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Background: The critical factors for optimal recovery of microbial pathogens from blood are not only the basis for how we perform blood cultures on a daily basis, but are also important for development of all current blood-culture systems. Because much of this research was conducted and published between 1975 and 2010 there is a general sense that many physicians and scientists may not be familiar with it, but it is critical for performing and interpreting blood cultures., Objectives: To review the critical factors in the laboratory recovery and isolation of pathogenic microorganisms in blood., Sources: Literature review of published papers, select reviews and updated guidelines., Content: This review focuses on the critical factors that affect the recovery isolation of pathogenic microorganisms from individuals with bloodstream infections. Contemporary blood-culture systems, and current methods for blood-culture collection, have been designed to incorporate these critical factors so as to optimize recovery and isolation of pathogens while minimizing contamination., Implications: It is important for microbiologists and practitioners to understand how and why these critical factors affect current blood-culture practices to improve patient management. Future research should emphasize which of these critical factors will still play a role in the era of molecular diagnostic tests, which will no longer be relevant, and which new critical factors have yet to be defined., (Copyright © 2019 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Enteric dysbiosis is a characteristic feature of progressive human immunodeficiency virus type 1 (HIV-1) infection but has not been observed in simian immunodeficiency virus (SIVmac)-infected macaques, including in animals with end-stage disease. This has raised questions concerning the mechanisms underlying the HIV-1 associated enteropathy, with factors other than virus infection, such as lifestyle and antibiotic use, implicated as playing possible causal roles. Simian immunodeficiency virus of chimpanzees (SIVcpz) is also associated with increased mortality in wild-living communities, and like HIV-1 and SIVmac, can cause CD4 + T cell depletion and immunodeficiency in infected individuals. Given the central role of the intestinal microbiome in mammalian health, we asked whether gut microbial constituents could be identified that are indicative of SIVcpz status and/or disease progression. Here, we characterized the gut microbiome of SIVcpz-infected and -uninfected chimpanzees in Gombe National Park, Tanzania. Subjecting a small number of fecal samples (N = 9) to metagenomic (shotgun) sequencing, we found bacteria of the family Prevotellaceae to be enriched in SIVcpz-infected chimpanzees. However, 16S rRNA gene sequencing of a larger number of samples (N = 123) failed to show significant differences in both the composition and diversity (alpha and beta) of gut bacterial communities between infected (N = 24) and uninfected (N = 26) chimpanzees. Similarly, chimpanzee stool-associated circular virus (Chi-SCV) and chimpanzee adenovirus (ChAdV) identified by metagenomic sequencing were neither more prevalent nor more abundant in SIVcpz-infected individuals. However, fecal samples collected from SIVcpz-infected chimpanzees within 5 months before their AIDS-related death exhibited significant compositional changes in their gut bacteriome. These data indicate that SIVcpz-infected chimpanzees retain a stable gut microbiome throughout much of their natural infection course, with a significant destabilization of bacterial (but not viral) communities observed only in individuals with known immunodeficiency within the last several months before their death. Am. J. Primatol. 80:e22515, 2018. © 2015 Wiley Periodicals, Inc., (© 2015 Wiley Periodicals, Inc.)

Simian immunodeficiency virus of chimpanzees (SIVcpz) is the ancestor of human immunodeficiency virus type 1 (HIV-1), the etiologic agent of acquired immunodeficiency syndrome (AIDS) in humans. Like HIV-1-infected humans, SIVcpz-infected chimpanzees can develop AIDS-like symptoms. Because SIVcpz/HIV-1 may disrupt regulation of the gut microbiome and because it has not been possible to sample individual humans pre- and postinfection, we investigated the influence of infection on gut communities through long-term monitoring of chimpanzees from Gombe National Park, Tanzania. SIVcpz infection accelerated the rate of change in gut microbiota composition within individuals for periods of years after the initial infection and led to gut communities marked by high frequencies of pathogen-containing bacterial genera absent from SIVcpz-negative individuals. Our results indicate that immune function maintains temporally stable gut communities that are lost when individuals become infected with SIVcpz., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Microbes inhabiting the human gastrointestinal tract tend to adopt one of three characteristic community structures, called 'enterotypes', each of which is overrepresented by a distinct set of bacterial genera. Here we report that the gut microbiotae of chimpanzees also assort into enterotypes and that these chimpanzee enterotypes are compositionally analogous to those of humans. Through the analysis of longitudinal samples, we show that the microbial signatures of the enterotypes are stable over time, but that individual hosts switch between enterotypes over periods longer than a year. These results support the hypothesis that enterotypic variation was present in populations of great apes before the divergence of humans and chimpanzees.

