A New Cell Line from the Brain of Red Hybrid Tilapia (Oreochromis spp.) for Tilapia Lake Virus Propagation.

Tilapia lake virus (TiLV) presents a substantial threat to global tilapia production. Despite the development of numerous cell lines for TiLV isolation and propagation, none have been specifically derived from red hybrid tilapia (Oreochromis spp.). In this study, we successfully established a new cell line, RHTiB, from the red hybrid tilapia brain. RHTiB cells were cultured for 1.5 years through over 50 passages and demonstrated optimal growth at 25 °C in Leibovitz-15 medium supplemented with 10% fetal bovine serum at pH 7.4. Morphologically, RHTiB cells displayed a fibroblast-like appearance, and cytochrome oxidase I gene sequencing confirmed their origin from Oreochromis spp. Mycoplasma contamination testing yielded negative results. The revival rate of the cells post-cryopreservation was observed to be between 75 and 80% after 30 days. Chromosomal analysis at the 25th passage revealed a diploid count of 22 pairs (2n = 44). While no visible cytopathic effects were observed, both immunofluorescence microscopy and RT-qPCR analysis demonstrated successful TiLV propagation in the RHTiB cell line, with a maximum TiLV concentration of 107.82 ± 0.22 viral copies/400 ng cDNA after 9 days of incubation. The establishment of this species-specific cell line represents a valuable advancement in the diagnostic and isolation tools for viral diseases potentially impacting red hybrid tilapia.

Short-term starvation at different feeding regimes on appetite responses, feeding utilization and physiological indices, of red hybrid tilapia (Oreochromis mossambicus × Oreochromis niloticus) fingerlings reared in brackish water.

A 42 day factorial trial (3x2) was designed to evaluate the effect of short-term starvation with different feeding frequencies on performance, feed utilization, physiological status and appetite responses of red hybrid tilapia fingerlings. Eighteen plastic tanks with a capacity of (55 L) were used to accomplish this work. Fingerlings with an average initial weight of 23 g ± 0.2 (SE) were randomly stocked at a rate of 8 fingerlings/aquarium. Six groups were designated as the following: II/ED: fish was fed twice every day; IV/ED: fish fed four times every day; II/EOD: fish fed twice every other day (alternate-day feeding or one day of feeding followed by another of fasting); IV/EOD: was fed four times every other day; II/EO3D: fish fed twice every other three days (three day of feeding followed by another of fasting) and IV/EO3D: fish fed four times every other three days. Fish were fed on commercial diets 30 % protein (4 % of biomass). Results showed insignificant differences between fish fed every other day and those fed every day in some growth indicators. In the same trend, the interaction between feed deprivation and feeding frequency cleared that fingerlings of IV/EOD did not significantly differ with those fed every day in growth indices. Moreover this treatment was the best in feed conversion efficiency and several physiological indicators.

Immunogenicity and Efficacy of a Feed-Based Bivalent Vaccine against Streptococcosis and Motile Aeromonad Septicemia in Red Hybrid Tilapia (Oreochromis sp.).

