Shigelloides Strains Research Articles

Horizontal Gene Transfer Clarifies Taxonomic Confusion and Promotes the Genetic Diversity and Pathogenicity of Plesiomonas shigelloides.

Plesiomonas shigelloides is an emerging pathogen that has been shown to be involved in gastrointestinal diseases and extraintestinal infections in humans. However, the taxonomic position, evolutionary dynamics, and pathogenesis of P. shigelloides remain unclear. We reported the draft genome sequences of 12 P. shigelloides strains representing different serogroups. We were able to determine a clear distinction between P. shigelloides and other members of Enterobacterales via core genome phylogeny, Neighbor-Net network, and average genome identity analysis. The pan-genome analysis of P. shigelloides revealed extensive genetic diversity and presented large flexible gene repertoires, while the core genome phylogeny exhibited a low level of clonality. The discordance between the core genome phylogeny and the pan-genome phylogeny indicated that flexible accessory genomes account for an important proportion of the evolution of P. shigelloides, which was subsequently characterized by determinations of hundreds of horizontally transferred genes (horizontal genes), massive gene expansions and contractions, and diverse mobile genetic elements (MGEs). The apparently high levels of horizontal gene transfer (HGT) in P. shigelloides were conferred from bacteria with novel properties from other taxa (mainly Vibrionaceae and Aeromonadaceae), which caused the historical taxonomic confusion and shaped the virulence gene pools. Furthermore, P. shigelloides genomes contain many macromolecular secretion system genes, virulence factor genes, and resistance genes, indicating its potential to cause intestinal and invasive infections. Collectively, our work provides insights into the phylogenetic position, evolutionary dynamic, and pathogenesis of P. shigelloides at the genomic level, which could facilitate the observation and research of this important pathogen.IMPORTANCE The taxonomic position of P. shigelloides has been the subject of debate for a long time, and until now, the evolutionary dynamics and pathogenesis of P. shigelloides were unclear. In this study, pan-genome analysis indicated extensive genetic diversity and the presence of large and variable gene repertoires. Our results revealed that horizontal gene transfer was the focal driving force for the genetic diversity of the P. shigelloides pan-genome and might have contributed to the emergence of novel properties. Vibrionaceae and Aeromonadaceae were found to be the predominant donor taxa for horizontal genes, which might have caused the taxonomic confusion historically. Comparative genomic analysis revealed the potential of P. shigelloides to cause intestinal and invasive diseases. Our results could advance the understanding of the evolution and pathogenesis of P. shigelloides, particularly in elucidating the role of horizontal gene transfer and investigating virulence-related elements.

Putative virulence factors of Plesiomonas shigelloides.

Plesiomonas shigelloides is a Gram-negative rod-shaped bacterium which has been isolated from humans, animals and the environment. It has been associated with diarrhoeal disease in humans and various epizootic diseases in animals. In this study P. shigelloides strains were isolated from the faecal material of a captive Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis; YFP) living in semi-natural conditions in China. Plesiomonas shigelloides strain EE2 was subjected to whole genome sequencing. The draft genome was then compared to the genome sequences of ten other P. shigelloides isolates using the Pathosystems Resource Integration Center pipeline. In addition to several virulence factors which have been previously reported, we are proposing new candidate virulence factors such as a repeats-in-toxin protein, lysophospholipase, a twin-arginine translocation system and the type VI secretion effector Phospholipase A1.

Plesiomonas shigelloides sipD mutant, generated by an efficient gene transfer system, is less invasive

Plesiomonas shigelloides is widely associated with human diarrheal disease. Research on this pathogen has been hampered by the absence of an effective genetic manipulation system. In the present study, an efficient and precise conjugation transfer procedure, mediated by suicide vector pRE112 was used to overcome this limitation. The efficiency of generating double recombinants was average 74.3%, and the conjugation protocol may be applied to other P. shigelloides strains. We also identified that the SipD protein of P. shigelloides G5884 (serotype O45) is 65% similar to the SipD in Salmonella pathogenicity island 1 (SPI-1), which is a key element of the type III secretion system related to Salmonella invasion. A P. shigelloides sipD null mutant was generated via the conjugation system, using the suicide vector pRE112. The isogenic mutant strain lacking sipD showed a 50% reduction in its capacity to invade Caco-2 cells.

