Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Clonal expansion of a virulent Streptococcus suis serotype 9 lineage distinguishable from carriage subpopulations
Willemse, Niels ; Ark, Kees C.H. van der; Stockhofe-Zurwieden, Norbert ; Smith, Hilde ; Picavet, Daisy I. ; Solt-Smits, Conny van; Wisselink, Henk J. ; Schultsz, Constance ; Greeff, Astrid de - \ 2019
Scientific Reports 9 (2019)1. - ISSN 2045-2322

Streptococcus suis is a porcine pathogen, causing severe invasive infections. S. suis serotype 9 is increasingly causing disease in Dutch and Chinese pig herds, but it is unknown whether all serotype 9 isolates are equally virulent and markers that can identify virulent strains are not available. Therefore, discrimination between virulent isolates and carriage isolates typically not associated with disease, is currently not possible. We collected tonsillar S. suis isolates from 6 herds not previously diagnosed with S. suis infections, and clinical S. suis isolates of previously diseased pigs. We confirmed the virulence of a virulent type strain and one representative clinical isolate, and the lack of virulence of two carriage isolates, in a pig infection model. Phylogenetic analysis of whole genome sequences of 124 isolates resulted in 10 groups, of which two were almost uniquely populated by clinical isolates. The population structure of S. suis serotype 9 appears highly diverse. However, analysis of the capsule loci sequences showed variation in a single region which fully correlated with a virulent genotype. Transmission electron microscopy suggested differences in capsule thickness between carriage and clinical genotypes. In conclusion, we found that that the S. suis serotype 9 population in the Netherlands is diverse. A distinct virulence-associated lineage was identified and could be discriminated based on the capsule locus sequence. Whilst the difference in virulence cannot be directly attributed to the DNA sequence, the correlation of capsule locus sequence with virulence could be used in the development of diagnostic tests to identify potential virulent S. suis serotype 9 in pigs.

Toll-like receptor agonists as adjuvants for inactivated porcine reproductive and respiratory syndrome virus (PRRSV) vaccine
Vreman, Sandra ; McCaffrey, Joanne ; Popma-de Graaf, Ditta J. ; Nauwynck, Hans ; Savelkoul, Huub F.J. ; Moore, Anne ; Rebel, Johanna M.J. ; Stockhofe-Zurwieden, Norbert - \ 2019
Veterinary Immunology and Immunopathology 212 (2019). - ISSN 0165-2427 - p. 27 - 37.
Adjuvant - PRRSV - Skin vaccination - Toll-like receptor agonist - Vaccine

Toll-like receptor (TLR) agonists can effectively stimulate antigen-presenting cells (APCs) and are anticipated to be promising adjuvants in combination with inactivated vaccines. In this study, the adjuvant potential of three different TLR-agonists were compared with an oil-in-water (O/W) adjuvant in combination with inactivated porcine reproductive and respiratory syndrome virus (iPRRSV) applied by different administration routes: intramuscular (i.m.) or into the skin using dissolving microneedle (DMN) patches. Pigs received a prime vaccination followed by a booster vaccination four weeks later. TLR1/2 (Pam3Cys), TLR7/8 (R848) or TLR9 (CpG ODN) agonists were used as adjuvant in combination with iPRRSV strain 07V063. O/W adjuvant (Montanide™) was used as reference control adjuvant and one group received a placebo vaccination containing diluent only. All animals received a homologous challenge with PRRSV three weeks after the booster vaccination. Antibody and IFN-γ production, serum cytokines and viremia were measured at several time-points after vaccination and/or challenge, and lung pathology at necropsy. Our results indicate that a TLR 1/2, 7/8 or 9 agonist as adjuvant with iPRRSV does not induce a detectable PRRSV-specific immune response, independent of the administration route. However, the i.m. TLR9 agonist group showed reduction of viremia upon challenge compared to the non-vaccinated animals, supported by a non-antigen-specific IFN-γ level after booster vaccination and an anamnestic antibody response after challenge. Montanide™-adjuvanted iPRRSV induced antigen-specific immunity after booster combined with reduction of vireamia. Skin application of TLR7/8 agonist, but not the other agonists, induced a local skin reaction. Further research is needed to explore the potential of TLR agonists as adjuvants for inactivated porcine vaccines with a preference for TLR9 agonists.

