Long-term administration of a commercial porcine reproductive and respiratory syndrome virus (PRRSV)-inactivated vaccine in PRRSV-endemically infected sows

The purpose of this study was to investigate the safety and efficacy of a commercial European porcine reproductive and respiratory syndrome virus (PRRSV)-inactivated vaccine after 18-month use in gilts/sows at a farm with high seroprevalence. In a farrow-to-finish farm with 1100 sows, all sows and gilts were systematically vaccinated with the PRRS-inactivated PROGRESSIS vaccine for a period of 18 months. Farm's reproductive and litter characteristics were longitudinally recorded for this period and historically compared with those of the year prior to vaccination. Serology, employing immunoperoxidase monolayer assay, had confirmed a high prevalence of PRRS-specific antibodies in most age groups within the farm prior to vaccination. Seroprevalence during the experiment ranged between 0% and 100% in weaners and growers, but remained at stable high levels (> 93%) in finishing pigs and gilts throughout all 2-year period of serology measurements. No local or systemic vaccine side effects were noted throughout the trial period. Vaccinations had resulted over time in a significant improvement of sow reproductive performance (e.g. reduction of premature farrowings, abortions and increase of farrowing rate) and litter characteristics (e.g. increase of the number of live born and weaned pigs and decrease of stillborn, mummified, weak and splay-legged piglets). It has also been observed that the higher the degree of immunization of a sow, the better the improvement of her reproductive parameters. Sows after vaccination have shown improved characteristics compared to homoparous sows prior to the application of vaccinations in the farm.     

Changes in macrophage and lymphocyte subpopulations of lymphoid tissues from pigs infected with the porcine reproductive and respiratory syndrome virus (PRRSV)

We used an immunohistochemical method to investigate changes in macrophage and lymphocyte subpopulations in various lymphoid tissues of pigs in the acute phase of porcine reproductive and respiratory syndrome virus (PRRSV) infection. The numbers of CD8+ cells and B-cells varied among lymphoid tissues after PRRSV infection. In the infected pigs, numbers of CD8+ cells increased in systemic lymphoid tissues whereas numbers of B-cells increased in mucosa-associated lymphoid tissues. There was no difference in the distribution of virus-infected cells and macrophages between lymphoid tissues of the infected pigs. These changes may be associated with the establishment of virus persistence or the emergence of concurrent infection in mucosal organs.     

猪对PRRSV易感性差异的研究进展

猪繁殖与呼吸综合征是当前危害养猪业最严重的传染病之一,其病原为猪繁殖与呼吸综合征病毒(porcine reproductive and respiratory syndromevirus,PRRSV)。由于PRRSV的不断变异,其感染力和致病力增强,现有疫苗的保护效果不佳,且弱毒活疫苗有变异成新强毒的风险。本文从宿主遗传变异的角度讨论猪对PRRSV的易感性,包括猪免疫应答的变异与易感性、猪对PRRSV易感性的品种(系)间差异以及PRRSV抗性育种亟待解决的问题,试图为PRRS的防制提供新的思路。     

Effects of different us isolates of porcine reproductive and respiratory syndrome virus (PRRSV) on blood and bone marrow parameters of experimentally infected pigs

Seventy five-week-old, crossbred, caesarean-derived, colostrum-deprived pigs were randomly divided into five groups of 14 pigs and assigned one of five treatments: the intranasal inoculation of 1 (5.7) TCID50 of one of four plaque-purified isolates of porcine reproductive and respiratory syndrome virus (PRRSV) (VR2385, VR2431, ISU-984 and ISU-22), or uninfected cell culture and media. Haematological variables were measured for 21 days and bone marrow was analysed when the pigs were killed three, seven, 10, 21 or 28 days after the inoculation. The PRRSV-infected pigs had non-regenerative anaemia and markedly increased myeloid:erythroid ratios from three to 21 days after inoculation. There was a significant (P < 0.05) difference in the severity of the anaemia induced by the four PRRSV isolates; the most highly pneumovirulent strains (VR2385, ISU-984 and ISU-22) induced more severe anaemia than the least virulent isolate (VR2431). The anaemia induced by PRRSV was probably due to a direct or indirect effect on erythroid precursor cells in the bone marrow.     

猪繁殖和呼吸综合征油乳剂灭活苗的研制

使用猪场自行分离的PRRSV,进行细胞转瓶培养,收获病毒液后灭活,乳化制成油乳苗,各项指标检测均合格,并且免疫试验效果较好.     

