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Farmers play a key role in conserving native livestock breeds, but without economic support, farms with native breeds may not be viable. We hypothesized that terminal crossbreeding can improve herd economy and decrease the economic support needed from society. Three scenarios were simulated using SimHerd Crossbred: a herd of purebred Swedish Polled Cattle, a herd of purebred Swedish Red, and a herd of 75% Swedish Polled Cattle and 25% F1 crossbreds. The results showed annual contribution margin per cow in the herd can be increased by €181 by crossbreeding compared with pure-breeding with the native breed, giving a 13.6% growth in contribution margin. However, the needed cost in subsidies paid by the government will remain unchanged if the population size of the native breed is to be maintained. Combining a crossbreeding strategy with the marketing of niche products may facilitate the conservation of native cattle.

Objectives This study was conducted to examine the molecular mechanisms responsible for the evolution of TEM-type extended-spectrum β-lactamases (ESBLs) following selective pressure from four third-generation cephalosporins, namely ceftazidime, cefotaxime, ceftriaxone and ceftibuten. In addition, selective enrichment for ESBL detection in environmental samples was investigated. Methods Using experimental evolution, resistant variants were isolated and mutations in TEM-1 were examined by DNA sequencing. Resistance levels and the development of cross-resistance were determined for ESBL-producing isolates by Etest and disk diffusion assay. Selective plating with or without prior growth in selective broth was used to examine the approach of selective enrichment for ESBL detection. Results The third-generation cephalosporins ceftazidime, cefotaxime and ceftriaxone selected for ESBLs, whereas ceftibuten did not. All ESBL variants additionally remained susceptible to ceftibuten. DNA sequencing of the TEM-1 coding sequence of mutants revealed mutations not previously isolated through selection. This indicates that the potential for ESBL evolution is much broader than can be inferred from sequence analysis of clinical samples alone. The results also indicate that selective enrichment for enhanced detection of ESBL-producers may give unreliable results owing to the selection of spontaneous mutations in narrow-spectrum β-lactamases resulting in TEM-type ESBL-producers. Conclusion These results help explain the molecular changes responsible for evolution of TEM-type ESBLs and meanwhile question the appropriate use of selective enrichment for detection of ESBLs in environmental samples.

Crossbreeding in dairy cattle has recently become of increased interest. However, farmers in Scandinavian countries are reluctant to implement crossbreeding in their herds, and one reason is the common opinion that only herds at a poor level of management can benefit from crossbreeding. The Danish Cattle Database (SEGES, Aarhus, Denmark) provided data on 14 traits regarding milk yield, udder health, fertility traits, stillbirth, and survival. The data were collected from 103,307 pure Holstein cows and 14,832 F1 crosses (Holstein dam and Nordic Red sire). The cows were born between 2008 and 2014 and originated from 424 herds that contributed data from at least 5 purebreds and 5 crossbreds across the years. We split the animals into 3 production levels: high, average, and low according to the herd's average production (kg) of 305-d fat plus protein in the given birth year of the cow. We estimated least squares means of breed group (purebred and crossbred) performance within each production level. Crossbred performance in 305-d fat yield in first-parity cows was greater than that of Holstein across all herd production levels; the gain was greater in high- (9 kg more than Holstein) and average-producing herds (7 kg more than Holstein) than in low-producing herds (3 kg more than Holstein). Regardless of production level or parity, crossbreds did not outperform Holstein in terms of 305-d protein yield (0 to 8 kg less). Crossbreds had relatively better udder health than Holstein in both first and second parity (up to 15% less mastitis) within any of the production levels. In terms of fertility, stillbirth, and survival, crossbreds performed better than purebreds, and improved performance was independent of herd production level. We conclude that differences in performance between F1 crossbreds and Holstein are independent of production level.

