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Background Outbreaks of cyclosporiasis, a diarrheal illness caused by Cyclospora cayetanensis, have been a public health issue in the USA since the mid 1990’s. In 2018, 2299 domestically acquired cases of cyclosporiasis were reported in the USA as a result of multiple large outbreaks linked to different fresh produce commodities. Outbreak investigations are hindered by the absence of standardized molecular epidemiological tools for C. cayetanensis. For other apicomplexan coccidian parasites, multicopy organellar DNA such as mitochondrial genomes have been used for detection and molecular typing. Methods We developed a workflow to obtain complete mitochondrial genome sequences from cilantro samples and clinical samples for typing of C. cayetanensis isolates. The 6.3 kb long C. cayetanensis mitochondrial genome was amplified by PCR in four overlapping amplicons from genomic DNA extracted from cilantro, seeded with oocysts, and from stool samples positive for C. cayetanensis by diagnostic methods. DNA sequence libraries of pooled amplicons were prepared and sequenced via next-generation sequencing (NGS). Sequence reads were assembled using a custom bioinformatics pipeline. Results This approach allowed us to sequence complete mitochondrial genomes from the samples studied. Sequence alterations, such as single nucleotide polymorphism (SNP) profiles and insertion and deletions (InDels), in mitochondrial genomes of 24 stool samples from patients with cyclosporiasis diagnosed in 2014, exhibited discriminatory power. The cluster dendrogram that was created based on distance matrices of the complete mitochondrial genome sequences, indicated distinct strain-level diversity among the 2014 C. cayetanensis outbreak isolates analyzed in this study. Conclusions Our results suggest that genomic analyses of mitochondrial genome sequences may help to link outbreak cases to the source.

Background Cyclospora cayetanensis is an emerging coccidian parasite that causes endemic and epidemic diarrheal disease called cyclosporiasis, and this infection is associated with consumption of contaminated produce or water in developed and developing regions. Food-borne outbreaks of cyclosporiasis have occurred almost every year in the USA since the 1990s. Investigations of these outbreaks are currently hampered due to lack of molecular epidemiological tools for trace back analysis. The apicoplast of C. cayetanensis, a relict non-photosynthetic plastid with an independent genome, provides an attractive target to discover sequence polymorphisms useful as genetic markers for detection and trace back analysis of the parasite. Distinct differences in the apicoplast genomes of C. cayetanensis could be useful in designing advanced molecular methods for rapid detection and, subtyping and geographical source attribution, which would aid outbreak investigations and surveillance studies. Methods To obtain the genome sequence of the C. cayetanensis apicoplast, we sequenced the C. cayetanensis genomic DNA extracted from clinical stool samples, assembled and annotated a 34,146 bp-long circular sequence, and used this sequence as a reference genome in this study. We compared the genome and the predicted proteome to the data available from other apicomplexan parasites. To initialize the search for genetic markers, we mapped the raw sequence reads from an additional 11 distinct clinical stool samples originating from Nepal, New York, Texas, and Indonesia to the apicoplast reference genome. Results We identified several high quality single nucleotide polymorphisms (SNPs) and small insertion/deletions spanning the apicoplast genome supported by extensive sequencing reads data, and a 30 bp sequence repeat at the terminal spacer region in a Nepalese sample. The predicted proteome consists of 29 core apicomplexan peptides found in most of the apicomplexans. Cluster analysis of these C. cayetanensis apicoplast genomes revealed a familiar pattern of tight grouping with Eimeria and Toxoplasma, separated from distant species such as Plasmodium and Babesia. Conclusions SNPs and sequence repeats identified in this study may be useful as genetic markers for identification and differentiation of C. cayetanensis isolates found and could facilitate outbreak investigations. Electronic supplementary material The online version of this article (doi:10.1186/s13071-016-1896-4) contains supplementary material, which is available to authorized users.

Background Horizontal gene transfer is an important source of genetic variation among Neisseria species and has contributed to the spread of resistance to penicillin and sulfonamide drugs in the pathogen Neisseria meningitidis. Sulfonamide resistance in Neisseria meningitidis is mediated by altered chromosomal folP genes. At least some folP alleles conferring resistance have been horizontally acquired from other species, presumably from commensal Neisseriae. In this work, the DNA sequence surrounding folP in commensal Neisseria species was determined and compared to corresponding regions in pathogenic Neisseriae, in order to elucidate the potential for inter-species DNA transfer within this region. Results The upstream region of folP displayed differences in gene order between species, including an insertion of a complete Correia element in Neisseria lactamica and an inversion of a larger genomic segment in Neisseria sicca, Neisseria subflava and Neisseria mucosa. The latter species also had DNA uptake signal sequences (DUS) in this region that were one base different from the DUS in pathogenic Neisseriae. Another interesting finding was evidence of a horizontal transfer event from Neisseria lactamica or Neisseria cinerea that introduced a novel folP allele to the meningococcal population. Conclusion Genetic recombination events immediately upstream of folP and horizontal transfer have resulted in sequence differences in the folP region between the Neisseria species. This variability could be a consequence of the selective pressure on this region exerted by the use of sulfonamide drugs.

Development of sulphonamide resistance in Neisseria meningitidis has been suggested to involve horizontal DNA-transfer from a commensal Neisseria species. In this study, we isolated commensal Neisseria from throat specimens and examined the isolates with respect to sulphonamide resistance. Three resistant clones were identified and the resistance phenotype could be explained by amino acid variations in their dihydropteroate synthase, the target molecule for sulphonamides. Some of these variations occurred in positions corresponding to previously detected variations in resistant N. meningitidis. Sulphonamide resistant commensal Neisseria were isolated from an environment not exposed to sulphonamides, suggesting that resistant Neisseria has become a natural part of the commensal throat flora.