Search Results (20)

Vibrio species are among the primary pathogenic bacteria affecting abalone aquaculture, posing significant threats to farming practices. Current clinical control predominantly relies on antibiotics, which can result in antibiotic residues in both abalone and the surrounding marine environments. Lactobacillus plantarum (LP) has been shown to release bioactive antagonistic substances and exhibits potent inhibitory effects against marine pathogenic bacteria. This study aimed to screen and characterize the probiotic properties of LP strains isolated from rice wine lees to develop a novel biocontrol strategy against Vibriosis in abalone. The methods employed included selective media cultivation, streak plate isolation, and single-colony purification for strain screening, followed by Gram staining, 16S rDNA sequencing, and phylogenetic tree construction using MEGA11 for identification. The resilience, antimicrobial activity, and in vivo antagonistic efficacy of the strains were evaluated through stress tolerance assays, agar diffusion tests, and animal experiments. The results demonstrated the successful isolation and purification of four LP strains (NDMJ-1 to NDMJ-4). Phylogenetic analysis revealed closer genetic relationships between NDMJ-3 and NDMJ-4, while NDMJ-1 and NDMJ-2 were found to be more distantly related. All strains exhibited γ-hemolytic activity, bile salt tolerance (0.3–3.0%), and resistance to both acid (pH 2.5) and alkali (pH 8.5), although they were temperature sensitive (inactivated above 45 °C). The strains showed susceptibility to most of the 20 tested antibiotics, with marked variations in hydrophobicity (1.91–93.15%) and auto-aggregation (13.29–60.63%). In vitro antibacterial assays revealed that cell-free supernatants of the strains significantly inhibited Vibrio parahaemolyticus, V. alginolyticus, and V. natriegens, with NDMJ-4 displaying the strongest inhibitory activity. In vivo experiments confirmed that NDMJ-4 significantly reduced mortality in abalone infected with V. parahaemolyticus. In conclusion, the LP strains isolated from rice wine lees (NDMJ-1 to NDMJ-4) possess robust stress resistance, adhesion capabilities, and broad antibiotic susceptibility. Their metabolites exhibit significant inhibition against abalone-pathogenic Vibrios, particularly NDMJ-4, which demonstrates exceptional potential as a candidate strain for developing eco-friendly biocontrol agents against Vibriosis in abalone aquaculture. Full article

The prevention of biofouling (biological fouling) became a major economic and environmental issue. In the present study, we designed a series of four cyclam and cyclen derivatives with a modulation of their lipophilicity by introducing either two benzyl (Bn) groups or two tetradecyl (C₁₄) chains in the structure to produce (Cyclam(Bn)₂, Cyclam(C₁₄)₂, Cyclen(Bn)₂ and Cyclen(C₁₄)₂). Additionally, copper (Cu) and zinc (Zn) complexes of each compound were prepared and evaluated as potential antifouling candidates against two models of Vibrio species (V. natriegens and V. aestuarianus). The results highlight that no significant antifouling activity was measured for the metal free polyazamacrocyclic derivatives. However, for the metal complexes, the nature of the cation (Cu²⁺ or Zn²⁺) modulates both the growth and adhesion capacities of the two bacteria. Overall, in most cases, Zn complexes showed better activity than the Cu complexes, revealing the importance of the metal cation. Moreover, in the cyclam series, the anti-adhesion properties could be linked to a biocidal effect while a full anti-adhesion activity was observed in the cyclen series. Full article

Vibrio natriegens (V. natriegens) is an emerging synthetic biology chassis characterized by rapid growth, and its potential for the synthesis of sesquiterpenes (such as pentalenene) has not been developed. In this study, heterologous pentalenene biosynthesis was successfully established in V. natriegens via metabolic engineering. The optimization of gene dosage and culture conditions led to an increase in pentalenene yield from 0.75 mg/L to 39.4 mg/L, representing the highest titer reported in V. natriegens to date, though still markedly lower than yields achieved in conventional microbial hosts. Transcriptome analysis demonstrated that the exogenous mevalonate (MVA) pathway effectively activated terpenoid precursor synthesis, as evidenced by the up-regulation of key pathway genes. However, the endogenous methylerythritol 4-phosphate (MEP) pathway remained inactive, and genes involved in oxidative phosphorylation, the pentose phosphate pathway, and thiamine biosynthesis were down-regulated, leading to limited availability of ATP, NADPH, and acetyl-CoA. Competition for cofactors, particularly NADPH, further constrained precursor supply and pathway efficiency. This study confirmed the potential of V. natriegens as a pentalenene production platform and revealed its metabolic bottleneck, providing a theoretical basis for subsequent engineering optimization. Full article

