Campylobacter  fetusis an important reproductive pathogen of ruminantsthat occasionally infects humans. Here, we describe the complete circularized genomeof a strain ofCampylobacter  fetussubsp.fetusisolated from a sheep. Thefinal assem-bly consisted of a unique contig with a length of 1,849,237 bp

Aráoz, Virginia

Barcellos Coitiño, Maila Sabrina

Caffarena, Rubén Darío

Calleros Basilio, Lucía

Costa, Daniela

Fraga Cotelo, Martín

Giannitti, Federico

Monesiglio, Cecilia

Newton, Irene L.G.

Pérez Crossa, Ruben Gustavo

Silva Silveira, Caroline da

Microbiology Resource Announcements

English

LAReferencia - Red Federada de Repositorios Institucionales de Publicaciones Científicas Latinoamericanas

Complete Genome Sequence of Campylobacter fetus Isolatedfrom a SheepDaniela Costa,a,b Virginia Aráoz,c Maila Barcellos,a Rubén Darío Caffarena,c Martín Fraga,c Federico Giannitti,c Cecilia Monesiglio,cRuben Pérez,a Caroline da Silva Silveira,c Lucía CallerosaaSección Genética Evolutiva, Instituto de Biología, Facultad de Ciencias, Universidad de la República, Montevideo, UruguaybLaboratorio de Genómica Microbiana, Institut Pasteur de Montevideo, Montevideo, UruguaycPlataforma de Investigación en Salud Animal, Instituto Nacional de Investigación Agropecuaria Estación Experimental La Estanzuela, Colonia, UruguayABSTRACT Campylobacter fetus is an important reproductive pathogen of ruminantsthat occasionally infects humans. Here, we describe the complete circularized genomeof a strain of Campylobacter fetus subsp. fetus isolated from a sheep. The final assem-bly consisted of a unique contig with a length of 1,849,237bp.C ampylobacter species are Epsilonproteobacteria adapted to vertebrate hosts. Manyof them cause disease in a wide range of livestock species and have extensive res-ervoirs in wildlife. Campylobacter fetus is an important animal pathogen and an oppor-tunistic human pathogen. It produces considerable economic losses as a major reproduc-tive pathogen of cattle and sheep. This species is currently divided into C. fetus subsp.fetus, C. fetus subsp. venerealis (1), and C. fetus subsp. testudinum (2). C. fetus subsp. fetuscauses abortion mainly in sheep (3) and, to a lesser extent, cattle.Here, we describe the closed whole genome of a strain of C. fetus subsp. fetus iso-lated from a sheep, representing an important resource for evolutionary and epidemio-logical studies of the species.Fresh placenta was received at the Instituto Nacional de Investigación Agropecuaria(INIA) Animal Health Platform (Uruguay) for bacteriological analyses. The sample wasspiked into Skirrow agar and incubated in a microaerobic atmosphere for 48 h at 37°Cusing CampyGen (Oxoid) (4). Biochemical testing was performed, including catalase,oxidase, and hydrogen sulfide production (triple-sugar iron medium), growth in thepresence of 3.5% sodium chloride or 1% glycine, and growth at 25 and 42°C in Skirrowagar (4). Results were consistent with those of Campylobacter fetus subsp. fetus.Bacterial colonies from five petri dishes of pure culture were suspended in 500mlof phosphate-buffered saline solution (pH 7.4), and DNA was extracted using theQIAamp DNA minikit (Qiagen, Inc., Valencia, CA). Whole-genome sequencing (WGS)was performed with Illumina and Pacific Biosciences (PacBio) sequencing technologies.An Illumina library was prepared with the Nextera XT library prep kit without modifica-tions. Illumina reads were obtained using a MiSeq platform at Institut Pasteur deMontevideo, obtaining 1,443,064 2  150-bp paired-end (PE) reads. All bioinformaticanalysis software was set to default values. Read quality was assessed using FastQCv0.11.7 (5). Bases at the ends of the reads with a quality score lower than 20 and readsshorter than 50 bases were trimmed using Trimmomatic v0.39 (TRAILING:20 MINLEN:50)(6). A total of 1,421,381 PE reads remained after filtration (mean length, 145 bases)(Table 1).Long reads were obtained using PacBio RS II single-molecule real-time (SMRT) tech-nology with P6-C4 chemistry (Pacific Biosciences) at Macrogen, Inc. (Seoul, SouthKorea). A library was prepared with the 20-kb SMRTbell template library kit withoutmodifications. DNA was sheared, and 20-kb fragments were selected. PreprocessingCitation Costa D, Aráoz V, Barcellos M,Caffarena RD, Fraga M, Giannitti F, MonesiglioC, Pérez R, da Silva Silveira C, Calleros L. 2020.Complete genome sequence of Campylobacterfetus isolated from a sheep. Microbiol ResourAnnounc 9:e01008-20. https://doi.org/10.1128/MRA.01008-20.Editor Irene L. G. Newton, Indiana University,BloomingtonCopyright © 2020 Costa et al. This is an open-access article distributed under the terms ofthe Creative Commons Attribution 4.0International license.Address correspondence to Lucía Calleros,calleros@fcien.edu.uy.Received 28 August 