Background
The burden of disease attributable to bloodstream infections (BSIs) is insufficiently studied in Africa where few healthcare facilities have the capacity to identify invasive diseases. BSIs are increasingly recognized as an important cause of hospitalisation and mortality in the region, and studies indicate that the prevalence of and mortality in bacteraemia parallels or exceeds that of malaria [
1]-[
4].
Due to symptom overlap bacteraemia is frequently misclassified as malaria and malaria is often over-diagnosed [
3],[
5]-[
7]. Malaria may occur concomitantly with and has been indicated as a risk factor for bacteraemia [
3],[
8],[
9]. Yet, the World Health Organization’s (WHO) guidelines for management of acute paediatric illness primarily focus on malaria and do not include BSIs in the diagnostic algorithm for fever in the absence of general danger signs [
10],[
11].
It would be valuable to identify children at risk of bacteraemia, where blood cultures should be performed. However, studies have demonstrated poor predictive values of clinical signs to detect bacteraemia [
2],[
4]. In a study from Kenya where blood cultures were performed on all children attending a hospital outpatient department, the sensitivity of fever to detect BSIs was only 73% [
2].
There are reported differences in the aetiology of BSIs between African regions that may limit the effectiveness of treatment recommendations and immunization programmes [
1],[
12]-[
16] and antibiotic resistance is an increasingly recognized problem [
3],[
4],[
17]. There is to our knowledge no study that have investigated the prevalence of bacterial blood stream infections in Guinea-Bissau and many previous studies of bacteraemia in Africa have been limited to febrile or hospitalized patients, which may limit the generalizability of the findings [
12]-[
15].
The objective of this study was to investigate the prevalence, species distribution and antibiotic susceptibility of BSIs in consecutive children presenting with systemic infection signs to a paediatric emergency department in Guinea-Bissau. We also aimed to investigate the predictive capacity of clinical parameters to identify BSIs.
Results
General characteristics of children in the study
Clinical data and blood samples were obtained from 414 children that fulfilled the inclusion criteria. During two weeks in July 2010, 39 consecutive children were excluded due to lack of freezing media for conservation of bacteria. Three further children were excluded since their isolates were not frozen according to protocol. Hence, 372 children were included in the analyses of which 48% (180) were admitted to hospital. Categorized by the inclusion criteria, 46% (172/372) had fever and tachycardia, 45% (167/372) had only tachycardia while 9% (33/372) had only fever. The mean age was 1.7 years and 44% (163/372) were females. Of the children from whom we obtained a report 2% (5/325) had been hospitalized during the month prior to enrolment.
Bacteraemia prevalence, molecular epidemiology and antibiotic susceptibility
Probable pathogens were isolated from the blood of 12% (46/372) of the children (Table
1).
S. aureus accounted for 54% (26/48) of the isolates while non-typhoidal
Salmonella (NTS) accounted for 10% (5/48),
S. pneumoniae for 8% (4/48) and
Salmonella Typhi for 6% (3/48). Two children had polymicrobial bacteraemia, one with
S. aureus and
Streptococcus pyogenes, and one with
S. aureus and
S. pneumoniae. Three out of five children under 60 days of age were infected with Enterobacteriaceae. Gram-negative bacteria caused 55% (6/11) of the BSIs in children aged <1 year, compared to 24% (9/37) in children aged 1–5 years (p = 0.07). Among hospitalized children 14% (25/180) had bacteraemia, compared to 11% (21/192) among outpatients (p = 0.40).
S. aureus may be part of the skin flora and hence it is possible that some of these findings were due to contamination. The corresponding figure for non-Staphylococcal bacteraemia was 8% (14/180) in the hospitalized group compared to 4% (8/191) of the outpatients (p = 0.14). There was no mortality among hospitalized children with Staphylococcal bacteremia (0/11), compared with 14% (2/14) in children with non-Staphylococcal bacteraemia (p = 0.49). The two fatalities in the bacteraemia group were diagnosed with neonatal sepsis and the isolated pathogens were
Escherichia coli and
Enterobacter cloacae.
