Phenoloxidase Research Articles

Enhanced resistance to Bacillus thuringiensis Cry1Ac toxin mediated by the activation of prophenoloxidase in a cosmopolitan pest

Plutella xylostella has evolved resistance to Bacillus thuringiensis Cry1Ac toxin over a long evolutionary period. Enhanced immune response is an important factor in insect resistance to a variety of insecticides, and whether phenoloxidase (PO), an immune protein, is involved in resistance to Cry1Ac toxin in P. xylostella remains unclear. Here, spatial and temporal expression patterns showed that prophenoloxidase (PxPPO1 and PxPPO2) in the Cry1S1000-resistant strain was more highly expressed in eggs, 4th instar, head, and hemolymph than those in G88-susceptible strain. The results of PO activity analysis showed that after treatment with Cry1Ac toxin PO activity was about 3 times higher than that before treatment. Furthermore, knockout of PxPPO1 and PxPPO2 significantly increased the susceptibility to Cry1Ac toxin. These findings were further supported by the knockdown of Clip-SPH2, a negative regulator of PO, which resulted in increased PxPPO1 and PxPPO2 expression and Cry1Ac susceptibility in the Cry1S1000-resistant strain. Finally, the synergistic effect of quercetin showed that larval survival decreased from 100 % to <20 % compared to the control group. This study will provide a theoretical basis for the analysis of immune-related genes (PO) genes involved in the resistance mechanism and pest control of P. xylostella.

Assessment of Antioxidant and Polyphenol Profile of Chrysobalanus icaco from Southern Nigeria

Due to the distinguished medicinal value of Chrysobalanus icaco, it was of great interest to&nbsp;conduct phytochemical and antioxidant investigations on it. In this study, the polyphenol profile and antioxidant activity of C. icaco methanol crude extract and fractions were investigated. The crude extract of C. icaco was subjected to fractionation using methanol, n-hexane, ethyl acetate, and n-butanol as solvents. 2, 2-diphenyl-1picryl-hydrazyl (DPPH), 2,2’-azino-bis-(3- ethylbenzothiazoline-6-sulfonic acid) (ABTS) and ferric ion reducing power (FRAP), ascorbate oxidase, total phenolic content, total flavonoids and polyphenol profile was determined using standard methods. A significant increase was observed in the DPPH radical scavenging activity, total flavonoids, total phenol, FRAP, ABTS, TAC and ascorbate oxidase of the crude extract when compared to methanol, ethyl acetate, n-hexane, n-butanol, and aqueous fractions. Polyphenols such as kaempferol, chlorogenic acid, catechin, quercetin, galic acid, pomolic acid, rutin, epicatechin, stigmasterol, sitosterol and caffeic acid were found in the different fractions. The total polyphenols content of different fractions of C icaco extract were as follows; methanol fraction &gt; n-hexane fraction &gt; ethyl acetate fraction &gt; n-butanol fraction &gt; aqueous fraction. The findings revealed that C icaco kernel could be used as a potent source of molecules with positive health attributes.

Evaluation of Phytosterols as an Alternative to Cholesterol in Practical Diets on Growth and Nonspecific Immunity of Litopenaeus vannamei

This study is aimed at evaluating the effectiveness of phytosterols as an alternative to cholesterol in practical diets of Pacific white shrimp Litopenaeus vannamei from the perspective of growth and nonspecific immunity. Five diets were formulated to contain different sterol sources and levels. Two diets were supplemented with 1 g/kg cholesterol (LC (low cholesterol)) or phytosterol (LP (low phytosterol)). Other three experimental diets were supplemented with 2 g/kg cholesterol (HC (high cholesterol)), 2 g/kg phytosterol (HP (high phytosterol)), or mixed sterol source (CP, 1 g/kg cholesterol + 1 g/kg phytosterol), respectively. A total of 750 healthy and uniform-sized shrimp (0.52 ± 0.008 g) were randomly distributed into 5 groups with 3 replicates and fed with the five experimental diets for 60 days. Results showed that the growth performance of shrimp was influenced by the sterol levels and supplementation with 2 g/kg sterol level facilitated the growth of shrimp. The inclusion of phytosterol has a cholesterol-lowering effect on shrimp, as evidenced by a reduction in hemolymph cholesterol and triglyceride contents in the HP group. Besides, supplementation with 2 g/kg phytosterol or mixed sterol sources had positive effects on the hemolymph superoxide dismutase, phenol oxidase, and lysozyme as well as hepatopancreas alkaline phosphatase activities, demonstrating that the nonspecific immunity and antioxidative capacity were improved. In conclusion, phytosterols could be an appropriate alternative to partially replace dietary cholesterol in shrimp feeds. This study preliminarily revealed the effects of different sterol sources and levels on the growth and nonspecific immunity of shrimp and provided a basis for further exploration of the mechanism of phytosterol.

