|Year : 2020 | Volume
| Issue : 2 | Page : 80-85
Neonatal fungal sepsis in Jos North-Central Nigeria
Okolo Mark Ojogba1, Ayanbimpe M Grace2, Toma O Bose3, Envuladu A Esther4, Olubukunnola Inyang5, Izang Abel5, Obishakin F Emmanuel6, Dashe Nanma2, Onyedibe Kenneth2, Echeonwu Bobmanuel7, Sagay A Solomon8, Egah Z Daniel2, Novitsky Vladimir9
1 Department of Medical Microbiology, College of Health Science, University of Jos; Department of Histopathology, National Veterinary Research Institute, Vom, Plateau State, Nigeria
2 Department of Medical Microbiology, College of Health Science, University of Jos, Nigeria
3 Department of Paediatrics, College of Health Science, University of Jos, Nigeria
4 Department of Community Medicine, College of Health Science, University of Jos, Nigeria
5 Department of Paediatrics, Plateau State Specialist Hospital, Jos, Nigeria
6 Department of Biotechnology, National Veterinary Research Institute, Vom, Plateau State, Nigeria
7 Department of Histopathology, National Veterinary Research Institute, Vom, Plateau State, Nigeria
8 Department of Obstetrics and Gynaecology, College of Health Science, University of Jos, Nigeria
9 Harvard School of Public Health, Boston, USA
|Date of Submission||12-Aug-2019|
|Date of Decision||21-May-2020|
|Date of Acceptance||01-Jun-2020|
|Date of Web Publication||11-Sep-2020|
Okolo Mark Ojogba
Department of Medical Microbiology, University of Jos, Plateau State
Source of Support: None, Conflict of Interest: None
Background: Fungal sepsis in neonates is still one of the major causes of morbidity and mortality despite advances in health care. This study aimed to characterize fungal agents of sepsis in neonates and their susceptibility pattern. Methods: This was a cross-sectional study among neonates in two tertiary health care facilities in Jos. Neonates with sepsis whose parents consented to the study were recruited based on the Integrated Management of Childhood Illnesses(IMCI) criteria. Blood sample was collected for culture, antifungal susceptibility test and molecular characterization of fungal agents isolated from blood culture of the neonates was performed using the ribosomal DNA (rDNA) of the internal transcribed spacer (ITS) region. Univariate and bivariate analysis was carried out using STATA statistical software (version 14 IC). Results: The prevalence of fungal sepsis in neonates was 5.5%. Candida albicans was responsible for 11 of the 20 cases of neonatal fungal sepsis. All the fungal isolates were susceptible to the antifungal agents used except for a little resistance by C. glabrata observed to amphotericin B (%R=0.3). Bayesian analysis confirmed the major phylogenetic relationships among the isolates and molecular identification of the different Candida species. Conclusion: Candida albicans are the major cause of neonatal fungal sepsis. The study highlights the need to evaluate the causes of neonatal fungal sepsis, their antifungal susceptibility pattern and molecular characterization for early implementation of medical intervention to reduce the morbidity and mortality.
Keywords: Antifungal agents, Candida albicans, neonates, neonatal fungal sepsis
|How to cite this article:|
Ojogba OM, Grace AM, Bose TO, Esther EA, Inyang O, Abel I, Emmanuel OF, Nanma D, Kenneth O, Bobmanuel E, Solomon SA, Daniel EZ, Vladimir N. Neonatal fungal sepsis in Jos North-Central Nigeria. J Med Trop 2020;22:80-5
|How to cite this URL:|
Ojogba OM, Grace AM, Bose TO, Esther EA, Inyang O, Abel I, Emmanuel OF, Nanma D, Kenneth O, Bobmanuel E, Solomon SA, Daniel EZ, Vladimir N. Neonatal fungal sepsis in Jos North-Central Nigeria. J Med Trop [serial online] 2020 [cited 2020 Sep 22];22:80-5. Available from: http://www.jmedtropics.org/text.asp?2020/22/2/80/294816
| Introduction|| |
Fungal sepsis (FS) is still one of the major causes of morbidity and mortality in neonates, despite the recent advances in health care. Neonatal fungal sepsis can be early when it occurs less than or equal to three days from birth or late when it occurs after three days of birth but less than 28 days from birth. Mortality from FS is between 21% and 76% among neonates.