Table 1
Blood culture findings from children in Guinea-Bissau in relation to age and hospitalization
Gram-positive | 33 (9) | 2 (7) | 3 (4) | 12 (9) | 16 (13) | 18 (5) | 15 (4) |
Staphylococcus aureus
| 26 (7) | 2 (7) | 3 (4) | 10 (7) | 11 (9) | 13 (3) | 13 (3) |
Streptococcus pneumoniae
| 4 (1) |
-
|
-
|
-
| 4 (3) | 4 (1) |
-
|
Streptococcus pyogenes
| 1 (0) |
-
|
-
|
-
| 1 (1) | 1 (0) |
-
|
Enterococcus faecalis
| 2 (1) |
-
|
-
| 2 (1) |
-
|
-
| 2 (1) |
Gram-negative | 15 (4) | 3 (11) | 3 (4) | 4 (3) | 5 (4) | 9 (2) | 6 (2) |
Klebsiella pneumoniae
| 3 (1) |
-
| 2 (2) |
-
| 1 (1) | 2 (1) | 1 (0) |
Escherichia coli
| 2 (1) | 2 (7) |
-
|
-
|
-
| 1 (0) | 1 (0) |
Salmonella typhi
| 3 (1) |
-
|
-
|
-
| 3 (2) | 2 (1) | 1 (0) |
Non-typhoidal Salmonella
| 5 (1) |
-
| 1 (1) | 3 (2) | 1 (1) | 2 (1) | 3 (1) |
Enterobacter cloacae
| 2 (1) | 1 (4) |
-
| 1 (1) |
-
| 2 (1) |
-
|
Total children positive | 46 (12) | 5 (23) | 6 (8) | 16 (14) | 21 (20) | 27 (8) | 21 (6) |
Probable contaminants were isolated in 31% (117/372) of the samples. Prevalent contaminants were coagulase-negative Staphylococci (n = 87), Sphingomonas (n = 4), Bacillus (n = 4), and viridans group Streptococci (n = 3). None out of four Acinetobacter was confirmed as Acinetobacter baumanii in MALDI-TOF analysis and they were hence regarded as contaminants. To assess if detection of S. aureus was associated with contaminants, the prevalence of contaminants was evaluated among patients with S. aureus, patients with other probable pathogens, and patients with no growth of probable pathogens. There was no difference in the frequency of contaminants, 5/26 (19%), 6/20 (30%), and 106/326 (33%), respectively (p = 0.39).
The prevalence of S. aureus was similar in children admitted to hospital (7%, 13/180) and outpatients (7%, 13/192) (p = 0.87). Frequent spa types were t084 (n = 7), t355 (n = 5), t127 (n = 2), t1476 (n = 2) and t4690 (n = 2). The spa types t008, t024, t314, t491, t571, t760, t939 and t1458 were found in one isolate, each. In total, 38% (10/26) of the isolates produced PVL. The four S. pneumoniae isolates belonged to serotypes 6B (n = 2), 5 (n = 1) and 23F (n = 1).
Antibiotic resistance was uncommon among Gram-positive isolates (Table
2). All
S. aureus isolates remained susceptible to methicillin. Both
Enterococcus faecalis isolates were susceptible to ampicillin and vancomycin. One out of four
S. pneumoniae isolates had a minimum inhibitory concentration for penicillin of 0.5 mg/l. Among Gram-negative isolates, two out of three
K. pneumoniae isolates produced ESBLs (one TEM and the other SHV type). Similar to one of the two
E. cloacae isolates, the ESBL-producing
K. pneumoniae isolates were resistant to all tested antibiotics but imipenem. One of the patients with ESBL-producing
K. pneumoniae had been hospitalized within the last month. The
Salmonella had uniformly low minimum inhibitory concentrations to ciprofloxacin and azithromycin, indicating susceptibility.
