Bacterial Growth Research Articles

Erythromycin disrupts Acinetobacter baumannii biofilms through destruction of the quorum sensing system.

This study was conducted to explore the effects of erythromycin on biofilms comprising Acinetobacter baumannii (A baumannii). To clarify the effect of erythromycin on the biofilms of A baumannii, we collected pure Ab strains isolated and identified from a variety of sample types extracted from patients in the microbiological laboratory of our hospital from April to August 2023, and divided them into an experimental group (treated with erythromycin) and a control group (without erythromycin). The morphology and quantity of A baumannii biofilm were observed at 24h, 48h, 72h, and 5d post-treatment, respectively, and the expression of quorum sensing (QS) system gene (abaI, abaR) mRNA was detected by fluorescence quantitative PCR. The results showed that A baumanniis are prone to form multiple drug-resistant (MDR) bacteria, against which the most commonly used clinical antibiotics are ineffective. Overall, we found that the number of bacteria, the number of bacteria in the biofilm, and the number of biofilms formed gradually increased over time, with a statistical difference (P < .05). After the addition of erythromycin, significant improvements in biofilm formation were achieved, indicating that erythromycin can destroy A baumannii biofilms, inhibiting bacterial growth to a certain extent. The expression levels of abaI and abaR gradually increased over time, indicating that the role of the QS system became more apparent over time. Biofilm formation is related to the QS system of A baumanniis. After erythromycin treatment, abaI and abaR mRNA expression was downregulated in the experimental group. Erythromycin disrupts A baumannii biofilms by destroying the quorum sensing system.

Calorie Restriction Decreases Competitive Fitness in Saccharomyces cerevisiae Following Heat Stress.

Experiments exposing Saccharomyces cerevisiae to glucose limitation (calorie restriction) are widely used to determine impacts on cell health as a model for aging. Using growth on plates and in liquid culture, we demonstrated that calorie restriction reduces fitness in subsequent nutrient-limited environments. Yeast grown in a calorie-restricted environment took longer to emerge from the lag phase, had an extended doubling time and had a lower percentage of culturability. Cells grown under moderate calorie restriction were able to withstand a gradual heat stress in a similar manner to cells grown without calorie restriction but fared less well with a sudden heat shock. Yeast grown under extreme calorie restriction were less fit when exposed to gradual heating or heat shock. Using RNAseq analysis, we provide novel insight into the mechanisms underlying this response, showing that in the absence of calorie restriction, genes whose products are involved in energy metabolism (glycolysis/gluconeogenesis and the citrate cycle) are predominantly overexpressed when yeasts were exposed to gradual heating, whereas this was not the case when they were exposed to shock. We show that both the culture history and the current environment must be considered when assaying physiological responses, and this has wider implications when developing strategies for the propagation, preservation or destruction of microbial cells.

Design and Evaluation of Microemulsion-Based Drug Delivery Systems for Biofilm-Based Infection in Burns

Normal skin is the first line of defense in the human body. A burn injury makes the skin susceptible to bacterial infection, thereby delaying wound healing and ultimately leading to sepsis. The chances of biofilm formation are high in burn wounds due to the presence of avascular necrotic tissue. The most common pathogen to cause burn infection and biofilm is Pseudomonas aeruginosa. The purpose of this study was to create a microemulsion (ME) formulation for topical application to treat bacterial burn infection. In the present study, tea tree oil was used as the oil phase, Tween 80 and transcutol were used as surfactants, and water served as the aqueous phase. Pseudo ternary phase diagrams were used to determine the design space. The ranges of components as suggested by the design were chosen, optimization of the microemulsion was performed, and in vitro drug release was assessed. Based on the characterization studies performed, it was found that the microemulsion were formulated properly, and the particle size obtained was within the desired microemulsion range of 10 to 300 nm. The I release study showed that the microemulsion followed an immediate release profile. The formulation was further tested based on its ability to inhibit biofilm formation and bacterial growth. The prepared microemulsion was capable of inhibiting biofilm formation.Graphical

Carvacrol and Thymol Enhance the Quality of Beni Arouss Buck Semen Stored at 4 °C Thanks to Their Antimicrobial Properties.

