Molecular diagnostics is increasingly embracing pharmacogenomics. Here we discuss the role of G protein-coupled receptors and their accessory proteins in disease, drawing on our experience addressing the role of the calcium-sensing receptor polymorphisms/variation in familial hypocalciuric hypercalcemia and autosomal dominant hypocalcemia in order to highlight the role that pharmacogenomics may play in personalized treatment.

by Dr M. D. Thompson, Dr D. E. C. Cole and Dr G. N. Hendy

Introduction
The identification and characterization of gene families encoding G protein-coupled receptors (GPCRs) and the proteins necessary for the processes of ligand binding, GPCR activation, inactivation and receptor trafficking facilitates the study of drug response in the context of human genetic disease. Thompson et al. reviewed these topics in Volume 1175 of Methods in Molecular Biology in 2014 [1–3].

With the advance of genomic technologies, there has been a substantial increase in the inventory of naturally occurring rare and common GPCR variants [2, 3]. In addition to functional GPCR variants, genetic variation has been found in a variety of G protein subunits and accessory proteins that normally modify or organize heterotrimeric G protein coupling. These include variants of the regulator of G protein signalling (RGS) protein associated with hypertension; variants of the activator of G protein signalling (AGS) proteins associated with various phenotypes (such as the type III AGS8 variant to hypoxia); variants in of the G protein-coupled receptor kinase (GRK) proteins, such as GRK4, associated with disorders such as hypertension [1]. Variation in GPCR, G protein and accessory protein structure and function provides the basis for examining the pharmacogenomics of GPCRs and the genetics of related monogenic disorders [1–3].

GPCR variants and variant G protein subunits associated with human disease
Diseases caused by the genetic disruption of GPCR functions may be selectively targeted by drugs that rescue altered receptors. The identification of variants in these receptors provides genetic reagents useful in drug screens. Examples of drugs developed as a result of targeting GPCRs mutated in disease include: the calcimimetics and calcilytics, drugs targeting melanocortin receptors in obesity and interventions that alter gonadotropin-releasing hormone receptor (GNRHR) loss from the cell surface in idiopathic hypogonadotropic hypogonadism [2, 3].

Inactive, overactive and constitutively active receptors
Genetic variations in GPCR genes disrupt GPCR function in a variety of human genetic diseases. In vitro studies and animal models have been used to identify the molecular pathologies underlying these GPCR mutations. Inactive, overactive, or constitutively active receptors have been identified. These receptor variants alter ligand binding, G protein coupling, receptor desensitization, or receptor recycling. Variant GPCRs disrupted in disease include rhodopsin, thyrotropin, parathyroid hormone (PTH), melanocortin, follicle-stimulating hormone (FSH), luteinizing hormone, GNRHR, adrenocorticotropic hormone, vasopressin, endothelin-β, purinergic, and the G protein associated with asthma [GPRA or neuropeptide S receptor 1 (NPSR1)] [2]. Data on the role of activating and inactivating calcium-sensing receptor (CASR) mutations provide examples that will be discussed in detail with respect to familial hypocalciuric hypercalcemia (FHH) and autosomal dominant hypocalcemia (ADH) [4].

Calcium-sensing receptor mutations and hypercalcemia/hypocalcemia
The CASR functions as an extracellular calcium sensor for the parathyroid gland and the kidney. The CASR itself is a plasma membrane GPCR that is abundantly expressed in the PTH secreting cells of the parathyroid gland and the cells lining the renal tubule lumen [2, 4]. The activity and/or expression levels of the CASR dictate the calcium set-point at which PTH is secreted from the parathyroid gland [2]. CASR gene variants may influence many physiological processes by contributing to individual differences in calcium metabolism [2].

Inherited abnormalities of the CASR gene give rise to a variety of disorders of mineral ion homeostasis [5]. Heterozygous loss-of-function mutations cause familial (benign) hypocalciuric hypercalcemia (FHH) in which the lifelong mild hypercalcemia is generally asymptomatic. Homozygous inactivating mutations give rise to neonatal severe hyperparathyroidism (NSHPT) with extreme hypercalcemia and marked skeletal changes [5–7]. Heterozygous activating mutations of the CASR cause ADH that may be asymptomatic or present with seizures in the neonatal period or childhood or later in life [2].

Familial hypocalciuric hypercalcemia and neonatal severe hyperparathyroidism
The syndrome known as familial hypocalciuric hypercalcemia (FHH), or familial benign hypercalcemia, results in mild primary hyperparathyroidism and relatively normal serum concentrations of PTH [8]. A key feature of FHH is the unusually high renal tubular reabsorption of calcium and magnesium in the face of hypercalcemia. However, some FHH families have affected members in which calcium excretion is increased and this may reflect the particular CASR mutation involved [2].

NSHPT involves multiglandular parathyroid hyperplasia. Children under the age of 6 months develop severe, symptomatic hypercalcemia with bony changes of hyperparathyroidism. Delay in treatment can lead to a devastating neurodevelopmental disorder. Some forms of neonatal hyperparathyroidism, involving either a de novo or paternal inheritance of a mutated CASR allele, present with milder symptoms [2].

Upwards of 200 unique inactivating, FHH/NSHPT type mutations in the CASR have been identified [2], as shown in Figure 1 (http://www.casrdb.mcgill.ca/). Although FHH is inherited in an autosomal dominant manner with almost 100% penetrance and variable expressivity, the population prevalence is not well defined. The FHH trait was initially mapped to chromosome 3q21, the locus of the CASR gene: two-thirds of FHH cases are due to mutations in the CASR gene and the disorder is FHH type 1 [2].

In some kindreds, however, the FHH trait maps to either chromosome 19p13.3 (FHH type 2) or 19q13.3 (FHH type 3). FHH2 is due to heterozygous loss-of-function mutations in GNA11, the gene encoding the alpha subunit of G11 that couples the activated CASR to intracellular signalling pathways [9]. FHH3 is due to inactivating mutations in the AP2S1 gene that encodes the sigma subunit of adaptor protein complex 2 critical for clathrin-mediated endocytosis of a variety of cell surface proteins including GPCRs such as the CASR [2].

Hypocalcemia, hypoparathyroidism, and hypocalcemic hypercalciuria
Gain-of-function mutations in the CASR gene have been identified in several families previously diagnosed with ADH, autosomal dominant hypoparathyroidism, and hypocalcemic hypercalciuria [2]. In the parathyroid gland, the activated CASR suppresses PTH secretion and in the kidney, it induces hypercalciuria [4]. De novo mutations are common [2]. Mosaicism for de novo mutation in an otherwise healthy parent has been described and this has important implications for counselling parents about the risk of recurrence [2].

