Browsing by Subject "Veterinary Medicine"

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69th Minnesota Nutrition Conference, University of Minnesota Research Update and Symposium: Role of Immune Tissue in Fighting Pathogen Invasion.Proceedings. Front Matter.
(University of Minnesota, Minnesota Extension Service, 2008-09) University of Minnesota, Extension Service; University of Minnesota, Dept. of Animal Science
Assessment of the demographics and network structure of swine populations in relation to regional disease transmission and contro.
(2011-06) Wayne, Spencer R
Commercial swine production has steadily evolved the into interconnected multi-site production systems of today. As a result, large numbers of growing pigs and breeding animals move from one location to another on a daily basis. The health of the national swine herd has improved dramatically, due in large part to this new production structure; but the increased network size and the long distances travelled pose obvious threats to swine health. As animal agriculture has become more sophisticated, our government resources have not kept up. Available datasets are inaccurate, fragmented, and offer limited definition of the population at risk and its nature. National efforts to improve livestock population data have met considerable public resistance, and as a result, progress has been limited. Knowledge of the populations at risk is of primary importance when trying to define the potential for disease to spread within and between these populations. Disease spreads by non-mechanical means (as in aerosol transmission of PRRS virus) potentiates the need for knowledge of the neighborhood. Given the dynamic and transient nature of our swine populations, the neighborhood's health status is constantly challenged by the most recent delivery of pigs into the neighborhood. The following dissertation seeks to expand the knowledge of swine populations. Current geographic datasets were assessed for accuracy and reliability. In the event of a foreign animal disease outbreak the usefulness of these datasets would be of prime importance, as they will dictate the distribution of resources. Additionally, the use of satellite-derived thermal imagery to verify the presence of commercial swine is described, along with its estimated sensitivity and specificity. Any regional disease elimination program must consider all swine populations, therefore non-commercial populations (specifically, 4H exhibition pigs) are analyzed. Population size, seasonality, caretaker knowledge, presumed and measured health status, and relationship with commercial swine are defined. The physical movement of infected animals across the landscape allows rapid spread of a pathogen to occur. Volume, frequency, and geographic scale of movements will dictate how quickly and thoroughly an epidemic will proceed. For this reason, these are defined and displayed for pig producing areas at different scales.
Assessment of the demographics and network structure of swine populations in relation to regional disease transmission and control.
(2011-06) Wayne, Spencer R.
Commercial swine production has steadily evolved the into interconnected multi-site production systems of today. As a result, large numbers of growing pigs and breeding animals move from one location to another on a daily basis. The health of the national swine herd has improved dramatically, due in large part to this new production structure; but the increased network size and the long distances travelled pose obvious threats to swine health. As animal agriculture has become more sophisticated, our government resources have not kept up. Available datasets are inaccurate, fragmented, and offer limited definition of the population at risk and its nature. National efforts to improve livestock population data have met considerable public resistance, and as a result, progress has been limited. Knowledge of the populations at risk is of primary importance when trying to define the potential for disease to spread within and between these populations. Disease spreads by non-mechanical means (as in aerosol transmission of PRRS virus) potentiates the need for knowledge of the neighborhood. Given the dynamic and transient nature of our swine populations, the neighborhood's health status is constantly challenged by the most recent delivery of pigs into the neighborhood. The following dissertation seeks to expand the knowledge of swine populations. Current geographic datasets were assessed for accuracy and reliability. In the event of a foreign animal disease outbreak the usefulness of these datasets would be of prime importance, as they will dictate the distribution of resources. Additionally, the use of satellite-derived thermal imagery to verify the presence of commercial swine is described, along with its estimated sensitivity and specificity. Any regional disease elimination program must consider all swine populations, therefore non-commercial populations (specifically, 4H exhibition pigs) are analyzed. Population size, seasonality, caretaker knowledge, presumed and measured health status, and relationship with commercial swine are defined. The physical movement of infected animals across the landscape allows rapid spread of a pathogen to occur. Volume, frequency, and geographic scale of movements will dictate how quickly and thoroughly an epidemic will proceed. For this reason, these are defined and displayed for pig producing areas at different scales.
Cellulitis in turkeys: characterization of causative agents and preventive measures.
(2011-08) Thachil, Anil Johny
Cellulitis continues to cause extensive losses in turkey production in USA due to severe mortality, carcass condemnation and treatment costs. Clostridium perfringens and Clostridium septicum have been recognized as the primary causative agents of cellulitis in turkeys. In this study, cellulitis lesions and mortality in turkeys were successfully reproduced with Clostridium perfringens and Clostridium septicum isolated from cellulitis cases. Clostridium perfringens and Clostridium septicum isolates varied in their ability to produce spores as well as toxins. We observed differences in the toxicity and biological effects of different strains of C. perfringens and C. septicum in vitro, and in vivo. .Though the spore count and hemolytic effects of C. perfringens were found to be higher than C. septicum in vitro, mortality studies in mice and turkeys showed that C. septicum was much more potent than C. perfringens. However, gross lesions produced by C. perfringens and C. septicum were almost identical. Surprisingly, the development of cellulitis lesions and mortality was markedly higher in 7-week-old birds than in 3-week-old birds. The results of our study demonstrated for the first time that both C. perfringens and C. septicum can multiply in the subcutaneous and muscle tissues and cause cellulitis lesions in turkeys. Our cellulitis disease model offers promise as a challenge model in the development of vaccines against cellulitis in turkeys. Both bivalent C. perfringens and C. septicum toxoid and C. septicum toxoid were found to be safe and offered complete protection against cellulitis following homologous challenge under experimental conditions. The use of these vaccines enabled us to reduce the mortality and antibiotic usage in preventing cellulitis in commercial turkeys. Multiple vaccinations or use of a day old vaccine followed by a booster dose probably will offer better protection than a single vaccination at 6-weeks of age against cellulitis due to C. perfringens and C. septicum in turkeys.
Characterization of the multidomain Nsp2 Replicase protein of porcine reproductive and respiratory syndrome virus.
