Opercular Activity and Temporal Organization of Surfacing Behaviour in Indian Catfishes,Clarias batrachus and Heteropneustes fossilis

Opercular and surfacing activity were observed in the Indian catfish Clarias batrachus and Heteropneustes fossilis. In the series A experiments, the opercular activity was monitored under two experimental conditions, viz., (1) surfacing allowed and (2) surfacing prevented. A statistically significant elevation of the rate of opercular activity was observed when air breathing was prevented in both species. In addition, a significant prevention effect and a time of day dependence of that were noticed in both species. In the series B experiments, temporal patterns of surfacing and air-gulping activities were examined under an artificial LD 12:12 schedule at 2-h intervals over a period of 48 h. The inter air-gulping interval in minutes between two consecutive bouts was also recorded four times each day in both species. A significant 24-h rhythm was found for the rate of surfacing activity and length of the inter air-gulping interval in both species. However, the overall activity appears to be much higher in Heteropneustes fossilis as compared to Clarias batrachus.     

Circadian Variation in Phototactic Behaviour of Walking Indian Catfish, Clarias batrachus

Temporal organization in phototactic rhythm in male Clarias batrachus was studied in a choice chamber consisting of a photic zone and an aphotic zone. Results indicate that most of the fishes exhibited a statistically significant 24-h variation in the frequency of visits to the photic zone with a peak frequency at late night or early morning. Cosinor analysis of phototactic index (PI) values also yielded comparable results. This behaviour of C. batrachus shows similarity with its other activities, like air gulping, surfacing, foraging or predation that occur chiefly at night. The light intensity at the surface of a natural water body is more than at the bottom, even at night. Therefore, the peak phototactic activity observed in C. batrachus in the night or early morning could be attributed to its daily migration from the bottom to the surface of the water bodies. The results of the present study might be useful in aquaculture practice and inland fishing.     

Effect of Starvation on the Characteristics of 24-h Rhythm in Surfacing Activity of an Indian Freshwater Catfish, Clarias batrachus

The present study was aimed to examine the effect of starvation on the surfacing activity of an Indian freshwater catfish, Clarias batrachus. Live Clarias batrachus of mixed sex were purchased from the local market and kept in stock aquaria under natural daylength. After proper acclimation, animals were randomly selected and divided into two groups. Animals were exposed under natural daylength and kept in specially designed glass aquaria for recording of the surfacing activity. Animals of one group were fed once a day at a fixed time (14:00 h) while the other group had no food at all for a total period of experiment (8 days). Results clearly indicate that the surfacing activity exhibits a significant 24-h rhythm in both fed and starved groups of animals. Further, the acrophases of the activity are located during the dark phase. A statistically significant effect of time on surfacing activity was validated with the help of ANOVA. The night hour mean of the activity was found significantly higher as compared to the day hour mean. Results of ANOVA also revealed a statistically significant effect due to starvation. The activity in the fed group is significantly increased as compared with starved group. Further, statistically significant rhythms with τ = 12 h and 6 h have also been noticed in both fed and starved groups. It may be concluded that starvation does not affect the multifrequency rhythms in air-breathing activity of Clarias batrachus under natural daylength.     

Effect of Pinealectomy on Re-entrainment of Air-gulping Activity in Indian Walking Catfish, Clarias batrachus

The present study was aimed to examine the effect of pinealectomy on re-entrainment of air-gulping activity in Indian walking catfish Clarias batrachus under various LD phases. Live Clarias batrachus of mixed sex (40-50g body weight) were procured from the local market and kept in the stock aquaria under the laboratory conditions for proper acclimation. Following 7 days of acclimation, animals were randomly selected from the stock aquaria and divided into 2 groups. The pineal was surgically removed from one group (P_xG) and the other group was treated as intact control group (IG). Both groups were exposed under LD 12:12 cycle inside the chronocubicles. In every 7 days LD schedule was delayed by 4 hours by lengthening of the light-on time. Both intact and pinealectomized animals are exhibited 24h rhythm in air-gulping behaviour and its entrainability irrespective of phase shifts of LD cycle. However, the phase angle of peak with reference to light on hour gradually decreased from phase 1 to phase 4, irrespective of treatment (intact/pinealectomized). Further, it has been noticed that the occurrence of multifrequency rhythm (24h, 12h and 6h) is being decreased from phase 1 to phase 4. It could be concluded that daily light-dark cycle plays an important role in modulating the rhythmic characteristics of air-breathing activity in Clarias batrachus and that pinealectomy does not modulate the effects of LD cycles on such activity.     