In a subset of ADH families, CASR gain-of-function mutations have been associated with the onset of tonic–clonic seizures. In ADH, brain calcifications – sometimes accompanied by seizures – suggest that activating mutations may alter calcium homeostasis in the brain. The abnormal set-point of calcium regulation complicates treatment with calcitriol and dietary calcium supplementation because the CASR expressed in the kidney may override other regulators of calcium excretion. The constitutively activated CASR mutant induces hypercalciuria, which may exacerbate the hypocalcemia [2, 10].

More than 100 activating mutations (virtually all missense) have been identified and appear almost equally divided between the amino-terminal third of the extracellular domain (ECD) and the transmembrane domain shown in Figure 1 (http://www.casrdb.mcgill.ca/).

GPCR pharmacogenomics
Pharmacogenetics investigates the influence of genetic variants on physiological phenotypes related to drug response and disease, while pharmacogenomics takes a genome-wide approach to advancing this knowledge. Both play an important role in identifying responders and non-responders to medication, avoiding adverse drug reactions, and optimizing drug dose for the individual.

The CASR provides an example of GPCR variability in the population. While CASR variants contribute to monogenic disorders such as FHH and ADH, common CASR polymorphisms also account for some of the population variation in calcium response that is a risk factor for a variety of disease susceptibilities. CASR single nucleotide polymorphisms (SNPs) have been associated with a number of complex phenotypes. For example, the Ala986Ser variant may contribute to bone mineral density, primary hyperparathyroidism, and Paget disease [11].
The cluster of missense polymorphisms located in the cytoplasmic tail of the receptor is associated with inter-individual population differences in Ca²⁺ metabolism [12]. Different haplotypes are associated with primary hyperparathyroidism and the frequency of kidney stones. More recent genome-wide association studies in ~33,000 individuals of European and Indian Asian ancestry confirmed that the blood calcium concentration associated most significantly with SNPs in the CASR gene [13].

CASR variants are known to alter the sensitivity of the CASR and result in altered extracellular calcium-concentration set points in tissues. Web sites such as http://www.casrdb.mcgill.ca/ document a number of SNPs scattered across the more than 100 kb region of genomic DNA that encompasses the CASR gene. Common missense SNPs (Ala986Ser and Arg990Gly) are clustered in the DNA region encoding the cytoplasmic tail of CASR. The most common of these, the Ala986Ser variant, is predictive of the unbound, extracellular calcium levels [11]. The Ala986Ser variant is thus a mild inactivating variant that may predispose to hypercalcemia without being fully predictive of hypocalciuria. By contrast, the Arg990Gly variant (activating) results in the increased calcium excretion that characterizes idiopathic hypercalciuria and is predictive of nephrolithiasis [2].

Conclusion
GPCRs are the primary target of therapeutic drugs and have been the focus of these studies. These variants include SNPs and insertion/deletions that have potential to alter GPCR expression of function. In vivo and in vitro studies have determined functional roles for many GPCR variants, but genetic association studies that define the physiological impact of the majority of these common variants are still limited. Despite the breadth of pharmacogenetic data available, GPCR variants have not been included in drug labelling and are only occasionally considered in optimizing clinical use of GPCR targeted agents. As the extent of GPCR pharmacogenomic data increases, the opportunity for routine assessment of GPCR variants to predict disease risk, drug response and potential adverse drug effects will no doubt become more commonplace.

References
1. Thompson MD, Cole DE, Jose PA, et al. G protein-coupled receptor accessory proteins and signaling: pharmacogenomic insights. Methods Mol Biol. 2014; 1175: 121-52.
2. Thompson MD, Hendy GN, Percy ME, et al. G protein-coupled receptor mutations and human genetic disease. Methods Mol Biol. 2014; 1175: 153-87.
3. Thompson MD, Cole DE, Capra V, et al. Pharmacogenetics of the G protein-coupled receptors. Methods Mol Biol. 2014; 1175: 189-242.
4. Zhang C, Zhuo Y, Moniz HA, et al. Direct determination of multiple ligand interactions with the extracellular domain of the calcium sensing receptor. J Biol Chem. 2014 Oct 10. pii: jbc.M114.604652.
5. Thim SB, Birkebaek NH, et al. Activating calcium-sensing receptor gene variants in children: a case study of infant hypocalcaemia and literature review. Acta Paediatr. 2014 Jul 10. doi: 10.1111/apa.12743.
6. Toka HR, Pollak MR.The role of the calcium-sensing receptor in disorders of abnormal calcium handling and cardiovascular disease. Curr Opin Nephrol Hypertens. 2014; 23: 494-501.
7. Grzegorzewska AE, Ostromecki G. Gene polymorphism of the vitamin D receptor, vitamin D-binding protein and calcium-sensing receptor in respect of calcium-phosphate disturbances in chronic dialysis patients. Przegl Lek. 2013; 70: 735-8.
8. Jakobsen NF, Rolighed L, Nissen PH, et al. Muscle function and quality of life are not impaired in familial hypocalciuric hypercalcemia: a cross- sectional study on physiological effects of inactivating variants in the calcium-sensing receptor gene (CASR). Eur J Endocrinol. 2013; 169: 349-57.
9. Nesbit MA, Hannan FM, Howles SA, et al. Mutations affecting G-protein subunit α11 in hypercalcemia and hypocalcemia. N Engl J Med. 2013; 368: 2476-86.
10. Ranieri M, Tamma G, Di Mise A, et al. Excessive signal transduction of gain-of-function variants of the calcium-sensing receptor (CaSR) are associated with increased ER to cytosol calcium gradient. PLoS One. 2013; 8: e79113.
11. Han G, Wang O, Nie M, et al. Clinical phenotypes of Chinese primary hyperparathyroidism patients are associated with the calcium-sensing receptor gene R990G polymorphism. Eur J Endocrinol. 2013; 169: 629-38.
12. Scillitani A, Guarnieri V, Battista C, et al. Primary hyperparathyroidism and the presence of kidney stones are associated with different haplotypes of the calcium-sensing receptor. J Clin Endocrinol Metab. 2007; 92: 277-83.
13. Kapur K1, Johnson T, Beckmann ND, et al. Genome-wide meta-analysis for serum calcium identifies significantly associated SNPs near the calcium-sensing receptor (CASR) gene. PLoS Genet. 2010; 6: e1001035.

