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1.
Five groups of eight fattening pigs were vaccinated and then infected with Aujeszky's disease virus. Viral excretion was evaluated by two means: deep nasal swabbing and air sampling. It appeared that infectious airborne virus could be recovered from day 1 to day 6 after infection in the isolated units where control animals were raised. In vaccinated animals, airborne particles were also detected but the amount and duration varied in relation to their immune status at the day of virulent challenge: viral excretion was significantly lower in pigs presenting a high antibody level (1/16 to 1/64) just before infection. Results obtained with nasal swabs and with air samples were closely related. Despite its low sensitivity, the air sampling procedure could be considered as an efficient tool for reflecting infectious viral pressure in a confined atmosphere.  相似文献   

2.
We examined nasal swab and lung homogenate samples collected from pigs experimentally and naturally infected with Mycoplasma hyopneumoniae for the detection of M. hyopneumoniae by the nested PCR (nPCR) and culture methods. In the 23 experimentally infected pigs, M. hyopneumoniae was commonly detected in nasal swabs by the nPCR and culture methods at 4 weeks after inoculation, and there was a significant correlation (P<0.01) between the titers of viable organisms in nasal swabs and in lung homogenates in the experimentally inoculated pigs. In the naturally infected pigs, on the other hand, discrepancies in detection were found between nasal swab and lung homogenate samples in 17 of 36 cases, although the presence of gross lung lesions correlated relatively well with the detection of organisms from the samples. Our results indicated that the diagnosis of mycoplasmal pneumonia by nPCR in individual pigs with nasal swabs is reliable under these experimental conditions. At present, nPCR with nasal swabs should only be used for monitoring the disease status at the herd level under field conditions.  相似文献   

3.
A PCR assay for the detection of Actinobacillus pleuropneumoniae was developed based on the amplification of a dsbE-like gene. All of 157 field isolates of A. pleuropneumoniae reacted in the PCR by the amplification of a 342bp product. No reaction was observed with related bacterial species or other bacterial species isolated from pigs, except for A. lignieresii. The lower detection limit of the PCR was 10(2) CFU per PCR test tube and was not affected by the addition of 10(6) CFU Escherichia coli. The PCR was evaluated on mixed bacterial cultures from nasal and tonsillar swabs as well as suspensions of nasal conchae and tonsils obtained from specific pathogen-free (SPF) pigs, experimentally infected pigs, and pigs from farrow-to-finish herds. The results of the new PCR were compared with a PCR based on the detection of the omlA gene coding for an outer membrane protein, with a commercially available PCR (Adiavet APP, Adiagène, Saint-Brieuc, France), and with conventional culturing. No positive reactions were observed with any of the PCR methods in samples of SPF animals. In samples of the other animals, no or low significant differences between nasal swabs and suspensions as well as tonsillar swabs and suspensions were observed in any method. In general, more positive results were obtained from tonsillar samples in comparison to nasal samples. Interassay sensitivity and specificity values were assessed for each test by pair wise comparisons between assays. The agreement between tests was evaluated by calculating Cohen's kappa coefficient. From these analyses the three PCR assays showed a good agreement. The dsbE-based PCR proved to be highly sensitive (95 and 93%) and specific (82 and 74%) in comparison to the omlA-based PCR and the commercially available PCR, respectively. It was concluded that the dsbE-like gene-based PCR is a reliable diagnostic assay for demonstration of A. pleuropneumoniae. Furthermore, it was demonstrated that tonsillar swabs can be used for the detection of the pathogen in healthy carrier animals.  相似文献   

4.
The purpose of this study was to determine the optimal route of infection and the optimal sampling sites for the recovery of M. hyopneumoniae, the etiological agent of enzootic porcine pneumonia. Virulence of two strains, BQ 14 and 116, isolated in France in 1975 and 2003, respectively, was also compared. Groups of specific pathogen free piglets were experimentally infected by the intratracheal or intranasal route. One non-inoculated pig was placed in each group of infected pigs to study direct transmission. Two groups were kept uninfected. Coughing was recorded daily. Blood samples, nasal, tonsillar and tracheal swabs and tracheobronchiolar washings were collected weekly. Pigs were killed 27-37 days post-infection. Lung lesions were scored and swabs were collected from nasal cavities, tonsils, trachea, lung, liver and spleen. All the samples, collected from live and dead pigs, were cultured for M. hyopneumoniae recovery. Results showed that both experimentally infected pigs and contact pigs developed enzootic pneumonia, whatever the route of infection and the strain tested. Direct contact transmission occurred quickly. No difference between the two routes of infection or between the two strains tested was evidenced, but high individual variations were observed between pigs. Tracheal swabs and tracheobronchiolar washings were the most effective samples to detect M. hyopneumoniae compared to nasal or tonsillar swabs. Our results also suggested that tracheobronchiolar washings could have an influence on the lesion extent observed at necropsy. M. hyopneumoniae could be re-isolated from liver and spleen of experimentally infected pigs and contact pigs.  相似文献   

