Impact of graded doses of enrofloxacin on the safety and biological responses of Nile tilapia Oreochromis niloticus.

The cultivation of tilapias, the third most farmed fish group globally, has been rapidly growing, especially in Southeast Asia. This surge in tilapia farming intensification has led to increased use of antibiotics to control bacterial diseases. This study investigated the safety implications of administering graded doses of enrofloxacin (ENF) at 0 (control), 10, 30, 50 and 100 mg/kg biomass/day orally to Oreochromis niloticus. The 43-day study comprised 7 days of pre-dosing, 15 days of ENF-dosing, and a 21-day recovery period with a periodical assessment of the biological responses of fish. The results revealed that the overdosed groups experienced up to 21% reduction in feed consumption, 11% mortalities, and adverse impacts on hematology, including a decrease in erythrocytes, and monocytes and an increase in leukocytes, thrombocytes, lymphocytes, and neutrophils. Haematological indices like mean corpuscular volume and mean corpuscular hemoglobin decreased, while mean corpuscular hemoglobin concentration increased. The plasma biochemical parameters including glucose and liver and kidney enzymes unveiled a significant dose- and time-dependent increase, while calcium and chloride levels decreased. Erythrocytes displayed several erythrocyte cellular and nuclear abnormalities. The frequency of micronucleus increased with dose and time, suggesting potential genotoxicity of ENF. Additionally, a dose-dependent increase in residues in the tissues with the highest accumulation in muscle was documented. Nevertheless, the recovery of the measured parameters upon dose termination indicated that the ENF-induced alterations are reversible. The study affirmed the safety of ENF at the recommended dose (10 mg) in O. niloticus and their adoptive responses to higher doses.

Safety, tolerability and biological responses of Oreochromis niloticus juveniles upon oral oxolinic acid administration.

In aquaculture, oxolinic acid (OA) is used as a second-line treatment at 12 mg/kg biomass/day for seven consecutive days. The present study evaluated the biosafety of 21 days of dietary administration of OA at 0, 12, 36, 60 and 120 mg by assessing the growth, biochemical, erythrocytic morphological and histopathological alterations and residue levels in Oreochromis niloticus. A significant dose-dependent reduction in feed intake and biomass and an increase in mortalities and erythrocytic cellular and nuclear changes were recorded. Significant elevations in plasma glucose, creatinine, alkaline phosphatase, alanine transaminase and aspartate transaminase and a decline in calcium and chloride levels were documented. The kidney, liver and intestine histoarchitecture showed mild to marked alterations. The edible tissue OA residues peaked on day 21 and decreased upon cessation of administration in all the dosing groups. The residue levels in the muscle of the recommended dose group were well within the maximum residue limit set by the European Medicines Evaluation Agency. Although the current study hinted at the safety and tolerability of OA even during long-term usage in O. niloticus in Indian conditions, care must be exercised for its aquacultural application because of its listing as a critically important medicine for humans.

Assessment of Single-Dose Pharmacokinetics of Oxolinic Acid in Rainbow Trout and Determination of In Vitro Antibacterial Activity Against Pathogenic Bacteria From Diseased Fish.

In response to the heightened risk of bacterial diseases in fish farms caused by increased demand for fish consumption and subsequent overcrowding, researchers are currently investigating the efficacy and residue management of oxolinic acid (OA) as a treatment for bacterial infections in fish. This research is crucial for gaining a comprehensive understanding of the pharmacokinetics of OA. The present study investigates pharmacokinetics of OA in juvenile rainbow trout. The fish were given a 12 mg kg-1 dose of OA through their feed, and tissue samples were collected of the liver, kidney, gill, intestine, muscle, and plasma for analysis using LC-MS/MS. The highest concentrations of the drug were found in the gill (4096.55 µg kg-1) and intestine (11592.98 µg kg-1), with significant absorption also seen in the liver (0.36 L/h) and gill (0.07 L/h) (p < 0.05). The liver (0.21 L/h) and kidney (0.03 L/h) were found to be the most efficient (p < 0.05) at eliminating the drug. The study also confirmed the drug antimicrobial effectiveness against several bacterial pathogens, including Shewanella xiamenensis (0.25 µg mL-1), Lactococcus garvieae (1 µg mL-1), and Chryseobacterium aquaticum (4 µg mL-1). The study concludes significant variations among different fish tissues, with higher concentrations and longer half-lives observed in the kidney and intestine. The lowest MIC value recorded against major bacterial pathogens demonstrated its therapeutic potential in aquaculture. It also emphasizes the importance of understanding OA pharmacokinetics to optimize antimicrobial therapy in aquaculture.

