RESUMEN
F4ac-positive enterotoxigenic Escherichia coli (ETEC) must attach to the intestinal mucosa to cause diarrhea in piglets. Prevention of bacterial attachment to the intestinal mucosa is the most effective defense against ETEC-induced diarrhea. Porcine milk fat globule membranes (MFGM) were shown to be able to inhibit attachment of ETEC to the intestinal brush border; however, the specific components of porcine MFGM that inhibited attachment of ETEC to enterocytes were not identified. Accordingly, the purpose of this study was to identify F4ac-binding MFGM proteins by overlay Western blot and affinity chromatography. The proteome of porcine MFGM was characterized and the following F4ac-binding proteins were detected by overlay Western blot and affinity chromatography: lactadherin, butyrophilin, adipophilin, acyl-CoA synthetase 3, and fatty acid-binding protein 3. The biological function of these proteins was not investigated but it is possible that their interaction with F4ac fimbria interferes with bacterial attachment and colonization.
Les Escherichia coli entérotoxinogénique (ETEC) positif pour F4ac doivent s'attacher à la muqueuse intestinale pour causer la diarrhée chez les porcelets. L'empêchement de l'attachement bactérien à la muqueuse intestinale est le moyen de défense le plus efficace contre la diarrhée induite par les ETEC. Les membranes de globules de gras de lait porcin (MFGM) ont été montré comme étant capable d'inhiber l'attachement des ETEC à la bordure en brosse intestinale; toutefois, les composantes spécifiques des MFGM porcines qui inhibaient l'attachement des ETEC aux entérocytes ne furent pas identifiées. Ainsi, le but de la présente étude était d'identifier les protéines des MFGM liant F4ac par immunobuvardage par superposition et chromatographie d'affinité. Le protéome des MFGM porcine fut caractérisé et les protéines liant F4as suivantes furent détectées par immunobuvardage par superposition et chromatographie d'affinité : lactadhérine, butyrophiline, adipophiline, acyl-CoA synthétase 3, et la protéine 3 liant les acides gras. La fonction biologique de ces protéines ne fut pas étudiée mais il est possible que leur interaction avec les fimbriae F4ac interfère avec l'attachement bactérien et la colonisation.(Traduit par Docteur Serge Messier).
Asunto(s)
Escherichia coli Enterotoxigénica/metabolismo , Proteínas de Escherichia coli/metabolismo , Glucolípidos/metabolismo , Glicoproteínas/metabolismo , Porcinos/fisiología , Animales , Adhesión Bacteriana , Western Blotting/veterinaria , Cromatografía de Afinidad/veterinaria , Proteínas de Escherichia coli/química , Femenino , Glucolípidos/química , Glicoproteínas/química , Gotas Lipídicas , Unión ProteicaRESUMEN
Absorption of colostral proteins ingested by neonatal piglets within 24 to 36 h after birth is generally considered to be non-selective. Nevertheless, the transfer of colostral proteins, except immunoglubulins, from gut to bloodstream after natural suckling is still poorly characterized. The purpose of this study was to investigate the changes in 2-dimensional electrophoretic plasma protein profiles of neonatal piglets before and after suckling, in order to characterize the gastrointestinal absorption of colostral proteins into the neonatal bloodstream. As expected, the most significant change in plasma after suckling is the presence of a large amount of immunoglobulin. However, while the concentration of a few proteins was mildly increased in post-suckling plasma, the evidence of absorption of colostral non-immunoglobulin proteins by neonatal piglets was not detected in this study.
