Microbial communities display alternative stable states in a fluctuating environment.

The effect of environmental fluctuations is a major question in ecology. While it is widely accepted that fluctuations and other types of disturbances can increase biodiversity, there are fewer examples of other types of outcomes in a fluctuating environment. Here we explore this question with laboratory microcosms, using cocultures of two bacterial species, P. putida and P. veronii. At low dilution rates we observe competitive exclusion of P. veronii, whereas at high dilution rates we observe competitive exclusion of P. putida. When the dilution rate alternates between high and low, we do not observe coexistence between the species, but rather alternative stable states, in which only one species survives and initial species' fractions determine the identity of the surviving species. The Lotka-Volterra model with a fluctuating mortality rate predicts that this outcome is independent of the timing of the fluctuations, and that the time-averaged mortality would also lead to alternative stable states, a prediction that we confirm experimentally. Other pairs of species can coexist in a fluctuating environment, and again consistent with the model we observe coexistence in the time-averaged dilution rate. We find a similar time-averaging result holds in a three-species community, highlighting that simple linear models can in some cases provide powerful insight into how communities will respond to environmental fluctuations.

Mortality causes universal changes in microbial community composition.

All organisms are sensitive to the abiotic environment, and a deteriorating environment can cause extinction. However, survival in a multispecies community depends upon interactions, and some species may even be favored by a harsh environment that impairs others, leading to potentially surprising community transitions as environments deteriorate. Here we combine theory and laboratory microcosms to predict how simple microbial communities will change under added mortality, controlled by varying dilution. We find that in a two-species coculture, increasing mortality favors the faster grower, confirming a theoretical prediction. Furthermore, if the slower grower dominates under low mortality, the outcome can reverse as mortality increases. We find that this tradeoff between growth and competitive ability is prevalent at low dilution, causing outcomes to shift dramatically as dilution increases, and that these two-species shifts propagate to simple multispecies communities. Our results argue that a bottom-up approach can provide insight into how communities change under stress.

Anti-pathogenic, antibiofilm, and technological properties of fermented food associated Staphylococcus succinus strain AAS2.

The present investigation was aimed to determine the anti-pathogenic, antibiofilm, and technological properties of fermented food associated Staphylococcus succinus strain AAS2. The anti-pathogenic attribute of cell-free neutralized supernatant (CFNS) of strain AAS2 was assessed against food-borne and enteric pathogens that revealed promising activity against Staphylococcus aureus and Enterobacter aerogenes with high arbitrary unit of 220.25 ± 3.3 and 170.2 ± 4.6 AU/mL, respectively. Furthermore, the antibiofilm and time-kill assay of CFNS of strain AAS2 depicted remarkable reduction in biofilm formation of indicator pathogens in a dose-dependent manner. Moreover, time-kill assay data revealed the drastic reduction in the viability (log cfu/mL) of S. aureus and E. aerogenes in the presence of varied minimum inhibitory concentration ranges of CFNS. The distinct technological properties of strain AAS2 were demonstrated using standard methodologies. Reported results estimated moderate level of exopolysaccharide (41.3 ± 0.6 mg/L) and lipase production (8.3 ± 0.3 mm), followed by remarkable autolytic (30.1 ± 1.2-43.1 ± 1.3%), catalase (13.82 ± 0.3 AU), and nitrate reductase (10.25 ± 0.3 mM nitrite/mg dry weight) activities under standard conditions. Most importantly, the strain cleared the specific in vitro safety assessment tests. The described anti-pathogenic and technological traits of strain AAS2 paved the way to utilize it in pharmaceutical as well as food processing industries as starter/adjunct culture.

Emergence of IMP-4-Producing Enterobacter aerogenes Clinical Isolate.

