Routine Quantitative Blood Cultures in Children With Haemophilus influenzae or Streptococcus pneumoniae Bacteremia

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Routine

Quantitative

Blood

Cultures

in Children

With

Haemophilus

influenzae

or

Streptococcus

pneumoniae

Bacteremia

Louis M. Bell, MD, Gershon Alpert, MD, Joseph M. Campos, PhD, and Stanley A. Plotkin, MD

From the Division of Infectious Diseases, Children’s Hospital of Philadelphia, Philadelphia

ABSTRACT. The potential clinical value of quantitative blood cultures determined by a commercially available lysis-direct plating method was studied in 50 children with either Haemophilus influenzae or Streptococcus pneumoniae bacteremia. The magnitude of bacteremia correlated with the severity of the infection; patients with 100 colony-forming units per milliliter were signifi-cantly more likely to have meningitis (P < .01,

x2

= 7.5).

On the other hand, all patients with S. pneumoniae

bacteremia with colony counts lower than 15 colony-forming units per milliliter had “occult bacteremia” with no focus of infection. The data suggest that patients with higher levels of bacteremia have more severe disease. Quantitative blood culture results may be helpful in iden-tifying which children are at risk for invasive disease.

Pediatrics 1985;76:901-904; quantitative blood culture, oc-cult bacteremia, Haemophilus influenzae, Streptococcus pneumonio.e, meningitis.

Bacteremia occurs in approximately 4% of febnile children in the outpatient setting.’ Some of these children have focal signs of infection and a serious

illness, whereas others have what has been termed “occult” bactememia and may suffer no sequelae of

the

bactenemia.2 Quantitative blood cultures have been shown to be helpful in a variety of clinical situations including endocarditis, catheter-related

sepsis, and persistent septicemia.36 In addition, there is evidence that the magnitude of bacteremia correlates with clinical disease, although only

lim-ited

information has been collected.7

In the present study, to determine the magnitude

of bacteremia caused by Haemophilus influenzae and Streptococcus pneumoniae in ill children, we

Received for publication Nov 5, 1984; accepted Jan 28, 1985. Reprint requests to (L.M.B.) The Children’s Hospital of Phila-delphia, Division of Infectious Diseases, 34th and Civic Center Blvd, Philadelphia, PA 19104.

used

a commercially

available

blood

culture

device

that enabled routine quantitation of blood cultures.

We attempted to correlate the magnitude of

bacten-emia with the presence of disease and prognosis.

MATERIALS AND METHODS

Patient Population

During the period August 1982 to January 1983

and again from April 1983 to July 1983, we

com-pared the Isolator 1.5 Microbial Tube (E. I. duPont

de Nemouns and Co, Wilmington, DE) system for blood culturing to a conventional broth culture

bottle method.8 After the evaluations, the Isolator

1.5 Microbial Tube was adopted for routine use in November 1983.

During the 18-month period when the Isolator

1.5 system was initially tested and then routinely used, the charts of all patients with bacteremia caused by H influenzae type b and S pneumoniae were reviewed. Cases were accepted for evaluation

if a single bacterial pathogen was identified from a

peripheral blood culture in an immunocompetent patient with no history of having received recent antibiotic therapy. Blood was drawn and cultured and treatment was begun at the discretion of the physician on patients with certain localized

infec-tions and on most children less than 2 years of age

with a high fever (39#{176}C), with or without an identifiable focus of infection.

Blood Cultures

Blood samples were obtained from patients by

the pediatric housestaff using standard aseptic

technique. The housestaff inoculated the specimens

into sterile vacuum tubes containing 0.7 mL of sodium polyanetholesulfonate (SPS)

(Becton-Dickinson, Rutherford, NJ) and these were

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10.0

8.0

0 1 2

C S. Pneumoniae

. H.Influenzae

902 QUANTITATIVE BLOOD CULTURES

During periods of the comparative blood culture

evaluation, blood samples obtained on the wards

were divided in the microbiology laboratory

be-tween Isolator 1.5 and conventional blood culture

broth bottles. Specimens added to Isolator 1.5 were

vortexed to facilitate lysis and 0.25 to 0.30 mL was

inoculated onto each agar medium (one to four

chocolate agar plates and one CDC anaerobic blood

agar plate). Chocolate agar plates were incubated

at 35#{176}Cunder aerobic conditions (5% to 10% CO2)

for at least two days, and CDC anaerobic blood agan plates were incubated at 35#{176}Cunder anaerobic con-ditions for at least four days.

