Plesiomonas: A Review on Food Safety, Fish-Borne Diseases, and Tilapia.

Fish and fish products are considered a fundamental part of the human diet due to their high nutritional value. Food-borne diseases are considered a major public health challenge worldwide due to their incidence, associated mortality, and negative economic repercussions. Food safety is the guarantee that foods will not cause harm to the health of those who consume them, and it is a fundamental property of food quality. Food safety can be at risk of being lost at any stage of the food chain if the food is contaminated by pathogenic microorganisms. Many diverse bacteria are present in the environment and as part of the microbiota of food that can be transmitted to humans during the handling and consumption of food. Plesiomonas shigelloides has been mainly associated with outbreaks of gastrointestinal diseases due to the consumption of fish. This bacterium inhabits the environment and aquatic animals and is associated with the microbiota of fish such as tilapia, a fish of importance in fishing, aquaculture, commercialization, and consumption worldwide. The purpose of this document is to provide, through a bibliographic review of databases (Scopus, Web of Science, and Google Scholar, among others), a general informative perspective on food-borne diseases and, in particular, the consumption of fish and tilapia. Diseases derived from contamination by Plesiomonas shigelloides are included, and control and prevention actions and sanitary regulations for fishery products established in several countries around the world are discussed to promote the safety of foods of aquatic origin intended for human consumption and to protect public health.

Modelling the effects of physicochemical variables and anthropogenic activities as ecological drivers of Plesiomonas shigelloides distribution and freshwaters quality.

Spatio-temporal distribution of pathogens in freshwater is driven by environmental variables (EVs), natural, and human-induced activities and the spread of infections and disease outbreaks are triggered significantly by these processes. The role of EVs and anthropogenic activities on the distribution of Plesiomonas shigelloides is not well understood; hence this study aimed at modelling the effects of EVs and anthropogenic gradients on the densities of Plesiomonas in freshwaters and freshwater quality. Three freshwaters were sampled from February to December 2017. The EVs and Plesiomonas densities of the freshwaters were determined using standard techniques, while partial least square path modelling and correlation analysis were performed on the data collected. Factors underpinning the quality of the freshwaters were identified through principal component analysis (PCA). Most EVs fell within the bounds of recommended permissible limits except turbidity, TSS, salinity and TDS. Results revealed a significant increase of Plesiomonas densities with an increase in the magnitude of path coefficients and intensities of anthropogenic activities along the freshwaters. The distribution of Plesiomonas correlated with temperature (r = 0.69, p < 0.01), TSS (r = 0.30, p < 0.01), TBS (r = 0.28, p < 0.01), and BOD (r = 0.39, p < 0.01). Similarly, a significant correlation existed between conductivity and TDS (r = 0.97, p < 0.01) and salinity (r = 0.99, p < 0.01). Network analysis of EVs identified three closed networks essential for freshwaters quality and Plesiomonas distribution with nodes of variables under synergistic latent influences. Overall, PCA identified four drivers of the freshwater quality and in part, Plesiomonas density; namely, nutrient loading; thermal and organic pollutions, aesthetic pollution, and pH modulators. Higher component score indicated a greater impact of nutrient loading on the freshwater quality. The study concluded that Plesiomonas distribution is largely shaped by anthropogenic gradients and EVs in rivers, and these may play a major role in its dissemination along freshwater milieus.

Plesiomonas shigelloides hugZ encodes an iron-regulated heme binding protein required for heme iron utilization.

Plesiomonas shigelloides is an intestinal pathogen that uses heme as an iron source. The P. shigelloides heme utilization system consists of 10 genes, 7 of which permit heme transport and 3 of which are associated with utilization of heme as an iron source once it is inside the cell. The goal of this study was to examine hugZ, 1 of the 3 genes associated with utilization of heme iron. DPH8, a hugZ mutant, failed to grow to full cell density in media containing heme as the iron source, indicating that hugZ is required for heme iron utilization. Western blots using antibodies against Vibrio cholerae HutZ to detect the P. shigelloides HugZ indicated that hugZ encodes an iron-regulated cytoplasmic protein, which is absent in DPH8. A heme affinity bead assay performed on soluble protein fractions from P. shigelloides DPH8/pHUG24.5 (pHUG24.5 encodes hugZ) indicated that HugZ binds heme. Heme utilization was restored in DPH8 by hox1, which encodes the alpha-heme oxygenase from Synechocystis sp. strain PCC6803. However, HugZ did not exhibit alpha-heme oxygenase activity in an assay that detects the conversion of heme to the bilin functional group present in phycobiliproteins. These results do not rule out that HugZ exhibits another type of heme oxygenase activity not detected in the assay.