RNAi-mediated CrebA silencing inhibits reproduction and immunity in Locusta migratoria manilensis

Locusta migratoria manilensis is a major agricultural pest that causes severe direct and indirect damage to several crops. Thus, to provide a theoretical foundation for pest control, the role of CrebA in the reproduction and immune regulation of L. migratoria was investigated. CrebA is a bZIP transcription factor that critically regulates intracellular protein secretion. In this study, CrebA was widely expressed in the brain, fat body, integument, midgut, and reproductive tissues of different maturity stages of adult locusts, especially in the female fat body. RNA interfering (RNAi)-mediated silencing of CrebA inhibited locusts ovarian development, and key reproduction gene expressions, Vgs, VgRs, Chico, and JHAMT were downregulated. After the locusts were injected with Micrococcus luteus or Escherichia coli, M. luteus activated lysozyme expression, while the E. coli activated both phenol oxidase cascade and lysozyme expression. Furthermore, both bacteria stimulated the upregulation of the antimicrobial peptide genes DEF3 and DEF4. However, CrebA silencing is fatal to locusts infection with E. coli, with a mortality rate of up to 96.3%, and resulted in a significant decrease in the expression of DEF3 and DEF4 and changes in the activities of phenol oxidase and lysozyme of locusts infected by bacteria. Collectively, CrebA may be involved in diverse biological processes, including reproduction and immunity. CrebA inhibited locusts reproduction by regulating JH signaling pathway and inhibits the expression of immune genes TLR6, IMD, and AMPs. These results demonstrate CrebA seems to play a crucial role in reproduction and innate immunity.

The Co-Invasion of the Three Asteraceae Invasive Plants Can Synergistically Increase Soil Phenol Oxidase Activity

The Co-Invasion of the Three Asteraceae Invasive Plants Can Synergistically Increase Soil Phenol Oxidase Activity

HMG-like DSP1 is a damage signal to mediate the western flower thrips, Frankliniella occidentalis, immune responses to tomato spotted wilt virus infection

Tomato spotted wilt virus (TSWV) causes a serious plant disease and is transmitted by specific thrips including the western flower thrips, Frankliniella occidentalis. The persistent and circulative virus transmission suggests an induction of immune defenses in the thrips. We investigated the immune responses of F. occidentalis to TSWV infection. Immunofluorescence assay demonstrated viral infection in the larval midguts at early stage and subsequent propagation to the salivary gland in adults. In the larval midgut, TSWV infection led to the release of DSP1, a damage-associated molecular pattern, from the gut epithelium into the hemolymph. DSP1 up-regulated PLA2 activity, which would lead to biosynthesis of eicosanoids that activate cellular and humoral immune responses. Phenoloxidase (PO) activity was enhanced following induction of PO and its activating protease gene expressions. Antimicrobial peptide genes and dual oxidase, which produces reactive oxygen species, were induced by the viral infection. Expression of four caspase genes increased and TUNEL assay confirmed apoptosis in the larval midgut after the virus infection. These immune responses to viral infection were significantly suppressed by the inhibition of DSP1 release. We infer that TSWV infection induces F. occidentalis immune responses, which are activated by the release of DSP1 from the infection foci within midguts.

