PPD批次与注射方式对PPD皮内变态反应检疫的影响

[目的]为了探究在牛结核病检疫中出现的使用不同批次的PPD和皮下注射PPD等情况是否会对检疫结果的判定造成干扰。[方法]试验首次通过在豚鼠背部采用皮下注射法和皮内交替注射同一厂家不同批次的结核菌素提纯蛋白衍化物(purified protein derivative of tuberculin, PPD)的方法,参照《结核菌素提纯蛋白衍化物(PPD)制造及检定规程》于每次注射后24 h, 48 h和72 h观察皮试处是否产生迟发型变态反应。[结果]皮内组(不同批次PPD皮内注射)和皮下组(同一批次PPD皮下注射)经3次注射后的皮试结果与标准参照组的皮试结果无差异,且在每次皮试后的24 h, 48 h和72 h皮试处均未出现局部红斑硬结。[结论]试验证明了PPD皮内变态反应检疫结果判定不受PPD皮下接种及PPD的不同批次的干扰。     

应用提纯牛型结核菌素(PPD)和结核菌素(OT)皮内试验对成年奶牛检疫的观察

我们自1981年5月对小群成年奶牛采用提纯牛型结核菌素(PPD)和结核菌素(OT)皮内对比试验后,发现PPD不仅可简化检疫程序(改二次注射、二次观察、二次判定为一次注射、一次观察和一次判定)、缩短检疫时间(由120小时缩短到72小     

不同厂家牛型结核菌素（PPD）在奶牛结核病检测中的效果对比

采用国产2个不同厂家的牛型提纯结核菌素进行皮内变态反应比对试验,以期了解不同厂家生产的牛型提纯结核菌素(PPD)对奶牛结核病皮内变态反应试验结果的影响。试验结果显示,2个厂家生产的试剂检测结果符合率较高,差异不显著。     

不同监测方法在奶牛结核病检疫中的应用

为了解牛型提纯结核菌素PPD皮内变态反应试验和牛结核γ-干扰素ELISA试验的相关性,对规模牛场先进行牛型PPD皮内变态反应试验,采集检疫阳性牛和部分阴性牛抗凝血进行牛结核γ-干扰素ELISA试验,通过试验结果找出两者之间的相关性,研究牛结核γ-干扰素ELISA试验在牛结核病检疫中的可行性。     

两种奶牛结核病检测方法的比较研究

牛结核病是由牛分枝杆菌引起的一种人兽共患的慢性消耗性传染病,严重威胁着人类的健康。因此,对该病的定期检疫对公共卫生和食品安全具有重大意义。本研究采用提纯结核菌素(PPD)皮内变态反应试验和γ-干扰素(IFN-γ)ELISA试验,对扬州市的892头奶牛进行结核病检测。结果表明,PPD皮内变态反应检出的31头牛型结核阳性牛和可疑牛中,经IFN-γELISA试验检测仅检出9头禽型结核可疑牛。PPD皮内变态反应试验特异性较差,出现了较高的假阳性率,其原因可能与禽结核分枝杆菌感染或其他非特异性因素的干扰有关。     

二种牛型提纯结核菌素变态反应试验结果比较

为了解不同厂家生产的牛型提纯结核菌素（PPD）对奶牛结核病检疫皮内变态反应试验结果的影响,随机选取二个不同规模奶牛场的100头青年母牛,分别采用国产和荷兰产PPD诊断液2000IU和3000IU的剂量,在每头参试牛的左右侧颈部同时进行皮内变态反应试验,并对判定结果进行比较;同时对所有阳性牛肺门淋巴结样品应用细菌培养的方法进行复核。结果显示：二种PPD诊断液均存在非特异性反应,但国产PPD的非特异性反应显著大于荷兰产PPD（P〈O．01）。     

γ-干扰素检测技术和皮内变态反应在奶牛结核病检测中的应用比较

笔者对牛型提纯结核菌素(PPD)皮内变态反应试验和新型牛结核杆菌γ-干扰素检测技术进行比较,分析其优劣。试验结果表明,γ-干扰素检测技术特异性好、敏感性高、操作简便、易于标准化,但价格较高。该方法可作为PPD皮内变态反应的平行试验,用于复核检查目前国家标准方法 PPD皮内变态反应阳性牛。     

