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Aeromonas species are enteropathogens that cause gastroenteritis with a unique three-peak infection pattern related to patient age. The contributions of individual Aeromonas species to age-related infections remain unknown. Multi-locus sequence typing (MLST) was performed to determine the species of Aeromonas strains from Australian patients with gastroenteritis. Public database searches were conducted to collect strains of enteric Aeromonas species, identified by either MLST or whole genome sequencing with known patient age. Violin plot analysis was performed to assess Aeromonas infection distribution across patients of different ages. Generalized additive model (GAM) analysis was employed to investigate the relationship between Aeromonas species and patient age. A total of 266 strains of seven Aeromonas species met the selection criteria, which were used for analyses. The violin plots revealed distinct patterns among individual Aeromonas species in relation to patient age. The GAM analyses identified a significant association between Aeromonas species and patient age (p = 0.009). Aeromonas veronii (153 strains) showed the highest probability of infection in most ages, particularly among young adults. Aeromonas caviae (59 strains) is more common in young children and adults over 60 years of age. The probability of infection for Aeromonas hydrophila (34 strains) and Aeromonas dhakensis (9 strains) was generally low, there was a slight increase in individuals aged 50–60 for A. hydrophila and over 60 years for A. dhakensis. These findings provide novel evidence of the varied contributions of different Aeromonas species to human enteric infections related to patient age, offering valuable insights for epidemiology and clinical management. [ABSTRACT FROM AUTHOR]

Background: Aeromonas dhakensis is associated with soft tissue infection, bacteremia and gastroenteritis. Involvement of respiratory system in adults is extremely rare. We report a case of fulminant pneumonia and bacteremia due to A. dhakensis in a patient without underlying diseases. Case presentation: A 26-year-old man became ill suddenly with pneumonia after swimming in a river. Despite intensive support measures in the intensive care unit, he died 13 hours after admission and 4 days after his first symptoms. Autopsy showed abundant Gram-negative bacteria, massive inflammatory cell infiltration, edema, necrosis and hemorrhage in lung tissue. A. dhakensis was isolated from blood culture taken at admission and bronchoalveolar lavage fluid (BALF) after intubation. Moreover, A. dhakensis was also detected in lung tissue by metagenomic next-generation sequencing (mNGS) assay. The infection may have come from river water. Conclusion: In patients who develop a fulminant pneumonia after contacting an aquatic environment, A. dhakensis should be alerted and mNGS may aid in the detection of aquatic pathogens by being more sensitive and specific versus traditional bacterial culture. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis is reported as an emerging pathogenic species within the genus Aeromonas and is widely distributed in tropical coastal areas. This study provided a detailed description and characterization of a strain of A. dhakensis (202108B1) isolated from diseased Ancherythroculter nigrocauda in an inland region of China. Biochemical tests identified the isolate at the genus level, and the further molecular analysis of concatenated housekeeping gene sequences revealed that the strain belonged to the species A. dhakensis. The isolated A. dhakensis strain was resistant to five antibiotics, namely, penicillin, ampicillin, clindamycin, cephalexin, and imipenem, while it was susceptible to or showed intermediate resistance to most of the other 15 tested antibiotics. The isolated strain of A. dhakensis caused acute hemorrhagic septicemia and tissue damage in artificially infected A. nigrocauda , with a median lethal dose of 7.76 × 104 CFU/fish. The genome size of strain 202108B1 was 5 043 286 bp, including 1 chromosome and 4 plasmids. This is the first detailed report of the occurrence of infection caused by an A. dhakensis strain causing infection in an aquaculture system in inland China, providing important epidemiological data on this potential pathogenic species. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis is an increasingly recognized human pathogen in recent years and was first isolated and reported in a sample of childhood diarrhea in Bangladesh. More and more cases of Aeromonas dhakensis infection have been reported in recent years. Here we report a case of severe pneumonia caused by Aeromonas dhakensis with severe atrial septal defect. The patient, a 56-year-old male, was admitted to the hospital with severe hypoxemia and severe septic shock. Detection of the patient’s bronchoalveolar lavage fluid (BALF) and peripheral blood by the metagenomic next generation sequencing (mNGS) indicated Aeromonas dhakensis infection.

Aeromonas species are enteropathogens that cause gastroenteritis with a unique three-peak infection pattern related to patient age. The contributions of individual Aeromonas species to age-related infections remain unknown. Multi-locus sequence typing (MLST) was performed to determine the species of Aeromonas strains from Australian patients with gastroenteritis. Public database searches were conducted to collect strains of enteric Aeromonas species, identified by either MLST or whole genome sequencing with known patient age. Violin plot analysis was performed to assess Aeromonas infection distribution across patients of different ages. Generalized additive model (GAM) analysis was employed to investigate the relationship between Aeromonas species and patient age. A total of 266 strains of seven Aeromonas species met the selection criteria, which were used for analyses. The violin plots revealed distinct patterns among individual Aeromonas species in relation to patient age. The GAM analyses identified a significant association between Aeromonas species and patient age (p = 0.009). Aeromonas veronii (153 strains) showed the highest probability of infection in most ages, particularly among young adults. Aeromonas caviae (59 strains) is more common in young children and adults over 60 years of age. The probability of infection for Aeromonas hydrophila (34 strains) and Aeromonas dhakensis (9 strains) was generally low, there was a slight increase in individuals aged 50–60 for A. hydrophila and over 60 years for A. dhakensis. These findings provide novel evidence of the varied contributions of different Aeromonas species to human enteric infections related to patient age, offering valuable insights for epidemiology and clinical management.

Abstract Aeromonas dhakensis (A. dhakensis) is becoming an emerging pathogen worldwide, with an increasingly significant role in animals and human health. It is a ubiquitous bacteria found in terrestrial and aquatic milieus. However, there have been few reports of reptile infections. In this study, a bacterial strain isolated from a dead Aldabra giant tortoise was identified as A. dhakensis HN-1 through clinical observation, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF/MS), and gene sequencing analysis. Subsequently, to evaluate its pathogenicity, the detection of virulence genes and mice infection experiments were performed. A. dhakensis HN-1 was found to contain seven virulence genes, including alt, ela, lip, act, aerA, fla, and hlyA. Mice infected with A. dhakensis HN-1 exhibited hemorrhage of varying degrees in multiple organs. The half-maximal lethal dose (LD50) value of A. dhakensis HN-1 for mice was estimated to be 2.05 × 107 colony forming units (CFU)/mL. The antimicrobial susceptibility test revealed that A. dhakensis HN-1 was resistant to amoxicillin, penicillin, ampicillin and erythromycin. This is the first report of A. dhakensis in Aldabra giant tortoises, expanding the currently known host spectrum. Our findings emphasize the need for One Health surveillance and extensive research to reduce the spread of A. dhakensis across the environment, humans, and animals.

Aeromonas dhakensis stands out as the most potent Aeromonas species causing a range of human diseases. This research marks the pioneering effort in isolating and characterizing virulent phages targeting A. dhakensis. Only the AM isolate among the Aeromonas isolates showed compatibility for phage isolation and was identified as A. dhakensis. Computational analysis identified the presence of virulence factors and antimicrobial resistance genes in A. dhakensis AM. Phage isolation was conducted using this particular strain as the host, resulting in the isolation of four virulent phages: vB_AdhM_DL, vB_AdhS_TS3, vB_AdhM_TS9, and vB_AdhS_M4. Bacterial numbers significantly decrease after both pre-treatment and post-treatment with individual phages and phage cocktails, ranging from 2.82 to 6.67 log CFU/mL and 4.01 to 6.49 log CFU/mL, respectively. Combining a phage cocktail with sub-MIC amoxicillin led to complete inactivation in both pre-treatment and post-treatment scenarios within a 200 µL volume. The complete genomes of phages vB_AdhM_DL, vB_AdhS_TS3, and vB_AdhM_TS9 were determined to be 42,388 bp, 115,560 bp, and 115,503 bp, respectively. This study establishes the effectiveness of using phages as an complement with sublethal antibiotic concentrations, presenting a potential and effective therapeutic approach.

