Blood Volume Collected for Cultures in Infants with Suspected Neonatal Sepsis

Blood Volume Collected for Cultures in Infants with Suspected Neonatal Sepsis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Blood Volume Collected for Cultures in Infants with Suspected Neonatal Sepsis Maria Rueda Altez, Lamia Soghier, Joseph Campos, Burak Bahar, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4474756/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 28 Sep, 2024 Read the published version in Journal of Perinatology → Version 1 posted 9 You are reading this latest preprint version Abstract Objectives To evaluate blood culture sample volumes, identify factors linked to insufficient samples, and compare volumes among neonates treated for culture-negative-sepsis, sepsis-rule-outs, and bloodstream infections (BSI). Methods Observational cohort of blood cultures collected during NICU stay. Association of age, weight, gender, source, and collection time with lower-than-recommended volumes was determined by logistic regression. Blood culture inocula of patients with culture-negative-sepsis, sepsis rule-out, and BSI were compared using ANOVA. Results 742 blood cultures were obtained from 292 neonates. Median inoculum was 1mL (IQR:0.6–1.4), and 259 bottles (35%) had inocula < 0.9mL. Night shift sample collection was associated with lower-than-recommended volumes (p = 0.006). No difference in sample volumes was observed between culture-negative-sepsis, sepsis-rule-outs, and BSI (p = 0.5). Conclusions Median NICU blood culture volumes align with recommendations. Night shift collections correlate with lower volumes. Sample volumes don’t differ in patients with culture-negative-sepsis, BSI, and sepsis-rule-out, and should not be a justification for longer duration of antibiotics. neonatal sepsis blood cultures culture-negative sepsis Figures Figure 1 Figure 2 INTRODUCTION The treatment of culture-negative suspected sepsis in neonatal intensive care units (NICUs) leads to 10-times more antibiotic use than culture-proven infections.[ 1 , 2 ] Blood cultures, the gold-standard for diagnosis of neonatal sepsis, have high sensitivity to detect bacteremia in concentrations of 1–10 colony-forming units (CFU)/mL when sufficient blood volume is collected.[ 3 ] The Infectious Diseases Society of America consensus recommendations for blood culture volume collection are weight based, with a minimum of 2 mL of blood for infants < 1 kg inoculated into one or two blood culture bottles, and 4 mL for infants < 2 kg.[ 4 ] In contrast, the latest update of the American Academy of Pediatrics (AAP) clinical practice guidelines for early onset sepsis management in premature infants recommend the inoculation of a minimum of 1 mL per bottle for all premature infants < 34 weeks undergoing a sepsis evaluation.[ 5 ] The availability of reliable blood culture results is a crucial element of antimicrobial stewardship in the NICU. However, the collection of appropriate volumes may not always be possible, given the sampling difficulties in preterm infants and clinical instability of neonates, with prior studies showing inappropriately low volumes in as many as 93% of samples.[ 6 – 9 ] Consequently, negative blood cultures are frequently met with mistrust by clinicians, leading to prolonged courses of antimicrobial therapy in neonates without a focus of infection.[ 2 ] The risk of inappropriate use of antibiotics is substantial, as it is associated with a higher incidence of antibiotic resistance, necrotizing enterocolitis, invasive candidiasis, alteration of gut microbiota and death.[ 10 – 12 ] Clinicians are in a constant need to find a balance between making diagnostic decisions with tests that may be suboptimal, and decreasing antibiotic use. Determining blood culture volumes for neonatal samples could allay concerns regarding the accuracy of the test and potentially decrease antimicrobial use. This study aims to (1) determine the blood culture sample volumes collected by weighing inoculated blood culture bottles, (2) identify clinical or environmental factors associated with lower than recommended volumes, and (3) compare the sample volumes of patients treated for culture-negative sepsis with those with confirmed bloodstream infections (BSI) and those in whom antibiotics were discontinued after negative cultures. We hypothesized that mean blood culture volumes would be lower for low-birth weight neonates and that samples collected from patients who were treated for culture-negative sepsis would not be significantly different than the other groups. METHODS Study Design and Setting An observational single-center cohort study was conducted with a retrospective period from September 2018 to January 2019 and a prospective period from February 2019 to November 2019 at the Level IV neonatal intensive care unit (NICU) of Children’s National Hospital (CNH) in Washington DC. The NICU is a 66-bed unit that serves as a referral center for the District of Columbia, Maryland, and Virginia. The CNH Institutional Review Board approved the study with a waiver of consent. Study Subjects All aerobic and anerobic blood cultures from infants hospitalized in the CNH NICU that were collected during the study period were included. Cultures with no documentation of the amount of blood inoculated were excluded. Blood culture results were reviewed for each patient. For the comparative analysis focused on suspected or confirmed bloodstream infections, positive blood cultures considered contaminants based on the study definition below were excluded. Additionally, negative blood cultures collected from patients diagnosed with focal infections (urinary tract infections, intraabdominal infections including necrotizing enterocolitis, pneumonia, tracheitis, skin and soft tissue infections and/or meningitis) were excluded, as they do not constitute culture-negative sepsis given the presence of a source of infection. Finally, repeated negative cultures for the same patient that were collected during an antibiotic course for a confirmed bloodstream infection or for culture negative sepsis were also excluded. Data Collection Demographic and clinical data were collected through retrospective chart review. The patient’s gender, chronological age at time of culture collection, weight at time of blood culture collection, time of sample collection (day versus night shift), and source of sample (peripheral versus central venous catheter) were documented. Information regarding antibiotic administration up to 48 hours prior to blood culture collection was collected. C- reactive protein (CRP) results and antibiotic therapy choice and duration were documented for patients with positive blood cultures for coagulase-negative Staphylococcus (CoNS). Laboratory data was obtained from the laboratory data warehouse. All data were recorded in a web-based secure REDCap database.[ 13 ] Inoculated Volume Calculation The inoculated volume was calculated by subtracting the weight of the uninoculated bottle from the measured weight of the inoculated bottle, and dividing the weight in grams by 1.06 g/mL to account for the density of blood.[ 14 ]. After blood culture collection by the neonatology providers, all inoculated culture bottles were weighed after arrival to the microbiology laboratory by the technologists and the weights in grams were documented to the first decimal place. The weight of the uninoculated aerobic and anaerobic blood culture bottles was obtained using a calibrated scale (Brand: OHAUS ®ฏ . Model: Ranger 3000). The uninoculated weight was determined by weighing a random selection of one hundred unused blood culture bottles without the disposable caps and obtaining an average weight and standard deviation. The mean weight for aerobic bottles was 62.5 g (standard deviation (SD): 0.09) and for anaerobic bottles was 72.7g (SD: 0.2). Results were further validated by the study team by weighing 50 unused aerobic and anaerobic blood culture bottles during two additional time points, with similar mean and SD results (62.5 ± 0.1g for aerobic bottles, and 72.7g ± 0.2 for anaerobic bottles) and narrow ranges (62.6-63.1g for aerobic bottles, and 72.8-73.4g for anaerobic bottles). The microbiology laboratory uses BACT/Alert FAN plus® blood culture system which includes the FA plus® aerobic blood culture bottle and the FN plus® anaerobic blood culture bottle. Definitions The following definitions were used throughout the study: Lower than recommended volumes Our planned definition was < 1mL. However, after the adjustment of the volumes to correct for the density of blood, many blood culture inocula had values between 0.9 and 1, therefore making the proportion of < 1 ml a less representative metric (Fig. 1 ). Therefore, we opted to modify the definition of lower than recommended volumes to < 0.9mL. Blood culture set Aerobic and anaerobic cultures collected and sent simultaneously from the same patient. To calculate the volume, the inoculated volumes of both bottles were added. Bloodstream infection the Centers for Disease Control National Healthcare Safety Network definition of bloodstream infection was used.