Cell-associated hemolytic activity in environmental strains of Plesiomonas shigelloides expressing cell-free, iron-influenced extracellular hemolysin.

Hemolysis is a means of providing pathogenic bacteria with heme iron in vivo. In a previous work, iron-influenced hemolytic activity against sheep erythrocytes was detected in cell-free supernatants, but not in the cell fraction of two environmental Plesiomonas shigelloides strains incubated without shaking. Both strains have the hugA gene, which encodes an outer membrane receptor required for heme iron utilization. The present study was undertaken to investigate the expression of a second hemolytic activity detected during aerated incubation in normal and iron-depleted tryptone soya broth (id-TSB). An agar overlay procedure and doubling dilution titrations were employed to detect the hemolytic activity against several erythrocyte species. The kinetics of growth and hemolytic activity were assayed at 35 degrees C in aerated normal and id-TSB and salmon extract. Overlaid colonies showed a cell-associated beta-hemolytic activity within 4 h. For aerated cell-free supernatants, titers above 16 were not attained until 30 to 48 h of incubation; the best activity was noted with dog and mouse erythrocytes. After 24 h of aerated incubation, sonicated cells yielded high hemolytic activity against dog erythrocytes without activity in supernatants, but after 48 h, only 28 to 30% of the total activity remained cell associated. The hemolytic factor was released in broths during the death phase. Hemolytic activity was not detected in fish extract. This and other studies suggest that P. shigelloides may produce at least two hemolytic factors, their expression and detection being influenced by environmental growth conditions and testing procedures. The overlay assay appears to be the best routine method for detecting hemolytic activity in P. shigelloides.

[Effect of perfluorodecalin, carbogal, and perfluoromethyldecalin on growth and ice-forming activity of bacteria].

The isolates of pseudomonads (56) and of Plesiomonas shigelloides (7) exhibiting ice-forming activity were isolated from plant leaves and rhizosphere. The theoretical possibility of the application of perfluorocarbons (PFC) with a gas-transporting function (perfluorodecalin, carbogal, and perfluoromethyldecalin) for the cultivation of bacteria with different levels of ice-forming activity (IFA) in order to enhance their growth rates, biomass yields, and IFA was demonstrated. Introduction of 5% perfluorodecalin, carbogal, or perfluoromethyldecalin resulted for two strains in a 1.7-3.1-fold increase in biomass and a 3.2-24.5-fold increase in ice-forming activity compared to the control (without PFC).

Serotypes and anti-microbial susceptibility of Plesiomonas shigelloides isolates from humans, animals and aquatic environments in different countries.

A total of 73 strains of Plesiomonas shigelloides isolated from humans (24 strains) animals (21 strains) and aquatic environment (28 strains) were determined for their O:H serotype and susceptibility to 18 anti-microbial substances and to the vibriostatic agent O/129. Of all strains, 86.3% were typeable by the O and 94.5% by the H anti-sera used. The serotype distribution was heterogeneous within a country and between the countries. Of the 57 different serotypes identified, O11:H2 (2 strains), O22:H3 (4 strains), O35:HH11 (2 strains), O52:H3 (2 strains) and O90:H6 (2 strains) were found among isolates from humans and animals (mainly in cats) in Finland and Cuba, and O23:H1a1b (3 strains) among isolates from environmental sources in Slovak Republic and Italy. Most (93-100%) of all strains were susceptible to all anti-microbials tested but resistant (92-96%) to the broad-spectrum penicillins (ampicillin, mezlocillin). No correlation between anti-microbial resistance patterns and serotypes was found.

Some properties of Plesiomonas shigelloides treated with aminoglycosides.

The effect of aminoglycoside antibiotics (amikacin, gentamicin, netilmicin and tobramycin) at sublethal concentrations (sub-MICs) on some properties of Plesiomonas shigelloides strains was evaluated. All agents decreased the bacterial surface hydrophobicity. Amikacin (1/4 of the MIC) and netilmicin (1/4 and 1/8 of the MIC) changed the hydrophobic character of P. shigelloides surface to a hydrophilic one. Treatment of the strains with aminoglycosides decreased also motility, netilmicin being the most effective. No significant changes were found in lipolytic activity of antibiotic-treated strains. In the majority of cases aminoglycosides increased sensitivity of bacteria to hydrogen peroxide. The tested antibiotics did not induce production of short-chained N-acylhomoserine lactones signal molecules. Aminoglycosides at sub-MICs affected important activities of P. shigelloides potentially associated with their virulence in dependence on strain, antibiotic and concentration.