In a descriptive study of Candidaemia among immunosuppressed patients in Ibadan, Nigeria, Candida (C) tropicalis was the most common fungal isolate (50%), followed by C. parapsilosis in 33.3% and C. albicans 8.3%.
Non-albicans Candida (NAC) isolates, including C. parapsilosis, are being isolated with increasing frequency.,, A number of studies have shown that NAC exhibit varying levels of susceptibility to fluconazole and other anti-fungal agents., Hence, the use of empirical fluconazole as first-line therapy may be inappropriate.
Early diagnosis of FS and appropriate treatment improves neonatal outcomes. However the definitive diagnosis of FS is difficult and pathogen isolation and identification takes long time, hence neonates are treated with empirical therapy while waiting for culture results., There is urgent need to have current data on neonatal FS and their antifungal susceptibility pattern to guide empiric treatment.
This study set out to determine the fungal agents of sepsis in neonates and their sensitivity patterns in Jos, North-Central Nigeria.
| Materials and methods|| |
In order to achieve the objective, a cross-sectional study was designed involving neonates diagnosed with sepsis admitted in the Special Care Baby Unit (SCBU) of two tertiary health care facilities in Jos, Plateau State, North-Central Nigeria. The institution research board committees for the two health care facility approved the study. Case definition of neonatal sepsis was based on the Integrated Management of Childhood Illnesses (IMCI) criteria. Blood volume of 1.5ml was collected from the neonates, added to the blood culture broth and incubated at 37°C. Positive samples were examined, antifungal susceptibility patterns were determined using clinical and laboratory standard institute (CLSI) guideline, and the isolates were stored at −20°C for molecular characterization.
Total genomic DNA extraction from the Candida isolates was performed using Zymo Quick-DNA kits reagents (Zymo Research Corporation, USA) in accordance with the manufacturer’s instructions. Polymerase chain reaction (PCR) assay optimization was performed using specific primer sequences for the internal transcribed spacer 1 and 4 (ITS 1 and ITS 4) regions; ITS1 (5’-TCCGTAGGTGAACCTGCGG-3’) as forward primer and ITS4 (5’-TCCTCCGCTTATTGATATGC-3’) as reverse primer. The PCR products were subjected to dideoxynucleotide sequencing with a Big Dye Terminator Reaction Kit v3.1 (Applied Biosystems, Inc., Foster City, CA, USA) according to manufacturer’s instructions. After the purification and denaturation of the products, the samples were run on an automated ABI 3500XL genetic analyser (Applied Biosystems, Inc., Foster City, CA, USA). The sequences obtained in our study were aligned and compared with sequences deposited in public genomic database (GenBank, NCBI, USA). To ensure high reliability of the results obtained using the nucleotide sequence alignment tools, an e-value of less than 10−5 and a maximum identity of equal to or higher than 98% were considered for the correct identification of Candida at the species level. The evolutionary history was inferred using the Neighbor-Joining method. The optimal tree with the sum of branch length = 17.83120213 is shown. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (500 replicates) are shown next to the branches. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Maximum Composite Likelihood method and are in the units of the number of base substitutions per site. The analysis involved 24 nucleotide sequences. All positions containing gaps and missing data were eliminated. There were a total of 324 positions in the final dataset. Evolutionary analyses were conducted in MEGA7.
The data obtained was analyzed by means of STATA (Version 14IC) statistical software and presented in tables and figure.
| Results|| |
Of the 363 neonates with median age 8 days, definitive diagnosis of neonatal fungal sepsis was made in 20 out of the 363 neonates with a prevalence rate of 5.5%.
Early onset neonatal fungal sepsis (less than or equal to 3 days old) was demonstrated in one (1) of the 20 neonates and late onset neonatal fungal sepsis (more than 3 days old) in 19 of the 20 neonates [Table 1].
|Table 1: Demographic characteristics of neonates with fungal sepsis in Jos, North-Central Nigeria|
Click here to view
Eleven of the 20 neonates with fungal sepsis were of birth weight ≤2.5kg while nine of the 20 neonates with fungal sepsis had birth weight >2.5kg [Table 1].
All fungal agents isolated were C. albicans (55%), C. glabrata (25%), C. tropicalis (15%), C. parapsillosis (5%) [Table 1].
