A supplement to the MLabs Handbook

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1 MLabs Spectrum A supplement to the MLabs Handbook June 2013 Volume 27, Number 2 Matrix-assisted laser desorption/ionization timeof-flight (MALDI-TOF) mass spectrometry: A new paradigm for diagnostic microbiology by Amanda O. Fisher-Hubbard M.D., House Officer IV, Department of Pathology and Duane W. Newton, Ph.D., D(ABMM), Associate Professor of Pathology & Director of Clinical Microbiology Laboratory Technological advances and their application in the clinical microbiology laboratory have contributed significantly to improved detection rates and decreased time-to-detection of important microbial pathogens. Technologies such as continuous-monitoring blood culture instruments, automated identification and susceptibility test systems, and platforms for the extraction, amplification, and detection of nucleic acid targets have shifted the paradigm defining routine approaches for pathogen detection and identification. Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry is a well-known technology in the clinical chemistry laboratory, but we will discuss how this technology is creating a paradigm-shifting revolution for clinical microbiology. BACKGROUND The primary goal of the clinical microbiology laboratory is to identify causative agents of infectious diseases in patient specimens. Traditionally in bacteriology, mycology, and mycobacteriology, this is achieved by first isolating an organism in culture and using a variety of different features of the organism to classify it. These features include macroscopic and microscopic appearances, growth characteristics, and results of various biochemical tests. Identification procedures can often be timeconsuming and cumbersome, as they generally require sub-cultures and multiple incubation steps. Efforts have been made to streamline and abbreviate this process by applying alternate technologies, including immunoassays, automated identification systems, and nucleic acid-based methodologies. MALDI-TOF mass spectrometry has recently emerged as an important, rapid, and accurate tool for microbiological identification, with a substantial upsurge of publications on the topic in the past decade (Figure 1). TECHNOLOGY All mass spectrometers consist of three main components an ion source to ionize and subsequently transfer ions into a gas phase, a mass analyzer to separate particles based on their mass-to-charge ratio (m/z), and a detector (Figure 2). MALDI is considered a soft ionization technique, meaning that DEPARTMENT OF PATHOLOGY in this issue 1 Matrix-assisted laser desorption/ionization timeof-flight (MALDI-TOF) mass spectrometry: A new paradigm for diagnostic microbiology 3 Spotlight on Duane W. Newton, Ph.D., D(ABMM) 6 Test Updates New Tests ATP7B Gene Sequencing MEF2C Gene Sequencing Test Methodology, Reference Range, and Specimen Handling Changes Hantavirus Antibodies Hexosaminidase A and Total Natural Killer Cell Functional Assay Parathyroid Hormone, Intact Polychlorinated Biphenyls Vitamin K1 Discontinued Tests Ketones, Serum Warfarin Sensitivity Analysis 8 MLabs News PECOS Enrollment Required for Laboratory Referrals MLabs Participating with GWH- Cigna U-M Department of Pathology News

2 Figure 1. Publications on MALDI applications in microbiology. A PubMed search of all articles containing both keywords MALDI and microbiology was performed and the number of articles was plotted by year of publication. it is gentler than some methods and does not result in the breakage of chemical bonds. It uses an acidic crystal matrix, in which analyte molecules are embedded, or co-crystallized, and spotted onto a target plate. The matrix is chosen based on properties such as ease of sublimation and wavelength of absorbance. A laser then irradiates a spot on the sample plate, which results in excitation of the matrix and ejection (desorption) of the analyte molecules surrounded by matrix. The analyte molecules then become ionized and enter the gas phase in which they are accelerated across an electric field within the ionization chamber. Next, the molecules enter the mass analyzer. With time-offlight (TOF) mass analyzers, the TOF of an ionized molecule is proportional to the mass-to-charge ratio of the molecule. Analytes with different mass-to-charge ratios are therefore separated based on their time of flight [1-2]. MALDI mass spectrometry has traditionally been used in a variety of life science fields to analyze and identify biomolecules such as proteins and sugars. The use of mass spectrometry for identification of