|Year : 2019 | Volume
| Issue : 3 | Page : 1094-1098
Evaluation of serum amyloid A protein as a marker in neonatal sepsis
Ghada M Elmashad1, Hanan M Elsayed1, Zein A Omar1, Eman A Badr2, Osama M Omran3
1 Department of Pediatric, Faculty of Medicine, Menoufia University, Menoufia, Egypt
2 Department of Medical Biochemistry and Molecular Biology, Faculty of Medicine, Menoufia University, Menoufia, Egypt
3 Department of Pediatric, Benha Children Specialized Hospital, Qaliobia, Egypt
|Date of Submission||30-Jan-2019|
|Date of Acceptance||30-Mar-2019|
|Date of Web Publication||17-Oct-2019|
Osama M Omran
Quesna, Menoufia Governorate
Source of Support: None, Conflict of Interest: None
To evaluate the serum amyloid A (SAA) as an early and accurate diagnostic marker in neonatal sepsis.
The early and efficient diagnosis of neonatal sepsis in high-risk neonates remains a difficult task as the clinical signs are nonspecific, complete blood count parameters and C-reactive protein (CRP) have low sensitivity, also difficulty of its diagnosis may be due to decreased positive values of blood culture and long time which is needed for detection of blood culture results. The SAA protein level in the blood increases earlier and up to 1000-fold in response to inflammation.
Participants and methods
A case–control study was carried out on 50 septic newborns who were admitted to the Neonatal ICU, Benha Specialized Children Hospital. Patients were subdivided into confirmed (35 cases) and clinical (15 cases) septic groups as well as 25 sex-matched and age-matched neonates as a control group during the study period from April to November 2018. Full assessment of history, clinical examination, complete blood count, CRP, blood culture, and SAA protein were performed for all neonates.
The mean value of SAA in the confirmed septic group (35 cases with elevated CRP and positive blood cultures) was 77.1 ± 5.5 μg/ml and that in the clinically septic group (15 cases with elevated CRP and negative blood cultures) was 18.5 ± 1.32 μg/ml compared with the control group (4.7 ± 1.1 μg/ml), and this represented a highly statistically significant difference (P = 0.000).
Serum amyloid A increases significantly in neonates with sepsis in comparison with healthy neonates. Therefore, SAA protein could aid the clinicians in diagnosing most cases of neonatal sepsis.
Keywords: markers, neonatal sepsis, serum amyloid A
|How to cite this article:|
Elmashad GM, Elsayed HM, Omar ZA, Badr EA, Omran OM. Evaluation of serum amyloid A protein as a marker in neonatal sepsis. Menoufia Med J 2019;32:1094-8
|How to cite this URL:|
Elmashad GM, Elsayed HM, Omar ZA, Badr EA, Omran OM. Evaluation of serum amyloid A protein as a marker in neonatal sepsis. Menoufia Med J [serial online] 2019 [cited 2019 Nov 19];32:1094-8. Available from: http://www.mmj.eg.net/text.asp?2019/32/3/1094/268824
| Introduction|| |
Neonatal sepsis is defined as a clinical syndrome in the first 28 days of life, manifested by systemic signs of infection and isolation of a bacterial pathogen from the bloodstream . The epidemiology of neonatal sepsis is important because it is the most common cause of neonatal mortality and is responsible for 30–50% of total neonatal deaths each year in developing countries. It is estimated that up to 20% of neonates develop sepsis and ∼1% die from sepsis-related causes . As the signs and symptoms of sepsis are nonspecific, early diagnosis poses a challenge to the clinicians. Blood culture is commonly used as a traditional approach for the detection of sepsis. However, blood cultures are time-consuming and at least 2–5 days of culture are necessary to identify the organism in the blood. Moreover, the sensitivity of blood cultures decreases significantly if antibiotic therapy has been initiated or when highly sensitive or slowly growing pathogens are cultured are cultured. Therefore, other tests are carried out along with the blood culture to identify sepsis . However, even with the current severity scores and biomarkers that are derived from complete blood count (CBC), prediction of outcomes of neonatal sepsis is still challenging. Conventional screening tests such as total and differential leukocyte counts, band cells, neutrophil counts, and rapid immunological tests such as C-reactive protein (CRP) assays may aid in the diagnosis of septicemia . The early and efficient diagnosis of neonatal sepsis in high-risk neonates remains a difficult task as the clinical signs are nonspecific. If treatment is delayed until the symptoms and signs of sepsis become obvious, the risk of preventable mortality will increase . A number of diagnostic tests are currently used in the assessment of a neonate suspected of having sepsis. The standard method for the diagnosis of bacterial infection is the culture of body fluids, especially blood; however, it often takes up to 72 h for the results of bacterial cultures to be known, also some cases of infected infants couldn't be diagnosed by blood culture . Serum amyloid A (SAA) (the precursor protein in inflammation-associated reactive amyloidosis, whose level in the blood increases ≤1000-fold in response to inflammation) is synthesized in the liver. SAA is also an acute-phase reactant like procalcitonin (PCT) and CRP . Therefore, the aim of this study was to evaluate the SAA as an early diagnostic marker in neonatal sepsis.
