Examination of Care Milestones for Preventing Congenital Syphilis Transmission Among Syphilis-Infected Pregnant Women in Alberta, Canada: 2017-2019.

BACKGROUND: An infectious syphilis outbreak in Alberta has resulted in increased congenital syphilis (CS) cases. To shed light on potential risk factors, we used administrative data sets to examine care milestones for the prevention of CS among pregnant women diagnosed with syphilis, as well as correlates of women giving birth to infants with CS. METHODS: Provincial administrative databases were used to identify and describe pregnant women diagnosed with any stage of infectious or noninfectious syphilis who gave birth in Alberta between January 1, 2017, and December 31, 2019. Data on prenatal care, syphilis screening, and syphilis medication dispensation were used to evaluate the care milestones. Clinical care and maternal demographics were assessed using logistic and linear regression analyses to determine correlates for missed care milestones or a newborn outcome of CS. RESULTS: Of 182 syphilis-infected pregnant women, 63 (34.6%) delivered a newborn with CS. Overall, in the first trimester, 136 (75.1%) women had a health care visit, 72 (39.6%) had a prenatal care visit, 71 (39.0%) were screened for syphilis, and 44 (24.2%) were treated. Gestational time to treatment initiation (adjusted odds ratio, 1.04; 95% confidence interval, 1.02-1.06) and older maternal age at diagnosis (adjusted odds ratio: 1.28, 95% confidence interval, 1.08-1.50) were independently associated with CS outcomes. No variables were found to be independently associated with a health care visit, prenatal screening, or initiation of treatment. CONCLUSIONS: Although nearly two-thirds of CS cases were prevented, there remained missed opportunities in the prevention of CS. Early treatment, which relies on timely access to prenatal care and screening, was the most important for the prevention of CS.

Comparative Analysis of Molecular and Serologic Testing for Primary Syphilis: A Population-Based Cohort Study.

Rising rates of syphilis (T. pallidum; Tp) requires rapid diagnosis and treatment to manage the growing epidemic. Syphilis serology is imperfect and requires interpretation of multiple tests while molecular diagnostics allows for potential yes-no identification of highly infective, primary anogenital lesions. Accuracy of this testing modality has thus far been limited to small, highly selective studies. Therefore, we retrospectively assessed a large, adult population of patients with anogenital lesions seen at Sexually Transmitted Infection (STI) clinics in Alberta, Canada who were screened for syphilis and herpes simplex (HSV) 1/2 using PCR to evaluate Tp-PCR versus serology to diagnose primary syphilis. 114 (3.1%) of the 3,600 adult patients had at least one Tp-PCR+ anogenital lesion with 99 (2.8%) patients having newly positive syphilis serology (new INNO-LIA positive or 4-fold RPR increase). Tp-PCR had a sensitivity of 49.3% (95% CI 42.6-56.1) and specificity of 99.9% (99.7-100.0). Positive predictive values and negative predictive values in the study population or when corrected for provincial prevalence were 97.4% (92.5-99.5) or 0.4% (0.4-1.2) and 96.7% (96.1-97.3) or 100.0% (100.0-100.0), respectively. Positive and negative likelihood ratios were estimated at 555 (178-1733) and 0.5 (0.4-0.6), respectively. Review of all Tp-PCR performed with or without exclusion of HSV-positive lesions resulted in no significant change in Tp-PCR characteristics. Interestingly, 12 of the Tp-PCR+ samples had negative serology at time of lesion sampling but became positive within our 28-day testing window. Overall, this study further supports the use of Tp-PCR as an accurate assay to rapidly identify, treat, and prevent the spread of primary syphilis.

Increasing Incidence of Invasive Group A Streptococcus Disease in First Nations Population, Alberta, Canada, 2003-2017.

The incidence of invasive group A Streptococcus (iGAS) disease in the general population in Alberta, Canada, has been steadily increasing. To determine whether rates for specific populations such as First Nations are also increasing, we investigated iGAS cases among First Nations persons in Alberta during 2003-2017. We identified cases by isolating GAS from a sterile site and performing emm typing. We collected demographic, social, behavioral, and clinical data for patients. During the study period, 669 cases of iGAS in First Nations persons were reported. Incidence increased from 10.0 cases/100,000 persons in 2003 to 52.2 cases/100,000 persons in 2017. The 2017 rate was 6 times higher for the First Nations population than for non-First Nations populations (8.7 cases/100,000 persons). The 5 most common emm types from First Nations patients were 59, 101, 82, 41, and 11. These data indicate that iGAS is severely affecting the First Nations population in Alberta, Canada.