5.
Hysterectomy-produced colostrum-deprived 5- and 27-day-old pigs were inoculated intramuscularly (IM) or intranasally (IN) with the temperature-sensitive and thymidine kinase-deficient ZHtsTK- strain of Aujeszky's disease virus (ADV), and the nasal swabs and organs of the pigs were periodically collected for virus isolation. No abnormal clinical signs were observed in these pigs, except for a mild febrile response. Viral shedding in the nasal swabs with low titers was detected in the pigs inoculated IN between postinoculation day (PID) 1 and 5, but not in those of the pigs inoculated IM. No contact infection, however, occurred in the cohabiting pigs. Viruses with low titers were isolated only from the muscles and lymph nodes at the site of inoculation in the pigs inoculated IM on PID 2 and 4, but not from any organs of the pigs inoculated IN. To investigate the ability of the ZHtsTK- strain to establish a latent infection in pigs, the pigs inoculated IM or IN with the ZHtsTK- strain were treated with prednisolone. No virus was detected in the trigeminal ganglia or the nasal swabs collected after prednisolone treatment by the cocultivation method. The immunological evaluation demonstrated that immunization of pigs with this strain was effective in preventing clinical signs caused by ADV infection. The duration of virus shedding was markedly shortened in immunized pigs, particularly in those immunized twice and the total quantity of virus recovered from immunized pigs was reduced in comparison with unimmunized pigs.  相似文献   

6.
A high prevalence of MRSA among farm animals, especially pigs, has been observed for some time. However, knowledge on transmission routes of MRSA in livestock production is still scarce. Therefore, the aim of this study was to determine the occurrence of MRSA in pig house air as well as in samples from pigs and their housing environment in 27 MRSA positive pig barns of different sizes and production types. In 85.2% of all barns MRSA was detected in the animal house air. Impingement turned out to be a more sensitive sampling technique than filtration. Other environmental samples such as boot swabs or faeces showed prevalences of MRSA from 55.6% to 85.2% at sample level. The level of MRSA was 88.3% for pooled and 82.1% for single nasal swabs, in skin swabs the one was 87.7%, the others was 78.7%. Spa typing of isolates from air and nasal swabs showed predominantly spa types t011 and t034. MRSA prevalences in pigs as well as in various environmental samples were significantly higher in fattening farms than in breeding farms. This study provides good reference that there could be an airborne transmission of MRSA within pig herds indicating a potential contamination of the environment of barns.  相似文献   

7.
The efficacy of detecting toxigenic Pasteurella multocida from nasal swabs of slaughtered and live pigs was assessed. The isolation of toxigenic P multocida from nasal cavities of slaughtered bacon pigs from two herds with atrophic rhinitis was reduced by immersion in the hot water tank by 25 per cent and 75 per cent. Individual sows from one of the infected herds were repeatedly swabbed to find the best method of isolating toxigenic P multocida. Toxigenic P multocida were isolated from 50 per cent of cotton swabs inoculated on to selective medium the same day. After 24 hours in the post, 45 per cent of cotton swabs placed in transport medium, 38 per cent of alginate swabs dissolved in transport medium and inoculated into mice, and 36 per cent of the dissolved swabs inoculated directly on to selective medium yielded toxigenic P multocida. These bacteria were isolated from only 25 per cent of cotton swabs held in transport medium at 10 degrees C for 48 hours to simulate prolonged postage times; from slaughtered pigs a similar reduction in isolation was seen with swabs kept for 24 or 48 hours. The reduced isolation caused by a delay before culture was associated with an overgrowth of other flora. The development of this flora was prevented by storage of swabs at 4 degrees C in the laboratory or by the use of cool boxes for postage.  相似文献   