[ 15 ] If a common commensal was identified, it was considered a bloodstream infection when isolated from two or more blood specimens collected on separate occasions. Additionally, as defined by the National Institute of Child Health and Human Development Neonatal Research Network[ 16 ], when presented with a single CoNS positive blood culture, a bloodstream infection was considered if the patient had a CRP of > 1 mg/dL within 2 days from blood culture collection or if it was treated by the clinician for ≥ 5 days with an appropriate anti-staphylococcal regimen. Contaminant A common commensal if it was identified in a single blood specimen[ 15 ]. For single positive blood culture with CoNS, those who do not meet the criteria mentioned above were considered contaminants. Culture-negative sepsis A patient without an identifiable source of infection that, in the context of a negative blood culture, was treated with antibiotics for > 72 hours. Sepsis rule-out Patients that, in the context of a negative blood culture, were treated with antibiotics for ≤ 72 hours. Statistical Analysis The distribution of blood culture volumes and summary statistics and a point estimate for median and interquartile range were described. For the comparative analyses, the variables analyzed included gender, chronological age, weight, time of collection (day shift from 7AM-7PM, night shift from 7PM-7AM), and source of sample (peripheral, central). We used a chi-squared or Fisher’s exact test for binary and t-test or Wilcoxon-Mann-Whitney for continuous variables. In addition, a multivariate analysis was conducted using logistic regression. A p < 0.05 was considered significant. To compare the mean blood volume collected for patients treated for culture-negative sepsis, bloodstream infections, and sepsis rule-out were analyzed using analysis of variance test. Since the blood culture inoculum volumes were not normally distributed, the mean volumes were converted to logarithmic scale for normalization. A p < 0.05 was considered significant. All analysis were conducted using Stata version 13 (StataCorp LLC). RESULTS During the study period, 742 blood cultures from 292 NICU patients were obtained. Most culture bottles were aerobic (604, 81%). The cohort’s demographic characteristics are summarized in Table 1 . The median volume of blood inoculated into blood cultures was 1 mL (IQR: 0.6–1.4) and the mean was 1.06 (SD: 0.7). A total of 259 cultures (35%) had inoculum volumes < 0.9 ml. (Fig. 1 ). Table 1 Demographic characteristics of the study population Clinical and environmental factors N (%) Median (IQR) Weight (g) 2700 (1340–3560) 7 days 614 (83%) Source of sample Peripheral 612 (82%) Central 130 (18%) Gender Female 321 (43%) Male 421 (57%) Time of collection Day shift 447 (60%) Night shift 295 (40%) A total of 226 bottles were collected as part of a blood culture set (123 blood culture sets). The mean volume per set was 1.7 ml (SD: 0.7) per set. We compared the volume inoculated in aerobic blood cultures when they were part of a set and when they were collected independently and found no significant difference (p = 0.32). In the univariate analysis, blood cultures obtained during the night shift were associated with the collection of lower than recommended sample volumes (p = 0.007). This difference remained in the multivariate analysis. No significant differences in blood culture volume were noted by patient gender, chronological age, weight at time of culture collection, or the source of the sample (Table 2 ). Table 2 Association of clinical and environmental factors with volume of blood inoculated in cultures Patient Characteristics < 0.9 ml ≥ 0.9 ml Univariate analysis p value Multivariate analysis p value Weight, kg (median) 2.68 2.70 0.52 0.51 Age, days (median) 27 30 0.32 0.87 Source of sample (n) 0.25 0.17 Peripheral 220 394 Central 39 89 Gender (n) 0.53 0.68 Male 143 278 Female 116 205 Time of collection (n) 0.007 0.006 Day shift 175 308 Night shift 120 139 Of the 742 blood cultures identified with documented volumes, 146 (20%) were from patients treated for culture-negative sepsis, 102 (14%) had confirmed bloodstream infections, 291 (39%) underwent a sepsis rule-out, and 194 (26%) were diagnosed with another infectious diagnosis (Fig. 2 ). The mean inoculated volume was 1.06 mL (SD: 0.7) for culture-negative sepsis, 1.03 mL (SD: 0.7) for sepsis rule-out, and 1.09 mL (SD: 0.7) for BSI, which were not significantly different (p = 0.5). The proportion of blood cultures with inoculum < 0.9 mL was 36% (106/291) for sepsis rule-out, 32% (47/146) for culture negative sepsis and 30% (31/102) for bloodstream infection. DISCUSSION This single-center study shows that the mean volume of blood cultures collected in a level IV NICU was consistent with prior recommendations of collecting > 1mL of blood/bottle, although a significant proportion of inocula are below the recommended volume. In addition, the volume of blood inoculated in blood cultures of patients treated for culture negative sepsis does not differ from those with bloodstream infections and with sepsis rule-out. Determining the ideal volume to be collected for neonatal blood cultures is challenging, as it needs to balance the greater diagnostic yield of larger samples[ 3 , 17 ] with the challenges of sample collection and small circulating blood volumes in premature neonates. The mean and median volume inoculated in neonatal blood cultures for our study were 1.06 and 1 mL respectively, which is consistent with most published recommendations[ 3 , 5 , 17 ], although there is significant room for improvement, given the large proportion of cultures with < 0.9 ml. The earliest study on this topic published in 1986 by Neal et al ., who reported that 56% of the collected samples were ≤ 0.5 mL.[ 6 ] Subsequent studies have shown variability in proportion of samples with lower-than-recommended volumes, ranging from 23 to 93%.[ 7 , 8 , 18 ] The highest compliance was reported by Woodford et al[ 18 ], who described their experience with blood culture collection in a Level III NICU. Their institutional guidance included collection of a minimum of 1 ml of blood per sample and required documentation of blood culture inoculum in the patient’s medical record by the clinician. A similar approach has been used in another recent study by Singh et al., who by implementing a local guideline, educating providers, and fomenting documentation, reduced inappropriate samples from 97–25%.[ 8 ] These results are encouraging and suggest that systematization of blood culture sampling can greatly optimize the collected volumes. This study did not find an association between weight and lower inoculated volumes, which is consistent with past reports.[ 6 , 8 ] These results challenge the concept that low-birth-weight is a justification for the collection of suboptimal sample volumes. Similarly, chronological age was not associated with lower volumes, which contrasts the findings by Woodford el at. 18 In their recently published article, the authors found that very-low-birth-weight infants older than 7 days had significantly lower sample volumes when compared to their counterparts that were evaluated for early onset sepsis. This was attributed to the reliable access to central lines during the first week of life (e.g., umbilical lines). However, in our cohort, central access sample collection was not associated with higher blood volumes and most blood cultures in our unit are obtained from a peripheral arterial source to the greatest extent possible. Samples collected during the night shift had significantly lower volumes, which has not been reported previously in the literature to our knowledge. This could be associated with different providers collecting samples (pediatric residents, fellows, advanced practice providers), or provider fatigue influencing sampling technique, but these factors were outside of our study’s scope and were not explored. Our study showed there’s no significant difference in the inoculated volume of aerobic bottles when collected as part of a blood culture set or as single bottles. AAP recommends, when presented with an infant with suspected sepsis, to collect aerobic and anaerobic cultures to optimize pathogen recovery.