Comparability of comparative toxicity: insect sensitivity to imidacloprid reveals huge variations across species but also within species.

Pesticides have been identified as major drivers of insect biodiversity loss. Thus, the study of their effects on non-pest insect species has attracted a lot of attention in recent decades. In general toxicology, the 'gold standard' to assess the toxicity of a substance is to measure mass-specific LD50 (i.e. median lethal dose per unit body mass). In entomology, reviews attempting to compare these data across all available studies are lacking. To fill this gap in knowledge, we performed a systematic review of the lethality of imidacloprid for adult insects. Imidacloprid is possibly the most extensively studied insecticide in recent times, yet we found that little is comparable across studies, owing to both methodological divergence and missing estimates of body mass. By accounting for body mass whenever possible, we show how imidacloprid sensitivity spans across an apparent range of approximately six orders of magnitude across insect species. Very high variability within species can also be observed owing to differences in exposure methods and observation time. We suggest that a more comparable and comprehensive approach has both biological and economic relevance. Ultimately, this would help to identify differences that could direct research towards preventing non-target species from being negatively affected.

RNA-seq analysis revealed the pathogenicity of Vibrio vulnificus to American eel (Anguilla rostrata) and the strategy of host anti-V. vulnificus infection.

Vibrio vulnificus is a commonly pathogenic bacterium in cultivated eels, but its pathogenicity to American eel (Anguilla rostrata) and the molecular mechanism of host anti-V. vulnificus infection remains uncertain. In this study, American eels were infected with different dose of V. vulnificus to determine the LD50. Then, bacterial load in the liver and kidney histopathology were assessed post the LD50 of V. vulnificus infection. Additionally, gene expressions of 18 immune related genes in the liver, spleen and kidney were detected. Furthermore, transcriptome sequencing and enrichment of differentially expressed genes (DEGs) were analyzed in the eel spleens between pre-infection (Con_0), post-36 h (Vv_36), and post-60 h (Vv_60) infection. The results showed that LD50 of V. vulnificus to American eels was determined to be 5.0 × 105 cfu/g body weight, and the bacterial load peaked at 24 and 12 h post the infection (hpi) in the kidney and liver, respectively. The histopathology was highlighted by necrotic hepatocytes and splenic cells, congestion blood vessels in liver and spleen, atrophied glomeruli and vacuolization of renal tubular epithelial cells. The results of RT-PCR revealed that 18 host immune-related genes showed significantly up or downregulated expression post-infection compare to that of pre-infection. Finally, results of the RNA-seq revealed 16 DEGs play essential role to the immunosuppression in American eels, and the protein-protein interactions shed light on the widespread upregulation GEGs related to metabolism and immune response maintained the host cell homeostasis post the V. vulnificus infection, shedding new light on our understanding of the V. vulnificus pathogenesis towards understudied American eel and the host anti-V. vulnificus infection strategies in gene transcript.

Transcriptome analysis of host anti-Aeromonas hydrophila infection revealed the pathogenicity of A. hydrophila to American eels (Anguilla rostrata).

Aeromonas hydrophila is a commonly pathogenic bacterium in cultivated eels, but its pathogenicity to American eel (Anguilla rostrata) and the molecular mechanism of host anti-A. hydrophila infection remains uncertain. In this study, LD50 of A. hydrophila to American eels was determined and bacterial load in the liver and kidney of eels was assessed post 2.56 doses of LD50 of A. hydrophila infection. The results showed that the LD50 of A. hydrophila to American eels was determined to be 3.9 × 105 cfu/g body weight (7.8 × 106 cfu/fish), and the bacterial load peaked at 36 h post the infection (hpi) in the liver. Then, the histopathology was highlighted by congestion in splenic blood vessels, atrophied glomeruli, and necrotic hepatocytes. Additionally, the results of qRT-PCR revealed that 18 host immune-related genes showed significantly up or downregulated post-infection compare to that of pre-infection. Finally, results of the RNA-seq revealed 10 hub DEGs and 7 encoded proteins play essential role to the anti-A. hydrophila infection in American eels. Pathogenicity of A. hydrophila to American eels and RNA-seq of host anti-A. hydrophila infection were firstly reported in this study, shedding new light on our understanding of the A. hydrophila pathogenesis and the host immune response to the A. hydrophila infection strategies in gene transcript.

