State University Veterinary Diagnostic Laboratory Research Articles

Selection of Syndromes and Algorithms for Monitoring Bovine Laboratory Test Data

ObjectiveStandardize selection of indicator data streams and corresponding alerting algorithms for syndromic, reportable disease, and confirmed diagnostic categories derived from veterinary laboratory test order data for bovines.IntroductionThe Johns Hopkins University Applied Physics Laboratory is collaborating with epidemiologists of the US Dept. of Agriculture's Animal and Plant Health Inspection Service (APHIS) Center for Epidemiology and Animal Health (CEAH) to increase animal health surveillance capacity. CEAH monitors selected syndromic animal health indicators for stakeholder reporting. This project’s goal was to extend this capacity to bovine veterinary laboratory test accession data.MethodsIndicators for weekly monitoring were derived from bovine test records from the Colorado State University Veterinary Diagnostic Laboratory System from 27 Jun 2010 - 29 May 2016. Selected indicator types were syndromic test orders, disease-specific orders, and disease-specific positive results. Indicators were adopted if APHIS epidemiologists considered them worth monitoring and if they were represented by at least 100 lab accessions.Ten syndromes were chosen for routine monitoring based on body systems, bovine-specific concerns (e.g. mastitis), and concepts to capture novel threats. Reportable diseases were chosen from the list published by the Colorado Dept. of Agriculture [1]. Based on APHIS concerns and test order frequencies, 4 diseases were chosen for weekly monitoring: Bluetongue, Brucellosis, Epizootic Hemorrhagic Disease, and Paratuberculosis. To monitor positives, we considered both the number and the ratio of herds with at least one positive result for each disease. For included tests (excluding results quantified with antibody levels), we counted an accession as “positive” if the result field contained strings “positive”, “suspect”, or “detect” without negation terms. For weekly counts, we added the number of herds with any positives after deduplication. Diseases adopted for monitoring of positive results were Bovine Viral Diarrhea, Trichomoniasis, and Paratuberculosis.From experience and literature, we compared variants of 4 algorithm types, including: the C2 method of the CDC Early Aberration Reporting System, a CuSUM control chart with a sliding baseline, the temporal scan statistic Gscan applied to hospital infection counts, and the CDC Historical Limits method.We adapted a semisynthetic simulation approach for algorithm comparison in which authentic disease count data are used as baseline, and simulated signals are added to the background as detection targets. In discussions about specific diseases and veterinary testing practice, CEAH required sensitivity to one-week data spikes as well as effects of health threats with multi-week incubation periods and more gradual test ordering. For such gradual signals, we chose the lognormal signal model of Sartwell applied to incidence data for many diseases. Incubation periods vary widely by disease, and for this project, we chose lognormal parameters such that 90% of reported cases would occur within 6 weeks. We conducted separate algorithm detection trials for spike and gradual signals.Calculations of sensitivity, alert rate, and timeliness were derived with sets of 1000 repeated trials for each combination of algorithm and syndrome or disease. We applied minimum performance requirements of 95% sensitivity, ≤1 alert per 8 weeks, and mean detection delays of <2 weeks. The rule adopted for recommending an alerting method was to seek the method with the lowest alert rate that satisfied the sensitivity, alert rate, and delay criteria.ResultsThe Table shows the syndromes with chosen algorithms and thresholds for detection of the gradual signals. The scan statistic Gscan and the historical limits method HistLim achieved consistently higher sensitivities with acceptable alert rates than the other methods applied . The presentation will extend the results to reportable disease and clinical positive indicators and to the spike signals for all indicators.ConclusionsAmong results for both signal types, the results yielded a few preferred methods covering all chosen indicator streams. Monitored indicators with median weekly counts = 0 remain a challenge requiring more background data and veterinarian judgment. From analysis of orders from the few available laboratories, manual review will be required to achieve accurate syndromic categorization for each lab. Monitoring of test positives will require combined analysis of positive herd counts and percentages (of all tested herds) due to routine variation in laboratory submissions.

Comparison of treatment history and antimicrobial sensitivity results of common bovine respiratory disease bacterial pathogens from cases submitted to the ISU-VDL from 2013-2015

Summarized susceptibility results from diagnostic laboratory cases are often used to evaluate trends in antibiotic resistance. The objective of this retrospective study was to evaluate summarized antimicrobial susceptibility data from bovine respiratory disease cases submitted to the Iowa State University Veterinary Diagnostic Laboratory in the context of reported antibiotic treatment history.