microorganisms was first described in 1975 [3-4]. The technology was employed to study biomarker profiles of certain bacterial species. The initial experiments were plagued by problems with reproducibility attributed to growth conditions and media. Further difficulties were related to the detection of ribosomal and membrane proteins released from the bacterial cells. At that time, it was challenging to isolate those proteins without their destruction, thus prohibiting 2 U-M Department of Pathology analysis of their profiles and attainment of mass pattern spectra [1, 4]. The use of MALDI in the field of microbiology made its resurgence in the mid-1990s, when soft ionization techniques were applied. These permitted the analysis of the larger, more fragile, organic molecules which were often damaged by older, conventional methods. In 1996, the first MALDI-TOF mass spectrometry experiment successfully identified bacteria from whole colonies based on protein content. Additionally, use of alternative matrices allowed for ionization primarily of ribosomal proteins. These proteins tend to be better conserved than surface proteins and thus provided a more reliable indicator of the identity of specific bacterial species [1]. APPLICATIONS TO MICROBIOLOGY Use of MALDI-TOF in the clinical microbiology laboratory requires culture isolation of an organism (bacteria, mycobacteria, or fungi) from a clinical specimen (Figure 3). Once a cultured isolate (e.g. a bacterial colony) can be picked off of a plate, the isolate can then be mixed with a matrix, applied to a metal plate, and allowed to dry. Generally, the plates can hold between 16 and 384 samples. The samples are then loaded into the mass spectrometer and subjected to laser pulses, which leads to desorption of the ionized, high-abundance, soluble proteins (including ribosomal and heat shock proteins). These ionized molecules are accelerated to the flight tube by an electromagnetic field. The TOF of the analytes to the detector is then measured and produces a characteristic spectrum from which an organism can be identified. This is accomplished by comparing the resultant spectrum to a database that contains the spectral patterns of various clinically-relevant organisms. Each result is given a score value based its similarities to the stored data. These score values indicate the strength of the identification, including whether the identification is considered valid to the species and/or genus level. Target plates are then removed from the analyzer and are either discarded or washed for reuse, depending on which vendor platform is used [1, 4]. MALDI-TOF mass spectrometry is suitable for the microbiology laboratory for several reasons, including ease, speed, and analytical performance (Table 1). Samples require minimal preparation, as the whole bacterial cell is analyzed. Starting from a colony of a positive culture, identification can often be made in less than one minute.

3 This technology also has excellent specificity, low operating cost, and minimal maintenance needs. Other than the instrument itself, the only requirements are growth medium and an appropriate matrix. Traditional, routine biochemical tests can essentially be replaced by MALDI- TOF. The database is continuously updated so that both the more common clinically-relevant organisms, as well as less common, more difficult to identify organisms, are included [1]. There are, however, drawbacks to MALDI-TOF mass spectrometry (Table 1). Currently, intact colonies from pure cultures are required for analysis, which may take time for slow-growing organisms. Although databases of organisms are constantly improved, they are certainly not complete. Due to proteomic similarities, there can be difficulty differentiating between closely-related organisms (e.g. Escherichia coli and Shigella sp.; Streptococcus pneumoniae and S. mitis group). This lack of discriminative power is Figure 2. Principle of MALDI-TOF mass spectrometry. For MALDI-TOF, laser impact causes thermal desorption of ribosomal proteins of bacteria embedded in matrix material and applied to the target plate (analytes shown as red, yellow, and blue particles; matrix shown in green). In an electric field, ions are accelerated according to their mass and electric charge. The drift path allows further separation and leads to measurable differences in time-of-flight of the desorbed particles that are detected on top of the flight tube. From the time-of-flight, a characteristic spectrum is produced (adapted from Reference 1). Spotlight on Duane W. Newton, Ph.D., D(ABMM) Duane W. Newton, Ph.D., D(ABMM), Associate