This study aims to investigate the impact of carvacrol and thymol on the quality of Beni Arouss buck semen stored in skim milk at 4 °C. Ejaculates were collected from eight Beni Arouss bucks weekly for 11 weeks, pooled, and then divided into three equal parts. Samples were diluted to 400 × 106 sperm/mL in skim milk (control) and skim milk supplemented with a single dose of 200 µM carvacrol and thymol each. Evaluations of sperm motility, viability, abnormalities, membrane integrity, lipid peroxidation, and bacterial growth were conducted at 0, 6, 24, and 48 h of liquid storage at 4 °C. After 48 h of storage, the results indicate that the addition of carvacrol positively influences total and progressive motility and viability. However, it also leads to a decrease in lipid peroxidation and bacterial growth compared to the control group (p < 0.05). Thymol showed similar results to carvacrol, except for progressive motility (p > 0.05). Bacterial growth was negatively correlated with total and progressive motility and viability (p < 0.05), while no correlation between lipid peroxidation and these parameters was observed (p > 0.05). In conclusion, the addition of carvacrol and thymol to skim milk extender moderately improves the quality of Beni Arouss buck semen after 48 h storage at 4 °C due to its antimicrobial activity.

Evaluation of the long-term protection conferred by an organosilicon-based disinfectant formulation against bacterial contamination of surfaces.

The aim of this work was to evaluate the efficacy of an organosilicon-based, commercially available antimicrobial formulation in the My-shield® product line against bacterial surface contamination. The antimicrobial product was tested in vitro for its long-term persistence on surfaces and effectiveness against Staphylococcus aureus biofilms in comparison to 70% ethanol and 0.1% or 0.6% sodium hypochlorite. Field testing was also conducted over 6 weeks at a university athletic facility. In vitro studies demonstrated the log reductions achieved by the test product, 70% ethanol, and 0.1% sodium hypochlorite were 3.6, 3.1, and 3.2, respectively. The test product persisted on surfaces after washing and scrubbing, and pre-treatment with this product prevented S. aureus surface colonization for up to 30 days. In comparison, pre-treatment with 70% ethanol or 0.6% sodium hypochlorite was not protective against S. aureus biofilm formation after seven days. The field test demonstrated that weekly applications of the test product were more effective at reducing surface bacterial load than daily applications of a control product. The test product conferred greater long-term protection against bacterial growth and biofilm formation by S. aureus than ethanol and sodium hypochlorite. Even with less frequent applications, the test product maintained a high level of antimicrobial activity.

The effect of epigallocatechin gallate on Streptococcus Gordonii biofilm formation

Background: Dental caries, as a primary disease in dentistry, is strongly influenced by the presence of biofilms. One of the Gram-positive bacteria that acts as an initiator in the biofilm formation process is Streptococcus gordonii. As the primary catechin in green tea, epigallocatechin gallate (EGCG) is easily found in our daily lives, and it has a broad spectrum of antimicrobial effects. Several studies have revealed that EGCG inhibited the growth of Gram-positive bacteria, including inhibiting biofilm formation by damaging the bacterial cell wall and reducing glucosyltransferase activity. However, there is still limited information that explains the effect of EGCG on S. gordonii bacterial biofilms. Purpose: This study aims to analyze the effect of EGCG in inhibiting the formation of S. gordonii bacterial biofilms. Methods: This study was an in-vitro experimental laboratory study, with samples divided into five groups, namely, the group containing BHIB-bacteria, the BHIB-bacteria-5% sucrose groups, and the treatment groups containing BHIB-bacteria-5% sucrose-EGCG with concentrations of 12.5%, 6.25%, and 3.125%, respectively, incubated for 24 hours. The data was analyzed using the Kruskal–Wallis test. Results: There was a significant difference in the formation of biofilms in S. gordonii bacteria with the addition of 5% sucrose in BHIB compared with the group of S. gordonii bacteria in BHIB. The highest biofilm formation in the group containing bacteria-BHIB-5% sucrose, while the lowest biofilm formation occurred in the treatment group containing bacteria-BHIB-5% sucrose-12.5% EGCG with significant difference between the group. Conclusion: The addition of EGCG 12.5% inhibits the formation of S. gordonii biofilms.

