A.J. Masot

Histopathological and Immunohistochemical Findings in the Lungs of Pigs Infected Experimentally with Mycoplasma hyopneumoniae

Pigs were infected intranasally with Mycoplasma hyopneumoniae and killed at intervals ranging from 7 to 35 days post-infection (dpi). Histopathological changes consisted of (1) exudates in airways and alveolar lumina, (2) peribronchial and peribronchiolar lymphoid hyperplasia, and (3) enlargement of alveolar septa. These changes were particularly marked from 7 to 28dpi, coinciding with significant increases in the expression, detected immunohistochemically, of cytokines (IL-1alpha, IL-1beta, IL-8, TNF-alpha and INF-gamma) and lymphoid markers (CD4+, CD8+, muramidase, IgG+, IgA+). Both the lesions and immunohistochemical signals declined in intensity beyond 35 days.

Induction of interleukin-8 and interleukin-12 in neonatal ovine lung following experimental inoculation of bovine respiratory syncytial virus.

This study aimed to determine the immunohistochemical expression of interleukin (IL)-1β, tumour necrosis factor alpha (TNF)-α, interferon (IFN)-γ, IL-4, IL-6, IL-8, IL-10 and IL-12 and to measure the concentrations of these cytokines in lung tissue from lambs infected experimentally with bovine respiratory syncytial virus (BRSV). Lambs (n = 15) were inoculated at 2 days of age with 20 ml of viral inoculum (1.26 × 10(6) TCID50 per ml) or sterile medium (n = 15). Rectal temperature, pulse and respiratory rates were monitored daily in control and infected lambs. Lambs were killed and subject to necropsy examination at 1, 3, 5, 7 and 15 days post inoculation (dpi). There was a temporal association between pulmonary expression of these cytokines and lung pathology in BRSV-infected lambs. The cytokines IL-4 and IL-10 were not elevated, but there was a significant increase in IL-1β, TNF-α, IFN-γ and IL-6 proteins and labelled cells, suggesting that these cytokines may play a role in the biological response to BRSV infection and contribute to the development of lung lesions. There was also a significant increase in the cytokine concentration and number of immunolabelled cells expressing IL-8 and IL-12 in infected lungs, suggesting that these cytokines might be used as therapeutic targets in the management of BRSV, in conjunction with measures to combat the causative pathogen and prophylactic methods aimed at preventing infection.

Dominant expression of interleukin-8 vs interleukin-1β and tumour necrosis factor alpha in lungs of lambs experimentally infected with Mannheimia haemolytica

Abstract AIMS: To quantify the number of cells infected with Mannheimia haemolytica and expressing interleukin (IL)-1β, tumour necrosis factor alpha (TNFα) and IL-8 using immunohistochemistry, and to measure the immunoreactivity of cytokines in pulmonary tissue extracts using ELISA, in the lung of lambs experimentally infected with M. haemolytica, and to compare the patterns of expression of cytokines in airways at different times post-infection (p.i.). METHODS: Twenty 3-month-old lambs of both sexes were randomly assigned to two groups, viz infected (n=15), and uninfected controls (n=5). Each lamb in the infected group was inoculated with 1.5 x 109 cfu M. haemolytica in 5 mL sterile nutrient broth, control lambs were inoculated with 5 mL sterile nutrient broth and clinical signs were monitored. Infected and control animals were killed at 1, 3, 5, 7, and 15 days p.i. Histopathology and immunohistochemistry were conducted to determine the number of immunolabelled cells in pneumonic lungs, and study the pattern of expression of IL-1β, TNFα and IL-8 in lung extracts using ELISA. RESULTS: Lesions in bronchi and bronchioles ranged from epithelial desquamation to bronchiolitis obliterans and necrosis. The alveoli had areas of seroproteinaceous fluid, fibrin and bacterial aggregates that evolved to foci of pyogranulomatous inflammation with clustered inflammatory cells, referred to as ‘oat cells’. M. haemolytica antigen was observed in the cytoplasm of inflammatory cells. Labelling of IL-1β, TNFα and IL-8 was observed in bronchial and bronchiolar epithelial cells, alveolar exudate, and in interstitial inflammatory infiltrate, with increased expression on 1 and 3 days p.i. for IL-1β and TNFα, and 1, 3, and 5 days p.i. for IL-8. In lung tissue extracts, peak concentrations of IL-1β (55 (SD 5) ng/mL), TNFα (92 (SD 6) pg/mL) and IL-8 (8 [SD 2] μg/mL) occurred at 3 days p.i. CONCLUSIONS: The results of this study suggested that the inflammatory cytokines IL-1β, TNFα and IL-8 may play an important role in enhancing the biological response to M. haemolytica, and contribute to the development of lesions in the lung in pulmonary pasteurellosis in sheep. Given that the expression of IL-8 in lung was much greater than that of IL-1β and TNFα, anti-cytokine agents directed at this mediator could be useful in the prevention and treatment of this disease.

Bovine respiratory syncytial virus in-situ hybridization from sheep lungs at different times postinfection

Se estudio, mediante la tecnica de hibridacion in situ, la distribucion del ARN viral del virus respiratorio Sincicial Bovino (VRSB) en pulmon de corderos infectados en forma experimental, a diferentes tiempos postinoculacion. La sonda usada para la hibridacion in situ se preparo mediante transcripcion reversa del ARN del VRSB, seguida de la amplificacion mediante PCR de cADN. 25 corderos de raza Merino de ambos sexos y de un peso vivo de 55 (+/- 10) Kg, fueron inoculados por via intratraqueal con 40 ml de solucion salina que contenia 1.26 x 10[elevado a 6] DIM[subindice 50] por ml (cepa viral NMK7). Los corderos se sacrificaron los dias 1, 3, 7, 11 y 15 postinoculacion. Las celulas epiteliales bronquiales y bronquiolares resultaron positivas a los acidos nucleicos de VRSB los dias 1, 3, 7 y 11 postinoculacion. A su vez, el epitelio alveolar contenia celulas positivas los dias 1, 3, y 7 postinoculacion. Se detectaron celulas que contenian el ARN viral en las luces bronquiales y bronquilares del dia 1 al 11 postinoculacion, y en el exudadado alveolar en los dias 3 y 7 postinoculacion. Se identificaron senales positivas de hibridacion desde el dia 3 al 11 postinoculacion, tanto en las celulas intersticiales mononucleares como en el tejido linfoide asociado a los bronquios. Las senales de hibridacion mas intensas se detectaron a los dias 3 y 7 postinoculacion, lo que coincidio con las lesiones histopatologicas de mayor consideracion.