Pathogenomics of Virulence Traits of Plesiomonas shigelloides That Were Deemed Inconclusive by Traditional Experimental Approaches.

One of the major challenges of modern medicine includes the failure of conventional protocols to characterize the pathogenicity of emerging pathogens. This is particularly apparent in the case of Plesiomonas shigelloides. Although a number of infections have been linked to this microorganism, experimental evidence of its virulence factors (VFs), obtained by traditional approaches, is somewhat inconclusive. Hence, it remains unclear whether P. shigelloides is a true or opportunistic one. In the current study, four publicly available whole-genome sequences of P. shigelloides (GN7, NCTC10360, 302-73, and LS1) were profiled using bioinformatics platforms to determine the putative candidate VFs to characterize the bacterial pathogenicity. Overall, 134 unique open reading frames (ORFs) were identified that were homologous or orthologous to virulence genes identified in other pathogens. Of these, 52.24% (70/134) were jointly shared by the strains. The numbers of strain-specific virulence traits were 4 in LS1; 7 in NCTC10360; 10 in 302-73; and 15 in GN7. The pathogenicity islands (PAIs) common to all the strains accounted for 24.07% ORFs. The numbers of PAIs exclusive to each strain were 8 in 302-73; 11 in NCTC10360; 14 in GN7; and 18 in LS1. A PAI encoding Vibrio cholerae ToxR-activated gene d protein was specific to 302-73, GN7, and NCTC10360 strains. Out of 33 antibiotic multi-resistance genes identified, 16 (48.48%) genes were intrinsic to all strains. Further, 17 (22.08%) of 77 antibiotic resistance islands were found in all the strains. Out of 23 identified distinct insertion sequences, 13 were only harbored by strain LS1. The number of intact prophages identified in the strains was 1 in GN7; 2 in 302-73; and 2 in NCTC10360. Further, 1 CRISPR element was identified in LS1; 2 in NCTC10360; and 8 in 302-73. Fifteen (78.95%) of 19 secretion systems and secretion effector variants were identified in all the strains. In conclusion, certain P. shigelloides strains might possess VFs associated with gastroenteritis and extraintestinal infections. However, the role of host factors in the onset of infections should not be undermined.

Plesiomonas shigelloides: A notable carrier of acquired antimicrobial resistance in small aquaculture farms

Small and low-cost family aquaculture tilapia (Oreochromis niloticus) production with regional commercial targets is currently common practice in continental production systems of many parts of the world. In the state of Rio de Janeiro, Brazil, fish are reared in natural ponds, with limited technology, using available water resources. Nutrition relies on pelleted feed, and production costs are minimized by adding untreated manure and other organic fertilizers directly into the water. The aim of this study was to describe antimicrobial resistance in gram-negative bacteria isolated from fish produced under these conditions in four watersheds of that state. Gram-negative aerobic bacteria isolated from fish bowel and gill samples were identified, and resistance phenotypes determined. Fecal and sediment samples were analysed for residual evidence of tetracycline, the most commonly used therapeutic drug in animal husbandry, and for the presence of tetracycline resistance genetic determinants. Gram-negative bacteria belonging to different genera were the majority of culturable bacteria obtained from these samples. Results revealed a high number of tetracycline, and to a lesser extent, chloramphenicol resistant P. shigelloides strains, by far the predominant isolated bacterial species. Tetracycline residues were detected in both, sediment and fish fecal samples, from all farms. Tetracycline resistance genetic determinants tet(A), tet(C), tet(D), tet(M) and tet(O) were detected in sediment and fecal samples, and tet(A), tet(D) and tet(B) were detected in P. shigelloides isolates. Chloramphenicol resistant P. shigelloides isolates carried the floR, catI and catII genetic determinants. These data suggest the putative role of P. shigelloides as a reservoir for antimicrobial resistance genes in traditional continental aquaculture reflecting the selective pressure imposed by a combination of varied and uncontrolled rearing practices.