Antixenosis of Thodan (AAB), a resistant Musa cultivar to Odoiporus longicollis (Olivier) and characterization of larvicide molecules

Banana pseudostem borer (BPB) Odoiporus longicollis (Olivier) (Coleoptera, Curculionidae) is a serious pest of Musa cultivars. Experimental maintenance of larvae in the live pseudostem of cultivar Thodan, a resistant AAB Musa cultivar has resulted antixenosis followed by death of larvae within a week. Antixenosis was characterized by significant decrease of total hemocytes and sharp changes on the proportionate distribution of different types of hemocytes in larvae. As the number of plasmatocytes, prohemocytes, splenocytes and adipohemocytes decreased the number of granulocytes and oenocytes increased. Antixenosis also caused accumulation of 20-hydroxyecdysone (20E) and significant inhibition on the activities of trypsin like serine protease (TlSP) and phenoloxidase (PO). Phytochemical analysis of Thodan resulted characterization of three larvicides such as Betulinic acid (BA), Stigmasterol-3-O-glucoside (SOG) and Sulfoquinovosyl diacyl glycerol (SQDG), and the content in the pseudostem ranged 0.0027 to 0.007 per cent. All the three larvicides were highly toxic to the larvae with LD50 of 0.38 ppm for SOG, 0.41 ppm for SQDG and 0.83 ppm for BA. Simultaneous action of three larvicides in the live pseudostem resulted resistance in Thodan against infestation by BPB. Susceptible Nendran showed negligibly low content of SOG (0.0011%) and SQDG (0.0013%) but no detectable quantity of BA. Intoxication by all the three larvicides caused significant changes on the proportionate distribution of hemocytes, accumulation of 20-hydroxy ecdysone (20E) and inhibition of TlSP and PO, the enzymes involved in larval metamorphosis and cuticle sclerotisation. This study demonstrated that resistance of Thodan against O. longicollis is due to adverse effect of these larvicides on endocrine system, cuticle development and cytotoxicity of hemocytes. As these larvicide molecules are stable compounds, there is scope for them to be used as substitutes in place of deleterious insecticides for the management BPB.

Co-application of entomopathogenic fungi with chemical insecticides against Culex pipiens

Culex pipiens (Diptera: Culicidae) is a vector of many human and animal diseases. Its control is regarded as a preventative approach that is focused on effectively managing such diseases. In this context, dose response assays of two insecticides, bendiocarb and diflubenzuron were performed with two entomopathogenic fungi, Beauveria bassiana and Metarhizium anisopliae against 3rd instar C. pipiens larvae. The most effective agents, combination experiments as well as enzymatic activities of phenoloxidase (PO) and chitinase (CHI) were also assessed.The results showed that diflubenzuron was more effective at low concentrations (LC50: 0.001 ppm) than bendiocarb (LC50: 0.174 ppm), whereas M. anisopliae was more effective (LC50: 5.2 × 105 conidia/mL) than B. bassiana (LC50: 7.5 × 107 conidia/mL). Synergistic interactions were observed when diflubenzuron was applied at 2- and 4-days post- exposure to M. anisopliae, with the highest degree of synergism observed when diflubenzuron was applied 2 days post-fungal exposure (χ2 = 5.77). In contrast, additive interactions were recorded with all other insecticide-fungal combinations. PO activities significantly (p ≤ 0.05) increased during 24 h after a single diflubenzuron treatment as well as when diflubenzuron was applied prior to M. anisopliae, whereas suppressed after 24 h when M. anisopliae applied prior to diflubenzuron as well as after 48 h from single and combined treatments. CHI activity increased 24 h after both single and combined treatments, the activity remained elevated 48 h after a single diflubenzuron treatment and when diflubenzuron was applied after M. anisopliae. Histological study of the cuticle by transmission electron microscopy revealed abnormalities following single and combined treatments. Germination of the conidia and production of the mycelium that colonizes the lysing cuticle was obvious when diflubenzuron was applied 48 h after M. anisopliae exposure. Overall, these results demonstrate that M. anisopliae is compatible with diflubenzuron at lower concentrations and that combined applications can improve C. pipiens management.

Effects of Panax notoginseng Water Extract on Immune Responses and Digestive Enzymes in White Shrimp Litopenaeus vannamei