BackgroundAeromonas dhakensis is a gram-negative bacterium. In recent years, Aeromonas dhakensis has gradually attracted increasing attention due to its strong virulence and poor prognosis. Clinical reports of pulmonary infection caused by Aeromonas dhakensis are rare.Case presentationA patient with acute T lymphoblastic leukemia experienced myelosuppression after chemotherapy, developed a secondary pulmonary infection with Aeromonas dhakensis and was hospitalized due to fever. The patient underwent testing for inflammatory markers, chest imaging, blood culture, bronchoalveolar lavage, pleural drainage, and metagenomic next-generation sequencing of alveolar lavage fluid and pleural fluid to obtain evidence of Aeromonas dhakensis infection, and was treated with four generations of cephalosporin combined with fluoroquinolone antibiotics. The patient’s condition significantly improved.DiscussionAmong pulmonary infectious pathogens, Aeromonas dhakensis is relatively rare. Once an Aeromonas strain is cultured in the clinical work, pathogenic sequencing should be performed on the detected samples for early accurate diagnosis and effective anti-infection treatment.

Aeromonas dhakensis (A. dhakensis) is becoming an emerging pathogen worldwide, with an increasingly significant role in animals and human health. It is a ubiquitous bacteria found in terrestrial and aquatic milieus. However, there have been few reports of reptile infections. In this study, a bacterial strain isolated from a dead Aldabra giant tortoise was identified as A. dhakensis HN-1 through clinical observation, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF/MS), and gene sequencing analysis. Subsequently, to evaluate its pathogenicity, the detection of virulence genes and mice infection experiments were performed. A. dhakensis HN-1 was found to contain seven virulence genes, including alt, ela, lip, act, aerA, fla, and hlyA. Mice infected with A. dhakensis HN-1 exhibited hemorrhage of varying degrees in multiple organs. The half-maximal lethal dose (LD50) value of A. dhakensis HN-1 for mice was estimated to be 2.05 × 107 colony forming units (CFU)/mL. The antimicrobial susceptibility test revealed that A. dhakensis HN-1 was resistant to amoxicillin, penicillin, ampicillin and erythromycin. This is the first report of A. dhakensis in Aldabra giant tortoises, expanding the currently known host spectrum. Our findings emphasize the need for One Health surveillance and extensive research to reduce the spread of A. dhakensis across the environment, humans, and animals. [ABSTRACT FROM AUTHOR]

To identify the cause of mass mortality of adult Macrobrachium rosenbergii in a farm in Gaoyou City, Jiangsu Province, China, a dominant strain named DKQ-1 was isolated from the hepatopancreas of dying M. rosenbergii and identified as Aeromonas dhakensis by purification culture, biochemical characterization, and 16S rRNA and gyrB gene sequence analysis. The results of the challenge test revealed that the strain was highly pathogenic and the 50% lethal dose (LD50) in 72 h to M. rosenbergii was 1.54×105 CFU/mL. The amplification results of virulence genes show that strain DKQ-1 carried 9 virulence genes, including ascV, aexT, aer, act, lip, ompAI, gcaT, acg, and exu, supporting the strong virulence of strain DKQ-1 to M. rosenbergii. Histopathological observation of the hepatopancreas, gills, and intestines indicated that DKQ-1 injection into M. rosenbergii could cause serious tissue damage, which further supported the strong virulence of this strain. In addition, a drug susceptibility test revealed that strain DKQ-1 was sensitive to 16 kinds of antibiotics, resistant to 9 kinds of antibiotics, and had intermediate resistance to spectinomycin and kanamycin. This study is the first report of A. dhakensis isolated from M. rosenbergii and provided a reference for the pathogen identification of bacterial diseases in M. rosenbergii, and for the prevention and treatment caused by A. dhakensis. [ABSTRACT FROM AUTHOR]

Introduction: Motile Aeromonas septicemia (MAS) is a burden for striped catfish (Pangasius hypophthalmus) farmers in Vietnam. MAS can be caused by several species of Aeromonas but Aeromonas hydrophila is seen as the leading cause of MAS in aquaculture, but recent reports suggest that A. dhakensis is also causing MAS. Methods: Here we investigated the bacterial etiology of MAS and compared the genomic features of A. hydrophila and A. dhakensis. We collected 86 isolates from diseased striped catfish fingerlings over 5 years from eight provinces in Vietnam. Species identification was done using PCR, MALDI-TOF and whole genome sequence (WGS). The MICs of commonly used antimicrobials was established. Thirty presumed A. hydrophila isolates were sequenced for species confirmation and genomic comparison. A phylogenetic analysis was conducted using publicly available sequences and sequences from this study. Results: A total of 25/30 isolates were A. dhakensis sequence type (ST) 656 and 5/30 isolates were A. hydrophila ST 251. Our isolates and all publicly available A. hydrophila isolates from Vietnam belonged to ST 251 and differed with <200 single nucleotide polymorphisms (SNP). Similarly, all A. dhakensis isolates from Vietnam belonged to ST 656 and differed with <100 SNPs. The tet(A) gene was found in 1/5 A. hydrophila and 19/25 A. dhakensis. All A. hydrophila had an MIC ≤2 mg/L while 19/25 A. dhakensis had MIC ≥8 mg/L for oxytetracycline. The floR gene was only found in A. dhakensis (14/25) which showed a MIC ≥8 mg/L for florfenicol. Key virulence genes, i.e., aerA/act, ahh1 and hlyA were present in all genomes, while ast was only present in A. dhakensis. Discussion: This study confirms previous findings where A. dhakensis was the dominating pathogen causing MAS and that the importance of A. hydrophila has likely been overestimated. The differences in antimicrobial susceptibility between the two species could indicate a need for targeted antimicrobial treatment plans. The lipopolysaccharide regions and outer membrane proteins did not significantly differ in their immunogenic potentials, but it remains to be determined with in vivo experiments whether there is a difference in the efficacy of available vaccines against A. hydrophila and A. dhakensis. [ABSTRACT FROM AUTHOR]

IntroductionMotile Aeromonas septicemia (MAS) is a burden for striped catfish (Pangasius hypophthalmus) farmers in Vietnam. MAS can be caused by several species of Aeromonas but Aeromonas hydrophila is seen as the leading cause of MAS in aquaculture, but recent reports suggest that A. dhakensis is also causing MAS.MethodsHere we investigated the bacterial etiology of MAS and compared the genomic features of A. hydrophila and A. dhakensis. We collected 86 isolates from diseased striped catfish fingerlings over 5 years from eight provinces in Vietnam. Species identification was done using PCR, MALDI-TOF and whole genome sequence (WGS). The MICs of commonly used antimicrobials was established. Thirty presumed A. hydrophila isolates were sequenced for species confirmation and genomic comparison. A phylogenetic analysis was conducted using publicly available sequences and sequences from this study.ResultsA total of 25/30 isolates were A. dhakensis sequence type (ST) 656 and 5/30 isolates were A. hydrophila ST 251. Our isolates and all publicly available A. hydrophila isolates from Vietnam belonged to ST 251 and differed with