Considerations for veterinary practitioners involved in feed-related toxicology cases

The objective of this paper is to identify key considerations for production animal veterinarians involved in legal cases regarding feed-related errors. Veterinarians are trained in the science of management and production, but are often ill-prepared for advising clients on potential litigation that may result when large monetary losses are incurred. The availability of modern analytical equipment and technology can provide practitioners with more information to understand the interactions of toxins with production medicine. This information may further elucidate the source of potential losses. The following considerations were derived from experiences at the Iowa State University Veterinary Diagnostic Laboratory when involved in several legal cases. Since it is likely that production veterinarians will be involved in a legal case at some point in their career, this paper is intended to help practitioners to be better prepared for cases involving feed-related toxicology cases.

Economic impact of university veterinary diagnostic laboratories: A case study

Veterinary diagnostic laboratories (VDLs) play a significant role in the prevention and mitigation of endemic animal diseases and serve an important role in surveillance of, and the response to, outbreaks of transboundary and emerging animal diseases. They also allow for business continuity in livestock operations and help improve human health. Despite these critical societal roles, there is no academic literature on the economic impact of VDLs. We present a case study on the economic impact of the Iowa State University Veterinary Diagnostic Laboratory (ISUVDL). We use economic contribution analysis coupled with a stakeholder survey to estimate the impact. Results suggest that the ISUVDL is responsible for $2,162.46 million in direct output, $2,832.45 million in total output, $1,158.19 million in total value added, and $31.79 million in state taxes in normal years. In an animal health emergency this increases to $8,446.21 million in direct output, $11,063.06 million in total output, $4,523.70 million in total value added, and $124.15 million in state taxes. The ISUVDL receives $4 million annually as a direct state government appropriation for operating purposes. The $31.79 million in state taxes in normal years and the $124.15 million in state taxes in an animal health emergency equates to a 795% and 3104% return on investment, respectively. Estimates of the economic impact of the ISUVDL provide information to scientists, administrators, and policymakers regarding the efficacy and return on investment of VDLs.

Treatment history and antimicrobial susceptibility results for Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni isolates from bovine respiratory disease cases submitted to the Iowa State University Veterinary Diagnostic Laboratory from 2013 to 2015.

Bovine respiratory disease is the most costly disease facing the cattle industry. Increasing resistance to antimicrobial treatment has been presented as a significant contributing factor, often through summarized susceptibility testing data. We assessed the relationship between previous antimicrobial treatment and antimicrobial susceptibility results from isolates of Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni cultured from bovine respiratory cases submitted to the Iowa State University Veterinary Diagnostic Laboratory from 2013 to 2015. Antimicrobial susceptibility data from 1,251 bacterial isolates were included for analysis. More bacterial isolates from cattle that received antimicrobial treatment showed resistance compared to isolates from untreated cattle, and the percentage of resistant isolates increased as the number of antimicrobial treatments increased. Resistance to enrofloxacin, spectinomycin, tilmicosin, and tulathromycin was present in >75% of M. haemolytica isolates from cattle that had received 3 or more antimicrobial treatments; resistance to each of those 4 antimicrobials was present in ≤10% of M. haemolytica isolates from untreated cattle. Similar but less dramatic trends were apparent for isolates of P. multocida and H. somni. The percentage of multi-drug resistant bacterial isolates also increased with the number of treatments. Results of our study suggest that previous antimicrobial treatment may have a profound effect on antimicrobial susceptibility testing. Summarized susceptibility results from diagnostic laboratories should not be used to make generalized statements regarding trends in antimicrobial resistance without providing context regarding antimicrobial treatment history.

