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Biomedical subjects

B Hoffmann

Publications and source records attributed to B Hoffmann.

At least 19 recordsLinked to original sources

Age-dependent expression and localization of the progesterone receptor in the boar testis.

Expression of the progesterone receptor (PR) was monitored in testes of groups of five boars aged 50, 100, 150, 200 and 250 days. The primary monoclonal antibody used for immunohistochemistry (IHC) was raised against a peptide mapping the amino acids 922-933 of the carboxy-terminus of the human PR, negative controls were set up using an irrelevant monoclonal isotype-specific antibody, porcine endometrium served as positive control tissue. In parallel, qualitative reverse transcription-polymerase chain reaction was applied. Based on the developmental status of spermatogenesis the 50- and 100-day-old boars were considered as immature, the boars aged 200 and 250 days as mature. Positive and negative controls confirmed specificity of IHC. In the 50-day-old boars 85.1% of the prespermatogonia that had reached the basal lamina and 18.2% of the centrally located prespermatogonia stained positive, while it was 92.1% respectively 2.1% in the 100-day-old boars. The effect of time and location was highly significant (p < 0.005 resp. 0.0001). In mature boars between 77 and 80% of the A and B spermatogonia stained positive, there was no effect of boar age and stage of spermatogenesis. In both groups also few peritubular myoid cells stained positive. It is hypothesized that Leydig cell-derived progesterone plays a functional role in spermatogoniogenesis in a synergistic manner with Leydig cell-derived oestrogens.

Aging↗

Persistence of viral RNA in rabbits which overcome an experimental RHDV infection detected by a highly sensitive multiplex real-time RT-PCR.

An internally controlled multiplex real-time RT-PCR using TaqMan probes and external standards for absolute RNA quantification was developed as a new diagnostic tool for the detection of rabbit haemorrhagic disease virus (RHDV). The test revealed a specificity of 100%, an analytical sensitivity of 10 copies/well and a linearity over a range from 10(1) to 10(10) copies. The viral loads in organs, leukocytes, sera and excretions of seropositive, convalescent rabbits which were overcoming an experimental infection with RHDV were determined using the validated assay. As a result, viral RNA was demonstrated and quantified for at least 15 weeks. Thus, a persistence of viral RNA after experimental infection of rabbits could be shown for the first time. In contrast, neither antigen nor infectious virus could be detected by antigen-ELISA, immunohistochemistry or experimental transmission. Therefore, further experiments are necessary to prove that the persistence of RNA is linked with the persistence of infectious virus particles.

Animals↗

Detection of small pulmonary nodules in high-field MR at 3 T: evaluation of different pulse sequences using porcine lung explants.

To evaluate two MR imaging sequences for the detection of artificial pulmonary nodules inside porcine lung explants. 67 agarose nodules ranging 3-20 mm were injected into ten porcine lungs within a dedicated chest phantom. The signal on T1-weighted images and radiopacity were adjusted by adding 0.125 mmol/l Gd-DTPA and 1.5 g/l of iodine. A T1-weighted three-dimensional gradient-echo (T1-3D-GRE; TR/TE:3.3/1.1 ms, slice:8 mm, flip-angle:10 degrees ) and a T2-weighted half-Fourier fast-spin echo sequence (T2-HF-FSE; TR/TE:2000/66 ms, slice:7 mm, flip-angle:90 degrees ) were applied in axial orientation using a 3-T system (Intera, Philips Medical Systems, Best, The Netherlands), followed by CT (16x0.5 mm) as reference. Nodule sizes and locations were assessed by three blinded observers. In nodules of >10 mm, sensitivity was 100% using 3D-GRE-MRI and 94% using the HF-FSE sequence. For nodules 6-10 mm, the sensitivity of MRI was lower than with CT (3D-GRE:92%; T2-HF-FSE:83%). In lesions smaller than 5 mm, the sensitivity declined to 80% (3D-GRE) and 53% (HF-FSE). Small lesion diameters were overestimated with both sequences, particularly with HF-FSE. This study confirms the feasibility of 3 T-MRI for lung nodule detection. In lesions greater than 5 mm, the sensitivity of the 3D-GRE sequence approximated CT (>90%), while sensitivity and PPV with the HF-FSE sequence were slightly inferior.

Animals↗

Characterization of the canine 3beta-hydroxysteroid dehydrogenase and its expression in the corpus luteum during diestrus.