The authors
Miles D. Thompson¹* PhD; David E. C. Cole² MD, PhD; Geoffrey N. Hendy³ PhD

¹Department of Pharmacology and Toxicology, Medical Sciences Building, University of Toronto, Toronto, ON, Canada. M5S 1A8.
²Departments of Laboratory Medicine and Pathobiology, Medicine and Genetics, University of Toronto, ON, Canada. M4N 3M5.
³Departments of Medicine, Physiology, and Human Genetics, McGill University, and Calcium Research Laboratory and Hormones and Cancer Unit, Royal Victoria Hospital, Montreal, QC, Canada. H3A 1A1.

*Corresponding author
E-mail: miles.thompson@utoronto.ca

A comparative analysis of serotyping and mass spectrometry (MS) methods for the determination of the flagellar type (H type) of clinically isolated Escherichia coli has been performed. In this analysis, it was shown that determination of the correct H type of a clinical E. coli strain was better achieved by MS than serotyping. Whole genome sequencing was used for the validation of this analysis.

by M. Chan, Dr H. Chui, D. Hernandez, Dr G. Wang and Dr K. Cheng

Background
During outbreaks of disease caused by pathogenic Escherichia coli, it is important to be able to identify the precise E. coli strain involved in order to track and prevent the spread of infection. Typically, the identification of E. coli strains has been based on the serotype of the cell surface antigens such as the lipopolysaccharide O antigen and the flagellar H antigen. Even though serotyping is viewed as the ‘gold standard’ for O- and H-type determination, this technique does have its downfalls. These conventional serotyping methods are based on antisera, which makes procedures costly and laborious to perform because of the variable quality of antibody preparations and the number of antibody agglutination reactions needed to assign a final classification [1, 2, 3]. Also, when bacterial cells do not generate lipopolysaccharide on the surface, the cultured colonies become ‘rough strains’. This makes both O- and H-antigen identification by antibody-based agglutination problematic despite the cellular motility and presence of the flagella H-antigen structure [1, 3, 4]. In addition, flagellar serotyping needs the induction of motility, which can take up to two weeks, and so does not result in the fast identification required in an E. coli outbreak situation [5].

A promising technique for E. coli H-type identification is the use of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, termed ‘MS-H’. In our own analytical assays, we harvest, enrich and digest the flagella proteins. LC-MS/MS uses liquid chromatography to separate flagellar fragments after trypsin digestion, and mass spectrometry to analyse and determine the protein sequences of these fragments. The MS results are then analysed against a curated database of H antigen variants to determine the H type. Compared with serotyping, MS-H has a high throughput, requires less labour, needs less time to perform, and provides sequence-level information.

Comparative H typing of E. coli with serotyping and MS-H
Using reference strains of all the different forms of E. coli flagella, it was shown that all 53 different types of H antigen can be determined through MS-H. Table 1 details the comparisons between H serotyping and MS-H [1]. It is important to note that both methods can reach 100% sensitivity and specificity for H-type determination. However, MS-H also provides sequence-level identification of the H type. This is important because this provides more reliable results, while antisera serotyping only provides a visual conformation, making there more of a chance for subjective and inaccurate determination of the H type.

Flagella motility induction is commonly used in serotyping. This increases the time-length of the procedure and requires more hands-on involvement [5]. In comparison, MS-H uses motility induction much less frequently, which makes MS-H identification of E. coli H type quicker and less labour-intensive.

Clinically isolated strains of E. coli that had already had their H types determined by serotyping were supplied from three provinces within Canada. A preliminary H-type analysis of the same E. coli isolates was performed by MS-H [5]. The workflow that was completed to analyse the H type of these strains is detailed in Figure 1.

Things to note within the workflow are that vortexing the suspended culture in Step 2 was to shear the flagella away from the E. coli. Also in Step 2, the flagella were trapped onto a filter membrane. This serves as an isolation step to separate the flagella from the supernatant as well as an enrichment process to concentrate the flagella. We then compared the flagellar sequence obtained by MS to a curated database formed from NCBI flagella protein sequence entries. A curated database was necessary because a public database can be problematic because of size, specificity and inconsistent annotation of the data for the specific flagellar protein entries. The curated database focuses on the flagellar proteins with fit-for-purpose annotations, thus allowing differentiation between the 53 different H types better than the public database [6].

On comparison of the H types identified by serotyping and MS-H, the majority of the results agreed with one another. However, there were some discrepancies between the two methods. In these cases, whole genome sequencing and polymerase chain reaction (PCR) detection was used to identify the correct H type [5] and these results predominantly agreed with those obtained by MS-H.
Even though PCR as a form of detection for the flagella gene is popular, the accuracy for determining all forms of flagella types through PCR is low and the use a many different primers had to be implemented, making it not ideal for the detection of an unknown H type [5]. Whereas whole genome sequencing may be more expensive then PCR detection, it was much more accurate at determining the correct H-type allele than PCR detection. Thus moving into the confirmational assays for the comparative analysis of MS-H determination and H serotyping, whole genome sequencing was only used to resolve disputed results between the two methods as it is not exclusively representative of a host’s flagella phenotype. On the other hand serotyping and MS-H is representative of the bacteria’s H type. Whole genome sequencing isn’t ideal for identifying clinical strains of E. coli for it is laborious and a long process compared to MS-H which is faster and contains less labour-intensive.

MS-H in clinical laboratories
Having shown that MS-H determined E. coli H type with high accuracy and in a short time frame, the application of H typing through MS-H could become very useful for the identification of unknown E. coli strains in a clinical laboratory. This is especially useful in the identification of an E. coli strain during a disease outbreak [7]. Faster identification of the pathogenic strain of E. coli would in turn also help combat the pathogenic E. coli more quickly.

Without the use of antibodies for agglutination and procedures for motility induction that are required in serotyping, MS-H is much less laborious in comparison. This would greatly benefit the professionals in the medical microbiology and public health laboratories who perform H typing.