Panax notoginseng (Burk) F. H. Chen is a traditional Chinese herbal medicine commonly used in clinical applications. This study examined the effects of the Panax notoginseng water extract (PNWE) on the immune responses and digestive enzyme activity of Litopenaeus vannamei (L. vannamei). The PNWE (50, 100, and 200 μg (g shrimp)-1) was injected into L. vannamei to analyze the immune response parameters, including the total haemocyte count (THC), granular haemocytes (GC), semi-granular haemocytes (SGC), hialin haemocyte (HC), the respiratory burst (RB), the phagocytic ratio (PR), the phagocytic index (PI), and phenoloxidase (PO). We evaluated the activity of the intestinal digestive enzymes (trypsin, chymotrypsin, amylase, and lipase), the histopathology, and the intestine Vibrio numbers. The results showed that different concentrations of the PNWE significantly increased THC, GC, SGC, PO and RB activity, the PR, and the PI of L. vannamei while reducing the HC. In addition, the PNWE also significantly increased the chymotrypsin, trypsin, and amylase activity of L. vannamei. Furthermore, 50 µg (g shrimp)-1 of PNWE regulated the lipase activity. Additionally, different concentrations of the PNWE significantly reduced the Vibrio numbers in the intestine without damaging the hepatopancreas and intestine tissues. These results indicate that the PNWE improves the immune responses of L. vannamei by increasing the haemocyte count and regulating intestinal digestive enzymes.

The Symbiotic Bacteria-Xenorhabdus nematophila All and Photorhabdus luminescens H06 Strongly Affected the Phenoloxidase Activation of Nipa Palm Hispid, Octodonta nipae (Coleoptera: Chrysomelidae) Larvae.

Symbiotic bacteria form a mutualistic relationship with nematodes and are pathogenic to many insect pests. They kill insects using various strategies to evade or suppress their humoral and cellular immunity. Here we evaluate the toxic effects of these bacteria and their secondary metabolites on the survival and phenoloxidase (PO) activation of Octodonta nipae larvae using biochemical and molecular methods. The results show P. luminescens H06 and X. nematophila All treatments caused significant reductions in the number of O. nipae larvae in a dose-dependent manner. Secondly, the O. nipae immune system recognizes symbiotic bacteria at early and late stages of infection via the induction of C-type lectin. Live symbiotic bacteria significantly inhibit PO activity in O. nipae whereas heat-treated bacteria strongly increase PO activity. Additionally, expression levels of four O. nipae proPhenoloxidase genes following treatment with P. luminescens H06 and X. nematophila All were compared. We found that the expression levels of all proPhenoloxidase genes were significantly down-regulated at all-time points. Similarly, treatments of O. nipae larvae with metabolites benzylideneacetone and oxindole significantly down-regulated the expression of the PPO gene and inhibited PO activity. However, the addition of arachidonic acid to metabolite-treated larvae restored the expression level of the PPO gene and increased PO activity. Our results provide new insight into the roles of symbiotic bacteria in countering the insect phenoloxidase activation system.

Plantations thinning: A meta-analysis of consequences for soil properties and microbial functions

Thinning is a widely-used management practice to reduce tree competition and improve wood production and quality in forest plantations. Thinning affects the soil ecosystem by changing the microclimate and plant growth, as well as litter inputs above and belowground, with all the resulting consequences for microbial communities and functions. Although many case studies have been carried out, a comprehensive understanding of the thinning effects on soil properties and microbial communities and functions in plantations remains to be explored. In this study, a meta-analysis was performed on 533 paired observations based on 90 peer-reviewed articles to evaluate the general responses of soil (mainly 0–20 cm depth) physicochemical properties, microbial biomass and community structure, and enzyme activities to thinning. Results showed that thinning increased soil temperature (13 %), moisture (8.0 %), electric conductivity (13 %), and the contents of total nitrogen (TN, 4.1 %), dissolved organic carbon (DOC, 9.7 %), nitrate N (NO3−–N, 27 %) and available phosphorous (22 %). For microbial properties, thinning decreased the fungi to bacteria ratio (F:B, −28 %) and the gram-positive bacteria to gram-negative bacteria ratio (G+:G−, −12 %), while increased microbial biomass C (7.1 %), microbial respiration (13 %), and nutrient-cycle related enzyme activities, including phenol oxidase (14 %), cellobiohydrolase (21 %), urease (10 %), and acid phosphatase (9 %). In particular, moderate thinning (30–60 % intensity) has higher conservation benefits for soil C and nutrients than light and heavy intensity, thus being recommended as the optimal thinning activity. This meta-analysis suggests that thinning consistently altered soil properties, shifted microbial community compositions from K- to-r strategist dominance, and stimulated microbial activities. These results are essential for optimizing plantation thinning management and provide evidence for applying the macro-ecology theory to ecosystem disturbance in soil microbial ecology.

