The unique N-terminal region of Mycobacterium tuberculosis sigma factor A plays a dominant role in the essential function of this protein.

SigA (σA) is an essential protein and the primary sigma factor in Mycobacterium tuberculosis (Mtb). However, due to the absence of genetic tools, our understanding of the role and regulation of σA activity and its molecular attributes that help modulate Mtb survival is scant. Here, we generated a conditional gene replacement of σA in Mtb and showed that its depletion results in a severe survival defect in vitro, ex vivo, and in vivo in a murine infection model. Our RNA-seq analysis suggests that σA either directly or indirectly regulates ∼57% of the Mtb transcriptome, including ∼28% of essential genes. Surprisingly, we note that despite having ∼64% similarity with σA, overexpression of the primary-like σ factor SigB (σB) fails to compensate for the absence of σA, suggesting minimal functional redundancy. RNA-seq analysis of the Mtb σB deletion mutant revealed that 433 genes are regulated by σB, of which 283 overlap with the σA transcriptome. Additionally, surface plasmon resonance, in vitro transcription, and functional complementation experiments reveal that σA residues between 132-179 that are disordered and missing from all experimentally determined σA-RNAP structural models are imperative for σA function. Moreover, phosphorylation of σA in the intrinsically disordered N-terminal region plays a regulatory role in modulating its activity. Collectively, these observations and analysis provide a rationale for the centrality of σA for the survival and pathogenicity of this bacillus.

Genomic and transcriptomic profiling of phoenix colonies.

Pseudomonas aeruginosa is a Gram-negative bacterium responsible for numerous human infections. Previously, novel antibiotic tolerant variants known as phoenix colonies as well as variants similar to viable but non-culturable (VBNC) colonies were identified in response to high concentrations of aminoglycosides. In this study, the mechanisms behind phoenix colony and VBNC-like colony emergence were further explored using both whole genome sequencing and RNA sequencing. Phoenix colonies were found to have a single nucleotide polymorphism (SNP) in the PA4673 gene, which is predicted to encode a GTP-binding protein. No SNPs were identified within VBNC-like colonies compared to the founder population. RNA sequencing did not detect change in expression of PA4673 but revealed multiple differentially expressed genes that may play a role in phoenix colony emergence. One of these differentially expressed genes, PA3626, encodes for a tRNA pseudouridine synthase which when knocked out led to a complete lack of phoenix colonies. Although not immediately clear whether the identified genes in this study may have interactions which have not yet been recognized, they may contribute to the understanding of how phoenix colonies are able to emerge and survive in the presence of antibiotic exposure.

Genetically stable CRISPR-based kill switches for engineered microbes.

Microbial biocontainment is an essential goal for engineering safe, next-generation living therapeutics. However, the genetic stability of biocontainment circuits, including kill switches, is a challenge that must be addressed. Kill switches are among the most difficult circuits to maintain due to the strong selection pressure they impart, leading to high potential for evolution of escape mutant populations. Here we engineer two CRISPR-based kill switches in the probiotic Escherichia coli Nissle 1917, a single-input chemical-responsive switch and a 2-input chemical- and temperature-responsive switch. We employ parallel strategies to address kill switch stability, including functional redundancy within the circuit, modulation of the SOS response, antibiotic-independent plasmid maintenance, and provision of intra-niche competition by a closely related strain. We demonstrate that strains harboring either kill switch can be selectively and efficiently killed inside the murine gut, while strains harboring the 2-input switch are additionally killed upon excretion. Leveraging redundant strategies, we demonstrate robust biocontainment of our kill switch strains and provide a template for future kill switch development.

Epigenetics and Helicobacter pylori.

Epigenetics regulates gene expression, cell type development during differentiation, and the cell response to environmental stimuli. To survive, bacteria need to evade the host immune response. Bacteria, including Helicobacter pylori (Hp), reach this target epigenetically, altering the chromatin of the host cells, in addition to several more approaches, such as DNA mutation and recombination. This review shows that Hp prevalently silences the genes of the human gastric mucosa by DNA methylation. Epigenetics includes different mechanisms. However, DNA methylation persists after DNA replication and therefore is frequently associated with the inheritance of repressed genes. Chromatin modification can be transmitted to daughter cells leading to heritable changes in gene expression. Aberrant epigenetic alteration of the gastric mucosa DNA remains the principal cause of gastric cancer. Numerous methylated genes have been found in cancer as well as in precancerous lesions of Hp-infected patients. These methylated genes inactivate tumor-suppressor genes. It is time for us to complain about our genetic and epigenetic makeups for our diseases.