Advantages and disadvantages of MS-H
It is significant that MS-H better determined the H type compared to conventional serotyping. As MS-H is faster, less laborious, provides sequence-level identification and has a higher throughput than serotyping, MS-H can be very useful in rapid and accurate identification of E. coli flagellar antigens. This may be a little over-idealistic as LC-MS/MS machines are very expensive and serotyping has been around for a very long time. It might be difficult currently for clinical laboratories to afford a LC-MS/MS machine or to change their workflow to incorporate MS-H as their method of H determination. However, mass spectrometer platforms are becoming more common in clinical laboratories. The uses of matrix-assisted laser desorption/ionization (MALDI) mass spectrometers have shown varying successes in microbiological identification. Also the use of MS isn’t just limited to H typing. MS could extend to the determination of the lipopolysaccharides (O antigen) of E. coli, toxins, and other relevant molecules within the spectrum of the mass spectrometer. This would not only determine the surface antigens of pathogenic E. coli, it would also give a broader profile of a pathogenic E. coli.

Conclusion and future directions
MS has potential to determine the H antigen type of E. coli better than serotyping, as shown in a comparative study between MS-H and serotyping through the use of a mass LC-MS/MS platform. MS-H provides sequence level identification of the flagellum type, whereas serotyping only provides visual agglutination assays for positive results and is therefore more prone to errors such as false positives and misidentification. Also, without the need for antisera, MS-H is less laborious, requires less time, and has a high throughput.
With the completion of the preliminary assay results of the comparative study between MS-H and serotyping in the determination of the flagella type, we are currently working on a country-wide thorough validation of the platform.

References
1. Cheng K, Drebot M, McCrea J, Peterson L, Lee D, McCorrister S, Nickel R, Gerbasi A, Sloan A, Janella D, Van Domselaar G, Beniac D, Booth T, Chui L, Tabor H, Westmacott G, Gilmour M, Wang G. MS-H: A novel proteomic approach to isolate and type the E. Coli H antigen using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS). PLoS One 2013; 8(2): 1–12.
2. Tenover FC, Arbeit RD, Goering RV. How to select and interpret molecular strain typing methods for epidemiological studies of bacterial infections: A review for healthcare epidemiologists. Molecular Typing Working Group of the Society for Healthcare Epidemiology of America. Infect Control Hosp Epidemiol. 1997; 18(6): 426–439.
3. Machado J, Grimont F, Grimont PA. Identification of Escherichia coli flagellar types by restriction of the amplified fliC gene. Res Microbiol 2000; 151(7): 535–546.
4. Edwards PR, Ewing WH. Edwards and Ewing’s Identification of Enterobacteriaceae, p536. Elsevier 1986. ISBN 978-0444009817.
5. Cheng K, Sloan A, Peterson L, McCorrister S, Robinson A, Walker M, Drew T, McCrea J, Chui L, Wylie J, Bekal S, Reimer A, Westmacott G, Drebot M, Nadon C. Knox D, Wang G. Comparative study of traditional flagellum serotyping and liquid chromatography-tandem mass spectrometry-based flagellum typing with clinical Escherichia coli isolates. J Clin Microbiol. 2014; 52(6): 2275-2278.
6. Cheng K, Sloan A, McCorrister S, Babiuk S, Bowden TR, Wang G, Knox D. Fit-for-purpose curated database application in mass spectrometry-based targeted protein identification and validation. BMC Res Notes 2014; 7: 444.
7. Cheng K, Sloan A, McCorrister S, Peterson L, Chui H, Drebot M, Nadon Celine, Knox D, Wang G. Quality evaluation of LC-MS/MS-based E. coli H antigen typing (MS-H) through label-free quantitative data analysis in a clinical sample setup. Proteomics Clin Appl. 2014; 8: 963–970.

The authors
Michael Chan^*1,2, Huixia Chui^1,3 MD, Drexler Hernandez^1,2, Gehua Wang^*1 MD and Keding Cheng^*1,4 MD
¹National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB, Canada
²University of Manitoba, Winipeg, MB, Canada
³Centre of Disease Control and Prevention, Henan Province, PR China
⁴Department of Human Anatomy and Cell Sciences, Faculty of Medicine, University of Manitoba, Winnipeg, MB, Canada

*Corresponding authors
E-mail: M. Chan, umchanm@myumanitoba.ca; G. Wang, gehua.wang@phac-aspc.gc.ca; K. Cheng, keding.cheng@phac-aspc.gc.ca.

Over the last few years, hepatitis C virus (HCV) infection has emerged as one of the most significant causes of chronic liver disease worldwide, with estimated prevalence ranging from 2.2 to 3.0%. Since January 2011, the Infectious Diseases Department of San Raffaele Scientific Institute in Milan carried out a prevention programme called ‘EASY test project’, to diagnose the HCV infection. In these four years a total of 35,000 subjects have been approached to inform them about HCV infection and other sexually transmitted diseases. Of the total eligible volunteers, 6500 (18,6% of contacted subjects) performed HCV tests on saliva and completed the interview in the alternative ‘street lab’. We believe that increasing awareness of these alternative tests among individuals at risk and providers may be an appropriate strategy to increase the number of people who know their serological status and who could be linked to care and engaged in care!

by M.R. Parisi, Dr L. Soldini, Dr G. Vidoni, Dr K. Schlusnus, Dr F. Dorigatti, and Prof. A. Lazzarin

Background
Over the last few years, hepatitis C virus (HCV) infection has emerged as one of the most significant causes of chronic liver disease worldwide, with estimated prevalence ranging from 2.2 to 3.0% (1).

In our country, the proportion of subjects infected with HCV is approximately 2% of the general population with a gradient that increases from the north to the south and the islands and with age (60% of patients with hepatitis C are over 65 years old). It is estimated that about 1 million people in Italy are ill with hepatitis C (2).
As acute HCV infection is usually asymptomatic, early diagnosis is rare. Those people who are developing chronic infection, even though undiagnosed, may suffer serious liver damage. In fact, a significant proportion of HCV infected subjects will ultimately progress to liver cirrhosis and/or hepatocellular carcinoma, making chronic HCV infection a major health problem (3, 4).

Despite the excellent accuracy of the tests currently available for the detection of anti-HCV antibodies (anti-HCV), the delay in reporting the results, the need for specialized equipment for processing the samples and interpreting the results, as well as the need to transfer individuals to sample collection and processing centres, limit their use as screening tools. Serologic points-of-care tests (POCTs) have several advantages, namely that they require little specialized apparatus, can be brought to the individuals who are to be tested and allow diagnosis in as little as a few minutes in different clinical settings (5). These advantages might be translated into increased testing opportunity and, ultimately, identification of more patients who could benefit from antiviral treatment (6). Over the last few years, several tests for rapid detection of anti-HCV have been developed and are currently in use in various countries; however, only recently, the first POCT was approved by the U.S. Food and Drug Administration (7). The investigation of the diagnostic accuracy of POCTs and rapid tests for the detection of anti-HCV is a highly relevant topic. As well as the great importance of the issue in terms of public health, there is a lack of studies evaluating the performance of several of the currently used tests.

