Plant Hormone Salicylic Acid Produced by a Malaria Parasite Controls Host Immunity and Cerebral Malaria Outcome

The apicomplexan parasite Toxoplasma gondii produces the plant hormone abscisic acid, but it is unclear if phytohormones are produced by the malaria parasite Plasmodium spp., the most important parasite of this phylum. Here, we report detection of salicylic acid, an immune-related phytohormone of land plants, in P. berghei ANKA and T. gondii cell lysates. However, addition of salicylic acid to P. falciparum and T. gondii culture had no effect. We transfected P. falciparum 3D7 with the nahG gene, which encodes a salicylic acid-degrading enzyme isolated from plant-infecting Pseudomonas sp., and established a salicylic acid-deficient mutant. The mutant had a significantly decreased concentration of parasite-synthesized prostaglandin E₂, which potentially modulates host immunity as an adaptive evolution of Plasmodium spp. To investigate the function of salicylic acid and prostaglandin E₂ on host immunity, we established P. berghei ANKA mutants expressing nahG. C57BL/6 mice infected with nahG transfectants developed enhanced cerebral malaria, as assessed by Evans blue leakage and brain histological observation. The nahG-transfectant also significantly increased the mortality rate of mice. Prostaglandin E₂ reduced the brain symptoms by induction of T helper-2 cytokines. As expected, T helper-1 cytokines including interferon-γ and interleukin-2 were significantly elevated by infection with the nahG transfectant. Thus, salicylic acid of Plasmodium spp. may be a new pathogenic factor of this threatening parasite and may modulate immune function via parasite-produced prostaglandin E₂.     

Distribution of NO-induced cGMP-like immunoreactive neurones in the abdominal nervous system of the crayfish,Procambarus clarkii

Nitric oxide (NO) acts as a signalling molecule by activating soluble guanylate cyclase and causing accumulation of the second messenger cyclic guanosine 3',5'-monophosphate (cGMP) in target cells. In order to detect the presence of NO-cGMP signalling pathway in the crayfish abdominal nervous system, accumulation of NO-induced cGMP was investigated by anti-cGMP immunochemistry. Some preparations were incubated in a high-K(+) saline containing an inhibitor of cGMP-degrading phosphodiesterase, 3-isobutyl-1-methyxanthine (IBMX), to activate NO generating neurones, which could release NO in the ganglion, and then immunohistochemistry using an anti-cGMP antibody was performed. The other preparations were incubated in NO donor, sodium nitroprusside (SNP) saline containing IBMX before anti-cGMP immunohistochemistry was performed. The distribution of cGMP-like immunoreactive neurones in high-K(+) treated preparations was similar to that of cGMP-like immunoreactive neurones in NO donor treated preparations. About 70-80 cell bodies and many neuronal branches in the neuropilar area of the ganglion were stained, although no neurones showed immunoreactivity unless preparations were activated by either high-K(+) or the NO donor. Some of them were identical neurones, and they were intersegmental ascending interneurones and motor neurones. Sensory afferents that innervates hind gut showed strong cGMP-like immunoreactivity, although no mechanosensory afferents showed any immunoreactivity. These results strongly suggest the presence of an NO-cGMP signalling pathway that regulates neuronal events in the abdominal nervous system of the crayfish.     

Nitric oxide regulates the levels of cGMP accumulation in the cricket brain

Cricket brains were incubated in a saline containing nitric oxide (NO)-donor and phosphodiesterase inhibitor IBMX, which could activate soluble guanylate cyclase (sGC) to increase cGMP levels in the targets of NO. The increase of cGMP was detected by immunohistochemistry and enzyme linked immunosorbent assay. NO-induced cGMP immunohistochemistry revealed that many cell bodies of cricket brain showed cGMP immunoreactivity when preparations were treated with a saline containing 10 mM NO-donor SNP and phosphodiesterase inhibitor IBMX, but only a few cell bodies showed immunoreactivity when preparations were incubated without NO-donor. The concentration of cGMP in cricket brains were then measured by using cGMP-specific enzyme linked immunosorbent assay. Cricket brains were treated with a saline containing 1 microM of NO-donor NOR3 and 1 mM IBMX. The cGMP levels in the brain were increased about 75% compared to control preparations that was treated with a cricket saline containing IBMX. The level of cGMP decreased about 40% when preparations were incubated NOR3 saline containing sGC inhibitor ODQ. These results indicate that NO activates sGC and increases the levels of cGMP in particular neurons of the cricket brain and that the level of cGMP would be kept a particular level, which might regulate synaptic efficacy in the neurotransmission.     

