Topological analysis of IFN-induced transmembrane proteins in porcine peripheral blood lymphocytes and changes of transcription level after infection with porcine reproductive and respiratory syndrome virus / 中国生物制品学杂志

@#Objective To analyze the topology of IFN-induced transmembrane(IFITM) protein in porcine peripheral blood lymphocytes(PBMCs) and detect the change of IFITM mRNA transcription in PBMCs after porcine reproductive and respiratory syndrome virus(PRRSV) infection in vitro.Methods PRRSV,porcine circovirus 2(PCV2) and Japanese encephalitis virus(JEV) negative anticoagulant blood of piglets were collected aseptically and isolated for PBMCs.Porcine IFITM CDS sequence was amplified by PCR,sequenced and analyzed for topology.PBMCs were infected with PRRSV in vitro.Cell samples were collected at 12,24,36 and 48 h after infection,detected for PRRSV infection by RT-PCR,and detected for mRNA transcription level changes of IFITM1,IFITM2 and IFITM3 by RT-PCR.Results The porcine PBMCs were successfully isolated and the full-length sequence of IFITM CDS derived from PBMCs was cloned.The porcine IFITM protein might have two topological structures.PBMCs inoculated with PRRSV for 24 h produced obvious cytopathic effect.PRRSV was replicated in PBMCs.The transcription levels of IFITM1,IFITM2 and IFITM3 mRNA in PBMCs were significantly up-regulated at the early stage of PRRSV infection,and reached the peak at 12h after infection,and then gradually decreased;The transcription level of IFITM1 mRNA increased at 36 h after virus infection and then declined rapidly.Conclusion PRRSV infection in vitro significantly up-regulated the transcription level of IFITM mRNA in PBMCs,indicating that IFITM was involved in the antiviral immune response of PBMCs.This study provided a reference for revealing the natural immune response against PRRSV in vivo.

Susceptibility screening of highly pathogenic porcine reproductive and respiratory syndrome virus in several strains of minipigs / 中国比较医学杂志

Objective To screen strains of minipigs sensitive to highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) for evaluation of HP-PRRS live vaccine.Methods Lantang pigs, Juema, Bama and Wuzhishan ( white) minipigs were inoculated with virulent strain NVDC-JXA1 of PRRSV, and local binary hybrid pigs were used as control.The animals were continuously observed for 5 weeks on mental status, appetite, survival, etc.after inoculation of virus.The dead pigs were autopsied and the lung tissue samples were collected for detecting virus by RT-PCR.By the end of the experiment, serum of survival animals were collected for detecting PRRSV antibody by ELISA assay.Result The animals showed depression, anorexia, and other clinical signs and death in each group after inoculation.Meanwhile, the testing results were all positive in the RT-PCR and ELISA detection.Bama and Wuzhishan ( white) minipigs were the most sensitive to virulent strain NVDC-JXA1 of PRRSV regarding mortality rate.Conclusions Bama and Wuzhishan ( white) minipigs are sensitive to HP-PRRSV, and can be used for the inspection of HP-PRRS live vaccine.

Interacción Candida albicans-Hospedero: un proceso complejo en el que la inmunidad innata juega un importante papel / Candida albicans-host interaction: a complex process in which the innate immunity play an important role

Candida albicans es una levadura comensal, que bajo ciertas circunstancia puede convertirse en patógeno. La capacidad de cambiar se forma constituye un factor de virulencia, permitiéndole a la levadura invadir y diseminarse. El sistema inmune innato es capaz de reconocer las diferentes morfologías de C. albicans activando receptores (PRRs) que reconocen PAMPs o patrones moleculares conservados. Las células inmunes más importantes son los macrófagos y los neutrófilos que desencadenan una respuesta efectora a través de la fagocitosis y la activación de estrés oxidativo contra C. albicans. Las células dendríticas, por su parte expresan la mayoría de los PRRs involucrados en el reconocimiento de C. albicans activando la secreción de citoquinas hacia una respuesta tipo TH1 (inducida por INF tipo1, IL-12, INFγ, ), Treg (inducida por TGFβ, IL-10) y TH17 (inducida por IL-23, IL6). Las células epiteliales no sólo constituyen una barrera física frente a la infección por C. albicans, sino son fundamentales en el reconocimiento primario de este microorganismo. Mediante una respuesta bifásica estas células activan diferencialmente, vías de transducción de señales que determinan que no se active una respuesta de citoquinas frente a la presencia de blastoconidios (forma comensal) y que se active frente a presencia de hifas (forma invasora). Por su parte, C. albicans es capaz de desarrollar mecanismos evasivos de la respuesta inmune. Esta compleja interacción y hongo-hospedero determina si el hospedero será capaz de eliminar a este microorganismo o si éste finalmente invadirá expresando su virulencia.

