Evaluation of potential novel biomarkers for feline hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most common cardiomyopathy in cats. The diagnosis can be difficult, requiring advanced echocardiographic skills. Additionally, circulating biomarkers (N-terminal pro-B type natriuretic peptide and cardiac troponin I) have several limitations when used for HCM screening. In previous work, we identified interleukin 18 (IL-18), insulin-like growth factor binding protein 2 (IGFBP-2), brain-type glycogen phosphorylase B (PYGB), and WNT Family Member 5 A (WNT5A) as myocardial genes that show significant differential expression between cats with HCM and healthy cats. The products of these genes are released into the circulation, and we hypothesized that IL-18, IGFBP-2, PYGB, and WNT5A serum RNA and protein concentrations differ between healthy cats, cats with subclinical HCM, and those with HCM and congestive heart failure (HCM + CHF). Reverse transcriptase quantitative polymerase chain reaction (RTqPCR) and enzyme-linked immunosorbent assay (ELISA) were applied to evaluate gene and protein expression, respectively, in the serum of eight healthy controls, eight cats with subclinical HCM, and six cats with HCM + CHF. Serum IGFBP-2 RNA concentrations were significantly different among groups and were highest in cats with subclinical HCM. Compared to healthy controls, serum IL-18 and WNT5A gene expression were significantly higher in cats with HCM + CHF, and WNT5A was higher in cats with subclinical HCM. No differences were observed for PYGB. These results indicate that further investigation via large scale clinical studies for IGFBP-2, WNT5A, and IL-18 may be valuable in diagnosing and staging feline HCM.

Effects of inflammatory stimuli on the development of Mycoplasma bovis pneumonia in experimentally challenged calves.

Many cattle infected with Mycoplasma bovis remain healthy while others develop severe chronic respiratory disease. We hypothesized that inflammatory stimuli such as co-pathogens worsen disease outcomes in M. bovis-infected calves. Calves (n=24) were intrabronchially inoculated with M. bovis and either killed bacterial lysate, transient M. haemolytica infection, or saline. Caseonecrotic lesions developed in 7/7 animals given M. haemolytica and M. bovis compared to 2/8 given M. bovis with no inflammatory stimulus, and 6/9 animals given bacterial lysate and M. bovis (P=0.01). Animals receiving M. haemolytica and M. bovis had more caseonecrotic foci in lungs than those receiving M. bovis with no inflammatory stimulus (median = 21 vs 0; P = 0.01), with an intermediate response (median = 5) in animals given bacterial lysate. In addition to caseonecrotic foci, infected animals developed neutrophilic bronchiolitis that appeared to develop into caseonecrotic foci, peribronchiolar lymphocytic cuffs that were not associated with the other lesions, and 4 animals with bronchiolitis obliterans. The data showed that transient lung inflammation at the time of M. bovis infection provoked the development of caseonecrotic bronchopneumonia, and the severity of inflammation influenced the number of caseonecrotic foci that developed. In contrast, caseonecrotic lesions were few or absent in M. bovis-infected calves without a concurrent inflammatory stimulus. These studies provide insight into how caseonecrotic lesions develop within the lung of M. bovis-infected calves. This and other studies suggest that controlling co-pathogens and harmful inflammatory responses in animals infected with M. bovis could potentially minimize development of M. bovis caseonecrotic bronchopneumonia.

Bovine astrovirus and its role in lymphocytic encephalitis in cattle in Ontario, Canada, 1988-2019.

Astroviruses have been found in cattle and other species with encephalitis. Our objective was to determine the frequency of neurotropic bovine astrovirus (BoAstV) in cases of encephalitis in cattle ≥ 4-mo-old. Of 56 cases of idiopathic lymphocytic encephalitis examined retrospectively (1988-2019), fixed brain from 11 cases (19%) tested positive by semi-quantitative RT-PCR for BoAstV CH13/NeuroS1. None of the control cases tested positive, including 32 with other forms of encephalitis and 40 with no neurologic disease. Most astrovirus-positive cases were 1-2-y-old, with a range of 7 mo to 7 y, and affected both beef and dairy breeds with wide geographic distribution. BoAstV-positive cases had acute onset of neurologic signs of 12 h to 7 d before death or euthanasia. Affected cattle had lymphocytic inflammation throughout the brain including cerebrum, thalamus, midbrain, cerebellum, medulla oblongata, and spinal cord, and affecting gray and white matter. Further PCR testing identified a possible cause in 9 of the 45 (20%) remaining idiopathic cases of lymphocytic encephalitis, including eastern equine encephalitis virus, Listeria monocytogenes, bovine viral diarrhea virus, bovine alphaherpesvirus 1, and ovine gammaherpesvirus 2 (malignant catarrhal fever); we found no cases of infection by West Nile virus, rabies virus, or Chlamydia spp. No cause was identified in 36 of 56 (64%) cases of lymphocytic encephalitis. We frequently identified neurotropic BoAstV in cases of lymphocytic encephalitis that had no previously identified cause. Neurotropic BoAstV infections had gone undetected for decades, but the frequency of BoAstV infections has not increased among contemporary cases.