Division of labor between SOS and PafBC in mycobacterial DNA repair and mutagenesis.

DNA repair systems allow microbes to survive in diverse environments that compromise chromosomal integrity. Pathogens such as Mycobacterium tuberculosis must contend with the genotoxic host environment, which generates the mutations that underlie antibiotic resistance. Mycobacteria encode the widely distributed SOS pathway, governed by the LexA repressor, but also encode PafBC, a positive regulator of the transcriptional DNA damage response (DDR). Although the transcriptional outputs of these systems have been characterized, their full functional division of labor in survival and mutagenesis is unknown. Here, we specifically ablate the PafBC or SOS pathways, alone and in combination, and test their relative contributions to repair. We find that SOS and PafBC have both distinct and overlapping roles that depend on the type of DNA damage. Most notably, we find that quinolone antibiotics and replication fork perturbation are inducers of the PafBC pathway, and that chromosomal mutagenesis is codependent on PafBC and SOS, through shared regulation of the DnaE2/ImuA/B mutasome. These studies define the complex transcriptional regulatory network of the DDR in mycobacteria and provide new insight into the regulatory mechanisms controlling the genesis of antibiotic resistance in M. tuberculosis.

Long-chain unsaturated fatty acids are involved in the viability and itraconazole susceptibility of Aspergillus fumigatus.

The prevalence of invasive aspergillosis with azole resistance is increasing, but the mechanisms underlying the development of resistance and treatment strategies are still limited. The present work is focused on finding a relationship between long-chain unsaturated fatty acids (LCUFAs), Aspergillus fumigatus development, and antifungal resistance. The effects of LCUFAs on antifungal agents in vitro were determined, and the stearic acid desaturase gene (sdeA) of A. fumigatus was characterized. In in vitro antifungal tests, LCUFAs antagonized the antifungal activity of itraconazole by extracting it from media, thereby preventing it from entering cells. The OA auxotrophic phenotype caused by an sdeA deletion confirmed that SdeA was required for OA biosynthesis in A. fumigatus. Furthermore, several low-level sdeA-overexpressing mutants with impaired vegetative growth phenotypes were successfully constructed. Additionally, an sdeA-overexpressing mutant, OEsdeA-5, showed lowered sensitivity levels to itraconazole. Moreover, RNA sequencing of OEsdeA-5 revealed that the altered gene-expression pattern. Through targeted metabolomics, decreased palmitic acid and stearic acid contents, accompanied by higher palmitoleic acid, margaroleic acid, and OA production levels, were found in OEsdeA-5. This study provides a novel insight of understanding of azole resistance and a potential target for drug development.

Inactivation of the riboswitch-controlled GMP synthase GuaA in Clostridioides difficile is associated with severe growth defects and poor infectivity in a mouse model of infection.

Clostridioides difficile is the main cause of nosocomial antibiotic-associated diarrhoea. There is a need for new antimicrobials to tackle this pathogen. Guanine riboswitches have been proposed as promising new antimicrobial targets, but experimental evidence of their importance in C. difficile is missing. The genome of C. difficile encodes four distinct guanine riboswitches, each controlling a single gene involved in purine metabolism and transport. One of them controls the expression of guaA, encoding a guanosine monophosphate (GMP) synthase. Here, using in-line probing and GusA reporter assays, we show that these riboswitches are functional in C. difficile and cause premature transcription termination upon binding of guanine. All riboswitches exhibit a high affinity for guanine characterized by Kd values in the low nanomolar range. Xanthine and guanosine also bind the guanine riboswitches, although with less affinity. Inactivating the GMP synthase (guaA) in C. difficile strain 630 led to cell death in minimal growth conditions, but not in rich medium. Importantly, the capacity of a guaA mutant to colonize the mouse gut was significantly reduced. Together, these results demonstrate the importance of de novo GMP biosynthesis in C. difficile during infection, suggesting that targeting guanine riboswitches with analogues could be a viable therapeutic strategy.

The extracellular vesicle generation paradox: a bacterial point of view.

