The bifunctional enzyme β-carotene hydroxylase (CrtZ) catalyzes the hydroxylation of carotenoid β-ionone rings at the 3, 3’ position regardless of the presence of keto group at 4, 4’ position, which is an important step in the synthesis of astaxanthin. The level and substrate preference of CrtZ may have great effect on the amount of astaxanthin and the accumulation of intermediates.crtZ from Paracoccus sp. PC1 and PAcrtZ from Pantoea Agglomerans were certified and were combined utilization for increase astaxanthin production. Firstly, PCcrtZ from Paracoccus sp. PC1 and PAcrtZ from P. Agglomerans were expressed in platform strains CAR032 (β-carotene producing strain) and Can004 (canthaxanthin producing strain) separately to identify their substrate preference for carotenoids with keto groups at 4,4’ position or not. The results showed that PCcrtZ led to a lower zeaxanthin yield in CAR032 compared to that of PAcrtZ. On the contrary, higher astaxanthin production was obtained in Can004 by PCcrtZ than that of PAcrtZ. This demonstrated that PCCrtZ has higher canthaxanthin to astaxanthin conversion ability than PACrtZ, while PACrtZ prefer using β-carotene as substrate. Finally, Ast010, which has two copies of PAcrtZ and one copy of PCcrtZ produced 1.82 g/L of astaxanthin after 70 h of fed-batch fermentation.In this study, the substrate preference of PCcrtZ genes, which have β-carotene and canthaxanthin substrate preference respectively, can greatly enhance the production of astaxanthin and increase the ratio of astaxanthin among total carotenoids.Combined utilization of The online version contains supplementary material available at 10.1186/s12934-022-01798-1. Carotenoids are organic pigments ranging in color from yellow to red that are naturally produced by certain organisms, including photosynthetic organisms , as well as some nonphotosynthetic bacteria, yeast and fungi. Carotenoids such as lutein, zeaxanthin or astaxanthin are important additives in food and feed industry as pigmenting substances and precursors of vitamin A derivatives –4. In adE. coli. The best recombinant strain reported in the literature expressed crtZ from Pantoea agglomerans and crtW from Nostoc sp. PCC 7120, producing an astaxanthin titter of 1.99 mg/g DCW [E. coli. The result showed that crtZ from P. agglomerans was found to be the most suitable for astaxanthin biosynthesis. Moreover, efficient astaxanthin biosynthesis also requires the balanced expression of β-carotene ketolase and β-carotene hydroxylase [crtW with respect to crtZ increased the efficiency of astaxanthin biosynthesis [crtZ [crtZ orthologs from different organisms were compared in the presence of both utilizable substrates in vitro [Paracoccus sp. PC1  and Agrobacterium aurantiacum preferentially converted canthaxanthin to astaxanthin, while CrtZ from the Pantoea ananatis (formerly Erwinia uredovora) preferentially catalyzed the formation of zeaxanthin from β-carotene. The specific enzyme activity of CrtZ from Paracoccus sp. PC1 was 28 pmol/h/mg protein when converted canthaxanthin into astaxanthin, and was 3.4 pmol/h/mg/ protein when β-carotene as substrate. Furthermore, CrtZ from Paracoccus sp. PC1 showed that canthaxanthin was the favored substrate for conversion, 6-fold for canthaxanthin compared to β-carotene, when both substrates were added to the enzyme preparation in an equal ratio. However, it was still unclear whether the same substrate preference was still present in vivo, or if the canthaxanthin preference of CrtZ could reduce canthaxanthin accumulation and improve astaxanthin production when combined with another β-carotene hydroxylase with β-carotene propensity.Astaxanthin, an important hydroxyl-carotenoid, has great potential commercial value in the feed, cosmetics, and nutraceutical industries due to its strong antioxidant capacity . The hetmg/g DCW . Subsequroxylase , 12. Zeaynthesis . Conversis [crtZ . We hypois [crtZ –14. The in vitro . The rescrtZ and crtW (including 2 copies of PAcrtZ and 1 copy of crtW*) (Table crtZ or crtW was added [P. agglomerans in E. coli.Ast007 is an astaxanthin producing strain in which as added . A largeBeta-carotene hydroxylase (CrtZ) introduces hydroxyl moieties at the 3, 3’ position of the β-ionone ring irrespective of the presence or absence of a keto group at the 4, 4’ position. In order to study the substrate preference of different CrtZ orthologs, platform strains that produce high levels of β-carotene (substrate without keto group) or canthaxanthin (substrate with keto group) were constructed.The numbers indicate the peaks of carotenoids in the HPLC chromatogram: 1, β-carotene; 2, canthaxanthin; 3, echinenone; 4, zeaxanthin; 5, β-cryptoxanthin; 6, adonixanthin; 7, astaxanthin; 8, lycopene.crtZ and a copy of crtW* were integrated into the chromosome of CAR026 at mgsA site, had much more lycopene accumulation then that of CAR026, even there wasn’t any modification on lycopene biosynthesis pathway [crtY can greatly reduce the accumulation of lycopene and increase the yield of astaxanthin [crtY gene into the chromosome of CAR026 at the mgsA site. The biomass and β-carotene production of CAR032 respectively decreased by 16 and 4% compared to that of CAR026 in shake flasks fermentation  and Zea002 (CAR032 with two copies of PCcrtZ) respectively produced 1.23 and 2.7 mg/L of zeaxanthin, while leaving a large amount of β-carotene  and Ast016 (Can004 with one copy of PCcrtZ) produced similar amounts of astaxanthin, which were 9.9 and 10.21 mg/L astaxanthin respectively  and Ast015 (Can004 with two copies of PAcrtZ) produced 19.84 mg/L and 16.4 mg/L astaxanthin, respectively. The astaxanthin production of Ast017 was 21% higher than that of Ast015  produced 13.12 mg/L of astaxanthin, which was a 34% decrease compared with Ast017  . Notably, P. agglomerans and P. anantis belong to the same genus, so the PAcrtZ has similar enzymatic characteristics to crtZ from P. anantis (β-carotene preference and substrate inhibition). Furthermore, high levels of canthaxanthin (40.7 mg/L in shake flask fermentation and 3.9 g/L in fed-batch fermentation) were produced by Can004, which has two copies of crtW*, and the major by-products of adonirubin (8%), canthaxanthin (5%), echinenone (5%), lycopene (12%) and β-carotene (17%) were accumulated besides astaxanthin. The low-level accumulation of zeaxanthin and adonixanthin may indicate insufficient CrtZ expression in Gro-46 [crtZ did not increase the astaxanthin yield. We speculated that PAcrtZ preferentially converts β-carotene into zeaxanthin, and cannot easily convert ketocarotenoids into astaxanthin. Accordingly, the low ratio of astaxanthin to total carotenoids in Gro-46 was probably caused by the substrate preference and substrate inhibition of PAcrtZ. To assess its substrate preference, PCcrtZ was expressed in platform stains producing β-carotene and canthaxanthin, respectively. Low levels of zeaxanthin was got by expressing PCcrtZ in CAR032, while high levels of astaxanthin was produced by expressing PCcrtZ in Can004, which suggested that PCcrtZ has a substrate preference for canthaxanthin over β-carotene, which was consistent with earlier reports.The most suitable biosynthetic genes that enable the efficient channeling of available metabolic intermediates towards the desired product are important for the  studies , 19–21. carotene . Furthertrations . In thisn Gro-46 . HowevercrtZ and two copies of crtW produced 1.73 g/L of astaxanthin, while Ast019 containing four copies of PCcrtZ and two copies of crtW* produced 1.75 g/L of astaxanthin along with certain amounts of canthaxanthin and adonirubin in fad-batch fermentation  using the primer pair CrtZ-RBS-99 A-CPEC-F/CrtZ-CPEC-R , and was amplified using the primer pair PC-crtZ-CPEC-RBS-F/PC-crtZ-CPEC-R . To determine the effects of E. coli gene insertions were performed using two-plasmid CRISPR/Cas9 technology as reported previously [All eviously , 25. ThecrtZ was described as an example to show how to integrate heterologous genes into the E. coli chromosome. The helper plasmid pCas9 and donor plasmid pManA-crtZ were co-electroporated into the β-carotene producing strain CAR032, and the resulting strain was processed using the Cas9 genome-editing protocol as described previously [crtZ integration at the manA site. Zea002, Zea003, and Zea004 were obtained by inserting PCcrtZ-PCcrtZ, PAcrtZ or PAcrtZ-PAcrtZ into the chromosome of CAR032 at the manA site respectively, using the same method.Integration of eviously –27, yielStrains Ast017, Ast018 and Ast010 were used for producing astaxanthin through fed-batch fermentation. Two stage systems were used for preparing seed. The first seed was prepared by inoculating fresh colonies into a 100 mL flask containing 10 mL medium at 30℃ with shaking at 250 rpm until OD600 was 0.7-1.0. Then 3 mL of culture from first seed stage was added to a 500 mL flask containing 120 mL medium and incubated at 30℃ with shaking at 250 rpm until OD600 reached 1.5. The second seed solution was transferred to a 5 L fermentation tank containing 3 L substrate medium. The fermentation substrate medium and fermentation progress were the same as those described previously .Carotenoids were extracted using acetone as described before . A sampl600) using the empirical formula 1 OD600 = 0.323 g DCW L− 1.The analytes were separated on a Symmetry C18 column , which was kept at 30 °C, and were detected at 476 nm using a Shimadzu UV-2550 spectrophotometer . The mobile phase and flow rate in measurement of carotenoids production were the same as that described previously . The resAdditional file : Text S1 Construction of plasmids for the chromosomal integration of heterologous genes. Text S2 The amino acid sequences of PCcrtZ. Text S3 The amino acid sequences of PacrtZ. Table S1. Primers used in this study. Table S2. Plasmids used in this study. Table S3. Caretenoids production by strains producing Zeaxanthin in shake flasks. Fig. S1. Flowchart illustrating the construction of the main strains used in this study Fig. S2. Carotenoids production of antxanthin producing strains. Fig. S3. Fed-batch fermentation results for canthaxanthin production by strains Can004 Fig. S4. Fed-batch fermentation results for astaxanthin production by strains Ast019.