Histoplasmosis acquired in Alberta, Canada: an epidemiological and genomic study.

BACKGROUND: The classic geographical range of histoplasmosis in North America primarily includes the states and provinces adjacent to the Ohio, Mississippi, and St Lawrence riverways. Although Alberta, Canada is not typically considered a region of risk for histoplasmosis, cases with suspected local acquisition have been reported. We aimed to investigate the epidemiology and geographical distribution of cases of histoplasmosis in Alberta to assess evidence for local acquisition of infections, using genomic analysis for corroboration. METHODS: We did an epidemiological and genomic investigation, in which laboratory-confirmed cases of histoplasmosis were reviewed in Alberta from 2011, when the disease became reportable, until 2018. We used data attained from Alberta Health. Travel and exposure histories and clinical features were reviewed. Definite local acquisition was defined as a case without previous travel outside Alberta or associated with a common-source outbreak within the province, whereas probable local acquisition was a sporadic case with travel outside Alberta but compelling local exposures. Genomes of selected case isolates were analysed, including those from cases suspected to have been locally acquired and imported. FINDINGS: Between Jan 1, 2011, and June 30, 2018, 45 cases of histoplasmosis were identified. Participants had a median age of 53 years (range 17-77) and 32 [71%] were male. Among 34 patients with documented travel histories, ten (29%) had never left the province. 11 cases were of definite local acquisition, including eight cases from three common-source outbreaks and three sporadic cases in patients who had never travelled outside Alberta. The common-source outbreaks all involved exposure to bats or their droppings in chimneys or attics of private dwellings or churches. Four patients had travelled outside Alberta but compelling evidence was seen for local exposure to bat guano. Genome sequencing showed that isolates from cases of definite and probable local acquisition clustered together and were genetically distinct from isolates from suspected imported cases and other published isolates. INTERPRETATION: Using epidemiological and genomic analyses, we established that cases of histoplasmosis have been acquired in Alberta, thus expanding the geographical range of Histoplasma spp much further northwest than was previously appreciated. Histoplasmosis should be considered in patients with compatible symptoms outside areas of classic geographical risk. FUNDING: None.

The use of laboratory-identified event surveillance to classify adverse outcomes due to Clostridioides difficile infection in Canadian long-term care facilities.

OBJECTIVE: Adverse outcomes following Clostridioides difficile infection (CDI) are not often reported for long-term care facility (LTCF) residents. We focused on the adverse outcomes due to CDI identified in Alberta LTCFs. METHODS: All positive Clostridioides difficile stool specimens identified by laboratory-identified (LabID) event surveillance in Alberta from 2011 to 2018, along with Alberta Continuing Care Information System, were used to define CDI in Alberta LTCFs. CDI cases were classified as long-term care onset, hospital onset, and community onset. Laboratory records were linked to provincial databases to analyze acute-care admissions and mortality within 30-day post CDI. Age, sex, case classification, episode, and operator type, were investigated using logistic regression. RESULTS: Overall, 902 CDI cases were identified in 762 LTCF residents. Of all CDI events, 860 (95.3%) were long-term care onset, 38 (4.2%) were hospital onset, and 4 (0.4%) were community onset. The CDI rate was 2.0 of 100,000 resident days. In total, 157 residents (20.6%) had 30-day all-cause mortality, 126 CDI cases (14.0%) had 30-day all-cause acute-care admissions. The 30-day all-cause mortality rate was significantly higher in residents aged >80 versus ≤80 years (24.9 vs 12.3 per 100 residents; P < .05). Residents aged >80 years, with hospital-onset CDI, and those staying in private or voluntary LTCFs were more likely to have 30-day all-cause acute-care admissions. CONCLUSIONS: The prevalence of CDI adverse outcomes is in LTCFs was found to be high using LabID event surveillance. Annual review of CDI adverse outcomes using LabID event can minimize the burden of surveillance and standardize the process across all Alberta LTCFs.

Epidemiology of Shiga Toxin-Producing Escherichia coli O157 in the Province of Alberta, Canada, 2009-2016.