新疆石河子地区牛结核病检测方法的比较研究

采用比较变态反应检测方法对石河子地区6家规模牛场牛型提纯结核菌素(PPD)皮内变态反应检出的475头阳性牛进行了检测,按照国家标准-动物结核病诊断技术(GB/T18645-2002)进行结果判定,结果表明:牛型结核阳性457头,符合率在93.18%～100%之间,平均96.21%。经反复试验对比分析采用牛型提纯结核菌素皮内变态反应进行结核病的检测,结果准确,既经济实用又便于操作,还可避免再采用比较变态反应要隔离阳性牛而导致牛结核病传染散播风险。项目组通过在四个牛场的对比分析,在未建立符合防疫条件的独立隔离圈舍的牛场检测阳性牛及可疑牛在隔离45d后再次检测时新增阳性牛数量增加;而采用牛型提纯结核菌素检测后对阳性牛采取扑杀、无害化处理、消毒等综合性措施后,间隔45d对全群再次检测时新增阳性牛数量明显减少。因此牛结核病检疫采用适宜基层实际的牛型提纯结核菌素皮内变态反应对及时检出阳性牛,并尽快进行扑杀、无害化处理、净化牛群、控制结核的传播扩散,实现公共卫生安全具有重要意义。     

上林县奶水牛结核病检疫报告

按国家规定采用牛型结核分枝杆菌提纯的结核菌素 (PPD)皮内变态反应试验对上林县某奶牛场水奶牛进行结核病检疫 ,被检奶水牛 46头 ,检出结核病阳性奶水牛 3头。文章分析了结核病阳性牛发生原因并提出净化奶水牛结核病的措施。     

不同接种方法对奶牛PPD变态反应结果的影响

通过对奶牛PPD点眼、皮内注射、浅层肌肉注射、多次进针接种等不同方法的变态反应，对比观察其反应结果，得出了一次性成功的皮内注射是PPD接种的最佳方式，而点眼法和浅层肌肉注射、多次进针接种，可使变态反应呈现出假阴性和假阳性事件，提出掌握娴熟的皮内注射方法和技能是提高PPD变态反应结果准确性的关键。     

Evaluation of relationship between plasma fibrinogen concentration and tuberculin testing in red deer

After intradermal injection of bovine purified derivative (PPD), increases in plasma fibrinogen concentration and plasma viscosity developed in red deer (Cervus elaphus) with a history of tuberculosis caused by Mycobacterium bovis. Serum haptoglobin concentrations were also found to increase under similar circumstances. The increases were reproducible and did not appear to be related to mustering, stress, or the handling associated with injection of PPD. A significant (P less than 0.05) direct relationship was found between the increase in plasma fibrinogen concentration and various markers of bovine tuberculosis infection, such as stimulation of lymphocyte transformation in response to bovine PPD and the diameter of intradermal tuberculin skin test reactions. A stronger correlation (P less than 0.01) was found with the volume of intradermal tuberculin skin test reactivity, and the strongest correlation (P less than 0.001) was with the presence of circulating antibovine PPD antibody.     

Evaluation of bovine cutaneous delayed-type hypersensitivity (DTH) to various test antigens and a mitogen using several adjuvants

