Abstract
This study investigated the occurrence, species, infectivity and removal efficiency of Cryptosporidium spp. across typical wastewater treatment train. Samples from different process units were collected seasonally and synchronously from four wastewater treatment plants (WWTPs) in Northeastern China. Live Cryptosporidium oocysts were identified in most samples from both influent (97.50%) and effluent (90.00%) wastewaters of the four WWTPs, at an average density of 26.34 and 4.15 oocysts/L, respectively. The overall removal efficiency was 84.25%, and oocysts were mainly removed (62.01%) by the modified secondary sedimentation process. Ten Cryptosporidium species were identified in the effluent samples. C. andersoni, C. bovis, and C. ryanae were the three most prevalent species. Oocyst viability assays indicated no reduction of excystation rate during the primary and secondary wastewater treatments (varied in the range of 63.08%–68.50%), but the excystation rate declined to 52.21% in the effluent after disinfection. Notably, the Cryptosporidium oocysts showed higher infection intensity in the cold season (winter and spring) than that in summer and autumn. The influences of environmental temperature on virulence factors of Cryptosporidium were further examined. It was observed that more extracellular secretory proteins were bound on the oocyst surface and several virulence genes were expressed relatively strongly at low temperatures, both of which could facilitate oocyst adhesion, invasion, and host immune evasion. This research is of considerable interest since it serves as an important step towards more accurate panoramic recognition of Cryptosporidium risk reduction in WWTPs, and especially highlights the potential health risk associated with Cryptosporidium in cold regions/seasons.
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References
Adeyemo F E, Singh G, Reddy P, Bux F, Stenström T A (2019). Efficiency of chlorine and UV in the inactivation of Cryptosporidium and Giardia in wastewater. PLoS One, 14(5): e0216040
Andreoli F C, Sabogal-Paz L P (2019). Coagulation, flocculation, dissolved air flotation and filtration in the removal of Giardia spp. and Cryptosporidium spp. from water supply. Environmental Technology, 40(5): 654–663
Arrowood M J, Sterling C R (1987). Isolation of Cryptosporidium oocysts and sporozoites using discontinuous sucrose and isopycnic Percoll gradients. Journal of Parasitology, 73(2): 314–319
Barudin M A, Isa M L M, Yusof A M (2018). Signal peptide sequence analysis of selected protein sequences from Cryptosporidium parvum. Trends in Bioinformatics, 11(1): 33–43
Bouzid M, Hunter P R, Chalmers R M, Tyler K M (2013). Cryptosporidium pathogenicity and virulence. Clinical Microbiology Reviews, 26(1): 115–134
Bradford S A, Kim H, Headd B, Torkzaban S (2016). Evaluating the transport of Bacillus subtilis spores as a potential surrogate for Cryptosporidium parvum oocysts. Environmental Science & Technology, 50(3): 1295–1303
Cama V A, Ross J M, Crawford S, Kawai V, Chavez-Valdez R, Vargas D, Vivar A, Ticona E, Navincopa M, Williamson J, Ortega Y, Gilman R H, Bern C, Xiao L (2007). Differences in clinical manifestations among Cryptosporidium species and subtypes in HIV-infected persons. The Journal of Infectious Diseases, 196(5): 684–691
Castro-Hermida J A, García-Presedo I, Almeida A, González-Warleta M, Da Costa J M, Mezo M (2009). Detection of Cryptosporidium spp. and Giardia duodenalis in surface water: A health risk for humans and animals. Water Research, 43(17): 4133–4142
Chauret C P, Radziminski C Z, Lepuil M, Creason R, Andrews R C (2001). Chlorine dioxide inactivation of Cryptosporidium parvum oocysts and bacterial spore indicators. Applied and Environmental Microbiology, 67(7): 2993–3001
