Chemical control of psychrophilic bacterial spoilage of fish 1. Isolation and identification of psychrophilic bacteria from marine fish

Making use of the streak plate technique and low temperature incubation, 28 cultures belonging to six genera namely, Achromobacter, Flavobacterium, Pseudomonas, Micrococcus, Vibrio and Alcaligenes were isolated from different varieties of marine fish. The growth studies indicated that all of them were able to grow between -5 and +5°C within a week's time and none of them showed growth at 37°C. The optimum temperature of growth for all these cultures was in the range 25-28°C. Among these only one, i.e., a Vibrio sp., was found to be an obligate psychrophile.

Isolation of bile from fish and identification by thin layer chromatography

The paper deals with the collection of gall bladders, isolation of bile and identification of the constituents of the bile salts from different fishes. The yield of bile contents from fresh water fishes rohu, mrigal and catla was compared with that from marine fishes seer, tuna, shark and sardine. Considerable variation in yield was showed between marine and fresh water fish as well as between the species in both groups. It ranged from 0.04 to 0.06% of the body weight of fish in calla, mrigal and rohu. The bile constituents from rohu and mrigal were analysed by thin layer chromatography. The result showed that bile of rohu and mrigal contains mainly taurine derivative of lithocholic acid.

Artificial fertilization of the eggs, and rearing and identification of the larvae of the anchoveta, Cetengraulis mysticetus

ENGLISH: The anchoveta, Cetengraulis mysticetus (Günther), is an important bait fish used to capture tunas in the Eastern Tropical Pacific Ocean. Contributions to the early life history of this species in the Gulf of Panama were made by Simpson (1959), who was able to identify deductively the planktonic egg of the anchoveta from 10 other anchovy eggs concurrently present. He also reared these planktonic eggs in the laboratory and described the resultant larvae to the age of 48 hours after hatching. Because of the lack of differences among the anchovy larvae, this description does not permit the identification of anchoveta larvae from those of other engraulid species. Furthermore, while adult specimens are easily recognized, up to the present it has not been possible to extend the identification of the juvenile anchoveta to specimens smaller than about 25 mm. The purpose of this study, therefore, was to identify anchoveta from the time of hatching to about 25 mm. SPANISH: La anchoveta, Cetengraulis mysticetus (Günther), es un importante pez de carnada que se emplea en la captura de los atunes en el Océano Pacífico Oriental Tropical. Simpson (1959) logró identificar deductivamente el huevo planctónico de la anchoveta al separarlo de otros diez huevos de anchoas que se encuentran al mismo tiempo, contribuyendo de esta manera a conocer los primeros estados de la historia natural de esta especie en el Golfo de Panamá. El también estableció un criadero en el laboratorio con estos huevos planctónicos y describió las larvas resultantes hasta la edad de 48 horas después de la eclosión. Debido a que no hay diferencias entre las larvas de las anchoas, esta descripción no permite identificar las larvas de la anchoveta de las otras especies de engráulidos. Más aun, a pesar de que los especímenes adultos son fácilmente reconocibles, hasta ahora no ha sido posible identificar la anchoveta juvenil de menos de unos 25 mm. Consecuentemente, el propósito del presente estudio ha sido el de identificar al anchoveta desde el momento de la eclosión hasta que tiene unos 25 mm.

Detection of cryptosporidium oocysts in water and environmental concentrates

Whilst current methods for the isolation and enumeration of Cryptosporidium spp. oocysts in water have provided some insight into their occurrence and significance, they are regarded as being inefficient, variable and time-consuming, with much of the interpretation being left to the expertise of the analyst. Two expectations of novel developments are to reduce the variability and subjectivity associated with the isolation and identification of oocysts. Flocculation, immunomagnetisable and flow cytometric techniques, for concentrating oocysts from water samples, should prove more reliable than current methods, whilst the development of more avid and specific monoclonal antibodies in conjunction with the use of nuclear fluorochromes will aid identification. Further insight into the viability, taxonomy, species identification, infectivity and virulence of the parasite should be forthcoming through the use of techniques such as the polymerase chain reaction, in situ hybridisation and non-uniform alternating current electrical fields. Such information is necessary in order to enable microbiologists, epidemiologists, engineers, utility operators and regulators to assess the safety of a water supply, with respect to Cryptosporidium contamination, more effectively.

