Pub Date : 2024-08-21Epub Date: 2024-07-31DOI: 10.1128/aem.00795-24
Robin Bonné, Ian P G Marshall, Jesper J Bjerg, Ugo Marzocchi, Jean Manca, Lars Peter Nielsen, Kartik Aiyer
Cable bacteria are filamentous bacteria that couple the oxidation of sulfide in sediments to the reduction of oxygen via long-distance electron transport over centimeter distances through periplasmic wires. However, the capability of cable bacteria to perform extracellular electron transfer to acceptors, such as electrodes, has remained elusive. In this study, we demonstrate that living cable bacteria actively move toward electrodes in different bioelectrochemical systems. Carbon felt and carbon fiber electrodes poised at +200 mV attracted live cable bacteria from the sediment. When the applied potential was switched off, cable bacteria retracted from the electrode. qPCR and scanning electron microscopy corroborated this finding and revealed cable bacteria in higher abundance present on the electrode surface compared with unpoised controls. These experiments raise new possibilities to study metabolism of cable bacteria and cultivate them in bioelectrochemical devices for bioelectronic applications, such as biosensing and bioremediation.
Importance: Extracellular electron transfer is a metabolic function associated with electroactive bacteria wherein electrons are exchanged with external electron acceptors or donors. This feature has enabled the development of several applications, such as biosensing, carbon capture, and energy recovery. Cable bacteria are a unique class of long, filamentous microbes that perform long-distance electron transport in freshwater and marine sediments. In this study, we demonstrate the attraction of cable bacteria toward carbon electrodes and demonstrate their potential electroactivity. This finding enables electronic control and monitoring of the metabolism of cable bacteria and may, in turn, aid in the development of bioelectronic applications.
{"title":"Interaction of living cable bacteria with carbon electrodes in bioelectrochemical systems.","authors":"Robin Bonné, Ian P G Marshall, Jesper J Bjerg, Ugo Marzocchi, Jean Manca, Lars Peter Nielsen, Kartik Aiyer","doi":"10.1128/aem.00795-24","DOIUrl":"10.1128/aem.00795-24","url":null,"abstract":"<p><p>Cable bacteria are filamentous bacteria that couple the oxidation of sulfide in sediments to the reduction of oxygen via long-distance electron transport over centimeter distances through periplasmic wires. However, the capability of cable bacteria to perform extracellular electron transfer to acceptors, such as electrodes, has remained elusive. In this study, we demonstrate that living cable bacteria actively move toward electrodes in different bioelectrochemical systems. Carbon felt and carbon fiber electrodes poised at +200 mV attracted live cable bacteria from the sediment. When the applied potential was switched off, cable bacteria retracted from the electrode. qPCR and scanning electron microscopy corroborated this finding and revealed cable bacteria in higher abundance present on the electrode surface compared with unpoised controls. These experiments raise new possibilities to study metabolism of cable bacteria and cultivate them in bioelectrochemical devices for bioelectronic applications, such as biosensing and bioremediation.</p><p><strong>Importance: </strong>Extracellular electron transfer is a metabolic function associated with electroactive bacteria wherein electrons are exchanged with external electron acceptors or donors. This feature has enabled the development of several applications, such as biosensing, carbon capture, and energy recovery. Cable bacteria are a unique class of long, filamentous microbes that perform long-distance electron transport in freshwater and marine sediments. In this study, we demonstrate the attraction of cable bacteria toward carbon electrodes and demonstrate their potential electroactivity. This finding enables electronic control and monitoring of the metabolism of cable bacteria and may, in turn, aid in the development of bioelectronic applications.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337825/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141854573","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-21Epub Date: 2024-07-31DOI: 10.1128/aem.00988-24
Siqi Li, Weiping Zhang
Shigella bacteria utilize the type III secretion system (T3SS) to invade host cells and establish local infection. Invasion plasmid antigen D (IpaD), a component of Shigella T3SS, has garnered extensive interest as a vaccine target, primarily due to its pivotal role in the Shigella invasion, immunogenic property, and a high degree of conservation across Shigella species and serotypes. Currently, we are developing an epitope- and structure-based multivalent vaccine against shigellosis and require functional epitope antigens of key Shigella virulence determinants including IpaD. However, individual IpaD B-cell epitopes, their contributions to the overall immunogenicity, and functional activities attributing to bacteria invasion have not been fully characterized. In this study, we predicted continuous B-cell epitopes in silico and fused each epitope to a carrier protein. Then, we immunized mice intramuscularly with each epitope fusion protein, examined the IpaD-specific antibody responses, and measured antibodies from each epitope fusion for the activity against Shigella invasion in vitro. Data showed that all epitope fusion proteins induced similar levels of anti-IpaD IgG antibodies in mice, and differences were noted for antibody inhibition activity against Shigella invasion. IpaD epitope 1 (SPGGNDGNSV), IpaD epitope 2 (LGGNGEVVLDNA), and IpaD epitope 5 (SPNNTNGSSTET) induced antibodies significantly better in inhibiting invasion from Shigella flexneri 2a, and epitopes 1 and 5 elicited antibodies more effectively at preventing invasion of Shigella sonnei. These results suggest that IpaD epitopes 1 and 5 can be the IpaD representative antigens for epitope-based polyvalent protein construction and protein-based cross-protective Shigella vaccine development.IMPORTANCEShigella is a leading cause of diarrhea in children younger than 5 years in developing countries (children's diarrhea) and continues to be a major threat to public health. No licensed vaccines are currently available against the heterogeneous Shigella species and serotype strains. Aiming to develop a cross-protective multivalent vaccine against shigellosis and dysentery, we applied novel multiepitope fusion antigen (MEFA) technology to construct a broadly immunogenic polyvalent protein antigen, by presenting functional epitopes of multiple Shigella virulence determinants on a backbone protein. The functional IpaD epitopes identified from this study will essentially allow us to construct an optimal polyvalent Shigella immunogen, leading to the development of a cross-protective vaccine against shigellosis (and dysentery) and the improvement of global health. In addition, identifying functional epitopes from heterogeneous virulence determinants and using them as antigenic representatives for the development of cross-protective multivalent vaccines can be applied generally in