Shiga toxin-producing Escherichia coli (STEC) infections are the product of the interaction between bacteria, phages, animals, humans, and the environment. In the late 1980s, Alberta had one of the highest incidences of STEC infections in North America. Herein, we revisit and contextualize the epidemiology of STEC O157 human infections in Alberta for the period 2009-2016. STEC O157 infections were concentrated in large urban centers, but also in rural areas with high cattle density. Hospitalization was often required when the Shiga toxin genotype stx2a stx2c was involved, however, only those aged 60 years or older and infection during spring months (April to June) independently predicted that need. Since the late 1980s, the rate of STEC O157-associated hemolytic uremic syndrome (HUS) in Alberta has remained unchanged at 5.1%, despite a marked drop in the overall incidence of the infection. While Shiga toxin genotypes stx1a stx2c and stx2a stx2c seemed associated with HUS, only those aged under 10 years and infection during spring months were independently predictive of that complication. The complexity of the current epidemiology of STEC O157 in Alberta highlights the need for a One Health approach for further progress to be made in mitigating STEC morbidity.

Increasing Rates of Invasive Group A Streptococcal Disease in Alberta, Canada; 2003-2017.

BACKGROUND: We present an analysis of increasing rates of invasive group A streptococci (iGAS) over a 15-year period in Alberta, Canada. METHODS: From 2003 to 2017, the emm type of iGAS isolates was identified from patients with iGAS disease in Alberta. Demographic, clinical, and risk factor data were collected. RESULTS: A total of 3551 cases of iGAS were identified in Alberta by isolation of a GAS isolate from a sterile site. The age-standardized incidence rates of iGAS increased from 4.24/100 000 in 2003 to 10.24 in 2017. Rates (SD) were highest in those age <1 (9.69) years and 60+ (11.15) years; 57.79% of the cases were male. Commonly identified risk factors included diabetes, hepatitis C, nonsurgical wounds, addiction, alcohol abuse, drug use, and homelessness. The overall age-standardized case fatality rate was 5.11%. The most common clinical presentation was septicemia/bacteremia (41.84%), followed by cellulitis (17.25%). The top 4 emm types from 2003-2017 were emm1, 28, 59, and 12. In 2017, the top 4 emm types (emm1, 74, 101, and 59) accounted for 46.60% of cases. CONCLUSIONS: The incidence of iGAS disease in Alberta, Canada, has increased from 2003 to 2017. This increase has been driven not by a single emm type, but rather what has been observed is a collection of common and emerging emm types associated with disease. In addition, it is also likely that societal factors are playing important roles in this increase as risk factors associated with marginalized populations (addiction, alcohol abuse, and drug use) were found to have increased during the survey period.

How well do ICD-9 physician claim diagnostic codes identify confirmed pertussis cases in Alberta, Canada? A Canadian Immunization Research Network (CIRN) Study.

BACKGROUND: Rates of Bordetella pertussis have been increasing in Alberta, Canada despite vaccination programs. Waning immunity from existing acellular component vaccines may be contributing to this. Vaccine effectiveness can be estimated using a variety of data sources including diagnostic codes from physician billing claims, public health records, reportable disease and laboratory databases. We sought to determine if diagnostic codes from billing claims (administrative data) are adequately sensitive and specific to identify pertussis cases among patients who had undergone disease-specific laboratory testing. METHODS: Data were extracted for 2004-2014 from a public health communicable disease database that contained data on patients under investigation for B. pertussis (both those who had laboratory tests and those who were epidemiologically linked to laboratory-confirmed cases) in Alberta, Canada. These were deterministically linked using a unique lifetime person identifier to the provincial billing claims database, which contains International Classification of Disease version 9 (ICD-9) diagnostic codes for physician visits. We examined visits within 90 days of laboratory testing. ICD-9 codes 033 (whooping cough), 033.0 (Bordetella pertussis), 033.1 (B. parapertussis), 033.8 (whooping cough, other specified organism), and 033.9 (whooping cough, other unspecified organism) in any of the three diagnostic fields for a claim were classified as being pertussis-specific codes. We calculated sensitivity, specificity, positive (PPV) and negative (NPV) predictive values. RESULTS: We identified 22,883 unique patients under investigation for B. pertussis. Of these, 22,095 underwent laboratory testing. Among those who had a laboratory test, 2360 tested positive for pertussis. The sensitivity of a pertussis-specific ICD-9 code for identifying a laboratory-confirmed case was 38.6%, specificity was 76.9%, PPV was 16.0%, and NPV was 91.6%. CONCLUSION: ICD-9 codes from physician billing claims data have low sensitivity and moderate specificity to identify laboratory-confirmed pertussis among persons tested for pertussis.