Memory trace in feeding neural circuitry underlying conditioned taste aversion in lymnaea

Background

The pond snail Lymnaea stagnalis can maintain a conditioned taste aversion (CTA) as a long-term memory. Previous studies have shown that the inhibitory postsynaptic potential (IPSP) evoked in the neuron 1 medial (N1M) cell by activation of the cerebral giant cell (CGC) in taste aversion-trained snails was larger and lasted longer than that in control snails. The N1M cell is one of the interneurons in the feeding central pattern generator (CPG), and the CGC is a key regulatory neuron for the feeding CPG.

Methodology/Principle Findings

Previous studies have suggested that the neural circuit between the CGC and the N1M cell consists of two synaptic connections: (1) the excitatory connection from the CGC to the neuron 3 tonic (N3t) cell and (2) the inhibitory connection from the N3t cell to the N1M cell. However, because the N3t cell is too small to access consistently by electrophysiological methods, in the present study the synaptic inputs from the CGC to the N3t cell and those from the N3t cell to the N1M cell were monitored as the monosynaptic excitatory postsynaptic potential (EPSP) recorded in the large B1 and B3 motor neurons, respectively. The evoked monosynaptic EPSPs of the B1 motor neurons in the brains isolated from the taste aversion-trained snails were identical to those in the control snails, whereas the spontaneous monosynaptic EPSPs of the B3 motor neurons were significantly enlarged.

Conclusion/Significance

These results suggest that, after taste aversion training, the monosynaptic inputs from the N3t cell to the following neurons including the N1M cell are specifically facilitated. That is, one of the memory traces for taste aversion remains as an increase in neurotransmitter released from the N3t cell. We thus conclude that the N3t cell suppresses the N1M cell in the feeding CPG, in response to the conditioned stimulus in Lymnaea CTA.     

Amelioration of the salt-stressed root growth of rice and normalization of the Na+ distribution between the shoot and root by (S)-alpha-methylbenzyl-2-fluoro-4-methylphenylurea

Optically active alpha-methylbenzyl phenyl ureas (MBPUs) show diverse plant physiological properties. Experiments were conducted to evaluate the salt-stress response of just-germinated rice seedlings supplemented with the S-enantiomer of MBPUs by assessing the growth and Na+ content. This study indicates that S-MBPUs served as a unique stress reliever for just-germinated young seedlings of rice injured by salinity. NaCl severely affected the root growth of rice seedlings. Concomitant treatment with S-MBPUs effectively ameliorated the growth inhibition of rice by NaCl. Glycine betaine (GB) did not act as a reliever of the NaCl stress. The addition of S-alpha-methylbenzyl 2-fluoro-4-methylphenyl urea (7, denoted as S-FM) to the saline medium ameliorated not only the root growth but also the protein content and dry weight of roots depending upon its concentration. The protein content, Na+ content and growth rate were correlated to each other with a positive relationship. The Na+ distribution ratio (S/R(Na+)) between the shoot and root increased with increasing concentration of NaCl when added alone, viz. with increasing growth reduction. A concomitant treatment with S-FM (7), however, resulted in the S/R(Na+) value becoming smaller with growth amelioration. This indicates that S-FM (7) controlled the translation of Na+ from the roots to shoots. S-FM (7) would have influenced some inherent functions connected with the Na+ behavior in the rice plant, although details of the mechanism for normalization of the S/R(Na+) ratio are still not clear.     

