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1.
Fifty‐seven isolates of mycobacteria comprising 10 reference strains, 47 field isolates and one non‐Mycobacterium isolate were screened using commercial INNO‐LiPA v2 assay kits. All mycobacteria isolates tested hybridized with the Mycobacterium genus probe on the LiPA strip. All M. marinum, M. fortuitum and M. chelonae reference and field strains and three out of the four M. gordonae isolates hybridized to their corresponding species‐ or complex‐specific probes. Two cultures (a type strain and a field isolate) yielded mixed growth of two mycobacterial species, i.e. M. chelonae and M. fortuitum. A Mycobacterium isolate from one of these cultures was subsequently purified and correctly identified with the kit. However, sequence analysis of the 16S–23S rRNA internal transcribed spacer (ITS) region of various mycobacteria isolates revealed a misidentification of M. shottsii and M. pseudoshottsii with the kit because these isolates reacted with the M. marinum/M. ulcerans probe. Moreover, nine of the 13 field isolates presumed to be M. fortuitum from the results of the kit had closer ITS sequence homology with M. conceptionense, a species which, to our knowledge, has never been reported in fish. These findings highlight the need to redesign the M. fortuitum–M. peregrinum probe included in the INNO‐LiPA assay and to introduce additional complex‐specific probes into the kit. Nevertheless, the kit proved to be a rapid and reliable method for identifying mycobacteria in the aquatic environment and would be particularly useful in laboratories without sequencing facilities.  相似文献   

2.
Fish are commonly infected with non‐tuberculous mycobacteria (NTM), which should be regarded as potential pathogens when handling aquarium fish and equipment. This study examined 107 aquarium fish from pet shops. Cultivation of the fish samples using different selective media was conducted for identification of NTM. Isolates were identified using the GenoType Mycobacterium common mycobacteria and additional species assays, sequencing of the 16S rRNA and rpoB genes, and real‐time PCR assay for identification of Mycobacterium (M.) marinum. Among the investigated fish, 79.4% (85/107) were positive for mycobacteria, with 8.2% (7 of 85) having two mycobacterial species present. Among the positive fish, the common pathogens M. marinum, Mycobacterium fortuitum (M. fortuitum group) and Mycobacterium chelonae were identified in approx. 90% of fish and other NTM species in 10%, including Mycobacterium peregrinum/septicum, Mycobacterium gordonae, Mycobacterium arupense, Mycobacterium kansasii, Mycobacterium ulcerans and Mycobacterium setense. The well‐known human pathogen M. marinum was present in 10.6% of the positive fish (9 of 85). The species of mycobacteria identified in the study are not only recognized as aquarium fish pathogens, but can also cause pathology in humans. Microbiological and clinical communities should therefore be sensitized to the role of NTM in infections associated with exposure to aquarium fish.  相似文献   

3.
Based on microbiological and histopathological examinations and DNA sequencing, several outbreaks of mycobacteriosis in the reared sturgeons, including Chinese sturgeon (Acipenser sinensis Gray) and Amur sturgeon (Acipenser schrencki), were identified during 2009 to 2010. Forty‐nine isolates of non‐tuberculous mycobacteria(NTM)were isolated from 19 diseased sturgeons. In total, seven species of Mycobacterium were identified, namely, Mycobacterium chelonae, Mycobacterium marinum, Mycobacterium gordonae, Mycobacterium fortuitum, Mycobacterium szulgai, Mycobacterium arupense and Mycobacterium porcinum. Among them, M. marinum was found to be more prevalent (89.5%) compared with the other mycobacterial species. When two molecular biological methods, PCR‐DGGE (denaturing gradient gel electrophoresis) analysis and rpoB gene library sequencing, were used to analyse the mycobacterial DNAs extracted from the diseased fish tissues, mixed infections of two or three mycobacterial species were found being the predominant infection form (94.7%) in sturgeon mycobacteriosis. M. marinum was the only one species that caused sturgeon mycobacteriosis alone. Virulence assay showed that M. marinum possessed stronger pathogenicity to zebrafish killing 100% of fish in 28 days at 103 cfu/fish than the other species. These results suggested that M. marinum is the major pathogenic bacteria in sturgeon mycobacteriosis. To the best of our knowledge, this study is the first report on mycobacteriosis in farmed Chinese and Amur sturgeons as well as the first isolation of M. porcinum and M. arupense from fish.  相似文献   

