Mechanical transmission of porcine reproductive and respiratory syndrome virus throughout a coordinated sequence of events during warm weather

Mechanical transmission of porcine reproductive and respiratory syndrome virus (PRRSV) throughout a coordinated sequence of events that replicated common farm worker behavior during warm weather (10°C to 16°C) was assessed using a field-based model. The model involved fomites (boots and containers), vehicle sanitation, transport, and personnel movement. In a previous study, the model successfully demonstrated mechanical transmission of PRRSV in 8 out of 10 replicates during cold weather. A field strain of PRRSV was inoculated into carriers consisting of soil samples, which were adhered to the undercarriage of a vehicle. The vehicle was driven approximately 50 km to a commercial truck washing facility where the driver's boots contacted the carriers during washing, introducing the virus to the vehicle interior. The vehicle was then driven 50 km to a simulated farm site, and the driver's boots mechanically spread virus into the farm anteroom. Types of containers frequently employed in swine farms contacted drippings from the footwear on the anteroom floor. The truck wash floor, vehicle cab floor mats, boot soles, anteroom floor, and the ventral surface of containers were sampled to track the virus throughout the model. Ten replicates were conducted, along with sham-inoculated controls, and control replicates. In 2 replicates, infectious PRRSV was detected on the anteroom floor and in 1 replicate, infectious PRRSV was detected on the surface of the container by swine bioassay. All sham-inoculated controls and protocol controls were negative. These results indicate that mechanical transmission of PRRSV throughout a coordinated sequence of events in warm weather can occur, but in contrast to data from studies conducted during cold weather, it appears to be a relatively infrequent event.     

Evaluation of 4 intervention strategies to prevent the mechanical transmission of porcine reproductive and respiratory syndrome virus

Four intervention strategies were tested for their ability to prevent the mechanical transmission of porcine reproductive and respiratory syndrome virus (PRRSV): the use of disposable plastic boots to prevent contamination of personal footwear, the use of boot baths to disinfect PRRSV-contaminated plastic boots, the use of plastic slatted (Polygrate) flooring in the anteroom to prevent PRRSV contamination of incoming personal footwear, and the use of bag-in-a-box shipping methods to prevent PRRSV contamination of the contents of a container destined for a swine farm. Ten PRRSV-positive replicates and 10 PRRSV-negative (sham-inoculated) replicates were used for each strategy. Swabs were collected from selected sites and tested by TaqMan polymerase chain reaction for PRRSV RNA and by swine bioassay to confirm the presence of infectious PRRSV. Results indicated that the use of disposable boots, bleach boot baths or bag-in-a-box shipping methods was highly efficacious in preventing mechanical transmission of PRRSV. In contrast, the use of Polygrate flooring in the anteroom did not prevent contamination of personal footwear. The numbers of PRRSV-positive samples from the Polygrate surface and the soles of incoming footwear placed directly on the Polygrate surface were not significantly different (P = 0.24) from those of footwear that directly contacted the floor of the contaminated anteroom. Although these results are promising, this study should be considered a pilot project and the intervention strategies not considered biosecurity protocols. The model used may or may not represent field conditions. Therefore, the information should be used to develop larger experimental studies, with sufficient statistical power, in combination with field-based epidemiologic studies to better assess the role of mechanical transmission of PRRSV under field conditions.     

Transmission of agents of the porcine respiratory disease complex (PRDC) between swine herds: a review. Part 2--Pathogen transmission via semen, air and living/nonliving vectors

The transmission of PRDC-pathogens (PRRSV, influenza virus A, PCV2, M. hyopneumoniae, A. pleuropneumoniae) between swine herds, which was summarized in the first part of the review, mainly occurs via pig movement. The risk of pathogen transmission by insemination with contaminated semen plays only a relevant role in the infection with PRRSV and PCV2. A risk of the aerogen transmission of pathogens between herds within a distance of 2 to 3 km is described for M. hyopneumoniae and PRRSV. Evidence for the other pathogens is not investigated. The PRDC-pathogens are frequently detected in wild boar populations. Therefore, the transmission between wild boars and domestic pigs seems possible by close contacts. PRRSV and M. hyopneumoniae can be transmitted by contaminated clothes and boots, but the use of sanitation protocols appears to limit their spread. Live vectors like rodents or birds seemed to have no special importance for the transmission of PRDC-pathogens.     