EASY test project
Since January 2011, the Infectious Diseases Department of San Raffaele Scientific Institute in Milan carried out a Prevention Program called “EASY test project”, using the new oral test (rapid and non-invasive) the OraQuick® HCV Rapid Antibody Test (OraSure technologies, Inc.) to diagnose the HCV infection. The test is a single-use, immunoassay for the qualitative detection of antibodies to hepatitis C virus (anti-HCV) in oral fluid, fingerstick whole blood, venipuncture whole blood and plasma specimens. The HCV rapid test received the FDA approval for use with oral fluid on 28 June 2010.

The clinical sensitivity and specificity of the OraQuick HCV test using oral fluid were 97.8% (95% confidence interval [CI]) and 100% (95% CI, 98.4-100%), respectively (8). 

The main objective of the project is to evaluate the acceptability of an alternative, free and anonymous HCV test offer, available in different settings (in Points of Care, STDs Prevention clinics and General Practitioner surgeries) (9, 10). Furthermore, contacting the ‘hard-to-reach’ people with this anonymous and free test offer could reduce or stop this public health problem, by making an easy link to healthcare.

Subjects who underwent the test were asked to complete an anonymous questionnaire, through which it has been possible to collect a series of data on risk behaviours of the population tested. The questionnaire was devised with the intention of collecting demographic and risk behaviour data, as well as previous HCV/HIV testing experience, information about sex, drug use, educational level, nationality, general behaviours, use of HIV/HCV prevention services, previous surgical practices, invasive diagnostic practices, dental cares, tattoos or sexually transmitted diseases. Post-test counselling has been provided to all HCV reactive and non-reactive subjects, by the Infectious Diseases Department physicians involved in the study. The test was been carried out by a biologist or a practitioner, following the manufacturer’s procedures.

If the HCV oral test provided a preliminary positive result, a venipuncture was performed immediately for standard test confirmation, supported by the post-test counselling.

The results were received in two working days. At this point, the HCV-positive patient was contacted directly by the infectious diseases specialist for the visit and the diagnostic procedures to define the liver disease status and eventually to start the treatment, according to the guidelines for when HCV viral load and genotype are identified.
In these four years a total of 35,000 subjects have been approached to inform them about HCV infection and other sexually transmitted diseases. Of the total eligible volunteers, 6500 (18.6%) performed HCV tests on saliva and completed the interview in the alternative ‘street lab’. From the questionnaires we know that this initiative has been much appreciated.

Discussion
In recent years, advances in detection technology made available a range of POCTs for different infectious diseases. It is now possible to screen and diagnose those conditions at primary healthcare settings, using minimally invasive tests. In the present study, a new POCT for HCV infection has been performed on oral fluid. The use of oral fluid is an attractive alternative based on the fact that collection of plasma or serum samples requires equipment and training, and is more time consuming.

The FDA-approved OraQuick HCV Rapid Antibody Test (OraSure Technologies) is one of the most studied rapid tests for the diagnosis of HCV infection.

The development of rapid alternative tests for the diagnosis of HCV infection is to facilitate access to testing to reduce the individual risk of disease progression and social costs.

Despite the excellent sensitivity and specificity of third-generation enzyme immunoassays (EIAs), the turnaround time for reporting test results is at least one day, thereby making it difficult to deliver the results to tested individuals at first visit. Rapid tests are formatted such that they do not require complicated instrumentation or testing by skilled technical staff. They potentially generate results within an hour and therefore may be used for point-of-care testing.  Rapid tests are obviously more expensive than conventional immunoassays and are not designed for testing large batches of specimens. However, in no-clinical settings and laboratories that conduct low-volume testing, adoption of rapid oral testing can be cost-effective. CDC guidelines formulated for confirming screening anti-HCV results remain to be refined to accommodate rapid anti-HCV testing. It is important to emphasize that OraQuick HCV test has not been approved for general screening. A positive result of a rapid anti-HCV positive test is indicative of the presence of anti-HCV and, again, does not indicate active infection (11).

We successfully conducted this rapid HCV testing and counselling programme with the goal of spreading the use of saliva test anonymously and free of charge. We aim to facilitate access to testing in alternative settings, in order to understand if the ‘hard-to access’ population would access salivary rapid testing versus the conventional settings.

Increasing awareness of these alternative tests among individuals at risk and providers may be an appropriate strategy to increase the number of people who know their serological status and who could be linked to care and engaged in care!

The recent introduction of rapid oral HCV antibody test could completely change the HCV diagnosis approach by facilitating the possibility of testing millions of people worldwide (in particular in the developing countries).

For these reasons, we hope the oral-fluid based rapid HCV tests could become the ‘gold standard’ to facilitate the HCV screening access and become the standard of care and the basis for the national HCV testing algorithm in many countries with spread HCV epidemic, also in the dental care surgeries.

References
1. Lavanchy D. The global burden of hepatitis C. Liver Int. 2009; 29: 74–81.
2. Istituto Superiore di Sanita (ISS). Available at: www.iss.it.
3. Hoofnagle JH. Hepatitis C: the clinical spectrum of disease. Hepatology 1997; 26: 15S–20S.
4. Hutin Y, Kitler M, Dore G, Perz J, Armstrong G, Dusheiko G, et al. Global burden of disease (GBD) for hepatitis C. J Clin Pharmacol. 2004; 44: 20–29.
5. Ferreira-Gonzales A, Shiffman ML. Use of diagnostic testing for managing hepatitis C virus infection. Semin Liver Dis. 2004; 24: 9–18.
6. Tucker JD, Bien CH, Peeling RW. Point-of-care testing for sexually transmitted infections: recent advances and implications for disease control. Curr Opin Infect Dis. 2013; 26: 73–79.
7. Food and Drug Administration. Available at: http://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfClia/detail
8. OraQuick HCV Rapid Antibody Test. Available from:
http://www.fda.gov/MedicalDevices/productsandMedicalProcedures/DeviceApprovalsandClearances/Recently-approved-Devices
9. Parisi MR, Soldini L, Di Perri G, Tiberi S, Lazzarin A, et al. Offer of rapid testing and alternative biological samples as practical tools to implement HIV screening programs. New Microbiol. 2009; 32(4): 391–396.
10. Parisi MR, Soldini L, Vidoni GM, Clemente F, Mabellini C, Belloni T, Nozza S, Brignolo L, Negri S, Rusconi S, Schlusnus K, Dorigatti F, Lazzarin A. Cross-sectional study of community serostatus to highlight undiagnosed HIV infections with oral fluid HIV-1/2 rapid test in non-conventional settings. New Microbiol. 2013; 36(2): 121–132.
11. Center for Disease Control and Prevention. Recommendations for prevention and control of hepatitis C virus (HCV) infection and HCV-related chronic disease. Available from: http://www.cdc.gov/hepatitis/hcv/Management.htm.

