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bioRxiv2020

FK506-binding protein FklB is involved in biofilm formation through its peptidyl-prolyl isomerase activity

FklB is a member of the FK506-binding proteins (FKBPs), a family that consists of five genes in Escherichia coli. Little is known about the physiological and functional role of FklB in bacterial movement. In the present study, FklB knock-out mutant {Delta}fklB presented an increased swarming and swimming motility and biofilm formation phenotype, suggesting that FklB is a negative regulator of these cellular processes. Complementation with Peptidyl-prolyl isomerase (PPIase)-deficient fklB gene (Y181A) revealed that the defects in biofilm formation were not restored by Y181A, indicating that PPIase activity of FklB is modulating biofilm formation in E. coli. The mean cell length of {Delta}fklB swarming cells was significantly smaller as compared to the wild-type BW25113. Furthermore, the mean cell length of swarming and swimming wild-type and {Delta}fklB cells overexpressing fklB or Y181A was considerably larger, suggesting that PPIase activity of FklB plays a role in cell elongation and/or cell division. A multi-copy suppression assay demonstrated that defects in motility and biofilm phenotype were compensated by overexpressing sets of PPIase-encoding genes. Taken together, our data represent the first report demonstrating the involvement of FklB in cellular functions of E. coli.

microbiology↗

bioRxiv2020

Thermogenic hydrocarbons sustain diverse subseafloor microbial communities in deep sea cold seep sediments

At marine cold seeps, gaseous and liquid hydrocarbons migrate from deep subsurface origins to the sediment-water interface. Cold seep sediments are known to host taxonomically diverse microorganisms, but little is known about their metabolic potential and depth distribution in relation to hydrocarbon and electron acceptor availability. In this work, we combined geochemical, metagenomic and metabolomic measurements in distinct sediment redox regimes to profile microbial activities within the uppermost 350 cm of a newly discovered cold seep in the NW Atlantic deep sea (2.3 km water depth). Depth-resolved metagenomic profiling revealed compositional and functional differentiation between near-surface sediments (dominated by Proteobacteria) and deeper subsurface layers (dominated by Atribacteria, Chloroflexi, Euryarchaeota and Lokiarchaeota). Metabolic capabilities of community members were inferred from 376 metagenome-assembled genomes spanning 46 phyla (including five novel candidate phyla). In deeper sulfate-reducing and methanogenic sediments, various community members are capable of anaerobically oxidizing short-chain alkanes (alkyl-CoM reductase pathway), longer-chain alkanes (fumarate addition pathway), and aromatic hydrocarbons (fumarate addition and subsequent benzoyl-CoA pathways). Geochemical profiling demonstrated that hydrocarbon substrates are abundant in this location, thermogenic in origin, and subject to biodegradation. The detection of alkyl-/arylalkylsuccinate metabolites, together with carbon isotopic signatures of ethane, propane and carbon dioxide, support that microorganisms are actively degrading hydrocarbons in these sediments. Hydrocarbon oxidation pathways operate alongside other deep seabed metabolisms such as sulfide oxidation, hydrogen oxidation, carbon fixation, fermentation and reductive dehalogenation. Upward migrated thermogenic hydrocarbons thus sustain diverse microbial communities with activities that affect subseafloor biogeochemical processes across the redox spectrum in deep sea cold seeps.

microbiology↗

bioRxiv2020

A screen for Plasmodium falciparum sporozoite surface protein binding to human hepatocyte surface receptors identifies novel host-pathogen interactions