(2008-12) Han, Jun
This dissertation focused on understanding the biology of the nonstructural protein 2 (nsp2) of porcine reproductive and respiratory syndrome virus (PRRSV), the etiological agent of porcine reproductive and respiratory syndrome (PRRS). PRRSV nsp2 is a multidomain protein, containing a putative N-terminal cysteine protease PL2 domain, a 500-700aa middle region of unknown function, a transmembrane domain and a C-terminal tail with uncertainty. In this dissertation, we report the following. (i) PRRSV nsp2 is undergoing rapid evolution in field strains exemplified by viral isolates MN184A and B. (ii) We showed that PRRSV nsp2 hypervariable regions aa12-35 and aa324-813 were not essential for viral replication in MARC-145 cells by using a reverse genetics system based on strain VR-2332. In contrast, deletion of the cysteine protease PL2 core domain, the PL2 downstream flanking sequence (aa181-323), the predicted transmembrane domain and the C-terminal domain were lethal to the virus. (iii) We provided evidence that the nsp2 protein encodes an active PL2 protease and mediates nsp2/3 processing in CHO cells with a substrate preference for the dipeptide G1196|G1197. The PL2 protease possessed both trans- and cis-cleavage activities, which could be distinguished by point mutations. Site-directed mutagenesis studies revealed that mutations that caused a specific loss of trans function of the PL2 protease, but not cis activity, were detrimental to the virus. In addition, we showed that the conserved aspartic acid residues (e.g., Asp89) played an important role in the PL2 trans-cleavage activity. (iv) We investigated the proteolytic processing of nsp2 in MARC-145 cells using recombinant PRRSV expressing foreign epitope-tagged nsp2 protein. We showed the presence of the nsp2 protein as different isoforms in PRRSV-infected cells, which appeared to share the same N terminus but differed in their respective C-termini. The nsp2 species emerged almost simultaneously in the early stage of PRRSV infection, were stable and had low turnover rates. Deletion mutagenesis suggested that the smaller nsp2 species (e.g. nsp2d, e and f) were not essential for viral replication in cell culture. Lastly, a cellular protein, heat shock 70kDa protein 5 (HSPA5), was identified as a coimmunoprecipitate of nsp2.
Compensatory force plate responses to single or multiple limb lameness induction in horses using a hoof clamp technique.
(2011-12) Swaab, Megan E.
Reasons for Performing Study: Previous equine research has combined subjective analysis with kinematic or kinetic data to describe lameness, but has only shown evidence of compensatory mechanisms for only single forelimb and hindlimb lameness, not multiple limb lameness. Objectives: To create a consistent, controlled and immediately reversible lameness using a circumferential hoof clamp technique and measure the resultant changes in ground reaction forces. To compare the changes in ground reaction forces in both the limb(s) in which lameness was induced and in the sound limbs among a variety of individual and multiple-limb lameness scenarios to detect possible patterns in lameness and compensation in the sound limbs. Materials and Methods: Lameness was induced in 8 horses by tightening a circumferential hoof clamp, on an individual forelimb, or hindlimb, an ipsilaterally paired forelimb/hindlimb, a contralaterally paired forelimb/hindlimb, and bilateral forelimbs and hindlimbs. Kinetic analysis was performed with a fore plate prior to (baseline) and after lameness induction. The percent change in ground reaction forces (vertical and longitudinal) from baseline were calculated for all limbs for each lameness scenario. Changes were examined within each of four ground reaction forces (peak vertical force, vertical impulse, breaking impulse and propulsion impulse) to determine whether consistent patterns emerged in the lame limbs or the sound limbs that might indicate compensation. The magnitude of percent change from baseline was also compared between lame and sound limbs. Results: Using the circumferential hoof clamp technique, we were able to induce a consistent, controlled, and immediately reversible grade 2 out of 5 lameness in individual and multiple limbs. In general, peak vertical force, vertical impulse, and braking impulse decreased, and propulsion impulse increased in the lame limb(s). The forelimbs tended to decrease most consistently in the peak vertical force and vertical impulse, while hindlimbs tended decrease most consistently in the braking impulse. The majority of compensation seemed to come from the contralateral limb, directly opposite the lame limb (i.e. the sound forelimb compensated for a lame forelimb, and the sound hindlimb compensated for a lame hindlimb). Compensation tended to occur through increased vertical and braking impulses in the sound limb(s). The magnitude of percent change from baseline was small for the majority of forces, but was consistent and was associated with a visible lameness. Conclusions and Potential Relevance: Definite patterns were seen in ground reaction forces in both lameness and compensation. This information may help equine practitioners understand how horses alter their ground reaction forces in response to single and multiple limb lameness by determining primary versus compensatory change, and to clarify some of the complexities of multiple limb lameness.