To evaluate the knowledge, attitudes and behavior regarding antibiotics, use of antibiotics, and antibiotic resistance in students and health care professionals of the district of Barranquilla, Colombia.Descriptive, cross-sectional.A sample of 399 respondents was selected, that included health professionals and medical students from 12 health institutions in the district of Barranquilla (Colombia), using an established stratified sampling method. Each of the respondent professionals completed a survey that included 43 items in the Likert scale. A descriptive analysis of the study variables was performed using the software SPSS version 25.Most of the respondents were women (64.4%), aged between 26 and 35 years (47.6%); 28.8% were nurses and 26.1% general practitioners, with ≤10 years of professional experience (63.4%). Overall, the survey revealed that the participants had considerable knowledge about antibiotic use (89.5%–98% correct answers) and the spread of antibiotic resistance (67.4%–89% correct answers). Approximately 74% of the respondents agreed or fully agreed with the questions related to the management of infections and the provision of advice.The present study revealed that most of the health care professionals surveyed had a good knowledge about antibiotic use, although strategies must be developed to strengthen knowledge regarding the spread of antibiotic resistance. Likewise, it is important to identify opportunities for improvement related with access to the guidelines and/or materials necessary to treat infections and to provide advice on antibiotic use and antibiotic resistance. Microbial resistance; Antibiotics; Knowledge; Attitudes and practices; Colombia. They retrieved that in Asia, Europe, America, Africa and Oceania, the prevalence of mobile colistin resistance (mcr)-harbouring colistin-resistant E. coli was 66.72%, 25.49%, 5.19%, 2.27% and 0.32 %, respectively. These results suggested that the dissemination of this strain is worldwide arising concerns between countries as well [Antibiotic resistance is one of the most serious public health problems worldwide . The cliBarranquilla City, along with its metropolitan area, concentrate the largest number of health care provider institutions in the Colombian Caribbean region, and therefore, baseline studies aimed at professionals working in these institutions are required that allow identifying intervention needs on knowledge of antibiotics, antibiotic use, and resistance, and changes in behavior related with the prescription of antibiotics and antibiotic resistance.The present study aimed to assess the knowledge, attitudes, and behavior regarding antibiotics, antibiotic use, and antibiotic resistance in students and health care professionals from health institutions in the district of Barranquilla and its metropolitan area in Colombia.2Nearly 86.5%–91.2% of the respondents agreed or strongly agreed with the questions related with the perceived capacity . With reApproximately 74% of respondents either agreed or fully agreed that they have easy access to the guidelines needed to treat infections, that they have easy access to the materials needed to provide advice on the cautious use of antibiotics and on antibiotic resistance and that they have good opportunities to offer advice to people about the cautious use of antibiotics .Table 2PBetween 82% and 86.2% of the individuals surveyed, considered that they played a key role in helping to control antibiotic resistance and that there was a connection between the prescription, dispensation or administration of antibiotics and the emergence and spread of antibiotic-resistant bacteria .Table 3PAmong the 399 professionals surveyed, 174 (43.6%) were eligible to prescribe antibiotics; most of them were general practitioners (52.3%), internists (10.3%), intensivists (6.9%), general surgeons (5.8%), and pediatricians (5.2%). The frequency of antibiotic prescription was 55.8% on a daily basis, 36.8% weekly, and 7.5% on a monthly or quarterly basis. The daily prescription of antibiotics was mainly given by general practitioners (51.5%), followed by internists (14.4%) and intensivists (9.2%).Regarding the decisions made when prescribing antibiotics, the support, and access to the guidelines by the prescribers, it was found that between 83.2% and 92.3% of respondents agreed or totally agreed that they felt confident when making decisions related with the prescription of antibiotics, relied on the antibiotic guidelines available, took into account antibiotic resistance when treating a patient, had easy access to the guidelines on the antibiotics needed to treat the infections, and felt supported not to prescribe antibiotics when these were not needed . The mos3There are few published studies in Colombia about the knowledge, attitudes, and behavior regarding antibiotics and antibiotic resistance in health professionals. There is a previous study that evaluated the knowledge about the prescription of antibiotics in general practitioners and specialists in three Colombian cities, including Barranquilla ; HoweverThe above could also have occurred in the present study regarding the questions about knowledge that were answered incorrectly by our respondents, as the sample population in our study was also a heterogeneous one in terms of professions, which could lead to incorrect answers, as the professionals included in the sample could have various levels of training on antibiotics, antibiotic use, and antibiotic resis-tance.When considering our results and those from these other studies, it is important to design education and communication strategies for our population that impact the strengthening of this knowledge.Both the ability and the opportunity to influence the motivation of an individual to generate positive behaviors , which cThis study found that more than 80% of the prescribers surveyed agreed with the questions related with confidence when making decisions about the prescription of antibiotics, the accessibility to the guidelines on antibiotics to treat infections, and about the support felt not to prescribe antibiotics when they are not necessary. Despite the latter, reasons for an inadequate prescription of antibiotics were identified by the surveyed prescribers, the main one being fear of a worsening of the patient status or fear of complications. The least frequent reason being to maintain the relationship with the patient. Both indulgence and fear have been the most frequent attitudes found to influence the improper prescription of antibiotics in previous studies both in Colombia and other countries of the world , 12, 13.This study has some limitations, because did not include all health institutions in the district of Barranquilla, as some of the institutions summoned decided not to participate and others did not meet the inclusion criteria. In the participating institutions, the largest number of physicians hired in the general hospitalization and emergency rooms are general practitioners. It is common to observe that in the daily practice of antibiotic prescription, the general practitioner prescribes under the directive of the specialist, and therefore, it was not possible to differentiate between the prescription roles of the general practitioner and the specialist physician. The data doesn't contain items regarding right antibiotic selection based on accurate diagnosis and right antibiotic dose and duration selection. Finally, are necessary future studies outside heath institutions, could help in understanding the general awareness in antibiotic and its use among people.4The present study revealed that most of the health care professionals surveyed had a good knowledge about antibiotic use, although strategies must be developed to strengthen knowledge regarding the spread of antibiotic resistance. Likewise, it is important to identify opportunities for improvement related with access to the guidelines and/or materials necessary to treat infections and to provide advice on antibiotic use and antibiotic resistance in order to control barriers that hinder accessibility to the guidelines and materials necessary to guide appropriate practices in the treatment of infections and in the prevention and control of antibiotic resistance.5The population included in this cross-sectional study was comprised of medical students and health care professionals from 12 health care institutions in the district of Barranquilla and its metropolitan area and included health professionals and specialists who prescribe antibiotics and are members of the operational and coordination teams for surveillance and control programs for antimicrobial resistance and infections associated with health care: Physicians, specialists in internal medicine, general surgery, pediatrics, intensive medicine, infectious diseases, epidemiology, auditing, management, nursing, chemistry and pharmacy, microbiology, with a minimum of 6 months of employment at the institutions included in the study. These health institutions were selected because they had microbiology laboratories equipped with automated methods for the identification of multidrug resistance and to use the Whonet 5.6 software.We exclude health institutions without microbiological data and health professionals who at the time of completing the survey were disabled, on maternity leave and vacations.The sample was calculated from an approximate population size of 3,448 health care professionals linked to the participating health care institutions, with a 95% confidence interval and a power of 80% for a sample of 346 professionals, which increased by approximately 15% for a final sample size of 399 respondents. Each of these subjects was selected according to a proportionate stratified sampling, in order to maintain the proportions observed in the study population.A survey was applied between april and may 2021; the process was carried out online using the Microsoft Office 365 forms application, on a voluntary and anonymous basis. The survey was designed and validated by a previous study and included 43 items, that included statements knowledge using a true or false answer and statements assessing attitudes and behaviours using a 5 point Likert scale: strongly agree, agree, neither agree, nor disagree, strongly disagree [Demographic variables and variables related to knowledge, attitudes, and behavior regarding antibiotics, antibiotic use, and antibiotic resistance were assessed.Descriptive statistics based on absolute and relative frequencies were used to analyze the information. All the analyses were performed in the open epi and SPSS version 25 software.Soraya Salcedo, Laura Mora, Ronald Maestre-Serrano : Conceived and designed the experiments; Analyzed and interpreted the data; Wrote the paper.Dinno Fernandez, Adriana Marín, Indira Berrío Medina: Conceived and designed the experiments; Wrote the paper.Humberto Mendoza Charris, Erika Paola Viana Cárdenas, Michelle Polo-Rodríguez, Lorena Muñoz-García, Juany Alvarez-Herrera, Gisselle Olivares-Goenaga, Yorleidis Jimenez-Castro, Violeta Castro del Portillo, Sylena Chiquillo-Gómez, Lidys Barrios-Matute, Jaime Villa-García, Yolima Gonzalez-Mattos, Jose Torres-Barraza, Rafael Jaraba-Coronado: Contributed reagents, materials, analysis tools or data; Wrote the paper.Pan American Health Organization/World Health Organization .This work was supported by Data included in article/supp. material/referenced in article.The authors declare no conflict of interest.No additional information is available for this paper.


