Review of methods for the detection of Lawsonia intracellularis infection in pigs.

Lawsonia intracellularis is an obligate intracellular bacterium associated with enteric disease in pigs. Clinical signs include weight loss, diarrhea, and, in some cases, sudden death. The hallmark lesion is the thickening of the intestinal mucosa caused by increased epithelial cell replication, known as proliferative enteropathy. The immune response to L. intracellularis is not well defined, and detection of the infection, especially in the early stages, is still a significant challenge. We review here the main approaches used to identify this important but poorly understood pathogen. Detection of L. intracellularis infection as the cause of clinical disease is confounded by the high prevalence of the pathogen in many countries and that several other pathogens can produce similar clinical signs. A single L. intracellularis-specific ELISA and several amplification assays are available commercially to aid detection and surveillance, although histopathology remains the primary way to reach a conclusive diagnosis. There are major gaps in our understanding of L. intracellularis pathogenesis, especially how the host responds to infection and the factors that drive infection toward different clinical outcomes. Knowledge of pathogenesis will increase the predictive value of antemortem tests to guide appropriate interventions, including identification and treatment of subclinically affected pigs in the early stages of disease, given that this important manifestation reduces pig productivity and contributes to the economic burden of L. intracellularis worldwide.

Oral fluid for detection of exposure to Lawsonia intracellularis in naturally infected pigs.

To demonstrate the utility of oral fluid (OF) for indirect diagnostic detection of Lawsonia intracellularis (Li), 15 pig farms were studied. Serum and fecal samples were collected from 20 animals from five different age groups on each farm. OF samples were collected from animals in two pens of the same age groups. Serum and OF samples were analyzed in an immunoperoxidase in monolayer assay (IPMA) for the detection of anti-Li immunoglobulin G (IgG) and A (IgA). Compatible results were found between PCR and IgG in OF in four of the five ages evaluated. Simultaneous detection of IgG in serum and OF was mainly observed on farms showing clinical signs suggestive of porcine proliferative enteropathy (PPE). These findings demonstrate the potential usefulness of OF in detecting anti-Li antibodies as a diagnostic tool that can be used to monitor PPE in herds with clinical signs compatible with the disease.

Lawsonia intracellularis associated equine proliferative enteropathy in Danish weanling foals.

BACKGROUND: Lawsonia intracellularis, an obligate intracellular bacterium, causes equine proliferative enteropathy, mainly in horses around weaning. This disease is rarely reported in the Scandinavian countries. RESULTS: Five cases of equine proliferative enteropathy were diagnosed between 2008-2016 at the University of Copenhagen Large Animal Teaching Hospital. Cases were Danish Warmbloods and a Friesian horse, aged 6-7 months, presenting with typical clinical signs of lethargy, poor body condition, pyrexia and diarrhea. Clinical pathology was consistent with previous reports of severe hypoalbuminemia and leukocytosis. Diagnosis was confirmed by fecal polymerase chain reaction, serum immunomonolayer peroxidase assay and/or immunofluorescence and fluorescence in situ hybridization performed on formalin-fixed ileum samples. Concurrent intestinal parasitism was present in all five cases. Treatment consisted of antimicrobial therapy, anti-inflammatories, intravenous crystalloids and plasma. Three foals were euthanised due to deterioration and poor response to treatment, one with complications of septic arthritis and Strongylus vulgaris associated intestinal infarct. The other two foals survived and were reported by the owners to be healthy on long-term follow-up. CONCLUSIONS: Equine proliferative enteropathy is a disease to consider in young horses presenting with diarrhea and hypoproteinemia in Denmark.

Tylosin injectable for the treatment of porcine proliferative enteropathy in experimentally inoculated pigs / Tilosina injetável no tratamento da enteropatia proliferativa suína em leitões experimentalmente inoculados