Asunto(s)
Proteínas Sanguíneas/análisis , Calostro/metabolismo , Porcinos/sangre , Animales , Animales Recién Nacidos , Animales Lactantes , Femenino , Proteómica/métodosRESUMEN
The emergence of severe porcine circoviral disease in North America is associated with Porcine circovirus-2 genotype b (PCV-2b), which has led to speculation that PCV-2b is more virulent than PCV-2a. The objectives of this study were to 1) correlate the PCV-2 DNA concentration and lesions in wasting (WST) and age-matched healthy (HLTH) pigs from 2 clinically affected farms, and unaffected (UNFCT) pigs from a farm with no prior clinical or diagnostic history of PCVD; and 2) to determine the initial estimates of sensitivity and specificity of PCV-2 quantitative polymerase chain reaction (qPCR). PCV-2b was confirmed in all 3 farms. Compared with HLTH pigs, WST pigs demonstrated significantly more prevalent thymic atrophy, failure of normal pulmonary collapse, and ascites (P < 0.017 for all). The HLTH and UNFCT pigs had significantly more pronounced lymphoid germinal centers and proliferative paracortical T-dependent zones, compared with WST pigs (P < 0.017). Across all tissues, PCV-2 DNA concentrations were significantly higher in WST compared with HLTH and UNFCT pigs (P < 0.017 for all). The PCV-2 DNA concentrations were strongly correlated with PCV-2 nucleocapsid staining intensity in lymph node, spleen, Peyer's patches, lung, liver, and kidney (0.60 < or = r < or = 0.84). In the current study, the PCV-2 DNA log10 cutoff concentrations best able to distinguish WST from HLTH and UNFCT pigs were between 7.0 and 8.0 per gram for tissues, and between 4.0 and 5.0 per milliliter for sera. The presence of PCV-2b in UNFCT pigs is evidence that PCV-2b by itself is not sufficient to induce severe disease.
Asunto(s)
Circovirus , ADN Viral/aislamiento & purificación , Síndrome Multisistémico de Emaciación Posdestete Porcino/virología , Animales , Ascitis/patología , Canadá , Estudios de Casos y Controles , Circovirus/genética , Genotipo , Pulmón/patología , Nucleocápside , Síndrome Multisistémico de Emaciación Posdestete Porcino/patología , Coloración y Etiquetado , Porcinos , Timo/patología , Carga ViralRESUMEN
Maedi-visna (MV) is a relatively common chronic infection of sheep in North America resulting in economic loss to the sheep industry. The objectives of this study were to: 1) measure the prevalence of MV infection in culled ewes in Alberta, by histologic examination (lungs and udder) and serologic testing using an agar gel immunodiffusion (AGID) test, 2) examine any geographic differences in its prevalence in the province, 3) evaluate the level of agreement between histopathologic examination and serologic testing, 4) grade the lesions and correlate the serologic results with the presence of severe histological lesions, and 5) correlate the presence of histological lesions in the lungs and udder in the same animal. Based on histologic findings, the prevalence of MV was 26.8%, compared with 13.0% using serologic testing. There were no significant geographical differences in prevalence, fair agreement (kappa = 42.0%) between histopathologic and serologic results, and poor agreement (kappa = 11.5%) between the presence of lung and udder histological lesions within the same animal. This study indicates that MV is relatively common in culled ewes in Alberta, with no significant geographic variation. The poor sensitivity of the AGID test, compared with histologic examination, should be taken into consideration when interpreting serologic results.
Asunto(s)
Anticuerpos Antivirales/sangre , Inmunodifusión/veterinaria , Neumonía Intersticial Progresiva de los Ovinos/epidemiología , Pruebas Serológicas/veterinaria , Virus Visna-Maedi/inmunología , Alberta/epidemiología , Animales , Diagnóstico Diferencial , Femenino , Inmunodifusión/métodos , Inmunodifusión/normas , Pulmón/patología , Pulmón/virología , Glándulas Mamarias Animales/patología , Glándulas Mamarias Animales/virología , Neumonía Intersticial Progresiva de los Ovinos/sangre , Neumonía Intersticial Progresiva de los Ovinos/diagnóstico , Neumonía Intersticial Progresiva de los Ovinos/patología , Prevalencia , Sensibilidad y Especificidad , Pruebas Serológicas/métodos , Pruebas Serológicas/normas , OvinosRESUMEN
An outbreak of malignant catarrhal fever (MCF) among bison sold at an auction market was studied for an 18-month period. Forty-five of 163 bison submitted for sale from 8 different bison farms died on 7 other destination farms. The outbreak began on day 50 after the sale, peaked between days 60 and 70, and ended on day 220. Twenty-one dead bison were confirmed to be MCF cases by clinical histories, pathology, and detection of ovine herpesvirus-2 DNA in their tissues with polymerase chain reaction assays. Twenty-four dead bison were classified as suspect MCF cases from clinical histories. No cases of MCF were observed among bison remaining on originating farms or resident bison mixed with sale bison on destination farms. There were no sheep reported within 3 km of originating or destination farms, limiting bison exposure to sheep to the auction facility, where sheep were present for less than 1 day. The outbreak provides an illustration of the temporal distribution of MCF mortality expected in bison and an estimate of the time from exposure until death from MCF after a single short exposure to sheep. The study provides evidence that bison with MCF do not transmit MCF to other bison.