The PhoBR two-component system is a critical regulator of virulence in many bacterial pathogens, but its role in Aeromonas dhakensis remains poorly characterized. In this study, three markerless knockout mutants (Δ pho B, Δ pho R and Δ pho BR) were constructed to explore the function of PhoBR. Under low phosphate conditions (0.2 mM Pi), physiological analyses showed that all mutants exhibited defects in growth, motility, biofilm formation and resistance to acid, with Δ pho BR also showing resistance to H 2 O 2 , while no significant changes were observed in extracellular protease secretion. Cell adhesion and zebrafish infection assays indicated that PhoBR is crucial for A. dhakensis adhesion to EPC cells. The Δ pho BR mutant had the highest LD 50 value (1.77 × 107 CFU per fish), which was 38.15 times higher than that of the wild type strain. Additionally, Δ pho BR conferred the highest relative percent survival (RPS) of 61.5 % in zebrafish following two immunizations. These findings suggest that PhoBR positively regulates virulence in A. dhakensis and that the Δ pho BR mutant could serve as a potential vaccine candidate for the prevention of aeromoniasis in aquaculture. • Δ pho BR exhibits defective in growth, motility, biofilm formation, adhesion to EPC cells, and resistance to acid and H 2 O 2. • Δ pho BR is more attenuated in zebrafish than both Δ pho B and Δ pho R mutants. • Double vaccination with Δ pho BR confers 61.5 % relative percent survival (RPS) to zebrafish against the isogenic wild type strain. [ABSTRACT FROM AUTHOR]

The development of biomaterials to enable application of antimicrobial peptides represents a strategy of high and current interest. In this study, a bioparticle was produced by the complexation between an antimicrobial polypeptide and the biocompatible and biodegradable polysaccharides chitosan-N-arginine and alginate, giving rise to a colloidal polyelectrolytic complex of pH-responsive properties. The inclusion of the polypeptide in the bioparticle structure largely increases the binding sites of complexation during the bioparticles production, leading to its effective incorporation. After lyophilization, detailed evaluation of colloidal structure of redispersed bioparticles evidenced nano or microparticles with size, polydispersity and zeta potential dependent on pH and ionic strength, and the dependence was not withdrawn with the polypeptide inclusion. Significant increase of pore edge tension in giant vesicles evidenced effective interaction of the polypeptide-bioparticle with lipid model membrane. Antibacterial activity against Aeromonas dhakensis was effective at 0.1% and equal for the isolated polypeptide and the same complexed in bioparticle, which opens perspectives to the composite material as an applicable antibacterial system. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis is ubiquitous in aquatic habitats and can cause life-threatening septicaemia in humans. However, limited data are available on their antimicrobial susceptibility testing (AST) profiles. Hence, we aimed to examine their AST patterns using clinical (n = 94) and non-clinical (n = 23) isolates with dehydrated MicroScan microdilution. Carbapenem resistant isolates were further screened for genes related to carbapenem resistance using molecular assay. The isolates exhibited resistance to imipenem (76.9%), doripenem (62.4%), meropenem (41.9%), trimethoprim/sulfamethoxazole (11.1%), cefotaxime (8.5%), ceftazidime (6%), cefepime (1.7%) and aztreonam (0.9%), whereas all isolates were susceptible to amikacin. Clinical isolates showed significant association with resistance to doripenem, imipenem and meropenem compared to non-clinical isolates. These blacphA were detected in clinical isolates with resistance phenotypes: doripenem (67.2%, 45/67), imipenem (65.9%, 54/82) and meropenem (65.2%, 30/46). Our findings showed that the MicroScan microdilution method is suitable for the detection of carbapenem resistance in both clinical (48.9–87.2%) and non-clinical (4.3–13.0%) isolates. This study revealed that A. dhakensis isolates had relatively high carbapenem resistance, which may lead to potential treatment failure. Continued monitoring of aquatic sources with a larger sample size should be carried out to provide further insights. [ABSTRACT FROM AUTHOR]

Abstract Background The study aimed to elucidate the species taxonomy, clinical manifestations, virulence gene profiles and antimicrobial susceptibilities of Aeromonas strains isolated from life-threatening bacteremia in southeastern China. Methods Clinical samples of Aeromonas causing bacteremia were isolated from a teaching hospital in Wenzhou from 2013 to 2018 and a retrospective cohort study was performed. Aeromonas strains were identified at species level by housekeeping gene gyrB. Virulence and drug resistance-associated genes were screened by polymerase chain reaction (PCR) and antimicrobial susceptibility testing (AST) was performed by the VITEK 2 Compact system. Results A total of 58 Aeromonas isolated from patients with bacteremia were collected during 6 years (2013–2018). 58 isolates were identified to five different species, where Aeromonas dhakensis appeared to be the predominant species (26/58), followed by Aeromonas veronii (13/58), Aeromonas caviae (10/58), Aeromonas hydrophila (7/58) and Aeromonas jandaei (2/58). 16 of 58 patients had poor prognosis. Poor prognosis was significantly associated with liver cirrhosis and inappropriate empirical antimicrobials therapy. The progression of bacteremia caused by Aeromonas was extremely fast, especially in A. dhakensis infections. Virulence genes aer, lip, hlyA, alt, ast, and act, were detected at ratios of 24.1% (14/58), 62.1% (36/58), 65.5% (38/58), 58.6% (34/58), 15.5% (9/58) and 65.5% (38/58), respectively. Antimicrobial susceptibility testing exhibited that 9 out of 58 isolates were identified as multi-drug resistant (MDR) organism. The bla TEM gene was identified in all 9 MDR isolates. bla SHV, bla AQU-1, bla MOX, bla CepH, bla CphA and aac(6′)-Ib-cr were detected in 4 isolates, 2 isolates, 1 isolate, 3 isolates, 8 isolates, and 3 isolates, respectively. The majority of Aeromonas strains maintained susceptible to 3rd generation cephalosporins, aminoglycosides, fluoroquinolones and furantoin. Conclusions The prevalence and dangerousness of Aeromonas infections, especially A. dhakensis, are underestimated in clinic. Continuous monitoring is essential to keep track of MDR Aeromonas due to the increasing prevalence recently and a more effective measure is required to control the spread of resistance determinants.

Species of Aeromonas are ubiquitous pathogens of fish and aquatic animals and can infect humans and other animals through the food chain. However, there are few reports of marine mammalian infections. In 2020, a sea lion (Zalophus californianus) died acutely at an aquarium in Harbin, Heilongjiang Province, China. In order to explore the cause of death, we dissected the animal and observed pathological changes. Ogans were aseptically collected and used for bacterial isolation and culture. This revealed that the sea lion had died of sepsis caused by a bacterial infection. Isolated bacteria were investigated by morphology, biochemical phenotype and molecular identification, and this determined the pathogen as A. dhakensis. The isolate contained six virulence genes, hlyA, aerA, act, lafA, ela, fla, and was susceptible to most antibiotics. This is the first report of A. dhakensis associated with septicaemia in pinnipeds and a description of its virulence and resistance profiles. Its presence in aquatic environments poses a potential threat to marine mammals. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis is an emerging human pathogen which causes fast and severe infections worldwide. Under the gradual pressure of lacking useful antibiotics, finding a new strategy against A. dhakensis infection is urgent. To understand its pathogenesis, we created an A. dhakensis AAK1 mini-Tn10 transposon library to study the mechanism of A. dhakensis infection. By using a Caenorhabditis elegans model, we established a screening platform for the purpose of identifying attenuated mutants. The uvrY mutant, which conferred the most attenuated toxicity toward C. elegans, was identified. The uvrY mutant was also less virulent in C2C12 fibroblast and mice models, in line with in vitro results. To further elucidate the mechanism of UvrY in controlling the toxicity in A. dhakensis, we conducted a transcriptomic analysis. The RNAseq results showed that the expression of a unique hemolysin ahh1 and other virulence factors were regulated by UvrY. Complementation of Ahh1, one of the most important virulence factors, rescued the pore-formation phenotype of uvrY mutant in C. elegans; however, complementation of ahh1 endogenous promoter-driven ahh1 could not produce Ahh1 and rescue the virulence in the uvrY mutant. These findings suggest that UvrY is required for the expression of Ahh1 in A. dhakensis. Taken together, our results suggested that UvrY controls several different virulence factors and is required for the full virulence of A. dhakensis. The two-component regulator UvrY therefore a potential therapeutic target which is worthy of further study.

