Abstract
Heavy metals have been severely polluting the environment. However, the response mechanism of microbial communities to short-term heavy metals stress remains unclear. In this study, metagenomics (MG) and metatranscriptomics (MT) was performed to observe the microbial response to short-term Cr(VI) stress. MG data showed that 99.1% of species were similar in the control and Cr(VI) treated groups. However, MT data demonstrated that 83% of the microbes were active in which 58.7% increased, while the relative abundance of 41.3% decreased after short-term Cr(VI) incubation. The MT results also revealed 9% of microbes were dormant in samples. Genes associated with oxidative stress, Cr(VI) transport, resistance, and reduction, as well as genes with unknown functions were 2–10 times upregulated after Cr(VI) treatment. To further confirm the function of unknown genes, two genes (314 and 494) were selected to detect the Cr(VI) resistance and reduction ability. The results showed that these genes significantly increased the Cr(VI) remediation ability of Escherichia coli. MT results also revealed an increase in the expression of some rare genera (at least two times) after Cr(VI) treatment, indicating these rare species played a crucial role in microbial response to short-term Cr(VI) stress. In summary, MT is an efficient way to understand the role of active and dormant microbes in specific environmental conditions.
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Data availability
All the raw datasets have been deposited into NCBI’s Sequence Read Achieve under accession number SRP154829.
References
Aanderud ZT, Vert JC, Lennon JT, Magnusson TW, Breakwell DP, Harker AR (2016) Bacterial dormancy is more prevalent in freshwater than Hypersaline Lakes. Front Microbiol 7. https://doi.org/10.3389/fmicb.2016.00853
Anantharaman K et al. (2018) Expanded diversity of microbial groups that shape the dissimilatory sulfur cycle. ISME J 12:1715–1728. https://doi.org/10.1038/s41396-018-0078-0
Baaziz H et al. (2017) ChrASO, the chromate efflux pump of Shewanella oneidensis, improves chromate survival and reduction. PLoS ONE 12:e0188516. https://doi.org/10.1371/journal.pone.0188516
Bhatt K, Timsit E, Rawlyk N, Potter A, Liljebjelke K (2018) Integrative conjugative element ICEHs1 encodes for antimicrobial resistance and metal tolerance in Histophilus somni. Front Vet Sci 5:153–153. https://doi.org/10.3389/fvets.2018.00153
Couradeau E et al. (2019) Probing the active fraction of soil microbiomes using BONCAT-FACS. Nat Commun 10:2770. https://doi.org/10.1038/s41467-019-10542-0
Curtis C et al. (2012) The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups. Nature 486:346–352. https://doi.org/10.1038/nature10983
Dai R, Chen J, Lin J, Xiao S, Chen S, Deng Y (2009) Reduction of nitro phenols using nitroreductase from E. coli in the presence of NADH. J Hazard Mater 170:141–143. https://doi.org/10.1016/j.jhazmat.2009.04.122
Damon C et al. (2011) A novel fungal family of oligopeptide transporters identified by functional metatranscriptomics of soil eukaryotes. ISME J 5:1871–1880. https://doi.org/10.1038/ismej.2011.67
Dewi Puspita I, Kamagata Y, Tanaka M, Asano K, Nakatsu CH (2012) Are uncultivated bacteria really uncultivable? Microbes Environ 27:356–366. https://doi.org/10.1264/jsme2.ME12092
Francis CA, Beman JM, Kuypers MMM (2007) New processes and players in the nitrogen cycle: the microbial ecology of anaerobic and archaeal ammonia oxidation. ISME J 1:19–27. https://doi.org/10.1038/ismej.2007.8
Gharechahi J, Salekdeh GH (2018) A metagenomic analysis of the camel rumen’s microbiome identifies the major microbes responsible for lignocellulose degradation and fermentation. Biotechnol Biofuels 11:216. https://doi.org/10.1186/s13068-018-1214-9