Asunto(s)
Bison/virología , Brotes de Enfermedades/veterinaria , Fiebre Catarral Maligna/epidemiología , Animales , Femenino , Masculino , Fiebre Catarral Maligna/transmisión , Saskatchewan/epidemiología , OvinosRESUMEN
Transmission electron microscopy (TEM) was used to characterize the colonization patterns of 3 pathogenic Escherichia coli strains: PD58 and PD149 of the AIDA-I/STb/EAST1 pathotype, serogroup O: ND (not determined), and PD31 of the LT/STb/EAST1 pathotype, serogroup O149. These strains were isolated from diseased piglets and caused diarrhea in experimentally inoculated, newborn, colostrum-deprived pigs. In this study, intestinal tissues from newborn pigs experimentally infected with a high inoculum (20 ml containing 10(10) cfu) were harvested and examined for bacterial colonization using light microscopy. A nonaqueous perfluorocarbon fixation method was used to preserve the glycocalyx of the microvillus border in tissues collected for TEM. Transmission electron micrographs revealed that E. coli strain PD149 displayed long flexible fimbria-like structures that intimately attached the bacteria both to the microvillus border of the upper colon and to adjacent bacteria. In vitro, this strain demonstrated the localized adherence pattern to HEp-2 cells characteristic of enteroaggregative E. coli (EAggEC). Both PD58 and PD31 strains colonized the upper colon through the formation of a biofilm, also characteristic of EAggEC. Strains PD58 and PD31 adhered poorly to HEp-2 cells in vitro, although these demonstrated a colonization pattern suggestive of diffuse and aggregative adherence, respectively. These findings suggest that strains PD58 and PD149, expressing the AIDA-I, factor and strain PD31 represents hybrid pathotypes of diarrheagenic E. coli and that they probably cause diarrhea in piglets through differing mechanisms.
Asunto(s)
Adhesinas de Escherichia coli/biosíntesis , Diarrea/veterinaria , Infecciones por Escherichia coli/veterinaria , Escherichia coli/crecimiento & desarrollo , Enfermedades de los Porcinos/microbiología , Animales , Animales Recién Nacidos , Adhesión Bacteriana/fisiología , Línea Celular , Colon/microbiología , Colon/patología , Colon/cirugía , Colon/ultraestructura , Diarrea/microbiología , Diarrea/patología , Escherichia coli/aislamiento & purificación , Escherichia coli/metabolismo , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/patología , Femenino , Humanos , Mucosa Intestinal/microbiología , Mucosa Intestinal/patología , Mucosa Intestinal/ultraestructura , Masculino , Microscopía Electrónica/veterinaria , Distribución Aleatoria , Enfermedades de los Porcinos/patología , VirulenciaRESUMEN
One potential reason for the enhancement of immune responses to DNA vaccines following electroporation is increased gene expression. However, the inflammatory response and accompanying cellular infiltration stimulated by electroporation may also be essential for enhancing immune responses to DNA vaccines. These parameters were investigated in pigs, using different electroporation conditions to induce different levels of gene expression and inflammation. Results indicated that the least effective strategy was conventional intramuscular injection where there was low gene expression and low inflammatory cell infiltration. The most efficacious strategy was plasmid administration immediately followed by electroporation. This latter set of conditions elicited a combination of high gene expression and high cellular infiltration. This indicates that electroporation enhances immune responses to DNA vaccines through increased gene expression and inflammatory cell infiltration.
