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Nature Communications Mar 2024Matrix-encapsulated communities of bacteria, called biofilms, are ubiquitous in the environment and are notoriously difficult to eliminate in clinical and industrial...
Matrix-encapsulated communities of bacteria, called biofilms, are ubiquitous in the environment and are notoriously difficult to eliminate in clinical and industrial settings. Biofilm formation likely evolved as a mechanism to protect resident cells from environmental challenges, yet how bacteria undergo threat assessment to inform biofilm development remains unclear. Here we find that population-level cell lysis events induce the formation of biofilms by surviving Vibrio cholerae cells. Survivors detect threats by sensing a cellular component released through cell lysis, which we identify as norspermidine. Lysis sensing occurs via the MbaA receptor with genus-level specificity, and responsive biofilm cells are shielded from phage infection and attacks from other bacteria. Thus, our work uncovers a connection between bacterial lysis and biofilm formation that may be broadly conserved among microorganisms.
Topics: Bacteriophages; Biofilms; Cell Aggregation; Cell Death; Vibrio cholerae
PubMed: 38443393
DOI: 10.1038/s41467-024-46399-1 -
Microbiology Spectrum Apr 2024Fluorescent proteins have revolutionized science since their discovery in 1962. They have enabled imaging experiments to decipher the function of proteins, cells, and...
Fluorescent proteins have revolutionized science since their discovery in 1962. They have enabled imaging experiments to decipher the function of proteins, cells, and organisms, as well as gene regulation. Green fluorescent protein and all its derivatives are now standard tools in cell biology, immunology, molecular biology, and microbiology laboratories around the world. A common feature of these proteins is their dioxygen (O)-dependent maturation allowing fluorescence, which precludes their use in anoxic contexts. In this work, we report the development and characterization of genetic circuits encoding the O-independent KOFP-7 protein, a flavin-binding fluorescent protein. We have optimized the genetic circuit for high bacterial fluorescence at population and single-cell level, implemented this circuit in various plasmids differing in host range, and quantified their fluorescence under both aerobic and anaerobic conditions. Finally, we showed that KOFP-7-based constructions can be used to produce fluorescing cells of , a facultative anaerobe, demonstrating the usefulness of the genetic circuits for various anaerobic bacteria. These genetic circuits can thus be modified at will, both to solve basic and applied research questions, opening a highway to shed light on the obscure anaerobic world.IMPORTANCEFluorescent proteins are used for decades, and have allowed major discoveries in biology in a wide variety of fields, and are used in environmental as well as clinical contexts. Green fluorescent protein (GFP) and all its derivatives share a common feature: they rely on the presence of dioxygen (O) for protein maturation and fluorescence. This dependency precludes their use in anoxic environments. Here, we constructed a series of genetic circuits allowing production of KOFP-7, an O-independant flavin-binding fluorescent protein. We demonstrated that cells producing KOFP-7 are fluorescent, both at the population and single-cell levels. Importantly, we showed that, unlike cells producing GFP, cells producing KOFP-7 are fluorescent in anoxia. Finally, we demonstrated that NS1, a facultative anaerobe, is fluorescent in the absence of O when KOFP-7 is produced. Altogether, the development of new genetic circuits allowing O-independent fluorescence will open new perspective to study anaerobic processes.
Topics: Green Fluorescent Proteins; Bacteria; Flavins; Oxygen; Vibrio
PubMed: 38441526
DOI: 10.1128/spectrum.04091-23 -
Scientific Reports Feb 2024A mathematical model that describes the dynamics of bacterium vibrio cholera within a fixed population considering intrinsic bacteria growth, therapeutic treatment,...
A mathematical model that describes the dynamics of bacterium vibrio cholera within a fixed population considering intrinsic bacteria growth, therapeutic treatment, sanitation and vaccination rates is developed. The developed mathematical model is validated against real cholera data. A sensitivity analysis of some of the model parameters is also conducted. The intervention rates are found to be very important parameters in reducing the values of the basic reproduction number. The existence and stability of equilibrium solutions to the mathematical model are also carried out using analytical methods. The effect of some model parameters on the stability of equilibrium solutions, number of infected individuals, number of susceptible individuals and bacteria density is rigorously analyzed. One very important finding of this research work is that keeping the vaccination rate fixed and varying the treatment and sanitation rates provide a rapid decline of infection. The fourth order Runge-Kutta numerical scheme is implemented in MATLAB to generate the numerical solutions.
