Detection of virulence Hemolysin gene (HlyA) in Photobacterium damselae subspecies damselae isolates

ABSTRACT Although autochthonous vibrio densities are known to be influenced by water temperature and salinity, little is understood about other environmental factors associated with their abundance and distribution. Densities of culturable Vibrio vulnificus containing vvh (V. vulnificus hemolysin gene) and V. parahaemolyticus containing tlh (thermolabile hemolysin gene, ubiquitous in V. parahaemolyticus), tdh (thermostable direct hemolysin gene, V. parahaemolyticus pathogenicity factor), and trh (tdh-related hemolysin gene, V. parahaemolyticus pathogenicity factor) were measured in coastal waters of Mississippi and Alabama. Over a 19-month sampling period, vibrio densities in water, oysters, and sediment varied significantly with sea surface temperature (SST). On average, tdh-to-tlh ratios were significantly higher than trh-to-tlh ratios in water and oysters but not in sediment. Although tlh densities were lower than vvh densities in water and in oysters, the opposite was true in sediment. Regression analysis indicated that SST had a significant association with vvh and tlh densities in water and oysters, while salinity was significantly related to vibrio densities in the water column. Chlorophyll a levels in the water were correlated significantly with vvh in sediment and oysters and with pathogenic V. parahaemolyticus (tdh and trh) in the water column. Furthermore, turbidity was a significant predictor of V. parahaemolyticus density in all sample types (water, oyster, and sediment), and its role in predicting the risk of V. parahaemolyticus illness may be more important than previously realized. This study identified (i) culturable vibrios in winter sediment samples, (ii) niche-based differences in the abundance of vibrios, and (iii) predictive signatures resulting from correlations between environmental parameters and vibrio densities.

Download Full-text

16S rRNA Gene Sequence Analysis ofPhotobacterium damselae and Nested PCR Method for Rapid Detection of the Causative Agent of Fish Pasteurellosis

Applied and Environmental Microbiology ◽

10.1128/aem.65.7.2942-2946.1999 ◽

1999 ◽

Vol 65 (7) ◽

pp. 2942-2946 ◽

Cited By ~ 67

Author(s):

Carlos R. Osorio ◽

Matthew D. Collins ◽

Alicia E. Toranzo ◽

Juan L. Barja ◽

Jesús L. Romalde

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Causative Agent ◽

Nested Pcr ◽

Gene Sequence ◽

16S Rrna Gene Sequence ◽

Rrna Genes ◽

Rrna Gene ◽

Rrna Gene Sequence ◽

Photobacterium Damselae

ABSTRACT The causative agent of fish pasteurellosis, the organism formerly known as Pasteurella piscicida, has been reclassified asPhotobacterium damselae subsp. piscicida on the basis of 16S rRNA gene sequence comparisons and chromosomal DNA-DNA hybridization data; thus, this organism belongs to the same species asPhotobacterium damselae subsp. damselae(formerly Vibrio damselae). Since reassignment of P. damselae subsp. piscicida was based on only two strains, one objective of the present work was to confirm the taxonomic position of this fish pathogen by sequencing the 16S rRNA genes of 26 strains having different geographic and host origins. In addition, a nested PCR protocol for detection of P. damselae based on 16S rRNA was developed. This PCR protocol was validated by testing 35 target and 24 nontarget pure cultures, and the detection limits obtained ranged from 1 pg to 10 fg of DNA (200 to 20 cells). A similar level of sensitivity was observed when the PCR protocol was applied to fish tissues spiked with bacteria. The PCR approach described in this paper allows detection of the pathogen in mixed plate cultures obtained from asymptomatic fish suspected to be carriers of P. damselae subsp. piscicida, in which growth of this bacterium cannot be visualized. Our results indicate that the selective primers which we designed represent a powerful tool for sensitive and specific detection of fish pasteurellosis.

Download Full-text

The Fish Pathogen Vibrio ordalii Under Iron Deprivation Produces the Siderophore Piscibactin

Microorganisms ◽

10.3390/microorganisms7090313 ◽

2019 ◽

Vol 7 (9) ◽

pp. 313 ◽

Cited By ~ 3

Author(s):

Pamela Ruiz ◽

Miguel Balado ◽

Juan Carlos Fuentes-Monteverde ◽

Alicia E. Toranzo ◽

Jaime Rodríguez ◽

...

Keyword(s):

Gene Cluster ◽

Iron Uptake ◽

Peptide Mass Fingerprinting ◽

Iron Limitation ◽

Fish Pathogen ◽

Fish Pathogens ◽

Iron Deprivation ◽

Peptide Mass ◽

Photobacterium Damselae ◽

Vibrio Ordalii

Vibrio ordalii is the causative agent of vibriosis, mainly in salmonid fishes, and its virulence mechanisms are still not completely understood. In previous works we demonstrated that V. ordalii possess several iron uptake mechanisms based on heme utilization and siderophore production. The aim of the present work was to confirm the production and utilization of piscibactin as a siderophore by V. ordalii. Using genetic analysis, identification by peptide mass fingerprinting (PMF) of iron-regulated membrane proteins and chemical identification by LC-HRMS, we were able to clearly demonstrate that V. ordalii produces piscibactin under iron limitation. The synthesis and transport of this siderophore is encoded by a chromosomal gene cluster homologous to another one described in V. anguillarum, which also encodes the synthesis of piscibactin. Using β-galactosidase assays we were able to show that two potential promoters regulated by iron control the transcription of this gene cluster in V. ordalii. Moreover, biosynthetic and transport proteins corresponding to piscibactin synthesis and uptake could be identified in membrane fractions of V. ordalii cells grown under iron limitation. The synthesis of piscibactin was previously reported in other fish pathogens like Photobacterium damselae subsp. piscicida and V. anguillarum, which highlights the importance of this siderophore as a key virulence factor in Vibrionaceae bacteria infecting poikilothermic animals.

