Sara Cannas

Detection and Isolation of Mycobacterium avium Subspecies paratuberculosis from Intestinal Mucosal Biopsies of Patients with and without Crohn's Disease in Sardinia

OBJECTIVES:Sardinia is an island community of 1.6 million people. There are also about 3.5 million sheep and one hundred thousand cattle in which Johnes disease and Mycobacterium avium subspecies paratuberculosis infection are endemic. The present study was designed to determine what proportion of people in Sardinia attending for ileocolonoscopy with or without Crohns disease were infected with this pathogen.METHODS:Mycobacterium avium subspecies paratuberculosis was detected by IS900 PCR on DNA extracts of fresh intestinal mucosal biopsies as well as by isolation in culture using supplemented MGIT media followed by PCR with amplicon sequencing.RESULTS:Twenty five patients (83.3%) with Crohns disease and 3 control patients (10.3%) were IS900 PCR positive (p = 0.000001; Odds ratio 43.3). Mycobacterium avium subspecies paratuberculosis grew in cultures from 19 Crohns patients (63.3%) and from 3 control patients (10.3%) (p = 0.00001; Odds ratio 14.9). All patients positive by culture had previously been positive by PCR. Mycobacterium avium subspecies paratuberculosis first appeared in the liquid cultures in a Ziehl Neelsen (ZN) staining negative form and partially reverted through a rhodamine-auramine positive staining form to the classical ZN positive form. This resulted in a stable mixed culture of all 3 forms illustrating the phenotypic versatility of these complex chronic enteric pathogens.CONCLUSIONS:Mycobacterium avium subspecies paratuberculosis was detected in the majority of Sardinian Crohns disease patients. The finding of the organism colonizing a proportion of people without Crohns disease is consistent with what occurs in other conditions caused by a primary bacterial pathogen in susceptible hosts.

Mycobacterium avium Subspecies paratuberculosis Infection in Cases of Irritable Bowel Syndrome and Comparison with Crohn's Disease and Johne's Disease: Common Neural and Immune Pathogenicities

ABSTRACT Mycobacterium avium subsp. paratuberculosis causes Johnes disease, a systemic infection and chronic inflammation of the intestine that affects many species, including primates. Infection is widespread in livestock, and human populations are exposed. Johnes disease is associated with immune dysregulation, with involvement of the enteric nervous system overlapping with features of irritable bowel syndrome in humans. The present study was designed to look for an association between Mycobacterium avium subsp. paratuberculosis infection and irritable bowel syndrome. Mucosal biopsy specimens from the ileum and the ascending and descending colon were obtained from patients with irritable bowel syndrome attending the University of Sassari, Sassari, Sardinia, Italy. Crohns disease and healthy control groups were also included. Mycobacterium avium subsp. paratuberculosis was detected by IS900 PCR with amplicon sequencing. Data on the potential risk factors for human exposure to these pathogens and on isolates from Sardinian dairy sheep were also obtained. Mycobacterium avium subsp. paratuberculosis was detected in 15 of 20 (75%) patients with irritable bowel syndrome, 3 of 20 (15%) healthy controls, and 20 of 23 (87%) people with Crohns disease (P = 0.0003 for irritable bowel syndrome patients versus healthy controls and P = 0.0000 for Crohns disease patients versus healthy controls). One subject in each group had a conserved single-nucleotide polymorphism at position 247 of IS900 that was also found in isolates from seven of eight dairy sheep. There was a significant association (P = 0.0018) between Mycobacterium avium subsp. paratuberculosis infection and the consumption of hand-made cheese. Mycobacterium avium subsp. paratuberculosis is a candidate pathogen in the causation of a proportion of cases of irritable bowel syndrome as well as in Crohns disease.

Evaluation of the Antimicrobial Properties of the Essential Oil of Myrtus communis L. against Clinical Strains of Mycobacterium spp.

Mycobacterium tuberculosis is the etiological agent of tuberculosis. The World Health Organization has estimated that 8 million of people develop active TB every year and the situation is complicated by an increase of Mycobacterium tuberculosis strains resistant to drugs used in antitubercular therapy: MDR and XDR-TB. Myrtle leaf extracts, used as an antiseptic in Sardinian traditional medicine, have strong antibacterial activity as several investigations showed. In this study we investigated the antimicrobial properties of the essential oil of Myrtus communis against clinical strains of M. tuberculosis and M. paratuberculosis.

Chemical composition, cytotoxicity, antimicrobial and antifungal activity of several essential oils

Essential oils (EOs) are known and used for their biological, antibacterial, antifungal and antioxidant properties. Numerous studies have shown that EOs exhibit a large spectrum of biological activities in vitro. The incidence of drug-resistant pathogens and the toxicity of antibiotics have drawn attention to the antimicrobial activity of natural products, encouraging the development of alternative treatments. The aim of this study was to analyse the phytochemical and the cytotoxic characteristic of 36 EOs; we then evaluated the antimicrobial activity of the less-toxic EOs on Gram-positive, Gram-negative and fungi strains. The results showed low cytotoxicity in seven EOs and good activity against Gram-negative and Candida spp. strains. Based on our results, EOs could be proposed as a novel group of therapeutic agents. Further experiments are necessary to confirm their pharmacological effectiveness, and to determine potential toxic effects and the mechanism of their activity in in vivo models.