Vibrio natriegens is widely used as a production host for biotechnological processes due to its superior maximum glucose consumption rate, high growth rate, and abundant ribosomes. Most bioprocesses also need a scalable biomass separation step. This can be achieved by cross-flow filtration with ceramic membranes, although the membrane pores are susceptible to fouling. However, the fouling characteristics of V. natriegens culture broth have not been investigated in detail. We therefore characterized membrane fouling during the separation of V. natriegens biomass from culture broth using a cross-flow filtration plant with ceramic membranes. The resistance in series model was used to quantify the fouling-induced resistance caused by the different components of the culture broth. The total fouling resistance was 4.1·10⁹ ± 0.6·10⁹ m⁻¹ for the culture broth and 5.4·10⁹ ± 0.7·10⁹ m⁻¹ for the summed broth components. Reversible resistance accounted for 86% and 81% of these totals, respectively. We then applied Hermia’s adapted filtration laws to determine the dominant fouling mechanism induced by the different broth components. In a further step, we established a setup to determine the compressibility index of the cells during cross-flow filtration, resulting in an estimated value of 0.55 ± 0.04. These results will facilitate the design of economic filtration plants and will help to establish V. natriegens as a production host for large-scale industrial processes. Full article

DNA amplification and reverse transcription enzymes have proven to be invaluable in fast and reliable diagnostics and research applications because of their processivity, specificity, and robustness. Our study focused on the production of mutant Taq DNA polymerase and mutant M-MLV reverse transcriptase in the expression hosts Vibrio natriegens and Escherichia coli under various expression conditions. We also examined nonspecific extracellular production in V. natriegens. Intracellularly, M-MLV was produced in V. natriegens at the level of 11% of the total cell proteins (TCPs) compared with 16% of TCPs in E. coli. We obtained a soluble protein that accounted for 11% of the enzyme produced in V. natriegens and 22% of the enzyme produced in E. coli. Taq pol was produced intracellularly in V. natriegens at the level of 30% of TCPs compared with 26% of TCPs in E. coli. However, Taq pol was almost non-soluble in E. coli, whereas in V. natriegens, we obtained a soluble protein that accounted for 23% of the produced enzyme. We detected substantial extracellular production of Taq pol. Thus, V. natriegens is a suitable alternative host with the potential for production of recombinant proteins. Full article

Agar oligosaccharides from the degradation of agar harbor great potential in the food and pharmaceutical industries. An agar-degrading bacterium, Vibrio natriegens WPAGA4, was isolated from the deep sea in our previous work. However, the agar-degrading activity of WPAGA4 remains to be improved for more production benefits of this strain. The aim of this study was to enhance the agar-degrading activity of WPAGA4 by using atmospheric and room temperature plasma (ARTP) mutagenesis. Three mutant strains, including T1, T2, and T3, with good genetic stability were obtained, and the agar-degrading activities of these strains increased by 136%, 141%, and 135%, respectively. The optimal temperature and pH for agar degradation were slightly changed in the mutant strains. No sequence mutation was detected in all the agarase genes of WPAGA4, including agaW3418, agaW3419, agaW3420, and agaW3472. However, ARPT mutagenesis increased the relative expression levels of agaW3418, agaW3419, and agaW3420 in the mutant strains, which could be the reason for the improvement of degradation activities in the mutant strains. Furthermore, T3 had the lowest consumption rate of agar oligosaccharide, which was 21% less than the wild-type strain. Therefore, T3 possessed a preferable production value due to its higher degrading activity and lower consumption of agar oligosaccharides. The current work enhanced the agar-degrading activity of WPAGA4 and offered strains with greater potential for agar oligosaccharide production, thereby laying the foundation for industrial applications. Full article

