Campylobacter

Cam-porA dendogram fragment
Typing Services
Cam-porA
Cam-flaA
Cam-porA / Cam-flaA

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Campylobacter species are thermophilic, microaerophilic, motile Gram-negative bacteria that commonly colonize the intestinal tracts of various animal species. Consequently, they frequently contaminate raw meat, and occassionally unpasterized milk and public water supplies. Ingestion results in gastroenteritis which is generally mild and self-limiting but may be accompanied by bloody diarrhea, and some cases are severe enough to require hospitalization. Moreover, roughly 1 in 3000 infections lead to autoimmune-based neuropathologies, particularly Guillian-Barré syndrome (GBS), due to antigenic similarities between surface lipooligosaccharides on some Campylobacter strains and gangliosides on human peripheral nerves. Since their culture requires specialized media and incubation conditions, it is only in recent years that Campylobacter species have been recognized as the leading cause of bacterial diarrhea in the U.S.

 

Although multiple Campylobacter species colonize birds and mammals, it has long been known that only two, C. jejuni and C. coli, are responsible for human illness (90% and 10%, respectively). Recognition of retail chicken as the primary source of these infections, however, required the development of molecular typing systems and their application to epidemiological studies. As recently reviewed (1), while conventional serological and gel pattern-based methods (e.g., PFGE) have recognized limitations, multilocus sequence typing (MLST) of conserved housekeeping genes has proved useful in identifying pathogenic Campylobacter ‘clones’ or ‘lineages’. For epidemiological analysis of outbreaks, however, the higher strain resolution afforded by sequence analysis of polymorphic antigen-encoding genes is required. The first gene used for this purpose was flagellin-encoding flaA, and more recently the major outer membrane protein-encoding porA. Sequence analysis of variable regions within these genes is used to define alleles with reference to a curated public database (2). Unfortunately, the MLST, flaA, and porA sequence types are arbitrarily numbered in order of database entry, and consequently relationships among strains can be difficult to discern.

 

MicrobiType PLST services for Campylobacter typing, Cam-porA and Cam-flaA, are based on these well validated targets, but go further by expanding their sequence coverage 1.4 to 2.4-fold, respectively. Furthermore, results are reported in terms of both numerical sequence type and more readily interpretable dendrogram.

 

As shown in the Cam-porA dendrogram, for example, all 21 C. jejuni and C. coli strains with fully sequenced genomes (from the NCBI Nucleotide database) are resolved, including strains S3 and 32488 which share porA type 11 but differ in their flanking sequences. Additional strengths of Cam-porA typing evident in this dendrogram include: (a) C. jejuni and C. coli form clearly distinct clusters; (b) passage of C. jejuni strain 11168GSv through a human (following accidental lab infection) and subsequently through a mouse (3) demonstrate that the porA sequence, despite its polymorphism, is epidemiologically stable, consistent with previous studies (4); (c) addition to the Cam-porA dendrogram of three additional ‘Walkerton’ strains (5) from the partially sequenced genome database WGS illustrate its potential for outbreak detection and investigation; and (d) GBS-associated strains from an outbreak in China (6) cluster together, along with a GBS-associated strain from South Africa (7).

Cam-flaA typing provides nearly equivalent resolution, as demonstrated in the Cam-flaA dendrogram (20 of 21 strains with fully sequenced genomes resolved). Again, stability is evidenced by clustering of the lab, human, and mouse isolates of strain 11168, and a GBS-associated strain cluster including China and South Africa isolates is present. (The WGS database lacks flaA sequences for the additional Walkerton outbreak strains and two of the GBS-associated strains, due to inherent limitations of ‘next generation’ sequencing technologies with respect to tandem repeats.) Two strain pairs sharing flaA sequence type (36 and 100) are resolved in Cam-flaA typing due to its extended sequence coverage. Note, however, that C. jejuni and C. coli  strains fail to cluster, consistent with previous studies showing that flaA is poorly correlated with Campylobacter evolution (1). Nevertheless, Cam-flaA typing can provide a useful complement to Cam-porA since it provides an independent test of strain relatedness (when ordered together, a discount is applied).

In addition to sequence type and dendrogram, a sequence alignment is provided illustrating relatedness of the submitted isolate to previously or concurrently submitted isolates and representative database strains.

 

1.  Colles & Maiden, 2012, Microbiology 158:2695
2.  Jolley & Maiden, 2010, BMC Bioinformatics 11:595 (http://pubmlst.org/campylobacter)
3.  Thomas et al., 2014, PLoS One 9:E88229
4.  Cody et al., 2009, Microbiology 155:4145.
5.  Clark et al., 2005, J Clin Microbiol 43:2080
6.  Zhang et al., 2013, Genome Announc 1:e00256
7.  Quiñones et al., 2008, PLoS One 3:e2015

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