Breeding technologies play a significant role in improving dairy cattle production. Scientifically proven tools for improved management and genetic gain in dairy herds, such as sexed semen, beef semen, genomic testing, dairy crossbreeding, and multiple ovulation embryo transfer (MOET), are readily available to dairy farmers. However, despite good accessibility, decreasing costs, and continuous development of these tools, their use in Sweden is limited. This study investigated Swedish dairy farmers' preferences for breeding tools through a survey including a discrete choice experiment. The survey was distributed online to 1 521 Swedish farmers and by an open link published through a farming magazine. In total, the study included 204 completed responses. The discrete choice experiment consisted of 10 questions with two alternative combinations, which gave 48 combinations in total. Utility values and part-worth values were computed using a conditional logit model based on the responses in the discrete choice experiment for nine groups of respondents: one group with all respondents, two groups based on respondents using dairy crossbreeding or not within the past 12 months, two based on herd size, two based on respondent age, and two based on whether respondents had used breeding advisory services or not. The strongest preferences in all groups were for using sexed semen and beef semen. Genomic testing was also significantly preferred by all groups of respondents. Except in large herds, MOET on own animals was significantly and relatively strongly disfavoured by all groups. Buying embryos had no significant utility value to any group. Dairy crossbreeding had low and insignificant utility values in the group of all respondents, but it was strongly favoured by the group that had used dairy crossbreeding within the past 12 months, and it was disfavoured by the group that had not. Part-worth values of combined breeding tools showed that combinations of sexed and beef semen, alone or with genomic testing without dairy crossbreeding, were the most preferred tools. Compared with the most common combinations of breeding tools used in the past 12 months, the part-worth values indicated that Swedish dairy farmers may prefer to use breeding tools more than they do today. Statements on the different breeding tools indicated that the respondents agreed with the benefits attributed to the breeding tools, but these benefits may not be worth the cost of genomic testing and the time consumption of MOET. These valuable insights can be used for further development of breeding tools. (c) 2021 The Author(s). Published by Elsevier B.V. on behalf of The Animal Consortium. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Mathematical and statistical modeling can be a very useful tool in understanding and fighting antimicrobial resistance (AMR). Here we present investigations of mixed effect models of varying complexity in order to identify and address possible management factors affecting the tetracycline AMR levels in Danish pig farms. Besides antimicrobial exposure during pigs life cycle, the type of production seems to also have an influence. The results concludes that not only fully integrated farms (CHR integrated) but also farms in a production network with a single ownership (CVR integrated) might have a preventive effect on levels of tetracycline AMR compared to more complex trading patterns.

This study simulated the consequences of crossbreeding between Swedish Holstein and Swedish Red on herd dynamics and herd profitability under Swedish conditions. Two base herds were simulated using a stochastic herd simulation model, SimHerd Crossbred. The herds reflected average Swedish conventional and organic herds having purebred Swedish Holstein. For each base herd, 3 breeding strategies were simulated: purebreeding, 2-breed terminal crossbreeding, and 2-breed rotational crossbreeding. The terminal crossbreeding strategy implied having a nucleus of Swedish Holstein and a proportion of F-1 Swedish Red x Swedish Holstein crossbred cows within the same herd. The crossbreds in this herd did not produce replacement heifers but exclusively beef x dairy crass calves. Beef semen was also used in the pure-breeding (10-20% in cows) and the rotational crossbreeding (40% in cows) strategies to retain a limited surplus of replacement heifers. To ensure an adequate number of crossbreds in the terminal crossbreeding strategy, X-sorted sexed semen was used for insemination in all the purebred heifers. The outcome was 67% purebred and 31% F-1 crossbreds in the herd. In addition, 31% heterosis was expressed compared with 67% heterosis expressed using a 2-breed rotational crossbreeding strategy. Compared with the pure-breeding strategy, crossbreeding increased the annual contribution margin per cow by (sic)20 to (sic)59, with the rotational crossbreeding strategy creating the largest profitability. The increased profitability was mainly due to improved functional traits, especially fertility. For the conventional production system, the replacement rate was 39.3% in the pure-breeding strategy and decreased to 35.8 and 30.1% in the terminal and rotational crossbreeding strategy, respectively. Similar changes happened in the organic production system. Additionally, the crossbreeding strategies earned (sic)22 to (sic)42 more annually per cow from selling live calves for slaughter due to the extended use of beef semen. Milk production was similar between pure-breeding and terminal crossbreeding, and only decreased 1 to 2% in rotational crossbreeding. These results show that crossbreeding between Swedish Holstein and Swedish Red can be profitable in both conventional and organic Swedish herds using the strategies we have simulated. However, some aspects remain to be investigated, such as the economically optimal breeding strategy, genetic improvement, and transition strategies.

