Friday, August 23, 2013

Use of bacteriotherapy to resolve Clostridium difficile infection

Clostridium difficile
Cara N. Wilder, Ph.D.

Antibiotic treatment of hospitalized patients poses a serious risk for the colonization of Clostridium difficile. Infection with this anaerobic, spore-forming bacterium is characterized by a wide spectrum of symptoms including diarrhea, fulminant pseudomembranous colitis, and death. Though treatment of C. difficile with antibiotics is often effective, a number of treated patients experience a relapse in infection following the cessation of antibiotic therapy. It is believed that these recurrent infections may be linked to a condition termed dysbiosis, which is characterized by a general imbalance of the intestinal microflora resulting from continuous antibiotic exposure.

In recent years, the use of a probiotic-based approach has been proposed as a promising therapy for the treatment of C. difficile infections. To determine the ideal combination of probiotic strains that both resolves C. difficile disease and restores a healthy intestinal microflora, Lawley et al. employed the use of a C. difficile murine infection model that parallels many aspects of human disease. In the study, the murine model was infected with an epidemic strain of C. difficile that was able to out-compete health-associated intestinal bacteria, allowing for the maintenance of dysbiosis. Following the establishment of infection, the group used fecal-transplantation as a model to identify a mixture of six phylogenetically-diverse bacteria that were able to trigger the expansion of health-associated intestinal microflora and resolve C. difficile infection within the mice. Overall, the results from this study highlight the therapeutic potential of probiotic bacteriotherapy in the treatment of C. difficile and other forms of dysbiosis.

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Wednesday, August 14, 2013

Preventing aspergillosis through immunization

Aspergillus fumigatus. Photo courtesy of
David Gregory and Debbie Marshall
Cara N. Wilder, Ph.D.

Over the last 20 years, Aspergillus fumigatus has become one of the most frequent causes of invasive fungal infection in immunologically compromised patients. This infection, termed aspergillosis, is associated with a wide spectrum of symptoms, including allergic reactions, organ failure, and lung infection. In most patients, A. fumigatus predominantly affects the lungs, resulting in an often fatal illness termed invasive pulmonary aspergillosis (IPA).

Current therapies for aspergillosis include treatment with antifungal drugs such as Amphotericin B or Triazole medications (Voriconazole, Posaconazole, Itraconazole). Unfortunately, these therapies have had limited success in treating IPA, and are often associated with serious toxicities. This is further compounded by the growing number A. fumigatus strains that have developed resistance against antifungal drugs, thus making the eradication of aspergillosis increasing more difficult. In response to these concerns, many research laboratories have focused their efforts on the development of new strategies targeted toward the prevention and treatment of such infections.

In one vaccine discovery program, Ito et al. used an immunochemical and mass spectrometric approach to identify an antigenic target for the development of a candidate vaccine that provides immunization against A. fumigatus. Following nasopulmonary inoculation of immunocompetent mice with viable A. fumigatus conida, Asp f 3 was identified as a potential antigenic target based on immunodominance during infection. Upon the analysis of this allergen, it was determined that subcutaneous immunization with full-length and truncated recombinant Asp f 3 offered protection against A. fumigatus infection. This was further confirmed following the examination of the lungs of vaccinated survivors, which were deemed free of hyphae and exhibited minimal infiltration of mononuclear cells. Overall, the findings from this study indicate that recombinant Asp f 3 is an effective immunogen and promising candidate for the development of a novel vaccine to prevent aspergillosis.



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Friday, August 9, 2013

Response of Campylobacter jejuni to Erythromycin Exposure

Campylobacter jejuni. Photo courtesy of Dr. Patricia Fields
and Dr. Collette Fitzgerald.
Cara N. Wilder, Ph.D.