Aeromonas dhakensis is ubiquitous in aquatic habitats and can cause life-threatening septicaemia in humans. However, limited data are available on their antimicrobial susceptibility testing (AST) profiles. Hence, we aimed to examine their AST patterns using clinical (n = 94) and non-clinical (n = 23) isolates with dehydrated MicroScan microdilution. Carbapenem resistant isolates were further screened for genes related to carbapenem resistance using molecular assay. The isolates exhibited resistance to imipenem (76.9%), doripenem (62.4%), meropenem (41.9%), trimethoprim/sulfamethoxazole (11.1%), cefotaxime (8.5%), ceftazidime (6%), cefepime (1.7%) and aztreonam (0.9%), whereas all isolates were susceptible to amikacin. Clinical isolates showed significant association with resistance to doripenem, imipenem and meropenem compared to non-clinical isolates. These blacphA were detected in clinical isolates with resistance phenotypes: doripenem (67.2%, 45/67), imipenem (65.9%, 54/82) and meropenem (65.2%, 30/46). Our findings showed that the MicroScan microdilution method is suitable for the detection of carbapenem resistance in both clinical (48.9–87.2%) and non-clinical (4.3–13.0%) isolates. This study revealed that A. dhakensis isolates had relatively high carbapenem resistance, which may lead to potential treatment failure. Continued monitoring of aquatic sources with a larger sample size should be carried out to provide further insights.

Essential oils are plant extracts that have been used for their antimicrobial properties for centuries. The keeping of turtles as pets exhibits a growing trend worldwide but these animals are known to harbour a range of pathogenic bacteria. In the current study, we assessed eight essential oils as alternative antibacterial agents against nine species of pet turtle-borne Gram-negative bacteria, namely Aeromonas caviae, A. dhakensis, A. hydrophila, Citrobacter freundii, Morganella morganii, Proteus mirabilis, P. vulgaris, Pseudomonas aeruginosa and Salmonella enterica. Except for Pseudomonas aeruginosa, all other bacterial species showed high susceptibility to six essential oils, namely oregano, cinnamon, clove, lemongrass, lavender and eucalyptus oils in descending order of efficacy. Minimum inhibitory concentrations and minimum bactericidal concentrations values of the essential oils against all tested species except for P. aeruginosa showed low heterogeneity, showing that these essential oils can effectively control the growth of nearly all the tested. However, most of the tested bacteria were multiple-antibiotic-resistant as determined in the antibiotic disc diffusion test, with multiple-antibiotic-resistant index values of ≥ 0.2 for most of the strains. Therefore, with regards to their in vitro activity in controlling growth of multi-drug resistant bacteria, we can classify oregano, cinnamon, clove, lemongrass, lavender and eucalyptus essential oils as effective antibacterial agents. Thus, prospective application of these essential oils in controlling and treating these bacteria should be considered.

Background: The study aimed to elucidate the species taxonomy, clinical manifestations, virulence gene profiles and antimicrobial susceptibilities of Aeromonas strains isolated from life-threatening bacteremia in southeastern China. Methods: Clinical samples of Aeromonas causing bacteremia were isolated from a teaching hospital in Wenzhou from 2013 to 2018 and a retrospective cohort study was performed. Aeromonas strains were identified at species level by housekeeping gene gyrB. Virulence and drug resistance-associated genes were screened by polymerase chain reaction (PCR) and antimicrobial susceptibility testing (AST) was performed by the VITEK 2 Compact system. Results: A total of 58 Aeromonas isolated from patients with bacteremia were collected during 6 years (2013–2018). 58 isolates were identified to five different species, where Aeromonas dhakensis appeared to be the predominant species (26/58), followed by Aeromonas veronii (13/58), Aeromonas caviae (10/58), Aeromonas hydrophila (7/58) and Aeromonas jandaei (2/58). 16 of 58 patients had poor prognosis. Poor prognosis was significantly associated with liver cirrhosis and inappropriate empirical antimicrobials therapy. The progression of bacteremia caused by Aeromonas was extremely fast, especially in A. dhakensis infections. Virulence genes aer, lip, hlyA, alt, ast, and act, were detected at ratios of 24.1% (14/58), 62.1% (36/58), 65.5% (38/58), 58.6% (34/58), 15.5% (9/58) and 65.5% (38/58), respectively. Antimicrobial susceptibility testing exhibited that 9 out of 58 isolates were identified as multi-drug resistant (MDR) organism. The blaTEM gene was identified in all 9 MDR isolates. blaSHV, blaAQU-1, blaMOX, blaCepH, blaCphA and aac(6′)-Ib-cr were detected in 4 isolates, 2 isolates, 1 isolate, 3 isolates, 8 isolates, and 3 isolates, respectively. The majority of Aeromonas strains maintained susceptible to 3rd generation cephalosporins, aminoglycosides, fluoroquinolones and furantoin. Conclusions: The prevalence and dangerousness of Aeromonas infections, especially A. dhakensis, are underestimated in clinic. Continuous monitoring is essential to keep track of MDR Aeromonas due to the increasing prevalence recently and a more effective measure is required to control the spread of resistance determinants. [ABSTRACT FROM AUTHOR]

Aeromonads are ubiquitous in aquatic environments and have been implicated in fish and human infections. In this study, we isolated, studied antimicrobial susceptibility patterns and screened the existence of 15 virulence genes in aeromonads from two famously consumed fish species—seven marine Tiger Grouper (Epinephelus fuscoguttatus) and eight freshwater Marble Goby (Oxyeleotris marmoratus) from the aquaculture hatchery in Sabah, Malaysia. A total of 30 aeromonads (17 A. caviae, 9 A. rivuli, 4 A. dhakensis) were identified using PCR targeting GCAT gene, rpoD‐restriction fragment length polymorphism and multi‐locus phylogenetic analysis. All 30 strains were resistant to amoxicillin and cephalothin and five strains were multidrug‐resistant. Nine virulence genes (lip, ela, eno, fla, aerA, hylA, dam, alt and ser) present in A. dhakensis, suggesting the virulence potential of this species as a fish pathogen. This study offers as a baseline for future studies in monitoring and managing these two fish in aquaculture industry. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis stands out as the most potent Aeromonas species causing a range of human diseases. This research marks the pioneering effort in isolating and characterizing virulent phages targeting A. dhakensis. Only the AM isolate among the Aeromonas isolates showed compatibility for phage isolation and was identified as A. dhakensis. Computational analysis identified the presence of virulence factors and antimicrobial resistance genes in A. dhakensis AM. Phage isolation was conducted using this particular strain as the host, resulting in the isolation of four virulent phages: vB_AdhM_DL, vB_AdhS_TS3, vB_AdhM_TS9, and vB_AdhS_M4. Bacterial numbers significantly decrease after both pre-treatment and post-treatment with individual phages and phage cocktails, ranging from 2.82 to 6.67 log CFU/mL and 4.01 to 6.49 log CFU/mL, respectively. Combining a phage cocktail with sub-MIC amoxicillin led to complete inactivation in both pre-treatment and post-treatment scenarios within a 200 µL volume. The complete genomes of phages vB_AdhM_DL, vB_AdhS_TS3, and vB_AdhM_TS9 were determined to be 42,388 bp, 115,560 bp, and 115,503 bp, respectively. This study establishes the effectiveness of using phages as an complement with sublethal antibiotic concentrations, presenting a potential and effective therapeutic approach. [ABSTRACT FROM AUTHOR]