3beta-Hydroxysteroid dehydrogenase (3betaHSD) is a key enzyme in the synthesis of bioactive steroid hormones. Objectives of the present study were to clone canine 3betaHSD and to investigate its expression in dog corpora lutea (CL) covering the periods of their formation, early and late regression (days 5, 15, 25, 35, 45, 65 after ovulation). Complete complementary DNA sequence was amplified by RACE PCR. Subsequent cloning revealed that the canine ovarian 3betaHSD transcript was composed of a 5'-untranslated region (5'-UTR) of 126 nucleotides, an open reading frame (ORF) of 1122 nucleotides and a 3'-UTR of 441 nucleotides. The putative ORF encoded a 374 amino acid protein which remains highly conserved (79-85% identity) between species. The transient expression of the cloned canine 3betaHSD in a mammalian heterologous cell expression system (HEK293T cells) identified the 3betaHSD activity as the only activity of this canine enzyme (absence of any detectable 17-hydroxysteroid dehydrogenase activity). Qualitative RT-PCR revealed expression of 3betaHSD on all days investigated and the signals were strongest on days 5 and 15, with day 25 intensity tending to decrease. However, variability between individual animals was high. The significant decrease in the expression of 3betaHSD towards the end of diestrus as indicated by Real Time PCR (p<0.01) and immunhistochemistry may indicate that the provision of progesterone is controlled by availability of the enzyme rather than the substrate.

17-Hydroxysteroid Dehydrogenases↗

Effect of ACTH (tetracosactide) on steroid hormone levels in the mare. Part A: effect in intact normal mares and mares with possible estrous related behavioral abnormalities.

Ovariectomized mares and mares with inactive ovaries may show signs of estrus. The reason behind this phenomenon is not clear; however, steroid hormones of adrenal origin have been suggested. Moreover, aberrant adrenal hormone production has been implied as a reason why some intact mares may change behavior. In the present study, the effect of ACTH on plasma levels of cortisol, progesterone, androstenedione and testosterone was investigated in intact mares with normal estrous behavior ('controls', n=5) and intact mares that according to their owners showed deviant estrous behavior ('problem' mares, n=7). Blood samples were collected hourly from 12:00 h until 14:00 h the following day (half-hourly between 14:00 and 17:00 h) on two occasions (at two estruses), with saline or ACTH treatment (tetracosactide) at 14:00 h (saline treatment day or ACTH treatment day). ACTH treatment caused a significant increase in plasma levels of cortisol, progesterone, androstenedione and testosterone in all mares (P<0.05). An overall significant difference in cortisol response to ACTH was found (P<0.05), with 'problem' mares showing a significantly lower increase in cortisol levels 30 min to 3h post ACTH treatment (P<0.001). The 'problem' mares also showed a significantly higher increase than controls in progesterone levels in the same time period (P<0.05). The reason for the reduced adreno-cortical reactivity, with a low cortisol response to the ACTH treatment, in the 'problem' mares is unknown, but may indicate a difference in adrenal function as compared to control mares.

Androstenedione↗

Effect of ACTH (tetracosactide) on steroid hormone levels in the mare. Part B: effect in ovariectomized mares (including estrous behavior).

The mare is the only non-primate species known to display estrous signs after ovariectomy and adrenal hormones have been implicated as a possible cause. Moreover, in several species, estradiol seems to have a stimulatory effect on the hypothalamic-pituitary-adrenal axis. The aim of the present study was to compare the effect of ACTH (tetracosactide) on pertinent hormones [cortisol, progesterone, androstenedione, testosterone (intact and ovariectomized mares) and estradiol (ovariectomized mares only)] in intact mares in estrus with the same mares after ovariectomy (n=5). Blood samples were collected hourly from 12:00 until 14:00 h the following day (half-hourly between 14:00 and 17:00 h) on two occasions, with saline or ACTH treatment at 14:00 h (saline treatment day or ACTH treatment day). The mares, both when intact and after ovariectomy, showed a significant increase in all measured hormones, except estradiol (not measured in intact mares), after ACTH treatment, lasting at least 3h post-treatment (P<0.001). On the saline treatment day, cortisol levels in ovariectomized mares were lower than in intact mares in the evening (18:00-23:00 h), but higher at night (24:00-05:00 h). No differences in cortisol response between mares, when intact and after ovariectomy, were found after ACTH treatment (P=0.3). Androstenedione levels were lower (P<0.001) and increased less after ACTH treatment in ovariectomized mares, as compared to when intact (P<0.05). Progesterone concentrations were lower in the ovariectomized mares at night (24:00-05:00 h) on the saline treatment day and at all times on the ACTH treatment day (P<0.05). Testosterone concentrations were lower in ovariectomized mares on both treatment days, as compared to when intact (P<0.001). It was concluded that ovariectomy affected basal cortisol pattern. Ovarian androstenedione and testosterone contributed to the basal circulating levels and, in the case of androstenedione, was stimulated by ACTH. Endogenous estradiol did not act stimulatory on adrenal gland hormone production in the mare.