All bacteria produce secreted vesicles that carry out a variety of important biological functions. These extracellular vesicles can improve adaptation and survival by relieving bacterial stress and eliminating toxic compounds, as well as by facilitating membrane remodeling and ameliorating inhospitable environments. However, vesicle production comes with a price. It is energetically costly and, in the case of colonizing pathogens, it elicits host immune responses, which reduce bacterial viability. This raises an interesting paradox regarding why bacteria produce vesicles and begs the question as to whether the benefits of producing vesicles outweigh their costs. In this review, we discuss the various advantages and disadvantages associated with Gram-negative and Gram-positive bacterial vesicle production and offer perspective on the ultimate score. We also highlight questions needed to advance the field in determining the role for vesicles in bacterial survival, interkingdom communication, and virulence.

Mycobacterium tuberculosis Rv0927c Inhibits NF-κB Pathway by Downregulating the Phosphorylation Level of IκBα and Enhances Mycobacterial Survival.

Through long-term coevolution with its host, Mycobacterium tuberculosis (M. tuberculosis) uses multiple strategies to escape host defenses. The M. tuberculosis Rv0927c protein is predicted to be a short-chain dehydrogenase/reductase related to bacterial metabolism. However, the role of Rv0927c during M. tuberculosis infection remains unclear. Here, we observed that Rv0927c inhibited the expression of IL-6, TNF-α, and IL-1ß, an effect dependent on NF-κB and p38 pathways. Western blot analysis of macrophages infected with recombinant Mycobacterium smegmatis strains showed that Rv0927c attenuated NF-κB activation by downregulating the phosphorylation of IκBα. Additionally, Rv0927c enhanced intracellular survival of M. smegmatis and pathological effects in mice. In conclusion, our findings demonstrate that Rv0927c functions as a regulator of inflammatory genes and enhances the survival of M. smegmatis.

Bacterial resistance to CRISPR-Cas antimicrobials.

In the age of antibiotic resistance and precise microbiome engineering, CRISPR-Cas antimicrobials promise to have a substantial impact on the way we treat diseases in the future. However, the efficacy of these antimicrobials and their mechanisms of resistance remain to be elucidated. We systematically investigated how a target E. coli strain can escape killing by episomally-encoded CRISPR-Cas9 antimicrobials. Using Cas9 from Streptococcus pyogenes (SpCas9) we studied the killing efficiency and resistance mutation rate towards CRISPR-Cas9 antimicrobials and elucidated the underlying genetic alterations. We find that killing efficiency is not correlated with the number of cutting sites or the type of target. While the number of targets did not significantly affect efficiency of killing, it did reduce the emergence of chromosomal mutations conferring resistance. The most frequent target of resistance mutations was the plasmid-encoded SpCas9 that was inactivated by bacterial genome rearrangements involving translocation of mobile genetic elements such as insertion elements. This resistance mechanism can be overcome by re-introduction of an intact copy of SpCas9. The work presented here provides a guide to design strategies that reduce resistance and improve the activity of CRISPR-Cas antimicrobials.

Role of a cryptic tRNA gene operon in survival under translational stress.

As compared to eukaryotes, bacteria have a reduced tRNA gene set encoding between 30 and 220 tRNAs. Although in most bacterial phyla tRNA genes are dispersed in the genome, many species from distinct phyla also show genes forming arrays. Here, we show that two types of arrays with distinct evolutionary origins exist. This work focuses on long tRNA gene arrays (L-arrays) that encompass up to 43 genes, which disseminate by horizontal gene transfer and contribute supernumerary tRNA genes to the host. Although in the few cases previously studied these arrays were reported to be poorly transcribed, here we show that the L-array of the model cyanobacterium Anabaena sp. PCC 7120, encoding 23 functional tRNAs, is largely induced upon impairment of the translation machinery. The cellular response to this challenge involves a global reprogramming of the transcriptome in two phases. tRNAs encoded in the array are induced in the second phase of the response, directly contributing to cell survival. Results presented here show that in some bacteria the tRNA gene set may be partitioned between a housekeeping subset, which constantly sustains translation, and an inducible subset that is generally silent but can provide functionality under particular conditions.

A method for the enrichment, isolation and validation of Mycobacterium smegmatis population surviving in the presence of bactericidal concentrations of rifampicin and moxifloxacin.

