Identification of Intracellular Bacteria in Adenoid and Tonsil Tissue Specimens: The Efficiency of Culture Versus Fluorescent In Situ Hybridization (FISH)

Abstract

Monocyte/macrophage cells from human nasopharyngeal lymphoid tissue can be a source of bacteria responsible for human chronic and recurrent upper respiratory tract infection. Detection and characterization of pathogens surviving intracellularly could be a key element in bacteriological diagnosis of the infections as well as in the study on interactions between bacteria and their host. The present study was undertaken to assess the possibility of isolation of viable bacteria from the cells expressing monocyte/macrophage marker CD14 in nasopharyngeal lymphoid tissue. Overall, 74 adenotonsillectomy specimens (adenoids and tonsils) from 37 children with adenoid hypertrophy and recurrent infections as well as 15 specimens from nine children with adenoid hypertrophy, which do not suffer from upper respiratory tract infections (the control group), were studied. The suitability of immunomagnetic separation for extraction of CD14(+) cells from lymphoid tissue and for further isolation of the intracellular pathogens has been shown. The coexistence of living pathogens including Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pyogenes with the bacteria representing normal nasopharyngeal microbiota inside CD14(+) cells was demonstrated. Twenty-four strains of these pathogens from 32.4 % of the lysates of CD14(+) cells were isolated. Concurrently, the fluorescent in situ hybridization (FISH) with a universal EUB388, and the species-specific probes demonstrated twice more often the persistence of these bacterial species in the lysates of CD14(+) cells than conventional culture. Although the FISH technique appears to be more sensitive than traditional culture in the intracellular bacteria identification, the doubts on whether the bacteria are alive, and therefore, pathogenic would still exist without the strain cultivation.

Cellular Pharmacokinetics and Intracellular Activity of Gepotidacin against Staphylococcus aureus Isolates with Different Resistance Phenotypes in Models of Cultured Phagocytic Cells

“…Pharmacodynamic studies in a model of murine lung infection have demonstrated the area under the concentration-time curve-dependent activity and a pharmacokinetic profile that supports further investigation of this compound for the treatment of infections caused by MRSA ( 30 ). The present study adds that gepotidacin may be of particular interest as an agent that acts upon intracellular S. aureus , which may play a critical role in persistent or recurrent skin or respiratory tract infections ( 21 , 31 , 32 ).…”

Adenotonsillar microbiome: an update

“…The bacteria have been shown to be located in various anatomical locations in the tonsils and adenoids and have been reported intracellularly 6. In biopsies taken from the middle ear in children with otitis media with effusion, S treptococcus pneumoniae and H. influenzae within biofilms on the tissue surface have been observed 18.…”

“…In biopsies taken from the middle ear in children with otitis media with effusion, S treptococcus pneumoniae and H. influenzae within biofilms on the tissue surface have been observed 18. The authors suggest that biofilm-embedded and intracellular pathogens may contribute to the pathogenesis of adenotonsillar hyperplasia 6. However, the localisation of intracellular bacteria is difficult, and only two approaches have so far proved successful.…”

“…The group with adenoid hyperplasia that did not suffer from recurrent infection was designated the control group. After demonstrating that immunomagnetic separation of CD14+ve cells and further isolation of intracellular pathogens was possible, the authors observed the coexistence of viable Staphylococcus aureus, Streptococcus pyogenes and H. influenzae inside CD14+ve cells 6. It was also observed by the use of the fluorescent in situ hybridisation (FISH) with a universal EUB388, and species-specific probes, that intracellular bacteria were detected twice as often than with conventional culture techniques.…”

Chronic tonsillitis, etiological and pathogenetic aspects of the development of metatonsillar complications

“…Культуральными методами не выделяются стрептококки, персистирующие внутриклеточно [21,22]. В исследовании В.Т.…”