Bacterial Copy Numbers Research Articles

Upper respiratory tract detection of Mycoplasma ovipneumoniae employing nasopharyngeal swabs

BackgroundFlock-level prevalence and characterization of Mycoplasma ovipneumoniae is determined almost exclusively using nasal swabbing followed by molecular detection with either quantitative PCR or multi-locus sequence typing. However, the diagnostic performance and efficiency of swabbing the nasal passage compared to other anatomical locations has not been determined within sheep populations. The goal of this research was to assess the diagnostic capability of nasopharyngeal swabs in comparison to nasal swabs for the detection of Mycoplasma ovipneumoniae.ResultsNasal and nasopharyngeal swabs were collected during a controlled exposure study of domestic sheep with Mycoplasma ovipneumoniae. Both swab types were then analyzed via conventional and quantitative PCR. This dataset showed that the use of nasopharyngeal swabs in lieu of nasal swabs resulted in higher sensitivity, reduced inhibition during quantitative PCR, and higher bacterial copy numbers per swab. Moreover, it was demonstrated that diagnostic sensitivity could be further increased during quantitative PCR via ten-fold dilution of the extracted DNA. To confirm these observations in naturally infected animals, we conducted a field study employing a production flock of domestic sheep using both nasal and nasopharyngeal swabbing techniques. Extracted DNA was assessed using the same molecular techniques, where detection of Mycoplasma ovipneumoniae was confirmed by sequencing of either the rpoB or 16S rRNA gene. Similar improvements were observed for nasopharyngeal swabs and template treatment methods within the naturally infected flock.ConclusionsResults demonstrate increased diagnostic sensitivity and specificity when sampling with nasopharyngeal swabs as compared to nasal swabs. Therefore, alternate field-testing strategies employing nasopharyngeal swabs should be considered for diagnosis of the presence of M. ovipneumoniae. Importantly, sample treatment following acquisition was found to affect the sensitivity of quantitative PCR, where dilution of eluted DNA template doubled the calculated sensitivity. This demonstrates that, in addition to anatomical location, the presence of inhibitory components in swab extracts also strongly influences diagnostic performance.

Rapid bacterial evaluation beyond the colony forming unit in osteomyelitis.

Examination of bacteria/host cell interactions is important for understanding the aetiology of many infectious diseases. The colony forming unit (CFU) has been the standard for quantifying bacterial burden for the past century, however, this suffers from low sensitivity and is dependent on bacterial culturability in vitro. Our data demonstrate the discrepancy between the CFU and bacterial genome copy number in an osteomyelitis-relevant co-culture system and we confirm diagnosis and quantify bacterial load in clinical bone specimens. This study provides an improved workflow for the quantification of bacterial burden in such cases.

Rapid bacterial evaluation beyond the colony forming unit in osteomyelitis

Examination of bacteria/host cell interactions is important for understanding the aetiology of many infectious diseases. The colony forming unit (CFU) has been the standard for quantifying bacterial burden for the past century, however, this suffers from low sensitivity and is dependent on bacterial culturability in vitro. Our data demonstrate the discrepancy between the CFU and bacterial genome copy number in an osteomyelitis-relevant co-culture system and we confirm diagnosis and quantify bacterial load in clinical bone specimens. This study provides an improved workflow for the quantification of bacterial burden in such cases.

Sub-optimal temperatures lead to altered expression of stress-related genes and increased 'Candidatus Liberibacter solanacearum' accumulation in potato psyllid.