An assessment of sanitation protocols for commercial transport vehicles contaminated with porcine reproductive and respiratory syndrome virus

The objective of this study was to develop and test a rapid (< 2 h) sanitation protocol designed for porcine reproductive and respiratory syndrome virus (PRRSV) positive commercial transport vehicles involving cold water washing and disinfection via fumigation using scale models of weaned pig trailers. The study consisted of 2 phases. Following experimental contamination of model trailers with PRRSV MN 30-100 (5 x 10(5)TCID50), phase 1 evaluated the presence or absence of PRRSV RNA by polymerase chain reaction (PCR) on swabs collected from the trailer interiors 0, 60, and 90 min after treatment. Phase 2 consisted of evaluating the infectivity of trailers 90 min posttreatment by monitoring changes in the PRRSV-status of naive sentinel pigs housed for 2 h. Treatments included washing only (treatment 1), washing plus formaldehyde fumigation (treatment 2), washing plus fumigation with glutaraldehyde-quaternary ammonium chloride (treatment 3), and washing plus overnight drying (treatment 4). Porcine reproductive and respiratory syndrome virus RNA was detected in all trailers (20 out of 20 replicates) at 60 and 90 min following the application of treatments 1 and 2. These trailers also contained infectious PRRSV, as determined by the infection of naive pigs housed in treated trailers and the testing of organic debris collected from the interior of trailers by swine bioassay. At 90 min posttreatment, all trailers treated with glutaraldehyde-quaternary ammonium chloride were PCR-negative, non-infectious to sentinel pigs, and swine bioassay negative. Similar results were observed in trailers allowed to dry for 8 h. Under the conditions of this study, it appears certain disinfectants may possess different levels of efficacy against PRRSV and PRRSV-positive models may be effectively sanitized in the absence of overnight drying.     

家蝇体内猪繁殖与呼吸综合征病毒的初步分离与鉴定

为调查养猪场环境中家蝇携带猪繁殖与呼吸综合征病毒(PRRSV)情况,2010年采集贵州某猪场家蝇样本,采用RT-PCR方法筛选阳性样本,接种Marc-145细胞分离培养病毒,对分离获得的家蝇样本的PRRSV Nsp2基因进行克隆和序列分析。针对PRRSV N基因家蝇样本扩增出377 bp的特异性片段,阳性家蝇样本接种的Marc-145细胞出现明显CPE现象,针对PRRSV Nsp2基因细胞培养物扩增出1064 bp的特异性片段,测序结果显示,家蝇样本的细胞培养物的PRRSV Nsp2基因片段与贵州猪场猪源PRRSV流行株相比具有较高的核苷酸同源性,高达97.1%~98.5%。从而初步推断家蝇也可能成为PRRSV携带者。     

Spatial dispersal of porcine reproductive and respiratory syndrome virus-contaminated flies after contact with experimentally infected pigs

OBJECTIVE: To determine whether flies can acquire porcine reproductive and respiratory syndrome virus (PRRSV) and disperse the virus throughout a designated area. ANIMALS: 60 four-month-old pigs. PROCEDURE: On day 0, 28 of 60 pigs were inoculated with PRRSV MN 30-100 (index variant). On the same day, 100,000 pupae of ochre-eyed houseflies and 100,000 pupae of red-eyed (wild-type) houseflies were placed in the swine facility for a release-recapture study. Flies were recaptured at 2 locations within the swine facility, 6 locations immediately outside the facility, and 30 locations 0.4, 0.8, 1.3, 1.7, 1.9, and 2.3 km from the facility. Traps were emptied on days 2, 7, 8, 10, and 14. Samples derived from flies were tested by use of a polymerase chain reaction assay, virus DNA was sequenced, and viruses were tested for infectivity by means of a swine bioassay. RESULTS: PRRSV RNA homologous to the index PRRSV was detected in trapped flies collected inside and immediately outside the facility and from 9 of 48 samples collected at 0.4 km, 8 of 24 samples collected at 0.8 km, 5 of 24 samples collected at 1.3 km, and 3 of 84 samples collected at > 1.7 km from the facility. Two samples collected at 0.8 km contained genetically diverse variants of PRRSV. Swine bioassays revealed the virus in flies was infectious. CONCLUSIONS AND CLINICAL RELEVANCE: Flies appeared to become contaminated with PRRSV from infected pigs and transported the virus > or = 1.7 km. Fly-born transmission may explain how PRRSV is seasonally transported between farms.     