Sporozoite invasion of hepatocytes is a necessary step prior to development of malaria, with similarities, at the cellular level, to merozoite invasion of erythrocytes. In the case of the malaria blood-stage, efforts to identify host-pathogen protein-protein interactions have yielded important insights including vaccine candidates. In the case of sporozoite-hepatocyte invasion, the host-pathogen protein-protein interactions involved are poorly understood. Here, we performed a systematic screen to identify such interactions. We substantially extended previous Plasmodium falciparum and human surface protein ectodomain libraries, creating new libraries containing 88 P. falciparum sporozoite protein coding sequences and 182 sequences encoding human hepatocyte surface proteins. Having expressed recombinant proteins from these sequences, we used a plate-based assay capable of detecting low affinity interactions between recombinant proteins, modified for enhanced throughput, to screen the proteins for interactions. We were able to test 7540 sporozoite-hepatocyte protein pairs under conditions likely to be sensitive for interaction. We report and characterise an interaction between human fibroblast growth factor receptor 4 (FGFR4) and the P. falciparum protein Pf34, and describe an additional interaction between human low-density lipoprotein receptor (LDLR) and the P. falciparum protein PIESP15. Strategies to inhibit these interactions may have value in malaria prevention, and the modified interaction screening assay and protein expression libraries we report may be of wider value to the community.

microbiology↗

bioRxiv2020

Ancient MAPK ERK7 is regulated by an unusual inhibitory scaffold required for Toxoplasma apical complex biogenesis

Apicomplexan parasites use a specialized cilium structure called the apical complex to organize their secretory organelles and invasion machinery. The apical complex is integrally associated with both the parasite plasma membrane and an intermediate filament cytoskeleton called the inner membrane complex (IMC). While the apical complex is essential to the parasitic lifestyle, little is known about the regulation of apical complex biogenesis. Here, we identify AC9 (apical cap protein 9), a largely intrinsically disordered component of the Toxoplasma gondii IMC, as essential for apical complex development, and therefore for host cell invasion and egress. Parasites lacking AC9 fail to successfully assemble the tubulin-rich core of their apical complex, called the conoid. We use proximity biotinylation to identify the AC9 interaction network, which includes the kinase ERK7. Like AC9, ERK7 is required for apical complex biogenesis. We demonstrate that AC9 directly binds ERK7 through a conserved C-terminal motif and that this interaction is essential for ERK7 localization and function at the apical cap. The crystal structure of the ERK7:AC9 complex reveals that AC9 is not only a scaffold, but also inhibits ERK7 through an unusual set of contacts that displaces nucleotide from the kinase active site. ERK7 is an ancient and auto-activating member of the mitogen-activated kinase family and we have identified its first regulator in any organism. We propose that AC9 dually regulates ERK7 by scaffolding and concentrating it at its site of action while maintaining it in an "off" state until the specific binding of a true substrate. Significance StatementApicomplexan parasites include the organisms that cause widespread and devastating human diseases such as malaria, cryptosporidiosis, and toxoplasmosis. These parasites are named for a structure, called the "apical complex," that organizes their invasion and secretory machinery. We found that two proteins, apical cap protein 9 (AC9) and an enzyme called ERK7 work together to facilitate apical complex assembly. Intriguingly, ERK7 is an ancient molecule that is found throughout Eukaryota, though its regulation and function are poorly understood. AC9 is a scaffold that concentrates ERK7 at the base of the developing apical complex. In addition, AC9 binding likely confers substrate selectivity upon ERK7. This simple competitive regulatory model may be a powerful but largely overlooked mechanism throughout biology.

microbiology↗

bioRxiv2020

Dynamics of mcr-1 prevalence and mcr-1-positive Escherichia coli after the cessation of colistin use as a feed additive for animals in China: a prospective cross-sectional and whole genome sequencing based molecular epidemiological study