Effects of parasitism by the braconid wasp, Bracon hebetor (Hymenoptera: Braconidae), on the host hemolymph and phenoloxidase activation of the rice moth, Corcyra cephalonica larvae (Lepidoptera: Pyralidae)

BackgroundBiological control of insect pests is an economic, eco-friendly and harmless approach to integrated pest management strategies. Bracon hebetor (Say) (Hymenoptera: Braconidae) considers a polyphagous ectoparasitoid of various pests of the order Lepidoptera. The parasitized host insects' defense mechanisms are triggered as a result of the parasitic wasps' injury and penetration. Thus, induce the host cellular and humoral immune responses through a blend of secretions injected into the host body. The present study was designed to evaluate the efficacy of B. hebetor on the immune response of the parasitized full grown larval instar of the rice moth, Corcyra cephalonica (Stainton) (Lepidoptera: Pyralidae) under natural envenomation.ResultsOn cellular level, the ultrastructure examination of the hemocytes displayed a considerable structural deformation in hemocyte morphology of the detected types of the hemocytes. Moreover, the effects of parasitism on both differential hemocyte counts (DHCs) and total hemocyte counts (THCs) were investigated. The number of prohemocytes (PR) (40.33 ± 5.61, 43 ± 8.33 and 26 ± 2.31) was statistically differed after (24, 48 and 72 h) of parasitism, respectively, compared to unparasitized (31.33 ± 6.49) larvae. Similar observations were recorded in plasmatocytes (PL) before and after the parasitism. However, Spherulocytes (SP) and Oenocytoids (OE) were recorded in the hemolymph with little abundance. On the other side, the quantitative analysis of total hemolymph proteins (THP) provoked a significant effect of considering parasitized and non-parasitized larvae. As the Phenoloxidase (PO) cascade plays a critical role in immune defenses, so the substantial activation of PO in the host's hemolymph following successive hours of parasitism compared to unparasitized larvae using L-DOPA as a substrate indicated the induction of larval immune system. The parasitized larvae showed a gradual increase in the PO activity (0.442 ± 0.103) after 24 h reached up to (1.482 ± 0.272) at the end of parasitism (72 h) in comparable to unparasitized larvae (0.177 ± 0.0.044).ConclusionThe present investigations clarified the efficacy of B. hebetor parasitism on the host immune mechanism, which will enable the progress of sustainable stored product protection approaches for the control of an important pest rice moth C. cephalonica.

Stimulation of shrimp (Penaeus monodon) hemocytes by lipopolysaccharide-like molecules derived from Novacq™

Immune stimulation through feed additives is a promising strategy that can help to combat disease in shrimp farming and reduce the use of antibiotics and other chemotherapeutics. The present study investigated the in vitro immunostimulatory effects of lipopolysaccharide (LPS)-like molecules isolated from the microbial based feed additive Novacq™ (N-LPS). The presence of LPS-like molecules was confirmed and quantified Novacq™ using a HEK-TLR4 reporter cell line. Primary hemocytes isolated from adult Penaeus monodon were used to measure the immunostimulatory of N-LPS compared with the control group that were treated with E. coli derived LPS (E-LPS). The N-LPS stimulated a rapid and significant induction of the phenoloxidase (PO) response in the hemocytes. The PO response increased with exposure time and LPS concentration and was significantly higher compared with an E. coli LPS (E-LPS) control. In addition, using gene expression data, we quantified the transcriptome response of the hemocytes at 15, 30 and 60 mins post stimulation. Compared with the controls, the N-LPS treated hemocytes had a significant up-regulation of genes involved in the immune system modulation and control at all time-points. Most noteworthy was the significant induction of transcripts that function as serine protease inhibitors (namely SERPINs), that regulate the overexpression of the PO system. Transcription factors from the Notch family which directly regulate the expression of many immune genes were also induced within the hemocytes. Additionally, we also saw a strong up-regulation of crustacean hyperglycemic hormone (CHH) transcripts, an important neuropeptide involved in immune function. Overall, the transcriptome profile of the hemocytes suggests that the LPS component of Novacq™ is highly immunostimulatory and generates a strong PO response in vitro. The subsequent transcriptional response appears to be directed towards preventing further activation of the PO system most likely in an attempt to limit cytoxicity to the host. Our study highlights the immunostimulatory ability of Novacq™ and provides further evidence of the positive health benefits this microbial based feed additive can have in shrimp.