Aeromonas spp. are commonly found in the aquatic environment and have been responsible for motile Aeromonas septicemia (MAS) in striped catfish, resulting in significant economic loss. These organisms also cause a range of opportunistic infections in humans with compromised immune systems. Here, we conducted a genomic investigation of 87 Aeromonas isolates derived from diseased catfish, healthy catfish and environmental water in catfish farms affected by MAS outbreaks in eight provinces in Mekong Delta (years: 2012-2022), together with 25 isolates from humans with bloodstream infections (years: 2010-2020). Genomics-based typing method precisely delineated Aeromonas species while traditional methods such as aerA PCR and MALDI-TOF were unable identify A. dhakensis. A. dhakensis was found to be more prevalent than A. hydrophila in both diseased catfish and human infections. A. dhakensis sequence type (ST) 656 followed by A. hydrophila ST251 were the predominant virulent species-lineages in diseased catfish (43.7 and 20.7 %, respectively), while diverse STs were found in humans with bloodstream infections. There was evidence of widespread transmission of ST656 and ST251 on striped catfish in the Mekong Delta region. ST656 and ST251 isolates carried a significantly higher number of acquired antimicrobial resistance (AMR) genes and virulence factors in comparison to other STs. They, however, exhibited several distinctions in key virulence factors (i.e. lack of type IV pili and enterotoxin ast in A. dhakensis ), AMR genes (i.e. presence of imiH carbapenemase in A. dhakensis ), and accessory gene content. To uncover potential conserved proteins of Aeromonas spp. for vaccine development, pangenome analysis has unveiled 2202 core genes between ST656 and ST251, of which 78 proteins were in either outer membrane or extracellular proteins. Our study represents one of the first genomic investigations of the species distribution, genetic landscape, and epidemiology of Aeromonas in diseased catfish and human infections in Vietnam. The emergence of antimicrobial resistant and virulent A. dhakensis strains underscores the needs of enhanced genomic surveillance and strengthening vaccine research and development in preventing Aeromonas diseases in catfish and humans, and the search for potential vaccine candidates could focus on Aeromonas core genes encoded for membrane and secreted proteins.

Aeromonas dhakensis, a newly recognized species, is often misidentified as A. hydrophila, A. veronii, or A. caviae by commercial phenotypic tests. Limited data about A. dhakensis are available in Japan. We retrospectively analyzed the patients with monomicrobial Aeromonas bacteremia at Hiroshima University Hospital from January 2011 to December 2017, and species re-identification was conducted using rpoD and gyrB gene sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system. Of the 19 strains from blood isolates, A. caviae (n = 9, 47.4%), A. dhakensis (n = 4, 21.1%), A. hydrophila (n = 3, 15.8%), and A. veronii (n = 3, 15.8%) were re-identified. A. dhakensis was phenotypically misidentified as A. hydrophila (n = 3, 75%) or A. sobria (n = 1, 25%). A. dhakensis was also misidentified as A. caviae (n = 2, 50%), A. hydrophila (n = 1, 25%), and A. jandaei (n = 1, 25%) in MALDI-TOF MS system. Malignancies (n = 12, 63.2%) and liver cirrhosis (n = 7, 36.8%) were common comorbidities. Biliary tract infection was the most frequent source of Aeromonas bacteremia (n = 11, 57.9%). The major source of A. dhakensis bacteremia was also biliary tract infection (n = 3, 75%), and the 14-day infection-related mortality of A. dhakensis was 25%. A. dhakensis isolates showed similar clinical characteristics, antimicrobial susceptibility, and mortality with those of other Aeromonas species isolates. This study demonstrated that A. dhakensis is not a rare cause of Aeromonas bacteremia, but is often misidentified as A. hydrophila in Hiroshima, Japan. Further studies should be conducted to identify the geographical distribution and clinical impact of A. dhakensis in Japan. [ABSTRACT FROM AUTHOR]

As one of the most important economic fishes, largemouth bass (Micropterus salmoides) is being widely farmed around the world. With the improper expansion and high-density culture, largemouth bass is susceptible to an increasing number of pathogens. Particularly, co-infection of multiple pathogens is ubiquitous, which has become a main obstacle to the development of largemouth bass industry. In the study, a co-infected case caused by Aeromonas dhakensis (A. dhakensis) and Chryseobacterium indologenes (C. indologenes) in largemouth bass was firstly reported. Diseased fish with clinical signs such as exophthalmia, hemorrhage on gill cover, rot or ulceration on gill, skin and fin were collected for further diagnostics. Tissue lesions of liver, spleen and kidney were observed, such as necrosis, hemorrhage, vacuolization and disintegration. Two bacteria strains were isolated from liver, one appeared as a white or yellowish colony, and the other was characterized by golden yellow color. Based on the conjoint analysis of 16S rRNA and gyrB gene sequences, the isolated strains were identified as A. dhakensis and C. indologenes , respectively. Antibiotic sensitivity testing indicated that both A. dhakensis and C. indologenes were sensitive to flumequine, enrofloxacin, ceftazidime and minocycline, and resistant to penicillin, sulfamethoxine sodium, ampicillin and cefamezin. Largemouth bass showed similar clinical signs and tissue lesions to naturally diseased fish following co-infection with the isolated strains, indicating that the isolated strains fulfill Koch's postulates. Besides, a slightly increased pathogenicity was observed following co-infection compared with single infection. The data firstly reveal a co-infected case caused by A. dhakensis and C. indologenes in largemouth bass, not only enriching the relevant information of largemouth bass diseases, but also promoting the diagnosis, prevention and control of A. dhakensis and C. indologenes co-infection. • Co-infection of A. dhakensis and C. indologenes in largemouth bass is firstly reported. • Conjoint analysis of 16S rRNA and gyrB is performed to identify the isolated strains. • Both A. dhakensis and C. indologenes show sensitive to flumequine, enrofloxacin, ceftazidime and minocycline. • The isolated A. dhakensis and C. indologenes fulfill Koch's postulates. [ABSTRACT FROM AUTHOR]

Some Aeromonas species, potentially pathogenic for humans, are known to express up to three different classes of chromosomal β-lactamases, which may become hyperproduced and cause treatment failure. The aim of this study was to assess the utility of these species-specific β-lactamase genes as phylogenetic markers using whole-genome sequencing data. Core-genome alignments were generated for 36 Aeromonas genomes from seven different species and scanned for antimicrobial resistance genes. Core-genome alignment confirmed the MALDI-TOF identification of most of the isolates and re-identified an A. hydrophila isolate as A. dhakensis. Three (B, C and D) of the four Ambler classes of β-lactamase genes were found in A. sobria, A. allosacharophila, A. hydrophila and A. dhakensis (blaCphA, blaAmpC and blaOXA). A. veronii only showed class-B- and class-D-like matches (blaCphA and blaOXA), whereas those for A. media, A. rivipollensis and A. caviae were class C and D (blaCMY, blaMOX and blaOXA427). The phylogenetic tree derived from concatenated sequences of β-lactamase genes successfully clustered each species. Some isolates also had resistance to sulfonamides, quinolones and aminoglycosides. Whole-genome sequencing proved to be a useful method to identify Aeromonas at the species level, which led to the unexpected identification of A. dhakensis and A.rivipollensis and revealed the resistome of each isolate.

Innate immunity is the primary defense mechanism against infection in metazoans. However, aberrant upregulation of innate immune-signaling pathways can also be detrimental to the host. The p38 MAPK/PMK-1 innate immune-signaling pathway has been demonstrated to play essential roles in cellular defenses against numerous infections in metazoans, including Caenorhabditis elegans. However, the negative regulators that maintain the homeostasis of this important innate immune pathway remain largely understudied. By screening a focused RNAi library against the kinome of C. elegans, we identified RIOK-1, a human RIO kinase homolog, as a novel suppressor of the p38 MAPK/PMK-1 signal pathway. We demonstrated that the suppression of riok-1 confers resistance to Aeromonas dhakensis infection in C. elegans. Using quantitative real time-PCR and riok-1 reporter worms, we found the expression levels of riok-1 to be significantly upregulated in worms infected with A. dhakensis. Our genetic epistasis analysis suggested that riok-1 acts on the upstream of the p38 MAPK/pmk-1 genetic pathway. Moreover, the suppression of riok-1 enhanced the p38 MAPK signal, suggesting that riok-1 is a negative regulator of this innate pathway in C. elegans. Our epistatic results put riok-1 downstream of skn-1, which encodes a p38 MAPK downstream transcription factor and serves as a feedback loop to the p38 MAPK pathway during an A. dhakensis infection. In conclusion, riok-1 is proposed as a novel innate immune suppressor and as a negative feedback loop model involving p38 MAPK, SKN-1, and RIOK-1 in C. elegans.