猪繁殖与呼吸道综合征病毒(PRRSV)单克隆抗体的研制

利用猪繁殖与呼吸道综合征病毒（ＰＲＲＳＶ）国内分离株Ｊ１,采用反复差速离心法制备免疫抗原,长程免疫法免疫ＢＡＬＢ／ｃ小鼠,用间接ＥＬＩＳＡ方法检测抗体,通过细胞融合技术,并经３次亚克隆获得了１０株能稳定分泌抗ＰＲＲＳＶ单抗的杂交瘤细胞单克隆株（Ａ１Ｄ７Ｈ１０,Ａ１Ｄ７Ｈ１１,Ａ１Ｅ７Ｈ９,Ａ１Ｅ７Ｄ９,Ａ２Ｄ８Ｅ７,Ａ２Ｄ８Ｂ１１,Ｂ３Ｄ１１Ｄ６,Ｂ２Ｇ９Ａ９,Ｂ２Ｇ９Ｆ２）。这些细胞经体外连续传     

Elevated serum haptoglobin in pigs infected with porcine reproductive and respiratory syndrome virus.

We examined the two acute phase proteins, alpha (alpha)-1 acid glycoprotein (AGP) and haptoglobin (HP), in serum of pigs following experimental porcine reproductive and respiratory syndrome (PRRS) virus infection. Increased levels of serum HP, but not AGP, were observed from 7 to 21 days post-inoculation in the infected pigs. Furthermore, serum IL-6 increased in the infected pigs, but TNF-alpha did not. The increase of serum IL-6 in pigs following PRRS virus infection may induce production of HP. Also, in the field investigation, serum HP in pigs was dramatically increased after exposure to the PRRS virus.     

Genetics and geographical variation of porcine reproductive and respiratory syndrome virus (PRRSV) in Thailand

The Thai isolates of porcine reproductive and respiratory syndrome virus (PRRSV) were obtained from the Chulalongkorn University-Veterinary Diagnostic Laboratory (CU-VDL). Virus isolation was confirmed by immunoperoxidase monolayer assay (IPMA) using SDOW-17. The virus genotype was determined using nested multiplex RT-PCR (nm RT-PCR) of ORF 1b. The nm RT-PCR was able to detect at least 10TCID50/ml of PRRSV. Of 137 Thai isolates, 66.42% belonged to the European (EU) genotype and 33.58% to the North American (US) genotype. ORF5 products of the eight US strains (00CS1, 01NP1, 01UD6, 02CB13, 02KK1, 02PB1, 02SP2 and 02SP3) and the six EU strains (01CB1, 01RB1, 02BR1, 02CB12, 02SB2 and 03RB1) were sequenced for genetic variation analysis. The US strains of the Thai isolates are clustered within the same group and are more closely related to the IAF-EXP91 from Canada (89-90% nucleotide identity), whereas the EU strains were very similar to the EU prototype, Lelystad virus (87-97.5% nucleotide identity). The ORF5 nucleotide identities within the US genotype tested in this study compared to the US prototype, VR-2332 varied from 83.7 to 85.2%, whereas 83.5-85.5% amino acid identities were found. Based on the phylogenetic tree, each pair of the Thai isolates (01NP1 and 02KK1, 00CS1 and 01UD6, and 01CB1 and 01RB1) was identical despite they were collected from different provinces. Therefore, there was no geographic influence on the spreading of PRRSV in Thailand. Interestingly, 02CB12 (EU genotype) shared over 99% similarity of the ORF5 nucleotide sequence and 98.6% of amino acid identity with the European vaccine, Porcillis (AF378819). However, modified live virus vaccines for PRRSV have not yet been used in the swine population in Thailand. The results suggested that both US and EU genotypes exist in Thailand, genetic variation does occur in both genotypes, and the sources of the viruses appear to be from Canada and Northern Europe, respectively. In addition, the spreading of PRRSV in Thailand might be due to introducing infected replacement pigs or infected semen into the farm.     