Larval development and identification of the genus Triglops (Scorpaeniformes: Cottidae)

Prior to Pietsch’s (1993) revision of the genus Triglops, identification of their larvae was difficult; six species co-occur in the eastern North Pacific Ocean and Bering Sea and three co-occur in the western North Atlantic Ocean. We examined larvae from collections of the Alaska Fisheries Science Center and Atlantic Reference Centre and used updated meristic data, pigment patterns, and morphological characters to identify larvae of Triglops forficatus, T. macellus, T. murrayi, T. nybelini, T. pingeli, and T. scepticus; larvae of T. metopias, T. dorothy, T. jordani, and T. xenostethus have yet to be identified and are thus not included in this paper. Larval Triglops are characterized by a high myomere count (42–54), heavy dorsolateral pigmentation on the gut, and a pointed snout. Among species co-occurring in the eastern North Pacific Ocean, T. forficatus, T. macellus, and T. pingeli larvae are distinguished from each other by meristic counts and presence or absence of a series of postanal ventral melanophores. Triglops scepticus is differentiated from other eastern North Pacific Ocean larvae by having 0–3 postanal ventral melanophores, a large eye, and a large body depth. Among species co-occurring in the western North Atlantic Ocean, T. murrayi and T. pingeli larvae are distinguished from each other by meristic counts (vertebrae, dorsal-fin rays, and anal-fin rays once formed), number of postanal ventral melanophores, and first appearance and size of head spines. Triglops nybelini is distinguished from T. murrayi and T. pingeli by a large eye, pigment on the lateral line and dorsal midline in flexion larvae, and a greater number of dorsal-fin rays and pectoral-fin rays once formed.

Improved handling and preservation of freshwater giant prawn, Macrobrachium rosenbergii for producing safe and wholesome products

Studies were undertaken to evaluate the quality changes in freshwater giant prawn, Macrobrachium rosenbergii during various storage conditions of handling and preservation and producing safe and quality products. The samples kept in ice immediately after catch with head-on and head-less condition were found to be acceptable for 6 days and 7 days, respectively. Delaying of icing considerably shortened the shelf-life. The pH value increased from 6.36 to 8.0 after 10 days in ice. The initial average TVB-N value of sample increased from below 10 mg/100 g to 25 mg/100 g with the lapse of storage period. The Ca++ ATPase activity in presence of 0.1M KCl slightly decreased at the end of 10 days of ice storage. Immediately after harvest, initial aerobic plate count (APC) was 2.88x10^6 CFU/g which gradually increased to 1.12x10^8 CFU/g after 6 days in ice storage and showed early signs of spoilage. Initial bacterial genera in the prawn iced at 0 hours were comprised of Coryneform (22.21 %), Bacillus (7.40%), Micrococcus (11.11 %), Achromobacter (48.14%), Flavobacterium/Cytophaga (7.40%), Pseudomonas (3.70%) and Aeromonas (3.70%). During ice storage Coryneforms and Bacillus were always dominating along with less prominent ones - Micrococcus, Achromobacter and Flavobacterium. Studies were conducted on the stability of myofibrillar protein of M. rosenbergii under different storage and pH conditions. The influence of a wide range of pH on the remaining Ca++ ATPase activity of M. rosenbergii muscle myofibrils after storage at -20°C for 2 days, at 0°C for 2 days and at 35°C for 30 minutes demonstrated that ATPase activities were lower in acidic and alkaline pH regions and the activity remained relatively high. Mg++ ATPase activities both in presence and absence of Ca++ remained high at neutral pH compared to those of acidic and alkaline region. The solubility of myofibrillar protein decreased gradually both in acidic and alkaline pH regions. The study also examined the bacteriological quality of freshly harvested M. rosenbergii, pond sediment and pond water from four commercial freshwater prawn farms at Fulpur and Tarakanda upazilas in the district of Mymensingh. The study included aerobic plate count (APC), total coliform count, detection, isolation and identification of suspected public health hazard bacteria and their seasonal variation, salt tolerance test, antibiotic sensitivity test of the isolates and washing effect of chlorinated water on the bacterial load in the prawn samples. APC in sediment soil and water of the farm and gill and hepatopancreas of freshly harvested prawns varied considerably among the farms and between summer and winter season. The range of coliform count in water, gill and hepatopancreas ranged between 6 - 2.8x10^2 CFU/ml, 1.2x10^2 - 3.32x10^2 CFU/g and 1.43x10^2 - 3.89 x10^3 CFU/g, respectively. No coliform was detected in pond sediment sample. Suspected health hazard bacteria isolated and identified from pond sediment, water, gill and hepatopancreas included Streptococcus, Bacillus, Escherichia coli, Klebsialla, Salmonella, Staphylococcus, Pseudomonas and Aeromonas. Bacillus, Salmonella and Staphyloccus [sic], and were found to be highly salt tolerant and capable of growing at 10% NaCl. The antibiotic discs with different concentration of antibiotics were used for the sensitivity test. The organisms were found to be most sensitive against Tetracyclin and Gentamycin.

