Meera C. Heller

Bluetongue virus infection activates bovine monocyte-derived macrophages and pulmonary artery endothelial cells.

Bluetongue virus (BTV) is the cause of bluetongue (BT), an emerging, arthropod-transmitted disease of ungulates. The cellular tropism of BTV in ruminants includes macrophages, dendritic cells and endothelial cells (ECs), and fulminant infection is characterized by lesions consistent with those of so-called viral hemorrhagic fevers. Specifically, BT is characterized by vascular injury with hemorrhage, tissue infarction and widespread edema. To further investigate the pathogenesis of vascular injury in BT, we evaluated the responses of cultured bovine pulmonary artery EC (bPAEC) and monocyte-derived macrophages (bMDM) to BTV infection by measuring transcript levels of genes encoding molecules important in mediating EC activation and/or endothelial barrier dysregulation. The data confirm that BTV infection of bPAEC resulted in increased transcription of genes encoding chemokine ligand 2 (CCL2) and E-selectin, and BTV infection of bMDM resulted in increased transcription of genes encoding TNF-alpha, IL-1beta, IL-8, and inducible nitric oxide synthase (iNOS). The data from these in vitro studies provide further evidence that cytokines and other vasoactive substances produced in macrophages potentially contribute to vascular injury in BTV-infected ruminants, along with direct effects of the virus itself on ECs.

Characterization of Brucella abortus infection of bovine monocyte-derived dendritic cells.

Brucella abortus is a Gram negative facultative intracellular pathogen of cattle, and an important zoonosis in humans worldwide. Previous studies have shown that dendritic cells (DC) from humans and mice are highly permissive for Brucella survival and proliferation. Impairment of DC activation and maturation by Brucella infection has also been reported in these two species. The aim of this study was to characterize infection of bovine DC with B. abortus. Monocyte-derived DC (mdDC) were cultured from bovine peripheral blood mononuclear cells (PBMC) using the recombinant bovine cytokines IL-4 and GM-CSF. The resulting mdDC were DEC205(+), MHC class II(hi). Approximately 70% of the cultured cells were DEC205(+), MHC II(+). MdDC were infected with B. abortus strain 2308 at an MOI of 1 and 100. Parallel infection experiments were performed in monocyte derived macrophages (mdM) isolated from the same subjects. Bacteria were successfully killed by mdDC by 24 hours post infection (PI) with high and low dose of B. abortus, bacteria persisted in mdM infected with a high dose. Expression of IL-1b, IL-6, IL-10, IL-12p40, IFNγ, iNOS and TNFα in B. abortus infected and LPS stimulated mdDC at 6 and 24 hours PI were evaluated using RT-qPCR. At 6 hours PI all transcripts were increased over control cells and significantly less IL-10, IL-12p40, and IFNγ were expressed in mdDC infected with B. abortus compared to LPS stimulation. Evaluation of mdDC cultures by flow cytometry was performed. Flow cytometric analysis of infected and LPS stimulated mdDC 24 hours PI showed expression of CD80 and CD86 was impaired in two of the three animals analyzed. MHC class II expression was equivocal between the groups. From these results we conclude that cultured bovine mdDC are not permissive for intracellular proliferation of B. abortus, and infected mdDC exhibit some signs of maturational and activational impairment.

A potential role for indoleamine 2,3-dioxygenase (IDO) in Rhodococcus equi infection.