Topics: Humans; Cholera; Models, Biological; Models, Theoretical; Vibrio cholerae; Sanitation
PubMed: 38409239
DOI: 10.1038/s41598-024-55240-0 -
BioRxiv : the Preprint Server For... Feb 2024In species, quorum sensing signaling culminates in the production of a TetR-type master transcription factor collectively called the LuxR/HapR family, which regulates...
UNLABELLED
In species, quorum sensing signaling culminates in the production of a TetR-type master transcription factor collectively called the LuxR/HapR family, which regulates genes required for colonization and infection of host organisms. These proteins possess a solvent accessible putative ligand binding pocket. However, a native ligand has not been identified, and the role of ligand binding in LuxR/HapR function in is unknown. To probe the role of the ligand binding pocket, we utilize the small molecule thiophenesulfonamide inhibitor PTSP (3- henyl-1-( hiophen-2-yl ulfonyl)-1 - yrazole) that we previously showed targets LuxR/HapR proteins. Amino acid conservation in the ligand binding pocket determines the specificity and efficacy of PTSP inhibition across species. Here, we used structure-function analyses to identify PTSP-interacting residues in the ligand binding pocket of SmcR - the LuxR/HapR homolog - that are required for PTSP inhibition of SmcR activity . Forward genetic screening combined with X-ray crystallography structural determination of SmcR bound to PTSP identified substitutions at eight residues that were sufficient to reduce or eliminate PTSP-mediated SmcR inhibition. Small-angle X-ray scattering and computational modeling determined that PTSP drives allosteric unfolding at the N-terminal DNA binding domain. We discovered that SmcR is degraded by the ClpAP protease in the presence of PTSP ; substitution of key PTSP-interacting residues stabilized or increased SmcR levels in the cell. This mechanism of inhibition is observed for all thiophenesulfonamide compounds tested and against other species. We conclude that thiophenesulfonamides specifically bind in the ligand binding pocket of LuxR/HapR proteins, promoting protein degradation and thereby suppressing downstream gene expression, implicating ligand binding as a mediator of LuxR/HapR protein stability and function to govern virulence gene expression in pathogens.
SIGNIFICANCE
LuxR/HapR proteins were discovered in the 1990s as central regulators of quorum sensing gene expression and later discovered to be conserved in all studied species. LuxR/HapR homologs regulate a wide range of genes involved in pathogenesis, including but not limited to genes involved in biofilm production and toxin secretion. As archetypal members of the broad class of TetR-type transcription factors, each LuxR/HapR protein has a predicted ligand binding pocket. However, no ligand has been identified for LuxR/HapR proteins that control their function as regulators. Here, we used LuxR/HapR-specific chemical inhibitors to determine that ligand binding drives proteolytic degradation , the first demonstration of LuxR/HapR function connected to ligand binding for this historical protein family.
PubMed: 38405947
DOI: 10.1101/2024.02.15.580527 -
MSphere Mar 2024Vaccination is important to prevent cholera. There are limited data comparing anti-O-specific polysaccharide (OSP) and anti-cholera toxin-specific immune responses...
Vaccination is important to prevent cholera. There are limited data comparing anti-O-specific polysaccharide (OSP) and anti-cholera toxin-specific immune responses following oral whole-cell with cholera toxin B-subunit (WC-rBS) vaccine (Dukoral, Valneva) administration in different age groups. An understanding of the differences is relevant because young children are less well protected by oral cholera vaccines than older children and adults. We compared responses in 50 adults and 49 children (ages 2 to <18) who were administered two doses of WC-rBS at a standard 14-day interval. All age groups had significant IgA and IgG plasma-blast responses to the OSP and cholera toxin B-subunit (CtxB) antigens that peaked 7 days after vaccination. However, in adults and older children (ages 5 to <18), antibody responses directed at the OSP antigen were largely IgA and IgG, with a minimal IgM response, while younger children (ages 2 to <5) mounted significant increases in IgM with minimal increases in IgA and IgG antibody responses 30 days after vaccination. In adults, anti-OSP and CtxB memory B-cell responses were detected after completion of the vaccination series, while children only mounted CtxB-specific IgG memory B-cell responses and no OSP-memory B-cell responses. In summary, children and adults living in a cholera endemic area mounted different responses to the WC-rBS vaccine, which may be a result of more prior exposure to in older participants. The absence of class-switched antibody responses and memory B-cell responses to OSP may explain why protection wanes more rapidly after vaccination in young children compared to older vaccinees.IMPORTANCEVaccination is an important strategy to prevent cholera. Though immune responses targeting the OSP of are believed to mediate protection against cholera, there are limited data on anti-OSP responses after vaccination in different age groups, which is important as young children are not well protected by current oral cholera vaccines. In this study, we found that adults mounted memory B-cell responses to OSP, which were not seen in children. Adults and older children mounted class-switched (IgG and IgA) serum antibody responses to OSP, which were not seen in young children who had only IgM responses to OSP. The lack of class-switched antibody responses and memory B-cell responses to OSP in younger participants may be due to lack of prior exposure to and could explain why protection wanes more rapidly after vaccination in young children.