8.
The objective of this study was to determine the effect of porcine reproductive and respiratory syndrome virus (PRRSV) infection on porcine circovirus type 2 (PCV2) subtypes a (PCV2a) or b (PCV2b) viremia and shedding characteristics in oral, nasal and fecal samples in experimentally infected pigs. Twenty-three, 2- to 6-week-old pigs were randomly divided into five groups: negative control (n=3), PCV2a-I (n=5), PCV2a-PRRSV-CoI (n=5), PCV2b-I (n=5), and PCV2b-PRRSV-CoI (n=5). Blood, oral, nasal and fecal swabs were collected in regular intervals from day post inoculation (dpi) 0 until dpi 70 and tested by quantitative real-time PCR for the presence and amount of PCV2 DNA and by ELISA for the presence of PCV2-specific antibodies. The results indicate that there were significantly (P<0.05) higher loads of PCV2a and PCV2b DNA in serum, oral swabs, nasal swabs and fecal swabs and a higher prevalence of detectable PCV2 antigen in tissues of pigs concurrently infected with PCV2 and PRRSV compared to pigs singularly infected with PCV2 further confirming that PRRSV enhances replication of PCV2. Moreover, PRRSV infection significantly prolonged the presence of PCV2 DNA in serum and increased the amount of PCV2 DNA in oral and nasal secretions and fecal excretions in the later stages of infection between dpi 28 and 70. Shedding patterns were similar between groups infected with PCV2a and PCV2b, indicating that there was no subtype-specific interaction with the PRRSV isolate used in this study. The results from this study highlight the interaction between PRRSV and PCV2 and the importance of controlling PRRSV infection in order to reduce PCV2 virus loads in pig populations.  相似文献   

9.
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10.
Several tests for Bovine viral diarrhea virus (BVDV) were applied to samples collected monthly from December 20, 2005, through November 27, 2006 (day 0 to day 342) from 12 persistently infected (PI) cattle with BVDV subtypes found in US cattle: BVDV-1a, BVDV-1b, and BVDV-2a. The samples included clotted blood for serum, nasal swabs, and fresh and formalin-fixed ear notches. The tests were as follows: titration of infectious virus in serum and nasal swabs; antigen-capture (AC) enzyme-linked immunosorbent assay (ELISA), or ACE, on serum, nasal swabs, and fresh ear notches; gel-based polymerase chain reaction (PCR) testing of serum, nasal swabs, and fresh ear notches; immunohistochemical (IHC) testing of formalin-fixed ear notches; and serologic testing for BVDV antibodies in serum. Of the 12 animals starting the study, 3 died with mucosal disease. The ACE and IHC tests on ear notches had positive results throughout the study, as did the ACE and PCR tests on serum. There was detectable virus in nasal swabs from all the cattle throughout the study except for a few samples that were toxic to cell cultures. The serum had a virus titer ≥ log10 1.60 in all samples from all the cattle except for 3 collections from 1 animal. Although there were several equivocal results, the PCR test most often had positive results. The BVDV antibodies were due to vaccination or exposure to heterologous strains and did not appear to interfere with any BVDV test. These findings illustrate that PI cattle may be identified by several tests, but differentiation of PI cattle from cattle with acute BVDV infection requires additional testing, especially of blood samples and nasal swabs positive on initial testing. Also, calves PI with BVDV are continual shedders of infectious virus, as shown by the infectivity of nasal swabs over the 11-mo study.  相似文献   

11.
The present study focused on Mycoplasma hyopneumoniae (M. hyopneumoniae) detection by nPCR in nasal swabs of 507 suckling pigs. These animals came from 69 sows (from 1 to 8 parity number) of a farrow-to-finish herd with Enzootic Pneumonia (EP) problems at finishing stages. At 1 and 3 weeks of age (still in the farrowing units), nasal swabs and blood samples were taken from all piglets. Moreover, from these 507 animals, 37 randomly selected pigs were necropsied at 3 weeks of age. From those necropsied pigs, M. hyopneumoniae presence was tested in bronchial and tonsillar swabs. At 1 week post-farrowing, blood samples from sows were collected and used to detect M. hyopneumoniae antibodies. From the 69 analysed sows, 19 (27.5%) were seropositive. Global percentage of pigs with M. hyopneumoniae detection in nasal swabs at 1 and 3 weeks of age was 1.5% (8 out of 507) and 3.8% (19 out of 507), respectively. From these nPCR positive pigs, 89% (24 out of 27) were seronegative and 11% were seropositive. From necropsied animals, the pathogen DNA was detected in two pigs at bronchus level and in another pig at tonsil. In this study, sow parity was not statistically related with sow seropositivity and piglet colonization. These results confirm that M. hyopneumoniae infection may be detected not only in nasal cavities of naturally infected suckling piglets but also at their low respiratory tract airways. Our results suggest that M. hyopneumoniae detection in lower and upper respiratory tract could be an indicator that respiratory problems associated to EP may start relatively early in the production system. In consequence, sow-to-piglet and/or piglet-to piglet transmission in farrowing barns should not be underestimated.  相似文献   