Rhodococcus equi is a facultative intracellular bacterial pathogen of foals and immunocompromised humans that infects and proliferates within host macrophages and dendritic cells (DC). Indoleamine 2,3-dioxygenase (IDO), the initial enzyme in the tryptophan catabolism pathway, is upregulated in R. equi infected equine monocyte-derived DC and alveolar macrophages. Tryptophan requirement of R. equi for extracellular and intracellular growth was assessed. Growth of R. equi in minimal media did not require tryptophan and pharmacologic inhibition of IDO had no effect on intracellular proliferation of R. equi in equine alveolar macrophages. To investigate an immune-regulatory role for INDO in R. equi infection, IDO(-/-) (B6.129-(Indotm1Alm)/J) (n=22) and strain matched control (C57BL/6J) (n=20) mice were infected with R. equi by intraperitoneal injection, for 3 and 6 days. There was no difference in bacterial counts in liver or spleen between the two groups. Histological sections of liver and spleen were assigned inflammation scores and RT-PCR for interferon-gamma (IFNγ), tumor necrosis factor-alpha (TNFα), IL-4, IL-6, IL-10, IL-12, IL-23, forkhead box P3 (FoxP3), and transforming growth factor-beta (TGFβ) was performed on liver and spleen. Liver tissue of IDO(-/-) had higher inflammation scores at 6 days post-infection (PI) (P=0.05) and had decreased expression of TGFβ at 3 days PI (P=0.01), and FOXP3 at 3 days (P=0.02) and 6 days (P=0.03) compared to control mice. Immunostaining for FOXP3 showed lower numbers of FOXP3+ regulatory T cells in liver of IDO(-/-) mice 6 days PI. Prolonged inflammation in the liver tissue of IDO(-/-) mice corresponded with lower expression of FOXP3 and TGFβ in that tissue, and also with lower numbers of FOXP3+ regulatory T cells. We conclude that IDO expression by activated macrophages and DC plays a role in dampening the inflammatory response to R. equi infection in mice.

Acute phase proteins in naturally occurring respiratory disease of feedlot cattle.

The aim of this study was to evaluate three acute phase proteins (APP) [haptoglobin (HPT), lipopolysaccharide binding protein (LBP) and transferrin (Tf)] in feedlot cattle with naturally occurring respiratory disease diagnosed by a calf health scoring chart (CHSC). Seventy-seven beef calves were observed for signs of Bovine Respiratory Disease (BRD) during the first 28 days after arrival at the feedlot. Fourteen cases and pen matched controls were selected based on the CHSC. BRD cases were defined as a score of ≥ 5, while controls were defined as a score ≤ 4. The mean CHSC score in cases was 6.9 which was significantly greater than the controls 2.8 (P < 0.01). Mean plasma LBP and HPT concentrations were significantly greater in cases than controls (P < 0.01). Our study results show that measurement of HPT and LBP could be useful in detecting respiratory disease in feedlot conditions. Transferrin concentrations between the two groups were not statistically different.

Acute phase proteins in healthy goats Establishment of reference intervals

Acute inflammatory processes can trigger increased production of acute phase proteins (APPs) that can be useful biomarkers of inflammation. APPs are diverse and include proteins involved in coagulation, opsonization, iron regulation, and limitation of tissue injury. Haptoglobin, serum amyloid A, and alpha-1 acid glycoprotein have been proposed as useful APPs in goats. APPs can differ markedly by species, therefore species-specific reference intervals and studies are necessary. The objective of this study was to determine species-specific reference intervals for 4 APPs in goats. Haptoglobin, serum amyloid A, lipopolysaccharide binding protein, and alpha-1 acid glycoprotein were measured in in 54 clinically normal adult goats. APPs were measured using goat-specific commercial enzyme-linked immunosorbent assay kits. Results were analyzed by 1-way analysis of variance to compare sexes and breeding status. Reference Value Advisor was used to calculate reference limits according to the IFCC-CLSI guidelines. Only 1 APP was found to vary in healthy animals; serum haptoglobin was increased in lactating animals and decreased in pregnant does in their second trimester when compared with open, nonlactating does. No sex-based differences were seen for any of the APPs measured. We report normal reference intervals for 4 serum APPs that may be useful as disease markers. Haptoglobin should be interpreted with caution in animals with unknown pregnancy status. Further studies are needed to determine whether these APPs are useful biomarkers in goat disease states.

Identification of immunologically relevant genes in mare and foal dendritic cells responding to infection by Rhodococcus equi.