BackgroundThe global dissemination of colistin resistance encoded by mcr-1 has been attributed to extensive use of colistin in livestock, threatening colistin efficacy in medicine. The emergence of mcr-1 in common pathogens, such as Escherichia coli, is of particular concern. Therefore, China banned the use of colistin in animal feed from May 1ST 2017. We investigated subsequent changes in mcr-1 prevalence, and the genomic epidemiology of mcr-1-positive Escherichia coli (MCRPEC). MethodsSampling was conducted pre- (October-December 2016) and post-colistin ban (October-December, 2017 and 2018, respectively). 3675 non-duplicate pig fecal samples were collected from 14 provinces (66 farms) in China to determine intervention-related changes in mcr-1 prevalence. 15193 samples were collected from pigs, healthy human volunteers, colonized and infected hospital inpatients, food and the environment in Guangzhou, to characterize source-specific mcr-1 prevalence and the wider ecological impact of the ban. From these samples, 688 MCRPEC were analyzed with whole genome sequencing (WGS), plasmid conjugation and S1-PFGE/Southern blots to characterize associated genomic changes. FindingsAfter the ban, mcr-1 prevalence decreased significantly in national pig farms, from 45{middle dot}0% (308/684 samples) in 2016, to 19{middle dot}4% (274/1416) in 2018 (p<0{middle dot}0001). This trend was mirrored in samples from most sources in Guangzhou (overall 19{middle dot}2% [959/5003 samples] in 2016; 5{middle dot}3% [238/4489] in 2018; p<0{middle dot}0001). The population structure of MCRPEC was diverse (23 sequence clusters [SCs]); ST10 clonal complex isolates were predominant (247/688 [36%]). MCRPEC causing infection in hospitalized inpatients were genetically more distinct and appeared less affected by the ban. mcr-1 was predominantly found on plasmids (632/688 [92%]). Common mcr-1 plasmid types included IncX4, IncI2 and IncHI2 (502/656 [76.5%]); significant increases in IncI2-associated mcr-1 and a distinct lineage of mcr-1-associated IncHI2 were observed post-ban. Changes in the frequency of mcr-1-associated flanking sequences (ISApl1-negative MCRPEC), 63 core genome SNPs and 30 accessory genes were also significantly different after the ban, consistent with rapid genetic adaptation in response to changing selection pressures. InterpretationA rapid, ecosystem-wide, decline in mcr-1 was observed after banning the use of colistin in animal feed, with associated genetic changes in MCRPEC. Genomic surveillance is key to assessing and monitoring stewardship interventions. FundingNational Natural Science Foundation of China

microbiology↗

bioRxiv2020

Poecivirus is present in individuals with beak deformities in seven species of North American birds

Avian keratin disorder (AKD), a disease characterized by debilitating beak overgrowth but with unknown etiology, has increasingly affected wild bird populations since the 1990s. We previously showed that a novel picornavirus, Poecivirus, is closely correlated with disease status in Black-capped Chickadees (Poecile atricapillus) in Alaska. However, our knowledge of the relationship between Poecivirus and beak deformities in other species and other geographic areas remains limited. The growing geographic scope and number of species affected by AKD-like beak deformities require a better understanding of the causative agent to evaluate the population-level impacts of this epizootic. Here, we tested eight individuals from six avian species with AKD-consistent deformities for the presence of Poecivirus: Mew Gull (Larus canus), Hairy Woodpecker (Picoides villosus), Black-billed Magpie (Pica hudsonia), Northwestern Crow (Corvus caurinus), Red-breasted Nuthatch (Sitta canadensis), and Blackpoll Warbler (Setophaga striata); individuals were sampled in Alaska and Maine (1999-2016). We used targeted PCR followed by Sanger sequencing to test for the presence of Poecivirus in each specimen, and to obtain viral genome sequence from virus-positive host individuals. We detected Poecivirus in all individuals tested, but not in negative controls. Furthermore, we used unbiased metagenomic sequencing to test for the presence of other pathogens in six of these specimens (Hairy Woodpecker, two Northwestern Crows, two Red-breasted Nuthatches, Blackpoll Warbler). This analysis yielded additional viral sequence from several specimens, including the complete coding region of Poecivirus from one Red-breasted Nuthatch, which we confirmed via targeted PCR followed by Sanger sequencing. This study demonstrates that Poecivirus is present in individuals with AKD-consistent deformities from six avian species other than Black-capped Chickadee. While further investigation will be required to explore whether there exists a causal link between this virus and AKD, this study demonstrates that Poecivirus is not geographically restricted to Alaska, but rather occurs elsewhere in North America.