BACKGROUND: IMP-4 class B metallo-ß-lactamase-producing Enterobacteriaceae are resistant to carbapenems. The aim of this study was to characterize of IMP-4 metallo-ß-lactamase (MBL)-producing Enterobacter aerogenes clinical isolate. METHODS: IMP-4 MBL-producing E. aerogenes clinical isolate was collected from a Korean Hospital in 2017. Antimicrobial susceptibility was determined by disk diffusion methods. Further, minimum inhibitory concentrations of ß-lactams were determined by Etest. Detection of bla genes was performed by PCR. The genetic organization of class 1 integron carrying the MBL gene cassette was investigated by PCR mapping and sequencing. RESULTS: E. aerogenes strain YN170501 exhibited resistance to penicillins, cephalosporins, and carbapenems and was susceptible to monobactam, aminoglycosides, fluoroquinolone, tigecycline, and trimethoprim-sulfamethoxazole. The blaIMP-4 gene was located in class 1 integron. CONCLUSIONS: The blaIMP-4 gene has never been reported in Enterobacter aerogenes clinical isolate from Korea.

Enterobacter aerogenes metabolites enhance Microcystis aeruginosa biomass recovery for sustainable bioflocculant and biohydrogen production.

We report a recycling bioresource involving harvesting of Microcystis aeruginosa using the bioflocculant (MBF-32) produced by Enterobacter aerogenes followed by the recovery of the harvested M. aeruginosa as the main substrate for the sustainable production of MBF-32 and biohydrogen. The experimental results indicate that the efficiency of bioflocculation exceeded 90% under optimal conditions. The harvested M. aeruginosa was further recycled as the main substrate for the supply of necessary elements. The highest yield (3.6±0.1g/L) of MBF-32 could be obtained from 20g/L of wet biomass of M. aeruginosa with an additional 20g/L of glucose as the extra carbon source. The highest yield of biohydrogen was 35mL of H2/g (dw) algal biomass, obtained from 20g/L of wet biomass of M. aeruginosa with an additional 10g/L of glycerol. Transcriptome analyses indicated that MBF-32 was mainly composed of polysaccharide and tyrosine/tryptophan proteins. Furthermore, NADH synthase and polysaccharide export-related genes were found to be up-regulated.

Donor-derived infections among Chinese donation after cardiac death liver recipients.

AIM: To investigate blood cultures of deceased donors and report the confirmed transmission of bacterial infection from donors to liver recipients. METHODS: We retrospectively studied the results of blood cultures among our donation after cardiac death (DCD) donors and calculated the donor-derived bacterial infection rates among liver recipients. Study participants underwent liver transplantation between January 1, 2010 and February 1, 2017. The study involved a total of 67 recipients of liver grafts from 67 DCD donors. We extracted the data of donors' and patients' characteristics, culture results and clinical outcomes, especially the post-transplant complications in liver recipients, from electronic medical records. We analyzed the characteristics of the donors and the corresponding liver recipients with emphasis put on donor-derived infections. RESULTS: Head trauma was the most common origin of death among our 67 DCD donors (46.3%). Blood taken prior to the procurement operation was cultured for 53 of the donors, with 17 episodes of bloodstream infections developing from 13 donors. The predominant organism isolated from the blood of donors was Gram-positive bacteria (70.6%). Only three (4.5%) of 67 liver recipients developed confirmed donor-derived bacterial infections, with two isolates of multidrug-resistant Klebsiella pneumoniae and one isolate of multidrug-resistant Enterobacter aerogenes. The liver recipients with donor-derived infections showed relation to higher crude mortality and graft loss rates (33.3% each) within 3 mo post transplantation, as compared to those without donor-derived infections (9.4% and 4.7%, respectively). All three liver recipients received appropriate antimicrobial therapy. CONCLUSION: Liver recipients have high occurrence of donor-derived infections. The liver recipients with donor-derived multidrug-resistant Enterobacteriaceae infections can have good outcome if appropriate antimicrobial therapy is given.

Physiological and molecular characteristics of carbapenem resistance in Klebsiella pneumoniae and Enterobacter aerogenes.