{"title":"Mapping the functional B-cell epitopes of <i>Shigella</i> invasion plasmid antigen D (IpaD).","authors":"Siqi Li, Weiping Zhang","doi":"10.1128/aem.00988-24","DOIUrl":"10.1128/aem.00988-24","url":null,"abstract":"<p><p><i>Shigella</i> bacteria utilize the type III secretion system (T3SS) to invade host cells and establish local infection. Invasion plasmid antigen D (IpaD), a component of <i>Shigella</i> T3SS, has garnered extensive interest as a vaccine target, primarily due to its pivotal role in the <i>Shigella</i> invasion, immunogenic property, and a high degree of conservation across <i>Shigella</i> species and serotypes. Currently, we are developing an epitope- and structure-based multivalent vaccine against shigellosis and require functional epitope antigens of key <i>Shigella</i> virulence determinants including IpaD. However, individual IpaD B-cell epitopes, their contributions to the overall immunogenicity, and functional activities attributing to bacteria invasion have not been fully characterized. In this study, we predicted continuous B-cell epitopes <i>in silico</i> and fused each epitope to a carrier protein. Then, we immunized mice intramuscularly with each epitope fusion protein, examined the IpaD-specific antibody responses, and measured antibodies from each epitope fusion for the activity against <i>Shigella</i> invasion <i>in vitro</i>. Data showed that all epitope fusion proteins induced similar levels of anti-IpaD IgG antibodies in mice, and differences were noted for antibody inhibition activity against <i>Shigella</i> invasion. IpaD epitope 1 (SPGGNDGNSV), IpaD epitope 2 (LGGNGEVVLDNA), and IpaD epitope 5 (SPNNTNGSSTET) induced antibodies significantly better in inhibiting invasion from <i>Shigella flexneri</i> 2a, and epitopes 1 and 5 elicited antibodies more effectively at preventing invasion of <i>Shigella sonnei</i>. These results suggest that IpaD epitopes 1 and 5 can be the IpaD representative antigens for epitope-based polyvalent protein construction and protein-based cross-protective <i>Shigella</i> vaccine development.IMPORTANCE<i>Shigella</i> is a leading cause of diarrhea in children younger than 5 years in developing countries (children's diarrhea) and continues to be a major threat to public health. No licensed vaccines are currently available against the heterogeneous <i>Shigella</i> species and serotype strains. Aiming to develop a cross-protective multivalent vaccine against shigellosis and dysentery, we applied novel multiepitope fusion antigen (MEFA) technology to construct a broadly immunogenic polyvalent protein antigen, by presenting functional epitopes of multiple <i>Shigella</i> virulence determinants on a backbone protein. The functional IpaD epitopes identified from this study will essentially allow us to construct an optimal polyvalent <i>Shigella</i> immunogen, leading to the development of a cross-protective vaccine against shigellosis (and dysentery) and the improvement of global health. In addition, identifying functional epitopes from heterogeneous virulence determinants and using them as antigenic representatives for the development of cross-protective multivalent vaccines can be applied generally in","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337796/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141854574","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Despite their low quantity and abundance, the cellulolytic bacteria that inhabit the equine large intestine are vital to their host, as they enable the crucial use of forage-based diets. Fibrobacter succinogenes is one of the most important intestinal cellulolytic bacteria. In this study, Fibrobacter sp. HC4, one cellulolytic strain newly isolated from the horse cecum, was characterized for its ability to utilize plant cell wall fibers. Fibrobacter sp. HC4 consumed only cellulose, cellobiose, and glucose and produced succinate and acetate in equal amounts. Among genes coding for CAZymes, 26% of the detected glycoside hydrolases (GHs) were involved in cellulolysis. These cellulases belong to the GH5, GH8, GH9, GH44, GH45, and GH51 families. Both carboxymethyl cellulase and xylanase activities of Fibrobacter sp. HC4 were detected using the Congo red method and were higher than those of F. succinogenes S85, the type strain. The in vitro addition of Fibrobacter sp. HC4 to a fecal microbial ecosystem of horses with large intestinal acidosis significantly enhanced fibrolytic activity as measured by the increase in gas and volatile fatty acids production during the first 48 h. According to this, the pH decreased and the disappearance of dry matter increased at a faster rate with Fibrobacter sp. HC4. Our data suggest a high specialization of the new strain in cellulose degradation. Such a strain could be of interest for future exploitation of its probiotic potential, which needs to be further determined by in vivo studies.IMPORTANCECellulose is the most abundant of plant cell wall fiber and can only be degraded by the large intestine microbiota, resulting in the production of volatile fatty acids that are essential for the host nutrition and health. Consequently, cellulolytic bacteria are of major importance to herbivores. However, these bacteria are challenged by various factors, such as high starch diets, which acidify the ecosystem and reduce their numbers and activity. This can lead to an imbalance in the gut microbiota and digestive problems such as colic, a major cause of mortality in horses. In this work, we characterized a newly isolated cellulolytic strain, Fibrobacter sp. HC4, from the equine intestinal microbiota. Due to its high cellulolytic capacity, reintroduction of this strain into an equine fecal ecosystem stimulates hay fermentation in vitro. Isolating and describing cellulolytic bacteria is a prerequisite for using them as probiotics to restore intestinal balance.
