MEDICAL

Dr. Judith Isaac-RentonPublic Health Laboratory Director, Program Head, Environmental Microbiology and Parasitology

Dr. Mel KrajdenProgram Head, Virology Services

Dr. Linda HoangProgram Head, Bacteriology and Mycology

Dr. Patrick TangProgram Head, Mycobacteriology/TB and Molecular Services

Dr. Muhammad MorshedProgram Head, Zoonotic Diseases and Emerging Pathogens

Dr. Martin PetricClinical Virologist

LABORATORY

Kathy KelemenMulti-Site Operations Director

Amelia TrinidadChief Technologist, BCCDC and Central Pro-cessing and Receiving Laboratory Operations

Anita KwongDirector, Laboratory Quality and Process Improvement

Yin ChangLaboratory Surveillance and Outbreak Coordinator

Kim MacDonaldLaboratory Liaison Technical Officer

PROGRAMS

Neil ChinPublic Health Lead, Biosafety, Biosecurity and Biohazard Containment

John ChanSection Head, Technical Support and Pre-Analytical, Central Processing & Receiving

Joe FungSection Head, Environmental Microbiology

Annie MakAnalytical Section Head, High-Volume Serology, Central Processing & Receiving

Alan McNabbSection Head, Molecular Servicesand Virology Program

Ana PaccagnellaSection Head, Bacteriology and Mycology

Dr. Mabel RodriguesSection Head, Mycobacteriology/TB

Quantine WongSection Head, Parasitology/Zoonotic Diseases and Emerging Pathogens

PUBLIC HEALTH MICROBIOLOGY & REFERENCE LABORATORY

NEWS OCTOBER 2010

As Dr. Petric notes in the Virology article on this page, we took part in some significant events this past year. In this, our first Newsletter since last Spring, I would like to first acknowledge all staff at the Public Health Microbiology & Reference Laboratory (PHMRL) at the BCCDC, who pulled efficiently and effectively, on behalf of the residents of British Columbia. Secondly, I would like to also acknowledge our medical microbiology partners throughout BC. You pulled along side us, from your own laboratory sites, during the Olympics and the pandemic. Our team salutes yours!

Hopefully we won’t see any comparable major events this year. Advances in technology, however, may offer opportunities to work together, hopefully with a paced and well planned approach. Genomics, built on our molecular microbiology network, could be one of the next opportunities to improve patient and population health. Education is our focus for a start; we plan a follow up session to last year’s Microbiology for Public Health/Reference Labs Lecture Series with one on Genomics for labs. Let us know if you are interested in video-conferencing with us.

In the Shadow of Pandemic Influenza -Virology Program Activity, Spring and Summer

The arrival of pandemic influenza made 2009 a very challenging year for the Virology (VI) Program and many of the other Programs as well at the Public Health Microbiology & Reference Laboratory. The end of the pandemic in early 2010 was followed by the excitement of the Olympic & Paralympic Winter Games which, despite extensive preparation, did not provide much of a demand on the laboratory system with both the athletes and spectators enjoying unprecedented good health. However, in the aftermath of the winter games BC experienced an outbreak of measles between March and May. The virus was diagnosed in patient specimens both by serology, which included the detection of measles specific IgM class antibody and by nucleic acid-based testing. The RT-PCR assay was validated with the assistance of the National Microbiology Laboratory (NML) and implemented at the PHMRL. This allowed the Virology Laboratory to diagnose the virus from respiratory specimens and urines with a turn-around time of 24 hours from receipt of specimen. Sequence analysis of the virus genome by the NML established that it was predominantly genotype H1, which is commonly present in Asia. Cases were detected across the province with the majority from Fraser Health and Vancouver Coastal Health authorities. In total 74 laboratory-confirmed cases were diagnosed.

In late April the laboratory began detecting enteroviruses from cerebrospinal fluid specimens. Over the spring and summer 69 cases of enterovirus infection have been diagnosed at a level well above what had been experienced in previous years. Enteroviruses diagnosed mainly in cerebrospinal fluid specimens included 54 cases of Coxsackievirus A9, 13 cases of Echovirus 30 and 1 case of Echovirus 14. The viruses were isolated in cell culture and typed by immunofluorescence microscopy. An RT-PCR assay for the detection of enteroviruses from the clinical specimen was validated with the help of the BC Children’s Hospital Microbiology Laboratory. In addition a genomic sequence based typing has been implemented and validated.

