- Pathogen detection in human samples and food
- Antibiotic resistance genes
- mRNA, miRNA, and lncRNA as disease biomarkers
- Genotyping using animal and blood samples
Antibiotic resistance genes
The discovery and use of antibiotics are two of the most significant breakthroughs in twentieth-century medicine—leading to dramatic reductions in human morbidity and mortality. Antibiotics and other antimicrobial agents are also used extensively in agriculture. Typically, livestock are given antibiotics to encourage growth and prevent illness, thereby increasing the amount of food produced. There are multiple classes of antibiotics, categorized based on their mechanism of action. Antibiotics can damage the cell wall of a bacterium, block DNA, RNA, or protein synthesis, or even inhibit the metabolic growth of bacteria. Some antibiotics are specific to certain species of bacteria (narrow-spectrum), whereas others can affect a wide range of bacteria (broad-spectrum).
One emerging problem with antibiotics is the development of antibiotic-resistant bacteria. Bacteria can acquire antibiotic resistance genes (ARGs) over time due to selective pressure. Excessive antibiotic use can lead to the development of spontaneous ARGs in certain strains. As these strains grow and expand, different antibiotics have to be developed to combat them. Ultimately, the current strategy is to prevent overuse of antibiotics, thereby limiting the exposure of bacteria to these agents and minimizing selective pressures. For human patients, overuse can result from incorrect dosing, over- or mis-prescribing, or improper disposal. Critically, antibiotic use in agriculture can also contribute to bacteria developing ARGs. Not only can antibiotic-resistant bacteria develop in livestock, but manure can act as a vector to transmit antibiotic-resistant strains into the environment via soil, water, and produce. Thus, antibiotic-resistant bacteria can be spread widely, and recent efforts have focused on identifying ARGs and tracking their prevalence in a variety of environments and sample types.
There are currently over 1,000 types of ARGs that have been identified to confer resistance to the hundreds of available antibiotics. Consequently, monitoring human and environmental samples for ARGs requires the ability to perform high-throughput real-time PCR (qPCR) screens for many different target sequences in numerous sample types. Further, because bacteria are constantly developing new ARGs, it is critical to be able to modify and add new targets to the screen. Thus, ARG research requires a flexible, high-throughput qPCR system to monitor gene expression in a wide range of sample types.
The SmartChip Real-Time PCR System has been used in numerous studies for profiling and tracking ARGs in a variety of samples. Initially, high-throughput screens were performed using panels of 296 targets, which supported the ability to screen up to 16 soil samples per chip. In recent years, the panels have evolved to contain 384 targets (see Table I), as well as being run on soil, water, sediment, manure, lettuce, fish, and sludge sample types. The flexibility and throughput of the SmartChip system have been instrumental in enabling these and future studies of the antibiotic resistome. In fact, the SmartChip system has been utilized in 75% of the publications using high-throughput qPCR for antibiotic resistance research (Waseem et al. 2019).
|Target gene category||Number of primer sets|
|Mobile genetic elements||52|
Table I. Antibiotic resistance gene categories that have been analyzed on the SmartChip system.
An, X.-L. et al. Tracking antibiotic resistome during wastewater treatment using high throughput quantitative PCR. Environ. Int. 117, 146–153 (2018).
Chen, Q. et al. Long-term field application of sewage sludge increases the abundance of antibiotic resistance genes in soil. Environ. Int. 92-93, 1–10 (2016).
Chen, Q.-L., An, X.-L., Zheng, B.-X., Ma, Y.-B. & Su, J.-Q. Long-term organic fertilization increased antibiotic resistome in phyllosphere of maize. Sci. Total Environ. 645, 1230–1237 (2018).
Chen, Q.-L. et al. Application of struvite alters the antibiotic resistome in soil, rhizosphere, and phyllosphere. Environ. Sci. Technol. 51, 8149–8157 (2017).
Chen, Q.-L. et al. Effect of biochar amendment on the alleviation of antibiotic resistance in soil and phyllosphere of Brassica chinensis L. Soil Biol. Biochem. 119, 74–82 (2018).
Chen, Y. et al. High-throughput profiling of antibiotic resistance gene dynamic in a drinking water river-reservoir system. Water Res. 149, 179–189 (2019).
Chen, Z. et al. Antibiotic resistance genes and bacterial communities in cornfield and pasture soils receiving swine and dairy manures. Environ. Pollut. 248, 947–957 (2019).