Candida albicans is a comensal microorganism that under certain circumstances is able to transform into a pathogen. This ability to switch constitute a virulence factor that C. albicans uses to invade and spread. The innate immune system recognize the different forms of C. albicans activating receptors (PRRs) that recognize PAMPs or conserved molecular patterns. The most important immune cells are macrophages and neutrophils that generate an effector response through phagocytosis and oxidative burst against C. albicans. Dendritic cells express the most of PRRs involved in the recognition of C. albicans activating the cytokines synthesis forward to a TH1 (induced by INF tipo1, IL-12, INFγ), Treg (induced by TGFβ, IL-10) y TH17 (induced by IL- 23, IL6) immune response. Epithelial cells not only constitute a physical barrier against C. albicans, but they are crucial in the first recognition of this microorganism. Through a biphasic response these cells differentially activate pathways that determine a no cytokine response when blastoconidia (commensal form) is present and an inflammatory response in presence of hyphae (pathogenic form). C. albicans develops immune evasive mechanism. This complex host-fungi interaction determines if the host will eradicate the infection or if this microorganism will invade the host, expressing it virulence.

Vacunas contra el virus del síndrome reproductivo y respiratorio porcino (PRRSV): escribiendo una historia / Vaccines against porcine reproductive and respiratory syndrome virus (PRRSV): writing a history

The porcine reproductive and respiratory syndrome virus (PRRSV) constitutes one of the most serious problems of the pig industry in the world. It affects pigs of all ages and causes reproductive and respiratory disorders that create great economic losses. These increase due to the rapid spread of the disease and the inefficiency of the commercial vaccines. Modified live and killed vaccines are the two types of commercial vaccines currently available on the market for American and European strains. Although these vaccines can reduce disease symptoms and viremia, they do not prevent infection in any way and cross-presentation is variable against heterologous virus. Furthermore, it has been reported that the vaccine's virus can spread to other susceptible animals. For these reason, many studies focused on the development of new vaccines that include different vectors for the development of DNA vaccines by evaluating various structural proteins. The use of PRRSV infectious clones, as well as the construction of viral chimeras between the vaccine virus and highly infectious virus have also been evaluated. All these strategies have failed to develop an effective vaccine against PRRSV; therefore it remains as a major challenge.

El virus del síndrome reproductivo y respiratorio porcino (PRRSV) constituye uno de los problemas más grandes de la industria porcina. Afecta a cerdos de todas las edades, causa problemas reproductivos y respiratorios que generan pérdidas económicas millonarias. Éstas van en aumento debido a la rápida diseminación del virus y a la poca eficiencia de las vacunas comerciales para su tratamiento. Actualmente existen dos tipos de vacunas contra el PRRSV: una utiliza el virus atenuado y otra el virus inactivado. Existen en el mercado ambos tipos, tanto para cepas americanas como europeas. Aun cuando dichas vacunas disminuyen los síntomas de la enfermedad y la viremia, no evitan la infección y la protección cruzada es variable frente a virus heterólogos. Además, se ha informado que el virus vacunal puede diseminarse a otros animales susceptibles. Por esta razón muchos estudios se han orientado al desarrollo de nuevas vacunas que incluyen diferentes vectores para el desarrollo de vacunas de ADN evaluando varias proteínas estructurales. También se ha evaluado el empleo de clones infecciosos de PRRSV, así como la construcción de quimeras virales entre el virus vacunal y un virus altamente infeccioso. Todas estas estrategias no han logrado desarrollar una vacuna eficaz contra el PRRSV, por lo que dicho propósito sigue siendo un reto muy importante.

