Cooccurrence of Five Pathogenic Legionella spp. and Two Free-Living Amoebae Species in a Complete Drinking Water System and Cooling Towers
Abstract
:1. Introduction
2. Results
3. Discussion
4. Materials and Methods
4.1. Site Location and Sampling
4.2. Chemical and Physical Analyses
4.3. Microbiological Analysis
4.4. Water Sample Processing and DNA Extraction
4.5. Molecular Analysis
4.6. Molecular Analysis of Acanthamoeba spp., N. fowleri, General Legionella spp., and Four Pathogenic Legionella Species
4.7. Statistical Analysis
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Free-Living Amoebae | Site Location | ||||
---|---|---|---|---|---|
Res_In | Res_EF | Fa | ERC | CT | |
Acanthamoeba spp. (%) | 0/6 (0%) | 0/6 (0%) | 3/15 (20%) | 0/9 (0%) | 5/6 (83%) |
Acanthamoeba spp. Min, and Max Geomean (Log10 GC/100 mL) | ND | ND | 1.1, 1.4 | ND | 2.0, 2.5 |
N.fowleri (%) | 5/6 (83%) | 1/6 (16%) | 0/15 (0%) | 6/9 (66%) | 5/6 (83%) |
N.fowleri Min, and Max Geomean (Log10 GC/100 mL) | 1.3, 1.7 | 1.5 a | ND | 1.1, 1.8 | 1.3, 2.4 |
Legionella Species | |||||
Legionella spp. (23S rRNA) (%) | (6/6) (100%) | (6/6) (100%) | 15/15 (100%) | 9/9 (100%) | 6/6 (100%) |
Legionella spp. (23S rRNA) Geomean (Log10 GC/100 mL) | 3.1 | 2.7 | 2.3 | 4.3 | 4.5 |
L. pneumophila (%) | 5/6 (83%) | 5/6 (83%) | 6/15 (40%) | 3/9 (33%) | 5/6 (83%) |
L. pneumophila Geomean (Log10 GC/100 mL) | 1.6 | 1.8 | 1.6 | 1.4 | 2.8 |
L. micdadei (%) | 1/6 (17%) | 0/6 (0%) | 3/15 (20%) | 4/9 (44%) | 2/6 (33%) |
L. micdadei Geomean (Log10 GC/100 mL) | 1.5 | ND | 1.5 | 1.6 | 2.4 |
L. bozemanii (%) | 4/6 (67%) | (6/6) (100%) | 2/15 (13%) | 4/9 (44%) | 6/6 (100%) |
L. bozemanii Geomean (Log10 GC/100 mL) | 1.5 | 1.7 | 1.4 | 1.5 | 2.9 |
L. longbeachae (%) | 0/6 (0%) | 0/6 (0%) | 8/15 (53%) | 4/9 (44%) | 3/6 (50%) |
L. longbeachae Geomean (Log10 GC/100 mL) | ND | ND | 1.4 | 1.4 | 1.5 |
L. anisa (%) | 0/6 (0%) | 0/6 (0%) | 5/15 (33%) | 0/6 (0%) | 4/6 (67%) |
L. anisa Geomean (Log10 GC/100 mL) | ND | ND | 1.4 | ND | 2.1 |
Temperature (°C) | Total Chlorine (mg/L) | Free Chlorine (mg/L) | Turbidity NTU | pH | Conductivity (mS) | HPC (CFU/100 mL) | Water Age (h) |
---|---|---|---|---|---|---|---|
RES_IN (N = 6) | |||||||
12.1 | 0 | 0 | 4.1 | 7.2 | 851 | 3.52 × 101 | 4.5 |
RES_EF (N = 6) | |||||||
11.9 | 0.64 | 0.33 | 3.85 | 7.2 | 855 | 2.10 × 100 | 3.4 |
Building F Influent (N = 3) | |||||||
26.8 | 0.41 | 0.35 | 8.4 | 7.3 | 897 | 8.57 × 104 | 9.2 |
Building Fa 1st Floor Cold; N = 3 (Hot Taps; N = 3) | |||||||
26.7 (28.6) | 0.16 (0.04) | 0.14 (0.02) | 3.06 (0.53) | 7.2 (7.1) | 867 (815) | 1.02 × 104 (7.3 × 103) | – |
Building Fa 2nd Floor Cold; N = 3 (Hot Taps; N = 3) | |||||||
26.8 (28.8) | 0.05 (0.02) | 0.03 (0) | 3.37 (0.67) | 7.0 (6.9) | 856 (822) | 2.00 × 104 (3.15 × 103) | – |
Building ERC Influent (N = 3) | |||||||
31.5 | 0.31 | 0.20 | 12.5 | 7.4 | 883 | 4.32 × 105 | 20.8 |
Building ERC 1st Floor Cold (N = 3) (Hot Taps; N = 3) | |||||||
23.5 (24.5) | 0.09 (0.04) | 0.03 (0) | 5.97 (6.27) | 7.6 (7.5) | 866 (847) | 4.38 × 105 (6.80 × 105) | – |