Further assessment of fomites and personnel as vehicles for the mechanical transport and transmission of porcine reproductive and respiratory syndrome virus

This study re-evaluated the role of fomites and personnel in the mechanical transport and transmission of porcine reproductive and respiratory syndrome virus (PRRSV) between pig populations. Swabs were collected from hands, boots, coveralls, and other fomites following contact with infected pigs and compared with identical samples collected in the absence of PRRSV exposure. Naïve pigs were provided contact with contaminated fomites/personnel and blood tested periodically post-exposure [positive exposure population (PEP)] and compared with populations that did not gain exposure via these routes [negative exposure population (NEP)]. The majority of swab samples from hands, coveralls, and boots from personnel and fomite samples (cable snare and bleeding equipment) following contact with the PRRSV-infected Source Population. Transmission of PRRSV to the PEP was observed (7/7) cases but not in the NEP. In conclusion, under the proper conditions, transport and transmission of PRRSV by fomites and personnel may occur between swine populations in the absence of intervention.     

Ultrastructural pathogenesis of the PRRS virus

Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) was first isolated in swine alveolar macrophages (SAMs) and has subsequently been reported to replicate in other cell lines. Entry of the virus inside the cell takes place by receptor-mediated endocytosis. Following the entry of the virus into the cell, several not completely understood changes take place. PRRSV has been reported to be an apoptotic-inductor virus both in vivo and in vitro. Interestingly, it has been suggested that PRRSV-induced apoptosis occurs in cells other than those in which PRRSV replicates by a bystander mechanism. In this paper the ultrastructural pathogenesis of PRRSV will be reviewed.     

某规模化猪场伪狂犬病的诊断

河南平顶山某猪场母猪出现较严重的流产和产死胎现象,且50日龄~70日龄仔猪出现神经症状,根据临床表现初步诊断为伪狂犬病。为排除猪繁殖与呼吸综合征和猪瘟,进行了实验室诊断。应用ELISA方法检测发病保育猪及母猪血清的伪狂犬病病毒野毒株gE抗体,并对发病仔猪病料进行了伪狂犬病病毒(PRV)、猪繁殖与呼吸综合征病毒(PRRSV)和猪瘟病毒(CSFV)的实时荧光定量PCR检测。结果显示,伪狂犬病病毒野毒抗体阳性,实时荧光定量PCR检测确定仔猪病料中PRV核酸阳性,PRRSV和CSFV核酸阴性。结合临床症状及实验室检测,确诊该猪场发生的是猪伪狂犬病。     

Identification of genetically diverse sequences (ORF 5) of porcine reproductive and respiratory syndrome virus in a swine herd.

The ability of genetically diverse strains of porcine reproductive and respiratory syndrome virus (PRRSV) to coexist in a 1750-sow farm was assessed through the case study describing a chronically infected farm, and also by an animal experiment involving the use of swine bioassay. The case study employed a program of monitoring sera from suckling, nursery, and finishing pigs for the presence of PRRSV by polymerase chain reaction (PCR) and virus isolation (VI). The swine bioassay tested homogenates, consisting of lymphoid and pulmonary tissues, collected from 60 breeding animals from the same farm. The open reading frame (ORF) 5 portion of selected positive PRRSV detected from sera or tissues were nucleic acid sequenced and their phylogenies compared. The results indicated the presence of 3 genetically diverse groups, designated PRRSV-A, -B, and -C. Sequence heterology ranged from 5.8 to 11% between groups. Sequence homology ranged from 98.7 to 99.8% within groups. Swine bioassay verified the presence of PRRSV-A in 1 of 60 animals, and no evidence of strains B or C were detected. This paper indicates that based on the evaluation of ORF 5, genetically diverse strains of PRRSV appear to coexist, although the frequency and significance of this observation is not understood.     

Isolation and Identification of Highly Pathogenic PRRSV from a Pig Farm

[Objective] To identify suspected cases of highly pathogenic PRRSV from a pig farm. [Method] The suspected cases of highly pathogenic PRRSV were conducted pathologic anatomy, the gene of PRRSV N protein was amplified by RT-PCR and its sequences were determined and analyzed. [Result] The nucleotide sequence of N protein of N7/N8 isolate shared the consistency of 97.0% with highly pathogenic strains(JXA1, Hu N4, SX-1 and TJ), and that of isolate N9 shared the consistency of 97.3% with highly pathogenic strains, indicating the infected virus in the pig farm was highly pathogenic PRRSV. [Conclusion] This research provides a reference for the diagnosis of highly pathogenic PRRSV infec-tion in swine production.     