Streptococcosis and motile Aeromonad septicemia (MAS) are the main bacterial diseases in tilapia culture worldwide, causing significant economic losses. Vaccination is an effective method of preventing diseases and contributes to economic sustainability. This study investigated the immuno-protective efficacy of a newly developed feed-based bivalent vaccine against streptococcosis and MAS in red hybrid tilapia. The feed-based bivalent vaccine pellet was developed by incorporating the formalin-killed S. agalactiae and A. hydrophila antigens into a commercial feed pellet with palm oil as the adjuvant. The bivalent vaccine was subjected to feed quality analyses. For immunological analyses, 900 fish (12.94 ± 0.46 g) were divided into two treatment groups in triplicate. Fish in Group 1 were unvaccinated (control), while those in Group 2 were vaccinated with the bivalent vaccine. The bivalent vaccine was delivered orally at 5% of the fish's body weight for three consecutive days on week 0, followed by boosters on weeks 2 and 6. Lysozyme and enzyme-linked immunosorbent assays (ELISAs) on serum, gut lavage, and skin mucus were performed every week for 16 weeks. Lysozyme activity in vaccinated fish was significantly (p ≤ 0.05) higher than in unvaccinated fish following vaccination. Similarly, the IgM antibody levels of vaccinated fish were significantly (p ≤ 0.05) higher after vaccination. The bivalent vaccine provided high protective efficacy against S. agalactiae (80.00 ± 10.00%) and A. hydrophila (90.00 ± 10.00%) and partial cross-protective efficacy against S. iniae (63.33 ± 5.77%) and A. veronii (60.00 ± 10.00%). During the challenge test, fewer clinical and gross lesions were observed in vaccinated fish compared with unvaccinated fish. Histopathological assessment showed less severe pathological changes in selected organs than the unvaccinated fish. This study showed that vaccination with a feed-based bivalent vaccine improves immunological responses in red hybrid tilapia, and thus protects against streptococcosis and MAS.

Tilapia lake virus (TiLV) causes severe anaemia and systemic disease in tilapia.

Tilapia lake virus disease (TiLVD) is an emerging disease in tilapia that is associated with mass mortality affecting global tilapia aquaculture. In this study, red hybrid tilapias (Oreochromis spp.) were experimentally infected by intracoelomic injection with Tilapia lake virus (TiLV) to gain a better understanding of the clinicopathological changes during infection. Pale bodies and gill were observed in infected fish after 7 days of post-challenge (dpc) associated with severe anaemia. Further haematological analysis in TiLV-infected fish revealed decreased levels of haemoglobin and haematocrit at 3 dpc. Common pathological findings included pale and friable liver, pale intestine with catarrhal content, and dark and shrunken spleen in TiLV-infected fish at 7 dpc and 14 dpc. Histologically, reduced numbers of red blood cells and accumulation of melano-macrophage centre in the spleen were found in infected fish at 3 dpc, and severe lesions were more commonly observed at 7 and 14 dpc. Lymphocyte infiltration, syncytial cell formation and multifocal necrotic hepatitis were the prominent pathological findings in the liver of infected fish. The severity of pathological changes was associated with TiLV-infection with higher viral loads and with the expression pattern of pro-inflammatory cytokines and antiviral genes, including interferon regulatory factor 1 (irf1), interleukin (il-8), radical s-adenosyl methionine domain containing 2 (rsad2) and mx. Our study provides a comprehensive analysis of the haematological profile and pathological changes in tilapia during TiLV infection. Overall, lesions present in various organs, together with alteration of host immune response in TiLV-infected fish, indicate the systemic infection of this virus. The knowledge gained from this study improves our understanding of how TiLV causes pathological and haematological changes in tilapia.

The effect of feeding black soldier fly larvae on growth performance, protein, and fat content of red hybrid tilapia (Oreochromis spp.).

Background and Aim: In the aquaculture industry, the crucial goal is to minimize production costs, especially feeding costs, without significant side effects. Black soldier fly larva (BSFL) is a locally available, eco-friendly, and sustainable source that is high in crude protein (42% dry matter [DM]) and fat (35% DM). This study aimed to determine the growth performance along with the composition of crude fat and protein in red hybrid fingerlings after the addition of BSFL into the diet. Materials and Methods: A total of 120 fingerlings of uniform size (mean initial weight of 1.46 ± 0.06 g) were randomly assigned to one of four groups (n = 10) (A, B, C, and D) per tank (1 m × 2 m × 1 m). For 21 days, Group A (control group) was fed with 100% commercial diet; Group B was fed with 90% commercial fish diet + 10% BSFL; Group C was fed with 80% commercial fish diet + 20% BSFL; and Group D was fed with 70% commercial fish diet + 30% BSFL. Feed efficiency, growth performance, and proximate composition analysis were performed on the fish. Results: The results displayed that the group with the highest BSFL percentage had a greater effect on protein and fat composition than the control group. The proximate composition analysis of fish-fed diet revealed that an increase in the level of BSFL inclusion increases the protein content in the fish. In comparison to the other groups, the experimental diet with 30% BSFL inclusion has the highest levels of crude protein (80.30% DM) and fat (2.90% DM). Conclusion: It is concluded that incorporating BSFL into a commercial diet for red hybrid tilapia fingerlings increased crude protein and fat composition, providing an alternative protein and fat source in fish diets.