4.
Zebrafish (Danio rerio) are a popular model organism used in a growing number of research fields. Maintaining healthy, disease‐free laboratory fish is important for the integrity of many of these studies. Mycobacteriosis is a chronic bacterial infection caused by several Mycobacterium spp. and is the second most common disease found in laboratory zebrafish. Current mycobacteriosis control measures recommend the removal of infected fish and in severe outbreaks, depopulation. These measures can be effective, but less disruptive measures should be assessed for controlling mycobacteriosis, particularly when valuable and rare lines of fish are affected. Here, the in vivo efficacy of two drug candidates, tigecycline (1 μg g?1) and clarithromycin (4 μg g?1), was tested in adult zebrafish experimentally infected with Mycobacterium chelonae. We assessed both short (14 day)‐ and long‐term (30 day) treatments and evaluated fecundity and pathological endpoints. Fecundity and histology results show that zebrafish tolerated antibiotics. Antibiotic treatments did not significantly impact the prevalence of acid‐fast granulomas; however, the severity of infections (acid‐fast granuloma intensity) was significantly decreased following treatments.  相似文献   

5.
Mycobacteriosis in fish is a chronic progressive ubiquitous disease caused by Mycobacterium marinum, M. gordonae and M. fortuitum in most cases. The aim of this study was to describe the morphology and distribution of lesions in 322 freshwater ornamental fish across 36 species. Granulomatous inflammation was diagnosed by gross examination and histopathology testing in 188 fish (58.4%); acid‐fast rods (AFR) were determined in only 96 (51.1%) fish from 19 species after Ziehl–Neelsen staining. The most often affected organs with AFR were the kidney (81.2%), digestive tract (54.1%), liver (48.2%), spleen (45.9%) and skin (21.2%); sporadically, AFR were found in the branchiae (9.4%) and gonads (4.7%). In 14 randomly selected fish originating from four different fish tanks, the distribution of mycobacterial infection was studied by culture examination of the skin, gills, muscle tissue, digestive tract, liver, spleen and kidney. In 12 fish, the species M. marinum, M. gordonae, M. fortuitum, M. triviale, and M. avium subsp. hominissuis (serotypes 6 and 8 and genotype IS901? and IS1245+) were detected; mixed infection caused by different mycobacterial species was documented in five of them.  相似文献   

6.
The isolation of mycobacteria from field samples is problematic, and isolation of the bacterium is sometimes not even attempted. The detection of mycobacteria through traditional histology using formalin‐fixed, paraffin‐embedded (FFPE) tissues is neither sensitive nor specific. However, detection of mycobacterial DNA from FFPE specimens, suspected of being infected with mammalian mycobacteriosis, is a routine clinical procedure. In the present study, a polymerase chain reaction (PCR)‐based method was used to detect and identify mycobacteria in FFPE specimens sampled from fish suspected of being infected with fish mycobacteriosis. A total of 45 fish tissue samples, comprising of 12 tissue samples obtained from experimentally infected fish and the remainder from fish naturally infected with mycobacteria, were analysed using a PCR protocol which amplifies a fragment of the mycobacterial 65 kDa heat‐shock protein (hsp65) gene. PCR‐restriction enzyme analysis and/or sequencing were employed to further analyse the PCR amplicons. The PCR results were compared with those obtained by histology and culture. Mycobacterial DNA was detected in 34 of the 45 samples examined, of which 16 samples (47%) showed granulomatous reactions on histological examination. Using histology as the gold standard, no false‐negative PCR results were obtained. Also, considering the presence or absence of granulomas as a diagnostic criterion, the sensitivity and specificity of PCR in 42 of the FFPE tissues were 16/16 (100%) and 8/26 (～30.8%), respectively. Corresponding microbiological cultures were available for 15 cases, of which 13 were pure Mycobacterium cultures. Of these, 13 were PCR positive (100% sensitivity and 50% specificity). The PCR‐based methods used here proved sensitive, specific and rapid for the detection of mycobacteria in routinely processed paraffin wax‐embedded and formalin‐fixed histological samples, and the results of the study suggest that this method has potential use in retrospective epidemiological studies.  相似文献   

