Bacterial and pathological study of caudal fin rot in brood stocks of Salmo trutta caspius from the propagation and breeding center of Shahid Bahonar of Kelardasht region

In order to study caudal fin rot with emphasis on Aeromonas hydrophila and Pseudomonas fluorescens in Salmo trutta caspius from the salmonids propagation and breeding center of Shahid Bahonar of kelardasht region, One hundred and eighty brood stocks having fin damage symptoms were chosen. Two bacterial samples from each fish were cultured on Aeromonas and Pseudomonas specific media. Biochemical tests, API2OE identification system and antibiogram test using six antibiotic disks were performed for diagnosing isolates bacteria and finding suitable antibiotic. Thirty samples from caudal fin of damaged fishes were fixed in 10% formalin and 51.tm microscopic sections were prepared using standard scatological methods and then stained by Haematoxylin-Eosin staining method to observe the pathological changes and also Maccallum-Goodpasture staining method to observe the bacterial colonies. In second stage of the study, bacterial samples were taken from thirty brood stocks using similar method at the first stage of sampling. For isolation and biochemical diagnosis of Aeromonas and Pseudormonas genus, the samples were analyzed by molecular research included PCR amplification (using 16S rDNA genes of the genus pseudomonas and 16S-23S rDNA intergenic spacer of the genus Aeromonas) and restriction analysis by four restriction enzymes for each genus. The results of biochemical tests showed that isolated bacteria were belonged to Aeromonas caviae and Aeromonas hydrophila (subspecies anaerogenes), Pseudomonas fluorescens, Pseudomonas putida and Pseudomonas alcaligenes while the results of API2OE identification system showed that the isolated bacteria belonged to Aeromonas hydrophila, Pseudomonas fluorescens, Pseudomonas putida and Pseudomonas aeruginosa. Restriction analysis of Aeromonas samples with Hin6l, Csp6I, Taql, and Tasl revealed three samples were different from others while restriction analysis of Pseudomonas samples with Alul, Hinfl, Rsal, and Trull showed at least five species or biovars. The results of antibiogram test showed all Aeromonas samples were sensitive to Trimethoprim, Chloramphenicol and Nitrofurazone, mostly to Nalidixic acid and Chloramphenicol, while most of samples were resistant to Erythromycin and Oxytetracycline. Pseudomonas samples were only sensitive to Nitrofurazone and mostly resistant to Oxytetracycline, Nalidixic acid, Erythromycin, Trimethoprim and Chloramphenicol. The results of light microscope study showed hyperplasia and spongiosis of the malpigian cells of epidermis, increasing of melanin pigments underlying epidermis; sever necrosis in both epidermis and dermis and also sloughing the epidermis in some cases. Occurrence of clefts through the epithelium, neovascularization, hyperemia and mild inflammatory response in dermis and separation of the fin rays also were observed. No bacterial colonies were found in the sections.

Role of Aeromonas hydrophila in bacterial septicemia of cultured carps in Khouzestan Province and investigation on protectivity of bacterin (s) prepared from acute isolate(s) of it

Motile Aeromonas are the most common bacteria of freshwater in the world that cause disease in fish and other cold-blooded and warm-blooded hosts. Among this group of bacteria, Aeromonas hydrophila is important in causing complications such as fin rot, skin ulcers and lethal hemorrhagic septicemia in fish. Several virulence factors involved in the pathogenesis of Aeromonas hydrophila, including extracellular enzymes (protease, lipase, elastase, gelatinase and nuclease) and toxins. From the exotoxins, hemolysin, aerolysin and cytolytic enterotoxin play an important role in pathogenesis. Detection of virulence markers by PCR as a key component of determining the pathogenesis of the bacteria and using indigenous vaccines for better immunization against this disease is important. In this study, a total of 200 fanned carps (126 common carp. 39 silver carp and 35 of grass carp) with symptoms suspected aeromonas septicemia were isolated from Khouzestan province farms. 125 bacteria belong to Aeromonas genus detected by biochemical and PCR methods. 31 of all isolates recognized as Aeromonas hydrophila with biochemical methods, I6srRNA detection and Lipase genes. Results showed that the role of Aeromonas sp. and Aeromonas hydrophila in fish with disease symptoms were 62.5% and 15.5% respectively. By using specific primers, three virulence genes including hemolysin, aerolysine and cytolytic enterotoxin were detected in these confirmed isolates, that 18 isolates (58/06%) hemolysin positive (hlyA +), 16 isolates (51/61%) aerolysine positive (aerA+) and 23 isolates (74/19%) for cytolytic enterotoxin gene (act+) were positive. The result of present study showed that most of the confirmed isolates genotype was hlyA+ act- with frequency equal to 51/61%. For investigating the protection effect of acut strain of bacteria, UV inactivated bacterin was used.