Applied and Environmental Microbiology最新文献

{"title":"Mycoidesin, a novel lantibiotic, exhibits potent bacteriostatic activity against <i>Listeria monocytogenes</i> and effectively controls its growth in beef.","authors":"Fei Zhang, Jiajia Ding, Shu Liu, Guoqiang Huang, Shulin Deng, Mengyu Gao, Hualin Liu, Wanjing Lv, Xin Zeng, Bingyue Xin, Congcong Xu","doi":"10.1128/aem.00067-25","DOIUrl":"https://doi.org/10.1128/aem.00067-25","url":null,"abstract":"<p><p><i>Listeria monocytogenes</i> can cause severe listeriosis, with the consumption of contaminated food being an important route of its transmission. Biopreservatives can be used for the prevention and control of <i>L. monocytogenes</i> in food. In this study, we identified a novel lantibiotic, mycoidesin, with potent bacteriostatic activity against <i>L. monocytogenes</i>. It exhibited 4- to 16-fold higher bacteriostatic activity against the <i>L. monocytogenes</i> strains than nisin A. Analysis of the mode of action of mycoidesin revealed that it exerted bacteriostatic activity against <i>L. monocytogenes</i> ATCC 19111 at low and high concentrations (1×-32× MIC, 0.39-12.5 µM). It blocked cell wall synthesis by binding to Lipid II and inhibiting the growth of <i>L. monocytogenes</i>. For other sensitive strains, such as <i>Bacillus cereus</i> CMCC 63301, mycoidesin exerted a bacteriostatic effect at a low concentration (1× MIC, 1.56 µM) via the same mechanism, whereas it exerted a bactericidal effect at high concentrations (2×-8× MIC, 3.13-12.5 µM), which can damage the cell membrane and cause cell death. The stability test showed that mycoidesin had increased stability compared to nisin A. Additionally, mycoidesin showed low cytotoxic and hemolytic activity. Furthermore, mycoidesin effectively inhibited the growth of <i>L. monocytogenes</i> in beef and delayed the decline in beef quality. Our study demonstrates the potential of mycoidesin as a biopreservative to prevent <i>L. monocytogenes</i> contamination and improve the safety of meat and meat products in the food industry.</p><p><strong>Importance: </strong>This study aimed to identify highly effective, stable, and safe natural bacteriocin preservatives with anti-<i>Listeria monocytogenes</i> activity. We isolated a novel class II lantibiotic, mycoidesin, which exhibited more efficient bacteriostatic activity against <i>L. monocytogenes</i> and increased stability compared to the applied bacteriocin food preservative, nisin A. Mycoidesin also showed favorable biosafety. Moreover, mycoidesin could be effectively used for controlling <i>L. monocytogenes</i> in beef, demonstrating its potential as a biopreservative to prevent <i>L. monocytogenes</i>-related contamination and improve the safety of meat and meat products in the agricultural and food industries.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0006725"},"PeriodicalIF":3.9,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699462","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"SGBP-B-like bimodular cellulose-binding protein CHU_1279 is essential for cellulose utilization by <i>Cytophaga hutchinsonii</i>.","authors":"Weixin Zhang, Lizhu Li, Tengxin Li, Xin Li, Xia Wang, Qiang Yao, Xuemei Lu, Guanjun Chen, Weifeng Liu","doi":"10.1128/aem.02471-24","DOIUrl":"https://doi.org/10.1128/aem.02471-24","url":null,"abstract":"&lt;p&gt;&lt;p&gt;The widespread cellulolytic specialist &lt;i&gt;Cytophaga hutchinsonii&lt;/i&gt; belonging to the phylum Bacteroidetes adopted a unique cellulose utilization strategy that did not conform to the known cellulose-degrading paradigms involving free cellulases or cellulosomes. The strategy used by &lt;i&gt;C. hutchinsonii&lt;/i&gt; still remains largely unclear. In this study, we showed that &lt;i&gt;chu_1279&lt;/i&gt; within the &lt;i&gt;chu_1276-chu_1280&lt;/i&gt; gene cluster, which has been previously shown to be important for cellulose utilization by &lt;i&gt;C. hutchinsonii&lt;/i&gt;, encodes an outer membrane protein, and its elimination prohibited bacterial growth on cellulose. Structural prediction revealed that CHU_1279 is a surface glycan-binding protein B (SGBP-B)-like protein comprising two putative carbohydrate-binding module (CBM)-like domains. Further analyses verified that recombinant CHU_1279 displayed significant cellulose-binding protein, and its C-terminal domain is predominantly responsible for cellulose binding. Expression of the C-terminal domain but not the N-terminal domain restored