Despite great improvements in infant mortality rates in recent years, the mortality of young infants, particularly neonates, remains high in most developing countries. In 1995, 47 countries were believed to have neonatal mortality rates in excess of 40/1000 live births. It is estimated currently that ∼5 million neonates die in developing countries each year.1, 2 Outside the first month of life most other infant deaths occur during the second and third months of life. By contrast, in industrialized countries, neonatal mortality rates are ∼5/1000 live births and most of those deaths are premature infants in intensive care units, a group not always included in developing country data. Infection is estimated to cause 30 to 40% of neonatal deaths in developing countries.2
There are several prevailing views of the etiology of neonatal infections in developing countries. Published series suggest that Staphylococcus aureus and Klebsiella spp. are the main pathogens,2, 3 although hospital series include many hospital-acquired infections, and the bacteriologic techniques in many places would not permit the growth of fastidious organisms like Streptococcus pneumoniae and Haemophilus influenzae. Before this study many believed that S. pneumoniae and H. influenzae were important pathogens in young infants in developing countries, although there were few published reports to support this view. In industrialized countries Escherichia coli and Streptococcus agalactiae (group B Streptococcus) are the most important neonatal pathogens, and it is unclear what role these organisms have in developing countries.
On the basis of available data WHO and other authorities have recommended that serious infections in very young infants in developing countries should be treated with penicillin and gentamicin initially.4 In practice many different combinations are used based on local interpretations of existing data; most regimens include a penicillin and an aminoglycoside. Some use chloramphenicol and, where they are available, third generation cephalosporins, particularly cefotaxime, are used.
Although treatment of established cases is important, control of infections in young infants ultimately rests on prevention.2 Although efforts are under way in many developing countries to improve perinatal care, attention is also being given to maternal immunization as a means of protecting young infants from infection with passively acquired maternal antibody. This has been used with great success to control neonatal tetanus in developing countries5 and has recently been tried to prevent neonatal infections caused by encapsulated bacteria such as S. agalactiae, S. pneumoniae and H. influenzae.6-9 Developments in this area provide another valid reason for investigating the etiology of infections in young infants in developing countries.
To address this question a multicenter project was set up to examine the etiology and clinical signs of serious infections in infants younger than 90 days of age in developing countries. The objective of the studies described in this paper was to determine the bacterial and viral causes of serious infections in young infants in developing countries. Only the bacteriology results are reported in this paper. Investigations that were not performed uniformly in all sites are reported in the site-specific papers in this supplement.10-13 These include all virology, urine culture, urine antibacterial activity and antimicrobial sensitivity results of isolates.
The four sites, Ethiopia, the Gambia, Papua New Guinea and The Philippines, were chosen to represent a range of developing country settings with high neonatal mortality rates.1 The age criterion for infants to be enrolled in the study was set as 90 days or less. In industrialized countries the transitional age when traditional neonatal pathogens give way to infant pathogens is thought to be at ∼4 to 6 weeks. Because there are no data on which to base such an estimate for developing countries, a broader age range was chosen which includes the period of highest mortality.
The methods are described in detail in an accompanying paper.14 Briefly infants younger than 90 days who presented ill to one of the study institutions underwent a formalized triage process. Those with any symptom or sign of infection who did not meet the exclusion criteria14 were enrolled and subjected to a standardized history and examination by a physician (all sites) or pediatric nurse (Papua New Guinea only). Those meeting predefined criteria suggestive of infection were investigated by blood culture, urine culture, hematologic examination, blood film for malaria, nasopharyngeal aspirate (where virology was performed) and lumbar puncture (where clinically indicated). Before the study laboratory procedures were optimized and standardized between sites. These are described in detail in an accompanying paper.14
Blood cultures were considered positive if a definite pathogen (e.g. S. pneumoniae, H. influenzae, Streptococcus pyogenes, S. agalactiae, Salmonella spp.) was isolated from either bottle. For organisms that could be either pathogens or contaminants (e.g. E. coli, Klebsiella pneumoniae, Staphylococcus aureus, Enterococcus faecalis (group D) a blood culture was considered positive if the organism grew from both (or one of one) bottles within 48 h of inoculation. Cultures that did not fulfil these criteria, or cultures of known contaminants such as coagulase-negative staphylococci, Micrococcus spp. and Bacillus spp. were regarded as negative. This strategy was supported by an analysis at the end of the study showing the mortality in those with contaminants to be the same as that in the culture-negative group. Nasopharyngeal swabs were cultured for Bordetella pertussis.
