Metagenomics-Guided Discovery of Potential Bacterial Metallothionein Genes from the Soil Microbiome That Confer Cu and/or Cd Resistance.

Metallothionein (MT) genes are valuable genetic materials for developing metal bioremediation tools. Currently, a limited number of prokaryotic MTs have been experimentally identified, which necessitates the expansion of bacterial MT diversity. In this study, we conducted a metagenomics-guided analysis for the discovery of potential bacterial MT genes from the soil microbiome. More specifically, we combined resistance gene enrichment through diversity loss, metagenomic mining with a dedicated MT database, evolutionary trace analysis, DNA chemical synthesis, and functional genomic validation to identify novel MTs. Results showed that Cu stress induced a compositional change in the soil microbiome, with an enrichment of metal-resistant bacteria in soils with higher Cu concentrations. Shotgun metagenomic sequencing was performed to obtain the gene pool of environmental DNA (eDNA), which was subjected to a local BLAST search against an MT database for detecting putative MT genes. Evolutional trace analysis led to the identification of 27 potential MTs with conserved cysteine/histidine motifs different from those of known prokaryotic MTs. Following chemical synthesis of these 27 potential MT genes and heterologous expression in Escherichia coli, six of them were found to improve the hosts' growth substantially and enhanced the hosts' sorption of Cu, Cd, and Zn, among which MT5 led to a 13.7-fold increase in Cd accumulation. Furthermore, four of them restored Cu and/or Cd resistance in two metal-sensitive E. coli strains.IMPORTANCE The metagenomics-guided procedure developed here bypasses the difficulties encountered in classic PCR-based approaches and led to the discovery of novel MT genes, which may be useful in developing bioremediation tools. The procedure used here expands our knowledge on the diversity of bacterial MTs in the environment and may also be applicable to identify other functional genes from eDNA.

Characterization of the In Vitro Chlamydia pecorum Response to Gamma Interferon.

Chlamydia pecorum is an important intracellular bacterium that causes a range of diseases in animals, including a native Australian marsupial, the koala. In humans and animals, a gamma interferon (IFN-γ)-mediated immune response is important for the control of intracellular bacteria. The present study tested the hypotheses that C. pecorum can escape IFN-γ-mediated depletion of host cell tryptophan pools. In doing so, we demonstrated that, unlike Chlamydia trachomatis, C. pecorum is completely resistant to IFN-γ in human epithelial cells. While the growth of C. pecorum was inhibited in tryptophan-deficient medium, it could be restored by the addition of kynurenine, anthranilic acid, and indole, metabolites that could be exploited by the gene products of the C. pecorum tryptophan biosynthesis operon. We also found that expression of trp genes was detectable only when C. pecorum was grown in tryptophan-free medium, with gene repression occurring in response to the addition of kynurenine, anthranilic acid, and indole. When grown in bovine kidney epithelial cells, bovine IFN-γ also failed to restrict the growth of C. pecorum, while C. trachomatis was inhibited, suggesting that C. pecorum could use the same mechanisms to evade the immune response in vivo in its natural host. Highlighting the different mechanisms triggered by IFN-γ, however, both species failed to grow in murine McCoy cells treated with murine IFN-γ. This work confirms previous hypotheses about the potential survival of C. pecorum after IFN-γ-mediated host cell tryptophan depletion and raises questions about the immune pathways used by the natural hosts of C. pecorum to control the widespread pathogen.

Heavy Metal and Metalloid Pollution of Soil, Water and Foods in Bangladesh: A Critical Review.

Bangladesh is a densely populated developing country. Both industrialization and geological sources have caused widespread heavy metal and metalloid pollution in Bangladesh, which is now posing substantial threats to the local people. In this review, we carried out one of the most exhaustive literature analyses on the current status of Bangladesh heavy metal and metalloid pollution, covering water, soil, and foods. Analysis showed that soils near high traffic and industrial areas contain high concentrations of heavy metals and metalloids. Agricultural land and vegetables in sewage-irrigated areas were also found to be heavy metal- and metalloid-contaminated. River water, sediment, and fish from the Buriganga, Turag, Shitalakhya, and Karnaphuli rivers are highly contaminated with cadmium (Cd), lead (Pb), and chromium (Cr). Particularly, groundwater arsenic (As) pollution associated with high geological background levels in Bangladesh is well reported and is hitherto the largest mass poisoning in the world. Overall, the contamination levels of heavy metals and metalloids vary among the cities, with industrial areas being most polluted. In all, this review provides a quantitative identification of the As, Pb, Cd, and Cr contamination hotspots in Bangladesh based on the literature, which may be useful to environmental restorationists and local policy makers.