7.
A real‐time PCR assay using a molecular beacon was developed and validated to detect the vapA (surface array protein) gene in the fish pathogen, Aeromonas salmonicida. The assay had 100% analytical specificity and analytical sensitivities of 5 ± 0 fg (DNA), 2.2 × 104 ± 1 × 104 CFU g?1 (without enrichment) and 40 ± 10 CFU g?1 (with enrichment) in kidney tissue. The assay was highly repeatable and proved to be robust following equivalency testing using a different real‐time PCR platform. Following analytical validation, diagnostic specificity was determined using New Zealand farmed Chinook salmon, Oncorhynchus tshawytscha (Walbaum), (n = 750) and pink shubunkin, Carassius auratus (L.) (n = 157). The real‐time PCR was run in parallel with culture and all fish tested were found to be negative by both methods for A. salmonicida, resulting in 100% diagnostic specificity (95% confidence interval). The molecular beacon real‐time PCR system is specific, sensitive and a reproducible method for the detection of A. salmonicida. It can be used for diagnostic testing, health certification and active surveillance programmes.  相似文献   

8.
Multiple greyish‐white visceral nodules containing abundant rapidly growing and acid‐fast bacteria, subsequently identified as Mycobacterium salmoniphilum, were detected in moribund and newly dead market‐sized fish during a period of increased mortality in an Atlantic salmon, Salmo salar, farm in western Norway. Isolates cultured from diseased fish were phenotypically consistent with Mycobacterium sp. previously isolated from Atlantic salmon [MT 1890 (= NCIMB13533), MT1892, MT1900 and MT1901] in the Shetland Isles, Scotland. Partial sequences of 16S rDNA, ribosomal RNA internal transcribed spacer (ITS1), 65‐kDa heat‐shock protein (Hsp65) and β subunit of RNA polymerase (rpoB) revealed 97‐99% similarity with M. salmoniphilum type strain ATCC 13758T. The source of infection was not confirmed. Koch’s postulates were fulfilled following experimental challenge of Atlantic salmon with field isolate NVI6598 ( FJ616988 ). Mortality was recorded in experimentally infected fish; however, the infection remained subclinical in the majority of affected fish over the 131‐day challenge period.  相似文献   

9.
The aim of this study was to screen Bacillus strains from the guts of Scophthalmus maximus, Paralichthys olivacues, Epinephelus coioides and Clupanodon punctatus, for use as probiotics in aquaculture. Eight Bacillus strains were screened, and strain M001 was selected for probiotic study based on its antagonistic activity against multiple aquatic bacterial pathogens including Vibrio anguillarum, V. campbellii, V. vulnificus, V. parahamolyticus, Streptococcus sp. and Edwardsiella tarda. M001 was identified as B. amyloliquefaciens based on the biochemical tests and 16S rRNA gene analysis. In vitro experiments revealed that M001 was able to grow at a wider range of temperature, pH and salinity and was capable to use turbot mucus as nutrient for growth. Additionally, M001 isolate greatly inhibited the growth of V. anguillarum by producing antibacterial substances and was acid tolerance, non‐antibiotic resistance and non‐harmful. Thereafter, the potential probiotic effect of M001 was tested in turbot by dietary administration of M001 at a dose of 108 CFU g?1 diet for 42 days. No significant differences of weight gain, specific growth rate and feed ratio were found in the M001‐diet group of fish compared with control fish, but which increased, respectively, by 5.5%, 4.7% and 7.0% after 42 days of feeding. Several digestive enzyme activities were found to increase significantly in the M001‐diet group, including protease and amylase activities in hepatopancreas, protease activity in intestine and lipase activity in stomach (P < 0.05). Sera superoxide dismutase activity and total protein content (P < 0.05) were also increased significantly (P < 0.05) in the M001‐diet group. The challenge experiment showed that the M001‐diet group of fish showed a relative per cent of survival of 62.7% against V. anguillarim infection. The Bacillus M001 identified from this study has good potential to provide vibriosis control as probiotic feed additive for turbot aquaculture.  相似文献   