Carbon fiber-reinforced concrete as a structural material is attractive for civil infrastructure because of its light weight, high strength, and resistance to corrosion. Ultra-high performance concrete, possessing excellent mechanical properties, utilizes randomly oriented one-inch long steel fibers that are 200 microns in diameter, increasing the concrete’s strength and durability, where steel fibers carry the tensile stress within the concrete similar to traditional rebar reinforcement and provide ductility. Virgin carbon fiber remains a market entry barrier for the high-volume production of fiber-reinforced concrete mix designs. In this research, the use of recycled carbon fiber to produce ultra-high-performance concrete is demonstrated for the first time. Recycled carbon fibers are a promising solution to mitigate costs and increase sustainability while retaining attractive mechanical properties as a reinforcement for concrete. A comprehensive study of process structure–properties relationships is conducted in this study for the use of recycled carbon fibers in ultra-high performance concrete. Factors such as pore formation and poor fiber distribution that can significantly affect its mechanical properties are evaluated. A mix design consisting of recycled carbon fiber and ultra-high-performance concrete was evaluated for mechanical properties and compared to an aerospace-grade and low-cost commercial carbon fiber with the same mix design. Additionally, the microstructure of concrete samples is evaluated non-destructively using high-resolution micro X-ray computed tomography to obtain 3D quantitative spatial pore size distribution information and fiber clumping. This study examines the compression, tension, and flexural properties of recycled carbon fibers reinforced concrete considering the microstructure of the concrete resulting from fiber dispersion. Concrete is the most widely used building material in the world and plays a critical role in the world’s infrastructure ,2. TradiVarious fiber types, including steel fiber, glass fiber, basalt fiber, and synthetic fibers, have been used as reinforcements in concrete ,10,11. SCarbon fiber has a higher modulus than steel, which allows it to develop higher stresses before concrete fails at relatively low tensile strains . SimilarrCF is a promising viable option because carbon waste from the automotive and aerospace industries can be utilized . Carbon Numerous studies have been performed on rCF-reinforced concrete and cement-based applications ,50,51,52Akbar et al. investigated microstructural milled rCF using scanning electron microscopy (SEM) coupled with energy x-ray dispersive spectroscopy (EDS), thermogravimetric analysis (TGA), x-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR) . The surOne of the major applications of FRC is Ultra-High-Performance Concrete (UHPC) . UHPC isOne of the key advantages of UHPC is that its tensile strength is significantly higher than that of normal concrete . Fiber rBoth carbon fiber and steel fibers have been studied for use in UHPC as reinforcement ,77. SimiOne method for evaluating the pore structure of UHPC is to utilize μ-XCT. μ-XCT is an invaluable non-destructive evaluation method for composite materials with increasingly complex geometry. There is growing interest in examining the microstructure of fiber-reinforced concrete materials. The key benefit of μ-XCT is the ability to evaluate in 3D spatially critical parameters, including cracks, fiber orientation, the bond between fiber and matrix, and porosity, which can affect the mechanical performance of FR concrete ,86,87,88In this study, the authors developed a multiscale experimental program to evaluate rCF-reinforced UHPC (rCF-UHPC) compared to two commercially available carbon fiber and steel fibers for mechanical properties, including compression and flexural and tensile strength. The carbon fiber types investigated included an aerospace-grade carbon fiber made by Hexcel and an industrial-grade carbon fiber made by Zoltek, along with the rCF provided by Carbon Fiber Recycling LLC. There are limited studies on rCF and its use in UPHC; therefore, it is critical to investigate how it performs in UHPC and how it compares to other commercially available fiber types to determine its viability as a reinforcement for UHPC.To this end, different carbon fiber types were investigated using TGA to assess the thermal stability and sizing content of the fibers. Scanning electron microscopy was used to analyze the surface morphology of the fibers, and energy X-ray dispersive spectroscopy was used to identify the elemental chemical composition of the fibers. This allowed the different fiber types to be analyzed and showed how rCF compared to the commercially available carbon fiber types physically and chemically. A μ-XCT technique was used to analyze pore geometry and pore size spatially in 3D for each of the UHPC mixes to investigate porosity effects in relation to the mechanical performance of each mixture. The microstructure properties were compared with the mechanical properties and evaluated to show how the pore structure of the different mixes affected the strength of the UHPC mixes. Scanning electron microscopy was also used to image the failed surface of the tensile samples and to provide insight into the failure mechanisms of the concrete and fibers. Overall, this study provides novel insights into how rCF compares to other fiber types, how rCF can be used in UHPC, how μ-XCT can be utilized to show how the microstructure affects the final UHPC mix performance, and how the fibers affect the failure mechanisms of the concrete.The multiscale analysis of the concrete mixes used in this study was characterized in three stages. The first stage consisted of characterizing the carbon fiber types for thermal and physical properties. The carbon fibers and steel fiber types were examined using scanning electron microscopy and X-ray energy dispersive spectroscopy to identify the chemical elemental composition of each fiber type. The second stage involved a non-destructive evaluation of the concrete mixes using μ-XCT to study the effect of void volume and size distribution spatially. We also investigated the effects of void formation from the added fiber volume content in the concrete mix, including the resulting reinforcement mechanisms of the fibers surrounded by the host concrete matrix. The third stage evaluated the concrete mix designs for compressive, tensile and flexural strength properties, correlating the porosity and microstructure of the fiber-reinforced concrete mixes.®Fluid Premia 150, Royse City, TX, USA), water, and steel fibers , Chicago, IL, USA) in accordance with ASTM A820 [®Fluid Premia 150 is particularly useful for UHPC due to its high flowability characteristics, according to CHRYSO [STM A820 . AccordiSTM A820 . These pSTM A820 ,93. The STM A820 . The CHRo CHRYSO ,96. The ® AS/1925 Type 2, West Valley City, UT, USA), representing the high-end of the carbon fiber market, were used for concrete mix design, where the fibers were sized 3–7% by weight with a 1925 sizing system [https://imagej.nih.gov/ij/index.html (accessed on 14 July 2022)).The properties and corresponding images of the fiber types used in this study are summarized in g system ,97,98. Ag system . Hexcel g system . The fib3 cell volume in a helium gas atmosphere. Five cycles were repeated per sample for accuracy and to obtain statistical density values.The densities  of the tThermogravimetric analysis (TGA) was performed for the carbon fiber types to observe the degradation behavior and thermal stability of the fibers using a TA Instruments TGA . Samples for each fiber type with an approximate mass of 8.76 to 16.6 mg were placed in a 100-microliter platinum pan and heated from room temperature to 900 °C at a heating rate of 10 °C/min under an air atmosphere. To calculate the fiber sizing content, isothermal TGA was performed for the carbon fiber types, where samples with an approximate mass of 10.82 to 22.2 mg were heated from room temperature to 400 °C at 10 °C/min and held at 400 °C for 30 min.® series, Oberkochen, Germany). The fiber samples were Au sputter-coated for 40 s using an Au sputter coater. Additionally, energy X-ray dispersive spectroscopy (EDS) was performed on Au sputter-coated fiber samples using a ThermoFisher Scientific Apreo S SEM  with an accelerating voltage of 20 kV to observe the chemical composition of the fibers. The surface topologies of the mechanically failed tensile briquette samples were imaged using an environmental scanning electron microscope  with an accelerating voltage of 5 kV and a secondary electron detector.The surface topography and morphology of the carbon fiber types were observed using a dual-beam scanning electron microscope (SEM) and a Focus Ion Beam (FIB) using a Zeiss Auriga SEM/FIB instrument  rotary stage, where the samples were scanned using a voltage of 150 kV and an amperage of 137 uA (Hamamatsu L8121-03). The 3001 2D projections  were collected over an angular range of 360 degrees at approximately 0.12 degrees per step at a voxel resolution of 13.89 μm. The 2D projections were normalized, and corresponding sinograms from the projections were used to obtain 2D reconstructed slices using reconstruction algorithm software . Correction methods to reduce artificial defects, such as beam hardening and ring artifacts, were applied to the collected projections using the above-mentioned reconstruction software . The 2D 3) was approximately 7.9% less than the neat mix design (2.49 g/cm3), approximately 5.1% lower than the steel fiber mix design (2.42 g/cm3), approximately 5.8% higher than the Hexcel carbon fiber mix design (2.17 g/cm3), and approximately 3.1% higher than the Zoltek carbon fiber (2.23 g/cm3). The neat concrete and steel fiber-based concrete mix designs were used as a baseline to compare the three carbon fiber-based concrete mixtures. 3. The steel fiber and neat concrete had a relatively even distribution of voids compared to the carbon fiber samples, with a peak at voids with a volume of 0.01–0.05 mm3. The carbon fiber samples had a higher peak in voids, with a volume below 0.05 mm3. There was also a significant concentration of voids greater than 1 mm3 in volume observed for the concrete design mix, with the exception of Zoltek carbon fibers, which had no voids larger than 0.75 mm3 in volume. The rCF mix had a relatively higher volume of small voids (39.6%), below 0.01 mm3, but fewer large voids than the other carbon fiber types.The primary objectives of the study included performing a multiscale analysis investigating the relationship between rheological behavior, microstructural features of interest, such as porosity, and mechanical performance for all the considered mixes, particularly the influence of carbon fiber reinforcement in the concrete mix design. Considering the inverse relationship between increased fiber volume content and flowability, introducing fiber into the mix reduced the flow, as expected, which can affect the mechanical performance of the specimens . It was 3) than the steel (2.42 g/cm3) and neat (2.49 g/cm3) concrete. This indicates that air voids were trapped in the concrete since the addition of the lighter-density carbon fiber did not change the density of the concrete as much as was observed. Thus, these factors suggest that the mechanical performance of the carbon fiber-reinforced concrete mixes was significantly influenced by increased air voids and poor microstructure, which had a significant influence on the mechanical strength properties , negating the full potential benefits of the carbon fiber reinforcement.The reduced flowability affects how the concrete compacts into the mold, leading to increased porosity, void size, and poor dispersion of fibers in the concrete. Increased porosity can be observed through a decrease in the density of the concrete. The carbon fiber-based concrete mixes all have a much lower density . However, the Hexcel carbon fiber had the highest percentage of void content over 1 mm3 (13.0%) and the lowest tensile strength (5.29 MPa) of the fiber-reinforced concrete. This observation suggests that the tensile strength of the concrete mix is not just affected by the strength of the fiber used but also the void content, particularly large voids over 1 mm3 (The μ-XCT void content results shown in ibutions  and compibutions , a few ker 1 mm3 . As showThe effect of voids on concrete strength can be