Porcine proliferative enteropathy (PPE) is one of the most common enteric diseases in growing and finishing pigs. PPE is characterized by reduced growth performance, accompanied or not by diarrhea. PPE is highly prevalent in several countries of the Americas, Europe and Asia, causing high economic losses in swine herds. The most common form of PPE control in pigs is antibiotic therapy. The objective of this study was to evaluate a new product based on tylosin injectable (Eurofarma Laboratórios S.A.) to control PPE in experimentally inoculated animals. Sixty 5-week-old pigs with mean weight of 9.5kg were divided into two experimental groups of 30 animals: medication and control. All pigs were challenged with Lawsonia intracellularis, the etiologic agent of PPE, on day zero. Fecal score, body condition score, and behavior were daily evaluated. Pigs were weighted on days -2, 13 and 21 of the experiment. Pigs in the Medication Group received tylosin injectable 13 days after inoculation, in three doses with a 12-hour interval between them. Pigs in the Control Group received injectable saline solution following the same protocol. In the Control Group, 23pigs presented with diarrhea before day 13. After day 13, the number of diarrheic animals in this group was reduced to 17. In the Medication Group, 26 pigs presented with diarrhea in the initial period, and in the period after medication, only 11 animals had diarrhea. The score of gross intestinal PPE lesions in the Medication Group was lower than that in the Control Group (p=0.031). The Medication Group also showed lower score for Lawsonia intracellularis antigen-labeling by immunohistochemistry compared with that of the Control Group (p=0.032), showing lower level of infection. These results demonstrate that tylosin injectable (Eurofarma Laboratórios S.A.), administrated in three doses (1mL/20kg) every 12 hours, was effective for the control of PPE in experimentally inoculated pigs.(AU)

Enteropatia proliferativa suína (EPS), causada pela bactéria Lawsonia intracellularis, é uma das doenças entéricas mais comuns em suínos de recria e terminação. A EPS caracteriza-se por redução no desempenho dos animais, acompanhada ou não por diarreia. É uma doença altamente prevalente em diversos países da América, Europa e Ásia, provocando elevados prejuízos econômicos nos rebanhos suínos. A forma de controle da EPS mais adotada em rebanhos suínos é a antibioticoterapia. O objetivo deste estudo foi avaliar um novo produto à base de tilosina (Eurofarma Laboratórios S.A.) na forma injetável para controlar a EPS em animais experimentalmente inoculados. Foram utilizados 60 leitões, de cinco semanas de idade, com peso médio de 9,5kg, divididos em dois grupos experimentais (n=30), medicados e não medicados. Todos os leitões foram desafiados com Lawsonia intracellularis no dia zero. Avaliações clínicas de escore fecal, escore corporal e comportamento foram realizadas diariamente além da pesagem individual dos animais realizada nos dias -2, 13 e 21 do experimento. Os leitões do grupo medicado receberam tilosina injetável 13 dias após a inoculação em três doses com intervalo de 12 horas cada. Já os leitões do grupo não medicado receberam solução salina injetável com o mesmo protocolo. O grupo não medicado apresentou 23 animais com diarreia antes do dia 13 e 17 após este período. No grupo medicado, 26 animais apresentaram diarreia previamente à medicação e apenas 11 após a medicação a partir do dia 13. Os leitões medicados apresentaram extensão de lesão macroscópica, caracterizada por espessamento de mucosa intestinal, menor em comparação com o grupo não medicado (p=0,031). A imunomarcação para Lawsonia intracellularis foi menor no grupo medicado (p<0,032), mostrando redução no grau de infecção por L. intracellularis nos animais medicados. Estes resultados demonstram que a tilosina injetável (Eurofarma Laboratórios S.A.) (1mL/20kg) em três doses, a cada 12 horas, foi eficaz no tratamento da enteropatia proliferativa suína em animais experimentalmente inoculados.(AU)

Isolation and in vitro antimicrobial susceptibility of porcine Lawsonia intracellularis from Brazil and Thailand.