INTRODUCTION: Bacterial resistance is a growing concern in the nosocomial environment in which Klebsiella pneumoniae and Enterobacter aerogenes play an important role due to their opportunism and carbapenemase-production. This work aimed to evaluate physiological and molecular characteristics of carbapenem-resistant K. pneumoniae and E. aerogenes isolated in a Brazilian tertiary hospital. METHODOLOGY: In total, 42 carbapenem-resistant bacteria isolated from clinical specimens were included (21 K. pneumoniae and 21 E. aerogenes). Drug-sensitive K. pneumoniae (n = 27) were also included. Antimicrobial susceptibility and biocide tolerance patterns, hemolytic activity, tolerance to oxidative stress, and aggregative ability were assessed. Genetic markers related to carbapenem resistance, or ESBL-production were screened by PCR. RESULTS: Compared to drug-sensitive strains, carbapenem-resistant K. pneumoniae were more tolerant to biocides and to oxidative stress, and they displayed an increase in biofilm formation. The genetic markers blaKPC (95.2%) and blaTEM (90.5%) were the most frequent. Among the carbapenem-resistant E. aerogenes strains, blaKPC, and blaTEM were detected in all bacteria. Drug-sensitive E. aerogenes were not isolated in the same period. blaSHV, blaVIM, and blaCTX markers were also observed among carbapenem-resistant bacteria. CONCLUSIONS: Results suggest that carbapenemase-producing enterobacteria might show peculiar characteristics regarding their physiology associated with their environmental persistency, virulence, and multidrug resistance. The observed phenomenon may have implications not only for antimicrobial chemotherapy, but also for the prognosis of infectious diseases and infection control.

The melatonin-sensitive circadian clock of the enteric bacterium Enterobacter aerogenes.

Circadian clocks are fundamental properties of all eukaryotic organisms and at least some prokaryotic organisms. Recent studies in our laboratory have shown that the gastrointestinal system contains a circadian clock that controls many, if not all, aspects of gastrointestinal function. We now report that at least one species of intestinal bacteria, Enterobacter aerogenes, responds to the pineal and gastrointestinal hormone melatonin by an increase in swarming activity. This swarming behavior is expressed rhythmically, with a period of approximately 24 hrs. Transformation of E. aerogenes to express luciferase with a MotA promoter reveals circadian patterns of bioluminescence that are synchronized by melatonin and whose periods are temperature compensated from 26°C to 40°C. Bioinformatics suggest similarities between the E. aerogenes and cyanobacterial clocks, suggesting the circadian clock may have evolved very early in the evolution of life. They also point to a coordination of host circadian clocks with those residing in the microbiota themselves.

Trapping and viability of swimming bacteria in an optoelectric trap.

Non-contact manipulation methods capable of trapping and transporting swimming bacteria can significantly aid in chemotaxis studies. However, high swimming speed makes the trapping of these organisms an inherently challenging task. We demonstrate that an optoelectric technique, rapid electrokinetic patterning (REP), can effectively trap and manipulate Enterobacter aerogenes bacteria swimming at velocities greater than 20 µm s(-1). REP uses electro-orientation, laser-induced AC electrothermal flow, and particle-electrode interactions for capturing these cells. In contrast to trapping non-swimming bacteria and inert microspheres, we observe that electro-orientation is critical to the trapping of the swimming cells, since unaligned bacteria can swim faster than the radially inward electrothermal flow and escape the trap. By assessing the cell membrane integrity, we study the effect of REP trapping conditions, including optical radiation, laser-induced heating, and the electric field on cell viability. When applied individually, the optical radiation and laser-induced heating have negligible effect on cells. At the standard REP trapping conditions fewer than 2% of cells have a compromised membrane after four minutes. To our knowledge this is the first study detailing the effect of REP trapping on cell viability. The presented results provide a clear guideline on selecting suitable REP parameters for trapping living bacteria.

Human Gut Bacteria Are Sensitive to Melatonin and Express Endogenous Circadian Rhythmicity.

Circadian rhythms are fundamental properties of most eukaryotes, but evidence of biological clocks that drive these rhythms in prokaryotes has been restricted to Cyanobacteria. In vertebrates, the gastrointestinal system expresses circadian patterns of gene expression, motility and secretion in vivo and in vitro, and recent studies suggest that the enteric microbiome is regulated by the host's circadian clock. However, it is not clear how the host's clock regulates the microbiome. Here, we demonstrate at least one species of commensal bacterium from the human gastrointestinal system, Enterobacter aerogenes, is sensitive to the neurohormone melatonin, which is secreted into the gastrointestinal lumen, and expresses circadian patterns of swarming and motility. Melatonin specifically increases the magnitude of swarming in cultures of E. aerogenes, but not in Escherichia coli or Klebsiella pneumoniae. The swarming appears to occur daily, and transformation of E. aerogenes with a flagellar motor-protein driven lux plasmid confirms a temperature-compensated circadian rhythm of luciferase activity, which is synchronized in the presence of melatonin. Altogether, these data demonstrate a circadian clock in a non-cyanobacterial prokaryote and suggest the human circadian system may regulate its microbiome through the entrainment of bacterial clocks.