Prevalencia y factores de riesgo asociados con el virus del síndrome reproductivo y respiratorio porcino en sementales de granjas porcinas en el sureste de México / Prevalence and risk factors associated with the porcine reproductive and respiratory syndrome virus in boars of pig farms at the southeastern Mexico

Los objetivos de este estudio fueron estimar la prevalencia y determinar algunos factores de riesgo asociados con el virus del Síndrome Reproductivo y Respiratorio Porcino (PRRS) en sementales de granjas porcinas de Yucatán, México. El estudio se realizó en 30 granjas con 28 a 2.000 vientres y con diferentes niveles de tecnificación. Los verracos al llegar a la granja fueron sometidos a un periodo de adaptación al manejo y al estatus sanitario. En las granjas se empleaba la monta natural, la inseminación artificial o ambas técnicas. La alimentación de los verracos consistió en alimento comercial. Se realizó un estudio transversal por conglomerados desde septiembre 2005 hasta febrero 2006, muestreándose 170 verracos. La presencia de anticuerpos y partículas virales se determinó mediante las pruebas de ELISA y RT-PCR (Reacción en Cadena de la Polimerasa- Transcriptasa Reversa), respectivamente. Cincuenta y seis verracos fueron positivos a la prueba de ELISA, 21 a la prueba de RT-PCR y 67 a una o ambas pruebas. La prevalencia en los verracos fue 39,4% (67/170) y dentro de granjas varió de 0 a 100%. Veintiún granjas tuvieron al menos un animal positivo a la prueba de ELISA, 13 a la prueba de RT-PCR y 25 a una o ambas pruebas. El riesgo de un animal positivo a PRRS fue 3,6 veces mayor en granjas positivas al virus de PRRS y 2,6 veces mayor en granjas donde no se realizaban pruebas diagnósticas antes de introducir los sementales. Las granjas que utilizaban sementales en la detección de celo tuvieron 7,0 veces mayor riesgo de un verraco positivo al virus de PRRS.

The objectives of this study were to estimate the prevalence of and to determine some risk factors associated with the Porcine Reproductive and Respiratory Syndrome (PRRS) virus in boars from pig farms in Yucatan, Mexico. The study was carried out in 30 farms with 28 to 2,000 sows and different levels of technology. On arrival, boars were kept for an adaptation period to the farm conditions and management. Natural mating, artificial insemination or both techniques were used. Commercial feed was provided to the boars. A cluster cross-sectional study was carried out from September 2005 to February 2006, and 170 boars were sampled. The presence of antibodies and virus particles was determined using ELISA and RT-PCR (Reverse transcriptase- polymerase chain reaction) tests respectively. Fifty six out of 170 boars were positive to ELISA test, 21 to RT-PCR and 67 to one or both tests. Boar prevalence was 39.4% (67/170) and within farm varied from 0 to 100%. Twenty one farms (70%) had at least one ELISA positive animal, 13 positives to RT-PCR test (43.3%) and 25 (83.3%) to one or both tests. The risk of a PRRS virus positive boar was 3.6 times greater in the positive farms and 2.6 when no diagnostic tests were carried out before introducing the boar to the farm. Farms using the boars for estrus detection had 7.0 times greater risk of a PRRS virus seropositive boar.

Presencia de animales seropositivos al síndrome reproductivo y respiratorio porcino en Nuevo León / Presence of seropositive animals to the porcine respiratory and reproductive syndrome virus in Nuevo Leon

A preliminary study was carried out to obtain serological evidence of the presence of the porcine respiratory and reproductive syndrome (PRRS) virus and the risk of infection in different areas and phases of production in five full cycle pig farms in the state of Nuevo Leon, Mexico. Sixty blood samples of each farm were obtained (10 for each phase of production: weaning, growing, finishing, pregnancy, lactation and mating-service). The detection of antibodies against the PRRS virus was carried out using a commercial kit. All farms were positives in all phases of production. The highest seroprevalences were found in the growing and finishing phases (36% y 56%, respectively).