| Participants and Methods|| |
The study protocol was approved by the Ethical Committee of Faculty of Medicine, Menoufia University. A case–control study was carried out on 50 septic newborns who were admitted to the Neonatal ICU, Benha Children Specialized Hospital. Patients were subdivided into confirmed (35 cases) and clinical (15 cases) septic groups as well as 25 sex-matched and age-matched neonates as a control group during the study period from April to November 2018. This study included infants in the neonatal period, both preterm and full-term neonates. Newborn infants with neonatal asphyxia, metabolic diseases, congenital malformations, and chromosomal abnormalities were excluded from our study. All the neonates were subjected to a full detailed assessment of history, meticulous clinical examination, and laboratory investigations including CBC using the electronic counter system Sysmex Corporation; Sawgrass Drive, Bellport, New York, USA, with its blood parameters used to calculate the hematological sepsis scoring system , CRP using the test kit (Cromatest), and blood culture with incubated media in the BECTON DICKINSON company, Sawgrass Drive, Bellport, New York, USA. SAA levels were determined for all neonates in the studyusing human SAA enzyme-linked immunosorbent assays kit (BioSource Europe SA. Rue de l'Industrie 8. Nivelles B-1400. Belgium). A total of 75 newborns were included in the study and categorized into two groups:group 1 and group 2. Group 1 included the cases admitted to the neonatal ICU and consisted of 50 newborn infants with suspected sepsis clinically  and CRP more than 6 mg/l. Group 1 was subdivided into two groups, group 1A and group 1B, depending on the results of blood culture. Group 1A [confirmed sepsis group (n = 35)] had positive blood cultures and group 1B [clinically septic group (n = 15)] had negative blood cultures. Group 2 was the control and included 25 healthy newborns. All parameters in this study were cross-matched among the septic and control groups. Statistical analysis: Results were tabulated and statistically analyzed using a personal computer using Microsoft Excel 2016 and SPSS, version 23 (SPSS Inc., Chicago, Illinois, USA). Statistical analysis was carried out using descriptive data, for example, percentage, mean, and SD and analytical data, which include the F test. The receiver-operating characteristic curve was constructed to determine the cut-off level of sepsis markers. A value of P less than 0.05 was considered statistically significant.
| Results|| |
The hemoglobin levels in groups 1A and 1B were significantly decreased compared with that in group 2. Groups 1A and 1B showed a significant increase in the total leukocytic count (TLC) (16.3 ± 7.6 and 17.2 ± 4.8 × 1000/mm 3) than in group 2 (11.4 ± 3.6 × 1000/mm 3). The immature to total neutrophils ratio (I: T) was significantly increased in groups 1A (0.27 ± 0.12) and 1B (0.21 ± 0.16) than in group 2 (0.08 ± 0.02), whereas the platelet count decreased significantly in groups 1A (113.3 ± 55) and 1B (169.5 ± 35) than in group 2 (261.2 ± 68) [Table 1].