Hundreds of farmed Siamese crocodiles (Crocodylus siamensis) died during July 2016 at a farm in Wenchang, Hainan, China. In two necropsied crocodiles, we observed symptoms of dermatorrhagia, hepatomegaly and hepatic congestion. Pulmonitis was diagnosed by pulmonary congestion and pulmonary fibrinous exudate. Septicaemia was diagnosed by isolation of three Aeromonas species from blood and visceral tissues; A. dhakensis, A. hydrophila and A. jandaei were identified by biochemical and molecular tests. We used a zebrafish model to determine the half‐maximal lethal dose (LD50), and A. dhakensis was found to be the most virulent species, with an LD50 of 8·91 × 105CFU per ml. The results of a drug sensitivity test indicated that these species were sensitive to 11 antibiotics. This is the first report of A. dhakensis, A. hydrophila and A. jandaei being isolated from a mixed infection in Siamese crocodiles. Significance and Impact of the Study: In this study, we isolated three species of Aeromonas (A. dhakensis, A. hydrophila and A. jandae) from farmed Siamese crocodiles with fatal fibrinous pneumonia and septicaemia. This is the first description of a mixed infection with three Aeromonas species among captive crocodilians. Significance and Impact of the Study: In this study, we isolated three species of Aeromonas (A. dhakensis, A. hydrophila and A. jandae) from farmed Siamese crocodiles with fatal fibrinous pneumonia and septicaemia. This is the first description of a mixed infection with three Aeromonas species among captive crocodilians. [ABSTRACT FROM AUTHOR]

Essential oils are plant extracts that have been used for their antimicrobial properties for centuries. The keeping of turtles as pets exhibits a growing trend worldwide but these animals are known to harbour a range of pathogenic bacteria. In the current study, we assessed eight essential oils as alternative antibacterial agents against nine species of pet turtle-borne Gram-negative bacteria, namely Aeromonas caviae, A. dhakensis, A. hydrophila, Citrobacter freundii, Morganella morganii, Proteus mirabilis, P. vulgaris, Pseudomonas aeruginosa and Salmonella enterica. Except for Pseudomonas aeruginosa, all other bacterial species showed high susceptibility to six essential oils, namely oregano, cinnamon, clove, lemongrass, lavender and eucalyptus oils in descending order of efficacy. Minimum inhibitory concentrations and minimum bactericidal concentrations values of the essential oils against all tested species except for P. aeruginosa showed low heterogeneity, showing that these essential oils can effectively control the growth of nearly all the tested. However, most of the tested bacteria were multiple-antibiotic-resistant as determined in the antibiotic disc diffusion test, with multiple-antibiotic-resistant index values of ≥ 0.2 for most of the strains. Therefore, with regards to their in vitro activity in controlling growth of multi-drug resistant bacteria, we can classify oregano, cinnamon, clove, lemongrass, lavender and eucalyptus essential oils as effective antibacterial agents. Thus, prospective application of these essential oils in controlling and treating these bacteria should be considered. [ABSTRACT FROM AUTHOR]

Innate immunity is the primary defense mechanism against infection in metazoans. However, aberrant upregulation of innate immune-signaling pathways can also be detrimental to the host. The p38 MAPK/PMK-1 innate immune-signaling pathway has been demonstrated to play essential roles in cellular defenses against numerous infections in metazoans, including Caenorhabditis elegans. However, the negative regulators that maintain the homeostasis of this important innate immune pathway remain largely understudied. By screening a focused RNAi library against the kinome of C. elegans, we identified RIOK-1, a human RIO kinase homolog, as a novel suppressor of the p38 MAPK/PMK-1 signal pathway. We demonstrated that the suppression of riok-1 confers resistance to Aeromonas dhakensis infection in C. elegans. Using quantitative real time-PCR and riok-1 reporter worms, we found the expression levels of riok-1 to be significantly upregulated in worms infected with A. dhakensis. Our genetic epistasis analysis suggested that riok-1 acts on the upstream of the p38 MAPK/pmk-1 genetic pathway. Moreover, the suppression of riok-1 enhanced the p38 MAPK signal, suggesting that riok-1 is a negative regulator of this innate pathway in C. elegans. Our epistatic results put riok-1 downstream of skn-1, which encodes a p38 MAPK downstream transcription factor and serves as a feedback loop to the p38 MAPK pathway during an A. dhakensis infection. In conclusion, riok-1 is proposed as a novel innate immune suppressor and as a negative feedback loop model involving p38 MAPK, SKN-1, and RIOK-1 in C. elegans. [ABSTRACT FROM AUTHOR]

The Macrobrachium rosenbergii industry is threatened by various Aeromonas , resulting in high mortality of adult prawns. However, there are few studies on the immune response of M. rosenbergii infected with Aeromonas dhakensis. In this study, we observed the hepatopancreas and gills histopathologically, performed a comparative transcriptome analysis of the hepatopancreas, and analyzed the candidate gene expression of immune-related genes in the hemolymph, hepatopancreas, and gills of M. rosenbergii that had been infected with A. dhakensis. Histopathology revealed the hepatopancreas was successively inflamed, followed by cellular vacuolation, lumen deformation, and finally tissue erosion; partial and severe inflammation of the gills occurred successively, and eventually the gill tissue atrophy and the gill filaments detached from the gill arch. Transcriptome analysis showed that a total of 77,742 unigenes and 8664 differentially expressed genes (DEGs), and the immune-related DEGs were mainly enriched in lysosome and phagosome pathways. In addition, 4 immune-related candidate genes (RhoA, CASP9, PKC, and DSCIGN) based on KEGG and PPI analysis were monitored at 6, 12, and 24h post injection (hpi) in hepatopancreas, hemolymph and gills. Their spatio-temporal expression results indicated that A. dhakensis have activated the immune system of M. rosenbergii. The present study may provide new information on the complex immune mechanism of M. rosenbergii. • It was the first report on the immune response of Macrobrachium rosenbergii infected with Aeromonas dhakensis. • The tissures of M. rosenbergii in the challenge group had various degrees of damage. • The candidate immune-related genes (Casp9 , PKC , RhoA , and DCSIGN) actively participated in immune activities. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is an important effector molecule which is involved in a myriad of biological processes, including immune responses against pathogens such as parasites, virus and bacteria. During the inflammatory processes in vertebrates, NO is produced by the inducible nitric oxide synthase (iNOS) enzyme in practically all nucleated cells to suppress or kill intracellular pathogens. The aim of the present study was to characterize the full coding region of the iNOS gene of pacu ( Piaractus mesopotamicus ), an economically and ecologically important South American fish species, and to analyze mRNA expression levels following intraperitoneal infection with the pathogenic bacterium Aeromonas dhakensis by means of quantitative real time PCR (qPCR). The results showed that the pacu iNOS transcript is 3237 bp in length, encoding a putative protein composed of 1078 amino acid residues. The amino acid sequence showed similarities ranging from 69.03% to 94.34% with other teleost fish and 57.70% with the human iNOS, with all characteristic domains and cofactor binding sites of the enzyme detected. Phylogenetic analysis showed that the iNOS from the red-bellied piranha, another South American characiform, was the closest related sequence to the pacu iNOS. iNOS transcripts were constitutively detected in the liver, spleen and head kidney, and there was a significant upregulation in the liver and spleen at 12, 24 and 48 h after infection with A. dhakensis . No significant variations were observed in the head kidney during the periods analyzed. These results show that iNOS expression was induced by A. dhakensis infection and suggest that this enzyme may be involved in the response to this bacterium in pacu. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis was first isolated from children with diarrhea in Dhaka, Bangladesh and described in 2002. In the past decade, increasing evidence indicate this species is widely distributed in the environment and can cause a variety of infections both in human and animals, especially in coastal areas. A. dhakensis is often misidentified as A. hydrophila, A. veronii or A. caviae by commercial phenotypic tests in the clinical laboratory. Correct identification relies on molecular methods. Increasingly used matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) may be able to identify Aeromonas specie rapidly and accurately. A. dhakensis has shown its potent virulence in different animal models and clinical infections. Although several virulence factors had been reported, no single mechanism is conclusive. Characteristically A. dhakensis is the principal species causing soft tissue infection and bacteremia, especially among patients with liver cirrhosis or malignancy. Of note, A. dhakensis bacteremia is more lethal than bacteremia due to other Aeromonas species. The role of this species in gastroenteritis remains controversial. Third generation cephalosporins and carbapenems should be used cautiously in the treatment of severe A. dhakensis infection due to the presence of AmpC β-lactamase and metallo-β-lactamase genes, and optimal regimens may be cefepime or fluoroquinolones. Studies of bacterial virulence factors and associated host responses may provide the chance to understand the heterogeneous virulence between species. The hypothesis A. dhakensis with varied geographic prevalence and enhanced virulence that compared to other Aeromonas species warrants more investigations.