Dreelin E A, Ives R L, Molloy S, Rose J B (2014). Cryptosporidium and Giardia in surface water: A case study from Michigan, USA to inform management of rural water systems. International Journal of Environmental Research and Public Health, 11(10): 10480–10503
Edzwald J K, Tobiason J E (2002). Fate and removal of Cryptosporidium in a dissolved air flotation water plant with and without recycle of waste filter backwash water. Water Science and Technology: Water Supply, 2(2): 85–90
Falohun O O, Ayinmode A B, Adejinmi J O (2021). Molecular characterisation of Cryptosporidium isolates from rivers, water treatment plants and abattoirs in Ibadan, Nigeria. Comparative Immunology, Microbiology and Infectious Diseases, 74: 101577
Fan W, Yang X, Wang Y, Huo M (2020). Loopholes in the current reclaimed water quality standards for clogging control during aquifer storage and recovery in China. Water Cycle, 1: 13–18
Farrell C, Hassard F, Jefferson B, Leziart T, Nocker A, Jarvis P (2018). Turbidity composition and the relationship with microbial attachment and UV inactivation efficacy. Science of the Total Environment, 624: 638–647
Fayer R, Orlandi P, Perdue M L (2009). Virulence factor activity relationships for hepatitis E and Cryptosporidium. Journal of Water and Health, 7(S1 Suppl 1): S55–S63
Foster T J, Geoghegan J A, Ganesh V K, Höök M (2014). Adhesion, invasion and evasion: the many functions of the surface proteins of Staphylococcus aureus. Nature Reviews. Microbiology, 12(1): 49–62
Fu C Y, Xie X, Huang J J, Zhang T, Wu Q Y, Chen J N, Hu H Y (2010). Monitoring and evaluation of removal of pathogens at municipal wastewater treatment plants. Water Science and Technology, 61(6): 1589–1599
Galván A L, Magnet A, Izquierdo F, Fernández Vadillo C, Peralta R H, Angulo S, Fenoy S, del Aguila C (2014). A year-long study of Cryptosporidium species and subtypes in recreational, drinking and wastewater from the central area of Spain. Science of the Total Environment, 468–469(468–469): 368–375
Gharpure R, Perez A, Miller A D, Wikswo M E, Silver R, Hlavsa M C (2019). Cryptosporidiosis outbreaks- United States, 2009–2017. Morbidity and Mortality Weekly Report, 68(25): 568–572
Hamilton K A, Waso M, Reyneke B, Saeidi N, Levine A, Lalancette C, Besner M C, Khan W, Ahmed W (2018). Cryptosporidium and Giardia in wastewater and surface water environments. Journal of Environmental Quality, 47(5): 1006–1023
Haramoto E, Kitajima M, Kishida N, Katayama H, Asami M, Akiba M (2012). Occurrence of viruses and protozoa in drinking water sources of Japan and their relationship to indicator microorganisms. Food and Environmental Virology, 4(3): 93–101
Hatam-Nahavandi K, Mohebali M, Mahvi A, Keshavarz H, Khanaliha K, Tarighi F, Molaei-Rad M, Rezaeian T, Charehdar S, Salimi M, Farnia S, Rezaeian M (2015). Evaluation of Cryptosporidium oocyst and Giardia cyst removal efficiency from urban and slaughter house wastewater treatment plants and assessment of cyst viability in wastewater effluent samples from Tehran, Iran. Journal of Water Reuse and Desalination, 5(3): 372–390
Headd B, Bradford S A (2016). Use of aerobic spores as a surrogate for Cryptosporidium oocysts in drinking water supplies. Water Research, 90: 185–202
Huang C, Hu Y, Wang L, Wang Y, Li N, Guo Y, Feng Y, Xiao L (2017). Environmental transport of emerging human-pathogenic Cryptosporidium species and subtypes through combined sewer overflow and wastewater. Applied and Environmental Microbiology, 83(16): e00682–e17
Jenkins M B, Eaglesham B S, Anthony L C, Kachlany S C, Bowman D D, Ghiorse W C (2010). Significance of wall structure, macro-molecular composition, and surface polymers to the survival and transport of Cryptosporidium parvum oocysts. Applied and Environmental Microbiology, 76(6): 1926–1934
Kihara T, Ito J, Miyake J (2013). Measurement of biomolecular diffusion in extracellular matrix condensed by fibroblasts using fluorescence correlation spectroscopy. PLoS One, 8(11): e82382