Development of Polioencephalomyelitis in Cesarean-Derived Colostrum-Deprived Pigs Following Experimental Inoculation with Either Teschovirus A Serotype 2 or Serotype 11

Teschovirus encephalomyelitis is a sporadic disease associated with Teschovirus A (PTV) serotype 1 and, less frequently, other serotypes. In recent years, the number of cases submitted to the Iowa State University Veterinary Diagnostic Laboratory with a history of posterior paresis has increased. Submission histories from various regions of the United States suggest a trend for clinical disease to persist in herds and affect a wider age-range of pigs than historically reported. Polioencephalitis and/or myelitis was consistently present and PTV was detected in affected neural tissue by PCR in a portion of cases. Sequencing from two clinical cases identified PTV-2 and PTV-11. To assess neuropathogenicity of these isolates, 5-week-old cesarean derived and colostrum-deprived pigs were assigned to three groups: negative control (n = 4), PTV-2-inoculated (n = 7), and PTV-11-inoculated (n = 7). Three PTV-2-inoculated pigs developed mild incoordination of the hind limbs, one of which progressed to posterior ataxia. While all PTV-11-inoculated pigs showed severe neurological signs consistent with Teschovirus encephalomyelitis, no evidences of neurological signs were observed in sham-inoculated animals. All PTV-2- and PTV-11-inoculated pigs had microscopic lesions consistent with Teschovirus encephalomyelitis. To our knowledge, this is the first description of PTV-11 and experimental study demonstrating the neuropathogenicity of PTV-11 in the United States.

PCV2d-2 is the predominant type of PCV2 DNA in pig samples collected in the U.S. during 2014–2016

Porcine circovirus type 2 (PCV2) vaccination was introduced in the US in 2006 and since has been adopted by most pig producers. While porcine circovirus associated disease (PCVAD) outbreaks are now relatively uncommon in the US, PCV2 remains a concern which is emphasized by increasing numbers of PCR and sequencing requests for PCV2. In the present study, randomly selected lung tissues from 586 pigs submitted in 2015 were tested for presence of PCV2 DNA. Positive samples were further characterized by sequencing and combined with available PCV2 open-reading-frame (ORF) 2 sequences from the client data base of the Iowa State University Veterinary Diagnostic Laboratory. The prevalence of PCV2 in the randomly selected lung tissues was 23% (135/586) with 11.3% PCV2a, 29% PCV2b and 71.8% for PCV2d subgroup PCV2d-2. A total of 455 ORF2 sequences obtained from 2014 through 2016 were analyzed and PCV2d accounted for 66.7% of the 2014 sequences, 71.8% of the 2015 sequences, and 72% of the 2016 sequences. Interestingly, only 1.9% (9/455) of the sequences belonged to the recently identified PCV2e genotype. The present data indicates that despite an almost 100% PCV2 vaccine coverage in the US, PCV2 DNA can still be detected in almost 1 of 4 randomly selected pig tissues. PCV2d-2 is now the predominant genotype in the USA suggesting that PCV2d-2 may have some advantage over PCV2a and PCV2b in its ability to replicate in pigs under vaccination pressure.

Outbreak of H5N2 highly pathogenic avian Influenza A virus infection in two commercial layer facilities: lesions and viral antigen distribution.

The largest outbreak of highly pathogenic avian Influenza A virus (HPAIV) infection in U.S. history began in December 2014 resulting in the euthanasia of millions of birds and collateral economic consequences to the U.S. poultry industry. We describe 2 cases of H5N2 HPAIV infection in laying hens in Iowa. Following a sharp increase in mortality with minimal clinical signs, 15 dead birds, from 2 unrelated farms, were submitted to the Iowa State University Veterinary Diagnostic Laboratory. Common lesions included diffuse edema and multifocal hemorrhage of the comb, catarrhal exudate in the oropharynx, and multifocal tracheal hemorrhage. Less common lesions included epicardial petechiae, splenic hemorrhage, and pancreatic necrosis. Influenza A virus nucleoprotein was detected by immunohistochemistry in multiple cell types including ependymal cells, the choroid plexus, neurons, respiratory epithelium and macrophages in the lung, cardiac myocytes, endothelial cells, necrotic foci in the spleen, Kupffer cells in the liver, and necrotic acinar cells in the pancreas. Real-time polymerase chain reaction and sequencing confirmed H5N2 HPAIV with molecular characteristics similar to other contemporary U.S. H5N2 HPAIVs in both cases.