Control and characterization of influenza A viruses in swine
(2011-07) Detmer, Susan Elisabeth
Between 1958 and 2005, there were 37 human cases of zoonotic swine-origin influenza A virus (IAV) infection reported (Myers et al 2007; Van Reeth 2007). A majority of these infections were with classical swine H1N1 viruses and these 37 cases did not include the Fort Dix cases in 1976 that resulted in 1 death and up to 230 soldiers infected (Myers et al. 2007; Van Reeth 2007). However, a recent report of pig to human transmission was at an Ohio County Fair in 2007 (Vincent et al. 2009b). The sequence analysis of the HA gene segment of the IAVs isolated from the humans and pigs in this case revealed that it was a strain that was currently circulating in the U.S. pig population. The internal genes of this isolate were determined to be of the triple-reassortant swine influenza lineage, including a conserved avian PB2 gene sequence (Vincent et al. 2009b). On June 11, 2009 the first influenza pandemic in 41 years was declared by the World Health Organization. This virus was like no virus previously seen in the human population with gene segments from both Eurasian and North American swine viruses. The 2009 pandemic H1N1 virus was called a "quadruple-reassortant" virus because it is composed of neuraminidase (NA) and matrix (M) gene segments from Eurasian swine influenza viruses combined with triple-reassortant proteins of North American swine influenza viruses (human-origin polymerase B1 (PB1), avian-origin polymerase B2 (PB2) and polymerase A (PA), and classical swine-origin hemagglutinin (HA), nucleoprotein (NP) and non-structural (NS) (Garten et al. 2009; Smith et al. 2009). The evolutionary analysis of the 2009 pandemic H1N1 shows that the generation of this strain was not likely a recent event. In fact, in order to facilitate human-to-human spread, it probably adapted to the human host through secondary reassortments in humans (Ding et al. 2009). However, the original source of this virus has not yet been determined. The emergence of the 2009 pandemic H1N1 virus and scattered reports of human infections with swine-origin isolates underscores the importance of fully understanding the genetic, antigenic, and pathogenic characteristics of influenza A viruses so that we may limit the introduction of novel IAVs to the swine population and monitor for newly emerging and evolving viruses. In order to improve our understanding of IAVs in swine, the goal of this dissertation is to address the ability of genetic characterization to predict variations in virus phenotype, such as viral binding and antigenicity. Understanding the genetic, antigenic and pathogenic features of viruses is important to prevent introduction of human and avian viruses into swine herds and the potential spillback of those viruses to the human population, as wells as preventing the sustained transmission of IAVs within an endemically infected herd. The control of influenza viruses in pig populations continues to be dynamic and complex, and is reliant on a number of factors. Two of these factors include appropriate selection and application of (1) diagnostic tests and (2) vaccines. Routine surveillance for influenza viruses at the farm level, either syndromic or active surveillance, is often accomplished using real time RT-PCR tests on nasal swabs from live pigs and lung tissue samples from post-mortem examinations. Easily collected sample methods, such as oral fluids, could provide additional viruses for characterization of IAVs in swine. Oral fluids have been used extensively for diagnostic tests in human medicine and are now being applied in swine herds for detecting pathogens and antibodies against the pathogens (Prickett et al. 2010). As part of this dissertation, porcine oral fluids were validated as a viable sample collection method for routine RT-PCR and virus isolation tests (chapter 2). Another important control measure for influenza viruses in pigs continues to be vaccines. In order to assure continued efficacy of vaccines against currently circulating strains of virus, vaccine challenge trials are performed. In this dissertation, the efficacy of a commercial vaccine was examined against challenge with a contemporary field isolate (chapter 3). To address the genetic and phenotypic characterization of influenza A viruses from swine, two sets of viruses were selected from the influenza database at the University of Minnesota, Veterinary Diagnostic Laboratory and sequenced. The first set was isolated from a group of endemically infected farms treated by the same veterinarian from 2005 to 2009. The selected viruses were either used to produce autogenous vaccines or they were the epizootic viruses found during outbreaks in the vaccinated herds (chapter 4). The second set of viruses were isolated from nursery pigs in one endemically infected multi-site swine production system (farm M) from 2007 to 2009 and either contained a distinct two amino acid insertion or were presumptive ancestral viruses without the insertion (chapter 5). The viruses from farm M were further characterized along with representative viruses that have been previously studied in vivo using a new technique called virus histochemistry to examine the patterns of virus attachment in the respiratory tract (chapter 6). For the purposes of this dissertation IAVs were classified as virulent increased virulence have some of the following clinical/case presentations: (a) morbidity approaching 90% and mortality approaching 10 percent, (b) sudden, unexpected deaths occurring early in the disease outbreak, (c) gross lesions that are not typical of swine influenza including profuse hemorrhage and/or edema, and (d) sufficient health and production records along with laboratory results that indicate that a highly virulent influenza virus is involved. The characteristics of highly virulent influenza viruses, such as A/swine/KS/77778/2007 H1N1 and A/swine/OH/511445/2007 H1N1, have been previously described in the literature (Ma et al. 2010; Vincent et al. 2009b). This classification was not related to the criteria for classification of avian viruses as having high or low pathogenicity.
Detection, characterization, and control of Bovine Viral Diarrhea Virus in dairy herds.
(2010-09) Schefers, Jeremy Math
Effective Bovine Viral Diarrhea Virus (BVDV) control on dairy farms is multifaceted and includes methods to accurately detect virus, remove BVDV persistently infected (PI) cattle, prevent virus introduction using comprehensive biosecurity plans, and optimize herd immunity through continuous vaccination against BVDV. The work in this thesis takes into consideration the above means to achieve effective BVDV control more specifically by attempting to: 1) determine the herd infection status by screening newborn calves for precolostral BVDV serum antibodies; 2) eradicate BVDV from a large commercial dairy herd through a combination of test and removal procedures and biosecurity measures; 3) characterize BVDV in PI calves from the Upper Midwestern United States by nucleic acid sequencing in order to more fully understand the changes that are occurring in the BVDV genome that may affect detection and elimination protocols, and; 4) implement a quantitative real-time RT-PCR (qRT-PCR) for quantification of BVDV RNA in a variety of clinical samples obtained from PI cattle. Although many tests have been developed to detect BVDV PI cattle, there are few strategies to detect endemic BVDV infections at the herd level, especially in those herds that routinely administer BVDV vaccines. Many BVDV infections result from direct exposure to BVDV PI cattle. The detection and removal of BVDV PI cattle are essential steps towards reducing virus exposure within the herd and are critical components of national BVDV eradication efforts, such as those in Scandinavia. Veterinary diagnosticians and researchers have developed a variety of accurate tests to detect BVDV PI cattle in dairy herds. For example, screening bulk milk from dairy herds for BVDV by RT-PCR is popular due to the ease of sample collection and the large number of animals that can be screened with one sample. A disadvantage of screening bulk milk is that it will not detect PI cattle in the non-lactating herd nor in