Bacterial responses to background organic pollutants in the northeast subarctic Pacific Ocean

Thousands of man-made synthetic chemicals are released to oceans and compose the anthropogenic dissolved organic carbon (ADOC). Little is known about the effects of this chronic pollution on marine microbiome activities. In this study, we measured the pollution level at three sites in the Northeast Subarctic Pacific Ocean (NESAP) and investigated how mixtures of three model families of ADOC at different environmentally relevant concentrations affected naturally occurring marine bacterioplankton communities' structure and metabolic functioning. The offshore northernmost site (North) had the lowest concentrations of hydrocarbons, as well as organophosphate ester plasticizers, contrasting with the two other continental shelf sites, the southern coastal site (South) being the most contaminated. At North, ADOC stimulated bacterial growth and promoted an increase in the contribution of some Gammaproteobacteria groups (e.g. Alteromonadales) to the 16 rRNA pool. These groups are described as fast responders after oil spills. In contrast, minor changes in South microbiome activities were observed. Gene expression profiles at Central showed the coexistence of ADOC degradation and stress-response strategies to cope with ADOC toxicities. These results show that marine microbial communities at three distinct domains in NESAP are influenced by background concentrations of ADOC, expanding previous assessments for polar and temperate waters.     

Ancient saltern metagenomics: tracking changes in microbes and their viruses from the underground to the surface

Microbial communities in hypersaline underground waters derive from ancient organisms trapped within the evaporitic salt crystals and are part of the poorly known subterranean biosphere. Here, we characterized the viral and prokaryotic assemblages present in the hypersaline springs that dissolve Triassic-Keuper evaporite rocks and feed the Añana Salt Valley (Araba/Alava, Basque Country, Spain). Four underground water samples (around 23% total salinity) with different levels of exposure to the open air were analysed by means of microscopy and metagenomics. Cells and viruses in the spring water had lower concentrations than what are normally found in hypersaline environments and seemed to be mostly inactive. Upon exposure to the open air, there was an increase in activity of both cells and viruses as well as a selection of phylotypes. The underground water was inhabited by a rich community harbouring a diverse set of genes coding for retinal binding proteins. A total of 35 viral contigs from 15 to 104 kb, representing partial or total viral genomes, were assembled and their evolutionary changes through the spring system were followed by SNP analysis and metagenomic island tracking. Overall, both the viral and the prokaryotic assemblages changed quickly upon exposure to the open air conditions.     

Therapeutic potential of periodontal ligament stem cells

Inflammatory periodontal disease known as periodontitis is one of the most common conditions that affect human teeth and often leads to tooth loss. Due to the complexity of the periodontium, which is composed of several tissues, its regeneration and subsequent return to a homeostatic state is challenging with the therapies currently available. Cellular therapy is increasingly becoming an alternative in regenerative medicine/dentistry, especially therapies using mesenchymal stem cells, as they can be isolated from a myriad of tissues. Periodontal ligament stem cells (PDLSCs) are probably the most adequate to be used as a cell source with the aim of regenerating the periodontium. Biological insights have also highlighted PDLSCs as promising immunomodulator agents. In this review, we explore the state of knowledge regarding the properties of PDLSCs, as well as their therapeutic potential, describing current and future clinical applications based on tissue engineering techniques.     

Optimization and validation of a cost‐effective protocol for biosurveillance of invasive alien species