Cheng, J.-H., Tang, X.-Y. & Cui, J.-F. Effect of long-term manure slurry application on the occurrence of antibiotic resistance genes in arable purple soil (entisol). Sci. Total Environ. 647, 853–861 (2019).
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Cui, E.-P. et al. Amendment soil with biochar to control antibiotic resistance genes under unconventional water resources irrigation: Proceed with caution. Environ. Pollut. 240, 475–484 (2018).
Ding, J. et al. Long-term application of organic fertilization causes the accumulation of antibiotic resistome in earthworm gut microbiota. Environ. Int. 124, 145–152 (2019).
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Gao, J.-F., Liu, X.-H., Fan, X.-Y. & Dai, H.-H. Effects of triclosan on performance, microbial community and antibiotic resistance genes during partial denitrification in a sequencing moving bed biofilm reactor. Bioresour. Technol. 281, 326–334 (2019).
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Gu, J. et al. High-throughput analysis of the effects of different fish culture methods on antibiotic resistance gene abundances in a lake. Environ. Sci. Pollut. Res. 26, 5445–5453 (2019).
Guo, X. et al. Antibiotic resistome associated with small-scale poultry production in rural Ecuador. Environ. Sci. Technol. 52, 8165–8172 (2018).
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Hu, H.-W. et al. Diversity of herbaceous plants and bacterial communities regulates soil resistome across forest biomes. Environ. Microbiol. 20, 3186–3200 (2018).
Huang, X. et al. Higher temperatures do not always achieve better antibiotic resistance gene removal in anaerobic digestion of swine manure. Appl. Environ. Microbiol. 85, e02878-18 (2019).
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Li, Y. et al. Prevalence of antibiotic resistance genes in air-conditioning systems in hospitals, farms, and residences. Int. J. Environ. Res. Public Health 16, 683 (2019).
Lin, W., Zhang, M., Zhang, S. & Yu, X. Can chlorination co-select antibiotic-resistance genes? Chemosphere 156, 412–419 (2016).
Liu, L. et al. Large-scale biogeographical patterns of bacterial antibiotic resistome in the waterbodies of China. Environ. Int. 117, 292–299 (2018).
Lu, X.-M., Li, W.-F. & Li, C.-B. Characterization and quantification of antibiotic resistance genes in manure of piglets and adult pigs fed on different diets. Environ. Pollut. 229, 102–110 (2017).
McCann, C. M. et al. Understanding drivers of antibiotic resistance genes in High Arctic soil ecosystems. Environ. Int. 125, 497–504 (2019).
Muurinen, J. et al. Influence of manure application on the environmental resistome under Finnish agricultural practice with restricted antibiotic use. Environ. Sci. Technol. 51, 5989–5999 (2017).
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Muziasari, W. I. et al. The resistome of farmed fish feces contributes to the enrichment of antibiotic resistance genes in sediments below Baltic Sea fish farms. Front. Microbiol. 7, 1–10 (2017).
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Tang, M. et al. Abundance and distribution of antibiotic resistance genes in a full-scale anaerobic-aerobic system alternately treating ribostamycin, spiramycin and paromomycin production wastewater. Environ. Geochem. Health (2017). doi:10.1007/s10653-017-9987-5
Tian, Z., Chi, Y., Yu, B., Yang, M. & Zhang, Y. Thermophilic anaerobic digestion reduces ARGs in excess sludge even under high oxytetracycline concentrations. Chemosphere 222, 305–313 (2019).
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Wang, F.-H. et al. High throughput profiling of antibiotic resistance genes in urban park soils with reclaimed water irrigation. Environ. Sci. Technol. 48, 9079–9085 (2014).
Wang, H. et al. The antibiotic resistome of swine manure is significantly altered by association with the Musca domestica larvae gut microbiome. ISME J. 11, 100–111 (2017).
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Waseem, H. et al. Contributions and challenges of high throughput qPCR for determining antimicrobial resistance in the environment: a critical review. Molecules 24, 163 (2019).
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Xie, W.-Y. et al. Long-term effects of manure and chemical fertilizers on soil antibiotic resistome. Soil Biol. Biochem. 122, 111–119 (2018).
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Zhang, Y.-J. et al. Salinity as a predominant factor modulating the distribution patterns of antibiotic resistance genes in ocean and river beach soils. Sci. Total Environ. 668, 193–203 (2019).
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