Galleria mellonella larvae as an alternative model to determine the pathogenicity of avian pathogenic Escherichia coli.

RESEARCH HIGHLIGHTS: Galleria mellonella larvae are a viable model for determining APEC pathogenicity.Larval disease score is the main variable for determining APEC pathogenicity.Response variables should be evaluated up to 24 h post-inoculation.

Interference between Escherichia coli genotypes from the E. coli peritonitis syndrome given simultaneously to productive SPF White Leghorn hens by intratracheal inoculation.

RESEARCH HIGHLIGHTS: All four or only two E. coli genotypes were found in groups of hens given mixes of four genotypes.In contrast, only one genotype was found in individual hens.E. coli genotypes interfere with each other in hens after given as a mix.Interference is likely based on a random process.Broad protection can best be assessed by challenging with single genotypes.

Variation in Pesticide Toxicity in the Western Honey Bee (Apis mellifera) Associated with Consuming Phytochemically Different Monofloral Honeys.

Insecticide toxicity to insect herbivores has long been known to vary across different host plants; this phenomenon has been widely documented in both foliage-feeders and sap-feeders. Species-specific phytochemical content of hostplant tissues is assumed to determine the pattern of induction of insect enzymes that detoxify insecticides, but specific phytochemicals have rarely been linked to host plant-associated variation in pesticide toxicity. Moreover, no studies to date have examined the effects of nectar source identity and phytochemical composition on the toxicity of insecticides to pollinators. In this study, we compared LD50 values for the insecticide bifenthrin, a frequent contaminant of nectar and pollen in agroecosystems, in the western honey bee, Apis mellifera, consuming three phytochemically different monofloral honeys: Nyssa ogeche (tupelo), Robinia pseudoacacia (black locust), and Fagopyrum esculentum (buckwheat). We found that bifenthrin toxicity (LD50) values for honey bees across different honey diets is linked to their species-specific phytochemical content. The profiles of phenolic acids and flavonoids of buckwheat and locust honeys are richer than is the profile of tupelo honey, with buckwheat honey containing the highest total content of phytochemicals and associated with the highest bifenthrin LD50 in honey bees. The vector fitting in the ordination analysis revealed positive correlations between LD50 values and two honey phytochemical richness estimates, Chao1 and Abundance-based Coverage Estimator (ACE). These findings suggest unequal effects among different phytochemicals, consistent with the interpretation that certain compounds, including ones that are rare, may have a more pronounced effect in mitigating pesticide toxicity.

Evaluation of in silico model predictions for mammalian acute oral toxicity and regulatory application in pesticide hazard and risk assessment.

The United States Environmental Protection Agency (USEPA) uses the lethal dose 50% (LD50) value from in vivo rat acute oral toxicity studies for pesticide product label precautionary statements and environmental risk assessment (RA). The Collaborative Acute Toxicity Modeling Suite (CATMoS) is a quantitative structure-activity relationship (QSAR)-based in silico approach to predict rat acute oral toxicity that has the potential to reduce animal use when registering a new pesticide technical grade active ingredient (TGAI). This analysis compared LD50 values predicted by CATMoS to empirical values from in vivo studies for the TGAIs of 177 conventional pesticides. The accuracy and reliability of the model predictions were assessed relative to the empirical data in terms of USEPA acute oral toxicity categories and discrete LD50 values for each chemical. CATMoS was most reliable at placing pesticide TGAIs in acute toxicity categories III (>500-5000 mg/kg) and IV (>5000 mg/kg), with 88% categorical concordance for 165 chemicals with empirical in vivo LD50 values ≥ 500 mg/kg. When considering an LD50 for RA, CATMoS predictions of 2000 mg/kg and higher were found to agree with empirical values from limit tests (i.e., single, high-dose tests) or definitive results over 2000 mg/kg with few exceptions.