In vivo transcriptomes of Streptococcus suis reveal genes required for niche-specific adaptation and pathogenesis
Arenas, Jesús ; Bossers-de Vries, Ruth ; Harders-Westerveen, José ; Buys, Herma ; Ruuls-van Stalle, Lisette M.F. ; Stockhofe-Zurwieden, Norbert ; Zaccaria, Edoardo ; Tommassen, Jan ; Wells, Jerry M. ; Smith, Hilde E. ; Greeff, Astrid de - \ 2019
Virulence 10 (2019)1. - ISSN 2150-5594 - p. 334 - 351.
infection - infectomics - pathogenesis - transcriptomics - zoonotic pathogen

Streptococcus suis is a Gram-positive bacterium and a zoonotic pathogen residing in the nasopharynx or the gastrointestinal tract of pigs with a potential of causing life-threatening invasive disease. It is endemic in the porcine production industry worldwide, and it is also an emerging human pathogen. After invasion, the pathogen adapts to cause bacteremia and disseminates to different organs including the brain. To gain insights in this process, we infected piglets with a highly virulent strain of S. suis, and bacterial transcriptomes were obtained from blood and different organs (brain, joints, and heart) when animals had severe clinical symptoms of infection. Microarrays were used to determine the genome-wide transcriptional profile at different infection sites and during growth in standard growth medium in vitro. We observed differential expression of around 30% of the Open Reading Frames (ORFs) and infection-site specific patterns of gene expression. Genes with major changes in expression were involved in transcriptional regulation, metabolism, nutrient acquisition, stress defenses, and virulence, amongst others, and results were confirmed for a subset of selected genes using RT-qPCR. Mutants were generated in two selected genes, and the encoded proteins, i.e., NADH oxidase and MetQ, were shown to be important virulence factors in coinfection experiments and in vitro assays. The knowledge derived from this study regarding S. suis gene expression in vivo and identification of virulence factors is important for the development of novel diagnostic and therapeutic strategies to control S. suis disease.

Neonatal porcine blood derived dendritic cell subsets show activation after TLR2 or TLR9 stimulation
Vreman, Sandra ; Auray, Gael ; Savelkoul, Huub F.J. ; Rebel, Annemarie ; Summerfield, Artur ; Stockhofe-Zurwieden, Norbert - \ 2018
Developmental and Comparative Immunology 84 (2018). - ISSN 0145-305X - p. 361 - 370.
Dendritic cell - Innate immunity - Neonate - Porcine - Toll like receptor ligand
The present study investigated the innate immune response in vitro to determine porcine neonate responses with Toll-like receptor (TLR)2 ligand (Pam3Cys) or TLR9 ligand (CpG) and compared these with adults. We identified the same phenotypically defined dendritic cell (DC) subsets and DC proportions in porcine neonate and adult blood by flow cytometry, which were plasmacytoid DCs (pDCs): CD14−CD4+CD172a+CADM1-) and conventional DCs (cDCs), being further divided into a cDC1 (CD14−CD4−CD172alowCADM1+) and a cDC2 (CD14−CD4−CD172a+CADM1+) subset. With neonatal cells, the TLR2 ligand induced a stronger TNF expression in monocytes and pDCs, and a stronger CD80/86 upregulation in cDC1, when compared to adult cells. Furthermore, in neonatal mononuclear cells TLR9 ligand was more potent at inducing IL12p40 mRNA expression. These results indicate clear responses of porcine neonatal antigen presenting cells after TLR2 and TLR9 stimulation, suggesting that corresponding ligands could be promising candidates for neonatal adjuvant application.
Environmental and social enrichment reduces susceptibility to co-infection in pigs
Dixhoorn, I.D.E. van; Reimert, I. ; Bolhuis, J.E. ; Groot Koerkamp, P.W.G. ; Kemp, B. ; Stockhofe-Zurwieden, Norbert - \ 2016
Supplementation of piglets with nutrient-dense complex milk replacer improves intestinal development and microbial fermentation
Greeff, A. de; Resink, J.W. ; Hees, H.M.J. van; Ruuls, L. ; Klaassen, G.J. ; Rouwers, S.M.G. ; Stockhofe-Zurwieden, N. - \ 2016
Journal of Animal Science 94 (2016)3. - ISSN 0021-8812 - p. 1012 - 1019.
Circular intestinal growth - Gene expression - Gut health - Nutrient-dense complex milk replacer - Pig