microbiology↗

bioRxiv2020

Human cytomegalovirus protein pUL36: a dual cell death pathway inhibitor

Human cytomegalovirus (HCMV) is an important human pathogen and a paradigm of intrinsic, innate and adaptive viral immune evasion. Here, we employed multiplexed tandem mass tag-based proteomics to characterise host proteins targeted for degradation late during HCMV infection. This approach revealed that mixed lineage kinase domain-like protein (MLKL), a key terminal mediator of cellular necroptosis, was rapidly and persistently degraded by the minimally passaged HCMV strain Merlin but not the extensively passaged strain AD169. The strain Merlin viral inhibitor of apoptosis pUL36 was necessary and sufficient both to degrade MLKL and to inhibit necroptosis. Furthermore, mutation of pUL36 Cys131 abrogated MLKL degradation and restored necroptosis. As the same residue is also required for pUL36-mediated inhibition of apoptosis by preventing proteolytic activation of pro-caspase 8, we define pUL36 as a multifunctional inhibitor of both apoptotic and necroptotic cell death. SIGNIFICANCE STATEMENTCell death is a key defence against viral infection, preventing spread from infected to uninfected cells. Correspondingly, certain viruses encode inhibitors of apoptotic and necroptotic cell death pathways in order to facilitate their persistence. Human cytomegalovirus (HCMV) is an important human pathogen that can block apoptosis, but hitherto it has been unclear whether or how the virus blocks necroptosis. Here, we used a proteomic screen to identify human proteins targeted for destruction by HCMV, finding that the key necroptosis mediator MLKL is degraded throughout infection. MLKL is targeted for degradation by HCMV protein pUL36, which is also instrumental in inhibiting apoptosis. Thus, pUL36 is a dual cell death pathway inhibitor, and may represent an important therapeutic target.

microbiology↗

bioRxiv2020

A lung-on-chip infection model reveals protective and permissive roles of alveolar epithelial cells in tuberculosis

Mycobacterium tuberculosis (Mtb) makes first contact with a host in the alveolar space, an interaction largely inaccessible to experimental observation. We establish a lung-on-chip model for early tuberculosis and use time-lapse imaging to reveal the dynamics of host-Mtb interactions at an air-liquid interface with a spatiotemporal resolution unattainable in animal models. By reconstituting host physiology in a modular manner, we probe the role of pulmonary surfactant secreted by alveolar epithelial cells (AECs) in early infection. This is difficult to study directly in animal models, as surfactant-deficient animals are either non-viable or develop acute lung pathologies. We demonstrate that surfactant deficiency results in rapid and uncontrolled Mtb growth in both macrophages and AECs. In contrast, under normal surfactant levels, a significant fraction of intracellular bacteria are non-growing. The surfactant-deficient phenotype is rescued by exogenous addition of surfactant replacement formulations, which have no effect on bacterial viability in the absence of host cells. Surfactant partially removes virulence-associated lipids and proteins 1,2 from the bacterial cell surface and consistent with this mechanism of action, we show that attenuation of bacteria lacking the virulence-associated ESX-1 secretion system is independent of surfactant levels. These findings may partly explain why individuals with compromised surfactant function, such as smokers and elderly persons, are at increased risk of developing active tuberculosis.

microbiology↗

bioRxiv2020

Microbial grazers can control chytridiomycosis caused by aquatic zoosporic fungi

Free-living eukaryotic microbes may reduce animal diseases. We evaluated the dynamics by which micrograzers (primarily protozoa) apply top-down control on the chytrid Batrachochytrium dendrobatidis (Bd) a devastating, panzootic pathogen of amphibians. Although micrograzers consumed zoospores ([~]3 {micro}m), the dispersal stage of chytrids, not all species grew monoxenically on zoospores. However, the ubiquitous ciliate Tetrahymena pyriformis, which likely co-occurs with Bd, grew at near its maximum rate (r = 1.7 d-1). A functional response (ingestion vs. prey abundance) for T. pyriformis, measured using spore-surrogates (microspheres) revealed maximum ingestion (Imax) of 1.63 x 103 zoospores d-1, with a half saturation constant (k) of 5.75 x 103 zoospores ml-1. Using these growth and grazing data we developed and assessed a population model that incorporated chytrid-host and micrograzer dynamics. Simulations using our data and realistic parameters obtained from the literature suggested that micrograzers could control Bd and potentially prevent chytridiomycosis (defined as 104 sporangia host-1). However, simulated inferior micrograzers (0.7 x Imax and 1.5 x k) did not prevent chytridiomycosis, although they ultimately reduced pathogen abundance to below levels resulting in disease. These findings indicate how micrograzer responses can be applied when modelling disease dynamics for Bd and other zoosporic fungi.