Characterization of the Plesiomonas shigelloides genes encoding the heme iron utilization system.

Plesiomonas shigelloides is a gram-negative pathogen which can utilize heme as an iron source. In previous work, P. shigelloides genes which permitted heme iron utilization in a laboratory strain of Escherichia coli were isolated. In the present study, the cloned P. shigelloides sequences were found to encode ten potential heme utilization proteins: HugA, the putative heme receptor; TonB and ExbBD; HugB, the putative periplasmic binding protein; HugCD, the putative inner membrane permease; and the proteins HugW, HugX, and HugZ. Three of the genes, hugA, hugZ, and tonB, contain a Fur box in their putative promoters, indicating that the genes may be iron regulated. When the P. shigelloides genes were tested in E. coli K-12 or in a heme iron utilization mutant of P. shigelloides, hugA, the TonB system genes, and hugW, hugX, or hugZ were required for heme iron utilization. When the genes were tested in a hemA entB mutant of E. coli, hugWXZ were not required for utilization of heme as a porphyrin source, but their absence resulted in heme toxicity when the strains were grown in media containing heme as an iron source. hugA could replace the Vibrio cholerae hutA in a heme iron utilization assay, and V. cholerae hutA could complement a P. shigelloides heme utilization mutant, suggesting that HugA is the heme receptor. Our analyses of the TonB system of P. shigelloides indicated that it could function in tonB mutants of both E. coli and V. cholerae and that it was similar to the V. cholerae TonB1 system in the amino acid sequence of the proteins and in the ability of the system to function in high-salt medium.

Effects of medium composition, calcium, iron and oxygen on haemolysin production by Plesiomonas shigelloides isolated from water.

AIMS: The effects of medium composition, calcium, iron and oxygen tension on the haemolytic activity of Plesiomonas shigelloides were investigated. METHODS AND RESULTS: The haemolytic activity of seven strains of Ple. shigelloides was tested on the surface of Luria Agar (LA), Brain Heart Infusion Agar (BHIA) and Trypitic Soy Agar (TSA) containing 5% (v/v) sheep blood, and in the Agar Overlay (AO) assay. All strains produced beta-haemolysis in the AO assay in three media, and on the surface of LA. The kinetics of growth and haemolytic activity of Ple. shigelloides 9P3-1 were evaluated in six different media, and the highest production of haemolysin occurred in Luria Broth (LB). The haemolytic activity of 9P3-1 was stimulated by Ca2+ and inhibited by EDTA. Addition of iron to the culture medium did not affect bacterial growth, although it reduced bacterial haemolytic activity. In the presence of an iron chelator, growth of the 9P3-1 was inhibited, but its haemolytic activity was enhanced. CONCLUSION: The haemolytic activity of Ple. shigelloides depends on medium composition, and that it is regulated by iron and is calcium-dependent. SIGNIFICANCE AND IMPACT OF THE STUDY: These results show the importance of optimization of media composition and oxygen tension for detection of Ple. shigelloides haemolytic activity.

31P-NMR and 13C-NMR studies of mannose metabolism in Plesiomonas shigelloides. Toxic effect of mannose on growth.

The metabolism of mannose was examined in resting cells in vivo using 13C-NMR and 31P-NMR spectroscopy, in cell-free extracts in vitro using 31P-NMR spectroscopy, and by enzyme assays. Plesiomonas shigelloides was shown to transport mannose by a phosphoenolpyruvate-dependent phosphotransferase system producing mannose 6-phosphate. However, a toxic effect was observed when P. shigelloides was grown in the presence of mannose. Investigation of mannose metabolism using in vivo 13C NMR showed mannose 6-phosphate accumulation without further metabolism. In contrast, glucose was quickly metabolized under the same conditions to lactate, ethanol, acetate and succinate. Extracts of P. shigelloides exhibited no mannose-6-phosphate isomerase activity whereas the key enzyme of the Embden-Meyerhof pathway (6-phosphofructokinase) was found. This result explains the mannose 6-phosphate accumulation observed in cells grown on mannose. The levels of phosphoenolpyruvate and Pi were estimated by in vivo 31P-NMR spectroscopy. The intracellular concentrations of phosphoenolpyruvate and Pi were relatively constant in both starved cells and mannose-metabolizing cells. In glucose-metabolizing cells, the phosphoenolpyruvate concentration was lower, and about 80% of the Pi was used during the first 10 min. It thus appears that the toxic effect of mannose on growth is not due to energy depletion but probably to a toxic effect of mannose 6-phosphate.