Professor of Pathology, is Director of the MLabs Clinical Microbiology Laboratory at the University of Michigan. Son of a career Air Force officer, Dr. Newton s family criss-crossed the U.S. and Europe and, although home is where the Air Force sends you, spent his formative years in Dayton, Ohio. Dr. Newton received a B.S. in Biology (1988) and a Ph.D. in Biology with an emphasis in Microbiology and Immunology (1993) from the University of Dayton, Ohio. His dissertation work was completed under the tutelage of Robert Kearns, Ph.D., a long-time mentor and friend. Dr. Newton completed his post-doctoral training at the University of Tennessee Medical School and the VA Medical Center in Memphis, Tennessee. He studied host and pathogen factors influencing severity of invasive disease caused by Streptococcus pyogenes ( flesh-eating disease ). This was followed by clinical training, where he was selected in 1998 for a Clinical and Public Health Microbiology Fellowship at the University of Rochester Medical Center, Clinical Microbiology Laboratories in Rochester, New York. Upon completion of his fellowship in 2000, Dr. Newton was appointed as Manager of the Virology/ Immunology Section of the Department of Community Health, Division of Infectious Diseases, Bureau of Laboratories for the State of Michigan and as Adjunct Assistant Professor for the Michigan State University Medical Technology Program. In 2002, he joined the faculty of the Department of Pathology at the University of Michigan Medical School. Dr. Newton was named Director of Microbiology/Virology Laboratories in 2003, and received his certification as a Diplomate by the American Board of Medical Microbiology in Dr. Newton has been very active and visible in the clinical, administrative, academic and educational arenas within the UMHS and far beyond! He has led many important initiatives including UMHS readiness and response to annual influenza epidemics as well as other high profile infectious disease outbreak challenges. Among Dr. Newton s more than 40 peer-reviewed publications and 70 invited external presentations are his particularly important contributions in the area of cost-effective practice of clinical microbiology and the application of leading edge technology to microbiology practice. He is also very active in numerous leading regional and national professional organizations as a thought leader, journal referee and educational policy leader. MLabs Spectrum 3

4 also related to the absence of sufficient reference spectra in the databases. Additionally, certain growth media can preclude an accurate identification [1, 4-5]. Though many of these issues are becoming less problematic as the technology improves, culture is still necessary for antimicrobial susceptibility testing [1, 4]. Table 1. Benefits and limitations of MALDI-TOF mass spectrometry for identification of microorganisms a Benefits Low cost/test High specificity Minimal sample prep Rapid time-to-result High-throughput Potential direct patient sample applications Limitations High up-front instrument costs Current databases are incomplete Proteomic similarities among organisms Lack of quantification Large inoculum required Maintenance - duration/downtime can be long Growth medium can affect identification Lack of ability to perform susceptibility testing a adapted from Reference 1 OUR EXPERIENCE There are currently two commercially-available MALDI- TOF instruments in the U.S. for the identification of microorganisms: MALDI Biotyper (Bruker Daltonics, Billerica, MA) and VITEK MS (biomérieux, Durham, NC). The main difference between the systems is that the Biotyper utilizes reusable steel plates for spotting of isolates for analysis and the VITEK MS uses disposable plates. Neither system has yet received FDA-approval, but their submissions are currently under review. The MALDI Biotyper has been in use by the MLabs Clinical Microbiology Laboratory since May, It was selected primarily because it was the sole instrument available at the time of implementation but also because it was the most studied in the literature. Extensive method verification was performed prior to implementation with several hundred isolates previously characterized by conventional or sequencebased methods tested by MALDI-TOF. Organism groups evaluated initially included gram-positive and gram-negative aerobic bacteria and yeast. We have subsequently added aerobic actinomycetes, mycobacteria, and non-fermenting gram-negative bacilli to our list of reportable organisms, and are completing our evaluation of anaerobic bacteria. This technology