Intestinal Histopathological Aberrations in Oreochromis niloticus Juveniles upon Dietary Florfenicol Administration.

The aquaculture use of antibiotics can cause detrimental effects on fish organs and gut microbial dysbiosis. The impact of florfenicol (FFC) on fish intestinal histology, an approved antibiotic, remains unclear. This study aimed to investigate the effects of FFC on Oreochromis niloticus juveniles by administering FFC at 10 mg and 30 mg/kg biomass/day for 30 consecutive days to mimic long-term use. A dose-dependent reduction in feed intake, survival and biomass, with an upsurge in mortalities was observed. Even the therapeutic dose instigated mortalities on day 30 of FFC dosing (FD). Histopathological analysis revealed mild to moderate alterations, including loss of absorptive regions, epithelial degeneration, necrotized areas, intercellular enterocytic space and swollen laminar propria. Post-dosing, the observation of the detachment of lamina propria from the epithelium indicated imminent irritability. Goblet cells reduced drastically on day 30 FD, accompanied by an increase in intraepithelial lymphocytes. However, cessation of dosing for 13 days resulted in the reclamation of goblet cells and absorptive regions, indicating that the intestinal tissues underwent considerable repair after lifting antibiotic pressure. These findings suggested that O. niloticus can tolerate dietary FFC but emphasize the need for responsible use of antibiotics in aquaculture.

Development of microbial enrichments for simultaneous removal of sulfur and nitrogenous metabolites in saline water aquaculture.

AIM: The aim of the study was to develop microbial enrichments from the nitrifying microbial consortia and the environment for simultaneous removal of ammonia, nitrate, and sulfide in aquaculture systems at varied salinities. METHODS AND RESULTS: Sulfur and nitrogen metabolites are the major factors affecting the farmed aquatic animal species and deteriorate the receiving environments causing ecological damage. The present study reports the development of microbial enrichments from the nitrifying microbial consortia and the environment. The enrichments used thiosulfate or thiocyanate as an energy source and simultaneously removed sulfur, ammonia, and nitrite in spiked medium (125 mg/l ammonia; 145 mg/l nitrite). Further, the microbes in the enrichments could grow up to 30 g/l salinity. Metagenomic studies revealed limited microbial diversity suggesting the enrichment of highly specialized taxa, and co-occurrence network analysis showed the formation of three micro-niches with multiple interactions at different taxonomic levels. CONCLUSIONS: The ability of the enrichments to grow in both organic and inorganic medium and simultaneous removal of sulfide, ammonia, and nitrite under varied salinities suggests their potential application in sulfur, nitrogen, and organic matter-rich aquaculture pond environments and other industrial effluents.

In-feed emamectin benzoate abuse affects the biological responses and erythrocytes of Nile tilapia Oreochromis niloticus.

In aquaculture, drugs are often abused to accomplish disease control without considering the negative effects on fish health. This study aimed at elucidating the pernicious effects of in-feed antiparasitic drug emamectin benzoate (EB) abuse on the haemato-biochemistry and erythro-morphometry of healthy Nile tilapia Oreochromis niloticus. The fish were fed EB at 50 µg (1×) and 150 µg/kg biomass/d (3×) for 14 d as against the recommended 7 d and periodically assessed the blood parameters. A significant dose- and time-dependent reduction in feed intake, survival, total erythrocytes (TEC), monocytes (MC), hemoglobin (Hb), hematocrit (Ht) and mean corpuscular Hb concentration were noted. The total leukocytes (TLC), thrombocytes (TC), lymphocytes (LC) and neutrophils (NC) markedly augmented. The EB-dosing altered the fish physiology by enhancing the glucose, alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and creatinine and reducing the calcium, chloride and acetylcholinesterase (AChE) levels dose-dependently. The fish recovered within 4 weeks in the 1× group post-dosing but persevered in the overdosed group. The erythro-cellular and nuclear dimensions were reduced with the increase in dose and normalized after the cessation of dosing, except for nuclear volume. The erythro-morphological alterations were more prominent in the overdosed group. The results implied the pernicious effect of oral EB medication on the biological responses of fish if abused.