During the period of routine use of the Isolator

1.5, aliquots (0.25 to 0.30 mL) of the lysed blood specimens were inoculated onto one 5% sheep blood

agan plate and one chocolate agam plate; the

remain-den of the lysate was inoculated into a tube

contain-ing 10 mL of modified eugonic broth (Remel

Lab-oratories, Lenexa, KS). Blood agan and chocolate

agan plates were incubated at 35#{176}Cfor three days

and the modified eugonic broth was incubated at

35#{176}Cfor seven days.

Transport Time and Quantitative Blood Cultures

The effect of SPS on colony counts was studied

by inoculating two SPS transport tubes containing

fresh human blood with recent isolates of either H influenzae type b (1.5 x 102 colony-forming units [cfu]/mL) on S. pneumoniae (3.0 x 102 cfu/mL).

The inoculated tubes were held at room

tempera-tune. At times zero, two hours, four hours and 6#{189}

hours, 0. 1 mL of blood was removed from each of the SPS transport tubes containing H influenzae type b on S pneumoniae and was plated onto

choc-olate agar or blood agar and incubated at 35#{176}Cfor

24 hours. Colonies were counted and the

colony-forming units pen milliliter were calculated for each

time point, in triplicate.

Statistics

x2

analysis with Yates’ connection was used to

compare discrete variables.

RESULTS

There were no significant changes in viable counts for either H influenzae on S pneumoniae

during the first four hours of incubation in the

0.35% SPS transport tube. After four hours,

how-even, the number of bacteria in both tubes began to

increase (Fig 1). At 6#{189}hours, H influenzae had

increased 4.5-fold from 1.5 x 102 cfu/mL to 7 x 102

cfu/mL. S pneumoniae increased as well from 3 x

102 cfu/mL to 9.8 x 102 cfu/mL, a threefold increase in number of organisms. On the basis of these results, only patient specimens that had a transport

time to the laboratory of less than four hours were

included in the study.

During the 18-month review period, a total of 40 patients were bactenemic with H influenzae, but

only 37 charts were available for review. Of these,

26

cases

met

the

criteria

of the

study.

Of the

pa-tients rejected, one was immunosuppressed, four

had no colony counts, three had been treated with

antibiotics prior to obtaining the blood culture, and three had unknown transport times. Of the 26 cases of H influenzae bacteremia that were evaluated, the median age of the patients was 10 months (mange 3

to 72 months); 17 were black and nine were whites;

eight were girls and 18 were boys. Diagnoses

in-cluded meningitis (18), epiglottitis (three), cellulitis

(three), osteomyelitis (one), and penicanditis (one).

The median colony count for all cases in this group was 130 cfu/mL (mange 12 to 2,000 cfu/mL) (Table).

In addition, 47 patients were bactenemic with S

pneumoniae and 43 charts were reviewed.

Twenty-four of the 43 cases were accepted for analysis. Of

the patients rejected, nine were immunosuppressed,

five had no colony counts or were treated with prior

antibiotics, two were bacteremic with multiple on-ganisms, and three had prolonged on unknown

transport times. The median age of the patients

accepted for study was 13 months (range 3 to 24

months); 16 were black and eight were whites; 16

were boys and eight were girls. Diagnoses included

meningitis (four), pneumonia (one), and occult

bac-teremia (19). The median colony count for all cases in this group was 8 cfu/mL (range 1 to 400 cfu/

mL).

There was a correlation between the magnitude

of H influenzae and S pneumoniae bacteremia and

, 6.0

E

3 4.0

-

i L I

3 4 5 6 7

Time in SPS (hours)

Fig I. Effect of time on colony-forming units per milli-liter (cfu/mL) ofSpneumoniae and H influenzae in blood, collected in 0.35% sodium polyanetholesulfonate (SPS). Each time point represents cfu/mL calculated from 0.1 mL of blood, removed from each SPS transport tube containing H influenzae type b or S pneumoniae. Sample points were performed in triplicate.

