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1.
Med Microbiol Immunol ; 209(6): 669-680, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32880037

RESUMO

The ability of bacteria to aggregate and form biofilms impairs phagocytosis by polymorphonuclear leukocytes (PMNs). The aim of this study was to examine if the size of aggregates is critical for successful phagocytosis and how bacterial biofilms evade phagocytosis. We investigated the live interaction between PMNs and Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli and Staphylococcus epidermidis using confocal scanning laser microscopy. Aggregate size significantly affected phagocytosis outcome and larger aggregates were less likely to be phagocytized. Aggregates of S. epidermidis were also less likely to be phagocytized than equally-sized aggregates of the other three species. We found that only aggregates of approx. 5 µm diameter or smaller were consistently phagocytosed. We demonstrate that planktonic and aggregated cells of all four species significantly reduced the viability of PMNs after 4 h of incubation. Our results indicate that larger bacterial aggregates are less likely to be phagocytosed by PMNs and we propose that, if the aggregates become too large, circulating PMNs may not be able to phagocytose them quickly enough, which may lead to chronic infection.


Assuntos
Biofilmes , Escherichia coli/fisiologia , Neutrófilos/fisiologia , Fagocitose , Pseudomonas aeruginosa/fisiologia , Staphylococcus aureus/fisiologia , Staphylococcus epidermidis/fisiologia , Escherichia coli/ultraestrutura , Humanos , Pseudomonas aeruginosa/ultraestrutura , Pele/microbiologia , Staphylococcus aureus/ultraestrutura , Staphylococcus epidermidis/ultraestrutura
2.
J Med Life ; 13(2): 200-205, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32742514

RESUMO

The work assessed the state of the intestinal microbiocenosis in 52 puerperae at the in whom the pregnancy developed against the background of the metabolic syndrome. The diagnosis of metabolic syndrome was determined according to the criteria approved by the World Health Organization for pregnant women. The state of intestinal microbiocenosis was assessed by a bacteriological examination of feces immediately after delivery. The content of the main representatives of the obligate microflora (bifidobacteria, lactobacilli, native intestinal bacilli, fecal streptococci) and facultative (conditionally pathogenic) microorganisms (representatives of the genus Prоteus, Klebsiella, pathogenic strains of E. coli, Staphylococcus epidermidis, Enterobacter, Citrobacter, Clostridium difficile, Candida fungi) was determined. Cultures were made on appropriate growth media. At the time of birth, all patients of group I showed signs of intestinal microbiocenosis disorder. At the same time, 13 (54.2%) puerperae were diagnosed signs of dysbiosis of II degree, 9 (37.5%) with signs of III degree, which were generally characterized by a significant decrease in the content of the main representatives of obligate microflora (Bifidobacterium, Lactobacillus, Escherichia coli, Fecal streptococci) with simultaneous high contamination of Candida albicans and Clostridium difficile. So, it can be considered as a possible predictor of very early preterm birth in women with MS. In pregnant women with MS, but who gave timely birth (group II), dysbiotic disorders were detected to a lesser extent. Thus, in 13 (46.4%) patients, initial signs of intestinal dysbiosis (first degree) were detected in 4 (14.3%) patients (second degree). In 11 (39.3%) puerperae of group II, microbial indices indicated normal eubiotic ratios.


Assuntos
Disbiose/microbiologia , Disbiose/patologia , Intestinos/microbiologia , Intestinos/patologia , Síndrome Metabólica/complicações , Trabalho de Parto Prematuro/microbiologia , Adulto , Bifidobacterium/fisiologia , Escherichia coli/fisiologia , Fezes/microbiologia , Feminino , Humanos , Recém-Nascido , Lactobacillus/fisiologia , Gravidez , Adulto Jovem
3.
Nat Commun ; 11(1): 4013, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32782250

RESUMO

Antibiotics that interfere with translation, when combined, interact in diverse and difficult-to-predict ways. Here, we explain these interactions by "translation bottlenecks": points in the translation cycle where antibiotics block ribosomal progression. To elucidate the underlying mechanisms of drug interactions between translation inhibitors, we generate translation bottlenecks genetically using inducible control of translation factors that regulate well-defined translation cycle steps. These perturbations accurately mimic antibiotic action and drug interactions, supporting that the interplay of different translation bottlenecks causes these interactions. We further show that growth laws, combined with drug uptake and binding kinetics, enable the direct prediction of a large fraction of observed interactions, yet fail to predict suppression. However, varying two translation bottlenecks simultaneously supports that dense traffic of ribosomes and competition for translation factors account for the previously unexplained suppression. These results highlight the importance of "continuous epistasis" in bacterial physiology.


