Differential somatic cell count as an additional indicator for intramammary infections in dairy cows.

Mastitis, often caused by intramammary infection (IMI), is a significant problem in dairy farming globally. Somatic cell count (SCC) is widely used as a parameter for screening IMI in cows that are then treated or culled. We investigated the potential of a new parameter, differential SCC (DSCC), to detect IMI at cow level when SCC is already known. We achieved this using bacterial culture (BC) and PCR to detect 4 categories of pathogens (major, minor, other, and any) in 2 Danish dairy herds. Quarter milk samples were collected from monthly dairy herd improvement samplings over 1 yr and analyzed with BC, whereas cow-level dairy herd improvement samples were analyzed using PCR. Days in milk, parity, and IMI status had a significant effect on DSCC. Using DSCC in addition to SCC significantly improved the indication of IMI compared with using only SCC in the any pathogen category in both herds as well as the minor pathogens category in herd 2 when BC was used for detection. When PCR was used to detect IMI, the use of DSCC in addition to SCC was significant for the other pathogens category in herd 1 and the minor pathogens category in herd 2. Thus, our data revealed that DSCC can add significant information describing IMI status even when SCC is already known; however, this depends on the causative pathogen. Future studies may address how to use DSCC in practice as well as consider the availability of temporal data to potentially gain insight into the course of infection.

Transmission dynamics of Staphylococcus aureus within two Danish dairy cattle herds.

Staphylococcus aureus is a major pathogen causing intramammary infections (IMI) in dairy cattle herds worldwide. Simulation models can be used to investigate the epidemiologic and economic outcomes of different control strategies against IMI. The transmission rate parameter is one of the most influential parameters on the outcomes of these simulation models. Very few studies have estimated the transmission rate parameter and investigated the transmission dynamics of Staph. aureus IMI in dairy cattle herds. The objective of our study was therefore to analyze the transmission dynamics of Staph. aureus in 2 Danish dairy herds participating in a longitudinal study. The 2 herds had 180 and 360 milking cows, and animals were tested at quarter level once per month over a period of 1 yr. We estimated the quarter-level prevalence to be 34% for herd 1 and 2.57% for herd 2. The daily quarter-level transmission rate was estimated to be 0.0132 and 0.0077 cases/quarter-day for herds 1 and 2, respectively, and the median duration of infection was estimated to be 91 and 64 d for herds 1 and 2, respectively. We also estimated the reproductive ratio at 1.21 for herd 1 and 0.52 for herd 2. The results can provide valuable information for simulation models to aid decision-making in terms of the prevention and control of Staph. aureus IMI in dairy cattle herds.

Spatial patterns of antimicrobial resistance genes in a cross-sectional sample of pig farms with indoor non-organic production of finishers.

Antimicrobial resistance (AMR) in pig populations is a public health concern. There is a lack of information of spatial distributions of AMR genes in pig populations at large scales. The objective of the study was to describe the spatial pattern of AMR genes in faecal samples from pig farms and to test if the AMR genes were spatially randomly distributed with respect to the geographic distribution of the pig farm population at risk. Faecal samples from 687 Danish pig farms were collected in February and March 2015. DNA was extracted and the levels of seven AMR genes (ermB, ermF, sulI, sulII, tet(M), tet(O) and tet(W)) were quantified on a high-throughput real-time PCR array. Spatial differences for the levels of the AMR genes measured as relative quantities were evaluated by spatial cluster analysis and creating of risk maps using kriging analysis and kernel density estimation. Significant spatial clusters were identified for ermB, ermF, sulII and tet(W). The broad spatial trends in AMR resistance evident in the risk maps were in agreement with the results of the cluster analysis. However, they also showed that there were only small scale spatial differences in the gene levels. We conclude that the geographical location of a pig farm is not a major determinant of the presence or high levels of AMR genes assessed in this study.

A prospective observational longitudinal study of new-onset seizures and newly diagnosed epilepsy in dogs.