16 S rRNA gene diversity and gut microbial composition of the Indian white shrimp (Penaeus indicus).

The endemic Indian white shrimp (Penaeus indicus) is an economically important crustacean species, distributed in the Indo-West Pacific region. Knowledge of its gut microbial composition helps in dietary interventions to ensure improved health and production. Here we analyzed V3-V4 hypervariable regions of the 16 S rRNA gene to examine intestinal microbiota in wild and domesticated farmed P. indicus. The study revealed that Proteobacteria, Fusobacteria, Tenericutes, and Bacteroidetes, were the dominant phyla in both the groups although there were differences in relative abundance. The dominant genera in case of the wild group were Photobacterium (29.5 %) followed by Propionigenium (13.9 %), Hypnocyclicus (13.7 %) and Vibrio (11.1 %); while Vibrio (46.5 %), Catenococcus (14 %), Propionigenium (10.3 %) and Photobacterium (8.7 %) were dominant in the farmed group. The results of the study suggest the role of environment on the relative abundance of gut bacteria. This is the first report characterizing gut microbial diversity in P. indicus, which can be used to understand the role of gut microbiota in health, nutrition, reproduction, and growth.

Influence of Fluctuating Water Temperature and Dietary Oxytetracycline on the Safety of Monosex Nile Tilapia Oreochromis niloticus Fries.

The influence of fluctuating water temperature and dietary oxytetracycline (OTC) at 0 (0X), 80 (1X), 240 (3X), 400 (5X) and 800 mg (10X)/kg biomass/day for 30 consecutive days on the safety of monosex (all male) Nile tilapia Oreochromis niloticus fries in terms of feeding, growth, survival and histopathology of vital organs were assessed. A dose-dependent decline in feed intake and biomass was recorded. The OTC-dosed groups recorded higher mortalities than the control. The therapeutic OTC-dosing (1X) in conjunction with low temperature caused 75.56 ± 8.01% mortality and 25.75% reduced feed intake in 30 days. The mortalities increased with increasing OTC-doses from 85.19 ± 3.39% (1X) to 95.56 ± 2.22% (10X) and fluctuating temperature (12.00-21.50°C) even after the withdrawal of OTC. Relatively mild to moderate histopathological lesions were observed in the kidney, liver and intestine of OTC-dosed fries. These results suggested that dietary OTC and low water temperature may cause adverse effects on monosex O. niloticus fries.

Vibrio harveyi biofilm as immunostimulant candidate for high-health pacific white shrimp, Penaeus vannamei farming.

The study was to develop Vibrio harveyi biofilm-based novel microbial product and its oral delivery for high health Penaeus vannamei farming. Yield of bacterial biofilm was optimized on chitin substrate (size: <360, 360-850 and 850-1250 µm; concentration: 0.3, 0.6 and 0.9%) in tryptone soy broth (0.15%). The biofilm was characterized by crystal violet assay, SEM and LSCM imaging; protein profiling by SDS-PAGE and LC-ESI-MS/MS. The immune stimulatory effect of the biofilm in yard experiments was evaluated by relative quantification of immune genes using real-time PCR effect on overall improvement on health status under field trials. The highest biofilm yield (6.13 ±â€¯0.2 × 107 cfu/ml) was obtained at 0.6% of <360 µm chitin substrate. The biofilm formation was stabilized by 96 h of incubation at 30 °C. Protein profiling confirmed expression of six additional proteins (SDS-PAGE) and 11 proteins were differentially expressed (LC-ESI-MS/MS) in biofilm cells over free cells of V. harveyi. Oral administration of the biofilm for 48 h confirmed to enhance expression of antimicrobial peptides, penaeidin, crustin and lysozyme in P. vannamei. Further Oral administration of biofilm for two weeks to P. vannamei (1.8 ±â€¯0.13 g) improved the growth (2.66 ±â€¯0.06 g) and survival (84.44 ±â€¯1.82%) compared to control (2.15 ±â€¯0.03 g; 70.94 ±â€¯0.66%) Nursery trials showed a significant reduction in occurrence of anatomical deformities like antenna cut (12.67 ±â€¯0.66%), rostrum cut (4.66 ±â€¯0.87%), and tail rot (3.33 ±â€¯0.88%), compared to animals fed with normal diet which was 24.33 ±â€¯2.72; 14 ±â€¯1.52 and 10.66 ±â€¯1.45% respectively. In vitro and in vivo studies suggest inactivated biofilm cells of V. harveyi on chitin substrate express additional antigenic proteins and when administered orally through feed at regular intervals stimulates immune response and improve growth, survival and health status of shrimp.