Epidemiology of pertussis in Alberta, Canada 2004-2015.

BACKGROUND: We describe the epidemiology of pertussis in Alberta, Canada by person, place, and time between 2004 and 2015, identify outbreak years, and examine vaccination coverage and vaccination timeliness. METHODS: We used health data from Alberta's Communicable Disease Registry System for the period of January 1, 2004 through August 31, 2015 to identify unique cases of pertussis. Unique cases were deterministically linked to data in Alberta's immunization repository and health care insurance plan registry. Population estimates and vaccination coverage were extracted from Alberta's online Interactive Health Data Application. We estimated pertussis incidence rates per 100,000 persons by year, age group, gender, and health zone. Outbreak years were identified using a one-sided cumulative sum (CUSUM) analysis by comparing annual incidence rates to baseline rates. RESULTS: Over the period, 3510 cases of pertussis were confirmed by laboratory testing or epidemiological linkage. Incidence rates per 100,000 persons were highest in 2004 (20.5), 2005 (13.6), and 2015 (10.4) for all age groups. Incidence rates were highest among the youngest age groups and decreased as age groups increased. Based on CUSUM analysis, 2008 and 2012 met the criteria for outbreak years. Vaccination coverage was over 90% among the general population, however only 61% of cases received at least one dose. About 60% of cases were diagnosed 5+ years after receiving the vaccine. Approximately 87-91% of vaccinated cases did not receive the first three vaccine doses in a timely manner. CONCLUSION: Pertussis incidence rates fluctuated over the period across all age groups. The majority of cases had no record of vaccination or were delayed in receiving vaccines. CUSUM analysis was an effective method for identifying outbreaks.

Use of laboratory and administrative data to understand the potential impact of human parainfluenza virus 4 on cases of bronchiolitis, croup, and pneumonia in Alberta, Canada.

BACKGROUND: Human Parainfluenza Virus (hPIV) causes severe respiratory illness in infants and adults. Our study describes the association of hPIV1-4 with bronchiolitis, croup, and pneumonia using retrospective laboratory, administrative and public health data. Due to issues including the historic lack of hPIV4 in some commercial respiratory virus panels, the description of the impact of hPIV4 on croup, bronchiolitis, and pneumonia at population levels has often been limited. This study will use routine clinical laboratory data, and administrative data to provide a preliminary description of the impact of hPIV4 on these diseases in our population. METHODS: A three year cohort of patients positive for hPIV was linked with data from physician visits and hospital admissions to define cases and hospitalization status. International Classification of Disease (ICD-9) codes were used to determine if cases had croup, bronchiolitis, and pneumonia. We also looked at differences in hospitalization status, age and gender among hPIV1-4. All statistical analysis was done using SPSS (Version 19.0.0, IBM Corp© 2010) and Graphpad Prism V6 (GraphPad Software, Inc., 2012). RESULTS: Only hPIV1 and hPIV4 specimens had positivity rates greater than 5 % of all specimens sent for respiratory virus panel testing. hPIV1 exhibited a biennial pattern while the pattern for hPIV3 was less interpretable due to lower positivity rates. Circulation patterns for hPIV2 and hPIV4 were not assessed due to the low positivity rates of theses specimens. From 2010 to 2013, there were 2300 hPIV cases with hPIV3 (46 %) being the most common, followed by hPIV1 (27 %), hPIV4 (16 %) and hPIV2 (11 %). The median age was 2 years for all hPIV types. Males were slightly greater than females for hPIV1 and hPIV2, with an equal distribution for hPIV3 and slightly more females than males for hPIV4. hPIV1 and hPIV2 had the highest or proportion of croup while hPIV3 and hPIV4 had the highest proportion of pneumonia. Within hPIV4 cases, distributions of diseases were; pneumonia (21 %, 95 % CI 17.1-25.7), bronchiolitis (18 %, 95 % CI 14.3-22.5), croup (2 %, 95 % CI 0.8-3.9), mixed illness of any of pneumonia, bronchiolitis or croup (4 %, 95 % CI 2.5-7.0) or other respiratory diseases (54 %, 95 % CI 49.1-59.6). CONCLUSIONS: We used laboratory and administrative data to undertake a descriptive analysis of the association of hPIV1-4 with croup, bronchiolitis and pneumonia. hPIV4 appears to be more associated more with bronchiolitis and pneumonia and less with croup in our population.