Assuntos
Antibacterianos/farmacologia , Modelos Teóricos , Biossíntese de Proteínas/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , Interações Medicamentosas , Epistasia Genética , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Biossíntese de Proteínas/fisiologia , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Ribossomos/efeitos dos fármacos , Ribossomos/metabolismo
4.
ACS Nano ; 14(9): 12045-12053, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32790338

RESUMO

Wearing face masks has been widely recommended to contain respiratory virus diseases, yet the improper use of masks poses a threat of jeopardizing the protection effect. We here identified the bacteria viability on common face masks and found that the majority of bacteria (90%) remain alive after 8 h. Using laser-induced graphene (LIG), the inhibition rate improves to ∼81%. Combined with the photothermal effect, 99.998% bacterial killing efficiency could be attained within 10 min. For aerosolized bacteria, LIG also showed superior antibacterial capacity. The LIG can be converted from a diversity of carbon precursors including biomaterials, which eases the supply stress and environmental pressure amid an outbreak. In addition, self-reporting of mask conditions is feasible using the moisture-induced electricity from gradient graphene. Our results improve the safe use of masks and benefit the environment.


Assuntos
Antibacterianos/farmacologia , Grafite/farmacologia , Lasers , Luz , Viabilidade Microbiana , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Staphylococcus epidermidis/efeitos dos fármacos , Staphylococcus epidermidis/fisiologia
5.
PLoS Pathog ; 16(7): e1008700, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32687537

RESUMO

With antibiotic resistance rates on the rise, it is critical to understand how microbial species interactions influence the evolution of resistance. In obligate mutualisms, the survival of any one species (regardless of its intrinsic resistance) is contingent on the resistance of its cross-feeding partners. This sets the community antibiotic sensitivity at that of the 'weakest link' species. In this study, we tested the hypothesis that weakest link dynamics in an obligate cross-feeding relationship would limit the extent and mechanisms of antibiotic resistance evolution. We experimentally evolved an obligate co-culture and monoculture controls along gradients of two different antibiotics. We measured the rate at which each treatment increased antibiotic resistance, and sequenced terminal populations to question whether mutations differed between mono- and co-cultures. In both rifampicin and ampicillin treatments, we observed that resistance evolved more slowly in obligate co-cultures of E. coli and S. enterica than in monocultures. While we observed similar mechanisms of resistance arising under rifampicin selection, under ampicillin selection different resistance mechanisms arose in co-cultures and monocultures. In particular, mutations in an essential cell division protein, ftsI, arose in S. enterica only in co-culture. A simple mathematical model demonstrated that reliance on a partner is sufficient to slow the rate of adaptation, and can change the distribution of adaptive mutations that are acquired. Our results demonstrate that cooperative metabolic interactions can be an important modulator of resistance evolution in microbial communities.


Assuntos
Adaptação Fisiológica/efeitos dos fármacos , Resistência Microbiana a Medicamentos/fisiologia , Escherichia coli/fisiologia , Interações Microbianas/fisiologia , Salmonella enterica/fisiologia , Adaptação Fisiológica/genética , Ampicilina/farmacologia , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Técnicas de Cocultura , Escherichia coli/efeitos dos fármacos , Interações Microbianas/efeitos dos fármacos , Modelos Teóricos , Mutação , Rifampina/farmacologia , Salmonella enterica/efeitos dos fármacos
6.
Proc Natl Acad Sci U S A ; 117(31): 18729-18736, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32669426

