Clinical decision support system supported interventions in hospitalized older patients: a matter of natural course and adequate timing.

BACKGROUND: Drug-related problems (DRPs) and potentially inappropriate prescribing (PIP) are associated with adverse patient and health care outcomes. In the setting of hospitalized older patients, Clinical Decision Support Systems (CDSSs) could reduce PIP and therefore improve clinical outcomes. However, prior research showed a low proportion of adherence to CDSS recommendations by clinicians with possible explanatory factors such as little clinical relevance and alert fatigue. OBJECTIVE: To investigate the use of a CDSS in a real-life setting of hospitalized older patients. We aim to (I) report the natural course and interventions based on the top 20 rule alerts (the 20 most frequently generated alerts per clinical rule) of generated red CDSS alerts (those requiring action) over time from day 1 to 7 of hospitalization; and (II) to explore whether an optimal timing can be defined (in terms of day per rule). METHODS: All hospitalized patients aged ≥ 60 years, admitted to Zuyderland Medical Centre (the Netherlands) were included. The evaluation of the CDSS was investigated using a database used for standard care. Our CDSS was run daily and was evaluated on day 1 to 7 of hospitalization. We collected demographic and clinical data, and moreover the total number of CDSS alerts; the total number of top 20 rule alerts; those that resulted in an action by the pharmacist and the course of outcome of the alerts on days 1 to 7 of hospitalization. RESULTS: In total 3574 unique hospitalized patients, mean age 76.7 (SD 8.3) years and 53% female, were included. From these patients, in total 8073 alerts were generated; with the top 20 of rule alerts we covered roughly 90% of the total. For most rules in the top 20 the highest percentage of resolved alerts lies somewhere between day 4 and 5 of hospitalization, after which there is equalization or a decrease. Although for some rules, there is a gradual increase in resolved alerts until day 7. The level of resolved rule alerts varied between the different clinical rules; varying from > 50-70% (potassium levels, anticoagulation, renal function) to less than 25%. CONCLUSION: This study reports the course of the 20 most frequently generated alerts of a CDSS in a setting of hospitalized older patients. We have shown that for most rules, irrespective of an intervention by the pharmacist, the highest percentage of resolved rules is between day 4 and 5 of hospitalization. The difference in level of resolved alerts between the different rules, could point to more or less clinical relevance and advocates further research to explore ways of optimizing CDSSs by adjustment in timing and number of alerts to prevent alert fatigue.

Immunohistochemical biomarkers are prognostic relevant in addition to the ESMO-ESGO-ESTRO risk classification in endometrial cancer.

OBJECTIVE: Pre-operative immunohistochemical (IHC) biomarkers are not incorporated in endometrial cancer (EC) risk classification. We aim to investigate the added prognostic relevance of IHC biomarkers to the ESMO-ESGO-ESTRO risk classification and lymph node (LN) status in EC. METHODS: Retrospective multicenter study within the European Network for Individualized Treatment of Endometrial Cancer (ENITEC), analyzing pre-operative IHC expression of p53, L1 cell-adhesion molecule (L1CAM), estrogen receptor (ER) and progesterone receptor (PR), and relate to ESMO-ESGO-ESTRO risk groups, LN status and outcome. RESULTS: A total of 763 EC patients were included with a median follow-up of 5.5-years. Abnormal IHC expression was present for p53 in 112 (14.7%), L1CAM in 79 (10.4%), ER- in 76 (10.0%), and PR- in 138 (18.1%) patients. Abnormal expression of p53/L1CAM/ER/PR was significantly related with higher risk classification groups, and combined associated with the worst outcome within the 'high and advanced/metastatic' risk group. In multivariate analysis p53-abn, ER/PR- and ESMO-ESGO-ESTRO 'high and advanced/metastatic' were independently associated with reduced disease-specific survival (DSS). Patients with abnormal IHC expression and lymph node metastasis (LNM) had the worst outcome. Patients with LNM and normal IHC expression had comparable outcome with patients without LNM and abnormal IHC expression. CONCLUSION: The use of pre-operative IHC biomarkers has important prognostic relevance in addition to the ESMO-ESGO-ESTRO risk classification and in addition to LN status. For daily clinical practice, p53/L1CAM/ER/PR expression could serve as indicator for surgical staging and refine selective adjuvant treatment by incorporation into the ESMO-ESGO-ESTRO risk classification.

