Plasma levels of TNF-α, IFN-γ, IL-4 and IL-10 during a course of experimental contagious bovine pleuropneumonia

<p>Abstract</p> <p>Background</p> <p>Contagious Bovine Pleuropneumonia (CBPP), caused by <it>Mycoplasma mycoides </it>subsp. <it>mycoides</it>, is widespread in sub-Saharan Africa. The current live vaccine T1/44 has limited efficacy and occasionally leads to severe side effects in the animals. A better understanding of the immune responses triggered by <it>Mycoplasma mycoides </it>subsp. <it>mycoides </it>and their role in disease progression will help to facilitate the design of a rational vaccine. Currently, knowledge of cytokines involved in immunity and immunopathology in CBPP is rather limited. The aim of this study was to characterize the <it>in vivo </it>plasma concentrations of the cytokines TNF-α, IFN-γ, IL-4, IL-10 and the overall role of CD4⁺T cells in the development of cytokine levels during a primary infection. Plasma cytokine concentrations in two groups of cattle (CD4⁺T cell-depleted and non-depleted cattle) experimentally infected with <it>Mycoplasma mycoides </it>subsp. <it>mycoides </it>were measured and their relationship to the clinical outcomes was investigated.</p> <p>Results</p> <p>Plasma cytokine concentrations varied between animals in each group. Depletion of CD4⁺T cells did not induce significant changes in plasma levels of TNF-α, IL-4, and IL-10, suggesting a minor role of CD4⁺T cells in regulation or production of the three cytokines during the time window of depletion (1-2 weeks post depletion). Unexpectedly, the IFN-γ concentrations were slightly, but statistically significantly higher in the depleted group (p < 0.05) between week three and four post infection. Three CD4⁺T cell-depleted animals that experienced severe disease, had high levels of TNF-α and IFN-γ. Only one severely diseased non-depleted animal showed a high serum concentration of IL-4 post infection.</p> <p>Conclusions</p> <p>Comparison of most severely diseased animals, which had to be euthanized prior to the expected date, versus less severe diseased animals, irrespective of the depletion status, suggested that high TNF-α levels are correlated with more severe pathology in concomitance with high IFN-γ levels.</p

Immunohistochemical investigations on Brucella ceti-infected, neurobrucellosis-affected striped dolphins (Stenella coeruleoalba)

Bacteria of the genus Brucella cause brucellosis, an infectious disease common to humans as well as to terrestrial and aquatic mammals. Since 1994 several cases of Brucella spp. infection have been reported in marine mammals worldwide. Indeed, since human brucellosis ranks as one of the most common bacterial zoonotic infections on a global scale, it is necessary to increase our knowledge about it also in the marine environment. Brucella ceti, which is phenotypically similar to other smooth brucellae as B. abortus and B. melitensis, shares with the latter two the same surface antigens that are routinely used for the serological diagnosis of Brucella spp. infection. Marine mammal Brucella spp. infections are characterized by a pathogenicity similar to their terrestrial counterparts, with the occurrence of abortion, stillbirth and orchitis and an involvement of the host’s central nervous system (CNS), similarly to what happens in mankind. While sero-epidemiological data suggest that Brucella spp. infection is widespread globally, detecting Brucella spp.-associated antigens by immunohistochemistry (IHC) in tissues from infected animals is often troublesome. The present study was aimed at investigating, by means of IHC based upon the utilization of an anti-Brucella LPS monoclonal antibody (MAb), the CNS immunoreactivity (IR) shown by B. ceti-infected, neurobrucellosis-affected striped dolphins

Development and validation of an antigen capture ELISA based on monoclonal antibodies specific for Listeria monocytogenes in food

A capture enzyme-linked immunosorbent assay (ELISA) for the identification of Listeria monocytogenes in food was standardised and validated. The assay was refined by analysing samples of meat, seafood, dairy products, pasta and flour. The method was found to be 100% specific for Listeria spp. tested, with a limit of sensitivity of 6.6 × 10(3) colony-forming units (cfu)/ml. Comparison of L. monocytogenes capture ELISA against the official International Organization for Standardization (ISO) method 11290-1:1996 for the isolation and identification of L. monocytogenes in food matrices produced a significant concordance index. The assay was validated on food matrices including meat, seafood and dairy products in line with ISO 16140:2003 concerning qualitative analytical methods. The assay was found to be accurate, specific, sensitive, selective, reproducible and fast, resulting in lower costs and faster turnaround in microbiological screening of foods

Sviluppo e validazione di un antigene-capture ELISA basato su anticorpi monoclonali specifici per Listeria monocytogenes negli alimenti