Environmental DNA (eDNA) metabarcoding has revolutionized biodiversity monitoring and invasive pest biosurveillance programs. The introduction of insect pests considered invasive alien species (IAS) into a non‐native range poses a threat to native plant health. The early detection of IAS can allow for prompt actions by regulating authorities, thereby mitigating their impacts. In the present study, we optimized and validated a fast and cost‐effective eDNA metabarcoding protocol for biosurveillance of IAS and characterization of insect and microorganism diversity. Forty‐eight traps were placed, following the CFIA''s annual forest insect trapping survey, at four locations in southern Ontario that are high risk for forest IAS. We collected insects and eDNA samples using Lindgren funnel traps that contained a saturated salt (NaCl) solution in the collection jar. Using cytochrome c oxidase I (COI) as a molecular marker, a modified Illumina protocol effectively identified 2,535 Barcode Index Numbers (BINs). BINs were distributed among 57 Orders and 304 Families, with the vast majority being arthropods. Two IAS (Agrilus planipennis and Lymantria dispar) are regulated by the Canadian Food Inspection Agency (CFIA) as plant health pests, are known to occur in the study area, and were identified through eDNA in collected traps. Similarly, using 16S ribosomal RNA and nuclear ribosomal internal transcribed spacer (ITS), five bacterial and three fungal genera, which contain species of regulatory concern across several Canadian jurisdictions, were recovered from all sampling locations. Our study results reaffirm the effectiveness and importance of integrating eDNA metabarcoding as part of identification protocols in biosurveillance programs.     

Arginine substitution by alanine at the P1 position increases the selectivity of CmPI-II,a non-classical Kazal inhibitor

CmPI-II is a Kazal-type tight-binding inhibitor isolated from the Caribbean snail Cenchritis muricatus. This inhibitor has an unusual specificity in the Kazal family, as it can inhibit subtilisin A (SUBTA), elastases and trypsin. An alanine in CmPI-II P1 site could avoid trypsin inhibition while improving/maintaining SUBTA and elastases inhibition. Thus, an alanine mutant of this position (rCmPI-II R12A) was obtained by site-directed mutagenesis. The gene cmpiR12A was expressed in P. pastoris KM71H yeast. The recombinant protein (rCmPI-II R12A) was purified by the combination of two ionic exchange chromatography (1:cationic, 2 anionic) followed by and size exclusion chromatography. The N-terminal sequence obtained as well as the experimental molecular weight allowed verifying the identity of the recombinant protein, while the correct folding was confirmed by CD experiments. rCmPI-II R12A shows a slightly increase in potency against SUBTA and elastases. The alanine substitution at P1 site on CmPI-II abolishes the trypsin inhibition, confirming the relevance of an arginine residue at P1 site in CmPI-II for trypsin inhibition and leading to a molecule with more potentialities in biomedicine.     

The citrus plant pathogen Xanthomonas citri has a dual polyamine-binding protein

ATP-Binding Cassette transporters (ABC transporters) are protein complexes involved in the import and export of different molecules, including ions, sugars, peptides, drugs, and others. Due to the diversity of substrates, they have large relevance in physiological processes such as virulence, pathogenesis, and antimicrobial resistance. In Xanthomonas citri subsp. citri, the phytopathogen responsible for the citrus canker disease, 20% of ABC transporters components are expressed under infection conditions, including the putative putrescine/polyamine ABC transporter, PotFGHI. Polyamines are ubiquitous molecules that mediate cell growth and proliferation and play important role in bacterial infections. In this work, we characterized the X. citri periplasmic-binding protein PotF (XAC2476) using bioinformatics, biophysical and structural methods. PotF is highly conserved in Xanthomonas sp. genus, and we showed it is part of a set of proteins related to the import and assimilation of polyamines in X. citri. The interaction of PotF with putrescine and spermidine was direct and indirectly shown through fluorescence spectroscopy analyses, and experiments of circular dichroism (CD) and small-angle X-ray scattering (SAXS), respectively. The protein showed higher affinity for spermidine than putrescine, but both ligands induced structural changes that coincided with the closing of the domains and increasing of thermal stability.     

