Field study on bovine paratuberculosis using real-time PCR and liquid culture for testing environmental and individual fecal samples implemented in dairy cow management.

Bovine paratuberculosis (Johne's disease) is a bacterial, chronic, and wasting intestinal disease caused by Mycobacterium avium ssp. paratuberculosis (MAP). Johne's disease causes severe losses in dairy farm productivity and is also suspected to be a potential trigger for Crohn's disease in humans. The fecal-oral infection of MAP to neonates is recognized as an important within-herd transmission route. Our objective was to recommend diagnostic methods for herds with suspected paratuberculosis requiring fast results, as well as for herds with breeding programs or others that aim at being nonsuspected of paratuberculosis infection. We determined a period of 8 wk from sampling to diagnostic findings suitable for testing of cows during the dry period. We therefore tested environmental and individual fecal samples with one rapid and one highly sensitive diagnostic method. Environmental samples (boot swabs) were taken as a first step in 3 herds and tested using a DNA extraction protocol for feces and subsequent real-time PCR (referred to as fecal PCR). Additionally, cultivation in liquid medium for 6 wk was performed and verified with real-time PCR (referred to as liquid culture). Automation of DNA extraction based on magnetic beads and the PCR setup was performed with pipetting robots. As a result, we successfully detected MAP in boot swabs of all herds by both methods. In a second step, 245 individual fecal samples from the 3 herds were examined using also fecal PCR and liquid culture. The results obtained by fecal PCR were compared with detection of MAP using cultivation in liquid medium for 6 wk. Testing individual cows, we identified MAP-specific DNA in 53 fecal samples using the liquid culture. Using fecal PCR, we revealed 43 positive samples of which 39 also tested positive in the liquid culture, revealing MAP-positive cows in all 3 herds. The fecal PCR procedure allows rapid detection of MAP-specific DNA with 74% of the sensitivity of liquid culture. For the purpose of testing with maximal sensitivity, cultivation in liquid medium is recommended. Cultivation of MAP in liquid medium M7H9C means a significant time gain in comparison to cultivation on solid media, which requires twice as much time. Thus, this testing fits within the 6- to 8-wk dry period of gravid cows and provides test results before calving, a prerequisite to prevent fecal-oral transmission to newborn calves.

Enhanced sensitivity and fast turnaround time in laboratory diagnosis for bovine paratuberculosis in faecal samples.

Bovine paratuberculosis (Johne's disease in cattle) is caused by the pathogen Mycobacterium avium subsp. paratuberculosis (MAP) and is a widespread chronic bacterial infectious disease in cattle. Due to the peculiarities of the pathogen, detection of MAP in faeces remains difficult. DNA extraction and real-time PCR for detection of MAP in bovine faeces (direct PCR) have been refined and feasible procedures for rapid, sensitive and automatable detection of the pathogen agent have been developed. Accordingly, in a first step we tested 20 faecal samples using two MAP complete kits (DNA extraction kits based on magnetic beads combined with real-time PCR assays) and six other DNA extraction kits for faeces. MAP-specific DNA was detected by real-time PCR assays. Cultivation of MAP on the solid medium HEYM and in the liquid medium M7H9C served as reference standards. The two complete kits detected significantly more MAP-DNA positive samples than the other procedures applied (p < 0.04). Ct values of 37 and 38 served as cut-off for the respective real-time PCR assays calculated on the basis of standard curves and droplet digital PCR (ddPCR). In a second step, the two MAP complete kits were employed for a comprehensive study including 107 positive and 50 negative faecal samples which had been previously tested on HEYM cultivation. The MAP complete kits yielded sensitivity values of 86% and 89% and specificity values of 100% compared to cultivation of MAP in the liquid medium M7H9C. In detail, cultivation of MAP in M7H9C detected the pathogen in 97% and 100% of the samples tested after an incubation period of six and twelve weeks, respectively. However, the cultivation of MAP on HEYM succeeded in only 74% after twelve weeks of incubation. In all these solid culture positive samples, MAP was also detected using the two complete kits. Additionally, the impact of repeated freezing and thawing of samples on re-cultivation of MAP was tested using 20 faecal samples and resulted in a reduction to 75% and 25% of bacterial growth when using liquid medium M7H9C and solid medium HEYM, respectively. The results of this study show that complete kits with refined automatable protocols for DNA extraction in combination with real-time PCR assays for detection of MAP can compete with sensitive cultivation of the pathogen in liquid medium.