Aeromonas and Plesiomonas species from scarlet ibis (Eudocimus ruber) and their environment: monitoring antimicrobial susceptibility and virulence.

The present study aimed at evaluating the role of captive scarlet ibises (Eudocimus ruber) and their environment as reservoirs of Aeromonas spp. and Plesiomonas spp., and analyzing the in vitro antimicrobial susceptibility and virulence of the recovered bacterial isolates. Thus, non-lactose and weak-lactose fermenting, oxidase positive Gram-negative bacilli were recovered from cloacal samples (n=30) of scarlet ibises kept in a conservational facility and from water samples (n=30) from their environment. Then, the antimicrobial susceptibility, hemolytic activity and biofilm production of the recovered Aeromonas spp. and Plesiomonas shigelloides strains were assessed. In addition, the virulence-associated genes of Aeromonas spp. were detected. Ten Aeromonas veronii bv. sobria, 2 Aeromonas hydrophila complex and 10 P. shigelloides were recovered. Intermediate susceptibility to piperacillin-tazobactam and cefepime was observed in 2 Aeromonas spp. and 1 P. shigelloides, respectively, and resistance to gentamicin was observed in 4 P. shigelloides. The automated susceptibility analysis revealed resistance to piperacillin-tazobactam and meropenem among Aeromonas spp. and intermediate susceptibility to gentamicin among P. shigelloides. All Aeromonas isolates presented hemolytic activity, while 3 P. shigelloides were non-hemolytic. All Aeromonas spp. and 3/10 P. shigelloides were biofilm-producers, at 28°C, while 10 Aeromonas spp. and 6/10 P. shigelloides produced biofilms, at 37°C. The most prevalent virulence genes of Aeromonas spp. were asa1 and ascV. Scarlet ibises and their environment harbour potentially pathogenic bacteria, thus requiring monitoring and measures to prevent contamination of humans and other animals.

Rapid and sensitive detection of Plesiomonas shigelloides by cross‑priming amplification of the hugA gene.

Plesiomonas shigelloides (P. shigelloides) is implicated as an aetiological agent of human gastroenteritis in humans, for which reliable laboratory detection of P. shigelloides is clinically and epidemiologically desirable. A simple molecular method for rapid detection of P. shigelloides using cross-priming amplification (CPA) has been developed, with hugA as the target. The hugA gene is required for haem iron utilisation and is critical for the survival and growth of P. shigelloides. The assay output was visualised as a colour change with no need to open the reaction tubes, and no false-positive results were detected for the 33 non- P. shigelloides strains examined to assess assay specificity. The limit of detection was 200 fg P. shigelloides DNA per reaction and 3×103 CFU per g in human stools, which was 100 and 10-fold more sensitive than polymerase chain reaction, respectively. The CPA method was used to detect the presence of P. shigelloides in stool specimens from 70 patients with diarrhoea and 30 environmental water samples, with no difference in accuracy between the CPA assay and the biological culture. The present study, therefore, suggests that the P. shigelloides hugA CPA assay may represent a valuable tool for rapid and sensitive detection of P. shigelloides in primary care facilities and clinical laboratories.

The polar and lateral flagella from Plesiomonas shigelloides are glycosylated with legionaminic acid.

Plesiomonas shigelloides is the unique member of the Enterobacteriaceae family able to produce polar flagella when grow in liquid medium and lateral flagella when grown in solid or semisolid media. In this study on P. shigelloides 302-73 strain, we found two different gene clusters, one exclusively for the lateral flagella biosynthesis and the other one containing the biosynthetic polar flagella genes with additional putative glycosylation genes. P. shigelloides is the first Enterobacteriaceae were a complete lateral flagella cluster leading to a lateral flagella production is described. We also show that both flagella in P. shigelloides 302-73 strain are glycosylated by a derivative of legionaminic acid (Leg), which explains the presence of Leg pathway genes between the two polar flagella regions in their biosynthetic gene cluster. It is the first bacterium reported with O-glycosylated Leg in both polar and lateral flagella. The flagella O-glycosylation is essential for bacterial flagella formation, either polar or lateral, because gene mutants on the biosynthesis of Leg are non-flagellated. Furthermore, the presence of the lateral flagella cluster and Leg O-flagella glycosylation genes are widely spread features among the P. shigelloides strains tested.