Identification of Novel Non-Nucleoside Inhibitors of Zika Virus NS5 Protein Targeting MTase Activity.

Zika virus (ZIKV) is a positive-sense single-stranded virus member of the Flaviviridae family. Among other arboviruses, ZIKV can cause neurological disorders such as Guillain Barré syndrome, and it can have congenital neurological manifestations and affect fertility. ZIKV nonstructural protein 5 (NS5) is essential for viral replication and limiting host immune detection. Herein, we performed virtual screening to identify novel small-molecule inhibitors of the ZIKV NS5 methyltransferase (MTase) domain. Compounds were tested against the MTases of both ZIKV and DENV, demonstrating good inhibitory activities against ZIKV MTase. Extensive molecular dynamic studies conducted on the series led us to identify other derivatives with improved activity against the MTase and limiting ZIKV infection with an increased selectivity index. Preliminary pharmacokinetic parameters have been determined, revealing excellent stability over time. Preliminary in vivo toxicity studies demonstrated that the hit compound 17 is well tolerated after acute administration. Our results provide the basis for further optimization studies on novel non-nucleoside MTase inhibitors.

Potentially active aspirin derivative to release nitric oxide: In-vitro, in-vivo and in-silico approaches.

The series of newer salicylate derivatives incorporating nitroxy functionality were synthesized and evaluated for their potential effect in gastrointestinal (GI) related toxicity produced by aspirin. The synthesized compounds (5a-j) were subjected to %NO (nitric oxide) release study, in-vitro anti-inflammatory potential, % inhibition of carrageenan-induced paw edema and the obtained results were validated by in-silico studies including molecular docking, MD simulations and in-silico ADME (absorption, distribution, metabolism, and elimination) calculations. Compounds 5a (20.86 %) and 5g (18.20 %) displayed the highest percentage of NO release in all the tested compounds. Similarly, 5a and 5h were found to have (77.11 % and 79.53 %) &(78.56 % and 66.10 %) inhibition in carrageenan induced paw edema in animal mode which were relatively higher than ibuprofen (standard used). The obtained results were validated by molecular docking and MD simulations studies. The molecular docking study of 5a and 5h revealed that docking scores were also obtained in very close proximity of -8.35, -9.67 and -8.48 for ibuprofen, 5g and 5h respectively. In MD simulations studies, the calculated lower RMSD (root mean square deviation) values 2.8 Å and 5.6 Å for 5g and 5h, respectively indicated the stability of ligand-protein complexes. Similarly lower RSMF (root mean square fluctuation) values indicated the molecules remained in the active pocket throughout the entire MD simulations run. Further, in-silico ADME calculations were determined and all compounds obey the Lipinski's rule of five and it was predicted that these molecules would be orally active without any serious toxic effect.

Pathogenicity of Edwardsiella anguillarum to American eels (Anguilla rostrata) and RNA-seq analysis of host immune response to the E. anguillarum infection.

Edwardsiella anguillarum is a commonly pathogenic bacterium in cultivated eels, but its pathogenicity to American eel (Anguilla rostrata) and the molecular mechanism of host anti-E. anguillarum infection remains uncertain. In this study, LD50 of E. anguillarum to American eels was determined and bacterial load in the liver and kidney of eels was assessed post the LD50 of E. anguillarum infection. The results showed that LD50 of E. anguillarum to American eels was determined to be 2.5 × 105 cfu/g body weight, and the bacterial load peaked at 36 and 72 h post the infection (hpi) in the kidney and liver, respectively. Then, the histopathology was highlighted by congestion in splenic blood vessels, atrophied glomeruli, and necrotic hepatocytes, as well as ultrastructural pathology in the kidney were charactered by acute nephritis, showing necrosis of the renal tubular epithelial cells, glomerular capillaries dilate, mitochondria swelling and ribosomes separate from the endoplasmic reticulum. Furthermore, the results of qRT-PCR revealed that 12 host immune-related genes showed significantly up or downregulated post-infection compare to that of pre-infection. Finally, results of the RNA-seq revealed 6 hub DEGs play essential role to the anti-E. anguillarum infection in American eels. Pathogenicity of E. anguillarum to American eels and hub genes related host anti- E. anguillarum infection were firstly reported in this study, shedding new light on our understanding of the E. anguillarum pathogenesis and the host immune response to the E. anguillarum infection strategies in gene transcript.