Detection of Mycobacterium avium subsp. paratuberculosis in faeces using different procedures of pre-treatment for real-time PCR in comparison to culture.

One of the most relevant aspects in the diagnosis of paratuberculosis (Johne's disease) in cattle is the availability of a method for the rapid and sensitive detection of Mycobacterium avium subsp. paratuberculosis (MAP) in order to facilitate the prompt removal of pathogen-shedding animals from a herd. To meet this requirement, methods for pre-treatment of bovine faecal samples and subsequent extraction of DNA for detection of MAP by real-time PCR were compared with MAP culture results. A total of 116 bovine faecal samples that showed weak (64.7%), moderate (18.1%) or strong (17.2%) growth of MAP on solid HEY medium were investigated. For PCR, supernatants, sediments or bacterial pellets were obtained from faecal samples by pre-treatment before extraction of MAP DNA based on silica membranes or magnetic particles. Samples then were tested by MAP IS900 and ISMav2 real-time PCR with an analytical sensitivity of 6 and 28 genome equivalents (GE) per mL, respectively. The best results were obtained by including a microfiltration step in the sample pre-treatment in combination with silica membrane-based mini-columns or magnetic particles for DNA extraction. This approach enhanced the detection rate of MAP in IS900 real-time PCR from 58.6% to 84.5% using silica membrane mini-columns and from 61.2% to 64.7% using magnetic particles.

Occurrence of vancomycin-resistant enterococci in turkey flocks.

In the present study, the prevalence of vancomycin-resistant enterococci (VRE) in turkeys in the southwest of Germany was investigated. For this purpose, 200 cloacal swab samples and 5 environmental dust samples (tested as a pooled sample) of each of the 20 flocks (10 female and 10 male flocks) included in this study were examined. The VRE could be isolated by means of a procedure combining bacterial cultivation in an enrichment broth and on a selective solid media. Enterococci were identified biochemically and subsequently tested on the presence of the vancomycin resistance genes vanA, vanB (B1/B2/B3), and vanC (C1/C2/C3) using real-time PCR assays. In 54 (27%) turkeys originating from 11 (55%) flocks and in 14 (70%) of the dust samples, exclusively vanA and vanC1 genes could be detected. Of the turkeys examined, 46 were colonized with VRE bearing the resistance gene vanC1 and 8 vanA, originating from 9 and 2 flocks, respectively. None of the birds carried vanB, vanC2, or vanC3 positive VRE. The results obtained from the birds are largely confirmed by the dust samples originating from 4 vanA and 10 vanC1 positive flocks. However, one flock housing animals colonized with vanC1 positive VRE could not be confirmed by the dust samples that revealed vanA bearing VRE. However, in one case vanA and in 3 cases vanC1 carrying VRE could be detected in dust samples of the turkey houses, but not in the turkeys of the associated flock. In 5 flocks the turkeys as well as the dust samples were free of VRE.

Prevalence of types of methicillin-resistant Staphylococcus aureus in turkey flocks and personnel attending the animals.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) have been isolated from a number of livestock species and persons involved in animal production. We investigated the prevalence of LA-MRSA in fattening turkeys and people living on farms that house fattening turkeys. Eighteen (90%) of 20 investigated flocks were positive for MRSA, and on 12 of the farms 22 (37·3%) of 59 persons sampled were positive for MRSA. People with frequent access to the stables were more likely to be positive for MRSA. In most flocks MRSA that could be assigned to clonal complex (CC) 398 were detected. In five flocks MRSA of spa-type t002 that is not related to CC398 were identified. Moreover, other methicillin-resistant Staphylococcus spp. were detected on 11 farms and in eight people working on the farms.

