Antiparasitic potentiality of ethanol and methanol extracts of Azadirachta indica leaf for eggs and copepodid stage of Argulus japonicus: in vitro study.

In the present study, eggs and copepodid stages of Argulus japonicus were treated with ethanol and methanol extract of Azadirachta indica (neem) leaf and its antiparasitic efficacy (AE %) was determined. The experiments were performed in triplicate along with the positive (2% DMSO) and negative (without DMSO and extract) control groups. The reduced cumulative hatching percentage of eggs by 13% (in ethanolic) and 17% (in methanolic) extract of neem leaf at 1.5 g L-1 was obtained during 15-day exposure compared to the control group showing 70-85% eggs hatching. The AE of 100% for ethanolic and 91.66% for methanolic extract against the copepodid stage was found at 1.25 and 1.5 g L-1 respectively in 6 h. The histological analysis of the eggs showed the undifferentiated decaying mass of cells with extensively damaged eggs when treated with ethanolic extract of neem leaf. Further, severe degeneration in the branchial region, digestive tract and eye cells was observed in the copepodids treated with ethanol extract than the methanol extract. The terpenoids a potential antiparasitic compound of ethanolic extract produced more AE than the methanolic extract. Thus, the ethanolic extract of neem leaf can be potentially utilized as a natural parasiticide to disrupt the egg and other life phases of A. japonicus.

Antibiotic resistance of culturable heterotrophic bacteria isolated from shrimp (Penaeus vannamei) aquaculture ponds.

Shrimp aquaculture is one of the fastest growing food-producing avenues, where antibiotics usage has become an issue of great concern due to the development of antimicrobial resistance in bacteria. A total of 2304 bacterial isolates from 192 samples (sediment, water, shrimp, and source water) from Andhra Pradesh, India were screened. Antibiotic resistance of bacterial isolates was highest for oxytetracycline (23.4%) followed by erythromycin (12.7%), co-trimoxazole (10%) ciprofloxacin (9.6%), and chloramphenicol (6%), of which 11.9% isolates were multi-drug resistant. Bacterial isolates from shrimp (26.7%), water (23.9%), and sediment (19.6%) samples exhibited more resistance (p ≤ 0.05) towards oxytetracycline. Higher antibacterial resistance was observed from samples of southern Andhra Pradesh (locations L6 and L7). Gram negative bacteria were more prevalent (64%) and showed significantly (p ≤ 0.01) higher resistance. This study indicated the wider distribution of antibiotic-resistant bacteria in shrimp aquaculture ponds with potential risk to humans and the environment.

Role of temperature and co-infection in mediating the immune response of goldfish.

Aquatic Pathogens are expected to encounter tremendous levels of variation in their environment - both abiotic and biotic. Here we examined the change in innate immune parameters and mortality pattern of Carassius auratus during the interaction of co-infection due to an ectoparasite, Argulus and bacteria Aeromonas hydrophila, along with a temperature gradient. Experimental fish were assigned randomly to six treatment groups (T1-T6). Fish of groups T1, T3 and T5 are assigned for healthy fishes kept at 23, 28 and 33°c temperature and served as control. T2, T4 and T6 groups are the co-infected groups kept at temperature gradient. For the haematological and enzyme parameter analysis, sampling was done at 24 h, 72 h and 168 h post challenge from 4 fish in all experimental groups. A temperature dependent increase in intensity of Argulus was observed in the experimental group. Both in control group and co-infected group a temperature dependent mortality pattern was observed, showing an increased mortality of 60% in T6 and 20% in T5 group. A significant decrease of RBC, Hb, and PCV values was observed in co-infected group when compared with control fish in each of the experimental group. Also a temperature dependent increase in WBC, neutrophil and monocyte value was observed in control fish. Whereas, a significant reduction in WBC, neutrophil and monocyte was observed in co-infected fish exposed to 33 °C during the progression of infection. Furthermore, T4 group showed a significantly higher Nitroblue tetrazolium test, Myelo peroxidase and lyzozyme activity compared to other co-infection group. A significantly increased activity of Superoxide dismutase, Glutathione peroxidise and catalase activity was recorded in control fish exposed to 33 °C (T5) whereas, there was no significant difference observed in the activity of catalase and Glutathione peroxidise in the other control fish (T1 and T3 group). This result implies that increase in temperature not only accelerates the intensity of co-infection but also imbalance the health status of the fish by hampering the immunological and physiological parameters towards more detrimental side.

Dose-dependent co-infection of Argulus sp. and Aeromonas hydrophila in goldfish (Carassius auratus) modulates innate immune response and antioxidative stress enzymes.