The potato psyllid Bactericera cockerelli is the insect vector of the fastidious bacterium 'Candidatus Liberibacter solanacearum'. The bacterium infects both B. cockerelli and plant species, causing zebra chip (ZC) disease of potato and vein-greening disease of tomato. Temperatures are known to influence the initiation and progression of disease symptom in the host plant, and seasonal transitions from moderate to high temperatures trigger psyllid dispersal migration to facilitate survival. 'Ca. L. solanacearum' -infected and uninfected psyllids were reared at previously established 'permissible', optimal, and 'non-permissible' and temperatures of 18°C, 24°C, and 30°C, respectively. Gene expression profiles for 'Ca. L. solanacearum'-infected and -uninfected adult psyllids reared at different temperatures were characterized by Illumina RNA-Seq analysis. Bacterial genome copy number was quantified by real-time quantitative-PCR (qPCR) amplification. Relative gene expression profiles varied in psyllids reared at the three experimental temperatures. Psyllids reared at 18°C and 30°C exhibited greater fold-change increased expression of stress- and 'Ca. L. solanacearum' invasion-related proteins. Quantification by qPCR of bacterial genome copy number revealed that 'Ca. L. solanacearum' accumulation was significantly lower in psyllids reared at 18°C and 30°C, compared to 24°C. Temperature is a key factor in the life history of potato psyllid and multiplication/accumulation of 'Ca. L. solanacearum' in both the plant and psyllid host, influences the expression of genes associated with thermal stress tolerance, among others, and may have been instrumental in driving the co-evolution of the pathosystem.

Fertilizer management methods affect bacterial community structure and diversity in the maize rhizosphere soil of a coal mine reclamation area

BackgroundThe filling or mixed stack mode is a frequently used coal mine reclamation engineering technique that results in changes in soil microbial community structure and nutrient content, which lead to considerable deviations from the characteristics of restored coal mine reclamation areas that can be used for farming. Fertilization is an effective strategy for improving soil fertility in such areas; however, the response of soil bacterial communities, especially in the crop rhizosphere soil, to different fertilization techniques in such soils remains unclear. Therefore, we investigated the effects of different fertilization management methods, including no fertilizer, farmers' practice, inorganic fertilizer, organic fertilizer, and organic–inorganic fertilizer, on maize yield, rhizosphere soil bacterial community and diversity, soil physicochemical properties, and nitrogen cycle-related gene abundance (nifH, AOB, and nirS).ResultsThe results showed that organic–inorganic fertilizer treatment significantly improved maize yield. The relative abundance of the dominant phyla did not significantly differ between the treatment groups. However, the Chao 1 and Shannon indices of the bacterial community significantly changed between the no fertilizer and organic–inorganic fertilizer treatments. Notably, organic–inorganic fertilizer application significantly increased the copy numbers of nifH and nirS. Further, moisture, bulk density, and available phosphorus content were identified as the major driving factors responsible for the changes in bacterial community structure, diversity, and copy numbers of nifH, nirS, and AOB.ConclusionsThe results of this study revealed that organic–inorganic fertilizer application improved soil bacterial diversity and the copy numbers of nifH and nirS in maize rhizosphere soil. Therefore, we concluded that organic–inorganic fertilizer is an effective strategy for the restoration of maize rhizosphere soil properties and bacterial communities in coal mine reclamation areas.

Effect of Cork Flour Supplementation on the Growth Indices and Rhizosphere Bacterial Communities of Quercus variabilis Seedlings

Substrate and rhizosphere microorganisms are key factors affecting seedling growth; however, the effects of seedling substrates and rhizosphere bacteria on the growth of Quercus variabilis are not completely understood. Here, Q. variabilis seedlings were grown in substrates with and without cork flour, as follows: H substrate (charcoal soil/cork flour/perlite, 1:1:2), S substrate (cork flour/perlite, 1:1), and the control (CK) substrate (charcoal soil/perlite, 3:2). High-throughput sequencing and qPCR were used to investigate the effects of these substrates on seedling growth, physiological indices, and rhizosphere bacterial communities. Root and shoot weights of seedlings grown in H and S substrates were significantly higher than those of seedlings grown in CK. Moreover, H was conducive to chlorophyll synthesis in seedling leaves, and the transpiration rate and intercellular CO2 concentration of the leaves of seedlings grown in H were higher than those of seedlings grown in CK. The number of rhizosphere bacterial 16S rRNA copies was significantly greater in the case of seedlings grown in S than for those grown in H and CK. As well, rhizosphere bacterial richness was higher in seedlings grown in H and S than in those grown in CK. Thus, cork-flour-supplemented substrates are beneficial for seedling growth and development, seedling rhizosphere bacterial abundance and diversity, and the abundance of nitrogen and phosphorus metabolism-promoting microbial taxa.

