Stimulatory effect of dietary clove,Eugenia caryophyllata,bud extract on growth performance,nutrient utilization,antioxidant capacity,and tolerance of African catfish,Clarias gariepinus (B.), to Aeromonas hydrophila infection

The occurrence of pathogenic bacteria, Aeromonas hydrophila, in farm‐raised fish requires urgent attention. Continuous and indiscriminate use of antibiotics as growth promoters and disease control agents in aquaculture have been discouraged because of the risk of development of antibiotic‐resistant bacterial strains. There is steady interest in the use of botanicals, such as clove, Eugenia caryophyllata, buds extract (ECBE), as alternatives. Hence, the present study evaluated the effect of dietary ECBE supplementation on the growth performance, physiological, antioxidant, and immunity biomarkers of African catfish, Clarias gariepinus. Fish (11.7 ± 0.5 g) were fed diets containing 0.0 (control), 5.0, 10.0, or 15.0 g ECBE/kg diet up to apparent satiation twice daily for 12 weeks. After the feeding trial, fish from each treatment were challenged with A. hydrophila infection by intraperitoneal injection and kept under observation for 14 days to record any abnormal clinical signs and daily mortality. The results demonstrated that fish performance and feed intake were significantly enhanced with increasing ECBE levels, and its optimum level is 15 g/kg diet. Further, the dietary ECBE increased significantly the intestinal villi length/width and absorption area in a dose‐dependent manner. There are significant progressive increases in the values of red blood cells, hemoglobin, hematocrit, white blood cells, platelets, lymphocytes, and heterocytes, while monocytes, eosinophil, and basophils decreased significantly due to dietary ECBE in a dose‐dependent manner. Highest glucose, cholesterol, total protein, globulin, and albumin‐globulin ratios, aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), urea, and creatinine values were found in fish fed 15 g ECBE/kg diets, while lowest values were recorded in fish fed the control diet. Despite the high AST and ALT values, no visible lesions or damage were observed in the liver cells of fish fed ECBE‐enriched diets. In addition, the inclusion of ECBE in fish diets enhanced the antioxidant and immunity capacity. Fish mortality after the bacterial challenge was higher in fish fed the control diet (82.3%) than those fed ECBE‐enriched diets. The lowest fish mortality was observed in fish fed the 15 ECBE/kg diet (4.7%) [Correction added on 16 November 2018: this section has been revised for clarity.].     

Control of tomato bacterial wilt and root-knot diseases by Bacillus thuringiensis CR-371 and Streptomyces avermectinius NBRC14893

Ralstonia solanacearum and Meloidogyne incognita are two soilborne pathogens that cause serious damage and great losses in the production of tomato. For this purpose, a bacterial isolate, Bacillus thuringiensis CR-371, and an actinomyces isolate, Streptomyces avermectinius NBRC14893, were examined for their ability to protect tomato from root-knot nematode and bacterial wilt diseases under glasshouse conditions. Treatment of tomato roots with B. thuringiensis CR-371 and S. avermectinius NBRC14893 followed by challenge inoculation with R. solanacearum and M. incognita significantly decreased disease severity of bacterial wilt alone, root-knot nematode alone, or mixed infection by both pathogens compared to the control. Furthermore, pretreatment of tomato roots with B. thuringiensis CR-371 and S. avermectinius NBRC14893 significantly reduced bacterial proliferation of R. solanacearum both in pathogen alone inoculated plants and in plants co-inoculated with R. solanacearum and M. incognita. In conclusion, our results suggest that the treatment of tomato roots with B. thuringiensis CR-371 and S. avermectinius NBRC14893 simultaneously suppresses bacterial wilt and root-knot nematode diseases. Therefore, B. thuringiensis CR-371 and S. avermectinius NBRC14893 could provide new options for integrated pest management strategies against plant diseases, especially against bacterial-nematode disease complexes that cause synergistic yield losses.     

Monitoring pyrethroid insecticide resistance in major malaria vector Anopheles culicifacies: comparison of molecular tools and conventional susceptibility test

BACKGROUND: Anopheles culicifacies is a main malaria vector in southeastern part of Iran, bordring Afghanistan and Pakistan. So far, resistance to DDT, dieldrin, malathion and partial tolerance to pyrethroids has been reported in An. stephensi, but nothing confirmed on resistance status of An. culicifacies in Iran. METHODS: In current study, along with WHO routine susceptibility test with DDT (4%), dieldrin (0.4%), malathion (5%), permethrin (0.25%), lambadacyhalothrin (0.1%), and deltamethrin 0.025, we cloned and sequenced segment VI of domain II (SII6) in voltage-gated sodium channel (vgsc) gene of An. culicifacies specimens collected in Sistan and Baluchistan province (Iran). RESULTS: A 221-bp amplified fragment showed 91% and 93% similarity with exon I and exon II of An. gambiae. The size of intron II in An. culicifacies is 62 bp, while in An. gambiae is 57 bp. The major difference within An. culicifacies specimens and also with An. gambiae is in position 29 of exon I, which led to substitution of Leu to His amino acid. CONCLUSION: This data will act as first report on partial sequence of vgsc gene and its polymorphism in An. culicifacies. A Leu to His amino acid substitution detected upstream the formerly known knockdown resistance (kdr) mutation site could be an indication for other possible mutations related to insecticide resistance. However, the result of WHO susceptibility test carried out in Baluchistan of Iran revealed a level of tolerance to DDT and dieldrin, but almost complete susceptibility to pyrethroids in An. culicifacies. We postulate that the molecular diagnostic tool developed for detection and identification of kdr-related mutations in An. culicifacies, could be useful in monitoring insecticide resistance in Iran and neighbouring countries such as Pakistan and Afghanistan. A phylogenetic tree also constructed based on the sequence of exon I and II, which readily separated An. culicifacies populations from An. stephensi, An. fluviatilis and An. gambiae.     