Does probability guided hysteroscopy reduce costs in women investigated for postmenopausal bleeding?

OBJECTIVE: To evaluate whether a model to predict a failed endometrial biopsy in women with postmenopausal bleeding (PMB) and a thickened endometrium can reduce costs without compromising diagnostic accuracy. DESIGN, SETTING, AND POPULATION: Model based cost-minimization analysis. METHODS: A decision analytic model was designed to compare two diagnostic strategies for women with PMB: (I) attempting office endometrial biopsy and performing outpatient hysteroscopy after failed biopsy and (II) predicted probability of a failed endometrial biopsy based on patient characteristics to guide the decision for endometrial biopsy or immediate hysteroscopy. Robustness of assumptions regarding costs was evaluated in sensitivity analyses. MAIN OUTCOME MEASURES: Costs for the different strategies. RESULTS: At different cut-offs for the predicted probability of failure of an endometrial biopsy, strategy I was generally less expensive than strategy II. The costs for strategy I were always € 460; the costs for strategy II varied between € 457 and € 475. At a 65% cut-off, a possible saving of € 3 per woman could be achieved. CONCLUSIONS: Individualizing the decision to perform an endometrial biopsy or immediate hysteroscopy in women presenting with postmenopausal bleeding based on patient characteristics does not increase the efficiency of the diagnostic work-up.

Evaluation of a multifaceted medication review in older patients in the outpatient setting: a before-and-after study.

BACKGROUND: The prevalence of medication-related emergency department visits and acute hospital admissions in older patients is rising due to the ageing of the population and increasing prevalence of multimorbidity and associated polypharmacy. AIM: To explore whether a combined medication review performed in the outpatient setting reduces the number of medication-related emergency department visits and hospital (re)admissions. METHOD: All consecutive patients visiting the geriatric outpatient clinic underwent a multifaceted medication review (i.e. evaluation by at least a geriatrician, and/or pharmacist and use of clinical decision support system). Subsequently, we analysed the number of, and reason for, emergency department visits, acute hospital admissions and readmissions in the year prior to and the year following the index-date (date of first presentation and medication review). RESULTS: A multifaceted medication review reduced the number of potentially medication-related emergency department visits (38.9% vs. 19.6%, p < 0.01), although the total number of ED visits or acute hospital admissions per patient in the year before and after medication review did not differ. CONCLUSION: A multifaceted medication review performed in the outpatient clinic reduced the number of potentially medication-related emergency department visits and could therefore reduce negative health outcomes and healthcare costs.

Evaluation of Potency and Duration of Immunity Elicited by a Multivalent FMD Vaccine for Use in South Africa.

South Africa (SA) experiences sporadic foot and mouth disease (FMD) outbreaks irrespective of routine prophylactic vaccinations of cattle using imported commercial vaccines. The problem could be mitigated by preparation of vaccines from local virus strains related to those circulating in the endemically infected buffalo populations in the Kruger National Park (KNP). This study demonstrates the individual number of protective doses (PD) of five vaccine candidate strains after homologous virus challenge, as well as the vaccines safety and onset of humoral immunity in naïve cattle. Furthermore, the duration of post-vaccination immunity over a 12-month period is shown, when a multivalent vaccine prepared from the five strains is administered as a primary dose with or without booster vaccinations. The five monovalent vaccines were shown to contain a 50% PD between 4 and 32, elicit humoral immunity with antibody titers ≥2.0 log10 from day 7 post-vaccination, and cause no adverse reactions. Meanwhile, the multivalent vaccine elicited antibody titers ≥2.0 log10 and clinical protection up to 12 months when one or two booster vaccinations were administered within 6 months of the primary vaccination. An insignificant difference between the application of one or two booster vaccinations was revealed. Owing to the number of PDs, we anticipate that the multivalent vaccine could be used successfully for prophylactic and emergency vaccinations without adjustment of the antigen payloads. Furthermore, a prophylactic vaccination regimen comprising primary vaccination of naïve cattle followed by two booster vaccinations 1.5 and 6 months later could potentially maintain herd immunity over a period of 12 months.

Time-resolved FRET fluorescence spectroscopy of visible fluorescent protein pairs.