RESUMO

Many microorganisms face a fundamental trade-off between reproduction and survival: Rapid growth boosts population size but makes microorganisms sensitive to external stressors. Here, we show that starved bacteria encountering new resources can break this trade-off by evolving phenotypic heterogeneity in lag time. We quantify the distribution of single-cell lag times of populations of starved Escherichia coli and show that population growth after starvation is primarily determined by the cells with shortest lag due to the exponential nature of bacterial population dynamics. As a consequence, cells with long lag times have no substantial effect on population growth resumption. However, we observe that these cells provide tolerance to stressors such as antibiotics. This allows an isogenic population to break the trade-off between reproduction and survival. We support this argument with an evolutionary model which shows that bacteria evolve wide lag time distributions when both rapid growth resumption and survival under stressful conditions are under selection. Our results can explain the prevalence of antibiotic tolerance by lag and demonstrate that the benefits of phenotypic heterogeneity in fluctuating environments are particularly high when minorities with extreme phenotypes dominate population dynamics.


Assuntos
Farmacorresistência Bacteriana , Escherichia coli , Viabilidade Microbiana , Antibacterianos/farmacologia , Evolução Biológica , Escherichia coli/genética , Escherichia coli/fisiologia , Modelos Biológicos , Fenótipo , Análise de Célula Única
7.
Int J Food Microbiol ; 331: 108715, 2020 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-32554040

RESUMO

Ground beef contamination with Escherichia coli is usually a result of carcass faecal contamination during the slaughter process. Carcasses are contaminated when they come into contact with soiled hides or intestinal leakage content during dressing and the evisceration processes. A more recent and compelling hypothesis is that, when lymph nodes are present in manufacturing beef trimmings, they can be a potential source of Enterobacteriaceae contamination of ground beef. The aim of this study was to investigate the occurrence of E. coli in lymph nodes from beef carcasses used for ground meat production, in six slaughter plants situated in central Italy A total of 597 subiliac (precrural) lymph nodes were obtained from 597 cattle carcasses and screened for E. coli by culture. Furthermore, E. coli isolates (one per positive carcass) were tested for stx1, stx2 eaeA and hlyA genes that are commonly used to identify and characterise shiga toxin-producing E. coli (STEC). In addition, the E. coli isolates were profiled for antimicrobial susceptibility. A proportion of 34.2% (204/597) carcasses were positive for E. coli. PCR revealed that 29% (59/204) of E. coli possessed stx1 or stx2 which corresponded to 9.9% of the cattle sampled. Moreover, a combination of stx1 or stx2 and eaeA was found in in 4 isolates (2% among E. coli positive samples and 1% among cattle sampled) and a combination of stx1 or stx2 and eaeA and hly in 1 isolate (0.5% and 0.2%). More than 95% of isolates were susceptible to gentamicin, ceftriaxone, cyprofloxacin and cefotaxime while high rates of resistance were recorded for cephalotin, ampicillin, tetracycline, tripe sulfa and streptomycin. The multivariate analysis identified "age" as the factor most closely related to E. coli positivity (either generic E. coli or STEC) in bovine lymph nodes. In conclusion, subiliac lymph nodes represent a source of E. coli for ground beef. These results are of major importance for risk assessment and improving good manufacturing practices during animal slaughter and ground meat production.


Assuntos
Escherichia coli/fisiologia , Linfonodos/microbiologia , Carne/microbiologia , Animais , Bovinos , Escherichia coli/genética , Itália , Reação em Cadeia da Polimerase
8.
PLoS One ; 15(6): e0228294, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32479536