BACKGROUND: Seizures are common in dogs and can be caused by non-epileptic conditions or epilepsy. The clinical course of newly diagnosed epilepsy is sparsely documented. The objective of this study was to prospectively investigate causes for seizures (epileptic and non-epileptic) in a cohort of dogs with new-onset untreated seizures, and for those dogs with newly diagnosed epilepsy to investigate epilepsy type, seizure type and the course of disease over time, including the risk of seizure recurrence. Untreated client-owned dogs experiencing new-onset seizures were prospectively enrolled in a longitudinal observational study including clinical investigations and long-term monitoring at the Copenhagen University Hospital for Companion Animals. A baseline clinical assessment was followed by investigator/owner contact every eight weeks from inclusion to death or end of study. Inclusion of dogs was conducted from November 2010 to September 2012, and the study terminated in June 2014. RESULTS: One hundred and six dogs were included in the study. Seventy-nine dogs (74.5%) were diagnosed with epilepsy: 61 dogs (77.2%) with idiopathic epilepsy, 13 dogs (16.5%) with structural epilepsy and five dogs (6.3%) with suspected structural epilepsy. A non-epileptic cause for seizures was identified in 13 dogs and suspected in 10 dogs. Four dogs in which no cause for seizures was identified experienced only one seizure during the study. In dogs with idiopathic epilepsy 60% had their second epileptic seizure within three months of seizure onset. Twenty-six dogs with idiopathic epilepsy (43%) completed the study without receiving antiepileptic treatment. The natural course of idiopathic epilepsy (uninfluenced by drugs) was illustrated by highly individual and fluctuating seizure patterns, including long periods of remission. Cluster seizures motivated early treatment. In a few dogs with a high seizure frequency owners declined treatment against the investigators advice. CONCLUSIONS: Epilepsy is the most likely diagnosis in dogs presenting with new-onset seizures. The course of idiopathic epilepsy is highly individual and might not necessarily require long-term treatment. This must be considered when advising owners about what to expect with regard to treatment and prognosis.

Effect of presampling procedures on real-time PCR used for diagnosis of intramammary infections with Staphylococcus aureus in dairy cows at routine milk recordings.

Aseptic procedures for milk sample collection are considered crucial for bacterial culture to avoid misdiagnosis and subsequently unnecessary treatment or culling. The objective of this field study was to investigate the effect of presampling procedures on the PCR-positivity at cycle threshold value ≤ 37 of real-time PCR assay to detect Staphylococcus aureus from composite milk samples at routine milk recordings while accounting for known cow-level risk factors. A total of 1,199 dairy cows from 6 herds with conventional milking parlors were sampled and tested by PCR in 2011. Following the farmers' routine premilking preparations, 624 cows of the 1,199 cows were randomly selected for bacterial culture preceded by presampling procedures. These procedures were: cleaning of udder teats, removing the first streams of milk, and 70% alcohol teat disinfection. Data on parity, somatic cell counts, days in milk, daily milk yield, fat %, and protein % were extracted from the Danish Cattle Database, whereas energy-corrected milk was calculated based on the latter 3. The within-herd prevalence of intramammary infections with Staph. aureus was 31%, ranging from 16 to 48% in the 6 herds. Univariable analysis showed that the presampling procedures, somatic cell counts, energy-corrected milk, and days in milk were significantly associated with PCR-positivity, whereas parity was not significant. A multivariable model with herd as random effect showed that presampling procedures decreased the chance of being PCR-positive to 0.75 (95% CI; 0.58-0.97) compared with cows where the presampling procedures were not carried out. In conclusion, presampling procedures decreased the cow's chance of being PCR-positive to Staph. aureus. Presampling procedures appeared to improve the specificity of PCR for detection of Staph. aureus by reducing false positives through destruction of Staph. aureus bacteria colonizing or contaminating the teat skin, orifice, and canal. Random herd effects accounted for only 8.9%, indicating that the cluster effect due to herd management on the PCR positivity to Staph. aureus was virtually negligible.

Output-based assessment of herd-level freedom from infection in endemic situations: Application of a Bayesian Hidden Markov model.