During the last decade, the use of systematic crossbreeding in dairy cattle herds has increased in several countries of the world. The aim of this study was to estimate the effect of breed proportion and heterosis on milk production traits and udder health traits in dairy cattle. The study was based on records on milk yield (MY), protein yield (PY), fat yield (FY), somatic cell score (SCS), and mastitis (MAST) from 73,695 first-lactation dairy cows in 130 Danish herds applying systematic crossbreeding programs. Around 45% of the cows were crosses between Danish Holstein (DH), Danish Red (DR), or Danish Jersey (DJ), and the remaining were purebred DH, DR, or DJ. The statistical model included the fixed effects of herd-year, calving month, and calving age and an effect representing the lactation status of the cow. In addition, the model included a regression on calving interval from first to second lactation, a regression on the proportion of DH, DR, and DJ genes, and a regression on the degree of heterozygosity between DH and DR, DH and DJ, and DR and DJ. Random effects were the genetic effect of the cow and a residual. The effect of breed proportions was estimated relatively to DH. For MY, a pure DR yielded 461 kg milk less than DH, whereas a pure DJ yielded 2,259 kg milk less than a pure DH. Compared with DH, PY was 41.7 kg less for DJ, whereas PY for DR was 4.0 kg less than for DH. For FY, a DR yielded 10.6 kg less than DH, whereas there was no significant effect of breed proportion between DJ and DH. A DR cow had lower SCS (0.13) than DH, whereas DJ had higher SCS (0.14) than DH. There was no significant effect of breed proportion on MAST between the 3 breeds. Heterosis was significant in all combinations of breeds for MY, FY, and PY. Heterosis for crosses between DH and DR was 257 kg (3.2%), 11.9 kg (3.2%), and 8.9 kg (3.2%) for MY, PY, and FY, respectively. Corresponding figures for crosses between DH and DJ were 314 kg (4.4%), 14.3 kg (4.4%), and 10.4 kg (4.0%), whereas heterosis between DR and DJ was 462 kg (6.7%), 19.6 kg (6.7%), and 13.9 kg (5.4%) for MY, PY, and FY, respectively. Heterosis was only significant for SCS in the crosses between DH and DR. Heterosis effects for MAST were nonsignificant for all the crosses. The results obtained in this study demonstrate that in first lactation cows, there is a positive effect of heterosis on milk production traits, but limited effect on udder health traits.

Bacterial antimicrobial resistance (AMR) in pigs is an important public health concern due to its possible transfer to humans. We aimed at quantifying the relationship between the lifetime exposure of antimicrobials and seven antimicrobial resistance genes in Danish slaughter pig farms. AMR gene levels were quantified by qPCR of total-community DNA in faecal samples obtained from 681 batches of slaughter pigs. The lifetime exposure to antimicrobials was estimated at batch level for the piglet, weaner, and finisher periods individually for the sampled batches. We showed that the effect of antimicrobial exposure on the levels of AMR genes was complex and unique for each individual gene. Several antimicrobial classes had both negative and positive correlations with the AMR genes. From 10–42% of the variation in AMR gene levels could be explained in the final regression models, indicating that antimicrobial exposure is not the only important determinant of the AMR gene levels.

Vancomycin-resistant Enterococcus spp. is a major health problem worldwide and livestock have been implicated in constituting a reservoir for the transmission of vancomycin resistance to zoonotic pathogens. Vancomycin resistance determinants can be situated on mobile genetic elements and transferred between bacterial species The livestock reservoir must therefore be included in a risk assessment of the vancomycin resistance burden. Avoparcin, a vancomycin analogue, has not been used in Danish pig production for over 20 years and vancomycin has never been used. The objective of this study was to screen faecal samples from Danish pig farms for nine selected vancomycin resistance determinants. We found at least four different vancomycin resistance determinants in all screened Danish pig farms (665 finisher farms and 78 sow farms). The vancomycin resistance determinants present in vanB or vanG clusters were found at significantly different levels in sow and finisher farms. However, vanA was not detected in any of the farms. In conclusion, vancomycin resistance determinants are still present in Danish pig production 25 years after the ban on avoparcin use.