microbiology↗

bioRxiv2020

A single-cell RNAseq atlas of the pathogenic stage of Schistosoma mansoni identifies a key regulator of blood feeding

Schistosomiasis is an ancient and chronic neglected tropical disease that infects over 240 million people and kills over 200,000 of the worlds poorest people every year1, 2. There are no vaccines and because there is only one drug available, the need for new therapeutics is great. The causative agents of this disease are flatworm parasites that dwell inside the hosts circulation, often for decades, where they feed on blood and lay eggs which are primarily responsible for disease pathology. As metazoans comprised of multiple tissue types, understanding the schistosomes tissues on a molecular level and their functions during what can be decades of successful parasitism could suggest novel therapeutic strategies. Here, we employ single-cell RNAseq to characterize 43,642 cells from the pathogenic (adult) stage of the schistosome lifecycle. From these data, we characterize 68 molecularly distinct cell populations that comprise nearly all tissues described morphologically, including the nervous and reproductive systems. We further uncover a lineage of somatic stem cells responsible for producing and maintaining the parasites gut - the primary tissue responsible for digestion of host blood. Finally, we show that a homologue of hepatocyte nuclear factor 4 (hnf4) is expressed in this gut lineage and required for gut maintenance, blood feeding and inducing egg-associated pathology in vivo. Together, the data highlight the utility of this single-cell RNAseq atlas to understand schistosome biology and identify potential therapeutic interventions.

microbiology↗

bioRxiv2020

Survival strategies of Enterococcus mundtii in the gut of Spodoptera littoralis : a live report

The complex interaction between a higher organism and its resident gut flora is a subject of immense interest in the field of symbiosis. Many insects harbor a complex community of microorganisms in their gut. Larvae of Spodoptera littoralis, a lepidopteran pest which is prevalent in tropical and subtropical regions of the world, have a tube-like gut structure containing a simple bacterial community. This community varies both spatially (along the length of the gut) and temporally (during the life cycle of the insect). To monitor the dynamics and rapid adaptation of microbes to the gut conditions, a GFP-tagged reporter E. mundtii was constructed. After feeding to early instar S. littoralis larvae, the tagged-microbes recovered from the fore and hind guts by flow cytometry. The fluorescent reporter confirmed the persistence of E. mundtii in the gut. RNA-sequencing of the sorted bacteria highlighted various strategies that the symbiont employs to survive, including upregulated pathways for tolerating alkaline stress, forming biofilms and two-component signaling systems, resisting oxidative stress and quorum sensing. Although these symbionts depend on the host for amino acid and fatty acids, differential regulation among various metabolic pathways points to an enriched lysine synthesis pathway in the hindgut of the larvae.

microbiology↗

bioRxiv2020

EslB is required for cell wall Integrity and cell division in Listeria monocytogenes