youngstock. Alternatively, non-vaccinated sentinel calves can be used to detect BVDV PI exposure in youngstock; however, the sensitivity of sentinel calves in large herds with multiple groups of cattle is not known. Chapter 2 of this thesis investigated a novel screening approach to detect BVDV by screening newborn calves for BVDV serum antibodies prior to colostrum feeding. Newborn calves that are seropositive for BVDV antibody prior to colostrum feeding indicate fetal infection during the last two trimesters of gestation. The number of newborn calves seropositive for BVDV serum antibodies at birth is estimated to be greater than the number of PI calves. Because the number of BVDV seropositive calves is greater than the number of BVDV PI calves fewer calves need to be tested by precolostral serum antibody screening to detect BVDV fetal infections and the probability of one or more PI cattle in the pregnant herd is likely. In addition to requiring fewer test animals, precolostral screening detects infections in lactating, non-lactating, and pregnant youngstock populations and is not confounded by vaccination. Rapid consolidation of the United States dairy industry has resulted in fewer and larger dairy herds. Chapter 3 of this thesis describes the elimination of BVDV PI animals in a large commercial dairy herd with a RT-PCR test. Previous testing in the study herd indicated that approximately 5% of the calves were born with BVDV precolostral serum antibodies. The birth of BVDV seropositive calves also roughly coincided with an increase in post-partum diseases that failed to respond to proven therapies. The herd owners elected to test all animals for BVDV PI with a serum BVDV RT-PCR test. Accurate detection of BVDV PI cattle is important in all herds, but less than perfect sensitivity and the potential of a false negative result are amplified in large herds with PI cattle. False negative test results would lead to the retention of one or more PI cattle and ultimately the continued persistence of BVDV within the herd. Serum samples from all cattle on the premises, and heifer calves born during the following 9 months, were tested for BVDV by RT-PCR and those determined to be BVDV PI on confirmatory tests were removed from the herd. Whether or not BVDV persisted in, or was eliminated from, this herd was determined by monitoring newborn calf precolostral serum antibodies for BVDV one year after the test and removal of all PI cattle. The chapter describes the detection of BVDV PI cattle, genetic characterization of BVDV isolated from the PI cattle, the detection of BVDV acute infections, and the precolostral monitoring results before and after the removal of PI cattle. Bovine viral diarrhea virus is a single-stranded RNA virus that lacks a proof-reading mechanism resulting in mutations and recombination of the viral genome. Point mutations and recombination of viral RNA can result in novel, unique viruses. Few animal disease laboratories perform nucleic acid sequencing for BVDV, thus changes in the BVDV genome are not well described. The objective of chapter 4 was to successfully sequence a portion of the viral RNA and compare the viral genome sequences of forty PI cattle detected on dairy farms in the Upper Midwestern United States. The 5' untranslated region (5'UTR) region of the BVDV genome contains conserved regions and is commonly used for PCR detection tests. This project described the use of primers targeting 5'UTR that produce a PCR product for nucleic acid sequence comparisons between vaccine and field strains and allow for differentiation between subgenotypes BVDV 1a, BVDV 2a, and BVDV 1b. Testing many animals for BVDV PI requires appreciable amount of supplies and labor. The ear notch (skin) sample is a convenient tissue for testing and detecting BVDV PI animals because it is an easy sample to collect and requires minimal amounts of supplies and equipment. Ear notch skin samples offer some flexibility because they can be tested for BVDV by immunohistochemistry (IHC), antigen-capture ELISA (ACE), or RT-PCR. Pooling ear notch phosphate buffered saline (PBS) supernatant for RT-PCR is a popular method to screen large numbers of animals at a reduced cost. This method involves soaking the ear notch in a small amount (~2 ml) of PBS and then pooling the supernatant. The pooled supernatant is then tested for BVDV by RT-PCR. If the pooled supernatant is positive, the originally submitted samples can be tested individually to determine the PI animal. While ear notches have become the sample of choice for PI testing, there is little information available on the quantity of viral RNA in ear notches and the PBS supernatant that contains the soaking ear notch. The objective outlined in Chapter 6 was to implement a quantitative real-time RT-PCR (qRT-PCR) for quantification of BVDV RNA in a variety of clinical samples obtained from PI cattle. Serum, whole blood, nasal swabs and skin samples were collected from PI cattle and analyzed by qRT-PCR. The data derived from qRT -PCR allowed for an estimation of RNA copies in the variety of samples obtained from PI calves. This thesis will give bovine veterinarians, diagnosticians, and researchers additional information on the dynamics and manifestations of BVDV in dairy herds. The precolostral screening method of newborn calves appears feasible and has potential application in commercial dairy herds. The performance and utility of a highly sensitive and specific RT-PCR test was assessed and appeared successful in a large commercial dairy herd. Additionally, nucleic acid sequence analysis of BVDV obtained from PI dairy cattle were compared to PI cattle from other farms and well-described viruses strains listed in GenBank. Quantification of BVDV RNA in clinical samples provided essential information needed to estimate the size of pools and potential variations in detectable RNA from diagnostics samples.
Effects of cigarette smoke condensate on microrna expression of human bronchial epithelial cells.
(2010-10) Endalew, Abaineh Dagne
Lung cancer is the leading cause of cancer death and accounts for 1.2 million deaths annually world-wide and its five year survival is less than 15%. The primary etiology of lung cancer is genetic and epigenetic alterations caused by tobacco smoke. Although smoking-related genetic changes have been well studied, much is not known about the association between lung cancer and epigenetic changes, in particular deregulation of microRNAs. Therefore, our objective in this study was to examine the association between early phenotypic changes and altered microRNA expression in cigarette smoke condensate (CSC)-treated immortalized human bronchial epithelial cells (HBECs). We hypothesized that extended treatment of HBECs with CSC causes morphological and growth alterations and these effects are mediated, at least partly, through deregulation of microRNA expression. The aims of this study were to: 1. Determine CSCinduced early phenotypic alterations in HBECs; 2. Identify microRNAs whose levels are altered in CSC-treated HBECs. HBECs were exposed to least-toxic dose of CSC (5 μg/ml) for 4 months and effects on cell proliferation, morphology, transformation, activation of AKT and ERK, and microRNA expression profile were determined. CSC exposure caused HBECs to become round and elongated and their proliferation is increased by 64%. Western blot analysis also indicated activation of ERK1/2 and AKT. Microarray analysis revealed changes in the expression of 89 microRNAs in CSC-exposed HBECs and 87 of them were down-regulated. Further qRT-PCR analysis revealed altered expression of miR-138, miR-921, miR-293-3p, & IVGN-novel-miR-3526. However only the change in IVGN-novel-miR-3526 was statistically significant (up-regulated by 2.4-fold) (P= 0.03). The expression level of IVGN-novel-miR-3526 in A549 cells found to be 4.6-fold higher iv than untreated HBECs. To our knowledge, this is the first report on the altered level of this microRNA in CSC-treated bronchial cells or lung cancer cells. Overall, this study has provided useful insight on CSC-induced phenotypic alterations and microRNA deregulation in HBECs. Functional assays are required to determine the association between IVGN-novel-miR-3526 deregulation and observed phenotypic changes in CSC-exposed HBECs.