The molecular epidemiology of incident methicillin-resistant Staphylococcus aureus cases among hospitalized patients in Alberta, Canada: a retrospective cohort study.

BACKGROUND: Infection Prevention and Control (IPC) surveillance for incident methicillin-resistant Staphylococcus aureus (MRSA) in hospitalized patients is performed in a complete provincial surveillance network of all acute care facilities in Alberta, Canada. IPC surveillance is centralized using a web-based data entry platform so that each patient is counted only once. All diagnostic laboratories submit the first clinical MRSA isolate associated with a patient without previous MRSA positive clinical cultures in the preceding year to the Provincial Laboratory for Public Health (ProvLab) for molecular typing. This study will investigate the relationship between the IPC epidemiological classification based on time of detection following admission to hospital (Hospital Acquired and Community Associated) and the matched laboratory MRSA surveillance data using a retrospective cohort study design. METHODS: Incident IPC MRSA cases were classified according to IPC epidemiologic definitions. DNA sequencing of the Staphylococcus protein A (spa) gene and pulsed-field gel electrophoresis (PFGE) typing was performed. IPC MRSA surveillance data were matched to the ProvLab molecular surveillance data. Univariate comparisons of proportions were performed for categorical variables and the Student's t test for continuous variables. RESULTS: MRSA molecular typing data were available for matching for 46.7 % (2248/4818) of incident IPC cases. There was agreement in definitions for traditional nosocomial clones (USA100/CMRSA2) with Hospital Acquired (HA)-MRSA (65.1 % of all IPC HA-MRSA) and traditional community clones (USA400/CMRSA7 and USA300/CMRSA10) with Community Acquired (CA)-MRSA (62.4 % of CA-MRSA). However, we observed discordance for both traditional nosocomial/CA-MRSA (30.4 % of CA-MRSA) and for traditional community/HA-MRSA (26.9 % of HA-MRSA). CONCLUSIONS: We note agreement between traditional nosocomial clones and HA-MRSA, and traditional community clones and CA-MRSA. However, approximately one-quarter of HA-MRSA are those of traditional community clones while approximately one-third of CA-MRSA are those of traditional nosocomial clones. Collaborative provincial MRSA surveillance is important as the distinction between IPC case attribution in acute care settings and the historical definitions of MRSA clones as community- or healthcare-associated have blurred.

Respiratory Outbreak Investigations: How Many Specimens Should Be Tested?

To determine the optimal number of specimens for virus detection in a respiratory outbreak, laboratory results from 2 Canadian public health laboratories were reviewed. The evidence suggests that 3 specimens are sufficient for detection of a virus in >95% of outbreaks, thereby reducing laboratory costs.

Dominance of two genotypes of Bordetella pertussis during a period of increased pertussis activity in Alberta, Canada: January to August 2012.

OBJECTIVES: The purpose of this study was to undertake an epidemiological analysis of an increase in Bordetella pertussis activity during the period January 1 to August 31, 2012 in Alberta, Canada. B. pertussis testing was done using an IS481 real-time PCR assay with PCR-positive and indeterminate specimens cultured and stored for further analysis. METHODS: Laboratory data were linked to Alberta Health (AH) cases that were reported in the Communicable Disease Reporting System (CDRS) to identify case isolates; exclusion criteria were used to avoid biases. Case isolates were analyzed at the National Microbiology Laboratory (NML) by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Pertussis immunization data were extracted from the Alberta Provincial Immunization Repository (Imm/ARI) and linked to the pertussis cases. RESULTS: Using PFGE and MLST, 52 case isolates could be divided into two main sequence type groups: 41 cases belonged to the ST-1 group (ST-1 and the novel ST-19) and 11 cases belonged to the ST-2 group (ST-2 and the novel ST-20). Of the total cases genotyped (N=52), 18 (34.6%) had a history of immunization, 28 (53.8%) were not immunized, and six (11.6%) had an unknown immunization history. Of the total non-immunized cases, 25/28 (89.2%) belonged to the ST-1 group. Furthermore, of the 41 ST-1 group cases, 25 were not immunized compared to only three of the ST-2 group cases (p=0.0004, Fisher's exact test). CONCLUSIONS: This study shows the dominance of two genotypes of B. pertussis in our jurisdiction and indicates less pertussis immunization in individuals infected with the ST-1 group.