Mannheimia haemolytica increases Mycoplasma bovis disease in a bovine experimental model of BRD.

Amongst the bacterial pathogens associated with the bovine respiratory disease syndrome (BRD) in cattle are Mannheimia haemolytica and Mycoplasma bovis. The interaction between these two pathogens has not been investigated before; thus, there are gaps in the knowledge of why and how a previous infection with M. haemolytica allows the development of M. bovis-related lesions. We hypothesized that upon M. haemolytica infection, inflammatory products are produced in the lung and that these inflammatory products stimulate M. bovis to produce proteases and lipases that degrade lipids and proteins important for lung function. In this work, we identified several M. bovis proteases and lipases whose expression was modulated by M. haemolytica products in vitro. We performed co-infection animal challenges to develop a model to test vaccine protection. A prior exposure to BHV-1 followed by infection with M. bovis and M. haemolytica resulted in severe pathology and the BHV-1 infection was abandoned. When M. bovis and M. haemolytica were introduced into the lungs by bronchoscopy, we found that M. haemolytica resulted in worsening of the respiratory disease caused by M. bovis. We performed a proof-of-concept trial where animals were immunized with the M. bovis proteins identified in this study and challenged with both pathogens. Despite detecting significant humoral immune responses to the antigens, the experimental vaccine failed to protect against M. bovis disease.

Bronchopneumonia with interstitial pneumonia in feedlot cattle: Epidemiologic characteristics of affected animals.

Bronchopneumonia with interstitial pneumonia (BIP) of feedlot cattle is characterized by gross and histologic lesions of cranioventral bronchopneumonia (BP) and caudodorsal interstitial pneumonia. This study described the characteristics and frequency of BIP in western Canadian feedlot cattle and identified epidemiologic differences between BIP and either BP or acute interstitial pneumonia (AIP). The study of 9909 deaths on 4 western Canadian feedlots included 1105 BIP, 1729 BP, and 878 AIP cases. A population of 55 cases with gross, histopathology, and microbiology data was used to validate the primary data set. BIP was the second most common reason for death (or euthanasia) from respiratory disease (1105/9909 cases), and the observed frequency was twice what was expected from random concurrence of BP and AIP. Based on logistic regression models, epidemiologic characteristics of BIP were comparable to those of BP, although BIP cases were more chronic with more instances of clinical illness prior to death. BIP was epidemiologically distinct from AIP. Specifically, BIP more frequently affected steers than heifers, deaths occurred earlier in the feeding period at lower body weights and lower daily weight gains, and BIP cases had longer durations from the first clinical illness to death and more separate instances of clinical illness prior to death. Furthermore, death from BIP mainly occurred in winter and fall, while death from AIP was most frequent in summer. These findings define BIP as a unique condition of feedlot cattle and suggest that chronic BP may promote the development of fatal interstitial lung disease in at-risk cattle.

Bronchopneumonia with interstitial pneumonia in beef feedlot cattle: Characterization and laboratory investigation.