Is XP-endo Finisher a better treatment option for its efficacy against intracanal bacteria for post-treatment apical periodontitis cases than EndoActivator?

To investigate the efficacy of the supplementary use of a rotary agitation method [XP-endo Finisher (XPF)] and sonically-activated irrigation [EndoActivator (EA)], using droplet digital PCR (ddPCR) on reducing the bacterial load in previously root canal treated teeth with apical periodontitis. Twenty patients with post-treatment apical periodontitis were allocated into two groups according to the irrigation activation method used: XPF and EA group. Total bacterial loads, as well as the amount of Enterococcus faecalis (E. faecalis) were determined before (S1) and after (S2) chemomechanical preparation, and after final irrigation activation (S3) by means of ddPCR. The bacterial copy numbers were compared between groups using the Friedman test (Nonparametric Repeated Measures ANOVA). When the groups were examined in terms of gender, age, number of root canals, periapical index score, sterility control total bacteria (SCTB), S1- and S2-total bacteria copy number, it was found that there was no statistical difference between the XPF group and the EA group (p > 0.05). Subsequent activation (S3) resulted in a significant microbial reduction in both XPF and EA groups, both of which reduced significantly more bacteria than chemomechanical instrumentation (S2) (p < 0.0001). On the contrary, S3-total bacteria copy number of the EA group was lower than the XPF group (p < 0.0147). There was no statistical difference between the XPF group and the EA group in terms of E. faecalis copy number (p > 0.05). Although both the XPF and the EA optimised the antibacterial efficiency of chemomechanical preparation in previously root canal-treated teeth with apical periodontitis, a lower total bacterial copy number was achieved with the EA application than the XPF application.

Shifts in soil prokaryotic and microeukaryotic communities following a translocation of wet meadows to derelict land

AbstractHuman exploitation of natural resources dictates the need for the sustainable use of lands already in use. To combine the conservation of meadows with land recycling, 13,000 m2 of turf were translocated from a built‐up area to an abandoned quarry. Three Molinion meadows were monitored for three seasons—before the translocation and for two years afterwards. The composition of the main groups of microorganisms—bacteria, archaea, fungi, microfauna, and unicellular algae—was examined to check the stability of soil ecosystem. After observing changes in the ratio of ammonia to nitrate in the translocated turf, the bacterial and archaeal ammonia monooxygenase gene (amoA) copy numbers were quantified. Following the translocation, the α‐diversity of prokaryotes and eukaryotes remained usually similar. The structure of the microbial communities has changed: relative abundance of Acidobacteria (Acidobacteriota) and Mortierellomycota increased, along with fungal saprotrophs, while several other phyla decreased. The number of bacterial amoA gene copies has risen 7‐fold in two years. The slight acidification of the soil had an influence; however, the presumed turf aeration and moisture drop initially impacted the soil communities. In conclusion, turf block translocation preserves diversity. However, cutting turf for transportation followed by dryer conditions at the receptor location lead to a functional switch in microbial communities. Increased activities of ammonia oxidisers and saprotrophs proceeded to nitrification and humus degradation, which may indicate degradation of soil. When translocating wet meadows, it is crucial to ensure the hydrological regime at receptor sites to support long‐term success.

Aquatic insects differentially affect lake sturgeon larval phenotypes and egg surface microbial communities.

Documentation of how interactions among members of different stream communities [e.g., microbial communities and aquatic insect taxa exhibiting different feeding strategies (FS)] collectively influence the growth, survival, and recruitment of stream fishes is limited. Considerable spatial overlap exists between early life stages of stream fishes, including species of conservation concern like lake sturgeon (Acipenser fulvescens), and aquatic insects and microbial taxa that abundantly occupy substrates on which spawning occurs. Habitat overlap suggests that species interactions across trophic levels may be common, but outcomes of these interactions are poorly understood. We conducted an experiment where lake sturgeon eggs were fertilized and incubated in the presence of individuals from one of four aquatic insect FS taxa including predators, facultative and obligate-scrapers, collector-filterers/facultative predators, and a control (no insects). We quantified and compared the effects of different insect taxa on the taxonomic composition and relative abundance of egg surface bacterial and lower eukaryotic communities, egg size, incubation time to hatch, free embryo body size (total length) at hatch, yolk-sac area, (a measure of resource utilization), and percent survival to hatch. Mean egg size varied significantly among insect treatments. Eggs exposed to predators had a lower mean percent survival to hatch. Eggs exposed to predators had significantly shorter incubation periods. At hatch, free embryos exposed to predators had significantly smaller yolk sacs and total length. Multivariate analyses revealed that egg bacterial and lower eukaryotic surface community composition varied significantly among insect treatments and between time periods (1 vs 4 days post-fertilization). Quantitative PCR documented significant differences in bacterial 16S copy number, and thus abundance on egg surfaces varied across insect treatments. Results indicate that lethal and non-lethal effects associated with interactions between lake sturgeon eggs and free embryos and aquatic insects, particularly predators, contributed to lake sturgeon trait variability that may affect population levels of recruitment.