To determine the optimal anaerobic companion bottle to pair with BACTEC Plus Aerobic/F medium for recovery of pathogenic microorganisms from adult patients with bacteremia and fungemia, we compared Plus Anaerobic/F bottles with Standard Anaerobic/F bottles, each of which was filled with 4 to 6 ml of blood. The two bottles were paired with a Plus Aerobic/F bottle filled with 8 to 12 ml of blood. A total of 14,011 blood culture sets were obtained. Of these, 11,583 sets were received with all three bottles filled adequately and 12,257 were received with both anaerobic bottles filled adequately. Of 818 clinically important isolates detected in one or both adequately filled anaerobic bottles, significantly more staphylococci (P < 0.001), streptococci (P < 0.005), Escherichia coli isolates (P < 0.02), Klebsiella pneumoniae isolates (P < 0.005), and all microorganisms combined (P < 0.001) were detected in Plus Anaerobic/F bottles. In contrast, significantly more anaerobic gram-negative bacilli were detected in Standard Anaerobic/F bottles (P < 0.05). Of 397 unimicrobial episodes of septicemia, 354 were detected with both pairs, 30 were detected with Plus Aerobic/F-Plus Anaerobic/F pairs only, and 13 were detected with Plus Aerobic/F-Standard Anaerobic/F pairs only (P < 0.05). Significantly more episodes of bacteremia caused by members of the family Enterobacteriaceae (P < 0.05) and aerobic and facultative gram-positive bacteria (P < 0.025) were detected with Plus Anaerobic/F bottles only. In a paired-bottle analysis, 810 of 950 isolates were recovered from both pairs, 90 were recovered from Plus Aerobic/F-Plus Anaerobic/F pairs only, and 50 were recovered from Plus Aerobic/F-Standard Anaerobic/F pairs only (P < 0.001). Paired Plus Aerobic/F-Plus Anaerobic/F bottles yielded significantly more staphylococci (P < 0.001), streptococci (P < 0.05), and members of the family Enterobacteriaceae (P <0.001). We conclude that Plus Anaerobic/F bottles detect more microorganisms and episodes of bacteremia and fungemia than Standard Anaerobic/F bottles as companion bottles to Plus Aerobic/F bottles in the BACTEC 9240 blood culture system.

Iodophor and alcohol pledgets were compared with the Medi-Flex Prep Kit II for skin disinfection before venipuncture. Of 12,367 blood cultures collected, 6,362 were done with conventional pledgets and 6, 005 were done with Medi-Flex kits. Contamination occurred in 351 of 6,362 blood cultures (5.5%; range, 3.7 to 8.1%) with conventional pledgets versus 328 of 6,005 (5.5%; range, 3.5 to 7.5%) with Medi-Flex kits.

The overall recovery of organisms and time to detection with the BACTEC 9050 and BACTEC 9240 systems were compared in a multicenter evaluation. In the first phase of the study, a total of 4,383 compliant aerobic (Plus Aerobic/F) blood culture sets were processed. There was no significant difference in the recovery of individual groups of organisms with the two systems, with the exception of Streptococcus pneumoniae which was isolated more frequently with BACTEC 9050. False-positive signals occurred more often with BACTEC 9240 (58 cultures) than with BACTEC 9050 (43 cultures), but false-negative cultures were uncommon with both systems (3 cultures for each system). Time to detection of positive cultures of clinically significant organisms was essentially the same with both instruments. In the second phase of the study, 2,431 compliant anaerobic (Plus Anaerobic/F) blood culture sets were processed. There was no significant difference in the recovery of organisms with BACTEC 9050 compared with BACTEC 9240. Significantly (P < 0.03) more false-positive signals occurred with BACTEC 9240 (15 cultures) than with BACTEC 9050 (4 cultures). Likewise, more false-negative cultures occurred with BACTEC 9240 (11 cultures) than with BACTEC 9050 (8 cultures). Time to detection of positive cultures of clinically significant organisms was essentially the same with both systems with the exception of anaerobes (N = 10), which were recovered earlier (P < 0.01) with BACTEC 9240 (35.0 h) than with BACTEC 9050 (61.4 h).