Lysozyme is an important component of the innate immune system. It functions by hydrolysing the peptidoglycan (PG) layer of bacteria. The human pathogen Listeria monocytogenes is intrinsically lysozyme resistant. The peptidoglycan N-deacetylase PgdA and O-acetyltransferase OatA are two known factors contributing to its lysozyme resistance. Furthermore, it was shown that the absence of components of an ABC transporter, here referred to as EslABC, leads to reduced lysozyme resistance. How its activity is linked to lysozyme resistance is still unknown. To investigate this further, a strain with a deletion in eslB, coding for a membrane component of the ABC transporter, was constructed in L. monocytogenes strain 10403S. The eslB mutant showed a 40-fold reduction in the minimal inhibitory concentration to lysozyme. Analysis of the PG structure revealed that the eslB mutant produced PG with reduced levels of O-acetylation. Using growth and autolysis assays, we show that the absence of EslB manifests in a growth defect in media containing high concentrations of sugars and increased endogenous cell lysis. A thinner PG layer produced by the eslB mutant under these growth conditions might explain these phenotypes. Furthermore, the eslB mutant had a noticeable cell division defect and formed elongated cells. Microscopy analysis revealed that an early cell division protein still localized in the eslB mutant indicating that a downstream process is perturbed. Based on our results, we hypothesize that EslB affects the biosynthesis and modification of the cell wall in L. monocytogenes and is thus important for the maintenance of cell wall integrity. IMPORTANCEThe ABC transporter EslABC is associated with the intrinsic lysozyme resistance of Listeria monocytogenes. However, the exact role of the transporter in this process and in the physiology of L. monocytogenes is unknown. Using different assays to characterize an eslB deletion strain, we found that the absence of EslB not only affects lysozyme resistance, but also endogenous cell lysis, cell wall biosynthesis, cell division and the ability of the bacterium to grow in media containing high concentrations of sugars. Our results indicate that EslB is by a yet unknown mechanism an important determinant for cell wall integrity in L. monocytogenes.

microbiology↗

bioRxiv2020

Prevalence of Staphylococcal Superantigens and their association among bacteremic and Infective Endocarditis patients in Egypt

AimInfective endocarditis (IE) is a major complication of Staphylococcus (S.) aureus infection in humans particularly those with bacteremia. Although Staphylococcus species are commensal on or in different parts of the human body, it is also known to be a serious pathogen causing bacteremia and sepsis that could lead to IE. Therefore, our aim was to assess the prevalence as well as phenotypic and genotypic association of the Staphylococcal superantigens (SAgs) among bacteremic and IE patients. MethodsThis study was conducted on Staphylococcus isolates recovered from bacteremic and IE patients. The isolates were screened phenotypically for the detection of SAgs including Staphylococcal enterotoxins (SEs) and toxic shock syndrome toxin-1 (TSST-1). Molecular detection and analysis of sea, seb, sec, sed, see and tsst-1, the major SAgs coding genes were performed using PCR and agarose gel electrophoresis, respectively. The obtained findings were statistically analyzed using standard methods. ResultsDetection of SAgs using ELISA revealed that 12 (46%) isolates were positive for enterotoxin production. However, the PCR revealed that 19 (73%) isolates were positive for enterotoxin genes with the highest prevalence of the sea gene (79%), followed by the seb (63%), tsst-1 (21%). The least frequent gene was the sed (5.3%). Accordingly, phenotypic and genotypic screening for prevalence of SAgs among Staphylococcal isolates showed significant difference (P value =0.046703), however, no significant correlation could be observed among the coagulase negative Staphylococci (CoNS) isolates (P value =0.248213). Statistical correlations between bacteremic and IE isolates with respect to prevalence of SAgs, showed no significant difference (P-value = 0.139, Effect size = 0.572) indicating no specific association between any of the detected SAgs and IE. Conclusionno significant difference has been found between Staphylococcal IE and bacteremia isolates regarding both phenotypic and genotypic detection of the most commonly SAgs. Therefore, all Staphylococcal bacteremic patients are suspected for IE. Also, future work should be conducted for analysis of SAgs gene expression.

microbiology↗

bioRxiv2020

Beef cattle that respond differently to fescue toxicosis have distinct gastrointestinal tract microbiota