A Message from the Public Health

Laboratory Director

West Nile Virus in 2010: A Quiet Year in BC

TB Laboratory imPROVEs Processes

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The Zoonotic Diseases and Emerging Pathogens (ZEP) Program reported the arrival of West Nile virus (WNv) in the October 2009 issue of this newsletter. More human cases were expected to be detected in 2010 as well as more positive mosquito pools and corvids (crows and relatives of crows). Surprisingly, this didn’t happen this year, most probably because of weather conditions.

In the 2010 season, ZEP tested over 2,078 mosquito pools. One pool consists of up to 50 pre-identified (in Parasitology) Culex spp mosquitoes. Insect RNA is extracted and the genetic material tested using real time PCR (for WNv RNA fragments). All pools tested this year were negative for WNv. The British Columbia (BC) Animal Health Centre, however, found 5 WNv positive corvids (233 tested); ZEP confirmed these results. At the end of the season only one BC endemic WNv human case was detected. The question is why?

As part of their surveillance and research activities, the WNv Team (Epidemiology and Labs) at the BC Centre for Disease Control closely monitors environmental trends (rainfall, temperature, humidity etc). The 2010 season was cooler and drier compared to the 2009 season. Last year, Culex tarsalis was found to be a carrier of WNv in BC but and populations of C. tarsalis in the Okanagan in 2010 were far lower this year. Mosquitoes require ideal environmental conditions (temperature, rainfall) for breeding as well as for the successful spread of WNv to humans.

The WNv story on the East Coast continues to be markedly different than that in BC. This year, the virus caused some concern in New York again (it was first detected in 1999). There were 103 (72 neurological and 31 non-neurological) cases, including 3 deaths. There were also 5 presumptive viremic blood donors, indicating that WNv continues to have a significant presence in that region.

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SNIPPETS FROM THE YEAR

The Mycobacteriology/Tuberculosis (TB) Laboratory carried out a PHSA imPROVE event early this year. Established in 2007 in PHSA, imPROVE is a Health Authority-wide program for improvement based on principles of the Toyota Production System. Staff are engaged to look at enhancing processes by decreasing waste. For PHSA and the public health laboratory, where service demands continue to increase in both volume and complexity (more molecular tests), a strategy for increasing capacity as well as for improving quality for patient care (faster, better tests), is needed.

The focus of the imPROVE Rapid Process Improvement Workshop (RPIW) in the TB Laboratory in April, 2010 was optimization of pre-analytical processes. Some of the outcomes included:

• Using smaller batch sizes to improve the flow of samples; • Redesigning the laboratory to improve efficiency of accessioning, including removing unused equipment and rearrangement of work stations; and, • Establishing dedicated benches with much less set up and restocking time needed.

All of these improvements resulted in reduction in pre-analytical time (from sample receipt in the laboratory to mycobacteriology culture) required. This translated to attention on other needed challenges such as increasing the molecularization of TB testing.

As with any RPIW, the challenge is not necessarily to maintain these changes but to continuously reevaluate practices and strive towards further optimization. The TB Laboratory is now carrying out a second imPROVE event. The objective of this second TB Laboratory RPIW, October, 2010 is to improve work flows for increased molecular testing capacity.

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RPIW participants (left to right): Dr. J. Isaac-Renton, L. Olaer, T. Kim, D. Johal, A. Trinidad, Dr. M. Rodrigues, P. Tsang, B. Abbott, Dr. P. Tang, N. Di Tomaso, and M.

Outbreak Activity Update

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Botulism is a potentially life-threatening disease. In adults it is caused by ingestion of preformed toxin. Within hours of consuming contaminated foods, patients most often present with cranial nerve dysfunction and rapidly descending, symmetrical muscle paralysis. Botulism Immune Globulin must be considered and given quickly. Notification by local public health workers will ensure rapid follow up on implicated food sources, if a potential case is identified. The Environmental Microbiology (EM) Program (Medical Microbiologist) must approve all botulism testing (bioassay).

Planning for possible events during the 2010 Winter Olympics and Paralympics, it became clear that expanding the Canadian Laboratory Response Network (CLRN) capacity for botulism testing was important. Partnering with the Public Health Agency’s Bureau of Microbial Hazards and the National Microbiology Laboratory, staff in the EM Program are now developing new molecular and immunological tests to screen foods potentially contaminated with the potent toxins of Clostridium botulinum. Mr. Joe Fung, Section Head, presented a summary of the British Columbia experience at a recent national conference: most cases in BC are infant botulism. See our Guide to Programs and Services for procedures if a case is suspected.