{"title":"<i>Fibrobacter</i> sp. HC4, a newly isolated strain, demonstrates a high cellulolytic activity as revealed by enzymatic measurements and <i>in vitro</i> assay.","authors":"Alicia Froidurot, Emmanuel Jacotot, Samy Julliand, Pauline Grimm, Véronique Julliand","doi":"10.1128/aem.00514-24","DOIUrl":"10.1128/aem.00514-24","url":null,"abstract":"<p><p>Despite their low quantity and abundance, the cellulolytic bacteria that inhabit the equine large intestine are vital to their host, as they enable the crucial use of forage-based diets. <i>Fibrobacter succinogenes</i> is one of the most important intestinal cellulolytic bacteria. In this study, <i>Fibrobacter</i> sp. HC4, one cellulolytic strain newly isolated from the horse cecum, was characterized for its ability to utilize plant cell wall fibers. <i>Fibrobacter</i> sp. HC4 consumed only cellulose, cellobiose, and glucose and produced succinate and acetate in equal amounts. Among genes coding for CAZymes, 26% of the detected glycoside hydrolases (GHs) were involved in cellulolysis. These cellulases belong to the GH5, GH8, GH9, GH44, GH45, and GH51 families. Both carboxymethyl cellulase and xylanase activities of <i>Fibrobacter</i> sp. HC4 were detected using the Congo red method and were higher than those of <i>F. succinogenes</i> S85, the type strain. The <i>in vitro</i> addition of <i>Fibrobacter</i> sp. HC4 to a fecal microbial ecosystem of horses with large intestinal acidosis significantly enhanced fibrolytic activity as measured by the increase in gas and volatile fatty acids production during the first 48 h. According to this, the pH decreased and the disappearance of dry matter increased at a faster rate with <i>Fibrobacter</i> sp. HC4. Our data suggest a high specialization of the new strain in cellulose degradation. Such a strain could be of interest for future exploitation of its probiotic potential, which needs to be further determined by <i>in vivo</i> studies.IMPORTANCECellulose is the most abundant of plant cell wall fiber and can only be degraded by the large intestine microbiota, resulting in the production of volatile fatty acids that are essential for the host nutrition and health. Consequently, cellulolytic bacteria are of major importance to herbivores. However, these bacteria are challenged by various factors, such as high starch diets, which acidify the ecosystem and reduce their numbers and activity. This can lead to an imbalance in the gut microbiota and digestive problems such as colic, a major cause of mortality in horses. In this work, we characterized a newly isolated cellulolytic strain, <i>Fibrobacter</i> sp. HC4, from the equine intestinal microbiota. Due to its high cellulolytic capacity, reintroduction of this strain into an equine fecal ecosystem stimulates hay fermentation <i>in vitro</i>. Isolating and describing cellulolytic bacteria is a prerequisite for using them as probiotics to restore intestinal balance.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337828/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141854566","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Second Correction for Gao et al., \"VqsA, a Novel LysR-Type Transcriptional Regulator, Coordinates Quorum Sensing (QS) and Is Controlled by QS To Regulate Virulence in the Pathogen <i>Vibrio alginolyticus</i>\".","authors":"Xiating Gao, Xuetong Wang, Qiaoqiao Mao, Rongjing Xu, Xiaohui Zhou, Yue Ma, Qin Liu, Yuanxing Zhang, Qiyao Wang","doi":"10.1128/aem.00867-24","DOIUrl":"10.1128/aem.00867-24","url":null,"abstract":"","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337822/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141615796","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-21Epub Date: 2024-07-16DOI: 10.1128/aem.00354-24
Kristina Ceres, Jordan D Zehr, Chloe Murrell, Jean K Millet, Qi Sun, Holly C McQueary, Alanna Horton, Casey Cazer, Kelly Sams, Guillaume Reboul, William B Andreopoulos, Patrick K Mitchell, Renee Anderson, Rebecca Franklin-Guild, Brittany D Cronk, Bryce J Stanhope, Claire R Burbick, Rebecca Wolking, Laura Peak, Yan Zhang, Rebeccah McDowall, Aparna Krishnamurthy, Durda Slavic, Prabhjot Kaur Sekhon, Gregory H Tyson, Olgica Ceric, Michael J Stanhope, Laura B Goodman
Infections caused by antimicrobial-resistant Escherichia coli are the leading cause of death attributed to antimicrobial resistance (AMR) worldwide, and the known AMR mechanisms involve a range of functional proteins. Here, we employed a pan-genome wide association study (GWAS) approach on over 1,000 E. coli isolates from sick dogs collected across the US and Canada and identified a strong statistical association (empirical P < 0.01) of AMR, involving a range of antibiotics to a group 1 capsular (CPS) gene cluster. This cluster included genes under relaxed selection pressure, had several loci missing, and had pseudogenes for other key loci. Furthermore, this cluster is widespread in E. coli and Klebsiella clinical isolates across multiple host species. Earlier studies demonstrated that the octameric CPS polysaccharide export protein Wza can transmit macrolide antibiotics into the E. coli periplasm. We suggest that the CPS in question, and its highly divergent Wza, functions as an antibiotic trap, preventing antimicrobial penetration. We also highlight the high diversity of lineages circulating in dogs across all regions studied, the overlap with human lineages, and regional prevalence of resistance to multiple antimicrobial classes.