further illustrated through other studies that have investigated how void content and void distribution affect the strength of concrete. Li et al. developed a model for how voids affect the tensile and compressive strengths of concrete. In their study, they examined samples with different void contents and void sizes ranging from 0.01 mm to 1.6 mm and found that increasing the total void volume and increasing the size of the voids at the same total volume decreased tensile and compression strength . Knab etFurthermore, the SEM micrographs  of the fBased on these characterizations, rCF is comparable to commercially available Zoltek carbon fiber in both chemical composition and thermal degradation. The rCFs mechanical performance was comparable to steel fiber and Zoltek carbon fiber, showing that it is a suitable reinforcement for concrete. The rCF had a lower porosity than the other two carbon fiber types and fewer large voids than the Hexcel carbon fiber. Based on the SEM micrographs of the tensile samples, the rCF had comparable fiber dispersion and fiber bonding to the Zoltek carbon fiber, which was sized for concrete applications. Thus, rCF could serve as a viable replacement for steel fiber as a low-cost carbon fiber alternative in UHPC.The thermal, density, and scanning electron microscopy coupled with energy X-ray dispersive elemental analysis characterizations revealed that the recycled carbon fibers were chemically and physically comparable to Zoltek and Hexcel carbon fibers.The addition of carbon fiber significantly reduced the flow and workability of ultra-high-performance concrete compared to steel fiber and neat concrete mix, leading to increased porosity.The addition of recycled carbon fibers can increase the tensile and compressive strength of ultra-high-performance concrete and is comparable to steel fibers.Micro X-ray computed tomography is an effective tool for the non-destructive evaluation of concrete mix design to obtain quantitative spatial information on microstructural features of interest, including formation porosity, void size distribution, and potential fiber clumping.All fiber types of reinforced concrete mix designs indicated fiber pull-out as the dominant failure mechanism for the tensile samples, indicating poor bonding with the concrete host matrix. However, the Zoltek carbon fiber and recycled carbon fiber showed reduced pullout and higher tensile strength, indicating improved bonding due to compatible sizing and rough surface morphology compared to the Hexcel carbon fiber.Carbon fibers did not provide strength post-cracking, potentially due to the fiber shearing once the cracks developed.The effect of using recycled carbon fibers in ultra-high-performance concrete was evaluated for its physical and mechanical properties. The recycled carbon fiber-reinforced concrete design mix was compared to two commercially available carbon fibers, aerospace-grade carbon fiber with platelet geometry and intermediate modulus chopped low-cost carbon fibers. The three carbon fiber reinforcements were evaluated and compared with neat concrete and steel fiber-reinforced concrete. The carbon fibers were evaluated for their flowability in concrete, and physical properties, including density. The carbon fibers and steel fibers were evaluated for thermal stability in air using thermogravimetric analysis. Additionally, the surface morphology of the fibers and elemental chemical composition were examined using scanning electron microscopy, coupled with X-ray dispersive spectroscopy analysis. Micro X-ray computed tomography was used to provide spatial quantitative information on the microstructure of the concrete mix designs used in this study, including void volume size and distribution relationship with the mechanical performances of the resulting concrete mix. The considered concrete mixes were evaluated for tensile, flexural, and compressive properties. Scanning electron microscopy was also used to investigate the failure modes of the tensile samples. The following conclusions can be drawn from the concrete mixes evaluated in this study:In this study, it was observed using carbon fiber as a replacement for steel fiber has been shown to provide comparable or better mechanical performance, with recycled carbon fiber being of particular interest because it is potentially more economically feasible compared to other types of carbon fiber. One major challenge with using carbon fiber in concrete is the decrease in workability, which leads to increased void content, which was observed with both simple density measurements and micro X-ray computed tomography scans. The total void volume and void size both negatively affect the strength of the concrete, and, in some cases, can completely negate the benefits of carbon fiber. The fibers pulled out of the concrete without developing their full strength, showing that there was poor bonding between the fibers and the concrete, thus further reducing the benefit of the fiber. The effects of fiber bonding are shown to significantly improve the tensile strength of concrete, with the sized Zoltek carbon fibers showing particular promise in tension. Unfortunately, the Zoltek fibers also showed poor workability due to their length. In future studies, the workability and void content of carbon fiber-reinforced concrete should be carefully evaluated to ensure that the fibers being used can achieve their full potential, and how well the fibers bond within the concrete should be examined. Overall, it was demonstrated that recycled carbon fibers as a reinforcement in ultra-high performance are comparable with the commercially available aerospace-grade carbon fiber and intermediate low-cost carbon fibers, along with steel fiber reinforcement and neat concrete mixes used in this study.


Background. Antibiotic treatment in emergency general surgery (EGS) is a major challenge for surgeons, and a multidisciplinary approach is necessary in order to improve outcomes. Intra-abdominal infections are at high risk of increased morbidity and mortality, and prolonged hospitalization. An increase in multi-drug resistance bacterial infections and a tendency to an antibiotic overuse has been described in surgical settings. In this clinical scenario, antibiotic de-escalation (ADE) is emerging as a strategy to improve the management of antibiotic therapy. The objective of this article is to summarize the available evidence, current strategies and unsolved problems for the optimization of ADE in EGS. Methods. A literature search was performed on PubMed and Cochrane using “de-escalation”, “antibiotic therapy” and “antibiotic treatment” as research terms. Results. There is no universally accepted definition for ADE. Current evidence shows that ADE is a feasible strategy in the EGS setting, with the ability to optimize antibiotic use, to reduce hospitalization and health care costs, without compromising clinical outcome. Many studies focus on Intensive Care Unit patients, and a call for further studies is required in the EGS community. Current guidelines already recommend ADE when surgery for uncomplicated appendicitis and cholecystitis reaches a complete source control. Conclusions. ADE in an effective and feasible strategy in EGS patients, in order to optimize antibiotic management without compromising clinical outcomes. A collaborative effort between surgeons, intensivists and infectious disease specialists is mandatory. There is a strong need for further studies selectively focusing in the EGS ward setting. Antibiotic resistance is a major global issue, and the appropriate use of antibiotics is a cutting-edge topic in the current literature. Abdominal acute care and emergency surgery is a particularly challenging situation for the proper management of antibiotic treatment. Intra-abdominal infections (IAI) are known to be associated with high morbidity and mortality, prolonged hospitalization and increased hospital costs. Surgeons often face complex clinical scenarios in emergency general surgery (EGS). Patients presenting with IAI have an increased fragility due to the infectious condition itself, that may be further aggravated by the presence of comorbidities or immunosuppression. Moreover, patients with IAI may be at an increased risk of multi-drug resistant bacterial infections in case of recent hospitalization or recent surgery. When facing an emergency surgery patient, antibiotic treatment should be balanced together with source control strategies, in a relationship often challenging to manage and with timing that is often difficult to define. Of course, antibiotics are a cornerstone in the treatment of emergency general surgery (EGS) conditions. However, in this complex setting, there is an increased risk of antibiotic misuse and over-use. A rate of up to 47% of inappropriate antibiotic prescriptions has been documented in surgical specialties ,2,3. ThiA literature search was performed on PubMed and Cochrane to identify suitable publications using the following search terms: “de-escalation” AND (“antibiotic therapy” OR “antibiotic treatment”). The searches were limited to papers fully published in English. The resulting outputs were combined, excluding duplicate results. Abstracts were scanned for suitability and the full text retrieved for all potentially relevant studies. Bibliographies and reference lists were reviewed to identify additional relevant studies.The inclusion criteria for the studies were as follows: (1) ADE of antimicrobial therapy, (2) application of any intervention or provision of substantial epidemiological data to judge the effects or determinants of ADE.The first issue when talking about ADE is the absence of a globally accepted consensus definition. Although commonly defined as a narrowing of the spectrum of antimicrobial treatment, ADE has also been identified in other forms. ADE has been described as a decrease in the number of antibiotics used in a treatment regimen or as a shortening in the duration of the therapy . MoreoveAccording to international guidelines, the use of broad spectrum antibiotics is the cornerstone of the empirical treatment of critically ill EGS patients, in order to minimize the risk for inadequate antimicrobial therapy ,9,10. HoDe-escalation strategies may differ in many aspects. Implementing an empirical antibiotic coverage is pivotal to cure patients with sepsis, but it has to be targeted as soon as possible. So, it is mandatory to obtain appropriate cultures before starting an antimicrobial therapy in order to identify the pathogens. In addition, broad spectrum therapy should treat the entire possible microorganism responsible for septic status. Obviously, once the pathogens are identified, the therapy has to be narrowed. De-escalation strategies include shortening the duration of the therapy and source control, as well as narrowing antimicrobial therapy on the results obtained through the cultures. Surgeons must maintain a high warning level when managing an EGS patient. Multiple cultures should be obtained intra and postoperatively. Ideally, antimicrobial therapy should be directed at the common microbiota of the suspected source of sepsis. For IAI, factors known to be associated with worse outcomes and nosocomial or multi-drug resistant pathogens are length of hospital stay or recent health-care exposure, recent antibiotic treatment and recent abdominal surgery . When reCultures results need usually at least 48 h to be ready. They can be deemed unreliable in case of previous antibiotic exposure. Moreover, a 30–80% rate of negative cultures in patients clinically considered infected has been reported. Molecular diagnostic solutions have been proposed to speed the process up such as PCR array merged for specific clinical contests . Two disadvantages of these technologies are costs and the need for blood cultures to establish antibiotic susceptibility, since only a limited number of acquired resistance genes can be screened.Antibiotics stewardship, ward rounds and frequent antibiotic time-outs are a fundamental part of ADE strategies. Evaluation of pharmacokinetics and pharmacodynamics are of utmost importance since they can fluctuate significantly in critically ill EGS patients. Optimization of dosing strategies include the optimization of the time the serum level of beta-lactams antibiotics are above the mean inhibitory concentration (MIC), as well as optimizing the peak serum level above the MIC for fluoroquinolones and aminoglycosides. It has been