Weaning of piglets causes stress due to environmental, behavioral, and nutritional stressors and can lead to postweaning diarrhea and impaired gut development. The diet changes experienced during weaning require extensive adaptation of the digestive system. A well-developed piglet that had creep-feed experience before weaning performs better after weaning. In the current study, the effect of providing sow-fed piglets with a supplemental nutrient-dense complex milk replacer (NDM) on gut development and growth performance was studied. Litters of sows with similar parities (3.6 ± 0.8) and similar numbers of live born piglets (13.5 ± 0.3) were assigned to 1 of 2 groups: 1 group of piglets had ad libitum access to NDM from Day 2 through 21 after birth, whereas the other group was used as controls. Nutrient-dense complex milk replacer–fed piglets were shown to be significantly heavier after 21 d of supplementation compared with the control piglets. At Day 21, 3 piglets from each litter were euthanized for morphological and functional analyses of the intestinal tract. The small intestines of NDM-fed piglets had significantly higher weights (g) as well as significantly higher relative weight:length ratios (g//cm) compared with the small intestines of control piglets (P <0.05). Morphometric analysis demonstrated that villi length and numbers of goblet cells did not differ between groups. However, NDM-fed piglets had deeper crypts (P <0.001) and an increased expression of the cell-proliferation marker proliferating cell nuclear antigen in crypts (P <0.05), suggesting higher cell-proliferation rates. The gene encoding IGF- 1 showed a tendency to higher gene expression in the jejunum from NDM-fed piglets (P = 0.07) compared with the jejunum from control piglets, suggesting that IGF-1 might be involved in the regulation of cell proliferation and intestinal growth. Finally, as a result of dietary fiber in NDM, piglets showed significantly increased concentrations of metabolic fermentation products. This suggests differences in metabolic activity in the colon between treatment groups. In conclusion, providing sow-fed piglets with NDM before weaning stimulates intestinal proliferation, leading to increased circular growth. Nutrient-dense complex milk replacer supplementation might, therefore, help piglets through the transition period at weaning by increased BW and increased capacity for uptake of nutrients.

Pneumococcal colonization and invasive disease studied in a porcine model
Greeff, Astrid de; Selm, Saskia van; Buys, Herma ; Harders-Westerveen, José F. ; Tunjungputri, Rahajeng N. ; Mast, Quirijn de; Ven, Andre J. van der; Stockhofe-Zurwieden, Norbert ; Jonge, Marien I. de; Smith, Hilde E. - \ 2016
BMC Microbiology 16 (2016)1. - ISSN 1471-2180
Animal model - Colonization - Pigs - Streptococcus pneumoniae

Background: Streptococcus pneumoniae, a Gram-positive bacterium carried in the human nasopharynx, is an important human pathogen causing mild diseases such as otitis media and sinusitis as well as severe diseases including pneumonia, meningitis and sepsis. There is a strong resemblance between the anatomy, immunology and physiology of the pig and human species. Furthermore, there are striking similarities between S. suis pathogenesis in piglets and S. pneumoniae pathogenesis in humans. Therefore, we investigated the use of piglets as a model for pneumococcal colonization and invasive disease. Results: Intravenous inoculation of piglets with an invasive pneumococcal isolate led to bacteraemia during 5 days, showing clear bacterial replication in the first two days. Bacteraemia was frequently associated with fever and septic arthritis. Moreover, intranasal inoculation of piglets with a nasopharyngeal isolate led to colonization for at least six consecutive days. Conclusions: This demonstrates that central aspects of human pneumococcal infections can be modelled in piglets enabling the use of this model for studies on colonization and transmission but also on development of vaccines and host-directed therapies. Moreover this is the first example of an animal model inducing high levels of pneumococcal septic arthritis.

Invasive pneumococcal disease leads to activation and hyperreactivity of platelets
Tunjungputri, Rahajeng N. ; Jonge, Marien I. de; Greeff, Astrid de; Selm, Saskia van; Buys-Bergen, Herma ; Harders-Westerveen, Jose F. ; Stockhofe-Zurwieden, Norbert ; Urbanus, Rolf T. ; Groot, Phillip G. De; Smith, Hilde E. ; Ven, Andre J. van der; Mast, Quirijn de - \ 2016
Thrombosis Research 144 (2016). - ISSN 0049-3848 - p. 123 - 126.
Infection - Myocardial infarction - Platelet activation - Platelets - Pneumonia - Streptococcus pneumoniae

Using a novel porcine model of intravenous Streptococcus pneumoniae infection, we showed that invasive pneumococcal infections induce marked platelet activation and hyperreactivity. This may contribute to the vascular complications seen in pneumococcal infection.

