Laboratory diagnostics of chronic kidney disease in Croatia: state of the art

Introduction Early identification and management of chronic kidney disease (CKD) is highly cost-effective and can reduce the risk of kidney failure progression and cardiovascular disease. In 2014, the Joint Croatian Working Group (JCWG) for laboratory diagnostic of CKD on the behalf of Croatian society of medical biochemistry and laboratory medicine (CSMBLM) and Croatian chamber of medical biochemists (CCMB) conducted a survey across Croatian medical-biochemistry laboratories to assess the current practice in this area of laboratory medicine. The aim of this study was to present the data collected through the survey and to give insight about laboratory diagnostics of chronic kidney disease in Croatia. Materials and methods An invitation to participate in the survey was sent to all Croatian medical-biochemistry laboratories (N = 196). The questionnaire was designed in a form of questions and statements, with possible multiple answers, comprising 24 questions. Results The response rate was 80/196 (40.8%). 39 answers were from primary medical-biochemistry laboratories. 31/78 (0.40) laboratories measure creatinine with non-standardized method (uncompensated Jaffe method). 58/78 (0.74) of laboratories that measure creatinine do not report eGFR values. Similar number of laboratories (58/80, 0.73) do not measure urine albumin or protein. Conclusions There is a large heterogeneity among Croatian laboratories regarding measuring methods, reporting units and reference intervals (cut-off values), both for creatinine and urine albumin or protein. The two key prerequisites for CKD screening, automatic reporting of eGFR and albuminuria or proteinuria assessment, are not implemented nationwide. There is a need for harmonization in laboratory diagnostics of CKD in Croatia.


Introduction
Chronic kidney disease (CKD) is a general term for heterogeneous disorders affecting kidney structure and function with variable clinical presentations. The Kidney Disease: Improving Global Outcomes (KDIGO) CKD Working Group released new updated KDIGO 2012 Clinical Practice Guideline for the Evaluation and Management of Chronic Kidney Disease which pointed on the relevance of laboratory medicine in diagnosis of CKD (1). Early identification and management of CKD is highly cost-effective and can reduce the risk of kidney failure progression and cardiovascular disease by up to 50% and increasing the recognition of kidney damage is a key part of improving health outcomes in the community (2). The prerequisite for screening policies for chronic kidney disease is standardization of measurement of serum creatinine and automatic reporting of estimated glomerular filtration rate (eGFR) consequently as a marker of kidney function, and uniform assessment of Radišić Biljak V. et al. Joint CSMBLM and CCMB TF-CKD Croatian survey study albuminuria (or proteinuria) type of sample used and reporting units as a marker of kidney damage, nationwide (3). In 2014 Joint Croatian Working Group (JCWG) for laboratory diagnostics of CKD on the behalf of Croatian society of medical biochemistry and laboratory medicine (CSMBLM) and Croatian chamber of medical biochemists (CCMB) initiated proceedings to provide easily applicable national recommendations in this area of laboratory medicine. The initial step was to assess the current practice among Croatian laboratories in CKD evaluation, in order to set the background for developing national recommendations. The aim of this study was to assess the degree of adherence of medical laboratories in Croatia with KDIGO 2012 Clinical Practice Guideline for the Evaluation and Management of Chronic Kidney Disease.

Materials and methods
Questionnaire An e-mail invitation to participate in the on-line survey was sent to all Croatian medical biochemistry laboratories (N = 196). The e-mail was addressed to laboratory managers in order to avoid duplicate answers from one laboratory. The online survey was performed using the SurveyMonkey tool (Palo Alto, CA, USA), and it was held from April 3 rd until May 20 th 2014, with two reminders in that period of time. The survey was anonymous.
Questionnaire was designed in a form of questions and statements, with possible multiple answers and comprised 24 questions (   emphasize that only one laboratory among primary health care laboratories, and none of the private laboratories, determine serum creatinine using enzymatic method. The majority of laboratories (73/78) are using reference intervals according to the recommendations from the CCMB (4). However, a large proportion (16/78, 0.20) of laboratories are using incorrect reference intervals according to the stated method used for serum creatinine measurement. Five laboratories measure creatinine with uncompensated Jaffé method, but their reference intervals are for standardized creatinine. Eleven laboratories determine creatinine with standardized method (compensated Jaffé or enzymatic), but they are using reference intervals for non-standardized creatinine.

eGFR
The majority of laboratories generally do not report results for eGFR (58/78). There was a statistically significant difference in distribution of laboratories that report eGFR by type of institution (P < 0.001), with the lowest number of laboratories from primary health care institutions (Figure 1.).
Description of equations used for calculating eGFR by type of medical biochemistry laboratory,