Countries have implemented control programmes (CPs) for cattle diseases such as bovine viral diarrhoea virus (BVDV) that are tailored to each country-specific situation. Practical methods are needed to assess the output of these CPs in terms of the confidence of freedom from infection that is achieved. As part of the STOC free project, a Bayesian Hidden Markov model was developed, called STOC free model, to estimate the probability of infection at herd-level. In the current study, the STOC free model was applied to BVDV field data in four study regions, from CPs based on ear notch samples. The aim of this study was to estimate the probability of herd-level freedom from BVDV in regions that are not (yet) free. We additionally evaluated the sensitivity of the parameter estimates and predicted probabilities of freedom to the prior distributions for the different model parameters. First, default priors were used in the model to enable comparison of model outputs between study regions. Thereafter, country-specific priors based on expert opinion or historical data were used in the model, to study the influence of the priors on the results and to obtain country-specific estimates. The STOC free model calculates a posterior value for the model parameters (e.g. herd-level test sensitivity and specificity, probability of introduction of infection) and a predicted probability of infection. The probability of freedom from infection was computed as one minus the probability of infection. For dairy herds that were considered free from infection within their own CP, the predicted probabilities of freedom were very high for all study regions ranging from 0.98 to 1.00, regardless of the use of default or country-specific priors. The priors did have more influence on two of the model parameters, herd-level sensitivity and the probability of remaining infected, due to the low prevalence and incidence of BVDV in the study regions. The advantage of STOC free model compared to scenario tree modelling, the reference method, is that actual data from the CP can be used and estimates are easily updated when new data becomes available.

Effect of management practices on paratuberculosis prevalence in Danish dairy herds.

A voluntary risk-based control program on paratuberculosis in dairy cattle was initiated in Denmark in 2006. Cows were categorized as high-risk (antibody-positive at least once within the last 3 tests) or low-risk animals based on the results of 3 to 4 annual milk ELISA detecting Mycobacterium avium ssp. paratuberculosis (MAP)-specific antibodies. High-risk animals require management practices aimed at decreasing calf exposure to MAP-contaminated colostrum and milk, and feces originating from these cows. Moreover, repeated test-positive cows are recommended for slaughter before next calving. The objective was to assess the effect of different management practices on the prevalence of MAP-specific antibodies. A questionnaire on management practices was distributed to 1,261 participating herds in December 2008. A total of 1,092 (87%) herd managers returned the questionnaire. Repeated prevalence data from 1,081 herds were available for a period up to 4.25 yr after the first test round. The changes in the prevalence of MAP-specific antibodies from the start of interventions were assessed using a hierarchical logistic model, where different management practices were assessed: a) culling of repeated test-positive cows, b) separation of high-risk from low-risk cows in calving areas, c) cleaning of calving areas after high-risk cows calved, d) removal of calves born to high-risk dams within 2h after calving, e) use of colostrum for feeding of heifer calves from low-risk cows only, f) use of waste milk for feeding of heifer calves from low-risk cows only, g) herd size, and h) proportion of purchased animals. Multivariable analyses suggested that only the proportion of purchased animals (>15% purchased animals as well as 0 to 15% purchased animals compared with no purchased animals in the herd), culling of repeated test-positive animals, and use of waste milk from specific cow groups influenced the decrease in prevalence of MAP-specific antibodies. The control program has been running for just 4.25 yr, and it is assumed that the full effect of the risk-based management practices will only be observed after 4 to 8 yr. Therefore, lack of association between some practices and decrease in prevalence may be a reflection of a short study period. Furthermore, decreases in the prevalence of MAP-specific antibodies may not reflect discontinued transmission of MAP in all age groups.

Latent class evaluation of a milk test, a urine test, and the fat-to-protein percentage ratio in milk to diagnose ketosis in dairy cows.

In this study, 3 commonly used tests to diagnose ketosis were evaluated with a latent class model to avoid the assumption of an available perfect test. The 3 tests were the KetoLac BHB (Sanwa Kagaku Kenkyusho Co. Ltd., Nagoya, Japan) test strip that tests milk for ß-hydroxybutyrate, the KetoStix (Bayer Diagnostics Europe Ltd., Dublin, Ireland) test strip that tests urine for acetoacetate, and the fat-to-protein percentage ratio (FPR) in milk. A total of 8,902 cows were included in the analysis. The cows were considered to be a random sample from the population of Danish dairy cattle under intensive management, thus representing a natural spectrum of ketosis as a disease. All cows had a recorded FPR between 7 and 21 d postpartum. The KetoLac BHB recordings were available from 2,257 cows and 6,645 cows had a KetoStix recording. The recordings were analyzed with a modified Hui-Walter model, in a Bayesian framework. The specificity of the KetoLac BHB test and the KetoStix test were both high [0.99 (0.97-0.99)], whereas the specificity of FPR was somewhat lower [0.79 (0.77-0.81)]. The best sensitivity was for the KetoStix test [0.78 (0.55-0.98)], followed by the FPR [0.63 (0.58-0.71)] and KetoLac BHB test [0.58 (0.35-0.93)].

