Sites of blood collection and topical disinfectants associated with contaminated cultures: An ambidirectional cohort study

Abstract Background We aimed to determine whether puncture sites for blood sampling and topical disinfectants are associated with rates of contaminated blood cultures in the emergency department (ED) of a single institution. Methods This single‐center, ambidirectional cohort study of 548 consecutive patients ≥20 years of age was performed in the ED of a university hospital in Japan over a 13‐month period. Pairs of blood samples were collected for aerobic and anaerobic cultures from patients in the ED. Physicians selected puncture sites and topical disinfectants according to their personal preference. Results Potential contamination was identified in 110 of the 548 patients (20.1%). One hundred fourteen (20.8%) patients showed true‐positive results for bacteremia, and 324 (59.1%) patients showed true‐negative results. Multivariate analysis revealed more frequent contamination when puncture sites were disinfected with povidone‐iodine (PVI) than with alcohol/chlorhexidine (ACHX) (adjusted risk difference, 19.1%; 95% confidence interval [CI]), 15.7–22.6; p < 0.001). In terms of blood collection sites, femoral and central venous (CV) catheter with PVI disinfection showed more frequent contamination than venous sites with ACHX (adjusted risk differences: 26.6%, 95% CI 21.3–31.9, p < 0.001 and 41.1%, 95% CI 22.2–59.9, p < 0.001, respectively). Conclusion Rates of contaminated blood cultures were significantly higher when blood was collected from the CV catheter or femoral sites with PVI as the topical disinfectant.

that only two methods can achieve this: sampling from a separate venipuncture site or a well-trained phlebotomy team. 3Although topical 1.0% alcohol/chlorhexidine gluconate (ACHX) reduces blood culture contamination more effectively than 10% aqueous povidone-iodine (PVI), 4,5 both agents are routinely applied at our institution as topical disinfectants before blood sampling.Rates of false-positive culture are significantly reduced when blood is sampled from various venipuncture sites compared with intravenous or central venous catheters. 6,7ysicians at our institution may sample blood from various sites, such as intravenous and central venous catheters, as well as femoral arteries and veins, according to personal preference.
The emergency department (ED) is the frontline to investigate unwell patients, with blood culture contamination rates above the recommended 3%. 3,8A previous study for a period of 6 months revealed that blood sampled from femoral arteries or veins and using PVI was associated with a significantly higher contamination rate in our ED. 9The study showed that rates of contamination of femoral sites, peripheral sites, and central venous catheter were 73.6%, 16.1%, and 7.7%, respectively. 9 thus aimed to investigate and validate in greater detail which puncture sites for blood sampling and which topical disinfectants are associated with blood culture contamination in patients and samples from a single ED over a longer period.

| Study design
This single-center, ambidirectional cohort study proceeded at the ED of a university hospital in Japan between August 1, 2018, and August 31, 2019.The hospital is an 882-bed university teaching hospital with 8000 adults presenting to the ED annually.Strengthening the Reporting of Observational studies in Epidemiology (STROBE) guidelines were used to design and report the results from this study. 10The institutional review board at our institution approved the study protocol (675(2476)) and waived the need to obtain written informed consent.

| Patients
This study included 548 consecutive patients ≥20 years of age from whom blood was sampled in the ED.Inclusion criteria were as follows: age ≥20 years and at least one pair of blood cultures collected in the ED.Patients were excluded if all blood samples were collected elsewhere.If one pair of blood samples was collected at our ED and another was collected elsewhere or no second pair was collected, then only the pair collected at our ED was analyzed.[13][14] Death data were also analyzed in February 2021.

| Blood cultures
Nurses and other medical staff at our institution are not permitted to collect blood for cultures.Only physicians, typically first-or secondyear residents, are permitted to collect blood samples for blood culture in the ED.Blood (14-20 mL) from peripheral veins or arteries was sampled for aerobic and anaerobic cultures (7-10 mL each) in BacT/Alert FA Plus and FN Plus resin bottles (bioMérieux Inc., Durham, NC, USA).The use of sterile gloves was mandatory at our institution when the physician took blood culture samples.Physicians selected the topical disinfectants such as ACHX, PVI, and alcohol available in the ED, according to their personal preferences.A blood culture was considered contaminated if one or more of the following organisms were identified in one of the two blood cultures: coagulase-negative staphylococci (CoNS), Propionibacterium acnes, Micrococci, Corynebacteria, Bacillus species other than Bacillus anthracis, or Clostridium perfringens. 7,15,16ridans group streptococci are regarded as contaminants based on the described criteria, 7,15 but are not considered contaminants in our institute.Polymicrobial cultures showing a mixture of contaminant and true pathogens were regarded as contaminated. 14A culture was defined as "negative" when bacterial growth was absent.The source of infection was identified based on a chart review with other cultures such as sputum, urine, and ascites or with other modalities including ultrasonography, X-ray, computed tomography (CT), and magnetic resonance imaging (MRI).Details are described in Table 1.