Acute and chronic toxicity of imidacloprid in the pollinator fly, Eristalis tenax L., assessed using a novel oral bioassay.

Imidacloprid is a neonicotinoid neurotoxin that remains widely used worldwide and persists in the environment, resulting in chronic exposure to non-target insects. To accurately map dose-dependent effects of such exposure across taxa, toxicological assays need to assess relevant modes of exposure across indicator species. However, due to the difficulty of these experiments, contact bioassays are most frequently used to quantify dose. Here, we developed a novel naturalistic feeding bioassay to precisely measure imidacloprid ingestion and its toxicity for acute and chronic exposure in a dipteran, Eristalis tenax L., an important member of an under-represented pollinator group. Flies which ingested imidacloprid dosages lower than 12.1 ng/mg all showed consistent intake volumes and learned improved feeding efficiency over successive feeding sessions. In contrast, at doses of 12.1 ng/mg and higher flies showed a rapid onset of severe locomotive impairment which prevented them from completing the feeding task. Neither probability of survival nor severe locomotive impairment were significantly higher than the control group until doses of 1.43 ng/mg or higher were reached. We were unable to measure a median lethal dose for acute exposure (72 h) due to flies possessing a relatively high tolerance for imidacloprid. However, with chronic exposure (18 days), mortality went up and an LD50 of 0.41 ng/mg was estimated. Severe locomotive impairment (immobilisation) tended to occur earlier and at lower dosages than lethality, with ED50s of 7.82 ng/mg and 0.17 ng/mg for acute and chronic exposure, respectively. We conclude that adult Eristalis possess a much higher tolerance to this toxin than the honeybees that they mimic. The similarity of the LD50 to other dipterans such as the fruitfly and the housefly suggests that there may be a phylogenetic component to pesticide tolerance that merits further investigation. The absence of obvious adverse effects at sublethal dosages also underscores a need to develop better tools for quantifying animal behaviour to evaluate the impact of insecticides on foraging efficiency in economically important species.

Tenebrio molitor as a Simple and Cheap Preclinical Pharmacokinetic and Toxicity Model.

The progression of drugs into clinical phases requires proper toxicity assessment in animals and the correct identification of possible metabolites. Accordingly, different animal models are used to preliminarily evaluate toxicity and biotransformations. Rodents are the most common models used to preliminarily evaluate the safety of drugs; however, their use is subject to ethical consideration and elevated costs, and strictly regulated by national legislations. Herein, we developed a novel, cheap and convenient toxicity model using Tenebrio molitor coleoptera (TMC). A panel of 15 drugs-including antivirals and antibacterials-with different therapeutic applications was administered to TMC and the LD50 was determined. The values are comparable with those already determined in mice and rats. In addition, a TMC model was used to determine the presence of the main metabolites and in vivo pharmacokinetics (PK), and results were compared with those available from in vitro assays and the literature. Taken together, our results demonstrate that TMC can be used as a novel and convenient preliminary toxicity model to preliminarily evaluate the safety of experimental compounds and the formation of main metabolites, and to reduce the costs and number of rodents, according to 3R principles.

In Vitro Evaluation of Antiprotozoal Properties, Cytotoxicity Effect and Anticancer Activity of New Essential-Oil Based Phytoncide Mixtures.

