Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods.
The study aimed to delineate the robustness of the culture-based and molecular biology methods to assess the total bacterial concentration and Campylobacter jejuni (C. jejuni) quantification in caecal content, analysed as fresh or after being stored immediately at ultra-low (-80°C) temperature at di...
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2022-01-01
|
| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0274682 |
| _version_ | 1811180729466355712 |
|---|---|
| author | Farina Khattak Salvatore Galgano Jos Houdijk |
| author_facet | Farina Khattak Salvatore Galgano Jos Houdijk |
| author_sort | Farina Khattak |
| collection | DOAJ |
| description | The study aimed to delineate the robustness of the culture-based and molecular biology methods to assess the total bacterial concentration and Campylobacter jejuni (C. jejuni) quantification in caecal content, analysed as fresh or after being stored immediately at ultra-low (-80°C) temperature at different time points (for 3, 7, 14, 28 and 62 days post collection). The caecal content was collected from birds that were artificially colonised with C. jejuni (in-vivo), and quantification was performed using both colony-forming unit (CFU) and qPCR. The results showed that storage time affected the output of culture-based analyses but mostly did not alter concentration retrieved via qPCR. After an initial ~4.5 log10 reduction in CFU observed from fresh (day 0) to frozen samples, bacterial concentration retrieved with culture-based methods seemed to be constant in samples frozen for 3 to 62 days, indicating a possible threshold for C. jejuni loss of viability due to effect of storage temperature. Ranking order analyses, revealed that the molecular biology technique was able to attribute somewhat the same relative C. jejuni concentrations to the samples analysed via qPCR. However, day 0 measurements from culture-based methods were associated with the absence of or negatively weak correlations with the rest of the time points, but ranking order was maintained from day 3 onwards. On the other hand, ranking order correlations were less constant when measuring total bacterial concentration through qPCR. The study suggests that if biological samples can't be analysed as fresh (immediately after collection) and have to be stored prior to analysis, then storage at -80°C samples be recommended to avoid the temporal-dependent effects on C. jejuni concentrations. In addition, irrespective of the method of analysis, an initial loss of CFU must be factored in when interpreting the results obtained from frozen samples. |
| first_indexed | 2024-04-11T09:08:14Z |
| format | Article |
| id | doaj.art-5c16274c253f4c02997878811d50a80d |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-04-11T09:08:14Z |
| publishDate | 2022-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-5c16274c253f4c02997878811d50a80d2022-12-22T04:32:34ZengPublic Library of Science (PLoS)PLoS ONE1932-62032022-01-01179e027468210.1371/journal.pone.0274682Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods.Farina KhattakSalvatore GalganoJos HoudijkThe study aimed to delineate the robustness of the culture-based and molecular biology methods to assess the total bacterial concentration and Campylobacter jejuni (C. jejuni) quantification in caecal content, analysed as fresh or after being stored immediately at ultra-low (-80°C) temperature at different time points (for 3, 7, 14, 28 and 62 days post collection). The caecal content was collected from birds that were artificially colonised with C. jejuni (in-vivo), and quantification was performed using both colony-forming unit (CFU) and qPCR. The results showed that storage time affected the output of culture-based analyses but mostly did not alter concentration retrieved via qPCR. After an initial ~4.5 log10 reduction in CFU observed from fresh (day 0) to frozen samples, bacterial concentration retrieved with culture-based methods seemed to be constant in samples frozen for 3 to 62 days, indicating a possible threshold for C. jejuni loss of viability due to effect of storage temperature. Ranking order analyses, revealed that the molecular biology technique was able to attribute somewhat the same relative C. jejuni concentrations to the samples analysed via qPCR. However, day 0 measurements from culture-based methods were associated with the absence of or negatively weak correlations with the rest of the time points, but ranking order was maintained from day 3 onwards. On the other hand, ranking order correlations were less constant when measuring total bacterial concentration through qPCR. The study suggests that if biological samples can't be analysed as fresh (immediately after collection) and have to be stored prior to analysis, then storage at -80°C samples be recommended to avoid the temporal-dependent effects on C. jejuni concentrations. In addition, irrespective of the method of analysis, an initial loss of CFU must be factored in when interpreting the results obtained from frozen samples.https://doi.org/10.1371/journal.pone.0274682 |
| spellingShingle | Farina Khattak Salvatore Galgano Jos Houdijk Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods. PLoS ONE |
| title | Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods. |
| title_full | Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods. |
| title_fullStr | Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods. |
| title_full_unstemmed | Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods. |
| title_short | Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods. |
| title_sort | bacterial concentration and campylobacter spp quantification differ when fresh or ultra frozen samples are analysed over time using molecular biology and culture based methods |
| url | https://doi.org/10.1371/journal.pone.0274682 |
| work_keys_str_mv | AT farinakhattak bacterialconcentrationandcampylobactersppquantificationdifferwhenfreshorultrafrozensamplesareanalysedovertimeusingmolecularbiologyandculturebasedmethods AT salvatoregalgano bacterialconcentrationandcampylobactersppquantificationdifferwhenfreshorultrafrozensamplesareanalysedovertimeusingmolecularbiologyandculturebasedmethods AT joshoudijk bacterialconcentrationandcampylobactersppquantificationdifferwhenfreshorultrafrozensamplesareanalysedovertimeusingmolecularbiologyandculturebasedmethods |