Uncovering the link between environmental factors and coral immunity: A study of fluorescent protein expression and phenoloxidase activity in Acropora cervicornis

Worsening environmental conditions due to climate change have profoundly affected the health of coral reefs worldwide. Thus, understanding how corals respond to fluctuating and/or extreme levels of temperature and solar irradiation will guide future protection and restoration efforts of this valuable ecosystem. Herein, we present a study of the immune responses of the endangered coral Acropora cervicornis to seasonal fluctuations in water temperature (WT), light intensity (LI), and water depth. Immune responses were observed by measuring the concentration of green and cyan fluorescent proteins (GFP and CyFP) and the activity of phenoloxidase (PO), an enzyme involved in the biosynthesis of the photoprotective protein melanin. To study these responses, visually healthy A. cervicornis fragments were placed at 8, and 12 m depth, and GFP, CyPF, and PO activity were measured at three-month intervals over a 12-month period. Seawater temperature and light intensity were also measured at each depth during this period. A general linear mixed model was used to determine the effects of seasonal variations of WT, LI, and water depth on the immune proteins. GFP, CyFP, and PO activity varied significantly across time – all higher in late summer/early fall and lower in late winter/early spring. Likewise, WT and LI significantly affected GFP, CyFP, and PO activity. On the other hand, water depth only had a significant effect on fluorescent protein concentrations but not PO activity. Our study demonstrates that corals can modulate these key immune-related proteins throughout natural seasonal fluctuations. That is, increasing in months of higher thermal and light conditions while decreasing in months with mild thermal and light conditions. The phenotypic plasticity of A. cervicornis in adapting to a changing environment underscores the importance that in future studies time of the year should be a meaningful consideration when evaluating the responses of A. cervicornis to the environment.

Sericin Ser3 Ectopic Expressed in Posterior Silk Gland Affects Hemolymph Immune Melanization Response via Reducing Melanin Synthesis in Silkworm

The transgenesis of silkworms is an important way to innovate genetic resources and silk function. However, the silk-gland (SG) of transgenic silkworms, which is the most concerned target tissue of sericulture, often suffers from low vitality, stunting and other problems, and the reasons are still unknown. This study trans engineered recombinant Ser3, a middle silk gland (MSG) specific expression gene, in the posterior silk gland (PSG) of the silkworm, and studied hemolymph immune melanization response changes in mutant pure line SER (Ser3+/+). The results showed that although the mutant had normal vitality, the melanin content and phenoloxidase (PO) activity in hemolymph related to humoral immunity were significantly reduced, and caused significantly slower blood melanization and weaker sterilization ability. The mechanism investigation showed that the mRNA levels and enzymatic activities of phenylalanine hydroxylase (PAH), tyrosine hydroxylase (TH) and dopamine decarboxylase (DDC) in the melanin synthesis pathway in mutant hemolymph, as well as the transcription levels of the PPAE, SP21 and serpins genes in the serine protease cascade were significantly affected. Moreover, the total antioxidant capacity, superoxide anion inhibition capacity and catalase (CAT) level related to the redox metabolic capacity of hemolymph were significantly increased, while the activities of superoxide dismutase (SOD) and glutathione reductase (GR), as well as the levels of hydrogen peroxide (H2O2) and glutathione (GSH), were significantly decreased. In conclusion, the anabolism of melanin in the hemolymph of PSG transgenic silkworm SER was inhibited, while the basic response level of oxidative stress was increased, and the hemolymph immune melanization response was decreased. The results will significantly improve the safe assessment and development of genetically modified organisms.

Bombyx mori as a model for Niallia circulans pathogenicity.