Effects of electroacupuncture in a mouse model of experimentally-induced osteosarcoma.
(2009-12) Al-Gizawiy, Mona Maria
Osteosarcoma (OSA) is a devastating form of musculoskeletal cancer that most commonly results in death due to pulmonary metastatic disease. The anti-inflammatory, immune-boosting, and analgesic effects of electroacupuncture (EA) are well-documented in a variety of animal models. To date, there are no studies investigating the gender effects of EA on OSA pain and tumor growth. We studied the effects of EA in a mouse model of experimentally-induced OSA. Electroacupuncture (4 Hz) was applied to the Zusanli (ST-36) acupuncture point at different time intervals. Each group was accompanied by a sham treatment group (no current). Primary hyperalgesia was evaluated using von Frey filaments and by quantification of spinal cfos expression. Tumor size was measured using calipers. Vaginal swabs were carried out in female mice to determine the stage of the estrous cycle during treatments. Estrous cycles in females varied greatly and were not synchronized within groups. They showed no correlation to EA or behavioral testing, indicating no direct hormonal influence. Hyperalgesia consistently increased with tumor growth, although less so in mice receiving early EA treatments. Hyperalgesia dropped slightly in all mice on the days EA was performed, but rose again 24 hours later. Tumors tended to grow slightly larger in males and resulted in higher von Frey scores and cfos expression across the groups. With few exceptions, however, there were no significant gender differences in tumor growth or von Frey scores. We conclude that early EA treatment has inhibitory effects on nociception and tumor growth that are not influenced by gender.
Efficacy of interventions and role of raw colostrum feeding programs in the transmission of Mycobacterium avium subsp. paratuberculosis.
(2009-06) Pithua, Patrick
This thesis described the role of raw bovine colostrum feeding programs and natural nursing practices in the transmission of Mycobacterium avium subsp. paratuberculosis (MAP), and the efficacy of commercially available colostrum replacement products in preventing MAP transmission, additional to their effect on production and longevity performance outcomes. Incidences of fecal excretion of MAP by calves following natural exposure were also evaluated. Calves fed CR (vs MC) had a lower risk of MAP infection when the serum ELISA (HR = 0.474, P = 0.081), bacterial fecal culture (HR = 0.572, P = 0.076) or both test combinations (HR = 0.559, P = 0.056) were used to define MAP status of study cohorts, suggesting that MC could be an important vehicle by which calves become exposed to MAP within hours following birth, and that CR feeding programs may be an effective management tool for use in dairy herds in a Johne’s disease control effort. From birth-to-54 months of follow-up, risk of death (HR = 1.22, P = 0.17), culling (HR =1.01, P = 0.95), and death and/or culling (HR = 1.1, P = 0.61) event outcomes did not significantly differ between groups (CR vs MC). Similarly there were no significant differences between groups (CR vs MC) with respect to the risk of death (HR = 1.22, P = 0.46), culling (HR =1.01, P = 0.98), and death and/or culling (HR = 1.05, P = 0.85) event outcomes when only heifers that entered the lactating herd (period from first calving date to 54 months of age) were considered. Feeding CR (vs MC) had no significant effect on age at first calving (P = 0.34), number of breedings per conception in the first (P = 0.83) and second (P = 0.32) lactations respectively, and calving-to-conception intervals in the first (P = 0.7) and second (P = 0.21) lactations, respectively. Considering the milk yield outcome, feeding CR (vs MC) significantly (P = 0.02) decreased first lactation milk by 429 kg, although there were no significant effects of feeding CR (vs MC) on second lactation (P = 0.18) and lifetime milk yields (P = 0.5), respectively. Risk of MAP infection was not significantly different between groups of calves that ingested MAP DNA positive colostrum (vs MAP DNA negative colostrum) when the serum ELISA (HR = 0.74, P = 0.65), bacterial fecal culture (HR = 0.92, P = 0.85) or both test combinations (HR = 0.82, P = 0.65) were used to define MAP status of study (HR = 0.74, P = 0.65), bacterial fecal culture (HR = 0.92, P = 0.85) or both test combinations (HR = 0.82, P = 0.65) were used to define MAP status of study cohorts, suggesting lack of an added risk of MAP infection associated with ingesting MAP DNA positive raw colostrum by Holstein calves. This finding contradicted several other reports which seem to provide evidence in support of colostrum as a possible early vehicle by which calves get exposed to MAP in infected herds. Cows that were fecal culture positive were significantly more likely to have detectable MAP in their colostrum (OR =2.02 , P < 0.001) and teat skin (OR =1.87 , P = 0.008) compared with fecal culture negative cows with the population attributable fraction estimates for exposure for each of the latter outcomes being 18% and 19.5%, respectively. Finally, MAP was not recovered from fecal samples collected between 1-to-90 d of age and tested using the sedimentation bacterial culture method suggesting that the calves studied did not excrete detectable levels of MAP in feces following natural exposure.
Efficacy of on-farm programs for the diagnosis and selective treatment of clinical and subclinical mastitis in dairy cattle.