Characterization of immune responses following homologous reinfection of pigs with European subtype 1 and 3 porcine reproductive and respiratory syndrome virus strains that differ in virulence
Weesendorp, Eefke ; Stockhofe-Zurwieden, Norbert ; Nauwynck, Hans J. ; Popma-De Graaf, D.J. ; Rebel, Johanna M.J. - \ 2016
Veterinary Microbiology 182 (2016). - ISSN 0378-1135 - p. 64 - 74.
Genetic subtypes - Homologous reinfection - Immune response - Porcine reproductive and respiratory syndrome virus - Protection

Porcine reproductive and respiratory syndrome virus (PRRSV) causes significant economic losses to the pork industry worldwide. Vaccination results often in limited protection. Understanding host immune responses elicited by different PRRSV strains could help to develop more efficacious vaccines. In the current study we characterized immunological responses and viral kinetics in pigs after primo infection and homologous challenge of the highly virulent European subtype 3 strain Lena, and the moderate to low virulent subtype 1 strain LV. Eighteen pigs were infected per strain, and 18 non-infected pigs served as control. Post mortem analysis was performed at days 7, 46 and 60 p.i. At day 46, pigs were challenged with the homologous strain. After the first inoculation, pigs infected with Lena developed fever and clinical symptoms, while this was not observed in pigs infected with LV. Virus titres in serum were about 100-fold higher in pigs infected with Lena than in pigs infected with LV. An inflammatory response was observed in pigs after primo infection with Lena with significantly higher levels of IL-12, IL-1β and TNF-α in the bronchoalveolar lavage. IFN-γ ELISPOT assay showed comparable responses between Lena and LV. Neutralizing antibodies were detected earlier in serum of pigs infected with Lena than in pigs infected with LV. After the challenge, a boost in antibody levels in both groups was observed. Challenge infection resulted in both groups in complete protection and sterile immunity, with no viraemia, clinical symptoms or viral RNA in tissues. In conclusion, although there were clear differences in immunological, clinical and virological responses to the primo infection, there were no differences observed in protection against homologous challenge.

Host-pathogen interaction at the intestinal mucosa correlates with zoonotic potential of Streptococcus suis
Ferrando, Maria Laura ; Greeff, Astrid De; Rooijen, W.J.M. Van; Stockhofe-Zurwieden, Norbert ; Nielsen, Jens ; Wichgers Schreur, P.J. ; Pannekoek, Yvonne ; Heuvelink, Annet ; Ende, Arie Van Der; Smith, Hilde ; Schultsz, Constance - \ 2015
The Journal of Infectious Diseases 212 (2015)1. - ISSN 0022-1899 - p. 95 - 105.
clonal complex - intestinal translocation - piglets. - serotype - Streptococcus suis - tight junctions - zoonotic infections

Streptococcus suis has emerged as an important cause of bacterial meningitis in adults. The ingestion of undercooked pork is a risk factor for human S. suis serotype 2 (SS2) infection. Here we provide experimental evidence indicating that the gastrointestinal tract is an entry site of SS2 infection. Methods. We developed a noninvasive in vivo model to study oral SS2 infection in piglets.We compared in vitro interaction of S. suis with human and porcine intestinal epithelial cells (IEC). Results. Two out of 15 piglets showed clinical symptoms compatible with S. suis infection 24-48 hours after ingestion of SS2. SS2 was detected in mesenteric lymph nodes of 40% of challenged piglets. SS2 strains isolated from patients showed significantly higher adhesion to human IEC compared to invasive strains isolated from pigs. In contrast, invasive SS9 strains showed significantly higher adhesion to porcine IEC. Translocation across human IEC, which occurred predominately via a paracellular route, was significantly associated with clonal complex 1, the predominant zoonotic genotype. Adhesion and translocation were dependent on capsular polysaccharide production. Conclusions. SS2 should be considered a food-borne pathogen. S. suis interaction with human and pig IEC correlates with S. suis serotype and genotype, which can explain the zoonotic potential of SS2.