10.
It is hypothesized that the frequency of bacterial coldwater disease outbreaks can be reduced through the detection of the aetiologic agent, Flavobacterium psychrophilum, in broodstock followed by culling of eggs from heavily infected broodstock. Before a culling programme can be instituted, however, it is necessary to determine the sensitivity and specificity of existing assays for the detection of F. psychrophilum. In this study, tissue and ovarian fluid samples were collected from 224 fish at five hatcheries and screened using an enzyme‐linked immunosorbent assay (ELISA), a membrane‐filtration fluorescent antibody test (MF‐FAT), bacteriological culture and nested PCR. Latent class analysis was used to estimate sensitivity and specificity of kidney culture, kidney ELISA, nested PCR and MF‐FAT. Analytical sensitivity of the ELISA varied but was greatest when bacteria were cultured under iron‐limiting conditions. Diagnostic sensitivity estimates ranged from 0.02 (kidney culture) to 0.97 (kidney ELISA). Specificity estimates ranged from 0.02 (MF‐FAT) to 0.98 (kidney ELISA). In a separate challenge experiment, the ELISA confirmed the presence of F. psychrophilum in sub‐clinically infected fish. Results from this study demonstrate that the ELISA is an appropriate tool to screen broodstock and provides an indication of infection severity, which is crucial for implementation of a screening/culling programme.  相似文献   

11.
Furunculosis, a septicaemic infection caused by the bacterium Aeromonas salmonicida subsp. salmonicida, currently causes problems in Danish seawater rainbow trout production. Detection has mainly been achieved by bacterial culture, but more rapid and sensitive methods are needed. A previously developed real‐time PCR assay targeting the plasmid encoded aopP gene of A. salmonicida was, in parallel with culturing, used for the examination of five organs of 40 fish from Danish freshwater and seawater farms. Real‐time PCR showed overall a higher frequency of positives than culturing (65% of positive fish by real‐time PCR compared to 30% by a culture approach). Also, no real‐time PCR‐negative samples were found positive by culturing. A. salmonicida was detected by real‐time PCR, though not by culturing, in freshwater fish showing no signs of furunculosis, indicating possible presence of carrier fish. In seawater fish examined after an outbreak and antibiotics treatment, real‐time PCR showed the presence of the bacterium in all examined organs (1–482 genomic units mg?1). With a limit of detection of 40 target copies (1–2 genomic units) per reaction, a high reproducibility and an excellent efficiency, the present real‐time PCR assay provides a sensitive tool for the detection of A. salmonicida.  相似文献   

12.
In the present study a multiplex real-time PCR method was developed for early detection of diseased fish infected by Aeromonas salmonicida, Vibrio anguillarum, and/or Tenacibaculum maritimum. The method consisted of the detection of three species-specific genes after DNA extraction with a commercial kit. Three types of samples were tested, and the results were compared with those of traditional diagnosis. The method obtained a limit of detection of 104 cfu/mL (2 x 102 cfu/tube). Additionally, 27 samples from fish showing signs of disease were correctly diagnosed by the developed methodology, demonstrating its suitability for implementation in aquaculture.  相似文献   

13.
This study investigated the effect of the replacement of fish oil (FO) with DHA‐Gold (DHA‐G)‐supplemented plant oils (PO) in rainbow trout fed plant‐protein‐based diets. Five diets (450 mg g?1 digestible protein and 150 mg g?1 crude lipid) were fed to rainbow trout (initial weight 37 ± 0.5 g) for 12 weeks in a 15 °C recirculating water system. The lipid inclusion types and levels were FO, PO and PO with DHA‐G supplemented at 30 mg g?1, 60 mg g?1 or 90 mg g?1 of the diet replacement for corn oil. Fish fed 90 mg g?1 DHA‐G were significantly larger and consumed more feed than fish‐fed PO or FO (218 g and 2.6% bwd?1 versus 181 g and 2.4% and 190 g and 2.3%, respectively). Feed conversion ratio was significantly increased in fish fed 90 mg g?1 DHA‐G (0.99) as compared to fish‐fed FO (0.90) and 30 mg g?1 DHA‐G (0.91). Panellists found trout fillets from fish fed the 90 mg g?1 DHA‐G diet to have significantly fishier aroma and flavour than fish fed the FO diet. Fatty acid analysis demonstrated that 60 mg g?1 or 90 mg g?1 DHA‐G supplementation increased PO fed fish fillet DHA to fatty acid levels equivalent or higher than those fish fed a FO diet.  相似文献   