Importance: Much of the human genomic epidemiology data available for E. coli mechanism discovery studies has been heavily biased toward shiga-toxin producing strains from humans and livestock. E. coli occupies many niches and produces a wide variety of other significant pathotypes, including some implicated in chronic disease. We hypothesized that since dogs tend to share similar strains with their owners and are treated with similar antibiotics, their pathogenic isolates will harbor unexplored AMR mechanisms of importance to humans as well as animals. By comparing over 1,000 genomes with in vitro antimicrobial susceptibility data from sick dogs across the US and Canada, we identified a strong multidrug resistance association with an operon that appears to have once conferred a type 1 capsule production system.
抗菌性大肠杆菌引起的感染是全球抗菌药耐药性(AMR)导致死亡的主要原因,而已知的 AMR 机制涉及一系列功能蛋白。在这里,我们采用了一种泛基因组关联研究(GWAS)方法,对从美国和加拿大各地收集的 1000 多份病犬大肠杆菌分离物进行了研究,并确定了 AMR 与第 1 组囊状(CPS)基因簇的强统计学关联(经验 P <0.01),其中涉及一系列抗生素。该基因簇包括处于宽松选择压力下的基因,有几个基因位点缺失,并有其他关键基因位点的假基因。此外,该基因簇广泛存在于多个宿主物种的大肠杆菌和克雷伯氏菌临床分离株中。早先的研究表明,八聚体 CPS 多糖输出蛋白 Wza 可将大环内酯类抗生素传输到大肠杆菌的外质中。我们认为,相关的 CPS 及其高度分化的 Wza 具有抗生素陷阱的功能,可阻止抗菌素渗透。我们还强调了在所研究的所有地区,狗体内流行的菌系的高度多样性、与人类菌系的重叠,以及对多种抗菌药物的耐药性的地区流行性:重要意义:用于大肠杆菌机制发现研究的人类基因组流行病学数据大多偏重于人类和家畜中产生志贺毒素的菌株。大肠杆菌占据了许多壁龛,并产生了多种其他重要的病原体,包括一些与慢性疾病有关的病原体。我们假设,由于狗往往与主人分享类似的菌株,并使用类似的抗生素进行治疗,因此它们的致病分离物将蕴藏着对人类和动物都很重要的、尚未探索的 AMR 机制。通过比较来自美国和加拿大病犬的 1000 多个基因组和体外抗菌药敏感性数据,我们发现了一个与操作子密切相关的多重耐药性,该操作子似乎曾经赋予了 1 型胶囊生产系统。
{"title":"Evolutionary genomic analyses of canine <i>E. coli</i> infections identify a relic capsular locus associated with resistance to multiple classes of antimicrobials.","authors":"Kristina Ceres, Jordan D Zehr, Chloe Murrell, Jean K Millet, Qi Sun, Holly C McQueary, Alanna Horton, Casey Cazer, Kelly Sams, Guillaume Reboul, William B Andreopoulos, Patrick K Mitchell, Renee Anderson, Rebecca Franklin-Guild, Brittany D Cronk, Bryce J Stanhope, Claire R Burbick, Rebecca Wolking, Laura Peak, Yan Zhang, Rebeccah McDowall, Aparna Krishnamurthy, Durda Slavic, Prabhjot Kaur Sekhon, Gregory H Tyson, Olgica Ceric, Michael J Stanhope, Laura B Goodman","doi":"10.1128/aem.00354-24","DOIUrl":"10.1128/aem.00354-24","url":null,"abstract":"<p><p>Infections caused by antimicrobial-resistant <i>Escherichia coli</i> are the leading cause of death attributed to antimicrobial resistance (AMR) worldwide, and the known AMR mechanisms involve a range of functional proteins. Here, we employed a pan-genome wide association study (GWAS) approach on over 1,000 <i>E. coli</i> isolates from sick dogs collected across the US and Canada and identified a strong statistical association (empirical <i>P</i> < 0.01) of AMR, involving a range of antibiotics to a group 1 capsular (CPS) gene cluster. This cluster included genes under relaxed selection pressure, had several loci missing, and had pseudogenes for other key loci. Furthermore, this cluster is widespread in <i>E. coli</i> and <i>Klebsiella</i> clinical isolates across multiple host species. Earlier studies demonstrated that the octameric CPS polysaccharide export protein Wza can transmit macrolide antibiotics into the <i>E. coli</i> periplasm. We suggest that the CPS in question, and its highly divergent Wza, functions as an antibiotic trap, preventing antimicrobial penetration. We also highlight the high diversity of lineages circulating in dogs across all regions studied, the overlap with human lineages, and regional prevalence of resistance to multiple antimicrobial classes.</p><p><strong>Importance: </strong>Much of the human genomic epidemiology data available for <i>E. coli</i> mechanism discovery studies has been heavily biased toward shiga-toxin producing strains from humans and livestock. <i>E. coli</i> occupies many niches and produces a wide variety of other significant pathotypes, including some implicated in chronic disease. We hypothesized that since dogs tend to share similar strains with their owners and are treated with similar antibiotics, their pathogenic isolates will harbor unexplored AMR mechanisms of importance to humans as well as animals. By comparing over 1,000 genomes with <i>in vitro</i> antimicrobial susceptibility data from sick dogs across the US and Canada, we identified a strong multidrug resistance association with an operon that appears to have once conferred a type 1 capsule production system.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337803/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141619123","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
