title: Multiplex PCR Assay for Identifi cation and Differentiation of Campylobacter jejuni and Campylobacter coli Isolates.
abstract: Campylobacter spp. are important causative agents of gastrointestinal infections in humans. The most frequently isolated strains of this bacterial genus are Campylobacter jejuni and Campylobacter coli. To date, genetic methods for bacterial identification have not been used in Bulgaria. We optimized the multiplex PSR assay to identify Campylobacter spp. and differentiate C. jejuni from C. coli in clinical isolates. We also compared this method with the routinely used biochemical methods.
  
  
  
  To identify Campylobacter spp. and discriminate C. coli from C. jejuni in clinical isolates using multiplex PCR assay.
  
  
  
  Between February 2014 and January 2015 we studied 93 stool samples taken from patients with diarrheal syndrome and identified 40 species of Campylobacter spp. in them. The clinical material was cultured in microaerophilic atmosphere, the isolated strains being biochemically diff erentiated (hydrolysis of sodium hippurate for C. jejuni, and hydrolysis of indoxyl acetate for C. coli). DNA was isolated from the strains using QiaAmp MiniKit (QIAGEN, Germany). Twenty strains were tested with multiplex PCR for the presence of these genes: cadF, characteristic for Campylobacter spp., hipO for C. jejuni and asp for C. coli.
  
  
  
  The biochemical tests identified 16 strains of C. jejuni, 3 strains of C. coli, and 1 strain of C. upsaliensis. After the multiplex PCR assay the capillary gel electrophoresis confirmed 16 strains of C. jejuni, 2 strains of C. coli and 2 strains of Campylobacter spp. - because of the presence of the gene cadF. C. jejuni has the gene hipO, and it is possible that this gene may not be expressed in the biochemical differentiation yielding a negative reaction as a result. In comparison, we can conclude that the genetic differentiation is a more accurate method than the biochemical tests.
  
  
  
  The multiplex PCR assay is a fast, accurate method for identifi cation of Campylobacter spp. which makes it quite necessary in the clinical diagnostic practice.
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Maria R
authors.initials: MR
authors.lastname: Pavlova
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Elina G
authors.initials: EG
authors.lastname: Dobreva
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Katucha I
authors.initials: KI
authors.lastname: Ivanova
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Galina D
authors.initials: GD
authors.lastname: Asseva
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Ivan N
authors.initials: IN
authors.lastname: Ivanov
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Peter K
authors.initials: PK
authors.lastname: Petrov
authors.affiliation: Hospital for Infectious and Parasitic Diseases (HIPD), Medical University of Sofia, Bulgaria
authors.forename: Valeri R
authors.initials: VR
authors.lastname: Velev
authors.affiliation: Hospital for Infectious and Parasitic Diseases (HIPD), Medical University of Sofia, Bulgaria
authors.forename: Ivelina I
authors.initials: II
authors.lastname: Tomova
authors.affiliation: Hospital for Infectious and Parasitic Diseases (HIPD), Medical University of Sofia, Bulgaria
authors.forename: Maida M
authors.initials: MM
authors.lastname: Tiholova
authors.affiliation: National Center of Infectious and Parasitic Diseases (NCIPD), Sofi a, Bulgaria
authors.forename: Todor V
authors.initials: TV
authors.lastname: Kantardjiev
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D015373:Bacterial Typing Techniques; D002031:Bulgaria; D002169:Campylobacter Infections / Q000175:diagnosis / Q000382:microbiology*; D017000:Campylobacter coli / Q000235:genetics*; D016123:Campylobacter jejuni / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics; D002648:Child; D002675:Child, Preschool; D003967:Diarrhea / Q000175:diagnosis / Q000382:microbiology*; D005243:Feces / Q000382:microbiology; D005260:Female; D006801:Humans; D007223:Infant; D008297:Male; D060885:Multiplex Polymerase Chain Reaction
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; C107872:CadF protein, Campylobacter jejuni; D002352:Carrier Proteins