(2009-08) Lago Vázquez, José Alfonso
The research reported in this dissertation includes two multi-state multi-herd clinical trials evaluating the efficacy of on-farm programs for the diagnosis and selective treatment of clinical and subclinical mastitis in dairy cattle. The use of an OFC system for the selective treatment of clinical mastitis during lactation reduced intramammary antibiotic use by half and tended to reduce withholding time by one day, without significant differences in days to clinical cure, bacteriological cure risk, new infection risk and ICR risk (where the ICR risk represented the presence of infection risk, clinical mastitis risk, or removal from herd risk) within 21 days after the clinical mastitis event. Similarly, there were no differences between both treatment programs in long-term outcomes such as recurrence of clinical mastitis in the same quarter, somatic cell count, milk production, and cow survival for the rest of the lactation after the clinical mastitis event. The treatment with intramammary Cephapirin Sodium of cows and quarters based on CMT results alone, or sequential testing using OFC to diagnose Gram-positives in CMT-positive quarters resulted in a higher bacteriological cure risk and reduced the ICR risk within 21 days after enrollment (significantly and only numerically, respectively for treatment each program). The implementation of both treatment programs required the administration of intramammary treatment and extended the time that milk is withhold from the market. Both programs resulted in a significantly lower clinical mastitis risk and lower milk SCC during lactation (significantly and only numerically, respectively for each treatment program). However, the implementation of both treatment programs did not result in higher milk production, improved reproductive performance or lower risk for removal from the herd. A secondary objective of both clinical trials was to validate the use of the Minnesota Easy Culture Bi-Plate System. This OFC system is a useful cow-side test to correctly identify bacterial growth, Gram-positive bacterial growth, or Gram-negative bacterial growth in quarter secretion samples from clinical mastitis cases and in CMT-positive quarter milk samples collected after parturition. Treatment decisions based on identification of bacterial growth, or Gram-positive bacterial growth specifically, were correct over 73% of the time.
Epidemiology of lameness in breeding female pigs.
(2011-03) Sukumaran Nair, Santhakumari Anil
A low level of sow retention in the herd is a cause for both economic as well as welfare concerns. The results of the study confirmed that a low lactation feed intake, incidence of lameness or health problems, as well as sow-level characteristics such as higher parity and fewer piglets born alive per litter may adversely affect sow longevity. Sows retained with periparturient health problems had reduced longevity and fewer live-born piglets, and fewer such sows had another farrowing. A prospective data analysis indicated that the overall performance of lame sows in terms of the number of pigs born alive during the period of the study was less, compared with that for non-lame sows. Retaining sows with less severe lameness may enable the producer to meet immediate production targets. The findings suggest that sow removal decisions should be judiciously evaluated after farrowing considering the potential long-term losses. Lameness in swine herds should be minimized and if treatment is not an option lame sows should be culled as soon as possible to reduce long-term losses. The results also confirmed the high prevalence of claw lesions in breeding female pigs and their association with lameness, specifically, white line and side wall lesions. The results indicate the possibility of nutritional intervention in minimizing claw lesions. However, there are other factors associated with claw lesion development in pigs. The quality of the floor as well as different bio-mechanical factors operating in lesion development are important here. The space between slats, roughness of the surface, and edge design are critical in claw lesion development. Those factors have not been addressed in this study. Further studies are required to understand the mechanism of lesion development in relation to the housing and management systems in place. This information is vital in formulating the appropriate intervention strategy to minimize the incidence of lameness and to improve sow longevity and performance. The studies in this thesis included data from single herds and therefore the generalization of the results may be restricted owing to the wide variations in management, housing and in genetic lines of sows.
Epidemiology of Mycobacterium avium subsp. paratuberculosis fecal shedding in Johne's disease infected dairy herds.
(2012-02) Espejo, L. A.
Johnes’s disease, also known as paratuberculosis, is a chronic enteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). The disease has a slowly progressing detrimental effect on cow health and production. In the United States most dairy cattle farms are infected, causing an economic impact to dairy herds in the short term by the association of the disease with low milk production, higher culling rates and low reproductive performance. Control of the disease has been focused on implementation of management practices that reduce the MAP transmission from infected cows to uninfected young calves, and on culling cattle that shed large amount of MAP in feces (heavy fecal shedding cows) as soon as detected. The objectives of these studies were to evaluate the association between use of recommended management practices on Johne’s disease incidence and to improve the understanding of the most commonly used diagnostic tests to identify heavy fecal shedders. The evaluation of the association between use of a standardized control program on the incidence of Johne’s disease was conducted in a prospective longitudinal observational study that in 8 dairy herds in Minnesota. Herds were followed during a period of 5 to 10 years. We found a reduction of the incidence of bacterial culture positivity, serum ELISA positivity, heavy fecal shedding status, and clinical Johne’s disease associated with higher levels of implementation of the recommended management practices. The evaluation of the analytical sensitivity of bacterial culture of feces and direct fecal PCR was performed in two separate experiments using MAP negative bovine fecal samples spiked with different concentrations of MAP. The analytical sensitivity of the bacterial culture of feces was 105 MAP/g of feces and the probability of a higher bacterial culture result increased with the concentration of MAP in the fecal sample. The analytical sensitivities of the direct fecal PCR in experiments 1 and 2 using different approaches were 107 and 102 MAP/g of feces, respectively. A latent class model using a Bayesian approach was fitted to estimate the posterior conditional probabilities that the results of the bacterial culture of feces and serum ELISA correctly identified cows as high positive, low positive or negative given that they were heavy, light and non-fecal shedders, respectively. The estimated conditional probabilities that bacterial culture of feces correctly identified heavy, light and non-fecal shedders were 70.8, 32.2 and 98.5%, respectively. The same values for the serum ELISA were 60.5, 18.8 and 99.5, respectively. Finally, we conducted a cross-sectional study to evaluate the association between bacterial culture of cow-level and pooled environmental fecal sample results for detection of MAP in dairy herds. The sensitivity and specificity of the parallel interpretation of bacterial cultures of pooled environmental fecal samples from the herd to detect at least one heavy fecal shedding cow in the herd was 98.2% and 43.5%, respectively. The sensitivity and specificity of the bacterial culture on pooled individual samples to detect at least one heavy fecal shedding cow in the pool was 100% and 91%, respectively, and these values did not change when pool size increased from 5 to 10 cows per pool. In summary, these studies shown that implementation of critical management practices are associated with a reduction on the incidence of Johne’s disease and diagnostic tests can be used to indentify heavy shedding cows using individual or pooled fecal samples.
