Cyanobacteria as a critical reservoir of the environmental antimicrobial resistome.

Antimicrobial resistance (AMR) is predicted to cause a worldwide annual toll of 10 million deaths by 2050. This looming public health threat has been linked to antibiotic overuse and pollution, which places selective pressures on AMR maintenance and transfer in and between microbial populations. We examined the distribution, diversity and potential mobility of AMR genes in cyanobacteria. While cyanobacteria are not pathogenic, we hypothesised that they could be a major environmental reservoir for AMR genes. Genes encoding AMR to seven antimicrobial drug classes were found in 10% of cyanobacterial genomes. AMR genes were found in 13% of freshwater, 19% of terrestrial, 34% of symbiotic, 2% of thermal spring, and 3% of marine genomes. AMR genes were found in five cyanobacterial orders with 23% of Nostocales and 8% of Oscillatoriales strains containing AMR genes. The most frequently observed alleles were ansamycin resistance genes, which were present in 7% of strains. AMR genes responsible for resistance to broad-spectrum ß-lactams, chloramphenicols, tetracyclines, macrolides, and aminoglycosides were associated with mobile genetic elements or plasmid replicons or both. These results suggest that cyanobacteria are an extensive reservoir, and potential vector, for AMR genes in diverse terrestrial and aquatic habitats.

Hive Transplantation Has Minimal Impact on the Core Gut Microbiome of the Australian Stingless Bee, Tetragonula carbonaria.

Bacteria residing in the guts of pollinating insects play a key role in nutrient acquisition, digestion, and resistance to pests and diseases. Imbalances in microbial flora in response to environmental change and stress can therefore impact insect health and resilience. This study is aimed at defining the core gut microbiome of the Australian native stingless bee, Tetragonula carbonaria, and exploring the impact of colony transplantation on gut health. The gut microbiomes of nine forager bees from natural (log) and manufactured (box) hives were examined via 16S rRNA gene amplicon sequencing. Some differences were observed at the ASV level between the microbiomes of log and box hive bees. However, a core microbiome, dominated by Lactobacillus spp., unclassified Acetobacteraceae spp., and Bombella spp., was maintained. Further, the inferred functional potential of the microbiomes was consistent across all individuals. This study highlights that although hive transplantation has an impact on the overall diversity of stingless bee gut microbiomes, it is unlikely to have a significant negative impact on the overall health and resilience of the colony.

Australian bush medicines harbour diverse microbial endophytes with broad-spectrum antibacterial activity.

AIMS: Microbial endophytes produce specialized metabolites, including antibiotics and other compounds of pharmaceutical and agricultural value. This study aimed to investigate the diversity and bioactivity of endophytes from medicinal plants used by the Dharawal People of Gamay (Botany Bay), Australia. METHODS AND RESULTS: Of the 48 endophytes isolated, 19 tested positive for polyketide synthase or non-ribosomal peptide synthetase genes via a PCR incorporating degenerate primers. The biosynthetically talented endophytes were identified by 16S rRNA gene sequencing and included 4 bacteria species belonging to the orders Bacillales, Rhizobiales and Burkholderiales and 15 Ascomycota fungi species belonging to the orders Botryosphaeriales, Cladosporiales, Glomerellales, Microascales and Eurotiales. Antimicrobial testing using the disc diffusion assay demonstrated that 15 of the 19 isolates had broad-spectrum activity against a range of Gram-positive and Gram-negative bacteria. CONCLUSIONS: Taken together, these results suggest that Australian bush medicines harbour diverse biosynthetically talented microbial endophytes capable of producing broad-spectrum antibacterial compounds. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that compounds produced by microbial endophytes likely contribute to the collective medicinal properties of Australian bush medicines. Significantly, it highlights that Indigenous botanical knowledge and modern molecular approaches can be used in tandem to prioritize microorganisms that produce pharmaceutically relevant compounds.

Functional characterization of the twin ZIP/SLC39 metal transporters, NpunF3111 and NpunF2202 in Nostoc punctiforme.

The ZIP family of metal transporters is involved in the transport of Zn(2+) and other metal cations from the extracellular environment and/or organelles into the cytoplasm of prokaryotes, eukaryotes and archaeotes. In the present study, we identified twin ZIP transporters, Zip11 (Npun_F3111) and Zip63 (Npun_F2202) encoded within the genome of the filamentous cyanobacterium, Nostoc punctiforme PCC73120. Sequence-based analyses and structural predictions confirmed that these cyanobacterial transporters belong to the SLC39 subfamily of metal transporters. Quantitative real-time (QRT)-PCR analyses suggested that the enzymes encoded by zip11 and zip63 have a broad allocrite range that includes zinc as well as cadmium, cobalt, copper, manganese and nickel. Inactivation of either zip11 or zip63 via insertional mutagenesis in N. punctiforme resulted in reduced expression of both genes, highlighting a possible co-regulation mechanism. Uptake experiments using (65)Zn demonstrated that both zip mutants had diminished zinc uptake capacity, with the deletion of zip11 resulting in the greatest overall reduction in (65)Zn uptake. Over-expression of Zip11 and Zip63 in an E. coli mutant strain (ZupT736::kan) restored divalent metal cation uptake, providing further evidence that these transporters are involved in Zn uptake in N. punctiforme. Our findings show the functional role of these twin metal uptake transporters in N. punctiforme, which are independently expressed in the presence of an array of metals. Both Zip11 and Zip63 are required for the maintenance of homeostatic levels of intracellular zinc N. punctiforme, although Zip11 appears to be the primary zinc transporter in this cyanobacterium, both ZIP's may be part of a larger metal uptake system with shared regulatory elements.

NtcA from Microcystis aeruginosa PCC 7806 is autoregulatory and binds to the microcystin promoter.

NtcA is a transcription factor that has been found in a diverse range of cyanobacteria. This nitrogen-controlled factor was focused on as a key component in the yet-to-be-deciphered regulatory network controlling microcystin production. Adaptor-mediated PCR was utilized to isolate the ntcA gene from Microcystis aeruginosa PCC 7806. This gene was cloned, and the recombinant (His-tagged) protein was overexpressed and purified for use in mobility shift assays to analyze NtcA binding to putative sites identified in the microcystin mcyA/D promoter region. Autoregulation of NtcA in M. aeruginosa was shown via NtcA binding in the upstream ntcA promoter region. The observation of binding of NtcA to the mcyA/D promoter region has direct relevance for the regulation of microcystin biosynthesis, as transcription of the mcyABCDEFGHIJ gene cluster appears to be under direct control of nitrogen.

The molecular genetics and regulation of cyanobacterial peptide hepatotoxin biosynthesis.

Over the last 10 years, we have witnessed major advances in our understanding of natural product biosynthesis, including the genetic basis for toxin production by numerous groups of cyanobacteria. Cyanobacteria produce an unparalleled array of bioactive secondary metabolites, including alkaloids, polyketides and non-ribosomal peptides, some of which are potent toxins. This review addresses the molecular genetics underlying the production of hepatotoxins, microcystin and nodularin in fresh and brackish water. These toxins pose a serious threat to human health and their occurrence in water supplies is increasing, because of the prevalence of toxic algal blooms worldwide. Toxin biosynthesis gene-cluster-associated transposition and the natural transformability of certain species suggest a broader distribution of toxic cyanobacterial taxa. The information gained from the discovery of these toxin biosynthetic pathways has enabled the genetic screening of various environments for drinking-water quality management. Understanding the role of cyanotoxins in the producing microorganisms and the environmental regulation of their biosynthesis genes may also suggest the means of controlling toxic-bloom events.