Criterios preanalíticos en la toma de muestras de hemocultivo J. Adv. Educ. Sci. Humanit. (January - June 2023) 1(1): 27-31 https://doi.org/10.5281/zenodo.14601930 ISSN: XXXX-XXXX REVIEW ARTICLE Pre-analytical criteria in blood culture sample collection  Ivón Howland ivon.howland@utm.edu.ec Received: 2 October 2022 / Accepted: 8 December 2022 / Published online: 18 January 2023 © The Author(s) 2023 Carlos E. Mera 1 · Jean P. Sanclemente 2 · Ivón Howland 3 Abstract The pre-analytical process in blood culture sam- ple collection is essential to ensure the accuracy in detect- ing pathogenic microorganisms in blood, which is crucial in diagnosing bacteremia and fungemia. This article reviews the main pre-analytical criteria associated with sample col- lection, considering aspects such as proper puncture site selection, strict aseptic techniques, blood volume collect- ed, timing with febrile episodes, and the optimal number of samples. Sample contamination remains a signiﬁcant chal- lenge, impacting both the validity of results and the patient’s clinical management. Evidence suggests that eﬀectively disinfecting the site with 70% alcohol and chlorhexidine, sterile materials, and staﬀ training are essential to minimize errors. Furthermore, collecting at least two blood cultures to increase diagnostic sensitivity is emphasized. This arti- cle emphasizes that strict adherence to pre-analytical criteria optimizes clinical laboratory resources and enhances patient care while improving the results’ quality. Keywords blood cultures, pre-analytical phase, good clin- ical laboratory practices. Resumen El proceso preanalítico en la toma de muestras para hemocultivos es fundamental para garantizar la pre- cisión en la detección de microorganismos patógenos en sangre, siendo crucial en el diagnóstico de bacteriemias y fungemias. Este artículo revisa los principales criterios preanalíticos asociados a la obtención de muestras, consi- derando aspectos como la selección adecuada del sitio de punción, técnicas de asepsia estrictas, volumen de sangre recolectado, sincronización con episodios febriles y el nú- mero óptimo de tandas. La contaminación de las muestras sigue siendo un desafío signiﬁcativo, impactando tanto en la validez de los resultados como en la gestión clínica del paciente. La evidencia sugiere que prácticas como la desin- fección efectiva del sitio con alcohol al 70 % y clorhexidina, el uso de materiales estériles, y la capacitación del personal sanitario, son esenciales para minimizar errores. Además, se destaca la importancia de la recolección de al menos dos tandas de hemocultivos para aumentar la sensibilidad diag- nóstica. Este artículo enfatiza que la adherencia rigurosa a los criterios preanalíticos, además de mejorar la calidad de los resultados, optimiza los recursos del laboratorio clínico y la atención del paciente. Palabras clave hemocultivos, fase preanalítica, buenas prácticas de laboratorio clínico. How to cite Mera, C. E., Sanclemente, J. P., & Howland, I. (2023). Pre-analytical criteria in blood culture sample collection. Journal of Advances Education, Sciences and Humanities, 1(1), 27-31. https://doi.org/10.5281/zenodo.14601930 1 Laboratorio Clínico Crisvem, Portoviejo, Ecuador. 2 Quality Labs, Riochico, Portoviejo, Ecuador. 3 Facultad de Ciencias de la Salud, Universidad Técnica de Manabí, Portoviejo, Ecuador.

J. Adv. Educ. Sci. Humanit. (January - June 2023) 1(1): 27-31 28 Introduction Blood cultures are a diagnostic tool in the clinical labora- tory to identify the presence of microorganisms in the blood. They are part of the standard recommendations for mana- ging sepsis, a condition with a mortality rate ranging from 25.5 to 32% in patients in intensive care units and 35% in hospitalized patients. Studies suggest that, in Latin America, the prevalence of sepsis and the associated mortality could be higher than in developed countries (Cohen et al., 2015; Dellinger et al., 2012; Vincent et al., 2014). One in ﬁve deaths worldwide is related to sepsis, dispro- portionately aﬀecting children in impoverished areas. A study revealed that in 2017, 48.9 million cases of sepsis were recorded globally, with 11 million deaths accounting for 20% of all deaths worldwide (WHO, 2020); 85% of sepsis cases occurred in low- and middle-income countries. Sepsis occurs when organs stop functioning properly due to an un- controlled immune response to an infection. Although sepsis does not always result in death, it can leave severe sequelae in survivors (WHO, 2020). Blood cultures, as part of the recommended measures for managing sepsis, have been associated with reduced morta- lity when performed appropriately (Levy et al., 2012). The clinical laboratory encompasses the pre-analytical, analyti- cal, and post-analytical phases. Some recommendations hi- ghlight key aspects such as incubation time, the number and type of bottles, blood volume, reporting contaminants, and the limitations of current systems for blood cultures (Weins- tein & Doern, 2011). Optimizing this process should focus on three main ob- jectives: improving the isolation and identiﬁcation of true pathogens, reducing the presence of contaminants, and en- hancing the detection of infections associated with central catheters. Ineﬃcient antisepsis processes have increased the likelihood of isolating contaminants in blood cultures, lea- ding to diagnostic