Effect of Pinealectomy on Re-entrainment of Air-gulping Activity in Indian Walking Catfish,Clarias batrachus

The present study was aimed to examine the effect of pinealectomy on re-entrainment of air-gulping activity in Indian walking catfish Clarias batrachus under various LD phases. Live Clarias batrachus of mixed sex (40-50g body weight) were procured from the local market and kept in the stock aquaria under the laboratory conditions for proper acclimation. Following 7 days of acclimation, animals were randomly selected from the stock aquaria and divided into 2 groups. The pineal was surgically removed from one group (P_xG) and the other group was treated as intact control group (IG). Both groups were exposed under LD 12:12 cycle inside the chronocubicles. In every 7 days LD schedule was delayed by 4 hours by lengthening of the light-on time. Both intact and pinealectomized animals are exhibited 24h rhythm in air-gulping behaviour and its entrainability irrespective of phase shifts of LD cycle. However, the phase angle of peak with reference to light on hour gradually decreased from phase 1 to phase 4, irrespective of treatment (intact/pinealectomized). Further, it has been noticed that the occurrence of multifrequency rhythm (24h, 12h and 6h) is being decreased from phase 1 to phase 4. It could be concluded that daily light-dark cycle plays an important role in modulating the rhythmic characteristics of air-breathing activity in Clarias batrachus and that pinealectomy does not modulate the effects of LD cycles on such activity.     

Diurnal and Infradian Rhythms in Lipid Parameters of Indian Catfish, Heteropneustes fossilis

Twentyfour-hour variation in ( 14 C) acetate incorporation in the liver lipid, and lipid concentrations in the liver and muscle tissues were studied during different phases of the annual reproductive cycle in the female freshwater catfish, H. fossilis . A 24-h rhythm in hepatic lipogenic activity was validated. Lipid concentration in liver and muscle also varied in a rhythmic fashion (t = 24 h). The mesor and amplitude of these rhythms appeared to be modulated by the phase of the annual gonadal cycle of H. fossilis . All three lipid parameters, in addition, exhibited low frequency rhythms with a t = 6 months or 3 months. The results indicate that the time of the day and month of the year for sampling in any experiment involving examination of lipid metabolism are of critical importance.     

Thermal tolerance and preference of the Indian catfish Heteropneustes fossilis

Synopsis The catfish Heteropneustes fossilis tolerates a wide range of temperatures. The minimal (7.9°C) and maximal (39.8°C) lethal temperature values obtained during summer are higher than the minimal (4°C) and maximal (37.7°C) lethal temperature values obtained during winter; gradual heating or cooling versus abrupt exposure to various temperatures did not produce significant differences. Catfish acclimated to temperatures of 28° (summer) or 16°C (winter) finally selected temperatures ranging from 31.3° to 32° C, when placed in a temperature gradient of 15° to 35° C. Catfish avoid temperatures below 25° C regardless of seasonal acclimatization.     

Diurnal and Infradian Rhythms in Lipid Parameters of Indian Catfish,Heteropneustes fossilis

Twentyfour-hour variation in (14 C) acetate incorporation in the liver lipid, and lipid concentrations in the liver and muscle tissues were studied during different phases of the annual reproductive cycle in the female freshwater catfish, H. fossilis. A 24-h rhythm in hepatic lipogenic activity was validated. Lipid concentration in liver and muscle also varied in a rhythmic fashion (t = 24 h). The mesor and amplitude of these rhythms appeared to be modulated by the phase of the annual gonadal cycle of H. fossilis. All three lipid parameters, in addition, exhibited low frequency rhythms with a t = 6 months or 3 months. The results indicate that the time of the day and month of the year for sampling in any experiment involving examination of lipid metabolism are of critical importance.     

Influence of temperature on ovarian recrudescence in the Indian catfish, Heteropneustes fossilis

Over a 60-day experiment during the preparatory phase of the reproductive cycle, ovarian weights increased with rise in temperature in Heteropneustes fossilis and oocyte diameters suggested an optimum temperature of 22° C for Stage II oocyte formation. The oocytes did not reach Stage II at 10° C. Atresia of Stage III oocytes occurred following 60 days of exposure at 30°C.     

Circadian Phase Dependent Thermal Stimulation of Ovarian Recrudescence in Indian Catfish, Clarias batrachus