Topics: Adult; Child; Humans; Adolescent; Child, Preschool; Aged; Infant, Newborn; Cholera Vaccines; Cholera; Cholera Toxin; O Antigens; Vibrio cholerae O1; Immunoglobulin M; Antibodies, Bacterial; Immunoglobulin A; Vaccination; Antibody Formation; Immunoglobulin G
PubMed: 38391226
DOI: 10.1128/msphere.00565-23 -
Inorganic Chemistry Mar 2024Photoxenobactin E () is a natural product with an unusual thiocarboxylic acid terminus recently isolated from an entomopathogenic bacterium. The biosynthetic gene...
Photoxenobactin E () is a natural product with an unusual thiocarboxylic acid terminus recently isolated from an entomopathogenic bacterium. The biosynthetic gene cluster associated with photoxenobactin E, and other reported derivatives, is very similar to that of piscibactin, the siderophore responsible for the iron uptake among bacteria of the family, including potential human pathogens. Here, the reisolation of from the fish pathogen RV22 cultured under iron deprivation, its ability to chelate Ga(III), and the full NMR spectroscopic characterization of the Ga(III)-photoxenobactin E complex are presented. Our results show that Ga(III)-photoxenobactin E in solution exists in a thiol-thione tautomeric equilibrium, where Ga(III) is coordinated through the sulfur (thiol form) or oxygen (thione form) atoms of the thiocarboxylate group. This report represents the first NMR study of the chemical exchange between the thiol and thione forms associated with thiocarboxylate-Ga(III) coordination, including the kinetics of the interconversion process associated with this tautomeric exchange. These findings show significant implications for ligand design as they illustrate the potential of the thiocarboxylate group as a versatile donor for hard metal ions such as Ga(III).
Topics: Animals; Humans; Thiones; Metals; Iron; Siderophores; Sulfhydryl Compounds
PubMed: 38387064
DOI: 10.1021/acs.inorgchem.3c04076 -
Nature Medicine Mar 2024Our understanding of cholera transmission and burden largely relies on clinic-based surveillance, which can obscure trends, bias burden estimates and limit the impact of...
Our understanding of cholera transmission and burden largely relies on clinic-based surveillance, which can obscure trends, bias burden estimates and limit the impact of targeted cholera-prevention measures. Serological surveillance provides a complementary approach to monitoring infections, although the link between serologically derived infections and medically attended disease incidence-shaped by immunological, behavioral and clinical factors-remains poorly understood. We unravel this cascade in a cholera-endemic Bangladeshi community by integrating clinic-based surveillance, healthcare-seeking and longitudinal serological data through statistical modeling. Combining the serological trajectories with a reconstructed incidence timeline of symptomatic cholera, we estimated an annual Vibrio cholerae O1 infection incidence rate of 535 per 1,000 population (95% credible interval 514-556), with incidence increasing by age group. Clinic-based surveillance alone underestimated the number of infections and reported cases were not consistently correlated with infection timing. Of the infections, 4 in 3,280 resulted in symptoms, only 1 of which was reported through the surveillance system. These results impart insights into cholera transmission dynamics and burden in the epicenter of the seventh cholera pandemic, where >50% of our study population had an annual V. cholerae O1 infection, and emphasize the potential for a biased view of disease burden and infection risk when depending solely on clinical surveillance data.
Topics: Humans; Cholera; Vibrio cholerae; Incidence
PubMed: 38378884
DOI: 10.1038/s41591-024-02810-4 -
Applied Microbiology and Biotechnology Feb 2024The gut microbial communities interact with the host immunity and physiological functions. In this study, we investigated the bacterial composition in Litopenaeus...