cellulose utilization of ∆&lt;i&gt;chu_1279&lt;/i&gt;. Moreover, site-directed mutagenesis analyses identified three aromatic residues important for cellulose binding of the recombinant CHU_1279 protein. The defective cellulose utilization of ∆&lt;i&gt;chu_1279&lt;/i&gt; cells otherwise could be recovered by CHU_1279 variants with significantly damaged cellulose-binding capability. Sequence analyses revealed that orthologs of CHU_1279 as well as the atypical polysaccharide utilization loci (PUL) constituted by the gene cluster &lt;i&gt;chu_1276-chu_1280&lt;/i&gt; are also present in two other cellulolytic Bacteroidetes bacteria, &lt;i&gt;Cytophaga aurantiaca&lt;/i&gt; and &lt;i&gt;Sporocytophaga myxococcoides&lt;/i&gt;, which are closely related to &lt;i&gt;C. hutchinsonii&lt;/i&gt;. Our results contribute to unveiling the unique mechanism underlying the efficient cellulose utilization by &lt;i&gt;C. hutchinsonii&lt;/i&gt; and similar cellulolytic bacteria.IMPORTANCEMost members of the phylum Bacteroidetes are highly competitive and efficient degraders of complex polysaccharides largely ascribed to their employment of a SusC-like system encoded by a polysaccharide utilization locus (PUL). However, characterization of PULs is limited to those responsible for utilization of (semi)soluble glycans. PULs involved in the utilization of cellulose, the most abundant renewable polymer, have not been identified and functionally characterized yet. We demonstrated that &lt;i&gt;chu_1279&lt;/i&gt; in the cellulolytic specialist &lt;i&gt;C. hutchinsonii&lt;/i&gt; encodes an SGBP-B-like protein that is required for cellulose utilization, supporting that the gene cluster &lt;i&gt;chu_1276-chu_1280&lt;/i&gt; in &lt;i&gt;C. hutchinsonii&lt;/i&gt; encodes an atypical PUL system dedicated to cellulose assimilation. Further analyses showed that this atypical PUL system is also present in two other cellulolytic Bacteroidetes bacteria. This study not only contributes to unveiling the unusual cellulose utilization strategy adopted by &lt;i&gt;C. hutchinsonii","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0247124"},"PeriodicalIF":3.9,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699481","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Overexpression of sulfide:quinone reductase (SQR) in <i>Acidithiobacillus ferrooxidans</i> enhances sulfur, pyrite, and pyrrhotite oxidation.","authors":"Heejung Jung, Yuta Inaba, Scott Banta","doi":"10.1128/aem.00170-25","DOIUrl":"https://doi.org/10.1128/aem.00170-25","url":null,"abstract":"<p><p>Hydrogen sulfide is produced during the dissolution of some sulfidic minerals and during the microbial metabolism of reduced sulfur compounds. The sulfide:quinone reductase (SQR) enzyme is able to oxidize H₂S, and the bioleaching cells <i>Acidithiobacillus ferrooxidans</i> have two SQR genes, only one of which has been characterized. We cloned and overexpressed the two SQR genes in <i>A. ferrooxidans</i> and show that they both have SQR activity. Both AFE_0267 and AFE_1792 are active under anaerobic conditions, but only AFE_1792 is active under aerobic conditions. The effect of the SQR overexpression and the expression of related genes on sulfur metabolism was investigated. The overexpression of SQR improved cell growth and sulfur oxidation, suggesting enhanced SQR activity led to a reduction in H₂S toxicity as well as providing additional energy through H₂S oxidation. Additionally, the impact on the oxidation of pyrite and pyrrhotite was investigated. The rate of oxidation of pyrite by the engineered cells was enhanced, and, furthermore, the rate of pyrrhotite oxidation was more than doubled.IMPORTANCEH₂S is a toxic sulfur intermediate, and the SQR enzyme has evolved to oxidize H₂S in <i>A. ferrooxidans</i>. In addition to detoxification, H₂S oxidation provides energy, and overexpression of SQR enhanced aerobic and anaerobic growth on sulfur. The SQR overexpression also enhanced pyrite and pyrrhotite oxidation, which may facilitate the pyrometallurgical processing of a number of critical materials including copper, nickel, and the platinum group metals.