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www.aappublications.org/news

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0 00 0 0000 0000 00 0 000 0

TABLE. Correlation of Clinical Disease and Magnitude of Bacteremia

Organism and Diagnosis No. of Pa-tients Me-dian Age (mo)

Col ony-F orming Units/mL .

Median Range

<15 15-30 31-100 101-1,000 >1,000 H influenzae

Meningitis 18 10 150 (16_2,000)* 0 2 5 5 6

Epiglottitis 3 20 500 (28-500) 0 1 0 2 0

Orbital cellulitis 1 8 800 0 0 0 1 0

Osteomyelitis 1 7 1,200 0 0 0 0 1

Pericarditis 1 6 >100* 0 1 0 0 0

Facial cellulitis 1 20 20 0 1 0 0 0

Periorbital cellulitis 1 9 12 1 1 0 0 0

S pneumoniae

Occult bacteremia 19 13 6 (1-16) 18 1 0 0 0

Meningitis 4 7.5 150 (32_160)* 0 0 1 3 0

Pneumonia 1 5 400 0 0 0 1 0

* In these groups, one or more patients had colony counts recorded as >100.

Meningitis Occult bacteremia Other invasive

diseases *

Clinical disease

Fig 2. Ranges of colony-forming units per milliliter (cfu/mL) compared with clinical disease for H influenzae and S pneumoniae bacteremia. *Other invasive diseases include: epiglottitis, orbital cellulitis, osteomyelitis, pen-carditis, facial cellulitis, and pneumonia.

the type of disease (Fig 2). Patients who had men-ingitis were more likely to have a magnitude of bactenemia of 100 cfu/mL (P < .01,

x2

= 7.5).

Eighteen patients had meningitis with H influ-enzae. Eleven (61%) of the 18 had bactenemia with >100 cfu/mL. Six (86%) of seven patients with

>1000 cfu/mL had H influenzae meningitis; the seventh patient had osteomyelitis of the humerus also caused by H influenzae. Four patients had

meningitis caused by S pneumoniae. Of these, three

had colony counts of >100 cfu/mL and one had

bactenemia with 32 cfu/mL.

Eighteen patients (3 to 24 months of age) who were bacteremic with S pneumoniae had colony counts of <15 cfu/mL. Sixteen of these patients

had a fever and no focus of infection. Two of the remaining patients had otitis media and one had right maxillary sinusitis. Fifteen (83%) of this group had bactenemias with <10 cfu/mL, the me-dian colony count being 6 cfu/mL (mange 1 to 14 cfu/mL). Therefore, most patients with S pneu-moniae bactememia and colony counts of <15 cfu/ mL had occult bacteremia without a focus as op-posed to invasive disease. Most of these patients were treated as outpatients and released; none

re-turned to our hospital with invasive disease.

Four patients were excluded because they had received prior antibiotics. All had been treated as outpatients for otitis media; three had H influenzae

meningitis, and one had occult bacteremia with S pneumoniae. One of the patients with meningitis

had a bactenemia of 4 cfu/mL; in the three

remain-ing cases, agam media revealed no growth and the organisms were detected in the modified eugonic broth only.

DISCUSSION

More than 50 years ago, the magnitude of bac-teremia was correlated to fever in patients with

subacute bacterial endocamditis.9 Since then, quan-titative blood cultures have been used to determine the site of night-sided endocarditis, as well as for identifying intravenous catheter-related infection in both children and adults.3’4’6 Others have come-lated the magnitude of bactememia with the occur-mence of meningitis in infant mats and mortality in

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904 QUANTITATIVE BLOOD CULTURES

Several investigators have accumulated infor-mation about the magnitude of childhood

bactere-mia by directly plating samples of blood onto agan

growth media. Santosham and Moxon’2 found a wide range of bacterial counts whereas Durbin and co-workers” concluded that the direct plating method permitted earlier detection of bacteremia than standard broth culture methods. More

me-cently, LaScolea et al’4 reported a quantitative di-rect plating technique that permits early detection of bacteremia. Although determining the level of

bacteremia provides the clinician valuable infor-mation, methods of quantitation such as pour plates or direct agan inoculation are tedious and not

rou-tinely undertaken.