BACKGROUND: Lawsonia intracellularis is an obligate intracellular bacterium which cannot be cultured by conventional bacteriological methods. Furthermore, L. intracellularis needs enriched medium and a unique atmosphere for isolation, cultivation and propagation. Because of this,there are only a few isolates of L. intracellularis available and few studies in vitro demonstrating the susceptibility of this bacterium to antimicrobial agents. The objectives of this study were to isolate South American and Southeast Asia strains of L.intracellularis and to determine the in vitro antimicrobial activity against these isolates. Tested antimicrobials included: chlortetracycline, lincomycin, tiamulin, tylosin and valnemulin(against both Brazilian and Thailand strains) and additionally, amoxicillin, zinc-bacitracin, carbadox, enrofloxacin, gentamicin, sulfamethazine, trimethoprim, spectinomycin and a combination (1:1) of spectinomycin and lincomycin were also tested against the Thai isolates. The minimum inhibitory concentration (MIC) was determined by the antimicrobial activity that inhibited 99% of L. intracellularis growth in a cell culture as compared to the control (antimicrobial-free). RESULTS: Two strains from Brazil and three strains from Thailand were successfully isolated and established in cell culture. Each antimicrobial was evaluated for intracellular and extracellular activity. Pleuromutilin group (valnemulin and tiamulin) and carbadox were the most active against L. intracellularis strains tested. Tylosin showed intermediate activity, chlortetracycline had variable results between low and intermediate activity, as well as spectinomycin, spectinomycin and lincomycin, amoxicillin, sulfamethazine and enrofloxacin. L. intracellularis was resistant to lincomycin, gentamicin, trimethoprim, colistin and bacitracin in in vitro conditions. CONCLUSIONS: This is the first report of isolation of L. intracellularis strains from South America and Southeast Asia and characterization of the antimicrobial susceptibility patterns of these new strains.

A real-time loop-mediated isothermal amplification method for rapid detection of Lawsonia intracellularis in porcine fecal samples.

Porcine proliferative enteritis is a common diarrheal disease characterized by thickening of the intestinal mucosa in swine due to enterocyte proliferation, which is caused by Lawsonia intracellularis. In this study, a real-time loop-mediated isothermal amplification (LAMP) assay was developed to detect L. intracellularis based on the conserved region of the 16S ribosomal RNA gene. The optimal reaction conditions of the real-time LAMP was 65 °C for 60 min. The LAMP products could be detected by both real-time turbidity and direct visual inspection. The assay was specific for L. intracellularis, as no cross-reaction was observed with other pathogens. The detection limit of the real-time LAMP assay was 1.4 × 10-1pg of L. intracellularis DNA, which was the same as that of real-time PCR and approximately 100 times more sensitive than that of conventional PCR. Of the 136 clinical samples, L. intracellularis DNA was identified in 60 samples by real-time LAMP, which was the same as real-time PCR and higher than conventional PCR (36.8%, 50/136). The specific, sensitive and rapid real-time LAMP assay developed in this study could be a useful alternative tool in point-of-care (POC) diagnosis of L. intracellularis infection.

Lawsonia intracellularis in the feces of wild rodents and stray cats captured around equine farms.

BACKGROUND: Proliferative enteropathy is a global enteric disease of particular importance in pigs. The causative bacterium, Lawsonia intracellularis, has a wide range of susceptible host species. Recently, L. intracellularis has been recognized as an etiologic agent of an emerging enteric disease in foals called equine proliferative enteropathy (EPE). The presence of L. intracellularis in nonruminant wildlife has raised questions regarding the role of these species in EPE transmission. RESULTS: This study investigated exposure to L. intracellularis in wild rodents and feral cats from eight farms with confirmed EPE. Serum (42) and fecal (40) samples from resident foals and fecal samples (131), intestinal mucosa tissues (14), and mesenteric lymph nodes (14) from wild and feral animals were collected for the evaluation of the farm status and the molecular detection of L. intracellularis following the diagnosis of EPE in index cases. Fresh feces from wild rodents and feral cats were collected from the ground while walking the premises or after trapping the animals using live traps. A total of 3 brown rats, 7 house mice, 1 striped field mouse, 2 grey red-backed voles, and 3 feral cats showed evidence of prior exposure to L. intracellularis. CONCLUSIONS: Our data add to increasing evidence demonstrating the potential for L. intracellularis transmission and infection in wild rodents and feral cats and provide possible evidence of interspecies transmission. The exposure of wild rodents and feral cats provides potential evidence for the spillover of L. intracellularis to wildlife species and raises the question of spillback to horses. Additionally, these animals may represent an indicator of environmental exposure or may be actively involved in the transmission of L. intracellularis to foals by acting as potential reservoir/amplifier hosts. This study is the first to demonstrate the magnitude of L. intracellularis shedding in the feces of wild rodents and feral cats and to indicate the significant infection risk that wild rodents and feral cats pose to naïve horses in South Korea.

Effect of Tetracycline Dose and Treatment Mode on Selection of Resistant Coliform Bacteria in Nursery Pigs.