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0017025"},"PeriodicalIF":3.9,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699464","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Product formulation and rubbing time impact the inactivation of enveloped and non-enveloped virus surrogates by foam-based hand sanitizers.","authors":"Francis Torko, Kristen E Gibson","doi":"10.1128/aem.02474-24","DOIUrl":"https://doi.org/10.1128/aem.02474-24","url":null,"abstract":"<p><p>Effective hand hygiene, such as hand washing and hand sanitizer use, is crucial in reducing infectious disease transmission via the hands. The efficacy of hand washing has been well-documented; however, relatively less is known regarding foam-based hand sanitizer efficacy, which is considered an effective alternative to washing hands with soap and water. Hand sanitizers are recommended by both the United States Centers for Disease Control and Prevention and the World Health Organization when the hands are not visibly dirty or greasy. This study examined the efficacy of five commercially available foam hand sanitizers-four alcohol-based and one non-alcohol-based-against enveloped and non-enveloped viruses using bacteriophage phi6 (Φ6) and bacteriophage MS2 as surrogates, respectively. A cocktail of MS2 and Φ6 (8 log PFU/mL) was inoculated on the hands and exposed to 3 or 6 mL of hand sanitizer product followed by rubbing the palmar surface of the hands together for 10 s or until dry. The results showed significant log reduction among the virus surrogates (<i>P</i> ≤ 0.05), with Φ6 consistently showing higher susceptibility across all factors compared with MS2 with log reductions of 2.83 ± 1.98 and 0.50 ± 0.53 log reduction, respectively. Although dosing volume did not significantly impact log reduction (<i>P</i> = 0.31), rubbing time significantly affected bacteriophage inactivation (<i>P</i> ≤ 0.05). Higher log reduction was observed when hands were rubbed until dry (2.69 ± 2.06), compared with the typical 10 s rubbing time (0.65 ± 0.75). This study revealed that the efficacy of commercially available foam hand sanitizers depends on rubbing time and overall product formulation, rather than exclusively on active ingredient concentration.IMPORTANCEHuman hands are a key factor in the transmission of viral diseases, and proper hand hygiene is regarded as the gold standard against the spread of such diseases. This study examined the effectiveness of a hand hygiene technique, that is, the application of foam-based hand sanitizers, against the inactivation of enveloped and non-enveloped virus surrogates on the hands. Factors such as virus type, rubbing time, volume of product used, and product formulation can significantly influence the efficacy of hand sanitizers. To assess these effects, we tested different rubbing times and product volumes across alcohol- and non-alcohol-based, foam hand sanitizer formulations, each with varying active ingredient concentrations and inactive ingredients. The study was performed on the palmar surface of human hands to realistically simulate real-world conditions, providing valuable evidence to inform future hand sanitizer practices aimed at maximizing the reduction of infectious viral pathogens on the hands.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0247424"},"PeriodicalIF":3.9,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699467","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Indole-3-acetic acid (IAA) protects <i>Azospirillum brasilense</i> from indole-induced stress.","authors":"Elena E Ganusova, Ishita Banerjee, Trey Seats, Gladys Alexandre","doi":"10.1128/aem.02384-24","DOIUrl":"https://doi.org/10.1128/aem.02384-24","url":null,"abstract":"<p><p><i>Azospirillum brasilense</i> is plant-growth promoting rhizobacteria that produces the phytohormone indole-3-acetic acid (IAA) to induce changes in plant root architecture. The major pathway for IAA biosynthesis in <i>A. brasilense</i> converts tryptophan into indole-3-pyruvic acid (I3P) and then, through the rate-limiting enzyme, indole-3-pyruvate decarboxylase (IpdC), into IAA. Here, we characterize the potential role for IAA biosynthesis in the physiology of these bacteria by characterizing the expression pattern of the <i>ipdC</i> promoter, analyzing an <i>A. brasilense ipdC</i> mutant using multiple physiological assays and characterizing the effect of I3P, which likely accumulates in the absence of <i>ipdC</i> and affects bacterial physiology. We found that the <i>ipdC</i> mutant derivative has a reduced growth rate and an altered physiology, including reduced translation activity as well as a more depolarized membrane potential compared to the parent strain. Similar effects could be recapitulated in the parent strain by exposing these cells to increasing concentrations of I3P, as