Gut Microbiome Changes among Undernutrition and Stunting Infants and Children under 2 Years: A Scoping Review

Background Undernutrition and stunting are significant public health concerns globally, particularly in low- and middle-income countries. Nutritional intervention is the cornerstone of the management and prevention of these conditions. However, the gut microbiome has recently emerged as an essential modulator of the effects of nutritional interventions in undernutrition and stunting. This scoping review aims to examine the impact of nutrition intake (including if there is any intervention) over time on gut microbiome changes in infants and children under the age of 2 who experience undernutrition and stunting. Methods A literature search was conducted in PubMed and Cochrane Library, including studies from 2013 to 2023, using terms related to malnutrition, stunting, failure to thrive, and gut microbiome. Inclusion criteria were applied to select eligible studies for review. Five studies were chosen to be included in this review. Results The findings indicate that nutrition intake over time, including dietary supplementation and prebiotics, can influence the gut microbiome composition, diversity, and functionality in undernutrition and stunting infants and children. These may promote the growth of beneficial bacteria while reducing the abundance of harmful pathogens. Moreover, improvements in nutritional status, growth parameters, and immune function were observed in association with positive changes in the gut microbiota. Conclusion Nutrition intake can positively modulate the gut microbiome in undernutrition and stunting infants by supporting the growth of beneficial gut bacteria such as Actinobacteria, Bacteroides, Streptococcus, Bifidobacterium, Prevotella sp, and other bacterial taxa that vary with age, ultimately contributing to enhanced growth and development outcomes. Further studies are needed to better understand the mechanisms underlying these effects and to develop targeted nutritional interventions that optimize the gut microbiome in undernutrition and stunting infants and children under 2 years.

Tip extension and simultaneous multiple fission in a filamentous bacterium

Organisms display an immense variety of shapes, sizes, and reproductive strategies. At microscopic scales, bacterial cell morphology and growth dynamics are adaptive traits that influence the spatial organization of microbial communities. In one such community-the human dental plaque biofilm-a network of filamentous Corynebacterium matruchotii cells forms the core of bacterial consortia known as hedgehogs, but the processes that generate these structures are unclear. Here, using live-cell time-lapse microscopy and fluorescent D-amino acids to track peptidoglycan biosynthesis, we report an extraordinary example of simultaneous multiple division within the domain Bacteria. We show that C. matruchotii cells elongate at one pole through tip extension, similar to the growth strategy of soil-dwelling Streptomyces bacteria. Filaments elongate rapidly, at rates more than five times greater than other closely related bacterial species. Following elongation, many septa form simultaneously, and each cell divides into 3 to 14 daughter cells, depending on the length of the mother filament. The daughter cells then nucleate outgrowth of new thinner vegetative filaments, generating the classic "whip handle" morphology of this taxon. Our results expand the known diversity of bacterial cell cycles and help explain how this filamentous bacterium can compete for space, access nutrients, and form important interspecies interactions within dental plaque.

Evaluation of Shunt-Related Ventriculitis in Infants

Aims: Shunt-related infections are resistant to antibiotics. This study examined patients aged under two years after ventriculitis. It is the first study on ventriculitis developed in a completely closed system (without skin defect infection, or puncture history), evaluating infants only. Method: The study included 13 subjects (four females) with ventriculitis who had undergone surgery for hydrocephalus during the neonatal period and were monitored for shunt infection, evaluating certain parameters. Results: Ventriculitis developed on an average of 2.8 months after surgery. The most common causes of hydrocephalus were spina bifida and intraventricular hemorrhage. Bacteria grown in culture included Staphylococcus epidermidis, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Serratia marcescens. Seven (53.8%) patients were treated without complications. Among other patients, three (23.1%) developed multilocular cystic hydrocephalus and three (23.1%) exhibited decreased cerebrospinal fluid production and did not require shunting. Conclusions: Patients receiving intraventricular therapy showed no complications, warranting further studies on early intraventricular therapy. In the presence of gram-negative bacterial growth, cerebrospinal fluid production may decrease and affected patients do not require shunts.

