- Volume 4, Issue 5, 2022

Meteoritic material accumulated on the surface of the anoxic Early Earth during the Late Heavy Bombardment around 4.0 Gya. These meteorites may have provided the Earth with extra-terrestrial nutrients and energy sources for early life. How could the presence of meteorites have affected the origin and evolution of early life on Earth? And what is the influence of geothermal activity on the Earth’s surface? This research investigates the growth of an anaerobic microbial community from pond sediment on non-pyrolyzed (pristine) or pyrolyzed (heat-treated) carbonaceous chondrite ‘Cold Bokkeveld’. A microbial community was grown anaerobically in batch cultures containing a liquid environment and powdered non-pyrolyzed or pyrolyzed Cold Bokkeveld. Cell concentrations were measured by Colony-Forming Units on agar plates. The community composition in the presence of non-pyrolyzed meteorite was determined by 16S rRNA amplicon sequencing. Non-pyrolyzed Cold Bokkeveld supported the growth of a stable, anaerobic community containing mainly the Deltaproteobacteria Geobacteraceae and Desulfuromonadaceae. Members of these families are known to use elemental sulfur and ferric iron as electron acceptors, and organic compounds as electron donors. Pyrolyzed Cold Bokkeveld however, was inhibitory to the growth of the microbial community. These results show that carbonaceous chondrites can host an anaerobic microbial community, but that pyrolysis, e.g. by geothermal activity, can inhibit microbial growth and potentially toxify the material. This indicates that extraterrestrial meteoritic material and the environment on Early Earth could have shaped the nature of early microbial ecosystems by enhancing growth of microorganisms with metabolic capabilities favored in the presence of this material.

Bacteriocins are proteinaceous antimicrobials produced by bacteria which are active against other strains of the same species. R-type pyocins are phage tail-like bacteriocins produced by Pseudomonas aeruginosa. Due to their anti-pseudomonal activity, R-pyocins have potential as therapeutics in infection. P. aeruginosa is a Gram-negative opportunistic pathogen and is particularly problematic for individuals with cystic fibrosis (CF). P. aeruginosa from CF lung infections develop increasing resistance to antibiotics, making new treatment approaches essential. P. aeruginosa populations become phenotypically and genotypically diverse during infection, however, little is known of the efficacy of R-pyocins against heterogeneous populations. R-pyocins vary by subtype (R1-R5), distinguished by binding to different residues on the lipopolysaccharide (LPS). Each type varies in killing spectrum, and each strain produces only one R-type. To evaluate the prevalence of different R-types, we screened P. aeruginosastrains from the International Pseudomonas Consortium Database (IPCD) and from our biobank of CF strains. We found that (i) R1-types were the most prevalent R-type among strains from respiratory sources; (ii) there are a large number of strains lacking R-pyocin genes, and (iii) isolates collected from the same patient have the same R-type. We then assessed the impact of diversity on R-pyocin susceptibility and found a heterogenous response to R-pyocins within populations, likely due to differences in the LPS core. Our work reveals that heterogeneous populations of microbes exhibit variable susceptibility to R-pyocins and highlights that there is likely heterogeneity in response to other types of LPS-binding antimicrobials, including phage.

The experience with SARS-CoV-2 that spread rapidly throughout the world makes us realize we need protocols to act quickly against unknown pathogens. The immune system protects recovering patients from any pathogen by producing antibodies against their immunogenic epitopes. Therefore, the strategy presented here is based on convalescent blood samples and a phage display platform for antibody and peptide drug discovery. Peptide libraries are screened against purified convalescent antibodies to identify immunogenic epitopes of the pathogen. Furthermore, the B cells of the recovery patients are used to amplify variable domains of antibody heavy and light chains expressed during the infection. These domains are cloned in a phagemid and produce free phage particles expressing the antibody fragments on their surfaces to select binders to pathogen immunogenic epitopes. These findings are essential in the identification of the unknown pathogen and the design of therapeutic molecules. In conclusion, this report describes a phage display strategy to combat outbreaks of unknown pathogens such as SARS-CoV-2.

Shigellosis is an intestinal infection caused by Shigella bacteria. Shigella cause an estimated ~200,000 global deaths annually. Antimicrobial resistant (AMR) shigellosis is a significant cause of morbidity in high-income nations, with both multidrug resistant (MDR) and extensively drug resistant (XDR) cases being increasingly reported in Australia, England, and the USA. Sexually transmissible shigellosis was first described in San Francisco, 1974, but it would be a further 30 years before its first description in England. In 2004, London experienced an outbreak of Shigella sonnei (S. sonnei) mediated sexually transmitted shigellosis, associated with men-who-have-sex-with-men (MSM). Since then, sexually transmissible shigellosis has become endemic in England, with a greater than two-fold increase in Shigella diagnoses within sexual health services from 2015 to 2019. Through genomic exploration of samples from the original 2004 outbreak (provided by Public Health England (PHE)), we identified that the 2004 London outbreak isolates clustered within the base of genotype 3.1, lineage III, a lineage which has since gone on to dominate the global epidemiology of S. sonnei. The isolates displayed early evidence of varying degrees of antimicrobial resistance to several drug classes: macrolides, tetracyclines, beta-lactams and sulphonamides. Reconstructing the chronological process of how shigellosis has arrived at its current position in AMR and transmissibility is critical. Further investigation is underway to link this outbreak with MSM-associated shigellosis outbreaks occurring in the early 2000s in other countries to establish whether this lineage globally disseminated; determine the timeframe for global connectivity of shigellosis; and examine the outbreak isolates for virulence determinants.