well as other indole intermediates of IAA biosynthesis. Our results also indicate a protective role for IAA against the harmful effects of indole derivatives, with exogenous IAA restoring the membrane potential of cells exposed to indole derivatives for prolonged periods. These protective effects appeared to restore cell physiology, including in the wheat rhizosphere. Together, our data suggest that the IAA biosynthesis pathway plays a major role in <i>A. brasilense</i> physiology by maintaining membrane potential homeostasis and regulating translation, likely to mitigate the potential membrane-damaging effects of indoles that accumulate during growth under stressful conditions.IMPORTANCEIAA is widely synthesized in bacteria, particularly in soil and rhizosphere bacteria, where it functions as a phytohormone to modulate plant root architecture. IAA as a secondary metabolite has been shown to serve as a signaling molecule in several bacterial species, but the role of IAA biosynthesis in the physiology of the producing bacterium remains seldom explored. Results obtained here suggest that IAA serves to protect <i>A. brasilense</i> from the toxic effect of indoles, including metabolite biosynthetic precursors of IAA, on membrane potential homeostasis. Given the widespread production of IAA in soil bacteria, this protective effect of IAA may be conserved in diverse soil bacteria.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0238424"},"PeriodicalIF":3.9,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699457","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Role of a single MCP in evolutionary adaptation of <i>Shewanella putrefaciens</i> for swimming in planktonic and structured environments.","authors":"Daniel B Edelmann, Anna M Jakob, Laurence G Wilson, Rémy Colin, David Brandt, Frederik Eck, Jörn Kalinowski, Kai M Thormann","doi":"10.1128/aem.00229-25","DOIUrl":"https://doi.org/10.1128/aem.00229-25","url":null,"abstract":"<p><p>Bacteria can adapt to their environments by changing phenotypic traits by mutations. However, improving one trait often results in the deterioration of another one, a trade-off that limits the degree of adaptation. The gammaproteobacterium <i>Shewanella putrefaciens</i> CN-32 has an elaborate motility machinery comprising two distinct flagellar systems and an extensive chemotaxis array with 36 methyl-accepting chemotaxis sensor proteins (MCPs). In this study, we performed experimental selection on S. <i>putrefaciens</i> for increased spreading through a porous environment. We readily obtained a mutant that showed a pronounced increase in covered distance. This phenotype was almost completely caused by a deletion of 24 bp from the chromosome, which leads to a moderately enhanced production of a single MCP. Accordingly, chemotaxis assays under free-swimming conditions and cell tracking in soft agar showed that the mutation improved navigation through nutritional gradients. In contrast, further increased levels of the MCP negatively affected spreading. The study demonstrates how moderate differences in the abundance of a single MCP can lead to an efficient upgrade of chemotaxis in specific environments at a low expense of cellular resources.IMPORTANCEExperimental evolution experiments have been used to determine the trade-offs occurring in specific environments. Several studies that have used the spreading behavior of bacteria in structured environments identified regulatory mutants that increase the swimming speed of the cells. While this results in a higher chemotaxis drift, the growth fitness decreases as the higher swimming speed requires substantial cellular resources. Here we show that rapid chemotaxis adaptation can also be achieved by modifying the chemotaxis signal input at a low metabolic cost for the cell.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0022925"},"PeriodicalIF":3.9,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699470","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effect of beaver dam analogs (BDAs) on waterborne protozoal pathogens <i>Giardia duodenalis</i> and <i>Cryptosporidium parvum</i>.","authors":"Ariel I Loredo, Andrea Packham, Nick Graham, Skylar Johnson, Stephanie Elliott, Brian H Bird, Carrie Monohan, Sarah Yarnell, Woutrina A Smith","doi":"10.1128/aem.01569-24","DOIUrl":"https://doi.org/10.1128/aem.01569-24","url":null,"abstract":"<p><p>Installing beaver dam analogs (BDAs) in freshwater ecosystems is a process-based restoration