Mechanical transmission of porcine reproductive and respiratory syndrome virus by mosquitoes, Aedes vexans (Meigen)

The objective of this study was to determine whether porcine reproductive and respiratory syndrome virus (PRRSV) could be transmitted to naïve pigs by mosquitoes following feeding on infected pigs. During each of 4 replicates, mosquito-to-pig contact took place on days 5, 6, and 7 after PRRSV infection of the donor pig. A total of 300 mosquitoes [Aedes vexans (Meigen)] were allowed to feed on each viremic donor pig, housed in an isolation room. After 30 to 60 s, feeding was interrupted, and the mosquitoes were manually transferred in small plastic vials and allowed to feed to repletion on a naïve recipient pig housed in another isolation room. Prior to contact with the recipient pig, the mosquitoes were transferred to clean vials. Swabs were collected from the exterior surface of all vials, pooled, and tested for PRRSV. Separate personnel handled the donor pig, the recipient pig, and the vial-transfer procedure. Transmission of PRRSV from the donor to the recipient pig occurred in 2 out of 4 replicates. The PRRSV isolated from the infected recipient pigs was nucleic-acid-sequenced and found to be 100% homologous with the virus used to infect the donor pigs. Homogenates of mosquito tissues collected in all replicates were positive by either polymerase chain reaction or swine bioassay. All control pigs remained PRRSV negative, and PRRSV was not detected on the surface of the vials. This study indicates that mosquitoes (A. vexans) can serve as mechanical vectors of PRRSV.     

Investigation of biosecurity risks associated with the feed delivery: A pilot study

This study explored potential biosecurity issues related to the delivery of feed to commercial farms. A pilot study was conducted to collect information about the day-to-day feed delivery, including biosecurity concerns at the level of the feed truck, the driver, and the farm. In addition, a reusable rubber boot was tested in an effort to increase the proportion of farms at which truck drivers wore clean footwear, and to explore an alternative to the standard plastic disposable boots that may be unsafe in winter conditions. Most farms did well in terms of proper dead-stock management and keeping the farm lane and feed bin areas clean. The provision of reusable rubber boots significantly increased the proportion of deliveries in which the driver wore clean footwear.     

Oral fluid samples used for PRRSV acclimatization program and sow performance monitoring in endemic PRRS-positive farms

An effective gilt acclimatization program is one of the most important management strategies for controlling porcine reproductive and respiratory syndrome virus (PRRSV) infection. Recently, oral fluid samples have been used as alternative diagnostic samples for various swine diseases. This study utilized oral fluids for PRRSV monitoring during the gilt acclimatization period in PRRSV endemic farms. The study was performed in two selected commercial breeding herds (farm A and farm B). PRRSV RNA and PRRSV-specific antibodies were monitored using oral fluid and serum samples. Sow performance parameters related to PRRSV infection were recorded and assessed. After PRRSV exposure during acclimatization, viral RNA was demonstrated in oral fluids from 1 to 10 weeks post-exposure (WPE). PRRSV RNA was detected in serum at 1 and 4 WPE in farm A and at 1, 4, 8, and 12 WPE in farm B. Prolonged viremia of gilts from farm B was possibly due to re-infection (within the herd) and later, reproductive problems were found in the breeding herd. The correlation of PRRSV RNA concentration in oral fluids and serum was evident. The S/P ratio values of PRRSV antibodies in oral fluid samples were higher and had similar patterns of antibody responses to the serum samples. The results suggest that the use of oral fluid samples for PRRSV monitoring during gilt acclimatization in endemic farms is effective, convenient, practical, and economical and would be most beneficial when used with other parameters.     

Further assessment of houseflies (Musca domestica) as vectors for the mechanical transport and transmission of porcine reproductive and respiratory syndrome virus under field conditions

The purpose of this study was to evaluate the potential for houseflies (Musca domestica) to mechanically transport and transmit porcine reproductive and respiratory syndrome virus (PRRSV) between pig populations under controlled field conditions. The study employed swine housed in commercial livestock facilities and a release-recapture protocol involving marked (ochre-eyed) houseflies. To assess whether transport of PRRSV by insects occurred, ochre-eyed houseflies were released and collected from a facility housing an experimentally PRRSV-inoculated population of pigs (facility A) and collected from a neighboring facility located 120 m to the northwest that housed a naïve pig population (facility B). All samples were tested for PRRSV RNA by polymerase chain reaction (PCR). To assess transmission between the 2 populations, blood samples were collected from naïve pigs in facility B at designated intervals and tested by PCR. A total of 7 replicates were conducted. During 2 of 7 replicates (1 and 5), PCR-positive ochre-eyed houseflies were recovered in facility B and pigs in this facility became infected with PRRSV. Chi-squared analysis indicated that the presence of PRRSV in an insect sample was significantly (P = 0.0004) associated with infection of facility B pigs. Porcine reproductive and respiratory syndrome virus was not recovered from other reported routes of transmission during the study period, including air, fomites, and personnel. In conclusion, while an insufficient number of replicates were conducted to predict the frequency of the event, houseflies may pose some level of risk for the transport and transmission of PRRSV between pig populations under field conditions.     