Effect of papaya (Carica papaya) leaf extract as dietary growth promoter supplement in red hybrid tilapia (Oreochromis mossambicus × Oreochromis niloticus) diet.

The purpose of this experiment was to examine the potential use of Carica papaya leaf extract as a supplement to promote growth and improve feed utilization in red hybrid tilapia. Five diets were formulated containing isolipidic (80 g/kg) and isonitrogenic (350 g/kg) levels. All feeds contained similar types and amounts of raw materials but differed in the inclusion of papaya leaf extract (0, 5, 10, 20 and 40 g/kg feed). The initial size of fish used was 2.3 ± 0.01 g. Each diet was performed in triplicate tanks, and the feeding period was 12 weeks. Fish fed diet containing 2% papaya leaf extract (PLE) had the highest final weight, 31.14 ± 1.47 g, followed by 1% PLE (27.27 ± 1.75 g). These two diets (1% and 2%) were also showed significant improvements of weight gain, SGR, and feed efficiency of the red hybrid tilapia (p < 0.05). However, papaya leaf extract did not affect the HSI, VSI, PER, digestive enzymes activity, blood composition, and survival rate. Supplementing the diets with papaya leaf extract lowered serum urea. Findings of this research suggest that adding papaya leaf extract to the diet of red hybrid tilapia improves growth and feed efficiency without adversely affecting blood parameters. Therefore, an inclusion level between 1% and 2% of the papaya leaf extract is recommended as a feed additive to promote red hybrid tilapia fry growth.

Effects of Probiotics on Growth, Survival, Water Quality and Disease Resistance of Red Hybrid Tilapia (Oreochromis spp.) Fingerlings in a Biofloc System.

Biofloc technology has shown positive effects in aquaculture, especially on the growth performance of cultured animals. The aims of this study were to evaluate the effects of adding different probiotic strains in a biofloc system on the growth performance and disease resistance of red hybrid tilapia (Oreochromis spp.). Three different probiotics (Lysinibacillus fusiformis SPS11, Bacillus amyloliquefaciens L9, and Enterococcus hirae LAB3), commercial probiotics (MG1) and a mixed probiotics (MP) combining all three strains were used in this study. The in vitro assay results showed that the mixed probiotic (MP) was able to inhibit the growth of Streptococcus agalactiae and Streptococcus iniae significantly compared to the single and commercial probiotic. The efficacy of MP was further tested in in vivo tilapia culture challenged with S. agalactiae. The best specific growth rate (3.73 ± 0.23% day-1) and feed conversion ratio (0.76 ± 0.04) were recorded in the group of biofloc with addition of MP. After being challenged with S. agalactiae, the group of biofloc with MP had significantly higher survival (83 ± 1.43%) compared to the other groups. Furthermore, the nitrogen concentration (NO2-N and NH4-N) was significantly lower in all the biofloc groups compared to the control. Hence, the addition of probiotics was able to provide beneficial effects to red hybrid tilapia culture in the biofloc system.

Comparative Pathogenicity of Aeromonas spp. in Cultured Red Hybrid Tilapia (Oreochromis niloticus × O. mossambicus).