Förster resonance energy transfer (FRET) is a powerful method for obtaining information about small-scale lengths between biomacromolecules. Visible fluorescent proteins (VFPs) are widely used as spectrally different FRET pairs, where one VFP acts as a donor and another VFP as an acceptor. The VFPs are usually fused to the proteins of interest, and this fusion product is genetically encoded in cells. FRET between VFPs can be determined by analysis of either the fluorescence decay properties of the donor molecule or the rise time of acceptor fluorescence. Time-resolved fluorescence spectroscopy is the technique of choice to perform these measurements. FRET can be measured not only in solution, but also in living cells by the technique of fluorescence lifetime imaging microscopy (FLIM), where fluorescence lifetimes are determined with the spatial resolution of an optical microscope. Here we focus attention on time-resolved fluorescence spectroscopy of purified, selected VFPs (both single VFPs and FRET pairs of VFPs) in cuvette-type experiments. For quantitative interpretation of FRET-FLIM experiments in cellular systems, details of the molecular fluorescence are needed that can be obtained from experiments with isolated VFPs. For analysis of the time-resolved fluorescence experiments of VFPs, we have utilised the maximum entropy method procedure to obtain a distribution of fluorescence lifetimes. Distributed lifetime patterns turn out to have diagnostic value, for instance, in observing populations of VFP pairs that are FRET-inactive.

Structural changes of yellow Cameleon domains observed by quantitative FRET analysis and polarized fluorescence correlation spectroscopy.

Förster resonance energy transfer (FRET) is a widely used method for monitoring interactions between or within biological macromolecules conjugated with suitable donor-acceptor pairs. Donor fluorescence lifetimes in absence and presence of acceptor molecules are often measured for the observation of FRET. However, these lifetimes may originate from interacting and noninteracting molecules, which hampers quantitative interpretation of FRET data. We describe a methodology for the detection of FRET that monitors the rise time of acceptor fluorescence on donor excitation thereby detecting only those molecules undergoing FRET. The large advantage of this method, as compared to donor fluorescence quenching method used more commonly, is that the transfer rate of FRET can be determined accurately even in cases where the FRET efficiencies approach 100% yielding highly quenched donor fluorescence. Subsequently, the relative orientation between donor and acceptor chromophores is obtained from time-dependent fluorescence anisotropy measurements carried out under identical conditions of donor excitation and acceptor detection. The FRET based calcium sensor Yellow Cameleon 3.60 (YC3.60) was used because it changes its conformation on calcium binding, thereby increasing the FRET efficiency. After mapping distances and orientation angles between the FRET moieties in YC3.60, cartoon models of this FRET sensor with and without calcium could be created. Independent support for these representations came from experiments where the hydrodynamic properties of YC3.60 under ensemble and single-molecule conditions on selective excitation of the acceptor were determined. From rotational diffusion times as found by fluorescence correlation spectroscopy and consistently by fluorescence anisotropy decay analysis it could be concluded that the open structure (without calcium) is flexible as opposed to the rather rigid closed conformation. The combination of two independent methods gives consistent results and presents a rapid and specific methodology to analyze structural and dynamical changes in a protein on ligand binding.

Survival of 125iodine-labeled Factor VIII in normals and patients with classic hemophilia. Observations on the heterogeneity of human Factor VIII.

Radiolabeled human Factor VIII was used to study its survival in normals and patients with classic hemophilia, and to study the heterogeneity of Factor VIII; Purified Factor VIII was radiolabeled with 125iodine (125I-VIII) without loss of its structural integrity. The survival of 125I-VIII was studied in six normals and six hemophiliacs of whom four of the hemophiliacs had received transfusions with normal cryoprecipitate before the 125I-VIII infusion. No significant difference was observed between the disappearance of Factor VIII coagulant activity and radioactivity in these hemophiliacs. 125I-VIII in plasma showed a biphasic disappearance with an average t1/2 of 2.9 +/- 0.4 h (SEM) for the first phase and 18.6 +/- 0.7 h (SEM) for the second phase, respectively. The survival of 125I-VIII was similar comparing normals and hemophiliacs. The highest molecular weight forms of Factor VIII disappear more rapidly than the lower molecular weight ones. This was established by analysis of the fractions obtained by gel chromatography of plasma collected at several times after infusion and by analysis of the in vivo disappearance of three subfractions of Factor VIII. The fraction of 125I-VIII binding to platelets in the presence of ristocetin (containing the highest molecular weight forms of Factor VIII including the ristocetin cofactor) represented about 50% of the radioactivity present in plasma after infusion and showed a t 1/2 of 11.7 +/- 0.9 h (SEM) for the second phase. The fraction, which was recovered in cryoprecipitate of the recipient's plasma, represented about 90% of the initial radioactivity and showed a t 1/2 of 16.3 +/- 0.8 h (SEM) for the second phase. The fraction of 125I-VIII remaining in the cryosupernatant plasma (containing low molecular weight forms of Factor (VIII) showed a t 1/2 of 27.2 +/- 1.1 h (SEM). The first phase of the disappearance of 125I-VIII is caused in part by the disappearance of the highest molecular weight forms, which are possibly removed by the reticuloendothelial system.