Protozoa, in both humans and animals, are one of the leading causes of disease. International programmes introduced in many countries have helped reduce the incidence of disease. However, it has recently become increasingly difficult to achieve the goals set for the coming years. One of the main reasons for this, as with other pathogenic organisms, such as bacteria and fungi, is the increasing resistance to current methods of treating and preventing infection. Therefore, new therapies with high efficacy are needed. In the present study, the novel mixtures of essential oils (EOs), clove, garlic, Ceylon cinnamon, and rosemary with organic acids (acetic, propionic, lactic) and metal ions (Cu, Mn, Zn) were tested against five selected model protozoa (Euglena gracilis, Gregarina blattarum, Amoeba proteus, Paramecium caudatum, Pentatrichomonas hominis). The cytotoxicity and potential anticancer activity of the obtained combinations were tested on the human fibroblasts (NHDF) and human cancer cell lines (A549, MCF7, LoVo, HT29). All of the mixtures showed very good antiprotozoal properties. The most efficient were the combination of clove and rosemary essential oils, mixtures of acids, and Mn ions. The LD50 values were in the range of 0.001-0.006% and the LD100 values were 0.002-0.008%. All of the tested mixtures did not show cytotoxicity against normal cells, but did show growth inhibition against cancer cell lines. The most cytotoxic against cancer cells were combinations with cinnamon essential oil. Nevertheless, the proposed combinations containing essential oils, organic acids, and metal ions have high antiprotozoal activity, with low toxicity to healthy human cells.

Acute and sub-acute toxicity study reveals no dentrimental effect of formononetin in mice upon repeated i.p. dosing.

AIM: Formononetin is a phytoestrogen which possess different pharmacological activities. The intraperitoneal route permits the identification of target organs involved in toxicity without compromising the molecule's bioavailability. The current study investigated the safety profile of intraperitoneal formononetin in Swiss albino mice. MATERIAL AND METHODS: For acute toxicity study, formononetin administered intraperitoneally to mice at the doses of 5, 50, 100, 150, 200, and 300 mg/kg for 14 days. For the subacute toxicity study, mice were intraperitoneally administered with formononetin (12.5, 25, and 50 mg/kg) daily for 28 days. RESULTS: During the acute study, no deteriorating effect was observed on body weight, food and water intake, no behavioral changes were observed in animals. The lethal dose 50% (LD50) of formononetin was determined to be 103.6 mg/kg of BW, with a no observed adverse effect level (NOAEL) of 50 mg/kg of BW. Mortality was observed in the 300 mg/kg dose group and histopathological changes such as a mild degree of diffuse granular degeneration in the liver but for rest all doses did not have any adverse effect. In subacute study, no signs of adverse effects, mortality, no changes in body weight, food and water intake, and hematological and biochemical parameters were observed. Histopathology of subacute study indicates, formononetin did not have any noxious effect on organs. CONCLUSION: Formononetin shows mortality at acute dose 300 mg/kg and LD50 at 103.6 mg/kg of BW, with a NOAEL of 50 mg/kg of BW, rest all doses for acute and sub-acute are safe when given intraperitoneally.

In Vivo Toxicity of Industrial Biocide Containing 2,2-Dibromo-3-nitrilopropionamide in Adult and Zebrafish Larvae.

2,2-Dibromo-3-nitrilopropionamide (DBNPA) is a widely used biocide with potential environmental implications due to its toxicity. This study aimed to investigate the in vivo toxicity of DBNPA in zebrafish (Danio rerio), a model organism in environmental toxicology. Both adult and larval zebrafish were exposed to varying concentrations of DBNPA, and significant morphological changes and mortality rates were observed. The study found that even relatively low concentrations of DBNPA can have detrimental effects on zebrafish embryonic development, and high concentrations resulted in rapid mortality in adult zebrafish and larvae. The LC50 values calculated from this study were 9.3 ppm for adults and 9.1 ppm for larvae, indicating the high toxicity of DBNPA to these organisms. These findings underscore the potential environmental impact of DBNPA and highlight the need for further research into its effects on aquatic ecosystems.