The genus Aeromonas has been recognised as an important pathogenic species in aquaculture that causes motile Aeromonas septicaemia (MAS) or less severe, chronic infections. This study compares the pathogenicity of the different Aeromonas spp. that were previously isolated from freshwater fish with signs of MAS. A total of 124 isolates of Aeromonas spp. were initially screened for the ability to grow on M9 agar with myo-inositol as a sole carbon source, which is a discriminatory phenotype for the hypervirulent A. hydrophila (vAh) pathotype. Subsequently, LD50 of six selected Aeromonas spp. were determined by intraperitoneal injection of bacterial suspension containing 103, 105, and 107 CFU/mL of the respective Aeromonas sp. to red hybrid tilapias. The kidneys, livers and spleens of infected moribund fish were examined for histopathological changes. The screening revealed that only A. dhakensis 1P11S3 was able to grow using myo-inositol as a sole carbon source, and no vAh strains were identified. The LD50-240h of A. dhakensis 1P11S3 was 107 CFU/mL, while the non-myo-inositol utilizing A. dhakensis 4PS2 and A. hydrophila 8TK3 was lower at 105 CFU/mL. Similarly, tilapia challenged with the myo-inositol A. dhakensis 1P11S3 showed significantly (p < 0.05) less severe signs, gross and histopathological lesions, and a lower mortality rate than the non-myo-inositol A. dhakensis 4PS2 and A. hydrophila 8TK3. These findings suggested that myo-inositol utilizing A. dhakensis 1P11S3 was not a hypervirulent Aeromonas sp. under current experimental disease challenge conditions, and that diverse Aeromonas spp. are of concern in aquaculture farmed freshwater fish. Therefore, future study is warranted on genomic level to further elucidate the influence of myo-inositol utilizing ability on the pathogenesis of Aeromonas spp., since this ability correlates with hypervirulence in A. hydrophila strains.

Herbal hydrogel-based encapsulated Enterococcus faecium ABRIINW.N7 improves the resistance of red hybrid tilapia against Streptococcus iniae.

AIMS: The streptococcal disease has been associated with serious mortality and significant global economic loss in the tilapia farming industry. The overall goal of this work was to test herbal hydrogels based on encapsulated Enterococcus faecium ABRIINW.N7 for potential probiotic anti-microbial activity against Streptococcus iniae in red hybrid tilapia. METHODS AND RESULTS: Abnormal behaviour, clinical signs, postinjection survival and histopathology (kidney, liver, eye and brain) were measured. Cumulative mortality of CON+ , free cells, ALG and treatments (F1-F7) was 30, 24, 22, 19, 17, 17, 16, 14, 14 and 12 out of 30 fish and the survival rates for E. faecium ABRIINW.N7 microencapsulated in an alginate-BS blend with 0·5, 1, 1·5, 2, 2·5 and 3% fenugreek were 43, 43, 47, 53, 53 and 60%, respectively. After the incorporation of fenugreek with the alginate-BS blend, there was an 8-21% increase in probiotic cell viability. Furthermore, the survival rate for the alginate-BS blend with 2·5 and 3% fenugreek (F6 and F7) was significantly (P ≤ 0·05) higher than other blends. The highest encapsulation efficiency, viability in gastrointestinal conditions and during storage time and excellent antipathogenicity against S. iniae were observed in alginate-BS +3% fenugreek formulation (F7). CONCLUSIONS: It is recommended that probiotic strains like E. faecium ABRIINW.N7 in combination with local herbal gums, such as BS and fenugreek plus alginate, can be used as a suitable scaffold and an ideal matrix for the encapsulation of probiotics. SIGNIFICANCE AND IMPACT OF THE STUDY: This study proposes models connecting process parameters, matrix structure and functionality.

Immuno-protective efficiency of feed-based whole-cell inactivated bivalent vaccine against Streptococcus and Aeromonas infections in red hybrid tilapia (Oreochromis niloticus × Oreochromis mossambicus).