14.
This study was aimed at quantifying methionine requirement of Indian major carp fry, Cirrhinus mrigala (2.2 ± 0.2 cm; 0.19 ± 0.02 g) by conducting a 12‐week feeding trial. Casein–gelatine‐based isonitrogenous (40 g 100 g?1 crude protein) and isoenergetic (15.42 kJ g?1 DE) amino acid test diets were prepared to contain six levels of l ‐methionine (1.1, 1.3, 1.5, 1.7, 1.9 and 2.1 g 100 g?1 dry diet) at a fixed level of cysteine (0.85 g 100 g?1 dry diet) and fed to apparent satiation thrice daily to triplicate groups of fish. When absolute weight gain (g per fish), feed conversion ratio, protein deposition (g per fish) and nitrogen retention efficiency data were subjected to broken‐line and second‐degree polynomial regression analysis, 95% of the plateau of above parameters was achieved at dietary methionine concentrations between 1.60 and 1.69 g 100 g?1 dry diet or 0.10 to 0.11 g methionine kJ?1 DE, corresponding to 4.1–4.22 g 100 g?1 protein or 0.44–0.47 g methionine kJ?1 DE. Based on these results, dietary methionine requirement of fry C. mrigala is recommended 1.60–1.69 g 100 g?1 diet or 0.10–0.11 g methionine kJ?1 DE.  相似文献   

15.
Sanguinarine (SA), with antimicrobial and antiparasitic activities against fish pathogens, exhibits great potential commercial use in aquaculture. However, little information on pharmacokinetics of SA restricts further application in aquaculture. In this study, pharmacokinetics of SA in common carp (Cyprinus carpio) following a single intraperitoneal administration [10 mg kg?1 BW (body weight)] was evaluated by high‐performance liquid chromatography (HPLC). The peak concentration (Cmax) of SA in kidney was 11.8 μg g?1, which was higher than in other tissues and plasma. The terminal half‐life in fish tissue and plasma was as follows: 42.3 h (kidney) > 37.2 h (liver) > 20.1 h (gill) > 18.8 h (muscle) > 10.9 h (spleen) > 10.0 h (plasma). Additionally, we determined the bacterial loads in tissues of common carp infected with Aeromonas hydrophila after i.p. administration of SA at 0, 5, 10 and 20 mg kg?1 BW. The results showed that i.p. administration of SA at 10 mg kg?1 BW significantly enhanced antibacterial efficacy against A. hydrophila, where the antibacterial ratio in the gill, kidney, spleen and liver on day 5 was 95.13%, 93.33%, 90.09% and 92.82%, respectively. Overall, these results suggested the potential of SA to treat A. hydrophila infection in common carp farming industry.  相似文献   

16.
Vibrio anguillarum is an aggressive and halophilic bacterial pathogen most commonly originating from seawater. Vibrio anguillarum presence in fisheries and aquaculture facilities causes significant morbidity and mortality among aquaculture species primarily from haemorrhaging of the body and skin of the infected fish that eventually leads to death, collectively recognized as the disease vibriosis. This study served to develop a non‐probe, multiplex real‐time PCR assay to rapidly detect V. anguillarum presence in seawater. Specific primers targeting genes vah1, empA and rpoN of V. anguillarum were selected for multiplex reaction among 11 different primer sets and the extension step was eliminated. Primer concentration, denaturation time as well as annealing time and temperature of DNA amplification were optimized, thus reducing reaction duration. The two‐step, non‐probed multiplex real‐time PCR set forth by this study detects as little as 3 CFU mL?1 of V. anguillarum presence in sea water, without enrichment cultivation, in 70 min with molecular precision and includes melting curve confirmation.  相似文献   

17.
The present study aimed to investigate leucocyte responses to inflammation as well as some innate immune parameters of Senegalese sole, Solea senegalensis, following challenge with two strains of Photobacterium damselae subsp. piscicida belonging to the European and Japanese clones described for this bacterium. Pathogenicity assays were performed to assess the virulence of each Photobacterium damselae subsp. piscicida strain for sole. Subsequently, fish were intraperitoneally injected with phosphate‐buffered saline (control) or two concentrations (2 × 102 and 2 × 106 CFU mL?1) of each bacterial strain and sampled after 6 and 24 h. Results showed that the European isolate induces a higher degree of response than the Japanese strain. While blood neutrophilia and monocytosis correlated well with the increase in neutrophil and macrophage numbers in the peritoneal cavity, fish infected with the European isolate presented higher peritoneal cell numbers than fish challenged with the Japanese strain. In addition, alternative complement pathway activity and respiratory burst of head kidney leucocytes increased significantly in fish infected with the European isolate. The enhanced innate immune response displayed by Senegalese sole challenged with the European isolate is probably due to the higher degree of virulence presented by this Photobacterium damselae subsp. piscicida strain.  相似文献   