Vibrio parahaemolyticus and the seafood industry

The role of Vibrio parahaemolyticus in food borne gastroenteritis outbreaks associated primarily with the consumption of contaminated seafoods has been well documented. Information pertaining to various aspects of its occurrence in seafoods, procedures for isolation and identification, generation time and inactivation profiles is discussed. Emphasis has been given to the response of V. parahaemolyticus to low temperatures, heating and antibacterial agents. The public health hazard posed by the pathogen is outlined and the guidelines for control are reviewed in detail.

Isolation and culture in artificial media of Lagenidium from Penaeus monodon larvae

Fungal infection of P. monodon larvae is a problem in hatchery operations. The fungus, which attacks the nauplius to postlarval stages and causes up to 100% mortality, has been tentatively identified as belonging to the genus Lagenidium . This pathogenic organism has recently been isolated and cultured. A description is given of the fungus, and features of its biology and pathology are discussed.

The study and production of fish sauce from Caspian Sea lam fish with traditional fermentation method and microbe and enzyme adding method and identification of important bacteria

Fish sauce is a popular fermented product used in south Asian countries which is made from different small fishes in this research work it was attempted to produce fish sauce from kilka of the Caspian sea, the fish sauce was made from three models of kilka ,such as whole kilka , cooked whole kilka and dressed kilka , each of these models treated it four different fashions of fermentation such as:1- Traditional method, 2- Enzymatic method 3- Microbial method, 4- Mixture of enzyme and microb The results of this investigation showed that time of fermentation for the traditional method was six month, enzymatic method one month, microbial method 3 month and the mixture of enzyme and microb 1 month. The rate of fermentation was least for dressed Kilka, microbial and biochemical changes of Kilka fish sauce were evaluated, total bacterial count was 2.1-6.15 log cfu/ml total volatile nitrogen (TVN) in samples recorded was 250 mg /100g, the amount of protein varied between 10-13 percent, the name of commercial enzymes added was Protamex and Flavourzyme, the bacteria added was L act ob acillus and Pediococous, fish sauce containers fish and 20% salt, temperature of keeping for fermentation was 37 degree c for 6 month.

Isolation and recognition Saprolegnia species from fungal infected eggs of the rainbow trout in Mazandaran province farms

Fungal infection in the eggs of freshwater fish is well known as a problematic disease. That isolation and recognition Saprolegnia fungi from fungal infected eggs of the rainbow trout in Mazandaran province was the aim of this research. For this purpose fungal infected eggs were examined from six fish farm in the fall and winter 2005-2006. The eggs with fungi were inoculated on SDA, CMA, GPagar and hemp seed and sesame seed cultures in sterile tap water at room temperature (18-24°C). In this study recognized three genera and six species Saprolegniaceae members, based on morphological characteristics which contain: Saprolegnia, Achlya, Brevilegnia. Four species were identified in the genus Saprolegnia; S.mixta, S.parasitica, S.moniliphera, S.lapponica and one species was identified in the genus Achlya; A.oblongata. S.parasitica was isolated from almost all the farms. In addition, another nine genera and species were identified; Penicillium, Aspergillus, Paeciliomyces, Acremonium, Fusarium oxysporum, F.solani , Alternaria, Helminthosporium, Mucor.

Separation and identification fresh water leeches of waters resources in Kordistan Province