RESUMO

Even though Escherichia coli are common bacteria of the bovine vaginal microbiota, they represent an important pathogen that causes diseases in the reproductive tract and subfertility. However, the actual endometrial virulence profile of E. coli is poorly understood. The present study aims to characterize the phylogenetic structure and virulence potential of native vaginal populations of E. coli from healthy heifers (H), and cows with postpartum uterine diseases (PUD), such as metritis/endometritis (MT) or repeat breeder cows (RB). To this end, the virulence repertoire of 97 E. coli isolates was genotypically and phenotypically assessed. Most of them were assigned to phylogenetic group A (74%), followed by B1 (17%) and D (9%); RB strains were significantly (p < 0.05) more represented by B1. Seven of the 15 evaluated virulence genes (VFG) were detected and the most prevalent were fimH (87%), agn43 (41%) and csgA (35%); while traT (27%), fyuA (11%), hlyA (5%) and kpsMT II (5%) were observed in a lower proportion. Particularly, fyuA was significantly higher (p < 0.05) in MT cows whereas csgA showed the same behavior in PUD animals (p < 0.05). When comparing H and PUD strains, these last ones were associated to positive expression of biofilm, fimbriae curli/cellulose and motility; yet RB strains did not show motility. Vaginal B1 E. coli populations, that possess VFG (fyuA and csgA) as well as the expression of motility, curli fimbriae/cellulose and biofilm, may represent risk factors for endometrial disorders; specifically, those that also, have kpsMT II may have a pathogenic potential for causing the RB syndrome. Future research focusing on the detection of these strains in the vaginal microbiota of cows with postpartum uterine diseases should be done since the control of their presence in vagina could reduce the risk that they access the uterus during the postpartum period.


Assuntos
Doenças dos Bovinos/microbiologia , Escherichia coli/isolamento & purificação , Doenças Uterinas/veterinária , Vagina/microbiologia , Animais , Biofilmes/crescimento & desenvolvimento , Bovinos , Escherichia coli/genética , Escherichia coli/fisiologia , Feminino , Regulação Bacteriana da Expressão Gênica , Variação Genética , Intestinos/microbiologia , Filogenia , Doenças Uterinas/microbiologia
9.
Nat Commun ; 11(1): 2851, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32503979

RESUMO

The colonization of surfaces by bacteria is a widespread phenomenon with consequences on environmental processes and human health. While much is known about the molecular mechanisms of surface colonization, the influence of the physical environment remains poorly understood. Here we show that the colonization of non-planar surfaces by motile bacteria is largely controlled by flow. Using microfluidic experiments with Pseudomonas aeruginosa and Escherichia coli, we demonstrate that the velocity gradients created by a curved surface drive preferential attachment to specific regions of the collecting surface, namely the leeward side of cylinders and immediately downstream of apexes on corrugated surfaces, in stark contrast to where nonmotile cells attach. Attachment location and rate depend on the local hydrodynamics and, as revealed by a mathematical model benchmarked on the observations, on cell morphology and swimming traits. These results highlight the importance of flow on the magnitude and location of bacterial colonization of surfaces.


Assuntos
Aderência Bacteriana , Biofilmes/crescimento & desenvolvimento , Escherichia coli/fisiologia , Modelos Biológicos , Pseudomonas aeruginosa/fisiologia , Fenômenos Biomecânicos , Hidrodinâmica , Técnicas Analíticas Microfluídicas , Movimento/fisiologia , Propriedades de Superfície
10.
PLoS One ; 15(6): e0228234, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32589639

RESUMO

A major issue in the surveillance of antimicrobial resistance (AMR) is "de-duplication" or removal of repeated isolates, for which there exist multiple methods. The World Health Organization (WHO) Global Antimicrobial Resistance Surveillance System (GLASS) requires de-duplication by selecting only the first isolate of a given bacterial species per patient per surveillance period per specimen type per age group, gender, and infection origin stratification. However, no study on the comparative application of this method has been reported. The objective of this study was to evaluate differences in data tabulation between the WHO GLASS and the Japan Nosocomial Infections Surveillance (JANIS) system, which counts both patients and isolates after removing repeated isolates of the same bacterial species isolated from a patient within 30 days, regardless of specimen type, but distinguishing isolates with change of antimicrobial resistance phenotype. All bacterial data, consisting of approximately 8 million samples from 1795 Japanese hospitals in 2017 were exported from the JANIS database, and were tabulated using either the de-duplication algorithm of GLASS, or JANIS. We compared the tabulated results of the total number of patients whose blood and urine cultures were taken and of the percentage of resistant isolates of Escherichia coli for each priority antibiotic. The number of patients per specimen type tabulated by the JANIS method was always smaller than that of GLASS. There was a small (< 3%) difference in the percentage of resistance of E. coli for any antibiotic between the two methods in both out- and inpatient settings and blood and urine isolates. The two tabulation methods did not show considerable differences in terms of the tabulated percentages of resistance for E. coli. We further discuss how the use of GLASS tabulations to create a public software and website that could help to facilitate the understanding of and treatment against AMR.