Co-infection with parasites and bacteria is of frequent occurrence in aquaculture, leads to growth impedance otherwise mortality in fish depending on the varying degree of a load of primary pathogen either parasite or bacteria. The mechanistic regulation of immune response during co-infection in fish has merely documented. The aim of this study was to determine the impact of co-infection with Aeromonas hydrophila at three exposure doses of Argulus sp. on the innate immune responses and antioxidative stress enzymes of goldfish (Carassius auratus). The experimental fish were randomly distributed into eight treatment groups viz. T1 (control group without Argulus and A. hydrophila infection), T2 (fish exposed to a sub-lethal dose of A. hydrophila), T3 (low Argulus-infested fish), T4 (T3 + sub-lethal dose of A. hydrophila), T5 (moderate Argulus-infested fish), T6 (T5 + sub-lethal dose of A. hydrophila), T7 (high Argulus-infested fish) and T8 (T7+ sub-lethal dose of A. hydrophila) in duplicates. After distributing experimental fish into their respective treatment group, A. hydrophila was injected to T2, T4, T6 and T8. After the bacterial challenge, four fish from each experimental group were randomly sampled on 24, 72, and 168 h and subjected to the hematological, innate immune parameters and enzymatic analysis. In the co-infection group T8, a high degree of enhanced pathogenicity of A. hydrophila was noticed with increased mortalities (84.2%) in comparison to other groups. The current study shows a declining pattern in RBC, PCV and Hb values with the degree of parasite infestation without co-infection groups. Moreover, in the T8 group, exposure of a sub-lethal dose of bacteria resulted in a drastic reduction of the recorded parameters. Furthermore, a decreased value for WBC, monocyte and neutrophil was found in higher parasite group co-infected with a sub-lethal dose of bacteria relative to other co-infected groups during the experimental period. Also, a decrease in innate immune parameters and antioxidative stress enzymes were observed in the T8 group compared to T7 and T2 groups throughout the trial period. These findings indicate that a rise in the dose of Argulus infection improves A. hydrophila colonization in goldfish and contributes to suppression of the innate immune system and increased mortality.

Modulation of innate immune responses and induction of oxidative stress biomarkers in Pangasianodon hypophthalmus following an experimental infection with dactylogyrid monogeneans.

Modulation of innate immune activity and oxidative stress response of Pangasianodon hypophthalmus through experimental infection with (Thaparocleidus sp.) dactylogyrid monogenean was studied. A standard cohabitation method was used to infect healthy experimental fish. After 14 days, dactylogyrid (gill monogenean) infested fish were sampled and categorised into three different infected groups namely (T1) low (<10 mean dactylogyrid per gill arch per fish), (T2) moderate (10-49 mean dactylogyrid per gill arch per fish) and (T3) high (>50 mean dactylogyrid per gill arch per fish) along with a control group T0 (un-infested fish). Serum and tissues (gills and liver) were collected from experimental fish and analyzed for markers of innate immune and oxidative stress, respectively. The results showed that respiratory burst activity, myeloperoxidase level, serum lysozyme, α-2 macroglobulin and total serum immunoglobulin level were significantly (p < 0.05) elevated in fish with different degrees of parasite infestation compared to the control (un-infested group). Similarly, cellular oxidative biomarkers superoxide dismutase, catalase, glutathione-S-transferase and Na+-K+-ATPase activities of gills and liver were significantly (p < 0.05) elevated in dactylogyrid infested fish in comparison to the control. However, significantly decreased level of albumin, albumin to globulin ratio, total serum antiprotease and ceruloplasmin were observed in fish infested with low degree of dactylogyrids, while no significant differences in these parameters were observed between moderately infested and the control groups. The results suggested that varying degree of gill monogenean dactylogyrid infestation not only modulated the innate immune response of P. hypophthalmus by lowering albumin, total serum antiprotease and ceruloplasmin and inducing respiratory burst activity, phagocytic activity, myeloperoxidase, lysozyme, α-2 macroglobulin and total immunoglobulins, but also the oxidative stress biomarkers. The baseline data obtained in the present study will be valuable in understanding the host-parasite relationship and the dynamics of innate, oxidative stress responses and susceptibility of P. hypophthalmus to different degrees of parasitosis.

Effect of orally administered azadirachtin on non-specific immune parameters of goldfish Carassius auratus (Linn. 1758) and resistance against Aeromonas hydrophila.