Ethical approval for the study was obtained from the WHO Secretariat Committee for Research in Human Subjects and from the relevant local ethical committee in each of the sites involved. Because no investigations were performed that would not have been included in the good clinical care of the infants with suspected infection, only verbal consent was obtained from parents or care givers before the enrollment of an infant into the study.
In the 4 sites a total of 8418 young infants were triaged, of whom 4552 satisfied the criteria for enrollment in the study and underwent a full history, physical examination and pulse oximetry. Of these, 2398 with clinical signs suggestive of infection were investigated, including blood culture (2452), lumbar puncture (507) and chest radiograph (1868), and this comprised the study sample for the analyses presented in this paper. Included in the above figures are 175 infants without signs of serious infection who were randomly chosen for investigation including blood culture. Not all investigated infants had blood cultures performed, usually because of technical difficulties. Overall 247 infants (5% of those enrolled) died, including 19 who were judged likely to have died because of being taken from the hospital by their families, against medical advice and in a serious condition. There were some important differences between the sites with respect to the study populations. The largest number of infants was enrolled in Papua New Guinea, but in that site the infants enrolled were less ill, fewer were admitted and few died (Table 1).14
Table 2 summarizes the 167 blood culture isolates that were obtained from 2452 infants who underwent this investigation. Of the isolates 102 (61%) were Gram-positive organisms, 96 of which were either S. pneumoniae, Staphylococcus aureus or S. pyogenes. The most frequent Gram-negative isolates were E. coli and Salmonella spp. Three infants grew both Staphylococcus aureus and S. pyogenes in the blood. One infant each grew: Staphylococcus aureus in the blood and S. pyogenes in the CSF; Staphylococcus aureus and Streptococcus spp. (group G) in the blood; Staphylococcus aureus and Enterococcus faecalis in the blood. One infant grew H. influenzae in the blood and had a positive nasopharyngeal culture for B. pertussis. In addition 3 other infants (2 in Ethiopia and 1 in PNG) grew B. pertussis from NP culture.
During the first month of life Staphylococcus aureus (19, 23%), S. pyogenes (17 isolates, 20%) and E. coli (15, 18%) were the most common organisms isolated from blood. S. pneumoniae was the most common organism in both the second and third month of life, accounting for 30% of blood isolates, with meningitis accounting for 47% (17 of 36) of all episodes of invasive pneumococcal disease in study infants.
Four of the positive blood cultures occurred amongst the 175 infants in the systematic sample of infants without signs of systemic infection. Three of the 4 isolates (all Staphylococcus aureus) were from The Gambia where an epidemic of scabies may have contributed to the high rate of isolation of this organism.
Table 3 lists the CSF isolates by age. Of the children with a positive CSF culture 28 (68%) also had a positive blood culture. These are shown in parentheses with the organisms listed in Table 3. In children younger than 7 days of age Gram-negative enteric organisms were the main causes of meningitis. During the remainder of the first month Gram-negative organisms continued to be important, but the pneumococcus was equally important. Thereafter during the second and third months of life the pneumococcus was the main cause of meningitis, accounting for 12 of 24 (50%) of cases.
Although the general patterns of organisms isolated were similar, there were some differences between the sites. Seventeen of 34 (50%) isolates of Staphylococcus aureus came from The Gambia, 10 of the 19 (52%) isolates of E. coli came from Ethiopia, and 13 of 29 (45%) S. pyogenes isolates came from Papua New Guinea.
Table 4 summarizes the clinical characteristics of infants with blood cultures that grew S. pneumoniae, S. pyogenes, Salmonella spp., Staphylococcus aureus, E. coli and H. influenzae compared with the clinical characteristics of all infants with positive blood cultures and all infants who were investigated but had negative blood cultures. The infants with S. pneumoniae infection were more likely to have fever and fast breathing. Most of the infants with S. pneumoniae infection who died had meningitis; the case fatality rate among infants with proven pneumococcal meningitis was 53%. The infants with S. pyogenes infection were also likely to be febrile with fast breathing; this group had a low case fatality rate, not significantly different to the infants who were culture-negative. Infants with Salmonella spp. infection were significantly more likely to be underweight. Infants with E. coli infection were likely to be young and underweight; the mortality in this group was high (53%).