technique mimicking natural beaver dams to increase stream channel complexity, floodplain connectivity, and hydrological residence time. BDAs have been applied in recent years by resource managers to improve riparian habitats with beneficial effects on ecosystem services, including improved water quality. While BDAs have been shown to reduce suspended sediments and nutrients in surface waters, research is needed to evaluate BDA effects on waterborne fecal pathogens such as <i>Giardia duodenalis</i> (syn. <i>G. lamblia</i>, <i>G. intestinalis</i>) and <i>Cryptosporidium parvum</i>. At a California riparian field site, this study used an <i>in situ</i> release trial of inactivated protozoal (oo)cysts upstream of BDAs to compare the concentrations and loads detected above and below the BDAs. The median percent decrease in load-based recovery rates was 78% for <i>G. duodenalis</i> and 80% for <i>C. parvum</i> at sites with BDAs. Mixed-effect linear regression analyses showed a significant reduction (by 81%) of the <i>G. duodenalis</i> adjusted cyst concentrations in downstream surface waters at sites with BDAs compared to similar sites without BDAs. While a reduction of <i>C. parvum</i> oocysts was noted, it was not statistically significant. The improved surface water quality downstream of BDAs could be because the BDA structures promote hyporheic exchange and act as a passive filter to remove pathogenic protozoa from streams. If BDAs are used to aid the restoration of riparian meadow ecosystems to slow water and raise the water table, they may also help to promote cleaner and safer waters for communities downstream.</p><p><strong>Importance: </strong>Beaver dam analogs (BDAs) are a cost-effective, low-environmental impact technique for stream and riparian meadow restoration that provide a variety of beneficial ecosystem services; however, their impact on waterborne fecal protozoal pathogens has not been evaluated. This study used an <i>in situ</i> protozoal release trial to quantify the effect of BDAs on the load of <i>Giardia duodenalis</i> and <i>Cryptosporidium parvum</i> in streams in California. Results showed that <i>G. duodenalis</i> concentrations below BDAs were significantly reduced by 81%. The median percent decrease in load-based recovery rates below a BDA for <i>G. duodenalis</i> and <i>C. parvum</i> was 78% and 80%, respectively. This finding indicates that BDAs may promote passive filtration of waterborne pathogens, thereby improving water quality in downstream reaches and adding to the beneficial outcomes associated with using BDAs in stream and meadow restoration efforts. The potential benefit to resource managers and communities is immense.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0156924"},"PeriodicalIF":3.9,"publicationDate":"2025-03-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143690506","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Efficient genetic manipulation of <i>Shewanella</i> through targeting defense islands.","authors":"Yilong Ruan, Huan Tang, Tongxuan Cai, Xiaofei Du, Tianlang Liu, Xiaoxue Wang, Pengxia Wang","doi":"10.1128/aem.02499-24","DOIUrl":"https://doi.org/10.1128/aem.02499-24","url":null,"abstract":"<p><p>The <i>Shewanella</i> genus is widely recognized for its remarkable respiratory adaptability in anaerobic environments, exhibiting potential for bioremediation and microbial fuel cell applications. However, the genetic manipulation of certain <i>Shewanella</i> strains is hindered by defense systems that limit their genetic modification in biotechnology processes. In this study, we present a systematic method for predicting, mapping, and functionally analyzing defense islands within bacterial genomes. We investigated the genetically recalcitrant strain <i>Shewanella putrefaciens</i> CN32 and identified several defense systems located on two genomic islands integrated within the conserved chromosomal genes <i>trmA</i> and <i>trmE</i>. Our experimental assays demonstrated that overexpression of excisionases facilitated the excision of these islands from the chromosome, and their removal significantly enhanced the genetic manipulation efficiency of <i>S. putrefaciens</i> CN32. Further analysis revealed that these defense islands are widespread across various <i>Shewanella</i> strains and other gram-negative bacteria. This study presents an effective strategy to circumvent genetic barriers and fully exploit the potential of <i>Shewanella</i> for environmental and microbial engineering applications.