Se realizó un estudio preliminar para obtener evidencia serológica de la presencia del virus del síndrome respiratorio y reproductivo porcino (PRRS, por sus siglas en inglés) y del riesgo de infección en las diferentes áreas y etapas de producción en cinco granjas porcinas de ciclo completo en Nuevo León, México. Se obtuvieron 60 muestras de sangre de cada granja (diez para cada etapa de producción: destete, iniciación, finalización, cerdas gestantes, lactando o en monta-servicio). La detección de anticuerpos antivirus del PRRS se realizó utilizando un equipo comercial. Todas las granjas resultaron positivas en las diferentes etapas de producción. La seroprevalencia fue mayor en las etapas de inicio y finalización (36% y 56%, respectivamente).

Seroprevalence of Antibody to Procine Reproductive and Respiratory Syndrome Virus in Diagnostic Submissions

During the period from January to December of 2001, a total of 3,391 swine sera were submitted to our laboratory from 256 farms for the diagnosis of porcine reproductive and respiratory syndrome (PRRS). The antibody to porcine reproductive and respiratory syndrome virus (PRRSV) was tested by the indirect immunofluorescent antibody (IFA) test. Of the 256 farms tested, 230 farms (89.8%) were positive for the PRRSV antibody. The overall seroprevalence of the PRRSV antibody was 52.1% (1765/3391). Most of the pigs seemed to be infected with PRRSV at around 50 to 60 days old. The seroprevalence of the antibody became higher with age, and peaked at around 100 days old. More than one-third of the adult pigs, including boars, gilts, and sows, was positive for the PRRSV antibody. The infection of PRRSV was chronic and confined to growers and/or finishers in most farms. However, the antibody was detected in all production phases at some farms.

Expression of Open Reading Frame 5 Protein of Porcine Reproductive and Respiratory Syndrome Virus Using Semliki Forest Virus Expression System

The ORF5 gene encodes a major envelope glycoprotein (GP5), which is one of the three major proteins of porcine reproductive and respiratory syndrome virus (PRRSV). The GP5 protein has been known to be a 24.5-26kDa N-glycosylated envelope protein. The GP5 is involved in inducing neutralizing antibodies. For this reason, the GP5 is primary candidate for the PRRSV subunit vaccine. To produce the native form of GP5 in mammalian cells, we have cloned the ORF5 gene from PRRSV CNV-1 into the Semliki Forest virus (SFV)-based expression vector, resulting in recombinant pSFV-ORF5. By the infection with recombinant pSFV-ORF5 to BHK-21 cells, the GP5 expression was confirmed by immunocytochemistry and immunoblotting assay. The recombinant virus particle harboring ORF5 gene was infectious to BHK-21 and MARC-145. The RNA synthesis and expression of GP5 in the infected cell was also confirmed by RT-PCR.

Study on targets for several traditional medical formula in innate immunity against M.tuberculosis / 中国免疫学杂志

Objective:To explore the potential targets for Traditional Chinese Medical Formula against tuberculosis infection.Methods:U937 cells were incubated with the drug-serum prepared with Clehnia root Ophioponis Decoction(COD),Lily Metal-Consolidating Decoction (LMCD) and Radix Ginseng Gecko Powder(RGGP) for 24 hours.The mRNA and protein level of CR1,CR3,CD14,CD43,TLR2 and TLR4 expressed in macrophages respectively were then measured by RT-PCR and FACS.The phagocytic and killing activity of the U937 cells interfered with the drug-serum for 24 h were then detected after BCG stimulation.Meanwhile,secretion of IL-10,IL-12 and the activity of iNOS in the macrophages were detected by ELISA and biochemistry method in each group.Results:The drug-serum prepared with COD,LMCD and RGGP had different regulatory effects on the expression of PRRs and the production of IL-10 and IL-12 in the macrophages,and could enhance the phagocytosis percentage and phagocytic index(P