Campylobacter jejuni is a Gram-negative, motile bacterium that is commonly present in the intestinal tract of both domestic and wild animals. In humans, C. jejuni causes a foodborne infection termed campylobacteriosis, which results in symptoms that range from mild enteritis to fever, headache, and bloody diarrhea. In some cases, Campylobacter infection has been associated with Guillain-Barré syndrome, which is a post-infection autoimmune disorder that damages nerve tissue.

Although most cases of campylobacteriosis are self-limiting, antibiotic treatment may be necessary for patients that are either immunologically compromised, or exhibit severe or persistent infection. The most common antimicrobial therapy used to treat campylobacteriosis is erythromycin, which is a macrolide antibiotic that inhibits bacterial protein translation. However, due the regular use of this antibiotic in animal production and veterinary medicine, an increasing number of C. jejuni isolates have become drug-resistant. In many of these strains, the mechanism of resistance is frequently conferred by target modification or through the expression of antibiotic efflux pumps.

Though the genetic basis of erythromycin resistance has been well-studied, the initial response and adaptive mechanisms directly following erythromycin exposure is not well understood. To analyze this, Xia et al. performed a competitive microarray hybridization study that examined the genome-wide transcriptional response of a sensitive and resistant strain of C. jejuni upon exposure to inhibitory and sub-inhibitory doses of erythromycin. Following treatment with erythromycin, the resistant strain of C. jejuni exhibited little to no differential gene expression. In contrast, a number of genes were differentially regulated in the sensitive strain, including the up-regulation of genes associated with motility, and the down-regulation of genes associated with energy production and conversion. Moreover, the inactivation of several differentially expressed genes appeared to negatively affect host colonization and the ability to tolerate high levels of oxygen. Overall, these results provide new insight into the adaptive response of C. jejuni to antibiotic treatment, and my help provide further understanding into the mechanisms underlying the emergence of antibiotic resistance.



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Friday, August 2, 2013

Survival mechanisms of Burkholderia cepacia complex cells grown in biofilms

Burkholderia cepacia. Photo courtesy of Janice Haney Carr
and CDC
Cara N. Wilder, Ph.D.

The Burkholderia cepacia complex (Bcc) is a group of Gram-negative bacteria composed of 17 closely related species. Of these strains, Burkholderia cenocepacia is an opportunistic pathogen frequently associated with high rates of transmission and mortality among immune-compromised people, such as those suffering from cystic fibrosis. Unfortunately, infections caused by B. cenocepacia and other Bcc strains are very difficult to eradicate due to a variety of  intrinsic antibiotic-resistance mechanisms including the expression of multidrug efflux pumps, inducible β-lactamases, altered penicillin-binding proteins, and the ability to form biofilms.

In particular, biofilms are multicellular microbial communities that can form on various environmental, clinical, and abiotic surfaces. These groups of sessile cells are often more tolerant to antibiotics than free-living, planktonic cells due to decreased growth rates, differential gene expression, and reduced penetration of the biofilm. Thus, upon exposure to antibiotic therapies, a small subpopulation of cells within the biofilm is able to persist by entering a dormant multidrug-tolerant state. Following the removal of the antibiotic, these “persister cells” are then able to reestablish growth and create a new biofilm.

To elucidate the mechanisms behind the emergence of persister cells in Bcc biofilms, Acker et al. analyzed B. cenocepacia biofilms following treatment with Tobramycin, a bactericidal antibiotic known to induce the formation of harmful reactive oxygen species (ROS). Through the use of transcriptome analysis, flow cytometry, ROS-staining, and inhibitor studies, the group discovered that surviving persister cells were able to escape cell death through the down-regulation of the tricarboxylic acid (TCA) cycle, allowing cells to avoid ROS production, and through the activation of the glyoxylate shunt, which is an anaplerotic pathway of the TCA cycle. This finding may provide novel approaches for the treatment of Bcc biofilms as the glyoxylate shunt is absent in humans, and inhibition of this pathway prior to treatment with Tobramycin was found to decrease the number of persisters. Thus, this pathway may be an ideal target for combination therapy.