available on reproduction of proliferative enteropathy (PE) using a virulent pure culture of Lawsonia intracellularis. Reproduction of the disease using PE-diseased mucosa homogenates, however, is well-characterized. The aims of this study were to evaluate and compare clinical signs, growth performance and the severity of lesions in pigs inoculated with intestinal mucosa homogenate or pure culture of the homologous L. intracellularis isolate. Five-week-old pigs were inoculated with pure culture of L. intracellularis (isolate PHE/MN1-00; n=10), PE-diseased mucosa (n=10), or control media (n=4). The L. intracellularis isolate PHE/MN1-00 used in the pure culture inoculum was extracted from a fragment of the same intestine used to prepare the mucosa homogenate. Clinical signs and growth performance were evaluated throughout the study. Fecal shedding was evaluated in all animals weekly during the experiment. All animals were euthanized 22 days post-inoculation, the intestines were examined grossly and histologically. Results showed that both the infection procedures reproduced clinical disease, macroscopic and histologic lesions typical of PE. Fecal shedding was detected in animals in both challenge groups. In conclusion, the L. intracellularis isolate PHE/MN1-00 reproduces typical clinical signs and lesions of PE similar to the homologous infection with an intestinal mucosa homogenate.     

Effects of oral vaccination against Lawsonia intracellularis on growing-finishing pig's performance in a pig production unit with endemic porcine proliferative enteropathy (PPE)

The aim of the present study was to evaluate the effect of oral vaccination against Lawsonia intracellularis (LI) on growing-finishing pig's performance. In a large Hungarian growing-finishing pig production unit, pigs with positive LI status were randomly divided into 2 groups and treated as follows: Group one: growing pigs (n = 4112) were LI vaccinated with an avirulent oral live vaccine (Enterisol Ileitis Boehringer Ingelheim Vetmedica, Inc., St. Joseph, USA). Group two: growing pigs (n = 4188 pigs) have not received LI vaccination. Culling and mortality rates, reasons for culling or mortality, and average daily weight gain were evaluated. Porcine proliferative enteropathy (PPE) caused culling and mortality rates were lower (0.2 % vs. 14.9 %, P < 0.001), and vaccinated pigs had lower none-PPE caused culling and mortality rates compared with the non-vaccinated ones (1.4 vs. 2.6 %, P > 0.05). While systemic infections and social stress or cannibalism related culling or mortality were significantly (P < 0.05) lower in vaccinated than in non-vaccinated pigs, reasons for culling or mortality due to non-LI caused diseases were non-significantly different between the vaccinated and non-vaccinated pigs. Average daily weight gain was higher (P < 0.05) in the LI vaccinated group of animals compared with the non-vaccinated ones (780 +/- 45 g vs. 660 +/- 71 g). The present results indicate that that LI vaccination does not only prevent PPE, but might result in more resistance and tolerance against other infectious and management caused losses.     

Cytologic diagnosis of Lawsonia intracellularis proliferative ileitis in a Japanese snow macaque (Macaca fuscata)

Diffuse ileal thickening and ileocecocolic lymphadenomegaly were observed during exploratory laparotomy in a 2-year-old male Japanese snow macaque (Macaca fuscata) that had flu-like signs and diarrhea. Cytologic examination of ileal biopsy imprints revealed many mature, mildly karyolytic neutrophils and fewer well-differentiated lymphocytes, eosinophils, macrophages, and plasma cells in a background containing amorphous, necrotic material. Tightly cohesive sheets of moderately pleomorphic epithelial cells also were seen. The cytologic diagnosis was chronic, active, mixed inflammation with atypical epithelial cells and necrosis. Histologically, the mucosal and crypt epithelium was moderately hyperplastic with a loss of goblet cells, increased mitoses, and frequent crypt abscesses. Within the lamina propria and extending into the submucosa was a marked neutrophilic infiltrate, with low numbers of lymphocytes, histiocytes, plasma cells, and eosinophils. The histologic diagnosis was chronic, diffuse, marked suppurative and lymphocytic ileitis. Warthin-Starry silver staining of the ileal biopsy and imprint specimens demonstrated numerous pleomorphic, curved bacilli consistent with Lawsonia intracellularis. Polymerase chain reaction (PCR) and immunohistochemistry confirmed the identity of the infectious agent. L intracellularis infection may be underdiagnosed because silver stain is required to visualize the organism with light microscopy and because the pathognomonic crypt hyperplasia may be complicated by secondary pathologic changes. Application of silver stain to cytologic specimens should be considered when distal intestinal lesions associated with hyperplastic epithelium, with or without inflammation, hemorrhage, or necrosis, are identified in animals with clinical signs of enteritis, especially in frequently affected species or in stressed or young animals.     

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.     

The effect of outdoor production on the seroprevalence of Lawsonia intracellularis in growing-finishing pigs in a large pig production unit infected with endemic porcine proliferative enteropathy

The study was performed in a large Croatian production unit from May 2000 till June 2002. Blood samples form late-pregnant gilts were tested by indirect immunofluorescence antibody (IFA) serum assay for Lawsonia intracellularis. The offspring of 301 positively tested gilts were dislocated after the nursery phase either to indoor or outdoor growing-finishing facilities. Ten percent of these animals (142 indoor, 143 outdoor raised pigs) were tested at 2, 6, 10, 14, 18, 22 and 26 weeks of age for seroprevalence of Lawsonia intracellularis. All offspring of IFA positive gilts were seronegative at 2 and 6 weeks of age. At 10 weeks of age 71.1% (101 animals) of indoor and 32.8% (47 animals) of outdoor pigs were tested positive (P < 0.05). While at 14 weeks of age 71.1% of indoor raised pigs showed seropositivity, the seropositivity declined in outdoor units to 7.6% (P < 0.01). At weeks 18 (52.1%), 22 (47.8%) and 26 (21.7%) indoor raised pigs still showed marked seropositivity and but their outdoor raised counterparts returned to seronegativity.