This study describes the results of a randomized clinical trial investigating the effect of oxytetracycline treatment dose and mode of administration on the selection of antibiotic-resistant coliform bacteria in fecal samples from nursery pigs. Nursery pigs (pigs of 4 to 7 weeks of age) in five pig herds were treated with oxytetracycline for Lawsonia intracellularis-induced diarrhea. Each group was randomly allocated to one of five treatment groups: oral flock treatment with a (i) high (20 mg/kg of body weight), (ii) medium (10 mg/kg), or (iii) low (5 mg/kg) dose, (iv) oral pen-wise (small-group) treatment (10 mg/kg), and (v) individual intramuscular injection treatment (10 mg/kg). All groups were treated once a day for 5 days. In all groups, treatment caused a rise in the numbers and proportions of tetracycline-resistant coliform bacteria right after treatment, followed by a significant drop by the time that the pigs left the nursery unit. The counts and proportions of tetracycline-resistant coliforms did not vary significantly between treatment groups, except immediately after treatment, when the highest treatment dose resulted in the highest number of resistant coliforms. A control group treated with tiamulin did not show significant changes in the numbers or proportions of tetracycline-resistant coliforms. Selection for tetracycline-resistant coliforms was significantly correlated to selection for ampicillin- and sulfonamide-resistant strains but not to selection for cefotaxime-resistant strains. In conclusion, the difference in the dose of oxytetracycline and the way in which the drug was applied did not cause significantly different levels of selection of tetracycline-resistant coliform bacteria under the conditions tested.IMPORTANCE Antimicrobial resistance is a global threat to human health. Treatment of livestock with antimicrobials has a direct impact on this problem, and there is a need to improve the ways that we use antimicrobials in livestock production. We hypothesized that antibiotic resistance development following treatment of diarrhea in nursery pigs could be reduced either by lowering the dose of oxytetracycline or by replacing the commonly used practice of flock treatment with individual or small-group treatments, since this would reduce the number of pigs treated. However, the study showed no significant difference between treatment groups with respect to the number or proportion of tetracycline-resistant coliforms selected. The most important conclusion is that under practical field conditions, there will be no added value, in terms of lowering resistance development, by exchanging flock treatment for individual or small-group treatment of nursery pigs. The reason for the lack of an effect of single-animal treatment is probably that such animals share the environment with treated animals and take up resistant bacteria from the environment.

Lawsonia intracellularis and Porcine Circovirus type-2 infection in Estonia.

The present study describes the reasons of post-weaning distress in Estonian pig herds. Here we examined the natural cases of Lawsonia intracellularis and porcine circovirus 2 (PCV2) infection and co-infections. The presence of L. intracellularis in swine herds were tested by PCR and by histopathological methods, whereas PCV2 was detected by real-time-PCR and immunohistochemical stainings. Seven of the 11 investigated herds with signs of post-weaning wasting were infected with L. intracellularis and all 11 herds with PCV2. From the analysed samples 22.2% were infected with L. intracellularis and 25% with PCV2. The results of microbiological studies suggested that the piglets suffered from enteritis and pneumonia. Escherichia coli and Pasteurella multocida often aggravated the process of illness. The frequency of L. intracellularis was high in pigs 7-12 weeks old (18.5-42.7%) and PCV2 infection was too high in pigs 7-12 weeks old (24.8-32.7%). E. coli was often a co-factor with L. intracellularis and PCV2. The primary reasons of post weaning wasting were PCV2 and E. coli, later aggravated by L. intracellularis and other pathogens. Our results indicated that different pathogens have an important role in developing post-weaning wasting. Proliferative intestinal inflammation caused by L. intracellularis is mainly characterised by its localization and morphological findings. The main gross lesions were the enlargement of mesenteric lymph nodes and thickening of the wall of ileum. In post-weaning multi-systemic wasting syndrome there are characteristic histological lesions in lymphoid tissues. They consist of a variable degree of lymphocyte depletion, together with histiocytic and/or multinucleate giant cell infiltration. This basic lymphoid lesions is observable in almost all tissues of a single severely affected animal, including lymph nodes, Peyer's patches and spleen. Sporadically, multifocal coagulative necrosis may be observed.