Epidemiology of struvite urolithiasis in ferrets.
(2010-08) Nwaokorie, Eugene Emeka E.
Objective--To test the hypothesis that the predominant mineral type in naturally occurring ferret uroliths was sterile struvite and to determine if age, breed, gender, reproductive status, geographic location of ferrets, season of the year of uroliths submission, and anatomic location within the urinary tract were risk factors associated with ferret sterile struvite uroliths formation. Design-- Case-control retrospective study Animals--408 case ferrets from the Minnesota Urolith Center (MUC) and 6528 control ferrets from the Veterinary Medical Data Base (VMDB) 1981 through 2007. Procedure-Historical information about age, gender, reproductive status, anatomic location within the urinary tract, and season of the year of urolith submission were obtained about each ferret. The association between these factors and outcome (sterile struvite urolith formation) was statistically assessed. Results--Sterile struvite was the predominant mineral in ferret uroliths. Cystine comprised the next most common type of urolith, followed by calcium oxalate. Neutered male ferrets had increased risk of developing sterile struvite uroliths. A significant association was also found between the ages of 2 years and < 7 years and the detection of struvite uroliths. Also a significant association was found between advancing age and the detection of struvite uroliths. Ferret struvite uroliths were more likely to be retrieved from the lower urinary tract (bladder and urethra, n = 254) than from the upper the urinary tract (kidney and ureter, n = 4) (the location of 14 uroliths was not recorded). vi Conclusion and Clinical Relevance-- Knowledge of predominant mineral type in uroliths along with insight into etiologic, demographic, and environmental risk and protective factors for urolithiasis may facilitate development of surveillance strategies that could result in earlier detection of uroliths in ferrets. Modification of risk factors including dietary risk factors may help to minimize urolith formation, dissolve existing uroliths and minimize urolith recurrence. In context of sterile struvite urolithiasis, ferrets and domestic cats are remarkably similar. Both species are true carnivores. Both species form sterile struvite uroliths. In both species, infection-induced uroliths are uncommon. In both species, bacterial urinary tract infections are apparently uncommon. The advent of safe and effective diet therapy to induce dissolution of sterile struvite uroliths in cats, and the striking parallels between ferret and feline urolithiasis, prompts the question as to the safety and efficacy of diet-induced dissolution and prevention of sterile struvite urolith formation in ferrets. The primary goal of this study was to compare features of sterile struvite in cats with those in ferrets. Results of these comparisons may provide a logical evidence-based rationale for clinical trials to determine the safety and efficacy of diet-induced dissolution of sterile struvite uroliths in ferrets.
Gene and protein expression in canine follicular thyroid carcinoma.
(2011-08) Metivier, Kristy Stacy
The major goal of this study was to investigate the molecular characteristics of canine follicular thyroid carcinoma (FTC). This is a rapidly growing and highly aggressive tumor in dogs, and many patients present with evidence of invasion or metastasis. Some smaller independent studies have attempted to evaluate the role of single molecules such as p53 and thyroid transcription factor-1 in tumor development, often with inconclusive results. In the present study, a genome-wide approach was employed to achieve the first objective of determining the gene expression profile of FTC compared to normal thyroid tissue. Microarray analysis was performed in a pilot study using five FTC tissues and four normal thyroid gland tissues, and this showed 489 transcripts as differentially expressed between the two groups; 242 were down-regulated and 247 were up-regulated. Some important biological functions that were affected include regulation of cell shape, cell adhesion, regulation of MAP kinase activity, angiogenesis, and regulation of cell migration. Osteopontin was a gene of interest as tumors consistently expressed it at high levels while it was expressed at low levels in all of the healthy samples. One of its up-stream regulators, VEGFA, was also differentially expressed but with a smaller fold change. The expression of osteopontin was validated by quantitative PCR using three groups: non-invasive FTC (tumors with capsular invasion only), invasive FTC (tumors with capsular and vascular invasion), and normal thyroid tissue. Both non-invasive FTC and invasive FTC had higher osteopontin gene expression than normal thyroid tissue but the two tumor groups were not different from each other. The second objective was to determine the protein expression of osteopontin and VEGFA in the same cases using semi-quantitative scoring of tissues stained by immunohistochemistry. The results were similar, with non-invasive and invasive FTC having higher osteopontin protein expression than normal thyroid tissue, but showing no difference from each other. With respect to VEGFA, there was no difference in gene or protein expression among the three groups. The final objective was to determine the plasma concentration of VEGFA and osteopontin in dogs diagnosed with FTC compared to clinically healthy dogs using a commercially available canine-specific ELISA. In this case, both VEGFA and osteopontin had higher plasma concentrations in dogs with FTC compared to healthy dogs. A small number of FTC cases were also measured two weeks after surgical removal of the tumor. Some cases showed a post-surgical decrease in VEGFA and osteopontin while others either remained the same or increased; however, the sample size for this comparison was small. The consistent expression of osteopontin in both tissues and blood suggest that it is a promising marker for identification of canine FTC. As in human studies of osteopontin in aggressive carcinomas, it is also possible to investigate it as a means of monitoring response to therapy, recurrence, and clinical outcome.
Growth dynamics of the canine proximal tibial physis
(2010-06) McBrien Jr., Charles S.