The authors
Maria Rita Parisi^*1 MSc, Laura Soldini² MD, Gianmarino Vidoni³ MD, Karin Schlusnus⁴ PhD, Fernanda Dorigatti² MD, Adriano Lazzarin¹ MD
¹Division of Immunology, Transplantation and Infectious Diseases, Vita-Salute University, San Raffaele Scientific Institute, Milan, Italy
²Laboraf Diagnostic and Research OSR S.p.A., San Raffaele Scientific Institute, Milan, Italy
³Prevention Department, Reference Centre for HIV and STDs, Local Public Health Unit, Milan, Italy
⁴ANLAIDS Lombardia Onlus, Milan, Italy

*Corresponding author
E-mail: parisi.mariarita@hsr.it

External Quality Assessment (EQA) is the cornerstone of quality assurance and method validation in clinical testing labs in the UK, ensuring that the results of patient investigations are reliable and comparable wherever they are produced. In this article we focus specifically on EQA for laboratories performing trace element measurements, although many of the points are applicable to the wider pathology areas.

by S.-J. Bainbridge and Dr C. F. Harrington

Introduction
External Quality Assessment (EQA), also termed proficiency testing (PT), involves the regular distribution of test materials to participating laboratories so that they may evaluate their analytical performance against a peer-group, detect any accuracy or other problems that may develop with the assay and so improve the results that they produce. The key elements that differentiate EQA from PT include: education and support; identification of method poor performance; and method evaluation [1].

Historically clinical science was one of the first disciplines to realize the usefulness of EQA and take steps to implement schemes that would be of use in the hospital laboratory. The first proficiency survey of UK clinical pathology laboratories was reported in 1953 and revealed a wide spectrum of results for the common tests [2]. Further surveys in the 1950s and 60s confirmed the need for regular PT. In 1969, the National Quality Control Scheme was initiated by the Wolfson Research Laboratories, Birmingham and involved the distribution of specimens every 14 days [2]. This is now known as the UK National External Quality Assessment Scheme (UKNEQAS) and is responsible for about 30 different schemes.

In 2013, the importance of EQA in the NHS pathology services was emphasized by Dr Ian Barnes in a Department of Health review into quality assurance [3]. The review assessed current NHS quality assurance frameworks and governance mechanisms for pathology services. It gathered a diverse range of evidence: examining expectations of pathology services; identifying areas for improvement; and recommending a system-wide way forward. It recommended strengthening and standardizing the current quality assurance structures that are in place, which are based on the Royal College of Pathologists (RCPath) Joint Working Group for Quality Assessment (JWGQA), which co-ordinates and oversees the standards and performance of EQA schemes for all schemes regardless of provider.

EQA for trace elements
The Trace Elements External Quality Assessment Scheme (TEQAS), which is part of UKNEQAS but based in Guildford, UK, was established in 1979 with distribution of specimens on a monthly schedule to UK hospital laboratories measuring copper and zinc in serum. During the next five years the scheme developed with inclusion of other participants and the introduction of additional analytes and specimen types. Following an international conference on Aluminium and Renal Disease in 1986, a two year arrangement was established with the EU Commission to fund participation in the serum aluminium programme for European laboratories involved with the monitoring of patients with chronic renal failure. An outcome of this work was the realization that analytical standards of performance for this measurement were very poor. In collaboration with the UK Department of Health it was proposed that the scheme should be linked to UKNEQAS in order to provide a mechanism for referral of poor performers to the Clinical Chemistry Advisory Panel. This link was formally established by the Advisory Committee on Analytical Laboratory Standards in 1988. The aims of TEQAS are consistent with the intentions of UK NEQAS, to:

• Provide professionally-led and scientifically-based schemes with a primarily educational objective.
• Provide regular distributions of specimens.
• Provide rapid feedback of performance.
• Support participants where problems occur.
• Stimulate the overall improvement in performance among all participating laboratories.

The main TEQAS scheme provides EQA for: Al, Cr, Co, Cu, Se and Zn in serum; As, Cd, Cr, Co, Pb, Mg, Mn, Hg, Se, Tl and Zn in whole blood; and As, Cd, Cr, Co, Cu, Fe, Pb, Mn, Hg, Ni, Tl and Zn in urine. This operates on a monthly cycle, with the results from the measurement of two specimens being returned on-line on the last day of each month. The report is then available five days later to download. Two smaller schemes providing Al in dialysis fluid and Cu and Fe in solid matrices are also available, but have a more limited number of participants.

The main steps in the EQA process
For a better appreciation of the overall EQA scheme it is useful to divide it into a number of process-based areas as shown in Figure 1. The activities that comprise these areas are also shown. Some of these areas come under particular focus as part of ISO 17043:

Design
Appropriate design of the PT scheme ensures that participants will have paid for a service that provides high quality comparable test items that are representative of patient samples they would normally expect to analyse. These test items will have undergone thorough assessment procedures in accordance with acceptable statistics to ensure a homogenous and stable test item.

Materials
The participants can be assured that the materials used in the production of the PT samples have been ethically and legally obtained and that the competence of the suppliers have been evaluated and verified to ensure their products or services do not affect the quality of the PT scheme.

Evaluation
ISO 17043:2010 ensures the evaluation of the participants performance is conducted fairly and consistently guaranteeing they receive an accurate evaluation calculated from the use of robust statistical methods.

Knowledge and Experience
In addition to all this ISO 17043:2010 ensures that the operations of the PT scheme are carried out by personnel that have the training, skills and competence necessary to professionally carry out their assigned tasks. Participants can feel secure in knowing that they have access to the specialist knowledge and expertise in the field of trace element testing to be able to discuss any concerns they may have.