Effects of solid feed level and roughage-to-concentrate ratio on ruminal drinking and passage kinetics of milk replacer, concentrates, and roughage in veal calves
Berends, H. ; Borne, J.J.G.C. van den; Stockhofe-Zurwieden, N. ; Gilbert, M.S. ; Zandstra, T. ; Pellikaan, W.F. ; Reenen, C.G. van; Bokkers, E.A.M. ; Gerrits, W.J.J. - \ 2015
Journal of Dairy Science 98 (2015)8. - ISSN 0022-0302 - p. 5621 - 5629.
Effects of solid feed (SF) level and roughage-to-concentrate (R:C) ratio on ruminal drinking and passage kinetics of milk replacer, concentrate, and roughage were studied in veal calves. In total, 80 male Holstein-Friesian calves (45 ± 0.2 kg of body weight) were divided over 16 pens (5 calves per pen). Pens were randomly assigned to either a low (LSF) or a high (HSF) SF level and to 1 of 2 R:C ratios: 20:80 or 50:50 on a dry matter (DM) basis. Roughage was composed of 50% corn silage and 50% chopped wheat straw on a DM basis. At 27 wk of age, measurements were conducted in 32 calves. During the measurement period, SF intake was 1.2 kg of DM/d for LSF and 3.0 kg of DM/d for HSF, and milk replacer intake averaged 2.3 kg of DM/d for LSF and 1.3 kg of DM/d for HSF. To estimate passage kinetics of milk replacer, concentrate, and straw, indigestible markers (CoEDTA, hexatriacontane C36, Cr-neutral detergent fiber) were supplied with the feed as a single dose 4, 24, and 48 h before assessment of their quantitative recovery in the rumen, abomasum, small intestine, and large intestine. Rumen Co recovery averaged 20% of the last milk replacer meal. Recoveries of Co remained largely unaffected by SF level and R:C ratio. The R:C ratio did not affect rumen recovery of C36 or Cr. Rumen fractional passage rate of concentrate was estimated from recovery of C36 in the rumen and increased from 3.3%/h for LSF to 4.9%/h for HSF. Rumen fractional passage rate of straw was estimated from Cr recovery in the rumen and increased from 1.3%/h for LSF to 1.7%/h for HSF. An increase in SF level was accompanied by an increase in fresh and dry rumen contents. In HSF calves, pH decreased and VFA concentrations increased with increasing concentrate proportion, indicating increased fermentation. The ratio between Cr and C36 was similar in the small and large intestine, indicating that passage of concentrate and straw is mainly determined by rumen and abomasum emptying. In conclusion, increasing SF level introduces large variation in passage kinetics of dietary components, predominantly in the rumen compartment. The SF level, rather than the R:C ratio, influences rumen recovery of concentrate and roughage. Our data provide insight in passage kinetics of milk (Co representing the milk replacer) and SF (Cr and C36 representing roughage and concentrate, respectively) and may contribute to the development of feed evaluation models for calves fed milk and SF.
Schmallenberg virus detection in bovine semen after experimental infection of bulls.
Poel, W.H.M. van der; Parlevliet, J.M. ; Verstraten, E.R.A.M. ; Kooi, E.A. ; Hakze-van der Honing, R.W. van der; Stockhofe-Zurwieden, N. - \ 2014
Epidemiology and Infection 142 (2014)07. - ISSN 0950-2688 - p. 1495 - 1500.
antibodies - cattle
To study Schmallenberg virus (SBV) excretion in bovine semen after experimental infection, two bulls were inoculated subcutaneously with a SBV isolate (1 ml Vero cell culture 106 TCID50). After inoculation (at day 0), semen was collected daily from both animals for 21 days and samples were tested for SBV by qRT–PCR assay. At 24 days post-inoculation both animals were subjected to necropsy and the genital organs and lymph nodes draining these organs were also tested for SBV RNA (qRT–PCR). After SBV infection both animals in the study showed viraemia (qRT–PCR) with fever and diarrhoea. SBV RNA could be detected in semen from both animals. The highest SBV RNA concentrations in semen were found in the first week (days 4–7 post-inoculation) but concentrations were relatively low (Ct values 30–39). Viable SBV was only isolated from blood samples and not from semen or genital tissues.
Schmallenberg virus antibody persistence in adult cattle after natural infection and decay of maternal antibodies in calves
Elbers, A.R.W. ; Stockhofe-Zurwieden, N. ; Poel, W.H.M. van der - \ 2014
BMC Veterinary Research 10 (2014). - ISSN 1746-6148
netherlands