The bacterial populations surviving in the presence of antibiotics contain cells that have gained genetic resistance, phenotypic resistance and tolerance to antibiotics. Isolation of live bacterial population, surviving against antibiotics, from the milieu of high proportions of dead/damaged cells will facilitate the study of the cellular/molecular processes used by them for survival. Here we present a Percoll gradient centrifugation based method for the isolation of enriched population of Mycobacterium smegmatis surviving in the presence of bactericidal concentrations of rifampicin and moxifloxacin. From the time of harvest, throughout the enrichment and isolation processes, and up to the lysis of the cells for total RNA preparation, we maintained the cells in the presence of the antibiotic to avoid changes in their metabolic status. The total RNA extracted from the enriched population of live antibiotic-surviving population showed structural integrity and purity. We analysed the transcriptome profile of the antibiotic-surviving population and compared it with the orthologue genes of Mycobacterium tuberculosis that conferred antibiotic tolerance on tubercle bacilli isolated from the tuberculosis patients under treatment with four antitubercular antibiotics. Statistically significant comparability between the gene expression profiles of the antibiotic tolerance associated genes of M. smegmatis and M. tuberculosis validated the reliability/utility of the method.

Role of the Staphylococcus aureus Extracellular Loop of GraS in Resistance to Distinct Human Defense Peptides in PMN and Invasive Cardiovascular infections.

GraS is a membrane sensor in Staphylococcus aureus that induces mprF and dltABCD expression to alter the surface positive charge upon exposure to cationic human defense peptides (HDPs). The sensing domain of GraS likely resides in the 9-residue extracellular loop (EL). In this study, we assessed a hospital-acquired methicillin-resistant S. aureus (HA-MRSA) strain (COL) for the specific role of two distinct EL mutations: F38G (bulk) and D/35/37/41K (charged inversion). Activation of mprF by polymyxin B (PMB) was reduced in the D35/37/41K mutant versus the D35/37/41G mutant, correlating with reduced surface positive charge; in contrast, these effects were less prominent in the F38G mutant but still lower than those in the parent. These data indicated that both electrostatic charge and steric bulk of the EL of GraS influence induction of genes impacting HDP resistance. Using mprF expression as a readout, we confirmed GraS signaling was pH dependent, increasing as pH was lowered (from pH 7.5 down to pH 5.5). In contrast to PMB activation, reduction of mprF was comparable at pH 5.5 between the P38G and D35/37/41K point mutants, indicating a mechanistic divergence between GraS activation by acidic pH versus cationic peptides. Survival assays in human blood and purified polymorphonuclear leukocytes (PMNs) revealed lower survival of the D35/37/41K mutant versus the F38G mutant, with both being lower than that of the parent. Virulence studies in the rabbit endocarditis model mirrored whole blood and PMN killing assay data described above. Collectively, these data confirmed the importance of specific residues within the EL of GraS in conferring essential bacterial responses for MRSA survival in infections.

A fructose/H+ symporter controlled by a LacI-type regulator promotes survival of pandemic Vibrio cholerae in seawater.

The bacterium Vibrio cholerae can colonize the human intestine and cause cholera, but spends much of its life cycle in seawater. The pathogen must adapt to substantial environmental changes when moving between seawater and the human intestine, including different availability of carbon sources such as fructose. Here, we use in vitro experiments as well as mouse intestinal colonization assays to study the mechanisms used by pandemic V. cholerae to adapt to these environmental changes. We show that a LacI-type regulator (FruI) and a fructose/H+ symporter (FruT) are important for fructose uptake at low fructose concentrations, as those found in seawater. FruT is downregulated by FruI, which is upregulated when O2 concentrations are low (as in the intestine) by ArcAB, a two-component system known to respond to changes in oxygen levels. As a result, the bacteria predominantly use FruT for fructose uptake under seawater conditions (low fructose, high O2), and use a known fructose phosphotransferase system (PTS, Fpr) for fructose uptake under conditions found in the intestine. PTS activity leads to reduced levels of intracellular cAMP, which in turn upregulate virulence genes. Our results indicate that the FruT/FruI system may be important for survival of pandemic V. cholerae in seawater.

Survival of the blaNDM-harbouring Escherichia coli in tropical seawater and conjugative transfer of resistance markers.