Botulism: A Public Health Emergency

STAY TUNED.

In addition to the measles outbreak mentioned earlier, this summer has also been a busy one for outbreaks and clusters of enteric illness. An outbreak of a rare type of Salmonella occurred in June and into July due to exposure to one brand of contaminated “head cheese”. The food source was determined through epidemiological investigations. This incident resulted in 30 cases of S. Chester, found mainly in elderly persons in Fraser Health and Interior Health Authorities; some other cases were also found throughout the province. The outbreak resulted in the voluntary recall of contaminated food product and investigation of other products manufactured in the same plant.

This outbreak is an example of the staff in the Bacteriology and Mycology (BAM) Program noticing firstly, that there were a number of cases of an unusual type of Salmonella, then applying Pulsed Field Gel Electrophoresis (PFGE) and finding a similar pattern amongst the positive specimens. Combined with epidemiological follow up, it was then determined that a point source outbreak was occurring. Such laboratory “clusters”, however, do not always resolve as outbreaks; background clustering is often the norm requiring careful follow up to laboratory molecular data.

An elderly Caucasian male presented with a clinical history of recurrent retinochoroiditis and vitritis in one eye. A retinochoroidal scar was observed on physical examination. Toxoplasma serological testing in the Zoonotic Diseases and Emerging Pathogens (ZEP) Program showed a reactive result with high avidity IgG. IgM was not reactive, however. Considering the clinical presentations, a PCR test was done. T. gondii DNA was identified in a vitreous sample using the PCR B1 gene (with subsequent DNA sequencing of the genetic product).

This is the first case reported detecting T. gondii DNA in vitreous fluid of patients in our laboratory. To date, only one case has been reported in the scientific literature that uses cytological methods. Serological methods are not useful in ruling out active ocular toxoplasmosis; PCR may be a useful adjunct/supplemental tool in confirming the diagnosis of active disease. Further work continues.

PCR Confirmation of Toxoplasma gondii Infection »

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Recent media reports brought attention to a family of gram negative bacteria called Enterobacteriaceae that harbour the New Delhi Metallo-beta-lactamase gene (NDM) endemic in South Asia. This recently emerged gene is carried as a plasmid conferring broad range resistance to beta-lactam antibiotics, including the carbapenem class antibiotics. In July 2010, a Klebsiella pneumoniae strain that produced a Verona integron-encoded metallo-beta-lactamase (VIM) carbapenemase was identified for the first time in the US from a patient who had required hospitalization while traveling in Greece. Currently, Klebsiella pneumoniae carbapenem (KPC) is the predominant metallo-beta-lactamase carbapenemase circulating in the US.

Microbiology laboratories across British Columbia (BC) have been on the alert for carbapenem resistant Enterobacteriaceae using phenotypic methods. To support this screening, the Bacteriology and Mycology (BAM) Program has recently evaluated and implemented a multiplex PCR panel to detect important mechanisms of antimicrobial resistance in gram negative bacteria. The multiplex assays target AmpC genes (MOX 1-2, CMY 1-11, LAT 1-4,BIL 1, DHA 1-2, ACC, MIR 1T, ACT 1, FOX 1-5b), ESBL genes (SHV, TEM, CTX-M and OXA-1) and the four carbapenemases KPC, IMP, VIM and NDM.

This work was done with the technical support of Drs. Michael Mulvey from the National Microbiology Laboratory (NML) and Johann Pitout from Calgary Laboratory Services. Dr. Edith Blondel-Hill in Interior Health Authority worked with BAM’s Dr. Linda Hoang on a screening study to differentiate between phenotypic and genotypic identification of carbapenemases. Similarly, collaborative studies with Dr. Sylvie Champagne at St. Paul’s Hospital are underway to evaluate mechanisms of resistance.

Following the confirmation of an NDM strain identified in BC, microbiology laboratories in the Lower Mainland have agreed

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NDM’s Arrival: Screening for ESBL and AmpC Carbapenemase Resistance