has performed very well, resulting, for example, in improved identification for nonfermenting gram-negative bacilli (including isolates from Cystic Fibrosis patients) which have historically been difficult to identify using commercial automated identification systems, as well as reduced time to identification for slow-growing organisms (e.g. aerobic actinomycetes and Mycobacterium sp.). There are, however, recognized technical challenges with MALDI-TOF that can impact workflow. Organisms with more robust cell walls (gram-positives, mycobacteria, yeast) may require more rigorous and manual extraction procedures in order to obtain acceptable identification scores. Our initial procedure for gram-positives and yeasts is to overlay formic acid ( plate extraction ) onto the organism spot prior to the addition of matrix. If an acceptable score is not obtained, organisms are subjected to an extensive offline procedure ( tube extraction ), and the resulting extract is spotted for analysis. This tube extraction is routinely required for aerobic actinomycetes and mycobacteria, and while laborintensive, batch processing reduces this burden and still substantially decreases the time-to-identification of these organisms by weeks. When implementing this technology into the routine workflow of the laboratory, we initially began with the positive blood cultures, using MALDI-TOF to identify all yeast, as well as aerobic gram-positive and gramnegative bacteria subcultured from positive bottles. The technologist was responsible for spotting the plate, performing the analysis, and reporting the results in the laboratory information system. When we expanded the use of MALDI-TOF to include identification of verified organism groups from all other sources, a traffic jam was created by technologists from each bench waiting for their opportunity to use the instrument. We changed the workflow such that each bench spotted their own plates, but those plates were transferred to the MALDI bench for analysis on the instrument by a technologist dedicated to this task. Results were then returned back to each technologist for reporting at their specific bench. 4 U-M Department of Pathology

5 Transitioning to this technology has substantially reduced time-to-identification and expenses compared to our previous systems (primarily VITEK 2, biomérieux), but other advantages have also been recognized. We have recently completed a study in collaboration with our institutional Antimicrobial Stewardship Team that also demonstrated clinically-significant benefits to our patients and health system overall [6]. The study compared patients with bloodstream infections caused by gram-positive bacteria, gram-negative bacteria, and yeast during two equivalent time periods: a three month period with standard practice before MALDI-TOF implementation and a three month period following MALDI-TOF implementation during which there was active intervention by a clinical pharmacist at each point of newly available clinical information (initial Gram stain, MALDI identification, susceptibility report). Not only were microbiology reporting times quicker, but patients in the intervention phase had significantly shorter times to placement on optimal antimicrobial therapy, had significantly reduced lengths-of-stay in intensive care units, and had significantly reduced mortality compared to patients in the pre-implementation phase [6]. NEW APPLICATIONS As MALDI-TOF still relies on culture isolates for analysis, a great deal of emphasis has been placed on adapting the technology for used on direct patient specimens. Recent studies have shown that MALDI-TOF can be used to identify organisms from positive blood culture bottles and shows promise when applied to direct urine samples, although polymicrobial infections still present a problem [1, 7-8]. MALDI-TOF also has the ability to detect some antibiotic resistance mechanisms, including differentiation between methicillin-susceptible and methicillin-resistant Staphylococcus aureus, detection of ß-lactamases and carbapenemases, and the identification of vancomycinresistant Enterococcus spp. [9-10]. In addition, virulence factors can be detected by examining the presence or absence of certain spectral peaks [1]. Finally, MALDI- TOF has shown promise as a tool for both taxonomic classification and strain typing of organisms, potentially supplanting more cumbersome and labor-intensive methods routinely used [11]. CONCLUSIONS Development and utilization of novel technologies has continued to generate an ever-changing landscape for diagnostic laboratories. Although