Bronchopneumonia with interstitial pneumonia (BIP) has been considered a variant of acute interstitial pneumonia (AIP) rather than a distinct disease. This study compared 18 BIP, 24 bronchopneumonia (BP), and 13 AIP cases in feedlot beef cattle. Grossly, BIP cases typically had cranioventral lung lesions of similar morphology and extent as BP cases, but the caudodorsal lung appeared overinflated, bulged on section, and had interlobular edema and emphysema. Gross diagnosis of BIP had 83% sensitivity and 73% specificity relative to histopathology. Histologic lesions of BIP in cranioventral areas were of chronic BP, while caudodorsal lesions included alveolar and bronchiolar damage and inflammation, interstitial hypercellularity, and multifocal hemorrhages. In BIP cases, cranioventral lung lesions were more chronic than caudodorsal lesions. Histologic scores and microbiology data were comparable in cranioventral lung of BIP versus BP cases and caudodorsal lung of BIP versus AIP cases, with differences reflecting a more chronic disease involving less virulent bacteria in BIP versus BP. Mycoplasma bovis infection was similarly frequent among groups, and a viral cause of BIP was not identified. Lesion morphology and similar blood cytokine concentrations among groups argued against sepsis as a cause of lung injury. Surfactant dysfunction was identified in BIP and BP, and was only partially the result of protein exudation. These and other findings establish BIP as a distinct condition in which chronic cranioventral BP precedes acute caudodorsal interstitial lung disease, supporting a role of chronic inflammation in heightened sensitivity to 3-methylindole or another lung toxicant.

Cardiac gene activation varies between young and adult cats and in the presence of hypertrophic cardiomyopathy.

Little is known about the difference of myocardial gene transcription in young and adult cats and how transcription is further modified in cats with hypertrophic cardiomyopathy (HCM) and with left atrial (LA) thrombus formation. We hypothesized that selected factors for coagulation, endothelial activation, inflammation, and remodelling are modified with age and are activated in the hearts of cats with HCM. Left atrial and ventricular (LV) samples from 12 cats with HCM (seven without (HCMwoAT] and five with LA thrombi [HCMwAT]), and six young (YC) and six adult (AC) control cats without cardiac disease were investigated for relative expression of the following genes using quantitative polymerase chain reaction: von Willebrand factor, a disintegrin and metalloproteinase with a thrombospondin type 1 motif member 13, platelet activating factor, E- and P-selectin, intercellular and vascular adhesion molecules-1, ß2-integrin, vascular endothelial growth factor, interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), heat shock protein-70, and myocyte-specific enhancer factor 2C. Significant differences in gene activation were found between YC and AC, and YC and cats with HCM. Compared to AC, MCP-1 and IL-6 were significantly higher in cats with HCM. The presence of an LA thrombus was associated with higher IL-6 expression. These results illustrate the relevance of age and/or lifestyle on gene expression in the feline heart. The gene transcription pattern found in AC hearts might predispose cats to their characteristic cardiac remodelling processes and thrombus formation if disease occurs. It further supports the involvement of inflammation, but not coagulation and endothelial activation, in HCM.

Laboratory investigation of cases of fatal bacterial pneumonia in dairy cows.

Objective: Bacterial bronchopneumonia occurs in mature dairy cows but much of the information is extrapolated from knowledge of the disease in calves. The study was prompted by perceptions of an increasing occurrence and a paucity of information on fatal Mannheimia haemolytica pneumonia in dairy cows in Ontario. The study objectives were to describe the seasonality, main pathogens involved, and suggested predisposing factors for cases of fatal bacterial bronchopneumonia in mature dairy cows submitted for postmortem examination to a diagnostic laboratory, and to evaluate if the frequency of such submissions has increased over time. Animals: Mature dairy cows. Procedure: Retrospective study of cases submitted for postmortem examination to a diagnostic laboratory from 2007-2020 that were diagnosed as bacterial bronchopneumonia. Results: Most of the postmortem cases of bacterial bronchopneumonia in dairy cows were submitted from November to February (54% of cases). Mannheimia haemolytica was isolated from lung of 61/101 cases. Viruses were only identified in 8/55 cases tested. A minority (29/92) of bacterial isolates had in vitro resistance to antimicrobials used to treat pneumonia. Frequently suggested predisposing factors included recent introductions or movement of animals, recent or imminent calving, inclement weather, concurrent diseases, and poor ventilation in barns. Conclusion and clinical relevance: This study describes seasonal and annual trends, major pathogens, antimicrobial resistance profiles, and suggested predisposing factors in Ontario dairy cows submitted to a diagnostic laboratory for postmortem investigation of pneumonia and provides insights for understanding why outbreaks occur.