12.
The time-dependent transmission rate of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) and the correlation between infectiousness, virological parameters and antibody responses of the infected pigs were studied in experimental conditions. Seven successive transmission trials involving a total of 77 specific pathogen-free piglets were carried out from 7 to 63 days post-inoculation (dpi). A semi-quantitative real time RT-PCR was developed to assess the evolution of the viral genome load in blood and nasal swabs from inoculated and contact pigs, with time. Virus genome in blood was detectable in inoculated pigs from 7 to 77 dpi, whereas viral genome shedding was detectable from nasal swabs from 2 to 48 dpi. The infectiousness of inoculated pigs, assessed from the frequency of occurrence of infected pigs in susceptible groups in each contact trial, increased from 7 to 14 dpi and then decreased slowly until 42 dpi (3, 7, 2, 1 and 0 pigs infected at 7, 14, 21, 28 and 42 dpi, respectively). These data were used to model the time-dependent infectiousness by a lognormal-like function with a latency period of 1 day and led to an estimated basic reproduction ratio, R0 of 2.6 [1.8, 3.3]. The evolution of infectiousness was mainly correlated with the time-course of viral genome load in the blood whereas the decrease of infectiousness was strongly related to the increase in total antibodies.  相似文献   

13.
A number of polymerase chain reaction (PCR)-based diagnostic tests have been developed for Mycoplasma hyopneumoniae, including one from this research group. This report presents further development, optimization, and standardization of a nested PCR test. Detection sensitivity was 1 fg of M. hyopneumoniae chromosomal DNA (approximately 1 organism). This exceeded the sensitivity of or compared favorably with other published PCR tests. Polymerase chain reaction primers to porcine beta2-microglobulin were included as internal controls for amplifiable chromosomal DNA from porcine samples. To standardize the test, a number of samples from experimentally infected pigs, including nasal, tonsil, tracheobronchial swabs, lung tissue, bronchial alveolar lavage (BAL) fluid, and tracheobronchial brush samples, were examined by PCR. Samples obtained from BAL fluid and tracheobronchial sites were most predictive of infection, whereas nasal swabs and lung tissue were not reliable indicators of experimentally induced infection. In conclusion, the nested PCR developed for this study was found to be a highly sensitive and specific diagnostic tool for M. hyopneumoniae, but the enhanced sensitivity may be unnecessary if the proper sites are sampled.  相似文献   

14.
The use of the four-layer enzyme immunoassay (EIA) for the detection of IgG, IgM and IgA antibodies against Aujeszky's disease virus in blood and oropharyngeal swabs of infected and vaccinated pigs is described. Mean antibody titres obtained using the four-layer EIA were 6.1 and 3829 times higher compared with the indirect enzyme-linked immunosorbent assay (ELISA) and virus neutralization (VN) test, respectively. The VN test detected mainly IgG antibodies, while the IgM antibodies did not react. Using the EIA, the first antiviral antibodies in sera were demonstrated on Days 5-7 after infection or vaccination. Up to the 7th day, demonstrable antibodies were almost exclusively of the IgM class. In infected pigs high titres of IgM antibodies were still detected on Day 18, while in vaccinated animals they were absent by this time. Antibodies of the IgG class appeared in infected pigs sooner (Day 7) than in vaccinated pigs (Day 10) and reached higher mean titres. Antibodies of the IgA class were demonstrable from Day 10 only in samples from infected pigs. Similar antibody dynamics and distribution were detected in oropharyngeal swabs, except that the IgG and IgM titres were roughly 100 times lower than in sera. However, titres of IgA antibodies in oropharyngeal swabs were two times higher than in sera. The greatest differences between both groups of animals were recorded on Day 18; in the infected pigs, IgG, IgM and IgA antibodies were present in sera and oropharyngeal swabs at that time, while in vaccinated pigs only IgG antibodies were demonstrable. The effect of infection and vaccination on the pattern of the immune response as well as the importance of the detection of individual immunoglobulin classes for the specificity of the enzyme immunoassay are discussed.  相似文献   