The Bacillus Calmette Guerin (BCG)-induced/purified protein derivative (PPD)-elicited tuberculin skin test is a reliable measure of cell-mediated immune response (CMIR), specifically delayed-type hypersensitivity (DTH); however, its use in livestock may confound diagnosis of Mycobacterium tuberculosis. Therefore, various alternative antigen/adjuvant combinations were evaluated as inducers of DTH that were compared to the BCG/PPD test system with the purpose of finding a skin DTH protocol that does not cross-react with the tuberculin test and allows identification of high and low CMIR responder phenotypes. Specifically, 30 non-lactating cows (five/treatment) were sensitized on day 0 with mycobacteria [BCG, M. tuberculosis or Mycobacterium phlei cell wall extract (MCWE)], and ovalbumin (OVA) emulsified in Freund's complete adjuvant (FCA), non-ulcerative Freund's adjuvant (NUFA), complete NUFA or MCWE. On day 21, cows were injected intradermally with various test antigens including PPD tuberculin, phlein, and OVA. Phosphate buffered saline was included as the negative control and the T-cell mitogen phytohemagglutinin (PHA) was also administered. Double skin-fold thickness was evaluated before and at 6, 24, and 48 h post-injection. Skin biopsies were taken at 24 and 48 h to assess oedema, necrosis, and inflammatory cell infiltration. BCG/PPD and M. phlei/phlein treatments when given with a Freund's adjuvant induced equivalent DTH with peak reactions at 24-48 h after antigen injection. Cows receiving NUFA had fewer injection site granulomas than FCA or CNUFA treatments. The change in skin thickness response to PHA peaked at 6 h. Only cows receiving mycobacteria in NUFA had skin response to OVA, which peaked 6-24 h post-injection. Only sites tested with PPD or phlein had significantly higher lymphocyte infiltration than control, whereas neutrophils were significantly higher at PHA test sites and eosinophils predominated at the PHA test sites. Macrophages were significantly more numerous at the PPD and/or phlein test sites in treatment groups that received killed mycobacteria in a Freund's adjuvant and/or with BCG, and at the PHA test sites in all treatment groups. It was concluded that the M. phlei/phlein system did induce DTH and was similar to the DTH induced by the BCG/PPD system when MCWE was administered with a Freund's adjuvant. Therefore, this protocol is suitable for detecting high/low CMIR responders in research herds. However, cross-reaction to PPD was evident following induction of DTH using M. phlei. Hence, this protocol does not alleviate the problem of artificial induction of DTH cross-reactivity and would not be suitable for commercial herds where tuberculin testing is required.     

COMPARISON OF THE EFFICIENCY OF TWO DOSES OF BOVINE PPD TUBERCULIN IN SINGLE CAUDAL FOLD TESTS ON AUSTRALIAN CATTLE

SUMMARY: The sensitivity and specificity of 0.2 mg and 0.4 mg doses of bovine PPD tuberculin were compared in Northern Territory beef cattle from tuberculous herds and herds with a prevalence of tuberculosis of less than 0.1%. Reactions were interpreted subjectively by observation and palpation, and were also measured to the nearest mm with calipers at 24 h, 48 h, 72 h and 96 h after injection of tuberculin. All cattle were examined post mortem for the presence of macroscopic and microscopic tuberculous lesions. The apparent specificity of caudal fold tests with 0.2 mg and 0.4 mg doses was determined in cattle in Victoria from tuberculosis-free dairy and beef herds. Victorian cattle reacting to the caudal fold tests were subjected to a comparative intradermal test with 0.1 mg bovine PPD and 2,500 IU avian PPD not less than 42 days later.
Tests with the 0.2 mg dose achieved the highest level of sensitivity of 95.6% at 48 h, 72 h and 96 h, while in tests with 0.4 mg the maximum reached was 94.7% at 72 h. The specificity of tests in Northern Territory cattle ranged from 85.0% to 88.3% with the 0.2 mg dose and from 80.6% to 82.3% with the 0.4 mg dose. The highest specificity was achieved with both doses at 96 h. The apparent specificity of 0.2 and 0.4 mg doses of bovine PPD in tuberculosis-free herds in Victoria was high, a false-positive reactor rate of only 0.6% occurring with caudal fold tests. All false-positive reactions were shown to be non-specific or due to previous experimental sensitisation.     