Micropropagation of Zeyheria montana Mart. (Bignoniaceae), an endangered endemic medicinal species from the Brazilian cerrado biome

Zeyheria montana Mart. has become endangered, primarily because of deforestation of its habitat, its use as a medicinal plant extract, and the strong endemism of the species. In this study, an efficient protocol was established for the micropropagation and conservation of Z. montana germplasm using isolated mature zygotic embryos as explants. Embryos germinated in vitro 4 d after isolation and inoculation on modified Murashige and Skoog (MS) medium containing 2.0 mg/l gibberellic acid (GA₃). The addition of GA₃ also improved the germination index and accelerated the process of germination. Nodal segments from seedlings were placed on modified ¼-strength MS medium containing 0.1 mg/l 6-benzyladenine and 0.5 mg/l GA₃. Nodal segments produced 7.3 shoots per explant within 60 d. Following transfer of shoots to a medium containing 1.5 mg/l indole-3-butyric acid, roots formed. All plantlets obtained were successfully acclimatized under greenhouse conditions, and approximately 68.5 acclimatized plants could be obtained per seed each year. This protocol provides a method to preserve this rare and endangered medicinal plant.     

Systematics and evolutionary history of butterflies in the “Taygetis clade” (Nymphalidae: Satyrinae: Euptychiina): Towards a better understanding of Neotropical biogeography

The so-called “Taygetis clade” is a group of exclusively Neotropical butterflies classified within Euptychiina, one of the largest subtribes in the subfamily Satyrinae. Since the distribution of the ten genera belonging to this group ranges throughout the entire Neotropics, from lowlands to lower montane habitats, it offers a remarkable opportunity to study the region’s biogeographic history as well as different scenarios for speciation in upland areas. We inferred a robust and well-sampled phylogeny using DNA sequences from four genes (4035 bp in total) using maximum parsimony and Bayesian inference. We estimated divergence times using the Bayesian relaxed clock method calibrated with node ages from previous studies. Ancestral ranges of distribution were estimated using the dispersal–extinction–cladogenesis (DEC) model as implemented in the program Lagrange. We propose several taxonomic changes and recognize nine well-supported natural genera within the “Taygetis clade”: Forsterinaria (subsuming Guaianaza syn. nov.), Parataygetis, Posttaygetis, Harjesia (excluding Harjesia griseola and Harjesia oreba), Pseudodebis (including Taygetomorpha syn. nov.,), Taygetina (subsuming Coeruleotaygetis syn. nov., Harjesia oreba comb. nov., Taygetis weymeri comb. nov. and Taygetis kerea comb. nov.), Taygetis (excluding Taygetis ypthima, Taygetis rectifascia, Taygetis kerea and Taygetis weymeri), and two new genera, one containing Harjesia griseola, and the other Taygetis ypthima and Taygetis rectifascia. The group diversified mainly during late Miocene to Pliocene, coinciding with the period of drastic changes in landscape configuration in the Neotropics. Major dispersals inferred from the Amazon basin towards northwestern South America, the Atlantic forests and the eastern slope of the Andes have mostly shaped the evolution and diversification of the group. Furthermore, expansion of larval dietary repertoire might have aided net diversification in the two largest genera in the clade, Forsterinaria and Taygetis.     

Phloroglucinol in plant tissue culture

In plant tissue culture research, there is a constant need to search for novel substances that could result in better or more efficient growth in vitro. A relatively unknown compound, phloroglucinol (1,3,5-trihydroxybenzene), which is a degradation product of phloridzin, has growth-promoting properties. Phloroglucinol increases shoot formation and somatic embryogenesis in several horticultural and grain crops. When added to rooting media together with auxin, phloroglucinol further stimulates rooting, most likely because phloroglucinol and its homologues act as auxin synergists or auxin protectors. Of particular interest is the ability of phloroglucinol—a precursor in the lignin biosynthesis pathway—to effectively control hyperhydricity through the process of lignification, thus maximizing the multiplication rate of woody species and other species that are difficult to propagate. Phloroglucinol has also been used to improve the recovery of cryopreserved Dendrobium protocorms, increasing the potential of cryopreservation for application in ornamental biotechnology. Phloroglucinol demonstrates both cytokinin-like and auxin-like activity, much like thidiazuron, and thus has considerable potential for application in a wide range of plant tissue culture studies.