Estimating test characteristics of somatic cell count to detect Staphylococcus aureus-infected dairy goats using latent class analysis.

The aim of this study was to estimate test properties of composite somatic cell count (cSCC) to detect subclinically Staphylococcus aureus-infected dairy goats. Staphylococcus aureus is the most prevalent major pathogen in goats, and responsible for the majority of clinical mastitis cases. Therefore, a diagnostic tool that detects subclinical Staph. aureus infections may be useful in decreasing the number of clinical cases. We collected samples from 384 animals in 4 herds for bacteriological culture and cSCC on 3 occasions in lactation: once in early lactation, once around peak lactation, and once in late lactation. Latent class models were used to estimate test properties of cSCC and bacteriological culture in the absence of a gold standard reference test under the assumption that both tests detect Staph. aureus intramammary infection. Estimates for test properties of cSCC in early lactation at a cut-off value of 1,500 × 10(3) cells/mL were 0.90 for sensitivity and 0.95 for specificity, making cSCC a useful screening tool for detection of Staph. aureus. An effect of lactation stage was observed, causing an increased sensitivity and decreased specificity in late lactation. The sensitivity of bacteriological culture was estimated to be very low in the latent class models and the models suggested that the true prevalence of Staph. aureus in dairy goat herds is much higher than what is commonly reported based on bacteriological culture. This implies that intramammary infection by Staph. aureus may be an underestimated problem in dairy goat herds, and that cSCC can be used to diagnose infected animals.

Stochastic models to simulate paratuberculosis in dairy herds.

Stochastic simulation models are widely accepted as a means of assessing the impact of changes in daily management and the control of different diseases, such as paratuberculosis, in dairy herds. This paper summarises and discusses the assumptions of four stochastic simulation models and their use in the design of certification, surveillance, and control strategies for paratuberculosis in cattle herds. A detailed comparison is made between the Dutch JohneSSim and the Danish PTB-Simherd, using the same context of a set of control strategies in a typical Dutch/Danish herd. The conclusion is that while the models are somewhat different in their underlying principles and do put slightly different values on the different strategies, their overall findings are similar. Therefore, simulation models may be useful in planning paratuberculosis strategies in dairy herds, although as with all models caution is warranted when interpreting and generalising the results.

Value of flow cytometry for MRD-based relapse prediction in B-cell precursor ALL in a multicenter setting.

PCR of TCR/Ig gene rearrangements is considered the method of choice for minimal residual disease (MRD) quantification in BCP-ALL, but flow cytometry analysis of leukemia-associated immunophenotypes (FCM-MRD) is faster and biologically more informative. FCM-MRD performed in 18 laboratories across seven countries was used for risk stratification of 1487 patients with BCP-ALL enrolled in the NOPHO ALL2008 protocol. When no informative FCM-marker was available, risk stratification was based on real-time quantitative PCR. An informative FCM-marker was found in 96.2% and only two patients (0.14%) had non-informative FCM and non-informative PCR-markers. The overall 5-year event-free survival was 86.1% with a cumulative incidence of relapse (CIR5y) of 9.5%. FCM-MRD levels on days 15 (HzR 4.0, p < 0.0001), 29 (HzR 2.7, p < 0.0001), and 79 (HzR 3.5, p < 0.0001) associated with hazard of relapse adjusted for age, cytogenetics, and WBC. The early (day 15) response associated with CIR5y adjusted for day 29 FCM-MRD, with higher levels in adults (median 2.4 × 10-2 versus 5.2 × 10-3, p < 0.0001). Undetectable FCM- and/or PCR-MRD on day 29 identified patients with a very good outcome (CIR5y = 3.2%). For patients who did not undergo transplantation, day 79 FCM-MRD > 10-4 associated with a CIR5y = 22.1%. In conclusion, FCM-MRD performed in a multicenter setting is a clinically useful method for MRD-based treatment stratification in BCP-ALL.

Association between bulk-tank milk Salmonella antibody level and high calf mortality in Danish dairy herds.

