The N -Acylneuraminate Cytidyltransferase Gene, neuA , Is Heterogenous in Legionella pneumophila Strains but Can Be Used as a Marker for Epidemiological Typing in the Consensus Sequence-Based Typing Scheme

Abstract

Sequence-based typing (SBT) is the internationally recognized standard method for genotyping Legionella pneumophila. To date all strains of serogroup 1 (SG1) and some of SGs 2 to 14 yield a seven-allele profile and can be assigned a sequence type (ST). However, for some strains belonging to SGs 2 to 14, the targeted region of the neuA gene could not be amplified using the published standard primers. We determined the DNA sequence of a neuA gene homolog located in the lipopolysaccharide synthesis locus of strain Dallas-1E. By using newly designed degenerate consensus primers based on the neuA homolog in strains Dallas-1E, Philadelphia-1, Paris, Lens, and Corby, we were able to obtain DNA sequences for all 48 non-SG1 strains which were untypeable by the standard method. Our data show that the neuA gene is present in all L. pneumophila strains but differs significantly in some non-SG1 strains at both the DNA and amino acid levels. The new primers can be used to amplify and sequence the neuA gene in all strains and can substitute for the standard primers. This offers the possibility of assigning an ST to all strains of L. pneumophila.

Prevalence of Sequence Types among Clinical and Environmental Isolates of Legionella pneumophila Serogroup 1 in the United States from 1982 to 2012

“…In this study, we identified for the first time two sporadic STs, ST1400 and ST1405, that had the neuAh allele previously only found in non-Lp1 strains of L. pneumophila (26). The ESGLI SBT database showed that ST1400 closely relates to ST12, ST150, ST159, ST221, and 1433, all of which are Lp1 STs that differ from ST1400 at only a single allele, the neuA or neuAh.…”

“…Two of these STs, ST1400 and ST1405, carried the N-acylneuraminate cytidyltransferase homolog (neuAh) typically found in non-Lp1 strains of L. pneumophila and not amplified with the standard neuA primers (26). Both ST1400 and ST1405 contained the same neuAh allele, 207, and this appears to be the first instance of the neuAh allele (in contrast to a typical neuA allele) in Lp1 isolates.…”

Sequence types diversity of Legionella pneumophila isolates from environmental water sources in Guangzhou and Jiangmen, China

“…For the strains that failed to get neuA amplification, primers targeting neuAh were used as suggested by ESGLI (Farhat et al, 2011). The PCR products were sent to BGI Beijing Inc. for purification and sequencing.…”

Genotypic diversity of Legionella pneumophila in environmental and clinical strains assessed by Sequence-Based Typing, in association with retrospective clinical surveillance in Northern Italy

“…These include monoclonal antibody (MAb) subgrouping as a rapid screening method, and various genotyping methods, such as restriction fragment length polymorphism (RFLP) analysis, amplified fragment length polymorphism (AFLP), and pulsed-field gel electrophoresis (PFGE). In contrast to band-based typing methods that are difficult to standardize, the sequence-based typing (SBT) system has the potential to achieve excellent typeability, interlaboratory reproducibility and epidemiologic concordance [3,5,6,7].…”