King B, Fanok S, Phillips R, Lau M, van den Akker B, Monis P (2017). Cryptosporidium attenuation across the wastewater treatment train: recycled water fit for purpose. Applied and Environmental Microbiology, 83(5): e03068–e16
Kitajima M, Haramoto E, Iker B C, Gerba C P (2014). Occurrence of Cryptosporidium, Giardia, and Cyclospora in influent and effluent water at wastewater treatment plants in Arizona. Science of the Total Environment, 484: 129–136
Kumar S, Stecher G, Tamura K (2016). MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Molecular Biology and Evolution, 33(7): 1870–1874
Lages M A, Balado M, Lemos M L (2019). The expression of virulence factors in Vibrio anguillarum is dually regulated by iron levels and temperature. Frontiers in Microbiology, 10: 2335
Lam O, Wheeler J, Tang C M (2014). Thermal control of virulence factors in bacteria: A hot topic. Virulence, 5(8): 852–862
Lapen D R, Schmidt P J, Thomas J L, Edge T A, Flemming C, Keithlin J, Neumann N, Pollari F, Ruecker N, Simhon A, Topp E, Wilkes G, Pintar K D M (2016). Towards a more accurate quantitative assessment of seasonal Cryptosporidium infection risks in surface waters using species and genotype information. Water Research, 105: 625–637
Li N, Xiao L, Wang L, Zhao S, Zhao X, Duan L, Guo M, Liu L, Feng Y (2012). Molecular surveillance of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi by genotyping and subtyping parasites in wastewater. PLoS Neglected Tropical Diseases, 6(9): e1809
Li X, Atwill E R, Dunbar L A, Jones T, Hook J, Tate K W (2005). Seasonal temperature fluctuations induces rapid inactivation of Cryptosporidium parvum. Environmental Science & Technology, 39(12): 4484–4489
Li X, Brasseur P, Agnamey P, Ballet J J, Clemenceau C (2004). Time and temperature effects on the viability and infectivity of Cryptosporidium parvum oocysts in chlorinated tap water. Archives of Environmental Health, 59(9): 462–466
Liang B, Kong D, Ma J, Wen C, Yuan T, Lee D J, Zhou J, Wang A (2016). Low temperature acclimation with electrical stimulation enhance the biocathode functioning stability for antibiotics detoxification. Water Research, 100: 157–168
Liu Y, Dong S, Kuhlenschmidt M S, Kuhlenschmidt T B, Drnevich J, Nguyen T H (2015). Inactivation mechanisms of Cryptosporidium parvum oocysts by solar ultraviolet irradiation. Environmental Science. Water Research & Technology, 1(2): 188–198
Medeiros R C, Sammarro Silva K J, Daniel L A (2020). Wastewater treatment performance in microbiological removal and oocysts viability assessed comparatively to fluorescence decay. Environmental Technology, 1811396: 1–9
Montemayor M, Valero F, Jofre J, Lucena F (2005). Occurrence of Cryptosporidium spp. oocysts in raw and treated sewage and river water in north-eastern Spain. Journal of Applied Microbiology, 99(6): 1455–1462
Morita S, Namikoshi A, Hirata T, Oguma K, Katayama H, Ohgaki S, Motoyama N, Fujiwara M (2002). Efficacy of UV irradiation in inactivating Cryptosporidium parvum oocysts. Applied and Environmental Microbiology, 68(11): 5387–5393
Nasser A M (2016). Removal of Cryptosporidium by wastewater treatment processes: A review. Journal of Water and Health, 14(1): 1–13
Nedwell D B (1999). Effect of low temperature on microbial growth: Lowered affinity for substrates limits growth at low temperature. FEMS Microbiology Ecology, 30(2): 101–111
Neumayerová H, Koudela B (2008). Effects of low and high temperatures on infectivity of Cryptosporidium muris oocysts suspended in water. Veterinary Parasitology, 153(3–4): 197–202
O’Hara S P, Chen X M (2011). The cell biology of cryptosporidium infection. Microbes and Infection, 13(8–9): 721–730
European Union (2020). Regulation (EU) 2020/741 of the European Parliament and of the Council of 25 May 2020 on minimum requirements for water reuse. Official Journal of the European Union. Brussels: European Union