Of the blood and CSF isolates of S. pneumoniae, 34 were serotyped at a reference laboratory. The serotypes were type 5 (9 cases), type 2 (9 cases), type 7F (2 cases) and 1 each of types 1, 9L, 9V, 12F, 18F, 19F, 19A, 23F, 27 and 33F. Four isolates were not factor-typed: one each of serogroups 6, 7, 10 and 12. Only 20 of 34 (59%) isolates of S. pneumoniae were of serogroups included in the current 11-valent pneumococcal conjugate vaccine. Of the 9 infants with type 2 infection 8 had meningitis. There were no clear differences between the sites with respect to serotype distribution. In particular type 2 isolates were obtained from all 4 sites.
At the time when this study was carried out published studies from developing countries suggested that Klebsiella spp. and Staphylococcus aureus were the most important neonatal pathogens in developing countries.2, 3 The main causes of serious infections in young infants in the four sites of this study were the classical Gram-positive primary pathogens, Staphylococcus aureus, S. pneumoniae and S. pyogenes. E. coli was the most important Gram-negative organism, followed by a wide range of enteric pathogens causing disease. Although this study confirmed the importance of Staphylococcus aureus, that organism was responsible for only 20% of blood isolates and one CSF isolate, whereas Klebsiella spp. was responsible for only 3% of blood isolates and no CSF isolates.
S. pneumoniae was a major pathogen in all age groups studied, particularly after the first week of life. It was also the most important cause of meningitis; in children older than 1 week of age it accounted for 50% of all cases. Surprisingly the outcome of pneumococcal meningitis in this group was not worse than that found with pneumococcal meningitis in older children in developing countries, with about one-half of all cases dying.15 Among the serotypes of S. pneumoniae found, the importance of type 2, which represented 26% of isolates serotyped, was a surprising finding. It is not included in any of the current generation of pneumococcal conjugate vaccines. Type 2 is a serotype rarely reported in previous series; eight of the nine type 2 isolates in our study were from meningitis cases. An earlier study from PNG found type 2 to be responsible for 11% of CSF isolates.16 It appears that this serotype has a predilection for causing meningitis in the very young. This observation is consistent with the view that there are important epidemiologic differences between pneumococcal serotypes, in both age and disease distribution.17 The serotypes usually associated with pediatric carriage and disease and most frequently associated with penicillin resistance (serotypes 6A, 6B, 14, 19F and 23F) were isolated infrequently in this study. The pneumococcus must be considered in any case of serious infection in a young infant in a developing country, particularly if signs of meningitis are present. The findings of this study support the continued investigation of maternal immunization with 23-valent pneumococcal polysaccharide vaccine as a strategy for controlling neonatal infections.7, 8
Earlier this century in industrialized countries, S. pyogenes was an important cause of puerperal sepsis. In the US during the 1930s and 1940s, S. pyogenes was a leading cause of neonatal sepsis and meningitis.18 In our study 10% of the S. pyogenes isolates were detected in the first week of life and thus could have been attributed to maternal infection. 42% of umbilical swabs in Papua New Guinea showed S. pyogenes carriage.12 Recent reports from industrialized countries suggest that there has been a global resurgence of S. pyogenes disease.19 It is not clear whether this is truly global, or whether S. pyogenes has always been an important neonatal pathogen in developing countries, and its resurgence is limited to industrialized countries. Our study does not help to clarify this.
The virtual absence of S. agalactiae (group B Streptococcus), which is the most important neonatal pathogen in industrialized countries, was striking. Another study from The Gambia found that rectovaginal carriage rates for S. agalactiae in Gambian women in labor were similar to those reported in the US, and significant infant carriage was demonstrated in the same study.20 Studies from other developing countries, particularly southern Africa, have shown that in those settings S. agalactiae is an important neonatal pathogen, mainly causing early onset neonatal disease (K Klugman, personal communication).21, 22 The absence of S. agalactiae in this study was not the result of small numbers of infants being seen during the first week of life given that 360 (8% of all infants enrolled and evaluated) were seen in the first week, compared with 505, 419, 389 and 384 in the 2nd, 3rd, 4th and 5th weeks, respectively. However, serious illness is much more common during the first week of life,1 so we can assume that many early sepsis cases did not reach health facilities.