One-step RT-qPCR with an internal control system for the detection of turkey rotaviruses in faecal samples.

Turkey rotaviruses are one of the major pathogens responsible for the poult enteritis syndrome (PES). In this study a one step real-time reverse transcription polymerase chain reaction (RT-qPCR) assay targeting the rotaviral non-structural protein 4 (NSP4) was developed. The NSP4 is a highly conserved gene inside the turkey rotavirus genome and contains an internal control system to monitor any potential RT-qPCR inhibitors. The detection limit of the optimized NSP4-RT-qPCR assay ranged from 8.15 to 8.15 × 10(5) copy numbers. In total 149 faecal samples were collected from eight different flocks of commercial turkey farms. Faecal samples from hens and toms were collected separately at 2-week intervals from the 2nd week of age through the 16th and 20th week of age (age of slaughter for female and male, respectively) and tested. One farm reared only hens. The samples were tested previously using conventional RT-PCR targeting the same gene. When the conventional RT-PCR was compared with the developed NSP4-RT-qPCR, the results revealed that 11% of the samples of the conventional RT-PCR were false negative. The results indicate that this NSP4-RT-qPCR is highly sensitive for the detection of turkey rotaviruses in faeces. In addition, it could be suitable for the development of high-throughput screening.

Corynebacterium ulcerans from diseased wild boars.

Two Corynebacterium strains were isolated from lymph nodes of wild boars showing severe alterations caused by caseous lymphadenitis. The wild boars came from different districts in southern Germany; one was found dead, the other had been shot. The two Corynebacterium strains obtained were both positive for phospholipase D. Further analysis of biochemical profiles did not allow unambiguous differentiation between Corynebacterium ulcerans and Corynebacterium pseudotuberculosis. Fourier-transformed infrared spectroscopy as well as partial sequencing of the genes for 16S rRNA and RNA polymerase beta subunit (rpoB) clearly identified both strains as Corynebacterium ulcerans. The tox gene for diphtheria toxin (DT) could be detected in both porcine isolates by PCR. Partial DNA sequencing of this tox gene showed significant differences from sequences described for other Corynebacterium ulcerans strains and a higher degree of similarity to that of Corynebacterium diphtheria. Production of diphtheria toxin could not be detected. These results indicate that wild game could be a reservoir for zoonotic Corynebacterium ulcerans.

Comparison of the Q-fever complement fixation test and two commercial enzyme-linked immunosorbent assays for the detection of serum antibodies against Coxiella burnetti (Q-fever) in ruminants : recommendations for use of serological tests on imported animals in New Zealand.

AIM: To make valid recommendations on the use of serological test methods for the detection of serum antibodies in ruminants against Coxiella burnetii (Q-fever), by comparing the performance of the complement fixation test (CFT) and two ELISA, and by identifying reasons for discrepancies between the test methods. METHODS: A total of 73 serum samples from infected cattle, 69 from infected goats, and 100 samples from non-infected cattle and 57 samples from non-infected sheep, as well as 95 samples from infected cattle herds (mix of seropositive and seronegative samples), were tested using the CFT, the IDEXX ELISA (I-ELISA) and the Pourquier ELISA (P-ELISA). A mixed panel of 12 serum samples from sheep from inter-laboratory proficiency testing (proficiency panel) was also tested using the CFT and both ELISA, and further investigated using IgG- and IgM-specific ELISA. RESULTS: Generally, the two commercial ELISA were more sensitive than the CFT for the detection of infected ruminants. Good agreement between ELISA for positive and negative results was found for samples from the infected herd, while results for the positive panels varied between the two ELISA. For the total of the positive serum panels, the I-ELISA detected 95% of samples as positive or suspicious, while the P-ELISA detected only 81%. In the P-ELISA, more samples were considered suspicious (18%) than in the I-ELISA (14%). All sera from non-infected sheep and cattle tested negative in the serological test methods employed, except for one positive sample from a sheep in the P-ELISA. Further investigation revealed that a CFT-positive but ELISA-negative result was due to high IgM and low IgG reactivity. CONCLUSIONS: The two commercial ELISA were more sensitive than the CFT in all panels from infected ruminants. However, they could only detect IgG. The I-ELISA should be the serological test method of choice for cattle, sheep and goats for import testing of animals into New Zealand because it was more sensitive than the P-ELISA and was equally specific to the PELISA and the CFT. For other animal species, such as deer and camelids, the CFT should still be used since none of the ELISA has been evaluated for these species. This study has shown that the two commercial ELISA will detect the majority of infected ruminants but may miss animals that have not developed an IgG response.