Insights into the global regulation of anaerobic metabolism for improved biohydrogen production.

To improve the biohydrogen yield in bacterial dark fermentation, a new approach of global anaerobic regulation was introduced. Two cellular global regulators FNR and NarP were overexpressed in two model organisms: facultatively anaerobic Enterobacter aerogenes (Ea) and strictly anaerobic Clostridium paraputrificum (Cp). The overexpression of FNR and NarP greatly altered anaerobic metabolism and increased the hydrogen yield by 40%. Metabolic analysis showed that the global regulation caused more reducing environment inside the cell. To get a thorough understanding of the global metabolic regulation, more genes (fdhF, fhlA, ppk, Cb-fdh1, and Sc-fdh1) were overexpressed in different Ea and Cp mutants. For the first time, it demonstrated that there were approximately linear relationships between the relative change of hydrogen yield and the relative change of NADH yield or ATP yield. It implied that cellular reducing power and energy level played vital roles in the biohydrogen production.

Inactivation of Enterobacter aerogenes in reconstituted skim milk by high- and low-frequency ultrasound.

The inactivation of Enterobacter aerogenes in skim milk using low-frequency (20kHz) and high-frequency (850kHz) ultrasonication was investigated. It was found that low-frequency acoustic cavitation resulted in lethal damage to E. aerogenes. The bacteria were more sensitive to ultrasound in water than in reconstituted skim milk having different protein concentrations. However, high-frequency ultrasound was not able to inactivate E. aerogenes in milk even when powers as high as 50W for 60min were used. This study also showed that high-frequency ultrasonication had no influence on the viscosity and particle size of skim milk, whereas low-frequency ultrasonication resulted in the decrease in viscosity and particle size of milk. The decrease in particle size is believed to be due to the breakup of the fat globules, and possibly to the cleavage of the κ-casein present at the surface of the casein micelles. Whey proteins were also found to be slightly affected by low-frequency ultrasound, with the amounts of α-lactalbumin and ß-lactoglobulin slightly decreasing.

Inactivation of microorganisms by low-frequency high-power ultrasound: 2. A simple model for the inactivation mechanism.

A simple theoretical model based on shear forces generated by the collapse of the ultrasound cavities near the surface of a microorganism is proposed. This model requires two parameters which take into account the number of acoustic cavitation bubbles, and the resistance of the cell wall of the microorganism to the shear forces generated by bubble collapse. To validate the model, high-power low frequency (20 kHz) ultrasound was used to inactivate two microorganisms with very different sizes, viz., a bacterium, Enterobacter aerogenes and a yeast, Aureobasidium pullulans. The inactivation ratio was experimentally measured as a function of sonication time for different ultrasound power and for different initial cell numbers. For both E. aerogenes and A. pullulans the Log of the inactivation ratio decreased linearly with sonication time, and the rate of inactivation increased (D-value decreased) with the increase in sonication power. The rate of inactivation was also found, for both microorganisms, to increase with a decrease in the initial cell number. The fits, obtained using the proposed model, are in very good agreement with the experimental data.

Acanthamoeba castellanii of the T4 genotype is a potential environmental host for Enterobacter aerogenes and Aeromonas hydrophila.