Extended lactation has shown economic benefits in dairy herds, but this strategy may affect the genetic lag between the production cows at herd level and the semen bulls unfavorably. We hypothesize, that a breeding strategy using sexed semen (SS) will reduce this effect and that SS and extended lactation has synergetic effects on the total economic return. A base scenario reflected an average Danish Holstein herd with 13 months calving interval (CInt) (BASE). Additionally, six scenarios with extended lactations: two with only first parity cows managed for a CInt of 15 (FIRST15) and 17 months (FIRST17), two with only cows in second parity or higher managed for a CInt of 15 (OLDER15) and 17 months (OLDER17), and two with all cows managed for a CInt of 15 (ALL15) and 17 months (ALL17). These were combined with four breeding schemes illustrating different uses of SS within each of the scenarios: SS in 50% genetically best heifers (SS50), 75% genetically best heifers (SS75), all heifers (SS100) and 70% genetically best heifers and 30% first parity cows (SS7030). Beef semen was used when necessary to equalize the number of replacement heifers within extended lactation scenarios. The scenarios were simulated using two stochastic simulation models for estimation of operational returns (SimHerd) and for genetic returns (ADAM). The genetic returns support that extended lactation has an unfavorable effect on genetic lag in SS0, because the generation interval among breeding females increases when extended lactation is introduced. Compared to BASE/SS0 most combinations of SS and extended lactation, with the exception of ALL17 combinations increased genetic returns as a result of decreased generation interval. Across extended lactation scenarios, reductions in genetic return relative to BASE was between €1 (FIRST15) and €12 (ALL17) per cow year within each scenario of SS. With increased use of SS, the genetic returns increased from €4 (SS50) to €12 (SS7030) per cow year, compared to SS0 within each scenario of extended lactation. In total economic return, the use of SS had effects between +€5 and +€29 per cow year due to increased use of beef semen, thus more sales of slaughter calves. Except for OLDER15 and OLDER17, the extended lactation scenarios within each scenario of SS increased total economic returns as well. In conclusion, the genetic lag caused by extended lactation can be reduced using SS, although there was no synergetic effect between SS and extended lactation on the total economic return.

Background The majority of antimicrobials given during the production of pigs are given to nursery pigs. The influence of antimicrobial use on the levels of antimicrobial resistant (AMR) genes is important to quantify to be able to assess the impact of resistance on the food chain and risk to human and animal health. Results This study investigated the response on the levels of nine AMR genes to five different treatment strategies with oxytetracycline, and the dynamics of gene abundance over time by following 1167 pigs from five different farms in Denmark. The results showed no significant difference between treatments and an increase in abundance for the efflux pump encoding tet(A) gene and the genes encoding the ribosomal protection proteins tet(O) and tet(W) tetracycline resistant genes following treatment, while tet(M) showed no response to treatment. However, it was also observed that the levels of tet(O), tet(W), and ermB in some farms would drift more over time compared to a single treatment-course with antibiotic. Conclusion This study underlines the large variation in AMR levels under natural conditions and the need for increased investigation of the complex interactions of antimicrobial treatment and other environmental and managerial practices in swine production on AMR gene abundance.

The primary objective of this study was to determine the minimum number of individual fecal samples to pool together in order to obtain a representative sample for herd level quantification of antimicrobial resistance (AMR) genes in a Danish pig herd, using a novel high-throughput qPCR assay. The secondary objective was to assess the agreement between different methods of sample pooling. Quantification of AMR was achieved using a high-throughput qPCR method to quantify the levels of seven AMR genes (ermB, ermF, sulI, sulII, tet(M), tet(O) and tet(W)). A large variation in the levels of AMR genes was found between individual samples. As the number of samples in a pool increased, a decrease in sample variation was observed. It was concluded that the optimal pooling size is five samples, as an almost steady state in the variation was observed when pooling this number of samples. Good agreement between different pooling methods was found and the least time-consuming method of pooling, by transferring feces from each individual sample to a tube using a 10μl inoculation loop and adding 3.5ml of PBS, approximating a 10% solution, can therefore be used in future studies.

