A practical method for preparation of pneumococcal and nontypeable Haemophilus influenzae inocula that preserves viability and immunostimulatory activity

BMC Research Notes

Peter C Richmond³  Andrew J Currie¹  Selma P Wiertsema²  Karli J Corscadden³  Lea-Ann S Kirkham³ 

[1] School of Veterinary and Life Sciences, Murdoch University, South Street, Perth, WA 6150, Australia;School of Paediatrics and Child Health, The University of Western Australia, 35 Stirling Highway, Perth, WA 6009, Australia;Telethon Institute for Child Health Research, Centre for Child Health Research, 100 Roberts Road, Perth, WA 6008, Australia

关键词: PBMCs;    Immunostimulation;    NTHi;    S. pneumoniae;    Cryopreservation;    Live bacteria;    Heat-killed;    Ethanol-killed;   

Others  :  1140535
DOI  :  10.1186/1756-0500-6-522

 received in 2013-08-07, accepted in 2013-12-04,  发布年份 2013

【 摘 要 】

Background

Convenience is a major reason for using killed preparations of bacteria to investigate host-pathogen interactions, however, host responses to such preparations can result in different outcomes when compared to live bacterial stimulation. We investigated whether cryopreservation of Streptococcus pneumoniae and nontypeable Haemophilus influenzae (NTHi) permitted investigation of host responses to infection without the complications of working with freshly prepared live bacteria on the day of experimental challenge.

Findings

S. pneumoniae and NTHi retained >90% viability following cryopreservation in fetal calf serum for at least 8 weeks. Host responses to live, cryopreserved (1 week and 4 weeks), heat-killed or ethanol-killed S. pneumoniae and NTHi were assessed by measuring cytokine release from stimulated peripheral blood mononuclear cells (PBMCs). We found that cryopreserved bacteria, in contrast to heat-killed and ethanol-killed preparations, resulted in comparable levels of inflammatory cytokine release from PBMCs when compared with fresh live bacterial cultures.

Conclusion

Cryopreservation of S. pneumoniae and NTHi does not alter the immunostimulatory properties of these species thereby enabling reproducible and biologically relevant analysis of host responses to infection. This method also facilitates the analysis of multiple strains on the same day and allows predetermination of culture purity and challenge dose.

【 授权许可】

   
2013 Kirkham et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

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