Next-generation sutures: Combining natural polymers and graphene for optimal wound healing

Sutures are essential for wound healing. Sutures cannot influence biological healing processes; hence their efficacy is still limited. Active healing sutures must be made. We created a biocompatible, biodegradable polymer suture material to reduce allergic reactions. Non-biocompatible materials that cause tissue rejection and inflammation impede wound healing. To improve wound healing, cellulose and chitosan (CS) were utilized to make a suture material. Dip coating with graphene oxide (GO) nanoparticles improved the suture material. The properties were assessed using FTIR, SEM, EDS, and other analytical methods. In an immersion investigation, coated suture material deterioration in simulated bodily fluid (SBF) was examined. When coated suture material inhibited bacterial growth, its antibacterial activity was tested. These properties suggest that GO nanoparticles, cellulose, and chitosan may be coupled for the first time to generate an aggressive mediator. Coated suture biocompatibility is assessed using in vitro cell survival and hemolysis tests. This paper reviews the latest suture material innovations and explores how natural polymers can advance this area.

UTILIZATION OF SOYBEAN POWDER AS AN ALTERNATIVE MEDIA FOR THE GROWTH OF ANAEROBIC BACTERIA

Background:Culture media or microbial growth media is a material consisting of a mixture of nutrients used by microorganisms for growth and reproduction. Anaerobic bacteria can thrive by requiring little or no oxygen for growth and metabolism. Purpose: The purpose to determine the potential of soybean powder as an alternative nitrogen source in Streptococcus mutans bacterial growth media and determine the optimum concentration of soybean powder in alternative media for Streptococcus mutans bacterial growth. Methods: The research method is a laboratory experiment to test soybean powder as an alternative medium for the growth of Streptococcus mutans bacteria using the Candle jar method. The population used is soybean powder with the brand soy flour, while the sample used is Streptococcus mutans bacteria. Results: The results obtained can be known that the average value of colony growth of Streptococcus mutans bacteria in alternative media of soybean powder with a concentration variation of 2 grams was 30 colonies, a concentration variation of 3 grams was 54 colonies, a concentration variation of 4 grams was 40 colonies, and a concentration variation of 5 grams was 72 colonies. From the average value, it can be seen that the growth of Streptococcus mutans on alternative media of soybean powder can grow well, but the most optimum growth is in the concentration variation of 5 grams. Conclusion:Based on the observations obtained, it shows that soybean powder (Glycine max (L.) Merr) can be used as an alternative medium for the growth of Streptococcus mutans bacteria.

Perception and Role of Indirect and Post-Discharge Surveillance in Predicting Surgical Site Infection in Surgical Patients: An Institutional Experience

Abstract Background: Surgical site infection (SSI) surveillance is a process of observing and reporting of infections following surgery. SSI surveillance studies are few in our country. Objectives: Present study was conducted to know the perception and role of SSI surveillance. Incidence of surgical site infection, identification of risk factors and bacterial spectrum of surgical wounds during surveillance were also studied. Methods: Patients who underwent a surgical procedure between July 2022 and June 2023 at our hospital were included. A prospective incidence surveillance for SSI was done to look for any signs of infection and record them in the surveillance form. Wound swabs or pus aspirates were sent for microbiological culture. Results: A total of 2312 patients underwent surgery during the study period, of whom 61 patients were diagnosed with SSI. The incidence of SSI was 2.6%. SSI was encountered more in female patients (57.3%). Age distribution had a significant association with SSI (P = 0.025). Most operative wounds were clean-contaminated (45.9%). Most SSIs were superficial (91.8%), followed by deep (8.2%). Most patients who developed SSI had a National Nosocomial Infection Surveillance (NNIS) patient risk index of 2 (45.9%). Bacterial growth was observed in 45.9% of cases. Multidrug resistance (MDR) among these isolates was 42.6%. Conclusion: This study helped to determine predictors of SSI like duration of surgery, type of wound, duration of hospital stays, and NNIS risk index. MDR isolates are now contributing to SSI. The perception of healthcare workers about SSI surveillance was that it was feasible to use and practice. Surveillance helped clinicians to strengthen the existing infection prevention and control practices for the future.