[ 5 ] This is supported by studies showing that, despite the low prevalence of bacteremia by strict anaerobes in neonates and children (< 1%), as many as 11–22% of clinically relevant isolates were only detectable in anaerobic bottles.[ 19 , 20 ] This could be secondary to the anaerobic culture’s more optimal conditions for growth of facultative anaerobic organisms that are prevalent in the neonatal population, like Streptococcus agalactiae. A study showed that there is additional value in the use of anaerobic bottles in very-low-birth-weight infants, where 16% of all cases of early-onset sepsis were secondary to obligate anaerobes.[ 21 ] The utility of anaerobic blood cultures in the neonatal population needs to be balanced with the decision to inoculum a larger volume into the aerobic culture, based on the sample that the clinician is able to obtain. To our knowledge, this is the first study to compare blood culture inoculum volumes from patients with culture-positive and culture-negative sepsis. Volume of samples collected in patients treated for culture negative sepsis did not differ from those with confirmed bloodstream infections and those that were treated for sepsis rule-outs. Cantey et al. evaluated the use of antibiotics in a Level III NICU, and found that over 70% of antibiotics were administered to patients undergoing “sepsis rule-outs” and patients diagnosed with culture-negative sepsis.[ 22 ] Similar results have been reported from Norway and Peru.[ 1 , 2 ] When presented with an ill-appearing neonate, the lack of documentation of collected volumes can play a role in clinician mistrust of negative blood culture results.[ 2 , 8 ] However, our results show that there was no significant difference in the diagnostic yield of the samples obtained in neonates treated for culture-negative sepsis. Our study has some limitations. First, documentation of direct volume measurements by syringe at the time of sample collection for blood cultures was not obtained which precluded us from comparing these values to the calculated blood volumes. However, weighing inoculated blood culture bottles has been successfully used in previous studies.[ 8 , 18 ] In addition, an average baseline weight for the uninoculated blood culture bottles was used and we did not weight each bottle before inoculation; however, baseline uninoculated bottle weights were validated by repeating the process with bottles from multiple lots during the study, with comparable results. This method would also be more practical and pragmatic for other institutions looking to apply this methodology to increase confidence in blood culture results. We were unable to analyze if low sample volumes increased the chances of false negative results, and this should be further investigated in future studies. Finally, our study population may not be representative of all NICU patients, as it is a single-center study. CONCLUSION Blood culture inocula in our NICU were generally consistent with guideline recommendations, and collection of samples during the night shift was associated with lower than recommended inoculum volumes. Quality improvement efforts are needed to increase the proportion of samples with adequate volume. The volume of blood sampled does not differ in patients with culture-negative sepsis, bloodstream infection, and sepsis rule-out, suggesting this should not be a justification for longer duration of antibiotic therapy. Declarations Conflict of Interest: Dr. Soghier is an editor for American Academy of Pediatric "Reference Range Values in Pediatric Care" and "Neonatal Simulation" textbooks. The remaining authors of this manuscript have no conflicts of interest to disclose. Funding: None Data Access Statement: Data supporting this study cannot be made fully available due to patient confidentiality. De-identified database can be provided upon request to corresponding author. References Fjalstad JW, Stensvold HJ, Bergseng H, Simonsen GS, Salvesen B, Ronnestad AE, et al. Early-onset Sepsis and Antibiotic Exposure in Term Infants: A Nationwide Population-based Study in Norway. Pediatr Infect Dis J. 2016;35:1. Rueda MS, Calderon-Anyosa R, Gonzales J, Turin CG, Zea-Vera A, Zegarra J, et al. Antibiotic Overuse in Premature Low-Birth-Weight Infants in a Developing Country. Pediatr Infect Dis J. 2018;38:302-7 Schelonka RL, Chai MK, Yoder BA, Hensley D, Brockett RM, Ascher DP. Volume of blood required to detect common neonatal pathogens. J Pediatr. 1996;129:275–8. Miller JM, Binnicker MJ, Campbell S, Carroll KC, Chapin KC, Gilligan PH, et al. A Guide to Utilization of the Microbiology Laboratory for Diagnosis of Infectious Diseases: 2018 Update by the Infectious Diseases Society of America and the American Society for Microbiology. Clin Infect Dis Off Publ Infect Dis Soc Am. 2018;67:e1–94. Puopolo KM, Benitz WE, Zaoutis TE; COMMITTEE ON FETUS AND NEWBORN; COMMITTEE ON INFECTIOUS DISEASES. Management of Neonates Born at ≤34 6/7 Weeks' Gestation With Suspected or Proven Early-Onset Bacterial Sepsis. Pediatrics. Pediatrics. 2018;142(6):e20182896. Neal PR, Kleiman MB, Reynolds JK, Allen SD, Lemons JA, Yu PL. Volume of blood submitted for culture from neonates. J Clin Microbiol. 1986;24:353–6. Connell TG, Rele M, Cowley D, Buttery JP, Curtis N. How reliable is a negative blood culture result? Volume of blood submitted for culture in routine practice in a children’s hospital. Pediatrics. 2007;119:891–6. Singh MP, Balegar V KK, Angiti RR. The practice of blood volume submitted for culture in a neonatal intensive care unit. Arch Dis Child Fetal Neonatal Ed. 2020;105:600–4. Jawaheer G, Neal TJ, Shaw NJ. Blood culture volume and detection of coagulase negative staphylococcal septicaemia in neonates. Arch Dis Child Fetal Neonatal Ed. 1997;76:F57-58. Kuppala VS, Meinzen-Derr J, Morrow AL, Schibler KR. Prolonged initial empirical antibiotic treatment is associated with adverse outcomes in premature infants. J Pediatr. 2011;159:720–5. Cotten CM, Taylor S, Stoll B, Goldberg RN, Hansen NI, Sanchez PJ, et al. Prolonged duration of initial empirical antibiotic treatment is associated with increased rates of necrotizing enterocolitis and death for extremely low birth weight infants. Pediatrics. 2009;123:58–66. Tripathi N, Cotten CM, Smith PB. Antibiotic use and misuse in the neonatal intensive care unit. Clin Perinatol. 2012;39:61–8. Harris PA, Taylor R, Thielke R, Payne J, Gonzalez N, Conde JG. Research electronic data capture (REDCap)--a metadata-driven methodology and workflow process for providing translational research informatics support. J Biomed Inform. 2009;42:377–81. Trudnowski RJ, Rico RC. Specific Gravity of Blood and Plasma at 4 and 37 °C. Clin Chem. 1974;20:615–6. Centers for Disease Control and Prevention (CDC). Bloodstream Infection Event (Central Line-Associated Bloodstream Infection and Non-central Line Associated Bloodstream Infection). Available at: https://www.cdc.gov/nhsn/PDFs/pscManual/4PSC_CLABScurrent.pdf. Accessed 16 June 2022. Stoll BJ, Hansen N, Fanaroff AA, Wright LL, Carlo WA, Ehrenkranz RA, et al. Late-onset sepsis in very low birth weight neonates: the experience of the NICHD Neonatal Research Network. Pediatrics. 2002;110:285–91. Yaacobi N, Bar-Meir M, Shchors I, Bromiker R. A prospective controlled trial of the optimal volume for neonatal blood cultures. Pediatr Infect Dis J. 2015;34:351–4. Woodford EC, Dhudasia MB, Puopolo KM, Skerritt L, Bhavsar M, DeLuca J, et al. Neonatal blood culture inoculant volume: feasibility and challenges. Pediatr Res. Nature Publishing Group; 2021;1–7. Gross I, Gordon O, Abu Ahmad W, Benenson S, et al. Yield of Anaerobic Blood Cultures in Pediatric Emergency Department Patients. Pediatr Infect Dis J. 2018;37:281–6. Gross I, Gordon O, Benenson S, Ahmad WA, Shimonov A, Hashavya S, et al. Using anaerobic blood cultures for infants younger than 90 days rarely showed anaerobic infections but increased yields of bacterial growth. Acta Paediatr Oslo Nor 1992. 