Dietary influences of oxytetracycline on the growth and serum biomarkers of Oreochromis niloticus (L.).

Tilapias are cultured globally and are rising in acceptance as the most important freshwater aquaculture species. Monitoring of serum biomarkers is a promising tool in aquaculture to screen the health status as they are virtuous indicators of extreme stress and organ dysfunction in fish. The present study examined the serum biomarkers of oxytetracycline (OTC)-dosed Nile tilapia Oreochromis niloticus at 0, 80 and 800 mg/kg biomass/day, i.e., 0X, 1X, and 10X the approved dose (X = 80 mg OTC/kg biomass/day) for 10 consecutive days. The fish biomass and levels of serum glucose, aspartate aminotransferase, alanine aminotransferase, creatinine and C-reactive protein (CRP) were determined at scheduled intervals. A significant dose-dependent reduction in fish biomass during the OTC-dosing (5.84%) and post-OTC dosing (8.16%) periods was observed. All the serum biomarkers of Nile tilapia increased significantly on day 10 OTC-dosing. Though their levels reduced significantly, normalcy was not achieved even after 42 days of cessation of OTC-dosing, except CRP. The CRP reached the normal level on day 25 post-OTC dosing in the 1X group. The results, thus, demonstrated that the oral OTC-dosing influences the physiological state of apparently healthy Nile tilapia in a dose-dependent manner. These changes were, however, reversible upon discontinuation of OTC-dosing. The set of data observed on growth reduction and elevated serum biomarker levels even after 42 days of cessation of OTC-dosing, thus, raises questions on the utility of oral OTC-dosing.

Effect of culture intensity and probiotics application on microbiological and environmental parameters in Litopenaeus vannamei culture ponds.

The present study examined the complex interaction among stocking density and extent of probiotic use with production and environmental parameters in Litopenaeus vannamei culture ponds to suggest suitable management strategies. The study was conducted inL. vannamei culture ponds with stocking density of 35 nos sq m(-1) (Group I) and 56 nos sq m(-1) (Group II) and probiotic application @16.5 kg ha(-1) and 157 kg ha(-1), respectively. There was no significant difference noted between the two groups of ponds in respect to ammonia oxidizing bacteria (AOB) in sediment and nitrite oxidizing bacteria (NOB) in water samples, whereas significantly higher levels of AOB in water samples of high intensity culture ponds (Group II) and NOB in sediment samples of Group I were observed. The levels of sulphur oxidizing bacteria (SOB) and sulphur reducing bacteria (SRB) in Group I pond water and in Group II sediment were significantly higher than their corresponding levels in the other group. In both the groups, ammonia, nitrite and sulphide concentrations were below toxic limits prescribed for shrimp farming. Comparing the production parameters at harvest revealed that low intensity culture ponds (Group I) had higher growth rate, average body weight and significantly lower FCR and higher survival rate than high intensity culture ponds (Group II). The results indicated that application of microbial products in higher quantities did not benefit significantly, and there is a need to regulate quantum and schedule of biological product usage for economically sustainable shrimp culture.

Pharmacokinetics and residues of florfenicol in Nile tilapia (Oreochromis niloticus) post-oral gavage.