Gourmelon V et al. (2016) Environmental and geographical factors structure soil microbial diversity in new caledonian ultramafic substrates: a metagenomic approach. PLoS ONE 11:e0167405. https://doi.org/10.1371/journal.pone.0167405
Guo J et al. (2016a) Metagenomic analysis of anammox communities in three different microbial aggregates. Environ Microbiol 18:2979–2993. https://doi.org/10.1111/1462-2920.13132
Guo Y et al. (2016b) Effect of short-term room temperature storage on the microbial community in infant fecal samples. Sci Rep 6:26648. https://doi.org/10.1038/srep26648
Haider B, Ahn T-H, Bushnell B, Chai J, Copeland A, Pan C (2014) Omega: an overlap-graph de novo assembler for metagenomics. Bioinformatics 30:2717–2722. https://doi.org/10.1093/bioinformatics/btu395
Hamasaki K, Taniguchi A, Tada Y, Kaneko R, Miki T (2016) Active populations of rare microbes in oceanic environments as revealed by bromodeoxyuridine incorporation and 454 tag sequencing. Gene 576:650–656. https://doi.org/10.1016/j.gene.2015.10.016
Han H, Ling Z, Zhou T, Xu R, He Y, Liu P, Li X (2017) Copper (II) binding of NAD(P)H- flavin oxidoreductase (NfoR) enhances its Cr (VI)-reducing ability. Sci Rep 7:15481. https://doi.org/10.1038/s41598-017-15588-y
He B, Jin S, Cao J, Mi L, Wang J (2019) Metatranscriptomics of the Hu sheep rumen microbiome reveals novel cellulases. Biotechnol Biofuels 12:153. https://doi.org/10.1186/s13068-019-1498-4
He H et al. (2013) Characterization of endophytic Rahnella sp. JN6 from Polygonum pubescens and its potential in promoting growth and Cd, Pb, Zn uptake by Brassica napus. Chemosphere 90:1960–1965. https://doi.org/10.1016/j.chemosphere.2012.10.057
He M, Li X, Guo L, Miller SJ, Rensing C, Wang G (2010) Characterization and genomic analysis of chromate resistant and reducing Bacillus cereus strain SJ1. BMC Microbiol 10:221. https://doi.org/10.1186/1471-2180-10-221
Hol WHG, de Boer W, de Hollander M, Kuramae EE, Meisner A, van der Putten WH (2015) Context dependency and saturating effects of loss of rare soil microbes on plant productivity. Front Plant Sci 6. https://doi.org/10.3389/fpls.2015.00485
Huang H et al. (2017) The naphthalene catabolic protein NahG plays a key role in hexavalent chromium reduction in Pseudomonas brassicacearum LZ-4. Sci Rep 7:9670. https://doi.org/10.1038/s41598-017-10469-w
Hugo W et al. (2016) Genomic and transcriptomic features of response to anti-PD-1 therapy in metastatic melanoma. Cell 165:35–44. https://doi.org/10.1016/j.cell.2016.02.065
Ignatov DV, Salina EG, Fursov MV, Skvortsov TA, Azhikina TL, Kaprelyants AS (2015) Dormant non-culturable Mycobacterium tuberculosis retains stable low-abundant mRNA BMC. Genomics 16:954. https://doi.org/10.1186/s12864-015-2197-6
Ishii S, Suzuki S, Tenney A, Nealson KH, Bretschger O (2018) Comparative metatranscriptomics reveals extracellular electron transfer pathways conferring microbial adaptivity to surface redox potential changes. ISME J 12:2844–2863. https://doi.org/10.1038/s41396-018-0238-2
Ishii SI et al. (2013) A novel metatranscriptomic approach to identify gene expression dynamics during extracellular electron transfer. Nat Commun 4:1601
Islam MS, Ahmed MK, Raknuzzaman M, Habibullah-Al-Mamun M, Islam MK (2015) Heavy metal pollution in surface water and sediment: a preliminary assessment of an urban river in a developing country. Ecol Indic 48:282–291. https://doi.org/10.1016/j.ecolind.2014.08.016
Jansson JK, Hofmockel KS (2018) The soil microbiome—from metagenomics to metaphenomics. Curr Opin Microbiol 43:162–168. https://doi.org/10.1016/j.mib.2018.01.013
Jiao S, Chen W, Wei G (2017) Biogeography and ecological diversity patterns of rare and abundant bacteria in oil-contaminated soils. Mol Ecol 26:5305–5317. https://doi.org/10.1111/mec.14218
