Status: this preprint is open for discussion and under review for Atmospheric Chemistry and Physics (ACP).
Measurement Report: Effects on viability, culturability, and cells fragmentation of two bioaerosol generators during aerosolization of E. coli bacteria
Abstract. Bioaerosol is a significant element of Particulate Matter (PM) and comprises various components, with bacterial species ranking as some of the most important. Reliable and consistent bioaerosol generators are essential for the investigation of bioaerosol in laboratory environments. Aerosol generators are utilized to evaluate the performance of bioaerosol collectors, explore the transport and deposition of biological particles, and study the health impacts and exposure to airborne microorganisms. The main goal of the bacteria experiments is to have an aerosol generator able to aerosolize the maximum number of viable and culturable cells at elevated particle concentrations. This study performs a comparative investigation of two bioaerosol generators: the Sparging Liquid Aerosol Generator (SLAG) by CH Technologies and the 1520 Flow Focusing Monodisperse Aerosol Generator (FMAG) by TSI. The analysis concentrated on the vitality, culturability, fragmentation, and nebulization efficiency of E. coli cells. The results indicated increased fragmentation using the SLAG nebulizer, and the size distribution varied according to the concentration of the injection fluid for FMAG. Both nebulizers imposed significant stress on bacteria during nebulization, halving their viability. Ultimately, the nebulization efficiency of FMAG is twenty times higher than that of SLAG.
Received: 12 Nov 2025 – Discussion started: 19 Nov 2025
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This work investigates the impact of different aerosolization techniques on E.coli within an atmospheric simulation chamber. The topic is surely interesting since it is relevant to many experimental protocols. Also, the analysis is quite thorough and bacteria viability is analyzed in multiple ways to better assess the impact of the aerosolization techniques. The reviewer suggests publication of the paper with only some minor modification as expressed below:
P. 3, Line 85: I suggest reporting also the aerodynamic diameter of E. coli (which should be smaller than 2 um) to make clearer the following choice of 0.5 um as a cut-off for the fragmentation range.Â
P.5-6: Were the aerosol generators cleaned between replicates to avoid a carry-over effect in bacterial cells?
P.5-6: While the droplet diameter of the FMAG is reported (0.8-12 um) the one from the SLAG is not, is there an approximate size of the droplets and is it compatible with E.coli size?
P. 9: The WIBS peak is centered around 0.8 um vs. the OPS peak at 0.5 um. Couldn't this be simply an effect on how the different samples define the particles' binning?Â
P. 9: Beside peak shifting WIBS seems to exhibit a higher (i.e.: non-overlapping error bars) maximum normalized concentration compared with OPS. Is this related to total particles counted by WIBS or to fluorescent-only particles? If so is that explainable by the dimensional shift or what alse could explain a higher number of fluorescent particles vs. total ones?
P. 10. Figure 3 is missing x-axis description (Midpoint bin (um)).
P. 14, L. 297-301. While the presented study is surely valuable and a thorough characterization, it is far from being the first step in this field. For example Thomas et al. (2011) already provided data on survival and site of damage of E. coli nebulized with different techniques. Rather, I think that the major strength of this work is the completeness of the analysis which goes way beyond simple viability (in a culturable sense). I suggest a rephrasing of this sentence.
Finally, this is not a comment on the quality of the paper itself, just an potential outlook for future work. Gram negative and gram positive bacteria differ in their structure, it would be interesting in the future to see if this also translates in differences when nebulized (in terms of viability, etc.). Â
Cited literature:
Thomas RJ Webber D, Hopkins R, Frost A, Laws T, Jayasekera PN, Atkins T.2011.The Cell Membrane as a Major Site of Damage during Aerosolization of Escherichia coli . Appl Environ Microbiol. 77:.https://doi.org/10.1128/AEM.01116-10
Effects on viability, culturability, and cells fragmentation of two bioaerosol generators during aerosolization of E. coli bacteriaFederico Mazzei, Marco Brunoldi, Elena Gatta, Franco Parodi, Paolo Prati, Virginia Vernocchi, and Dario Massabò https://doi.org/10.17632/j962hdhc96.1
The goal of this study was to compare the performance of two aerosol generators in terms of nebulization efficiency, size distribution, bacterial fragmentation, and the stresses produced on bacteria during nebulization, including loss of viability and culturability. Our findings indicated that bacterial viability is significantly influenced by nebulization procedures, and nebulization efficiency varies among different nebulizers.
The goal of this study was to compare the performance of two aerosol generators in terms of...
This work investigates the impact of different aerosolization techniques on E.coli within an atmospheric simulation chamber. The topic is surely interesting since it is relevant to many experimental protocols. Also, the analysis is quite thorough and bacteria viability is analyzed in multiple ways to better assess the impact of the aerosolization techniques. The reviewer suggests publication of the paper with only some minor modification as expressed below:
P. 3, Line 85: I suggest reporting also the aerodynamic diameter of E. coli (which should be smaller than 2 um) to make clearer the following choice of 0.5 um as a cut-off for the fragmentation range.Â
P.5-6: Were the aerosol generators cleaned between replicates to avoid a carry-over effect in bacterial cells?
P.5-6: While the droplet diameter of the FMAG is reported (0.8-12 um) the one from the SLAG is not, is there an approximate size of the droplets and is it compatible with E.coli size?
P. 9: The WIBS peak is centered around 0.8 um vs. the OPS peak at 0.5 um. Couldn't this be simply an effect on how the different samples define the particles' binning?Â
P. 9: Beside peak shifting WIBS seems to exhibit a higher (i.e.: non-overlapping error bars) maximum normalized concentration compared with OPS. Is this related to total particles counted by WIBS or to fluorescent-only particles? If so is that explainable by the dimensional shift or what alse could explain a higher number of fluorescent particles vs. total ones?
P. 10. Figure 3 is missing x-axis description (Midpoint bin (um)).
P. 14, L. 297-301. While the presented study is surely valuable and a thorough characterization, it is far from being the first step in this field. For example Thomas et al. (2011) already provided data on survival and site of damage of E. coli nebulized with different techniques. Rather, I think that the major strength of this work is the completeness of the analysis which goes way beyond simple viability (in a culturable sense). I suggest a rephrasing of this sentence.
Finally, this is not a comment on the quality of the paper itself, just an potential outlook for future work. Gram negative and gram positive bacteria differ in their structure, it would be interesting in the future to see if this also translates in differences when nebulized (in terms of viability, etc.). Â
Cited literature:
Thomas RJ Webber D, Hopkins R, Frost A, Laws T, Jayasekera PN, Atkins T.2011.The Cell Membrane as a Major Site of Damage during Aerosolization of Escherichia coli . Appl Environ Microbiol. 77:.https://doi.org/10.1128/AEM.01116-10