Synergism between porcine reproductive and respiratory syndrome virus (PRRSV) and Salmonella choleraesuis in swine

Porcine reproductive and respiratory syndrome virus (PRRSV) and Salmonella choleraesuis are two leading causes of economic loss in the swine industry. While respiratory disease is common in both S. choleraesuis and PRRSV infections, the factors that contribute to its development remain largely undefined. We investigated the interaction of PRRSV, S. choleraesuis, and stress in 5-week-old swine. All combinations of three factors (inoculation with S. choleraesuis on Day 0, PRRSV on Day 3, and treatment with dexamethasone on Days 3-7) were used to produce eight treatment groups in two independent trials. Fecal samples, tonsil and nasal swabs, serum samples and postmortem tissues were collected for bacteriologic and virologic examinations. No clinical signs were observed in pigs inoculated with only PRRSV or only S. choleraesuis. In contrast, pigs which were dually infected with S. choleraesuis and PRRSV exhibited unthriftiness, rough hair coats, dyspnea, and diarrhea. The pigs which received all three treatment factors were the most severely affected and 43% (three of seven) of the animals in this group died. Individuals in this group shed significantly higher quantities of S. choleraesuis in feces and had significantly higher serum PRRSV titers compared to other treatments (p < or = 0.05). In addition, S. choleraesuis and PRRSV were shed longer and by more pigs in this group than other groups and S. choleraesuis was recovered from more tissues in this group on Day 21 post inoculation. These results suggested that PRRSV, S. choleraesuis, and dexamethasone acted synergistically to produce a syndrome similar to that observed in the field.     

Polyclonal activation of B cells occurs in lymphoid organs from porcine reproductive and respiratory syndrome virus (PRRSV)-infected pigs

Porcine reproductive and respiratory syndrome virus (PRRSV) induces a persistent viral infection associated with an inefficient humoral immune response. A study of lymphoid B cells and specific humoral immune response was performed in blood and several lymphoid organs collected from PRRSV experimentally-infected pigs. Groups of specific pathogen-free (SPF) pigs were infected with the LHVA-93-3 isolate of PRRSV, and blood, tonsils, spleen and mediastinal lymph nodes (MLN) were collected at various times postinfection (p.i.) (3-60 days). Lymphoid cells were isolated, immunolabeled for cytofluorometric determination of B cell percentages, used for counting specific anti-PRRSV antibody secreting B cells by an ELISPOT assay, or cultured for metabolic activity. The presence of anti-PRRSV antibodies in the serum of infected pigs was determined using a commercial ELISA assay. Virus detection was performed in all tissues, including lungs, by virus isolation and RT-PCR. The results show that percentages of B cells increased in tonsils as soon as 3 days until 17 days p.i. in PRRSV-infected pigs while they increased in spleen at 3 days p.i. only, due to an increase of larger Ig(high)-producing B cells. Metabolic activity of lymphoid cells from blood and spleen increased at 3 days p.i. only while lymphoid cells from tonsils and MLN transiently decreased at that time and increased thereafter up to 60 days p.i. Anti-PRRSV antibody-secreting B cells occurred in tonsils after 10 days p.i. and strongly increased up to 60 days p.i. However, specific anti-PRRSV-secreting B cells were detected in blood and spleen after 17 days p.i and in MLN only after 45 days p.i. Specific antibodies were detectable in serum at 10 days p.i., reached the maximum level at 45 days and remained high up to 60 days p.i. Infectious virus was detected in lungs and MLN as soon as 3 days p.i., and remained detectable up to 45 days p.i. in tonsils of one pig while viral RNA was detected in most organs up to 60 days p.i. In vitro experiments revealed that inactivated virus induced a stimulation of lymphoid cells isolated from PRRSV-infected pigs while it was cytotoxic for lymphoid cells from control pigs. Taken together, these results indicate that viral infection induced simultaneously a polyclonal activation of B cells, mainly in tonsils, and an exaggerated and prolonged specific humoral immune response due to persistent viral infection in lymphoid organs.     

种公猪精液中猪瘟和蓝耳病病毒混合感染的快速检测

参考GenBank公布的猪蓝耳病病毒(PRRSV)VL2332株、LV株以及猪瘟兔化弱毒(CSFV)C株的基因序列,各设计合成了一对引物,建立了在相同PCR扩增条件下能同时检测PRRSV和CSFV的RT-PCR方法。对2003~2004年期间江苏、浙江、安徽、福建、上海等省市的17个大中型猪场送检的186份种公猪精液进行了检测,结果18份呈PRRSV阳性,24份呈CSFV阳性,其中有11份为PRRSV和CSFV的混合感染,约占送检精液样品的5.91%。试验结果表明,所建立的RT-PCR方法可用于精液中这2种野毒感染的快速鉴定和分子流行病学调查。     