estimated that one out of six patients receiving beta-lactams does not reach the minimal concentration target and many more do not reach the target associated with maximal bacterial killing . UtilizaAnother fundamental part of ADE strategies in EGS is proper source control. Source control is defined as elimination of infective foci through drainage, debridement, device removal, compartmental syndrome decompression and deferred definitive restoration of anatomy and function. Source control can be both surgical and interventional in abdominal emergencies. The appropriateness of the source control does not only rely on the completeness of the abdominal infection control but also on a timely strategy. Source control should be obtained as soon as possible, especially in critically ill and septic patients. In-hospital delay of source control has demonstrated a worsening of the outcome in several clinical scenarios. The efficacy of source control is time-dependent and is a major determinant of outcome regardless of the effectiveness of antimicrobial therapy .Biomarkers can help in identifying bacterial infections and guiding the treatment strategy. Procalcitonin (PCT) levels are high in bacterial infections, while they remain low in viral infections or non-infectious systemic inflammatory syndromes. PCT could be a useful aid in choosing the proper duration of antibiotic therapy . ProlongImmuno-compromised patients deserve a dedicated mention when discussing about ADE. These patients could have sepsis or septic shock without any typical warning sign such as leukocytosis or fever. Some examples of these patients are solid organ transplant recipients receiving immunosuppressive medications, haematological solid cancer patients receiving cytoablative chemotherapy and HIV-infected patients. In addition, some authors suggest ageing is associated with a decreased immunity response predisposing these patients to severe bacterial infections . These pIn patients with uncomplicated IAI such as uncomplicated appendicitis and uncomplicated cholecystitis, where the source of infections is treated definitively, postoperative antibiotic therapy is not necessary . GuideliSimilarly, uncomplicated acute calculous cholecystitis patients can be treated without postoperative antibiotics when the focus of infection is controlled by cholecystectomy. In a recently published prospective randomized controlled trial, patients were randomized to either no antibiotics after surgery or continuation with the preoperative antibiotic regimen three times daily for 5 days. The postoperative infection rates were 17% (35/207) in the non-treatment group and 15% 31/207) in the antibiotic group  [/207 in tNo clear consensus exists on ADE components and various definitions have been used. Moreover, most studies focus on ICU patients and there is a lack of high quality data selectively investigating ADE strategies in EGS. ADE has been reported to be applied in only 40–50% of inpatients with bacterial infection . IAI are


Neonatal sepsis is an important cause of neonatal morbidity and mortality. A significant proportion of bacteria causing neonatal sepsis is resistant to multiple antibiotics, not only to the usual empirical first-line regimens, but also to second- and third-line antibiotics in many neonatal intensive care units (NICUs). NICUs have unique antimicrobial stewardship goals. Apart from antimicrobial resistance, NICUs have to deal with another problem, namely biofilm infections, since neonates often have central and peripheral lines, tracheal tubes and other foreign bodies for a prolonged duration. The aim of this review is to describe traditional and novel ways to fight antibiotic-resistant bacteria and biofilm infections in NICUs. The topics discussed will include prevention and control of the spread of infection in NICUs, as well as the wise use of antimicrobial therapy and ways to fight biofilm infections. Neonatal sepsis is an important cause of neonatal morbidity and mortality. The incidence is 1 to 5 cases per 1000 live births , and it Escherichia coli, while the main isolates of late-onset sepsis (LOS) were Klebsiella pneumoniae and Escherichia coli. Almost 65% of Escherichia coli isolates and 78% of LOS isolates were MDR [Staphylococcus aureus isolates [Pseudomonas aeruginosa were multidrug-resistant [A significant proportion of bacteria causing neonatal sepsis is multidrug-resistant (MDR), not only to the usual empirical regimens, but also to second- and third-line antibiotics in many neonatal intensive care units (NICUs) ,7,8. A pisolates . Anotheresistant .In NICUs, antibiotics represent the most prescribed drugs and this renders them a high-risk environment for MDR-organism colonization and spread. Furthermore, the prolonged hospitalization of patients and the common use of foreign bodies, such as central and peripheral lines, tracheal tubes and other foreign bodies, are risk factors for antimicrobial resistance and biofilm infections .There is an ongoing effort from several health organizations, such as the Centers for Disease Control and Prevention and the World Health Organization, to address the problem of antibiotic resistance. Many antibiotic stewardship programs are aiming to deal with this problem. However, for common conditions requiring antibiotics in the NICU, there is still a paucity of data to guide day-to-day practice and antibiotic stewardship problems .This review aims to describe traditional and novel ways to fight antibiotic-resistant bacteria and biofilm infections in the NICU. The topics discussed will include prevention and control of the spread of infection in the NICU, the wise use of antimicrobial therapy, and ways to fight biofilm infections.We searched Pubmed and Google Scholar for relevant studies on traditional and novel ways of fighting antibiotic resistance and biofilm infections in NICUs. The following terms when used: “neonatal sepsis”, “neonate”, “infection”, and “antimicrobial resistance”, “multidrug resistance”, “antibiotic resistance”, or “biofilm infections”, up to December 2022. The initial literature search identified almost 1100 relevant studies. The titles, and then abstracts of the retrieved articles, were scanned for relevance. We also reviewed the reference lists of the retrieved articles in search of other relevant articles that could have been missed in the initial search. We included in this narrative review only papers written in English and papers referring to the population hospitalized in NICUs. Finally, 100 articles were included in the main text and specifically systematic reviews, meta-analyses, randomized controlled trials, observational studies, and narrative reviews were included.Antimicrobial therapy is a common practice in the everyday routine of NICUs and antibiotics are the most prescribed drugs in NICUs for many reasons. Firstly, neonates, and specifically preterm ones, are more vulnerable to infections and their survival relies on antibiotics. Many risk factors, such as prematurity, low birth weight, and immature immune system, require invasive procedures and prolonged hospitalization, which renders neonates a high-risk group for infection .Moreover, there are many diagnostic challenges in NICUs. Neonatal sepsis can present with non-specific clinical signs, such as apnea, respiratory distress, tachycardia and temperature instability rendering the diagnosis difficult, while on the other hand, the absence of clinical findings does not exclude infection . Many otStaphylococcus aureus (MRSA) in NICUs [The risk factors for developing multidrug-resistant pathogens are also different in NICUs compared to adult and pediatric ICUs. Gestational age below 32 weeks, birth weight below 1500 g and exposure to vancomycin or carbapenems were found to be independent risk factors for methicillin-resistant in NICUs ,19,20. Min NICUs ,19,20.Therefore, NICUs have special antimicrobial stewardship needs that deal with these diagnostic challenges and guide clinical practice.A traditional, simple and important way to reduce the spread of resistant bacteria is to reduce the number of infections, since fewer infections means fewer prescribed drugs. Simple and effective methods for infection control, such as safe delivery, avoidance of unnecessary invasive procedures, sanitation, proper cleaning of NICU equipment, hand hygiene before and after interaction with patients or their environment and restricted entry to NICUs are the milestones of prevention and should be strictly adhered to. Hand hygiene is crucial for the restriction of horizontal transfer of MDR microorganisms . AlthougCentral-line-related infections are also an important problem in NICUs. A prospective registry of 6215 very low birth weight infants found that central line catheters were commonly used in hospitalized infants and were associated with an increased risk of late-onset sepsis . HoweverAntifungal prophylaxis is also administered in many NICUs, especially if there is a high incidence of invasive candidiasis. According to the Clinical Practice Guidelines for the Management of Candidiasis in 2016, neonates with a birth weight below 1000 g could be administered intravenous or oral fluconazole twice weekly for 6 weeks. Alternatively, for neonates with a birth weight below 1500 g, oral nystamycin can be used .NICUs face many diagnostic challenges, as already described. Accurate diagnosis of sepsis and prompt identification of high-risk infants that need antibiotic therapy is an important first step in limiting the inappropriate use of antibiotics. The main challenge is that neonatal sepsis can present with various nonspecific signs and symptoms, while many of these findings overlap widely with those of other noninfectious causes . As a reThe gold standard for diagnosing sepsis is blood cultures. Ideally, two blood cultures of sufficient blood volume (1–2 mL) should be collected with a sterile technique, before antimicrobial therapy initiation ,29.Biomarkers, such as C-reactive protein (CRP), procalcitonin (PCT), white blood cell count and erythrocyte sedimentation rate, can serve as useful adjunctive tools in combination with clinical symptoms and signs in the diagnosis of infection. A systematic review and meta-analysis of 22 cohort studies including 2255 infants, found that CRP was not helpful at the initial evaluation of possible LOS, and was insufficient alone to guide diagnostic and treatment decisions . HoweverAnother area of active research is the development and validation of sepsis calculators to better predict EOS. In 2014, Escobar G et al. published a retrospective nested case-control study of 600,000 neonates from 14 hospitals from 1993 to 2007, and developed a risk classification scheme for EOS. This calculator, which is perhaps the most promising and widely used, applies to neonates 34-week-old and older and is a method of estimating the risk of EOS. This calculator uses both maternal risk factors, as well as factors from the clinical examination of the newborn, and the neonates are stratified into three risk groups: treat empirically, observe and evaluate, and continued observation ,37.The accuracy of this calculator, provided by the Kaiser Permanente Division of Research, has been tested in many studies. A meta-analysis of 13 studies, analyzing a total of 175,752 newborns, showed that the use of this calculator limited the use of antibiotic therapy for suspected EOS without missing cases of EOS, while evidence regarding safety was limited . The numFor LOS, there is a score, namely, the RALIS score ,41, thatExcept for the RALIS score, there is a variety of predictive scores for LOS that has been studied in the literature. The available scores were based exclusively on clinical variables, laboratory variables or risk factors, or were based on combinations of these variables. Sofouli et al. conducted a systematic review of the available scores and concluded that the majority of them may assist in early diagnosis, but almost all had a limited diagnostic accuracy . The scoTherefore, accurate diagnosis of sepsis is of paramount importance in order to restrict antibiotic administration. A combination of clinical signs and symptoms, properly drawn blood cultures and serum biomarkers can help promptly identify suspected sepsis. The sepsis calculators are also adjunct useful tools. Serum biomarkers are also useful in limiting the duration of antibiotic therapy.Streptococcus and Escherichia coli, while LOS is also caused by other Gram-negative