Tall fescue (Lolium arundinaceum) is a widely used forage grass which shares a symbiosis with the endophytic fungus Epichloe coenophiala. The endophyte produces an alkaloid toxin that provides herbivory, heat and drought resistance to the grass, but can cause fescue toxicosis in grazing livestock. Fescue toxicosis can lead to reduced weight gain and milk yields resulting in significant losses to the livestock industry. In this trial, 149 Angus cows across two farms were continuously exposed to toxic, endophyte-infected, fescue for a total of 13 weeks. Of those 149 cows, 40 were classified into either high (HT) or low (LT) tolerance groups according to their growth performance (weight gain). 20 HT and 20 LT cattle balanced by farm were selected for 16S rRNA gene and ITS1 region Illumina MiSeq amplicon sequencing to compare the fecal microbiota of the two tolerance groups. This study reveals significantly (P<0.05) different bacterial and fungal microbiota between HT and LT cattle, and indicates that fungal phylotypes may be important for an animals response to fescue toxicosis: We found that fungal phylotypes affiliating to the Neocallimastigaceae, which are known to be important fiber-degrading fungi, were consistently more abundant in the HT cattle. Whereas fungal phylotypes related to the genus Thelebolus were more abundant in the LT cattle. This study also found more pronounced shifts in the microbiota in animals receiving higher amounts of the toxin. We identified fungal phylotypes which were consistently more abundant either in HT or LT cattle and may thus be associated with the respective animals response to fescue toxicosis. Our results thus suggest that some fungal phylotypes might be involved in mitigating fescue toxicosis. The results of our study provide a basis for investigating the functional role of the gastrointestinal microbiota in fescue toxicosis in more detail in the future.

microbiology↗

bioRxiv2020

Genome-resolved metagenomics and detailed geochemical speciation analyses yield new insights into microbial mercury cycling in geothermal springs

Geothermal systems emit substantial amounts of aqueous, gaseous and methylated mercury, but little is known about microbial influences on mercury speciation. Here we report results from genome-resolved metagenomics and mercury speciation analysis of acid warm springs in the Ngawha Geothermal Field (<55 {degrees}C, pH < 4.5), Northland Region, Aotearoa (New Zealand). Our aim was to identify the microorganisms genetically equipped for mercury methylation, demethylation, or Hg(II) reduction to volatile Hg(0) in these springs. Dissolved total and methylated mercury concentrations in two adjacent springs with different mercury speciation ranked among the highest reported from natural sources (250-16000 ng L-1 and 0.5-13.9 ng L-1, respectively). Total solid mercury concentrations in spring sediments ranged from 1273 to 7000 {micro}g g-1. In the context of such ultra-high mercury levels, the geothermal microbiome was unexpectedly diverse, and dominated by acidophilic and mesophilic sulfur- and iron-cycling bacteria, mercury- and arsenic-resistant bacteria, and thermophilic and acidophilic archaea. Integrating microbiome structure and metagenomic potential with geochemical constraints, we constructed a conceptual model for biogeochemical mercury cycling in geothermal springs. The model includes abiotic and biotic controls on mercury speciation, and illustrates how geothermal mercury cycling may couple to microbial community dynamics and sulfur and iron biogeochemistry. IMPORTANCELittle is currently known about biogeochemical mercury cycling in geothermal systems. This manuscript presents an important new conceptual model, supported by genome-resolved metagenomic analysis and detailed geochemical measurements. This work provides a framework for studying natural geothermal mercury emissions globally. Specifically, our findings have implications for mercury speciation in wastewaters from geothermal power plants and the potential environmental impacts of microbially and abiotically formed mercury species, particularly where mobilized in spring waters that mix with surface- or ground-waters. Furthermore, in the context of thermophilic origins for microbial mercury volatilisation, this report yields new insights into how such processes may have evolved alongside microbial mercury methylation/demethylation, and the environmental constraints imposed by the geochemistry and mineralogy of geothermal systems.

microbiology↗

bioRxiv2020

Although host-related factors are important for the formation of gut microbiota, environmental factors cannot be ignored

The gut microbiome is essential to human health. However, little is known about the influence of the environment versus host-related factors (e.g. genetic background, sex, age, and body mass) in the formation of human intestinal microflora. Here, we present evidence in support of the importance of host-related factors in the establishment and maintenance of individual gut assemblages. We collected fecal samples (n = 249) from 44 Korean naval trainees and 39 healthy people living in Korea over eight weeks and sequenced the bacterial 16S rRNA genes. The following hypotheses were tested: 1) microbiome function is linked to its diversity, community structure, and genetic host-related factors, and 2) preexisting host-related factors have a more significant effect on gut microbiome formation and composition than environmental factors. For each individual, the difference between the initial gut microbiota and that after eight weeks was negligible even though the 44 naval trainees lived in the same area and received the same diet, the same amount of exercise, and the same amount of physical stress during the study. This suggests that host-related factors, rather than environmental factors, is a key determinant of individual gut microflora. Moreover, eight weeks of physical training and experiencing the same environmental conditions resulted in an increase in the species Bifidobacterium, Faecalibacterium, and Roseburia in most trainees, suggesting a healthier intestinal environment. IMPORTANCEIn order to understand the role of human gut microbiome, it is important to know how individuals gut microbiota are formed. In this study, we tested the host-related factors versus environmental factors to affect gut microbiome and found that the former have a more association. However, we also found that the controlled environment give an effect on the gut microflora as well. This study provides preliminary evidence that differences in the formation and diversity of gut microbiota within a population could be determined by host-related factors rather than environmental factors.