Assuntos
Infecção Hospitalar/epidemiologia , Farmacorresistência Bacteriana , Organização Mundial da Saúde , Adolescente , Adulto , Sangue/microbiologia , Infecção Hospitalar/microbiologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/isolamento & purificação , Escherichia coli/fisiologia , Feminino , Humanos , Japão/epidemiologia , Masculino , Pessoa de Meia-Idade , Urina/microbiologia , Adulto Jovem
11.
PLoS One ; 15(6): e0234239, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32525884

RESUMO

Hummingbirds are essential pollinators in many ecosystems, making their conservation critical. As is the case with many species, hummingbirds are now facing a variety of challenges resulting from anthropogenic changes. As populations shift and species interactions change, disease is likely to pose a significant threat. There is a basic understanding of which pathogens currently affect a variety of hummingbird species, however there is a paucity of information about their immune systems capacity to kill pathogens and what specific factors may affect immunity. The objective of this study was to gain a basic understanding of the effect of age, sex, and molt on the constitutive innate immunity of hummingbirds. An in vitro assay was used to assess the microbiocidal capacity of the whole blood of Anna's Hummingbirds (Calypte anna) against three different microbes: Escherichia coli (E. coli), Staphylococcus aureus (S. aureus) and Candida albicans (C. albicans). The effect of age, sex and molt on anti-microbial capacity varied based on the microbe type. After-hatch-year birds tended to have better anti-microbial capacity compared to hatch-year birds. Male birds had higher anti-microbial activity than female birds, although this was not observed against C. albicans. Molting birds had a weaker antimicrobial activity against E. coli and S. aureus than birds that were not molting. These results represent an important first step towards defining the parameters of constitutive innate immunity of Anna's Hummingbirds as well as providing important knowledge about factors that should be considered when evaluating the health of wild populations.


Assuntos
Aves/sangue , Plasma/metabolismo , Animais , Aves/crescimento & desenvolvimento , Candida albicans/fisiologia , Escherichia coli/fisiologia , Feminino , Masculino , Viabilidade Microbiana , Muda , Caracteres Sexuais , Staphylococcus aureus/fisiologia
12.
Proc Natl Acad Sci U S A ; 117(27): 15609-15619, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32571953

RESUMO

Ribosome biogenesis is a complex process, and dozens of factors are required to facilitate and regulate the subunit assembly in bacteria. The 2'-O-methylation of U2552 in 23S rRNA by methyltransferase RrmJ is a crucial step in late-stage assembly of the 50S subunit. Its absence results in severe growth defect and marked accumulation of pre50S assembly intermediates. In the present work, we employed cryoelectron microscopy to characterize a set of late-stage pre50S particles isolated from an Escherichia coli ΔrrmJ strain. These assembly intermediates (solved at 3.2 to 3.8 Å resolution) define a collection of late-stage particles on a progressive assembly pathway. Apart from the absence of L16, L35, and L36, major structural differences between these intermediates and the mature 50S subunit are clustered near the peptidyl transferase center, such as H38, H68-71, and H89-93. In addition, the ribosomal A-loop of the mature 50S subunit from ΔrrmJ strain displays large local flexibility on nucleotides next to unmethylated U2552. Fast kinetics-based biochemical assays demonstrate that the ΔrrmJ 50S subunit is only 50% active and two times slower than the WT 50S subunit in rapid subunit association. While the ΔrrmJ 70S ribosomes show no defect in peptide bond formation, peptide release, and ribosome recycling, they translocate with 20% slower rate than the WT ribosomes in each round of elongation. These defects amplify during synthesis of the full-length proteins and cause overall defect in protein synthesis. In conclusion, our data reveal the molecular roles of U2552 methylation in both ribosome biogenesis and protein translation.