Core Oligosaccharide of Plesiomonas shigelloides PCM 2231 (Serotype O17) Lipopolysaccharide — Structural and Serological Analysis

The herein presented complete structure of the core oligosaccharide of lipopolysaccharide (LPS) P. shigelloides Polish Collection of Microorganisms (PCM) 2231 (serotype O17) was investigated by 1H, 13C NMR spectroscopy, mass spectrometry, chemical analyses and serological methods. The core oligosaccharide is composed of an undecasaccharide, which represents the second core type identified for P. shigelloides serotype O17 LPS. This structure is similar to that of the core oligosaccharide of P. shigelloides strains 302-73 (serotype O1) and 7-63 (serotype O17) and differs from these only by one sugar residue. Serological screening of 55 strains of P. shigelloides with the use of serum against identified core oligosaccharide conjugated with bovine serum albumin (BSA) indicated the presence of similar structures in the LPS core region of 28 O-serotypes. This observation suggests that the core oligosaccharide structure present in strain PCM 2231 could be the most common type among P. shigelloides lipopolysaccharides.

Rapid and Sensitive Detection of Plesiomonas shigelloides by Loop-Mediated Isothermal Amplification of the hugA Gene

Plesiomonas shigelloides is one of the causative agents of human gastroenteritis, with increasing number of reports describing such infections in recent years. In this study, the hugA gene was chosen as the target to design loop-mediated isothermal amplification (LAMP) assays for the rapid, specific, and sensitive detection of P. shigelloides. The performance of the assay with reference plasmids and spiked human stools as samples was evaluated and compared with those of quantitative PCR (qPCR). No false-positive results were observed for the 32 non-P. shigelloides strains used to evaluate assay specificity. The limit of detection for P. shigelloides was approximately 20 copies per reaction in reference plasmids and 5×103 CFU per gram in spiked human stool, which were more sensitive than the results of qPCR. When applied in human stool samples spiked with 2 low levels of P. shigelloides, the LAMP assays achieved accurate detection after 6-h enrichment. In conclusion, the LAMP assay developed in this study is a valuable method for rapid, cost-effective, and simple detection of P. shigelloides in basic clinical and field laboratories in the rural areas of China.

Molecular evidence of Plesiomonas shigelloides as a possible zoonotic agent

The most frequently used method for establishing epidemiological relationships between Plesiomonas shigelloides strains is O:H serotyping. However, a number of strains are not serotypeable and isolates from diverse sources can display the same serovar. Moreover, since the zoonotic nature of Plesiomonas has been suggested and this hypothesis is based on the identical serovars found in animals and humans, we intend to use four DNA-based techniques: random amplified polymorphic DNA-PCR, enterobacterial repetitive intergenic consensus-PCR, repetitive extragenic palindromic-PCR, and pulsed field gel electrophoresis in order to screen 24 strains belonging to nine O:H serovars isolated from humans, animals, and the environment. In general, P. shigelloides showed a high genetic heterogeneity. Three pairs of strains, each containing a human and an animal isolate, displayed similar genotypes. This is the first report that provides molecular evidence that P. shigelloides may be zoonotic.

O41 Percutaneous laser disc decompression (PLDD) with 970 nm wavelength laser: a minimal invasive surgery for PLID, first time in Bangladesh

Ten new Czechoslovak O and four H antigens of Plesiomonas shigelloides are described and compared with fifteen new Japanese O and ten H serovars. The Czechoslovak O serovars are as follows: “O35”-11440, “O36”- 13101, “O37”- 24827, “O38”- 27159, “O39”- 27492, “O40”- 27919, “O41 -28384, “O42”- 28720, “O43”- 29021 and “O44”- 29470, H antigens t, u, v and w.Two of the new Czechoslovak O antigens were identical with the Japanese ones: “O36”- 13101 with O64 and “O39”- 27492 with O66, but the latter Czechoslovak O serovar possessed a different H antigen t. The new Japanese serovar O61 was identical with the previously reported Czechoslovak serovar O4. No relationship was found between the four new Czechoslovak H antigens and the ten new Japanese ones.Three of the Czechoslovak serovars occurred in strains of previously unknown antigenic structure: “O35”- 11440, “O37”- 24827 and “O39”- 27492. Two recently described Japanese serovars O56 : H18 and O62 but with a distinct Czechoslovak H antigen d, and three H antigens H9, H22 and H28, were found in the Czechoslovak Plesiomonas shigelloides strains.

