Evidence for Elizabethkingia anophelis transmission from mother to infant, Hong Kong

Abstract

Elizabethkingia anophelis, recently discovered from mosquito gut, is an emerging bacterium associated with neonatal meningitis and nosocomial outbreaks. However, its transmission route remains unknown. We use rapid genome sequencing to investigate 3 cases of E. anophelis sepsis involving 2 neonates who had meningitis and 1 neonate's mother who had chorioamnionitis. Comparative genomics revealed evidence for perinatal vertical transmission from a mother to her neonate; the 2 isolates from these patients, HKU37 and HKU38, shared essentially identical genome sequences. In contrast, the strain from another neonate (HKU36) was genetically divergent, showing only 78.6% genome sequence identity to HKU37 and HKU38, thus excluding a clonal outbreak. Comparison to genomes from mosquito strains revealed potential metabolic adaptations in E. anophelis under different environments. Maternal infection, not mosquitoes, is most likely the source of neonatal E. anophelis infections. Our findings highlight the power of genome sequencing in gaining rapid insights on transmission and pathogenesis of emerging pathogens.

Figure 1

Clinical course of illness in 3 patients infected with Elizabethkingia anophelis in whom sepsis developed and the mother of patient 1, who had culture-negative postpartum fever, Hong Kong, 2012. Locations where patients were treated at the hospital and times when they were home are noted.CSF, cerebrospinal fluid; leaking, leaking of amniotic fluid (membrane rupture).

Figure 2

Comparison of draft genome sequence data of the 3 Elizabethkingia anophelis strains from patients in Hong Kong (HKU36–38), E anophelis type strain R26^T, and E. meningoseptica type strain ATCC 13253^T. A) Distributions of predicted coding sequence function in genomes of E. anophelis strains HKU36–38, E. anophelis type strain R26^T, and E. meningoseptica type strain ATCC 13253T according to SEED Subsystems are shown. The columns indicate the number of proteins in different subsystems. B) Circular representation of sequence comparison between the draft genome of strain HKU37 and other draft genomes as labeled. Comparison generated in Rapid Annotations using Subsystem Technology (27). Intensity of color indicates degree of protein identity.

Figure 3

Phylogenetic trees constructed by using draft genome sequences and concatenated sequences of 69 housekeeping genes of 3 Elizabethkingia anophelis strains from patients in Hong Kong (HKU36–38). A) Neighbor-joining tree constructed on the basis of draft genome sequences using by using Genome-to-Genome Distance Calculator 2.0 (http://ggdc.dsmz.de/distcalc2.php; formula 1) and Chryseobacterium gleum ATCC 35910 as the root. Arrow indicates route of mother-to-neonate transmission. B) Maximum-likelihood tree constructed on the basis of 69 housekeeping genes, showing the relationship of E. anophelis strains HKU36–38 to related bacterial species, using RAxML version 7.2.8 (http://sco.h-its.org/exelixis/software.html) and Weeksella virosa DSM 16922 as the root. A total of 78,520 nt positions were included in the analysis. Bootstrap values were calculated from 1,000 replicates. Scale bars indicate mean number of nucleotide substitutions per site on the respective branches. Gene names and accession numbers are given as cited in GenBank (Technical Appendix Table 2). ‘E. meningoseptica’ strain 502 is a misidentified isolate that actually belongs to E. anophelis on the basis of draft genome sequencing.

Figure 4

Pulsed-field gel electrophoresis (PFGE) analysis of samples from patients in Hong Kong showing 3 Elizabethkingia anophelis strains compared with reference Elizabethkingia isolates. A) PFGE performed by using CHEF Mapper XA system (Bio-Rad, Hercules, CA, USA) and restriction endonuclease XbaI shows that isolates from patient 2 and patient 3 are indistinguishable, wheras isolates from patient 1 possess distinct PFGE patterns. Lane 1, E. anophelis strain HKU37 from uterine swab specimen of patient 2; lane 2, placental swab specimen from patient 2; lane 3, E. anophelis strain HKU38 from blood of patient 3; lane 4, cerebrospinal fluid from patient 3; lane 5, E. anophelis strain HKU36 from blood of patient 1; lane 6, cerebrospinal fluid from patient 1; lane 7, E. anophelis type strain R26^T; lane 8, E. meningoseptica type strain ATCC 13253^T; lane 9, E. miricola type strain LMG22470^T. B) Dendrogram constructed with PFGE data by similarity and clustering analysis using the Dice coefficient (1% tolerance and 0.5% optimization) and the unweighted pair-group method using average linkages with GelCompar II (Applied Maths, Sint-Martens-Latem, Belgium).