Table 2
Antibiotic resistance in bacteraemia isolates from children in Guinea-Bissau
1
Gram-positive | 33 | | | | | | | | | | | | | |
Staphylococcus aureus
| 26 | - | - | - | 0 | 0 | 2 (8) | - | 0 | 0 | 0 | 0 | 0 | 0 |
Streptococcus pneumoniae
| 4 | 0 | - | 1 (25)2
| - | 0 | 3 (75) | - | 0 | - | 0 | 4 (100) | - | 3 (75) |
Streptococcus pyogenes
| 1 | 0 | - | 0 | 0 | 0 | 0 | - | 0 | - | 0 | 0 | - | 1 (100) |
Enterococcus faecalis
3
| 2 | 0 | 0 | - | - | - | - | - | - | 0 | - | 2 (100)4
| - | 1 (50) |
Gram-negative | 15 | | | | | | | | | | | | | |
Escherichia coli
| 2 | 1 (50) | 0 | - | - | 0 | - | - | - | 0 | - | 1 (50) | 0 | 0 |
Klebsiella pneumoniae
| 3 | 3 (100) | 2 (67) | - | - | 2 (67) | - | - | - | 2 (67) | - | 2 (67) | 2 (67) | 1 (33) |
Salmonella Typhi | 3 | 0 | 0 | - | - | 0 | - | 0 | - | 0 | - | 1 (33) | 0 | 1 (33) |
Non-typhoidal Salmonella
| 5 | 2 (40) | 0 | - | - | 0 | - | 0 | - | 0 | - | 2 (40) | 0 | 2 (40) |
Enterobacter cloacae
5
| 2 | - | 2 (100) | - | - | 1 (50) | - | - | - | 1 (50) | - | 1 (50) | 1 (50) | 2 (100) |
Predictors of bacteraemia
In analysis of all 46 children with growth of a pathogen in their blood sample, no difference in age between children with and without bacteraemia was observed (p = 0.54). The bacteraemia prevalence was 10% (16/163) among females and 14% (30/209) among males (p = 0.19). Fever had 54% (25/46) sensitivity to detect bacteraemia and its positive predictive value (PPV) was only 12% (Table
3). Nearly all bacteraemic children (96%, 44/46) had tachycardia. However, the specificity of tachycardia as defined by our inclusion criteria was only 10% and the PPV was 13%. At higher tachycardia cut-offs, the specificity increased at the expense of reduced sensitivity while the PPV remained low. The other investigated clinical parameters were also poorly predictive of bacteraemia (Table
3).
Table 3
Clinical predictors of bacteraemia in children in Guinea-Bissau with fever and/or tachycardia
Temperature | | | | | | | | |
≥38° | - | 25 | 181 | 0.88 | 54 | 44 | 12 | 87 |
≥39° | - | 9 | 45 | 0.30 | 20 | 86 | 17 | 88 |
≥40° | - | 3 | 6 | 0.09 | 7 | 98 | 33 | 88 |
Tachycardia4
| | | | | | | | |
≥120 ≥ 160 | - | 44 | 295 | 0.25 | 96 | 10 | 13 | 94 |
≥130 ≥ 170 | - | 39 | 265 | 0.57 | 85 | 19 | 13 | 90 |
≥140 ≥ 180 | - | 23 | 167 | 0.88 | 50 | 49 | 12 | 87 |
Saturation5
| 4 | 3 | 10 | 0.20 | 7 | 97 | 23 | 88 |
Respiratory distress6
| 5 | 20 | 218 | <0.01 | 44 | 32 | 8 | 81 |
Anaemia7
| - | 7 | 49 | 0.97 | 15 | 85 | 13 | 88 |
Convulsions | 10 | 1 | 11 | 1.00 | 2 | 97 | 8 | 87 |
Lung crepitation | - | 13 | 130 | 0.13 | 28 | 60 | 9 | 86 |
Diarrhoea | - | 16 | 106 | 0.76 | 35 | 67 | 13 | 88 |
Reduced consciousness8
| 17 | 4 | 29 | 1.00 | 9 | 91 | 12 | 89 |
Vomiting | - | 15 | 97 | 0.69 | 33 | 70 | 13 | 88 |
MUAC9
| | | | | | | | |
<135 mm | - | 9 | 78 | 0.51 | 23 | 72 | 10 | 87 |
<125 mm | - | 3 | 35 | 0.60 | 8 | 87 | 8 | 87 |
<115 mm | - | 0 | 12 | 0.37 | 0 | 96 | 0 | 87 |
Leukocyte count 10
| | | | | | | | |
>11*109/L | 85 | 20 | 117 | 0.41 | 54 | 53 | 15 | 89 |
