Plesiomonas shigelloides, a Gram-negative bacillus, is the only member of the Enterobacteriaceae family able to produce polar and lateral flagella and cause gastrointestinal and extraintestinal illnesses in humans. The flagellar transcriptional hierarchy of P. shigelloides is currently unknown. In this study, we identified FlaK, FlaM, FliA, and FliAL as the four regulators responsible for polar and lateral flagellar regulation in P. shigelloides. To determine the flagellar transcription hierarchy of P. shigelloides, the transcriptomes of the WT and ΔflaK, ΔflaM, ΔfliA, and ΔfliAL were carried out for comparison in this study. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) and luminescence screening assays were used to validate the RNA-seq results, and the Electrophoretic Mobility Shift Assay (EMSA) results revealed that FlaK can directly bind to the promoters of fliK, fliE, flhA, and cheY, while the FlaM protein can bind directly to the promoters of flgO, flgT, and flgA. Meanwhile, we also observed type VI secretion system (T6SS) and type II secretion system 2 (T2SS-2) genes downregulated in the transcriptome profiles, and the killing assay revealed lower killing abilities for ΔflaK, ΔflaM, ΔfliA, and ΔfliAL compared to the WT, indicating that there was a cross-talk between the flagellar hierarchy system and bacterial secretion system. Invasion assays also showed that ΔflaK, ΔflaM, ΔfliA, and ΔfliAL were less effective in infecting Caco-2 cells than the WT. Additionally, we also found that the loss of flagellar regulators causes the differential expression of some of the physiological metabolic genes of P. shigelloides. Overall, this study aims to reveal the transcriptional hierarchy that controls flagellar gene expression in P. shigelloides, as well as the cross-talk between motility, virulence, and physiological and metabolic activity, laying the groundwork for future research into P. shigelloides\' coordinated survival in the natural environment and the mechanisms that infect the host.

A 21-year-old previously healthy Japanese woman visited an outpatient clinic because of abdominal pain, watery diarrhea, vomiting, and mild fever that had started on the previous day. She traveled to rural and urban areas of Rwanda and returned to Japan 3 days before. Stool culture yielded the Plesiomonas shigelloides strain TMCH301018, against which minimum inhibitory concentrations of cefotaxime and cefotaxime-clavulanate were 128 and ≤0.12/4 μg/mL, respectively. The strain had the blaCTX-M-27 gene and an IncA/C replicon-type plasmid. Moreover, a transformant produced by introduction of an IncA/C plasmid extracted from TMCH301018 into Escherichia coli DH5α was positive for the blaCTX-M-27 gene and fulfilled the criteria of extended-spectrum β-lactamase (ESBL) production described by the Clinical and Laboratory Standards Institute, indicating that TMCH301018 produced ESBL of CTX-M-27 and the ESBL-encoding gene was located on an IncA/C plasmid. Pathogenicity of TMCH301018 for the patient\'s complaints was uncertain because a molecular assay detected other enteropathogens in the stool specimen and the symptoms improved within 2 days with administration of oral ciprofloxacin, to which TMCH301018 was not susceptible. To our knowledge, this is the first report describing the isolation of ESBL-producing P. shigelloides.

Plesiomonas shigelloides, an aquatic bacterium belonging to the Enterobacteriaceae family, is a frequent cause of gastroenteritis with diarrhea and gastrointestinal severe disease. Despite decades of research, discovering a licensed and globally accessible vaccine is still years away. Developing a putative vaccine that can combat the Plesiomonas shigelloides infection by boosting population immunity against P. shigelloides is direly needed. In the framework of the current study, the entire proteome of P. shigelloides was explored using subtractive genomics integrated with the immunoinformatics approach for designing an effective vaccine construct against P. shigelloides. The overall stability of the vaccine construct was evaluated using molecular docking, which demonstrated that MEV showed higher binding affinities with toll-like receptors (TLR4: 51.5 ± 10.3, TLR2: 60.5 ± 9.2) and MHC receptors(MHCI: 79.7 ± 11.2 kcal/mol, MHCII: 70.4 ± 23.7). Further, the therapeutic efficacy of the vaccine construct for generating an efficient immune response was evaluated by computational immunological simulation. Finally, computer-based cloning and improvement in codon composition without altering amino acid sequence led to the development of a proposed vaccine. In a nutshell, the findings of this study add to the existing knowledge about the pathogenesis of this infection. The schemed MEV can be a possible prophylactic agent for individuals infected with P. shigelloides. Nevertheless, further authentication is required to guarantee its safeness and immunogenic potential.