Phthalate esters (PEs) are environmentally ubiquitous micropollutants that are used as plasticizers and additives in diverse consumer products. Considerable concern relates to their reported xenoestrogenicity and the microbial-based attenuation of environmental PE concentrations is of interest to combat harmful downstream effects. Fungal PE catabolism has received less attention than that by bacteria, and particularly marine fungal species remain largely overlooked in this respect. We have compared the biocatalytic and biosorptive removal rates of di-n-butyl phthalate (DBP) and diethyl phthalate (DEP), chosen as two environmentally prominent PE representatives (exhibiting differing structures and hydrophobicities), by marine- and freshwater fungal strains. Bisphenol A, both an extensively used plastic additive and prominent environmental xenoestrogen, was included as a reference compound due to its previously well-documented fungal degradation. Partial pathways for DBP metabolization by these ecophysiologically diverse ascomycetes were proposed with the help of UPLC-QTOF-MS analysis. Species-specific biochemical reaction steps contributing to DBP metabolism were also observed. The involved reactions include initial cytochrome P450-dependent monohydroxylations of DBP with subsequent further oxidation of related metabolites, de-esterification via either hydrolytic cleavage or cytochrome P450-dependent oxidative O-dealkylation, transesterification, and demethylation steps - finally yielding phthalic acid as a central intermediate in all pathways. Beyond previous research into fungal PE metabolism which emphasises hydrolytic de-esterification as the primary catabolic step, a prominent role of cytochrome P450 monooxygenase-catalysed reactions is established.

In bacteria, adaptation to changes in the environment is mainly controlled through two-component signal transduction systems (TCSs). Most bacteria contain dozens of TCSs, each of them responsible for sensing a different range of signals and controlling the expression of a repertoire of target genes (regulon). Frequently, TCS control key physiological changes required for pathogenesis and/or antimicrobial resistance. Over the years, identification of the regulon controlled by each individual TCS in different bacteria has been a recurrent question. However, limitations associated with the classical approaches used have left our knowledge far from complete. In this report, using a pioneering approach in which a strain devoid of the complete nonessential TCS network was systematically complemented with the constitutively active form of each response regulator, we have reconstituted the regulon of each TCS of S. aureus in the absence of interference between members of the family. Transcriptome sequencing (RNA-Seq) and proteomics allowed us to determine the size, complexity, and insulation of each regulon and to identify the genes regulated exclusively by one or many TCSs. This gain-of-function strategy provides the first description of the complete TCS regulon in a living cell, which we expect will be useful to understand the pathobiology of this important pathogen.

Enteric pathogens exploit the versatility of cytoskeletal system for internalization into non-phagocytic cells, as a crucial step in their pathogenic life cycle. Enteropathogenic Escherichia coli(EPEC) andEnterohemorrhagic Escherichia coli (EHEC) were shown to form actin pedestals in host cells using type-III secretion system. Earlier, we reported that EAEC induced increase in intracellular calcium ions might have crucial role in F-actin rearrangements in INT-407 cells leading to its invasion. It was suggested that EGFR might contribute in Rck-mediated adherence and invasion of Salmonella in host cells. In the present study, we assessed the role of EAEC induced activated EGFR in human intestinal epithelial cell lines (INT-407 & HCT-15). The presence of activated EGFR was detected in membrane fractions ofeach cell line, infected with two different strains of EAEC (EAEC-T8 & EAEC-O42) separatelyfor 3h in presence and absence of Tyrphostin AG1478 (EGFR-inhibitor), by western immunoblotting using p-EGFR (Y1068) antibody. Adherence and invasion of EAEC-T8 to each cell line were checked in presence of Tyrphostin AG1478. Further, the effect of Tyrphostin AG1478 on cytoskeletal F-actin rearrangement of EAEC-T8 infected cells was assessed under confocal microscope following staining with TRITC-phalloidin. EAEC-T8 induced maximum increase in EGFR autophosphorylation at Y1068. Adherence and invasion of EAEC-T8 as well as this organism induced cytoskeletal F-actin polymerization were found to be inhibited in presence of Tyrphostin AG1478. Our study revealed that EAEC induced activated EGFR might play a major role in host cell adherence and cytoskeletal rearrangements leading to invasion of the organism in these cells.