Corrigendum: Control and Eradication Programs for Six Cattle Diseases in the Netherlands.

[This corrects the article DOI: 10.3389/fvets.2021.670419.].

Control and Eradication Programs for Six Cattle Diseases in the Netherlands.

Within the European Union, infectious cattle diseases are categorized in the Animal Health Law. No strict EU regulations exist for control, evidence of disease freedom, and surveillance of diseases listed other than categories A and B. Consequently, EU member states follow their own varying strategies for disease control. The aim of this study was to provide an overview of the control and eradication programs (CPs) for six cattle diseases in the Netherlands between 2009 and 2019 and to highlight characteristics specific to the Dutch situation. All of these diseases were listed as C,D or E in the New Animal Health Law. In the Netherlands, CPs are in place for six endemic cattle diseases: bovine viral diarrhea, infectious bovine rhinotracheitis, salmonellosis, paratuberculosis, leptospirosis, and neosporosis. These CPs have been tailored to the specific situation in the Netherlands: a country with a high cattle density, a high rate of animal movements, a strong dependence on export of dairy products, and a high-quality data-infrastructure. The latter specifically applies to the dairy sector, which is the leading cattle sector in the Netherlands. When a herd enters a CP, generally the within-herd prevalence of infection is estimated in an initial assessment. The outcome creates awareness of the infection status of a herd and also provides an indication of the costs and time to achieve the preferred herd status. Subsequently, the herd enrolls in the control phase of the CP to, if present, eliminate the infection from a herd and a surveillance phase to substantiate the free or low prevalence status over time. The high-quality data infrastructure that results in complete and centrally registered census data on cattle movements provides the opportunity to design CPs while minimizing administrative efforts for the farmer. In the CPs, mostly routinely collected samples are used for surveillance. Where possible, requests for proof of the herd status are sent automatically. Automated detection of risk factors for introduction of new animals originating from a herd without the preferred herd status i.e., free or unsuspected, is in place using centrally registered data. The presented overview may inspire countries that want to develop cost-effective CPs for endemic diseases that are not (yet) regulated at EU level.

Self-Assembly of a Trithioorthoformate-Capped Cyclophane and Its Endohedral Inclusion of a Methine Group.

An unusual trithioorthoformate-capped cyclophane cage was assembled via antimony-activated iodine oxidation of thiols as confirmed by 1 H-NMR spectroscopy and X-ray crystallography. The disulfide bridges can undergo desulfurization with hexamethylphosphorous triamide (HMPT) at ambient temperature to capture a trithioether cyclophane cage capped by the trithioorthoformate. In both cages a methine proton points directly into the small cavity. This unexpected structure is hypothesized to have formed as a result of haloform insertion during oxidation.

Spatial and risk factor analysis of bovine viral diarrhoea (BVD) virus after the first-year compulsory phase of BVD eradication programme in Northern Ireland.

Bovine viral diarrhoea virus (BVDV) causes bovine viral diarrhoea (BVD), which is a contagious pathogen that can have a significant economic impact on cattle industries. In Northern Ireland (NI), the compulsory phase of a BVD eradication programme was implemented in 2016. The aim of this retrospective population based study was to utilize herd-level data after the first year of the compulsory phase (March 2016-March 2017) to determine the spatial distribution and variation of BVDV, to identify clusters of infection, and to quantify some risk factors associated with BVD in NI. Global spatial clustering (autocorrelation) and local spatial hot-spot analyses were used to specify the clustering areas (hot- and cold-spot). A suite of multivariable logistic analyses was performed to estimate the associations of spatial and non-spatial factors (relating to herd characteristics) with the risk of being a BVDV positive herd. Final models were compared by evaluating the model fit and the ability to account for spatial autocorrelation in the study area. There were 17,186 herds included in the analysis. The herd-level prevalence of BVDV was 11.31%. Significant spatial clustering of BVDV positive herds was presented in the central region of NI. A mixed effects logistic model, with a spatial random effect term, was considered the best model. The final model showed that a positive BVDV status during the voluntary phase prior to the compulsory phase started (OR = 2.25; CI 95% = 1.85-2.73), larger herd size (OR = 6.19; CI 95% = 5.22-7.34 for herd size > 100 animals) and a larger number of positive nearest neighbours within 4 km radius (OR = 1.24; CI 95% = 1.05-1.47 for 8-9 neighbours and OR = 1.41; CI 95% = 1.20-1.65 for 10-12 neighbours) were significantly related to the risk of a herd being tested positive for BVDV. The clear spatial pattern from the local spatial clustering analyses could be used for targeted surveillance and control measures by focusing on the central region of NI.