</p><p><strong>Importance: </strong>Efficiently modifying bacterial genomes is critical for advancing their industrial applications. However, bacteria in complex environments naturally develop defense mechanisms in response to bacteriophages and exogenous DNA, which pose significant challenges to their genetic modification. Several methods have emerged to tackle these challenges, including <i>in vitro</i> methylation of plasmid DNA and targeting specific restriction-modification (R-M) and CRISPR loci. Nevertheless, many bacteria harbor multiple, often uncharacterized defense mechanisms, limiting these strategies. Our study differs from previous approaches by specifically targeting defense islands-clusters of defense systems located within mobile genetic elements. Here, we investigated <i>Shewanella putrefaciens</i> CN32 and identified two key defense islands responsible for these protective functions. By selectively deleting these defense islands, we significantly enhanced the efficiency of genetic manipulation in <i>S. putrefaciens</i>. Our findings not only demonstrate a promising strategy for improving genetic engineering in <i>Shewanella</i> but also suggest broader applicability across other bacterial species. This work opens new opportunities for optimizing microbial processes in biotechnology, highlighting the potential of defense island-targeted genetic modification.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0249924"},"PeriodicalIF":3.9,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143673193","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"<i>Salmonella enterica</i> serovar Braenderup shows clade-specific source associations and a high proportion of molecular epidemiological clustering.","authors":"Harleen K Chaggar, Lauren K Hudson, Kelly Orejuela, Linda Thomas, Maya Spann, Katie N Garman, John R Dunn, Thomas G Denes","doi":"10.1128/aem.02594-24","DOIUrl":"https://doi.org/10.1128/aem.02594-24","url":null,"abstract":"&lt;p&gt;&lt;p&gt;&lt;i&gt;Salmonella enterica&lt;/i&gt; serovar Braenderup (&lt;i&gt;S. enterica&lt;/i&gt; ser. Braenderup) is an important clinical serovar in the United States. This serovar was reported by the CDC in 2017 as the fifth most common &lt;i&gt;Salmonella enterica&lt;/i&gt; serovar associated with outbreaks in the United States, which have been linked to both fresh produce and food animal products. The goals of this study were to compare the relatedness of human clinical isolates from southeastern USA (Tennessee (&lt;i&gt;n&lt;/i&gt; = 106), Kentucky (&lt;i&gt;n&lt;/i&gt; = 48), Virginia (&lt;i&gt;n&lt;/i&gt; = 252), South Carolina (&lt;i&gt;n&lt;/i&gt; = 109), Georgia (&lt;i&gt;n&lt;/i&gt; = 159), Alabama (&lt;i&gt;n&lt;/i&gt; = 8), Arkansas (&lt;i&gt;n&lt;/i&gt; = 26), and Louisiana (&lt;i&gt;n&lt;/i&gt; = 91)) and global clinical (&lt;i&gt;n&lt;/i&gt; = 5,153) and nonclinical (&lt;i&gt;n&lt;/i&gt; = 1,053) isolates obtained from the NCBI. Additionally, we also examined the population structure of &lt;i&gt;S. enterica&lt;/i&gt; ser. Braenderup strains (&lt;i&gt;n&lt;/i&gt; = 3,131) on EnteroBase and found that all the strains of this serovar are associated with a single cgMLST eBurst group (ceBG 185), confirming that this serovar is monophyletic. We divided the &lt;i&gt;S. enterica&lt;/i&gt; ser. Braenderup population into two clades (Clade I and Clade II) and one clade group (Clade Group III). The composition of distinct environmental isolates in the clades differed: Clade I was significantly associated with produce (90.7%; &lt;i&gt;P&lt;/i&gt; &lt; 0.0001) and water, soil, and sediment (76.9%; &lt;i&gt;P&lt;/i&gt; &lt; 0.0001), and Clade II was significantly associated with poultry environments (62.8%; &lt;i&gt;P&lt;/i&gt; &lt; 0.0001). The clade-specific gene associations (e.g., Clade I-associated competence proteins and cytochrome_c_asm protein and Clade II-associated heme-exporter protein and dimethyl sulfoxide [DMSO] reductase-encoding genes) provide potential insights into possible mechanisms driving environmental adaptation and host-pathogen interaction. Phylogenetic analyses identified 218 molecular epidemiological clusters in the current study, which represented a greater proportion of potentially outbreak-related isolates than previously estimated.