The gut microbial communities interact with the host immunity and physiological functions. In this study, we investigated the bacterial composition in Litopenaeus vannamei shrimp's gut and rearing water under different host (developmental stage: juvenile and adult; health status: healthy and diseased) and environmental factors (temperature 25 °C and 28 °C; and light intensity: low and high). The PCoA analysis showed that all water samples were clustered together in a quarter, whereas the gut samples spread among three quarters. In terms of functional bacteria, gut samples of adult shrimp, healthy adult shrimp, adult shrimp raised at 28 °C, and juvenile shrimp under high light intensity exhibited a higher abundance of Vibrionaceae compared to each other opposite group. Gut samples of juvenile shrimp, infected adult shrimp, juvenile shrimp with low light intensity, and adult shrimp with a water temperature of 25 °C showed a higher abundance of Pseudoaltromonadaceae bacteria compared to each other opposite group. Gut samples of juvenile shrimp, healthy adult shrimp, adult shrimp raised at a water temperature of 28 °C, and juvenile shrimp with high light intensity showed the higher abundance of Firmicutes/Bacteroidota ratio compared to each other opposite group. Our results showed that L. vannamei juveniles are more sensitive to bacterial infections; besides, water temperature of 28 °C and high light intensity groups were both important conditions improving the shrimp gut bacterial composition under industrial indoor farming systems. KEY POINTS: • Bacteria diversity was higher among shrimp intestinal microbiota compared to the rearing water. • Shrimp juveniles are more sensitive to bacterial infection compared to adults. • Water temperature of 28 °C and high light intensity are recommended conditions for white shrimp aquaculture.
Topics: Animals; Gastrointestinal Microbiome; Agriculture; Farms; Microbiota; Penaeidae; Water
PubMed: 38376561
DOI: 10.1007/s00253-024-13046-0 -
Scientific Reports Feb 2024In this study, three generations of polymerase chain reaction (PCR) assays: (i) conventional PCR, (ii) qPCR and (iii) droplet digital PCR (ddPCR), were systematically...
In this study, three generations of polymerase chain reaction (PCR) assays: (i) conventional PCR, (ii) qPCR and (iii) droplet digital PCR (ddPCR), were systematically tested for their abilities to detect non-pathogenic and pathogenic populations of Vibrio parahaemolyticus. The limit of detection (LOD) for the ddPCR was 1.1 pg/µL of purified DNA, followed by the qPCR (5.6 pg/µL) and the conventional PCR (8.8 pg/µL). Regarding the LOD for V. parahaemolyticus cells, the ddPCR assay was able to detect 29 cells, followed by the conventional PCR assay (58 cells) and the qPCR assay (115 cells). Regarding the sensitivities to detect this pathogen from PCR inhibition prone samples (naturally contaminated mussels), the ddPCR assay significantly outperformed the conventional PCR and qPCR. The ddPCR assay was able to consistently detect non-pathogenic and pathogenic populations of V. parahaemolyticus from naturally contaminated mussels, indicating its tolerance to various PCR inhibitors. This study also revealed the significant difference between conventional PCR and qPCR. The conventional PCR assay showed significantly greater sensitivity than that of the qPCR assay in detecting V. parahaemolyticus in crude samples, whereas the qPCR assay showed better sensitivity in detecting the presence of V. parahaemolyticus in purified DNA samples.
Topics: Vibrio parahaemolyticus; Sensitivity and Specificity; Polymerase Chain Reaction; Seafood; DNA
PubMed: 38374337
DOI: 10.1038/s41598-024-54753-y -
Infection, Genetics and Evolution :... Apr 2024Vibrio parahaemolyticus is a Gram-negative, halophilic and polymorphic coccobacillus. It is world-widely distributed and has resulted in great economic losses since its...
Vibrio parahaemolyticus is a Gram-negative, halophilic and polymorphic coccobacillus. It is world-widely distributed and has resulted in great economic losses since its first appearance. In this study, a pathogenic strain was isolated from diseased pearl gentian grouper and identified as V. parahaemolyticus based on the sequencing results of 16S rDNA gene. In order to gain a comprehensive understanding of this isolation, the whole genome sequencing was conducted. Phylogenetic analysis of the complete genomes of 16 Vibrio species showed that LF1113, ATCC17802, ATCC33787, 2210633, FORC 004, and 160807 were the most closely related. Animal experiments demonstrated that the isolated LF1113 strain was pathogenic in a fish model. This study is the first study to describe the complete genome sequence of a V. parahaemolyticus isolate, which infected pearl gentian grouper from an outbreak in a fish factory farm in Hainan. The results will expand our understanding of genetic characteristics, pathogenesis, diagnostics and disease prevention of V. parahaemolyticus, and lay the foundation for further study.
Topics: Animals; Vibrio parahaemolyticus; Phylogeny; Genomics; Whole Genome Sequencing; Fishes; Vibrio Infections
PubMed: 38373468
DOI: 10.1016/j.meegid.2024.105574