The pyruvate dehydrogenase complex regulator (PdhR) was originally identified as a repressor of the pdhR-aceEF-lpd operon, which encodes the pyruvate dehydrogenase complex (PDHc) and PdhR itself. According to previous reports, PdhR plays a regulatory role in the physiological and metabolic pathways of bacteria. At present, the function of PdhR in Plesiomonas shigelloides is still poorly understood. In this study, RNA sequencing (RNA-Seq) of the wild-type strain and the ΔpdhR mutant strains was performed for comparison to identify the PdhR-controlled pathways, revealing that PdhR regulates ~7.38% of the P. shigelloides transcriptome. We found that the deletion of pdhR resulted in the downregulation of practically all polar and lateral flagella genes in P. shigelloides; meanwhile, motility assay and transmission electron microscopy (TEM) confirmed that the ΔpdhR mutant was non-motile and lacked flagella. Moreover, the results of RNA-seq and quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) showed that PdhR positively regulated the expression of the T3SS cluster, and the ΔpdhR mutant significantly reduced the ability of P. shigelloides to infect Caco-2 cells compared with the WT. Consistent with previous research, pyruvate-sensing PdhR directly binds to its promoter and inhibits pdhR-aceEF-lpd operon expression. In addition, we identified two additional downstream genes, metR and nuoA, that are directly negatively regulated by PdhR. Furthermore, we also demonstrated that ArcA was identified as being located upstream of pdhR and lpdA and directly negatively regulating their expression. Overall, we revealed the function and regulatory pathway of PdhR, which will allow for a more in-depth investigation into P. shigelloides pathogenicity as well as the complex regulatory network.

The consumption of raw seafood, generally considered to be a healthy food, has greatly increased worldwide. Pathogens of fish can cause foodborne illnesses in humans, especially following the consumption of raw seafood from contaminated water.Foodborne illness in pregnant women is seldom the cause of neonatal infection, but, as in the reported cases, it has been associated with a high degree of morbidity and mortality.We present the case of a newborn with septicemia and meningitis caused by Plesiomonas shigelloides acquired via the transplacental route. There was a maternal history of ingestion of raw seafood 1 week prior to delivery. A few similar cases are described in the existing literature, which reports 7 neonatal deaths.Therefore, the primary objective of this paper is to highlight the fact that the popularity of raw seafood such as sushi, sashimi, and oysters, requires an improvement in dietary advice regarding unsafe choices in pregnancy in order to avoid preventable foodborne diseases, sometimes fatal for the newborn.

RpoN, also known as σ54, first reported in Escherichia coli, is a subunit of RNA polymerase that strictly controls the expression of different genes by identifying specific promoter elements. RpoN has an important regulatory function in carbon and nitrogen metabolism and participates in the regulation of flagellar synthesis, bacterial motility and virulence. However, little is known about the effect of RpoN in Plesiomonas shigelloides.
To identify pathways controlled by RpoN, RNA sequencing (RNA-Seq) of the WT and the rpoN deletion strain was carried out for comparison. The RNA-seq results showed that RpoN regulates ~ 13.2% of the P. shigelloides transcriptome, involves amino acid transport and metabolism, glycerophospholipid metabolism, pantothenate and CoA biosynthesis, ribosome biosynthesis, flagellar assembly and bacterial secretion system. Furthermore, we verified the results of RNA-seq using quantitative real-time reverse transcription PCR, which indicated that the absence of rpoN caused downregulation of more than half of the polar and lateral flagella genes in P. shigelloides, and the ΔrpoN mutant was also non-motile and lacked flagella. In the present study, the ability of the ΔrpoN mutant to kill E. coli MG1655 was reduced by 54.6% compared with that of the WT, which was consistent with results in RNA-seq, which showed that the type II secretion system (T2SS-2) genes and the type VI secretion system (T6SS) genes were repressed. By contrast, the expression of type III secretion system genes was largely unchanged in the ΔrpoN mutant transcriptome and the ability of the ΔrpoN mutant to infect Caco-2 cells was also not significantly different compared with the WT.
We showed that RpoN is required for the motility and contributes to the killing ability of P. shigelloides and positively regulates the T6SS and T2SS-2 genes.

The outbreak of mass mortality of M. salmoides occurred in an aquaculture farm in Jiangsu province of China, showing signs of skin ulceration and haemorrhages. The bacteria were isolated from diseased largemouth bass, and identified as Plesiomonas shigelloides based on morphological, physiological and biochemical features, as well as 16S rRNA gene sequence analysis. The pathogenicity of P. shigelloides was determined by challenge experiments, and the median lethal dosage (LD50) of the isolate NJS1 for M. salmoides was calculated as 1.6 × 105 CFU/mL at 7 d post-infection. Histopathological analysis revealed that extensive necrosis, vacuolization and inflammation were presented in the kidney, liver and gill of the diseased fish. Detection of virulence-related genes showed that P. shigelloides NJS1 was positive for astA, astB, astD, astE, actP and 6 ahpA. Additionally, the host defensive response of M. salmoides infected by P. shigelloides was analyzed by quantitive real-time PCR (qRT-PCR), and the results showed that the expression levels of Cas3, Hep1, HIF, IgM, IL15 and TGF were significantly up-regulated in head kidney, liver and spleen in different hours post-infection, which revealed varying expression profiles and clear transcriptional activation of immune related genes. The results suggested that P. shigelloides was an etiological element in the mass mortalities of M. salmoides and this study provided deeper insights for the pathogenesis and host defensive system in P. shigelloides invasion.