Diagnostic potential of the peptide-mediated magnetic separation (PMS)-phage assay and PMS-culture to detect Mycobacterium avium subsp. paratuberculosis in bovine milk samples.

Controlling the spread of Johne's disease, caused by Mycobacterium avium subsp. paratuberculosis (MAP), in domestic livestock is challenging. Current diagnostic methods lack sufficient sensitivity to detect subclinically infected animals, and thus, better diagnostic methods are needed. This study was carried out to investigate the diagnostic potential of two novel peptide-mediated magnetic separation (PMS)-based tests-a PMS-phage assay and PMS-culture-both of which have been developed and optimized to detect viable MAP cells in bovine milk. Individual milk samples (50 ml) were obtained from 105 "non-infected" and 40 "MAP-infected" animals (classified as such on the basis of prior faecal culture and serum-ELISA results) in three dairy herds and tested in parallel by the PMS-phage assay and PMS-culture. Diagnostic sensitivity (DSe) and specificity (DSp) of the PMS-phage and PMS-culture methods were determined relative to the MAP infection status of the animal contributing the milk sample. The PMS-based tests applied individually showed moderate DSe (PMS-culture 0.250 and PMS-phage assay 0.325) and high DSp (0.962 and 1.000, respectively). When results of the two PMS-based tests were combined, DSe increased substantially to 0.525, and the DSp was calculated to be 0.962. It was concluded that combined application of the PMS-phage assay and PMS-culture provided the most complete picture regarding the presence of viable MAP in bovine milk samples. A comprehensive validation of the PMS-based assays relative to currently used diagnostic methods (faecal culture and serum-ELISA) would be the next step in assessment of the diagnostic potential of these novel PMS-based methods.

Assessment of concurrent infection with bovine viral diarrhoea virus (BVDV) and Mycobacterium bovis: A herd-level risk factor analysis from Northern Ireland.

Bovine viral diarrhoea virus (BVDV) is a significant pathogen of cattle, leading to severe economic and animal-welfare impacts. Furthermore, the pathogen has been associated with impacting the progression or spread of other pathogens (e.g. Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB)). During this study we investigated (i) risk factors for BVDV at a herd-level and (ii) whether there was any association between BVDV and herd-level bTB risk. The data for this study were gathered from a voluntary BVDV control programme in Northern Ireland (2013-2015) based on the identification of virus positive animals through tissue tag testing of calves. We assigned a herd-level BVDV status to 2827 participating herds, where a herd was assumed "infected" if one or more animals tested positive for BVDV. Two model suites were developed. Firstly, we assessed risk factors for BVDV herd status using multivariable logit random-effects modelling, aggregating to the calendar year level (2013-2015; n=4828; model 1). Secondly, we aggregated data across the three years of the study to give an overall status for the whole study period (n=2827; logistic model 2). Risk factors included year, herd-type, herd size, number of births, inward trade moves, calf mortality, and region. Furthermore, the herd-level bovine tuberculosis status (based on the single intradermal comparative cervical tuberculin (SICCT) test outcomes, or confirmation at post-mortem), or the size of bTB breakdowns (number of SICCT test positive animals), of herds was also investigated to assess whether there was an association (co-infection) with herd BVDV status. The final models suggested that BVDV herd status was positively associated with increased levels of calf mortality, herd size, number of births, the number of BVDV tests undertaken and the number of animals introduced to the herd. There was a significant univariable positive association between BVDV status, and SICCT breakdown risk, breakdown size and confirmed bTB status in model 2. However, there was no evidence of significant associations between bTB status (using SICTT status, confirmed status or herd breakdown size) and BVDV status in final multivariable models when controlling for other significant confounders. These results provide information for action for the future control and eradication of BVDV in Northern Ireland, though these data provide little support for the hypothesised association between BVDV and bTB status at herd-level. Further animal-level analyses are necessary to investigate whether there is support for a BVD-bTB co-infection association, including the impact of co-infection on the severity of infection.

Evaluation of testing strategies to identify infected animals at a single round of testing within dairy herds known to be infected with Mycobacterium avium ssp. paratuberculosis.