Background: Schmallenberg virus (SBV) has swept through the major part of Europe in the period 2011–2013. A vaccine against SBV has been developed and may be a possible preventive instrument against infection. Presently, there is no data available to refute the assumption that natural SBV infection results in long-term immunity. In that respect, it is of interest to know how long (protecting) virus-neutralizing antibodies are present in naturally infected animals. New-born calves acquire passive immunity from their dams by ingestion and absorption of antibodies present in colostrum, which can block the production of serum antibodies when vaccine is administered to calves with maternally derived antibodies. In that respect, it is useful to know how long it takes for maternal antibodies against SBV to disappear in young animals born from infected dams. Results: Longitudinal whole-herd serological monitoring using virus neutralization test (VNT) indicated that 80% of adult dairy cows still had measurable antibodies against SBV at least 24 months after the estimated introduction of the virus into the herd. Median 2Log VNT titer of the adult dairy cows (=1 year) dropped from 8.6 to 5.6 in a period of 17 months. Median 2Log VNT maternal antibodies titers of calves sampled within 30 days after birth was 8. Calves lost their maternally-derived antibodies after 5–6 months. There was a definite positive relationship between the VNT titer of the dam and the VNT titer of the corresponding calf (age = 30 days) of dam-calf combinations sampled on the same day: the higher the VNT titer of the dam, the higher the VNT titer (maternal antibodies) of the calf. Conclusions: Our field data support the assumption that natural SBV infection in adult cows results in persistence of specific antibodies for at least two years. Based on the observed decay of maternally-derived antibodies in calves, it is presumed safe to vaccinate calves against SBV at an age of approximately 6 months
Intratypic heterologous vaccination of calves can induce an antibody response in presence of maternal antibodies against foot-and-mouth disease virus
Dekker, A. ; Eble, P.L. ; Stockhofe-Zurwieden, N. ; Chenard, G. - \ 2014
BMC Veterinary Research 10 (2014). - ISSN 1746-6148
Background - Maternal antibodies can interfere with foot-and-mouth disease vaccination. In this study we determined whether intratypic heterologous vaccination could help to improve herd immunity. Results - In unvaccinated calves, a half-life of maternal antibodies of 21 days was determined. At two weeks of age, calves without maternal antibodies showed a good antibody response against both vaccines used in the trial, while in calves with maternal antibodies no antibody response to homologous vaccination (A Turkey 14/98) but a limited antibody response to intratypic heterologous vaccination (A22 Iraq) was observed. Conclusion - Two weeks old calves without maternal antibodies respond well to vaccination, but when emergency vaccination is carried out in a region that uses prophylactic vaccination, using an intratypic heterologous vaccine strain may improve the immunity in calves with maternal antibodies
Effects of roughage source, amount, and particle size on behavior and gastrointestinal health of veal calves
Webb, L.E. ; Bokkers, E.A.M. ; Heutinck, L.F.M. ; Engel, B. ; Buist, W.G. ; Rodenburg, T.B. ; Stockhofe-Zurwieden, N. ; Reenen, C.G. van - \ 2013
Journal of Dairy Science 96 (2013)12. - ISSN 0022-0302 - p. 7765 - 7776.
growth-performance - rumen development - abomasal ulcers - iron-deficiency - beef-calves - dry feed - welfare - forestomach - stomach - damage
The European Union 1997 Directive, stipulating that veal calves should be fed a minimum of 50 to 250 g of fibrous feed from 8 to 20 wk of age, is vague. A fibrous feed ration maximum of 250 g has been implicated in welfare issues, namely the occurrence of abnormal oral behaviors and poor gastrointestinal health. Past research suggests that this amount is insufficient to prevent the development of abnormal oral behaviors and enabling good rumen development. Different sources and particle sizes of roughage could lead to very different welfare outcomes. In a 3 × 2 × 2 factorial design, 240 group-housed calves (10 ± 1 d; 46.1 ± 0.1 kg) were fed different roughage sources (straw, maize silage, or maize cob silage; the latter 2 were dried and provided no extra moisture compared with straw) in 2 amounts (250 or 500 g of dry matter per day), and 2 particle sizes (chopped or ground). Roughage was supplemented to milk replacer (MR) from 2 wk after arrival. In addition, 60 calves were fed 1 of 3 additional control treatments: MR only (n = 20), MR plus an iron supplement (n = 20), or MR plus ad libitum hay (n = 20). Oral behaviors were recorded using instantaneous scan sampling at 2-min intervals for 2 h in 3 periods per day, at 12 and 22 wk of age. Calves were slaughtered at 24 wk of age and rumen and abomasal health parameters were recorded. Limited provision of straw resulted in behavior comparable with that from unlimited provision of hay, with reduced tongue playing and oral manipulation of the environment, as well as increased chewing compared with diets with no roughage supplement. Straw prevented ruminal hairballs, but impaired rumen development and increased abomasal damage. A higher ration of roughage increased chewing (12 wk), decreased oral manipulation of the trough (12 and 22 wk) and the pen (22 wk), and increased rumen weight. However, more roughage led to increased abomasal damage for certain parameters. Longer feed particles had no obvious benefits for behavior, but decreased hairball prevalence. Overall, unlimited hay had the highest benefit for both behavior and gastrointestinal health. Adding iron to the MR did not alter behavior or gastrointestinal health compared with MR without iron supplement. This study demonstrated that different roughage sources, amounts, and particle sizes have different effects on veal calf behavior and gastrointestinal health, and hence on veal calf welfare. Key words: veal calf , behavior , gastrointestinal health , roughage