BACKGROUND: Acanthamoeba can interact with a wide range of microorganisms such as viruses, algae, yeasts, protists and bacteria including Legionella pneumophila, Pseudomonas aeruginosa, Vibrio cholerae, Helicobacter pylori, Listeria monocytogenes, Mycobacterium spp., and Escherichia coli. In this capacity, Acanthamoeba has been suggested as a vector in the transmission of bacterial pathogens to the susceptible hosts. METHODS: Here, we used a keratitis isolate of A. castellanii of the T4 genotype and studied its interactions with two bacterial genera which have not been tested before, Enterobacter aerogenes, and Aeromonas hydrophila, as well as E. coli. Assays were performed to determine bacterial association with and invasion of A. castellanii. Additionally, bacterial survival intracellular of A. castellanii trophozoites as well as cysts was determined. RESULTS: All three bacterial isolates tested, associated, invaded, and survived inside A. castellanii trophozoites as well as A. castellanii cysts. However, E. aerogenes and E. coli exhibited significantly reduced association with and invasion of A. castellanii as compared with A. hydrophila (P < 0.01 using paired T-test, one tail distribution). In the long term survival assays, all three bacterial isolates tested remained viable inside A. castellanii trophozoites, while amoeba remained intact; however A. hydrophila exhibited higher survival inside amoebae (14.54 ± 3.3 bacteria:amoeba ratio) compared with E. aerogenes (3.96 ± 0.7 bacteria:amoeba ratio) and E. coli (5.85 ± 1.1 bacteria:amoeba ratio). A. hydrophila, E. coli, and E. aerogenes remained viable during the encystment process and exhibited higher levels of recovery from mature cysts (14.13 ± 0.89 A. hydrophila:amoeba ratio, 10.13 ± 1.17 E. aerogenes:amoeba ratio, and 11.95 ± 0.7 E. coli:amoeba ratio). CONCLUSIONS: A. hydrophila and E. aerogenes also joined the ranks of other bacteria that could benefit from A. castellanii. Because cysts can be airborne, these findings suggest that Acanthamoeba is a potential vector in the transmission of A. hydrophila and E. aerogenes to susceptible hosts.

Nonhomologous end-joining promotes resistance to DNA damage in the absence of an ADP-ribosyltransferase that signals DNA single strand breaks.

ADP-ribosylation of proteins at DNA lesions by ADP-ribosyltransferases (ARTs) is an early response to DNA damage. The best defined role of ADP-ribosylation in the DNA damage response is in repair of single strand breaks (SSBs). Recently, we initiated a study of how ADP-ribosylation regulates DNA repair in Dictyostelium and found that two ARTs (Adprt1b and Adprt2) are required for tolerance of cells to SSBs, and a third ART (Adprt1a) promotes nonhomologous end-joining (NHEJ). Here we report that disruption of adprt2 results in accumulation of DNA damage throughout the cell cycle following exposure to agents that induce base damage and DNA SSBs. Although ADP-ribosylation is evident in adprt2(-) cells exposed to methylmethanesulfonate (MMS), disruption of adprt1a and adprt2 in combination abolishes this response and further sensitises cells to this agent, indicating that in the absence of Adprt2, Adprt1a signals MMS-induced DNA lesions to promote resistance of cells to DNA damage. As a consequence of defective signalling of SSBs by Adprt2, Adprt1a is required to assemble NHEJ factors in chromatin, and disruption of the NHEJ pathway in combination with adprt2 increases sensitivity of cells to MMS. Taken together, these data indicate overlapping functions of different ARTs in signalling DNA damage, and illustrate a critical requirement for NHEJ in maintaining cell viability in the absence of an effective SSB response.

Hydrogen production from biodiesel byproduct by immobilized Enterobacter aerogenes.

The recent rapid growth of the biodiesel industry has generated a significant amount of glycerol as a byproduct. As a result, the price of glycerol is currently relatively low, making it an attractive starting material for the production of chemicals with higher values. Crude glycerol can be directly converted through microbial fermentation into various chemicals such as hydrogen. In this study, we optimized immobilization of a facultative hydrogen producing microorganism, Enterobacter aerogenes, with the goal of developing biocatalysts that was appropriate for the continuous hydrogen production from glycerol. Several carriers were tested and agar was found to be the most effective. In addition, it was clearly shown that variables such as the carrier content and cell loading should be controlled for the immobilization of biocatalysts with high hydrogen productivity, stability, and reusability. After optimization of these variables, we were able to obtain reusable biocatalysts that could directly convert the byproduct stream from biodiesel processes into hydrogen in continuous processes.

Deposition behavior of self-assembled monolayers and bacteria on metallic surfaces using an electrochemical quartz crystal nanobalance.