Use of flow cytometry and PCR analysis to detect 5-carboxyfluoroscein-stained obligate intracellular bacteria Lawsonia intracellularis invasion of McCoy cells.

In this study, we describe a method to quantify invasion of obligate intracellular bacteria, Lawsonia intracellularis, inside McCoy cells. In immunological research, the cell-permeable fluorescent dye 5'-carboxyfluoroscein succidyl ester (CFSE) is commonly used to quantify eukaryotic cellular proliferation. Instead of using it in this traditional way, we stained L. intracellularis with CFSE dye prior to infection of McCoy cells. Flow cytometry was performed to quantify the percentage of eukaryotic cells which had taken up or were associated with fluorescent bacteria. As obligate intracellular bacteria, they cannot replicate outside of eukaryotic cells and thus qPCR analysis was used to quantify bacterial growth. Indirectly, PCR analysis confirmed invasion rather than adherence to the McCoy cell surface. Fluorescent activated cell sorting (FACS) was used to sort the CFSE(+) (i.e. infected) McCoy cells from the CFSE(-) (i.e. non-infected) McCoy cells and confocal microscopy was used to confirm bacterial invasion and cytosolic localization of CFSE-L. intracellularis. To show that this approach could be used in conjunction with functional assays, we investigated the effect that serum antibodies had on CFSE-bacterial invasion and growth. Instead of blocking invasion, rabbit hyperimmune serum augmented invasion of the bacteria inside McCoy cells and qPCR analysis confirmed bacterial growth over the course of 5days. We conclude that CFSE-labeling of bacteria and qPCR can be used to track and quantify bacterial invasion and may be a valuable tool for studying the invasive properties of bacteria, especially if commercial antibodies are not available. This approach may be adapted for use in other obligate intracellular bacteria and intracellular pathogens.

Occurrence of diarrhoea and intestinal pathogens in non-medicated nursery pigs.

BACKGROUND: Intestinal disease in nursery pigs is the most common cause of antibiotic usage in pigs in Denmark. The decision to initiate batch medication of intestinal diseases in nursery pigs is typically made by the stock personnel based on clinical assessments of pigs and counting of diarrhoeic faecal pools on the pen floor. The target population of this study was batches of nursery pigs (10-66 days after weaning) where the stock personnel assessed the pigs to be without signs of intestinal disease and therefore did not needed treatment. The objective was to determine the within-herd prevalence of diarrhoea, and to determine the prevalence of Escherichia coli F4 and F18, Lawsonia intracellularis and Brachyspira pilosicoli by quantitative PCR in pigs with and without diarrhoea. RESULTS: The overall apparent prevalence of diarrhoeic pigs across sixteen herds was 32.6 % (CI 95 % 27.9-37.3). The prevalence of diarrhoea increased (p ≤ 0.001) with age of the pigs (days after weaning) with an odds ratio of 1.04 (CI 95 % 1.02-1.05) per extra day. Diarrhoeic pools were observed in 51 % of the pens. L. intracellularis, B. pilosicoli, E. coli F4 and F18 were detected in 20, 17, 13 and 11 % of the 256 faecal samples analysed by quantitative PCR respectively. There was no association between detection of pathogens and diarrhoea status of the individual pigs and between detection of pathogens in a pen and diarrhoea floor pools. In 51 % of the samples from diarrhoeic pigs, pathogens were not detected. Only 5 % of the 3060 pigs examined had clinical signs of diseases other than diarrhoea. CONCLUSIONS: One-third of non-medicated nursery pigs had diarrhoea when clinically examined even though they were assessed as healthy by stock personnel. Diarrhoeic status of the pigs and diarrhoeic pools in pen was a poor indicator of intestinal infections with E. coli F4 and F18, L. intracellularis and B. pilosicoli and subclinical infections were common. Therefore, clinical examination and counting of diarrhoea pools should be supported by microbiological testing as decision tools for initiation of batch treatments of intestinal infections in nursery pigs.

Clinical utility and performance of sock sampling in weaner pig diarrhoea.