Objectif: La bronchopneumonie bactérienne survient chez les vaches laitières matures, mais une grande partie de l'information est extrapolée à partir de la connaissance de la maladie chez les veaux. L'étude a été motivée par la perception d'une occurrence croissante et d'un manque d'information sur la pneumonie mortelle à Mannheimia haemolytica chez les vaches laitières en Ontario. Les objectifs de l'étude étaient de décrire la saisonnalité, les principaux agents pathogènes impliqués et les facteurs prédisposants suggérés pour les cas de bronchopneumonie bactérienne mortelle chez les vaches laitières matures soumises à un examen post-mortem à un laboratoire de diagnostic, et d'évaluer si la fréquence de telles soumissions a augmenté au fil du temps. Animaux: Vaches laitières matures. Procédure: Étude rétrospective des cas soumis pour examen post-mortem à un laboratoire de diagnostic, entre 2007 et 2020, qui ont été diagnostiqués comme une bronchopneumonie bactérienne. Résultats: La plupart des cas post-mortem de bronchopneumonie bactérienne chez les vaches laitières ont été soumis de novembre à février (54 % des cas). Mannheimia haemolytica a été isolée du poumon de 61/101 cas. Des virus n'ont été identifiés que dans 8/55 cas testés. Une minorité (29/92) d'isolats bactériens présentaient une résistance in vitro aux antimicrobiens utilisés pour traiter la pneumonie. Les facteurs prédisposants fréquemment suggérés comprenaient des introductions ou des déplacements récents d'animaux, un vêlage récent ou imminent, des conditions météorologiques défavorables, des maladies concomitantes et une mauvaise ventilation dans les étables. Conclusion et pertinence clinique: Cette étude décrit les tendances saisonnières et annuelles, les principaux agents pathogènes, les profils de résistance aux antimicrobiens et les facteurs prédisposants suggérés chez les vaches laitières de l'Ontario soumises à un laboratoire de diagnostic pour une enquête post-mortem sur la pneumonie et fournit des informations pour comprendre pourquoi les épidémies se produisent.(Traduit par Dr Serge Messier).

Acute interstitial pneumonia and the biology of 3-methylindole in feedlot cattle.

Acute interstitial pneumonia (AIP) of cattle has been recognized for many decades. While the pathogenesis and risk factors for this condition in pastured cattle are relatively well characterized, there remains a poor understanding of the disease as it occurs in intensively fed cattle such as in beef feedlots. Specifically, in pastured cattle, AIP results from excessive ruminal production of the pneumotoxicant 3-methylindole (3-MI). In feedlot cattle, the evidence to substantiate the role of 3-MI is comparatively deficient and further investigations into the cause, pathogenesis, and control are sorely needed. This review highlights our current understanding of AIP with a focus on the disease as it occurs in feedlot cattle. Additionally, it illustrates the need for further work in understanding the specific animal factors (e.g. the ruminal microbiome, and the role of concurrent diseases), management factors (e.g. animal stocking and vaccination protocols), and dietary factors (e.g. dietary supplements) that may impact the development of AIP and which are relatively unique to the feedlot setting. All stakeholders in the beef industry stand to benefit from a greater understanding of what remains a pressing yet poorly understood issue in beef production.

The effect of aerosolized bacterial lysate on experimentally induced Mannheimia haemolytica pneumonia in calves.

Bovine respiratory disease (BRD) often occurs during specific periods of increased susceptibility when stress, viral infection, or reduced air quality are thought to suppress respiratory defences. The innate immune system is rapidly responsive and broadly protective and could be a target for preventing BRD during these periods of increased susceptibility. This study tested the hypothesis that stimulation of pulmonary innate immune responses by aerosol delivery of a lysate of killed Escherichia coli and Staphylococcus aureus bacteria would protect calves against Mannheimia haemolytica pneumonia. Ten clean-catch colostrum-deprived Holstein calves were randomly assigned to receive either aerosolized bacterial lysate or saline 24 hours before M. haemolytica challenge. Effects of this treatment on clinical, hematologic, microbiologic, and pathologic outcomes were assessed. Compared to controls, lysate-treated calves had lower serum haptoglobin and blood leukocyte and neutrophil concentrations following M. haemolytica challenge. There were no differences in temperature, heart and respiratory rates, clinical scores, ultrasound lesions, or number of M. haemolytica in the nasal cavity or lung. Thus, treatment with bacterial lysate prior to M. haemolytica challenge appeared to ameliorate early measures of inflammation but did not provide sufficient protection to substantially alter the course of disease.