Streptococcosis and motile aeromonad septicemia (MAS) are well-known diseases in tilapia culture, which cause mass mortality with significant economic losses. The development of feed-based bivalent vaccines in controlling these diseases has been initiated, however, the mechanisms of immunities and cross-protection in fish remain unclear. This study was conducted to assess the immuno-protective as well as the cross-protective efficacy of a newly developed feed-based bivalent vaccine against Streptococcus and Aeromonas infections in red hybrid tilapia. A total of five groups of fish were vaccinated orally through two different techniques; bivalent vaccine (inactivated Streptococcus iniae and Aeromonas hydrophila) sprayed on feed pellets (BS group); bivalent vaccine (inactivated S. iniae and A. hydrophila) incorporated in feed (BI group); monovalent inactivated S. iniae and A. hydrophila vaccine separately incorporated into feed as monovalent S. iniae (MS group) and monovalent A. hydrophila (MA group); and control group (without vaccine). The feed-based vaccine was delivered orally at 5% of body weight for five consecutive days. The booster doses were given in the same manner on weeks 2 and 6. Serum and skin mucus samples were collected to assess the IgM responses using indirect ELISA. The first administration of the feed-based vaccine stimulated the IgM levels that lasted until week 3, while the second booster ensured that the IgM levels remained high for a period of 16 weeks in the BI, MS and MA groups. The BI group developed a strong and significantly (P < 0.05) higher systemic and mucosal IgM responses against both S. iniae and A. hydrophila and also cross-protective antigen S. agalactiae and A. veronii compared to the BS and control groups. Quantitative real-time PCR results also showed that the relative expressions of IL-8, INF-γ and IgM in the BI immunized fish spleen, head kidney and hindgut exhibited various significant (P < 0.05) rising trends following both the vaccination and the challenge phase. On weeks 10, all fish were challenged through the intraperitoneal route, where relative percent survivals (RPS) of 82.22 ± 3.85% when challenged with S. iniae, 77.78 ± 3.85% when challenged with A. hydrophila and 77.78 ± 3.85% when co-challenged with both S. iniae and A. hydrophila were observed in the BI group, which were significantly higher (P < 0.05) compared to the other groups. The BI group also showed significantly (P < 0.05) higher partial cross-protections following challenges with S. agalactiae (RPS at 60.00 ± 6.67%) and A. veronii (RPS at 57.78 ± 7.70%). This study demonstrated that immunization with feed-based BI vaccine elicited immune responses that were capable of protecting red hybrid tilapia against streptococcosis and MAS.

Efficacy of Feed-Based Formalin-Killed Vaccine of Streptococcus iniae Stimulates the Gut-Associated Lymphoid Tissues and Immune Response of Red Hybrid Tilapia.

Red hybrid tilapia were fed a formalin-killed oral Streptococcus iniae vaccine (FKV) in the present study was assessed. Three hundred Red hybrid tilapia 80 ± 10 g were divided into five groups (1A, 1B, 2A, 2B, and Cx), each consisting of 60 fish. Fish from Groups 1A, 1B, 2A, and 2B were fed with FKV over different periods of administration, while Group 2B was the only group of fish to receive an oral booster vaccination on day 14- and 21-days post-vaccination (dpv). Group Cx was fed with normal pellets containing no vaccine as a control group. At four weeks post-vaccination (wpv), all fish were experimentally infected with S. iniae. Groups 2A and 2B had the lowest level of mortalities following vaccination (45% and 30%, respectively) compared to Groups 1A and 1B (80% and 55%, respectively), while the level of mortalities in Group Cx was 100%. All vaccinated groups showed a significant increase in anti-S. iniae IgM levels (p < 0.05) in serum, mucus, and gut-lavage, while Group Cx did not (p > 0.05) and all fish in this group died by five weeks post-infection. In conclusion, fish fed with the S. iniae FKV had a greater level of protection against S. iniae, with increased specific antibody response to the vaccine and there was also evidence of GALT stimulation by the vaccine.

Efficacy of Whole Cell Inactivated Vibrio harveyi Vaccine against Vibriosis in a Marine Red Hybrid Tilapia (Oreochromis niloticus × O. mossambicus) Model.