Bordetella pertussis in sporadic and outbreak settings in Alberta, Canada, July 2004-December 2012.

BACKGROUND: ProvLab Alberta provides all laboratory testing for Bordetella pertussis including sporadic cases and outbreak investigations through collaborations with provincial public health partners. We describe B. pertussis activity in Alberta from July 2004 to December 2012. METHODS: Laboratory testing for pertussis was analyzed using interpreted laboratory data that was generated by DIAL, a secure web-based platform. Duplicate specimens from the same individual ≤90 days were excluded to generate a case-based dataset. Immunization status of confirmed pertussis cases from the provincial immunization repository was reviewed. RESULTS: Overall, 7.1% of suspected pertussis cases tested positive with a higher positivity rate in outbreak as compared to sporadic setting. Annual variations in sporadic pertussis cases were observed across the province with higher positivity rates in 2005, 2008, 2009 and 2012. A significantly higher positivity rate was observed in a northern region of Alberta. While the positivity rate in sporadic setting was highest in adolescents aged 10 to <15 years old (14.8%), population-based disease burden was highest in young children <5 years old. Of the 81.6% (n = 1,348) pertussis cases with immunization records, 48.3% were up-to-date with immunization. The pertussis cases that were up-to-date with their immunization were older (median age 12.9 years) as compared to those with incomplete (median age 9.7 years) or no pertussis immunization (median age 3.8 years). CONCLUSIONS: Cyclic pattern of annual pertussis activity with geographic variation was observed in Alberta with no obvious case finding effect from outbreak investigations. The high positivity rates in adolescents suggested an underestimation of disease burden in this age group.

Use of an innovative web-based laboratory surveillance platform to analyze mixed infections between human metapneumovirus (hMPV) and other respiratory viruses circulating in Alberta (AB), Canada (2009-2012).

We investigated the proportions of mono vs. mixed infections for human metapneumovirus (hMPV) as compared to adenovirus (ADV), four types of coronavirus (CRV), parainfluenza virus (PIV), RSV, and enterovirus/rhinovirus (ERV) in Alberta, Canada. Using the Data Integration for Alberta Laboratories (DIAL) platform, 26,226 respiratory specimens at ProvLab between 1 July 2009 and 30 June 2012 were selected and included in the study. Using the Respiratory Virus Panel these specimens tested positive for one or more respiratory virus and negative for influenza A and B. From our subset hMPV was the fourth most common virus (n=2,561) with 373 (15%) identified as mixed infection using DIAL. Mixed infection with hMPV was most commonly found in infants less than 6 months old and ERV was most commonly found in mixed infection with hMPV (230/373, 56%) across all age groups. The proportion of mixed-infection vs. mono-infection was highest for ADV (46%), followed by CRV 229E (32%), CRV HKU1 (31%), CRV NL63 (28%), CRV OC43 (23%), PIV (20%), RSV (17%), hMPV (15%) and ERV (13%). hMPV was significantly more likely to be identified in mono infection as compared with ADV, CRV, PIV, and RSV with the exception of ERV [p < 0.05].

Multi-analyte suspension arrays for the detection of common viruses: how viable are these assays in clinical laboratories?

This article assesses the viability of a recently described multi-analyte suspension array for the detection of herpes simplex viruses-1 and -2, cytomegalovirus, Epstein-Barr virus, human papillomavirus and hepatitis B virus. This methodology was identified by the authors as a means of providing rapid, high-throughput multiplex assays that were easy to use. When paired with PCR assays, multi-analyte suspension arrays have the ability to overcome drawbacks associated with conventional detection methods such as long turnaround time, detection sensitivity and the ability to detect only one pathogen in each round of testing. However, the assays described in this article are still hampered by some key issues including limit of detection, the fact that median fluorescence intensity is not truly a quantitative diagnostic method, and that open molecular diagnostic systems can lead to contamination and/or increased operator-based errors. Although modern pressures on clinical virology laboratories have increased the need to develop a system that can detect pathogens in multiplexed assays, in the future these assays will only become more clinically relevant if they are designed with greater stakeholder input.