Table 2.
Advantages and limitations of various enumeration techniques.
| Bioaerosol sampling techniques | Advantages of the technique | Limitations of the technique |
|---|---|---|
| Culturable method | ||
| Classical microscopy | Cost effective and easy to handle | Only viable and culturable micro-organisms can be identified and not nonviable bioaerosol |
| Can be used to identify specific taxa of micro-organisms | Thus do not act as a representative of the microorganisms in the bioaerosol | |
| Poor precision of measurement. | ||
| Most probable number | Relatively swift and easy to perform | Being a statistical test it does not measure actual numbers of micro-organisms. |
| As the micro-organisms are grown in liquid media such technique is less susceptible to the culturability issues that affect selective isolation plate methods . |
Aggregates of cells may affect the result, thereby limiting the suitability of this method to analysis of bioaerosols. | |
| LIF | Sensitive | Sometimes difficult to quantify due to collisional quenching of the excited state and potential photochemical effects. |
| Measurements are spatially resolved and can be further extended to laser imaging. | Not all excited species fluoresces causing improper measurements. | |
| MALDI-TOF | Cheap technique and easy to operate | The compound (such as proteins) to be analyzed should be in the databases. |
| Highly sensitive | This technique is generally not suitable for compounds less than 600 Da in size due to intense matrix signal. | |
| Very mild ionization technique used, thereby making analysis of mixture possible | There is limitation in the resolution of this technique which can only be increased significantly by a reflector and or a delayed extraction. | |
| LIBS | Very little or no sample preparation is required that results in increased throughput, greater convenience and fewer opportunities for contamination to occur. | Limited usage due to increased cost and system complexity |
| Very sensitive and requires very small amount of sample (thus sometimes referred to as “nondestructive” method) | Sometimes regarded as semi quantitative technique as obtaining suitable standards is difficult. | |
| Possibility of multi-elemental analysis simultaneously | There are possibilities of large interference effects that include matrix interference as well as potential interference of particle size in case of aerosol. | |
| Has the potential for direct detection in aerosols | Less precision ranging from 5 to 10% depending upon excitation properties of laser, sample homogeneity and sample matrix | |
| Simple process with rapid analytical capability as in a single step ablation and excitation process is carried out. | ||
| Nonculturable methods | ||
| Epifluorescence microscopy | Both culturable and nonculturable cells can be counted making the results more representative of total numbers of micro-organisms in the bioaerosol. | Restricted ability to identify specific taxa of micro-organisms |
| Relatively cheap operating costs | Fluorochromes if binds to abiotic particles may result into false positive results. | |
| High throughput of samples possible if image analysis system used | Image analysis system may count abiotic particles within the same size parameters as microbial cells. | |
| Not suitable for counting aggregates of cells | ||
| Overestimation due to binding to abiotic | ||
| Material may take place | ||
| PCR technique | Remarkably sensitive technique | The efficiency and size ranges of bioaerosol high volume samplers should be completely characterized which can otherwise affect the quantification by Quantitative PCR. |
| Applicable to any biological matter containing nucleic acid | Possibility of inaccurate bioaerosol quantification due to improper sample preparation steps like filter elution/concentration and nucleic acid extraction | |
| Detection and identification can be made independent of culturing thereby removing the need of specialized labs to perform cell cultures which require extensive biosafety infrastructure. | Results may get affected by the presence of inhibitory PCR compounds in the samples. | |
| Results are provided rapidly on the order of hours as compared to days or weeks. | ||
| Flow cytometry | Same as for epifluorescence microscopy | Same as for epifluorescence microscopy |
| Next generation sequencing | Very sensitive technique | In general experiment runs at al large scale |
| Can be applied to any biological sample containing nucleic acid. | Has high startup cost | |
| A significantly quicker sequencing technique for DNA and RNA in comparison to the traditional ones | Multiple days of run time is usually required (except for 454 Roche sequencing) | |
| Works on short read lengths (usually < 600 bp) thereby limiting the phylogenetic characterization | ||
| DGGE | Simultaneous analysis of multiple samples are possible. | Time consuming technique |
| Can monitor shift in the community with passage of time | Multiple bands of single species may result in overestimation of community diversity due to microheterogeneity of rRNA. | |
| Is very sensitive to any form of variation in DNA sequence | Is a semi quantitative technique due to variation in 16S rRNA gene copy number in different species | |
| Can analyze any microbial community without prior knowledge of species due to universal primers | Limits phylogenetic characterization as it works with only short fragments | |
| GC clamp can be variable every time it is synthesized potentially causing same 16S rRNA having different DGGE profiles. | ||
| Biomarkers | Certain taxa of micro-organisms can be identified. | No standard approach available for monitoring of biomarkers so as to provide certain information. |
| As whole cells are not measured, this technique is not prone to many of the limitations of culturable or nonculturable methods | LAL assay types biomarker tests which are significantly used in bacterial bioaerosol analysis are affected by dust or other microbial cell components. This is likely to be a significant problem. | |
LIF, Laser Induced Fluorescence; MALDI-TOF, Matrix-assisted laser desorption/Ionization time of flight; LIB, Laser induced breakdown Spectroscopy.