Salmonella enterica ssp. enterica Dublin is the most common Salmonella serotype found in the dairy sector in Denmark. Salmonella antibody level in bulk-tank milk (BTM), indicative of Salmonella Dublin infection in the herd, has been recorded regularly in all Danish dairy herds through a surveillance program since 2002. The objective of this study was to investigate whether high BTM Salmonella antibody level was associated with high calf mortality at herd level. Other risk factors for high calf mortality were also investigated: breed, production type (organic vs. conventional), number of animals purchased, herd size, and number of neighbor herds within a 4.9-km radius. Data from the Danish Cattle Database including the Salmonella surveillance program from September 2007 through August 2008 were used. Dairy herds with more than 20 cows were included (n=4,337). Because of a highly right-skewed distribution of calf mortality with many zero values, calf mortality had to be dichotomized for the analysis. Therefore, in this study, high calf mortality was defined as calf mortality of more than 6.5% for calves aged 1 to 180 d. A logistic analysis was performed to identify risk factors associated with the probability of a herd having high calf mortality. The following factors were significantly associated with high calf mortality: high BTM Salmonella antibody level, odds ratio (OR)=2.0 (95% confidence interval=1.6-2.4), organic production OR=1.4 (95% confidence interval=1.1-1.7) for organic versus conventional production, and breed. Purchase of 8 or more animals increased the OR of high calf mortality more than purchase of 1 to 7 animals, which again had a higher OR compared with purchase of 0 animals. Because only 14.3% of the population consisted of herds with high BTM Salmonella status, the estimated proportion of herds with high calf mortality could only be reduced from 38.7 to 36.5% by eradicating Salmonella from the Danish cattle population (i.e., a population attributable risk of 2.2%). This showed that although there is a strong association between BTM Salmonella status and calf mortality, the problem with high calf mortality will not be solved by eradicating Salmonella. All other things equal, a population with more Salmonella-infected herds would gain a larger reduction in calf mortality from a Salmonella control campaign. Nevertheless, individual herds with a high within-herd prevalence of Salmonella are likely to benefit, both economically and regarding animal welfare, from controlling pathogenic Salmonella types in cattle.

Correction: Minimal residual disease quantification by flow cytometry provides reliable risk stratification in T-cell acute lymphoblastic leukemia.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

T-cell acute lymphoblastic leukemia in patients 1-45 years treated with the pediatric NOPHO ALL2008 protocol.

The NOPHO ALL2008 is a population-based study using an unmodified pediatric protocol in patients 1-45 years of age with acute lymphoblastic leukemia. Patients with T-ALL were given a traditional pediatric scheme if fast responding (minimal residual disease (MRD) < 0.1% day 29), or intensive block-based chemotherapy if slow responding (MRD > 0.1% day 29). Both treatment arms included pediatric doses of high-dose methotrexate and asparaginase. If MRD ≥ 5% on day 29 or ≥0.1% after consolidation, patients were assigned to allogeneic hematopoietic stem cell transplantation. The 5-year overall survival of the 278 T-ALL patients was 0.75 (95% CI 0.69-0.81), being 0.82 (0.74-0.88) for patients 1.0-9.9 years, 0.76 (0.66-0.86) for those 10.0-17.9 years, and 0.65 (0.55-0.75) for the older patients. The risk of death in first remission was significantly higher in adults (12%) compared with the 1-9 years group (4%). The MRD responses in the three age groups were similar, and only a nonsignificant increase in relapse risk was found in adults. In conclusion, an unmodified pediatric protocol in patients 1-45 years is effective in all age groups. The traditional pediatric treatment schedule was safe for all patients, but the intensive block therapy led to a high toxic death rate in adults.

Mechanical sensory threshold in Cavalier King Charles spaniels with syringomyelia-associated scratching and control dogs.

