Pre-Analytical Phase in Mycology

Pre-Analytical Phase in Mycology

*Fatiha Bousnina, Oumaima Nassiri, Oumaima kharkhach, Asmae Rhoubi, Ayoub Barroudi, Imad Mahi, Samira Laghzaoui, Aziza Hami

Parasitology-Mycology Laboratory, Mohammed VI University Hospital of Oujda. Faculty of Medicine and Pharmacy, Mohammed First University. Oujda, Morocco.

*Corresponding Author

DOI: https://dx.doi.org/10.47772/IJRISS.2024.803109

Received: 04 March 2024; Revised: 17 March 2024; Accepted: 22 March 2024; Published: 12 April 2024

ABSTRACT

Fungal infections remain a frequent cause for medical consultation, representing a prevalence of 20-25%. Mycology laboratory represents the key to the management of fungal diseases, whose objective lies not only in providing accurate analysis results, as well as completing them within a satisfactory time, maintaining traceability for all laboratory procedures, in accordance with ethical principles, while ensuring the safety of both patients and staff. The pre-analytical phase is the most complex part of the biological process, and some steps may escape the attention of biologists, since this phase involves several external factors beyond the laboratory, which requires the management of a massive flow of information. or this reason, the implementation of a quality system allows the detection and minimization of errors occurred during each stage of this phase, in particular the ISO15189 standard and the Guide of proper execution of analysis. Through this work, we have reviewed the main stages of the pre-analytical phase in mycology, from prescription to sample preservation, focusing on the requirements of this process, as well as the primary investigations to be conducted for each type of mycological specimen.

Keywords: pre-analytical phase, mycology laboratory, samples, steps.

RESUME

Les infections mycosiques demeurent un motif fréquent de consultation médicale avec une prévalence de 20 à 25 %. Le laboratoire de mycologie représente la clé de la prise en charge des maladies mycologiques dont leur objectif ne réside pas seulement dans la fourniture de résultats d’analyses exactes, mais également dans le fait de les réaliser dans un délai satisfaisant, en assurant une traçabilité de toutes les procédures du laboratoire, dans le respect de l’éthique, en assurant à la fois la sécurité des patients et du personnel. La phase pré-analytique est la phase la plus complexe du processus biologique, certaines étapes peuvent échapper aux biologistes puisque cette phase fait intervenir plusieurs acteurs extérieurs au laboratoire et nécessite la gestion d’un flux énorme d’informations. Pour cela la mise en place d’un système qualité permet de reconnaître et de minimiser les erreurs produites lors de chaque étape de cette phase notamment la norme ISO15189 et le Guide de bonne exécution des analyses (GBEA). A travers ce travail, nous avons rappelé les principales étapes de la phase pré-analytique en mycologie avec une mise au point sur les exigences de ce processus, ainsi que les principales recherches à effectuer sur chaque type de prélèvement mycologique.

MOTS CLES : Phase pré-analytique, laboratoire mycologie, prélèvements, étapes.

INTRODUCTION

Fungal infections remain a frequent cause for medical consultation. Superficial fungal infections of the skin and nails, as an example, have a high prevalence of 20 to 25% of the world population, that suffers from fungal diseases, while the most cases remain benign [1].

The mycology laboratory assumes a crucial role in the management of fungal diseases, by the identification of the mycosic agent of the infection [2]. The laboratory’s objective lies not only in providing accurate analysis results, but also in completing them within a satisfactory period of time, ensuring traceability of all laboratory procedures, respecting ethical standards and ensuring the safety of both patients and staff. Therefore, every medical analysis laboratory must have appropriate procedures to ensure the quality of the results delivered. However, biologists still face discrepancies between results and clinical practice [3]. These inaccuracies, which account approximately to 93% of errors detected during the biological diagnosis process, are mainly caused by the absence of standardization of pre-analytical procedures [4].

The pre-analytical phase includes all steps from the medical prescription to the receiving of the sample in the laboratory sampling, routing, reception, non-conformity management, recording, pre-processing, and preservation [5,6]. The pre-analytical phase is the most complex part of the biological process, and some steps may escape the attention of biologists, since this phase involves several external factors beyond the laboratory, which requires the management of a massive flow of information [7,8,9].

The implementation of a quality system allows the detection and minimization of errors occurred during each stage of this phase. Various international standards have been established to optimize error control in biological analyses and standardize practices across different laboratories, notably ISO 15189 and the Guide of Proper Execution of Analysis (GPEA). The GPEA represents the first regulatory reference concerning quality in Morocco [10]. It focuses on the general standards for managing biological samples regarding sampling, identification, and preservation (Chapter II) [11]. The latest states that every laboratory must have procedures for the pre-analytical phase to ensure the quality of the results provided [9,12].  Furthermore, the advent of ISO 15189 as a mandatory reference for medical biology laboratories has led to a significant improvement in the quality of the pre-analytical phase (Chapter 4.7 and 5.4) [13,14]. For this reason, various requirements should be considered to control all the stages of this process [10].

Through this work, we have reviewed the main stages of the pre-analytical phase in mycology, from prescription to sample preservation, focusing on the requirements of this process, as well as the primary investigations to be conducted for each type of mycological specimen.