Duplex real-time PCR assays for rapid detection of virulence genes in E. coli isolated from post-weaning pigs and calves with diarrhoea.

Duplex real-time PCR assays were used as modules to cover partially automated detection of 12 genes encoding adhesins, enterotoxins and Shiga toxins in faecal E. coli isolates. For this a total of 194 E. coli isolates from pigs suffering from post-weaning diarrhoea (PWD), including 65 isolates with haemolytic activity, and 83 isolates from calves with diarrhoea were examined. Data obtained by PCR were compared with O-typing and with haemolytic activity as indirect virulence markers. E. coli O-types O139:K82, O141:K85, and O149:K91 accounted for 43.8% (n = 85) of all porcine strains and for 55.4% (n = 36) of the porcine strains, which exhibited haemolytic activity. These strains carried virulence genes by 65.9% (n = 56) and 80.6% (haemolytic E. coli, n = 29), respectively. The E. coli O-types O139:K82 and O141:K85 were significantly associated with the adhesin gene F18, and O149:K81 with the F4 gene. In this context, detection of the gene encoding F18 was coupled predominantly with the genes responsible for the production of the toxins ST-I, ST-II and Stx2, and the F4 gene with those of the enterotoxins ST-I, ST-II and LT. Both virulence patterns were detected more pronounced in E. coli strains with haemolytic activity. Fifty-six of a total of 83 E. coli isolates originating from calves were O-typed as O101 (O101:K28, O101:K30, O101:K32; n = 29), O78:K80 (n = 23), and O9:K35 (n = 4). Most of the E. coli O78:K80 strains carried the F17 gene (69.6%, n = 16). Virulence genes encoding for F4, F5 or ST-I were detected only in single cases. Intimin and Shiga toxin genes that are present in enterohaemorrhagic E. coli (EHEC) were not detected.

[Comparative studies on detection of Chlamydophila psittaci and Chlamydophila abortus in meat turkey flocks using cell culture, ELISA, and PCR]. / Vergleichende Untersuchungen zum Nachweis von Chlamydophila psittaci und Chlamydophila abortus in Putenmastbetrieben mittels Zelikultur, ELISA und PCR.

The prevalence of chlamydia in 10 meat turkey flocks was investigated. As samples served of each moment of collection and sex of the animals 10 cloacal swabs which were taken at the age of 1, 4, 8 and 12 (females) or 16 weeks (males) and at the time of slaughter at the age of 16 or 20 weeks. Spleen samples were taken at the time of slaughter, additionally. These were pooled making 1 pool out of 5 individual samples. The cloacal and spleen pools were examined by nested PCR (nPCR), Capture-ELISA and Capture Blocking-ELISA directly as well as after isolation attempts in cell cultures. The most sensitive method to detect chlamydia, with 6 isolates proved to be the isolation by cell culture followed by detection using nPCR. Not corresponding to the results of the nPCR were 4 positive reactions found by the Capture-ELISA which could in no case be affirmed by Capture-Blocking-ELISA. The direct examination of cloacal swab pools by nPCR proved positive in only 2 cases. In contrast to this the examination of these samples by Capture-ELISA showed a high percentage of 71.9% positive results, of which only 2 cases were confirmed by nPCR and none by Capture-Blocking-ELISA. Of the 8 Chlamydia positive results in the nPCR 7 could be classified by DNA sequencing to Cp. abortus and only one to Cp. psittaci.