Multi-ligand molecular docking, simulation, free energy calculations and wavelet analysis of the synergistic effects between natural compounds baicalein and cubebin for the inhibition of the main protease of SARS-CoV-2.

Combination drugs have been used for several diseases for many years since they produce better therapeutic effects. However, it is still a challenge to discover candidates to form a combination drug. This study aimed to investigate whether using a comprehensive in silico approach to identify novel combination drugs from a Chinese herbal formula is an appropriate and creative strategy. We, therefore, used Toujie Quwen Granules for the main protease (Mpro) of SARS-CoV-2 as an example. We first used molecular docking to identify molecular components of the formula which may inhibit Mpro. Baicalein (HQA004) is the most favorable inhibitory ligand. We also identified a ligand from the other component, cubebin (CHA008), which may act to support the proposed HQA004 inhibitor. Molecular dynamics simulations were then performed to further elucidate the possible mechanism of inhibition by HQA004 and synergistic bioactivity conferred by CHA008. HQA004 bound strongly at the active site and that CHA008 enhanced the contacts between HQA004 and Mpro. However, CHA008 also dynamically interacted at multiple sites, and continued to enhance the stability of HQA004 despite diffusion to a distant site. We proposed that HQA004 acted as a possible inhibitor, and CHA008 served to enhance its effects via allosteric effects at two sites. Additionally, our novel wavelet analysis showed that as a result of CHA008 binding, the dynamics and structure of Mpro were observed to have more subtle changes, demonstrating that the inter-residue contacts within Mpro were disrupted by the synergistic ligand. This work highlighted the molecular mechanism of synergistic effects between different herbs as a result of allosteric crosstalk between two ligands at a protein target, as well as revealed that using the multi-ligand molecular docking, simulation, free energy calculations and wavelet analysis to discover novel combination drugs from a Chinese herbal remedy is an innovative pathway.

An update on the current status and prospects of nitrosation pathways and possible root causes of nitrosamine formation in various pharmaceuticals.

Over the last two years, global regulatory authorities have raised safety concerns on nitrosamine contamination in several drug classes, including angiotensin II receptor antagonists, histamine-2 receptor antagonists, antimicrobial agents, and antidiabetic drugs. To avoid carcinogenic and mutagenic effects in patients relying on these medications, authorities have established specific guidelines in risk assessment scenarios and proposed control limits for nitrosamine impurities in pharmaceuticals. In this review, nitrosation pathways and possible root causes of nitrosamine formation in pharmaceuticals are discussed. The control limits of nitrosamine impurities in pharmaceuticals proposed by national regulatory authorities are presented. Additionally, a practical and science-based strategy for implementing the well-established control limits is notably reviewed in terms of an alternative approach for drug product N-nitrosamines without published AI information from animal carcinogenicity testing. Finally, a novel risk evaluation strategy for predicting and investigating the possible nitrosation of amine precursors and amine pharmaceuticals as powerful prevention of nitrosamine contamination is addressed.

Development of fusarium wilt resistant mutants of Musa spp. cv.Rasthali (AAB, Silk subgroup) and comparative proteomic analysis along with its wild type.