Low pathogen diarrhoea is a group-level diagnosis, characterised by non-haemorrhagic diarrhoea. In the current study, the apparent prevalence of low pathogen diarrhoea outbreaks in Danish herds was investigated along with the clinical utility of a laboratory examination for intestinal disease, agreement between three consecutive herd examinations from the same herd and agreement between quantitative PCR results from pooled faecal samples and sock samples. Twenty-four veterinarians submitted faecal and sock samples for quantitative PCR testing from outbreaks of diarrhoea in nursery pigs (n=38 herds) where the farmer or veterinarian had decided that antimicrobial treatment was necessary. The veterinarians were asked to fill in a questionnaire and participate in telephone interviews. The apparent prevalence of low pathogen diarrhoea was 0.18 (95% CL: 0.08-0.34). Agreement between the veterinarians' clinical aetiological diagnosis and the pooled faecal sample was 0.18 (95% CL: 0.08-0.34), and Cohen's Kappa was 0.03 (95% CL: -0.08 to 0.14). Antibiotic treatment or prevention strategies were changed in 0.63 (95% CL: 0.46-0.78) of the herds, and the veterinarians indicated that, for 0.32 (95% CL: 0.18-0.50) of the herds, changes were related to the diagnostic results from the laboratory examination performed in the study. In 0.16 (95% CL: 0.05-0.36) of the herds, the same infections were demonstrated at all three consecutive examinations. No herds had three consecutive diarrhoea outbreaks classified as low pathogen diarrhoea. For the quantitative results (log10 of the summed amounts of Lawsonia intracellularis, Brachyspira pilosicoli, Escherichia coli F4 and F18) agreement between pooled faecal samples and sock samples was evaluated. Lin's concordance correlation coefficient was 0.69 (95% CL: 0.48-0.82), and the mean difference between the two types of samples was -0.38 log10 bacteria/g faeces (SD=1.59log10 bacteria/g faeces; 95% CI: -0.90 to 0.14log10 bacteria/g faeces). Agreement for the dichotomised results was 0.89 (95% CI: 0.75-0.97) when test results were classified as low pathogen diarrhoea or not, and Cohen's Kappa was 0.61 (95% CI: 0.26-0.95). In relation to detection of the individual infections, agreement was 0.63 (95% CI: 0.46-0.78), and Cohen's Kappa was 0.53 (95% CI: 0.34-0.71). In conclusion, low pathogen diarrhoea is a common finding amongst diarrhoea outbreaks that are subjected to antibiotic batch treatment in Danish nursery pigs. Sock samples seem to offer a reliable diagnostic method with impact on clinical decisions for treatment and prevention. However, both the diarrhoea type and the aetiology change with time in the majority of herds, indicating a potential need for frequent diagnostic examinations.

Lawsonia intracellularis and equine proliferative enteropathy.

Lawsonia intracellularis is the etiologic agent for equine proliferative enteropathy (EPE), which typically affects weanling and yearling horses. In North America, EPE cases often occur between August and January, although cases outside of this time frame have been reported. Clinical signs of EPE are usually nonspecific and include lethargy, pyrexia, anorexia, peripheral edema, weight loss, colic, and diarrhea. Diagnosis is based on the presence of hypoproteinemia and hypoalbuminemia along with clinical signs and positive commercial serologic and/or molecular testing. Treatment requires the use of antimicrobials with good intracellular penetration and supportive care to prevent or decrease secondary complications.

Detection of enteric pathogens in Turkey flocks affected with severe enteritis, in Brazil.

Twenty-two flocks of turkeys affected by enteric problems, with ages between 10 and 104 days and located in the Southern region of Brazil, were surveyed for turkey by PCR for turkey astrovirus type 2 (TAstV-2), turkey coronavirus (TCoV), hemorrhagic enteritis virus (HEV), rotavirus, reovirus, Salmonella spp., and Lawsonia intracellularis (Li) infections. Eleven profiles of pathogen combination were observed. The most frequently encountered pathogen combinations were TCoV-Li, followed by TCoV-TAstV-2-Li, TCoV-TastV-2. Only TCoV was detected as the sole pathogen in three flocks. Eight and 19 flocks of the 22 were positive for TAstV-2 and TCoV, respectively. Six were positive for Salmonella spp. and L. intracellularis was detected in 12 turkey flocks. Reovirus and HEV were not detected in this survey. These results throw new light on the multiple etiology of enteritis in turkeys. The implications of these findings and their correlation with the clinical signs are comprehensively discussed, illustrating the complexity of the enteric diseases.