Jousset A et al. (2017) Where less may be more: how the rare biosphere pulls ecosystems strings. ISME J 11:853–862. https://doi.org/10.1038/ismej.2016.174
Kearns PJ, Angell JH, Howard EM, Deegan LA, Stanley RHR, Bowen JL (2016) Nutrient enrichment induces dormancy and decreases diversity of active bacteria in salt marsh sediments. Nat Commun 7:12881. https://doi.org/10.1038/ncomms12881
Kell DB, Pretorius E (2018) No effects without causes: the Iron Dysregulation and Dormant Microbes hypothesis for chronic, inflammatory diseases. Biol Rev 93:1518–1557. https://doi.org/10.1111/brv.12407
Klaus-Joerger T, Joerger R, Olsson E, Granqvist C-G (2001) Bacteria as workers in the living factory: metal-accumulating bacteria and their potential for materials science. Trends Biotechnol 19:15–20. https://doi.org/10.1016/S0167-7799(00)01514-6
Kohler C, Lourenço RF, Avelar GM, Gomes SL (2012) Extracytoplasmic function (ECF) sigma factor σF is involved in Caulobacter crescentus response to heavy metal stress. BMC Microbiol 12:210. https://doi.org/10.1186/1471-2180-12-210
Lettat A, Benchaar C (2013) Diet-induced alterations in total and metabolically active microbes within the rumen of dairy cows. PLoS ONE 8:e60978. https://doi.org/10.1371/journal.pone.0060978
Li H et al. (2016) Pika gut may select for rare but diverse environmental bacteria. Front Microbiol 7. https://doi.org/10.3389/fmicb.2016.01269
Lin K-H et al. (2015) Metabolic characteristics of dominant microbes and key rare species from an acidic hot spring in Taiwan revealed by metagenomics. BMC Genom 16:1029. https://doi.org/10.1186/s12864-015-2230-9
Long PE, Williams KH, Hubbard SS, Banfield JF (2016) Microbial metagenomics reveals climate-relevant subsurface biogeochemical processes. Trends Microbiol 24:600–610. https://doi.org/10.1016/j.tim.2016.04.006
Marietou A, Røy H, Jørgensen BB, Kjeldsen KU (2018) Sulfate transporters in dissimilatory sulfate reducing microorganisms: a comparative genomics analysis. Front Microbiol 9. https://doi.org/10.3389/fmicb.2018.00309
McClary JS, Boehm AB (2018) Transcriptional response of Staphylococcus aureus to sunlight in oxic and anoxic conditions. Front Microbiol 9. https://doi.org/10.3389/fmicb.2018.00249
Mende DR et al. (2012) Assessment of metagenomic assembly using simulated next generation sequencing data. PLoS ONE 7:e31386. https://doi.org/10.1371/journal.pone.0031386
Miroshnichenko ML et al. (2002) Nautilia lithotrophica gen. nov., sp. nov., a thermophilic sulfur-reducing epsilon-proteobacterium isolated from a deep-sea hydrothermal vent. Int J Syst Evolut Microbiol 52:1299–1304. https://doi.org/10.1099/00207713-52-4-1299
Mu D-S, Liang Q-Y, Wang X-M, Lu D-C, Shi M-J, Chen G-J, Du Z-J (2018) Metatranscriptomic and comparative genomic insights into resuscitation mechanisms during enrichment culturing. Microbiome 6:230. https://doi.org/10.1186/s40168-018-0613-2
Mueller RC, Gallegos-Graves L, Zak DR, Kuske CR (2016) Assembly of active bacterial and fungal communities along a natural environmental gradient. Microb Ecol 71:57–67. https://doi.org/10.1007/s00248-015-0655-y
Nishiyama E et al. (2018) The relationship between microbial community structures and environmental parameters revealed by metagenomic analysis of hot Spring water in the Kirishima Area, Japan. Front Bioeng Biotechnol 6. https://doi.org/10.3389/fbioe.2018.00202
Ogg CD, Patel BKC (2009) Thermotalea metallivorans gen. nov., sp. nov., a thermophilic, anaerobic bacterium from the Great Artesian Basin of Australia aquifer. Int J Syst Evolut Microbiol 59:964–971. https://doi.org/10.1099/ijs.0.004218-0
Palomo A, Jane Fowler S, Gülay A, Rasmussen S, Sicheritz-Ponten T, Smets BF (2016) Metagenomic analysis of rapid gravity sand filter microbial communities suggests novel physiology of Nitrospira spp. ISME J 10:2569–2581. https://doi.org/10.1038/ismej.2016.63