Whole-Genome Sequences of Novel Porcine Circovirus Type 2 Viruses Detected in Swine from Mexico and the United States

A unique porcine circovirus type 2 capsid protein (ORF2) sequence was detected in swine samples submitted to the Iowa State University Veterinary Diagnostic Laboratory. The complete genome sequences of four viruses, one from Mexico and three from the United States, were determined to further characterize this novel PCV2 genotype.

PCR-based retrospective evaluation of diagnostic samples for emergence of porcine deltacoronavirus in US swine

Porcine deltacoronavirus (PDCoV) was first identified in Hong Kong in a regional surveillance study for Coronaviruses in 2012 and was detected for the first time in United States (US) swine in February 2014. However, it remains unknown if PDCoV had been introduced into the US prior to that time period. In the present study, 1734 clinical samples (903 cases) submitted to the Iowa State University Veterinary Diagnostic Laboratory (ISU VDL) for enteric disease diagnosis between October 2012 and December 2013 were tested retrospectively for PDCoV using a virus-specific real-time reverse transcription (RT) PCR targeting conserved region of the membrane gene. PDCoV genome was first detected in a fecal sample collected on August 19th 2013 from Minnesota. Subsequently, PDCoV was observed in samples collected on August 20th and August 27th from Iowa and on August 29th from Illinois. Therefore, with available samples submitted to the ISU VDL, it can be inferred that PDCoV has been present in US swine at least since August 2013.

Anti–glucagon-like peptide-1 immunoreactivity in samples of blood and ileum obtained from neonatal and adult alpacas

To compare numbers of L cells in intestinal samples and blood concentrations of glucagon-like peptide (GLP)-1 between neonatal and mature alpacas. Intestinal samples from carcasses of 4 suckling crias and 4 postweaning alpacas for immunohistochemical analysis and blood samples from 32 suckling crias and 19 healthy adult alpacas for an ELISA. Immunohistochemical staining was conducted in accordance with Oregon State University Veterinary Diagnostic Laboratory standard procedures with a rabbit polyclonal anti-GLP-1 primary antibody. Stained cells with staining results in ileal tissue were counted in 20 fields by 2 investigators, and the mean value was calculated. For quantification of GLP-1 concentrations, blood samples were collected into tubes containing a dipeptidyl peptidase-4 inhibitor. Plasma samples were tested in duplicate with a commercial GLP-1 ELISA validated for use in alpacas. Counts of stained cells (mean ± SD, 50 ± 18 cells) and plasma GLP-1 concentrations (median, 0.086 ng/mL; interquartile range, 0.061 to 0.144 ng/mL) were higher for suckling alpacas than for postsuckling alpacas (stained cells, 26 ± 4 cells; plasma GLP-1 concentration, median, 0.034 ng/mL; interquartile range, 0.015 to 0.048 ng/mL). Older alpacas had lower numbers of L cells in intestinal tissues and lower blood concentrations of GLP-1 than those in neonates. These findings suggested that there may be a decrease in the contribution of GLP-1 to insulin production in adult alpacas, compared with the contribution in neonates.

Detection and isolation of Influenza A virus subtype H1N1 from a small backyard swine herd in Colorado

Influenza A virus subtype H1N1 A(H1N1)pdm09 was first confirmed in pigs in the United States in October 2009. In November 2010, lungs and intestines from 2 York piglets from a small, privately owned herd were submitted to the Colorado State University Veterinary Diagnostic Laboratory. The submitting veterinarian reported rapid weight loss and signs of pneumonia in the piglets. Gross lesions included caudoventral pneumonia in both piglets, and histologic lesions in the lungs showed characteristics consistent with influenza virus and bacterial infection. Ribonucleic acid extracted from fresh lung homogenates from both piglets was positive for influenza A(H1N1)pdm09 by a real-time reverse transcription polymerase chain reaction. Virus was isolated from lung homogenates from both piglets in Madin-Darby canine kidney cells, as well as in 10-day-old specific pathogen-free embryonated chicken eggs. Sequence analysis showed 98% homology with 2009 H1N1 human isolates from across the United States and 98% homology against two 2009 and 2010 swine isolates from Nebraska and Minnesota. The current report documents the possible transmission of pandemic influenza A(H1N1)2009 virus [A(H1N1)pdm09] from a human being to a small, privately owned backyard swine herd. The owner was employed as a pharmacist, making occupational exposure to the pandemic influenza A(H1N1)pdm09 a possibility.