La maladie respiratoire bovine (BRD) survient souvent pendant des périodes spécifiques de sensibilité accrue lorsque le stress, une infection virale ou une qualité de l'air réduite sont censés supprimer les défenses respiratoires. Le système immunitaire inné est rapidement réactif et largement protecteur et pourrait être une cible pour prévenir la BRD pendant ces périodes de sensibilité accrue. Cette étude a testé l'hypothèse selon laquelle la stimulation des réponses immunitaires innées pulmonaires par la délivrance d'aérosols d'un lysat de bactéries Escherichia coli et Staphylococcus aureus tuées protégerait les veaux contre la pneumonie à Mannheimia haemolytica. Dix veaux Holstein dont on a limité la contamination bactérienne et privés de colostrum ont été répartis au hasard pour recevoir soit un lysat bactérien en aérosol, soit une solution saline 24 heures avant une infection défi par M. haemolytica. Les effets de ce traitement sur les résultats cliniques, hématologiques, microbiologiques et pathologiques ont été évalués. Comparativement aux témoins, les veaux traités au lysat présentaient des concentrations sériques d'haptoglobine et de leucocytes et de neutrophiles sanguins plus faibles après la provocation par M. haemolytica. Il n'y avait aucune différence dans la température, les fréquences cardiaques et respiratoires, les scores cliniques, les lésions échographiques ou le nombre de M. haemolytica dans la cavité nasale ou les poumons. Ainsi, le traitement avec un lysat bactérien avant la provocation par M. haemolytica a semblé améliorer les réactions précoces de l'inflammation mais n'a pas fourni une protection suffisante pour modifier substantiellement l'évolution de la maladie.(Traduit par Docteur Serge Messier).

Effects of inflammatory stimuli on responses of macrophages to Mycoplasma bovis infection.

Inflammation in the respiratory tract is thought to worsen the disease response to Mycoplasma bovis infection. This study investigated the cells involved in this response with a focus on proteases and cytokines as harmful effector mechanisms. By immunohistochemistry, Mac387-positive macrophages were the main cell type comprising the foci of caseous necrosis in cattle with M. bovis pneumonia. Thus, the study evaluated how priming of different types of macrophages with bacterial lysate (or pro-inflammatory cytokines induced by the bacterial lysate) affected their responses to M. bovis infection. Inducible responses were detected in monocyte-derived macrophages (M1-MDMs and M2-MDMs), whereas pulmonary alveolar macrophages (PAMs) were minimally affected by priming or infection. M. bovis-infected MDMs secreted MMP-12 and SPLA2, and priming with pro-inflammatory cytokines increased the secretion of cathepsin B in response to M. bovis infection. Of these, there were higher concentrations of cathepsin B and SPLA2 in lungs with M. bovis pneumonia compared to healthy lungs, and these are potential mechanisms for macrophage-induced lung damage in M. bovis infection. Priming of MDMs with either bacterial lysate or with pro-inflammatory cytokines caused an enhanced response to M. bovis infection with respect to IL-8 and IL-1ß secretion. The findings of this study suggest proteases, lipases and cytokines derived from monocyte-derived macrophages as possible mediators by which prior inflammation in the respiratory tract worsen disease outcomes from M. bovis infection.

Protective and Anti-Inflammatory Effects of Protegrin-1 on Citrobacter rodentium Intestinal Infection in Mice.

Infectious intestinal colitis, manifesting as intestinal inflammation, diarrhea, and epithelial barrier disruption, affects millions of humans worldwide and, without effective treatment, can result in death. In addition to this, the significant rise in antibiotic-resistant bacteria poses an urgent need for alternative anti-infection therapies for the treatment of intestinal disorders. Antimicrobial peptides (AMPs) are potential therapies that have broad-spectrum antimicrobial activity due to their (1) unique mode of action, (2) broad-spectrum antimicrobial activity, and (3) protective role in GI tract maintenance. Protegrin-1 (PG-1) is an AMP of pig origin that was previously shown to reduce the pathological effects of chemically induced digestive tract inflammation (colitis) and to modulate immune responses and tissue repair. This study aimed to extend these findings by investigating the protective effects of PG-1 on pathogen-induced colitis in an infection study over a 10-day experimental period. The oral administration of PG-1 reduced Citrobacter rodentium intestinal infection in mice as evidenced by reduced histopathologic change in the colon, prevention of body weight loss, milder clinical signs of disease, and more effective clearance of bacterial infection relative to challenged phosphate-buffered saline (PBS)-treated mice. Additionally, PG-1 treatment altered the expression of various inflammatory mediators during infection, which may act to resolve inflammation and re-establish intestinal homeostasis. PG-1 administered in its mature form was more effective relative to the pro-form (ProPG-1). To our knowledge, this is the first study demonstrating the protective effects of PG-1 on infectious colitis.