We developed a nondestructive three-dimensional microbial visualization method utilizing synchrotron radiation X-ray microscale computed tomography to better understand the relationship between microorganisms and their surrounding habitats. The method was tested and optimized using a mixture of axenic Escherichia coli and Comamonas testosteroni. The osmium-thiocarbohydrazide-osmium method was used to stain all the microbial cells, and gold in situ hybridization was used to detect specific phylogenetic microbial groups. The stained samples were embedded in epoxy resin for microtomographic analysis. Differences in X-ray absorbances were calculated by subtracting the pre-L3-edge images from the post-L3-edge images to visualize the osmium and gold signals. Although we successfully detected cells stained with osmium, those labeled with gold were not detected, probably because of the insufficient density of gold atoms in the microbial cells. We then applied the developed technique to anaerobic granules and visualized the distribution of microbial cells and extracellular polymeric substances. Empty spaces were highlighted to determine the cavity distribution in granules. Numerous independent cavities of different sizes were identified in the granules. The developed method can be applied to various environmental samples for deeper insights into microbial life in their habitats.
Importance: Microorganisms inhabit diverse environments and often form biofilms. One factor that affects their community structure is the surrounding physical environment. The arrangement of residential space within the formed biofilm plays a crucial role in the supply and transportation of substances, as well as the discharge of metabolites. Conventional approaches, such as scanning electron microscopy and confocal laser scanning microscopy combined with fluorescence in situ hybridization, have limitations as they provide information primarily from the biofilm surface and cross-sections. In this study, we developed a method for detecting microorganisms in biofilms using synchrotron radiation X-ray microscale computer tomography. The developed method allows nondestructive three-dimensional observation of biofilms at a single-cell resolution (voxel size of approximately 200 nm), facilitating an understanding of the relationship between microorganisms and their physical habitats.
我们利用同步辐射 X 射线微尺度计算机断层扫描技术开发了一种无损三维微生物可视化方法,以更好地了解微生物与其周围栖息地之间的关系。我们使用腋生大肠杆菌和睾丸酵母菌的混合物对该方法进行了测试和优化。锇-硫代羧肼-锇法用于染色所有微生物细胞,金原位杂交法用于检测特定的系统发育微生物群。染色后的样品嵌入环氧树脂中进行显微形貌分析。将 L3 边沿前的图像与 L3 边沿后的图像相减,计算 X 射线吸光度的差异,以观察锇和金的信号。虽然我们成功地检测到了被锇染色的细胞,但却没有检测到被金标记的细胞,这可能是因为微生物细胞中金原子的密度不够。然后,我们将开发的技术应用于厌氧颗粒,观察微生物细胞和细胞外聚合物质的分布。空隙被突出显示,以确定颗粒中的空腔分布。在颗粒中发现了许多不同大小的独立空腔。所开发的方法可用于各种环境样本,以深入了解微生物在其栖息地的生活情况:重要性:微生物栖息在不同的环境中,并经常形成生物膜。影响其群落结构的一个因素是周围的物理环境。已形成的生物膜内居住空间的排列对物质的供应和运输以及代谢物的排放起着至关重要的作用。传统的方法,如扫描电子显微镜和激光共聚焦扫描显微镜结合荧光原位杂交技术,主要从生物膜表面和横截面提供信息,具有一定的局限性。在这项研究中,我们开发了一种利用同步辐射 X 射线微尺度计算机断层成像技术检测生物膜中微生物的方法。所开发的方法可在单细胞分辨率(体素大小约为 200 纳米)下对生物膜进行无损三维观测,有助于了解微生物与其物理栖息地之间的关系。
{"title":"Nondestructive and three-dimensional visualization by identifying elements using synchrotron radiation microscale X-ray CT reveals microbial and cavity distributions in anaerobic granular sludge.","authors":"Kampachiro Urasaki, Yuki Morono, Go-Ichiro Uramoto, Kentaro Uesugi, Masahiro Yasutake, Manato Akishiba, Guangze Guo, Yu-You Li, Kengo Kubota","doi":"10.1128/aem.00563-24","DOIUrl":"10.1128/aem.00563-24","url":null,"abstract":"<p><p>We developed a nondestructive three-dimensional microbial visualization method utilizing synchrotron radiation X-ray microscale computed tomography to better understand the relationship between microorganisms and their surrounding habitats. The method was tested and optimized using a mixture of axenic <i>Escherichia coli</i> and <i>Comamonas testosteroni</i>. The osmium-thiocarbohydrazide-osmium method was used to stain all the microbial cells, and gold <i>in situ</i> hybridization was used to detect specific phylogenetic microbial groups. The stained samples were embedded in epoxy resin for microtomographic analysis. Differences in X-ray absorbances were calculated by subtracting the pre-L<sub>3</sub>-edge images from the post-L<sub>3</sub>-edge images to visualize the osmium and gold signals. Although we successfully detected cells stained with osmium, those labeled with gold were not detected, probably because of the insufficient density of gold atoms in the microbial cells. We then applied the developed technique to anaerobic granules and visualized the distribution of microbial cells and extracellular polymeric substances. Empty spaces were highlighted to determine the cavity distribution in granules. Numerous independent cavities of different sizes were identified in the granules. The developed method can be applied to various environmental samples for deeper insights into microbial life in their habitats.