|Table 1: Hemoglobin level, total leukocytic count, immature to total neutrophils ratio, and platelet count among the groups studied|
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Also, CRP increased significantly in group 1A (30.34 ± 2.44 mg/l) than in group 1B (7.20 ± 2.48 mg/l). SAA levels increased significantly in group 1A (77.1 ± 5.5 μg/ml) and 1B (18.7 ± 1.32 μg/ml) than in group 2 (4.5 ± 1.1 μg/ml), with P value less than 0.001 [Table 2].
|Table 2: C-reactive protein levels and serum amyloid A among the studied groups|
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The most common organisms in the infected group were Gram-negative organisms. Suspected sepsis was confirmed in 35 neonates; 24 of these cases (65.59%) had Gram-negative organisms and 11 (34.41%) cases had Gram-positive organisms. Klebsiella pneumoniae was the most common organism (37.1%) isolated from blood cultures, followed by Escherichia More Details coli (17.1%), coagulase-negative staphylococci (14.3%), Staphylococcus aureus (8.6%), Pseudomonas aeruginosa (5.7%), Enterobacter spp. (8.6%), group B streptococci (GBS, 5.7%), and Staphylococcal epidermidis (2.9%) [Table 3].
|Table 3: Causative organisms in 35 septic neonates with positive blood cultures|
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The receiver-operating characteristic curve was constructed to determine the cut-off level of sepsis markers. The results showed that the SAA protein was the most sensitive marker [91.42%, area under curve (AUC)=0.99, cut-off point 10 μg/ml] compared with the sensitivity of CRP (90%, AUC = 0.69, cut-off point 12 mg/l), TLC (80%, AUC = 0.45, cut-off point 10 × 1000 cm 3), I: T (80%, AUC = 0.74, cut-off point 0.20), and platelet (40%, AUC = 0.11 cut-off point 148 × 1000 cm 3). Also, the SAA protein was the most specific (86.66%) compared with CRP (80%), I: T (75.3%), TLC (53.33%), and platelet (34%). The positive predictive value (PPV) of SAA was 94.11%, which was higher than CRP (67%), TLC (63.16%), I: T (87.20%), and platelet (37.50%). The highest negative predictive value (NPV) of SAA was 81.25% compared with TLC (72.73%), I: T (77.00%), CRP (69%), and platelet (35.71%) [Table 4] and [Figure 1].
|Table 4: Comparison between serum amyloid A and other markers of sepsis in sensitivity, specificity, positive predictive value, negative predictive value, and accuracy according to the receiver-operating characteristic curve|
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|Figure 1: Receiver-operating characteristic curve for markers of sepsis.|
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| Discussion|| |
This study was carried out to evaluate the diagnostic value of SAA in neonatal sepsis. The results indicated that the hemoglobin levels in groups 1A and 1B were significantly decreased compared with that in group 2. Groups 1A and 1B showed a significant increase in the total leukocytic count than group 2. The I: T was significantly increased in groups 1A and 1B than group 2, whereas the platelet count decreased significantly in groups 1A and 1B than group 2. This is in agreement with the results of Fathy et al.  and Abou El-Ela et al.  Neutrophils are types of white blood cells that defend the body against organisms that cause infection. When the demand for the neutrophils exceeds the supply, immature neutrophils are released into the blood to help fight off the infection, which is called shift to left . Engle et al.  reported that the I: T ratio can provide some diagnostic information on sepsis, but has limitations in sensitivity and specificity. A ratio of 0.16 or greater suggests bacterial infection, whereas a ratio higher than 0.2 has been correlated with an increased risk of bacterial infection . Also, Powell and Marcy  found that there was a highly significant difference between septic and control neonates in terms of the platelet count, with a mean level in the confirmed sepsis group of 113.3 ± 55 and the clinically sepsis group of 169.5 ± 35, whereas in the control group, the mean level was 261.2 ± 68. Also, only 10–60% of newborns with proven neonatal sepsis had thrombocytopenia. The cause of thrombocytopenia may be due to toxic destruction of the infants' platelets, megakaryocytic suppression, increased peripheral consumption as in disseminated intravascular coagulopathy, or the presence of immune component because of increased levels of platelet-associated immunoglobulins .