Effect of esophageal distention on basal and stimulated gastric acid secretion in rats

BACKGROUND: It is well established that the esophageal distention (ED) leads to gastric relaxation, partly by vago-vagal reflex, but till now, the effect of ED on gastric acid secretion has not been investigated. The aim of this study was to investigate the effect of ED on basal and stimulated gastric acid secretion. METHODS: Adult male Wistar rats (200-240 g) were deprived of food but not the water 24 h before the experiments. Under urethane anesthesia (1.2 g/kg, i.p.), animals underwent tracheostomy and laparotomy. A catheter was inserted in the stomach through duodenum for gastric distention and gastric washout and the esophagus was cannulated with a distensible balloon orally to distend esophagus (0.3 ml, 10 min). Gastric acid secretion was stimulated by gastric distention, carbachol (4 microg/kg, i.p.) or histamine (5 mg/kg, s.c.). Effects of vagotomy, NG-nitro-L-arginine methyl ester (L-NAME, 10 mg/kg, i.v.) and also hexamethonium were investigated. RESULTS: Basal and gastric distention- and carbachol, histamine-stimulated acid secretion were reduced by the ED (P<0.05, P<0.0001, P<0.01 and P<0.02, respectively). L-NAME (10 mg/kg, i.v.) elevated the acid output (P<0.002). Vagotomy reduced the inhibitory effect of the esophagus distention on gastric distention-induced acid secretion (P<0.01). CONCLUSION: These results indicate that the vagus nerves are involved in the inhibitory effect of the ED on the basal and stimulated gastric acid secretion. Furthermore, nitric oxide could be involved.     

Comparative effects of copper, iron, vanadium and titanium on low density lipoprotein oxidation in vitro

INTRODUCTION: Oxidation of low density lipoprotein (LDL) has been strongly implicated in the phathogenesis of atherosclerosis. The use of oxidants in dietary food stuff may lead to the production of oxidized LDL and may increase both the development and the progression of atherosclerosis. The present work investigated the effects of some elements including: copper (Cu), iron (Fe), vanadium (V) and titanium (Ti) on in vitro LDL oxidation quantitatively. METHODS: The first LDL fraction was isolated from fresh plasma by single vertical discontinuous density gradient ultracentrifugation. The formation of conjugated dienes and thiobarbituric acid reactive substances and increase in electrophoretic mobility of LDL were monitored as markers of the oxidation of LDL. RESULTS: It was demonstrated that Cu, Fe, V and Ti exhibited strong oxidant activity in this respect (P<0.001). Oxidation of LDL in the presence of Cu was more and appeared to be in this order Cu>Fe>V>Ti. DISCUSSION: Cu, Fe, V and Ti are redox-active transition metals that may cause oxidative damage to lipids, proteins and DNA molecules. We suggest that these elements may also influence the oxidation of LDL in vivo, which could increase both the development and progression of atherosclerosis.     

Effects of grazing on natural regeneration of tree and herb species of Kheyroud forest in northern Iran

We investigated the effects of grazing on natural regeneration, quantity, and diversity of woody species and dominant herb species in Kheyroud forest in northern Iran. We sampled vegetation in 5m2 plots in custom units, which are demarcated resource areas traditionally used by local livestock producers. The authors quantified number of species, height of seedlings, and diameter of seedlings. Height classes were 0-30 cm, 30-130 cm, and >130 cm, and diameter classes were 0-2.5 cm, 2.5-5 cm and 5-7.5 cm. The density of seedlings declined with distance from corral until reaching the custom unit boundary. Most seedlings had diameters of 0-2.5 cm and heights of 0-30 cm. Predominant species, Carpinus betulus and Acer capadocicum, were in plots near the centers of custom units, Fagus orientalis, Acer velutinum, Quercus castanifolia species were dominant in plots near the custom unit boundary. Plant species such as Oplismenus undulatifolius, Euphorbia amygdaloides, Rubus fruticos and Pteridium aquilinum were dominant in plots nearer to forest corral. Healthy seedlings were more numerous in plots nearest the corral, while defective and deformed seedlings were more abundant away from the corral. We conclude that grazing had negative effects on the quantity and quality of vegetative regeneration. Continuation of overgrazing will not only endanger the sustainability of forest ecosystems, but also will increase the challenge of sustainable forest management.     