Effect of aerosolized bacterial lysate on development of naturally occurring respiratory disease in beef calves.

BACKGROUND: Bovine respiratory disease (BRD) is a major problem affecting beef cattle after arrival to feedlots. Alternatives to antibiotics are needed for prevention. HYPOTHESIS: Stimulation of pulmonary innate immune responses at the time of arrival to a feedlot reduces the occurrence and severity of BRD. ANIMALS: Sixty beef steers at high risk of BRD. METHODS: Randomized, double-blinded, placebo-controlled study. Calves received saline or a lysate of Staphylococcus aureus and Escherichia coli by aerosol, at 16 hours after feedlot arrival. Calves were monitored for 28 days for disease outcomes and levels of Mycoplasma bovis and Mannheimia haemolytica in nasal swabs. RESULTS: Death from M bovis pneumonia was significantly greater in lysate-treated animals (6/29, 24%) compared to controls (1/29, 3%; odds ratio = 10.2; 95% confidence interval [CI] = 1.1-96.0; P = .04). By 28 days after arrival, 29/29 lysate-treated calves had ultrasonographic pulmonary consolidation compared to 24/29 control calves (P = .05). Lysate-treated calves had lower weight gain compared to control calves (-8.8 kg, 95% CI = -17.1 to -0.5; P = .04), and higher body temperatures on days 4, 7, and 21 (0.19°C; 95% CI = 0.01-0.37; P = .04). Nasal M bovis numbers increased over time and were higher in lysate-treated calves (0.76 log CFU, 95% CI = 0.3-1.2; P = .001). CONCLUSIONS AND CLINICAL IMPORTANCE: Aerosol administration of a bacterial lysate exacerbated BRD in healthy high-risk beef calves, suggesting that respiratory tract inflammation adversely affects how calves respond to subsequent natural infection with M bovis and other respiratory pathogens.

Host Tolerance to Infection with the Bacteria that Cause Bovine Respiratory Disease.

Calves vary considerably in their pathologic and clinical responses to infection of the lung with bacteria. The reasons may include resistance to infection because of pre-existing immunity, development of effective immune responses, or infection with a minimally virulent bacterial strain. However, studies of natural disease and of experimental infections indicate that some calves develop only mild lung lesions and minimal clinical signs despite substantial numbers of pathogenic bacteria in the lung. This may represent "tolerance" to pulmonary infection because these calves are able to control their inflammatory responses or protect the lung from damage, without necessarily eliminating bacterial infection. Conversely, risk factors might predispose to bovine respiratory disease by triggering a loss of tolerance that results in a harmful inflammatory and tissue-damaging response to infection.

Pulmonary and systemic responses to aerosolized lysate of Staphylococcus aureus and Escherichia coli in calves.

BACKGROUND: Constitutive and inducible defenses protect the respiratory tract from bacterial infection. The objective of this study was to characterize the response to an aerosolized lysate of killed bacteria, as a basis for studying the regulation and in vivo effects of these inducible innate immune responses. RESULTS: Bacterial lysate consisting of heat-killed and sonicated Staphylococcus aureus and Escherichia coli was aerosolized to 6 calves and systemic and pulmonary innate immune and inflammatory responses were measured in the first 24 h relative to baseline. Evaluated parameters included clinical parameters (body temperature and heart and respiratory rates), blood acute phase proteins and leukocyte counts, and leukocytes and proteins in bronchoalveolar lavage fluid. Mild clinical signs with increased heart rates and rectal temperatures developed following administration of the lysate, with resolution by 24 h. Serum haptoglobin and plasma fibrinogen concentrations were elevated at 24 h relative to baseline. Bronchoalveolar lavage fluid (BALF) had increased cellularity and increased proportion of neutrophils, as well as higher concentrations of interleukin (IL)-8, IL-10 and total protein at 24 h relative to baseline. Mass spectrometry identified 965 unique proteins in BALF: 19 proteins were increased and 26 proteins were decreased relative to baseline. The upregulated proteins included those involved in innate immunity including activation of complement, neutrophils and platelets. At postmortem examination, calves receiving higher doses of lysate had areas of lobular consolidation and interlobular edema. Histologically, neutrophils were present within bronchioles and to a lesser extent within alveoli. Calves receiving highest doses of lysate had patchy areas of neutrophils, hemorrhage and hyaline membranes within alveoli. CONCLUSIONS: Aerosolization of bacterial lysate stimulated an innate immune response in lungs and airways, with alveolar damage observed at higher doses. Such a stimulus could be of value for investigating the effects of inducible innate immune responses on occurrence of disease, or for evaluating how stress, drugs or genetics affect these dynamic responses of the respiratory tract.