2018;107:1043–8. Mukhopadhyay S, Puopolo KM. Clinical and Microbiologic Characteristics of Early-onset Sepsis Among Very Low Birth Weight Infants: Opportunities for Antibiotic Stewardship. Pediatr Infect Dis J. 2017;36:477–81. Cantey JB, Wozniak PS, Sánchez PJ. Prospective surveillance of antibiotic use in the neonatal intensive care unit: results from the SCOUT study. Pediatr Infect Dis J. 2015;34:267–72. Additional Declarations Yes there is potential conflict of interest. Cite Share Download PDF Status: Published Journal Publication published 28 Sep, 2024 Read the published version in Journal of Perinatology → Version 1 posted Editorial decision: revise 10 Jun, 2024 Review # 1 received at journal 08 Jun, 2024 Review # 2 received at journal 03 Jun, 2024 Reviewer # 2 agreed at journal 03 Jun, 2024 Reviewer # 1 agreed at journal 30 May, 2024 Reviewers invited by journal 28 May, 2024 Submission checks completed at journal 28 May, 2024 Editor assigned by journal 24 May, 2024 First submitted to journal 24 May, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4474756","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":307941623,"identity":"aba122aa-848f-4923-a430-85bd3182c89f","order_by":0,"name":"Maria Rueda Altez","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA2klEQVRIiWNgGAWjYLACHgYGGSDF+ICB4QBYQIIYLTxAzGxAshY2CaK06M4+Y/jgTY0djz17+7Nqnpo70fwNzAdv8+DRYnYux9hwzrFkHh6eA2m3eY49y51xgC3ZGq+WMzxm0jxszDw8EgnHbvOwHc7dwAASwa/F/DfPv3oeHvmHbcU8/0Ba+L8R0mLGzNt2GGgLMxuIAbKFjYAWtmLJuX3HeXjOpDEDGUC/HGYztpyDVwvzxg9vvlXLsbcffwhk3Mntb29+eOMNHi1YADNpykfBKBgFo2AUYAEAh7tHj4iSX6AAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0003-1152-6019","institution":"University of Alabama at Birmingham","correspondingAuthor":true,"prefix":"","firstName":"Maria","middleName":"Rueda","lastName":"Altez","suffix":""},{"id":307941624,"identity":"db48e191-660f-4512-9521-ed80bcad0c95","order_by":1,"name":"Lamia Soghier","email":"","orcid":"https://orcid.org/0000-0002-0597-1397","institution":"Children's National Medical Center","correspondingAuthor":false,"prefix":"","firstName":"Lamia","middleName":"","lastName":"Soghier","suffix":""},{"id":307941625,"identity":"7d237ffd-6105-49bf-bb89-59c9549353fb","order_by":2,"name":"Joseph Campos","email":"","orcid":"","institution":"Children's National Hospital","correspondingAuthor":false,"prefix":"","firstName":"Joseph","middleName":"","lastName":"Campos","suffix":""},{"id":307941626,"identity":"82ecbdce-1d7b-49de-a414-2e39175a112f","order_by":3,"name":"Burak Bahar","email":"","orcid":"https://orcid.org/0000-0002-7779-7973","institution":"Children's National Hospital","correspondingAuthor":false,"prefix":"","firstName":"Burak","middleName":"","lastName":"Bahar","suffix":""},{"id":307941627,"identity":"82546353-f7b9-4596-ae5c-f06f12798c0a","order_by":4,"name":"Jiaxiang Gai","email":"","orcid":"","institution":"Children's National Hospital","correspondingAuthor":false,"prefix":"","firstName":"Jiaxiang","middleName":"","lastName":"Gai","suffix":""},{"id":307941628,"identity":"ebb3834f-793a-4fd0-b084-0c98a469980e","order_by":5,"name":"James Bost","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"James","middleName":"","lastName":"Bost","suffix":""},{"id":307941629,"identity":"49093d25-ae6c-4d41-9178-d10140f53e6f","order_by":6,"name":"Rana Hamdy","email":"","orcid":"","institution":"Childern's National","correspondingAuthor":false,"prefix":"","firstName":"Rana","middleName":"","lastName":"Hamdy","suffix":""}],"badges":[],"createdAt":"2024-05-25 01:55:12","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4474756/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4474756/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1038/s41372-024-02120-0","type":"published","date":"2024-09-28T04:00:00+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":58243712,"identity":"35137e04-3693-4c44-9367-2c6167dae53f","added_by":"auto","created_at":"2024-06-13 02:05:59","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":16629,"visible":true,"origin":"","legend":"\u003cp\u003eHistogram of blood culture inoculum volume distribution\u003c/p\u003e","description":"","filename":"JPFigure1.png","url":"https://assets-eu.researchsquare.com/files/rs-4474756/v1/8877d960a8ebf781e20e17ff.png"},{"id":58243711,"identity":"ed509769-da20-4bd4-9c29-54a1e93937ec","added_by":"auto","created_at":"2024-06-13 02:05:59","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":116590,"visible":true,"origin":"","legend":"\u003cp\u003eNumber of blood cultures by positivity and patient diagnosis. The 3 groups included in the comparative analysis are highlighted in grey\u003c/p\u003e","description":"","filename":"JPFigure2.png","url":"https://assets-eu.researchsquare.com/files/rs-4474756/v1/16d6b9af76dada8c6ba1a6d4.png"},{"id":65523856,"identity":"12614742-86f1-4008-867a-054a29d7d02c","added_by":"auto","created_at":"2024-09-29 07:06:42","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":555776,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4474756/v1/1c65c5cf-aa9e-40ac-8c3c-ec745151c53d.pdf"}],"financialInterests":"\u003cb\u003eYes\u003c/b\u003e there is potential conflict of interest.","formattedTitle":"Blood Volume Collected for Cultures in Infants with Suspected Neonatal Sepsis","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eThe treatment of culture-negative suspected sepsis in neonatal intensive care units (NICUs) leads to 10-times more antibiotic use than culture-proven infections.[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e] Blood cultures, the gold-standard for diagnosis of neonatal sepsis, have high sensitivity to detect bacteremia in concentrations of 1\u0026ndash;10 colony-forming units (CFU)/mL when sufficient blood volume is collected.[\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e] The Infectious Diseases Society of America consensus recommendations for blood culture volume collection are weight based, with a minimum of 2 mL of blood for infants\u0026thinsp;\u0026lt;\u0026thinsp;1 kg inoculated into one or two blood culture bottles, and 4 mL for infants\u0026thinsp;\u0026lt;\u0026thinsp;2 kg.[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e] In contrast, the latest update of the American Academy of Pediatrics (AAP) clinical practice guidelines for early onset sepsis management in premature infants recommend the inoculation of a minimum of 1 mL per bottle for all premature infants\u0026thinsp;\u0026lt;\u0026thinsp;34 weeks undergoing a sepsis evaluation.[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]\u003c/p\u003e \u003cp\u003eThe availability of reliable blood culture results is a crucial element of antimicrobial stewardship in the NICU. However, the collection of appropriate volumes may not always be possible, given the sampling difficulties in preterm infants and clinical instability of neonates, with prior studies showing inappropriately low volumes in as many as 93% of samples.[\u003cspan additionalcitationids=\"CR7 CR8\" citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e] Consequently, negative blood cultures are frequently met with mistrust by clinicians, leading to prolonged courses of antimicrobial therapy in neonates without a focus of infection.[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e] The risk of inappropriate use of antibiotics is substantial, as it is associated with a higher incidence of antibiotic resistance, necrotizing enterocolitis, invasive candidiasis, alteration of gut microbiota and death.[\u003cspan additionalcitationids=\"CR11\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e] Clinicians are in a constant need to find a balance between making diagnostic decisions with tests that may be suboptimal, and decreasing antibiotic use. Determining blood culture volumes for neonatal samples could allay concerns regarding the accuracy of the test and potentially decrease antimicrobial use.\u003c/p\u003e \u003cp\u003eThis study aims to (1) determine the blood culture sample volumes collected by weighing inoculated blood culture bottles, (2) identify clinical or environmental factors associated with lower than recommended volumes, and (3) compare the sample volumes of patients treated for culture-negative sepsis with those with confirmed bloodstream infections (BSI) and those in whom antibiotics were discontinued after negative cultures. We hypothesized that mean blood culture volumes would be lower for low-birth weight neonates and that samples collected from patients who were treated for culture-negative sepsis would not be significantly different than the other groups.