Antifungal susceptibility pattern shows that the organisms were still sensitive to the antifungal agents used [Table 2].
|Table 2: Antifungal susceptibilities of Candida species isolated from neonates with fungal sepsis in Jos, North-Central Nigeria|
Click here to view
Based on the Bayesian analysis [Figure 1] results of the rDNA ITS sequence, we identified different Candida species isolated from neonates with fungal sepsis. Large numbers of ITS sequences were generated based on quality control designed to ensure accuracy of molecular data used to identify the Candida species.
|Figure 1: Evolutionary relationships of Candida species isolated from neonates with fungal sepsis in Jos, North-Central Nigeria.|
Click here to view
| Discussion|| |
Candida species have emerged as a leading cause of neonate fungal sepsis. The prevalence of fungaemia in our study was 5.5%. This finding broadly supports the work by Oladele et al. who reported 5.2% in Ibadan, Nigeria. The result is not consistent with some reports with documented rates from 10% to 20%., A possible explanation for this might be due to differences in the sample size, variations in health care services and laboratory diagnostics between our study and those with higher prevalence.
Another important finding was the higher rate of isolation of C. albicans (55%) compared with the non-albicans Candida species (45%). Previous studies in China and some western Countries, have reported that C. albicans was the most common causative agent of neonatal candidaemia. Although these results differ from some published studies who reported 76.7% and 60.4% respectively for non-albicans Candida species., Our finding is similar to that reported at a hospital in China where C. albicans (57.8%) play a dominant role in candidaemia.
Historically, it is known that C. tropicalis is the second most common Candida species recovered from blood. In our study C. glabrata surpassed the other non-albicans Candida species to become the second most common species as reported by Chen et al. and Xia et al. We believe that this finding may simply be due to our geographic location and the small sample size; hence further studies with larger sample size are required to verify it.
Antifungal susceptibility testing is not routinely performed hence, empiric therapy approach is commonly followed in prescribing the drugs. When Candida enters and invades the host tissues, blood poisoning and systemic infections may occur. These infections are difficult to treat with antifungal drugs, with attendant high mortality rate (42%–63%). Several studies in the past showed that empiric use of antifungal agents, particularly fluconazole, is almost entirely responsible for changes in Candida ecology and antifungal susceptibility pattern. In accordance with the present results, previous studies have demonstrated that C. albicans were susceptible to fluconazole, voriconazole and amphotericin B. In many centres fluconazole is the first line agent used for empiric therapy, whereas in others it is amphotericin B. It has been shown that prior exposure to fluconazole was a predisposing factor for C. glabrata infection,, which may partly explain the variations observed regarding the distribution of Candida species among neonates. Many Neonatal Intensive Care Units (NICUs) use liposomal amphotericin B that is not usually affordable for empiric treatment whereas some others use fluconazole for empiric treatment.,, The accurate identification of Candida species is essential for effective therapeutic strategies to control fungal sepsis in neonates. For diagnostic purposes, DNA-based techniques have been used to differentiate Candida species. Worthy of note is the fact that all of the ITS sequences used for the analysis in our study were generated from the PCR products. The provision of reliable ITS sequences was based on quality control criteria that were applied from PCR to sequence analysis, such as, the amplification of products with the entire ITS region and the use of standard computational tools for sequence assembly and editing. In a recent study, nosocomial clusters have been shown to represent 33% of total isolates causing candidaemia in some hospitals in Canada.,,, Epidemiological surveys that determines relationship among species of Candida and geographic distribution of clusters of isolates enables the identification of outbreaks in fungal blood stream infections., Our study also reinforces the utility of DNA sequence analysis for evaluating the relationship among Candida species and its application for determining the molecular epidemiology of fungal sepsis in neonates.
Candida albicans are a significant cause of neonatal fungal sepsis. This study highlights the non C. albicans species that cause neonatal fungal sepsis and their antifungal susceptibility pattern; hence emphasizes the need for early detection, diagnosis and treatment of neonates with fungal sepsis. Molecular studies based on ITS sequencing are necessary for the identification of Candida species. However, establishment of surveillance on neonatal fungal sepsis is necessary to develop and evaluate preventive strategies.
This research was supported by the Fogarty International Centre (FIC); Office of the Director (OD/NIH); National Institutes of Neurological Disorders and Stroke (NINDS/NIH); and the National Institute of Nursing Research (NINR/NIH) of the National Institutes of Health under award number D43TW010130. The content is solely the responsibility of the authors and does not necessarily represent the views of the National Institutes of Health.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Xavier PCN, Chang MR, Nunes MO, Palhares DB, Silva RA e, Bonfim GF et al.