Aeromonas species is ubiquitously present in aquatic habitats, can cause motile Aeromonas septicemia (MAS). However, it is very complex to identify Aeromonas spp. based on its phenotypes. In the present study in mid-November 2018, we investigated a case of severe mortality of wild caught mascara barb (Dawkinsia assimilis) endemic to the Western Ghats, India. No cytopathic effect was observed in DRG and CSK fish cell line after inoculating filtrate of the tissue homogenates and found negative for all major viral pathogens. Here, aetiological agent was identified as bacteria, presumed as Aeromonas hydrophila , and Escherichia coli based on biochemical tests. Later, PCR assay was performed for the molecular confirmation by gyrB gene resulted in confirmation of Aeromonas as A. dhakensis with 99.63% similarity with A. dhakensis from SA1 of diseased pacu. Multiple antibiotic resistance (MAR) index of A. dhakensis was 0.9112. Further, A. dhakensis was positive for ten antibiotic resistance and seven virulence genes with β-hemolysis colonies. The LD50 value was found to be 3.42 × 105 CFU/mL in experimental trails. In the present study, a tetracycline, colistin, and ESBL resistant hypervirulent A. dhakensis was isolated from the diseased endemic mascara barb. Ten natural ingredients and five disinfectants found to be effective against A. dhakensis. To the best of our knowledge, this is the first evidence of extensively drug-resistant (XDR) virulent A. dhakensis associated with disease outbreak in endemic mascara barb in India. • Aeromonas dhakensis were producing β-hemolysis colonies on 5% sheep blood agar and positive for seven virulence genes. • DNA gyrB gene sequencing of the isolates exhibited 99.63% similarity with A. dhakensis from SA1 of diseased pacu. • A. dhakensis was sensitive to only aminoglycosides with MAR index of 0.9112 with ten positive antibiotic resistance genes. • The LD50 value was found to be 3.42 × 105 CFU/mL in experimental trails and found to be highly virulent. • This study is the first evidence of extensively drug-resistant virulent A. dhakensis in endemic mascara barb in India [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis is an emerging pathogen that is harmful to humans and aquaculture in tropical areas. The aim of this study was to understand the role of the two-component signal transduction system KdpDE in regulating the virulence of this species. A Δ kdp E mutant was generated via a markerless knockout. Phenotype analysis showed that the mutant was attenuated in growth, biofilm formation, mobility, ECPase activity, and resistance to H 2 O 2 and antimicrobial peptides under conditions of potassium depletion. A challenge experiment to analyze the virulence of A. dhakensis in zebrafish, via intraperitoneal injection, demonstrated that the clinicopathological signs of infection were dropsy, tail and/or fin bleeding, hepatonecrosis, spleen congestion, and nephrohemia. The LD 50 value of the mutant (9.02 × 106 CFU/ml) was 15.6-fold higher than that of the wild-type strain (5.77 × 105 CFU/ml). The survival rate of zebrafish was higher after double immunization (76.7%) than after single immunization with the mutant (36.7%). These data reveal for the first time that KdpDE is associated with the virulence of A. dhakensis. The Δ kdp E mutant is attenuated in virulence and shows high protective efficacy in adult zebrafish after double immunization, and is therefore a promising attenuated vaccine candidate. • dhakensis C160501 caused zebrafish hepatonecrosis, hyperaemia in skin, spleen and kidney. • △ kdp E attenuated A. dhakensis in biofilm formation, ECPase, mobility, as well as resistance to H 2 O 2 and AMPs of crocodile. • Δ kdp E resulted in the LD 50 of A. dhakensis increased 15.6-fold from 5.77 × 105 CFU/ml to 9.02 × 106 CFU/ml. • Double immunization of zebrafish with Δ kdp E mutant provided 76.7% of RPS against the challenge of isogenous C160501. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis possesses dual flagellar systems for motility under different environments. Flagella-mediated motility is necessary for biofilm formation through an initial attachment of bacteria to the surface, but this has not been elucidated in A. dhakensis. This study investigates the role of polar (flaH , maf1) and lateral (lafB , lafK and lafS) flagellar genes in the biofilm formation of a clinical A. dhakensis strain WT187 isolated from burn wound infection. Five deletion mutants and corresponding complemented strains were constructed using pDM4 and pBAD33 vectors, respectively, and analyzed for motility and biofilm formation using crystal violet staining and real-time impedance-based assays. All mutants were significantly reduced in swimming (p < 0.0001), swarming (p < 0.0001) and biofilm formation using crystal violet assay (p < 0.05). Real-time impedance-based analysis revealed WT187 biofilm was formed between 6 to 21 h, consisting of early (6–10 h), middle (11–18 h), and late (19–21 h) stages. The highest cell index of 0.0746 was recorded at 22–23 h and biofilms began to disperse starting from 24 h. Mutants Δ maf1 , Δ lafB , Δ lafK and Δ lafS exhibited reduced cell index values at 6–48 h when compared to WT187 which indicates less biofilm formation. Two complemented strains c maf1 and c lafB exhibited full restoration to wild-type level in swimming, swarming, and biofilm formation using crystal violet assay, hence suggesting that both maf1 and lafB genes are involved in biofilm formation through flagella-mediated motility and surface attachment. Our study shows the role of flagella in A. dhakensis biofilm formation warrants further investigations. • Flagella-mediated motility involved in Aeromonas dhakensis biofilm formation. • Polar Δ flaH and Δ maf1 were reduced 64–67% in swimming. • Lateral Δ lafB , Δ lafK and Δ lafS were abolished 85–86% in swarming. • Biofilm formation of all mutants was reduced 21–40% using crystal violet assay. • RTCA: maf1 and lafB genes involved in A. dhakensis biofilm formation. [ABSTRACT FROM AUTHOR]

A variety of bacterial infections cause muscle necrosis in humans. Caenorhabditis elegans has epidermis and bands of muscle that resemble soft-tissue structures in mammals and humans. Here, we developed a muscle necrosis model caused by Aeromonas dhakensis infection in C. elegans. Our data showed that A. dhakensis infected and killed C. elegans rapidly. Characteristic muscle damage in C. elegans induced by A. dhakensis was demonstrated in vivo. Relative expression levels of host necrosis-associated genes, asp-3, asp-4, and crt-1 increased significantly after A. dhakensis infection. The RNAi sensitive NL2099 rrf-3 (pk1426) worms with knockdown of necrosis genes of crt-1 and asp-4 by RNAi showed prolonged survival after A. dhakensis infection. Specifically knockdown of crt-1 and asp-4 by RNAi in WM118 worms, which restricted RNAi only to the muscle cells, conferred significant resistance to A. dhakensis infection. In contrast, the severity of muscle damage and toxicity produced by the A. dhakensis hemolysin-deletion mutant is attenuated. In another example, shiga-like toxin-producing enterohemorrhagic E. coli (EHEC) known to elicit toxicity to C. elegans with concomitant enteropathogenicty, did not cause muscle necrosis as A. dhakensis did. Taken together, these results show that Aeromonas infection induces muscle necrosis and rapid death of infected C. elegans, which are similar to muscle necrosis in humans, and then validate the value of the C. elegans model with A. dhakensis infection in studying Aeromonas pathogenicity. [ABSTRACT FROM AUTHOR]