Microbial resistance and resilience to drought across a European climate gradient

Drought and rainfall events will become more frequent and intense with climate change. At the same time, soil moisture is one of the major factors controlling soil microbial processes such as carbon cycling. When challenged with drought there are two main growth responses microorganisms can use: (1) they can maintain growth rates during drought (i.e., resistance) and (2) they can recover growth rates faster when the drought ends (i.e., resilience). Microbial communities are shaped by multiple other factors in the soil environment, however how those impact drought responses remain unclear. Here we investigate how climate (estimated as aridity index) and soil properties determine microbial growth resistance and resilience to drought across a climate gradient in Europe. To test this, we exposed the different soils to a standardised drought cycle in controlled conditions. We assessed bacterial growth, fungal growth and respiration during soil drying to determine resistance and in high resolution during three days after rewetting to estimate resilience to drought. We found that alpha diversity was the strongest driver of both bacterial drought resistance and resilience, which occurred via changes in soil pH. This shows the importance of diversity for sustaining bacterial functions during drought stress. A secondary driver of bacterial drought resistance and resilience was the aridity index was also an important driver, where bacterial communities from more arid climates had higher resistance and resilience to drought. Fungal communities were both more resistant and resilient compared to bacteria, but this was independent of other measured environmental factors. Bacterial resilience was partly linked with differences in community composition. Our results suggest that if sites are exposed to increased aridity due to climate change or are managed to promote bacterial diversity, they will have higher bacterial growth rates during drought perturbations, which could potentially promote soil carbon storage.

Hypoglycemic peptide preparation from Bacillus subtilis fermented with Pyropia: Identification, molecular docking, and in vivo confirmation

Hypoglycemic foods have attracted increasing research interest. This study prepared a hypoglycemic product from Bacillus subtilis fermented with Pyropia (PBP), which has promising industrial potential, and elucidated its hypoglycemic mechanism. The aqueous PBP solution was orange, with protein as the main functional component. In vivo experiments demonstrated that PBP could increase insulin secretion and inhibit α-glucosidase activity, resulting in a hypoglycemic effect superior to that of acarbose at the same dose. Molecular docking revealed that the peptides APPVDID, GPPDSPY, PPSSPRP, and SPPPPPA from PBP could inhibit both α-glucosidase and dipeptidyl peptidase-IV (DPP-IV) activities. Pro residues promoted PBP peptide binding to the hydrophobic pocket S1 of DPP-IV. Additionally, PBP reduced inflammation and promoted the growth of beneficial gut bacteria (Prevotellaceae_UCG_003, Lachnospiraceae_UCG_001). This study presents a novel approach for the high-value utilization of Pyropia and a new option for the production of hypoglycemic functional foods and medicines.

Innovative Strategies in Drug Repurposing to Tackle Intracellular Bacterial Pathogens.

Intracellular bacterial pathogens pose significant public health challenges due to their ability to evade immune defenses and conventional antibiotics. Drug repurposing has recently been explored as a strategy to discover new therapeutic uses for established drugs to combat these infections. Utilizing high-throughput screening, bioinformatics, and systems biology, several existing drugs have been identified with potential efficacy against intracellular bacteria. For instance, neuroleptic agents like thioridazine and antipsychotic drugs such as chlorpromazine have shown effectiveness against Staphylococcus aureus and Listeria monocytogenes. Furthermore, anticancer drugs including tamoxifen and imatinib have been repurposed to induce autophagy and inhibit bacterial growth within host cells. Statins and anti-inflammatory drugs have also demonstrated the ability to enhance host immune responses against Mycobacterium tuberculosis. The review highlights the complex mechanisms these pathogens use to resist conventional treatments, showcases successful examples of drug repurposing, and discusses the methodologies used to identify and validate these drugs. Overall, drug repurposing offers a promising approach for developing new treatments for bacterial infections, addressing the urgent need for effective antimicrobial therapies.

