Downloaded from www.microbiologyresearch.org by IP: 54.70.40.11 On: Wed, 17 Oct 2018 18:10:00 Exiguobacterium alkaliphilum sp. nov. isolated from alkaline wastewater drained sludge of a beverage factory Niha Mohan Kulshreshtha, 1 Rita Kumar, 1 Zareena Begum, 2 S. Shivaji 2 and Anil Kumar 3 Correspondence Anil Kumar anilk@nii.ac.in 1 Institute of Genomics and Integrative Biology, Mall Road, New Delhi 110007, India 2 Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad 500007, India 3 National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110067, India A facultatively anaerobic, alkaliphilic, Gram-stain-positive, rod-shaped bacterium, designated strain 12/1 T , isolated from alkaline wastewater drained sludge of a beverage industry facility located near New Delhi, India, was subjected to a polyphasic taxonomic study. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain 12/1 T belonged to the genus Exiguobacterium and was most closely related to Exiguobacterium aurantiacum DSM 6208 T (99.46 %), E. aquaticum IMTB-3094 T (99.18 %), E. mexicanum 8N T (99.06 %), E. profundum 10C T (98.17 %), E. aestuarii TF-16 T (98.1 %) and E. marinum TF-80 T (98.03 %). The DNA G+C content of strain 12/1 T was 55.6 mol%, major respiratory isoprenoid quinone was MK-7, major polar lipids were phosphatidylglycerol, diphosphatidylglycerol and phosphatidy- lethanolamine and the cell-wall peptidoglycan was of the A3a L-Lys–Gly type, characteristics consistent with its affiliation to the genus Exiguobacterium. Strain 12/1 T showed levels of DNA– DNA hybridization of less than 70 % with the closely related species of the genus Exiguobacterium. Overall, the phenotypic, chemotaxonomic and phylogenetic data presented in this study suggest that strain 12/1 T represents a novel species of the genus Exiguobacterium, for which the name Exiguobacterium alkaliphilum sp. nov. is proposed. The type strain is 12/1 T (5CCM 8459 T 5DSM 21148 T ). The genus Exiguobacterium (type species Exiguobacterium aurantiacum) was described by Collins et al. (1983) representing some slightly alkaliphilic strains isolated from potato wastewater effluent. The genus is characterized by distinct chemotaxonomic properties such as A3a L-Lys–Gly type of peptidoglycan in the cell wall, MK7 as the major respiratory quinone and the presence of diphosphatidyl- glycerol, phosphatidylglycerol and phosphatidylethanola- mine as the major polar lipids. At the time of writing, 14 species with validly published names were included in the genus Exiguobacterium: E. aurantiacum (Collins et al., 1983), E. acetylicum (Farrow et al., 1994), E. antarcticum and E. undae (Fru ¨hling et al., 2002), E. oxidotolerans (Yumoto et al., 2004), E. aestuarii and E. marinum (Kim et al., 2005), E. mexicanum and E. artemiae (Lo ´ pez-Corte ´s et al., 2006), E. indicum (Chaturvedi & Shivaji, 2006), E. sibiricum (Rodrigues et al., 2006), E. profundum (Crapart et al., 2007), E. soli (Chaturvedi et al., 2008) and E. aquaticum (Raichand et al., 2012). In this study, a facultatively alkaliphilic bacterial strain (12/1 T ) capable of neutralizing highly alkaline wastewater which had been isolated from alkaline wastewater drainage sludge of a soft-drink beverage factory located near New Delhi (Kulshreshtha et al., 2010) is described. The 16S rRNA gene sequence comparisons indicated that the isolate belongs to the genus Exiguobacterium. The aim of the present work was to determine the exact taxonomic position of the isolate by detailed characterization combining phenotypic, chemo- taxonomic and phylogenetic analyses. Strain 12/1 T was routinely maintained on alkaline Bacillus medium (ABM) pH 10.5 (Atlas, 2005) or Zobell marine broth 2216 (ZMB; HiMedia). ABM was used for the determination of growth at various temperatures (4, 5, 15, 22, 28, 35, 37, 40 and 42 u C), pH values (5–12 with 0.5-unit intervals) and NaCl concentrations (0–20 % with 0.5 % intervals). The pH of the medium was adjusted as described previously (Kulshreshtha et al., 2010). Cell morphology was examined by light microscopy (E600; Nikon) and transmission electron microscopy. Phenotypic characteristics such as colony morphology and sensitivity The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain 12/1 T is EU379016. A supplementary table is available with the online version of this paper. International Journal of Systematic and Evolutionary Microbiology (2013), 63, 4374–4379 DOI 10.1099/ijs.0.039123-0 4374 039123 G 2013 IUMS Printed in Great Britain