Anthropogenic contamination of coastal-marine water is responsible for introducing multidrug-resistant bacteria such as the pNDM-harbouring Escherichia coli into the seafood chain. This study was conducted to understand the survivability of a multidrug-resistant, the New Delhi Metallo-ß-lactamase-producing E. coli (AS-EC121) in tropical seawater at room temperature (28-32 °C) compared to E. coli K12 strain. The experimental and control strains were inoculated at 6 log CFU/ml level into seawater. After an initial sharp decline in counts, AS-EC121 and K12 strains showed a gradual loss of viability after week-1 of inoculation. AS-EC121 was undetectable after day-56, while K12 colonies disappeared a week later, from day-63. The conjugation experiment revealed that pNDM was transferable to a recipient E. coli strain in seawater. This study suggests that the multidrug-resistant, pNDM-harbouring E. coli is able to survive in seawater for over 2 months stably maintaining the resistance plasmid. The resistance genotypes do not seem to compromise the survivability of MDR E. coli and the stability of plasmid provides ample opportunities for dissemination of plasmids among co-inhabiting bacteria in the coastal-marine environments.

Bacteria autoaggregation: how and why bacteria stick together.

Autoaggregation, adherence between identical bacterial cells, is important for colonization, kin and kind recognition, and survival of bacteria. It is directly mediated by specific interactions between proteins or organelles on the surfaces of interacting cells or indirectly by the presence of secreted macromolecules such as eDNA and exopolysaccharides. Some autoaggregation effectors are self-associating and present interesting paradigms for protein interaction. Autoaggregation can be beneficial or deleterious at specific times and niches. It is, therefore, typically regulated through transcriptional or post-transcriptional mechanisms or epigenetically by phase variation. Autoaggregation can contribute to bacterial adherence, biofilm formation or other higher-level functions. However, autoaggregation is only required for these phenotypes in some bacteria. Thus, autoaggregation should be detected, studied and measured independently using both qualitative and quantitative in vitro and ex vivo methods. If better understood, autoaggregation holds the potential for the discovery of new therapeutic targets that could be cost-effectively exploited.

Mycobacterium tuberculosis Rv2145c Promotes Intracellular Survival by STAT3 and IL-10 Receptor Signaling.

Although Mycobacterium tuberculosis (Mtb) is an intracellular pathogen in phagocytic cells, the factors and mechanisms by which they invade and persist in host cells are still not well understood. Characterization of the bacterial proteins modulating macrophage function is essential for understanding tuberculosis pathogenesis and bacterial virulence. Here we investigated the pathogenic role of the Rv2145c protein in stimulating IL-10 production. We first found that recombinant Rv2145c stimulated bone marrow-derived macrophages (BMDMs) to secrete IL-10, IL-6 and TNF-α but not IL-12p70 and to increase the expression of surface molecules through the MAPK, NF-κB, and TLR4 pathways and enhanced STAT3 activation and the expression of IL-10 receptor in Mtb-infected BMDMs. Rv2145c significantly enhanced intracellular Mtb growth in BMDMs compared with that in untreated cells, which was abrogated by STAT3 inhibition and IL-10 receptor (IL-10R) blockade. Expression of Rv2145c in Mycobacterium smegmatis (M. smegmatis) led to STAT3-dependent IL-10 production and enhancement of intracellular growth in BMDMs. Furthermore, the clearance of Rv2145c-expressing M. smegmatis in the lungs and spleens of mice was delayed, and these effects were abrogated by administration of anti-IL-10R antibodies. Finally, all mice infected with Rv2145c-expressing M. smegmatis died, but those infected with the vector control strain did not. Our data suggest that Rv2145c plays a role in creating a favorable environment for bacterial survival by modulating host signals.

Yeast Fin1-PP1 dephosphorylates an Ipl1 substrate, Ndc80, to remove Bub1-Bub3 checkpoint proteins from the kinetochore during anaphase.

The spindle assembly checkpoint (SAC) prevents anaphase onset in response to chromosome attachment defects, and SAC silencing is essential for anaphase onset. Following anaphase onset, activated Cdc14 phosphatase dephosphorylates the substrates of cyclin-dependent kinase to facilitate anaphase progression and mitotic exit. In budding yeast, Cdc14 dephosphorylates Fin1, a regulatory subunit of protein phosphatase 1 (PP1), to enable kinetochore localization of Fin1-PP1. We previously showed that kinetochore-localized Fin1-PP1 promotes the removal of the SAC protein Bub1 from the kinetochore during anaphase. We report here that Fin1-PP1 also promotes kinetochore removal of Bub3, the Bub1 partner, but has no effect on another SAC protein Mad1. Moreover, the kinetochore localization of Bub1-Bub3 during anaphase requires Aurora B/Ipl1 kinase activity. We further showed that Fin1-PP1 facilitates the dephosphorylation of kinetochore protein Ndc80, a known Ipl1 substrate. This dephosphorylation reduces kinetochore association of Bub1-Bub3 during anaphase. In addition, we found that untimely Ndc80 dephosphorylation causes viability loss in response to tensionless chromosome attachments. These results suggest that timely localization of Fin1-PP1 to the kinetochore controls the functional window of SAC and is therefore critical for faithful chromosome segregation.