>20*109/L | 85 | 3 | 12 | 0.42 | 8 | 95 | 20 | 88 |
>11*109/L and/or fever | 85 | 29 | 183 | 0.56 | 78 | 27 | 14 | 89 |
Malaria positive | 52 | 4 | 13 | 0.25 | 10 | 95 | 24 | 88 |
Similarly, in a subgroup analysis of the 22 non-Staphylococcal bacteraemia cases we detected no difference in the prevalence of bacteraemia with regard to age (p = 0.32), sex (p = 0.47) nor the presence of fever (p = 0.21). There was weak evidence that fever of 39°C (p = 0.08) and a leukocyte particle concentration of ≥20 ×109/l (p = 0.07) was more common in non-Staphylococcal bacteraemia while PPV was low at 11% and 20%, respectively.
We further assessed predictors of bacteraemia in the 167 non-febrile children (Table
4). All parameters had a poor trade-off between sensitivity and specificity. Fifteen non-febrile children had
S. aureus infection, compared to 11 febrile children (p = 0.17). Also,
E. faecalis (2)
, ESBL-producing
K. pneumoniae (2)
, E. coli (1), NTS (1) and
S. pneumoniae (1) were isolated from non-febrile children.
Table 4
Clinical predictors of bacteraemia in non-febrile children in Guinea-Bissau
Tachycardia4
| | | | | | | | |
≥120 ≥ 160 | - | 21 | 145 | N/A | 100 | 0 | 13 | N/A |
≥130 ≥ 170 | - | 17 | 132 | 0.15 | 81 | 9 | 11 | 76 |
≥140 ≥ 180 | - | 10 | 76 | 0.68 | 48 | 48 | 12 | 86 |
Saturation5
| 1 | 1 | 3 | 0.42 | 5 | 98 | 25 | 88 |
Respiratory distress6
| 2 | 7 | 91 | 0.01 | 33 | 36 | 7 | 79 |
Anaemia7
| - | 2 | 18 | 1.00 | 10 | 87 | 10 | 88 |
Convulsions | 5 | 1 | 0 | 0.13 | 5 | 100 | 100 | 88 |
Lung crepitation | - | 6 | 55 | 0.41 | 29 | 62 | 10 | 86 |
Diarrhoea | - | 10 | 55 | 0.40 | 48 | 62 | 15 | 89 |
Reduced consciousness8
| 10 | 1 | 3 | 0.43 | 5 | 98 | 25 | 88 |
Vomiting | - | 7 | 46 | 0.88 | 33 | 68 | 13 | 88 |
MUAC9
| | | | | | | | |
<135 mm | - | 5 | 36 | 1.00 | 28 | 72 | 12 | 88 |
<125 mm | - | 2 | 15 | 1.00 | 11 | 88 | 12 | 88 |
<115 mm | - | 0 | 5 | 1.00 | 0 | 96 | 0 | 87 |
Leukocyte count10
| | | | | | | | |
>11*109/L | 44 | 8 | 39 | 0.41 | 50 | 63 | 17 | 89 |
>20*109/L | 44 | 1 | 5 | 0.58 | 6 | 95 | 17 | 87 |
Malaria positive | 23 | 1 | 1 | 0.24 | 6 | 99 | 50 | 88 |
Clinical management
Empirical antibiotic therapy with adequate coverage of the isolated pathogen was prescribed to 61% (28/46) of the children. The treatment regimen was selected individually by the paediatrician, commonly ampicillin supplemented with a single dose of gentamicin. Out of the bacteraemic children, 22% (10/46) did not receive any antibiotic. Among the 18 children that received no or inadequate antibiotic treatment S. aureus caused 12 (67%) episodes and S. Typhi two, while E. faecalis, NTS, E. cloacae and K. pneumoniae caused one episode each. There was no difference in bacteraemia prevalence with regards to clinical diagnosis. Although the bacteraemia prevalence was similar in children with and without a clinical malaria diagnosis (p = 0.67), malaria-diagnosed children were less likely to receive antibiotic treatment (p = 0.002). Only 18% (5/28) of the malaria-diagnosed children with bacteraemia received adequate antibiotic treatment.
