- Volume 166, Issue 4, 2020

Different model systems have, over the years, contributed to our current understanding of the molecular mechanisms underpinning the various types of interaction between bacteria and their animal hosts. The genus Photorhabdus comprises Gram-negative insect pathogenic bacteria that are normally found as symbionts that colonize the gut of the infective juvenile stage of soil-dwelling nematodes from the family Heterorhabditis. The nematodes infect susceptible insects and release the bacteria into the insect haemolymph where the bacteria grow, resulting in the death of the insect. At this stage the nematodes feed on the bacterial biomass and, following several rounds of reproduction, the nematodes develop into infective juveniles that leave the insect cadaver in search of new hosts. Therefore Photorhabdus has three distinct and obligate roles to play during this life-cycle: (1) Photorhabdus must kill the insect host; (2) Photorhabdus must be capable of supporting nematode growth and development; and (3) Photorhabdus must be able to colonize the gut of the next generation of infective juveniles before they leave the insect cadaver. In this review I will discuss how genetic analysis has identified key genes involved in mediating, and regulating, the interaction between Photorhabdus and each of its invertebrate hosts. These studies have resulted in the characterization of several new families of toxins and a novel inter-kingdom signalling molecule and have also uncovered an important role for phase variation in the regulation of these different roles.

Myxobacteria exhibit complex social behaviors such as predation, outer membrane exchange and fruiting body formation. These behaviors depend on coordinated movements of cells on solid surfaces that involve social (S) motility. S-motility is powered by extension-retraction cycles of type 4 pili (Tfp) and exopolysaccharides (EPS) that provide a matrix for group cellular movement. Here, we characterized a new class of S-motility mutants in Myxococcus xanthus . These mutants have a distinctive phenotype: they lack S-motility even though they produce pili and EPS and the phenotype is temperature-sensitive. The point mutations were mapped to a single locus, MXAN_3284, named sglT. Similar to pilT mutants, sglT mutants are hyperpiliated and, strikingly, the temperature-sensitive phenotype is caused by null mutations. Our results indicate that SglT plays a critical role in Tfp function associated with pilus retraction and that the block in pili retraction is caused by a Tfp assembly defect in the absence of SglT at high-temperature growth.

Abstract

Iron (Fe), which is a necessary micronutrient for algal growth, plays an important role in the physiological metabolism and enzymatic reactions of algae. This study aimed to investigate the absorption process of four kinds of complexing iron absorbed by Anabaena flos-aquae. Results showed that the absorptive capacity of A. flos-aquae to complex iron was inversely proportional to the stability of the complex bond of complex iron. Complex iron with weak binding ability can be quickly adsorbed by A. flos-aquae. The absorptive rate was as follows: ferric humate, ferric oxalate >ammonium ferric citrate >EDTA Fe. For EDTA-Fe with a strong binding ability, a moderate iron concentration (e.g. 0.6 mg l−1) is favourable for iron uptake by A. flos-aquae. Our experiments also revealed that the process of separating iron from complex iron before entering algal cells was probably as follows: iron complexed with organic ligands were firstly adsorbed on the surface of algae cells; afterwards, iron ions were captured by organic matter on the surface of algae cells, accompanied by the rupture of the bond between Fe3+ and ligand; finally, the Fe3+ entered into the cell of algae while the organic ligands returned to the medium.

The ciprofloxacin-resistance crpP gene, encoded by the pUM505 plasmid, isolated from a P. aeruginosa clinical isolate, confers an enzymatic mechanism of antibiotic phosphorylation, which is ATP-dependent, that decreases ciprofloxacin susceptibility. Homologous crpP genes are distributed across extended spectrum beta-lactamase (ESBL)-producing isolates obtained from Mexican hospitals and which confer decreased susceptibility to CIP. The analysis of sequences of the CrpP of proteins showed that the residues Gly7, Thr8, Asp9, Lys33 and Gly34 (located at the N-terminal region) and Cys40 (located at the C-terminal region) are conserved in all proteins, suggesting that these residues could be essential for CrpP function. The aim of this study was to investigate the amino acids essential to ciprofloxacin resistance, which is conferred by the CrpP protein of pUM505 plasmid. Mutations in the codons encoding Gly7, Asp9, Lys33 and Cys40 of CrpP protein from pUM505 were generated by PCR fusion. The results showed that all mutations generated in CrpP proteins increased ciprofloxacin susceptibility in Escherichia coli . In addition, the CrpP modified proteins were purified and their enzymatic activity on ciprofloxacin was assayed, showing that these modified proteins do not exert catalytic activity on ciprofloxacin. Moreover, by infrared assays it was determined that the modified proteins were are not able to modify the ciprofloxacin molecule. Our findings are the first report that indicate that the amino acids, namely Gly7, Asp9, Lys33 and Cys40, which are conserved in the CrpP proteins, possess an essential role for the enzymatic mechanism that confers ciprofloxacin resistance.

