tuf-PCR-temporal temperature gradient gel electrophoresis for molecular detection and identification of staphylococci: Application to breast milk and neonate gut microbiota

• Published in: Journal of microbiological methods Vol. 98; pp. 67 - 75
• Main Authors: Filleron, Anne, Simon, Margaux, Hantova, Stefaniya, Jacquot, Aurélien, Cambonie, Gilles, Marchandin, Hélène, Jumas-Bilak, Estelle
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2014; Elsevier
• Subjects: Bacterial Proteins - genetics; Biochemistry, Molecular Biology; Breast milk; Denaturing Gradient Gel Electrophoresis - methods; Elongation factor Tu gene (tuf); Feces - microbiology; Gastrointestinal Tract - microbiology; Humans; Infant, Newborn; Life Sciences; Microbiology and Parasitology; Microbiota - genetics; Milk, Human - microbiology; Neonate; PCR-temporal temperature gel electrophoresis; Polymerase Chain Reaction - methods; Staphylococcal Infections - diagnosis; Staphylococcal Infections - microbiology; Staphylococcus; Staphylococcus - isolation & purification; Stool; Stool; PCR-temporal temperature gel electrophoresis; CCM; Staphylococcus; CoNS; MALDI-TOF MS; CFU; Neonate; TAE; TTGE; TEMED; Elongation factor Tu gene (tuf); BLAST; DGGE; RFLP; Breast milk; PCR-temporal temperature gel elctrophoresis; Staphylococus aureus; Elongation factor Tu gene
• ISSN: 0167-7012; 1872-8359
• DOI: 10.1016/j.mimet.2013.12.022

Coagulase negative staphylococci (CoNS) are a leading cause of infections in preterm infants, mostly involved in late-onset infection in low birth weight neonates. The epidemiology and pathophysiology of these infections remain unclear, notably because the causing agents are gathered in the artificial CoNS group. The aim of this work was to optimize the study of Staphylococcus species diversity in human breast milk and neonate stool, two sample types with bacterial communities dominated by CoNS, using PCR-temporal temperature gel electrophoresis based on the tuf gene. The optimized protocol identified 18 Staphylococcus species involved in neonate gut microbiota and infections and was applied to cultivation-independent study of breast milk and neonate stool. The efficiency, sensitivity, specificity and species discrimination of the proposed protocol appears suitable for patient follow-up in order to link microbiological data at the community level in milk and stool and interpret them from epidemiological and pathophysiological points of view. •The role of breast milk in gut colonization and neonatal infections is unclear.•We develop a PCR-temporal temperature gel electrophoresis based on tuf gene.•We study species diversity of Staphylococcus in breast milk and in neonate stools.•The tool allows the rapid follow-up of staphylococci in complex microbiotae.•The tool would be useful to explore the pathophysiology of neonatal infections.