Different bacterial species have dramatically different generation times, from 20–30 min in Escherichia coli to about two weeks in Mycobacterium leprae. The translation machinery in a cell needs to synthesize all proteins for a new cell in each generation. The three subprocesses of translation, i.e., initiation, elongation, and termination, are expected to be under stronger selection pressure to optimize in short-generation bacteria (SGB) such as Vibrio natriegens than in the long-generation Mycobacterium leprae. The initiation efficiency depends on the start codon decoded by the initiation tRNA, the optimal Shine–Dalgarno (SD) decoded by the anti-SD (aSD) sequence on small subunit rRNA, and the secondary structure that may embed the initiation signals and prevent them from being decoded. The elongation efficiency depends on the tRNA pool and codon usage. The termination efficiency in bacteria depends mainly on the nature of the stop codon and the nucleotide immediately downstream of the stop codon. By contrasting SGB with long-generation bacteria (LGB), we predict (1) SGB to have more ribosome RNA operons to produce ribosomes, and more tRNA genes for carrying amino acids to ribosomes, (2) SGB to have a higher percentage of genes using AUG as the start codon and UAA as the stop codon than LGB, (3) SGB to exhibit better codon and anticodon adaptation than LGB, and (4) SGB to have a weaker secondary structure near the translation initiation signals than LGB. These differences between SGB and LGB should be more pronounced in highly expressed genes than the rest of the genes. We present empirical evidence in support of these predictions. Full article

An innovative antifouling composite coating comprising dopamine (DA), polyethyleneimine (PEI), and silica (SiO₂) was developed through a straightforward and environmentally friendly approach. Initially, silica nanoparticles comodified with DA and PEI were meticulously deposited onto 304 stainless steel surfaces pretreated with dopamine to achieve a uniformly distributed nanocomposite surface. Comprehensive analytical techniques, including Fourier transform infrared spectrometry (FTIR), X-ray photoelectron spectrometry (XPS), field emission scanning electron microscopy (SEM), contact angle measurement (CA), and 3D optical profilometry, were employed to affirm the successful preparation of the silica nanocomposite coatings and the effective grafting of MAG II. The antibacterial and antibiofilm performance of the DA/PEI/SiO₂-modified surface was rigorously assessed using Vibrio natriegens (V. natriegens), yielding compelling results indicating a substantial 51.4% reduction in biofilm formation on the SS-DA/PEI/SiO₂ sample surfaces, coupled with an impressive 95.2% decrease in V. natriegens adhesion. This pioneering research introduces an innovative strategy for the development of antimicrobial surfaces with promising applications in medical devices, aquaculture, and related fields. Full article

Vibrio natriegens is the fastest-growing bacterium, with a doubling time of approximately 12–14 min. It has a high potential for basic research and biotechnological applications, e.g., it can be used for the cell-free production of (labeled) heterologous proteins, for synthetic biological applications, and for the production of various compounds. However, the ploidy level in V. natriegens remains unknown. At nine time points throughout the growth curve, we analyzed the numbers of origins and termini of both chromosomes with qPCR and the relative abundances of all genomic sites with marker frequency analyses. During the lag phase until early exponential growth, the origin copy number and origin/terminus ratio of chromosome 1 increased severalfold, but the increase was lower for chromosome 2. This increase was paralleled by an increase in cell volume. During the exponential phase, the origin/terminus ratio and cell volume decreased again. This highly dynamic and fast regulation has not yet been described for any other species. In this study, the gene dosage increase in origin-adjacent genes during the lag phase is discussed together with the nonrandom distribution of genes on the chromosomes of V. natriegens. Taken together, the results of this study provide the first comprehensive overview of the chromosome dynamics in V. natriegens and will guide the optimization of molecular biological characterization and biotechnological applications. Full article

The Pacific oyster (Crassostrea gigas) aquaculture industry increased rapidly in China with the introduction and promotion of triploid oysters in recent years. Mass mortalities affecting different life stages of Pacific oysters emerged periodically in several important production areas of Northern China. During 2020 and 2021, we conducted a passive two-year investigation of infectious pathogens linked to mass mortality. Ostreid herpesvirus-1 (OsHV-1) was detected to be associated with mass mortalities of hatchery larvae, but not juveniles and adults in the open sea. Protozoan parasites, such as Marteilia spp., Perkinsus spp. and Bonamia spp. were not detected. Bacterial isolation and identification revealed that Vibrio natriegens and Vibrio alginolyticus were the most frequently (9 out of 13) identified two dominant bacteria associated with mass mortalities. Pseudoalteromonas spp. was identified as the dominant bacteria in three mortality events that occurred during the cold season. Further bacteriological analysis was conducted on two representative isolates of V. natriegens and V. alginolyticus, designated as CgA1-1 and CgA1-2. Multisequence analysis (MLSA) showed that CgA1-1 and CgA1-2 were closely related to each other and nested within the Harveyi clade. Bacteriological investigation revealed faster growth, and more remarkable haemolytic activity and siderophore production capacity at 25 °C than at 15 °C for both CgA1-1 and CgA1-2. The accumulative mortalities of experimental immersion infections were also higher at 25 °C (90% and 63.33%) than at 15 °C (43.33% and 33.33%) using both CgA1-1 and CgA1-2, respectively. Similar clinical and pathological features were identified in samples collected during both naturally and experimentally occurring mortalities, such as thin visceral mass, discolouration, and connective tissue and digestive tube lesions. The results presented here highlight the potential risk of OsHV-1 to hatchery production of larvae, and the pathogenic role of V. natriegens and V. alginolyticus during mass mortalities of all life stages of Pacific oysters in Northern China. Full article