microbiology↗

bioRxiv2020

Polar opposites; bacterioplankton susceptibility and mycoplankton resistance to ocean acidification

Microorganisms form the basis of ocean ecosystems yet the effects of perturbations such as decreasing pH on microbial community structure, interactions and functionality remain compared to multicellular organisms. Using an experimental manipulation of Southern Ocean seawater, we subjected bacterioplankton and mycoplankton to artificial pH decreases, which are predicted to occur in the future. We show that acidification led to substantial increases of bacterioplankton diversity, while in contrast it had no effect on mycoplankton diversity. Our analyses revealed a loss of putative keystone taxa and a decrease in predicted community interactions as a response to lower pH levels. Bacterioplankton shifted from generalist to specialist community members, suggesting a specific stress response to unfavourable conditions. In addition, enzyme activities involved in nitrogen acquisition were lower at reduced pH levels, suggesting altered organic matter cycling in a more acidic ocean. Our findings suggest that bacterioplankton and mycoplankton may respond differentially to future ocean acidification, with potentially negative impacts on community structure and biogeochemical cycling in the Southern Ocean. IMPORTANCEOceans absorb the majority of anthropogenically produced CO2, the consequence of which is ocean acidification, a phenomenon already negatively impacting key marine organisms. Marine microbial communities form the basis of ocean food webs by generating nutrients for higher trophic levels, yet the response of these key microbial drivers to acidification remains unclear. This knowledge deficit is particularly true for understudied marine ecosystems such as the Southern Ocean. Using a mesocosm approach, we found that acidification severely impacts microbial community stability, by altering bacterioplankton community structure, reducing network complexity, and augmenting enzyme activities associated with nitrogen acquisition. This study adds to our understanding of the effects of ocean acidification on microbial communities, particularly within an environment expected to be largely effected by future anthropogenically driven climate change.

microbiology↗

bioRxiv2020

Dynamic Laser Speckle Imaging meets Machine Learning to enable Rapid Antibacterial Susceptibility Testing (DyRAST)

Rapid antibacterial susceptibility testing (RAST) methods which measure change of a bacterial phenotype in response to a given treatment are of significant importance in healthcare, as they can assist care-givers in timely administration of the right treatment. Various RAST techniques have been reported for tracking bacterial phenotypes, such as size, shape, motion, and metabolic activity. However, they still require bulky and expensive instruments (which hinders their application in resource-limited environments) and/or utilize labeling reagents (which can interfere with antibiotics and add to cost). Furthermore, the existing ultra-rapid methods do not address possible adaptation of gradual adaptation of bacteria to antibiotics, which can lead to false interpretation of resistance when using ultra-rapid methods. In this work, we present a RAST approach leveraging machine learning analysis of time-resolved dynamic laser speckle imaging (DLSI) results to accurately predict the minimum inhibitory concentration (MIC) of a model strain of Escherichia coli in 60 minutes, compared to 6 hours using the currently FDA-approved phenotype-based RAST technique. To demonstrate the DLSI performance, we studied the effect of a {beta}-lactam ampicillin and an aminoglycoside gentamicin on Escherichia coli strain K-12. DLSI captures change of bacterial motion/division in response to treatment. The machine learning algorithm was trained and validated using the overnight results of gold standard, broth microdilution method. Empowered by machine learning, DyRAST can predict MIC with high accuracy comparable to gold standard methods through a voting strategy.

microbiology↗