猪繁殖与呼吸综合征病毒和猪瘟病毒混合感染的检测

2005年3月,江苏某猪场仔猪发生体温升高,呼吸困难,四肢末端、耳尖发绀,站立不稳和淋巴结出血为主要症状的疾病。4头发病仔猪的淋巴结、脾、肺脏组织用RT-PCR方法分别检测猪繁殖与呼吸综合征病毒和猪瘟病毒为阳性,用猪瘟ELISA试剂盒检测猪瘟病毒野毒为阳性。结合本病的临床症状和病理剖检,病例确诊为猪繁殖与呼吸综合征和猪瘟野毒的混合感染。     

新疆北疆地区某规模化猪场主要传染病抗体检测与分析

为了解新疆北疆地区某规模化猪场几种主要传染病的抗体水平,便于及时发现猪场潜在的疾病风险。本试验采用间接酶联免疫吸附试验（ELISA）方法对某规模化猪场各阶段猪群进行猪瘟病毒（CSFV）抗体、猪繁殖与呼吸障碍综合征病毒（PRRSV）抗体、猪圆环病毒2型（PCV2）抗体、猪O型口蹄疫病毒（FMDV-O）抗体,及猪伪狂犬病病毒（PRV）gB与gE蛋白抗体进行检测。试验结果表明,该场的CSFV、PRRSV、PCV-2、FMDV-O、PRV-gB蛋白、PRV-gE蛋白的平均抗体阳性率分别为83.57%、90.56%、90.29%、71.86%、82.84%、18.29%;不同类别猪群间抗体水平参差不齐,种猪群的PRRSV抗体阳性率仅为52.63%,保育猪群的CSFV抗体阳性率仅为44.12%,育肥猪群的PRV-gB抗体阳性率为30.00%,种公猪的PRV-gE抗体阳性率为30.00%,育肥猪群的FMDV-O抗体阳性率为6.25%,基于以上试验结果,为该场免疫程序的制定和优化提供参考依据,以期达到有效控制及逐渐净化疫病的目的。     

一例猪圆环病毒2型和大肠杆菌、溶血葡萄球菌混合感染的诊断

山东德州某猪场发生猪高热、呼吸系统疾病甚至死亡的疫情。采集病料提取病变组织总DNA或RNA进行猪繁殖与呼吸综合征病毒、猪圆环病毒2型、猪细小病毒、猪伪狂犬病病毒、猪瘟病毒的PCR或RT-PCR检测。PCR扩增出353 bp的猪圆环病毒2型特异性条带。同时进行细菌分离培养、生化鉴定等试验,诊断为猪圆环病毒2型和大肠杆菌、溶血葡萄球菌混合感染。     

Diagnosis and Pathogen Analysis of Mixed Infection of Porcine Reproductive and Respiratory Syndrome Virus,Porcine Circovirus Type 2 and Haemophilus parasuis

To identify the causative agent of porcine respiratory disease complex (PRDC) occurred at a pig farm in Kaifeng city,Henan province,we identified the potential causative bacteria of the morbid nursing piglets by bacterial test and drug sensitivity test of the clinical samples (lung,liver and spleen),and detected the common potential pathogens causing PRDC diseases by PCR and RT-PCR assays,including porcine reproductive and respiratory syndrome virus (PRRSV),porcine circovirus type 2 (PCV2),swine influenza virus (SIV),pseudorabies virus (PRV),classical swine fever virus (CSFV) and Mycoplasma hyopneumoniae (Mhp),and then sequcing and phylogenetic analysis of the structural gene of positive causative pathogens were carried out.The results showed that a Haemophilus parasuis (Hps) strain were isolated and identified by the observation of bacterial morphology and satellite phenomenon,and the sequence analysis of 16S rRNA gene.The drug sensitivity test showed that the Hps strain was sensitive to ampicillin,amoxicillin-clavulanic acid,ceftiofur and tetracycline.Meanwhile,PCR and RT-PCR assays indicated that all the samples were positive for PRRSV and PCV2,and named the involved strain as PRRSV/HN-2019 and PCV2/HN11-2019,respectively.Sequencing and phylogenetic analysis of the ORF5 gene of the newly identified PRRSV revealed that the PRRSV/HN-2019 strain was closely related to the NADC30-like strains and grouped into NADC30-like genotype clade.And cap gene of the newly identified PCV2 strain the PCV2/HN11-2019 strain was closely related to the PCV-2d strains and grouped into PCV-2d genotype clade.In conclusion,this study demonstrated that the morbid piglets were co-infected with PRRSV,PCV2 and Hps,which provided a basis for the development of effective control strategies in the pig farm.