The outbreaks caused by Vibrio spp. are a notable threat to the potential growth of the economy of penaeid culture, which is still controlled by the administration of antibiotics. At first, the infected group was subjected to phenotypic bacteriological examination with subsequent molecular identification via 16S rRNA gene sequencing, which confirmed four strains of Vibrio spp., V. atlanticus, V. natriegens, V. alginolyticus, and V. harveyi, from moribund-infected shrimp during mortality events in an Egyptian hatchery. To better understand the defense mechanism of the most effective antibiotic against Vibrio strains, the immune responses were compared and evaluated in infected Litopenaeus vannamei broodstock after being fed 5 mg kg⁻¹ of florfenicol antibiotic, which was first determined through in vitro antibiogram tests. Therefore, our study aimed to determine the immune response of L. vannamei during Vibrio spp. infection in Egyptian hatcheries and after antibiotic medication. The parameters assessed were the total and differential hemocyte count (THC), granular cells (GC), semi-granular cells (SGC), and hyaline cells (HC). As well as the metabolic and immune enzymes: alanine aminotransferases (ALT), aspartate aminotransferases (AST), alkaline phosphatase (ALP), acid phosphatase (ACP), and lysozyme activity; an antioxidant index, such as superoxide dismutase (SOD) and glutathione (GSH); a phagocytic assay; changes in reactive oxygen species (ROS); and bactericidal activity in the hemolymph of the control, infected, and treated groups. Further evaluation of the mRNA expression levels of the prophenoloxidase (LvproPO), toll-like receptor 1 (LvToll1), and haemocyanin (LvHc) genes were performed in the hepatopancreas of the same groups. A significant drop in the THC, GC, SGC, and HC counts, as well as lysozyme and bactericidal activities, phagocytic assay, ROS, SOD, and GSH index, were represented in infected shrimp compared to control shrimp; however, a marked increase in the activity of ALT, AST, ALP, and ACP was observed. These activities were significantly restored in the treated shrimp compared to the infected shrimp. Nevertheless, no significant changes were noted in the transcriptional levels of the LvproPO and LvToll1 genes in the treated shrimp when compared to the infected shrimp; however, a significant suppression of the LvHc gene was noted. Our study aimed to determine the immune response of L. vannamei during Vibrio spp. infection in Egyptian hatcheries and after antibiotic medication. We concluded that florfenicol in medicated feed could be effective in controlling vibriosis and ameliorating the immune response of shrimp. Full article

The development of MFC using electroactive industrial microorganisms has seen a surge of interest because of the co-generation for bioproduct and electricity production. Vibrio natriegens as a promising next-generation industrial microorganism chassis and its application for microbial fuel cells (MFC) was first studied. Mediated electron transfer was found in V. natriegens MFC (VMFC), but V. natriegens cannot secrete sufficient electron mediators to transfer electrons to the anode. All seven electron mediators supplemented are capable of improving the electronic transfer efficiency of VMFC. The media and carbon sources switching study reveals that VMFCs have excellent bioelectricity generation performance with feedstock flexibility and high salt-tolerance. Among them, 1% glycerol as the sole carbon source produced the highest power density of 111.9 ± 6.7 mW/cm². The insight of the endogenous electronic mediators found that phenazine-1-carboxamide, phenazine-1-carboxylic acid, and 1-hydroxyphenazine are synthesized by V. natriegens via the shikimate pathway and the phenazine synthesis and modification pathways. This work provides the first proof for emerging industrial biotechnology chassis V. natriegens as a novel high salt-tolerant and feedstock flexibility electroactive microorganism for MFC, and giving insight into the endogenous electron mediator biosynthesis of VMFC, paving the way for the application of V. natriegens in MFC and even microbial electrofermentation (EF). Full article