Assuntos
Escherichia coli/fisiologia , Elongação Traducional da Cadeia Peptídica , Iniciação Traducional da Cadeia Peptídica , RNA Ribossômico 23S/metabolismo , Subunidades Ribossômicas Maiores de Bactérias/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Microscopia Crioeletrônica , Técnicas de Inativação de Genes , Metilação , Metiltransferases/genética , Metiltransferases/metabolismo , Modelos Moleculares , Subunidades Ribossômicas Maiores de Bactérias/genética , Subunidades Ribossômicas Maiores de Bactérias/ultraestrutura , Uridina/metabolismo
13.
PLoS One ; 15(6): e0235059, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574199

RESUMO

BACKGROUND: To support effective antibiotic selection in empirical treatments, infection control interventions, and antimicrobial resistance containment strategies, many medical institutions collect antimicrobial susceptibility test data conducted at their facilities to prepare cumulative antibiograms. AIM: To evaluate how the setpoints of duplicate isolate removal period and data collection period affect the calculated susceptibility rates in antibiograms. METHODS: The Sakai City Medical Center is a regional core hospital for tertiary emergency medical care with 480 beds for general clinical care. In this study, all the Pseudomonas aeruginosa, Escherichia coli, and Klebsiella pneumoniae isolates collected at the Sakai City Medical Center Clinical Laboratory between July 2013 and December 2018 were subjected to antimicrobial susceptibility tests and the resulting data was analyzed. FINDINGS: The longer the duplicate isolate removal period, the fewer the isolates are available for every bacterial species. Differences in the length of the duplicate isolate removal period affected P. aeruginosa susceptibility rates to ß-lactam antibiotics by up to 10.8%. The setpoint of the data collection period affected the antimicrobial susceptibility rates by up to 7.3%. We found that a significant change in susceptibility could be missed depending on the setting of the data collection period, in preparing antibiogram of ß-lactam antibiotics for P. aeruginosa. CONCLUSIONS: When referring to antibiograms, medical professionals involved in infectious disease treatment should be aware that the parameter values, such as the duplicate isolate removal period and the data collection period, affect P. aeruginosa susceptibility rates especially to ß-lactam antibiotics. And antibiogram should be updated within the shortest time period that is practically possible, taking into account restrictions such as numbers of specimen.


Assuntos
Antibacterianos/farmacologia , Escherichia coli/efeitos dos fármacos , Klebsiella pneumoniae/efeitos dos fármacos , Testes de Sensibilidade Microbiana/métodos , Testes de Sensibilidade Microbiana/normas , Pseudomonas aeruginosa/efeitos dos fármacos , Algoritmos , Serviço Hospitalar de Emergência , Escherichia coli/isolamento & purificação , Escherichia coli/fisiologia , Hospitalização/estatística & dados numéricos , Humanos , Klebsiella pneumoniae/isolamento & purificação , Klebsiella pneumoniae/fisiologia , Pseudomonas aeruginosa/isolamento & purificação , Pseudomonas aeruginosa/fisiologia , Centros de Atenção Terciária , Fatores de Tempo
14.
Proc Biol Sci ; 287(1926): 20200569, 2020 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-32370668

RESUMO

Gene expression and growth rate are highly stochastic in Escherichia coli. Some of the growth rate variations result from the deterministic and asymmetric partitioning of damage by the mother to its daughters. One daughter, denoted the old daughter, receives more damage, grows more slowly and ages. To determine if expressed gene products are also allocated asymmetrically, we compared the levels of expressed green fluorescence protein in growing daughters descending from the same mother. Our results show that old daughters were less fluorescent than new daughters. Moreover, old mothers, which were born as old daughters, produced daughters that were more asymmetric when compared to new mothers. Thus, variation in gene products in a clonal E. coli population also has a deterministic component. Because fluorescence levels and growth rates were positively correlated, the aging of old daughters appears to result from both the presence of both more damage and fewer expressed gene products.


Assuntos
Escherichia coli/fisiologia , RNA , Escherichia coli/genética , Saccharomyces cerevisiae
15.
Nat Commun ; 11(1): 2097, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350281

RESUMO

Astroviruses are a global cause of pediatric diarrhea, but they are largely understudied, and it is unclear how and where they replicate in the gut. Using an in vivo model, here we report that murine astrovirus preferentially infects actively secreting small intestinal goblet cells, specialized epithelial cells that maintain the mucus barrier. Consequently, virus infection alters mucus production, leading to an increase in mucus-associated bacteria and resistance to enteropathogenic E. coli colonization. These studies establish the main target cell type and region of the gut for productive murine astrovirus infection. They further define a mechanism by which an enteric virus can regulate the mucus barrier, induce functional changes to commensal microbial communities, and alter host susceptibility to pathogenic bacteria.