| Statistical analysis
Categorical variables are described as frequencies and percentages (%), and continuous variables are shown as mean with standard deviation (SD).Data were compared using one-way analysis of variance, the Chi-squared test, and Fisher's exact test, as appropriate.Differences in risk and robust 95% confidence intervals (CIs) of contamination according to sites and topical disinfectants were estimated using uni-and multivariate analyses with modified leastsquares regression and a robust standard error estimator. 17,18The same patients were considered to be a random effect in the above model.Age, sex, and doctors' experience were adjusted as confounders in multivariate analyses.Because blood can be sampled from few sites, we also included blood in five categories: CV catheter, blood sampled from a newly inserted central venous (CV) catheter; femoral, blood sampled from the femoral artery or vein; other, blood sampled from a newly inserted arterial catheter and implanted port; venous, blood sampled from venipuncture without catheter insertion; and venous catheter, blood sampled from a newly inserted venous catheter.Because we did not have many topical disinfectants to assess, we included only PVI, ACHX, and other types (alcohol and benzalkonium) in analyses.We did not impute missing values.For all statistical investigations, values of p < 0.05 were taken as significant.
All analyses were performed using STATA version 16.1 (Stata Corp., College Station, TX, USA).

| Baseline characteristics
We analyzed data from 548 patients and 1066 pairs of blood cultures between August 1, 2018, and August 31, 2019.A total of 138 (12.9%) potential contaminants from 110 patients (20.1%) were found in blood cultures, the most common of which was Staphylococcus epidermidis in 55 of the 138 blood samples (39.9%), followed by Staphylococcus hominis in samples (18.1%).Four patients (2.9%) had a mixture of true bacteremia and contaminating isolates.We identified true bacteremia in patients (20.8%) and 224 blood samples (21.0%), with Escherichia coli as the most prevalent microorganism, in 59 of the 224 blood samples (26.3%).Cultured blood samples from 324 patients (59.1%) and 704 blood samples (66.0%) were identified as true negative.The most common source of infection in 47 patients (41.2%) with true bacteremia was urinary tract infection, whereas a pulmonary source was the most prevalent in 41 patients (37.3%) with contaminated cultures and 94 patients (29.0%) with true-negative cultures.
These two sources significantly differed among the three groups (pulmonary and urinary tract; both p < 0.001).Only one pair of blood samples was cultured from 31 patients.Neither total number of deaths nor death within 30 days differed significantly between the three groups.
Tables 1 and 2 show other baseline characteristics of the three groups of patients and blood cultures.Status of physicians collecting blood samples for blood culture in the ED is shown in Table 3.

| Sites and topical disinfectants
Femoral arteries and veins tended to be sampled in most patients in all three groups, but at different ratios (Tables 1 and 2).Over 80% of blood samples from patients with contaminants were collected from femoral sites (mostly the femoral artery), whereas <50% of blood samples from patients with negative cultures were collected from these sites.Blood sampled from a newly inserted central venous catheter conferred the greatest risk of contamination when taken as an independent factor.However, the number of samples was small (n = 30).
Topical disinfectants also differed significantly among groups (Table 2).Disinfection with PVI was performed for sites in >90% of patients with contaminated blood cultures, compared with <65% among patients in the other two groups.Other topical disinfectants comprising alcohol or benzalkonium are also shown in Table 2.

| Proportion of contamination by sites and topical disinfectants
With reference to ACHX, univariate analysis using modified least- bers of these combinations were 25 and 1, respectively (Table 5).
With reference to first-year residents, second-year residents, physicians experienced less than or equal to10 years, and physicians experienced more than 10 years were not significantly associated with contamination (Table S1).