Papp K, Mau RL, Hayer M, Koch BJ, Hungate BA, Schwartz E (2018) Quantitative stable isotope probing with h2 18 o reveals that most bacterial taxa in soil synthesize new ribosomal rna. ISME J 12:3043–3045
Pattanapipitpaisal P, Brown N, Macaskie L (2001) Chromate reduction and 16S rRNA identification of bacteria isolated from a Cr (VI)-contaminated site. Appl Microbiol Biotechnol 57:257–261
Pei Y et al. (2020a) The guanidine thiocyanate-high EDTA method for total microbial RNA extraction from severely heavy metal-contaminated soils. Microb Biotechnol 0:1–14. https://doi.org/10.1111/1751-7915.13615
Pei Y et al. (2020b) Exploring novel Cr(VI) remediation genes for Cr(VI)-contaminated industrial wastewater treatment by comparative metatranscriptomics and metagenomics. Sci Total Environ 742:140435. https://doi.org/10.1016/j.scitotenv.2020.140435
Pei Y, Yu Z, Ji J, Khan A, Li X (2018) Microbial community structure and function indicate the severity of chromium contamination of the yellow river. Front Microbiol 9. https://doi.org/10.3389/fmicb.2018.00038
Portillo MC, Gonzalez JM, Saiz-Jimenez C (2008) Metabolically active microbial communities of yellow and grey colonizations on the walls of Altamira Cave, Spain. J Appl Microbiol 104:681–691. https://doi.org/10.1111/j.1365-2672.2007.03594.x
Pradhan SK, Singh NR, Rath BP, Thatoi H (2016) Bacterial chromate reduction: a review of important genomic, proteomic, and bioinformatic analysis. Crit Rev Environ Sci Technol 46:1659–1703. https://doi.org/10.1080/10643389.2016.1258912
Probst AJ et al. (2017) Genomic resolution of a cold subsurface aquifer community provides metabolic insights for novel microbes adapted to high CO2 concentrations. Environ Microbiol 19:459–474. https://doi.org/10.1111/1462-2920.13362
Pu Y et al. (2016) Enhanced efflux activity facilitates drug tolerance in dormant bacterial cells. Mol Cell 62:284–294. https://doi.org/10.1016/j.molcel.2016.03.035
Puente-Sánchez F, Olsson S, Aguilera A (2016) Comparative transcriptomic analysis of the response of Dunaliella acidophila (Chlorophyta) to short-term cadmium and chronic natural metal-rich water exposures. Microb Ecol 72:595–607. https://doi.org/10.1007/s00248-016-0824-7
Qian J, Wei L, Liu R, Jiang F, Hao X, Chen G-H (2016) An exploratory study on the pathways of Cr (VI) reduction in sulfate-reducing up-flow anaerobic sludge bed (UASB). React Sci Rep 6:23694. https://doi.org/10.1038/srep23694
Qian J, Zhou J, Wang L, Wei L, Li Q, Wang D, Wang Q (2017) Direct Cr (VI) bio-reduction with organics as electron donor by anaerobic sludge. Chem Eng J 309:330–338. https://doi.org/10.1016/j.cej.2016.10.077
Qin Z et al. (2020) Coupled photocatalytic-bacterial degradation of pyrene: removal enhancement and bacterial community responses. Environ Res 183:109135. https://doi.org/10.1016/j.envres.2020.109135
Ray S, Pahan K, Gachhui R, Chaudhuri J, Mandal A (1993) Studies on the mercury volatilizing enzymes in nitrogen-fixing Beijerinckia mobilis. World J Microbiol Biotechnol 9:184–186. https://doi.org/10.1007/BF00327833
Reyes-Gallegos RI, Ramírez-Díaz MI, Cervantes C (2016) chr genes from adaptive replicons are responsible for chromate resistance by Burkholderia xenovorans LB400. World J Microbiol Biotechnol 32:45. https://doi.org/10.1007/s11274-015-1996-x
Robins KJ, Hooks DO, Rehm BHA, Ackerley DF (2013) Escherichia coli NemA is an efficient chromate reductase that can be biologically immobilized to provide a cell free system for remediation of hexavalent chromium. Plos One 8:e59200
Schiavon M, Wirtz M, Borsa P, Quaggiotti S, Hell R, Malagoli M (2007) Chromate differentially affects the expression of a high-affinity sulfate transporter and isoforms of components of the sulfate assimilatory pathway in Zea mays (L.). Plant Biology 9:662–671