Vibrio harveyi causes vibriosis in various commercial marine fish species. The infection leads to significant economic losses for aquaculture farms, and vaccination is an alternative approach for the prevention and control of fish diseases for aquaculture sustainability. This study describes the use of formalin-killed Vibrio harveyi (FKVh) strain Vh1 as a vaccine candidate to stimulate innate and adaptive immunities against vibriosis in a marine red hybrid tilapia model. Tilapia are fast growing; cheap; resistant to diseases; and tolerant to adverse environmental conditions of fresh water, brackish water, and marine water and because of these advantages, marine red hybrid tilapia is a suitable candidate as a model to study fish diseases and vaccinations against vibriosis. A total of 180 healthy red hybrid tilapias were gradually adapted to the marine environment before being divided into two groups, with 90 fish in each group and were kept in triplicate with 30 fish per tank. Group 1 was vaccinated intraperitoneally with 100 µL of FKVh on week 0, and a booster dose was similarly administered on week 2. Group 2 was similarly injected with PBS. Skin mucus, serum, and gut lavage were collected weekly for enzyme-linked immunosorbent assay (ELISA) and a lysozyme activity assay from a total of 30 fish of each group. On week 4, the remaining 60 fish of Groups 1 and 2 were challenged with 108 cfu/fish of live Vibrio harveyi. The clinical signs were monitored while the survival rate was recorded for 48 h post-challenge. Vaccination with FKVh resulted in a significantly (p < 0.05) higher rate of survival (87%) compared to the control (20%). The IgM antibody titer and lysozyme activities of Group 1 were significantly (p < 0.05) higher than the unvaccinated Groups 2 in most weeks throughout the experiment. Therefore, the intraperitoneal exposure of marine red hybrid tilapia to killed V. harveyi enhanced the resistance and antibody response of the fish against vibriosis.

Probiotics Modulate Tilapia Resistance and Immune Response against Tilapia Lake Virus Infection.

Tilapia lake virus (TiLV) causes an emerging viral disease associated with high mortality and economic damage in tilapia farming around the world. The use of probiotics in aquaculture has been suggested as an alternative to antibiotics and drugs to reduce the negative impact of bacterial and viral infections. In this study, we investigate the effect of probiotic Bacillus spp. supplementation on mortality, viral load, and expression of immune-related genes in red hybrid tilapia (Oreochromis spp.) upon TiLV infection. Fish were divided into three groups, and fed with: control diet, 0.5% probiotics-supplemented diet, and 1% probiotics-supplemented diet. After 21 days of experimental feeding, the three groups were infected with TiLV and monitored for mortality and growth performances, while organs were sampled at different time points to measure viral load and the transcription modulation of immune response markers. No significant difference was found among the groups in terms of weight gain (WG), average daily gain (ADG), feed efficiency (FE), or feed conversion ratio (FCR). A lower cumulative mortality was retrieved from fish fed 0.5% and 1% probiotics (25% and 24%, respectively), compared to the control group (32%). Moreover, fish fed with 1% probiotic diet had a significantly lower viral load, than those fed with 0.5% probiotic and control diet at 5, 6, 9, and 12 days post infection-challenge (dpc). The expression patterns of immune-related genes, including il-8 (also known as CXCL8), ifn-γ, irf-3, mx, rsad-2 (also known as VIPERIN) showed significant upregulation upon probiotic treatment during the peak of TiLV pathogenesis (between 9 and 12 dpc) and during most of the study period in fish fed with 1% probiotics-supplemented diet. Taken together, these findings indicate that dietary supplementation using Bacillus spp. probiotics may have beneficial effects to strengthen tilapia immunity and resistance against TiLV infections. Therefore, probiotic treatments may be preventively administered to reduce losses caused by this emerging viral infection in tilapia aquaculture.

Antibiotic and heavy metal resistance of Aeromonas hydrophila and Edwardsiella tarda isolated from red hybrid tilapia (Oreochromis spp.) coinfected with motile aeromonas septicemia and edwardsiellosis.