In the primary studying, known leeches have included into free living and parasitic which the parasitic group, besides of direct impacts like: growth detraction, anemia, making wound in the connecting part with the skin, with making plat for secondary bacterial and molding infections in the toll place, is able to cause to transfer blood flagellate and virus to the fish too. Therefore, by accusing information related to fauna leeches of each area a risk factor from the viewpoint of the possibility of being or accusing transferred diseases by these leeches, one can predict and forehand about them. Freshwater leeches of Iran to present accurate recognition (morphological, molecular) have not been, and there are some limited reports from different parts of the country about them. One of the areas that its leeches have not been identified yet is Kurdistan, By having five latrines and big permanent rivers and 32000 springs and a lot of deep and semi-deep wells and this province is a convenient bed for growing aquatics in the country. Therefore, identifying risk factors for development of aquaculture on water resources is one important factor to access achieving development goals. For recognizing leeches of this province, some samples from 10 stands were token. Samples from under stones, sticking to the fish, turtles, plants and solid substances in the water were separated and after recording their physical characteristics, calming with 10% ethylic alcohol with 10% formalin become fixed and after painting with Carmen acetic acid by standard keys for 7 species of Helobdella stagnalis, Placobdella costata, Hemiclepsis marginata, Erpobdella octoculata, Hirudo medicinalis, Dina lineate lineata have been identified and described. Which Helobdella stagnalis has the highest distribution in the province and the minimum one is Hirudo medicinalis. However, that the data obtained in leeches in Kurdistan is a relatively complete collection in this research, recognizing fauna of these areas needs more studying. The Placobdella costata and Hemiclepsis marginata sticking to the fish were separated among identified species which showed that these are parasites for the fish. The sticking area of those leeches to the skin was accompanied with scales cast, damage to mucous membranes beneath the parasite and bleeding Was associated with Histopathology studying effect includes observing break and disconnection in the leech connecting place to the epithelial layer of epidermis in the skin, destroyed nucleus in skin Epithelial cells with observing necrosis in ulcerative place become of the leech and the sub acute inflammated penetration until acute necrosis with opening in Dermis layer is observable. Kidney of this fish have changes such as: proliferation, like proliferative kidney disease with increasing proliferative glomerular cells and increasing in membranous cells in Capillary corpuscle, observing necrotic cells in haematopoietic tissue of kidney along with increasing in infiltration of leukocyte's cells generally mono nucluars such as lymphocytes and less poly morpho nucluars such as neutrophiles that are symptoms of disorders causing anemia become of nourishing and sucking blood by the leech and creating a chronic kidney infection that originally root is in another place like the skin. Also Hemorrhagic anemia causes losing RBC's is because of using the host blood by the leech. (In this situation, one can see immature RBC red cells in Peripheral blood. To identify potential carriers of the leech to the viruses, after finding them in recorded stands and putting them in 75% ethanol for viruses cause IPN, VHS, IHN, they were tested by PCR that the conclusion of these experiments approved IPN virus in Hemiclepsis marginata and Hirudo medicinalis. This kind of leeches can act like a mechanical carrier and causing spreading the agent of this disease. It is worth mentioning that studying the pathogenicity of this virus for aquaculture sources, mentioned before needs more research. During the study of infected fish with leeches that was done after preparing bloody slides and staining them, no case blood parasites was observed. During a research about infecting fish experimentally to known leeches it become clear that 5 days after being in aquarium including leeches, samples of sticking Hirudo medicinalis leech to the golden carp with scales cast were observed. Including leeches to the fish started with molting the scales in the sticking area in the fish and fish become too uneasy and by rubbing themselves to the malls and things inside the aquarium, tried to separate them. Finally, after around 30 hours, leeches penetrate the skin, feeding from blood and tissue liquids and cause mortality the fish and then they become separated from them. If the corpse of these fish stayed in the aquarium, the Helobdella stagnalis and Erpobdella octoculata would start feeding them.

Automatic identification and enumeration of algae

A good understanding of the population dynamics of algal communities is vital in many ecological and pollution studies of freshwater and oceanic systems. Present methods require manual counting and identification of algae and can take up to 90 min to obtain a statistically reliable count on a complex population. Several alternative techniques to accelerate the process have been tried on marine samples but none have been completely successful because insufficient effort has been put into verifying the technique before field trials. The objective of the present study has been to assess the potential of in vivo fluorescence of algal pigments as a means of automatically identifying algae. For this work total fluorescence spectroscopy was chosen as the observation technique.

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.

Identification of larval sea basses (Centropristis spp.) using ribosomal DNA-specific molecular assays

The identification of sea bass (Centropristis) larvae to species is difficult because of similar morphological characters, spawning times, and overlapping species ranges. Black sea bass (Centropristis striata) is an important fishery species and is currently considered to be overfished south of Cape Hatteras, North Carolina. We describe methods for identifying three species of sea bass larvae using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) assays based on species-specific amplification of rDNA internal transcribed spacer regions. The assays were tested against DNA of ten other co-occurring reef fish species to ensure the assay's specificity. Centropristis larvae were collected on three cruises during cross-shelf transects and were used to validate the assays. Seventy-six Centropristis larva were assayed and 69 (91%) were identified successfully. DNA was not amplified from 5% of the larvae and identification was inconclusive for 3% of the larvae. Those assays can be used to identify sea bass eggs and larvae and will help to assess spawning locations, spawning times, and larval dispersal.

Chaetognatha of the Caribbean Sea and adjacent areas

This illustrated manual is a guide to the distribution and identification of the 6 genera and 28 species of benthic and planktonic Chaetognatha known to occur in the Caribbean Sea, the Gulf of Mexlco, the Florida Straits, and the southwestern North Atlantic Ocean. As background, previous studies of chaetognaths in these areas are reviewed, gross morphology of the different forms is described, and instructions on methods of preserving and handling specimens preparatory to identification are provided. The key to genera and species is preceeded by a discussion of chaetognath taxonomy. A description of each species, consisting of an abbreviated synonymy, a summary of taxonomically important morphological features, and horizontal and vertical distribution follows the key. The occurrence of species in relation to water masses in the Caribbean and adjacent areas is noted. (PDF file contains 39 pages.)