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July-September 2013 Research Highlights
Transferring Antimicrobial Resistance
Collaborators from NCTR, the Marshfield Clinic Research Foundation, and China Agricultural University have characterized multiple antimicrobial genes located in the plasmids and integrons from Salmonella enterica, a well-known foodborne pathogen. Plasmids and their associated integrons are mobile genetic elements that can replicate independently from chromosomal DNA and can often be transferred from cell to cell, allowing for the spread of antimicrobial resistance. Integrons are genetic structures that can integrate within plasmids and transfer their genes into chromosomal DNA. This ability (characteristic) further optimizes the potential for gene transfer from one bacteria to the next. Based on the DNA sequence analysis of the plasmids in these studies, the antimicrobial resistance genes carried on the plasmids were consistent with the observed strain resistance; and in several instances, there were multiple genes associated with resistance to the same antimicrobial, thus indicating the complexity of resistance phenomenon. Likewise, one type of integron sequenced was unique, due to its large size and complexity, containing a surprisingly high number of resistance genes conferring resistance to the bacterium for at least six antimicrobials. The potential mobility and high number of resistance genes in integrons, and the plasmids where they often reside, can provide mechanisms for antimicrobial resistance to be transferred from resistant organisms to those that were not previously resistant, leading to an increased public-health concern. The results of these studies were recently published in PLoS One (7(12): e51160, doi:10.137/journal.pone.0051160) and Foodborne Pathogens and Diseases (doi: 10.1089/pd.2012.1455), and the DNA sequences from these studies have been deposited in the GenBank database to serve as a resource for other researchers.
For additional information, please contact Steven Foley, Ph.D., Division of Microbiology, FDA/NCTR.
Science Advisory Board Site Visit
On July 17-18, 2013, a subject-matter expert subcommittee of the NCTR Science Advisory Board (SAB) and their consultants conducted an in-depth review of the current research program and future plans of the Division of Microbiology. Representatives from each of FDA's Product Centers and the Office of Regulatory Affairs participated in the evaluation. The site-visit report will be presented at the full meeting of the NCTR SAB scheduled for December 10-11, 2013. The NCTR SAB advises the NCTR Director in establishing, implementing, and evaluating the Center’s research program.
For additional information, please contact Margaret Miller, Ph.D., Associate Director for Regulatory Activities, or Carl E. Cerniglia, Ph.D., Director, Division of Microbiology, FDA/NCTR.
Goldenseal, A Dietary Supplement
NCTR scientists in collaboration with the Shanghai Institute for Food and Drug Control (China) have shown that goldenseal and one of its major alkaloid constituents, berberine, are potent inducers of DNA damage in in vitro human cell cultures In cells treated with goldenseal, the extent of DNA damage was correlated to the berberine content and was directly associated with the inhibition of topoisomerase II (an essential enzyme for DNA replication). Goldenseal is an herbal product used to remedy a wide variety of ailments including gastrointestinal disturbances, urinary tract disorders, and inflammation; and was shown to increase liver tumors in rodents in a National Toxicology Program two-year carcinogenicity study. Toxicology Letters (2013, 221: 64-72).
For additional information, please contact Lei Guo, Ph.D., Division of Biochemical Toxicology, FDA/NCTR.
Sex Differences in Kidney Gene Expression
NCTR scientists have identified numerous genes that show sex-related expression differences in the kidneys, including a number of genes that code for urinary protein biomarkers (KIM-1, Clu, Tff3, and Lcn2) previously qualified by FDA for preclinical diagnostics of drug-induced kidney injury. This comprehensive in-life time-course study, which assessed the sex differences in global gene expression in the kidneys from untreated male and female rats, revealed a complex sexually dimorphic response. This response varied at multiple stages of development and may underlie sex-specific susceptibility to acute kidney injury and chronic kidney disease. The results of such studies could improve the use of current and future renal biomarkers and inform our assessments of kidney injury and disease. This study was supported by FDA's Office of Women’s Health and is published in Biology of Sex Differences(2013, 4:14).
For additional information, please contact Joshua Kwekel, Ph.D., Personalized Medicine Branch of the Division of Systems Biology, or James Fuscoe, Ph.D., Director, Personalized Medicine Branch of the Division of Systems Biology, FDA/NCTR.