This paper presents an investigation on the deposition behavior of self-assembled monolayers (SAMs) and bacteria on titanium and gold surfaces using an electrochemical quartz crystal nanobalance (EQCN). The tests included alkanethiol and alkanesilane and three bacteria, namely: E. coli, P. fluorescens and K. aerogenes. The mass change with respect to immersion time was measured by EQCN. The results showed SAM formation on both titanium and gold surfaces, but SAM formation on gold was generally higher by 26-74% as compared to that of titanium. On the other hand, bacteria also adhered well on the two metal surfaces. E. coli and P. fluorescens had high affinity on gold and titanium surfaces, respectively, while K. aerogenes was more adherent to titanium. The results showed that titanium and gold are good metals for biomaterials yet at the same time, their bioinert property provide excellent condition for bacterial adhesion. Therefore, there is a need for proper surface preparation to optimize the use of titanium and gold as biomaterials.

Characterization of six antibacterial response genes from the European corn borer (Ostrinia nubilalis) larval gut and their expression in response to bacterial challenge.

Six cDNAs encoding putative antibacterial response proteins were identified and characterized from the larval gut of the European corn borer (Ostrinia nubilalis). These antibacterial response proteins include four peptidoglycan recognition proteins (PGRPs), one ß-1,3-glucanase-1 (ßglu-1), and one lysozyme. Tissue-specific expression analysis showed that these genes were highly expressed in the midgut, except for lysozyme. Analysis of expression of these genes in different developmental stage showed that they were expressed in larval stages, but little or no detectable expression was found in egg, pupa and adult. When larvae were challenged with Gram-negative bacteria (Enterobacter aerogenes), the expression of all six genes was up-regulated in the fatbodies. However, when larvae were challenged with Gram-positive bacteria (Micrococcus luteus), only PGRP-C and lysozyme genes were up-regulated. This study provides additional insights into the expression of antibacterial response genes in O. nubilalis larvae and helps us better understand the immune defense response in O. nubilalis.

Physiological characterisation of the efflux pump system of antibiotic-susceptible and multidrug-resistant Enterobacter aerogenes.

Enterobacter aerogenes predominates amongst Enterobacteriaceae species that are increasingly reported as producers of extended-spectrum beta-lactamases. Although this mechanism of resistance to beta-lactams is important, other mechanisms bestowing a multidrug-resistant (MDR) phenotype in this species are now well documented. Amongst these mechanisms is the overexpression of efflux pumps that extrude structurally unrelated antibiotics prior to their reaching their targets. Interestingly, although knowledge of the genetic background behind efflux pumps is rapidly advancing, few studies assess the physiological nature of the overall efflux pump system of this, or for that matter any other, bacterium. The study reported here evaluates physiologically the efflux pump system of an E. aerogenes ATCC reference as well as two strains whose MDR phenotypes are mediated by overexpressed efflux pumps. The activities of the efflux pumps in these strains are modulated by pH and glucose, although the effects of the latter are essentially restricted to pH 8, suggesting the presence of two general efflux pump systems, i.e. proton-motive force-dependent and ABC transporter types, respectively.

Sepsis resulting from Enterobacter aerogenes resistant to carbapenems after liver transplantation.

BACKGROUND: Sepsis due to Enterobacter aerogenes (E. aerogenes) is rare after liver transplantation but is also a serious infection that may cause liver abscess. The purpose of this case report is to relate an unusual presentation of liver transplantation to show how successive treatment can be an appropriate option in septic patients after liver transplantation. METHOD: We report on a patient with liver transplantation who developed sepsis due to extended spectrum beta-lactamases and AmpC-producing E. aerogenes. RESULTS: A 39-year-old man had a biliary fistula and then was found to have multiple liver abscesses through abdominal ultrasound and an abdominal computed tomography scan, and carbapenem-sensitive E. aerogenes infection was confirmed. The patient was not successfully treated with conservative treatment consisting of intravenous carbapenems, percutaneous transhepatic cholangial drainage, and biliary stent placement by endoscopic retrograde cholangiopancreatography, so a second liver transplantation followed. Carbapenem-resistant E. aerogenes was detected in bile and blood after a five-week course of carbapenem therapy. The patient developed septic shock and multiple organ dysfunction syndrome. CONCLUSIONS: We first report an unusual case of sepsis caused by E. aerogenes after liver transplantation in China. Carbapenem-resistant E. aerogenes finally leads to uncontrolled sepsis with current antibiotics. We hypothesize that the infection developed as a result of biliary fistula and predisposing immunosuppressive agent therapy. Further research is progressing on the aspect of immunomodulation therapy.