this dynamic environment can be a burden emotionally, operationally, and financially, a willingness and ability to embrace these changes can present exciting opportunities to improve operations and have significant impacts on patient care. MALDI-TOF has not only provided the MLabs Clinical Microbiology Laboratory the opportunity to improve laboratory efficiency, but by partnering with clinicians to improve utilization of this information, we have positively impacted patient care. This integration of rapid reporting and active intervention on results will likely create the next paradigm shift for clinical laboratories as the availability of rapid detection/identification technologies increase. REFERENCES 1. Lavigne JP, Espinal P, Dunyach-Remy C, Messad N, Pantel A, Sotto A. Mass spectrometry: a revolution in clinical microbiology? Clin Chem Lab Med. 2013; 51: Sauer S, Kliem M. Mass spectrometry tools for the classification and identification of bacteria. Nat Rev Microbiol. 2010; 8: Anhalt JP, Fenselau C. Identification of bacteria using mass-spectrometry. Anal Chem. 1975; 47: Figure 3. Workflow of isolate identification. Subcultured colonies are applied on a clean target (a). Samples are overlaid with matrix solution and air dried (b). Measurement in the MALDI-TOF mass spectrometer is conducted (c); an analyte-specific mass spectrum is obtained (d); analyte identification is performed through automatic matching of the generated mass spectrum with spectra in the database (e). (With kind permission from Springer Science+Business Media: Reference 4, Fig. 2, Springer-Verlag 2011) MLabs Spectrum 5

6 4. Wieser A, Schneider L, Jung J, Schubert S. MALDI-TOF MS in microbiological diagnosticsidentification of microorganisms and beyond (mini review). Appl Microbiol Biotechnol. 2012; 93: Bizzini A, Greub G. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry, a revolution in clinical microbial identification. Clin Microbiol Infect. 2010; 16: Huang AM, Newton DW, Kunapuli A, Gandhi TN, Washer LL, Isip J, Collins CD, Nagel JL. Rapid Organism Identification via Matrix-Assisted Laser Desorption Ionization Time-of-Flight Combined with Antimicrobial Stewardship Team Intervention Decreases Mortality and Improves Time to Clinical Response in Adult Patients with Bacteremia and Candidemia. Clin Infect Dis 2013; submitted. 7. Lagacé-Wiens PR, Adam HJ, Karlowsky JA, Nichol KA, Pang PF, Guenther J, Webb AA, Miller C, Alfa MJ. Identification of blood culture isolates directly from positive blood cultures by use of matrixassisted laser desorption ionization-time of flight mass spectrometry and a commercial extraction system: analysis of performance, cost, and turnaround time. J Clin Microbiol. 2012; 50: Ferreira L, Sánchez-Juanes F, González-Avila M, Cembrero-Fuciños D, Herrero-Hernández A, González-Buitrago JM, Muñoz-Bellido JL. Direct identification of urinary tract pathogens from urine samples by matrix-assisted laser desorption ionization-time of flight mass spectrometry. J Clin Microbiol. 2010; 48: Edwards-Jones V, Claydon MA, Evason DJ, Walker J, Fox AJ, Gordon DB. Rapid discrimination between methicillin-sensitive and methicillinresistant Staphylococcus aureus by intact cell mass spectrometry. J Med Microbiol. 2000; 49: Hrabák J, Chudácková E, Walková R. Matrixassisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry for detection of antibiotic resistance mechanisms: from research to routine diagnosis. Clin Microbiol Rev. 2013; 26: Murray PR. Matrix-assisted laser desorption ionization-time of flight mass spectrometry: usefulness for taxonomy and epidemiology. Clin Microbiol Infect Sep;16: U-M Department of Pathology Test Updates New Tests atp7b gene SEQUENCING The MLabs Molecular Genetics Laboratory began sequencing the ATP7B gene (OMIM: ) effective March 25, Mutations in ATP7B are associated with Wilson disease (OMIM:277900), an autosomal recessive disorder characterized by excessive copper accumulation in the body, mainly in the liver and brain (De Bie et al. J Med Genet 44: , 2007). ATP7B encodes a plasma-membrane copper-transporting ATPase that is suggested to be involved in exporting copper from cells (Tanzi et al. Nature Genet 5: , 1993). Accumulation of toxic levels of copper causes progressive liver disease and degenerative changes in the basal ganglia. Early detection of Wilson disease, prior to irreversible tissue damage, is important for a successful therapy (Brewer et al. J Lab Clin Med 132: , 1998). mef2c gene sequencing The MLabs Molecular Genetics Laboratory began sequencing the MEF2C gene (OMIM: ) effective March 25, Heterozygous mutations