Estrogenic Activity Database (EADB) for Assessing Potential Endocrine Activity
NCTR scientists in collaboration with FDA's CDER and CFSAN, U.S. Environmental Protection Agency, National Institute of Environmental Health Sciences, Texas A&M University, and Chang’an University (China) have developed the publicly available Estrogenic Activity Database (EADB) of in vitro and in vivo estrogenic activity data for a diverse set of over 8,000 chemicals, including drugs, food additives, and environmental chemicals. The EADB enables the user to search and evaluate the estrogenic activity of chemicals of interest as well as create models to predict the estrogenic activity of chemicals for which that data is not available. Estrogenic chemicals can interfere with the body’s normal estrogen signaling pathways and can have adverse effects. The development of the EADB and its applications are described in a recent publication in Toxicological Sciences.
For additional information, please contact Huixiao Hong, Ph.D., Bioinformatics Branch of the Division of Bioinformatics and Biostatistics, or Weida Tong, Ph.D., Director, Division of Bioinformatics and Biostatistics, FDA/NCTR.
Rapid, High-Sensitivity Detection of E. coli O157:H7
NCTR scientists have improved a rapid method (RAPID-B™) to identify E. coli O157:H7 with 92% accuracy and a detection sensitivity of one viable cell in 25 grams of raw spinach after a six-hour growth period. The improved method reduces background interference from naturally fluorescent food-matrix particles. This is done by utilizing a chemical photosensitizer (phloxine B) to photobleach the sample along with gradient centrifugation to separate target cells from matrix debris. The improved procedure outperformed the reference method—which includes RT-PCR, selective enrichment, and culture plating—in both accuracy and speed. RAPID-B™ is based on a rugged platform employing flow-cytometric principles that is readily field deployable. Food Microbiology (2013, 36: 416-425).
For additional information, please contact Jon Wilkes, Ph.D., Associate Co-Director, or Dan Buzatu, Ph.D., Associate Co-Director, Innovative Safety and Technologies Branch, Division of Systems Biology, FDA/NCTR.
Nanotechnology Training at NCTR
The second annual nanotechnology “Hands-On” training course was held at the NCTR/ORA Nanotechnology Core Facility on August 12-15, 2013. The training consisted of participants from the six FDA Product Centers, ORA and NCTR and introduced them to the most commonly used methods for characterization of nanoparticle size with an emphasis on their strengths and limitations, and included "hands-on" operation of the analytical equipment. The course was sponsored by the FDA Nanotechnology Task Force to continue its charge to identify and recommend ways to enable agency investigators to evaluate possible adverse effects from FDA-regulated products that use nanomaterials. The NCTR/ORA Nanotechnology Core Facility fulfills part of this mission by supporting FDA research and regulatory needs through proper equipment and trained personnel, and by conducting training courses that enable FDA scientists to understand nanomaterial characterization and detection methods.
For additional information, please contact Paul C. Howard, Ph.D., Director, Office of Scientific Coordination, FDA/NCTR, or Sean Linder, Ph.D., Arkansas Regional Laboratory, FDA.
Advances in Preclinical Models to Evaluate Anesthesia-Related Toxicity
NCTR scientists have published a review article in Molecular Neurobiologythat presents the use of human embryonic neural stem cells to detect anesthetic-induced neurotoxicity and to develop potential neuroprotection strategies. The review discusses stem cell biology with respect to the nervous system and neural development, the involvement of neural cell types in developmental neurotoxicity associated with anesthetic exposure, and the potential for imaging endogenous neural stem cells in vivo. The goal is to use neural stem cell models in conjunction with biochemical, pathological, imaging, and pharmacokinetic assessments to develop improved approaches to extrapolate preclinical data to the human.
For additional information, please contact Cheng Wang, M.D., Ph.D., Division of Neurotoxicology, FDA/NCTR.
Food Safety—Tools to Discriminate S. Enteritidis
Scientists from NCTR, the Center for Disease Control and Prevention, and the Arkansas Department of Health found all 60 Salmonella enterica serovar Enteritidis isolates obtained in 2010 from poultry houses (28) and clinics (32) were positive for 12 of the 17 virulence genes found mostly in S. enterica. The techniques used to identify the virulence genes included pulsed-field gel electrophoresis (PFGE) complemented with multiple-locus variable-number tandem repeat analysis (MLVA) and plasmid profiling. The similarities in virulence genotypes between poultry and clinical isolates indicate that the isolates from poultry are capable of causing human infection through contaminated shell eggs, which are the most common source for S. Enteritidis. The rate of salmonellosis in the United States has remained relatively steady over the past 10 years, and it has been difficult to discriminate causative strains using PFGE technology alone. These studies also demonstrate that PFGE supplemented with MLVA and plasmid profiling are important epidemiological tools to discriminate S. Enteritidis isolates during outbreaks. Food Microbiology, http://dx.doi.org/10.1016/j.fm.2013.08.003.