Also, CRP increased significantly in group 1A than in group 1B. SAA levels increased significantly in groups 1A and 1B than group 2. This is in agreement with the results ofFathy et al.  and Arnon et al. , who found that the mean value of SAA in the septic group was 187.6 ± 78.3 μg/ml compared with 10.2 ± 8.3 μg/ml in the nonseptic group. Yuan et al., 2013. reported that serum level of Amyloid A protein is highly sensitive and specific in diagnosis of inflammatory process in neonatal sepsis.
This study indicated that the most common organisms in the infected group were Gram-negative organisms. Suspected sepsis was confirmed in 35 neonates; 24 (65.59%) of these cases had Gram-negative organisms and 11 (34.41%) cases had Gram-positive organisms. Klebsiella pneumoniae was the most common organism (37.1%) isolated from blood cultures, followed by E. coli (17.1%), coagulase-negative staphylococci (14.3%), S. aureus (8.6%), P. aeruginosa (5.7%), Enterobacter spp. (8.6%), GBS (5.7%), and S. epidermidis (2.9%). These results were in agreement with Hashim et al. , who reported that Klebsiella is the most common isolated organism in septic newborns, with a ratio ranging from 35 to 56% of all isolated organisms. In contrast, several studies have shown that GBS were the most common organism isolated in early-onset sepsis in 28% of cases , and E. coli was the most common isolated organism in 52 and 25% of cases according to Hashim et al.  The difference between our results and other studies may be because of the different infection control measures, small sample size, prolonged invasive procedures, intravenous line, central catheters, and parenteral nutrition.
Also, the results showed that the SAA protein was the most sensitive marker (91.42%, AUC = 0.99, cut-off point 10 μg/ml) compared with the sensitivity of CRP (90%, AUC = 0.69, cut-off point 12 mg/l), TLC (80%, AUC = 0.45, cut-off point 10 × 1000 cm 3), I: T (80%, AUC = 0.74, cut-off point 0.20), and platelet (40%, AUC = 0.11 cut-off point 148 × 1000 cm 3). Also, the SAA protein was the most specific (86.66%) compared with CRP (80%), I: T (75.3%), TLC (53.33%), and platelet (34%). The PPV of SAA was 94.11%, which was higher than CRP (67%), TLC (63.16%), I: T (87.20%), and platelet (37.50%). The highest NPV of SAA was 81.25% compared with TLC (72.73%), I: T (77.00%), CRP (69%), and platelet (35.71%). This is in agreement with the results of Fathy et al. , found that cut-off value of SAA of at least 10 μg/ml with sensitivity 96%, specificity 95%, PPV 85%, and NPV 99% at onset of sepsis. However, the study of Arnon et al.  found that the PPV of SAA was 96% and Arnon et al.  found that the PPV of SAA was 87% in comparison with the PPV of CRP (86%) and the PPV of IL-6 (64%). In a study carried out by Cetinkaya et al. , the role of SAA in the diagnosis of neonatal sepsis in comparison with CRP and PCT was determined. They found that SAA is more sensitive (76.4%) than PCT (74.8%) and CRP (72.3%). However, in the study carried out by Walliullah et al. , it was found that the specificity of the I: T ratio was 56% in the diagnosis of sepsis.
Also, Remington and Klein  found that the I: T ratio can provide some diagnostic information on sepsis, but had limitations in sensitivity and specificity. Finally, further studies on SAA on a larger scale are needed to verify its diagnostic value in neonatal sepsis. Also studies should be conducted on neonates in different gestational ages for detection of relation between serum Amyloid A level and fetal maturity. SAA should be studied in other pathological conditions of neonates such as trauma and asphyxia to verify the specificity of SAA release to neonatal sepsis. More studies are also recommended to discover more sensitive and cheaper tools to identify neonates with sepsis.
| Conclusion|| |
This study showed that SAA increases significantly in neonates with sepsis in comparison with healthy neonates. Therefore, SAA protein could help the clinicians to diagnose most cases of neonatal sepsis.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Table 1], [Table 2], [Table 3], [Table 4]