The swift emergence of antibiotic resistance (AR) in bacterial pathogens to make themselves adaptable to changing environments has become an alarming health issue. To prevent AR infection, many ways can be accomplished such as by decreasing the misuse of antibiotics in human and animal medicine. Among these AR bacterial species, Plesiomonas shigelloides is one of the etiological agents of intestinal infection in humans. It is a gram-negative rod-shaped bacterium that is highly resistant to several classes of antibiotics, and no licensed vaccine against the aforementioned pathogen is available. Hence, substantial efforts are required to screen protective antigens from the pathogen whole genome that can be subjected easily to experimental evaluations. Here, we employed a reverse vaccinology (RV) approach to design a multi-antigenic epitopes based vaccine against P. shigelloides. The complete genomes of P. shigelloides were retrieved from the National Center for Biotechnological Information (NCBI) that on average consist of 5226 proteins. The complete proteomes were subjected to different subtractive proteomics filters, and in the results of that analysis, out of total proteins, 2399 were revealed as non-redundant and 2827 as redundant proteins. The non-redundant proteins were further checked for subcellular localization analysis, in which three were localized in the extracellular matrix, eight were outer membrane, and 13 were found in the periplasmic membrane. All surface localized proteins were found to be virulent. Out of a total of 24 virulent proteins, three proteins (flagellar hook protein (FlgE), hypothetical protein, and TonB-dependent hemoglobin/transferrin/lactoferrin family receptor protein) were considered as potential vaccine targets and subjected to epitopes prediction. The predicted epitopes were further examined for antigenicity, toxicity, and solubility. A total of 10 epitopes were selected (GFKESRAEF, VQVPTEAGQ, KINENGVVV, ENKALSQET, QGYASANDE, RLNPTDSRW, TLDYRLNPT, RVTKKQSDK, GEREGKNRP, RDKKTNQPL). The selected epitopes were linked with each other via specific GPGPG linkers in order to design a multi-epitopes vaccine construct, and linked with cholera toxin B subunit adjuvant to make the designed vaccine construct more efficient in terms of antigenicity. The 3D structure of the vaccine construct was modeled ab initio as no appropriate template was available. Furthermore, molecular docking was carried out to check the interaction affinity of the designed vaccine with major histocompatibility complex (MHC-)I (PDB ID: 1L1Y), MHC-II (1KG0), and toll-like receptor 4 ((TLR-4) (PDB: 4G8A). Molecular dynamic simulation was applied to evaluate the dynamic behavior of vaccine-receptor complexes. Lastly, the binding free energies of the vaccine with receptors were estimated by using MMPB/GBSA methods. All of the aforementioned analyses concluded that the designed vaccine molecule as a good candidate to be used in experimental studies to disclose its immune protective efficacy in animal models.

Plesiomonas shigelloides is a gram-negative facultative anaerobic bacillus, usually found in soil and freshwater, which causes self-limited diarrhea, although reports of bacteremia are rare. Here, we report the first case of an intratumoral abscess with mixed bacteremia caused by P. shigelloides, Citrobacter freundii, Streptococcus mitis/oralis, Clostridium perfringens, and Candida albicans in a patient with recurrent postoperative cholangiocarcinoma. A 77-year-old man with hilar cholangiocarcinoma and hypertension was admitted to our hospital with fever and abdominal pain. He had visited Vietnam for 3 years, 20 years ago. Abdominal computed tomography showed air within the recurrent tumor at the left liver lobectomy resection margin site, which was diagnosed as an intratumor abscess perforating the intestinal tract. P. shigelloides, C. freundii, S. mitis/oralis, C. perfringens, and C. albicans were isolated in blood culture. P. shigelloides was identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and 16S ribosomal RNA (16S rRNA) sequencing. Piperacillin-tazobactam was administered for almost a week, ampicillin-sulbactam and levofloxacin for almost 3 weeks, and antifungal agents for almost 2 weeks, and the patient was discharged thereafter. Although bloodstream infections caused by P. shigelloides in patients with cancer are extremely rare, long-term colonization and the potential for future intra-abdominal infections were implicated.

The genus Plesiomonas, represented by a single species, Plesiomonas shigelloides, is a gram-negative bacillus associated with gastrointestinal and extraintestinal diseases in humans. In this study, 44 clinical isolates (gastrointestinal, n = 41; extraintestinal, n = 3) were genetically confirmed to be P. shigelloides using the hug gene. All 20 virulence genes were detected in the gastrointestinal isolates, ranging from 7.7% to 100%; however, only 12 genes were detected in the extra-gastrointestinal isolates, ranging from 33.3% to 100%. The phlA gene was significantly associated with the gastrointestinal isolates (P = 0.0216). The results of this study suggest that phlA may play a role in gastrointestinal infections. However, pilF, tolC, and fur were detected in both gastrointestinal and extraintestinal clinical isolates, and further investigations are warranted to elucidate their role in the pathogenesis of P. shigelloides.