Antifungal, anti-biofilm and adhesion activity of the essential oil of Myrtus communis L. against Candida species.

Candida species belong to the normal microbiota of the oral cavity, gastrointestinal tract and vagina. The increasing incidence of drug-resistant pathogens and the toxicity of the antifungal compounds have drawn the attention towards the antimicrobial activity of natural products, an inexpensive alternative. The aim of this work was to evaluate the adhesion activity, the biofilm formation and the action of the Myrtus communis L. essential oil (EO) on the biofilm formation towards three species isolated from clinical samples: Candida albicans, Candida parapsilosis and Candida tropicalis. Furthermore, we evaluated the antimycotic activity of the EO towards the three species, and the results were compared with the minimum inhibitory concentration of six antimycotics. The activity of the EO against C. albicans and C. parapsilosis was better than that obtained against C. tropicalis; moreover, the strains used in the assay were adhesive and biofilm producer, and the effect of myrtle EO on the biofilm formation yielded encouraging results.

Tuberculosis patients are characterized by a low–IFN-γ/high–TNF-α response to methylated HBHA produced in M. smegmatis

Whole blood from Mycobacterium tuberculosis-infected subjects was stimulated with heparin-binding hemagglutinin (HBHA). Tuberculosis (TB) patients showed an HBHA-specific T-cell response characterized by low-IFN-γ/high-TNF-α secretion, while asymptomatic subjects with latent infection (LTBI) and TB patients under therapy showed a pattern with high IFN-γ/low TNF-α. These results underscore the usefulness of HBHA in helping to distinguish LTBI subjects versus TB patients.

In vitro activities of antimycobacterial agents against Mycobacterium avium subsp. paratuberculosis linked to Crohn's Disease and Paratuberculosis

Crohns disease, a human disease similar to paratuberculosis in animals is the most painful and devastating disease that may involve infection with M. avium subsp. paratuberculosis (MAP), different genetic polymorphisms and an immune dysregulation syndrome. Treatment of Crohns disease is most commonly based on 5-aminosalicylic acid (5-ASA) compounds, corticosteroids, and immunosuppressive agents. Recently, biological therapies using monoclonal antibodies against inflammatory cytokines have shown some positive results. However, all these therapies treat the symptoms not the cause of the disease.

Molecular identification of Mycobacterium avium subspecies paratuberculosis in oral biopsies of Crohn’s disease patients

Oral lesions may be found in patients with Crohn’s disease (CD), in a percentage up to 20%. The aim of this study was to investigate a possible relationship between Mycobacterium avium subsp. paratuberculosis (MAP) and oral lesions in CD patients. 23 oral biopsies were examined performing IS900 Nested PCR; 9 of them were positive: 8 from CD patients and 1 from a control. Our purpose is to go on with this study, amplifying the number of subjects examined and testing subjects with oral lesions related to diseases other than CD to verify the specific association between MAP and oral lesions in CD patients.

Essential oils in ocular pathology: an experimental study

INTRODUCTION The antimicrobial activity of essential oils (EOs) has been known for ages; in particular, the EOs of Melaleuca alternifolia, Thymus vulgaris, Mentha piperita, and Rosmarinus officinalis have been used for the treatment of fungal and bacterial infections. METHODOLOGY This study focused on the in vitro cytotoxicity to normal human conjunctiva cells and antimicrobial activity of 20 EOs. RESULTS The oils tested showed no cytotoxic effect at very low concentrations. Rosmarinus officinalis, Melaleuca alternifolia, and Thymus vulgaris L. red thyme geraniol sel oils had good antimicrobial activity against Gram-positive and Gram-negative strains. CONCLUSIONS The results of this study are of great interest and may have a major impact on public health, providing useful tips to optimize the therapeutic use of some natural drugs.

Change in Caco-2 cells following treatment with various lavender essential oils

Abstract Lavender is an aromatic evergreen shrub diffused in the Mediterranean basin appreciated since antiquity. The genus Lavandula is part of Lamiaceae family and includes more than 20 species, among which true lavender (L. vera D.C. or L. angustifolia Miller.) and spike lavender (L. latifolia Medikus); there are also numerous hybrids known as lavandins (L. hybrida Rev.). L. vera, spike lavender and several hybrids are the most intensely used breeding species for the production of essential oils. Lavender and lavandin essential oils have been applied in food, pharmaceutical and other agro industries as biological products. In their chemical composition, terpenes linalool and linalyl acetate along with terpenoids such as 1,8-cineole are mostly responsible for biological and therapeutic activities. This study evaluates cytotoxic activity of essential oils derived from four lavender species on human epithelial colorectal adenocarcinoma cells. Analysis of pre- and post-treatment cell morphology has been performed using scanning electron microscope.