O61 Assumptions about the integrating role of the hypothalamus in processes of light control and light fluctuations filtering, with application in endocrine and neurologic conditions

Ten new Czechoslovak O and four H antigens of Plesiomonas shigelloides are described and compared with fifteen new Japanese O and ten H serovars. The Czechoslovak O serovars are as follows: “O35”-11440, “O36”- 13101, “O37”- 24827, “O38”- 27159, “O39”- 27492, “O40”- 27919, “O41 -28384, “O42”- 28720, “O43”- 29021 and “O44”- 29470, H antigens t, u, v and w.Two of the new Czechoslovak O antigens were identical with the Japanese ones: “O36”- 13101 with O64 and “O39”- 27492 with O66, but the latter Czechoslovak O serovar possessed a different H antigen t. The new Japanese serovar O61 was identical with the previously reported Czechoslovak serovar O4. No relationship was found between the four new Czechoslovak H antigens and the ten new Japanese ones.Three of the Czechoslovak serovars occurred in strains of previously unknown antigenic structure: “O35”- 11440, “O37”- 24827 and “O39”- 27492. Two recently described Japanese serovars O56 : H18 and O62 but with a distinct Czechoslovak H antigen d, and three H antigens H9, H22 and H28, were found in the Czechoslovak Plesiomonas shigelloides strains.

Virulence factors and in vitro susceptibility of Plesiomonas shigelloides isolated from diarrhea episodes in Cuba

Acute diarrhea is still one of the main causes of disease in developing countries. At the National Reference Laboratory for Acute Diarrhoeal Diseases, of the "Pedro Kourí" Tropical Medicine Institute, 54 Plesiomonas shigelloides strains were serotyped. As a result, the circulation of four new serotypes (O20:H2, 056:H18, 071 :H31,081 :H22) was detected. The most common antigenic variant was 094:H3. In addition, the susceptibility pattern to 21 antimicrobial agents (AA) was studied. Highest percentages of resistance were observed to ampicillin (77.7%), tetracycline (29.2%), and erythromycin (18.5%) while, to all other AA the resistance percentages were under 33%. There was no evidence of association between the serotypes and the multi-resistance patterns present in the strains under study. The frequency of virulence factors was determined: 21.15% of the strains turned out to be ss-haemolytic; 73.7% showed the presence of biofilm and 92.31% were hydrophobic. In 17.3% of the strains, the three virulence factors analysed were present. When establishing the relationship among the different virulence factors and the serotypes of the strains through cross reaction with the Shigella genus, there was evidence that 93.3% of the strains showed at least one of the virulence factors studied. Likewise, at least one of the virulence factors analysed was present in 90.9% of the multi-resistant strains.

Caracterización de factores de virulencia y susceptibilidad antimicrobiana en cepas de Plesiomonas shigelloides aisladas de pacientes con diarrea aguda en Cuba

Acute diarrhea is still one of the main causes of disease in developing countries. At the National Reference Laboratory for Acute Diarrhoeal Diseases, of the "Pedro Kourí" Tropical Medicine Institute, 54 Plesiomonas shigelloides strains were serotyped. As a result, the circulation of four new serotypes (O20:H2, 056:H18, 071 :H31,081 :H22) was detected. The most common antigenic variant was 094:H3. In addition, the susceptibility pattern to 21 antimicrobial agents (AA) was studied. Highest percentages of resistance were observed to ampicillin (77.7%), tetracycline (29.2%), and erythromycin (18.5%) while, to all other AA the resistance percentages were under 33%. There was no evidence of association between the serotypes and the multi-resistance patterns present in the strains under study. The frequency of virulence factors was determined: 21.15% of the strains turned out to be ss-haemolytic; 73.7% showed the presence of biofilm and 92.31% were hydrophobic. In 17.3% of the strains, the three virulence factors analysed were present. When establishing the relationship among the different virulence factors and the serotypes of the strains through cross reaction with the Shigella genus, there was evidence that 93.3% of the strains showed at least one of the virulence factors studied. Likewise, at least one of the virulence factors analysed was present in 90.9% of the multi-resistant strains.