This study detailed the responses of Galleria mellonella larvae to disseminated infection caused by co-infection with Candida albicans and Staphylococcus aureus . Doses of C. albicans (1×105 larva−1) and S. aureus (1×104 larva−1) were non-lethal in mono-infection but when combined significantly (P<0.05) reduced larval survival at 24, 48 and 72 h relative to larvae receiving S. aureus (2×104 larva−1) alone. Co-infected larvae displayed a significantly higher density of S. aureus larva−1 compared to larvae infected solely with S. aureus . Co-infection resulted in dissemination throughout the host and the appearance of large nodules. Co-infection of larvae with C. albicans and S. aureus (2×104 larva−1) resulted in an increase in the density of circulating haemocytes compared to that in larvae infected with only S. aureus . Proteomic analysis of co-infected larval haemolymph revealed increased abundance of proteins associated with immune responses to bacterial and fungal infection such as cecropin-A (+45.4-fold), recognition proteins [e.g. peptidoglycan-recognition protein LB (+14-fold)] and proteins associated with nodule formation [e.g. Hdd11 (+33.3-fold)]. A range of proteins were also decreased in abundance following co-infection, including apolipophorin (−62.4-fold), alpha-esterase 45 (−7.7-fold) and serine proteinase (−6.2-fold). Co-infection of larvae resulted in enhanced proliferation of S. aureus compared to mono-infection and an immune response showing many similarities to the innate immune response of mammals to infection. The utility of G. mellonella larvae for studying polymicrobial infection is highlighted.

Chemolithotrophic sulfur oxidation represents a significant part of the biogeochemical cycling of this element. Due to its long evolutionary history, this ancient metabolism is well known for its extensive mechanistic and phylogenetic diversification across a diverse taxonomic spectrum. Here we carried out whole-genome sequencing and analysis of a new betaproteobacterial isolate, Pusillimonas ginsengisoli SBSA, which is found to oxidize thiosulfate via the formation of tetrathionate as an intermediate. The 4.7 Mb SBSA genome was found to encompass a soxCDYZAXOB operon, plus single thiosulfate dehydrogenase (tsdA) and sulfite : acceptor oxidoreductase (sorAB) genes. Recombination-based knockout of tsdA revealed that the entire thiosulfate is first converted to tetrathionate by the activity of thiosulfate dehydrogenase (TsdA) and the Sox pathway is not functional in this bacterium despite the presence of all necessary sox genes. The ∆soxYZ and ∆soxXA knockout mutants exhibited a wild-type-like phenotype for thiosulfate/tetrathionate oxidation, whereas ∆soxB, ∆soxCD and soxO::KanR mutants only oxidized thiosulfate up to tetrathionate intermediate and had complete impairment in tetrathionate oxidation. The substrate-dependent O2 consumption rate of whole cells and the sulfur-oxidizing enzyme activities of cell-free extracts, measured in the presence/absence of thiol inhibitors/glutathione, indicated that glutathione plays a key role in SBSA tetrathionate oxidation. The present findings collectively indicate that the potential glutathione : tetrathionate coupling in P. ginsengisoli involves a novel enzymatic component, which is different from the dual-functional thiol dehydrotransferase (ThdT), while subsequent oxidation of the sulfur intermediates produced (e.g. glutathione : sulfodisulfane molecules) may proceed via the iterative action of soxBCD .

The transcriptional regulator PlcR, its cognate cell-cell signaling heptapeptide PapR7, and the oligopeptide permease OppABCDF, required for PapR7 import, form a quorum-sensing system that controls the expression of virulence factors in Bacillus cereus and Bacillus thuringiensis species. In B. cereus strain ATCC 14579, the transcriptional regulator PlcRa activates the expression of abrB2 gene, which encodes an AbrB-like transcriptional regulator involved in cysteine biosynthesis. PlcRa is a structural homolog of PlcR: in particular, its C-terminal TPR peptide-binding domain could be similarly arranged as in PlcR. The signaling peptide of PlcRa is not known. As PlcRa is a PlcR-like protein, the cognate PapR7 peptide (ADLPFEF) is a relevant candidate to act as a signaling peptide for PlcRa activation. Also, the putative PapRa7 peptide (CSIPYEY), encoded by the papRa gene adjacent to the plcRa gene, is a relevant candidate as addition of synthetic PapRa7 induces a dose-dependent increase of abrB2 expression. To address the issue of peptide selectivity of PlcRa, the role of PapR and PapRa peptides in PlcRa activity was investigated in B. thuringiensis 407 strain, by genetic and functional complementation analyses. A transcriptional fusion between the promoter of abrB2 and lacZ was used to monitor the PlcRa activity in various genetic backgrounds. We demonstrated that PapR was necessary and sufficient for PlcRa activity. We showed that synthetic PapRs from pherogroups II, III and IV and synthetic PapRa7 were able to trigger abrB2 expression, suggesting that PlcRa is less selective than PlcR. Lastly, the mode of binding of PlcRa was addressed using an in silico approach. Overall, we report a new role for PapR as a signaling peptide for PlcRa activity and show a functional link between PlcR and PlcRa regulons in B. thuringiensis .