Female Clarias batrachus acclimated to long photoperiod (13L:11D), were subjected to 30º ± 1ºC thermopulses of either 6-hour or 12-hour duration at different phases of the LD cycle during the late resting phase (first week of January) of their annual reproductive cycle. Six-hour pulses were given either at 0600 or 1200 or 1800 or 0000. Twelve-hour thermopulses were given at 0600 or 1800. The long photoperiods were started at 0530 and that of ambient at 0630 coinciding with the average timing of sunrise that prevailed during the period of the study. The results indicate that exposure to long photoperiod or constant high temperature induced gonadal growth (GSI) and elevated testosterone and oestradiol levels in plasma. The high temperature was significantly more effective in its action. Further, combination of long photoperiod and high temperature produced the strongest gonadal stimulation as gauged from GSI and the levels of steroid hormones. Interestingly, 30ºC thermopulses of 12-h duration when given at 0600 to fish held under long photoperiod induced gonadal development of comparable magnitude as observed in response to constant high temperature under long photoperiod. Thermopulses (30ºC) of 6-h duration given at 0600 or 1200 also induced significant gonadal recrudescence but of much lesser magnitude. Thermopulses either of 6-h or 12-h duration at 1800 failed to elicit any change in the variables under study. The results of cosinor analysis performed on the responses to 6-h thermopulses also substantiate that there is a rhythm in the sensitivity of C. batrachus to thermopulses. Thus it appears that in this species temperature-induced gonadal recrudescence would occur only following coincidence of high temperature with the thermoinducible phase. The underlying mechanism of this phenomenon might be circadian in nature.     

Efficacy of bi-valent whole cell inactivated bacterial vaccine against Motile Aeromonas Septicemia (MAS) in cultured catfishes (Heteropneustes fossilis,Clarias batrachus and pangasius pangasius) in Bangladesh

The Motile Aeromonas Septicemia (MAS) is an important disease of cultured catfishes (Heteropneustes fossilis, Clarias batrachus and Pangasius pangasius), caused by different species of Aeromonas bacteria which have been documented to be higher death rates (≤70%) in Bangladesh since 2016. Present study was conducted to develop bi-valent vaccine using A. hydrophila and A. veronii, and to validate their efficacy via intra-muscular (IM) and oral-routes of immunization in selected species of fishes. Brood fishes of the three species were immunized with three doses of inactivated vaccine (10⁷ CFU /2.3 mg/ml). Hematological parameters of brood fishes and antibody levels (IgM) of broods, their larvae and eggs were determined by ELISA. Additionally, Relative Percent Survivability (RPS) and the IgM levels of the larvae after challenge with virulent A. hydrophila and A. veronii were also evaluated. Findings of this study showed that the lymphocytes, monocytes, granulocytes counts and antibody (IgM) titre of brood fishes, larvae and eggs from the vaccinated fishes were found significantly higher (p< 0.05) compared to the un-vaccinated control groups. Alternatively, antibody levels (IgM) in the larvae of vaccinated group of brood fishes fed with antigen coated feed was exhibited to be remarkably higher (p< 0.05) than the antigen non-fed group. The RPS of larvae of Shing (91.24 ± 2.00%), Magur (88.09 ± 2.88%) and Pangas (93.17 ± 1.52%) was found to be higher in the larvae at 20-day age of vaccinated group compared to non-vaccinated brood fishes group. Findings of this study indicated that the active immunization of brood fishes followed by oral immunization of their larvae feeding with antigen coated feed showed synergistic effect in protecting cultured Shing, Magur and Pangas fishes from frequent attack with Aeromonas spp at any age of their lifetime.     

Circadian Phase Dependent Thermal Stimulation of Ovarian Recrudescence in Indian Catfish,Clarias batrachus

Female Clarias batrachus acclimated to long photoperiod (13L:11D), were subjected to 30º ± 1ºC thermopulses of either 6-hour or 12-hour duration at different phases of the LD cycle during the late resting phase (first week of January) of their annual reproductive cycle. Six-hour pulses were given either at 0600 or 1200 or 1800 or 0000. Twelve-hour thermopulses were given at 0600 or 1800. The long photoperiods were started at 0530 and that of ambient at 0630 coinciding with the average timing of sunrise that prevailed during the period of the study. The results indicate that exposure to long photoperiod or constant high temperature induced gonadal growth (GSI) and elevated testosterone and oestradiol levels in plasma. The high temperature was significantly more effective in its action. Further, combination of long photoperiod and high temperature produced the strongest gonadal stimulation as gauged from GSI and the levels of steroid hormones. Interestingly, 30ºC thermopulses of 12-h duration when given at 0600 to fish held under long photoperiod induced gonadal development of comparable magnitude as observed in response to constant high temperature under long photoperiod. Thermopulses (30ºC) of 6-h duration given at 0600 or 1200 also induced significant gonadal recrudescence but of much lesser magnitude. Thermopulses either of 6-h or 12-h duration at 1800 failed to elicit any change in the variables under study. The results of cosinor analysis performed on the responses to 6-h thermopulses also substantiate that there is a rhythm in the sensitivity of C. batrachus to thermopulses. Thus it appears that in this species temperature-induced gonadal recrudescence would occur only following coincidence of high temperature with the thermoinducible phase. The underlying mechanism of this phenomenon might be circadian in nature.     