In 1978, Dorn and Smith’5 described a

blood-culturing system based on lysis of blood cells fol-lowed by centnifugation and inoculation of the sed-iment onto agar media. This system, in adult

pa-tients, increased the isolation rate, required less

time to obtain a pure culture, and enabled colony

counts to be determined routinely. However, a sys-tern that requires 6 to 7 mL of blood was not practical for use with pediatric patients.

Our investigation used a commercially available

lysis-direct plating blood culture system requiring less blood (marketed as the Isolator 1.5 Microbial

Tube). Carey’6 recently compared this system with

a radiometnic broth culture method and found it to

be an equally sensitive but more rapid method for

detection of bacteremic children with the added

benefit of routinely providing bacterial counts.

Likewise, Campos and Spainour8 reached the same

conclusion in a comparison of the Isolator 1.5 method with a conventional broth culture method.

In our report, the quantitative results that are

obtained from the Isolator 1.5 system were only

valid in patient specimens that had transport times

of less than four hours.

Our study confirms the results of Sullivan et a!.7

A greater magnitude of bacteremia correlated with

invasive disease. A significant number of patients with colony counts of 100 cfu/mL had meningitis (P < .01,

x2

= 7.5).

Furthermore, if S pneumoniae bacteremia is

con-sidered, colony counts of <15 cfu/mL were

associ-ated with occult bacteremia rather than invasive

disease. All of the 26 patients with H influenzae bacteremia in our study had colony counts of >10 cfu/mL and all had invasive disease. However, more than half of the patients with S pneumoniae

bac-teremia had colony counts <10 cfu/mL and all had

occult bacteremia without invasive disease.

SUMMARY

In summary, quantitative blood culture results

obtained with a commercially available method

confirmed that the magnitude of bacteremia come-lated with invasive disease. Cleanly, clinical

judg-ment remains the most important aspect in the evaluation of an ill child with a positive blood culture. However, knowledge of the magnitude of

bacteremia at the time of evaluation may be helpful

in identifying which children are likely to have

invasive disease.

ACKNOWLEDGMENT

The authors wish to dedicate this work to the memory of Joseph Ritter, MD, a gifted pediatrician and teacher.

REFERENCES

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2. Bratton L, Teele DW, Klein JO: Outcome of unsuspected pneumococcemia in children not initially admitted to the hospital. J Pediatr 1977;90:703-706

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1975;82:746-750

4. Wing EJ, Norden CW, Shadduck RW, et al: Use of quanti-tative bacteriologic techniques to diagnose catheter-related sepsis. Arch Intern Med 1979;139:482-483

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7. Sullivan TD, LaScolea U, Neter E: Relationship between the magnitude of bacteremia in children and the clinical disease. Pediatrics 1982;69:699-702

8. Campos JM, Spainour JR: Comparison of the Isolator 1.5 Microbial Tube with a conventional blood culture system for detection of bacteremia in children. Diagn Microbiol

Infect Dis 1985;3:167-174

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1932;50:61-68

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11. Kluge RM, DuPont HL: Factors affecting mortality of pa-tients with bacteremia. Surg Gynecol Obstet 1973;137:267-269

12. Santosham M, Moxon ER: Detection and quantitation of bacteremia in childhood. J Pediatr 1977;91:719-721

13. Durbin WA, Szymczak EG, Goldmann DA: Quantitative blood cultures in childhood bacteremia. J Pediatr 1978; 92:778-780

14. LaScolea U, Dryja D, Sullivan TD, et al: Diagnosis of bacteremia in children by quantitative direct plating and a radiometric procedure. J Clin Microbiol 1981;13:478-482 15. Dorn GL, Smith, K: New centrifugation blood culture device.

,J Clin Microbiol 1978;7:52-54

16. Carey RB: Clinical comparison of the Isolator 1.5 Microbial Tube and the BACTEC radiometric system for detection of bacteremia in children. J Clin Microbiol 1984;19:634-638 17. Todd JK: Childhood infections: Diagnostic value of

periph-eral white blood cell and differential cell counts. Am J Dis

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18. Marshall R, Teele DW, Klein JO: Unsuspected bacteremia due to Haemophilus influenzae: Outcome in children not initially admitted to hospital. J Pediatr 1979;95:690-695

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1985;76;901

Pediatrics

Louis M. Bell, Gershon Alpert, Joseph M. Campos and Stanley A. Plotkin

Bacteremia

Streptococcus pneumoniae

Haemophilus influenzae or