Rapid emergence of a virulent PB2 E627K variant during adaptation of highly pathogenic avian influenza H7N7 virus to mice
Jong, M.C. de; Stockhofe-Zurwieden, N. ; Verheij, E.S. ; Boer-Luijtze, E.A. de; Ruiter, S.J.M. ; Leeuw, O.S. de; Cornelissen, A.H.M. - \ 2013
Virology journal 10 (2013). - ISSN 1743-422X
a virus - amino-acid - viral polymerase - molecular-basis - h5n1 viruses - host-range - mouse lung - humans - determinants - transmission
Background Highly pathogenic avian influenza (HPAI) viruses pose a potential human health threat as they can be transmitted directly from infected poultry to humans. During a large outbreak of HPAI H7N7 virus among poultry in The Netherlands in 2003, bird to human transmission was confirmed in 89 cases, of which one had a fatal outcome. Methods To identify genetic determinants of virulence in a mammalian host, we passaged an avian H7N7/03 outbreak isolate in mouse lungs and evaluated the phenotype of the mouse-adapted variant in animal models and in vitro. Results Three passages in mouse lungs were sufficient to select a variant that was highly virulent in mice. The virus had a MLD50 that was >4.3 logs lower than that of its non-lethal parental virus. Sequence analysis revealed a single mutation at position 627 in PB2, where the glutamic acid was changed to a lysine (E627K). The mouse-adapted virus has this mutation in common with the fatal human case isolate. The virus remained highly pathogenic for chickens after its passage in mice. In ferrets, the mouse-adapted virus induced more severe disease, replicated to higher titers in the lower respiratory tract and spread more efficiently to systemic organs compared with the parental virus. In vitro, the PB2 E627K mutation had a promoting effect on virus propagation in mammalian, but not in avian cells. Conclusions Our results show that the E627K mutation in PB2 alone can be sufficient to convert an HPAI H7N7 virus of low virulence to a variant causing severe disease in mice and ferrets. The rapid emergence of the PB2 E627K mutant during mouse adaptation and its pathogenicity in ferrets emphasize the potential risk of HPAI H7N7 viruses for human health.
Early host response in the mammary gland after experimental Streptococcus uberis challenge in heifers
Greeff, A. de; Zadoks, R.N. ; Ruuls, L. ; Toussaint, M. ; Nguyen, T.K. ; Downing, A. ; Rebel, J.M.J. ; Stockhofe-Zurwieden, N. ; Smith, H.E. - \ 2013
Journal of Dairy Science 96 (2013)6. - ISSN 0022-0302 - p. 3723 - 3736.
innate immune-response - lipopolysaccharide-binding protein - clinical mastitis - intramammary infections - staphylococcus-aureus - lipoteichoic acid - bovine mastitis - dairy-cattle - 2 strains - epidemiology
Streptococcus uberis is a highly prevalent causative agent of bovine mastitis, which leads to large economic losses in the dairy industry. The aim of this study was to examine the host response during acute inflammation after experimental challenge with capsulated Strep. uberis. Gene expression in response to Strep. uberis was compared between infected and control quarters in 3 animals. All quarters (n=16) were sampled at 16 different locations. Microarray data showed that 239 genes were differentially expressed between infected and control quarters. No differences in gene expression were observed between the different locations. Microarray data were confirmed for several genes using quantitative PCR analysis. Genes differentially expressed due to early Strep. uberis mastitis represented several stages of the process of infection: (1) pathogen recognition; (2) chemoattraction of neutrophils; (3) tissue repair mechanisms; and (4) bactericidal activity. Three different pathogen recognition genes were induced: ficolins, lipopolysaccharide binding protein, and toll-like receptor 2. Calgranulins were found to be the most strongly upregulated genes during early inflammation. By histology and immunohistochemistry, we demonstrated that changes in gene expression in response to Strep. uberis were induced both in infiltrating somatic milk cells and in mammary epithelial cells, demonstrating that the latter cell type plays a role in milk production as well as immune responsiveness. Given the rapid development of inflammation or mastitis after infection, early diagnosis of (Strep. uberis) mastitis is required for prevention of disease and spread of the pathogen. Insight into host responses could help to design immunomodulatory therapies to dampen inflammation after (early) diagnosis of Strep. uberis mastitis. Future research should focus on development of these early diagnostics and immunomodulatory components for mastitis treatment.