</p><p><strong>Importance: </strong>Microorganisms inhabit diverse environments and often form biofilms. One factor that affects their community structure is the surrounding physical environment. The arrangement of residential space within the formed biofilm plays a crucial role in the supply and transportation of substances, as well as the discharge of metabolites. Conventional approaches, such as scanning electron microscopy and confocal laser scanning microscopy combined with fluorescence in <i>situ</i> hybridization, have limitations as they provide information primarily from the biofilm surface and cross-sections. In this study, we developed a method for detecting microorganisms in biofilms using synchrotron radiation X-ray microscale computer tomography. The developed method allows nondestructive three-dimensional observation of biofilms at a single-cell resolution (voxel size of approximately 200 nm), facilitating an understanding of the relationship between microorganisms and their physical habitats.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337819/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141632460","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-21Epub Date: 2024-07-12DOI: 10.1128/aem.00553-24
Sriya Sunil, Tamara Walsky, Mikayla Henry, Leonie Kemmerling, Magdalena Pajor, Xiaodong Guo, Sarah I Murphy, Renata Ivanek, Martin Wiedmann
In the U.S., baby spinach is mostly produced in Arizona (AZ) and California (CA). Characterizing the impact of growing region on the bacterial quality of baby spinach can inform quality management practices in industry. Between December 2021 and December 2022, baby spinach was sampled after harvest and packaging for microbiological testing, including shelf-life testing of packaged samples that were stored at 4°C. Samples were tested to (i) determine bacterial concentration, and (ii) obtain and identify bacterial isolates. Packaged samples from the Salinas, CA, area (n = 13), compared to those from the Yuma, AZ, area (n = 9), had a significantly higher bacterial concentration, on average, by 0.78 log10 CFU/g (P < 0.01, based on aerobic, mesophilic plate count data) or 0.67 log10 CFU/g (P < 0.01, based on psychrotolerant plate count data); the bacterial concentrations of harvest samples from the Yuma and Salinas areas were not significantly different. Our data also support that an increase in preharvest temperature is significantly associated with an increase in the bacterial concentration on harvested and packaged spinach. A Fisher's exact test and linear discriminant analysis (effect size), respectively, demonstrated that (i) the genera of 2,186 bacterial isolates were associated (P < 0.01) with growing region and (ii) Pseudomonas spp. and Exiguobacterium spp. were enriched in spinach from the Yuma and Salinas areas, respectively. Our findings provide preliminary evidence that growing region and preharvest temperature may impact the bacterial quality of spinach and thus could inform more targeted strategies to manage produce quality.
Importance: In the U.S., most spinach is produced in Arizona (AZ) and California (CA) seasonally; typically, spinach is cultivated in the Yuma, AZ, area during the winter and in the Salinas, CA, area during the summer. As the bacterial quality of baby spinach can influence consumer acceptance of the product, it is important to assess whether the bacterial quality of baby spinach can vary between spinach-growing regions. The findings of this study provide insights that could be used to support region-specific quality management strategies for baby spinach. Our results also highlight the value of further evaluating the impact of growing region and preharvest temperature on the bacterial quality of different produce commodities.