Two recent multicenter blood culture studies found that BacT/Alert FAN (FAN) bottles (Organon Teknika, Durham, N.C.) had increased yields in detecting bacteremia and fungemia compared with standard BacT/Alert (STD) bottles. Because the clinical importance of this increase in microbial recovery is unknown, we performed a retrospective analysis to determine the frequency with which FAN bottles were the sole means of detecting an episode of bacteremia. There were 1,047 positive blood cultures in which both study bottles were adequately filled and the organism isolated was judged to be the cause of sepsis: 240 (23%) were positive only in FAN bottles and 73 (7%) were positive only in STD bottles. Of a total of 664 episodes of bacteremia, 126 (19%) were identified only by FAN bottles and 43 (7%) were identified only by STD bottles (P < 0.0001). Episodes detected only by FAN bottles more often were recurrent events (23 of 126, or 18%) than episodes detected only by STD bottles (2 of 43, or 5%) (P < 0.05) and more commonly occurred in patients receiving theoretically effective antibiotic therapy (33 of 126 [26%] versus 4 of 43 [9%]) (P < 0.05). The medical records for patients with 127 of these episodes (92 FAN bottles only; 35 STD bottles only) were available for review. More than half of both FAN bottle-only (60 of 92, or 65%) and STD bottle-only (20 of 35, or 57%) episodes were judged to be clinically important. We conclude that FAN bottles improve the detection of bacteremia and that the majority of the additional episodes detected are clinically important. The benefits of the greater yield in specific patient populations must be balanced against the higher costs of FAN bottles.

FAN medium was formulated to improve microbial recovery, particularly for fastidious microorganisms and for microorganisms causing sepsis in patients receiving antimicrobial therapy. In a controlled clinical evaluation performed at four university-affiliated hospitals, FAN anaerobic bottles were compared with standard anaerobic bottles for yield, speed of detection of microbial growth, and detection of septic episodes. A total of 10,431 blood culture sets were received; both anaerobic bottles of 7,694 blood culture sets were adequately filled with blood. Altogether, 925 isolates were recovered: 557 that were the cause of sepsis, 99 that were indeterminate as the cause of sepsis, and 269 contaminants. More Staphylococcus aureus (P < 0.001), coagulase-negative staphylococci (P < 0.001), Escherichia coli (P < 0.02), and all microorganisms combined (P < 0.005) were recovered from FAN bottles; more nonfermentative gram-negative bacilli (P < 0.05), Torulopsis glabrata (P < 0.001), and other yeasts (P < 0.01) were recovered from standard bottles. Growth of S. aureus (P < 0.001), coagulase-negative staphylococci (P < 0.001), Enterococcus faecalis (P < 0.025), streptococci other than Streptococcus pneumoniae (P < 0.01), and all microorganisms combined (P < 0.001) was detected earlier in standard bottles; growth of more isolates of E. coli (P < 0.05) and anaerobic bacteria (P < 0.01) was detected earlier in FAN bottles. The mean times to detection were 14.2 and 16.1 h for standard and FAN bottles, respectively. More septic episodes caused by S. aureus (P < 0.001), coagulase-negative staphylococci (P < 0.005), members of the family Enterobacteriaceae (P < 0.02), and all microorganisms combined (P < 0.02) were detected in FAN bottles; more septic episodes caused by nonfermentative gram-negative bacilli (P < 0.025) and yeasts (P < 0.005) were detected in standard bottles. In summary, more isolates (except for strict aerobes) were recovered from FAN bottles than from standard anaerobic bottles. Similarly, significant more septic episodes (except for those caused by strict aerobes) were detected with FAN bottles than with standard anaerobic bottles. With the exception of E. coli and anaerobic bacteria, growth of more isolates was detected earlier in standard anaerobic bottles.

Bottles developed for use in the BacT/Alert automated blood culture system (Organon Teknika Corp., Durham, N.C.) can accept up to 10 ml of blood without falling below a 1:5 ratio of blood to broth. We compared the yield and speed of detection of microorganisms in 13,128 adequately filled, paired, aerobic bottles inoculated with 5 versus 10 ml of blood at three university hospitals. A total of 798 microorganisms causing sepsis grew in one or both bottles. The overall recovery of microorganisms from 10-ml samples exceeded that from 5-ml samples (P < 0.001); the increased yield attributed to the additional 5 ml in the samples was 7.2%. The increased yield from 10-ml inocula was most marked for Escherichia coli (P < 0.01) and other members of the family Enterobacteriaceae (P < 0.001). Ten-milliliter samples did not yield more gram-positive bacteria, nonfermentative gram-negative rods, or yeasts. When both bottles were positive, the bottles inoculated with 10 ml of blood showed growth sooner (P < 0.001). Earlier detection with 10-ml inocula was especially notable for coagulase-negative staphylococci (P < 0.001), streptococci (P < 0.001), E. coli (P < 0.025), and other members of the family Enterobacteriaceae (P < 0.025). We conclude that an increase in the volume of blood inoculated into BacT/Alert aerobic blood culture bottles from 5 to 10 ml will increase the overall yield and the speed of detection of clinically important blood pathogens.