MAIN CONCLUSION: Induced mutagenesis using embryogenic cell suspension (ECS) explants with toxin based screening is an effective tool to create non-chimeral Fusarium wilt resistant mutants in banana. Global proteomics unravel the molecular mechanism behind resistance. Race 1 of Fusarium wilt is a serious threat to Musa spp. cv.Rasthali (AAB, Silk subgroup) which is a choice variety traditionally grown in most of the south East Asian countries. Resistant gene introgression into susceptible varieties through conventional breeding has several limitations and the predominant ones being sterility and long generation time. Under such circumstances, induced mutagenesis combined with toxin based in vitro screening remains as the viable alternative for the development of fusarium wilt resistant Rasthali. Therefore, induced mutagenesis was attempted by using ethylmethane sulfonate (EMS) in embryogenic cell suspension (ECS) of Rasthali followed by in vitro screening for fusarium wilt resistance using new generation toxins and pot screening through challenge inoculation with Foc race 1. This ultimately resulted in the identification of 15 resistant lines. Global proteomic analysis in one of the resistant mutant lines namely NRCBRM15 and its wild type revealed 37 proteins, of which 20 showed differential expression. Out of 20 proteins, nineteen were significantly abundant in NRCBRM15 and only one was abundant in wild Rasthali. A total of nine genes based on protein expression were further validated using quantitative real time polymerase chain reaction (qRT-PCR). Annotation results revealed that some of the genes namely Enolase, ATP synthase-alpha subunit, Actin 2, Actin 3,-glucanase, UTP-glucose-1-phosphate uridylyltransferase, Respiratory burst oxidase homolog, V type proton ATPase catalytic subunit A and DUF292 domain containing protein are involved in diverse functions such as carbohydrate metabolism, energy production, electron carrier, response to wounding, binding proteins, cytoskeleton organization, extracellular region, structural molecule and defense.

Pathogenicity of Plesiomonas shigelloides and Citrobacter freundii isolated from the endangered Chinese sturgeon (Acipenser sinensis).

Chinese sturgeon (Acipenser sinensis) is an indigenous species of China and is listed as a critically endangered species. Recently, second filial generations of Chinese sturgeon in the Yangtze River Fisheries Research Institute suffered from a severe disease. In this study, two kinds of pathogenic bacteria were isolated from diseased sturgeon and identified as Plesiomonas shigelloides and Citrobacter freundii, based on 16S rDNA gene sequence alignment analysis. Antimicrobial susceptibility testing showed that P. shigelloides was resistant to ampicillin, penicillin, midecamycin, oxacillin, and clindamycin; and sensitive to tocefatriaxone, piperacillin, cefoperazone, cefazolin, and ciprofloxacin. C. freundii was resistant to ampicillin, penicillin, midecamycin, oxacillin, and clindamycin; and sensitive to chloramphenicol, cefuroxime, norfloxacin, ciprofloxacin, and ceftazidime. The median lethal dose (LD50) values of P. shigelloides and C. freundii were 4.50 × 103 colony forming units (CFU)/g and 3.20 × 103 CFU/g, respectively. Clinical symptoms of challenged sturgeons were the same as those of naturally infected sturgeons. Histopathological examination disclosed severe damage in the viscera of P. shigelloides and C. freundii-infected sturgeons. This is the first report suggesting that P. shigelloides infection is associated with mortality of Chinese sturgeon. The results of this study revealed the pathogenesis and severe pathogenicity of P. shigelloides and C. freundii in cultured Chinese sturgeon, and offer insights into the prevention and treatment of bacterial infection caused by P. shigelloides and C. freundii in cultured sturgeons.

First description of Vibrio harveyi as the causative agent of morbidity and mortality in farmed flathead grey mullet Mugil cephalus.

Flathead grey mullet Mugil cephalus is an important species in the aquaculture industry in the Mediterranean basin and throughout the world. During the last 10 yr, M. cephalus breeding stocks, larvae, and juveniles cultured in Eilat (Israel) have shown neurological signs such as uncoordinated circular swimming, while also presenting oral hemorrhages. Death follows days after the onset of the clinical signs, and mortality rates may reach 80% in some cases, causing high economical losses. Bacteriology isolations from different organs, including the brain, and a Koch's postulate experiment, confirmed Vibrio harveyi as the causative agent. Histological analyses showed the presence of the bacterium in different organs. However, in the brain, the bacterium was observed only within blood vessels and meninges. In some samples, mild to severe brain tissue damage was seen. In order to understand the virulence and lethality of V. harveyi, a median lethal dose was calculated, and the result was 106 colony-forming units fish-1. To the best of our knowledge, this is the first report that describes V. harveyi isolated from the brain of M. cephalus and validates it as an etiological agent causing neurological signs in this fish species.