Okhuysen P C, Chappell C L (2002). Cryptosporidium virulence determinants—Are we there yet? International Journal for Parasitology, 32(5): 517–525
Phadtare S, Inouye M (2004). Genome-wide transcriptional analysis of the cold shock response in wild-type and cold-sensitive, quadruple csp-deletion strains of Escherichia coli. Journal of Bacteriology, 186(20): 7007–7014
Prasad A S B, Shruptha P, Prabhu V, Srujan C, Nayak U Y, Anuradha C K R, Ramachandra L, Keerthana P, Joshi M B, Murali T S, Satyamoorthy K (2020). Pseudomonas aeruginosa virulence proteins pseudolysin and protease IV impede cutaneous wound healing. Laboratory Investigation, 100(12): 1532–1550
Rajapandi T (2020). Apicomplexan lineage-specific polytopic membrane proteins in Cryptosporidium parvum. Journal of Parasitic Diseases: Official Organ of the Indian Society for Parasitology, 44(2): 467–471
Ramamurthy T, Ghosh A, Pazhani G P, Shinoda S (2014). Current perspectives on viable but non-culturable (VBNC) pathogenic bacteria. Frontiers in Public Health, 2: 103–111
Ramo A, Del Cacho E, Sánchez-Acedo C, Quílez J (2017). Occurrence and genetic diversity of Cryptosporidium and Giardia in urban wastewater treatment plants in north-eastern Spain. Science of the Total Environment, 598: 628–638
Ran Z, Li S, Huang J, Yuan Y, Cui C, Williams C D (2010). Inactivation of Cryptosporidium by ozone and cell ultrastructures. Journal of Environmental Sciences-China, 22(12): 1954–1959
Razzolini M T P, Breternitz B S, Kuchkarian B, Bastos V K (2020). Cryptosporidium and Giardia in urban wastewater: A challenge to overcome. Environmental Pollution, 257: 113545
Reid S D, Green N M, Buss J K, Lei B, Musser J M (2001). Multilocus analysis of extracellular putative virulence proteins made by group A Streptococcus: population genetics, human serologic response, and gene transcription. Proceedings of the National Academy of Sciences of the United States of America, 98(13): 7552–7557
Rider S D Jr, Zhu G (2010). Cryptosporidium: genomic and biochemical features. Experimental Parasitology, 124(1): 2–9
Rizk N, Herrawy A, Gad M, Shaheen M, Elmahdy E (2019). Existence and removal of Rotaviruses group A and Cryptosporidium species in a wastewater treatment plant. Polish Journal of Environmental Studies, 28(6): 4331–4339
Robertson L J, Paton C A, Campbell A T, Smith P G, Jackson M H, Gilmour M A, Black S E, Stevenson D A, Smith H V (2000). Giardia cysts and Cryptosporidium oocysts at sewage treatment works in Scotland, UK. Water Research, 34(8): 2310–2322
Ryan U, Power M (2012). Cryptosporidium species in Australian wildlife and domestic animals. Parasitology, 139(13): 1673–1688
Sammarro Silva K J, Sabogal-Paz L P (2021). Cryptosporidium spp. and Giardia spp. (oo)cysts as target-organisms in sanitation and environmental monitoring: A review in microscopy-based viability assays. Water Research, 189: 116590
Santos P R, Daniel L A (2017). Occurrence and removal of Giardia spp. cysts and Cryptosporidium spp. (oo)cysts from a municipal waste-water treatment plant in Brazil. Environmental Technology, 38(10): 1245–1254
Schmitz B W, Moriyama H, Haramoto E, Kitajima M, Sherchan S, Gerba C P, Pepper I L (2018). Reduction of Cryptosporidium, Giardia, and fecal indicators by bardenpho wastewater treatment. Environmental Science & Technology, 52(12): 7015–7023
Song J, Zhao J, Gao H, Liu Y, Yue H, Zhang J, Su Y, Yin H (2011). Serological detection of Cryptosporidium spp. infection in outpatients in Changchun. Chinese Journal of Parasitology and Parasitic Diseases, 29(3): 239–241 (in Chinese)
Song X, Jing L (2015). Annual observations of climatic impacts in the Songhua River Basin, China. Advances in Meteorology, 2015: 1–12
Su Y (2011). Molecular epidemiology of Cryptosporidium and Giardia infection in a part of children and cattle in northeast China. Dissertation for the Master Degree. Changchun: Jilin University (in Chinese)