Candidemia neonatal, em hospital público do Mato Grosso do Sul. Revista da Sociedade Brasileira de Medicina Tropical. scielo; 2008;41:459-63.
Fu J, Ding Y, Wei B, Wang L, Xu S, Qin P et al.
Epidemiology of Candida albicans and non-C.albicans of neonatal candidemia at a tertiary care hospital in western China. BMC Infect Dis [Internet] 2017;17:329. Available from: https://www.ncbi.nlm.nih.gov/pubmed/28477628
Benjamin DK, Stoll BJ, Fanaroff AA, McDonald SA, Oh W, Higgins RD et al.
Neonatal candidiasis among extremely low birth weight infants: risk factors, mortality rates, and neurodevelopmental outcomes at 18 to 22 months. Pediatrics [Internet] 2006;117:84LP-92. Available from: http://pediatrics.aappublications.org/content/117/1/84.abstract
Rodriguez D, Almirante B, Park BJ, Cuenca-Estrella M, Planes AM, Sanchez F et al.
Candidemia in neonatal intensive care units: Barcelona, Spain. Pediatr Infect Dis J 2006;25:224-9.
Pandya H, Slemming W, Saloojee H. Health system factors affecting implementation of integrated management of childhood illness (IMCI): qualitative insights from a South African province. Health Policy Plan 2018;33:171-82.
Castanheira M, Deshpande LM, Davis AP, Rhomberg PR, Pfaller MA. Monitoring antifungal resistance in a global collection of invasive yeasts and molds: application of clsi epidemiological cutoff values and whole-genome sequencing analysis for detection of azole resistance in Candida albicans. Antimicrob Agents Chemother 2017;61.
Karabicak N, Alem N. [Antifungal susceptibility profiles of Candida species to triazole: application of new CLSI species-specific clinical breakpoints and epidemiological cutoff values for characterization of antifungal resistance]. Mikrobiyol Bul 2016;50:122-32.
Saitou N, Nei M. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 1987;4:406-25.
Felsenstein J. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 1985;39:783-91.
Roilides E. Invasive candidiasis in neonates and children. Early Hum Dev 2011;87:S75-6.
Ma C, Li F, Shi L, Hu Y, Wang Y, Huang M et al.
Surveillance study of species distribution, antifungal susceptibility and mortality of nosocomial candidemia in a tertiary care hospital in China. BMC Infect Dis 2013;13:1-9.
Fu J, Ding Y, Jiang Y, Mo S, Xu S, Qin P. Persistent candidemia in very low birth weight neonates: risk factors and clinical significance. BMC Infect Dis 2018;18:4-9.
Xu B, Wu J-Q, Weng X-H, Hu X-P, Wang X, Ou X-T et al.
Epidemiology and risk factors for non-Candida albicans candidemia in non-neutropenic patients at a Chinese teaching hospital. Med Mycol [Internet] 2011;49:552-5. Available from: https://dx.doi.org/10.3109/13693786.2010.541948
Chen J, Jiang Y, Wei B, Ding Y, Xu S, Qin P et al.
Epidemiology of and risk factors for neonatal candidemia at a tertiary care hospital in western China. BMC Infect Dis [Internet] 2016;16:1-5. Available from: http://dx.doi.org/10.1186/s12879-016-2042-9
Xia H, Wu H, Xia S, Zhu X, Chen C, Qiu G et al.
Invasive Candidiasis in preterm neonates in China: a retrospective study from 11 NICUS during 2009-2011. Pediatr Infect Dis J 2014;33:106-9.
Filioti J, Spiroglou K, Roilides E. Invasive candidiasis in pediatric intensive care patients: epidemiology, risk factors, management, and outcome. Intensive Care Med 2007;33:1272-83.
Brion LP, Uko SE, Goldman DL. Risk of resistance associated with fluconazole prophylaxis: systematic review. J Infect 2007;54:521-9.
Lortholary O, Desnos-Ollivier M, Sitbon K, Fontanet A, Bretagne S, Dromer F. Recent exposure to caspofungin or fluconazole influences the epidemiology of candidemia: a prospective multicenter study involving 2,441 patients. Antimicrob Agents Chemother 2011;55:532-8.
Sarvikivi E, Lyytikäinen O, Soll DR, Pujol C, Pfaller MA, Richardson M et al.
Emergence of fluconazole resistance in a Candida parapsilosis strain that caused infections in a neonatal intensive care unit. J Clin Microbiol 2005;43:2729-35.
[Table 1], [Table 2]