18.
This study evaluated the effect of sublethal doses of antifungal drug miconazole nitrate (MCZ) on immunological responses and its role as a prophylactic drug against S. parasitica in Labeo rohita fingerlings. Fish were fed with sublethal doses of MCZ, that is, T1—6.30 mgMCZ kgBW?1, T2—12.61 mgMCZ kgBW?1 and T3—25.22 mgMCZ kgBW?1, and sampling was done at different time intervals for 240 hr. Immunological parameters viz. lysozyme activity, oxygen radical production and plasma antiprotease activity showed significant enhancement (p < 0.05) in fish fed with T2 and T3 doses. Expression of immune‐relevant genes such as TLR‐22 and β2‐M showed significantly higher expression at 6 hr and 24 hr of sampling in both liver and head kidney. However, these genes showed a downregulation after 120 hr of sampling in both the tissues. Preventive efficacy study showed that single dose of MCZ provides protection against oomycetes up to the fourth day of infection. Significantly higher mortality was observed in control diet‐fed fish as compared to fish fed with MCZ medicated diet. Thus, it can be concluded that the MCZ can act as a potent antifungal agent for preventing oomycetes infection as well as to enhance the immune response.  相似文献   

19.
Asian seabass (Lates calcarifer) has been recognized as an economically important aquaculture species which can be adapted to and cultivated in wide range of salinities. The number of freshwater intensive seabass farms in Thailand is increasing annually. Here, we first describe the susceptibility of Asian seabass, which were cultured in freshwater, to Streptococcus inae (SI) and their pathological changes. Three isolates of putative SI were identified using a combination of standard biochemical assays and species‐specific PCR prior subjected to in vivo challenge. Accumulated mortalities of the fish which received 107 CFU fish?1 of either SI1J, SI SGSA or SI2J were 90%, 90% and 100% at 7 days‐post infection (dpi), respectively, and mortalities increased sharply between 3 and 5 dpi. Clinical signs such as erratic swimming and opaque eyes were identified from a few infected fish, while most died rapidly without any abnormal signs. Histopathological manifestations were observed in the multiple organs (kidney, liver and brain). Haemorrhage, hyperhemia, cellular degeneration and inflammatory cells infiltration were commonly found within the internal organs. Notably, the formation of numerous encyst‐like lesion aggregated by eosinophilic cells, resembling macrophages, were typically found in the brain of the infected fish. Summarily, this study first revealed that freshwater reared Asian seabass is highly susceptible to SI infection and haemorrhagic septicaemia was a major pathological change that could be found in the infected fish.  相似文献   

20.
A 35 days feeding trial was conducted to assess the haemato‐immunological response of Labeo rohita fingerlings fed ethanolic leaf extracts of Psidium guajava and Mangifera indica, and infected with Aeromonas hydrophila. Six iso‐nitrogenous (354.6–361.6 g kg?1) purified diets were prepared with graded level of leaf extracts viz., control (basal feed without any extract); TG‐5 (5 g kg?1 guava extract); TG‐10 (10 g kg?1 guava extract); TM‐5 (5 g kg?1 mango extract); TM‐10 (10 g kg?1 mango extract); and TGM (5 g kg?1 guava extract +5 g kg?1 mango extract). Haematological, immunological, biochemical, along with antioxidant enzyme activities were examined after a 35 day‐feeding trial and following a 7 day challenge with A. hydrophila. The haemoglobin, total leucocyte and erythrocyte counts, respiratory burst activity, lysozyme, total protein, albumin and globulin contents increased significantly (P < 0.05) in leaf extracts fed groups compared with the control in pre‐ and post‐challenge conditions. A significant (P < 0.05) decrease was observed in SOD (superoxide dismutase) and catalase activities of the treatment groups compared with the higher value in control. The trends in mortality indicated that groups of fish showing significantly elevated haemato‐immunological responses had the lowest mortality following challenge with A. hydrophila. The results showed that extracts of P. guajava and M. indica appear to be potential immunostimulant at an inclusion level of 5 g kg?1 in the diet of rohu. But, mixing of both the extract at similar level did not show any synergistic effect, which needs to be tested at its lower level of inclusion.  相似文献   

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