{"title":"A longitudinal study on the bacterial quality of baby spinach cultivated in Arizona and California.","authors":"Sriya Sunil, Tamara Walsky, Mikayla Henry, Leonie Kemmerling, Magdalena Pajor, Xiaodong Guo, Sarah I Murphy, Renata Ivanek, Martin Wiedmann","doi":"10.1128/aem.00553-24","DOIUrl":"10.1128/aem.00553-24","url":null,"abstract":"<p><p>In the U.S., baby spinach is mostly produced in Arizona (AZ) and California (CA). Characterizing the impact of growing region on the bacterial quality of baby spinach can inform quality management practices in industry. Between December 2021 and December 2022, baby spinach was sampled after harvest and packaging for microbiological testing, including shelf-life testing of packaged samples that were stored at 4°C. Samples were tested to (i) determine bacterial concentration, and (ii) obtain and identify bacterial isolates. Packaged samples from the Salinas, CA, area (<i>n</i> = 13), compared to those from the Yuma, AZ, area (<i>n</i> = 9), had a significantly higher bacterial concentration, on average, by 0.78 log<sub>10</sub> CFU/g (<i>P</i> < 0.01, based on aerobic, mesophilic plate count data) or 0.67 log<sub>10</sub> CFU/g (<i>P</i> < 0.01, based on psychrotolerant plate count data); the bacterial concentrations of harvest samples from the Yuma and Salinas areas were not significantly different. Our data also support that an increase in preharvest temperature is significantly associated with an increase in the bacterial concentration on harvested and packaged spinach. A Fisher's exact test and linear discriminant analysis (effect size), respectively, demonstrated that (i) the genera of 2,186 bacterial isolates were associated (<i>P</i> < 0.01) with growing region and (ii) <i>Pseudomonas</i> spp. and <i>Exiguobacterium</i> spp. were enriched in spinach from the Yuma and Salinas areas, respectively. Our findings provide preliminary evidence that growing region and preharvest temperature may impact the bacterial quality of spinach and thus could inform more targeted strategies to manage produce quality.</p><p><strong>Importance: </strong>In the U.S., most spinach is produced in Arizona (AZ) and California (CA) seasonally; typically, spinach is cultivated in the Yuma, AZ, area during the winter and in the Salinas, CA, area during the summer. As the bacterial quality of baby spinach can influence consumer acceptance of the product, it is important to assess whether the bacterial quality of baby spinach can vary between spinach-growing regions. The findings of this study provide insights that could be used to support region-specific quality management strategies for baby spinach. Our results also highlight the value of further evaluating the impact of growing region and preharvest temperature on the bacterial quality of different produce commodities.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337821/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141589491","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pseudomonas protegens can serve as an agricultural biocontrol agent. P. protegens often encounters hyperosmotic stress during industrial production and field application. The ability of P. protegens to withstand hyperosmotic stress is important for its application as a biocontrol agent. AlgU is a global regulator responsible for stress response and biocontrol ability. However, the specific regulatory role of AlgU in the hyperosmotic adaptation of P. protegens is poorly understood. In this study, we found that the AlgU mutation disrupted the hyperosmotic tolerance of P. protegens. Many genes and metabolites related to cell envelope formation were significantly downregulated in ΔalgU compared with that in the wild-type (WT) strain under hyperosmotic conditions, and we found that the algU mutation caused membrane integrity to be compromised and increased membrane permeability. Further experiments revealed that the cell envelope integrity protein TolA, which is regulated by AlgU, contributes to cell membrane stability and osmotic tolerance in P. protegens. In addition, several genes related to oxidative stress response were significantly downregulated in ΔalgU, and higher levels of intracellular reactive oxygen species were found in ΔalgU. Furthermore, we found that the synthesis of N-acetyl glutaminyl glutamine amide is directly regulated by AlgU and contributes to the hyperosmotic adaptation of P. protegens. This study revealed the mechanisms of AlgU's participation in osmotic tolerance in P. protegens, and it provides potential molecular targets for research on the hyperosmotic adaptation of P. protegens.IMPORTANCEIn this study, we found that the extracytoplasmic function sigma factor AlgU is essential for the survival of P. protegens under hyperosmotic conditions. We provided evidence supporting the roles of AlgU in influencing cell membrane stability, intracellular reactive oxygen species (ROS) accumulation, and dipeptide N-acetylglutaminylglutamine amide (NAGGN) synthesis in P. protegens under hyperosmotic conditions. Our findings revealed the mechanisms of AlgU's participation in hyperosmotic stress tolerance in P. protegens, and they provide potential molecular targets for research on the hyperosmotic adaptation of P. protegens, which is of value in improving the biocontrol ability of P. protegens.