Hemolytic and elastolytic activities influenced by iron in Plesiomonas shigelloides.

The aim of this study was to determine the presence of hemolytic and elastolytic enzymes in several strains of Plesiomonas shigelloides in relation to the availability of iron in culture media. Hemolytic activity and elastolytic activity were detected in strains of P. shigelloides and were enhanced when the strains were grown in an iron-depleted medium and lost after thermal treatment at 100 degrees C for 10 min. Also, elastolytic activity was inactivated by phenylmethyl sulfonyl fluoride, an inhibitor of serine proteases. Hemolytic activity was detected extracellularly in cell-free supernatants, whereas elastin degradation activity was cell associated. Both activities may be related to the virulence of P. shigelloides.

Occurrence of Plesiomonas shigelloides in surface water: relationship with faecal pollution and trophic state.

Plesiomonas shigelloides is an autochthonous inhabitant of freshwater and has been associated with several waterborne outbreaks of acute gastro-enteritis. One of these outbreaks occurred in the summer of 1990, at a recreational beach near Amsterdam. In the summer of 1991, a survey was conducted to examine the presence of P. shigelloides in recreational fresh waters and marine water in the Netherlands. During the survey became clear that the specificity of current methods for the isolation and enumeration of P. shigelloides in water is low and requires extensive confirmation to obtain accurate data. P. shigelloides was ubiquitously present (30 of 42 samples positive) in fresh waters, but absent in sea water. Geometric mean Plesiomonas densities ranged from 140-340 per 100 ml; the range for individual positive samples was 33-840 per 100 ml. The Plesiomonas density was significantly correlated with index parameters for the trophic state (Secchi depth and chlorophyll A) and for faecal pollution (Escherichia coli). In winter, only 1 of 8 sites harboured P. shigelloides and in low numbers (4.4 per 100 ml). The ubiquitous presence of P. shigelloides in freshwater in the recreational season may result in cases of gastro-enteritis among bathers.

Optimal growth temperature for the isolation of Plesiomonas shigelloides, using various selective and differential agars.

The growth characteristics of known strains of Plesiomonas shigelloides were compared with those of Aeromonas species (the major competing species in environmental waters) on plesiomonas differential agar, inositol brilliant green bile salt, and modified salmonella-shigella agar at incubation temperatures of 37, 42, and 44 degrees C. Using local isolates from clinical and environmental sources, optimal growth conditions, as determined by colony counts and the colony characteristics, plesiomonas differential agar proved to be ideal when incubated at 44 degrees C. Contrary to earlier recommendations for 48 h incubation, the colonies could be recognized readily after an incubation of 24 h.

Distribution of Plesiomonas shigelloides in various components of pond ecosystems in Dhaka, Bangladesh.

Plesiomonas shigelloides is considered to be a waterborne agent of human gastroenteritis. An ecological study was carried out in five ponds in Dhaka city over a period of one year to elucidate the distribution and seasonality of this organism in various components of pond ecosystems. Samples were collected from hydrophytes, water, phytoplankton and sediment every 15 days over 12 months and cultured for P. shigelloides. P. shigelloides was isolated from a total of 120 samples including 25 (20.8%), 16 (13.3%), 22 (18.3%) and 35 (29.2%) of hydrophytes, water, phytoplankton and sediment samples, respectively. Distinct seasonal patterns of isolation of P. shigelloides were observed in the four components with two distinct peaks. The highest peaks were observed in hydrophytes and water samples in May and in phytoplankton and sediment in November. P. shigelloides was isolated from all components from all ponds during the study period. These results suggest that P. shigelloides is an autochthonous member in the freshwater pond ecosystems in Dhaka, Bangladesh.

[Mixed culture of Enterococcus faecium SF 68 with pathogenic intestinal bacterial strains]. / Colture miste diEenterococcus faecium SF 68 con ceppi batterici patogeni intestinali.

An antagonistic activity of Enterococcus faecium SF 68 towards Plesiomonas shigelloides, Aeromonas sp., enteropathogenic Escherichia coli and Yersinia enterocolitica has been studied and demonstrated.