bacilli, Candida spp., and Staphylococcus [Once the clinical decision that antibiotic therapy should be initiated has been made, the wise selection of antibiotics is mandatory. Antibiotics are the most prescribed drugs in the NICU and account for one-third of the most frequently used medications ,47. The lococcus ,23. Empilococcus .In LOS, there is a wide variation of antibiotic use among centers and NICUs. In a study of ten units in the Netherlands, seven different empiric regimens were used for LOS, six for meningitis, and seven for necrotizing enterocolitis (NEC) . A retroStaphylococcus Aureus, vancomycin usage should also be reserved and nafcillin or oxacillin should be preferred instead. The most commonly used antibiotic regimens in neonatal sepsis are shown in The most widely recommended regimen for LOS is the combination of a semisynthetic penicillin with an aminoglycoside. For Gram-negative organisms, piperacillin-tazobactam or cefepime can be used instead, especially in infants known to be colonized with resistant Gram-negative organisms . Third-gFor NEC there are no guidelines to clearly state the optimal empirical therapy. The empirical regimen should include antibiotics that are effective against the most common pathogens associated with NEC, but also preserve the normal microbiota . StudiesA wide variability in antimicrobial therapy was also identified in neonates that needed surgical intervention. In a study of 191 neonates with gastroschisis, some sites used the combination of ampicillin and gentamicin, while other sites used the combination of cefotaxime and metronidazole .The high variability of antibiotic regimens in common neonatal condition, such as LOS and NEC, highlights the need for treatment guidelines for these conditions. The goal of antimicrobial stewardship programs is to optimize antimicrobial prescribing habits. To limit the risk of emerging antibiotic-resistant microbes, the narrowest spectrum antibiotic regimen should be given for the shortest duration of time. Longer antibiotic courses are associated with an increased risk of colonization with multidrug-resistant organisms, necrotizing enterocolitis, and/or death ,63. ContFurthermore, in newborns, it is difficult to administer the optimal dose of antimicrobials and accurately monitor the drug levels since the glomerular filtration rate is lower and drugs exhibit different pharmacodynamics and pharmacokinetics .It is very important that antibiotic administration is constantly reevaluated and ceased when there is no further indication. When the culture results become available within 48 h, the antimicrobial therapy should be modified according to the isolated pathogen and its sensitivities. This practice is described as a de-escalation of therapy . SpecifiLimiting patient exposure to broad-spectrum antibiotics can result in decreased antimicrobial resistance and cost . FurtherWhile step-down or de-escalation of therapy is common advice in stewardship programs, it is often not followed, since clinicians tend to continue the treatment that was proven to be effective ,68.The duration of empiric treatment for possible EOS in the NICU varies significantly ,69,70. AThe duration of antibiotic therapy should be well established for many clinical conditions, such as sepsis, NEC, and perioperative prophylaxis, and should be constantly re-evaluated for proper discontinuation . For NECIn asymptomatic newborns born to mothers with chorioamnionitis, the American Committee of Fetus and Newborn recommends a discontinuation of antibiotics within 48 to 72 h, if blood cultures remain negative ,74. If tFor other neonatal infections, the recommended antimicrobial duration is 7 to 10 days for hospital-acquired pneumonia, 10 to 14 days for uncomplicated bloodstream infection, 14 to 21 days for complicated bloodstream infections, 7 to 14 days for catheter-associated UTI and 48 to 72 h for culture-negative chorioamnionitis . RegardiThe duration of therapy for culture-negative sepsis varied widely in a retrospective study of 30 academic NICUs, while the duration of the antibiotic course was not associated with clinical signs or antepartum risk factors for sepsis . An examThere is a scarcity of data regarding the duration of perioperative chemoprophylaxis in neonates undergoing surgery. A European study of 1799 children showed that surgical prophylaxis was administered for more than 1 day in 67% of children and neonates . AnotherAntibiotic stewardship efforts should focus on standardizing the duration and the empirical therapy of common neonatal conditions, to prevent antibiotic resistance, since longer antimicrobial therapy duration favors the development of resistant pathogens. The summary of the available ways to fight antibiotic resistance is shown in Biofilm formation is a severe health concern in NICUs and a persistent threat, as it increases both morbidity and mortality. Bacterial biofilms are immobile microbial communities with diverse bacterial cell colonies in groups, which colonize and grow on the surfaces of almost all medical devices or prostheses . The devIt has been estimated that most bacterial infections in humans are correlated with biofilms and about 50% of the nosocomial infections are indwelling-device associated . NeonateEnterococcus faecalis, Pseudomonas aeruginosa, Staphylococcus aureus and Staphylococcus epidermidis, while numerous other bacteria have been associated with biofilm infections in other adherent surfaces [Bacteria that have been associated with biofilm infections in central venous catheters are surfaces .Although it is well known that antibiotic treatment is currently the most important and effective measure of control for microbial infections, antibiotic treatments are almost impossible to eradicate biofilm infections . The devAs already described, biofilm infections are chronic and can have exacerbations. Antimicrobial therapy can be helpful in mitigating an exacerbation, but cannot fully eradicate the bacteria in a biofilm . Since bThere are many novel approaches under development for the prevention and treatment of biofilm infections. Specific agents may inhibit the initial biofilm attachment, result in the removal of biofilms or inhibit biofilm formation by quorum quenching (the communication process among cells) . InitialAt present, the prudent use in the NICU of all those foreign materials that act as substrates for biofilm infections is very important. For example, catheters should be removed as soon as possible, and less invasive ventilation modalities should be used . Traditional and novel ways of fighting antibiotic resistance are shown in The neonatal population has unique characteristics and antibiotic stewardship needs in order to restrict antibiotic resistance and biofilm infections. Simple and effective methods for infection control should be strictly followed by clinicians, and are milestones in the prevention of infections. Accurate diagnosis of sepsis, use of the narrowest-spectrum of antibiotics and for the shortest necessary duration, as well as de-escalation and proper discontinuation of therapy are important steps in the wiser use of antibiotics, that will limit antibiotic resistance. Lastly, biofilm infections are very difficult to treat and therefore, proper handling and early withdrawal of foreign materials is mandatory in order to prevent them.


Antibiotic resistance is a complex phenomenon heavily influenced by social, cultural, behavioural, and economic factors that lead to the misuse, overuse and abuse of antibiotics. Recent research has highlighted the role that norms and values can play for behaviours that contribute to resistance development, and for addressing such behaviours. Despite comparatively high antibiotic consumption in Greece, both at the community and healthcare level, Greeks have been shown to be relatively aware of the connection between antibiotic overuse and antibiotic resistance. This suggests that Greeks’ non-judicious use cannot simply be explained by lack of awareness but may relate to other factors specific to Greek society. The present study aimed to explore attitudes, perceived norms, and values in relation to antibiotics, in order to improve understanding of socio-cultural determinants of antibiotic resistance in Greece.Data were collected through online focus group discussions in 2021. Twenty Greeks were recruited through purposive sampling, aiming for as heterogeneous groups as possible regarding gender , age , and education level. Interview transcripts were analysed inductively using thematic content analysis.Participants considered antibiotic overconsumption as a consolidated habit influenced by ease of access, social expectations and, more generally, cultural practices. While critical of such norms and practices, participants opposed stewardship measures that would prioritize the societal interest in maintaining antibiotic effectiveness over individual needs. Participants considered responsibility for antibiotic resistance to be shared by the whole society, but the role of government actors and health professionals as well as of food producers was emphasized. Notably, scepticism about the prospect of effectively managing antibiotic resistance in Greece was commonly expressed.The study makes explicit attitudes, perceived norms and values that, besides limited awareness, may contribute to non-judicious antibiotic use in Greece. These socio-cultural determinants of antibiotic resistance warrant further research and should be considered when designing measures aimed to mitigate this problem.The online version contains supplementary material available at 10.1186/s12889-022-13855-w. Antibiotic resistance (ABR) enables bacteria to survive and proliferate despite exposure to antibiotic treatment. Resistance can be an innate feature of certain bacteria, or acquired through mutations in chromosomal genes or horizontal gene transfer . ABR is Worldwide, healthcare is already facing the effects of ABR; the diminishing effectiveness of antibiotics jeopardizes treatment of infections as well as prevention of complications to tertiary care associated with surgeries, cancer treatment, and other modern health procedures . It has The latest report on antibiotic consumption of the European Centre for Disease Prevention and Control ranked Greece the second highest country in EU/EES for total antibiotic use both at the community and healthcare level in 2020 . PreviouAlthough consistently regarded as a pre-requisite for judicious antibiotic use and for other behaviours that can impact ABR, information-giving in itself is often considered insufficient to change people’s behaviour , 11. ABRIn Greece, electronic prescription was legally mandated as the only antibiotic prescription option with the latest (2020) national plan against ABR . This caThe present study aimed to explore attitudes, perceived norms, and values in relation to antibiotics, in order to improve understanding of socio-cultural determinants of ABR in Greek society. Previous quantitative studies have focussed on the Greek population , 20–22, A qualitative and explorative design was used to collect data through focus group discussions (FGDs). FGDs provide insight into behaviour by generating a process that helps participants to self-disclose and are particularly useful to explore a group’s norms as well as the range of viewpoints that exist within a population , 30. ThiThe semi-structured interview guide was adapted from Ancillotti et al.  after a Consolidated criteria for reporting qualitative studies (COREQ) were adhered to  through purposive sampling, aiming for the composition of a sample as heterogeneous as possible regarding gender, age, and education level. Even before the COVID-19 pandemic limited the possibility of face-to-face encounters with participants, social media platforms and online video platforms (such as Zoom) had been recognised as valuable tools for recruitment and data collection in health research .The FGDs were conducted by the first author, a male student of Public Health Science (MSc) with a background in Molecular Biology and Genetics (BSc), using Zoom in February 2021. Participants were informed about the topic of the discussion. They had no prior relationship with the interviewers nor among themselves. The FGDs lasted between 45 and 60 minutes. After about 10 minutes, participants watched a six-minute animated video presenting basic facts about ABR mechanisms, how antibiotics work, and global issues connected to