The comparative performance of the single intradermal test and the single intradermal comparative tuberculin test in Irish cattle, using tuberculin PPD combinations of differing potencies

In national bovine tuberculosis (BTB) control programmes, testing is generally conducted using a single source of bovine purified protein derivative (PPD) tuberculin. Alternative tuberculin sources should be identified as part of a broad risk management strategy as problems of supply or quality cannot be discounted. This study was conducted to compare the impact of different potencies of a single bovine PPD tuberculin on the field performance of the single intradermal comparative tuberculin test (SICTT) and single intradermal test (SIT). Three trial potencies of bovine PPD tuberculin, as assayed in naturally infected bovines, namely, low (1192IU/dose), normal (6184IU/dose) and high (12,554IU/dose) were used. Three SICTTs (using) were conducted on 2102 animals. Test results were compared based on reactor-status and changes in skin-thickness at the bovine tuberculin injection site. There was a significant difference in the number of reactors detected using the high and low potency tuberculins. In the SICTT, high and low potency tuberculin detected 40% more and 50% fewer reactors, respectively, than normal potency tuberculin. Furthermore, use of the low potency tuberculin in the SICTT failed to detect 20% of 35 animals with visible lesions, and in the SIT 11% of the visible lesion animals would have been classified as negative. Tuberculin potency is critical to the performance of both the SICTT and SIT. Tuberculin of different potencies will affect reactor disclosure rates, confounding between-year or between-country comparisons. Independent checks of tuberculin potency are an important aspect of quality control in national BTB control programmes.     

Method for evaluation of Mycobacterium bovis purified protein derivative tuberculin in experimentally infected cattle.

A method for the evaluation of Mycobacterium bovis purified protein derivative (PPD) tuberculin in experimentally infected cattle is presented. The development of skin test responses in M bovis-infected cattle was determined for International Standard PPD-S, M bovis PPD-2, and M bovis PPD-5 at 24, 48, and 72 hours. Significantly larger reactions (dermal thickness) were observed at 48 and 72 hours than at 24 hours (P = 0.001). Statistically significant differences were not detected in the responses obtained with M bovis PPD-2, M bovis PPD-5, and International Standard PPD-S if comparisons were made at approximately the same concentrations in M bovis-infected cattle (P greater than 0.25). In Mycobacterium avium-infected cattle, M bovis PPD-2 produced skin test responses that were significantly smaller than responses obtained using M avium PPD-2 (P = 0.001). Significant variation was not observed in the PPD-S responses in 2 groups of M bovis-infected cattle (P greater than 0.1).     

The effect of the tuberculin test and the consequences of a delay in blood culture on the sensitivity of a gamma-interferon assay for the detection of Mycobacterium bovis infection in cattle

The strategic use of the gamma-interferon (IFN-gamma) assay (Bovigam) can provide a means for the early identification of Mycobacterium bovis infected cattle, thus ensuring their removal from an infected herd. It has been reported that performance of the test can be influenced by various factors including a recent tuberculin skin test and the length of delay between collection and processing of blood samples. In this study, single intradermal comparative tuberculin test (SICTT) reactor and non-reactor cattle were recruited from herds infected with M. bovis and grouped according to their SICTT responses. Group 1 comprised reactor cattle selected on the basis of their SICTT response to PPD-bovine (purified protein derivative of tuberculin) exceeding that of PPD-avian by at least 12mm. Group 2 animals were selected from herds undergoing routine surveillance for bovine tuberculosis and contained standard SICTT reactor cattle (PPD-bovine exceeding that of PPD-avian by at least 4mm) and non-reactors. We investigated the effects of the SICTT on the assay results by measuring the in vitro IFN-gamma responses of Group 1 reactor cattle at time intervals pre- and post-skin test. No significant differences were measured in the IFN-gamma responses of the reactor animals to PPD-bovine and PPD-avian for up to 65 days. To investigate if a delay in processing of blood affected the performance of the assay, we compared results using duplicate blood samples from Group 1 and Group 2 cattle stimulated with PPD antigen at 8h and at 24h after collection. In both groups of animals the mean optical density (OD) values of the assay at 24h post-collection were significantly lower than those at 8h. Our results demonstrated that a delay in processing of the blood samples from cattle subjected to routine surveillance could significantly impact on the outcome of the IFN-gamma assay resulting in a change of the IFN-gamma status of the animals.     