Veterinary vaccines for public health and prevention of viral and bacterial zoonotic diseases.

To meet with the increasing demand for food, the scale of world food production is increasing, as is the transport of animals and food products. At the same time, the contact of animals with the environment remains unchanged or, in the case of free-ranging animals, is even increasing. A number of microorganisms have established themselves in farmed animals, which although relatively harmless to animals are pathogenic to man. In this article, the options for reducing the risk of transferring zoonotic agents from animals (particularly farm animals) to man using veterinary vaccines against viral and bacterial diseases are described.

Intra-luminal gene therapy in the porcine artery using a recombinant adeno-associated virus 9.

The ability to improve or restore blood flow and promote healing in ischemic tissue has many potential clinical applications. Augmentation by direct delivery of growth factors may further enhance results, but requires a method for sustained delivery. In this study, we have tested the ability of adeno-associated virus 9 (AAV9) delivered within the lumen of a porcine artery to transfect the vessel and produce a desired product. The marker chosen was green fluorescent protein (GFP) (Ke et al., 2011). In 4 farm pigs the cranial tibial artery was surgically exposed. The vessel was temporarily clamped proximally, and divided distally. A cannula was placed intraluminally, and the arterial segment was injected with 1×10E13 particles of AAV9.CB7.CI.GFP·WPRE.rBG. At 14days the transfected cranial tibial artery as well as the liver, spleen and kidneys were harvested. ELISA and reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) were used to analyze the artery for GFP production. Significant GFP expression was seen in all transfected cranial tibial vessels, as determined by both GFP protein production (ELISA) and mRNA (RT-qPCR). No GFP was identified in liver, spleen or kidney, nor in the no-GFP control animal artery. Adeno-associated virus 9 is an appropriate vector for gene therapy experiments in the porcine artery model. This vector, and the intraluminal deliver method described result in robust gene expression at 2weeks without evident systemic spill of the virus. The ability to limit delivery of the gene to an isolated segment of vessel is desirable for future research applications.

A Structured Assessment to Decrease the Amount of Inconclusive Endometrial Biopsies in Women with Postmenopausal Bleeding.

OBJECTIVE: To determine whether structured assessment of outpatient endometrial biopsies decreases the number of inconclusive samples. DESIGN: Retrospective cohort study. SETTING: Single hospital pathology laboratory. POPULATION: Endometrial biopsy samples of 66 women with postmenopausal bleeding, collected during the usual diagnostic work-up and assessed as insufficient for a reliable histological diagnosis. METHODS: Endometrial biopsy samples were requested from the pathology laboratories. The retrieved samples were systematically reassessed by a single pathologist specialized in gynecology. MAIN OUTCOME MEASURE: Disagreement between initial assessment and conclusion after structured reassessment. RESULTS: We retrieved 36 of 66 endometrial biopsy samples from six different pathology laboratories. Structured reassessment of the retrieved samples by a single pathologist specialized in gynecology did not change the conclusion in 35 of the 36 samples. The remaining sample contained a large amount of endometrial tissue and the diagnosis at reassessment was endometrial hyperplasia without atypia. All other samples contained insufficient material for a reliable diagnosis. CONCLUSION: A structured reassessment of endometrial biopsies samples, which were classified as inconclusive due to insufficient material, did not change the conclusion. Although it might be helpful for pathologists to have diagnostic criteria for adequacy and/or inadequacy of an endometrial biopsy sample, the gain in efficiency is likely to be small.

Oxidation of unsaturated phospholipids in membrane bilayer mixtures is accompanied by membrane fluidity changes.