Effect of tracheal antimicrobial peptide on the development of Mannheimia haemolytica pneumonia in cattle.

Bacterial pneumonia causes significant economic loss to the beef industry and occurs at times of stress and viral infection. Administering antibiotics to at-risk calves is often used to prevent the disease, but alternatives to mass treatment with antibiotics are needed. Tracheal antimicrobial peptide (TAP), a ß-defensin naturally produced by bovine airways, has bactericidal activity against the pathogens that cause pneumonia in cattle. However, TAP expression is suppressed by glucocorticoid (stress) and viral infection. We hypothesized that delivering TAP to the respiratory tract would prevent development of pneumonia in calves infected with Mannheimia haemolytica. Clean-catch calves (i.e. obtained prior to contact with the dam) were challenged by aerosol with M. haemolytica, and TAP or water was delivered to the respiratory tract at 0.3, 2 and 6 hours post-infection. TAP treatment did not protect against development of disease. Calves treated with TAP had similar bacterial loads in the nasal cavity and lung compared to calves treated with water. Similarly, TAP treatment did not affect the development of clinical signs, elevated rectal temperatures, or increased levels of blood neutrophils, haptoglobin and fibrinogen that occurred after bacterial challenge. Postmortem gross and histologic lung lesions were also similar in the two groups. To determine why there was a lack of protective effect, we tested the effect of substances in respiratory lining fluid on the bactericidal activity of TAP. Physiologic concentrations of sodium chloride inhibited TAP bactericidal activity in vitro, as did serum at concentrations of 0.62 to 2.5%, but concentrated bronchoalveolar lavage fluid had no consistent effect. These findings suggest that TAP does not have in vivo bactericidal activity against M. haemolytica because of interference by physiological sodium chloride levels and by serum. Thus, administration of TAP may not be effective for prevention of M. haemolytica pneumonia.

Development of an aerosolized Mannheimia haemolytica experimental pneumonia model in clean-catch colostrum-deprived calves.

Mannheimia haemolytica is an important cause of bovine respiratory disease (BRD). BRD is usually a multifactorial disease with host factors and viral infections influencing pathogenesis. Previous studies that have attempted to experimentally induce pneumonia using aerosolized M. haemolytica alone have produced inconsistent results, yet an aerosol model would be useful to study the details of early infection and to investigate the role of innate defences in pathogenesis. The objective of these studies was to develop and characterize an aerosolized M. haemolytica disease model. In an initial study, conventionally raised calves with higher levels of antibody against M. haemolytica leukotoxin developed acute respiratory distress and diffuse alveolar damage, but did not develop bronchopneumonia, following challenge with M. haemolytica serotype 1. Clean-catch colostrum-deprived calves challenged with 1 × 1010 colony forming units of M. haemolytica serotype 1 consistently developed bronchopneumonia, with elevations in rectal temperature, serum haptoglobin, plasma fibrinogen, and blood neutrophils. Mannheimia haemolytica serotype 1 was consistently isolated from the nasal cavities and lungs of challenged calves. Despite distribution of aerosol and isolation of M. haemolytica in all lung lobes, gross lesions were mainly observed in the cranioventral area of lung. Gross and histologic lesions included neutrophilic bronchopneumonia and fibrinous pleuritis, with oat cells (necrotic neutrophils with streaming nuclei), and areas of coagulative necrosis, which are similar to lesions in naturally occurring BRD. Thus, challenge with M. haemolytica serotype 1 and use of clean-catch colostrum-deprived calves with low or absent antibody titres allowed development of an effective aerosol challenge model that induced typical clinical disease and lesions.

