Elizabethkingia anophelis Is the Dominant Elizabethkingia Species Found in Blood Cultures in Singapore

Shortly after the discovery of Elizabethkingia anophelis from the mosquito Anopheles gambiae, clinically significant infections attributed to this species were repeatedly described (1); they include a large outbreak involving 69 isolates in Wisconsin, USA (2), and also an intensive-care unit outbreak that occurred at our hospital (3). E. anophelis is genetically distinct from Elizabethkingia meningoseptica and Elizabethkingia miricola (4, 5). However, factory default databases accompanying the matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) identification systems, such as Vitek mass spectroscopy (MS) (bioMérieux, Marcy l’Etoile, France) and the Bruker MALDI Biotyper (Bruker Daltonics, Bremen, Germany), cannot accurately distinguish between E. anophelis and E. meningoseptica unless the user supplies his/her own mass spectrometry data set of characterized isolates (1, 6, 7). In this study, we were interested to see if there were misidentified cases of E. anophelis among our Elizabethkingia spp. Seventy-nine retrospective blood isolates spanning the time period from 2009 to May 2017 were analyzed. All isolates were identified as either E. meningoseptica (96.2%) or E. miricola (3.8%) by the Bruker MALDI Biotyper (bioMérieux). Almost-full-length 16S rRNA gene sequencing was performed using the universal primers fD1 (5=-AGAGTTTGATCCTGGCTCAG-3=) and rP2 (5=-ACGGC TACCTTGTTACGACTT-3=), with sequencing results indicating that 78/79 (98.7%) isolates were E. anophelis and that the remaining single isolate was E. meningoseptica. Specifically, 73 isolates had 99.8 to 100% nucleotide identity to E. anophelis strain 0422, 2 isolates had 100% nucleotide identity to E. anophelis strain 107618, and 1 isolate each had 100% nucleotide identity to E. anophelis strain HKU36, E. anophelis strain F3543, or E. endophytica strain F3201. E. endophytica has since been recognized as an additional strain of E. anophelis and not as a separate species (8). These results reiterate the genomic heterogeneity of this species (5). The E. meningoseptica isolate had 100% 16S sequence identity to E. meningoseptica strain KC1913. Strains NUHP1 (GenBank accession number NZ_CP007547.1) and E. meningoseptica ATCC 13253 (GenBank accession number ASAN00000000.1) were used as controls for both the sequencing and the PCR validation assay. For rapid PCR differentiation between E. anophelis and E. meningoseptica, speciesspecific gene targets were identified based on the comparative analysis of the complete genomes (as of 31 August 2017) of E. anophelis, E. meningoseptica, and E. miricola using the MAUVE software (9). The gene encoding lipid A-disaccharide synthase (GenBank locus tag BD94_RS01570) was utilized for E. anophelis detection with primers anoR (5=-TGCGTTATTACCAGGTAGTCGG-3=) and anoF (5=-GACTTCCGCGGTAGCAAACAA-3=). For the detection of E. meningoseptica, a putative sodium-proton antiporter (GenBank locus tag BBD35_RS10505) was targeted, using primers mengF (5=-TGGGACCTATTGCT GTTGGTT-3=) and mengR (5=-ACCACTTCCTGTGTACCTGC-3=). PCR amplifications were Accepted manuscript posted online 13 December 2017 Citation Chew KL, Cheng B, Lin RTP, Teo JWP. 2018. Elizabethkingia anophelis is the dominant Elizabethkingia species found in blood cultures in Singapore. J Clin Microbiol 56:e01445-17. https://doi.org/10.1128/JCM.01445-17. Editor Sandra S. Richter, Cleveland Clinic Copyright © 2018 American Society for Microbiology. All Rights Reserved. Address correspondence to Jeanette W. P. Teo, Jeanette_Teo@nuhs.edu. sg. LETTER TO THE EDITOR