Steady-state and time-resolved fluorescence spectroscopy has been used to obtain information on oxidation processes and associated dynamical and structural changes in model membrane bilayers made from single unilamellar vesicles (SUV's) of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) mixed with increasing amounts of 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (SAPC). The highly unsaturated arachidonoyl chain containing four double bonds is prone to oxidation. Lipid oxidation was initiated chemically by a proper oxidant and could be followed on line via the fluorescence changes of an incorporated fluorescent lipophilic fatty acid: 4,4-difluoro-5-(4-phenyl-1,3-butadienyl)-4-bora-3a,4a-diaza-s-indacene-3-undecanoic acid (BP-C11). The oxidation rate increases with an increasing amount of SAPC. Size measurements of different SUV's incorporated with a trace amount of a phosphatidylcholine analogue of BP-C11 using fluorescence correlation spectroscopy have demonstrated that an increase of lipid unsaturation results in smaller sized SUV's and therefore to a larger curvature of the outer bilayer leaflet. This suggests that the lipid-lipid spacing has increased and that the unsaturated fatty acyl chains are better accessible for the oxidant. Oxidation results in some characteristic physical changes in membrane dynamics and structure, as indicated by the use of specific fluorescence probes. Fluorescence measurements of both dipyrenyl- and diphenylhexatriene-labelled PC introduced in non-oxidised and oxidised DOPC-SAPC membranes clearly show that the microfluidity (local fluidity at the very site of the probes) significantly decreases when the oxidised SAPC content increases in the lipid mixture. A similar effect is observed from the lateral diffusion experiments using monopyrenyl PC in the same membrane systems: the lateral diffusion is distinctly slower in oxidised membranes.

Static and dynamic studies of the potential-sensitive membrane probe RH421 in dimyristoylphosphatidylcholine vesicles.

The dynamics of the potential-sensitive styryl dye RH421 in dimyristoylphosphatidylcholine vesicles have been investigated above and below the main phase transition temperature using iodine-laser temperature-jump relaxation spectrophotometry and time-resolved fluorescence lifetime measurements. Equilibrium fluorescence titrations have shown that the affinity of the dye for the membrane is much higher in the liquid-crystalline state than in the gel state. The interaction can be described by either a partition or a binding model and a theory is presented providing a relation between these two approaches. In the liquid-crystalline state bound dye exhibits steady-state fluorescence relaxation processes in the submicrosecond and millisecond time range following a temperature jump. Time-resolved fluorescence measurements show a variation in the fluorescence lifetime across the emission spectrum, suggesting an excited-state process occurring on the subnanosecond time scale. These processes are most likely related to dye and/or lipid reorientation following the temperature jump or excitation pulse. Temperature-dependent changes in the fluorescence excitation spectrum of bound dye suggest that the dye exists in at least two different sites within the membrane.

A high-molecular-weight (greater than 8.10(5)) non-collagenous glycoprotein is synthesized by bovine cartilage in vitro.

A high-molecular-weight (greater than 8.10(5)) glycoprotein was detected in [3H]glucosamine-labeled bovine cartilage. Extraction with varying amounts of guanidinium chloride showed that the molecule was not tightly bound to other matrix substances. Enzyme digestions identified the molecule as a non-collagenous glycoprotein. This glycoprotein constituted 10-20% of the [3H]glucosamine-labeled macromolecular material that was released into culture medium on the first day after labeling. The 3H-labeled glycoprotein was purified by anion-exchange chromatography, CsCl gradient centrifugation and gel filtration. The purified glycoprotein appeared on an SDS-polyacrylamide gel as one slightly polydisperse band, which could not be reduced by beta-mercaptoethanol.

Fluorescence dynamics of staphylococcal nuclease in aqueous solution and reversed micelles.

The dynamical fluorescence properties of the sole tryptophan residue (Trp-140) in Staphylococcus aureus nuclease (EC 3.1.31.1) have been investigated in aqueous solution and reversed micelles composed of either sodium bis(2-ethylhexyl)sulfosuccinate (AOT) in isooctane or cetyltrimethylammonium chloride (CTAC) in isooctane/hexanol (12:1 by volume). The fluorescence decay of nuclease in the different environments can be described by a trimodal distribution of fluorescence lifetimes at approx. 0.5, 1.5 and 5.0 ns. The relative amplitudes depend on the environment. For pH 9.0 solutions the contribution of the two shortest lifetime components in the distribution is largest for AOT and smallest for CTAC reversed micelles. There is reasonable agreement between the average fluorescence lifetime and the fluorescence quantum efficiency confirming a significant fluorescence quenching in AOT reversed micelles. Fluorescence anisotropy decay revealed that the tryptophan environment in aqueous nuclease solutions is rigid on a nanosecond timescale. When nuclease was entrapped into reversed micelles the tryptophan gained some internal flexibility as judged from the distinct presence of a shorter correlation time. The longer correlation time reflected the rotational properties of the protein-micellar system. Modulation of the overall charge of nuclease (isoelectric point pH 9.6) by using buffer of pH 9.0 and pH 10.4, respectively, and of the size of empty micelles by selecting two values of the water to surfactant molar ratio, had only a minor effect on the rotational properties of nuclease in the positively charged reversed micelles. Encapsulation of nuclease in anionic reversed micelles resulted in the development of protein bound to aggregated structures which are immobilised on a nanosecond timescale. According to far UV circular dichroism results the secondary structure of nuclease only followed the already published pH-dependent changes. Encapsulation had no major effect on the overall secondary structure.