Evaluation of two cocktails containing ESAT-6, CFP-10 and Rv-3615c in the intradermal test and the interferon-γ assay for diagnosis of bovine tuberculosis

The intradermal tuberculin tests and the interferon-gamma (IFN-γ) assay are the principal tests used worldwide for the ante-mortem diagnosis of bovine tuberculosis. The conventional reagent currently in use in these tests is purified protein derivative (PPD) tuberculin obtained from Mycobacterium bovis culture. The components of PPD are poorly characterized and difficult to standardize. To overcome this issue, antigens specific to the Mycobacterium tuberculosis complex are being studied. Here we have assessed the biological potency of ESAT-6, CFP-10 and Rv-3615c presented as peptide or recombinant protein cocktails in comparison with the standard bovine PPD used routinely in Spanish eradication campaigns. The study was performed in cattle (n=23) from a herd with natural M. bovis infection. Animals were simultaneously injected with PPD and the peptide and protein cocktails. The percentages of cattle reacting positively to single intradermal test were 60.9% (bovine PPD), 47.8% (peptide cocktail) and 60.9% (protein cocktail), with no significant difference between the actual skin fold thickness increases (p>0.05). The IFN-γ assay detected 60.9% of animals when stimulation was performed with bovine PPD, but decreased to 52.2% when stimulation was performed with the peptide cocktail and to 47.8% when stimulation was performed with the protein cocktail. However, no significant differences were found between IFN-γ responder frequencies (p>0.05). These results show a potential use of these defined reagents for in vivo tuberculosis diagnosis.     

COMPARISON OF THE EFFICIENCY OF 4 DOSES OF BOVINE PPD TUBERCULIN IN CATTLE AND GUINEA PIGS: RECOMMENDATIONS FOR THE MOST SUITABLE DOSE FOR USE IN THE SINGLE CAUDAL FOLD TEST IN CATTLE

The potencies of 4 batches of Australian bovine PPD tuberculin relative to the standard British bovine PPD preparation 291 were determined from guinea pig assays.
Using the assayed potencies, it was concluded that doses of 0.08, 0.17, 0.24 and 0.47 mg of bovine PPD were used in single caudal fold tests in 2 trials of Northern Territory cattle. The sensitivity, specificity and test efficiency of the Australian preparations were determined.
At 72 h after injection, the time recommended to read the test, the 0.08 mg dose gave the lowest sensitivity (79.2%) which was significantly different from that of the other 3 doses. This dose also gave the highest specificity (88.9%). However, the 0.24 mg dose gave a specificity of 85.0% and a sensitivity of 95.6% resulting in the highest test efficiency (87.9%).
Change in caudal fold thickness, subjectively assessed reaction and the amount of induration were measured in individual cattle. For tuberculous cattle, regression analysis showed significant relationships (p < 0.01) between the change in caudal fold thickness, subjectively assessed reaction and log concentration of each preparation. However, there was no significant difference between the mean responses observed with the 2 highest doses. Predicted responses for untested doses were determined from the regression equations.
When the results of the trials were considered in relation to factors such as batch variation, minimisation of injection error, likely levels of desensitisation, cost and international implications, it was considered that a dose of 0.3 mg bovine PPD relative to the British standard 291 would fulfil optimum requirements for use in all parts of Australia.     

Comparison of the specificity of human and bovine tuberculin PPF for testing cattle. 3. National trial in Great Britain.

A field trial on a country-wide basis was undertaken to compare the specificity for bovine tuberculosis of single and comparative tuberculin tests in cattle using either Weybridge human or Weybridge bovine PPD. The tests were made on 10,305 cattle in 179 herds distributed throughout all regions of England, Scotland and Wales. Results showed that a comparative tuberculin test using avian PPD with either human or bovine PPD had a much higher efficiency than a single injection of mammalian tuberculin in the neck of cattle, and confirmed that a comparative test is still essential in the British environment. Weybridge bovine PPD gave significantly better discrimination between tuberculous and non-tuberculous cattle than Weybridge human PPD when used together with avian PPD in a comparative tuberculin test. The diameter of induration gave an absolute measure of the extent of oedema, if present, and induration diameter used in conjunction with skin thickening increased the sensitivity and specificity of the test. Rules of interpretation were developed and are presented for an intradermal comparative tuberculin test in cattle using Weybridge avian and bovine PPDs.