REQUIREMENTS OF THE PRE-ANALYTICAL PROCESS IN MYCOLOGY

• Analysis request:

This sheet must include all the information required for accurate identification of the patient and the prescriber, particularly the identity of the patient, the prescriber, and the sampler. This last mention is required by the GPEA and is part of the care traceability process. A specific case should be provided for this purpose on the transmission sheet. To provide credible results, the information concerning the sample should be mentioned [15], such as the specimen type, sampling site, the nature of prescribed analyses, as well as the date and, if necessary, the time of sampling. Certain clinical data relating to the patient are essential for interpreting and processing the samples. Such as a notion of travel to foreign country or contact with animals, for example, contact with cattle, should be investigated in cases of suspected dermatophytes infection of Trichophyton verrucosum [16]. The notion of immunodeficiency is also crucial for interpreting results in mycology, especially to differentiate colonization from infection. An antifungigram is more likely to be performed for fungi that would have been considered simple contaminants in an immunocompetent subject. All this information must be stated clearly and precisely to minimize possible errors. Finally, the samples should be ideally taken prior any antifungal treatment that may inhibit the in vitro growth of the fungus [17,18]. The intake of antifungals, such as for nails infection, must be mentioned on the prescription sheet.

• Sampling procedure:
  • Sample collection:

Sampling is a decisive step in establishing a mycological diagnosis [2]. Therefore, mycological examination must be carried out at a distance from any application of antifungal treatment [17,18]. For a nail examination a therapeutic time window of approximately 15 days must be respected after prescribing a filmogenic solution or a nail polish with ciclopirox as the active substance, and at least 3 months after taking antifungal oral medication [17]. In the case of tinea corporis or tinea capitis a window of 4 weeks after applying an antifungal treatment must be respected [15]. A toilet with a PH neutral soap is recommended before sampling [15]. Cosmetic nail polish should be avoided [15].  As well as Avoid applying cream or other substances: before taking any samples from the nails and skin, those substances will interfere with the direct examination. Most of the samples must be taken inside a laboratory by an expert biologist, such as in the case of dermatophytes research, and some samples must be taken as soon as possible if the prognosis is life-threatening, as example, the research cryptococcosis infection [19].

• • Identification of sample: The sample should be identified by the patient’s first and last name, date of birth, and patient ID [4]. For skin samples and biopsies, the sampling site must also be indicated.
  • Sampling methods: Detailed methods for mycological sampling are summarized in tables (Tables I and II).

To standardize practices, it is essential to provide prescribers with a documented sampling manual. This will entail general recommendations for patient preparation, sample identification, sampling steps, sample type and quality, and the time of sampling [4,9]. This meets the requirement of ISO 15189 in chapter 5.4.4: “The specimen collection manual must be part of the document control system”.

Material: The quality of mycological sampling is essential for the quality of the results obtained during the examination. This condition is particularly important for skin samples, for which many false-negative results are due to improper execution of the sampling. A high mycological quality of sample cannot be taken without a minimum of equipment, including sterile Brocq curette or sterile Vidal scrapers.

The choice of equipment will depend on the location of the lesion. Similarly, a preliminary examination under a Wood’s lamp is necessary to identify lesions of pityriasis versicolor and differentiate them from erythrasma, while it is rarely used to localize tinea capitis, thus differentiate microsporosis from trichophytosis and favus tinea capitis [2,17,20]. Although they have little influence on the quality of the analysis, sterile Petri dishes made of glass are preferred instead polystyrene Petri dishes for the collection of epidermal scales [21]. Similarly, in the case of onychosis to use scissors to trim the upper nail plate, which is typically not very contributory for dermatophytes investigation [2]. And it’s essential to always remember that universal guidelines recommend wearing gloves for every sampling procedure [22].

• Sample routing:

The transport of samples to the laboratory must respect the time and temperature appropriate to each type of analysis in the sampling manual [4]. In this regard, the prescription sheet, which includes the date and time of sampling, ensures traceability to the laboratory and thus confirms the absence of any non-conformity [4]. Each non-conform sample to the recommendations is managed in a special way, both the sampler and the prescriber are informed through a non-conformity form. Furthermore, most samples do not present any particular constraints for their routing to the laboratory. A maximum delay of 24 hours at room temperature is conceivable for most samples. It can even be extended if the sample is stored at 4°C [2,12,15]. Finally, some types of samples should be carried in a specific way [23].

• Sample reception:

The received samples must be recorded and examined by authorized staff. In addition, samples must be checked for conformity. The laboratory is required to have a documented procedure with acceptance and rejection criteria. If a non-conformity sample is accepted, they must specify the actions taken to correct the problem, and the persons responsible for accepting the sample. The final report should specify the nature of the non-conformity as well as any reservations concerning interpretation of the result [4,9]. Samples must be stored for a specific period determined by the manual for each type of sampling. Most mycological samples are stored at a temperature of 4°C [2,12,15].

MYCOLOGY SAMPLE COLLECTION PROTOCOL AND PRESERVATION

We have summarized all mycological analyses in tables form (Table I). These tables are presented according to the context or location of the sampling. They contain the necessary information on conditioning, routing times, preservation of samples in the event of delayed processing, and the investigations that can be carried out on each type of sample. Furthermore, Table II corresponds to the protocols used in our laboratory for conducting mycological sampling.

CONCLUSION

The pre-analytical stage is the main step in the mycological examination process and must be mastered by biologists in order to provide the most reliable results. To this end, everyone involved in this phase must be aware of the importance of the pre-analytical stage, of the risks posed by making errors, and their potential consequences for the patient.  Thus, the implementation, respect, and exact application of the procedures and the quality improvement systems for this stage are necessary to avoid all errors that may occur during all mycological examinations conducted.

TABLES

Table I : mycological samples [2,15,24]

Table II: protocol used in our laboratory for mycological sampling.

Declaration of Interest

The authors declare to have no interest in relation to this article

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