\u003c/p\u003e"},{"header":"METHODS","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy Design and Setting\u003c/h2\u003e \u003cp\u003eAn observational single-center cohort study was conducted with a retrospective period from September 2018 to January 2019 and a prospective period from February 2019 to November 2019 at the Level IV neonatal intensive care unit (NICU) of Children\u0026rsquo;s National Hospital (CNH) in Washington DC. The NICU is a 66-bed unit that serves as a referral center for the District of Columbia, Maryland, and Virginia. The CNH Institutional Review Board approved the study with a waiver of consent.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eStudy Subjects\u003c/h2\u003e \u003cp\u003eAll aerobic and anerobic blood cultures from infants hospitalized in the CNH NICU that were collected during the study period were included. Cultures with no documentation of the amount of blood inoculated were excluded. Blood culture results were reviewed for each patient.\u003c/p\u003e \u003cp\u003eFor the comparative analysis focused on suspected or confirmed bloodstream infections, positive blood cultures considered contaminants based on the study definition below were excluded. Additionally, negative blood cultures collected from patients diagnosed with focal infections (urinary tract infections, intraabdominal infections including necrotizing enterocolitis, pneumonia, tracheitis, skin and soft tissue infections and/or meningitis) were excluded, as they do not constitute culture-negative sepsis given the presence of a source of infection. Finally, repeated negative cultures for the same patient that were collected during an antibiotic course for a confirmed bloodstream infection or for culture negative sepsis were also excluded.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eData Collection\u003c/h2\u003e \u003cp\u003e Demographic and clinical data were collected through retrospective chart review. The patient\u0026rsquo;s gender, chronological age at time of culture collection, weight at time of blood culture collection, time of sample collection (day versus night shift), and source of sample (peripheral versus central venous catheter) were documented. Information regarding antibiotic administration up to 48 hours prior to blood culture collection was collected. C- reactive protein (CRP) results and antibiotic therapy choice and duration were documented for patients with positive blood cultures for coagulase-negative \u003cem\u003eStaphylococcus\u003c/em\u003e (CoNS). Laboratory data was obtained from the laboratory data warehouse. All data were recorded in a web-based secure REDCap database.[\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eInoculated Volume Calculation\u003c/h2\u003e \u003cp\u003eThe inoculated volume was calculated by subtracting the weight of the uninoculated bottle from the measured weight of the inoculated bottle, and dividing the weight in grams by 1.06 g/mL to account for the density of blood.[\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. After blood culture collection by the neonatology providers, all inoculated culture bottles were weighed after arrival to the microbiology laboratory by the technologists and the weights in grams were documented to the first decimal place. The weight of the uninoculated aerobic and anaerobic blood culture bottles was obtained using a calibrated scale (Brand: OHAUS\u003csup\u003e\u0026reg;ฏ\u003c/sup\u003e. Model: Ranger 3000). The uninoculated weight was determined by weighing a random selection of one hundred unused blood culture bottles without the disposable caps and obtaining an average weight and standard deviation. The mean weight for aerobic bottles was 62.5 g (standard deviation (SD): 0.09) and for anaerobic bottles was 72.7g (SD: 0.2). Results were further validated by the study team by weighing 50 unused aerobic and anaerobic blood culture bottles during two additional time points, with similar mean and SD results (62.5\u0026thinsp;\u0026plusmn;\u0026thinsp;0.1g for aerobic bottles, and 72.7g\u0026thinsp;\u0026plusmn;\u0026thinsp;0.2 for anaerobic bottles) and narrow ranges (62.6-63.1g for aerobic bottles, and 72.8-73.4g for anaerobic bottles). The microbiology laboratory uses BACT/Alert FAN plus\u0026reg; blood culture system which includes the FA plus\u0026reg; aerobic blood culture bottle and the FN plus\u0026reg; anaerobic blood culture bottle.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eDefinitions\u003c/h2\u003e \u003cp\u003eThe following definitions were used throughout the study:\u003c/p\u003e \u003cp\u003e \u003cstrong\u003eLower than recommended volumes\u003c/strong\u003e \u003cp\u003eOur planned definition was \u0026lt;\u0026thinsp;1mL. However, after the adjustment of the volumes to correct for the density of blood, many blood culture inocula had values between 0.9 and 1, therefore making the proportion of \u0026lt;\u0026thinsp;1 ml a less representative metric (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Therefore, we opted to modify the definition of lower than recommended volumes to \u0026lt;\u0026thinsp;0.9mL.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eBlood culture set\u003c/strong\u003e \u003cp\u003eAerobic and anaerobic cultures collected and sent simultaneously from the same patient. To calculate the volume, the inoculated volumes of both bottles were added.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eBloodstream infection\u003c/strong\u003e \u003cp\u003ethe Centers for Disease Control National Healthcare Safety Network definition of bloodstream infection was used.[\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e] If a common commensal was identified, it was considered a bloodstream infection when isolated from two or more blood specimens collected on separate occasions. Additionally, as defined by the National Institute of Child Health and Human Development Neonatal Research Network[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e], when presented with a single CoNS positive blood culture, a bloodstream infection was considered if the patient had a CRP of \u0026gt;\u0026thinsp;1 mg/dL within 2 days from blood culture collection or if it was treated by the clinician for \u0026ge;\u0026thinsp;5 days with an appropriate anti-staphylococcal regimen.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eContaminant\u003c/strong\u003e \u003cp\u003eA common commensal if it was identified in a single blood specimen[\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. For single positive blood culture with CoNS, those who do not meet the criteria mentioned above were considered contaminants.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eCulture-negative sepsis\u003c/strong\u003e \u003cp\u003eA patient without an identifiable source of infection that, in the context of a negative blood culture, was treated with antibiotics for \u0026gt;\u0026thinsp;72 hours.\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eSepsis rule-out\u003c/strong\u003e \u003cp\u003ePatients that, in the context of a negative blood culture, were treated with antibiotics for \u0026le;\u0026thinsp;72 hours.\u003c/p\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eStatistical Analysis\u003c/h2\u003e \u003cp\u003eThe distribution of blood culture volumes and summary statistics and a point estimate for median and interquartile range were described. For the comparative analyses, the variables analyzed included gender, chronological age, weight, time of collection (day shift from 7AM-7PM, night shift from 7PM-7AM), and source of sample (peripheral, central). We used a chi-squared or Fisher\u0026rsquo;s exact test for binary and t-test or Wilcoxon-Mann-Whitney for continuous variables. In addition, a multivariate analysis was conducted using logistic regression. A p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was considered significant.\u003c/p\u003e \u003cp\u003eTo compare the mean blood volume collected for patients treated for culture-negative sepsis, bloodstream infections, and sepsis rule-out were analyzed using analysis of variance test. Since the blood culture inoculum volumes were not normally distributed, the mean volumes were converted to logarithmic scale for normalization. A p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was considered significant. All analysis were conducted using Stata version 13 (StataCorp LLC).\u003c/p\u003e \u003c/div\u003e"},{"header":"RESULTS","content":"\u003cp\u003eDuring the study period, 742 blood cultures from 292 NICU patients were obtained. Most culture bottles were aerobic (604, 81%). The cohort\u0026rsquo;s demographic characteristics are summarized in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. The median volume of blood inoculated into blood cultures was 1 mL (IQR: 0.6\u0026ndash;1.4) and the mean was 1.06 (SD: 0.7). A total of 259 cultures (35%) had inoculum volumes\u0026thinsp;\u0026lt;\u0026thinsp;0.9 ml. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDemographic characteristics of the study population\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eClinical and environmental factors\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eN (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMedian (IQR)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eWeight (g)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2700 (1340\u0026ndash;3560)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026lt;1500 g\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e203 (27%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026ge;1500 g\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e539 (73%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eAge (days)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e29 (11\u0026ndash;63)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026le;7 days\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e128 (17%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u0026gt;7 days\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e614 (83%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eSource of sample\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePeripheral\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e612 (82%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCentral\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e130 (18%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eGender\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFemale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e321 (43%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e421 (57%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eTime of collection\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eDay shift\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e447 (60%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNight shift\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e295 (40%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eA total of 226 bottles were collected as part of a blood culture set (123 blood culture sets). The mean volume per set was 1.7 ml (SD: 0.7) per set. We compared the volume inoculated in aerobic blood cultures when they were part of a set and when they were collected independently and found no significant difference (p\u0026thinsp;=\u0026thinsp;0.32). In the univariate analysis, blood cultures obtained during the night shift were associated with the collection of lower than recommended sample volumes (p\u0026thinsp;=\u0026thinsp;0.007). This difference remained in the multivariate analysis. No significant differences in blood culture volume were noted by patient gender, chronological age, weight at time of culture collection, or the source of the sample (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAssociation of clinical and environmental factors with volume of blood inoculated in cultures\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePatient Characteristics\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026thinsp;0.9 ml\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;0.9 ml\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eUnivariate analysis p value\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eMultivariate analysis p value\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eWeight, kg (median)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.68\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2.70\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.52\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.51\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAge, days (median)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e27\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e30\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.32\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.87\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSource of sample (n)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.25\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.17\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePeripheral\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e220\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e394\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCentral\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e39\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e89\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGender (n)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.53\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.68\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e143\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e278\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFemale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e116\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e205\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTime of collection (n)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003e0.007\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003e0.006\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eDay shift\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e175\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e308\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eNight shift\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e120\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e139\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eOf the 742 blood cultures identified with documented volumes, 146 (20%) were from patients treated for culture-negative sepsis, 102 (14%) had confirmed bloodstream infections, 291 (39%) underwent a sepsis rule-out, and 194 (26%) were diagnosed with another infectious diagnosis (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). The mean inoculated volume was 1.06 mL (SD: 0.7) for culture-negative sepsis, 1.03 mL (SD: 0.7) for sepsis rule-out, and 1.09 mL (SD: 0.7) for BSI, which were not significantly different (p\u0026thinsp;=\u0026thinsp;0.5). The proportion of blood cultures with inoculum\u0026thinsp;\u0026lt;\u0026thinsp;0.9 mL was 36% (106/291) for sepsis rule-out, 32% (47/146) for culture negative sepsis and 30% (31/102) for bloodstream infection.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eThis single-center study shows that the mean volume of blood cultures collected in a level IV NICU was consistent with prior recommendations of collecting\u0026thinsp;\u0026gt;\u0026thinsp;1mL of blood/bottle, although a significant proportion of inocula are below the recommended volume. In addition, the volume of blood inoculated in blood cultures of patients treated for culture negative sepsis does not differ from those with bloodstream infections and with sepsis rule-out.\u003c/p\u003e \u003cp\u003eDetermining the ideal volume to be collected for neonatal blood cultures is challenging, as it needs to balance the greater diagnostic yield of larger samples[\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e] with the challenges of sample collection and small circulating blood volumes in premature neonates. The mean and median volume inoculated in neonatal blood cultures for our study were 1.06 and 1 mL respectively, which is consistent with most published recommendations[\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e], although there is significant room for improvement, given the large proportion of cultures with \u0026lt;\u0026thinsp;0.9 ml. The earliest study on this topic published in 1986 by Neal \u003cem\u003eet al\u003c/em\u003e., who reported that 56% of the collected samples were \u0026le;\u0026thinsp;0.5 mL.[\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e] Subsequent studies have shown variability in proportion of samples with lower-than-recommended volumes, ranging from 23 to 93%.[\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e] The highest compliance was reported by Woodford et al[\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], who described their experience with blood culture collection in a Level III NICU. Their institutional guidance included collection of a minimum of 1 ml of blood per sample and required documentation of blood culture inoculum in the patient\u0026rsquo;s medical record by the clinician. A similar approach has been used in another recent study by Singh et al., who by implementing a local guideline, educating providers, and fomenting documentation, reduced inappropriate samples from 97\u0026ndash;25%.