Androstenedione↗

Foot-and-mouth disease virus: a first inter-laboratory comparison trial to evaluate virus isolation and RT-PCR detection methods.

Five European reference laboratories participated in an exercise to evaluate the sensitivity and specificity of their routinely employed RT-PCR tests and cell cultures for the detection and isolation of foot-and-mouth disease (FMD) virus. Five identical sets of 20 coded samples were prepared from 10 vesicular epithelia, which were derived from submissions from suspect cases of FMD or swine vesicular disease (SVD). Sixteen samples were derived from six FMD virus positive epithelia representing four different serotypes (two each of types O and A and one each of types Asia 1 and SAT 2), two from samples which had been found to be negative by antigen ELISA and virus isolation (VI) in cell culture and two from SVD virus positive epithelia. Some of the FMD virus positive samples were prepared from 10-fold serial dilutions of three of the initial suspensions. Each laboratory tested the samples by one or more of its available RT-PCR procedures and inoculated cell cultures that it routinely uses for FMD diagnosis in attempts to isolate virus, the specificity of which was confirmed by antigen ELISA. The best of the RT-PCR assays used in each laboratory gave comparable results while the sensitivity of cell cultures was variable from high in one laboratory, moderate in two and low in two others. This prototype panel of samples would appear suitable for external quality assurance of these tests but would benefit from the inclusion of more negative samples and an extension in the serial dilution range of one or more of the FMD positive sample titration series.

Animals↗

A universal heterologous internal control system for duplex real-time RT-PCR assays used in a detection system for pestiviruses.

A heterologous in vitro transcript based on a specific primer-probe HEX system was generated as a universal internal control (IC) to improve virus-specific real-time reverse-transcriptase PCR (RT-PCR) assays. By using a set of different primers, several PCR fragments of desired sizes of an in vitro transcript of the enhanced green fluorescent protein (EGFP) gene were generated, and the fragments were detected using a HEX-labelled probe. For long-term storage of the in vitro transcript a special RNA-safe buffer (RSB) was developed. Freezing and thawing of the IC diluted in RSB did not result in any substantial loss of detectable IC copy numbers. The new IC system was used for the first time in a duplex real-time RT-PCR assay for the detection of pestivirus-derived RNA, in particular from bovine viral diarrhea virus (BVDV). Primers and TaqMan probes for the 'panpesti' assay were selected by analysing the consensus sequence of the 5' non-translated region (5' NTR) of more than 600 different pestiviruses. Finally, the optimised primer probe combination showed an analytical sensitivity of less than 10 copies/reaction. In the duplex set-up, the analytical sensitivity of the validated real-time RT-PCR was identical to the sensitivity of the single assay without IC, and the diagnostic sensitivity of the duplex assay was equal or higher if compared to virus isolation.

Animals↗

[Ultrasound education by simulator training high acceptance by ultrasound trainees of all qualification levels].

Ultrasound education improves by simulator training, however, the acceptance of ultrasound simulator training has not been investigated. Therefore we evaluated the participants response to simulator education during an ultrasound course of abdominal emergencies at an international ultrasound congress, at a regular ultrasound course and during courses at a regional hospital and an university hospital. Altogether, 62.3% of the participants judged the simulator image quality to be good. 84.3% considered the case selection to be good and 92.7% of the participants viewed the educational benefit to be good. 98.5% whished to have further ultrasound simulator courses to be developed and 95.3% of the participants opted for the integration of simulator training into the conventional ultrasound education.