[The occurrence of Coxiella burnetii in sheep and ticks of the genus Dermacentor in Baden-Wuerttemberg]. / Untersuchungen zum Vorkommen von Coxiella burnetii bei Schafen und Zecken der Gattung Dermacentor in Baden-Württemberg.

This study examined the occurrence of Coxiella burnetii (C. burnetii), the infectious agent of Q-fever, in sheep and sheep-ticks in Baden-Wuerttemberg, Germany, as a possible source of infection in Q-fever outbreaks. Using PCR, we examined a total of 1066 Dermacentor ticks from 23 herds and 49 samples of tick excrement from 18 herds for C. burnetii. We found the infectious agent in one non-engorged tick and in one sample of tick excrement from the same herd, in Efringen-Kirchen (district Loerrach). Sequencing the PCR-products confirmed the amplifications as specific for C. burnetii. Further serological tests of random samples of the four districts of Baden-Wuerttemberg showed a seroprevalence from 0 to 1.4% using complement fixation test (CFT), as well as a 0.9 to 10.2% seroprevalence, using ELISA test. Serum samples from a Q-fever-suspicious herd resulted, however, in 6% (CFT) and 53% (ELISA) positive reactions. A comparison between CFT and ELISA showed both a correlation of the two test methods that increased with higher CFT titration levels and positive reactions using ELISA for 9.4% of the serums that had tested negative using CFT. The results of the present study reveal that ticks and their excrements are important vectors of transmission of Q-fever in Baden-Wuerttemberg. Investigations on C. burnetii using PCR as well as serological surveys of sheep are important instruments for diagnosis and disease control of Q-fever.

Efficacy of ELISA for detection of antibodies against several Ornithobacterium rhinotracheale serotypes.

The present investigation were carried out to compare self-made ELISA based on SDS-Antigen extraction of serotype B and one commercial ELISA-kit (Biocheck, Gouda, The Netherlands) for their ability to detect antibodies against 12 ORT serotypes (A-L). Using both ELISA systems, antibodies against all serotypes were detected. Examination of serum samples collected from commercial flocks showed similar results on flock bases. However, some minor variations on sample basis could be demonstrated. In conclusion using both ELISA system antibodies against different serotypes could be detected.

[Demonstration of Chlamydia from an equine abortion]. / Nachweis von Chlamydien bei einem Stutenabort.

The isolation and identification of a chlamydial agent from an equine fetus is reported. The fetus was aborted by a mare with respiratory disease and fever in the 9th month of pregnancy. The serum of the mare was investigated by the compliment fixation test. Specific antibodies were detected for chlamydial antigen in a titer of > 1:40 and for equine herpes virus 1 antigen in a titer of 1:32. Pathological lesions were not found in the organs of the fetus. Chlamydiae were detected in the placenta by ELISA and subsequently isolated by cell culture. Using PCR technique the agent was identified as Chlamydophila psittaci.

[Experience with simple ELISA test systems for Brucella serology in cattle, sheep and goats]. / Erfahrungen mit einfachen ELISA-Testsystemen für die Brucellose-Serologie bei Rind, Schaf und Ziege.

The objective of this work was to use the ELISA technique for the serological surveillance for freedom of brucellosis of cattle, sheep and goats. By comparing 28 cattle sera taken after a brucellosis outbreak, 15 bovine sera supplied by the Federal Institute for Health Protection of Consumers and Veterinary Medicine (BgVV) and 497 serum slow agglutination test (SSAT) and complement fixation test (CFT) negative bovine sera from herds officially declared free of brucellosis, the ELISA technique not only shows higher sensitivity as compared to SSAT and CFT but also distinguishes clearly between positive and negative reactions. The serological comparison by SSAT, CFT and ELISA of 615 cattle, 624 sheep and 630 goat sera from herds acknowledged as brucellosis free showed equivalent specificities for both CFT and ELISA. The specificity of the SSAT was much lower, 81.1% in cattle and 96.2% in goat sera. The examination of 5796 cattle, 1337 calf, 5031 sheep and 1796 goat sera demonstrates the advantage of the ELISA technique as routine method. The possible application of the ELISA technique as a screening method for serological brucellosis tests in sheep, goats and possibly also in pigs is discussed.