AIM: The aim of this study is to identify antibiogram and heavy metal resistance pattern of Aeromonas hydrophila and Edwardsiella tarda isolated from red hybrid tilapia (Oreochromis spp.) coinfected with motile aeromonas septicemia and edwardsiellosis in four commercial fish farms. MATERIALS AND METHODS: A. hydrophila and E. tarda were isolated using glutamate starch phenol red and xylose lysine deoxycholate (Merck, Germany) as a selective medium, respectively. All the suspected bacterial colonies were identified using conventional biochemical tests and commercial identification kit (BBL Crystal, USA). Susceptibility testing of present bacterial isolates to 16 types of antibiotics (nalidixic acid, oxolinic acid, compound sulfonamides, doxycycline, tetracycline, novobiocin, chloramphenicol, kanamycin, sulfamethoxazole, flumequine, erythromycin, ampicillin, spiramycin, oxytetracycline, amoxicillin, and fosfomycin) and four types of heavy metals (mercury, chromium, copper, and zinc) were carried out using disk diffusion and two-fold agar dilution method, respectively. RESULTS: Three hundred isolates of A. hydrophila and E. tarda were successfully identified by biochemical tests. Antibiotic susceptibility testing results showed that 42.2% of the bacterial isolates were sensitive to compound sulfonamides, sulfamethoxazole, flumequine, oxytetracycline, doxycycline, and oxolinic acid. On the other hand, 41.6% of these isolates were resistant to novobiocin, ampicillin, spiramycin, and chloramphenicol, which resulted for multiple antibiotic resistance index values 0.416. Among tested heavy metals, bacterial isolates exhibited resistant pattern of Zn2+ > Cr6+ > Cu2+ > Hg2+. CONCLUSION: Results from this study indicated that A. hydrophila and E. tarda isolated from coinfected farmed red hybrid tilapia were multi-resistant to antibiotics and heavy metals. These resistant profiles could be useful information to fish farmers to avoid unnecessary use of antimicrobial products in the health management of farmed red hybrid tilapia.

Peperomia pellucida leaf extract as immunostimulator in controlling motile aeromonad septicemia due to Aeromonas hydrophila in red hybrid tilapia, Oreochromis spp. farming.

AIM: This study was revealed the potential of Peperomia pellucida leaf extract as an immunostimulator agent in controlling motile aeromonad septicemia due to Aeromonas hydrophila in red hybrid tilapia, Oreochromis sp. MATERIALS AND METHODS: In the present study, minimum inhibitory concentration (MIC) of P. pellucida leaf extract against A. hydrophila was determined through two-fold microbroth dilution method. The plant extract was screening for its active compound using a gas chromatograph mass spectrometer, and the effectiveness of P. pellucida leaf extract as an immunostimulator agent was evaluated. The experimental fish were fed with medicated feed at three different concentrations (25 mg/kg, PP-25; 50 mg/kg, PP-50; and 100 mg/kg, PP-100) of P. pellucida leaf extract for 1 week before they were intraperitoneally exposed to A. hydrophila. Enzyme-linked immunosorbent assay was carried out to determine the value of antibody response to A. hydrophila in fish from a group of fish that received medicated feed, and the percentage of total cumulative mortality of the experimental fish were observed at the end of the experiment. RESULTS: The results showed that the major bioactive compound is phytol (40%), and the MIC value was 31.5 mg/L. The value of antibody response to A. hydrophila in fish from a group of fish which received medicated feed (PP-25, 0.128±0.014 optical density [OD]; PP-50, 0.132±0.003 OD; and PP-100, 0.171±0.02 OD) was found significantly higher (p<0.05) compared to fish did not receive medicated feed (0.00 OD). Whereas, percentage cumulative mortality of fish from all groups of fish received medicated feed (PP-25, 18.0±3.2%; PP-50, 18.2±2.8%; and PP-100, 17.7±1.8%) were found significantly lower (p<0.05) compared to a group of fish did not receive medicated feed (83.2±1.4%). CONCLUSION: The findings of the present study indicated the huge potential of P. pellucida leaf extract as natural immunostimulator agent for aquaculture uses.