Asunto(s)
Electroporación , Vacunas de ADN/administración & dosificación , Vacunas de ADN/inmunología , Animales , Femenino , Expresión Génica , Técnicas de Transferencia de Gen , Antígenos de Superficie de la Hepatitis B/genética , Antígenos de Superficie de la Hepatitis B/metabolismo , Inflamación/patología , Inyecciones Intramusculares , Interferón gamma/biosíntesis , Leucocitos/inmunología , Luciferasas/metabolismo , Masculino , Músculo Esquelético/inmunología , Músculo Esquelético/patología , Necrosis , Plásmidos/administración & dosificación , Plásmidos/genética , Plásmidos/inmunología , Porcinos , Factores de Tiempo , Regulación hacia Arriba , Vacunas de ADN/genética , Proteínas del Envoltorio Viral/genética , Proteínas del Envoltorio Viral/metabolismo , Proteínas ViralesRESUMEN
CpG oligonucleotides (ODNs) are potent mucosal and systemic adjuvants. For practical applications however, improvements in delivery need to be developed. A mouse model was used to determine if the biological activity of CpG ODNs could be enhanced using a novel delivery system of biphasic lipid vesicles (Biphasix Vaccine-Targeting Adjuvant; VTA). Immunization studies were performed to evaluate the potential of VTA formulations to enhance the immunoadjuvant activity of CpG ODNs following systemic or mucosal administration with gD. Immune responses following immunization were assessed by protection from HSV-1 viral challenge and characterization of serum gD-specific antibody responses using ELISA. VTA formulations in combination with CpG and glycoprotein D (gD) were able to increase gD-specific IgG in serum compared to gD alone, and protect from a lethal HSV-1 challenge following subcutaneous immunization. Following mucosal immunization, VTA formulations in combination with CpG and antigen enhanced mucosal IgA responses compared to CpG and antigen administered in PBS.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Membrana Dobles de Lípidos/inmunología , Oligonucleótidos/farmacología , Vacunas/inmunología , Adyuvantes Inmunológicos/efectos adversos , Animales , Anticuerpos Antivirales/análisis , Anticuerpos Antivirales/biosíntesis , Química Farmacéutica , Islas de CpG , Citocinas/biosíntesis , Citocinas/metabolismo , Portadores de Fármacos , Sistemas de Liberación de Medicamentos , Femenino , Herpes Simple/patología , Herpes Simple/prevención & control , Vacunas contra el Virus del Herpes Simple/inmunología , Herpesvirus Humano 1/inmunología , Inmunización , Ratones , Ratones Endogámicos BALB C , Membrana Mucosa/inmunología , Oligonucleótidos/efectos adversos , Piel/patologíaRESUMEN
To identify emerging Escherichia coli that have the potential to cause diarrhea in pigs, the prevalence of E. coli pathotypes was determined among 170 and 120 isolates from diarrheic and nondiarrheic piglets, respectively. The isolates were tested for F4, F5, F6, F18, and F41 fimbriae, for E. coli attaching and effacing (EAE), porcine attaching and effacing-associated (Paa), and adhesin involved in diffuse adherence (AIDA-I) factors, for LT, STa, STb, and enteroaggregative heat-stable (EAST1) enterotoxins, and for Shiga toxins (Stxl, Stx2, and Stx2e), using DNA hybridization and polymerase chain reaction. All isolates were O-serotyped and tested for antibiotic resistance against 10 drugs. Seventeen different pathotypes, accounting for 40.0% of the isolates, were recovered from diarrheic piglets. The main pathotypes included EAST1 (13.5%), F4/LT/STb/EAST1 (6.5%), AIDA-I/STb/EAST1 (4.1%), F5/STa (2.9%), EAE/EAST1 (2.9%), and AIDA-I/F18 (2.3%). Only 3 pathotypes, EAE (11.7%), EAST1 (10.8%), and EAE/EAST1 (3.3%), were recovered from nondiarrheic piglets. Paa factor was detected in 8.8% and 7.5% of isolates from diarrheic and nondiarrheic piglets, respectively, and always was associated with other virulence determinants. Overall, 22.9% of isolates from diarrheic piglets appeared to be enteropathogens: enterotoxigenic E. coli (11.7%), enteropathogenic E. coli (3.5%), and E. coli isolates (3.0%) for which none of the above adherence factors was detected. Pathotypes AIDA-I/STb/EAST1 and AIDA-I/STb were isolated only from diarrheic piglets and accounted for 4.7% of isolates. Strains of these pathotypes induced diarrhea when inoculated into newborn colostrum-deprived pigs, in contrast to an isolate positive only for EAST1, which did not induce diarrhea. Antibiotic sensitivity test showed that isolates of the AIDA-I/STb/EAST1 and AIDA-I/STb pathotypes were the only strains sensitive to enrofloxacin, gentamicin, neomycin, and trimethoprim-sulfamethoxazole. This study showed that at least 20.5% of isolates from diarrheic piglets appeared to be associated with AIDA-I/STb pathotype and that EAST1 pathotype is probably not an important marker for diarrhea in piglets.