Outbreak of central nervous system infections among children in Thai Binh, Viet Nam

Figures & data

Figure 1. Monthly distribution of CNS infections among children in Thai Binh Pediatric Hospital, Viet Nam from 01/01/2015 to 31/12/2020 (black bars: period of current study) (A) and daily distribution of CNS infections among children in Thai Binh Pediatric Hospital, Viet Nam during the study period (B)

Table 1. Clinical definition and classification of CNS infection based on laboratory features of CNS infections among children.

Clinical definition (Included criteria)*	Features
Age	0–16 years
Fever	≥38°C
Clinical features	At least one:
	Stiff neck Altered/reduced consciousness Focal neurological signs Convulsion Bulging fontanelle if <12 months of age Irritability if <5 years of age Headache if <5 years of age Prostration • Petechial or purpuric rash
Laboratory investigation	Lumbar puncture performed, or planned at the time of assessment, by clinical team
CNS infection classification based on laboratory feature	Features (in addition to meeting clinical case definition)
Confirmed	Pathogen detected in CSF by culture and/or PCR OR Positive serology in CSF OR Positive blood culture AND purulent CSF (WBC ≥100 cells/μL OR [WBC 10–99 cells/μL AND glucose <2.2 mmol/L or protein >1.0 g/L]) OR Purulent CSF AND positive CSF Gram stain
Probable	Purulent CSF AND absence of identifiable pathogens by culture, Gram stain, PCR, or serology** OR Abnormal CSF (WBC 10–99 cells/μL AND normal protein/glucose) AND positive blood culture and/or positive CSF Gram stain
Suspected	Non-purulent CSF AND absence of identifiable pathogens by culture, PCR, or serology

Note: *For study enrolment, all four criteria had to be met (age, fever, clinical features, and laboratory).

**Pathogen-negative probable cases were re-categorized as suspected cases if laboratory testing was incomplete.

CNS: central nervous system, CSF: cerebrospinal fluid, WBC: white blood cell, PCR: Polymerase Chain Reaction.

Table 2. Characteristics of included patients.

Characteristics	N = 99 n (%)
Age
Median [range]	5 (0–12)
<12 months	26 (26.3)
1–4 years	23 (23.2)
≥ 5 years	50 (50.5)
Gender
Male	58 (58.6)
Female	41 (41.4)
History
Fever	99 (100)
Vomiting	52 (52.5)
Diarrhea	2 (2.0)
Reduced feeding/eating & drinking	3 (3.0)
Lethargic	18 (18.2)
Headache (N = 50)	46 (92.0)
Seizure	2 (2.0)
Antimicrobial auto-medication prior to admission	12 (12.1)
Symptoms duration before admission (days), median (range)	2 (1–7)
Transfer from district hospitals	24 (24.2)
Physical findings
Stiff neck	45 (45.4)
Altered/reduced consciousness	3 (3.0)
Focal neurological symptoms	2 (2.0)
Irritability (N = 50)	6 (12.0)
Bulging fontanelle (N = 26)	23 (88.5)
Purpuric rash	1 (1.0)
Laboratory findings
Blood examination
White blood cells
Normal	5 (5.1)
Elevated CRP	94 (94.9)
Normal	30 (30.3)
Elevated	69 (69.7)
CSF laboratory examination
WBC
<10	16 (16.2)
10–<100	25 (25.2)
≥ 100	58 (58.6)
Protein
≤ 1.0 g/L	87 (87.9)
>1.0 g/L	12 (12.1)
Glucose
≥ 2.2 mmol/L	94 (95.0)
< 2.2 mmol/L	5 (5.0)

CRP: C-reactive protein; CSF: cerebrospinal fluid; WBC: white blood cell.

Table 3. Pathogens identified in 99 included children.¹

Pathogens	Viral culture n (%)	BioFire multiplex PCR* n (%)	Overall n (%)
CSF
Enterovirus	11 (11.1)	62 (62.6)	66 (66.7)
Adenovirus	1 (1.0)		1 (1.0)
Enterovirus and mumps virus coinfection	1 (1.0)	–	1 (1.0)
Herpes simplex virus 1	0 (0)	1 (1.0)	1 (1.0)
Herpes simplex virus 6	0 (0)	1 (1.0)	1 (1.0)
Haemophilus influenzae	–	1 (1.0)	1 (1.0)
H. influenzae and Streptococcus pneumoniae coinfection	–	1 (1.0)	1 (1.0)
At least one pathogen	13 (13.1)	66 (66.7)	72 (72.7)
Blood culture
Burkholderia cepacia	–	–	1 (1.0)
Candida parapsilosis	–	–	1 (1.0)
Staphylococcus aureus	–	–	2 (2.0)
Streptococcus agalactiae	–	–	1 (1.0)
At least one pathogen	–	–	5 (5.1)

Notes: *No sample was positive for Neisseria meningitidis, Escherichia coli K1, S. agalactiae, Listeria monocytogenes, herpes simplex virus type 2, cytomegalovirus, varicella zoster virus, human parechovirus, and Cryptococcus neoformans/gattii.