Malaria was diagnosed with microscopy in 13/227 (6%) children for which we obtained a slide and with PCR in 14/311 (5%) children for which we obtained a filter paper. In total, malaria was confirmed with either method in 5% (17/320) of the investigated children. In comparison, 64% (237/372) received a clinical malaria diagnosis by the attending physician. The bacteraemia prevalence in children with and without laboratory-confirmed malaria was 24% (4/17) and 12% (37/303), respectively (p = 0.25).
Discussion
This is to our knowledge the first study on the prevalence of bacteraemia in children in Guinea-Bissau. The prevalence of bacteraemia was 12% in this paediatric population and malaria was over-diagnosed (found in 5% using laboratory tests, but in 64% using clinical criteria). As many as 46% of the children with bacteraemia did not have fever at presentation to the hospital and no clinical parameters predicted bacteraemia with high sensitivity and specificity.
The prevalence of bacteraemia in previous studies from Africa has ranged from 2% to 46% [
1]-[
4],[
13],[
14],[
28], however most of these studies either did not use reproducible enrolment criteria [
14],[
28] or were focused on inpatients [
1],[
4],[
17]. We observed a poor sensitivity of 54% for fever to detect bacteraemia. Excluding
S. aureus bacteraemia the sensitivity improved slightly to 68% which is comparable to the 73% reported in a benchmark study by Brent
et al. where children at a Kenyan outpatient department were blood cultured regardless of clinical presentation [
2]. This implies that studies limited to febrile children may underestimate bacteraemia incidence and that the clinical threshold to perform blood cultures in children in Africa presenting to hospitals needs to be low. Similarly to the above mentioned study in Kenya, we found that tachycardia had higher sensitivity to detect bacteraemia but its specificity was poor [
2]. In comparison, Nadjm
et al. reported a 67% sensitivity of the current WHO criteria to detect invasive bacterial disease in a Tanzanian area of high malaria transmission [
4]. Although our record of clinical parameters reflects the rudimentary conditions of paediatric care in Guinea-Bissau, neither clinical examinations nor prescribed treatments were harmonized with WHO recommendations. Therefore we could not evaluate the capacity of WHO criteria to predict bacteraemia. This may limit the generalizability of our analyses of clinical parameters in relation to bacteraemia. Future studies that assess the predictive capacity of clinical parameters and other screening tests need to assess these in relation to current WHO guidelines. Also, the epidemiological design needs to be carefully assessed to avoid selection bias both with regards to pathogen detection as well as in evaluation of screening algorithms.