Cooling Towers (N = 6) | |||||||
25.3 | 0.49 | 0.08 | 1.94 | 8.2 | 2564 | 2.35 × 107 | 23.2 |
ap Value = 0.0286 * | L. anisa Present | L. anisa Absent | Total |
---|---|---|---|
Acanthamoeba spp. Present | 4 (9.52%) | 4 (9.52%) | 8 |
Acanthamoeba spp. Absent | 5 (11.90%) | 29 (69.05%) | 34 |
Total | 9 (21.42%) | 33 (78.57%) | 42 |
ap Value = 0.0293 * | L. micdadei Present | L. micdadei Absent | Total |
---|---|---|---|
N.fowleri Present | 7 (16.67%) | 10 (23.81%) | 17 |
N.fowleri Absent | 3 (7.14%) | 22 (52.38%) | 25 |
Total | 10 (23.8%) | 32 (76.19%) | 42 |
ap Value = 0.0099 ** | L. bozemanii Present | L. bozemanii Absent | Total |
---|---|---|---|
N.fowleri Present | 13 (30.95%) | 4 (9.52%) | 17 |
N.fowleri Absent | 9 (21.43%) | 16 (38.10%) | 25 |
Total | 22 (52.38%) | 20 (47.61%) | 42 |
ap Value = 0.0369 * | L. pneumophila Present | L. pneumophila Absent | Total |
---|---|---|---|
N.fowleri Present | 13 (30.95%) | 4 (9.52%) | 17 |
N.fowleri Absent | 11 (26.19%) | 14 (33.33%) | 25 |
Total | 24 (57.14%) | 18 (42.85%) | 42 |
Target Species | Primer/Probe Name | Primer/Probe Sequence | Amplicon Length (bp) | Reference |
---|---|---|---|---|
Acanthamoeba spp. | 18S rRNAF 18S rRNAR 18S rRNAP | 5′-CGACCAGCGATTAGGAGACG-3′ 5′-CCGACGCCAAGGACGAC-3′ 5′-FAM-TGAATACAAAACACCACCATCGGCGC-BHQ1-3′ | 63 | [65] |
N. fowleri | ITSF ITSR ITSP | 5′-GTGAAAACCTTTTTTCCATTTACA-3′ 5′-AAATAAAAGATTGACCATTTGAAA-3′ 5′-HEX-GTGGCCCACGACAGCTTT-BHQ1-3′ | 69 | [66,67] |
Legionella species | 23SF 23SR 23SP | 5′-CCCATGAAGCCCGTTGAA-3′ 5′-ACAATCAGCCAATTAGTACGAG TTAGC-3′ 5′-HEX-TCCACACCTCGCCTATCAACGTCGTAGT-BHQ1-3′ | 92 | [68] |
L. pneumophila | mipF mipR mipP | 5′-AAAGGCATGCAAGACGCTATG-3′ 5′-GAAACTTGTTAAGAACGTCTTTCATTTG-3′ 5′-FAM-TGGCGCTCAATTGGCTTTAACCGA-BHQ1-3′ | 78 | [68] |
L. micdadei L. anisa L. bozemanii L. longbeachae | Pan-Legionella F Pan-Legionella R LmicdadeiP LanisaP Lbozemanii LlongbeachaeP | 5′-GTACTAATTGGCTGATTGTCTTG-3′ 5′-TTCACTTCTGAGTTCGAGATGG-3′ 5′-FAM-AGCTGATTGGTTAATAGCCCAATCGG-BHQ1-3′ 5′-HEX-CTCAACCTACGCAGAACTACTTGAGG-BHQ1-3′ 5′-FAM-TACGCCCATTCATCATGCAAACCAGnT-BHQ1-3′ 5′-HEX-CTGAGTATCATGCCAATAATGCGCGC-BHQ1-3′ | Not available | [69] |
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Logan-Jackson, A.; Rose, J.B. Cooccurrence of Five Pathogenic Legionella spp. and Two Free-Living Amoebae Species in a Complete Drinking Water System and Cooling Towers. Pathogens 2021, 10, 1407. https://doi.org/10.3390/pathogens10111407
Logan-Jackson A, Rose JB. Cooccurrence of Five Pathogenic Legionella spp. and Two Free-Living Amoebae Species in a Complete Drinking Water System and Cooling Towers. Pathogens. 2021; 10(11):1407. https://doi.org/10.3390/pathogens10111407
Chicago/Turabian StyleLogan-Jackson, Alshae, and Joan B. Rose. 2021. "Cooccurrence of Five Pathogenic Legionella spp. and Two Free-Living Amoebae Species in a Complete Drinking Water System and Cooling Towers" Pathogens 10, no. 11: 1407. https://doi.org/10.3390/pathogens10111407