Sulaiman I M, Morgan U M, Thompson R C, Lal A A, Xiao L (2000). Phylogenetic relationships of Cryptosporidium parasites based on the 70-kilodalton heat shock protein (HSP70) gene. Applied and Environmental Microbiology, 66(6): 2385–2391
Swaffer B A, Vial H M, King B J, Daly R, Frizenschaf J, Monis P T (2014). Investigating source water Cryptosporidium concentration, species and infectivity rates during rainfall-runoff in a multi-use catchment. Water Research, 67: 310–320
Tai L, Li J, Yin J, Zhang N, Yang J, Li H, Yang Z, Gong P, Zhang X (2019). A novel detection method of Cryptosporidium parvum infection in cattle based on Cryptosporidium parvum virus 1. Acta Biochimica et Biophysica Sinica, 51(1): 104–111
Trevors J T, Bej A K, Mojib N, van Elsas J D, Van Overbeek L (2012). Bacterial gene expression at low temperatures. Extremophiles, 16(2): 167–176
Wanyiri J, Ward H (2006). Molecular basis of Cryptosporidium-host cell interactions: Recent advances and future prospects. Future Microbiology, 1(2): 201–208
Xiao F, Huang J H, Zhang B, Cui C (2009). Effects of low temperature on coagulation kinetics and floc surface morphology using alum. Desalination, 237(1–3): 201–213
Xiao L, Alderisio K, Limor J, Royer M, Lal A A (2000). Identification of species and sources of Cryptosporidium oocysts in storm waters with a small-subunit rRNA-based diagnostic and genotyping tool. Applied and Environmental Microbiology, 66(12): 5492–5498
Xing B S, Guo Q, Jiang X Y, Chen Q Q, Li P, Ni W M, Jin R C (2016). Influence of preservation temperature on the characteristics of anaerobic ammonium oxidation (anammox) granular sludge. Applied Microbiology and Biotechnology, 100(10): 4637–4649
Xu S, Wu X, Lu H (2021). Overlooked nitrogen-cycling microorganisms in biological wastewater treatment. Frontiers of Environmental Science & Engineering, 15(6): 133
Zacharia A, Outwater A H, Ngasala B, Van Deun R (2018). Pathogenic parasites in raw and treated wastewater in Africa: A review. Resources and Environment, 8: 232–240
Zahedi A, Gofton A W, Greay T, Monis P, Oskam C, Ball A, Bath A, Watkinson A, Robertson I, Ryan U (2018). Profiling the diversity of Cryptosporidium species and genotypes in wastewater treatment plants in Australia using next generation sequencing. Science of the Total Environment, 644: 635–648
Zhang T, Gao X, Wang D, Zhao J, Zhang N, Li Q, Zhu G, Yin J (2021). A single-pass type I membrane protein from the Apicomplexan parasite Cryptosporidium parvum with nanomolar binding affinity to host cell surface. Microorganisms, 9(5): 1015–1028
Acknowledgements
This work was funded by the National Natural Science Foundation of China (Nos. 51908062 and 51978135). It was also supported by the Scientific and Technological Development Plan Project of Jilin Province (China) (No. 20200201042JC). We thank Blessing Ifeoluwa Ogunniran for her linguistic and editing assistance during the revision of this manuscript.
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Highlights
• Cryptosporidium in WWTPs in a cold region was investigated in different seasons.
• The overall removal efficiency of Cryptosporidium in WWTPs was over 84%.
• The infectivity rate declined below 53% in effluents mainly due to disinfection.
• The infectivity of Cryptosporidium increased with a seasonal drop in temperature.
• Low temperature promotes binding protein retention and virulence genes expression.
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Xiao, D., Lyu, Z., Chen, S. et al. Tracking Cryptosporidium in urban wastewater treatment plants in a cold region: Occurrence, species and infectivity. Front. Environ. Sci. Eng. 16, 112 (2022). https://doi.org/10.1007/s11783-022-1533-8
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DOI: https://doi.org/10.1007/s11783-022-1533-8