The importance of Salmonella spp. as a pathogen in this age group is an important finding of this study. The association of this pathogen with malnutrition is striking. This pathogen must be considered in any sick young infant in a developing country, especially if malnutrition is also present.
Most of the isolates of Staphylococcus aureus were from the Gambia (69%) which was undergoing an epidemic of scabies at the time of the study. Scabies in very young infants typically presents with pustular eruptions on the palms and soles which are almost always secondarily infected. This may have been the source of many of the staphylococcal infections in our study. Historically there is great variation in neonatal staphylococcal infection by country and time, although most documented outbreaks have been hospital-associated. Staphylococcal bacteremia in neonates correlates with skin disease and tends to be self-curing. In this study the case-fatality rate in infants who were blood culture-positive for Staphylococcus aureus was 21%. This study supports the view that it is reasonable to include an antistaphylococcal penicillin such as cloxacillin in the initial therapy of neonatal sepsis in infants with pustular skin sores or in communities where many infants contract scabies. The finding of Salmonella spp. in 10% (17 of 167) of positive blood isolates is troubling as infection with this species of bacteria is not well covered with the penicillin-gentamicin combination. Ampicillin, chloramphenicol or third generation cephalosporins would be required to cover Salmonella spp.
The case-fatality rate of young infants with clinical signs suggestive of serious bacterial infection and a positive blood culture was 30%, despite hospital admission for standard treatment (usually with benzylpenicillin or ampicillin and gentamicin) in well-functioning hospitals. The number of deaths despite hospital care points to the importance of prompt care-seeking, early detection of cases and effective treatment and high-lights the overwhelming importance of prevention. Overall clinical symptoms were not particularly helpful in distinguishing among infections caused by different organisms. Microbiology data are rarely available to guide patient care in developing country hospitals. Therefore empiric treatment should be adequately effective against the likely pathogens.
The studies described in this paper represent the largest prospective study of early infant infections in developing countries. The findings suggest that in developing countries initial therapy of neonatal sepsis with ampicillin and gentamicin is suitable, covering most of the likely Gram-positive and Gram-negative organisms. The main problem with this regimen is the poor coverage of Staphylococcus aureus, which is now usually resistant to penicillin. The growing problem of penicillin-resistant S. pneumoniae makes this combination unsuitable for the empiric therapy of neonatal meningitis in many areas, which should be treated initially with a third generation cephalosporin such as cefotaxime once the diagnosis has been confirmed by lumbar puncture. This would also be a suitable first line antibiotic in this age group where it is available, although poor antistaphylococcal activity is of concern. If skin sepsis suggests likely staphylococcal disease, initial therapy should include an antistaphylococcal agent such as cloxacillin, but if this is used to replace ampicillin, the reduced efficacy of the combination against Salmonella spp. and S. pneumoniae should be kept in mind. If a child has not improved within 48 h, or if deterioration is evident within that time, treatment should be changed to second line antibiotic therapy, although cost will limit this in many developing country settings. Where possible this should be guided by bacteriology. However, if this is not available the addition of an antistaphylococcal agent such as cloxacillin to ampicillin and gentamicin would constitute a logical second line. Where the initial regimen included an antistaphylococcal agent, the second line therapy should strive to improve the Gram-negative cover by the use of cefotaxime if available, or chloramphenicol. The use of chloramphenicol in young infants has been associated in the past with serious adverse effects caused by the high doses used. If appropriate doses and dose intervals are used and they are calculated and administered with great care, chloramphenicol succinate can be safely administered intramuscularly to infants in this age group.
Better treatment of seriously ill young infants in developing countries is urgently needed. In most cases this can still be achieved with relatively inexpensive antibiotics. However, this study has shown that, even with adequate treatment, there is a high mortality. Prevention must be central to efforts to control this problem.
The investigators thank the medical and nursing staff of the participating hospitals, the bacteriology laboratory staff at each of the study centers and most importantly the families of young infants who participated in the study.
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Publication of this supplement was supported by a grant from the World Health Organisation Programme for Control of Acute Respiratory Infections.