Rhodococcus equi is a facultative intracellular bacterial pathogen of horses; infected foals develop pyogranulomatous pneumonia, however adult horses are largely unaffected. R. equi infects and proliferates within host macrophages and dendritic cells (DCs). DCs initiate the appropriate adaptive immune response, thereby playing a critical role in determining the outcome of infection. Our aim was to identify genes that are differentially expressed in R. equi infected monocyte-derived DCs (mdDCs). Peripheral blood monocytes from mares and foals were used to derive mdDCs by culturing with recombinant equine IL-4 and recombinant human GM-CSF. RNA harvested 24h after infection with R. equi (ATCC 33701+) was used to perform suppression subtractive hybridization (SSH) experiments. Approximately 38 unique sequences were obtained from these experiments. Differential expression of 19 immunologically relevant genes was validated by PCR. These genes are characterized by the following functions: cell adhesion, chemotaxis/migration, immune/inflammatory response, ion transport, signal transduction, T-cell regulation, and vesicular transport. In summary, we identified several novel genes that are differentially expressed in foal and adult mdDCs in response to R. equi infection. These genes provide promising targets for further research into the host response to R. equi, and the susceptibility of foals to this disease.

Use of a percutaneous transabdominal catheter for management of obstructive urolithiasis in goats, sheep, and potbellied pigs: 69 cases (2000-2014).

OBJECTIVE To evaluate the use of a percutaneous transabdominal catheter (PTC) for urinary bladder drainage in goats, sheep, and potbellied pigs with obstructive urolithiasis. DESIGN Retrospective case series. ANIMALS 43 goats, 10 sheep, and 16 potbellied pigs (all males) with obstructive urolithiasis evaluated at the University of California-Davis Veterinary Medical Teaching Hospital. PROCEDURES Medical records of goats, sheep, and potbellied pigs examined because of obstructive urolithiasis from January 2000 through December 2014 were reviewed. Records of animals for which a standard PTC had been placed into the urinary bladder as part of disease management were selected. Data were collected regarding signalment, complications associated with PTC placement, and duration of PTC placement prior to removal. RESULTS 42 of 43 goats, 5 of 10 sheep, and all potbellied pigs were castrated. Median (range) duration of PTC placement was 2 (1 to 4) days for goats, 1 (1 to 4) day for sheep, and 1 (1 to 3) day for potbellied pigs. Complications associated with PTC placement included blockage of the catheter by urine sediment, perforation of the cecum, and migration of the catheter out of the urinary bladder. CONCLUSIONS AND CLINICAL RELEVANCE Placement of a PTC into the urinary bladder allowed for effective stabilization of goats, sheep, and potbellied pigs with obstructive urolithiasis while acid-base and electrolyte imbalances were corrected. Use of a PTC should be considered for urinary bladder drainage during medical management or prior to surgical management of obstructive urolithiasis for these species.

Efficacy of Oral Administration of Sodium Iodide to Prevent Bovine Respiratory Disease Complex

Background The prevention of bovine respiratory disease complex (BRD) in beef cattle is important to maintaining health and productivity of calves in feeding operations. Objective Determine whether BRD bacterial and viral pathogens are susceptible to the lactoperoxidase/hydrogen peroxide/iodide (LPO/H2O2/I−) system in vitro and to determine whether the oral administration of sodium iodide (NaI) could achieve sufficient concentrations of iodine (I) in the respiratory secretions of weaned beef calves to inactivate these pathogens in vivo. Animals Sixteen weaned, apparently healthy, commercial beef calves from the University of Missouri, College of Veterinary Medicine teaching herd. Methods In vitro viral and bacterial assays were performed to determine susceptibility to the LPO/H2O2/I− system at varying concentrations of NaI. Sixteen randomly selected, healthy crossbred beef weanlings were administered 70 mg/kg NaI, or water, orally in a blinded, placebo‐controlled trial. Blood and nasal secretions were collected for 72 hours and analyzed for I− concentration. Results Bovine herpesvirus‐1, parainfluenza‐3, Mannheimia haemolytica and Bibersteinia trehalosi were all inactivated or inhibited in vitro by the LPO/H2O2/I− reaction. Oral administration of NaI caused a marked increase in nasal fluid I concentration with a C max = 181 (1,420 μM I), T12, a sufficient concentration to inactivate these pathogens in vitro. Conclusions and Clinical Importance In vitro, the LPO/H2O2/I− system inactivates and inhibits common pathogens associated with BRD. The administration of oral NaI significantly increases the I concentration of nasal fluid indicating that this system might be useful in preventing bovine respiratory infections.