Download Full-text

Development of a molecular methodology for fast detection of Photobacterium damselae subspecies in water samples

Aquaculture ◽

10.1016/j.aquaculture.2014.09.028 ◽

2015 ◽

Vol 435 ◽

pp. 137-142 ◽

Cited By ~ 7

Author(s):

Patrícia Martins ◽

Rafael V.V. Navarro ◽

Francisco J.R.C. Coelho ◽

Newton C.M. Gomes

Keyword(s):

Water Samples ◽

Fast Detection ◽

Photobacterium Damselae

Download Full-text

Sensitive and Rapid Detection of Edwardsiellosis in Fish by a Loop-Mediated Isothermal Amplification Method

Applied and Environmental Microbiology ◽

10.1128/aem.70.1.621-624.2004 ◽

2004 ◽

Vol 70 (1) ◽

pp. 621-624 ◽

Cited By ~ 62

Author(s):

Ram Savan ◽

Arisa Igarashi ◽

Satoru Matsuoka ◽

Masahiro Sakai

Keyword(s):

Rapid Detection ◽

Japanese Flounder ◽

Isothermal Amplification ◽

Edwardsiella Tarda ◽

Fish Disease ◽

Fish Pathogen ◽

Loop Mediated Isothermal Amplification ◽

Diagnostic Protocol ◽

Amplification Method ◽

Hemolysin Gene

ABSTRACT Here we report a rapid and sensitive method (using loop-mediated isothermal amplification [LAMP]) for the diagnosis of edwardsiellosis, a fish disease caused by Edwardsiella tarda, in Japanese flounder. A set of four primers was designed, and conditions for the detection were optimized for the detection of E. tarda in 45 min at 65°C. No amplification of the target hemolysin gene was detected in other related bacteria. When the LAMP primers were used, detection of edwardsiellosis in infected Japanese flounder kidney, and spleen and seawater cultures was possible. We have developed a rapid and sensitive diagnostic protocol for edwardsiellosis detection in fish. This is the first report of the application of LAMP for the diagnosis of a fish pathogen.

Download Full-text

Complete Genome Sequence ofPhotobacterium damselaeSubsp.damselaeStrain SSPD1601 Isolated from Deep-Sea Cage-CulturedSebastes schlegeliiwith Septic Skin Ulcer

International Journal of Genomics ◽

10.1155/2019/4242653 ◽

2019 ◽

Vol 2019 ◽

pp. 1-7 ◽

Cited By ~ 1

Author(s):

Yongxiang Yu ◽

Zheng Zhang ◽

Yingeng Wang ◽

Meijie Liao ◽

Xiaojun Rong ◽

...

Keyword(s):

Antibiotic Resistance ◽

Deep Sea ◽

Clinical Symptoms ◽

Antibiotic Resistance Genes ◽

Aquatic Organisms ◽

Skin Ulcer ◽

Rrna Genes ◽

Trna Genes ◽

Photobacterium Damselae ◽

Sea Cage

Photobacterium damselaesubsp.damselae(PDD) is a Gram-negative bacterium that can infect a variety of aquatic organisms and humans. Based on an epidemiological investigation conducted over the past 3 years, PDD is one of the most important pathogens causing septic skin ulcer in deep-sea cage-culturedSebastes schlegeliiin the Huang-Bohai Sea area and present throughout the year with high abundance. To further understand the pathogenicity of this species, the pathogenic properties and genome of PDD strain SSPD1601 were analyzed. The results revealed that PDD strain SSPD1601 is a rod-shaped cell with a single polar flagellum, and the clinical symptoms were replicated during artificial infection. The SSPD1601 genome consists of two chromosomes and two plasmids, totaling 4,252,294 bp with 3,751 coding sequences (CDSs), 196 tRNA genes, and 47 rRNA genes. Common virulence factors including flagellin,Fur,RstB,hcpA, OMPs,htpB-Hsp60,VasK, andvgrGwere found in strain SSPD1601. Furthermore, SSPD1601 is a pPHDD1-negative strain containing the hemolysin genehlyAchand three putative hemolysins (emrA, yoaF, and VPA0226), which are likely responsible for the pathogenicity of SSPD1601. The phylogenetic analysis revealed SSPD1601 to be most closely related to Phdp Wu-1. In addition, the antibiotic resistance phenotype indicated that SSPD1601 was not sensitive to ceftazidime, pipemidic, streptomycin, cefalexin, bacitracin, cefoperazone sodium, acetylspiramycin, clarithromycin, amikacin, gentamycin, kanamycin, oxacillin, ampicillin, and trimethoprim-sulfamethoxazole, but only the bacitracin resistance genebacAwas detected based on Antibiotic Resistance Genes Database. These results expand our understanding of PDD, setting the stage for further studies of its pathogenesis and disease prevention.

Download Full-text