Changes in antimicrobial susceptibility profiles of Mycoplasma bovis over time.

Mycoplasma bovis is a major cause of pneumonia, arthritis, and mastitis in cattle and can lead to significant economic losses. Antimicrobial resistance is a concern and further limits the already short list of drugs effective against mycoplasmas. The objective of this study was to examine changes in in vitro minimum inhibitory concentrations (MICs) of antimicrobials of aminoglycoside, fluoroquinolone, lincosamide, macrolide, pleuromutilin, phenicol, and tetracycline classes for 210 M. bovis isolates collected from 1978 to 2009. The MIC50 values of the various antimicrobials were also compared. The MIC50 levels for enrofloxacin and danofloxacin remained low (0.25 µg/mL) across all 3 decades. MIC50 levels for tetracyclines, tilmicosin, and tylosin tartrate were low in the 1980s, then increased in the 1990s and remained high. In the 1980s, MIC50 levels were low for clindamycin, spectinomycin, and tulathromycin, increased in the 1990s to 8 µg/mL (clindamycin) and 32 µg/mL (spectinomycin and tulathromycin), then decreased again in the 2000s. Members of the fluoroquinolone class of antimicrobials had the lowest MIC50 levels across all 3 decades, which suggests in vitro susceptibility of M. bovis to this class of antimicrobials. Statistically significant associations were observed between MIC values for chlortetracycline, oxytetracycline, tylosin tartrate, and tilmicosin; between clindamycin, tulathromycin, spectinomycin, and tiamulin; and between tylosin tartrate and clindamycin. Changes in MIC levels of various antimicrobials over time show the importance of monitoring the susceptibility of mycoplasmas to antimicrobials. The number of antimicrobials that showed elevated MIC50 levels, and therefore possibly reduced in vitro effectiveness against M. bovis, supports initiatives that promote prudent use of antimicrobials in agriculture.

Mycoplasma bovis est une cause majeure de pneumonie, d'arthrite, et mammite chez les bovins et peut entrainer des pertes économiques significatives. La résistance antimicrobienne est une préoccupation et réduit encore plus la courte liste déjà existante de médicaments efficaces contre les mycoplasmes. L'objectif de la présente étude était d'examiner in vitro les changements des concentrations minimales inhibitrices (CMI) des antimicrobiens des classes des aminoglycosides, des fluoroquinolones, des lincosamides, des macrolides, des pleuromutilines, des phénicoles, et des tétracyclines envers 210 isolats de M. bovis collectionnés entre 1978 et 2009. Les valeurs de CMI50 des différents antimicrobiens ont également été comparées. Les valeurs de CMI50 de l'enrofloxacine et de la danofloxacine sont demeurées faibles (0,25 µg/mL) au cours des trois décennies. Les valeurs de CMI50 pour les tétracyclines, le tilmicosin et le tartrate de tylosine étaient basses dans les années 1980s, puis augmentèrent dans les années 1990s et sont demeurées élevées. Durant les années 1980s, les valeurs de CMI50 étaient basses pour la clindamycine, la spectinomycine, et la tulathromycine, augmentèrent dans les années 1990s jusqu'à 8 µg/mL (clindamycine) et 32 µg/mL (spectinomycine et tulathromycine), puis ont diminué encore dans les années 2000s. Les membres de la classe des fluoroquinolones avaient les valeurs de CMI50 les plus faibles au cours des trois décennies examinées, ce qui suggère une sensibilité in vitro de M. bovis à cette classe d'antibiotiques. Des associations statistiquement significatives furent notées entre les valeurs de CMI de la chlortétracycline, l'oxytétracycline, le tartrate de tylosine, et le tilmicosin; entre la clindamycine, la tulathromycine, la spectinomycine, et la tiamulin; et entre le tartrate de tyloosine et la clindamycine. Les changements dans les valeurs de CMI de différents antibiotiques dans le temps démontrent l'importance de suivre la sensibilité des mycoplasmes aux antimicrobiens. Le nombre d'antimicrobiens qui a démontré des valeurs élevées de CMI50, et ainsi une efficacité in vitro réduite envers M. bovis, encourage les initiatives qui font la promotion de l'usage prudent des antimicrobiens en agriculture.(Traduit par Docteur Serge Messier).