Emergence of Porcine epidemic diarrhea virus in the United States: clinical signs, lesions, and viral genomic sequences

During the 10 days commencing April 29, 2013, the Iowa State University Veterinary Diagnostic Laboratory received the first 4 of many submissions from swine farms experiencing explosive epidemics of diarrhea and vomiting affecting all ages, with 90-95% mortality in suckling pigs. Histology revealed severe atrophy of villi in all segments of the small intestines with occasional villus-epithelial syncytial cells, but testing for rotaviruses and Transmissible gastroenteritis virus (Alphacoronavirus 1) were negative. Negative-staining electron microscopy of feces revealed coronavirus-like particles and a pan-coronavirus polymerase chain reaction (PCR) designed to amplify a conserved region of the polymerase gene for all members in the family Coronaviridae produced expected 251-bp amplicons. Subsequent sequencing and analysis revealed 99.6-100% identity among the PCR amplicons from the 4 farms and 97-99% identity to the corresponding portion of the polymerase gene of Porcine epidemic diarrhea virus (PEDV) strains, with the highest identity (99%) to strains from China in 2012. Findings were corroborated at National Veterinary Services Laboratories using 2 nested S-gene and 1 nested N-gene PCR tests where the sequenced amplicons also had the highest identity with 2012 China strains. Whole genome sequence for the virus from 2 farms in 2 different states using next-generation sequencing technique was compared to PEDV sequences available in GenBank. The 2013 U.S. PEDV had 96.6-99.5% identity with all known PEDV strains and the highest identity (>99.0%) to some of the 2011-2012 Chinese strains. The nearly simultaneous outbreaks of disease, and high degree of homology (99.6-100%) between the PEDV strains from the 4 unrelated farms, suggests a common source of virus.

Prevalence of Campylobacter spp. relative to other enteric pathogens in grow-finish pigs with diarrhea

Salmonella spp., Lawsonia intracellularis, and Brachyspira spp. are pathogens commonly associated with diarrhea in growing and finishing pigs. Brachyspira spp. infection has recently reemerged as a significant concern due to an increase in the incidence of swine dysentery; however, the mechanisms underlying this increase in dysentery remain largely unknown. Pigs are also well-recognized as potential carriers of Campylobacter spp., particularly Campylobacter coli, yet enteric disease in swine associated with infection by these bacteria is considered uncommon and diagnosis has historically been based upon exclusion of other causes. Accordingly, Campylobacter culture is often excluded in routine diagnostic testing of cases of porcine enterocolitis and the incidence of infection is therefore largely unknown. In this study, feces from 155 cases of clinical diarrhea in grow-finish pigs submitted to the Iowa State University Veterinary Diagnostic Laboratory were cultured for Campylobacter spp. in addition to other testing as indicated for routine diagnostic investigation. Campylobacter culture was positive from 82.6% (128/155) of samples with C. coli accounting for 75% of isolates and Campylobacter jejuni for the remaining 25%. In 14.8% (23/155) of cases a Campylobacter spp. was the sole infectious agent detected; however, there was no association with any particular Campylobacter spp. Interestingly, for those cases with a laboratory diagnosis of Brachyspira-associated disease, 100% (15/15) were also culture positive for Campylobacter spp. suggesting a possible interrelationship between these bacteria in the pig gut. No association was noted between Campylobacter culture results and infection with either Salmonella spp. or L. intracellularis.

Rickets

Rickets can be attributed to nutritional, genetic, hormonal, or toxic disturbances and is classified as a metabolic bone disease. Rickets is most often associated with inappropriate dietary levels of calcium, phosphorus, and/or vitamin D. During a 27-month period (January 2010 through March 2012), the Iowa State University Veterinary Diagnostic Laboratory investigated causes of sudden, unexpected death and lameness in growing pigs throughout the Midwestern United States. Clinical observations from 17 growing pig cases included weakness, lameness, reluctance to move, muscle fasciculations and/or tremors, tetany, and death. Ribs were weak, soft, and bent prior to breaking; rachitic lesions were apparent at costochondral junctions in multiple cases. Acute and/or chronic bone fractures were also noted in multiple bones. Failure of endochondral ossification, expanded physes, infractions, thin trabeculae, and increased osteoclasts were noted microscopically. Decreased bone ash and serum 25(OH)D(3), combined with clinical and microscopic evaluation, confirmed a diagnosis of vitamin D-dependent rickets in all cases. In 3 cases, disease was linked to a specific nutrient supplier that ultimately resulted in a voluntary feed recall; however, most cases in the current investigation were not associated with a particular feed company. The present report describes vitamin D-associated rickets and its importance as a potential cause of weakness, lameness, muscle fasciculations, recumbency or sudden unexpected death in swine, and describes appropriate samples and tests for disease diagnosis.