In the study on Oreochromis niloticus, singular oral gavage of florfenicol (FFC) at 15 mg/kg biomass/day was conducted, mimicking approved aquaculture dosing. Samples of plasma, bile, muscle, intestine, skin, liver, kidney, gill, and brain tissues were collected at 0, 2, 3, 4, 6, 8, 12, 16, 24, 32, 48, 64, 96, and 128 hours (h) after oral gavage. LC-MS/MS analysis revealed FFC concentrations peaked at 12.15 µg/mL in plasma and 77.92 µg/mL in bile, both at 24 hours. Elimination half-lives were 28.17 h (plasma) and 26.88 h (bile). The residues of FFC ranked muscle>intestine>skin>liver>kidney>gill. In contrast, the residues of florfenicol amine (FFA) ranked kidney>skin>liver>muscle>gill>intestine>brain, particularly notable in tropical summer conditions. The minimum inhibitory concentration of FFC was elucidated against several bacterial pathogens revealing its superior efficacy. Results highlight bile's crucial role in FFC elimination. Further investigation, especially during winter when fish susceptibility to infections rises, is warranted.

Assessing the effect of therapeutic level of oxytetracycline dihydrate on pharmacokinetics and biosafety in Oncorhynchus mykiss (Walbaum, 1792).

The aim of the experiment was to investigate the pharmacokinetics of oxytetracycline dihydrate after a single oral administration of 80 mg kg-1 day-1 in rainbow trout and assess its biosafety at concentration of 80, 240, 400, and 800 mg kg-1 day-1 over 30 days, focusing on various aspects such as effective feed consumption, physiological responses, drug tolerance, and detection of low drug concentrations in rainbow trout. The pharmacokinetics study spanned a duration of 5 days, while the assessment of biosafety extended for a 30-day safety margin, followed by a subsequent 10-day residual analysis. Pharmacokinetic analysis revealed slow absorption with low-rate constant in tissues. Absorption rates vary among tissues, with the gill showing the highest rate (0.011 h-1) and plasma exhibiting the slowest (0.0002 h-1). According to pharmacokinetic analysis, the highest concentration, Cmax (µg kg-1) was observed in the kidney (9380 µg kg-1) and gill (8710 µg kg-1), and lowest in muscle (2460 µg kg-1). The time (Tmax) to reach peak concentration (Cmax) varied among tissues, ranging from 3 h in the gill to 32 h in the muscle, with 24 h in plasma, 32 h in the kidney, and 16 h in both the liver and skin. The liver and kidney had the highest area under the concentration-time curve (AUC(0-128)), indicating widespread drug distribution. Prolonged elimination occurred at varying rates across tissues, with the gill showing the highest rate. The study found that OTC concentrations exceeded the LOD and LOQ values. Biosafety evaluation showed effective feed consumption, physiological responses, and low drug concentrations in muscle at the recommended dosage of 80 mg kg-1 fish day-1.

Assessing safety, efficacy and residue depletion in golden mahseer, Tor putitora (Hamilton, 1822): biochemical and physiological responses to graded concentrations of oxytetracycline dietary supplementation.