Dual transcriptional analysis reveals adaptation of host and pathogen to intracellular survival of Pseudomonas aeruginosa associated with urinary tract infection.

Long-term survival of bacterial pathogens during persistent bacterial infections can be associated with antibiotic treatment failure and poses a serious public health problem. Infections caused by the Gram-negative pathogen Pseudomonas aeruginosa, which can cause both acute and chronic infections, are particularly challenging due to its high intrinsic resistance to antibiotics. The ineffectiveness of antibiotics is exacerbated when bacteria reside intracellularly within host cells where they can adopt a drug tolerant state. While the early steps of adherence and entry of P. aeruginosa into mammalian cells have been described, the subsequent fate of internalized bacteria, as well as host and bacterial molecular pathways facilitating bacterial long-term survival, are not well defined. In particular, long-term survival within bladder epithelial cells has not been demonstrated and this may have important implications for the understanding and treatment of UTIs caused by P. aeruginosa. Here, we demonstrate and characterize the intracellular survival of wild type (WT) P. aeruginosa inside bladder epithelial cells and a mutant with a disruption in the bacterial two-component regulator AlgR that is unable to survive intracellularly. Using simultaneous dual RNA-seq transcriptional profiling, we define the transcriptional response of intracellular bacteria and their corresponding invaded host cells. The bacterial transcriptional response demonstrates that WT bacteria rapidly adapt to the stress encountered in the intracellular environment in contrast to ΔalgR bacteria. Analysis of the host transcriptional response to invasion suggests that the NF-κB signaling pathway, previously shown to be required for extracellular bacterial clearance, is paradoxically also required for intracellular bacterial survival. Lastly, we demonstrate that intracellular survival is important for pathogenesis of P. aeruginosa in vivo using a model of murine urinary tract infection. We propose that the unappreciated ability of P. aeruginosa to survive intracellularly may play an important role in contributing to the chronicity and recurrence of P. aeruginosa in urinary tract infections.

Genomic insights into the molecular mechanisms of a Pseudomonas strain significant in its survival in Kongsfjorden, an Arctic fjord.

Whole-genome sequence of Pseudomonas sp. Kongs-67 retrieved from Kongsfjorden, an Arctic fjord, has been investigated to understand the molecular machinery required for microbial association and survival in a polar fjord. The genome size of Kongs-67 was 4.5 Mb and was found to be closely related to the Antarctic P. pelagia strain CL-AP6. This genome encodes for chemotaxis response regulator proteins (CheABB1RR2VWYZ), chemoreceptors (methyl-accepting chemotaxis proteins), and flagellar system proteins (FliCDEFGOPMN, FlhABF, FlgBCDEFGHIJKL, and MotAB proteins) vital in cellular interactions in the dynamic fjord environment. A high proportion of genes were assigned to biofilm formation (pgaABCD operon) and signal transduction protein categories (EnvZ/OmpR, CpxA/CpxR, PhoR/PhoB, PhoQ) indicating that the biofilm formation in Kongs-67 could be tightly regulated in response to the availability of signalling-metabolites. The genome of Kongs-67 encoded for HemBCD, CbiA, CobABNSTOQCDP, and BtuBFR proteins involved in cobalamin biosynthesis and transport along with proteins for siderophore-mediated iron channelling (PchR, Fur protein, FpvA); crucial in a microbial association. The genomes of Arctic strain Kongs-67 and Antarctic strain CL-AP6 were similar which is indicative of retainment of the core genes in the polar Pseudomonas strains that could be vital in conferring evolutionary adaptation for its survival in a polar fjord. Thus, our study contributes to the knowledge on the genetics of a polar Pseudomonas member exhibiting biosynthetic potentials and suggest Pseudomonas sp. Kongs-67 as a suitable candidate for the investigation of functional aspects of molecular adaptations in the polar marine environment.