More than half (54%) of the bacterial findings were
S. aureus, which predominated also in studies from Malawi [
12] and Nigeria [
14],[
28]. Several other studies of bacteraemia in Africa, including a systematic review, have reported
S. pneumoniae [
1]-[
3] and NTS [
4],[
13],[
17] as the most prevalent species (in our study the second and third most prevalent). In spite of similar findings in two previous studies from the region [
14],[
28]
S. aureus may be part of the normal skin flora, and hence our finding of a high prevalence of
S. aureus bacteraemia needs to be interpreted with caution. To investigate the pathogenicity of the
S. aureus isolates they were subjected to
spa typing. PVL-production has been reported to impair neutrophil function [
29] and is associated with severe necrotizing pneumonia [
30] and the high prevalence of PVL-production in our study is a concern. Furthermore, the
spa type distribution closely resembled that of clinical isolates in a study from Gabon [
31] and unlike studies from industrialized countries, types 084 and 355 (corresponding to MLST types 15 and 152) were the most prevalent.
The contamination frequency in previous studies of bacteremia in Africa has varied from 9-34% [
1],[
2],[
8],[
9],[
12]-[
15],[
32], however some studies did not report the prevalence of contamination [
4],[
17],[
28] and there is variability with regards to what species are considered pathogens. Viridans group
Streptococci,
Citrobacter,
Aeromonas,
Cryptococcus and Morganella are bacterial geni that may be considered pathogenic [
12]-[
14] but that we regarded as contaminants. Unlike previous studies [
1],[
2], we used MALDI-TOF to differentiate the often pathogenic
Acinetobacter baumanii from low-pathogenic species
. The wide range in prevalence of both bacteremia and contamination can thus partly be explained by differences in study design and reporting customs. In Guinea-Bissau blood cultures had not been routinely performed at the hospital or analyzed in the laboratory prior to our study, and the high frequency of contaminants implies that our protocol was not sufficient to avoid contamination. In future studies of bacteraemia in Guinea-Bissau, thorough training of hospital and laboratory staff is warranted. Also, since established as well as several new molecular methods are used in identifying microorganisms, epidemiological designs such as case–control studies are needed to obtain results on the clinical importance of microbe detection [
33],[
34].
Previous studies of invasive bacterial isolates in Africa have found varying prevalence of antibiotic resistance [
3],[
4],[
12],[
17]. In our study the overall antibiotic resistance was low, however two out of three
K. pneumoniae produced ESBLs. We have recently reported that 33% of 408 children in Guinea-Bissau (from whom the 372 children in the present study originate) carried ESBL-producing Enterobacteriaceae in their intestines [
35] and one of the two children with ESBL-producing
K. pneumoniae bacteraemia in the present study was also a carrier. The presence of ESBLs also in bacteraemia is worrying considering the limited access to antibiotic drugs and the increased mortality in BSIs with ESBL-producing species [
17]. The WHO’s empirical treatment recommendations for septicaemia is chloramphenicol and/or benzylpenicillin [
10],[
11]. The high prevalence of chloramphenicol-susceptible
S. aureus in our material indicates this as an adequate treatment combination for suspected bacteraemia in Guinea-Bissau. Of the bacteraemia isolates in our study, 83% (40/48) were susceptible to at least one of these drugs.
There are limitations to our study. As discussed above the high prevalence of S. aureus may reflect contamination from the skin flora rather than infection. Interestingly however, removing S. aureus from the analysis did not alter our main finding that fever is an insufficient marker of bacteraemia. Since we did not measure CD4 counts or test for tuberculosis and other underlying conditions, we could not evaluate the influence of these parameters on the occurrence of bacteraemia. The study period coincided with the rainy season and was conducted at a national referral centre, which may have influenced the distribution of pathogens. Standardized laboratory testing with automated blood cultures and leukocyte counts would have strengthened the internal validity of our study. Finally we did not obtain follow-up data on outpatients, which hampered our analyses about mortality.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
PN and BHN conceived and designed the study. CM was the study clinician. PN and AR arranged the study infrastructure in Guinea-Bissau. PN and JI supervised the sampling and subsequent laboratory analyses on site. CG, WX and JI performed laboratory analyses in Sweden. PN and JI performed the statistical analyses. CG, PN and JI prepared the manuscript. All authors contributed intellectually to the interpretation of the study findings. All authors read and approved the final manuscript.