The tetracycline resistance gene tet (K) was shown to be integrated within the predominant staphylococcal cassette chromosome mec (SCC mec ) element of Danish livestock-associated methicillin-resistant Staphylococcus aureus CC398 (LA-MRSA CC398). These LA-MRSA CC398 isolates already possessed tet (M), but the acquisition of tet (K) significantly improved their fitness at sublethal concentrations of tetracycline. Because tet (K) is genetically linked to SCC mec , the use of tetracycline in food animals may have contributed to the successful spread of LA-MRSA CC398.

Antimicrobial resistance in pigs has been under scrutiny for many years. However, many questions remain unanswered, including whether the initial antimicrobial resistance level of a pig will influence the antimicrobial resistance found at slaughter. Faecal samples from finishers pigs from 681 farms and from sows from 82 farms were collected, and levels of seven antimicrobial resistance genes, ermB, ermF, sulI, sulII, tet(M), tet(O), and tet(W), were quantified by high-capacity qPCR. There were 40 pairs of observations where the finishers were born in the farms of the sows. The objective of this study was to evaluate whether the levels of AMR genes found in finisher pigs at slaughter were associated with the levels in the farm where the finishers were born, and whether the levels of the AMR genes were equal in the sow and finisher pig populations. We found a significant positive correlation between the levels of AMR genes in finishers and the sows in the farms where the pigs were born for some of the genes (ermB (ρ=0.47, p-value=0.002), ermF (ρ=0.41, p-value=0.03), and tet(O) (ρ=0.33, p-value=0.04)). Furthermore, there were significant differences between AMR gene levels for the sow and finisher populations for ermB, ermF, sulI, sulII, tet(O), and tet(W), though not for tet(M).

SUMMARYAntimicrobial 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.

Crossbreeding has been shown to improve the longevity of dairy cattle in countries across the world. The aim of this study was to estimate heterosis, breed effects, and genetic parameters for longevity in crossbred dairy cattle among Danish Holstein (DH), Danish Red (DR), and Danish Jersey (DJ) breeds. Data were provided from 119 Danish commercial herds that use systematic crossbreeding (i.e., rotational crossbreeding). Additional data from 11 mixed-breed herds with DH and DJ were included to estimate reliable breed effects for DJ. Survival information on 73,741 cows was analyzed with a linear animal model using the artificial insemination–REML algorithm in the DMU package. Five longevity (L) traits were defined: days from first calving until the end of first lactation or culling (L1), days from first calving until the end of second lactation or culling (L2), days from first calving until the end of third lactation or culling (L3), days from first calving until the end of fourth lactation or culling (L4), and days from first calving until the end of fifth lactation or culling (L5). Heritabilities ranged between 0.022 and 0.090. Additive breed effects in units of days were estimated relative to DH for DR as −0.5 (L1), +10.5 (L2), +18.5 (L3), +11.9 (L4), and +28.6 (L5), and corresponding figures for DJ were +2.0, +0.5, +14.2, +27.7, and +44.0. Heterosis effects in L1 were low (1.2%) but favorable in crosses between DH and DR, whereas negative heterosis effects were estimated for crosses between DH and DJ (−2.5%) and DR and DJ (−1.2%). The largest heterosis effects for L2, L3, L4, and L5 were found in DH × DR and were favorable (+3.3, +5.7, +7.7, and +8.5%, respectively). Corresponding figures for heterosis effects in DH × DJ and DR × DJ were favorable as well: +2.3, +4.1, +5.6, and +6.2% in DH × DJ and +3.1, +7.3, +6.9, and +7.2% in DR × DJ. The favorable heterosis effects show that crossbreeding is an efficient tool for improving longevity in Danish dairy cattle.