Assuntos
Infecções por Astroviridae/patologia , Infecções por Astroviridae/virologia , Astroviridae/fisiologia , Trato Gastrointestinal/patologia , Trato Gastrointestinal/virologia , Células Caliciformes/virologia , Muco/virologia , Animais , Células Epiteliais/patologia , Células Epiteliais/virologia , Escherichia coli/fisiologia , Feminino , Trato Gastrointestinal/microbiologia , Trato Gastrointestinal/ultraestrutura , Masculino , Camundongos Endogâmicos C57BL , Muco/microbiologia , Transcriptoma/genética , Replicação Viral/fisiologia , Eliminação de Partículas Virais/fisiologia
16.
Proc Natl Acad Sci U S A ; 117(21): 11597-11607, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32385156

RESUMO

The distribution of fitness effects of mutation plays a central role in constraining protein evolution. The underlying mechanisms by which mutations lead to fitness effects are typically attributed to changes in protein specific activity or abundance. Here, we reveal the importance of a mutation's collateral fitness effects, which we define as effects that do not derive from changes in the protein's ability to perform its physiological function. We comprehensively measured the collateral fitness effects of missense mutations in the Escherichia coli TEM-1 ß-lactamase antibiotic resistance gene using growth competition experiments in the absence of antibiotic. At least 42% of missense mutations in TEM-1 were deleterious, indicating that for some proteins collateral fitness effects occur as frequently as effects on protein activity and abundance. Deleterious mutations caused improper posttranslational processing, incorrect disulfide-bond formation, protein aggregation, changes in gene expression, and pleiotropic effects on cell phenotype. Deleterious collateral fitness effects occurred more frequently in TEM-1 than deleterious effects on antibiotic resistance in environments with low concentrations of the antibiotic. The surprising prevalence of deleterious collateral fitness effects suggests they may play a role in constraining protein evolution, particularly for highly expressed proteins, for proteins under intermittent selection for their physiological function, and for proteins whose contribution to fitness is buffered against deleterious effects on protein activity and protein abundance.


Assuntos
Evolução Molecular , Aptidão Genética/genética , Mutação de Sentido Incorreto/genética , Mutação de Sentido Incorreto/fisiologia , Escherichia coli/enzimologia , Escherichia coli/genética , Escherichia coli/fisiologia , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , beta-Lactamases/química , beta-Lactamases/genética , beta-Lactamases/metabolismo
17.
Nat Commun ; 11(1): 2418, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32415107

RESUMO

The spatial organization of microbial communities arises from a complex interplay of biotic and abiotic interactions, and is a major determinant of ecosystem functions. Here we design a microfluidic platform to investigate how the spatial arrangement of microbes impacts gene expression and growth. We elucidate key biochemical parameters that dictate the mapping between spatial positioning and gene expression patterns. We show that distance can establish a low-pass filter to periodic inputs and can enhance the fidelity of information processing. Positive and negative feedback can play disparate roles in the synchronization and robustness of a genetic oscillator distributed between two strains to spatial separation. Quantification of growth and metabolite release in an amino-acid auxotroph community demonstrates that the interaction network and stability of the community are highly sensitive to temporal perturbations and spatial arrangements. In sum, our microfluidic platform can quantify spatiotemporal parameters influencing diffusion-mediated interactions in microbial consortia.