¹ Additional real-time PCR tests were performed to identify Dengue virus, two bacteria: (Klebsiella pneumoniae and Staphylococcus aureus), and five main Candida species (C. parapsilosis, C. tropicalis, C. albicans, C. glabrata, and C. rugosa) on supernatant after viral culture. But no sample was positive for these pathogens.

CSF: cerebrospinal fluid; PCR: Polymerase Chain Reaction.

Figure 2. Phylogenetic tree based on partial VP1 gene sequences obtained in the present study. The sequences with the highest BLAST scores recovered from the NCBI GenBank nucleotide sequence database (http://www.ncbi.nlm.nih.gov/nucleotide/), indicated by BBH for best BLAST hit, for each sequence obtained here (indicated by a label starting by “Sample no.”) were incorporated in the phylogeny reconstruction. Alignment positions corresponded to coordinates 2462–2636 of sequence GenBank accession no. AF132497.1. Nucleotide alignments were performed using the ClustalW program with the BioEdit software v7.0.9.0 [11], and the evolutionary history was inferred in the MEGAX v10.2.6 software (http://www.megasoftware.net/) using the Neighbour-Joining method and the Kimura 2-parameter method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) is 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 scale bars indicate the number of nucleotide substitutions per site. Bootstrap values >50% are labelled on the tree.

Table 4. Univariate and multivariate analysis of potential factors associated with the detection of a pathogen in CSF samples.

Variable	Confirmed CNS infection (N = 99) n (%)	Univariate analysis		Multivariate analysis
		OR [95%CI]	p-value	OR [95%CI]	p-value
Age
<12 months	13 (50.0)	ref	ref	ref	ref
1 - 4 years	17 (73.9)	2.83 [0.85–9.47]	0.09	1.74 [0.45–6.74]	0.42
≥ 5 years	42 (84.0)	5.25 [1.79–15.43]	0.003	3.64 [1.11–11.95]	0.03
Gender
Female	34 (82.9)	ref	ref	ref	ref
Male	38 (65.5)	0.39 [0.12–1.12]	0.06	–	–
Vomiting
No	30 (63.8)	2.38 [0.96–5.92]	0.06	–	–
Yes	42 (80.8)
Lethargic
No	60 (74.1)	0.70 [0.23–2.10]	0.53	–	–
Yes	12 (66.7)
Stiff neck
No	37 (68.5)	1.61 [0.65–3.99]	0.31	–	–
Yes	35 (77.8)
Increase in WBC
No	28 (80.0)	0.55 [0.21–1.47]	0.23	–	–
Yes	44 (68.8)
Increase in CRP
No	22 (73.3)	0.96 [0.36–2.52]	0.93	–	–
Yes	50 (72.5)
WBC in CSF
< 10	5 (31.3)	ref	ref	ref	ref
10 - <100	19 (76.0)	6.97 [1.72–28.25]	0.007	6.38 [1.49–27.24]	0.01
≥ 100	48 (82.8)	10.56 [3.00–37.14]	<0.001	7.90 [2.07–30.13]	0.002
Increase of proteinorachia
No	65 (74.7)	0.47 [0.14–1.65]	0.24	–	–
Yes	7 (58.3)
Decrease of glycorrhachia
No	70 (74.5)	0.23 [0.04–1.45]	0.12	–	–
Yes	2 (40.0)

CRP: C-reactive protein; CSF: cerebrospinal fluid; WBC: white blood cell.

Table 5. Treatment and outcomes of children admitted for central nervous system infection.

Characteristics	N = 99 n (%)
Empiric antimicrobial treatment
Yes	97 (98.0)
No	2 (2.0)
Multiple empiric antibiotics	35 (35.3)
Beta-lactam	17 (17.2)
Cephalosporin	88 (88.9)
Carbapenem	6 (6.1)
Aminoglycosides	21 (21.2)
Glycopeptide	1 (1.0)
Antifungal	1 (1.0)
Duration (days), median (range)	9 (1–39)
Corticosteroids
Yes	60 (60.6)
No	39 (39.4)
Hospitalization duration (days), median (range)	10 (1–40)
Outcomes
Discharge	84 (84.9)
Transfer to national hospital	11 (11.1)
Discharge against medical advice	4 (4.0)
Death	0 (0)

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