[\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] These results are encouraging and suggest that systematization of blood culture sampling can greatly optimize the collected volumes.\u003c/p\u003e \u003cp\u003eThis study did not find an association between weight and lower inoculated volumes, which is consistent with past reports.[\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] These results challenge the concept that low-birth-weight is a justification for the collection of suboptimal sample volumes. Similarly, chronological age was not associated with lower volumes, which contrasts the findings by Woodford el at.\u003csup\u003e18\u003c/sup\u003e In their recently published article, the authors found that very-low-birth-weight infants older than 7 days had significantly lower sample volumes when compared to their counterparts that were evaluated for early onset sepsis. This was attributed to the reliable access to central lines during the first week of life (e.g., umbilical lines). However, in our cohort, central access sample collection was not associated with higher blood volumes and most blood cultures in our unit are obtained from a peripheral arterial source to the greatest extent possible.\u003c/p\u003e \u003cp\u003eSamples collected during the night shift had significantly lower volumes, which has not been reported previously in the literature to our knowledge. This could be associated with different providers collecting samples (pediatric residents, fellows, advanced practice providers), or provider fatigue influencing sampling technique, but these factors were outside of our study\u0026rsquo;s scope and were not explored.\u003c/p\u003e \u003cp\u003eOur study showed there\u0026rsquo;s no significant difference in the inoculated volume of aerobic bottles when collected as part of a blood culture set or as single bottles. AAP recommends, when presented with an infant with suspected sepsis, to collect aerobic and anaerobic cultures to optimize pathogen recovery.[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e] This is supported by studies showing that, despite the low prevalence of bacteremia by strict anaerobes in neonates and children (\u0026lt;\u0026thinsp;1%), as many as 11\u0026ndash;22% of clinically relevant isolates were only detectable in anaerobic bottles.[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e] This could be secondary to the anaerobic culture\u0026rsquo;s more optimal conditions for growth of facultative anaerobic organisms that are prevalent in the neonatal population, like \u003cem\u003eStreptococcus agalactiae.\u003c/em\u003e A study showed that there is additional value in the use of anaerobic bottles in very-low-birth-weight infants, where 16% of all cases of early-onset sepsis were secondary to obligate anaerobes.[\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e] The utility of anaerobic blood cultures in the neonatal population needs to be balanced with the decision to inoculum a larger volume into the aerobic culture, based on the sample that the clinician is able to obtain.\u003c/p\u003e \u003cp\u003eTo our knowledge, this is the first study to compare blood culture inoculum volumes from patients with culture-positive and culture-negative sepsis. Volume of samples collected in patients treated for culture negative sepsis did not differ from those with confirmed bloodstream infections and those that were treated for sepsis rule-outs. Cantey \u003cem\u003eet al.\u003c/em\u003e evaluated the use of antibiotics in a Level III NICU, and found that over 70% of antibiotics were administered to patients undergoing \u0026ldquo;sepsis rule-outs\u0026rdquo; and patients diagnosed with culture-negative sepsis.[\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e] Similar results have been reported from Norway and Peru.[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e] When presented with an ill-appearing neonate, the lack of documentation of collected volumes can play a role in clinician mistrust of negative blood culture results.[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] However, our results show that there was no significant difference in the diagnostic yield of the samples obtained in neonates treated for culture-negative sepsis.\u003c/p\u003e \u003cp\u003eOur study has some limitations. First, documentation of direct volume measurements by syringe at the time of sample collection for blood cultures was not obtained which precluded us from comparing these values to the calculated blood volumes. However, weighing inoculated blood culture bottles has been successfully used in previous studies.[\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e] In addition, an average baseline weight for the uninoculated blood culture bottles was used and we did not weight each bottle before inoculation; however, baseline uninoculated bottle weights were validated by repeating the process with bottles from multiple lots during the study, with comparable results. This method would also be more practical and pragmatic for other institutions looking to apply this methodology to increase confidence in blood culture results. We were unable to analyze if low sample volumes increased the chances of false negative results, and this should be further investigated in future studies. Finally, our study population may not be representative of all NICU patients, as it is a single-center study.\u003c/p\u003e"},{"header":"CONCLUSION","content":"\u003cp\u003e Blood culture inocula in our NICU were generally consistent with guideline recommendations, and collection of samples during the night shift was associated with lower than recommended inoculum volumes. Quality improvement efforts are needed to increase the proportion of samples with adequate volume. The volume of blood sampled does not differ in patients with culture-negative sepsis, bloodstream infection, and sepsis rule-out, suggesting this should not be a justification for longer duration of antibiotic therapy.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eConflict of Interest:\u003c/strong\u003e Dr. Soghier is an editor for American Academy of Pediatric \u0026quot;Reference Range Values in Pediatric Care\u0026quot; and \u0026quot;Neonatal Simulation\u0026quot; textbooks. \u0026nbsp; The remaining authors of this manuscript have no conflicts of interest to disclose.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003cstrong\u003eFunding:\u003c/strong\u003e None\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Access Statement:\u0026nbsp;\u003c/strong\u003eData supporting this study cannot be made fully available due to patient confidentiality. De-identified database can be provided upon request to corresponding author.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eFjalstad JW, Stensvold HJ, Bergseng H, Simonsen GS, Salvesen B, Ronnestad AE, et al. Early-onset Sepsis and Antibiotic Exposure in Term Infants: A Nationwide Population-based Study in Norway. Pediatr Infect Dis J. 2016;35:1. \u003c/li\u003e\n\u003cli\u003eRueda MS, Calderon-Anyosa R, Gonzales J, Turin CG, Zea-Vera A, Zegarra J, et al. Antibiotic Overuse in Premature Low-Birth-Weight Infants in a Developing Country. Pediatr Infect Dis J. 2018;38:302-7 \u003c/li\u003e\n\u003cli\u003eSchelonka RL, Chai MK, Yoder BA, Hensley D, Brockett RM, Ascher DP. Volume of blood required to detect common neonatal pathogens. J Pediatr. 1996;129:275\u0026ndash;8. \u003c/li\u003e\n\u003cli\u003eMiller JM, Binnicker MJ, Campbell S, Carroll KC, Chapin KC, Gilligan PH, et al. A Guide to Utilization of the Microbiology Laboratory for Diagnosis of Infectious Diseases: 2018 Update by the Infectious Diseases Society of America and the American Society for Microbiology. Clin Infect Dis Off Publ Infect Dis Soc Am. 2018;67:e1\u0026ndash;94. \u003c/li\u003e\n\u003cli\u003ePuopolo KM, Benitz WE, Zaoutis TE; COMMITTEE ON FETUS AND NEWBORN; COMMITTEE ON INFECTIOUS DISEASES. Management of Neonates Born at \u0026le;34 6/7 Weeks\u0026apos; Gestation With Suspected or Proven Early-Onset Bacterial Sepsis. Pediatrics. Pediatrics. 2018;142(6):e20182896. \u003c/li\u003e\n\u003cli\u003eNeal PR, Kleiman MB, Reynolds JK, Allen SD, Lemons JA, Yu PL. Volume of blood submitted for culture from neonates. J Clin Microbiol. 