Pharmacokinetics and efficacy of intravenous famotidine in adult cattle

Background Abomasal ulceration is recognized in neonatal and adult cattle, but research regarding treatment is limited. Histamine‐2 receptor antagonists (H2RA), such as famotidine, are used clinically with little evidence‐based research about efficacy in adult cattle. Hypothesis and Objectives Intravenous famotidine administered at 0.4 mg/kg will increase the pH of abomasal outflow digesta compared to saline control in adult cattle. The objectives were to assess the effect of famotidine, administered as a single dose and as multiple doses, on abomasal outflow fluid pH in adult cattle. A third objective was to describe the pharmacokinetic parameters of IV famotidine in cattle. Animals Four clinically healthy adult Angus‐cross steers previously fitted with duodenal cannulae placed orad to the biliary and pancreatic ducts. Methods Randomized, 2‐way cross‐over clinical trial. Steers received IV famotidine (0.4 mg/kg) as a single and 3‐dose regimen (every 8 hours) versus saline control. Blood for analysis of serum famotidine concentration was collected intermittently for 12 hours, and abomasal outflow fluid pH was measured at intervals for a 24‐hour period. After a 34‐hour washout period, the opposite treatments were administered and the sampling repeated. Results Abomasal outflow fluid pH was higher in steers treated with famotidine for up to 4 hours after a single dose but the effect decreased with subsequent doses. The median (range) elimination half‐life was 3.33 (3.21‐3.54) hours. Conclusions and Clinical Importance Famotidine may be useful for treatment or prevention of abomasal ulceration in adult cattle, but the duration of effect may decrease with time.

Transpalpebral exenteration in cattle: a retrospective study of 115 cases

OBJECTIVE To describe the indications for exenteration and complications associated with the procedure. ANIMALS STUDIED 115 cattle. PROCEDURES Medical records of cattle presented for unilateral exenteration evaluated at the University of California, Davis Veterinary Medical Teaching Hospital from January 1985 through December 2015 were reviewed. RESULTS Median (range) age at presentation for all cattle was 6 (0.2-30) years. The most prevalent (80.9%) indication for exenteration was squamous cell carcinoma (SCC). Cattle >5 years had higher odds (OR = 11.2, 95% CI, 2.8-45.8) for undergoing exenteration due to SCC compared to cattle ≤5 years. Herefords had higher odds (OR = 4.6, 95% CI, 1.5-14.6) for undergoing exenteration for SCC compared to other breeds. Holsteins had higher odds (OR = 140.7, 95% CI, 7.5-2644) for undergoing exenteration for retrobulbar lymphoma compared to other breeds. Complications following exenteration were reported in 15 cases (13.0%). The postsurgical complications were orbital abscesses (6/15), recurrence of SCC (5/15), wound dehiscence (3/15), and excessive hemorrhage (1/15). Median (range) time to occurrence of postsurgical complications was 19 (5-205) days. There was no significant association (P > 0.05) between ocular diagnosis, age, anesthetic technique or the suture pattern used to close the skin postsurgically, and occurrence of postsurgical complications. CONCLUSIONS Early clinical diagnosis of SCC by owners and veterinarians may prevent the need for exenteration. Owners should be made aware of the possible postsurgical complications following exenteration in cattle.