Antiviral effect of dietary germanium biotite supplementation in pigs experimentally infected with porcine reproductive and respiratory syndrome virus

Germanium biotite (GB) is an aluminosilicate mineral containing 36 ppm germanium. The present study was conducted to better understand the effects of GB on immune responses in a mouse model, and to demonstrate the clearance effects of this mineral against Porcine reproductive and respiratory syndrome virus (PRRSV) in experimentally infected pigs as an initial step towards the development of a feed supplement that would promote immune activity and help prevent diseases. In the mouse model, dietary supplementation with GB enhanced concanavalin A (ConA)-induced lymphocyte proliferation and increased the percentage of CD3+CD8+ T lymphocytes. In pigs experimentally infected with PRRSV, viral titers in lungs and lymphoid tissues from the GB-fed group were significantly decreased compared to those of the control group 12 days post-infection. Corresponding histopathological analyses demonstrated that GB-fed pigs displayed less severe pathological changes associated with PRRSV infection compared to the control group, indicating that GB promotes PRRSV clearance. These antiviral effects in pigs may be related to the ability of GB to increase CD3+CD8+ T lymphocyte production observed in the mice. Hence, this mineral may be an effective feed supplement for increasing immune activity and preventing disease.     

Respiratory function and pulmonary lesions in pigs infected with porcine reproductive and respiratory syndrome virus

Pulmonary dysfunction was evaluated in pigs infected with porcine reproductive and respiratory syndrome virus (PRRSV, isolate VR-2332) and compared to clinical and pathological findings. Infected pigs developed fever, reduced appetite, respiratory distress and dullness at 9 days post-inoculation (dpi). Non-invasive pulmonary function tests using impulse oscillometry and rebreathing of test gases (He, CO) revealed peripheral airway obstruction, reduced lung compliance and reduced lung CO-transfer factor. PRRSV-induced pulmonary dysfunction was most marked at 9–18 dpi and was accompanied by a significantly increased respiratory rate and decreased tidal volume. Expiration was affected more than inspiration. On histopathological examination, multifocal areas of interstitial pneumonia (more severe and extensive at 10 dpi than 21 dpi) were identified as a possible structural basis for reduced lung compliance and gas exchange disturbances.     

Immunisation of pigs with a major envelope protein sub-unit vaccine against porcine reproductive and respiratory syndrome virus (PRRSV) results in enhanced clinical disease following experimental challenge

Disease exacerbation was observed in pigs challenged with virulent porcine reproductive and respiratory syndrome virus (PRRSV) following immunisation with a recombinant GP5 sub-unit PRRSV vaccine (rGP5) produced in E. coli. Eighteen animals were divided into three experimental groups: group A were immunised twice IM with rGP5, 21 days apart; group B acted as positive controls (challenged but not immunised); and group C were negative controls. Pigs in groups A and B were challenged 21 days after the second immunisation of the group A animals. Following challenge, three pigs given rGP5 exhibited more severe clinical signs than the positive controls, including respiratory distress and progressive weight-loss. Although not statistically significant, the more severe disease exhibited by group A animals may suggest previous immunisation as a contributory factor. The mechanisms of these findings remain unclear and no association could be established between the severity of disease, non-neutralising antibody concentrations and tissue viral loads.     

间接免疫酶组织化学法检测猪繁殖与呼吸综合征病毒感染和抗原定位

以蔗糖密度梯度离心法提纯的猪繁殖与呼吸综合征（Porcine reproductive and respiratory syndrome,PRRS）病毒SC1株作为抗原,免疫家兔制备兔抗PRRS病毒IgG,成功建立了检测PRRS病毒抗原的间接免疫酶组织化学法。该方法只与PRRS病毒感染猪组织呈现阳性反应,与猪瘟病毒、猪细小病毒、猪伪狂犬病毒、猪乙脑病毒人工感染并致死仔猪的肝脏组织呈现阴性反应。用该方法检测PRRS SC-1株人工感染28日龄仔猪,在感染后7 d即可在肺门淋巴结、胸腺、扁桃体、十二指肠、肺、大脑和肾脏检测到PRRS病毒抗原。PRRS病毒抗原主要分布于胸腺和十二指肠感染细胞的胞浆内、肺门淋巴结小梁周围的淋巴窦和弥散的淋巴组织、扁桃体淋巴结的隐窝及其周边。该法具有特异、直观和敏感的特点,可用于PRRS病毒感染的实验室诊断、抗原定位及甲醛固定样本的回顾性诊断。     