1986;24:353\u0026ndash;6. \u003c/li\u003e\n\u003cli\u003eConnell TG, Rele M, Cowley D, Buttery JP, Curtis N. How reliable is a negative blood culture result? Volume of blood submitted for culture in routine practice in a children\u0026rsquo;s hospital. Pediatrics. 2007;119:891\u0026ndash;6. \u003c/li\u003e\n\u003cli\u003eSingh MP, Balegar V KK, Angiti RR. The practice of blood volume submitted for culture in a neonatal intensive care unit. Arch Dis Child Fetal Neonatal Ed. 2020;105:600\u0026ndash;4. \u003c/li\u003e\n\u003cli\u003eJawaheer G, Neal TJ, Shaw NJ. Blood culture volume and detection of coagulase negative staphylococcal septicaemia in neonates. Arch Dis Child Fetal Neonatal Ed. 1997;76:F57-58. \u003c/li\u003e\n\u003cli\u003eKuppala VS, Meinzen-Derr J, Morrow AL, Schibler KR. Prolonged initial empirical antibiotic treatment is associated with adverse outcomes in premature infants. J Pediatr. 2011;159:720\u0026ndash;5. \u003c/li\u003e\n\u003cli\u003eCotten CM, Taylor S, Stoll B, Goldberg RN, Hansen NI, Sanchez PJ, et al. Prolonged duration of initial empirical antibiotic treatment is associated with increased rates of necrotizing enterocolitis and death for extremely low birth weight infants. Pediatrics. 2009;123:58\u0026ndash;66. \u003c/li\u003e\n\u003cli\u003eTripathi N, Cotten CM, Smith PB. Antibiotic use and misuse in the neonatal intensive care unit. Clin Perinatol. 2012;39:61\u0026ndash;8. \u003c/li\u003e\n\u003cli\u003eHarris PA, Taylor R, Thielke R, Payne J, Gonzalez N, Conde JG. Research electronic data capture (REDCap)--a metadata-driven methodology and workflow process for providing translational research informatics support. J Biomed Inform. 2009;42:377\u0026ndash;81. \u003c/li\u003e\n\u003cli\u003eTrudnowski RJ, Rico RC. Specific Gravity of Blood and Plasma at 4 and 37 \u0026deg;C. Clin Chem. 1974;20:615\u0026ndash;6. \u003c/li\u003e\n\u003cli\u003eCenters for Disease Control and Prevention (CDC). Bloodstream Infection Event (Central Line-Associated Bloodstream Infection and Non-central Line Associated Bloodstream Infection). Available at: https://www.cdc.gov/nhsn/PDFs/pscManual/4PSC_CLABScurrent.pdf. Accessed 16 June 2022.\u003c/li\u003e\n\u003cli\u003eStoll BJ, Hansen N, Fanaroff AA, Wright LL, Carlo WA, Ehrenkranz RA, et al. Late-onset sepsis in very low birth weight neonates: the experience of the NICHD Neonatal Research Network. Pediatrics. 2002;110:285\u0026ndash;91. \u003c/li\u003e\n\u003cli\u003eYaacobi N, Bar-Meir M, Shchors I, Bromiker R. A prospective controlled trial of the optimal volume for neonatal blood cultures. Pediatr Infect Dis J. 2015;34:351\u0026ndash;4. \u003c/li\u003e\n\u003cli\u003eWoodford EC, Dhudasia MB, Puopolo KM, Skerritt L, Bhavsar M, DeLuca J, et al. Neonatal blood culture inoculant volume: feasibility and challenges. Pediatr Res. Nature Publishing Group; 2021;1\u0026ndash;7. \u003c/li\u003e\n\u003cli\u003eGross I, Gordon O, Abu Ahmad W, Benenson S, et al. Yield of Anaerobic Blood Cultures in Pediatric Emergency Department Patients. Pediatr Infect Dis J. 2018;37:281\u0026ndash;6. \u003c/li\u003e\n\u003cli\u003eGross I, Gordon O, Benenson S, Ahmad WA, Shimonov A, Hashavya S, et al. Using anaerobic blood cultures for infants younger than 90 days rarely showed anaerobic infections but increased yields of bacterial growth. Acta Paediatr Oslo Nor 1992. 2018;107:1043\u0026ndash;8. \u003c/li\u003e\n\u003cli\u003eMukhopadhyay S, Puopolo KM. Clinical and Microbiologic Characteristics of Early-onset Sepsis Among Very Low Birth Weight Infants: Opportunities for Antibiotic Stewardship. Pediatr Infect Dis J. 2017;36:477\u0026ndash;81. \u003c/li\u003e\n\u003cli\u003eCantey JB, Wozniak PS, S\u0026aacute;nchez PJ. Prospective surveillance of antibiotic use in the neonatal intensive care unit: results from the SCOUT study. Pediatr Infect Dis J. 2015;34:267\u0026ndash;72. \u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-perinatology","isNatureJournal":false,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"jp","sideBox":"Learn more about [Journal of Perinatology](http://www.nature.com/jp/)","snPcode":"41372","submissionUrl":"https://mts-jper.nature.com/cgi-bin/main.plex","title":"Journal of Perinatology","twitterHandle":"@jperinatology","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"neonatal sepsis, blood cultures, culture-negative sepsis ","lastPublishedDoi":"10.21203/rs.3.rs-4474756/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4474756/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cb\u003eObjectives\u003c/b\u003e\u003c/p\u003e \u003cp\u003eTo evaluate blood culture sample volumes, identify factors linked to insufficient samples, and compare volumes among neonates treated for culture-negative-sepsis, sepsis-rule-outs, and bloodstream infections (BSI).\u003c/p\u003e\u003cp\u003e\u003cb\u003eMethods\u003c/b\u003e\u003c/p\u003e \u003cp\u003eObservational cohort of blood cultures collected during NICU stay. Association of age, weight, gender, source, and collection time with lower-than-recommended volumes was determined by logistic regression. Blood culture inocula of patients with culture-negative-sepsis, sepsis rule-out, and BSI were compared using ANOVA.\u003c/p\u003e\u003cp\u003e\u003cb\u003eResults\u003c/b\u003e\u003c/p\u003e \u003cp\u003e742 blood cultures were obtained from 292 neonates. Median inoculum was 1mL (IQR:0.6\u0026ndash;1.4), and 259 bottles (35%) had inocula\u0026thinsp;\u0026lt;\u0026thinsp;0.9mL. Night shift sample collection was associated with lower-than-recommended volumes (p\u0026thinsp;=\u0026thinsp;0.006). No difference in sample volumes was observed between culture-negative-sepsis, sepsis-rule-outs, and BSI (p\u0026thinsp;=\u0026thinsp;0.5).\u003c/p\u003e\u003cp\u003e\u003cb\u003eConclusions\u003c/b\u003e\u003c/p\u003e \u003cp\u003eMedian NICU blood culture volumes align with recommendations. Night shift collections correlate with lower volumes. Sample volumes don\u0026rsquo;t differ in patients with culture-negative-sepsis, BSI, and sepsis-rule-out, and should not be a justification for longer duration of antibiotics.\u003c/p\u003e","manuscriptTitle":"Blood Volume Collected for Cultures in Infants with Suspected Neonatal Sepsis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-06-13 02:05:54","doi":"10.21203/rs.3.rs-4474756/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"revise","date":"2024-06-10T13:22:43+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"This content is not available.","date":"2024-06-08T12:18:08+00:00","index":1,"fulltext":"This content is not available."},{"type":"editorInvitedReview","content":"This content is not available.","date":"2024-06-03T18:14:22+00:00","index":2,"fulltext":"This content is not available."},{"type":"reviewerAgreed","content":"This content is not available.","date":"2024-06-03T15:39:48+00:00","index":2,"fulltext":"This content is not available."},{"type":"reviewerAgreed","content":"This content is not available.","date":"2024-05-30T17:17:38+00:00","index":1,"fulltext":"This content is not available."},{"type":"reviewersInvited","content":"","date":"2024-05-29T00:06:48+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-05-28T15:15:59+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-05-25T01:50:43+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Perinatology","date":"2024-05-25T01:50:43+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-perinatology","isNatureJournal":false,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"jp","sideBox":"Learn more about [Journal of Perinatology](http://www.nature.com/jp/)","snPcode":"41372","submissionUrl":"https://mts-jper.nature.com/cgi-bin/main.plex","title":"Journal of Perinatology","twitterHandle":"@jperinatology","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"6fd7849e-20d2-44c5-8e76-a8096c6d938a","owner":[],"postedDate":"June 13th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-09-29T07:06:38+00:00","versionOfRecord":{"articleIdentity":"rs-4474756","link":"https://doi.org/10.1038/s41372-024-02120-0","journal":{"identity":"journal-of-perinatology","isVorOnly":false,"title":"Journal of Perinatology"},"publishedOn":"2024-09-28 04:00:00","publishedOnDateReadable":"September 28th, 2024"},"versionCreatedAt":"2024-06-13 02:05:54","video":"","vorDoi":"10.1038/s41372-024-02120-0","vorDoiUrl":"https://doi.org/10.1038/s41372-024-02120-0","workflowStages":[]},"version":"v1","identity":"rs-4474756","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4474756","identity":"rs-4474756","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}