Attitude of Health Personnel↗

Expression of cyclooxygenase-II (COX-II) and 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD)/prostaglandin F-synthase (PGFS) in bovine placentomes: implications for the initiation of parturition in cattle.

The chain of events leading to prepartal luteolysis in cattle is not yet fully understood. Prostaglandin F(2alpha) (PGF(2alpha)) seems to be a major factor involved. However, only little information is available about the underlying regulatory mechanisms. Consequently, the expression of cyclooxygenase-II (COX-II) and 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD), an enzyme recently shown to be most likely responsible for the production of luteolytic PGF(2alpha) in the endometrium of cyclic cows, was investigated in bovine placentomes. Immunohistochemical methods were applied to placentomes from 17 pregnant cows between days 100 and 284, from three cows during the prepartal progesterone decrease (days 273-282) and from five parturient cows. COX-II was found in uninucleated trophoblast cells (UTC) from day 100 until parturition. However, between days 100 and 235 expression was only weak to moderate, focal and mainly restricted to the chorionic plate and adjacent basal parts of chorionic stem villi. In placentomes from a 270 and a 284 day pregnant cow, in which the prepartal decline of progesterone levels had not started yet, staining had substantially increased and extended to secondary and tertiary chorionic villi. In prepartal and parturient cows strong to intense staining was present in UTC all over the villous tree. Real time RT-PCR confirmed an extensive pre- and intrapartal rise of COX-II expression in bovine placentomes with a 70-100-fold increase of COX-II-mRNA levels. 20alpha-HSD/PGFS was widely expressed in UTC of chorionic villi. Like COX-II it was down-regulated in UTC differentiating into trophoblast giant cells. Immunostaining pattern did not change remarkably during the period under investigation, and 20alpha-HSD/PGFS-mRNA levels increased only 2.6-fold in the prepartal phase. Thus, in UTC PGF(2alpha) may be produced via COX-II and 20alpha-HSD/PGFS, but only COX-II may be substantially involved in the control of a putative prepartal placentomal output of luteolytic PGs, whereas 20alpha-HSD/PGFS seems to be expressed in a more constitutive manner.

20-alpha-Hydroxysteroid Dehydrogenase↗

Acute neuronopathic Gaucher disease complicated by fatal gastrointestinal bleeding.

Gaucher disease, a rare lysosomal storage disease caused by deficiency of glucocerebrosidase, may present with gastrointestinal bleeding. We report about an 11-month-old boy suffering from acute neuronopathic Gaucher disease who died after massive gastrointestinal bleeding. A gastric ulcer was found as the sole bleeding source. The gastric mucosa showed marked infiltration with Gaucher cells, in particular around the ulcer. Alterations of the gastrointestinal mucosa offer a new explanation for gastrointestinal bleedings in this disease.

Fatal Outcome↗

A virological survey in migrating waders and other waterfowl in one of the most important resting sites of Germany.

Wild birds are considered a potential reservoir or a carrier of viral diseases and may therefore play a role in the epidemiology of economically important or zoonotic diseases. In 2001 and 2002, a survey with special emphasis on virus isolation in migrating waders and some other birds were conducted. In one of the most important inland resting sites for migratory waterfowl, tracheal and cloacal swabs were collected from 465 waders representing 19 different species, and 165 other birds that were not captured on purpose. A total of 42 avian viruses were isolated, 34 of these were identified as paramyxoviruses (PMVs). The majority of isolates came from waders and wild ducks, and were characterized as PMV-1. In contrast, PMV-4 was found in wild ducks only, PMV-6 was mainly detected in wader species. Four avian influenza viruses (AIVs), belonging to H4 and H3 haemagglutinin subtype, were isolated from wild duck species. Furthermore, four reo-like viruses were isolated from one particular wader species for the first time. The majority of virus positive birds were <1 year old and did not show any clinical symptoms. There was no evidence for the presence of West Nile virus in these birds. These results confirm that the restricted resting sites in Western Europe must be considered as important locations for the intra- and interspecies transmission of avian viruses.

Animal Migration↗

Diagnostic evaluation of a real-time RT-PCR assay for routine diagnosis of classical swine fever in wild boar.