leading to haploinsufficiency of the MEF2C gene have been associated with intellectual disability, stereotypic repetitive movements, and epilepsy, with or without cerebral malformations (OMIM:277900; Zweier et al. Hum Mutat 31(6):722-33, 2010). The MEF2C gene encodes a transcription factor that is suggested to have a crucial function in the homeostatic control of activity-dependent synaptogenesis, which plays a role in the establishment of functional neuronal circuits during development and memory storage (Barbosa et al. Proc Nat Acad Sci 105: , 2008). Test Methodology, Reference Range, and Specimen Handling Changes hantavirus antibodies Please note that the reference range for Hantavirus Antibodies, IgG and IgM has been revised as follows effective January 10, 2013: Reference Range: <2.00

7 HEXOSAMINIDASE A AND TOTAL Effective February 21, 2013, the reference ranges and units of measure for the Hexosaminidase A and Total assays have been updated as follows: HEXOSAMINIDASE TOTAL, LEUKOCYTES 15 yrs 16 yrs 20 nmol/min/mg nmol/min/mg HEXOSAMINIDASE PERCENT A, LEUKOCYTES 15 yrs 16 yrs 20-80% of total 63-75% of total HEXOSAMINIDASE TOTAL, SERUM 15 yrs 16 yrs: 20 nmol/min/ml nmol/min/ml HEXOSAMINIDASE PERCENT A, SERUM 15 yrs 20-90% of total 16 yrs: 56-80% of total natural killer cell functional assay Focus Diagnostics has discontinued their Natural Killer Cell Functional assay effective January 23, Requests for this test will be sent to Cincinnati Children s Hospital. Collection Instructions: Collect sufficient specimens in green top (sodium heparin) tubes to obtain 10 ml of whole blood. Send intact specimens at room temperature. Do not refrigerate or freeze. Specimens are accepted by MLabs Monday through Thursday only and must be received by performing laboratory within 24 hours of collection. PARATHYROID HORMONE, INTACT Effective January 8, 2013, Parathyroid Hormone (PTHI) testing was moved from Special Chemistry to Automated Chemistry. The methodology remains the same - Immunometric Chemiluminescense. The preferred specimen type has changed to SST (serum); only one collection tube is necessary. PTHI testing will combine with other automation line testing in a single collection tube. EDTA collection will be accepted, but an SST tube will need to be drawn for other automation line testing and the PTHI calcium determination. Collection Instructions: Collect specimen in an SST tube. Centrifuge, aliquot 0.5 ml (minimum 0.3 ml) of serum into a plastic vial and refrigerate up to 48 hours or freeze for longer storage. EDTA plasma is also acceptable. If sending EDTA plasma, include a copy or note on the requisition the patient s calcium result performed on a specimen collected within the previous 48 hours OR send 0.5 ml serum, refrigerated, for calcium level to be performed by MLabs. Clearly mark aliquots plasma or serum as appropriate. Add-on test requests for previously processed in-lab samples will be honored for 24 hours. Polychlorinated biphenyls Effective February 11, 2013, Polychlorinated Biphenyls (Aroclors 1254 and 1260) referred to MedTox Laboratories has been replaced by Polychlorinated Biphenyls (Congeners/Aroclors). Collection Instructions: Collect specimen in a red top or SST tube. Centrifuge, aliquot 3 ml of serum into a plastic vial and refrigerate. VITAMIN K1 Effective April 3, 2013, Vitamin K1 serum testing is performed by Mayo Medical Laboratories New England using a Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) test methodology. Collection Instructions: Patient should abstain from alcohol for 24 hours prior to specimen collection. Collect specimen in a red top (preferred) or SST tube from a fasting patient (12-hour fast); do not use SST tube. Centrifuge, aliquot serum into a plastic vial and refrigerate. Protect specimen from light. Reference Range: ng/ml Discontinued Tests ketones, serum Effective February 5, 2013, the MLabs Chemical Pathology Laboratory has discontinued the semiquantitative serum ketones assay. The quantitative Beta Hydroxybutyrate assay is recommended for diagnosis and monitoring of therapy for diabetic ketoacidosis. WARFARIN SENSITIVITY ANALYSIS Effective January 23, 2013, the MLabs Molecular Diagnostics Laboratory no longer offers Warfarin Sensitivity Analysis (CYP2C9 and VKORC1 genotypes). Requests for this test will be sent to Mayo Medical Laboratories. MLabs Spectrum 7