Modulation of the immune responses using active bio-ingredients as a possible prophylaxis measure has been novel prospect for aquaculture. The present study evaluated the effects of azadirachtin EC 25% on non-specific immune responses in goldfish Carassius auratus and resistance against pathogenic bacteria Aeromonas hydrophila. The experimental trial for effects of azadirachtin on immuno-haematoloical parameters in goldfish was conducted by feeding the various levels of azadirachtin as control T(0) (without azadirachtin), T(1) (0.1%), T(2) (0.2%), T(3) (0.4%), T(4) (0.8%) and T(5) (1.6%) for a period of 28 days. Fishes were challenged with A. hydrophila 28 days post feeding and relative percentage survival (%) was recorded over 14 days post infection. Immuno-haematoloical (total erythrocyte count, total leukocyte count, haemoglobin, packed cell volume, NBT activity, phagocytic activity, serum lysozyme activity, myeloperoxidase activity, total immunoglobulin) and serum biochemical parameters (serum glutamate oxaloacetate transaminase (SGOT), serum glutamate pyruvate transaminase (SGPT) and blood glucose) of fishes were examined at 14 and 28 days of feedings. Fish fed with azadirachtin, showed significantly (p < 0.05) enhanced TEC, TLC, Total Ig, total protein, NBT activity, serum lysozyme activity and myeloperoxidase level in different treatment groups in comparison with control group. Similarly, SGOT, SGPT and blood glucose level were found to be significantly (p < 0.05) high but PCV and Hb did not differ significantly (p > 0.05) in the treatment groups compared to control groups. Azadirachtin at the concentration of 4 g kg(-1) showed significantly (p < 0.05) higher relative percentage survival (42.60%) when compared with the control against A. hydrophila infection. This study indicated that azadirachtin EC 25% (4 g kg(-1)) showed higher NBT activity, serum lysozyme, protein profiles, leukocyte counts and resistance against A. hydrophila infection and thus, can be used as a potential immunostimulant in aquaculture.

Effect of azadirachtin on haematological and biochemical parameters of Argulus-infested goldfish Carassius auratus (Linn. 1758).

Argulosis hampers aquaculture production and alters the host physiology and growth. Azadirachtin is recognized as a potential antiparasitic agent against Argulus sp. The present study aimed to investigate the effect of different concentration of azadirachtin solution on haematological and serum biochemical parameters of Argulus-infested goldfish Carassius auratus. Ninety Argulus-infested goldfish were randomly divided into six equal groups. Fish of group 1-5 were treated with azadirachtin solution through bath of 1, 5, 10, 15 and 20 mg L(-1) as T1, T2, T3, T4 and T5, respectively, and group 6 was exposed to 2% DMSO solution without azadirachtin and considered as negative control T0(-). Along with six treatment groups, a positive control T0(+) of healthy goldfish free from Argulus infestation was also maintained. Parasitic mortality was evaluated after 3 days of consecutive bath treatment. After 7 days of post-treatment, the blood and serum were drawn from each of the treatment groups and haematological and serum biochemical parameters were evaluated. Total leucocyte count (TLC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), blood glucose, total protein (TP), globulin, serum glutamate oxaloacetate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) were significantly (p < 0.05) high in negative control group when compared with positive control group. It could be concluded that Argulus infestation altered marked haematological and serum biochemical parameters. However, in treated groups complete elimination of Argulus was found in T4 and T5 groups. Also significant (p < 0.05) reduction in haematological and serum biochemical parameters of all the treatment groups were recorded in comparison with negative control group. In addition, T4 and T5 groups showed significantly (p < 0.05) high superoxide dismutase (SOD), catalase, total erythrocyte count (TEC) and haemoglobin (Hb). However, higher mean corpuscular haemoglobin concentration (MCHC), blood glucose and lactate dehydrogenase (LDH) levels in T5 group revealed that higher concentration of azadirachtin have notable effects on activity of vital tissues function and physiology of the host. Argulus spp. from infested goldfish could be eliminated using bath treatment with solution of azadirachtin having concentration of 15 mg L(-1) and that also shifted haematological and serum biochemical parameters towards homeostasis.

Antiparasitic efficacy of piperine against Argulus spp. on Carassius auratus (Linn. 1758): in vitro and in vivo study.