Development and efficacy of feed-based recombinant vaccine encoding the cell wall surface anchor family protein of Streptococcus agalactiae against streptococcosis in Oreochromis sp.

This study was carried out to determine the antibody responses and protective capacity of an inactivated recombinant vaccine expressing the cell wall surface anchor family protein of Streptococcus agalactiae following oral vaccination against streptococcosis in tilapia. Tilapia were vaccinated orally with 10(6) CFU/mL of the recombinant vaccine incorporated in feed (feed-based recombinant vaccine) (vaccinated group or Group 1), 10(6) CFU/mL of pET-32 Ek/LIC vector without cell wall surface anchor family protein (control group or Group 2), 10(6) CFU/mL of formalin-killed cells of S. agalactiae vaccine incorporated in feed was also prepared (feed-based vaccine) (vaccinated group or Group 3), and unvaccinated control group or Group 4 (fed with commercial pellets). During the course of study, serum, mucus and gut lavage fluid were collected to evaluate the antibody levels via enzyme-linked immunosorbent assay (ELISA). The results showed that tilapia immunized with the feed-based recombinant vaccine developed a strong and significantly (P < 0.05) higher IgM antibody response in serum, mucus and gut lavage fluid samples compared to groups 2, 3 and 4. Following heat intervenes and intraperitoneal challenge, the rate of survivors (RPS) was 70% for the vaccinated group, and 0% for the rest of the groups. Therefore, the study revealed that the feed-based recombinant vaccine significantly provides high protection against high dose challenge in heat stress environment and enhances the production of the mucosal and humoral immunity.

Agonistic and reproductive behaviors in males of red hybrid tilapia, Oreochromis niloticus (Linnaeus, 1758) x O. mossambicus (Peters, 1852) (Osteichthyes: Cichlidae)

The red hybrid tilapia, Oreochromis niloticus (Linnaeus, 1758) x O. mossambicus (Peters, 1852) is a fertile hybrid used in the semi-intensive level of fish culture in the Northeast of Brazil. It is a territorial cichlid and is highly aggressive towards conspecifics during the breeding season. The purpose of this study was to investigate and describe the aggressive behaviour displayed by the males of this hybrid in non-reproductive and reproductive contexts. Behavioural observations revealed that aggression displayed by the reproductive males of red hybrid tilapia included threatening, undulation, parallel, lateral and frontal attacks, chasing, escape and submission. Possession of a territory influenced male aggressiveness, which was more intense in their own territory than that observed in a neutral situation. The males built nests, irrespective of female presence. All the behavioural patterns were in accordance with those previously described for one parental species, the Nile tilapia, O. niloticus.

O híbrido vermelho de tilápia Oreochromis niloticus (Linnaeus, 1758) x O. mossambicus (Peters, 1852) é um híbrido fértil utilizado na piscicultura numa escala semi-intensiva no Nordeste brasileiro. É um ciclídeo territorial e altamente agressivo frente a coespecíficos na época reprodutiva. O objetivo deste estudo foi investigar e descrever os comportamentos agressivos exibidos pelos machos deste híbrido em contextos não-reprodutivos e reprodutivos. As observações comportamentais mostraram que as agressões exibidas pelos machos de híbrido vermelho de tilápia incluíram ameaça, ondulação, confronto paralelo, lateral e frontal, perseguição, fuga e submissão. A posse do território influenciou a agressividade dos machos, cuja intensidade foi maior no próprio território em relação àquela observada em situação neutra. Os ninhos foram escavados pelos machos independente da presença das fêmeas. Todos os padrões comportamentais descritos estavam de acordo com os padrões previamente descritos para uma das espécies parentais, a Tilápia do Nilo, O. niloticus.