This systematic review and meta-analysis identified seven independent risk factors for EOS, three and four of which were maternal and neonatal risk factors, respectively. Based on the significance of the combined after-effect, the most independent risk factor for EOS was at least three PV examinations. From our analysis, the incidence of EOS in neonates with this risk factor was approximately 7.18 times that of other neonates from our analysis.13,19 At least three PV examinations during labor and delivery were performed in 212 mothers, who were from cities in developing countries. This may be due to a low level of medical care, the non-practice of strict aseptic procedures, and the direct introduction of vaginal microbes into the uterus through the cervical canal during PV examination or under aseptic conditions. Additionally, repeated PV examination can easily cause an imbalance of the vaginal microecology and eventually lead to a significant increase in the risk of neonatal infection.19 Moreover, frequent PV examination may also lead to the occurrence of acute or subacute chorioamnionitis (7.4% in our study population). Chorioamnionitis is mainly caused by microorganisms invading the amniotic fluid, and the most common causative bacterium is Ureaplasma urealyticum. Amniotic fluid infection may cause vasospasm in the umbilical cord and placenta, and reduce fetal perfusion; further, fetal inhalation or ingestion of bacteria in the amniotic fluid may cause congenital pneumonia or systemic infection, and increase perinatal asphyxia, which increases the incidence of EOS and related mortality.34 In addition, chorioamnionitis can be caused by PROM, which is directly related to the duration of rupture.1 From our findings, when PROM lasts for more than 18 h, microorganisms tend to enter the amniotic cavity through the cervix, thereby causing chorioamnionitis, fetal distress, and asphyxia. This leads to intrauterine infection and induces a 2.74-fold increase in EOS risk.10,35,36 PROM may also lead to premature delivery. In neonates with lower GA, innate and adaptive immunity may be more immature, and hence, the risk of infection is 4.08 times that in full-term infants.37 Premature neonates may have an impaired immune function due to a lack of maternal IgG placental transfer, which occurs in late pregnancy.38 Moreover, the colonization of Ureaplasma in the respiratory tract of premature neonates with maternal chorioamnionitis after PROM can lead to bronchopulmonary dysplasia, which increases the risk of neonatal respiratory tract infection and perinatal asphyxia.39 It is necessary to resuscitate neonates with respiratory failure; however, neonatal peripheral airway stenosis, rich respiratory secretions, atelectasis, and effective resuscitation treatment may lead to neonatal vulnerable mucosa secondary injury and become the entry point of microorganisms. In addition, during resuscitation, catheters inserted into the trachea or blood vessels may not be sterile, and these equipments may introduce microorganisms into the lungs of neonates whose immune system is not well developed. This exposes the neonate to bacteria from the mother or exogenous pathogens during resuscitation and increases the risk of infection.1,10 Currently, few studies have focused on the bacterial colonization of an inserted intravenous catheter by added catheter culture to a scoring system for predicting neonatal sepsis occurrence, which significantly improves the discrimination of the latter.13,40 However, from the current research, bacterial culture of an implanted catheter in asymptomatic neonates has a certain predictive value for the occurrence of hospital-acquired neonatal sepsis. Furthermore, male sex was an independent risk factor of EOS. From a genetic perspective, innate immune cells carry a gene pool of alleles. In females, the difference of innate immune cells in X-linked parental alleles with different regulatory and activation abilities can represent a more effective immune system, which can better adapt to the dynamic changes during the inflammatory response.41 Moreover, different sexes have different levels of sex hormones, especially estrogen. Animal studies have shown that exogenous estrogen injection can improve the immunosuppressive state of male animals, and a high serum concentration of estrogen can eliminate the toxic effects of active free radicals, and reduce inflammatory damage.1 Therefore, male sex is a risk factor of EOS. In addition to genetic effects, sex hormone levels are an important cause of differences in EOS occurrence in males and females.38
Although birth weight, APGAR score, and cesarean section were also included in our meta-analysis, these variables showed strong heterogeneity after combination, which could not guarantee the reliability of the combined results. Further clinical studies are needed to confirm the role of these variables in EOS occurrence. In particular, many studies reported birth weight as an important risk factor for EOS. Five of the included studies (except the study by Levy et al., which did not explicitly present neonates with low birth weight), 11,15,16,24,25 mainly divided the birth weights into two groups (< 1500 g and < 2500 g); nevertheless, the combined results had strong heterogeneity. Although it is impossible to determine the influence of low birth weight in EOS occurrence by meta-analysis, this risk factor cannot be ignored, and further clinical research is needed.