Construction of a genetic linkage map with SSR,AFLP and morphological markers to locate QTLs controlling pathotype-specific powdery mildew resistance in diploid roses

Disease resistance is a sought-after trait in plant breeding programmes. One strategy to make resistance more durable is to increase the number of resistance genes, thereby increasing the number of pathotypes withstood. One of the most important diseases on roses is powdery mildew (PM) (Podosphaera pannosa). Recent studies show that pathotypes of PM and different types of resistances in roses exist. The results of this study aim to contribute to PM resistance in roses by the development of pathotype-specific markers on a genetic map. A diploid rose population (90 genotypes) derived from a cross between Rosa wichurana and Rosa ‘Yesterday’ was used to construct a genetic linkage map encompassing 20 AFLP primer combinations, 43 SSR, and 2 morphological markers. By applying the F1 pseudo test cross population strategy, two parental linkage maps were constructed (parent ‘Yesterday’ 536 cM; parent R. wichurana 526 cM). Both parental maps consisted of seven linkage groups with an average length of 70 cM (Kosambi) corresponding to the seven haploid rose chromosomes. These new maps were used to identify QTLs controlling disease resistance. The offspring population was screened for resistance to two PM pathotypes, R–E and R–P. QTLs for controlling pathotype-specific disease resistance were mapped by applying Kruskal–Wallis rank-sum tests and simple interval mapping. With two pathotypes analysed, nine QTL loci were detected on linkage groups 2, 3, 5 and 6, explaining 15–73% of the phenotypic variance for pathotype-specific disease response. The genetic maps developed here will be useful for future rose breeding, pathotype-specific resistance research and development of a consensus map for roses.     

Immunohistochemical and polymerase chain reaction studies in Neospora caninum experimentally infected broiler chicken embryonated eggs

The diagnostic characteristics of immunohistochemistry (IHC) and polymerase chain reaction (PCR) methods were studied in the tissues of broiler chicken embryos experimentally infected by Neospora caninum. An infection with N. caninum NC-1 isolate was conducted in 70 broiler chicken embryonated eggs randomly divided into seven equal groups. After 8 days of incubation, six groups were inoculated with 10, 10(2), 10(3), 10(4), 10(5), and 10(6) doses of tachyzoites/embryonated egg. The 7th group was considered as control. The mortality rate and pathological changes of the dead embryos and hatched chickens up to 60 days old were noticed. Consecutive sections to those used for histopathological examination including the liver, heart, brain, and chorioalantoic (CA) membrane were subjected to IHC. The intensity and distribution of the immunostaining was graded as highly to mildly positive. For PCR procedure, DNA was extracted from 50mg of the tissues and primer pair Np21/Np6 was used for amplification of the Nc-5 gene. The results of the immunosignaling ranged from variable degrees of mild to moderate staining as dark-brown to brown and coarsely to finely granular, mostly within the cytoplasm of infected cells such as the endothelial cells of blood vessels. The parasite aggregation was more predominant in the heart than other tissues. Immunoreactivity for N. caninum antigen was multifocally moderate positive in the heart, liver and CA of the 10(3) dose, and also heart, liver, brain and CA of the 10(4) dose. IHC showed mildly positive in the liver and heart of the chicken embryos infected with 10 and 10(2) tachyzoites, as well. The results of the PCR confirmed the existence of the parasite in all of the examined tissues from the 10(3) and 10(4) doses. In conclusion, the results indicate a good agreement between IHC and PCR in diagnosis of neospora antigen in the infected tissues.     

Identification of genes expressed by Phakopsora pachyrhizi,the pathogen causing soybean rust,at a late stage of infection of susceptible soybean leaves

Soybean is one of the top five agricultural products in the United States and is highly susceptible to Phakopsora pachyrhizi, an exotic obligate biotrophic fungus. The little amount of genomic information about P. pachyrhizi limits understanding of the soybean–soybean rust pathogen interaction and the possibility of engineering resistance to this pathogen in soybean. Illumina mRNA‐Seq analysis revealed P. pachyrhizi genes expressed during a biotrophic interaction between P. pachyrhizi and soybean during fungal sporulation 10 days after inoculation. Approximately 2·4 million DNA sequences representing portions of potential P. pachyrhizi genes were assembled into 32 940 contigs that were used to search against expressed sequence tag (EST), protein and conserved domain databases. About 7500 contigs represent newly discovered P. pachyrhizi sequences. Of these, 527 shared similarity to genes encoding fungal proteins involved in different metabolic pathways such as galactose and glycogen metabolism, glycolysis, the citrate cycle, fatty acid metabolism, amino acid metabolism, proteolysis, protein synthesis, cell cycle division and mitosis, and cell wall biogenesis. Almost 7000 potential P. pachyrhizi genes are still of unknown function. Such information may be useful in the development of new methods of broadening resistance of soybean to P. pachyrhizi, including the silencing of important P. pachyrhizi genes, and also to understand the molecular basis of soybean–P. pachyrhizi interactions.