The tetracycline resistance gene tet(K) was shown to be integrated within the predominant staphylococcal cassette chromosome mec (SCCmec) element of Danish livestock-associated methicillin-resistant Staphylococcus aureus CC398 (LA-MRSA CC398). These LA-MRSA CC398 isolates already possessed tet(M), but the acquisition of tet(K) significantly improved their fitness at sublethal concentrations of tetracycline. Because tet(K) is genetically linked to SCCmec, the use of tetracycline in food animals may have contributed to the successful spread of LA-MRSA CC398.

Combination therapy with several antibiotics is one strategy that has been applied in order to limit the spread of antimicrobial resistance. We compared the de novo evolution of resistance during combination therapy with the β-lactam ceftazidime and the fluoroquinolone ciprofloxacin with the resistance evolved after single-drug exposure. Combination therapy selected for mutants that displayed broad-spectrum resistance, and a major resistance mechanism was mutational inactivation of the repressor gene mexR that regulates the multidrug efflux operon mexAB–oprM. Deregulation of this operon led to a broad-spectrum resistance phenotype that decreased susceptibility to the combination of drugs applied during selection as well as to unrelated antibiotic classes. Mutants isolated after single-drug exposure displayed narrow-spectrum resistance and carried mutations in the MexCD–OprJ efflux pump regulator gene nfxB conferring ciprofloxacin resistance, or in the gene encoding the non-essential penicillin-binding protein DacB conferring ceftazidime resistance. Reconstruction of resistance mutations by allelic replacement and in vitro fitness assays revealed that in contrast to single antibiotic use, combination therapy consistently selected for mutants with enhanced fitness expressing broad-spectrum resistance mechanisms.

ObjectivesReliable methods for monitoring antimicrobial resistance (AMR) in livestock and other reservoirs are essential to understand the trends, transmission and importance of agricultural resistance. Quantification of AMR is mostly done using culture-based techniques, but metagenomic read mapping shows promise for quantitative resistance monitoring.MethodsWe evaluated the ability of: (i) MIC determination for Escherichia coli; (ii) cfu counting of E. coli; (iii) cfu counting of aerobic bacteria; and (iv) metagenomic shotgun sequencing to predict expected tetracycline resistance based on known antimicrobial consumption in 10 Danish integrated slaughter pig herds. In addition, we evaluated whether fresh or manure floor samples constitute suitable proxies for intestinal sampling, using cfu counting, qPCR and metagenomic shotgun sequencing.ResultsMetagenomic read-mapping outperformed cultivation-based techniques in terms of predicting expected tetracycline resistance based on antimicrobial consumption. Our metagenomic approach had sufficient resolution to detect antimicrobial-induced changes to individual resistance gene abundances. Pen floor manure samples were found to represent rectal samples well when analysed using metagenomics, as they contain the same DNA with the exception of a few contaminating taxa that proliferate in the extraintestinal environment.ConclusionsWe present a workflow, from sampling to interpretation, showing how resistance monitoring can be carried out in swine herds using a metagenomic approach. We propose metagenomic sequencing should be part of routine livestock resistance monitoring programmes and potentially of integrated One Health monitoring in all reservoirs.

The neuroleptic antipsychotic derivate thioridazine has been shown to increase the susceptibility of a methicillin-resistant Staphylococcus aureus (MRSA) isolate towards dicloxacillin. The aim of this study was to investigate the combinatorial effect of the two drugs on a broad selection of staphylococcal strains by analyzing a large collection of MRSA strains carrying different types of SCCmec, as well as MSSA strains. Transcription and translation of the resistance marker PBP2a encoded by mecA within the SCCmec cassette were analyzed by primer extension and western blotting. We observed increased susceptibility to dicloxacillin in the presence of thioridazine in all tested MRSA isolates. In contrast to previously published results, the synergistic effect was also applicable to methicillin-susceptible S. aureus (MSSA). We conclude that the combination of dicloxacillin and thioridazine potentiates the killing effect against S. aureus in a broad selection of clinical isolates. Additionally, the study indicates that the killing effect by the combinatorial treatment is independent of PBP2a-mediated resistance mechanisms.