[Coxiella burnetii infections and infections with bacteria of the genus Chlamydia in dairy cattle]. / Untersuchungen zu Coxiella burnetii-Infektionen und Infektionen mit Bakterien der Gattung Chlamydia in Milchviehbetrieben.

Comparative studies on the prevalence of infections caused by Coxiella burnetii (C. burnetii) and Chlamydia were carried out with 592 cattle older than 2 years and 234 cattle younger than 2 years. Of these 477 originated from 24 dairy herds with considerable fertility problems (positive herds) and 349 from 14 dairy herds without major fertility problems (control herds). For the direct detection of these pathogens in the genitals capture ELISAs were employed, for the demonstration of antibodies the complement fixation test (CFT). Direct detection of C. burnetii and Chlamydia single as well as mixed infection revealed significant higher values for cattle from positive herds compared with those from the control herds. Animals revealing insemination ratios of > or = 2 showed significantly more frequent excretion of Chlamydia via the genitals and antibodies against C. burnetii than cattle with an insemination ratio of < 2. Investigations of cows which had had an abortion showed no indications of significantly more frequent C. burnetii or chlamydial infections. Inseminated but non-pregnant cows excreted significantly more C. burnetii and Chlamydia than pregnant cows. Clinical signs of endometritis were associated with an enhanced excretion of Chlamydia. Animals younger than 2 years excreted significantly more frequently C. burnetii but not Chlamydia via the genitals than animals older than 2 years. Indirect test showed results vice versa.

Investigations on different Ornithobacterium rhinotracheale "ORT" isolates.

The aim of the present investigation was to determine the antigenic relationship between different Ornithobacterium rhinotracheale (ORT) isolates and to serotype field isolates obtained from turkey and chickens. Different antigen extractions (heat-stable, proteinase K-stable [lipopolysaccharide], and sodium dodecyl sulfate [SDS] extractions) were prepared from each serotype (A, B, C, D, E, and G) as well as from 21 ORT field isolates and examined in agar gel precipitation (AGP) and enzyme-linked immunosorbent assay (ELISA) tests. The field isolates were cultured from turkey (16 isolates) and chicken (5 isolates) flocks showing respiratory manifestations. Monospecific reactions were obtained with heat-stable as well as proteinase K-stable antigens prepared from serotypes A, C, D, E, and G in AGP tests. On the other hand, with the same antigen preparations from a strain of serotype B in AGP tests, cross-reactions with antisera prepared against serotypes A and E could be detected. The cross-reactions were observed mostly between 48 and 72 hr. In applications of SDS-antigen preparations in AGP tests, cross-reactions between all serotypes except serotype C were detected between 24 and 72 hr. Testing all antigen preparation in ELISA, different cross-reactions were observed and the evaluation of the results is very difficult. Serotyping of the field isolates in AGP tests by using heat-extracted antigens showed after 24 hr that 10 out of 16 isolates from turkey belonged to serotype B, five to serotype A, and one to serotype E. Results obtained after 48-72 hr revealed cross-reactions between serotype B and E in 11 cases and between A and B in two cases. All five isolates obtained from chicken reacted after 24 hr only with serum against serotype A. After 48-72 hr, two isolates showed cross-reaction with antiserum against serotype B. Similar results were obtained with proteinase K-stable antigen.

Serological studies on Corynebacterium pseudotuberculosis infections in goats using enzyme-linked immunosorbent assay.

