Classification of Culturable Bifidobacterial Population from Colonic Samples of Wild Pigs (Sus scrofa) Based on Three Molecular Genetic Methods Radko Pechar 1,2 · Jiří Killer 1,3 · Chahrazed Mekadim 1 · Martina Geigerová 1 · Vojtěch Rada 1 Received: 22 May 2017 / Accepted: 25 July 2017 © Springer Science+Business Media, LLC 2017 Abstract Occurrence of bifidobacteria, known as health- promoting probiotic microorganisms, in the digestive tract of wild pigs (Sus scrofa) has not been examined yet. One hundred forty-nine fructose-6-phosphate phosphoketolase positive bacterial strains were isolated from colonic content of twenty-two individuals of wild pigs originated from four localities in the Czechia. Based on PCR-DGGE technique targeting the variable V3 region of the 16S rRNA genes, strains were initially differentiated into four groups repre- sented by: (i) probably a new Bifidobacterium species (89 strains), (ii) B. boum/B. thermophilum/B. thermacidophilum subsp. porcinum/B. thermacidophilum subsp. thermaci- dophilum (sub)species (49 strains), (iii) Pseudoscardovia suis (7 strains), and (iv) B. pseudolongum subsp. globosum/ B. pseudolongum subsp. pseudolongum (4 strains), respectively. Given the fact that DGGE technique did not allow to differentiate the representatives of thermophilic bifidobacteria and B. pseudolongum subspecies, strains were further classified by the 16S rRNA and thrS gene sequences. Primers targeting the variable regions of the latter gene were designed to be applicable in identification and phylogeny of Bifidobacteriaceae family. The 16S rRNA-derived phylogenetic study classified members of the first group into five subgroups in a separated cluster of thermophilic bifidobacteria. Comparable results were obtained by the thrS-derived phylogenetic analysis. Remarkably, variability among thrS sequences was higher compared with 16S rRNA gene sequences. Overall, molecular genetic techniques application allowed to iden- tify a new Bifidobacterium phylotype which is predominant in the digestive tract of examined wild pigs. Introduction Symbiotic intestinal microbiota influences specific function in host nutrient metabolism, maintenance of structural integrity of the gut mucosal barrier, immunomodulation, and protection against pathogens [10]. Many factors such as diet, age, health, host genetics, individuality, geo- graphical location, environment, presence of biologically active substances play an important role in shaping of intestinal microbiota [22]. Family Bifidobacteriaceae represented by bifidobacteria and other fructose-6-phosphate phosphoketolase positive genera is classified into the phylum Actinobacteria [14]. Genus Bifidobacterium is the most numerous one within the family. Representatives of bifidobacteria, which are fre- quently applied and studied as human and animal probiotics [33], can be predominant in intestines of humans [9], some primates [6], cattle [34], and bumblebees [12]. Although the numbers of bifidobacteria in intestines of domestic pigs range only from 10 6 to 10 8 /g of digesta and represent a minor component of the intestinal microbiota Electronic supplementary material The online version of this article (doi:10.1007/s00284-017-1320-0) contains supplementary material, which is available to authorized users. & Jirˇı ´ Killer killer.jiri@seznam.cz; killer@iapg.cas.cz 1 Department of Microbiology, Nutrition and Dietetics, Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences, Kamy ´cka ´ 129, Suchdol, 165 00 Prague 6, Czech Republic 2 Food Research Institute Prague, Radiova ´ 1285/7, Hostivarˇ, 102 00 Prague 10, Czech Republic 3 Institute of Animal Physiology and Genetics v.v.i, Czech Academy of Sciences, Vı ´den ˇska ´ 1083, Krc ˇ, 142 20 Prague 4, Czech Republic 123 Curr Microbiol DOI 10.1007/s00284-017-1320-0