This study provides a detailed description and characterization of a strain of Aeromonas dhakensis isolated from a diseased juvenile Piaractus mesopotamicus obtained from the fish farm of the National Center for Continental Fish Research and Conservation ( CEPTA/ ICMBio), in the state of São Paul, Brazil. Biochemical tests using the VITEK 2 automated bacterial identification system identified the isolate to genus level; however, further molecular analysis of the 16S rRNA, gyrB and rpoD genes showed that the strain belonged to the species A. dhakensis. As expected, the isolated A. dhakensis strain was resistant to ampicillin and ampicillin/sulbactam, as resistance to ampicillin is a typical characteristic of the genus Aeromonas. Resistance to cefoxitin and meropenem was also observed, but the strain was susceptible to most of the tested antibiotics. The isolated strain of A. dhakensis caused acute haemorrhagic septicaemia in experimentally infected P. mesopotamicus, with a fifty per cent lethal dose of 1.14 × 105 CFU/fish. This is the first report of the occurrence of an A. dhakensis strain causing an infection in a fish species of South America, providing important epidemiologic data relating to this important pathogenic species. [ABSTRACT FROM AUTHOR]

Aeromonas dhakensis, often phenotypically identified as Aeromonas hydrophila, is an important human pathogen. The present study aimed to compare the clinical and biological features of A. dhakensis and A. hydrophila isolates from human wounds. A total of 80 Aeromonas wound isolates collected between January 2004 and April 2011 were analysed. The species was identified by the DNA sequence matching of rpoD and gyrB (or rpoB if necessary). Most of the Aeromonas isolates were identified as A. dhakensis (37, 46.3%), and 13 (16.3%) as A. hydrophila. Both species alone can cause severe skin and soft-tissue infections. More A. dhakensis isolates were found in wounds exposed to environmental water (32.4% vs 0%, p 0.042). More biofilm formation was noted among A. dhakensis isolates (mean optical density at 570 nm, 1.23 ± 0.09 vs 0.78 ± 0.21, p 0.03). The MICs of ceftriaxone, imipenem and gentamicin for A. dhakensis isolates were higher (p <0.0001, <0.04, and <0.01, respectively). The survival rates of Caenorhabditis elegans co-incubated with A. dhakensis from day 1 to day 3 were lower than those of worms infected with A. hydrophila in liquid toxicity assays (all p values <0.01). Isolates of A. dhakensis exhibited more cytotoxicity, as measured by the released leucocyte lactate dehydrogenase levels in human normal skin fibroblast cell lines (29.6 ± 1.2% vs 20.6 ± 0.6%, p <0.0001). The cytotoxin gene ast was primarily present in A. hydrophila isolates (100% vs 2.7%, p <0.0001). In summary, A. dhakensis is the predominant species among Aeromonas wound isolates, and more virulent than A. hydrophila. [ABSTRACT FROM AUTHOR]

Abstract: Aeromonas dhakensis, a recently described Aeromonas sp. formerly called Aeromonas aquariorum, is associated with human infections. In this study, a chromosomal gene, bla AQU-1, was identified in A. dhakensis AAK1 that constitutes a 1143-bp open reading frame and is 87% identical to the gene encoding CepH in Aeromonas hydrophila. An Escherichia coli TOP10 cell transformant harbouring bla AQU-1 was resistant to cefotaxime but not to cefepime. mRNA expression of bla AQU-1 in the cefotaxime-resistant mutant strain AAK1m was 70-fold higher than in the wild strain AAK1. In all 16 A. dhakensis isolates (the major species of 51 consecutive Aeromonas blood isolates collected from June 1999 to June 2001) as well as in A. aquariorum MDC47T and A. hydrophila subsp. dhakensis LMG 19562T, but not in the reference strains or clinical isolates of other A. hydrophila subspecies, Aeromonas caviae, Aeromonas veronii or Aeromonas enteropelogenes, bla AQU-1-related genes were detected by PCR. Overall, 13 (81%) of the 16 A. dhakensis blood isolates exhibited either cefotaxime resistance or the in vitro emergence of derepressed cefotaxime-resistant mutants. In vivo selection of an A. dhakensis resistant mutant was noted in a burn patient undergoing cefotaxime monotherapy. These observations suggest that AQU-1 is a chromosomal cephalosporinase in A. dhakensis. Cefotaxime monotherapy for severe A. dhakensis infections should be used cautiously. [Copyright &y& Elsevier]

Aims To identify bacterial pathogens of diseased NiIe tilapia Oreochromis niloticus and determine their virulence. Methods and Results Sixteen bacterial isolates were recovered from diseased Nile tilapias ( O. niloticus) reared in floating cages in Adolfo Lopez Mateos (ALM), Sanalona and Dique IV dams in Sinaloa, Mexico, from February to May 2009. The bacterial isolates were identified by phenotypic and molecular (rep-PCR and 16S rRNA sequencing) methods and were mostly isolated from the kidneys and the brain of tilapias. Bacterial cells and extracellular products (ECPs) of strains were characterized and used in experimental infections with sole Solea vulgaris and Mozambican tilapia Oreochromis mossambicus. The fish challenged with Aeromonas dhakensis sp. nov. comb nov , Pseudomonas mosselii and Microbacterium paraoxydans (3·1 × 106 CFU g−1) exhibited mortality between 40 and 100% starting at 6 h postinoculation. The ECPs displayed gelatinase, haemolytic and cytotoxic activity, causing the total destruction of the HeLa cell lines. Conclusions Aeromonas dhakensis and Ps. mosselii were virulent to O. mossambicus, whereas Mic. paraoxydans displayed virulence to S. vulgaris. Significance and Impact of the Study This the first time that Aeromonas dhakensis and Ps. mosselii are reported as pathogens to tilapia and Mic. paraoxydans was isolated from fish; then, these fish pathogens could be a threat to farmed Nile tilapia in Mexico. [ABSTRACT FROM AUTHOR]

Some Aeromonas species, potentially pathogenic for humans, are known to express up to three different classes of chromosomal β-lactamases, which may become hyperproduced and cause treatment failure. The aim of this study was to assess the utility of these species-specific β-lactamase genes as phylogenetic markers using whole-genome sequencing data. Core-genome alignments were generated for 36 Aeromonas genomes from seven different species and scanned for antimicrobial resistance genes. Core-genome alignment confirmed the MALDI-TOF identification of most of the isolates and re-identified an A. hydrophila isolate as A. dhakensis. Three (B, C and D) of the four Ambler classes of β-lactamase genes were found in A. sobria, A. allosacharophila, A. hydrophila and A. dhakensis (blaCphA, blaAmpC and blaOXA). A. veronii only showed class-B- and class-D-like matches (blaCphA and blaOXA), whereas those for A. media, A. rivipollensis and A. caviae were class C and D (blaCMY, blaMOX and blaOXA427). The phylogenetic tree derived from concatenated sequences of β-lactamase genes successfully clustered each species. Some isolates also had resistance to sulfonamides, quinolones and aminoglycosides. Whole-genome sequencing proved to be a useful method to identify Aeromonas at the species level, which led to the unexpected identification of A. dhakensis and A.rivipollensis and revealed the resistome of each isolate. [ABSTRACT FROM AUTHOR]