Silver S, Schottel J, Weiss A (2001) Bacterial resistance to toxic metals determined by extrachromosomal R factors. Int Biodeterior Biodegrad 48:263–281
Sorokin DY et al. (2014) Genome analysis of Chitinivibrio alkaliphilus gen. nov., sp. nov., a novel extremely haloalkaliphilic anaerobic chitinolytic bacterium from the candidate phylum Termite Group 3. Environ Microbiol 16:1549–1565. https://doi.org/10.1111/1462-2920.12284
Stoeck T, Zuendorf A, Breiner H-W, Behnke A (2007) A molecular approach to identify active microbes in environmental eukaryote clone libraries. Microb Ecol 53:328–339. https://doi.org/10.1007/s00248-006-9166-1
Suzuki T, Miyata N, Horitsu H, Kawai K, Takamizawa K, Tai Y, Okazaki M (1992) NAD(P)H-dependent chromium (VI) reductase of Pseudomonas ambigua G-1: a Cr(V) intermediate is formed during the reduction of Cr(VI) to Cr(III). J Bacteriol 174:5340–5345. https://doi.org/10.1128/jb.174.16.5340-5345.1992
Trivedi P et al. (2016) Microbial regulation of the soil carbon cycle: evidence from gene–enzyme relationships. ISME J 10:2593–2604. https://doi.org/10.1038/ismej.2016.65
Trutneva K, Shleeva M, Nikitushkin V, Demina G, Kaprelyants A (2018) Protein composition of Mycobacterium smegmatis differs significantly between active cells and dormant cells with ovoid morphology. Front Microbiol 9. https://doi.org/10.3389/fmicb.2018.02083
Vogel C, Marcotte EM (2012) Insights into the regulation of protein abundance from proteomic and transcriptomic analyses. Nat Rev Genet 13:227–232. https://doi.org/10.1038/nrg3185
White RA et al. (2016) Metagenomic analysis suggests modern freshwater microbialites harbor a distinct core microbial community. Front Microbiol 6. https://doi.org/10.3389/fmicb.2015.01531
Worden AZ, Follows MJ, Giovannoni SJ, Wilken S, Zimmerman AE, Keeling PJ (2015) Rethinking the marine carbon cycle: factoring in the multifarious lifestyles of microbes. Science 347:1257594. https://doi.org/10.1126/science.1257594
Yan X, Luo X, Zhao M (2016) Metagenomic analysis of microbial community in uranium-contaminated soil. Appl Microbiol Biotechnol 100:299–310. https://doi.org/10.1007/s00253-015-7003-5
Yang Q, Li Z, Lu X, Duan Q, Huang L, Bi J (2018) A review of soil heavy metal pollution from industrial and agricultural regions in China: pollution and risk assessment. Sci Total Environ 642:690–700. https://doi.org/10.1016/j.scitotenv.2018.06.068
Yu Z et al. (2016) The shifts of sediment microbial community phylogenetic and functional structures during chromium (VI) reduction. Ecotoxicology 25:1759–1770. https://doi.org/10.1007/s10646-016-1719-6
Zhai Q, Xiao Y, Zhao J, Tian F, Zhang H, Narbad A, Chen W (2017) Identification of key proteins and pathways in cadmium tolerance of Lactobacillus plantarum strains by proteomic analysis. Sci Rep 7:1182. https://doi.org/10.1038/s41598-017-01180-x
Zhu Y, Ma N, Jin W, Wu S, Sun C (2017) Genomic and transcriptomic insights into calcium carbonate biomineralization by marine actinobacterium Brevibacterium linens BS258. Front Microbiol 8. https://doi.org/10.3389/fmicb.2017.00602
Funding
This study was funded by the National Natural Science Foundation grant (Grant no. 31870082), the Gansu Province Major Science and Technology Projects (Grant no. 17ZD2WA017), and the Fundamental Research Funds for the Central Universities (Grant no. lzujbky-2018-it47, lzujbky-2018-103).
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Yu, Z., Pei, Y., Zhao, S. et al. Metatranscriptomic analysis reveals active microbes and genes responded to short-term Cr(VI) stress. Ecotoxicology 30, 1527–1537 (2021). https://doi.org/10.1007/s10646-020-02290-5
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DOI: https://doi.org/10.1007/s10646-020-02290-5
Keywords
- Short-term Cr(VI) stress
- Active microbes
- Dormant microbes
- Function-unknown genes
- Rare genera