Most, if not all, rare or fastidious microbial pathogens can be recovered from standard blood culture bottles and media using routine laboratory methods. Because they are only rarely recovered from blood cultures, relatively few data exist concerning the ability of different blood culture systems to recover these microorganisms. Thus, clinical microbiologists should take a conservative approach when processing blood cultures that may contain one of the microorganisms reviewed in this article, including the use of additional media, media supplements, blind and terminal subcultures, or prolonged incubation and testing.

The BACTEC high-blood-volume fungal medium (HBV-FM) (Becton Dickinson Diagnostic Instrument Systems, Sparks, Md.) was compared with the Isolator (IS) tube and the BACTEC Plus 26 (BP26) blood culture bottle for the ability to recover fungi from the blood of adult patients suspected of having fungemia. A total of 6,836 blood culture sets that fulfilled criteria for inclusion in the study were received. Three separate comparisons were performed: 4,907 HBV-FM versus IS, 4,886 BP26 versus HBV-FM, and 4,949 BP26 versus IS. For the HBV-FM versus IS comparison, 218 isolates were recovered: 125 (57.3%) were bacteria and 93 (42.7%) were fungi. HBV-FM was comparable to IS for recovery of yeasts, but IS was superior for recovery of Histoplasma capsulatum (25 versus 0 isolates recovered [P < 0.001]). Growth of Torulopsis glabrata was detected earlier (P < 0.05) in HBV-FM bottles. For the BP26 versus HBV-FM comparison, 229 isolates were recovered: 161 (70.3%) were bacteria, and 68 (29.7%) were fungi. HBV-FM was superior for recovery of T. glabrata (P < 0.025) and all fungi combined (P < 0.025). There were no statistically significant differences in the speed of detection of microbial growth. For the BP26 versus IS comparison, 251 isolates were recovered: 165 (65.7%) were bacteria, and 86 (34.2%) were fungi. IS was superior for recovery of H. capsulatum (P < 0.001), T. glabrata (P < 0.05), and fungi other than H. capsulatum (P < 0.025). BP26 was superior for recovery of all bacteria combined (P < 0.001) and viridans group streptococci (P < 0.01). Growth of T. glabrata (P < 0.05) was detected earlier in IS tubes. Growth of Staphylococcus aureus (P < 0.01), viridans group streptococci (P < 0.01), Pseudomonas aeruginosa (P < 0.05), and all microorganisms combined (P < 0.05) was detected earlier in BP26 bottles. For yeast, 57 of 59 (96.6%), 79 of 80 (98.7%), and 64 of 67(95.5%) were recovered from BP26 bottles, HBV-FM bottles, and IS tubes, respectively, by day 14; for H. capsulatum, 14 of 36 (38%) isolates were recovered from IS tubes by day 14. Mean times of recovery were similar for BACTEC bottles and IS. We conclude that (i) for recovery of fungi from blood cultures, HBV-FM is equivalent to IS (with the exception of H. capsulatum); (ii) for recovery of bacteria, BP26 is superior to IS; (iii) BP26 bottles are inferior to both HBV-FM bottles and IS tubes for recovery of T. glabrata; and (iv) HBV-FM bottles must be paired with another blood culture bottle or system to optimize detection of bacteremia.

Recently, we published a comparison of the BacT/Alert blood culture system with the BACTEC 660/730 nonradiometric blood culture system using blood inocula of 5 ml per bottle. By reanalyzing data collected during that study, we found that, for true-positive isolates causing bacteremia or fungemia, 363 (97.6%) of 376 and 341 (97.7%) of 349 isolates were recovered by the end of day 5 of testing, and 364 (97.9%) of 376 and 343 (98.3%) of 349 isolates were recovered by the end of day 6 of testing for aerobic and anaerobic bottles, respectively. Most isolates recovered on days 6 (24 of 27) and 7 (20 of 25) of testing were either contaminants or indeterminate as a cause of sepsis. When used as recommended by the manufacturer, only six (1.3%) of 464 clinically important isolates recovered on test days 6-7 would have gone undetected had testing been limited to 5 days and four (0.9%) of 464 had testing been limited to 6 days. We conclude that BacT/Alert bottles can be tested for as few as 5 days and then discarded with minimal loss of true-positive isolates and maximal reduction of contaminants.