Structure of the Core Region from the Lipopolysaccharide of Plesiomonas shigelloides Strain 302‐73 (Serotype O1)

AbstractPlesiomonas shigelloides is a Gram‐negative pathogenic bacterium belonging to the Enterobacteriaceae family. To date, only few lipopolysaccharide (LPS) structures from P. shigelloides strains are known. In particular, three core oligosaccharides have been found. Recently, we elucidated the structure of the O‐antigen of P. shigelloides 302‐73 (serotype O1) and in this paper we present the characterization of the core structure from the LPS of the same strain. The LPS was hydrolyzed under both alkaline and mildly acidic conditions. In both cases, a mixture of oligosaccharides was obtained, which was purified by gel filtration and HPAEC. The oligosaccharides were characterized by chemical analysis, 2D NMR spectroscopy and MALDI‐TOF mass spectrometry. A new core structure was found for P. shigelloides. In particular, from the analysis of the acid hydrolysed product it was possible to reveal the presence of a of D‐glycero‐D‐talo‐2‐octulopyranosonic acid (Ko) residue, which substitutes in part the terminal 3‐deoxy‐D‐manno‐oct‐2‐ulosonic acid (Kdo) unit. The Ko residue is not frequently found in core structures.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2009)

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°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.

Potential virulence-associated properties of Plesiomonas shigelloides strains.

Serotyping and some potential virulence-associated markers were investigated in Plesiomonas shigelloides strains isolated from humans, animals and aquatic environments. Surface properties of these strains were evaluated using Congo red binding, salt-aggregation test, bacterial adherence to xylene and motility. Production of pancreatic elastase, proteinase (consistent with subtilisin Carlsberg), triacylglycerol lipase, histidine decarboxylase and beta-hemolysin was also determined. In addition, detection of signal molecules such as C4-C8 unsubstituted N-acylhomoserine lactones (AHLs) was performed. The serological typing of the P. shigelloides strains showed that the isolates belonged to 13 different serovars. The majority of the strains were hydrophobic and motile. The strains produced low levels of elastase, proteinase and histidine decarboxylase whereas triacylglycerol lipase activity was relatively high. Only 23.3 % of the strains produced hemolysin. The AHLs signal molecules were not detected. P. shigelloides strains were able to produce a variety of potential virulence markers which may be involved in the pathogenesis of Plesiomonas-associated infections.

Effect of inoculum density on susceptibility of Plesiomonas shigelloides to cephalosporins.

Resistance of Plesiomonas shigelloides to cephalosporins at higher cell densities has been reported. We investigated whether these inoculum effects are due to the production of beta-lactamases. beta-Lactamase production of five P. shigelloides strains was characterized by activity tests, SDS-PAGE and isoelectric focusing. For all strains, MIC values of different cephalosporins were determined by microdilution methodology using inocula of 1 x 10(5) cfu/mL and 1 x 10(6) cfu/mL. Subsequently, the morphology of cells was determined by light microscopy. For one isolate, kill kinetics of cefpodoxime were determined using batch cultures with the lower and higher inocula. Four of five P. shigelloides strains were shown to be beta-lactamase-positive, producing different amounts of constitutively expressed non-inducible enzymes. Inoculum effects for cephalosporin susceptibility were observed for all strains. Examination of cells revealed a very strong filamentation, with filament sizes ranging from 100 microm up to 2 mm. The kill kinetics with cefpodoxime showed similar killing capacities of the antibiotic at both inoculum sizes. The reported resistance of P. shigelloides to cephalosporins at higher cell densities is not due to an inoculum-dependent regulation of beta-lactamases, but can be explained by the formation of extensive filaments.

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.