2020Accepted 20 October 2020Published 5 November 2020November 2020 Volume 9 Issue 45 e01008-20 mra.asm.org 1GENOME SEQUENCESDownloaded from https://journals.asm.org/journal/mra on 16 May 2022 by 2800:a4:31a4:d000:7482:ad70:fa7c:24b7.was done using the SMRT Portal (smrtanalysis_2.3.0.140936.p5.167094) RS Subreads 1protocol. It obtained 1,608,855,393 bases from 150,292 reads, with an average readlength of 10,704 bp and an N50 value of 16,155 bp. After filtering, 97% of these dataremained (Table 1).A hybrid assembly approach was performed with Unicycler v0.4.7 assembler (7),using boldmode. The quality of the assembly was addressed using Quast v4.3 (8).Assembly resulted in a unique 1,849,237-bp contig with an average G1C content of33.3%, as expected for C. fetus subsp. fetus genomes.The sequences were annotated with NCBI Prokaryotic Genome Annotation Pipeline(PGAP v4.13) (9). The genome contains 1,884 coding sequences: 1,811 hypotheticalproteins, 44 tRNA genes, 3 rRNA operons as in all other Campylobacter species, and 2CRISPR arrays.Data availability. These whole-genome sequencing, assembly, and raw data havebeen deposited in DDBJ/ENA/GenBank under the BioProject number PRJNA554155.ACKNOWLEDGMENTSThis work was supported by the Uruguayan Agencia Nacional de Innovacióne Investigación (grant ANII FSSA-105252 and Ph.D. fellowship for D.C.) and theInstituto Nacional de Investigación Agropecuaria (grants N-15156_PL_15_0_00 andN-23398).The funders had no role in study design, data collection and interpretation, or thedecision to submit the work for publication. We declare no conflict of interest.L.C., F.G., M.F., and R.P. conceived the study. V.A. and R.D.C. collected the sample.M.B., C.M., C.D.S.S., and L.C. performed the experiments. D.C. assembled the genome.L.C. and D.C. wrote the manuscript. All the authors revised the manuscript.REFERENCES1. Veron M, Chatelain R. 1973. Taxonomic study of the genus Campylo-bacter Sebald and Veron and designation of the neotype strain for thetype species, Campylobacter fetus (Smith and Taylor) Sebald and Veron.Int J Syst Bacteriol 23:122–134. https://doi.org/10.1099/00207713-23-2-122.2. Fitzgerald C, Tu ZC, Patrick M, Stiles T, Lawson AJ, Santovenia M, Gilbert MJ,van Bergen M, Joyce K, Pruckler J, Stroika S, Duim B, Miller WG, Loparev V,Sinnige JC, Fields PI, Tauxe RV, Blaser MJ, Wagenaar JA. 2014. Campylobacterfetus subsp. testudinum subsp. nov., isolated from humans and reptiles. Int JSyst Evol Microbiol 64:2944–2948. https://doi.org/10.1099/ijs.0.057778-0.3. Sahin O, Yaeger M, Wu Z, Zhang Q. 2017. Campylobacter-associated diseasesin animals. Annu Rev Anim Biosci 5:21–42. https://doi.org/10.1146/annurev-animal-022516-022826.4. OIE. 2008. Bovine genital campylobacteriosis, p 661–670. In Manual ofdiagnostic tests and vaccines for terrestrial animals (mammals, birds andbees), 6th ed, vol 2. OIE, Paris, France.5. Andrews S, Lindenbaum P, Howard B, Ewels P. 2010. FastQC high throughputsequence version 0.11.7. http://www.bioinformatics.babraham.ac.uk/projects/fastqc/.6. Bolger AM, Lohse M, Usadel B. 2014. Trimmomatic: a flexible trimmer forIllumina sequence data. Bioinformatics 30:2114–2120. https://doi.org/10.1093/bioinformatics/btu170.7. Wick RR, Judd LM, Gorrie CL, Holt KE. 2017. Completing bacterial genomeassemblies with multiplex MinION sequencing. Microb Genomics 3:e000132. https://doi.org/10.1099/mgen.0.000132.8. Gurevich A, Saveliev V, Vyahhi N, Tesler G. 2013. QUAST: quality assessmenttool for genome assemblies. Bioinformatics 29:1072–1075. https://doi.org/10.1093/bioinformatics/btt086.9. Tatusova T, Dicuccio M, Badretdin A, Chetvernin V, Nawrocki EP, ZaslavskyL, Lomsadze A, Pruitt KD, Borodovsky M, Ostell J. 2016. NCBI prokaryoticgenome annotation pipeline. Nucleic Acids Res 44:6614–6624. https://doi.org/10.1093/nar/gkw569.TABLE 1 Short- and long-read preprocessing summarySequencing platform Metric Prefilter PostfilterIllumina MiSeq No. of reads 1,443,064 1,421,381Read length range (bp) 30–151 50–151Read Q-score range 21–36 22–36PacBio RS II No. of polymerase read bases 1,608,855,393 1,563,018,757No. of polymerase reads 150,292 101,447Polymerase read N50 (bp) 21,336 21,464Polymerase read length (bp) 10,704 15,407Polymerase read quality 0.583 0.844Costa et al.November 2020 Volume 9 Issue 45 e01008-20 mra.asm.org 2Downloaded from https://journals.asm.org/journal/mra on 16 May 2022 by 2800:a4:31a4:d000:7482:ad70:fa7c:24b7.

Complete genome sequence of Campylobacter fetus isolated from a sheep

https://www.colibri.udelar.edu.uy/jspui/bitstream/20.500.12008/31667/1/10.1128MRA.01008-20.pdf

Complete genome sequence of Campylobacter fetus isolated from a sheep

Abstract

Similar works

Full text

Available Versions

LAReferencia - Red Federada de Repositorios Institucionales de Publicaciones Científicas Latinoamericanas