上海地区野鸟和家禽中PRRSV抗体的血清学调查

2005年3～5月，采用ELISA法对来自于上海地区的208份野鸟和357份家禽的血清样品进行了猪繁殖和呼吸综合征病毒（porcine reproductive andrespiratory syndrome virus，PRRSV）抗体的血清学调查。结果表明，受检的5种家禽存有不同程度的抗体阳性率，而野鸟的阳性样品集中于绿头鸭和麻雀。     

Occurrence of swine salmonellosis in postweaning multisystemic wasting syndrome (PMWS) affected pigs concurrently infected with porcine reproduction and respiratory syndrome virus (PRRSV)

Fourteen diseased pigs from four farms in which there had been an outbreak of salmonellosis were investigated. Granulomatous inflammation with depletion of lymphocytes was observed in the swollen lymph nodes in these pigs. Antigens to porcine circovirus type 2 (PCV2) were immunolabeled in the lesions along with detection of viral DNA as PCV2 by polymerase chain reaction (PCR). In addition, antigens to porcine reproductive respiratory syndrome virus (PRRSV) were immunodetected in the lungs and Salmonella Choleraesuis was isolated from the affected pigs. The nine salmonellosis affected pigs, five (55.6%) with salmonellosis and PMWS concurrently infected with PRRSV were much higher than those infected with salmonellosis and PMWS (22.2%) or with salmonellosis and PPPRV (22.2%).     

Oral transmission of porcine reproductive and respiratory syndrome virus by muscle of experimentally infected pigs

The current study was performed to determine if porcine reproductive and respiratory syndrome virus (PRRSV) could be transmitted to pigs by feeding muscle tissue obtained from recently infected pigs. Muscle obtained from pigs infected with either a European strain (EU donor pigs) or American strain (US donor pigs) of PRRSV was fed to PRRSV-free receiver pigs. The donor pigs were slaughtered 11 days post-infection (dpi). PRRSV was detected by conventional virus isolation in muscle at 11 dpi from 7 of 12 EU donor pigs and 5 of 12 US donor pigs. In contrast to conventional virus isolation, all muscle samples from infected pigs were positive for viral nucleic acid by PCR, except for muscle from one animal infected with the American strain of PRRSV. Five hundred grams of raw semimembranosus muscle from each of the donor pigs was fed over a 2 days period (250 g per day) to each of two receiver pigs (48 receiver pigs). The receiver pigs were housed separately in five groups. One of the five groups was fed muscle obtained from US donor pigs that was also spiked with the American strain of PRRSV. Sentinel pigs were placed in-contact with the group of receiver pigs fed spiked muscle. All receiver pigs became viraemic by 6 days post-feeding (dpf). There was evidence of horizontal transmission with sentinel pigs, in-contact with receiver pigs, becoming viraemic. The study demonstrates that PRRSV could be infectious through the oral route via the feeding of meat obtained from recently infected pigs.     

Effect of porcine reproductive and respiratory syndrome virus (PRRSV) on alveolar lung macrophage survival and function

Porcine reproductive and respiratory syndrome virus (PRRSV) recently emerged as an important cause of reproductive disorders and pneumonia in domestic pigs throughout the world. Acute cytocidal replication of PRRSV in alveolar lung macrophages causes the acute pneumonia; however, it remains largely unresolved whether there may also be a predisposition to longer-term local immunodeficiency in the PRRSV-convalescent lung. We applied various flow cytometric techniques to study the interplay between PRRSV replication and macrophage viability/function in pure cultures of porcine alveolar lung macrophages. Monitored by flow cytometric detection of intracellular PRRSV nucleocapsid protein, acute (24 h post infection) PRRSV replication did not impede the ability of alveolar macrophages to ingest fluorescently labelled Escherichia coli. At 48 h post infection, PRRSV-induced cytotoxicity (quantitated by flow analysis of cell size and membrane integrity) led to 40% reduction in the total number of phagocytozing cells. However, viable/uninfected macrophages in PRRSV-infected cultures exhibited normal phagocytic ability at 48 h, indicating that no soluble phagocytosis-suppressive mediators were induced by PRRSV infection in this system. In short, in our minimal system containing only a single cell type, phagocytosis-suppressive effects of PRRSV infection were detected, that acted at the culture level by reducing the total number of alveolar lung macrophages.