As part of a broader control strategy within herds known to be infected with Mycobacterium avium ssp. paratuberculosis (MAP), individual animal testing is generally conducted to identify infected animals for action, usually culling. Opportunities are now available to quantitatively compare different testing strategies (combinations of tests) in known infected herds. This study evaluates the effectiveness, cost, and cost-effectiveness of different testing strategies to identify infected animals at a single round of testing within dairy herds known to be MAP infected. A model was developed, taking account of both within-herd infection dynamics and test performance, to simulate the use of different tests at a single round of testing in a known infected herd. Model inputs included the number of animals at different stages of infection, the sensitivity and specificity of each test, and the costs of testing and culling. Testing strategies included either milk or serum ELISA alone or with fecal culture in series. Model outputs included effectiveness (detection fraction, the proportion of truly infected animals in the herd that are successfully detected by the testing strategy), cost, and cost-effectiveness (testing cost per true positive detected, total cost per true positive detected). Several assumptions were made: MAP was introduced with a single animal and no management interventions were implemented to limit within-herd transmission of MAP before this test. In medium herds, between 7 and 26% of infected animals are detected at a single round of testing, the former using the milk ELISA and fecal culture in series 5 yr after MAP introduction and the latter using fecal culture alone 15 yr after MAP introduction. The combined costs of testing and culling at a single round of testing increases with time since introduction of MAP infection, with culling costs being much greater than testing costs. The cost-effectiveness of testing varied by testing strategy. It was also greater at 5 yr, compared with 10 or 15 yr, since MAP introduction, highlighting the importance of early detection. Future work is needed to evaluate these testing strategies in subsequent rounds of testing as well as accounting for different herd dynamics and different levels of herd biocontainment.

The effect of alternative testing strategies and bio-exclusion practices on Johne's disease risk in test-negative herds.

Herd classification is a key component of national Johne's disease (JD) control programs. Herds are categorized on the basis of test results, and separate sub-programs are followed for test-positive and test-negative herds. However, a test-negative herd result does not necessarily equate to JD freedom for reasons relating to disease pathogenesis and available diagnostic tests. Thus, in several countries, JD control programs define test-negative herds as having a "low risk" of infection below a specified prevalence. However, the approach is qualitative, and little quantitative work is available on herd-level estimates of probability of freedom in test-negative herds. This paper examines the effect over time of alternative testing strategies and bio-exclusion practices on JD risk in test-negative herds. A simulation model was developed in the programming language R. Key model inputs included sensitivity and specificity estimates for 3 individual animal diagnostic tests (serum ELISA, milk ELISA, and fecal culture), design prevalence, testing options, and testing costs. Key model outputs included the probability that infection will be detected if present at the design prevalence or greater (herd sensitivity; SeH), the probability that infection in the herd is either absent or at very low prevalence (i.e., less than the design prevalence; ProbF), the probability of an uninfected herd producing a false-positive result [P(False+)], and mean testing cost (HerdCost) for different testing strategies. The output ProbF can be updated periodically, incorporating data from additional herd testing and information on cattle purchases, and could form the basis for an output-based approach to herd classification. A high ProbF is very difficult to achieve, reflecting the low sensitivity of the evaluated tests. Moreover, ProbF is greatly affected by any risk of introduction of infection, decreasing in herds with poor bio-exclusion practices despite ongoing negative test results. The value of P(False+) was substantial when tests with imperfect specificity were used. Testing strategies can substantially influence testing costs but with little effect on test performance. This study illustrates an output-based approach to herd classification, with potential for national and field applications.

Co-Infection of cattle with Fasciola hepatica and Mycobacterium bovis- immunological consequences.

Fasciola hepatica, the liver fluke, is a common parasite of cattle in much of the world. Previously, we have shown that cattle infected with F. hepatica have altered responsiveness (delayed type hypersensitivity reaction and cytokine responses) to M. bovis BCG infection. We hypothesized that co-infection with F. hepatica would, likewise, alter the immune response of cattle to virulent M. bovis infection, with possible implications for disease diagnosis and disease progression. Our previous work with F. hepatica/M. bovis BCG-infected cattle demonstrated a reduction in interferon (IFN)-gamma responsiveness in co-infected animals. Similar findings are reported here with virulent M. bovis following aerosol infection. The epidemiological significance of these findings, also, require exploration, particularly in view of the considerable resources devoted to the diagnosis and eradication of bovine tuberculosis, and the high prevalence of F. hepatica infection in areas where eradication has proved difficult.