The aim of the study was to evaluate a real-time reverse transcription-polymerase chain reaction (RT-PCR) assay for routine diagnosis of classical swine fever (CSF) in wild boar by using 1000 spleen homogenates that were tested previously negative by virus isolation and 26 homogenates from which CSF virus could be isolated. All 26 positive samples in the virus isolation assay also were found to be positive in real-time RT-PCR. Additionally further 10 samples were detected by real-time RT-PCR out of the 1000 negative samples in the virus isolation. With a commercial CSF antigen-ELISA only 14 out of the 36 real-time RT-PCR positive samples could be detected. The sequence analysis of all ten samples that tested positive by real-time RT-PCR and negative by virus isolation revealed CSF virus-specific sequences. Based on the assumption that all samples with a CSF virus-specific sequence or positive in the virus isolation test originated from truly CSF virus-infected wild boar, the following sensitivity values were calculated as antigen-ELISA, 39%; virus isolation, 72% and real-time RT-PCR, 100%. The use of real-time RT-PCR instead of antigen-ELISA and virus isolation as a routine tool for control and eradication of CSF in wild boar populations is recommended.

Animals↗

Reciprocal expression of 17alpha-hydroxylase-C17,20-lyase and aromatase cytochrome P450 during bovine trophoblast differentiation: a two-cell system drives placental oestrogen synthesis.

No definitive information is yet available on the steroidogenic capacity of the two morphologically distinct cell types forming the bovine trophoblast, the uninucleated trophoblast cells (UTCs) and the trophoblast giant cells (TGCs). Hence, in order to localise 17alpha-hydroxylase-C17,20-lyase (P450c17) on a cellular level and to monitor its expression as a function of gestational age, placentomes from pregnant (days 80-284; n = 19), prepartal (days 273-282; 24-36 h prior to the onset of labour; n = 3) and parturient cows (n = 5) were immunostained for P450c17 using an antiserum against the recombinant bovine enzyme. At all stages investigated, P450c17 was exclusively found in the UTCs of chorionic villi (CV), where staining was ubiquitous between days 80 and 160, but was largely restricted to primary CV and the branching sites of secondary CV between days 160 and 240. Thereafter, a distinct ubiquitous staining reoccurred in the UTCs of all CV in late pregnant, prepartal and parturient animals. Using an antiserum against human aromatase cytochrome P450 (P450arom), specific cytoplasmic staining was observed in TGCs. In placentomes from pregnant cows, staining intensity was higher in mature compared with immature TGCs and was more pronounced in the trophoblast covering big stem villi compared with the trophoblast at other sites of the villous tree. In placentomes of a parturient cow, specific staining was only found in mature TGCs that survived the normal, but substantial, prepartal decline in TGC numbers. These results clearly showed that bovine UTCs and TGCs exhibit different steroidogenic capacities, constituting a 'two-cell' organisation for oestrogen synthesis. P450c17 expression appears to be quickly down-regulated and P450arom is up-regulated when UTCs enter the TGC differentiation pathway.

Animals↗

Case report: the significance of genotyping for the epidemiological tracing of classical swine fever (CSF).

In Germany, eleven outbreaks of CSF in domestic pig holdings were reported in 2002. They occurred exclusively in regions where CSF virus circulated in the wild boar population. In ten cases the phylogenetic analysis revealed that the isolates from domestic pigs and wild boar had identical sequences in the 5' non-translated region (5'NTR). However, in one case a subtype was isolated which was slightly different from the virus subtype found in the wild boar population of that region. This case is decribed in detail. The epidemiological significance of different diagnostic methods is discussed, in particular the genetic typing of CSF virus isolates.

Animals↗

Influence of sex on the resistance of sheep lambs to an experimental Haemonchus contortus infection.

36 intact male and 69 female lambs of two breeds (n = 63 Merinoland; n = 42 Rhön) aged 12 weeks were orally infected with 5000 infective-stage larvae, L3, of the nematode Haemonchus contortus. After 4 and 8 weeks faecal egg counts (FEC), haematocrit values and plasma testosterone levels were determined. All lambs were slaughtered at 20 weeks of age. The gastrointestinal tracts were examined for the presence of adult stages of H. contortus. Male lambs showed significantly higher log FEC (p < 0.001), higher mean establishment rates (p < 0.05), higher worm burdens (p < 0.01) and lower haematocrit values (p < 0.001) when compared with female lambs. Correlations between economically important traits (body weight, daily weight gain) and parasitological parameters were significantly higher in male animals. Testosterone level was 4 weeks after infection significantly positive correlated with worm burden. The results suggest that female lambs are more resistant against an experimental H. contortus infection when compared with male lambs. Testosterone seems to play an important role in resistance. This approach can be of importance if parasite resistance is incorporated into breeding programs and the estimated breeding values for rams are only based on male offspring information. Therefore male breeding values are probably not representative for the whole population.

