Comprehensive Analysis of Antimicrobial, Heavy Metal, and Pesticide Residues in Commercial Organic Fertilizers and Their Correlation with Tigecycline-Resistant tet(X)-Variant Genes

An antimicrobial resistance gene (ARG) is considered a novel contaminant for the environment. Most animal feces are usually made into commercial organic fertilizers in China and will pose a threat to the farmland soil and agricultural product if fertilizers harboring clinically significant antimicrobial-resistant (AMR) genes are applied on farmland. With the issue of the antimicrobial additive ban in feed in Chinese animal husbandry, it is important to determine the potential drivers of the spread of the newly discovered tigecycline-resistant tet(X)-variant genes. Here, we investigated the correlations between residues of heavy metals, antimicrobials, and pesticides and the relative abundance of tet(X)-variant genes in 94 commercial organic-fertilizer samples collected from 9 Chinese provinces. A total of 5 heavy metals (mercury, lead, arsenic, chromium, and cadmium), 10 antimicrobials, and 18 pesticides were detected. The tet(X)-variant genes, including tet(X)/(X2), tet(X3), tet(X4), tet(X5), and tet(X6) were detected in 39 (41.5%) samples. Although tet(X)-variant-carrying bacteria were not isolated from these samples, the tet(X4)-carrying plasmids could be captured by exogenous Escherichia coli. Correlation analysis revealed that heavy metals, other than antimicrobials, showed a significant positive association with the relative abundance of the tet(X)-variant genes, especially tet(X3) and tet(X4) (R = 0.346 to 0.389, P < 0.001). The correlation was attributed to the coselection of the tet(X3)/tet(X4) gene on the same plasmid and the conjugation-promoting effect of tet(X3)/tet(X4)-carrying plasmids by subinhibitory concentrations of heavy metals. The heavy metals increased the permeability of the bacterial outer membrane and upregulated the transcription of type IV secretion system (T4SS)-encoding genes on tet(X)-variant-carrying plasmids, therefore enhancing the bacterial conjugation rates. Taken together, our findings have indicated that heavy metals may play an important role in spreading tet(X)-variant genes within the animal manure-related environment.IMPORTANCE An antimicrobial resistance gene (ARG) is considered a novel contaminant for the environment. Most animal feces are usually made into commercial organic fertilizers in China and will pose a threat to the farmland soil and agricultural product if fertilizers harboring clinically significant antimicrobial-resistant (AMR) genes are applied on farmland. This study has indicated that heavy metals may play an important role in the transmission of transferable tigecycline resistance genes [tet(X3) and tet(X4)]. The mechanism was that heavy metals posed a coselection effect of the tet(X3)/tet(X4) gene on the same plasmid and could increase the conjugation ability of tet(X3)/tet(X4)-carrying plasmids. The conjugation-promoting concentrations of heavy metals are lower than the maximal limits defined in the national standard for fertilizers, indicating a high transmission risk of tet(X3)/tet(X4) genes within the animal manure-related environment. The findings in this study will provide scientific evidence for the future development of effective measures to reduce AMR dissemination.

Automated summary

This study found that heavy metals may play an important role in the transmission of transferable tigecycline resistance genes (tet(X3) and tet(X4)) within the animal manure-related environment in China. Heavy metals have a coselection effect on the tet(X3)/tet(X4) gene on the same plasmid and can increase the conjugation ability of tet(X3)/tet(X4)-carrying plasmids. The conjugation-promoting concentrations of heavy metals are lower than the maximal limits defined in the national standard for fertilizers, indicating a high transmission risk of tet(X3)/tet(X4) genes. These findings will provide scientific evidence for the future development of effective measures to reduce antimicrobial resistance dissemination.