The relationship between IgA activity against 4th-stage larvae and density-dependent effects on the number of 4th-stage larvae of Teladorsagia circumcincta in naturally infected sheep.

The number of Teladorsagia circumcincta 4th-stage larvae in naturally infected lambs from a single farm varied among lambs and among different years. Within each year the distribution of 4th-stage larvae among lambs was similar to that expected from a negative binomial distribution. The ratio of 4th-stage larvae to adult T. circumcincta was low in two years with a low mean intensity of infection but high in two years with a higher mean intensity of infection. The negative binomial distribution is defined by the mean and by k, a parameter that measures dispersion; k was low when mean infection intensity was low but higher when mean infection intensity was high. As k is an inverse index of overdispersion this indicated that the distribution of 4th-stage larvae was more overdispersed at low levels of infection. In a combined analysis, the number of adult T. circumcincta and the plasma IgA activity against 4th-stage larvae were both associated with increased numbers of 4th-stage larvae. There was a statistical interaction between the number of adults and IgA activity that moderated their combined effect.

The genetic control of IgA activity against Teladorsagia circumcincta and its association with parasite resistance in naturally infected sheep.

Previous studies in deliberately infected sheep have shown an association between IgA activity against 4th-stage larvae of Teladorsagia circumcincta and parasite growth, development and fecundity. The purpose of this research was to determine if these results could be confirmed in naturally infected sheep and to explore the hypothesis that plasma IgA activity could help to identify resistant lambs with shorter adult nematodes. Plasma IgA activity was skewed with most animals having relatively low levels of IgA activity. Plasma IgA activity was repeatable and highly heritable. Animals with increased IgA activity had lower egg counts and shorter adult female T. circumcincta. Therefore, under conditions of natural parasite challenge, plasma IgA activity may help to identify lambs resistant to T. circumcincta.

The influence of protein supplementation on the immune response to Haemonchus contortus.

A major protective mechanism in lambs against the abomasal parasite Ostertagia (Teladorsagia) circumcincta appears to be the immunoglubulin (Ig)A-mediated suppression of worm growth and fecundity. The present study indicates that IgA may play a similar role in the control of another abomasal parasite Haemonchus contortus. Hampshire Down lambs were offered one of two diets: (i) a basal diet and (ii) a diet supplemented with additional protein. Lambs were then 'trickle' infected with H. contortus and killed 10 weeks after the start of infection. Those lambs on the supplemented diet had shorter adult worms and produced significantly more antiparasite IgA. There was a significant association between reduced female adult worm length and increased IgA against third-stage larvae. Most of the difference between the two groups in worm length could be accounted for by differences in IgA responses. Therefore, IgA may be the major mechanism controlling fecundity of H. contortus and the magnitude of the IgA response is influenced by the quality of the diet.

ROCK: a spreadsheet-based program for the generation and analysis of random oligonucleotide primers used in PCR.

Oligonucleotide primers used to amplify target DNA regions via PCR should meet certain design criteria to maximize the potential for efficient priming. The Random Oligonucleotide Construction Kit (ROCK), a spreadsheet-based program that runs under Microsoft Excel 97 or later version for Microsoft Windows, was developed to facilitate the design of efficient random oligonucleotide primers. Primer sequences are generated that meet user-defined criteria with regard to G + C content, size of a 3' GC clamp, maximum intramolecular/intermolecular complementation potential, and maximum intersequence similarity. The user can analyze the intramolecular/intermolecular complementation potential of program-generated primer sequences or of sequences entered manually. The latter may contain any of the standard nucleotide symbols, including ambiguous bases. Primer sequence length, GC%, individual base composition, molecular weight, approximate melting temperature, and mass/volume/concentration relationships can be determined for any sequence generated by ROCK or entered manually.

Comparative behavioural toxicity of domoic acid and kainic acid in neonatal rats.