The safety and effectiveness of oxytetracycline can potentially manage bacterial infections in fish. This, in turn, might reduce the concerns related to its use in aquaculture and human consumption, such as toxicity, antimicrobial resistance, and other associated risks. The primary objective of this study was to assess how adding oxytetracycline dihydrate to the diet affects its effectiveness, safety, and the presence of residues in T. putitora. T. putitora fingerlings, subjected to experimental infection with Aeromonas hydrophila at a concentration of 108 CFU mL- 1, received an oral administration of oxytetracycline dihydrate. The oxytetracycline dihydrate was added to the feed (corresponding to 2% of the fish body weight) at concentrations of 44.1, 88.2, 132.3 and 176.4 mg Kg- 1 fish body weight per day. This treatment was carried out for 10 consecutive days. The biochemical and physiological responses of T. putitora and efficacy of oxytetracycline dihydrate were determined through estimation of microbial load (CFU mL- 1), haematogram, serum biomarkers, behavioral characteristics, non-specific immunity and residue depletion. Experimentally infected fish showed disease progression and induced histopathological conditions with highest microbial load (CFU mL- 1) in the muscle of both control and treated fish. The fish haematogram showed increased leucocyte and haemoglobin content, influenced by dietary oxytetracycline dihydrate. The fish demonstrated adaptive physiological response to oxytetracycline dihydrate at 44.1 to 88.2 mg and resulted in increased albumin and globulin content. The serum-enzyme assay showed significant increase in aspartate aminotransferase (AST), alanine aminotransferase (ALT) and plasma alkaline phosphatase (ALP) activities in the test fish (< 0.05). Oxytetracycline dihydrate at 88.2 to 132.3 mg Kg- 1 fish body weight per day recorded higher feed intake (75%), significant survivability (66-68%) and histopathological recovery. The suppressed immune response was manifested with decreased respiratory burst and lysozyme activity. The palatability, treatment of bacterial infection, histopathological changes and survivability by fingerlings of golden mahseer determined the safety and optimized the therapeutic potential of the oxytetracycline dihydrate at 88.2 mg Kg- 1 fish body weight per day for 10 days to contain the infection by A. hydrophila. A withdrawal period of 8-d was recommended as oxytetracycline dihydrate concentration depleted below the legal maximum residue limit (MRL 2.0 mg g- 1) in the edible muscle of the golden mahseer reared at an average water temperature of 20 °C. This is considered safe for human consumption.

Impacts of Oral Florfenicol Medication and Residues on the Kidney and Liver of Nile Tilapia Oreochromis niloticus (L.)

Florfenicol (FFC), an approved aquaculture antibiotic, is administered in feed at doses of 10-15 mg kg biomass-1 day-1 for 10 successive days. In this study, healthy Oreochromis niloticus were fed with 0-10 times the therapeutic dose of 15 mg kg biomass-1 day-1 for 10 days and tracked for 43 days post dosing. Assessments of residue accrual and depletion, oxidative stress, serum biochemistry, histopathology and extent of kidney and liver damages were made. FFC dosing reduced the feed intake significantly. The therapeutic dose produced no mortalities on day 10. Dose-dependent alterations in serum biochemistry were noted upon dosing. Several histopathological alterations were observed in the kidney and liver, which vindicated the toxic potentials of FFC. The residual FFC and florfenicol amine (FFA) accrual, depletion and oxidative stress responses, such as increased malondialdehyde, total nitric oxide, ferric reducing antioxidant power and reduced glutathione S-transferase activity, were documented. The dietary FFC persuaded the physiological state of O. niloticus, the effects of which normalized sparsely with time upon cessation of dosing at the higher doses. The study provided a brief outlook on the physiological responses upon oral FFC administration, which should be kept in mind during its application for fish health safety purposes.

Pharmacokinetics and biosafety evaluation of a veterinary drug florfenicol in rainbow trout, Oncorhynchus mykiss (Walbaum 1792) as a model cultivable fish species in temperate water.

In two experimental trials; florfenicol pharmacokinetics following a single dose oral administration at 15 mg kg-1 fish body weight and biosafety through extended medicated feeding were studied in the rainbow trout, Oncorhynchus mykiss. The pharmacokinetic trial was conducted for 5 days, whereas the biosafety experiment lasted for a 30-day safety margin followed by a 20-day residual period analysis at 3, 5 and 10 times greater than the therapeutic dose 10 mg kg-1 biomass day-1. C max µg kg-1 calculated for florfenicol were found to be 5,360 in intestine, 2,890 in gill, 2,250 in kidney, 973 in liver and 273 in plasma, obtained at T max of 16 h. Intestine had utmost area under the concentration-time curve (tissue/plasma) of 13.83 h µg kg-1 and a prolonged half life (t1/2ß) of 28.62 h. The highest apparent metabolic rate value in the kidney (0.327) showed a high level of biotransformation of florfenicol to its metabolite florfenicol amine. The apparent distribution rate of florfenicol amine in muscle, in comparison to the parent drug florfenicol, indicated elimination of the medication mostly in the form of florfenicol amine with t1/2 of 16.75 h. The biosafety of florfenicol orally administered to rainbow trout recorded effective feed consumption, physiological responses, drug tolerance and significantly low drug concentrations in muscle of rainbow trout, thus its usage at 10 mg kg-1 fish body weight is recommended. In the study, the rapid absorption, greater bioavailability, enhanced dispersion, slower elimination and biosafety of the drug form a significant basis for the florfenicol and its metabolite florfenicol amine as a useful antibacterial agent in aquaculture.