Microbial molecule ingress promotes neuroinflammation and brain CCR5 expression in persons with HIV-associated neurocognitive disorders

BackgroundSystemic inflammation accompanies HIV-1 infection, resulting in microbial translocation from different tissues. We investigated interactions between lentivirus infections, neuroinflammation and microbial molecule presence in the brain. MethodsBrain tissues from adult humans with (n = 22) and without HIV-1 (n = 11) infection as well as adult nonhuman primates (NHPs) with (n = 11) and without (n = 4) SIVmac251 infection were investigated by RT-PCR/ddPCR, immunofluorescence and western blotting. Studies of viral infectivity, host immune gene expression and viability were performed in primary human neural cells. FindingsAmong NHPs, SIV DNA quantitation in brain showed increased levels among animals with SIV encephalitis (n = 5) that was associated with bacterial genomic copy number as well as CCR5 and CASP1 expression in brain. Microbial DnaK and peptidoglycan were immunodetected in brains from uninfected and SIV-infected animals, chiefly in glial cells. Human microglia infected by HIV-1 showed increased p24 production after exposure to peptidoglycan that was associated CCR5 induction. HIV-1 Vpr application to human neurons followed by peptidoglycan exposure resulted in reduced mitochondrial function and diminished beta-III tubulin expression. In human brains, bacterial genome copies (250–550 copies/gm of tissue), were correlated with increased bacterial rRNA and GroEL transcript levels in patients with HIV-associated neurocognitive disorders (HAND). Glial cells displayed microbial GroEL and peptidoglycan immunoreactivity accompanied by CCR5 induction in brains from patients with HAND. InterpretationIncreased microbial genomes and proteins were evident in brain tissues from lentivirus-infected humans and animals and associated with neurological disease. Microbial molecule translocation into the brain might exacerbate neuroinflammatory disease severity and represent a driver of lentivirus-associated brain disease.

Unexpected high carbon losses in a continental glacier foreland on the Tibetan Plateau

Closely related with microbial activities, soil developments along the glacier forelands are generally considered a carbon sink; however, those of continental glacier forelands remain unclear. Continental glaciers are characterized by dry conditions and low temperature that limit microbial growth. We investigated the carbon characteristics along a chronosequence of the Laohugou Glacier No. 12 foreland, a typical continental glacier on the Tibetan Plateau, by analyzing soil bacterial community structure and microbial carbon-related functional potentials. We found an unexpected carbon loss in which soil organic carbon decreased from 22.21 g kg−1 to 10.77 g kg−1 after receding 50 years. Structural equation modeling verified the important positive impacts from bacterial community. Lower carbon fixation efficiency along the chronosequence was supported by less autotrophic bacteria and carbon fixation genes relating to the reductive tricarboxylic acid cycle. Lower carbon availability and higher carbon requirements were identified by an increasing bacterial copy number and a shift of the dominant bacterial community from Proteobacteria and Bacteroidetes (r-strategists) to Actinobacteria and Acidobacteria (K-strategists). Our findings show that the carbon loss of continental glacier foreland was significantly affected by the changes of bacterial community, and can help to avoid overestimating the carbon sink characteristics of glacier forelands in climate models.