Increasing incidences of resistance to antibiotics by pathogenic bacteria is a worldwide concern and isolation of antibiotic-resistant strains of Niallia circulans (formerly known as Bacillus circulans), an opportunistic human pathogen, has been reported. Due to their lack of ethical constraints as well as their cost-effective rearing, invertebrates have been commonly used to study infection by bacteria pathogenic to humans. In this study, we demonstrate that a foodborne strain of N. circulans kills larvae of the silkworm, Bombyx mori within 48 h after hemolymph injection. The infected larvae turned black with an increase in the phenoloxidase (PO) activity in the hemolymph. Midgut injection of N. circulans resulted in the killing of larvae within 96 h. A significant increase in bacterial load was observed in the hemolymph 12 h after infection. The viable hemocyte number decreased to 48% within 12 h of injection. RT-qPCR analysis revealed that upon hemolymph infection with N. circulans the expression of the antimicrobial peptide (AMP) genes, Bmdefensin-B and Bmgloverin-3, were upregulated 2.5- and 1.8-fold, respectively, whereas 1.6-fold upregulation was observed for BmToll-2 in the larval fat body. Therapeutic effects of antibiotics like tetracycline, imipenem, ceftriaxone, ampicillin, and clindamycin were observed against N. circulans in the Bombyx larvae with varying efficacies. Results from this study suggest that larvae of B. mori can be used as infection models for screening therapeutics that are effective against N. circulans.

Effects of Shelter on the Hatching, Immune Performance, and Profitability of the Ovigerous Red Swamp Crayfish Procambarus clarkii under High Stocking Density

To develop the intensive breeding technology of the seed of the red swamp crayfish Procambarus clarkii, the survival rates, hatching effects (hatching rate, incubation level, and number of juveniles), and immune performance of ovigerous P. clarkia as well as economic benefits are evaluated under different shelter conditions under a high stocking density in this study. The experimental design includes three different forms of shelter treatments (D1: experiment without any shelters; D2: experiment with closed shelters; D3: experiment with open shelters), each with three replicates. The results show that the concentration of the total antioxidant capacity (T-AOC) and activities of phenoloxidase (PO), catalase (CAT), and acid phosphatase (ACP) in the D3 treatment are higher than those in the D1 treatment (all p &lt; 0.05), with the highest concentrations of total antioxidant capacity (T-AOC) and malondialdehyde (MDA) and the highest activities of phenoloxidase (PO), superoxide dismutase (SOD), catalase (CAT), acid phosphatase (ACP), and alkaline phosphatase (AKP) among the treatments being present in the ovigerous P. clarkii in the D3 treatment. The hatching rates of the three treatments vary from 69.51% to 94.28%, with the highest rate found in the D3 treatment and the lowest in the D1 treatment, but there is no significant difference among them (p &gt; 0.05). The highest incubation level (ind.·m−2) and the highest number of juveniles (ind.·m−2) among treatments are found in the D3 treatment, with the incubation level (ind.·m−2) in the D3 treatment being significantly higher than that in the D1 treatment (p &lt; 0.05). The benefit–cost ratios (BCRs) of the D2 and D3 treatments remain significantly higher than that of the D1 treatment when P. clarkii prices change (all p &lt; 0.05). Our results indicate that a high stocking density habitat with open shelters could effectively improve the hatching and immune performance of ovigerous P. clarkii. Our findings are relevant for the indoor aquaculture management of ovigerous P. clarkii.

Elevated CO2 alters aggregate‑carbon and microbial community but does not affect total soil organic C in the semi-arid tropics

Elevated CO2 increases biomass productivity, however, the retention of added C under such condition is less known in the arable soils of the Tropics. Hence, an experiment was conducted in free-air atmospheric carbon-dioxide enrichment (FACE) rings to assess the impacts of eight years of elevated CO2levels (550 ppm) on soil aggregation, soil organic carbon (SOC), microbial community and greenhouse gas (GHG) emission in a sandy loam soil. Aboveground biomass productivity of all crops increased significantly under elevated CO2as compared to the ambient CO2, however, soils under elevated and ambient CO2 had similar total SOC in all layers, up to 0–90 cm profile. Soils under elevated CO2 had ~54 % higher large macroaggregates, but had ~14 % fewer microaggregates than ambient CO2 in the 0–5 cm layer. Elevated CO2 led to ~25 % less microaggregate-associated C when compared to the ambient CO2 in the topsoil. Elevated CO2 led to 10 % more large macroaggregates than ambient CO2 in the 5–15 cm soil layer. β-glucosidase activity within microaggregates in the topsoil under elevated CO2 was 25 % higher than ambient CO2. Moreover, soil macro- and microaggregates under elevated CO2 had ~31 and 35 % higher phenol oxidase activities, respectively, than those under ambient CO2in the topsoil. Microaggregates under elevated CO2 also contained ~13 and 21 % more total glomalin in the topsoil and 5–15 cm soil layers, respectively, as compared to the ambient CO2. The functional diversity (0–15 cm soil) was significantly higher under elevated CO2 than ambient CO2. Soils exposed to elevated CO2 had higher relative abundances of fungi than ambient CO2. A 23.4 % higher qCO2 and higher global warming potential was observed in elevated CO2 exposed soil as compared to ambient. These results reveal that under elevated CO2, carbon sequestration is hindered by higher fungal activity and higher activities of soil C degrading enzymes.