Evaluation of a commercial rapid test kit for detecting bovine enteric pathogens in feces

Recently a commercial antigen-capture enzyme-linked immunosorbent assay kit in the form of a dipstick (Bovine Enterichek, Biovet Inc.) was made available to bovine practitioners and producers for the rapid detection of Betacoronavirus 1 (BCV-1), Rotavirus A (RV-A), Escherichia coli K99(+), and Cryptosporidium parvum in feces from diarrheic calves. The diagnostic performance of Bovine Enterichek was evaluated in comparison with a multiplex real-time polymerase chain reaction assay (mrtPCR). One hundred fecal samples were procured from diagnostic submissions to Iowa State University Veterinary Diagnostic Laboratory and were used for the assessment. The agreement quotient (kappa) in results for each pathogen between Bovine Enterichek and mrtPCR were 0.095 (BCV-1), 0.521 (RV-A), 0.823 (E. coli K99( + )), and 0.840 (C. parvum). In comparison to mrtPCR, the diagnostic sensitivity of Bovine Enterichek was 60.0%, 42.3%, 71.4%, and 81.5%, and the diagnostic specificity was 51.4%, 100%, 100%, and 98.6% for BCV-1, RV-A, E. coli K99( + ), and C. parvum, respectively. The current study suggested that Bovine Enterichek can be a rapid test tool in the field for detection of RV-A, C. parvum, or E. coli K99(+) in feces from calves at acute stage of clinical disease. However, test results for BCV-1 by the kit should be interpreted with caution due to low specificity and sensitivity of the kit.

Naturally occurring Influenza A virus subtype H1N2 infection in a Midwest United States mink (Mustela vison) ranch

Influenza A virus (FLUAV) causes acute respiratory disease in humans and a variety of animal species. The virus tends to remain within the species of origin; nonetheless, naturally occurring cross-species transmission of FLUAV has been periodically documented. Multiple cross-species transmissions of FLUAV have been reported from companion animals and captive wild animals, neither of which is historically considered as natural hosts of FLUAV. In the fall of 2010, mink (Mustela vison) inhabiting a 15,000-head mink farm in the Midwest United States experienced persistent severe respiratory distress and nose and/or mouth bleeding. Mink losses averaged approximately 10 animals per day. Six dead mink at 6 months of age were submitted to the Iowa State University Veterinary Diagnostic Laboratory for diagnostic investigation. Gross and microscopic examinations revealed that all 6 mink had hemorrhagic bronchointerstitial pneumonia. Hemolytic Escherichia coli was isolated from lungs, probably accounting for hemorrhagic pneumonia. All animals tested negative for Canine distemper virus and Aleutian mink disease virus. Interestingly, FLUAV of H1N2 subtype, which contained the matrix gene of swine lineage, was detected in the lungs. Serological follow-up on mink that remained in the ranch until pelting also confirmed that the ranch had been exposed to FLUAV of H1 subtype (δ clade). The case study suggests that FLUAV should be included in the differential diagnosis when mink experience epidemics of respiratory disease. Since the source of FLUAV appeared to be uncooked turkey meat, feeding animals fully cooked ration should be considered as a preventive measure.