Antibiotics for lower respiratory tract infection in children presenting in primary care (ARTIC-PC): the predictive value of molecular testing

ObjectivesThis study aimed to assess whether the presence of bacteria or viruses in the upper airway of children presenting with uncomplicated lower respiratory tract infection (LRTI) predicts the benefit of antibiotics. MethodsChildren between 6 months and 12 years presenting to UK general practices with an acute LRTI were randomized to receive amoxicillin 50 mg/kg/d for 7 days or placebo. Children not randomized (ineligible or clinician/parental choice) could participate in a parallel observational study. The primary outcome was the duration of symptoms rated moderately bad or worse. Throat swabs were taken and analyzed for the presence of bacteria and viruses by multiplex PCR. ResultsSwab results were available for most participants in the trial (306 of 432; 71%) and in the observational (182 of 326; 59%) studies. Bacterial pathogens potentially sensitive to amoxicillin (Haemophilus influenzae, Moraxella catarrhalis, Streptococcus pneumoniae) were detected among 51% of the trial placebo group and 49% of the trial antibiotic group. The median difference in the duration of symptoms rated moderately bad or worse between antibiotic and placebo was similar when potentially antibiotic-susceptible bacteria were present (median: –1 day; 99% CI, –12.3 to 10.3) or not present (median: –1 day; 99% CI, –4.5 to 2.5). Furthermore, bacterial genome copy number did not predict benefit. There were similar findings for all secondary outcomes and when including the data from the observational study. DiscussionThere was no clear evidence that antibiotics improved clinical outcomes conditional on the presence or concentration of bacteria or viruses in the upper airway. Before deploying microbiologic point-of-care tests for children with uncomplicated LRTI in primary care, rigorous validating trials are needed.

Rumen-protected glucose supplementation in transition dairy cows shifts fermentation patterns and enhances mucosal immunity.

Manipulation of perinatal diets, such as supplementing feed with rumen-protected glucose (RPG), has been positively regarded as a strategy to improve milking performance. This study was conducted to assess the effects of RPG on the fermentation profiles, resident microbiota and mucosal immunity in the cecum. Ten Holstein dairy cows were randomly assigned to either a 25 g/kg RPG diet (DM basis) or a 11 g/kg coating fat diet (control, CON). Compared with the CON group, the acetate-to-propionate ratio was lower in the RPG group. Gene expression analysis indicated that RPG supplementation tended to upregulate the expression of Na+/H+ hydrogen exchanger 3 (NHE3) (P = 0.076). RPG supplementation downregulated the expression of genes involved in self-rehabilitation such as matrix metalloproteinase 1 (MMP1), MMP3, MMP9 and MMP13. Additionally, the mRNA expression of genes involved in immunity including Toll-like receptors (TLR4, TLR6 and TLR7) and proinflammatory cytokines (immune interferon gamma [IFNG] and interleukins interleukin 17A [IL17F], IL17A, IL22), was downregulated by RPG supplementation. Nonetheless, no differences existed in the bacterial copy number and beta diversity between the 2 groups. Overall, supplementation with RPG would probably cause a shift towards propionate production in the cecal digesta, and promote the immune homeostasis of the cecal mucosa in transition dairy cows. Our results extended the basic understanding of RPG supplementation and utilization in transition dairy cows in terms of host microbe interplay in the cecum.

Soil microbial communities are driven by the declining availability of cations and phosphorus during ecosystem retrogression

Development of soil microbial communities is driven by local abiotic and biotic conditions, yet our current understanding of their ecology is limited to studies in modified or young and relatively fertile ecosystems. In nutrient-impoverished soils, microbial communities may be predominantly structured by availability of key elements such as phosphorus (P) or nitrogen, regardless of the taxa (bacteria, archaea or fungi). Here, we determined if shifts in bacterial communities and biomass occurred along a long-term soil chronosequence in south-western Australia, and whether such shifts were related to changes in soil nutrient availability along the retrogressive phase of this chronosequence. We quantified shifts in the biomass and abundance of major soil microbial groups using phospholipid fatty acids and quantitative polymerase chain reaction. We found that declining soil P availability limited microbial growth and community structure. Common cations, notably magnesium, showed strong relationships with the abundance of most bacterial and archaeal taxa. In the oldest most severely impoverished soils, the fungal to bacteria ratio was higher than in the young soils, indicating fungi are better adapted to such environments. Finally, archaeal copy numbers decreased more strongly than bacterial copy numbers with increasing nutrient-impoverishment of soils. The study enhances our understanding of the ecology of soil bacteria, archaea, and fungi during ecosystem development and retrogression.