| DISCUSS ION
This single-center, ambidirectional cohort study found that blood samples collected from femoral areas and CV catheter insertion at the internal jugular vein disinfected with PVI were significantly associated with contaminated blood cultures in the ED.As revealed by the previous study, the most common source of infection among patients with true bacteremia was urinary tract infection, and most such patients had pyelonephritis. 9In contrast, the most prevalent source of infection among patients with contamination was pulmonary disease, with most such patients having aspiration pneumonia, again similar to ).Physicians experienced less than or equal to 10 years mean that physicians who had finished their postgraduate clinical training for 2 years, and their experience was more than 2 years and less than or equal to 10 years.Physicians experienced more than 10 years mean that physicians who had finished their residency program, and their experience was more than 10 years.
the previous study. 9Mortality rates were not significantly different among contamination, true-negative results and true bacteremia.
We also found that three combinations (femoral puncture site with PVI, CV catheter insertion at internal jugular vein with PVI, and venous venipuncture with PVI) comprised independent risk factors for blood culture contamination.A previous similar study showed that the femoral site is unsuitable for blood cultures, 9,19 but this investigation suggested that the femoral site may be acceptable only if ACHX is used.Other disinfectants (alcohol or benzalkonium or both) might be effective to prevent contamination when blood samples were collected from the femoral site.However, the combination of the femoral site and ACHX should not be selected because this combination was only seen for 14 samples, and the data were thus insufficient for reliable statistical analysis.Furthermore, as femoral sites are colonized more often than other sites, 20 they are associated with catheter-related bloodstream infection. 210][21][22] In addition, the CV catheter is not suitable as a route for blood culture sampling compared with arterial lines or direct peripheral venipuncture. 23,24The contamination rate in the present study was highest (40.0%) for the 30 blood samples collected from central catheters newly inserted into the internal jugular vein in the ED.
However, this number was relatively low.Only one case of CV catheter insertion using alcohol was not contaminated, and this combination was significantly less frequent than the combination of ACHX with venous venipuncture without catheter insertion.However, we should not use this combination because only one case showed this combination, and the data were thus insufficient for reliable statistical analysis.ACHX has been recommended when a CV catheter is inserted. 25Handling and disinfection of access points, such as needleless connectors, antiseptic barrier caps, and catheter hubs, are complicated with CV catheters.Hence, the number of manipulations per blood collection through a newly inserted CV catheter is higher than that with arterial catheters or venous catheters. 24,26veral reports have described associations between blood samples collected from newly inserted peripheral venous catheter and blood culture contamination. 6,27According to those reports, collecting blood culture specimens through a newly inserted peripheral venous catheter increases the risk of contamination compared with venipuncture.Our results suggested that blood culture from a newly inserted peripheral venous catheter might be the best procedure to prevent blood culture contamination in the ED, and a review article also emphasized the need to balance the practical advantages of this method against the risk of increased contamination.Several reports have described associations between topical disinfectants and blood culture contamination. 29,30A meta-analysis found that blood culture contamination was more significantly reduced by ACHX than by PVI. 4 However, physicians have tended to disinfect puncture sites with PVI more frequently than with ACHX at our hospital, for various reasons.First, physicians and ED staff are more familiar with PVI than with ACHX because it has been applied as a skin disinfectant for many years.Second, residents and medical students are not educated about blood culture procedures while at university.
Pneumonia was the most common illness among patients with contaminated and true-negative blood cultures.2][33] One reason for the very high contamination rate at our hospital was the high proportion of blood samples collected from femoral areas disinfected with PVI.Blood samples should be cultured from selected immunocompromised patients, those with complicating urinary tract infection under antibiotic therapy at the time of blood collection, and patients with suspected endocarditis. 32,34l physicians are mandated to complete 2 years of accredited postgraduate clinical training in Japan with rotations in major clinical specialties, including emergency medicine. 35Our result and previous study showed that experience status of the physician performing sample collection was not associated with contaminated blood cultures in the ED (Table 3; Table S1). 22Initially, sample collection by residents was considered a likely risk factor for blood culture contamination; however, no such significant association with blood culture contamination was identified.Residents and physicians who chose femoral area with PVI were associated with contaminated blood cultures in the ED.
Several strategies have been advocated to reduce rates of blood culture contamination.Sampling from various venipuncture sites and reliance on a well-trained phlebotomy team can reduce these rates. 3Furthermore, switching from PVI to ACHX or other topical disinfectants and informational intervention and feedback might reduce rates of blood culture contamination, even when physicians conduct phlebotomies. 36,37is study shows several limitations.Our patient cohort was not large, and some parameters could not be conclusively determined.
Nevertheless, specific injection sites and PVI were associated with significantly increased contamination rates.Some physicians were aware of this study proceeding within the ED and might have been more attentive when collecting blood than they might have been in wards.Physicians could select their preferred topical disinfectant for blood sampling, which also have represented a confounder, because many studies have associated contamination more with PVI than with ACHX.Blood collection sites prepared using the same disinfectant should be compared under the same conditions.

CO N FLI C T O F I NTE R E S T S TATE M E NT
The authors have stated explicitly that there are no conflicts of interest in connection with this article.

E TH I C S A PPROVA L S TATE M E NT
The Institutional Review Board (IRB) at Osaka Medical College approved the study protocol (No. 675(2476)).

PATI E NT CO N S E NT S TATE M E NT
Patient consent statement was waived the need to obtain written, informed consent.

CLI N I C A L TR I A L R EG I S TR ATI O N
The study protocol (No. 675(2476)) at Osaka Medical College (Osaka Medical and Pharmaceutical University).

R E FE R E N C E S
This single-center, ambidirectional cohort study found that femoral puncture sites and PVI as well as CV catheter insertion and PVI were independent risk factors for blood culture contamination.Newly inserted venous catheters with ACHX should be used to obtain blood samples to reduce culture contamination.AUTH O R CO NTR I B UTI O N SKoshi Ota and Daisuke Nishioka designed the study and wrote the initial draft of the manuscript.Kanna Ota, Daisuke Nishioka, Emi Hamada, Yuriko Shibatad, and Akira Takasu contributed to the analysis and interpretation of data and assisted in the preparation of the manuscript.All authors contributed to data collection and interpretation and critically reviewed the manuscript.All authors approved the final version of the manuscript and agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.
Characteristics of blood cultures in the emergency department.Status of physicians collecting blood samples for blood culture in the emergency department.
TA B L E 2 a Fisher's exact test was performed because of the small numbers of patients in several cells.Other comparisons were analyzed using the Chi-squared test.TA B L E 3 28 Univariate analysis using modified least-squares regression.Multivariate analysis using modified least-squares regression.
TA B L E 4