access, excess, and antibiotic pollution . Data saThe transcripts were coded manually and analysed inductively using thematic content analysis . In the The second stage meant eliminating duplications and overlapping or too similar categories and was carried out by the whole research team. In the final stage, distinctions were refined by sorting the remaining categories into groups. The research team discussed the results to find consensus. The categories are descriptive and use the participants’ terms. The themes are the result of a final abstraction process and are thus interpretative. Transcripts were not returned to participants, who were not further involved after the FGDs.Twenty members of the general public participated in four FGDs  Everyone takes antibiotics like caramels and turns to antibiotics without necessarily visiting a physician to prescribe them, just because a person took the same antibiotic when he/she had the same symptoms, and therefore this will do the same for me. (Group(G)2 Woman(W)3)(Q2) And sometimes even physicians are telling you to take an antibiotic and you will be fine. So … doctors also drive you to this pathway. (G2W2)Participants considered it a major problem that antibiotics are easily accessible and that unduly obtaining antibiotics is not hindered significantly by either cost or the prescription procedure:(Q3) I also think the problem is the rather easy access, because of the cost and because no serious prescription is required. (G2W1)Participants explicitly mentioned that the Greek culture may be considered the main antagonist to engagement in behaviour beneficial to the greater good:(Q4) Unfortunately, it is deeply rooted inside our culture, the Greek culture that antibiotics are used as a precaution … What matters is to fight against a culture that is deeply rooted inside us. And even now that prescription is more controlled, antibiotics will be given with abandon. (G1W3)Participants looked upon antibiotic overconsumption as a consolidated practice in Greece. Individuals’ social circle but also physicians were referred to as exerting considerable influence towards excessive use of antibiotics:Under ‘values’ are categorized general, abstract views expressed by the participants about what is important and what should matter in relation to ABR.(Q5) I believe that human life, whether it concerns an individual or millions, has the same value … I do not think that many will agree to strict measures. (G2W1)As a consequence, they regarded it as morally questionable to limit individuals’ access to potentially beneficial treatment in the name of the greater good:(Q6) We cannot say that a part of society will suffer for the good of the whole society. Because if parts of society suffer, society can be dissolved. If the patient will certainly survive but will suffer for the good of all the others, in the end, who is responsible for making this decision? Who has the right to say that you will suffer for the good of others? (G2Man(M)1)Nonetheless, a few participants claimed that they would forego antibiotics for themselves and close ones, as long as this would not put their life in danger:(Q7) [When it is not a life-threatening infection], I would avoid using antibiotics for myself and the people around me. (G1W3)Elaborating on the values underpinning cultural norms, the role of emotions in decision making, including resource allocation, was emphasized:(Q8) I just think that I could not stand it personally and emotionally because every human being is important to the society in which he/she lives. It may have a logic to help the common good, but if the person in danger was one of my close people, I would be against it. So, I believe that we Greeks work more with emotion than with logic. (G2W4)Participants were critical of the idea of withholding antibiotic treatment to prioritize the societal interest in maintaining antibiotic effectiveness over individual needs. In doing so, they often supported their arguments by referring to the role of sentimental and cultural values as well as to inalienable individual rights. Respondents sympathized with each individual who would need antibiotic treatment and emphasized the equal value of all human life:Under ‘responsibility’ are categorized participants’ views about who is responsible for ABR, what such responsibility entails, and where blame should be directed.(Q9) Obviously we have a personal responsibility. Antibiotics are needed in our lives … It depends on how somebody wants to overcome an illness and how much one wants to suffer. It is a personal matter if I take antibiotics. No one obliges me in the end. The state is not responsible for my reckless use. (G2W2)However, for the individual to act responsibly there would be a need for the state to set conditions conducive to individual responsibility:(Q10) I focus on our personal responsibility but also on the state that must set the limits and then, certainly, individual responsibility. I believe that first the state must function properly. (G2W4)The notion of collective responsibility was a vague concept for most of the participants: a scenario in which society would share and act on the idea of a common purpose did not seem realistic:(Q11) So our society does not have the [features] to deal with it. Beyond antibiotics, there is a public confusion, in general. We have lost track of things. (G4M1)In contrast, a couple of participants grasped the notion of collective responsibility and highlighted its centrality:(Q12) I only see it as collective. My individual practice is not enough. Resistance is something that happens and is transmitted by all social groups. In a globalized society and economy where people travel, you cannot talk only about individual actions, subjectively leading to relative results. So obviously the society must make judicious antibiotic use because overconsumption is creating the problem. (G3M3)Furthermore, physicians were held to have a special responsibility because of their profession. Patients were also considered to have their share of responsibility, especially in the prescribing process:(Q13) When we do use and not misuse, we have a better result. Now to what extent can I be responsible for another patient? I believe that the issue is purely a matter of physician responsibility because each of us individually cannot make such a decision. (G3M2)(Q14) You cannot force a prescription on an adult. If antibiotics are a necessity, then both the patient and the physician should be really responsible. (G1W4)Participants held every individual responsible for ABR based on the idea that everyone contributes to it:Under ‘scepticism’ are categorized participants’ expressions of disbelief about the prospects of effectively managing ABR.(Q15) The mindset towards antibiotics should change but unfortunately, that is not feasible for Greece, I am referring to the corruption in the health system. (G4M1)Some participants voiced the concern that the use of antibiotics in meat and dairy production is uncontrollable, actually overshadowing human medical use:(Q16) I was surprised that there are animals in big farms crammed and fed with antibiotics. I did not expect it, personal use is possible to be controlled, but I think you cannot control this. Therefore, possibly this is the biggest problem. (G4M3)Corruption in the Greek healthcare system was regarded as an obstacle to the adaption of measures for the mitigation of ABR in the country:Under ‘alternative practices’ are categorized specific attitudes towards antibiotic treatment of participants who expressed their preference for alternatives to it.(Q17) I looked it up on my own and found health practices that daily help me to deal with the health problems I have/had. I'm trying not to get to the point of taking antibiotics for an infection. (G4W1)The use of other medical remedies but also homeopathic pills was mentioned as preferred over antibiotics, which were considered as the final option:(Q18) I use homeopathic pills for my children and myself. Antibiotics are my last resort. (G2W4)It was suggested that alternative health practices exist, which help preventing or coping with bacterial infections and avoiding antibiotics:Against the background of comparatively high levels of antibiotic use in Greece and limited understanding of related socio-cultural factors, we explored Greeks’ attitudes, perceived norms, and values regarding antibiotics and ABR. While previous research has emphasized ignorance as a major driver of improper antibiotic use in Greece and recommended informing prescribers and the public in response –9, our fIn general, participants recognized the severity of ABR, but stressed the complexity and difficulty of effectively mitigating it. They were sensitive to the tension between individuals’ interests and the common good that is often considered central to ABR mitigation , 38, genSome participants mentioned resorting to alternative health practices. This may be due to anti-medicine or anti-establishment convictions , 48 or, Some comparisons can be made with similar studies performed in other countries with relatively high rates of antibiotic use and antibiotic resistance in the WHO European region . The cenIn comparison to previous studies performed in Sweden (the interview guide used in this study was adapted from the Swedish research) , 32, somThis study reinforces the growing recognition of the importance of norms and values for antibiotic use and related mitigation strategies , 15 and Some limitations should be noted. In general, the results from a qualitative study are not strictly speaking representative of the studied population as a whole. Moreover, over half of the participants were from Northern Greece, most had received university-level education, and the oldest was 55 years. It is possible that a more heterogenous sample would have yielded more nuanced data, potentially affecting the relative prominence of certain themes. In addition, the focus group design together with the online format may have limited some participants’ willingness to interact and communicate. However, such effects were counteracted by limiting group size and posing probing questions. In addition to the relative homogeneity of the sample and the online setting, the structure of the interview guide and the informational video shown 10 minutes into discussions may have contributed to delimit the space within which the discussions happened, partly explaining the relatively early data saturation. The fact that FGDs took place during the COVID-19 pandemic may have affected the views expressed by the participants, in particular their disbelief about the prospects for effectively curbing ABR.This study highlights a complex set of attitudes, perceived norms and values that, besides limited knowledge, potentially contribute to non-judicious antibiotic use in Greek society. These include an experienced social expectation to retain inappropriate practices, reluctance to accept the imposition of burdens on individuals in the pursuit of collective goals, scepticism about the effectiveness of individual efforts in a context where institutional actors are seen as shirking their responsibilities, and a willingness to engage in alternative health practices. Future research, including quantitative studies, is needed to gain a more detailed understanding of the role of these and other socio-cultural determinants of antibiotic resistance in Greece and to design effective, contextually sensitive mitigation measures.Additional file 1. Interview guide.Additional file 2. COREQ