uncertainty, prolonged hospital stays, and increased healthcare costs (Maldonado et al., 2018). This article reviews the main pre-analytical criteria asso- ciated with sample collection, considering aspects such as appropriate site selection for puncture, strict aseptic techni- ques, blood volume collected, synchronization with febrile episodes, and the optimal number of sets. Pre-analytical phase The pre-analytical phase is considered the critical stage in the blood culture process. During this phase, failures are most likely to occur due to errors made by the responsible staﬀ, which compromises the reliability of the results. Pre- viously, this process was entirely manual, but in developed cities, automation and robotization have started to be imple- mented to improve the accuracy and eﬃciency of this stage (Aray-García et al., 2023). Despite technological advances, errors during this phase persist globally, aﬀecting the quality of blood culture re- sults. A reduction in the incidence of these errors has been observed thanks to technological innovations (Aray-García et al., 2023). Maldonado et al. (2018), in a study at Hospi- tal Ángeles Pedregal in Mexico, found that of 1,598 blood cultures, only 213 were positive, representing a success rate of 13%. The most frequent microorganisms found were Escherichia coli (43%), of which 35 (16%) were resistant to extended-spectrum beta-lactamases (ESBL), Burkhol- deria cepacia (6%), and Enterococcus faecalis (5%) in the Gram-negative group, and Staphylococcus epidermidis (9%) and Staphylococcus aureus (6%) in the Gram-positive group (Enberg et al., 2020). The research at Hospital Ángeles Pedregal in Mexico City concluded that the rate of positive blood cultures was slight- ly higher than that reported in previous studies. They obser- ved a higher frequency of positivity for E. coli, supporting the idea that microbial populations vary by hospital (Enberg et al., 2020). In this context, Maldonado et al. (2018) empha- sized the need for all institutions to have written protocols that include detailed procedures for sample collection and the techniques and products used in antisepsis. Maldonado et al. (2018) evaluated various institutional protocols and noted that 66.7% of the protocols speciﬁed the number of bottles and blood volume per blood culture set, and 60.0% included sample rejection criteria. However, these studies revealed that not all protocols contain the in- formation necessary to ensure a correct pre-analytical phase (Maldonado et al., 2018). On the other hand, the SEIMC (Spanish Society of Infec- tious Diseases and Clinical Microbiology), in its Clinical Microbiology Procedures, highlighted that the detection of bacteremia and fungemia is a priority in the Clinical Micro- biology Service of health institutions due to their diagnostic and prognostic relevance. These processes are associated with high mortality (10 to 30%), depending on the type of patient, the origin of the infection, and initial management (Aray-García et al., 2023). Sepsis requires a blood culture as a ﬁrst-line test for diag- nosis; however, literature indicates that the sensitivity for detecting bacteremia in blood cultures is low, with a growth rate of less than 10% (Maldonado et al., 2018).

J. Adv. Educ. Sci. Humanit. (January - June 2023) 1(1): 27-31 29 The laboratory study process begins with the clinician’s request for analysis, who must provide detailed and accurate information to ensure the quality of the results. The request should include essential data such as a unique identiﬁcation code (request number), type of request (routine or urgent), and patient identiﬁcation details (name, surname, medical record number, etc.) (Plebani, 2015). It is essential to include relevant clinical and demographic information, such as date of birth, gender, and diagnosis, to facilitate proper interpre- tation of results (Rojo et al., 2010). Administrative data must also be included, such as the re- questing physician, the report destination, and the adminis- trative person responsible for the request (Vélez-Cevallos et al., 2023). Transcribing this data into the laboratory informa- tion system (LIS) can lead to errors if not done carefully, hi- ghlighting the importance of clear protocols and agreements between clinicians and the laboratory (Rodríguez, 2021). The Clinical Laboratory Best Practices Guide by the Mi- nistry of Public Health (MSP) of Ecuador, in its Form No. 15: Request for Examinations, establishes that the basic re- quired information includes patient data (name, surname, ID number, age, gender, contact address or phone), requesting physician’s information (name, phone, address, and profes- sional code), and details of the requested examinations (such as previous studies, medication use, drug monitoring, type of biological sample, and date/time of sample collection) (Rojo et al., 2010). In institutions with digital systems, pro- fessionals can make direct requests through the laboratory information system (LIS) or web platforms to facilitate the management process and avoid transcription errors (Maldo- nado et al., 2018). Once the analysis request and patient appointment are made, the next step is sample collection, which can be per- formed by nursing staﬀ if the patient is already admitted (Aray-García et al., 2023). According to the guidelines of the “Dr. Abel Gilbert” Specialty Hospital