Pathogen presence in feral pigs and their movement around two commercial piggeries in Queensland, Australia.

Feral pigs are wild animal reservoirs of infectious pathogens transmissible to other species, all of which are transmissible to domestic pigs. The objective of this study was to detect the presence of harmful production-limiting pathogens; Brucella suis, Leptospira species, Lawsonia intracellularis, Mycoplasma hyopneumoniae and Actinobacillus pleuropneumoniae in a feral pig population within a 10 km radius of two large-scale commercial piggeries in Southern Queensland, Australia. The movement pattern of six pigs within the feral population was also investigated using geographic positioning system collars. All pathogens were present in the feral pig population except for A pleuropneumoniae. The true seroprevalence (TP) from 83 serum samples was 10.5 per cent for B suis, 48.6 per cent for Leptospira species, 100 per cent for L intracellularis and 42.1 per cent for M hyopneumoniae. Of 72 lung samples, 27.6 per cent were positive for M hyopneumoniae. Serum samples from 86 domestic sows within the study region were positive for Leptospira species (TP 2.1 per cent), L intracellularis (TP 100 per cent) and M hyopneumoniae (TP 100 per cent). The majority of feral pig movement was within 5 km of the piggeries, with one approaching to 100 m of the free-range piggery. The presence of pathogens in feral pigs in such close proximity to commercial piggeries could pose a biosecurity risk.

Species-specificity of equine and porcine Lawsonia intracellularis isolates in laboratory animals.

Lawsonia intracellularis infection causes proliferative enteropathy (PE) in many mammalian species, with porcine and equine proliferative enteropathy (PPE and EPE) known worldwide. Hamsters are a well-published animal model for PPE infection studies in pigs. There is no laboratory animal model for EPE infection studies and it is not known whether there is species-specificity for equine or porcine isolates of L. intracellularis in animal models. The objective of this study was to determine whether it is possible to generate typical EPE lesions in hamsters after inoculation with an equine strain of L. intracellularis (EPE strain) and whether it is comparatively possible to generate PPE lesions in rabbits after inoculation with a porcine strain of L. intracellularis (PPE strain). In 2 separate trials, 4-week-old and 3-week-old weanling golden Syrian hamsters were challenged with EPE strains and compared to uninfected (both trials) and PPE-infected controls (Trial 2 only). Concurrently, 6 female New Zealand white juvenile rabbits were infected with PPE strain and observed concomitantly to 8 similar rabbits infected with EPE strain for a different experiment. Hamsters and rabbits were observed for 21 to 24 days post-infection (DPI), depending on the experiment. Neither infected species developed clinical signs. The presence of disease was assessed with diagnostic techniques classically used for pigs and horses: immune-peroxidase monolayer assay on sera; quantitative polymerase chain reaction (qPCR) detection of molecular DNA in feces; and hematoxylin and eosin (H&E) stain and immunohistochemistry (IHC) on intestinal tissues. Our results showed that EPE-challenged hamsters do not develop infection when compared with PPE controls (IHC, P = 0.009; qPCR, P = 0.0003). Conversely, PPE-challenged rabbits do not develop typical intestinal lesions in comparison to EPE-challenged rabbits, with serological response at 14 DPI being significantly lower (P = 0.0023). In conclusion, PPE and EPE strains appear to have different host-specificities for hamsters and rabbits, respectively.