Streptomyces coelicolor, where specialized cells are generated through large‐scale genomic re‐organization. We investigate how the evolution of a genome architecture enables such mutation‐driven division of labor, using a multiscale computational model of bacterial evolution. In this model, bacterial behavior—antibiotic production or replication—is determined by the structure and composition of their genome, which encodes antibiotics, growth‐promoting genes, and fragile genomic loci that can induce chromosomal deletions. We find that a genomic organization evolves, which partitions growth‐promoting genes and antibiotic‐coding genes into distinct parts of the genome, separated by fragile genomic loci. Mutations caused by these fragile sites mostly delete growth‐promoting genes, generating sterile, and antibiotic‐producing mutants from weakly‐producing progenitors, in agreement with experimental observations. This division of labor enhances the competition between colonies by promoting antibiotic diversity. These results show that genomic organization can co‐evolve with genomic instabilities to enable reproductive division of labor.Division of labor can evolve when social groups benefit from the functional specialization of its members. Recently, a novel means of coordinating the division of labor was found in the antibiotic‐producing bacterium  Streptomyces reveals that if antibiotic production trades off with replication, genome architecture can evolve to support a mutation‐driven division of labor.A computational model of the multicellular bacteria  Similar divisions of labor are found in colonies of social insects, where one or few individuals are responsible for all of the reproduction, whereas the other individuals perform tasks that mirror those of somatic cells in multicellular organisms. Recently, several striking examples of reproductive and other divisions of labor have been described in the microbial world —after which we assume that resources are depleted and colonies sporulate. We model sporulation by randomly selecting a small fraction ξ of the bacteria (each cell is chosen with the same per‐capita probability), which will seed the next growth cycle. Spores do not disperse between cycles . In practice, we end the growth cycle by killing at random a fraction 1−ξ of all the cells, leaving the remaining cells to initiate the next growth cycle.We model individual nies Fig . ColonieS. coelicolor, as follows. We model the Streptomyces linear chromosome with a beads‐on‐a‐string model, which represents genomes as a linear sequence of genes and other genetic elements . In summary, this assumption allows us to study the genetic contributions to the division of labor and neglect regulatory contributions. It results in a trade‐off because bacteria cannot simultaneously maximize both growth and antibiotic production, since the number of growth‐promoting genes g trades antibiotic production for replication. For low values of βg, both growth and antibiotic production can be optimized at once, corresponding to an unrealistic situation where bacteria have arbitrary amounts of available energy. Therefore, βg must be sufficiently large to make the model realistic.We make the simplifying assumption that the metabolic strategy of each cell, i.e., the amount of resources dedicated to growth vs. antibiotic production, is determined solely by its genotype—and ignore that these strategies may be regulated by density‐dependent or other secreted cues from other bacteria  in the model, the distinction between centromeric and telomeric regions depends solely on fragile sites, whereas Streptomyces telomeres are specific genomic structures that can be recognized independently from the mutational dynamics; (iii) in Streptomyces, fragile site‐induced mutations transiently cause a complex interplay of duplications and deletions, which only after several mutational events result in the large‐scale elimination of DNA  duplications and deletions with probability some Fig . We set τs=2500 time steps. Over this time, bacteria incorporate about 10 fragile sites in their genome  without fragile sites, containing homogeneously distributed antibiotic genes and growth‐promoting genes, we let bacteria evolve over at least 500 vegetative growth cycles, with each growth cycle consisting of nome Fig , evolve nome Fig . After e100×103 time steps), antibiotics are collectively produced by the whole population. By contrast, at a later stage  nearly all antibiotics are produced by a small fraction of the bacteria . We also observe that replication and antibiotic production are performed by genetically distinct bacteria in the later stages: Bacteria with few growth‐promoting genes produce most antibiotics but do not replicate frequently, whereas bacteria with a larger number of growth‐promoting genes replicate frequently but do not produce antibiotics , under shallow trade‐off conditions  are pitted against generalists with a broad range of antibiotic production and growth rates. These “artificial” generalists do not pay a fitness cost for being able to replicate and produce antibiotics at the same time, and thus do not experience a trade‐off. A generalist with similar growth and antibiotic production rate to a wildtype only barely outperforms the wildtype . We observed that the genome of the evolved colony founder had two distinct regions: a region on the right of the chromosome that contains growth‐promoting genes but lacks fragile sites, and a region on the left side that lacks growth‐promoting genes and has abundant fragile sites Fig . After cTo assess whether genome architecture is required for division of labor, we initiated an evolutionary run where we shuffled the order of the genes in the genome of each spore at the beginning of every growth cycle. This disrupts the genomic architecture of colony founders without changing its composition, in terms of the number and type of genes and fragile sites. Starting from an evolved genome, we observe a rapid decrease in antibiotic genes and in growth‐promoting genes, resulting in small genomes Fig . At an enh=10 copies for the cell to remain alive. Any number of genes lower than nh is lethal and the bacterium carrying such genome is removed from the lattice, while a larger number is neutral. We find that the mutation‐driven division of labor evolves, and that the centromeric part of the evolved genomes contains always more than nh homeostatic genes, ensuring the survival of the antibiotic‐producing species that arise through mutation . We modeled these genes as an additional gene type and assumed that they must be present in at least et al, et al, et al, et al, ≥1000 time steps; Appendix Fig de novo from a simulation initialized with random genomes  core region located at their centromere and telomeric regions, which are prone to various forms of recombination and instability and contain much of the accessory genes . This suggests that the sensing of antibiotics increases mutation rates at contact points between colonies, so that mutants appear preferentially where colonies compete with one another. Investigating the consequences of this additional layer of evolutionary dynamics will be interesting in future extensions of the model. A more complete model of the Streptomyces genome could also include both telomeres and their complex recombination dynamics  unless explicitly stated.The model is a lattice‐based stochastic simulation system. We consider a population of bacteria that can replicate and produce antibiotics. The model is inspired by the life cycle of Λ1 and Λ2. Both lattices have size L×L and toroidal boundaries to avoid edge effects. Every site of Λ1 can either be occupied by one bacterium or be empty. Every site of Λ2 can be occupied by multiple antibiotics. Each bacterium possesses a genome that determines its replication rate, as well as the rate and type of antibiotics it produces and is resistant to. The genome consists of a linear sequence of three types of genetic elements :hg controls the curvature of the function and is equal to the number of growth‐promoting genes producing half maximum replication rate: Ghg=αg/2. We expect that other functions that phenomenologically reproduce G, e.g., an exponential function of the form Gg=αg1−exp−g/hg, will not change the results.Replication rate R depends on whether a genome has at least an antibiotic gene sufficiently similar to the antibiotics present on Λ2 at the corresponding location of the bacterium, where similarity is determined from the bit‐strings of the antibiotics and the antibiotic genes. For each antibiotic n on Λ2, the Hamming Distance D (the number of different bits) between the antibiotic and the gene with the minimum distance from n is calculated. All these minimum distances are summed into a susceptibility score S=∑D, and the overall resistance is a decreasing function of S:D=0, and thus R=1. Moreover, the Gaussian function ensures that small mutations in antibiotic types do not decrease resistance too rapidly.Antibiotic resistance Λ1. These bacteria compete on the basis of their probability of replication. For each competing bacterium i, the probability of replication is kreplicationi/η, with η the neighborhood size . Dividing by η scales the probability of replication so that a bacterium with maximum replication rate surrounded by empty sites replicates on average once per time step. Upon replication, the new bacterium inherits the genome of its parent, with possible mutations.Replication occurs when bacteria are adjacent to an empty site on Mutations occur during replication, and can expand or shrink genomes, and diversify antibiotics.μd. When a gene or a fragile site is duplicated, the copy is inserted at a random genomic location. This ensures that gene clustering is not a trivial consequence of neutral mutational dynamics, and must instead be selected upon to evolve. We expect that there is an upper limit for μd above which the genome becomes effectively scrambled (therefore no genome structure could evolve) because fast duplications randomize gene position and fast deletions decrease local genomic correlations.Duplication and deletion of genes and fragile sites occur with equal per‐gene probability μf. We take into account that large‐scale mutations in Streptomyces preferentially disrupt telomeric regions  probability μa. This changes the antibiotic type, and thus the antibiotic repertoire of the bacterium.The antibiotic genes consist of a genetic sequence modeled as a bit‐string of length Streptomyces. Because they are easily copied (or translocated) we assume that they can also be spontaneously generated with a small probability μn (independent of genome size). The new fragile site is inserted at a random location in the genome.Fragile genomic sites, such as inverted repeats or transposable elements are common in the genome of A of the number of antibiotic genes a a bacterium has. In analogy with the function derived for replication, we model antibiotic production as a Hill function. At the same time, production is strongly inhibited by growth‐promoting genes—a function Ig with g the number of growth‐promoting genes in the genome, in accordance with a likely trade‐off between growth and secondary metabolism . Analogous to the Gillespie algorithm . This makes antibiotic production independent from the deposition radius and allows us to compare results across simulations with different ra. If a bacterium has multiple antibiotic genes, each antibiotic deposited is chosen randomly from the antibiotic genes with uniform probability.Antibiotic production rate is modeled as an increasing function 1−R, where R is the bacterial resistance to the antibiotic defined above. When a bacterium dies, it is removed from the lattice, leaving behind an empty site.Death can occur when a bacterium is sensitive to an antibiotic located at the same site as the bacterium itself. The probability of a bacterium dying is calculated as pmov of moving if there is an empty site adjacent to them. This speeds up colony expansion and competition between colonies and avoids strong grid effects that could make spatial patterns too rigid.Bacteria have a small probability t=0 a small population of spores is seeded on the lattice. The initial spores have a small genome of length 10, i.e., a random sequence of growth‐promoting genes and antibiotic genes (of random types) but no fragile sites. The lattice is updated asynchronously: Over one time step, each lattice site is updated in random order.Unless differently specified, at time et al, The source code is written in c and uses the CASH libraries (de Boer & Staritsky, Enrico Sandro Colizzi: Conceptualization; software; formal analysis; investigation; methodology; writing – original draft; writing – review and editing. Bram van Dijk: Conceptualization; software; investigation; methodology; writing – original draft; writing – review and editing. Roeland M H Merks: Supervision; investigation; methodology; writing – review and editing. Daniel E Rozen: Supervision; investigation; methodology; writing – review and editing. Renske M A Vroomans: Conceptualization; software; supervision; investigation; methodology; writing – review and editing.The authors declare that they have no conflict of interest.Appendix S1Click here for additional data file.Movie EV1Click here for additional data file.Movie EV2Click here for additional data file.