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Importance: &lt;/strong&gt;This study provides insights into the genomic diversity of &lt;i&gt;S. enterica&lt;/i&gt; ser. Braenderup by revealing distinct clade-specific source attribution patterns and showing that a greater proportion of isolates were associated with epidemiological clusters based on the genomic relatedness than previously estimated. Specifically, we analyzed the diversity of human clinical isolates from southeastern USA and compared them with the global clinical and nonclinical isolates. Our analysis showed different clades of &lt;i&gt;S. enterica&lt;/i&gt; ser. Braenderup linked to different environments, providing insights on the potential source of human sporadic infection and outbreaks. These findings can enhance public health surveillance and response strategies targeting &lt;i&gt;S. enterica&lt;/i&gt; serovar Braenderup by expanding our understanding of potential transmission pathways and the genomic diversity of clinical ","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0259424"},"PeriodicalIF":3.9,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143673190","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Alternate routes to acetate tolerance lead to varied isoprenol production from mixed carbon sources in <i>Pseudomonas putida</i>.","authors":"Guilherme M V de Siqueira, Aparajitha Srinivasan, Yan Chen, Jennifer W Gin, Christopher J Petzold, Taek Soon Lee, María-Eugenia Guazzaroni, Thomas Eng, Aindrila Mukhopadhyay","doi":"10.1128/aem.02123-24","DOIUrl":"https://doi.org/10.1128/aem.02123-24","url":null,"abstract":"<p><p>Lignocellulose is a renewable resource for the production of a diverse array of platform chemicals, including the biofuel isoprenol. Although this carbon stream provides a rich source of sugars, other organic compounds, such as acetate, can be used by microbial hosts. Here, we examined the growth and isoprenol production in a <i>Pseudomonas putida</i> strain pre-tolerized (\"PT\") background where its native isoprenol catabolism pathway is deleted, using glucose and acetate as carbon sources. We found that PT displays impaired growth in minimal medium containing acetate and often fails to grow in glucose-acetate medium. Using a mutant recovery-based approach, we generated tolerized strains that overcame these limitations, achieving fast growth and isoprenol production in the mixed carbon feed. Changes in the glucose and acetate assimilation routes, including an upregulation in PP_0154 (SpcC, succinyl-CoA:acetate CoA-transferase) and differential expression of the gluconate assimilation pathways, were key for higher isoprenol titers in the tolerized strains, whereas a different set of mechanisms were likely enabling tolerance phenotypes in media containing acetate. Among these, a coproporphyrinogen-III oxidase (HemN) was upregulated across all tolerized strains and in one isolate required for acetate tolerance. Utilizing a defined glucose and acetate mixture ratio reflective of lignocellulosic feedstocks for isoprenol production in <i>P. putida</i> allowed us to obtain insights into the dynamics and challenges unique to dual carbon source utilization that are obscured when studied separately. Together, this enabled the development of a <i>P. putida</i> bioconversion chassis able to use a more complex carbon stream to produce isoprenol.IMPORTANCEAcetate is a relatively abundant component of many lignocellulosic carbon streams and has the potential to be used together with sugars, especially in microbes with versatile catabolism such as <i>P. putida</i>. However, the use of mixed carbon streams necessitates additional optimization. Furthermore, the use of <i>P. putida</i> for the production of the biofuel target, isoprenol, requires the use of engineered strains that have additional growth and production constraints when cultivated in acetate and glucose mixtures. In this study, we generate acetate-tolerant <i>P. putida</i> strains that overcome these challenges and examine their ability to produce isoprenol. We show that acetate tolerance and isoprenol production, although independent phenotypes, can both be optimized in a given <i>P. putida</i> strain. Using proteomics and whole genome sequencing, we examine the molecular basis of both phenotypes and show that tolerance to acetate can occur via alternate routes and result in different impacts on isoprenol production.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0212324"},"PeriodicalIF":3.9,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143662120","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}