Argulus are common aquatic ectoparasites that create one of the major threats to aquaculture due to absence of suitable therapy. Piperine, a bioactive component of Piper longum, has medicinal properties and acts as anti-inflammatory, antibacterial, and antifungal, considering eco-friendliness and cost-effectiveness. The present study aimed to evaluate antiparasitic effect of piperine against an ectoparasite Argulus spp. on Carassius auratus. Artificial Argulus infection was carried out by cohabitation method, and the fishes were selected for in vivo study when intensity of Argulus infestation was observed to be 15-20 Argulus per fish. In vitro and in vivo studies were performed at different concentration 1.0 (T (1)), 3.0 (T (2)), 5.0 (T (3)), 7.0 (T (4)), and 9.0 mg l(-1) (T (5)) of piperine solution to treat Argulus for 3 and 72 h, respectively. The acute toxicity test for piperine EC 97 % against goldfish was performed for 96 h. The 96 h median lethal concentration (LC(50)) for piperine was found to be 52.64 mg l(-1). In vitro effect of piperine solution led to 100 % mortality of Argulus at 9.0 mg l(-1) in 3 h whereas, under in vivo test, the 100 % antiparasitic efficacy of piperine solution was found at 9.0 mg l(-1) in 48 h. The EC(50) for 48 h was 9.0 mg l(-1), and thus, therapeutic index is 5.8. The results revealed that piperine at a concentration of 9.0 mg l(-1) can be used as a potential natural agent for controlling Argulus parasite.

In vitro and in vivo antiparasitic activity of Azadirachtin against Argulus spp. in Carassius auratus (Linn. 1758).

Argulus is one of the most common and predominant ectoparasites which cause serious parasitic disease and is a potent carrier of viruses and bacteria in the ornamental fish industry. In recent years, organic (herbs)-based medicines are widely used to cure the disease, and neem (Sarbaroganibarini) medicine is very popular and effective throughout the world. The present study was conducted to find the effects of Azadirachtin against Argulus spp. on Carassius auratus under in vitro and in vivo conditions. The 96-h median lethal concentration (LC(50)) for Azadirachtin EC 25% against Carassius auratus was found to be 82.115 mg L(-1). The antiparasitic activity test under in vitro and in vivo was evaluated at 1 (T1), 5 (T2), 10 (T3), 15 (T4) and 20 mg L(-1) (T5) to treat Argulus for 3 h and 72 h, respectively. In vitro effect of Azadirachtin solution led to 100% mortality of Argulus at 20 and 15 mg L(-1) for 2.5 and 3 h, respectively. Whereas, under in vivo test, the 100% antiparasitic efficacy of Azadirachtin solution was found at 15 and 20 mg L(-1) for 72 and 48 h, respectively. The EC(50) for 48 h was 20 mg L(-1), and thus, therapeutic index is 4.10. The results provided evidence that Azadirachtin can be used as a potential agent for controlling Argulus.

Is bipolar II disorder misdiagnosed as major depressive disorder in children?

OBJECTIVES: To estimate the lifetime prevalence of bipolar II disorder in children and adolescents presenting with DSM-IV major depressive disorder (MDD). METHODS: Sixty-one consecutive subjects aged < or =18 years attending the outpatient services of the Child and Adolescent Psychiatric (CAP) services of the National Institute of Mental Health and Neurosciences (NIMHANS), Bangalore, India with a diagnosis of MDD were evaluated using the Missouri Assessment of Genetics Interview for children (MAGIC). Two psychiatrists, one of whom was a child psychiatrist diagnosed hypomania by consensus. RESULTS: Twelve children had a past episode of hypomania (20%), which was hitherto undiagnosed clinically. LIMITATIONS: We recruited subjects from a psychiatric hospital, thus limiting the generalizability of the finding. Sample size was relatively small and assessments were cross-sectional. CONCLUSIONS: : Our study shows that bipolar II disorder is often misdiagnosed as MDD in children. The study also highlights that the chance of diagnosing bipolarity is enhanced by using semi-structured interview in routine clinical practice.

A customised portable LogMAR chart with adjustable chart illumination for use as a mass screening device in the rural population.

AIM: To develop a customised, portable, cost-effective (logarithmic minimal angle resolution) LogMAR chart with adjustable illumination for use as a mass vision-screening device in the rural population. MATERIALS AND METHODS: Visual acuity of 100 individuals was evaluated with a customised chart and compared with the standard Early Treatment Diabetic Retinopathy Study (ETDRS) chart and Snellen's Chart. Bland and Altman analytical techniques were used for analysis. RESULTS: Test-retest variability of the customised chart was just a one-line difference (95% CI for agreement), and so were the results with the standard ETDRS charts; a variability of 3-line was noted with Snellen's chart. Two-line differences were observed when comparison was made with Standard ETDRS chart and 2 to 3-line differences with Snellen's chart. CONCLUSION: The customised portable LogMAR chart with adjustable illumination shows less test-retest variability and better agreement with standard ETDRS chart; therefore, it can be used as a mass vision-screening device in rural settings.