in Guayaquil, ba- sed on the Ministry of Public Health (MSP) guidelines, it is recommended to take two blood cultures within 24 hours, with intervals of at least 20 to 30 minutes between them, and from diﬀerent puncture sites. Samples should be collected one hour before the febrile peak or at the onset of fever. They should be taken before administering antibiotics or at least one hour before the next dose. The recommended volume is 8 to 10 mL for adults and 1 to 3 mL for children, and each sample should be properly labeled with its number and time of collection. It is essential to choose the appropriate type of bottle according to the sus- pected microorganism (Acosta-Gnass, 2011). The procedure includes hand hygiene, selecting the punc- ture site, and disinfecting the area with 70° alcohol and povi- done-iodine solution. The bottle cap must be disinfected, and the samples should be inoculated without introducing air. Fi- nally, the samples should be sent immediately to the labora- tory or, if impossible, placed in an incubation oven at 35 °C in the on-call laboratory. They should never be refrigerated (Acosta-Gnass, 2011). It is important to note that there are several diﬀerences between the recommendations described by the Ministry of Public Health (MSP) of Ecuador and the SEIMC (Table 1). Proper sample collection is crucial, as errors in extraction or handling can signiﬁcantly aﬀect the analysis results (Ro- dríguez, 2021). According to SEIMC, despite no universal guidelines, blood cultures are generally recommended for patients with fever (≥38 ºC), chills, leukocytosis, leukopenia, thrombocytopenia, signs of sepsis, or suspected endocardi- tis. Blood cultures should also be performed when bactere- mia originating from a catheter is suspected (Rodríguez et al., 2017). Once obtained, samples must be organized and transported correctly, ensuring correct identiﬁcation of the samples, the request form, and the patient (Rojo et al., 2010). Transport Table 1. Diﬀerences between the sample collection recommendations from the Ministry of Public Health of Ecuador and SEIMC Indicator MSP Ecuador SEIMC Sample volume In children 1 to 3 mL and 8 to 10 mL in adults Small children require between 1 and 5 mL (1:5 dilution), adults 10-20 mL (1:10 dilution). Packaging Does not specify Children: 1 bottle (aerobic), Adults: 2 bottles (aerobic, anaerobic). Antiseptic action Application of povidone iodine solution, concentrically around the puncture site. Allow each to act for 30 to 60 seconds. Approximately 30 seconds for chlorhexidine and between 1.5 and 2 minutes if povidone is used.

J. Adv. Educ. Sci. Humanit. (January - June 2023) 1(1): 27-31 30 should be as quick as possible, avoiding interferences such as agitation (which can cause hemolysis) or exposure to light (to prevent degradation of components like bilirubin) (Vé- lez-Cevallos et al., 2023). In the case of blood cultures, it is recommended to extract blood from veins, not from catheters or umbilical cord blood, and to take samples from diﬀerent puncture sites to avoid contamination (Rojo et al., 2010). The main challenge in interpreting blood cultures is pos- sible contamination with the skin’s microbial ﬂora during extraction. To reduce this risk, it is essential to prepare the skin meticulously, use appropriate equipment like masks and gloves, and, if necessary, perform the extraction with additional protective measures (Callejas-Díaz et al., 2022). Microorganisms commonly associated with contamination include coagulase-negative Staphylococcus, Bacillus spp., and Propionibacterium acnes, among others, although their presence in a single sample per patient should not compro- mise the results (Callejas-Díaz et al., 2022). Conclusions The pre-analytical phase in the blood culture process is crucial to ensure the quality and reliability of results, but it is also one of the most error-prone stages. Despite techno- logical advances such as automation and robotics, errors resulting from human handling and the need for clear and universal protocols persist. The differences between the recommendations of organizations like the Ministry of Public Health of Ecuador and SEIMC highlight the need to standardize procedures and improve staff training. Pro- per collection, transport, and processing of samples, as well as adequate antisepsis and optimal volumes, are key elements to minimize errors and prevent contamination. Additionally, institutions need to implement detailed wri- tten protocols that cover all aspects of this phase, from sample collection to handling in the laboratory. Adopting these best practices will improve the detection of bacte- remia and fungemia, optimizing diagnoses and clinical treatments related to sepsis and other severe infections. References Acosta-Gnass, S.I. (2011). Manual de control de infecciones y epidemiología hospitalaria. Organización Panameri- cana de la Salud. https://iris.paho.org/bitstream/hand- le/10665.2/51545/ControlInfecHospitalarias_spa.pdf Aray-García, A.M., García-Pérez, J.P., & Lino-Villacre- ses, W. (2023). Errores en los hemocultivos: cau- sas y repercusiones para el paciente. MQRInves- tigar, 7(1), 1425-1447. https://doi.org/10.56048/ MQR20225.7.1.2023.1425-1447 Callejas-Díaz, A., Calderón-Parra, J., & Fernández-Cruz, A. (2022). Hemocultivos: indicaciones e interpretación. 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