Yersinia pseudotuberculosis growth arrest during type-III secretion system expression is associated with altered ribosomal protein expression and decreased gentamicin susceptibility.

Slow-growing bacterial cells have reduced antibiotic susceptibility, rendering them very difficult to eliminate during antibiotic treatment. However, for many key virulence factors (bacterial factors required to promote infection), it remains unclear whether expression is sufficient to slow bacterial growth and impact antibiotic susceptibility. Using Yersinia pseudotuberculosis , we found ribosomal protein expression fluctuated based on growth rate, and we generated a fluorescent reporter construct to detect altered ribosomal protein expression within individual bacterial cells. We then asked if expression of a key virulence factor in Yersinia , the type-III secretion system (T3SS), is sufficient to lower ribosomal protein expression, since it has been well established that T3SS induction results in growth arrest. We found high levels of T3SS expression promotes slowed growth and antibiotic tolerance, and bacterial cells that survive treatment with a ribosome-targeting antibiotic, gentamicin, have heightened levels of T3SS and lower levels of S10 expression.

Antimicrobial and antibiofilm effects of cyclic dipeptide-rich fraction from Lactobacillus plantarum loaded on graphene oxide nanosheets.

One effective method to combat bacterial infections is by using bacteria itself as a weapon. Lactobacillus is a type of fermenting bacterium that has probiotic properties and has demonstrated antimicrobial benefits against other bacteria. Cyclodipeptides (CDPs), present in the supernatant of Lactobacillus, possess several antimicrobial properties. In this study, the CDP fraction was isolated from the supernatant of Lactobacillus plantarum (L. plantarum). This fraction was then loaded onto graphene oxide nanosheets (GO NSs). The study assessed the substance's ability to inhibit bacterial growth by using the minimum inhibitory concentration (MIC) method on A. baumannii and S. aureus strains that were obtained from clinical samples. To determine the substance's impact on biofilm formation, the microtiter plate method was used. Moreover, the checkerboard technique was employed to explore the potential synergistic effects of these two substances. According to the study, the minimum inhibitory concentration (MIC) of the desired compound was found to be 1.25 mg/mL against S. aureus and 2.5 mg/mL against A. baumannii. Furthermore, at a concentration of 10 mg/mL, the compound prevented 81.6% (p < 0.01) of biofilm production in A. baumannii, while at a concentration of 1.25 mg/mL, it prevented 47.5% (p < 0.05) of biofilm production in S. aureus. The study also explored the synergistic properties of two compounds using the checkerboard method. In general, we found that GO NSs possess antimicrobial properties and enhance cyclodipeptides' activity against S. aureus and A. baumannii.

Antibacterial evaluation of garlic extracts on chitosan/starch packaging film using response surface methodology and its application for shelf-life extension of bell peppers (Capsicum annuum).

In this study, garlic extract (GE) was assessed as a potential additive in chitosan/starch (Ch/De) coatings, focusing on phenolic and flavonoid content analyses and antibacterial properties. Using response surface methodology approach, an optimization method was employed to achieve the optimal antibacterial formulation, with Ch, De, and GE identified as key variables in the Design of Experiment. Fourier transform infrared spectroscopy and X-ray diffraction analyses elucidated interactions among these primary components within the films, while thermogravimetric analysis confirmed the enhanced thermal stability of GE-coated film formulations (Ch/De/GE). The Ch/De/GE exhibited antibacterial efficacy against Escherichia coli (ATCC 25922) with an inhibition zone of 7.2mm at optimized concentrations of 2% w/v Ch, 1.5% w/v starch, and 0.5% v/v GE. In silico molecular docking studies provided insights into GE's inhibitory role as an antibacterial agent. Evaluation of green and yellow bell peppers (Capsicum annuum) over 18 days showed that coated peppers maintained better visual appearance and mass stability, with a weight loss decrease of 40.54%-48.96%, compared to uncoated ones. Additionally, the Ch/De/GE coating effectively inhibited bacterial growth, reducing it by 1-1.23 log CFU, during the storage period. In conclusion, the Ch/De/GE coating effectively extends the shelf-life of bell peppers and maintains their quality, demonstrating its potential for use in food packaging to preserve perishable items. PRACTICAL APPLICATION: The optimized chitosan/starch/garlic extract (Ch/De/GE) film developed in this study shows promising potential for application in the food packaging industry, particularly in extending the shelf life of perishable items like bell peppers. Its enhanced antibacterial properties, along with its ability to maintain visual appearance and reduce weight loss, make it an effective natural preservative that could replace synthetic additives in food packaging. By incorporating this biodegradable film into packaging solutions, producers can offer safer, more sustainable products that meet consumer demand for natural and environmentally friendly options.

