In vitro efficacy of aquaculture antimicrobials and genetic determinants of resistance in bacterial isolates from tropical aquaculture disease outbreaks.

Understanding the efficacy of antimicrobials against pathogens from clinical samples is critical for their responsible use. The manuscript presents in vitro efficacy and antimicrobial resistance (AMR) genes in seven species of fish pathogens from the disease outbreaks of Indian aquaculture against oxytetracycline, florfenicol, oxolinic acid, and enrofloxacin. In vitro efficacy was evaluated by minimum inhibitory concentration and minimum bactericidal concentration. The gene-specific PCR screened AMR genes against quinolones (qnrA, qnrB, and qnrS) and tetracyclines (tetM, tetS, tetA, tetC, tetB, tetD, tetE, tetH, tetJ, tetG, and tetY). The results showed that Aeromonas veronii (45%) showed the maximum resistance phenotype, followed by Streptococcus agalactiae (40%), Photobacterium damselae (15%), Vibrio parahaemolyticus (10%), and Vibrio vulnificus (5%). There was no resistance among Vibrio harveyi and Vibrio alginolyticus against the tested antimicrobials. The positive association between tetA, tetB, tetC, tetM, or a combination of these genes to oxytetracycline resistance and qnrS to quinolone resistance indicated their potential in surveillance studies. The prevalence of resistance phenotypes (16.43%) and evaluated AMR genes (2.65%) against aquaculture antimicrobials was low. The resistance phenotype pattern abundance was 0.143. All the isolates showed susceptibility to florfenicol. The results help with the appropriate drug selection against each species in aquaculture practices.

Description and development of Auerbachia ignobili n. sp. (Cnidaria: Myxosporea: Bivalvulida) from the giant trevally, Caranx ignobilis (Forsskål, 1775) from Indian waters.

The present study describes a new species of myxosporean, Auerbachia ignobili n. sp., infecting the hepatic bile ducts of Caranx ignobilis (Forsskål, 1775). Myxospores are club-shaped with a broad anterior region and a narrow, slightly curved and blunt caudal extension, measuring 17.4 ± 1.5 µm in length and 7.5 ± 7.4 µm in width. Shell valves asymmetrical, with a faint suture line, and enclosed a single, elongate-elliptical polar capsule with a ribbon-like polar filament, arranged in 5-6 coils. Developmental stages included early and late presporogonic stages, pansporoblast, and sporogonic stages with monosporic and disporic plasmodia. A. ignobili n. sp. differs from the other described species of Auerbachia in the shape and dimensions of the myxospores and polar capsules. The molecular analysis generated ∼1400 bp long SSU rDNA sequences and the present species exhibited a maximum similarity 94.04-94.91% with A. chakravartyi. Genetic distance analysis indicated the lowest interspecies divergence of 4.4% with A. chakravartyi. In phylogenetic analysis, A. ignobili n. sp. was positioned independently with a high bootstrap value (1/100) and appeared as sister to A. maamouni and A. chakravartyi. Fluorescent in situ hybridization and histology indicates that the parasite develops within the hepatic bile ducts. Histological studies did not reveal any pathological changes. Considering the morphological, morphometric, molecular, and phylogenetic differences coupled with the differences in host and geographic locations, the present myxosporean is treated as a new species and named A. ignobili n. sp.

Pharmacokinetics and tissue distribution of florfenicol and florfenicol amine in snubnose pompano (Trachinotus blochii) following oral administration.

The present study reports the comparative pharmacokinetic profiles of florfenicol and its metabolite (florfenicol amine, FFA) in Trachinotus blochii under tropical marine conditions (salinity: 35 ± 1.4‰; temperature: 28.8 ± 0.54 °C) following a single in-feed oral administration of the recommended dose (15 mg/Kg). Furthermore, the study investigated the distribution of these two compounds in nine different tissues. The maximum florfenicol concentrations (Cmax) in plasma and tissues were observed within five hours (Tmax), except for bile. The Cmax ranged from 572 to 1954 ng/g or ml and was in the intestine > bile > muscle + skin > liver > gill = heart > plasma > kidney = spleen. The elimination half-life of FFC was significantly slower in the bile (38.25 ± 4.46 h). The AUC tissue/plasma was highest for bile (3.77 ± 0.22), followed by intestine > muscle + skin > heart > liver > kidney = gill = spleen. Tmax and t1/2ß were slower, and Cmax was lower for FFA than florfenicol in all tissues except Cmax of the kidney and bile. FFA t1/2ß was exceptionally slower in the kidney (46.01 ± 8.2 h). Interestingly, reaching an apparent distribution rate of > 0.5 was comparatively faster in the kidney, liver, and gills than in other tissues. The highest apparent metabolic rate was in the kidney (0.95 ± 0.01) and the lowest in plasma (0.41 ± 0.01). The generated data can be applied for formulating efficient therapeutic protocols in T. blochii, a promising mariculture species.

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.

Histopathological evaluation of bivalves from the southwest coast of India as an indicator of environmental quality.

Bivalve molluscs have been regarded as excellent bioindicators of environmental pollution as they persistently accumulate toxic contaminants present in their ecosystem. Histological alterations in the digestive gland and gills of three bivalve sp., Viz. edible oyster (Magallana bilineata), green mussel (Perna viridis) and black clam (Villorita cyprinoides) from ecologically sensitive regions of international significance on the southwest coast of India were evaluated using a semi-quantitative histopathological index to assess the environmental quality. The prominent tissue alterations included tubular vacuolation, haemocytic infiltration, parasitosis, lamellar disorganization, and the presence of prokaryotic inclusions.  The presence of ten trace metals was also evaluated in the digestive gland of bivalves. The histopathological indices were evaluated season-wise and region-wise. Seasonal variation in all the reaction patterns was observed in the digestive gland across sampling zones, with the highest indices observed during post-monsoon. The indices for all the reaction patterns in the digestive gland were significantly higher in bivalves from Vembanad Lake (Z4), followed by Periyar River (Z5). The indices for cellular changes and parasitosis in gills were the highest in the Ashtamudi estuary (Z1) and Z5, respectively. The global histopathological indices of the digestive gland and gills were also the highest in Z4, followed by Z5. Principal component analysis revealed that Z4 was distinct with the highest metal pollution index. A positive relation was observed with heavy metals, digestive gland histological alterations, and season and region of sampling.