It is assumed that Cavalier King Charles spaniels with Chiari-like malformation and syringomyelia experience central neuropathic pain. An association between spinal cord parenchymal lesions and specific clinical signs (e.g. spontaneous and evoked scratching, withdrawal, and paroxysmal pain manifestations with vocalisation) has been suggested. This led to the hypothesis that mechanical sensory threshold is altered in clinical cases. The aim of this study was to quantify the cervical mechanical sensory threshold using Semmes-Weinstein monofilaments in nine Cavalier King Charles spaniels with Chiari-like malformation and assumed syringomyelia-associated central neuropathic pain compared to eight control dogs. Clinical and neurological examination including magnetic resonance imaging was undertaken. Mean mechanical sensory threshold was not significantly different between case and control dogs (t-test on log10 transformed data; P=0.25). Substantial variation within and between dogs was seen, with individual thresholds ranging from 0.04 to 26g in case dogs and from 0.02 to 10g in control dogs. Based on these results, it is unlikely that Cavalier King Charles spaniels with Chiari-like malformation and syringomyelia have increased mechanical sensation characterised by lower mechanical sensory threshold when quantified with Semmes-Weinstein monofilaments. Whether clinical cases experience central neuropathic pain remains unknown. The assessment of sensory function in dogs with assumed central neuropathic pain should be multimodal and include not only mechanical but also tactile and thermal threshold quantification. The use of threshold quantification in a clinical setting is challenging due to an insufficient signal relative to the biological background noise within and between dogs.

Composite or aseptic quarter milk samples: Sensitivity and specificity of PCR and bacterial culture of Staphylococcus aureus based on Bayesian latent class evaluation.

Bacterial culture (BC) and quantitative polymerase chain reaction (qPCR) are widely used as diagnostic tests for pathogens causing intra-mammary infections (IMI) and it is therefore important to evaluate their performance to optimize pathogen detection. However, with no reference standard, their sensitivity (Se) and specificity (Sp) are unknown. Furthermore, the Se and Sp can differ between quarter samples and composite samples, both of which can be used to test for IMI. Latent class analysis (LCA) offers a method for estimating the Se and Sp of two tests on the same population, but to our knowledge this has not yet been applied to qPCR and BC for the detection of Staph. aureus in both composite and aseptically collected quarter milk samples. This allows for a performance evaluation of not only the two diagnostic methods, but also the two different samples -both cow and quarter level. In this study, we used Bayesian LCA on a dataset from one sampling day at a Danish dairy herd to estimate the Se and Sp for detecting Staph. aureus at cow and quarter level. We used Ct cut-offs of 32 and 37 cycles for the qPCR. When using a cut-off of 37 cycles, the estimated Se and Sp for BC were 62.2% and 90.5% at cow level, and 90.7% and 93.5% at quarter level, respectively. Similarly, the estimated Se and Sp for qPCR at cow level were 82.7% and 84.1%, respectively, and 80.0% and 96.8% at quarter level. Sp was therefore higher for both BC and qPCR at quarter level. Se was also highest for BC at quarter level, but the opposite was true for qPCR. The same pattern was found using a Ct cut-off at 32 cycles, but with different estimates. The results show that qPCR with a Ct cut-off at 37 had a higher Se than BC for composite DHI samples and it is therefore more suitable as a routine screening test for Staph. aureus. However, BC on quarter samples gave the highest Se and Sp and should therefore be used for confirmatory testing.

Minimal residual disease quantification by flow cytometry provides reliable risk stratification in T-cell acute lymphoblastic leukemia.

Minimal residual disease (MRD) measured by PCR of clonal IgH/TCR rearrangements predicts relapse in T-cell acute lymphoblastic leukemia (T-ALL) and serves as risk stratification tool. Since 10% of patients have no suitable PCR-marker, we evaluated flowcytometry (FCM)-based MRD for risk stratification. We included 274 T-ALL patients treated in the NOPHO-ALL2008 protocol. MRD was measured by six-color FCM and real-time quantitative PCR. Day 29 PCR-MRD (cut-off 10-3) was used for risk stratification. At diagnosis, 93% had an FCM-marker for MRD monitoring, 84% a PCR-marker, and 99.3% (272/274) had a marker when combining the two. Adjusted for age and WBC, the hazard ratio for relapse was 3.55 (95% CI 1.4-9.0, p = 0.008) for day 29 FCM-MRD ≥ 10-3 and 5.6 (95% CI 2.0-16, p = 0.001) for PCR-MRD ≥ 10-3 compared with MRD < 10-3. Patients stratified to intermediate-risk therapy on day 29 with MRD 10-4-<10-3 had a 5-year event-free survival similar to intermediate-risk patients with MRD < 10-4 or undetectable, regardless of method for monitoring. Patients with day 15 FCM-MRD < 10-4 had a cumulative incidence of relapse of 2.3% (95% CI 0-6.8, n = 59). Thus, FCM-MRD allows early identification of patients eligible for reduced intensity therapy, but this needs further studies. In conclusion, FCM-MRD provides reliable risk prediction for T-ALL and can be used for stratification when no PCR-marker is available.