Abstract MP38: Identification Of A Gut Microbe That Attenuates The Blood Pressure Lowering Effect Of ACEi Quinapril

Introduction: Treatment resistant hypertension (rHTN) is present in ~20% of all hypertensive patients. rHTN is critical in African American patients who experience early onset, severe outcomes, and weak responsiveness to angiotensin converting enzyme inhibitor (ACEi). The mechanism for drug resistance is unknown. Gut microbiota harbors biotransformative enzymes such as esterase, which may hydrolyze ACEis, reducing absorption. Our study was to identify microbe responsible for ACEi resistance. Methods: 16-week-old male Spontaneously Hypertensive Rats (SHR) were gavaged with (N=12) or without (N=6) Vancomycin, Meropenem, and Omeprazole (VMO) 50 mg/kg/day for five days to deplete the gut microbiota. A single 8mg/kg dose of quinapril was given to SHR and SHR+VMO before blood pressure (BP) recording via telemetry. Quinapril catabolism was quantified by liquid chromatography-mass spectrometry. Bacterial esterase activity was monitored by hydrolysis of p-nitro-phenylbutyrate. Cecal microbiota was analyzed by 16S rDNA. Fecal microbiota were analyzed by metagenomics in 29 (16 black, 13 white) HTN patients. Results: Quinapril lowered BP more in the SHR+VMO than SHR ( P &lt;0.0001). With a 50% reduction in bacterial 16S copy numbers ( P &lt;0.0001), the SHR+VMO group showed (1) reduced Coprococcus ( P &lt;0.0001); (2) lower esterase activity per gram of cecal microbiota to hydrolyze quinapril ( P =0.0065); (3) a 50% lower reduction in quinapril quantity (nmol) after incubation with 1mg of cecal lysate for 3 hr ( P &lt;0.0001); (4) decreased bacterial genes in KEGG drug metabolism pathway ( P &lt;0.0001). The abundance of Coprococcus positively correlated with genes in drug metabolism ( P &lt;0.0001). Importantly, co-administration of quinapril with C. comes, a species in Coprococcus genus, reduced the BP-lowering effects of quinapril in the SHR ( P &lt;0.0001). Comparison of human microbiota demonstrated a higher abundance of C. comes in the black hypertensives (poor ACEi responder) than the white (ACEi responder) ( P =0.0167). Conclusion: We, for the first time, demonstrate a previously unrecognized role of gut microbes in reducing ACEi effectiveness. This serves a foundation for expanding clinical management of antihypertensive drug resistance via manipulation of gut microbiota.

Distribution and localization of Streptococcus agalactiae in different tissues of artificially infected tilapia (Oreochromis niloticus)

China's tilapia industry has the most complete industrial chain in the world, but, in recent years, tilapia streptococcosis have frequently occurred, which has become a bottleneck restricting the development of tilapia industry. Although plenty of studies have been conducted on Streptococcus agalactiae (Group B Streptococcus, GBS) in tilapia, the mechanism of GBS invasion is still unclear. In order to further understand the invasion route, we used the qPCR method to study bacteria load in liver, intestine, gill, spleen, head kidney, kidney, and brain tissues of GIFT tilapia infected by a GBS virulent strain WC1535 at different infection time and the histological method to systematically analyze bacterial distribution and localization in above infected tissue. Through the dynamic analysis of bacteria load, the bacterial copy number of WC1535 reached the peak in each tissue at 24 h post-infection (hpi). Therefore, tissues at 24 hpi were taken as the research object to investigate the distribution and localization of WC1535 by paraffin section and electron microscopy. The results of tissue section showed that WC1535 existed in three forms in blood: free scattering without attachment to any structure, phagocytized by phagocytes (mainly macrophages) and a few red blood cells, and attached to the inner wall of blood vessels. The results of qPCR and tissue section both showed that the density of bacteria in head kidney, spleen and gill was high, at the same time, these tissues had abundant blood and capillaries, so this suggested that the S. agalactiae existed mainly in the blood of tilapia. There were almost no pathogens in the parenchymal cells of each tissue, but the surface of epithelial cells such as vascular endothelial cells and fibroblasts adhered a lot, which indicated that S. agalactiae had epithelial cell affinity. The results of electron microscopy showed that S. agalactiae can survive and proliferate intracellularly after being phagocytosed by macrophages, suggesting that macrophages might play as pathogen carriers or “Trojan horses” to achieve the immune escape, break the blood-brain barrier (BBB), and lead the meningitis in tilapia. Since GBS of tilapia can be internalized by macrophages, antibiotics with strong tissue distribution, cell penetration and good meningeal penetration are recommended for the treatment in the early stage of streptococcosis.