Biosafety, histological alterations and residue depletion of feed administered anti-parasitic drug emamectin benzoate in golden mahseer, Tor putitora (Hamilton, 1822) as a model candidate fish for sport fishery and conservation in temperate waters.

In the present experiment, the attempt has been made to study the biosafety, toxicity, residue depletion and drug tolerance of graded doses of emamectin benzoate (EB) in juveniles of golden mahseer, Tor putitora as a model candidate fish for sport fishery and conservation in temperate waters through an extended medicated feeding. The graded doses of EB viz., 1× (50 µg/kg fish/day), 2 × (100 µg/kg fish/day), 5 × (250 µg/kg fish/day) and 10 × (500 µg/kg fish/day) were administered to golden mahseer juveniles through medicated diet for 21 days at water temperature of 18.6°C. The higher doses of EB did not cause any mortality during and 30 days after the end of medication period, but considerable variations in feeding and behavior were observed. Severe histological alterations observed after EB-diets (5 × and 10×) were vacuolation, pyknotic nuclei, melanomacrophage centre and necrosis in liver; Bowman's capsule dilation, degenerated renal tubules in kidney; myofibril disintegration, muscle oedema, splitting of muscle fibres, migration of inflammatory cells in muscle; and abundant goblet cells, dilated lamina propria and disarrangement of mucosa in intestine tissues. The residual concentrations of EB metabolites Emamectin B1a and B1b were analyzed using muscle extracts and were found to be peaked during medication period followed by gradual depletion in post-medication period. The outcome of this study showed that the Emamectin B1a residual concentration in fish muscle in 1×, 2×, 5×, and 10× EB treatment groups were 1.41 ± 0.49, 1.2 ± 0.7, 9.7 ± 3.3, and 37.4 ± 8.2 µg/kg at 30 days of post-medication period, respectively, which falls under the maximum residue limits (MRLs) of 100 µg/kg. The results support the biosafety of EB at recommended dose of 50 µg/kg fish/day for 7 days. As residue of EB is recorded falling within the MRL, no withdrawal period is recommended for golden mahseer.

Natural infestation of an anchor worm, Lernaea sp. in cage culture of Asian Seabass, Lates calcarifer juveniles and its control using an anti-parasitic drug, emamectin benzoate.

Parasitic infestations and their control programmes are one among the challenges to be considered the most significant in aquaculture. A parasitic infestation was studied elaborately in Asian Seabass, Lates calcarifer juveniles with clinical signs, post-mortem findings, morphological and molecular identifications. In addition, those fish were also treated with emamectin benzoate (EMB) @ 50 µg kg-1 of fish body weight (BW) d-1 for 10 consecutive days under the controlled wet lab facility by feeding through the medicated feed at 4% BW. Results showed that the parasitic prevalence, parasitic intensity (PI) and mortality were 45.5%, 8.17 ± 0.15 per fish and 40% over a period of one week in that existing cage culture. The parasite was identified as a crustacean bloodsucker, anchor worm Lernaea sp. and EMB was found to be 100% effective with significant reduction in PI over a period of 10 days with improved survival rate of 90% against the untreated group. Infested but treated group revealed substantial haematological improvement in parameters such as RBC, WBC, Hb, PCV, large lymphocytes, small lymphocytes and total lymphocytes (P < 0.01). Similarly, comparative histopathology of vital organs also revealed no discernible lesions between the healthy and treated fish juvenile as compared to that of infested untreated group. Hence, EMB can be used to control the Lernaea sp. infestation in Asian Seabass.

The effects of extended feeding of florfenicol coated medicated diets on the safety, serum biomarkers and blood cells morphology of Nile tilapia Oreochromis niloticus (L.).