8 Technical Editor: Jeffrey Myers, M.D. Managing Editor: Deirdre Fidler For additional clarification concerning any of the information contained in this Spectrum, please contact the MLabs Client Services Center at (local) or Address correspondence to: MLabs Spectrum PO Box 976 Ann Arbor, MI To keep our Spectrum circulation records accurate and up to date, please send any name or address changes, corrections, additions or deletions to the address listed above. Thank you. Executive Officers of the University of Michigan Health System: Ora Hirsch Pescovitz, M.D., Executive Vice President for Medical Affairs; James O. Woolliscroft, M.D., Dean, U-M Medical School; Douglas Strong, Chief Executive Officer, U-M Hospitals and Health Centers; Kathleen Potempa, Dean, School of Nursing The Regents of the University of Michigan: Mark J. Bernstein, Julia Donovan Darlow, Laurence B. Deitch, Shauna Ryder Diggs, Denise Ilitch, Andrea Fischer Newman, Andrew C. Richner, Katherine E. White, Mary Sue Coleman (ex officio). The University of Michigan, as an equal opportunity/affirmative action employer, complies with all applicable federal and state laws regarding nondiscrimination and affirmative action. The University of Michigan is committed to a policy of equal opportunity for all persons and does not discriminate on the basis of race, color, national origin, age, marital status, sex, sexual orientation, gender identity, gender expression, disability, religion, height, weight, or veteran status in employment, educational programs and activities, and admissions. Inquiries or complaints may be addressed to the Senior Director for Institutional Equity, and Title IX/Section 504/ADA Coordinator, Office of Institutional Equity, 2072 Administrative Services Building, Ann Arbor, Michigan , , TTY For other University of Michigan information call , The Regents of the University of Michigan. MLabs News pecos enrollment required for laboratory referrals Effective May 1, 2013, the Centers for Medicare & Medicaid Services (CMS) will turn on edits to deny claims that fail ordering or referring provider edits. This means that Medicare will deny claims for laboratory and pathology services if the ordering clinician is not in Medicare s enrollment records. Additional information and instructions regarding PECOS ordering and referring can be found at MedicareProviderSupEnroll/MedicareOrderingandReferring.html. Please verify that you have a current Medicare enrollment record that contains your NPI. If you order laboratory services for Medicare beneficiaries and you do not have a Medicare enrollment record, you need to submit an enrollment application to Medicare. You can do this using the Internet-based Provider Enrollment, Chain, and Ownership System (PECOS) or by completing the paper enrollment application. MLABS PARTICIPATING with gwh-cigna Effective December 1, 2012, the University of Michigan Health System is an innetwork provider with GWH-Cigna (formerly Great West Healthcare) through the HAP Preferred network. Note that GWH-Cigna utilizes the same plan authorization requirements as Cigna. u-m department of pathology news Tomasz Cierpicki, Ph.D., Assistant Professor of Pathology, is the senior author of a plenary publication in the November 29, 2012 issue of Blood entitled Structural insights into inhibition of the bivalent menin-mll interaction by small molecules in leukemia. A figure related to the paper was also selected for the cover. For a complete list of authors and more information see Joel K. Greenson, M.D. is co-author and editor of the recently published Morson and Dawson s Gastrointestinal Pathology, 5th edition. Congratulations to Jolanta Grembecka, Ph.D., Assistant Professor of Pathology, who has been awarded a five year Lymphoma and Leukemia Society Scholar Award effective July 1, Dr. Grembecka s research focuses on the development of small molecule-based targeted therapies for acute leukemia. David Lombard, M.D., Ph.D., Assistant Professor of Pathology, is a senior author of The Histone Deacetylase SIRT6 Is a Tumor Suppressor that Controls Cancer Metabolism, which was published in the December 7, 2012 issue of Cell. Other co-authors from the Department include Bernadette Zwaans, M.S., a Molecular and Cellular Pathology student, and Joel Greenson, M.D., Professor of Pathology. The work was done in collaboration with senior author Raul Mostoslavsky, M.D., Ph.D., and colleagues at Massachusetts General Hospital. A recent publication in Nature Genetics from the Michigan Center for Translational Pathology (MCTP) by Dan R. Robinson, Ph.D. (first author) and Arul Chinnaiyan, M.D., Ph.D. (senior author) and others, reports recurrent NAB2-STAT6 gene fusions in 100% of solitary fibrous tumor cases. For more information see edu/news/genetic-mutation-for-cancer-2013.shtml. 8 U-M Department of Pathology