Corynebacterium pseudotuberculosis was isolated from a goat suffering from caseous lymphadenitis and used for preparation of cell wall antigens and exotoxin for detection of specific antibodies in enzyme-linked immunosorbent assay (ELISA). In immunoblotting sodium dodecyt sulphate (SDS) extracted cell wall antigens revealed molecular weights ranging from 20 to 120 kDa. The raw exotoxin showed molecular weights of 30 and 55 kDa and an inhibition of the haemolysis of a Staphylococcus aureus strain. For validation of the ELISA 109 goats of known clinical status were examined reaching a sensitivity of 96% and a specificity of 76% for this test. No serological reactions showed in 191 goats originating from 11 flocks which never had suffered from caseous lymphadenitis. Using this ELISA test 24 goats originating from flocks suffering from caseous lymphadenitis were examined serologically before and after vaccination with a bacterin. Before vaccination one of the five goats with clinical signs showed no positive reaction in ELISA. After vaccination all 24 animals showed positive reactions. Of a total of 1868 goats sampled in Baden-Wuerttemberg 41 (2.2%) in 22 (10%) flocks showed positive reactions in ELISA.

[Detection methods for Salmonella abortus ovis and examinations in sheep flocks in northern Baden-Württemberg]. / Nachweismethoden für Salmonella abortus ovis sowie Untersuchungen in Schafherden im nördlichen Baden-Württemberg.

Investigations into the incidence of Salmonella abortus ovis (S. abortus ovis) infections should not be neglected in diagnosis of ovine abortion cases. For detection of this pathogen agent, direct cultivation, as well as pre-enrichment combined with enrichment procedures in tetrathionate or Rappaport-Vassiliadis medium, should be performed with respect to the features of S. abortus ovis. The detection limit of S. abortus ovis using pre-enrichment and enrichment media could be determined using 6.5 x 10(3)-6.5 x 10(4) bacteria. For subsequent cultivation of S. abortus ovis Gassner, XLD or Rambach agar are suitable. In infected sheep showing no excretion of S. abortus ovis, the pathogen can be detected by serological studies using microagglutination or the ELISA test. The ELISA test proved to be more sensitive than the microagglutination test, detecting antibodies against S. abortus ovis in 17% of the 814 sheep tested. The microagglutination test revealed positive results in only 2% of the sheep tested.

[Significance of causes of infectious abortion in sheep flocks in northern Baden-Württemberg with special reference to Chlamydia psittaci]. / Die Bedeutung infektiöser Abortursachen in Schafherden im nördlichen Baden-Württemberg unter besonderer Berücksichtigung von Chlamydia psittaci.

Investigations on the reasons of infectious abortion cases in sheep flocks in northern parts of Baden-Wuerttemberg eludicate the wide-spreading of Chlamydia psittaci (C. psittaci) and its significance as the most frequent cause of abortions in sheep. Another important pathogen agent causing abortions is Salmonella abortus ovis (S. abortus ovis) which could be demonstrated by using a serological ELISA test. A less important role than C. psittaci and S. abortus ovis plays Coxiella burnetii in abortion of sheep. A high prevalence of Toxoplasma gondii and border disease infections in sheep flocks could be revealed by serological studies, too. Statistical evaluations of the obtained results demonstrate a significantly increased number of positively reacting female sheep in flocks suffering from abortions in comparison to those deriving from flocks devoid of abortion cases. Serological studies on leptospirosis and brucellosis exclude a participation of these pathogen agents in abortion cases in the investigated flocks at present. The significance of zoonosis originated from sheep is emphasized.

Isolation of serovar-specific leptospiral antigens for use in an enzyme-linked immunosorbent assay (ELISA) compared with the microscopic agglutination test and immunofluorescence.

A total of 10 different detergents were used with the intention of extracting serovar-specific antigens from eight leptospiral serovars for use in an ELISA. Extractions with 2% sodium taurocholate at 50 degrees C proved to be best suited for this purpose. Taurocholate extracted antigens could be separated in SDS-PAGE and antigenic activities demonstrated in immunoblotting. In comparative studies on sera taken from cattle, pigs, horses and dogs, the self-made ELISA proved to be more sensitive than the microscopic agglutination test (MAT) and the immunofluorescence test (IFT). The IFT gave different results from those obtained using MAT or ELISA.