For additional information, please contact Ashraf A. Khan, Ph.D., Division of Microbiology, FDA/NCTR.
Can microRNAs Function as Biomarkers of Human Disease?
NCTR scientists have authored a chapter titled “Cell-free microRNAs as Biomarkers in Human Diseases” in the book microRNAs in Toxicology and Medicine (John Wiley & Sons, Inc.). MicroRNAs (miRNAs) are short, single-stranded, non-coding RNAs that regulate gene expression and thus influence a wide variety of basic cellular functions. In this chapter, the authors provide the following:
- an overview of the release of miRNAs from cells
- the stability of extracellular miRNAs present in human body fluids such as blood, urine, and saliva
- the role of extracellular miRNAs in cell-to-cell communications
- technical challenges in the analysis of extracellular miRNAs
- miRNAs potential use as biomarkers of human disease and injury
For additional information, please contact Xi Yang, Ph.D., Innovative Safety and Technologies Branch, Division of Systems Biology, FDA/NCTR.
2013 Global Summit on Regulatory Science (GSRS13)—Nanotechnology Focus
FDA/NCTR hosted GSRS13, the third annual Global Summit on Regulatory Science, wth a focus on Nanotechnology in Jefferson and Little Rock, Arkansas on September 10-11, 2013. The GSRS13 included roundtable discussions and speaker presentations, including FDA Commissioner Dr. Margaret Hamburg, that explored the applications and regulatory aspects of nanotechnology. The international conference provided an opportunity for scientists from government, industry, and academia from 10 countries to discuss innovative technologies and partnerships to enhance translation of basic science into regulatory applications within the global context.
For additional information, please contact William Slikker, Jr., Ph.D., Director, FDA/NCTR, or Paul C. Howard, Ph.D., Director, Office of Scientific Coordination, FDA/NCTR.
Global Coalition for Regulatory Science Research
FDA/NCTR hosted the First Meeting of the Global Coalition for Regulatory Science Research on September 10, 2013. The Coalition was established to foster the development of regulatory science through collaborative research and scientific exchange. After introductory remarks from FDA Commissioner Dr. Margaret Hamburg, Mary Lou Valdez (Associate Commissioner, International Programs) and Dr. William Slikker, Jr. (Director, NCTR), the Coalition participants engaged in roundtable discussions to elect a co-chair, develop operating procedures, determine the Coalition membership, and outline a strategic agenda for the development and sustainability of the Coalition.
For additional information, please contact William Slikker, Jr., Ph.D., Director, FDA/NCTR.
Expert Group Report on Anesthetic Neurotoxicity
An NCTR scientist coauthored an article outlining the results of the British Journal of Anaesthesia Salzburg Seminar on Anaesthetic Neurotoxicity and Neuroplasticity; an expert workshop focused on the review of current anesthetic research from animal and human studies and the future direction of the field. The article highlighted recent NCTR studies using positron emission tomography (PET) to characterize the time course of anesthetic-induced injury, as well as behavioral studies that explore the long-term cognitive consequences (learning and memory) of the pediatric anesthetic ketamine. The consensus was that although clinical data is indecisive, preclinical data demonstrates the potential adverse effects of anesthetics on neural development and function. Thus, it is imperative to gain a better understanding of the mechanisms of anesthetic-induced neurotoxicity to aid in the development of protective strategies. The report was published in the British Journal of Anaesthesia (2013, 111:143-151).
For additional information, please contact William Slikker, Jr., Ph.D., Director, FDA/NCTR.
Environmental Mutagenesis and Genomics Society Meeting
NCTR scientists presented their research results at the 44th Annual Meeting of the Environmental Mutagenesis and Genomics Society that introduced:
- an improved high-throughput methodology to evaluate chemical-induced chromosomal damage (in vitro micronucleus assay)
- a high-sensitivity methodology to quantify breast cancer gene mutations (ACB-PCR) which could improve individualized therapies
For additional information, please contact Robert Heflich, Ph.D., Division of Genetic and Molecular Toxicology, FDA/NCTR.