Tilapia is one of the most consumed farmed fish, which requires the use of antibiotics in certain phases of its production. This study assessed the safety of 30 days of oral florfenicol (FFC) dosing at 0-10 times the therapeutic dose (1 × : 10 mg/kg biomass/day) in Oreochromis niloticus juveniles. Behavioural changes, feed consumption, mortality and biomass were evaluated. Besides, the levels of serum glucose, calcium, chloride, creatinine, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and blood cell morphology were determined at scheduled intervals. The 30 days of oral FFC dosing caused 3.33% (1 ×) to 18.33% (10 ×) mortalities, reduced feed intake and biomass in a dose-dependent manner. The fish fed the therapeutic dose recorded 1.25-fold increase in biomass, while the control group recorded 1.45-fold increase in 30 days. No significant erythrocyte morphological alterations were observed in the 1 × group compared to the control. However, marked morphological alterations like tear-shaped, spindle-shaped and degenerative erythrocytes in higher dosing groups indicated FFC cytotoxicity. All the serum biomarkers of O. niloticus increased significantly on day 10 and day 30 FFC dosing in a dose-dependent manner, except for calcium and chloride, which reduced significantly during the dosing period. Within 2 weeks of suspension of FFC dosing, the serum biomarker levels became normal except for alkaline phosphatase and creatinine. The recovery of biomass, feed intake, serum biomarker levels and erythrocyte morphological changes suggested that the FFC-induced changes are reversible. This study has, thus, proclaimed the safety of FFC at the therapeutic dose in O. niloticus.

Dysbiosis and Restoration Dynamics of the Gut Microbiome Following Therapeutic Exposure to Florfenicol in Snubnose Pompano (Trachinotus blochii) to Aid in Sustainable Aquaculture Production Strategies.

Information on unintended effects of therapeutic exposure of antibiotics on the fish gut microbiome is a vital prerequisite for ensuring fish and environmental health during sustainable aquaculture production strategies. The present study forms the first report on the impact of florfenicol (FFC), a recommended antibiotic for aquaculture, on the gut microbiome of snubnose pompano (Trachinotus blochii), a high-value marine aquaculture candidate. Both culture-dependent and independent techniques were applied to identify the possible dysbiosis and restoration dynamics, pointing out the probable risks to the host and environment health. The results revealed the critical transient dysbiotic events in the taxonomic and functional metagenomic profiles and significant reductions in the bacterial load and diversity measures. More importantly, there was a complete restoration of gut microbiome density, diversity, functional metagenomic profiles, and taxonomic composition (up to class level) within 10-15 days of antibiotic withdrawal, establishing the required period for applying proper management measures to ensure animal and environment health, following FFC treatment. The observed transient increase in the relative abundance of opportunistic pathogens suggested the need to apply proper stress management measures and probiotics during the period. Simultaneously, the results demonstrated the inhibitory potential of FFC against marine pathogens (vibrios) and ampicillin-resistant microbes. The study pointed out the possible microbial signatures of stress in fish and possible probiotic microbes (Serratia sp., Methanobrevibacter sp., Acinetobacter sp., and Bacillus sp.) that can be explored to design fish health improvisation strategies. Strikingly, the therapeutic exposure of FFC neither caused any irreversible increase in antibiotic resistance nor promoted the FFC resistant microbes in the gut. The significant transient increase in the numbers of kanamycin-resistant bacteria and abundance of two multidrug resistance encoding genes (K03327 and K03585) in the treated fish gut during the initial 10 days post-withdrawal suggested the need for implementing proper aquaculture effluent processing measures during the period, thus, helps to reduce the spillover of antibiotic-resistant microbes from the gut of the treated fish to the environment. In brief, the paper generates interesting and first-hand insights on the implications of FFC treatment in the gut microbiome of a marine aquaculture candidate targeting its safe and efficient application in unavoidable circumstances. Implementation of mitigation strategies against the identified risks during the initial 15 days of withdrawal period is warranted to ensure cleaner and sustainable aquaculture production from aquatic animal and ecosystem health perspectives.