In partnership with the UBC Centre for Operations Excellence (COE) at the Sauder School of Business, the PHMRL hosted a master’s student’s research this summer. The recent H1N1 pandemic was selected as the subject for this modeling exercise which included observations on laboratory processes for influenza testing, model validation using real and estimated data, and assessing various outbreak scenarios (using an optimal approach based on sample volume) using the newly developed tool. The project demonstrated that upfront data entry and accessioning activities are key areas (no surprise – pre-analytical is always key). The computer simulation also showed that when new Flow Cells (developed during a rapid imPROVE event) were triggered, high volume extraction equipment, RT-PCR equipment and with cross-trained staff from the Virology Program (as well as from Molecular Services, Environmental Microbiology and Zoonotic Diseases & Emerging Pathogens) available, the response structure’s capacity could meet pandemics that produced up to 1000 respiratory samples a day (more severe pandemic then occurred in 2009). Staff illness, however, was not factored in in this summer student’s project. The study underlined the need to protocolize management response and the need for planning. Simulation modeling has been shown to be a useful tool for such planning. A project with staff in the Bacteriology and Mycology (BAM) Program and the Environmental Microbiology (EM) Program, funded by the Canadian Laboratory Response Network (CLRN), is planned for the next months. Contact Yin Chang if you are interested in this project.

Influenza Computer Simulation Modeling: Operational Research Tools

Klebsiella pneumoniae, one of the bacteria that produce the NDM enzyme.(Photo Credit: CDC Image Library)

to forward carbapenem resistant and intermediate isolates to the Public Health Microbiology & Reference Laboratory (PHMRL) for genotypic analysis. A total of 23 resistant gram negative bacilli have been submitted since October, 2009. Of these, 5 NDM isolates from 4 patients have been identified in BC to date. All submitted isolates were negative for KPC, IMP and VIM. Instead, they carried various resistance genes associated with ESBL and AmpC mechanisms. We will maintain a central repository and surveillance role on behalf of the BC Association of Medical Microbiologists (BCAMM) and in communication with NML. This collaboration between BCAMM and PHMRL will allow for timely identification and confirmation of highly resistant bacteria for patient care and infection control management.

PROJECTS AND SURVEILLANCE WORK

The public health laboratory is requesting your assistance in reducing the significant workload that is created when locum physicians include their name or billing number on the requisition. Each unnecessary report copy (there are many) results in returned reports from the locum’s principal office, which then triggers an investigation from the public health laboratory.

Clients are asked to indicate only those ordering physicians requiring a copy of the report be identified on the requisition. Locum physicians are encouraged to select the notation on the requisition stating that they do not require a copy of the report (see above).

Online requisitions and the Sample Container Order Form used for ordering all of our laboratory requisition forms as well as sample kits continue to be found on our website at www.phsa.ca/

4 A sample Bacteriology & Mycology Requisition form with “I do not require a copy of the report” selected.

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Note that our updated Guide to Programs and Services is also online on our website at www.phsa.ca/bccdcpublichealthlab. Information on Programs at the Public Health Microbiology & Reference Laboratory may be found with key contact names and numbers. Sample collection instructions for our menu of tests available are detailed in this manual.

GUIDE TO PROGRAMS AND SERVICES

Chagas’ disease, caused by the parasite Typanosoma cruzi, is a zoonosis in the American continents (hence, also known as American trypanosomiasis). The disease is acquired through the bite wound caused by triatomid (reduviid) insects. Trypomastigote forms of the parasite are released in the insect faeces during its blood meal, and the faeces are rubbed or scratched into the bite wound or onto mucosal surfaces of the human host. Some of these disease-carrying insects are known as “kissing bugs” due to the fact that bites often occur on the lips of their human victims. Human infections mainly occur in rural areas of South America where there are poor sanitary and socioeconomic conditions. Poor housing provide excellent breeding grounds for the reduviid bugs and allow maximum contact between humans and the vector. T. cruzi infects 8-11 million people in Mexico, Central and South America with over 45,000 deaths per year. Morbidity is significant as well. Diagnosis is usually made from the microscopic examination of blood samples. The parasites may be detected using think and thick stained blood films or buffy coat concentration techniques. Aspirates from “chagomas” or enlarged lymph nodes may also be examined and cultured, as may histologic examination of biopsy samples. Chagas’ disease may also be acquired transplacentally or as a result of sharing contaminated needles, or receiving organs or blood from positive T. cruzi donors. This Fall, the Canadian Blood Services (CBS) implemented selective Chagas’ disease screening (questionnaire) their antibody testing for donors who may be at risk of infection with T. cruzi. CBS will advise family physicians of these individuals (CBS Notice to Family Physicians, CMAJ, June 21, 2010). BCCDC Public Health Microbiology and Reference Laboratory will be referring these CBS related samples to the National Centre for Parasitology for diagnostic testing.

LABORATORY REQUISITIONS

PROGRAM INFORMATION

Chagas’ Disease

NOW UPDATED!