Unusual hepatic mitochondrial arginase in an Indian air-breathing teleost,Heteropneustes fossilis: Purification and characterization

A functional urea cycle with both cytosolic (ARG I) and mitochondrial (ARG II) arginase activity is present in the liver of an ureogenic air-breathing teleost, Heteropneustes fossilis. Antibodies against mammalian ARG II showed no cross-reactivity with the H. fossilis ARG II. ARG II was purified to homogeneity from H. fossilis liver. Purified ARG II showed a native molecular mass of 96 kDa. SDS–PAGE showed a major band at 48 kDa. The native enzyme, therefore, appears to be a homodimer. The pI value of the enzyme was 7.5. The purified enzyme showed maximum activity at pH 10.5 and 55 °C. The K_m of purified ARG II for l-arginine was 5.25 ± 1.12 mM. l-Ornithine and N^ω-hydroxy-l-arginine showed mixed inhibition with K_i values 2.16 ± 0.08 and 0.02 ± 0.004 mM respectively. Mn^+ 2 and Co^+ 2 were effective activators of arginase activity. Antibody raised against purified H. fossilis ARG II did not cross-react with fish ARG I, and mammalian ARG I and ARG II. Western blot with the antibodies against purified H. fossilis hepatic ARG II showed cross reactivity with a 96 kDa band on native PAGE and a 48 kDa band on SDS–PAGE. The molecular, immunological and kinetic properties suggest uniqueness of the hepatic mitochondrial ARG II in H. fossilis.     

Organization and size class homogeneity of ribosomal RNA genes of catfish Heteropneustes fossilis

The ribosomal RNA genes of catfish Heteropneustes fossilis Bloch were examined by Southern blot analysis of genomic DNA digested with restriction enzymes and probed with labelled catfish ribosomal RNA. The major repeat length is 12 kb and about 300 copies per haploid genome are tandemly arranged. The repeat lengths are homogeneous in size in different tissues and individuals. A restriction site polymorphism exists in some of the repeats.     

Phenol induced changes in the carbohydrate metabolism of the Indian catfish Heteropneustes fossilis

Synopsis The Indian catfish Heteropneustes fossilis when subjected to a high sublethal concentration of 26 mg 1^–1 phenol (0.8 of the 96-h LC50 value) for 3, 6, 12, 48 and 96 h elicited muscle glycogenolysis at all time intervals and hepatic glycogenolysis at 3, 6 and 12 h. Hyperglycemia was observed at 3, 6 and 12 h post exposure to phenol. Blood pyruvate content was increased markedly at 3 h. Hyperlacticemia occurred at 3, 6 and 12 h, but hypolacticemia resulted at 48 and 96 h. The results are discussed in relation to acute stress syndrome.     

Cloning, sequencing and expression of cDNA encoding growth hormone from Indian catfish (Heteropneustes fossilis)

A tissue-specific cDNA library was constructed using polyA⁺ RNA from pituitary glands of the Indian catfishHeteropneustes fossilis (Bloch) and a cDNA clone encoding growth hormone (GH) was isolated. Using polymerase chain reaction (PCR) primers representing the conserved regions of fish GH sequences the 3′ region of catfish GH cDNA (540 bp) was cloned by random amplification of cDNA ends and the clone was used as a probe to isolate recombinant phages carrying the full-length cDNA sequence. The full-length cDNA clone is 1132 bp in length, coding for an open reading frame (ORF) of 603 bp; the reading frame encodes a putative polypeptide of 200 amino acids including the signal sequence of 22 amino acids. The 5′ and 3′ untranslated regions of the cDNA are 58 bp and 456 bp long, respectively. The predicted amino acid sequence ofH. fossils GH shared 98% homology with other catfishes. Mature GH protein was efficiently expressed in bacterial and zebrafish systems using appropriate expression vectors. The successful expression of the cloned GH cDNA of catfish confirms the functional viability of the clone.     

EST-based identification of immune-relevant genes from spleen of Indian catfish, Clarias batrachus (Linnaeus, 1758)

A normalized cDNA library from spleen of Indian catfish, Clarias batrachus, was constructed with a redundancy factor of 2.29. A total of 2045 clones from the library were single-pass sequenced, which generated 1937 high quality ESTs with an average read length of approximately 700 bp. Based on sequence similarities, 65 ESTs were found to be associated with immune functions, which were mainly associated with response to stress, response to chemical stimulus, cellular response to stimulus, response to external stimulus, immune response and regulation of response to stimulus. The immune-relevant gene for CD141, thrombomodulin, has been identified in Teleosts for the first time. Six EST-SSRs and three SNPs were found associated with eight immune-relevant genes. These markers associated with important immune genes would be useful for the identification of trait associated alleles for marker-assisted selection. The identification of the putative immune-related genes provides a meaningful framework to understand the Indian catfish immune system and defense mechanisms.