Cumulative behavioural toxicity was measured in groups of male and female rat pups (n=6/sex) at different stages of postnatal development. Dose-response curves (DRCs) for toxicity produced by domoic acid (DOM) were generated using animals on postnatal days (PND) 0, 5, 14, and 22, using a behavioural rating scale. In a subsequent experiment, DRCs for toxicity generated by either DOM or kainic acid were produced in rats at PND 8 and 14 for comparison between the two toxins. DOM was found to be a very potent neurotoxin in newborn rats and the potency of DOM progressively decreased with increasing age (interpolated ED(50)=0.12, 0.15, 0.30, and 1.06 mg/kg at PND 0, 5, 14, and 22, respectively). In addition, the patterns of behavioural expression were found to differ with age. Comparisons between DOM and kainic acid revealed that DOM was approximately six-fold more potent than kainate at both PND 8 and PND 14 and that both toxins were approximately two-fold less potent in PND 14 rats, compared to PND 8. This implies that the mechanism(s) responsible for reduced potency is/are similar between the two compounds. Consistent with previous reports, however, there were both similarities and differences in the observed patterns of behavioural toxicity produced by the two toxins at both ages.

The influence of age on the variation among sheep in susceptibility to natural nematode infection.

A longitudinal study of faecal nematode egg counts was made in naturally infected Scottish Blackface sheep over two grazing seasons to 75 weeks of age. Although egg counts were lower in the second grazing season the variation among animals was greater. Egg counts were repeatable from 3 months of age. The repeatability of faecal egg counts within and between grazing seasons was about 0.3. Animals with lower than average egg counts in the first grazing season tended to have lower than average egg counts in the second grazing season. Therefore lambs with relatively low faecal egg counts after 3 months of age are likely to retain their advantage in the following year.

How lambs control infection with Ostertagia circumcincta.

Two of the most important questions for immunologists studying nematode infections are what effect does the host response have on the parasite and which components of the host response are responsible for these effects. The number of nematodes and the mean length of adult female Ostertagia circumcincta was measured in over 500 6-7 month old lambs of the Scottish Blackface breed. Quantitative genetic analyses indicated that there was substantial genetic variation among lambs in faecal egg counts and in worm length but the analyses provided no evidence for genetic variation in worm numbers. Separate analyses have shown a strong relationship between decreased worm length and decreased worm fecundity. Therefore, genetic resistance in lambs appears to be due to control of worm growth and not to control of worm numbers. The only immune response that is consistently associated with reduced worm length is the IgA response to fourth-stage larvae. The association is remarkably strong. After allowing for the influence of worm number on worm length (density-dependence) essentially all of the variation among deliberately infected lambs in worm length can be accounted for, in a statistical sense, by variation in the strength and specificity of the local IgA response. Therefore, the IgA mediated suppression of worm growth and fecundity appears to be the major mechanism of resistance to O. circumcincta in lambs.

The relationship between the number and size of nematodes in the abomasum and the concentration of pepsinogen in ovine plasma.

Six-month-old lambs that had been naturally infected with predominantly Teladorsagia (Ostertagia) circumcincta were tested for plasma pepsinogen concentrations because pepsinogen concentrations may reflect the extent of damage to the abomasum. The distribution of pepsinogen concentrations among these lambs was positively skewed with most individuals having relatively low concentrations. Pepsinogen concentrations were more strongly associated with variation in the mean length of the adult female worms than with variation in the number of nematodes present. Previous trials have suggested that genetic variation in the growth of lambs is strongly influenced by genetic variation in worm length. Together these results imply that variation among lambs in the pathogenic effects of T. circumcincta depends upon the mean size of the worms as well as the number of worms present.

The recognition of molecules from fourth-stage larvae of Ostertagia circumcincta by IgA from infected sheep.

The recognition of antigens from fourth-stage Ostertagia circumcincta by plasma IgA was studied in a group of 30, ten-month-old Scottish Blackface sheep which had been naturally, then deliberately infected. Western blotting revealed 49 bands that were recognized by antibody from one or more of the 30 sheep. There was a heterogeneous pattern of antigen recognition amongst the sheep. No individual recognized all the 49 bands and only one of the 49 bands was recognized by all sheep. Two antigens with approximate molecular weights 87,000 Da and 129,000 Da were significantly associated with a reduction in mean adult worm lengths. The observed variation in recognition of these two antigens on fourth-stage larval preparations accounted, in a statistical sense, for nearly 40% of the total variation in worm length. In addition to the variation in antibody mediated recognition of these two parasite molecules, three other components have been implicated in regulating worm length. They are a 37,000 Da band from adult worms, the amount of fourth-stage larval specific IgA in the abomasal mucosa and the density-dependent influence of adult worm burden. Together, these components and their interactions accounted for over 90% of the observed variation in worm length. These results indicate that the parasite-specific IgA response, or something extremely closely associated with it, is the major immunological mechanism controlling worm length.