15.
The purpose of this study was to describe the dynamics (shedding and transmission) of Mycoplasma hyopneumoniae infection within a population of swine and to determine the duration of the infection (persistence) through the identification of the agent in bronchial samples. Sixty-three 2-month-old pigs were used in this study. The pigs (n = 28) were experimentally infected by the intratracheal route with M. hyopneumoniae and considered as seeder pigs. The remaining pigs (n = 32) were not inoculated and randomly allocated to 2 different groups: direct contact exposure pigs (n = 12) and indirect contact exposure pigs (n = 20). Blood samples and nasal swabs were collected throughout the study on days 0, 28, 35, 42, 49, 63, 91, and 119 postinfection. To assess the duration of M. hyopneumoniae infection, 9 seeder and 6 contact exposure pigs were slaughtered at days 155 (group 1), 170 (group 2), and 185 (group 3) postinfection. Direct contact pigs showed evidence of infection on day 28 by polymerase chain reaction (PCR) and on day 35 by serology. The indirect contact exposure pigs presented a very delayed and slow seroconversion pattern; they did not present evidence of transmission until 42 d after the infection of seeder pigs. Identification of M. hyopneumoniae in bronchial swabs was confirmed by nested-PCR from days 155 to 185 postinfection. At the last slaughter date, 77.7% and 100% of the seeders and contact exposure pigs, respectively, tested positive. The results of this study reconfirmed direct infection of M. hyopneumoniae and suggest that indirect transmission can occur in a population. Finally, duration of the infection in this study was longer than previously described.  相似文献   

16.
Hendra virus (HeV) is a zoonotic virus from the family Paramyxoviridae causing fatal disease in humans and horses. Five-week-old Landrace pigs and 5-month-old Gottingen minipigs were inoculated with approximately 107 plaque forming units per animal. In addition to fever and depression exhibited in all infected pigs, one of the two Landrace pigs developed respiratory signs at 5 days post-inoculation (dpi) and one of the Gottingen minipigs developed respiratory signs at 5 dpi and mild neurological signs at 7 dpi. Virus was detected in all infected pigs at 2–5 dpi from oral, nasal, and rectal swabs and at 3–5 dpi from ocular swabs by real-time RT-PCR targeting the HeV M gene. Virus titers in nasal swab samples were as high as 104.6 TCID50/mL. The viral RNA was mainly distributed in tissues from respiratory and lymphoid systems at an early stage of infection and the presence of virus was confirmed by virus isolation. Pathological changes and immunohistochemical staining for viral antigen were consistent with the tissue distribution of the virus. This new finding indicates that pigs are susceptible to HeV infections and could potentially play a role as an intermediate host in transmission to humans.  相似文献   

17.
Objective Investigate the diversity of serovars of Haemophilus parasuis (Hps) present in Australian pig herds. Design Nasal swabs were used to obtain multiple isolates of Hps, which were grouped first by genotyping and then by serotyping representative isolates. Procedure Swabs were taken from the nasal cavity of just-weaned healthy pigs from multiparous sows on 12 farms and from post-weaned pigs of multiparous sows on 1 farm. On 5 of the 13 farms, nasal swabs were also obtained from pigs showing clinical signs suggestive of Glässer's disease. On a further 7 farms, nasal swabs were obtained only from pigs with clinical signs suggestive of Glässer's disease. Results A total of 556 Hps isolates were genotyped, and 150 isolates were serotyped. Hps was detected on 19 of the 20 farms, including 2 farms with a long history of freedom from Glässer's disease. Isolates of Hps belonging to serovars regarded as potentially pathogenic were obtained from healthy pigs at weaning on 8 of the 10 farms with a history of Glässer's disease outbreaks. Sampling 213 sick pigs yielded 115 isolates of Hps, 99 of which belonged to serovars that were either potentially pathogenic or of unknown pathogenicity. Only 16 isolates from these 213 sick pigs were of a serovar known to be non-pathogenic. Conclusion Healthy pigs contain a range of Hps serovars, even on farms free of Glässer's disease. Nasal swabbing of both healthy and sick pigs seems a useful method of serovar profiling of farms.  相似文献   