Animals↗

Fibroblast growth factor (FGF)-1, FGF2, FGF7 and FGF receptors are uniformly expressed in trophoblast giant cells during restricted trophoblast invasion in cows.

The bovine placenta is characterized by a limited invasion of trophoblast giant cells (TGC). In contrast to mononuclear trophoblast cells (MTC), TGC are non-polarized cells, which migrate and fuse with single uterine epithelial cells throughout gestation. Fibroblast growth factors (FGF) were shown to be associated with the migratory activity of cells, cell differentiation and angiogenesis, and due to its localization in trophoblast cells were proposed as important regulating factors in hemochorial placentae of rodents and humans, and the (syn)epitheliochorial placenta of pig and sheep. Since migrating bovine TGC are of epithelial origin, but exhibit similarities to mesenchymal cells we hypothesize that the restricted trophoblast invasion in cattle is characterized by a specific FGF expression pattern. Therefore, the spatiotemporal expression of specific FGF factor:receptor pairs, either acting on cells of mesenchymal origin or on epithelial cells was examined in bovine placental tissues throughout gestation and prepartum by immunohistochemistry, semiquantitative RT-PCR and in situ hybridization. FGF1 protein was found in trophoblast, caruncular epithelium (CE) and stroma (CS), stroma of chorionic villi (SCV), and in fetal and maternal blood vessels. FGF2 signals dominated in maternal vascular endothelia (VE), immature TGC, and MTC, whereas staining in other cell types was clearly weaker. FGF7 protein was detected in fetal and maternal blood vessel as well as in immature TGC and MTC predominantly at the chorionic plate. FGFR immunoreaction was localized in immature TGC, MTC, and to a clearly lesser extent in CS, CE and fetal and maternal blood vessels. Mature TGC stained negatively for all examined factors and FGFR. The corresponding mRNAs specific for FGF1, -2, -7, total FGFR, and FGFR2 isoforms IIIb and IIIc were colocalized in immature TGC, whereas hybridization was substantially lower in CE and absent in CS, SCV and mature TGC throughout gestation, but switched to CS and VE immediately prepartum. Semiquantitative RT-PCR revealed higher mRNA levels for FGF1, FGFR, and FGFR2IIIc in cotyledons compared to caruncles (p<0.05), whereas it was the opposite with FGF2 (p<0.001). FGF7 and FGFR2IIIb mRNA levels did not differ between caruncles and cotyledons. Significant changes (p<0.05) of mRNA levels related to gestational age were found for FGF1 and FGFR2IIIc, but not for FGF2, -7, total FGFR, and FGFR2IIIb. The specific localization of all examined FGF family members in TGC suggests that TGC, apart from their classical function as producers of hormonal products, play other important roles in the regulation of bovine placentomal growth, differentiation and angiogenesis.

Animals↗

Validation of a real-time RT-PCR assay for sensitive and specific detection of classical swine fever.

A fully validated, ready-to-use, real-time reverse transcription-polymerase chain reaction (RT-PCR) assay, multiplexed for simultaneous detection of an internal control, for the simple and rapid diagnosis of classical swine fever (CSF) was developed. Primers and FAM-labeled TaqMan-probes specific for classical swine fever virus (CSFV) were selected from the consensus sequence of the 5' non-translated region (5' NTR) of 78 different CSFV strains. For determining analytical sensitivity, an in vitro transcript (T7-PC3alf) of the 5' NTR was constructed and tested. In addition, the T7-PC3alf transcript was further used as a positive control and a standard for quantitation of CSFV genome copies. A second heterologous in vitro transcript based on a specific primer-probe HEX-system was designed as an internal positive control for the RNA isolation step and RT-PCR. By using limited primer concentrations for the internal control, no adverse effects on the sensitivity of the CSF-system could be observed, and the newly designed duplex real-time RT-PCR proved to have a sensitivity of approximately eight copies. The primer-probe combination selected was strictly CSFV-specific and no amplification was observed in all non-CSFV pestiviruses tested.

5' Untranslated Regions↗