The importance of moisture in regulating soil organic carbon content based on a comparison of “enzymic latch” and “iron gate” in Zoige Plateau peatland

Due to the enormous amount of carbon (C) stored in peatlands, understanding the regulator of soil organic carbon (SOC) in peatlands is critical for global C cycling. Currently, “enzymic latch” and “iron gate” are the frequently debated mechanisms that propose phenol oxidase (PHO) and Fe2+ as important regulators driving SOC cycling in peatland. Therefore, this study compared “enzymic latch” and “iron gate” in the same peatland (Zoige Plateau, the largest peatland in China) to figure out the crucial regulator of SOC content via investigating the interrelationships among soil properties, C-related enzymes, phenolics, and iron (Fe) oxides. Moreover, waterlogged and non-waterlogged soils were chosen to take samples in both dry and wet season to understand the potential role of moisture in regulating SOC content. It was found that the contents of dissolved organic C, phenolics, SOC, Fe oxides, and β-glucosidase (BG) activities in waterlogged soils were higher than those in non-waterlogged soils. Compared to the dry season, soils in the wet season had higher phenolics, SOC, and Fe oxides contents. Also, this study observed an insignificant relationship between PHO and SOC (p > 0.05) and the small amounts of Fe2+ to total Fe (<2‰) in soils. Meanwhile, the highest effect of moisture (0.89) and negligible effect of PHO (0.04) and Fe2+ (-0.08) on SOC based on pathway analysis suggested that moisture might be the crucial regulator of SOC. Furthermore, pathway analysis implied that moisture could indirectly affect SOC by adjusting BG (p < 0.05) and soluble phenolics (p < 0.05). This study provided valuable information for understanding the importance of moisture in driving C cycling and helping against soil C loss by moisture management in Zoige Plateau.

Functional homology of tachykinin signalling: The influence of human substance P on the immune system of the mealworm beetle, Tenebrio molitor L.

Tachykinin-related peptides (TRPs) are one of the most prominent families of neuropeptides in the animal kingdom. Insect TRPs display strong structural and functional homology to vertebrate tachykinins (TKs). To study functional homologies between these two neuropeptide families, the influence of human substance P (SP, one of the essential vertebrate TKs) on the immune system of the mealworm beetle, Tenebrio molitor L., was analysed.Human SP influences the phagocytic abilities of T. molitor haemocytes. Peptide injection leads to an increase in the number of haemocytes participating in the phagocytosis of latex beads. In contrast, incubation of haemocytes from non-injected beetles in a solution of physiological saline and SP causes a decrease in phagocytic activity. Treatment with human SP also led to increased adhesion of haemocytes, but no changes in the arrangement of the F-actin cytoskeleton were observed. Interestingly, 6 h after human SP injection, increased DNA integrity in T. molitor haemocytes was reported. The opposite effects were observed 24 h after SP injection.Human SP caused the upregulation of humoral immune responses, such as phenoloxidase (PO) activity in the T. molitor haemolymph, and the downregulation of immune-related genes encoding coleoptericin A, tenecin 3 and Toll receptor. However, genes encoding attacin 2 and cecropin were upregulated. Despite these differences, the antimicrobial activity of T. molitor haemolymph was significantly lower in beetles injected with SP than in control beetles. Moreover, an analysis of the direct influence of SP on lysozyme activity was performed. Our results suggest that SP at a concentration of 10−6 M can directly inhibit lysozyme activity. However, an opposite effect was reported after the application of SP at a concentration of 10−4 M.The presented results suggest structural and functional homology between TK signalling in vertebrates and insects. Primarily, this was visible in the context of the humoral response and general antimicrobial activity of T. molitor haemolymph. However, some of the results related to haemocyte function may also indicate the importance of the TK and TRP sequences for evoking immunological effects.