Fluorescent protein-based Zn2+ sensors reveal distinct responses of aerobic and anaerobic Escherichia coli cultures to excess Zn2+

Zinc ions are required by all known organisms. Maintaining zinc homeostasis by preventing toxic overload while ensuring sufficient acquisition for cellular functions is crucial for survival and growth of bacteria. Bacteria, however, frequently encounter and must survive in various environments. During infection in host animals, for example, bacteria are exposed to acidic conditions in the stomach and anaerobic conditions in the intestines, but the effects of oxygen on zinc homeostasis in E. coli have not been well-studied. Previously, we reported a flavin-binding fluorescent protein-based zinc sensor, CreiLOVN41C, which can respond to changes in labile Zn2+ levels in bacteria under both aerobic and anaerobic conditions. Here, we combined the use of CreiLOVN41C with established oxygen-dependent fluorescent protein-based sensors, inductively coupled plasma-mass spectrometry (ICP-MS), and growth curves to evaluate how oxygen levels affect zinc uptake in E. coli. ICP-MS results showed that cells grown aerobically with added zinc acquired more zinc, but no additional zinc was accumulated when cells were grown anaerobically. Using oxygen-independent CreiLOVN41C and the oxygen-dependent ZapCY series of sensors, intracellular labile zinc was detected in E. coli grown with varied zinc under varied conditions. Although little to no endogenous zinc was detected by any sensor in E. coli cells grown with up to 2 mM added zinc, CreiLOVN41C revealed that when Zn2+ was added and detected by cells in real-time, anaerobic cells required more Zn2+ to similarly saturate the sensor. Overall, this work reveals that zinc uptake in E. coli is impacted by oxygen levels during cell growth.

Oxygen-dependent regulation of F-box proteins in Toxoplasma gondii is mediated by Skp1 glycosylation

A dynamic proteome is required for cellular adaption to changing environments including levels of O2, and the SKP1/CULLIN-1/F-box protein/RBX1 (SCF) family of E3 ubiquitin ligases contributes importantly to proteasome-mediated degradation. We examine, in the apicomplexan parasite Toxoplasma gondii, the influence on the interactome of SKP1 by its novel glycan attached to a hydroxyproline generated by PHYa, the likely ortholog of the HIFα PHD2 oxygen-sensor of human host cells. Strikingly, the representation of several putative F-box proteins (FBPs) is substantially reduced in PHYaΔ parasites grown in fibroblasts. One, FBXO13, is a predicted lysyl hydroxylase related to the human JmjD6 oncogene except for its F-box domain. The abundance of FBXO13, epitope-tagged at its genetic locus, was reduced in PHYaΔ parasites thus explaining its diminished presence in the SKP1 interactome. A similar effect was observed for FBXO14, a cytoplasmic protein of unknown function that may have co-evolved with PHYa in apicomplexans. Similar findings in glycosylation-mutant cells, rescue by proteasomal inhibitors, and unchanged transcript levels, suggested the involvement of the SCF in their degradation. The effect was selective, because FBXO1 was not affected by loss of PHYa. These findings are physiologically significant because the effects were phenocopied in parasites reared at 0.5% O2. Modest impact on steady-state SKP1 modification levels suggests that effects are mediated during a lag phase in hydroxylation of nascent SKP1. The dependence of FBP abundance on O2-dependent SKP1 modification likely contributes to the reduced virulence of PHYaΔ parasites owing to impaired ability to sense O2 as an environmental signal.