L'infection par Lawsonia intracellularis provoque une entéropathie proliférative chez de nombreuses espèces de mammifères; celle des porcins (EPP) et des équidés (EEP) sont connues mondialement. Les hamsters sont un modèle animal bien connu pour l'étude de l'EPP. Il n'existe pas de modèle animal de laboratoire pour étudier l'EEP, et on ne sait pas s'il y a spécificité d'espèce pour les isolats équins ou porcins de L. intracellularis dans des modèles animaux. L'objectif de la présente étude était de déterminer s'il est possible de générer des lésions typiques d'EEP chez les hamsters après inoculation d'une souche équine de L. intracellularis (souche EEP) et s'il est également possible de générer des lésions d'EPP chez des lapins après inoculation d'une souche porcine de L. intracellularis (souche EPP). Dans 2 essais séparés, des hamsters dorés syriens sevrés âgés de 4 semaines et de 3 semaines ont été inoculés avec des souches EEP, et ont été comparés à des témoins non infectés (les deux essais) et à des témoins infectés avec EPP (essai 2 seulement). Parallèlement, 6 jeunes lapines Nouvelle-Zélande ont été infectées par la souche EEP et observées de façon concomitante à 8 lapins similaires infectés par la souche EPP pour une expérience différente. Les hamsters et les lapins ont été observés pendant 21 à 24 jours après l'infection (JAI), en fonction de l'expérience. Aucune des espèces infectées n'a développé de signes cliniques. La présence de maladie a été évaluée par des techniques classiques de diagnostic utilisées pour les porcs et les chevaux : l'essai par immuno-peroxydase sur monocouche pour les sérums; la détection par réaction d'amplification en chaîne par la polymérase quantitative (qPCR) de l'ADN moléculaire dans les selles; la coloration hématoxyline-éosine et l'immunohistochimie (IHC) sur des tissus intestinaux. Nos résultats ont montré que les hamsters inoculés avec EEP ne développent pas d'infection comparativement aux EPP témoins (IHC P = 0,009; qPCR P = 0,0003). À l'inverse, les lapins inoculés avec EPP ne développent pas des lésions intestinales typiques comparativement aux lapins inoculés avec EEP, avec une réponse sérologique à 14 JAI significativement plus faible (P = 0,0023). En conclusion, les souches d'EPP et d'EEP semblent avoir des spécificités d'hôte différentes chez les hamsters et les lapins, respectivement.(Traduit par Dr. J.M. Dhillon).

Risk factors associated with Lawsonia intracellularis in English pig farms.

Porcine proliferative enteropathy (PPE) caused by the bacterium Lawsonia intracellularis causes considerable economic loss to the pig industry. The objective of this study was to evaluate the seroprevalence of L. intracellularis exposure in different age groups of pigs (growers to finishers) within English farms and to identify potential risk factors. Samples were obtained in a cross-sectional study of 147 farms between 2008 and 2009. Twelve samples (six growers and six finishers) from each farm were tested for L. intracellularis by antibody ELISA. At animal level there was a significant positive linear trend between seroprevalence and age in weeks (r(2)=2.65, P<0.001), with seroprevalence lowest (24.73%) at 11 weeks and highest (93.33%) at 24 weeks. At farm level, seroprevalence was significantly lower in growers than finishers (56.80% vs. 94.26%, P<0.001). Farms reporting minor Salmonella problems and those that brought boars onto the farm had higher odds of testing positive in growers (OR 5.69 and 4.31, respectively. On the other hand, farms where producers considered temperature as an important stress factor (OR=0.3) and which had more than two sites on which pigs are kept (OR=0.16) were less likely to test positive in growers. The current study confirmed the high prevalence of L. intracellularis in English pig farms. The potential risk factors and further information of the disease impact on the farm productivity will aid the development of appropriate control strategies through better understanding of the disease.

Investigation of the association of growth rate in grower-finishing pigs with the quantification of Lawsonia intracellularis and porcine circovirus type 2.

As a part of a prospective cohort study in four herds, a nested case control study was carried out. Five slow growing pigs (cases) and five fast growing pigs (controls) out of 60 pigs were selected for euthanasia and laboratory examination at the end of the study in each herd. A total of 238 pigs, all approximately 12 weeks old, were included in the study during the first week in the grower-finisher barn. In each herd, approximately 60 pigs from four pens were individually ear tagged. The pigs were weighed at the beginning of the study and at the end of the 6-8 weeks observation period. Clinical data, blood and faecal samples were serially collected from the 60 selected piglets every second week in the observation period. In the killed pigs serum was examined for antibodies against Lawsonia intracellularis (LI) and procine circovirus type 2 (PCV2) and in addition PCV2 viral DNA content was quantified. In faeces the quantity of LI cells/g faeces and number of PCV2 copies/g faeces was measured by qPCR. The objective of the study was to examine if growth rate in grower-finishing pig is associated with the detection of LI and PCV2 infection or clinical data. This study has shown that diarrhoea is a significant risk factor for low growth rate and that one log(10) unit increase in LI load increases the odds ratio for a pig to have a low growth rate by 2.0 times. Gross lesions in the small intestine and LI load>log(10)6/g were significant risk factors for low growth. No association between PCV2 virus and low growth was found.