The distribution of the pathogenic nematode Nematodirus battus in lambs is zero-inflated.

Understanding the frequency distribution of parasites and parasite stages among hosts is essential for efficient experimental design and statistical analysis, and is also required for the development of sustainable methods of controlling infection. Nematodirus battus is one of the most important organisms that infect sheep but the distribution of parasites among hosts is unknown. An initial analysis indicated a high frequency of animals without N. battus and with zero egg counts, suggesting the possibility of a zero-inflated distribution. We developed a Bayesian analysis using Markov chain Monte Carlo methods to estimate the parameters of the zero-inflated negative binomial distribution. The analysis of 3000 simulated data sets indicated that this method out-performed the maximum likelihood procedure. Application of this technique to faecal egg counts from lambs in a commercial upland flock indicated that N. battus counts were indeed zero-inflated. Estimating the extent of zero-inflation is important for effective statistical analysis and for the accurate identification of genetically resistant animals.

Ante mortem diagnosis of paratuberculosis: a review of accuracies of ELISA, interferon-gamma assay and faecal culture techniques.

Infections with Mycobacterium avium subsp. paratuberculosis (MAP) can be latent for years without affecting the animal, but the animal may become infectious or clinical at some point. Diagnosis of paratuberculosis can be a challenge primarily in latent stages of the infection, and different diagnosis interpretations are usually required by the variety of decision makers. The objective of this paper was to provide a critical review of reported accuracies of ELISA tests, interferon-gamma assays (IFN-gamma) and faecal culture (FC) techniques used for diagnosis of three defined target conditions: MAP infected, MAP infectious and MAP affected animals. For each animal species, target condition and diagnostic test-type, sensitivities (Se) and specificities (Sp) were summarised based on a systematic, critical review of information in literature databases. The diagnostic test information often varied substantially for tests of the same type and make, particularly ELISA, which was the most frequently reported test-type. Comparison of the various tests accuracies was generally not possible, but stratification of test-evaluations by target condition improved the interpretation of the test accuracies. Infectious and affected animals can often be detected, but Se for infected animals is generally low. A main conclusion of the review was that the quality of design, implementation and reporting of evaluations of tests for paratuberculosis is generally poor. Particularly, there is a need for better correspondence between the study population and target population, i.e. the subjects chosen for test evaluation should reflect the distribution of animals in the population where the test is intended to be used.

Evaluation of test-strategies for estimating probability of low prevalence of paratuberculosis in Danish dairy herds.

Paratuberculosis is a chronic infection affecting cattle and other ruminants. In the dairy industry, losses due to paratuberculosis can be substantial in infected herds and several countries have implemented national programmes based on herd-classification to manage the disease. The aim of this study was to develop a method to estimate the probability of low within-herd prevalence of paratuberculosis for Danish dairy herds. A stochastic simulation model was developed using the R programming environment. Features of this model included: use of age-specific estimates of test-sensitivity and specificity; use of a distribution of observed values (rather than a fixed, low value) for design prevalence; and estimates of the probability of low prevalence (PrLow) based on a specific number of test-positive animals, rather than for a result less than or equal to a specified cut-point number of reactors. Using this model, five herd-testing strategies were evaluated: (1) milk-ELISA on all lactating cows; (2) milk-ELISA on lactating cows4 years old; (4) faecal culture on all lactating cows; and (5) milk-ELISA plus faecal culture in series on all lactating cows. The five testing strategies were evaluated using observed milk-ELISA results from 19 Danish dairy herds as well as for simulated results from the same herds assuming that they were uninfected. Whole-herd milk-ELISA was the preferred strategy, and considered the most cost-effective strategy of the five alternatives. The five strategies were all efficient in detecting infection, i.e. estimating a low PrLow in infected herds, however, PrLow estimates for milk-ELISA on age-cohorts were too low in simulated uninfected herds and the strategies involving faecal culture were too expensive to be of practical interest. For simulated uninfected herds, whole-herd milk-ELISA resulted in median PrLow values>0.9 for most herds, depending on herd size and age-structure. None of the strategies provided enough power to establish a high PrLow in smaller herds, or herds with a younger age-structure. Despite this, it appears as if the method is a useful approach for herd-classification for most herds in the Danish dairy industry.