Human factor VIII: purification from commercial factor VIII concentrate, characterization, identification and radiolabeling.

Human factor VIII was purified from commercial factor VIII concentrate with a 12% yield. The specific coagulant activity of purified factor VIII was 8,000 units/mg. In the presence of SDS the purified factor VIII consisted of a variety of polypeptides on polyacrylamide gels, ranging between Mr 80,000 and Mr 208,000. In the absence of SDS the purified factor VIII showed an apparent molecular weight of 270,000 upon Sephadex G200 gel-filtration. The purified factor VIII could be activated by thrombin, which resulted in the disappearance of Mr 108,000-208,000 polypeptides in favor of an Mr 92,000 polypeptide. Treatment with factor Xa also activated factor VIII, whereas treatment with activated protein C resulted in the inactivation of coagulant activity. Coagulant-active 125I-factor VIII was prepared using a lactoperoxidase radioiodination procedure. This 125I-factor had the same characteristics as unlabeled factor VIII. All polypeptides could be precipitated with monoclonal antibodies directed against factor VIII. With 125I-factor VIII a pIapp of 5.7 was found in the presence of urea.

Convergence of circadian and sleep regulatory mechanisms on hypocretin-1.

Hypocretin is a potential regulator of sleep and wakefulness and its levels fluctuate with the day-night cycle with high levels during the animal's activity period. Whether the daily fluctuations are driven endogenously or by external light cycles is unknown. We investigated the circadian and homeostatic regulation of hypocretin in the absence of environmental light cycles. To this purpose we performed repetitive samplings of cerebrospinal fluid in rats through implanted microcannulas in the cisterna magna and determined hypocretin-1 levels by radioimmunoassay. These experiments were also performed in rats that received a lesion of the suprachiasmatic nucleus (SCN), a major pacemaker for circadian rhythms in mammals. The results showed sustained rhythmicity of hypocretin in constant dim red light in control animals. SCN-lesioned animals showed no circadian rhythms in hypocretin and mean hypocretin levels were remarkably low. The results indicate that the SCN is indispensable for rhythmicity in hypocretin and induces a daily increase in hypocretin levels during the animal's active phase. Additional sleep deprivation experiments were carried out to investigate homeostatic regulation of hypocretin. Hypocretin levels increased in response to sleep deprivation in both control and SCN-lesioned animals, demonstrating that sleep homeostatic control of hypocretin occurs independently from the SCN. Our data indicate that the circadian pacemaker of the SCN and sleep homeostatic mechanisms converge on one single sleep regulatory substance.

Detection of porcine reproductive respiratory syndrome virus by the polymerase chain reaction.

A polymerase chain reaction (PCR) is described for the detection of the porcine reproductive respiratory syndrome virus (PRRSV). Using the PCR we were able to detect about 30 infectious units of PRRSV resolved in tissue culture medium or sperm. For the detection of PRRSV in sperm the PCR is 10-times more sensitive than culturing in alveolar macrophages. For the detection of PRRSV, PCR provides a good alternative to cell culture methods.

A mouse model for testing the pathogenicity of equine herpes virus-1 strains.

A mouse model was developed for testing the pathogenicity of equine herpes virus-1 (EHV-1) strains. The model was validated with EHV-1 strains that are known to be of a low or high pathogenicity in horses. From all parameters tested, the safety index, which was calculated from the body weights of the mice after infection, proved to be the best predictive parameter. When this parameter was used, good and reliable correlations were found with the pathogenicity of the EHV-1 strains in horses. This method enabled the differentiation between the two experimental EHV-1 strains whose genetic backgrounds were supposedly equal.