È stato standardizzato e validato un dosaggio immunoenzimatico capture ELISA per l’identificazione di Listeria monocytogenes negli alimenti. Il dosaggio è stato messo a punto analizzando campioni di prodotti carnei, ittici e lattiero-caseari, pasta di semola e di farina di grano. Il metodo è risultato specifico al 100% per Listeria spp., con limite di rivelazione di 6,6 × 10(3) cfu/ml. Il metodo L. monocytogenes capture ELISA è stato confrontato con il metodo ufficiale ISO 11290-1:1996 per l’isolamento e l’identificazione di L. monocytogenes in matrici alimentari ottenendo un indice di concordanza significativo. Il dosaggio è stato validato in base alle indicazioni della norma ISO 16140:2003 relativamente ai metodi di analisi qualitativi. Il dosaggio è risultato accurato, specifico, sensibile, selettivo, riproducibile e rapido da eseguire, consentendo nello screening degli alimenti la riduzione di tempi e costi dell’indagine microbiologica

Development and evaluation of diagnostic tests for the serological diagnosis of brucellosis in swine

A competitive enzyme-linked immunosorbent assay (c-ELISA), an indirect ELISA (i-ELISA) and a dissociation-enhanced lanthanide fluorescence immunoassay (DELFIA) were developed to test for antibodies to Brucella suis in pig and wild boar sera. An anti-Brucella-LPS monoclonal antibody (MAb 4B5A) (c-ELISA and DELFIA) and an anti-swine IgG monoclonal antibody (MAb 10C2G5) (i-ELISA) were used for the three assays. The specificity (Sp) and sensitivity (Se) of the assays gave the following results: Se and Sp = 100% at a cut-off value of 61.0% (B/B0%) for c-ELISA; Sp = 99.1% and Se = 100% at a cut-off value of 21.7% (percentage positivity: PP%) for i-ELISA; Sp = 91.0% and Se = 75% at a cut-off value of 37.0% (B/B0%) for DELFIA. In addition, the performance of a commercial fluorescence polarisation assay (FPA), standardised for bovine sera, was evaluated in swine sera. The specificity and sensitivity obtained were both 100% at a cut-off value of 99.5 (millipolarisation unit values). These results suggest that the combination of c-ELISA, i-ELISA and FPA can be used to improve the serological diagnosis of swine brucellosis

Sviluppo e valutazione di test diagnostici per la sierodiagnosi di brucellosi suina

Sono stati sviluppati una ELISA competitiva (c-ELISA), una ELISA indiretta (i-ELISA) e un test immunologico DELFIA (Dissociation-Enhanced Lanthanide Fluorescence Immunoassay) per la ricerca di anticorpi verso Brucella suis in sieri di maiale e cinghiale. I tre test prevedono l’utilizzo di un anticorpo monoclonale (MAb 4B5A) verso l’LPS di Brucella (c-ELISA e DELFIA) e di un anticorpo monoclonale (MAb 10C2G5) verso le IgG suine (i-ELISA). La specificità (Sp) e la sensibilità (Se) dei tre test sono le seguenti: per la c-ELISA Se e Sp = 100% con un valore di cut-off pari al 61.0% (B/B0%); per la i-ELISA Sp = 99.1% e Se = 100% con un valore di cut-off di 21.7% (PP%); per il DELFIA Sp = 91.0% e Se = 75% ponendo il valore di cut-off al 37.0% (B/B0%). Inoltre sono state valutate le performance, nei confronti di sieri suini, di un test FPA (Fluorescence Polarization Assay) commerciale sviluppato per la ricerca di anticorpi anti-Brucella in sieri bovini; la specificità e la sensibilità ottenute sono entrambe del 100% al valore di cut-off di 99.5 (mP). Questi risultati suggeriscono che la combinazione di c-ELISA, i-ELISA e FPA può essere utilizzata per migliorare la diagnosi di brucellosi suina

Stress Adaptation Responses of a Listeria monocytogenes 1/2a Strain via Proteome Profiling

Listeria monocytogenes is a foodborne pathogen that is ubiquitous and largely distributed in food manufacturing environments. It is responsible for listeriosis, a disease that can lead to significant morbidity and fatality in immunocompromised patients, pregnant women, and newborns. Few reports have been published about proteome adaptation when L. monocytogenes is cultivated in stress conditions. In this study, we applied one-dimensional electrophoresis and 2D-PAGE combined with tandem mass spectrometry to evaluate proteome profiling in the following conditions: mild acid, low temperature, and high NaCl concentration. The total proteome was analyzed, also considering the case of normal growth-supporting conditions. A total of 1,160 proteins were identified and those related to pathogenesis and stress response pathways were analyzed. The proteins involved in the expression of virulent pathways when L. monocytogenes ST7 strain was grown under different stress conditions were described. Certain proteins, particularly those involved in the pathogenesis pathway, such as Listeriolysin regulatory protein and Internalin A, were only found when the strain was grown under specific stress conditions. Studying how L. monocytogenes adapts to stress can help to control its growth in food, reducing the risk for consumers