Objective- To determine growth of the proximal tibial physis in the Labrador Retriever, a breed of dog at risk for rupture of the cranial cruciate ligament (RCCL). Animals- 6 male Labrador Retriever dogs Methods- 0.5 mm tantalum markers were implanted in the right proximal tibial epiphysis and metaphysis of each dog at sixteen weeks of age. Lateral and cranio-caudal radiographs of the tibia were made monthly and longitudinal growth was assessed from the radiographs. A growth curve was generated from the data. Data from previous patients that had undergone proximal tibial epiphysiodesis (PTE) was compared to the growth curve to demonstrate if the growth curve accurately predicted changes in growth associated with this procedure. Results- Growth rate decreased slowly and non-linearly over the first year of age. Growth from the proximal tibial physis is described. Conclusions- The growth curve generated here follows the model of saltation and stasis. The growth curve generated here predicted the change in tibial plateau angle (TPA) for two Labrador Retrievers that underwent PTE (+/- 1°). Clinical relevance- The growth curve generated in the present study may be considered for use for the surgical planning of PTE in Labrador Retrievers. Key Words- Proximal tibial epiphysiodesis, growth, saltation and stasis, Labrador Retriever
Immunopathogenesis of avian metapneumovirus in the Turkeys
(2009-09) Cha, Ra Mi
Avian metapneumovirus subtype C (aMPV/C) causes a severe upper respiratory tract (URT) disease in turkeys. The disease is characterized by viral replication and extensive lymphoid cell infiltrations in the URT. The identity of infiltrating cells and their possible involvement in the immunopathogenesis of the disease are not known. The role of local mucosal immunity in viral defense has not been examined for aMPV/C. The overall objective of the study was to examine the immunopathogenesis of aMPV/C in ovo and hatched turkeys, with emphasis on the involvement of local mucosal immunity in viral defense. Three specific objectives were pursued. First, the immune cells, especially mucosal T cells that infiltrate the URT of turkeys following aMPV/C exposure were characterized. Two-week-old aMPV/C antibody-free turkeys were inoculated oculonasally (O/N) with live aMPV/C. At 5 and 7 days post inoculation (DPI), lymphoid cells infiltrating the mucosal lining of the turbinates of the virus-exposed and untreated control turkeys were isolated by enzymatic treatment. In the URT, aMPV/C exposure increased the proportion of CD8+ T cells but not of CD4+ T cells. In addition, CD8 gene expression was upregulated after virus exposure whereas CD4 gene expression remained unchanged. At 5 and 7 DPI, aMPV/C-exposed turkeys showed upregulated gene expression of IFN-gamma and IL-10 in the turbinate tissue. These results suggested that aMPV/C modulated local cellular immunity in the URT of turkeys. Secondly, the ability of an adjuvanted inactivated aMPV/C (Ad-iaMPV/C) inoculated by the respiratory route to induce protective mucosal immunity in the URT was examined. aMPV/C antibody-free turkeys were inoculated via the O/N route with inactivated virus adjuvanted with synthetic double-stranded RNA polyriboinosinic polyribocytidylic acid (Poly IC). Ad-iaMPV/C immunized turkeys showed an increased number of mucosal IgA+ cells in the URT and increased levels of virus-specific IgG and IgA in the lachrymal fluid and serum. After 7 or 21 days post immunization, turkeys were challenged with pathogenic aMPV/C via the O/N route. Turkeys immunized with Ad-iaMPV/C were protected against microscopic lesions and the replication of the challenge virus in the URT. These observations revealed that inactivated aMPV/C administered by the respiratory route induced protective immunity against challenge with the pathogenic virus. As the last objective, we studied the immunopathogensis and protective immunity of aMPV/C in turkeys following in ovo exposure. aMPV/C was inoculated into commercial aMPV/C antibody-free turkey eggs via the amniotic route at embyronation day (ED) 24. Hatchability of eggs was not affected by the virus inoculation. At the day of hatch (ED 28) (4DPI) and 5 days post hatch (9DPI), the virus genome was detected by qRT-PCR in the turbinate, trachea and lung but not in the thymus or the spleen. Turbinate mucosa had mild lymphoid cell infiltration, and there were no detectable lesions in the lung. Spleen cells and thymus cells from virus-exposed turkeys responded poorly to T cell mitogens. In addition, IFN-gamma and IL-10 gene expression was increased in the turbinate tissue of virus-exposed turkeys. In ovo virus exposure increased the levels of aMPV/C-specific IgG in the serum and the lachrymal fluid. At 3 weeks of age, the in ovo immunized turkeys were protected against a challenge with pathogenic aMPV/C. These data indicated that in ovo vaccination may be used in turkeys to control aMPV/C.
Injectable levetiracetam use in the dog.
(2012-03) Hardy, Brian Thomas
Abstract summary not available
Lawsonia intracellularis disease control and epidemiology.
(2009-09) Wattanaphansak, Suphot
Proliferative enteropathy (PE) is an important enteric disease in grower and finisher pigs caused by an obligate intracellular bacterium, Lawsonia intracellularis. The overall aims of this thesis were to obtain more information about the in vitro activity of antimicrobials and disinfectants against L. intracellularis and to develop a new quantitative PCR assay. To determine the minimum inhibitory concentration (MIC) of antimicrobials against L. intracellularis, ten isolates obtained from North America and Europe were tested against 6 antimicrobials using a modified tissue culture assay. In vitro results found that carbadox, tiamulin, and valnemulin were the most active antimicrobials, chlortetracycline and tylosin were intermediately active, lincomycin was the least active against L. intracellularis, and the antimicrobial sensitivity patterns differed across isolates. We next evaluated the effectiveness of the modified tissue culture and direct count method with special fluorescence to determine in vitro disinfectant activity against L. intracellularis. The outcomes of both methods predicted similar in vitro bactericidal activities against L. intracellularis with a high degree of correspondence. This suggests that either assay would be appropriate in determining the bactericidal activity of disinfectants against L. intracellularis. Based on our in vitro results, we predict that a powder disinfectant (Stalosan® F), DC&R®, Roccal®-D, Synergize®, and Virkon®-S would perform well under field conditions, while Certi-Dine®, Nolvasan®-S, and Tek-Trol® would be less active against L. intracellularis. Finally, a new quantitative PCR assay was developed using a SYBR green system. The assay showed negative results when DNA from 16 other species of enteric bacteria as well as 20 negative control samples of pig feces was tested. Quantitative estimates from the assay were roughly 2-fold lower than the expected values across all dilutions of pure culture and spiked feces. Validation results indicated that this new qPCR is sensitive, specific, reliable, accurate, and precise for the detection and quantification of L. intracellularis in different types of samples. In conclusion, the results of these investigations update and expand upon existing information about the in vitro antimicrobial and disinfectant activities against L. intracellularis. Further, the development of a new, accurate, and precise qPCR assay will facilitate future epidemiology studies.

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