Phage-based pathogen control (i.e., phage therapy) has received increasing scientific attention to reduce and prevent the emergence, transmission, and detrimental effects of antibiotic resistance. In the current study, multidrug-resistant Vibrio natriegens strain AbY-1805 was isolated and tentatively identified as a pathogen causing the death of juvenile Pacific abalones (Haliotis discus hannai Ino). In order to apply phage therapy, instead of antibiotics, to treat and control V. natriegens infections in marine aquaculture environments, a lytic phage, vB_VnaS-L3, was isolated. It could effectively infect V. natriegens AbY-1805 with a short latent period (40 min) and high burst size (~890 PFU/cell). Treatment with vB_VnaS-L3 significantly reduced the mortality of juvenile abalones and maintained abalone feeding capacity over a 40-day V. natriegens challenge experiment. Comparative genomic and phylogenetic analyses suggested that vB_VnaS-L3 was a novel marine Siphoviridae-family phage. Furthermore, vB_VnaS-L3 had a narrow host range, possibly specific to the pathogenic V. natriegens strains. It also exhibited viability at a wide range of pH, temperature, and salinity. The short latent period, large burst size, high host specificity, and broad environmental adaptation suggest that phage vB_VnaS-L3 could potentially be developed as an alternative antimicrobial for the control and prevention of marine animal infections caused by pathogenic V. natriegens. Full article

Up until now, the characterizations of GH50 agarases from Vibrio species have rarely been reported compared to GH16 agarases. In this study, a deep-sea strain, WPAGA4, was isolated and identified as Vibrio natriegens due to the maximum similarity of its 16S rRNA gene sequence, the values of its average nucleotide identity, and through digital DNA–DNA hybridization. Two circular chromosomes in V. natriegens WPAGA4 were assembled. A total of 4561 coding genes, 37 rRNA, 131 tRNA, and 59 other non-coding RNA genes were predicted in the genome of V. natriegens WPAGA4. An agarase gene belonging to the GH50 family was annotated in the genome sequence and expressed in E. coli cells. The optimum temperature and pH of the recombinant Aga3420 (rAga3420) were 40 °C and 7.0, respectively. Neoagarobiose (NA2) was the only product during the degradation process of agarose by rAga3420. rAga3420 had a favorable stability following incubation at 10–30 °C for 50 min. The Km, Vmax, and kcat values of rAga3420 were 2.8 mg/mL, 78.1 U/mg, and 376.9 s⁻¹, respectively. rAga3420 displayed cold-adapted properties as 59.7% and 41.2% of the relative activity remained at 10 3 °C and 0 °C, respectively. This property ensured V. natriegens WPAGA4 could degrade and metabolize the agarose in cold deep-sea environments and enables rAga3420 to be an appropriate industrial enzyme for NA2 production, with industrial potential in medical and cosmetic fields. Full article

Vibrio is the most common bacterium associated with diseases in crustaceans. Outbreaks of vibriosis pose a serious threat to shrimp production. Therefore, antibiotics are commonly used as preventative and therapeutic measures. Unfortunately, improper use of antibiotics leads to antibiotic resistance. Nevertheless, information on the occurrence of Vibrio spp. and antibiotic use in shrimp, particularly in Malaysia, is minimal. This study aimed to provide information on the occurrence of Vibrio spp., its status of antibiotic resistance and the plasmid profiles of Vibrio spp. isolated from cultured shrimp in Peninsular Malaysia. Shrimp were sampled from seven farms that were located in different geographical regions of Peninsular Malaysia. According to the observations, 85% of the shrimp were healthy, whereas 15% were unhealthy. Subsequently, 225 presumptive Vibrio isolates were subjected to biochemical tests and molecular detection using the pyrH gene. The isolates were also tested for antibiotic susceptibility against 16 antibiotics and were subjected to plasmid profiling. Eventually, 13 different Vibrio spp. were successfully isolated and characterized using the pyrH gene. They were the following: V. parahaemolyticus (55%), V. communis (9%), V. campbellii (8%), V. owensii (7%), V. rotiferianus (5%), Vibrio spp. (4%), V. alginolyticus (3%), V. brasiliensis (2%), V. natriegens (2%), V. xuii (1%), V. harveyi (1%), V. hepatarius (0.4%) and P. damselae (3%). Antibiotic susceptibility profiles revealed that all isolates were resistant to penicillin G (100%), but susceptible to norfloxacin (96%). Furthermore, 16% of the isolates revealed MAR of less than 0.2, while 84% were greater than 0.2. A total of 125 isolates harbored plasmids with molecular weights between 1.0 and above 10 kb, detected among the resistant isolates. The resistant isolates were mediated by both chromosomal and plasmid factors. These findings support the use of surveillance data on the emerging patterns of antimicrobial-resistance and plasmid profiles of Vibrio spp. in shrimp farms. The findings from this study can be used to develop a better disease management strategy for shrimp farming. Full article