Evaluating the effects of pelleting, corn dried distillers grains with solubles source, and supplementing sodium metabisulfite in nursery pig diets contaminated with deoxynivalenol (2011)

A total of 360 barrows (PIC 1050, initially 24.7 lb ± 0.3 lb BW and 35 d of age) were used in a 28-d trial examining the effects of pelleting, pelleting dried distillers grains with solubles (DDGS), and supplementing sodium metabisulfite4 (SMB) in diets containing deoxynivalenol (DON) on nursery pig performance. Pigs were allotted to 1 of 10 treatments with 7 replications per treatment (pens) and 5 pigs per pen. Naturally contaminated DDGS were used to incorporate DON at desired concentrations. Ingredients were tested for mycotoxins by the North Dakota State University Veterinary Diagnostic Laboratory (NDSU; Fargo, ND) and served as the basis for diet formulation. The 5 experimental diets were fed in meal and pellet form: (1) positive control, (2) negative control (NC, 5.3 ppm DON), (3) NC with 0.5% SMB, (4) pelleted and reground DDGS (5.3 ppm DON), and (5) pelleted and reground DDGS with 2.5% SMB (final diet contained 0.5% SMB). Experimental diets were fed from d 0 to 21 with a common diet fed from d 21 to 28 to evaluate performance after DON was removed. Due to the variability of DON assays when levels exceed 8 ppm, final diets were lower in DON than predicted from analysis of the DDGS. As a result, expected reductions in performance due to DON were not as significant as anticipated, and may have affected results. From d 0 to 21, pigs fed diets with high-DON levels had decreased (P < 0.03) ADG, but the reduction in ADG was only 4%. Pelleting high-DON diets decreased (P < 0.04) ADFI and improved (P < 0.02) F/G compared with diets fed in meal form; however, pelleting DDGS prior to manufacturing final diets had no effect on growth performance. Supplementing SMB tended (P < 0.08) to decrease ADFI, and had no effect on ADG or F/G.

Evaluating the effects of pelleting Deoxynivalenol-contaminated dried distillers grains with solubles in the presence of sodium metabisulfite on analyzed DON levels (2011)

Deoxynivalenol (DON), also known as vomitoxin, was prevalent in the 2009 U.S. corn crop and subsequently present in dried distillers grains with solubles (DDGS), in which DON levels are about 3 times higher than the original corn source. One method shown to reduce DON levels was by increasing moisture and temperature when sodium bisulfite was added to DON-contaminated corn (Young et al., 19874). Therefore, a pilot study aimed first to replicate these results by placing DON-contaminated DDGS in an autoclave (60 min at 250°F) in the presence of sodium metabisulfite (SMB). The study used 6 treatments: (1) control, (2) 0.5% SMB, (3) 1.0% SMB, (4) 2.5% SMB, (5) 5.0% SMB, and (6) 5.0% SMB with 100 mL/kg water added to evaluate the role of water. After drying, samples were analyzed at North Dakota State University Veterinary Diagnostic Laboratory (NDSU; Fargo, ND). Autoclaving reduced DON levels (R2 = 0.99) with increasing SMB, justifying a follow-up study that aimed to assess whether SMB has the same detoxifying effects on corn DDGS in a commercial pellet mill.

A matrix gene–based multiplex real‐time RT‐PCR for detection and differentiation of 2009 pandemic H1N1 and other influenza A viruses in North America

The emergence in humans of pandemic (H1N1) 2009 (pH1N1) with similarities to swine influenza viruses (SIVs) caused much concern for both human and animal health as potential for interspecies transmission was initially unknown. The goal of this study was to develop a real-time RT-PCR test for the detection and differentiation of 2009 pH1N1 and endemic influenza A viruses in North America. Matrix (M) gene sequences from U.S. human pH1N1 cases and U.S. SIVs were aligned to determine a suitable region for an assay target. Primers were selected to amplify all influenza A. Two probes were designed to differentiate pH1N1 (EA matrix) from endemic (NA matrix) SIVs. The assay was validated using the first U.S. pH1N1 strain, 10 human pH1N1-positive specimens and nine U.S. SIV isolates, then evaluated on 165 specimens of swine and other animal origin submitted to the Iowa State University Veterinary Diagnostic Laboratory. Results were compared to other influenza A PCR assays. Sequences from additional pH1N1 strains and contemporary H1N1 SIVs were used to assess robustness of the selected primers and probes for the intended purpose. The new assay's results from clinical specimens concurred with confirmatory PCR testing. The additional probe designed from sequence analysis improved detection of the NA matrix subtype when added to the reaction mixture. This assay detects and differentiates pH1N1 and US influenza A viruses in various sample matrices and species. Good bioinformatics support is critical when designing RT-PCR assays and monitoring their performance.