{"title":"AlgU mediates hyperosmotic tolerance in <i>Pseudomonas protegens</i> SN15-2 by regulating membrane stability, ROS scavenging, and osmolyte synthesis.","authors":"Jian Wang, Yaping Wang, Shouquan Lu, Haibo Lou, XiaoBing Wang, Wei Wang","doi":"10.1128/aem.00596-24","DOIUrl":"10.1128/aem.00596-24","url":null,"abstract":"<p><p><i>Pseudomonas protegens</i> can serve as an agricultural biocontrol agent. <i>P. protegens</i> often encounters hyperosmotic stress during industrial production and field application. The ability of <i>P. protegens</i> to withstand hyperosmotic stress is important for its application as a biocontrol agent. AlgU is a global regulator responsible for stress response and biocontrol ability. However, the specific regulatory role of AlgU in the hyperosmotic adaptation of <i>P. protegens</i> is poorly understood. In this study, we found that the AlgU mutation disrupted the hyperosmotic tolerance of <i>P. protegens</i>. Many genes and metabolites related to cell envelope formation were significantly downregulated in Δ<i>algU</i> compared with that in the wild-type (WT) strain under hyperosmotic conditions, and we found that the <i>algU</i> mutation caused membrane integrity to be compromised and increased membrane permeability. Further experiments revealed that the cell envelope integrity protein TolA, which is regulated by AlgU, contributes to cell membrane stability and osmotic tolerance in <i>P. protegens</i>. In addition, several genes related to oxidative stress response were significantly downregulated in Δ<i>algU</i>, and higher levels of intracellular reactive oxygen species were found in Δ<i>algU</i>. Furthermore, we found that the synthesis of N-acetyl glutaminyl glutamine amide is directly regulated by AlgU and contributes to the hyperosmotic adaptation of <i>P. protegens</i>. This study revealed the mechanisms of AlgU's participation in osmotic tolerance in <i>P. protegens</i>, and it provides potential molecular targets for research on the hyperosmotic adaptation of <i>P. protegens</i>.IMPORTANCEIn this study, we found that the extracytoplasmic function sigma factor AlgU is essential for the survival of <i>P. protegens</i> under hyperosmotic conditions. We provided evidence supporting the roles of AlgU in influencing cell membrane stability, intracellular reactive oxygen species (ROS) accumulation, and dipeptide N-acetylglutaminylglutamine amide (NAGGN) synthesis in <i>P. protegens</i> under hyperosmotic conditions. Our findings revealed the mechanisms of AlgU's participation in hyperosmotic stress tolerance in <i>P. protegens,</i> and they provide potential molecular targets for research on the hyperosmotic adaptation of <i>P. protegens</i>, which is of value in improving the biocontrol ability of <i>P. protegens</i>.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337839/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141632447","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Quorum sensing (QS) orchestrates many bacterial behaviors, including virulence and biofilm formation, across bacterial populations. Nevertheless, the underlying mechanism by which QS regulates capsular polysaccharide (CPS)-dependent phage-bacterium interactions remains unclear. In this study, we report that QS upregulates the expression of CPS-dependent phage receptors, thus increasing phage adsorption and infection rates in Vibrio alginolyticus. We found that QS upregulated the expression of the ugd gene, leading to increased synthesis of Autographiviridae phage receptor CPS synthesis in V. alginolyticus. The signal molecule autoinducer-2 released by Vibrio from different sources can potentially enhance CPS-dependent phage infections. Therefore, our data suggest that inhibiting QS may reduce, rather than improve, the therapeutic efficacy of CPS-specific phages.
Importance: Phage resistance is a direct threat to phage therapy, and understanding phage-host interactions, especially how bacteria block phage infection, is essential for developing successful phage therapy. In the present study, we demonstrate for the first time that Vibrio alginolyticus uses quorum sensing (QS) to promote capsular polysaccharide (CPS)-specific phage infection by upregulating ugd expression, which is necessary for the synthesis of Autographiviridae phage receptor CPS. Although increased CPS-specific phage susceptibility is a novel trade-off mediated by QS, it results in the upregulation of virulence factors, promoting biofilm development and enhanced capsular polysaccharide production in V. alginolyticus. This suggests that inhibiting QS may improve the effectiveness of antibiotic treatment, but it may also reduce the efficacy of phage therapy.
{"title":"Quorum sensing positively regulates CPS-dependent <i>Autographiviridae</i> phage infection in <i>Vibrio alginolyticus</i>.","authors":"Xixi Li, Chen Zhang, Shenao Li, Sixuan Liang, Xuefei Xu, Zhe Zhao","doi":"10.1128/aem.02210-23","DOIUrl":"10.1128/aem.02210-23","url":null,"abstract":"<p><p>Quorum sensing (QS) orchestrates many bacterial behaviors, including virulence and biofilm formation, across bacterial populations. Nevertheless, the underlying mechanism by which QS regulates capsular polysaccharide (CPS)-dependent phage-bacterium interactions remains unclear. In this study, we report that QS upregulates the expression of CPS-dependent phage receptors, thus increasing phage adsorption and infection rates in <i>Vibrio alginolyticus</i>. We found that QS upregulated the expression of the <i>ugd</i> gene, leading to increased synthesis of <i>Autographiviridae</i> phage receptor CPS synthesis in <i>V. alginolyticus</i>. The signal molecule autoinducer-2 released by <i>Vibrio</i> from different sources can potentially enhance CPS-dependent phage infections. Therefore, our data suggest that inhibiting QS may reduce, rather than improve, the therapeutic efficacy of CPS-specific phages.</p><p><strong>Importance: </strong>Phage resistance is a direct threat to phage therapy, and understanding phage-host interactions, especially how bacteria block phage infection, is essential for developing successful phage therapy. In the present study, we demonstrate for the first time that <i>Vibrio alginolyticus</i> uses quorum sensing (QS) to promote capsular polysaccharide (CPS)-specific phage infection by upregulating <i>ugd</i> expression, which is necessary for the synthesis of <i>Autographiviridae</i> phage receptor CPS. Although increased CPS-specific phage susceptibility is a novel trade-off mediated by QS, it results in the upregulation of virulence factors, promoting biofilm development and enhanced capsular polysaccharide production in <i>V. alginolyticus</i>. This suggests that inhibiting QS may improve the effectiveness of antibiotic treatment, but it may also reduce the efficacy of phage therapy.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11337841/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141787062","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Articles of Significant Interest in This Issue.","authors":"","doi":"10.1128/aem.01565-24","DOIUrl":"10.1128/aem.01565-24","url":null,"abstract":"","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11338310/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142016131","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}