Albuminuria/proteinuria
Majority of laboratories that participated in the survey do not measure urine albumin or protein (58/80). Out of those who measure urine albumin or protein the most prevalent method is immunoturbidimetric method (12/14) for urine albumin

Discussion
This is the first study to give insight about laboratory diagnostics of chronic kidney disease in Croatian medical biochemistry laboratories. The motive for this study was the new updated KDIGO 2012 Clinical Practice Guideline for the Evaluation and Management of Chronic Kidney Disease in which the role of laboratory medicine is of great importance. The key prerequisite for the screening of CKD are two simple laboratory tests: eGFR and assessment of albuminuria (or proteinuria) (5). The American Society of Nephrology strongly recommends all adults to undergo routine screening for CKD and some data suggest that screening for CKD is cost-effective in patients with diabetes and hypertension (6,7). However, there are no national Croatian guidelines regarding this area of laboratory medicine, as in some other countries throughout the world (2,3,8,9). This study describes current status of harmonization of creatinine and albuminuria (proteinuria) measurement in Croatian medical biochemistry laboratories.
Harmonization process regarding creatinine standardization in Croatia started in 2010 with the recommendations for revision of creatinine methods and reference intervals published by CCMB (4). This initial process was followed by a course about harmonization in laboratory medicine in 2011 (10). However, it is obvious from the survey results that creatinine standardization process in Croatian laboratories is not finished and only minority of laboratories automatically report eGFR values. The biggest problem raised from this survey results is a fact that there is discrepancy between methods used for creatinine measurement and creatinine reference intervals, i.e. reference intervals for standardized creatinine were used for creatinine results measured by non-standardized methods, and vice versa. Similar problem was observed with equations used for calculating eGFR. Also, reporting values for eGFR are not as recommended (all values greater than 60 mL/min/1.73 m 2 report as "> 60 mL/min/1.73 m 2 ", but are given as the exact calculated number for MDRD equation). Very small number of laboratories use CKD-EPI equation recommended by KDIGO guideline for calculating eGFR. These results can have serious implications on patient health and there is a need for better regulation in this area of laboratory work. One of the possible approaches can be through pre-and post-analytical modules of Croatian national external quality assessment scheme which is in the domain of Croatian Centre for Quality Assessment in Laboratory Medicine (CROQALM).
Survey results about biomarkers of kidney damage show even higher heterogeneity among Croatian laboratories. There are no recommendations about preferred type of urine sample for an initial screening, and large part of laboratories perform analyses in 24-hour urine samples, regardless of all known disadvantages (11). There is a high variability of reporting units; some laboratories still report absolute concentrations of urine albumin, regardless of the type of urine sample, although there were some efforts made in the area of laboratory diagnostics of diabetic nephropathy to standardize type of samples and reporting units used for measuring urine albumin (12). One of the major problems is the fact that laboratories in primary health care institutions do not measure urine albumin or protein at all.
Some of these problems were acknowledged at a course of laboratory diagnostics of chronic kidney disease held in 2013. However this survey shows that in spite of numerous education efforts from our national associations, the policies and practices concerning CKD in Croatian laboratories are neither uniform nor harmonized (13). For this reason we concluded that education must start at the beginning and provide simple and understandable recommendations and guidelines for initiative screening of CKD followed by comprehensive education through our national associations. Such conclusion is supported by a high number of responders that stated they are interested in further education in the field of implementation of clinical guidelines into everyday routine practice. However, the most troubling fact is that all 4 survey participants that declared against further education, had discrepancy between serum creatinine measurement methods and used reference intervals. One of the goals of this study is to raise awareness amongst Croatian laboratories about critical points in this area of laboratory medicine.
This study has several limitations, similar to those observed in a study conducted by Dukić and Šimundić (14): the survey was self reported and participants might not be providing the accurate and reliable information about the real situation; there was a low number of responders and majority were from primary health care institutions.

Conclusions
It is evident from the survey results that laboratory diagnostics of chronic kidney disease in Croatia is not standardized. Laboratories still use non-standardized methods for creatinine measurement and they do not report eGFR values. Most laboratories still measure urine albumin or protein in 24-hour urine sample, which is also the only recommended sample in some laboratories. There is a large heterogeneity in reference intervals and types of units in albuminuria (or proteinuria) measurement. It is essential to provide a framework for nationwide harmonization and standardization of laboratory procedures in this area of laboratory medicine.