Establishment of performance criteria
Measurements of performance are based on deviations of results from target values, which are used to calculate a Z-score. As EQA has developed, various organizations have produced documents that summarize best practice. Those from authoritative international bodies include:

• ISO 17043 (Conformity assessment – General requirements for PT).
• ISO 13528 (Statistical methods for use in PT by interlaboratory comparisons).
• IUPAC (The international harmonized protocol for the PT of analytical chemistry laboratories) [4].

All these documents recommend that assessment of performance should be based upon calculation of a Z-score (or a derivative which takes uncertainty into consideration).
The Z-score is calculated as:
x-X/ SD_PT
where     x = laboratory result,
             X = target value, and
             SD_PT = standard deviation for PT (also represented as σ)

The ‘standard deviation for PT’ is set by the scheme organizer but should ideally be a value that will allow the score to demonstrate whether or not the performance is fit for the purpose for which the assay is being used. It is recommended that this value be set so that a Z-score of up to ±2 indicates acceptable performance and a score of more than ±3 indicates unsatisfactory performance.
In the TEQAS scheme, we have used quality specifications based on biological variation for the ‘standard deviation for PT’ and the determination of these quality specifications has been published [5]. For assays where there is insufficient data to prepare specifications in this way we have produced values that are related to performance within the scheme during recent years.
The quality specifications and their corresponding SDPT for some illustrative elements are shown in Table 1. These are presented as either a percentage of the target value or a fixed value depending on the concentration of the target value, and the one used is whichever is the greater. This allows for the increase in imprecision at low concentrations and conforms to a ‘funnel’ shape.

Scheme accreditation
Accreditation is fast becoming a preferred mechanism for delivering confidence in UK Healthcare and with the application of BS EN ISO 15189:2012 into Medical Laboratories and its requirement for the laboratories to seek confirmation for confidence in their results, the need for EQA schemes in the relevant fields of medical laboratories is ever increasing. Participation in a suitable scheme can be an effective way of demonstrating the laboratories’ technical competence. ISO 15189:2012 requires laboratories to evaluate their PT providers and a recognized acceptable basis of their evaluation recommended by the UK Accreditation Service (UKAS) is the participation in PT schemes with those providers that have been accredited to ISO/IEC 17043:2010. This International Standard specifies criteria and the general requirements for the competence of the PT providers and their responsibility for all tasks in the development and operation of the PT scheme. Some of the main differences introduced with ISO 17043 are summarized in Table 2.
 
Assessment of conformance
When conducting an assessment of a PT scheme for conformance to ISO 17043:2010, the assessors will take a holistic approach looking at the management system as a whole. The assessment will include areas such as scheme organization, scheme management, evaluation processes, technical competence and impartiality and integrity. Each separate area of the PT scheme are all interlinked and therefore when accreditation is granted by the accreditation body (UKAS in the UK) it will not be given on a single fact but the overall competence of the PT provider. Accreditation to ISO 17043:2010 can be a hard and thorough task for PT providers to undertake but once accreditation is granted it provides the necessary assurance of a competent and professional scheme which can provide an open and honest service whilst maintaining confidentiality for all those participants enrolled in the scheme.

Summary
The 2013 Barnes review into quality assurance in the NHS pathology services reinforced the importance of quality assurance and this article has discussed the implications of recently introduced ISO standards for clinical pathology departments (ISO 15189:2012) as well as for EQA scheme providers (ISO 17043:2010). This strengthens and standardizes the systems used in clinical testing laboratories and ensures high quality and comparable results for patient tests.

References
1. James D, Ames D, Lopez B, Still R, Simpson W, Twomey. External quality assessment: best practice. J Clin Pathol. 2014; doi: 10.1136/jclinpath-2013-20621.
2. Bullock DG. External quality assessment schemes for clinical chemistry in the United Kingdom. Ann Ist Super Sanita 1995; 31: 61–69.
3. Barnes I. Pathology Quality Assurance Review 2014. www.england.nhs.uk/wp-content/uploads/2014/01/path-qa-review.pdf
4. Thompson M, Ellison SLR, Wood R. The International Harmonized Protocol for the proficiency testing of analytical chemistry laboratories (IUPAC Technical Report). Pure Appl Chem. 2006; 78: 145–196.
5. Arnaud J, Weber J-P, Weykamp CW, Parsons PJ, Angerer J, Mairiaux E, Mazarrasa O, Valkonen S, Meditto A, Patriarca M, Taylor A. Quality specifications for the determination of copper, zinc, and selenium in human serum or plasma: evaluation of an approach based on biological and analytical variation. Clin Chem. 2008; 54(11): 1892-1899.
6. Summary of ISO 15189 additional requirements. CPA UK Ltd, 2012. http://www.ukas.com/Library/Services/CPA/Summary%20of%20Idifferences%20betwen%20ISO%2015189%20&%20CPA.pdf

The authors
Sarah-Jane Bainbridge and Chris F. Harrington* PhD
TEQAS, Trace Element Centre, Surrey Research Park, Guildford GU2 7YD, UK

*Corresponding author
E-mail: Chris.harrington1@nhs.net

Two opposing agendas confront clinical labs in terms of electronic health records (EHRs): privacy/security on the one side, and interoperability, on the other. The former involves an inward push for isolation, while the latter tends to pull technology in the other direction.
There also is a major financial challenge. While healthcare providers have been given a host of incentives to adopt EHRs (especially in the US), labs have been pretty much left out on their own.

EHRs and lab systems populate different worlds
Clearly, lab-compatible EHR systems which meet both (privacy and interoperability) criteria promise the quickest returns. EHR developers have however shown little enthusiasm, until recently, to incorporate clinical lab requirements as a sufficient driver, while laboratory system vendors have tended to ignore EHRs or postpone taking them into account until EHR development has matured sufficiently. 

US EHR adoption drives lab applications
In the US, this limbo is being shaken up by healthcare providers, who are compelling vendors to take account of their need for EHR-friendly clinical lab systems.
At end 2012, the US Centers for Disease Control and Prevention (CDC) released a survey which found 72 percent of office-based physicians using EHR systems, up from 48 percent in 2009 and 18 percent in 2001. 
The reason for the dramatic increase in EHR adoption lies in the Meaningful Use requirements of the 2009 Health Information Technology for Economic and Clinical Health Act, also known as the HITECH Act. The Act provides billions of dollars in incentive payments through the Medicare and Medicaid programmes to increase physician adoption of EHR systems.