Phenotypic modulation and cytokine profiles of antigen presenting cells by European subtype 1 and 3 porcine reproductive and respiratory syndrome virus strains in vitro and in vivo
Weesendorp, E. ; Stockhofe-Zurwieden, N. ; Popma-de Graaf, D.J. ; Fijten, H.P.D. ; Rebel, J.M.J. - \ 2013
Veterinary Microbiology 167 (2013)3-4. - ISSN 0378-1135 - p. 638 - 650.
dendritic cells - immune-responses - pigs - prrsv - infection - apoptosis - genotype - disease - lungs - interleukin-10
Porcine reproductive and respiratory syndrome virus (PRRSV) causes continuous problems in the pig industry, due to high costs of outbreaks and reduced welfare of diseased pigs. The severity of infection is, partly, dependent on the virus strain. Recently isolated Eastern-European subtype 3 strains are more pathogenic than the widespread subtype 1 strains. There is, however, almost no information available about the mechanisms involved in the pathogenicity of these subtype 3 strains. The objective of the present study was to characterize the in vitro and in vivo response of two European subtype 1 strains, Belgium A and Lelystad-Ter Huurne (LV), and a virulent subtype 3 strain, Lena, in bone marrow-derived dendritic cells (BM-DC) (in vitro) and alveolar macrophages (in vitro and in vivo). It was shown that infection with the Lena strain resulted in a higher apoptosis of cells in vitro and a higher level of infectivity in vitro and in vivo than the other virus strains. Furthermore, infection with Lena resulted in a small downregulation of the immunologically relevant cell surface molecules SLA-I, SLA-II and CD80/86 in vitro, and SLA-II in vivo. In spite of these differences, in vitro cytokine responses did not differ significantly between strains, except for the absence of IL-10 production by Lena in BM-DC. The higher infectivity, apoptosis and downregulation of the cell surface molecules, may have contributed to the increased pathogenicity of Lena, and have dampened specific immune responses. This could explain the delayed and decreased adaptive immune responses observed after infections with this strain.
Comparative analysis of immune responses following experimental infection of pigs with European porcine reproductive and respiratory syndrome virus strains of differing virulence
Weesendorp, E. ; Morgan, S. ; Stockhofe-Zurwieden, N. ; Popma-de Graaf, D.J. ; Graham, S.P. ; Rebel, J.M.J. - \ 2013
Veterinary Microbiology 163 (2013)1-2. - ISSN 0378-1135 - p. 1 - 12.
classical swine-fever - immunological responses - circovirus type-2 - t-lymphocytes - syndrome prrs - cells - disease - expression - antibodies - cytokines
Porcine reproductive and respiratory syndrome virus (PRRSV) is difficult to control due to a high mutation rate and the emergence of virulent strains. The objective of this study was to analyze the immunological and pathological responses after infection with the European subtype 3 strain Lena in comparison to subtype 1 strains Belgium A and Lelystad-Ter Huurne (LV). Sixteen pigs were inoculated per strain, and sixteen pigs with PBS. At days 7 and 21 post-inoculation (p.i.), four pigs per group were immunized with an Aujeszky disease vaccine (ADV) to study the immune competence after PRRSV infection. Infection with the Lena strain resulted in fever and clinical signs. This was not observed in the Belgium A or LV-infected pigs. Infection with the Lena strain resulted in high virus titers in serum, low numbers of IFN-¿ secreting cells, a change in leukocyte populations and a delayed antibody response to immunization with ADV. Levels of IL-1ß, IFN-a, IL-10, IL-12, TNF-a and IFN-¿ mRNA of the Lena-infected pigs were increased during the first week of infection. For pigs infected with the Belgium A or LV strain, the effects of infection on these parameters were less pronounced, although for the Belgium A-infected pigs, the level of the analyzed cytokines, except for TNF-a, and leukocyte populations were comparable to the Lena-infected pigs. These results suggest that while the outcome of infection for the three strains was comparable, with mostly clearance of viremia at day 33 p.i, differences in immune responses were observed, perhaps contributing to their virulence.
Garlic impairs Actinobacillus pleuropneumoniae in vitro and alleviates pleuropneumonia in a pig model
Becker, P.M. ; Wikselaar, P.G. van; Mul, M.F. ; Pol, J. ; Engel, B. ; Wijdenes, J.W. ; Peet-Schwering, C.M.C. van der; Wisselink, H.J. ; Stockhofe-Zurwieden, N. - \ 2012
In: Book of Abstracts of the International Symposium on Alternatives to Antibiotics: Challenges and Solutions in Animal Productions. - Paris : The World Organisation for Animal Health (OIE) - p. 1.21 - 1.21.
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