Apart from the abovementioned risk factors, many risk factors were not included in the meta-analysis due to the limitations of the current research results. However, these factors are very important, such as the presence of meconium-stained amniotic fluid, early maternal age, and the absence of crying immediately after birth. Anna et al. suggested that the OR of the presence of meconium-stained amniotic fluid as a risk factor was 3.63 (95% CI: 1.04–12.65). Compared to neonates without fecal infection, those with meconium-stained amniotic fluid have a higher risk of infection, probably because the meconium-stained amniotic fluid enters into fetal lungs, leading to dyspnea, hypoxemia, pneumonia, and finally EOS.42 In addition, maternal age ≤ 15 years is a risk factor for EOS, which is related to differences in the microbial flora of the vagina. Lactobacilli in the vaginal microbiota of normal adult women can prevent infection by reducing vaginal pH and producing antimicrobial substances, such as hydrogen peroxide. However, the dominant bacteria in the vaginal microbiota of young women under age 15 were different; most of them had E. coli, which alters the acidic environment of the vagina that is important against infections, weakens the ability of the vagina to inhibit pathogenic bacterial action, and increases the probability of neonatal infection.11 A case-control study conducted by Alemu et al. found that neonates who did not cry at birth were 2.85 times more likely to develop sepsis than those who cried at birth.10 Crying at birth may be the manifestation of a series of changes in lung function when the neonate takes the first breath, and it is the most critical part in the process of physiological adjustment of the neonate to survive outside the uterus after the umbilical cord is cut off. Initial breathing is the result of reflexes triggered by changes in pressure, temperature, noise, light, and other sensations associated with the birth process. Neonates may not be able to cry due to respiratory interference. Failure to breathe and/or cry would require immediate resuscitation, increase the chances of clinically invasive procedures, and increase the risk of infection.10
From the perspective of causative microorganisms, a few patients in this study had multiple positive blood culture results; thus, the actual proportion of patients with positive blood culture results should be lower. Among them, 160 patients had CONS, which can produce false-positive blood culture results since it is a skin-colonizing bacterium. The number of infections caused by CONS in the included studies may have been overestimated. In addition, CONS infection usually shows low virulence. Therefore, in the blood culture diagnosis of neonatal sepsis, some scholars treat CONS as a bacterial contamination of samples, not as an infectious cause of neonatal sepsis.3,43 Group B Streptococcus, a vagina-colonizing bacterium, has always been considered to constitute the main bacterial group causing EOS.14 Nonetheless, we found that E. coli accounted for the largest proportion in each flora, followed by CONS and Klebsiella pneumoniae; Group B Streptococcus ranked fourth. This probably indicates that, due to the frequent long-term use of antibiotics against gram-positive bacteria and the use of medical procedures such as prenatal PV examination, neonatal resuscitation, and central venous catheterization, the causative pathogen of EOS has gradually changed from gram-positive to gram-negative bacteria. Shane et al. and Stoll et al. reported that E. coli was the most common cause of EOS, which was akin to our findings.1,3 In addition, we found that fungi caused approximately 3% of EOS, all of which were caused by Candida. However, because three articles on fungal infection in this study did not distinguish between EOS and LOS, the fungal infection rate is probably underestimated in the current study. Compared with the LOS study, fungal infections seem to occur more frequently in patients with LOS.44 During hospitalization, due to the application of broad-spectrum antibiotics, normal body floras are out of balance. Many antibiotic-sensitive bacteria are killed, and therefore fungi can reproduce and grow. Thus, fungal infections can easily occur in patients with LOS. Compared to infections caused by gram-positive and gram-negative bacteria, sepsis caused by fungal infections are often more severe and result in higher mortality.45–47
Strengths and limitations
Our study had the following strengths. First, we evaluated the risk factors of EOS, which provides an effective prevention and treatment strategy to reduce the serious damage caused by EOS in the clinic. Seven independent risk factors of EOS were identified through quantitative meta-analysis. Second, we also systematically summarized the causative bacteria of EOS, and found that E. coli, not Group B Streptococcus, was the main cause of EOS.
We acknowledge that this study has some shortcomings. First, in the two studies on CVC, the analysis of neonatal sepsis did not distinguish between EOS and LOS. Therefore, although there was no heterogeneity in the results of our combination, it cannot be ascertained whether CVC is a risk factor for EOS or LOS. Second, to ensure the accuracy of our results, we eliminated most of the studies with heterogeneity through sensitivity analysis, resulting in a small number of studies, which can weaken the results of the current meta-analysis for each risk factor. Furthermore, the inclusion of the neonatal sepsis population had a certain degree of skewness, as two, four, and one studies were based on very low birth weight neonates (< 1500 g), premature neonates (< 37 weeks), and neonates with sepsis and congenital disease (diaphragmatic hernia), respectively; thus, our results may have publication bias.