Clinical labs are now being lifted by the rising tide of EHR adoption. According to the US Office of the National Coordinator for Health Information Technology (ONC), the “availability of structured lab results within the EHR contributes to office efficiencies while also assisting providers in the ability to make real time decisions about the patient’s care.”
The ONC explicitly specifies the threshold for EHR-friendly clinical lab practices in Stage 1 – of over 40 percent of all lab test results ordered by a provider and incorporated in certified EHR technology as structured data.
Stage 2 Meaningful Use requirements, finalised in August 2012, increase the clinical lab results threshold to 50 percent. The ONC has subsequently announced plans to assess health information exchange (HIE) in clinical laboratories.

Labs left to own resources
While healthcare providers have the financial incentives of the HITECH Act, clinical labs have been left to their own resources to set up interfaces from their laboratory information systems (LIS) to providers.
Compounding this has been inconsistencies in the way different EHR systems generate lab test orders.
However, the alternative has been stark – to be left out of referrals from tests.

EHR systems remain heterogeneous
The US EHR landscape is however hardly uniform. As of September 2013, there were 3,652 non-enterprise certified ambulatory EHR software systems, almost half of which were classified as “complete” to qualify for Meaningful Use Stage 1 or Stage 2.
In spite of efforts to set standards for semantic interoperability of healthcare data, standards so far are only syntactic (based on HL7 and XML). 
The alternative, to develop a common US-wide EHR system, has been accepted as being technically insurmountable – due to hurdles in specifying, developing, testing and deploying standardized tools, common architectures and vocabularies, within secure, real-time and scalable networks, and doing all this within the fast-changing world of information and communications technologies.
For proponents of a decentralized approach to EHR technology, in the US in particular, the sharp increase in offtake of EHR systems has shown that it has delivered – as far as healthcare IT objectives are concerned.

EHR faces teething problems
Still, teething troubles for EHRs also clearly remain.
In early September 2013, one of the leading EHR systems, from EPIC, crashed across seven major healthcare facilities of Sutter Health, a nearly 100 year-old healthcare provider in California. Some suspect the role of a routine upgrade a few days earlier in the EHR system, which was launched by Sutter at a cost of $1.2 billion in 2004, but has so far reached only a halfway mark.

EHR challenges for labs remain to be resolved
Such issues with the evolution of maturity of EHRs pose especially major problems for labs, who (as mentioned) have to develop and fund interfaces between their LISs and the EHRs of their client physicians but are also forced to cope with the lack of uniform EHR standards.

Some vendors have nevertheless sought to fill the gap.
A leading example is HDD Access, a joint initiative by the US Department of Defense, the Department of Veterans Affairs and 3M Health Information Systems to create a public use version of 3M’s Healthcare Data Dictionary (HDD). HDD Access consists of a relational database and Application Programming Interface (API) runtime services to which other applications can interface. The terminology is organized as a controlled medical vocabulary – a comprehensive set of clinical and other concepts used in healthcare.
HDD Access offers specific benefits for integrating LIS and EHR platforms. Independent of source system, it can track local fields and translate them into laboratory concepts. Nevertheless, HDD Access warns that it is “not a standard terminology and is not a replacement for standard terminologies.

In effect, in the US, clinical labs are likely to continue to face a host of technical challenges with respect to EHRs in the years to come.

EHR Big Bang fizzles in Europe
Unlike the US, Europe made a massive effort in 2004 to devise common semantic standards for EHR interoperability as part of its Single eHealth Area. The EU’s EHR objectives sought to integrate all patient information – from primary to tertiary settings, and include emergency and in-patient care. Also on the radar were ambitious plans to connect pharmacies as well as the web of disparate billing/reimbursement procedures, and do so across Europe.
In mid–2008, the EU Commission set 2015 as the target year for EHR interoperability, to ensure that key EHR datasets could cross European borders, and do so in conformity with medical rules and other relevant legal frameworks.

In January 2011, however, these ambitions were put on the backburner, after an official report criticized the effort as being both impractical and ‘grandiose’. The report found that a pan-EU EHR system would neither be technically feasible, cost-effective or even medically justified, and instead urged more emphasis on decentralized efforts – in other words, just like the US.
Technical challenges aside, massive differences in physician and medical cultures across Europe played a major role in derailing efforts toward a common EHR. Or, as EuroRec, an umbrella organization tasked with pan-EU EHR implementation, states: it was “widely recognized that social and organizational aspects are as likely to ruin an implementation process as technical factors are.”

European focus shifts to national efforts
The EHR focus in Europe has now totally shifted to national efforts. A new eHealth Governance Initiative (eHGI) encourages cooperation “between Member States” and “between national authorities and standardization bodies”, and seeks to “enable the recommendation of standards and (harmonized) profiles based on selected use cases.” On the technical side, compared to the Big Bang efforts of the Single eHealth Area, it also aims to “link and harmonize coding systems” and “facilitate access to existing standards and medical vocabularies.”
The second area for Europe’s EHR focus is a minimalistic intra-EU/regional approach embodied in a project called epSOS, which dates back to 2008, but was (temporarily) eclipsed by the ambitions of the Single eHealth Area. epSOS, which went live in April 2012, has the modest goal of connecting 20 EU nations (and 3 non-EU members) to a secure database, and sharing only Patient Summaries and ePrescription records via IHE X* profiles. Its target consists of Europeans holidaying overseas.

Today, EHR adoption varies considerably in Europe. The Nordic countries have been using the technology for over a decade and are fairly advanced as a result in EHR implementation.
However, adoption in France, Germany, Spain and the UK is ‘on course’ with the US.

Shift from Single eHealth Area encourages new EHR-directed lab applications
The shift away from forcing through a Single eHealth Area has also opened the way for innovative working approaches aimed at clinical labs. One good example of this is Valle de los Pedroches Hospital at Cordoba, Spain, which has  designed and implemented a unified lab test request module for the Andalusian regional EHR.
In spite of some outstanding issues (such as rigidity in error solving, and the need to adapt to a new nomenclature), implementation of the laboratory module in the EHR improved the analytical process, with better patient safety and less programming or container errors and shorter response times. Clinical professionals gave a rating of 7.8 out of 10, positively highlighting the speed at which results are delivered and their integration in the EHR.
Such efforts are likely to grow with time.