18.
Many farrow-to-finish herds are endemically infected with Actinobacillus pleuropneumoniae. In order to control the disease efficiently, a better knowledge of the ages at which pigs become infected is necessary. Furthermore, no information is available concerning the influence of maternally derived antibodies on the colonization of the upper respiratory tract. Therefore, A. pleuropneumoniae infection patterns were studied in five farrow-to-finish pig herds (A-E) with a history of pleuropneumonia. A longitudinal study was carried out in herds A and B. In these herds, piglets from sows carrying A. pleuropneumoniae in their noses or tonsils were sampled. Nasal and tonsillar swabs as well as sera, were collected from these animals at the age of 4, 8, 12, 16 (herds A and B) and 23 weeks (herd B). At these ages other pigs from the same sows were euthanized. The lungs were macroscopically examined and samples from nose, tonsils and lungs were collected at necropsy. A cross-sectional study was performed in herds C-E. In these herds nasal and tonsillar swabs, as well as sera, were taken from 10 animals of 4, 8, 12 and 16 weeks of age. Lung, nasal and tonsillar samples were tested for the presence of A. pleuropneumoniae by routine bacteriology and PCR with mixed bacterial cultures. The sera were examined for the presence of Apx toxin neutralizing antibodies. In herd A, A. pleuropneumoniae serotype 2 and 10 strains were isolated, whereas serotype 2, 3, 5b and 8 strains were demonstrated in herd B. In most herds, A. pleuropneumoniae was detected in mixed bacterial cultures of tonsillar and/or nasal samples by PCR from the age of 4 weeks onwards. Colonization of the lungs and development of lung lesions was observed in 12- and 16-week-old animals of herd A and 23-week-old animals of herd B. In most herds, high antibody titres were detected in 4-week-old piglets. These titres decreased during the first 12 weeks of age, but thereafter, increased. It was concluded that PCR with mixed bacterial cultures from tonsillar swabs is a valuable tool for the detection of infected animals. It was also concluded that colonization of tonsils and nasal mucosae can occur in the presence of maternally derived antibodies. Infection of the upper respiratory tract without lung involvement did not result in development of Apx toxin neutralizing antibodies. Therefore, such serological assays cannot be used for the detection of subclinically infected animals.  相似文献   

19.
Both conventional and specific pathogen free pigs were inoculated intranasally with a strain of Aujeszky's disease virus (ADV). Nasal cells were collected daily by swab, aspiration or wash. The nasal cells were examined for ADV by isolation on cell culture, direct or indirect immunofluorescence and immunoperoxidase staining by monoclonal antibodies. The infected pigs were studied for nasal shedding of infected cells until 30 days after infection. The study was also extended to naturally infected farm pigs. Swabbing, washing and aspiration proved effective methods of collecting between 10(5) and 10(8) pavement or columnar epithelial cells and non-epithelial cells. Macrophages and polymorphonuclear leucocytes were also identified. Infected nasal cells were detected by immunofluorescence and immunoperoxidase from one to 21 days after infection. The viral antigen was detected in both epithelial and non-epithelial cells, the fluorescence was nuclear and, or, 'cytoplasmic', in the latter case only the cell membrane was stained. ADV antigens were detected in nasal cavity cells in pigs infected with a virulent and a hypovirulent strain. Nasal swabs proved effective in confirming infection both by virus isolation and immunological assay, and the latter was shown to be a useful experimental tool for the rapid diagnosis of Aujeszky's disease virus infection in fattening pigs suffering from acute respiratory distress.  相似文献   

20.
The porcine respiratory disease complex (PRDC) is an increasingly important cause of decreased swine productivity and is characterized by slow growth, decreased feed efficiency, anorexia, cough, and dyspnea. Mycoplasma hyopneumoniae is among the most prevalent and important infectious agents associated with PRDC. Understanding of mycoplasmal pneumonia has been hindered by inadequate diagnostic methods. Many of the currently available tests are relatively insensitive or nonspecific when used in a diagnostic laboratory setting or are too costly or difficult for routine diagnostic use. Several polymerase chain reaction (PCR) assays have been described, but they are not sensitive enough to detect the microorganisms in live pigs, from either nasal or tracheal swabs. A nested PCR using 2 species-specific sets of primers from the 16S ribosomal DNA gave positive results with as little as 80 microorganisms and did not cross-react with other mycoplasma species or with other microorganisms commonly found in the respiratory tract of pigs. This assay was better suited for detection of M. hyopneumoniae from nasal swabs than was conventional PCR. Nasal swab samples were taken at different time periods following experimental challenge of 10 susceptible pigs. Only 2 of the 55 swabs examined gave a positive result with conventional PCR, whereas 30 of the 55 swabs gave a positive result using the nested PCR. Twenty of 40 (50%) nasal swabs from pigs experiencing a respiratory disease outbreak where M. hyopneumoniae had been diagnosed also gave a positive result with the nested PCR. To confirm that the amplified product was specific, 4 nested PCR products were purified, sequences were determined and aligned, and they were confirmed to be from M. hyopneumoniae.  相似文献   

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