Depletion of Esterase‐Harboring Bacteria Increases Antihypertensive Efficacy of ACE Inhibitor Quinapril

Introduction Resistant hypertension (rHTN) is a critical impediment to successful control of high blood pressure (BP) and hypertension-linked pathophysiology in 15-20% of all hypertensive patients. rHTN is of importance in African American patients because they experience early onset, increased prevalence and severe outcomes. The mechanisms for drug resistance in rHTN remain unknown. Eggerthellaceae family harbors biotransformative enzymes including esterase, which influences drug catabolism in the gut prior to absorption. Thus, we hypothesized that an increased abundance of bacteria with catabolic capacity for an antihypertensive drug will decrease its efficacy (Fig. A) and may thereby serve as a novel treatment target for rHTN. Methods Ten human feces (4 controlled HTN (cHTN), 6 rHTN) were analyzed by metatranscriptomics. 16-week old male Spontaneously Hypertensive Rats (SHR) were gavaged with (N=12) or without (N=6) Vancomycin, Meropenem, and Omeprazole (VMO), at 50 mg/kg/day for five days to deplete the gut microbiota. To test the quinapril efficacy, a single 8mg/kg dose was gavaged to both SHR and SHR+VMO before BP recording via telemetry. Quinapril catabolism was quantified by liquid chromatography-mass spectrometry. Bacterial esterase activity was monitored by the hydrolysis of p-nitro-phenylbutyrate. Cecal microbiota was analyzed by 16S rDNA. Pearson r correlation was plotted using GraphPad. Results Patients with rHTN showed ~3-fold more Eggerthellaceae compared to cHTN (p=0.0027, Fig. B). With 50% reduction in bacterial 16S copy numbers (P<0.0001, Fig. C), the SHR+VMO group showed (1) reduced Eggerthellaceae (P<0.0001, Fig. D); (2) lower esterase activity per gram of cecal microbiota to hydrolyze quinapril (p=0.0065, Fig. E); (3) a 50% lower reduction in quinapril quantity (Δquinapril, nmol) after incubation with 1mg of cecal lysate (P<0.0001, Fig. F); (4) decreased bacterial genes in KEGG drug metabolism pathway (P<0.0001, Fig. G). The abundance of Eggerthellaceae positively correlated with Δquinapril (P=0.029, Fig. H) and genes in drug metabolism (P<0.0001, Fig. I). Importantly, administration of quinapril to the SHR+VMO led to lowering of BP, compared to SHR (Fig. J). Conclusion Eggerthellaceae was enriched ~3-fold in rHTN. Using SHR, we showed that depletion of Eggerthellaceae and esterase activity of the gut microbiota by VMO increased the antihypertensive efficacy of quinapril, due to protection of quinapril from catabolism in the gut. Therefore, our study, for the first time, provides evidence for a previously unrecognized role of gut microbiota in rHTN. Innovative approaches can now be considered for rHTN based on strategies to protect antihypertensive drugs from catabolism by gut bacteria before reaching the systemic circulation for improved efficacy.