Assuntos
Dispositivos Lab-On-A-Chip , Consórcios Microbianos , Transdução de Sinais , Ecologia , Ecossistema , Desenho de Equipamento , Escherichia coli/fisiologia , Microbioma Gastrointestinal , Regulação Bacteriana da Expressão Gênica , Microfluídica/instrumentação , Modelos Genéticos , Oscilometria , Percepção de Quorum
18.
Proc Natl Acad Sci U S A ; 117(21): 11207-11216, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32424102

RESUMO

Bacteria frequently encounter selection by both antibiotics and lytic bacteriophages. However, the evolutionary interactions between antibiotics and phages remain unclear, in particular, whether and when phages can drive evolutionary trade-offs with antibiotic resistance. Here, we describe Escherichia coli phage U136B, showing it relies on two host factors involved in different antibiotic resistance mechanisms: 1) the efflux pump protein TolC and 2) the structural barrier molecule lipopolysaccharide (LPS). Since TolC and LPS contribute to antibiotic resistance, phage U136B should select for their loss or modification, thereby driving a trade-off between phage resistance and either of the antibiotic resistance mechanisms. To test this hypothesis, we used fluctuation experiments and experimental evolution to obtain phage-resistant mutants. Using these mutants, we compared the accessibility of specific mutations (revealed in the fluctuation experiments) to their actual success during ecological competition and coevolution (revealed in the evolution experiments). Both tolC and LPS-related mutants arise readily during fluctuation assays, with tolC mutations becoming more common during the evolution experiments. In support of the trade-off hypothesis, phage resistance via tolC mutations occurs with a corresponding reduction in antibiotic resistance in many cases. However, contrary to the hypothesis, some phage resistance mutations pleiotropically confer increased antibiotic resistance. We discuss the molecular mechanisms underlying this surprising pleiotropic result, consideration for applied phage biology, and the importance of ecology in evolution of phage resistance. We envision that phages may be useful for the reversal of antibiotic resistance, but such applications will need to account for unexpected pleiotropy and evolutionary context.


Assuntos
Colífagos/fisiologia , Farmacorresistência Bacteriana/fisiologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Pleiotropia Genética , Antibacterianos/farmacologia , Proteínas da Membrana Bacteriana Externa/genética , Escherichia coli/virologia , Proteínas de Escherichia coli/genética , Biblioteca Gênica , Genes Bacterianos , Especificidade de Hospedeiro , Lipopolissacarídeos/genética , Lipopolissacarídeos/metabolismo , Proteínas de Membrana Transportadoras/genética , Mutação
19.
Nat Commun ; 11(1): 2340, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32393772

RESUMO

Living organisms often display adaptive strategies that allow them to move efficiently even in strong confinement. With one single degree of freedom, the angle of a rotating bundle of flagella, bacteria provide one of the simplest examples of locomotion in the living world. Here we show that a purely physical mechanism, depending on a hydrodynamic stability condition, is responsible for a confinement induced transition between two swimming states in E. coli. While in large channels bacteria always crash onto confining walls, when the cross section falls below a threshold, they leave the walls to move swiftly on a stable swimming trajectory along the channel axis. We investigate this phenomenon for individual cells that are guided through a sequence of micro-fabricated tunnels of decreasing cross section. Our results challenge current theoretical predictions and suggest effective design principles for microrobots by showing that motility based on helical propellers provides a robust swimming strategy for exploring narrow spaces.


Assuntos
Escherichia coli/fisiologia , Fenômenos Biomecânicos , Flagelos/fisiologia , Movimento , Fótons , Polimerização , Fatores de Tempo
20.
J Vis Exp ; (159)2020 05 10.
Artigo em Inglês | MEDLINE | ID: mdl-32449734

RESUMO

Motility is crucial to the survival and success of many bacterial species. Many methodologies exist to exploit motility to understand signaling pathways, to elucidate the function and assembly of flagellar parts, and to examine and understand patterns of movement. Here we demonstrate a combination of three of these methodologies. Motility in soft agar is the oldest, offering a strong selection for isolating gain-of-function suppressor mutations in motility-impaired strains, where motility is restored through a second mutation. The cell-tethering technique, first employed to demonstrate the rotary nature of the flagellar motor, can be used to assess the impact of signaling effectors on the motor speed and its ability to switch rotational direction. The "border-crossing" assay is more recent, where swimming bacteria can be primed to transition into moving collectively as a swarm. In combination, these protocols represent a systematic and powerful approach to identifying components of the motility machinery, and to characterizing their role in different facets of swimming and swarming. They can be easily adapted to study motility in other bacterial species.


Assuntos
Escherichia coli/fisiologia , Flagelos/metabolismo , Técnicas Microbiológicas/métodos , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Genes Supressores , Movimento , Mutação/genética
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