Application of an indigenous microorganisms-based fixed-bed GAC-biofilm reactor for passive and sustainable treatment of oil sands process water through combined adsorption and biodegradation processes

In this study, a fixed-bed biofilm reactor (biofilter) was developed and applied for oil sands process water (OSPW) remediation by using granular activated carbon (GAC) as packing media. Using quantitative polymerase chain reaction (qPCR) detection, the total bacterial copy number (16S) in the GAC biofiltration system was found to reach a relatively stable level (1.3 ± 0.2 × 109 copies/g GAC) after 62 days of operation, and the thickness of biofilm on GAC surface was 26.7 ± 4.3 μm based on the scan of confocal laser scanning microscopy (CLSM). The established GAC-biofilter showed 95.4% naphthenic acids (NAs) removal from raw OSPW after 2 months of operation. The GAC-biofilter also showed 88.3% NAs removal after a long operation time (2 years), indicating its sustainable bioremediation capacity for OSPW. 16S and 18S rRNA gene-targeted metagenomic sequencing showed that the microbial community in the GAC biofilter had higher diversity and richness than that found in the sand biofilter which was used for OSPW treatment previously. Comamonadaceae and Saccharomycotina were found to be the dominant bacterial and fungal families in the GAC biofilter, respectively. Xenobiotic metabolism function of the microbial community may contribute significantly to the biodegradation of NAs. The GAC biofiltration process is a promising passive OSPW treatment approach that can be used in-situ.

Assessment of the Impact of No-Till and Conventional Tillage Technologies on the Microbiome of Southern Agrochernozems

Quantitative characteristics of microbial communities in southern agrochernozems of the Stavropol region managed with the use of no-till technology and moldboard plowing under different crops with and without application of mineral have been studied. The number of eco-trophic groups of microorganisms (nitrogen fixers, ammonifiers, denitrifiers, amylolytics, cellulolytics) was estimated using the classical method of inoculation on elective media; and the number of copies of ribosomal genes of archaea, bacteria, and fungi was determined using real-time polymerase chain reaction (PCR). The numbers of cultivated aerobic ammonifiers, denitrifiers, aerobic cellulolytics, actinomycetes, and micromycetes increase in the soils under conventional tillage, whereas no-till technology leads to an increase in the numbers of anaerobic cellulolytics and nitrogen fixers and aerobic diazotrophs and amylolytics. Corn stimulates the development of ammonifiers and amylolytics in the soil microbial community; soybeans, aerobic nitrogen fixers and aerobic cellulolytics; and sunflower, denitrifiers and anaerobic cellulolytics. The largest number of bacterial and fungal gene copies is characteristic of no-till plots and unmanaged long-term fallow, whereas gene copies of archaea have only been identified in the soil of long-term fallow. The data obtained attest to different directions and intensities of soil biological processes in agrochernozems under the conditions of no-till and conventional tillage technologies.

First insights to the microbial communities in the plant process water of the multi-metal Kevitsa mine

Metallurgical processes demand large quantities of water. However, in many locations, water is becoming scarce and process water recycling is needed. Closing water loops can be challenging due to build-up of flotation chemicals, metal ions and microorganisms in the recycled water affecting the flotation performance. Here, we have characterized the microbial communities over a 2-month period in different locations of the multi-metal Kevitsa mine in Northern Finland, by microbiome sequencing, enumeration of bacteria, archaea and fungi by qPCR, and cultivation. The microbial communities showed high diversity, but were dominated by Alpha- and Gammaproteobacteria. In addition, various fungal taxa were detected, whereas the archaeal taxa were only sparsely detected from the sequence data. The number of bacterial 16S rRNA gene copies in Process water and Ni thickener overflow varied between 0.5–3.3 × 105 mL−1, whereas the Flotation tailings showed two orders of magnitude lower amounts. Fungi were present at 3.0 × 102–8.1 × 104 5.8S rRNA gene copies mL−1 in all samples, while the number of archaea fluctuated between 8.8 × 101–3.2 × 105 16S rRNA gene copies mL−1. The number of all microbial groups were generally lower in September than in August. When tested on 8 different cultivation media, the microorganisms generally responded positively to organic carbon, and were also shown to oxidize thiosulfate, which may indicate that build-up of organic flotation chemicals and sulfur species from the ore may cause the microbial numbers to increase. This study is part of the H2020 ITERAMS project (Grant agreement# 730480), which strives to improve the recycling of water and minimize the environmental impact of mines.