Suppressor of cytokine signaling 1 DNA administration inhibits inflammatory and pathogenic responses in autoimmune myocarditis

Myocarditis and subsequent dilated cardiomyopathy are major causes of heart failure in young adults. Myocarditis in humans is highly heterogeneous in etiology. Recent studies have indicated that a subgroup of myocarditis patients may benefit from immune-targeted therapies, because autoimmunity plays an important role in myocarditis as well as contributing to the progression to cardiomyopathy and heart failure. Suppressor of cytokine signaling (SOCS) 1 plays a key role in the negative regulation of both TLR- and cytokine receptor-mediated signaling, which is involved in innate immunity and subsequent adaptive immunity. In this study, we investigated the therapeutic effect of SOCS1 DNA administration on experimental autoimmune myocarditis (EAM) in mice. EAM was induced by s.c. immunization with cardiac-specific peptides derived from α myosin H chain in BALB/c mice. In contrast to control myocarditis mice, SOCS1 DNA-injected mice were protected from development of EAM and heart failure. SOCS1 DNA administration was effective for reducing the activation of autoreactive CD4(+) T cells by inhibition of the function of Ag-presenting dendritic cells. Our findings suggest that SOCS1 DNA administration has considerable therapeutic potential in individuals with autoimmune myocarditis and dilated cardiomyopathy.     

Rapid recruitment of innate immunity regulates variation of intracellular pathogen resistance in Drosophila

Genetic variation in susceptibility to pathogens is a central concern both to medicine and agriculture and to the evolution of animals. Here, we have investigated the link between such natural genetic variation and the immune response in wild-type Drosophila melanogaster, a major model organism for immunological research. We found that within nine wild-type strains, different Drosophila genotypes show wide-ranging variation in their ability to survive infection from the pathogenic bacteria Listeria monocytogenes. Canton-S, a resistant strain, showed increased capacity to induce stronger innate immune activities (antimicrobial peptides (AMPs), phenol oxidase activity, and phagocytosis) compared to the susceptible strain (white) at early time points during bacterial infection. Moreover, PGRP-LE-induced innate immune activation immediately after infection greatly improves survival of the susceptible strain strongly suggesting a mechanism behind the natural genetic variation of these two strains. Taken together we provide the first experimental evidence to suggest that differences in innate immune activity at early time points during infection likely mediates infection susceptibility in Drosophila.     

Characterization of reconstituted high-density lipoprotein particles formed by lipid interactions with human serum amyloid A

The acute-phase human protein serum amyloid A (SAA) is enriched in high-density lipoprotein (HDL) in patients with inflammatory diseases. Compared with normal HDL containing apolipoprotein A-I, which is the principal protein component, characteristics of acute-phase HDL containing SAA remain largely undefined. In the present study, we examined the physicochemical properties of reconstituted HDL (rHDL) particles formed by lipid interactions with SAA. Fluorescence and circular dichroism measurements revealed that although SAA was unstructured at physiological temperature, α-helix formation was induced upon binding to phospholipid vesicles. SAA also formed rHDL particles by solubilizing phospholipid vesicles through mechanisms that are common to other exchangeable apolipoproteins. Dynamic light scattering and nondenaturing gradient gel electrophoresis analyses of rHDL after gel filtration revealed particle sizes of approximately 10 nm, and a discoidal shape was verified by transmission electron microscopy. Thermal denaturation experiments indicated that SAA molecules in rHDL retained α-helical conformations at 37 °C, but were almost completely denatured around 60 °C. Furthermore, trypsin digestion experiments showed that lipid binding rendered SAA molecules resistant to protein degradation. In humans, three major SAA1 isoforms (SAA1.1, 1.3, and 1.5) are known. Although these isoforms have different amino acids at residues 52 and 57, no major differences in physicochemical properties between rHDL particles resulting from lipid interactions with SAA isoforms have been found. The present data provide useful insights into the effects of SAA enrichment on the physicochemical properties of HDL.     

Soldier-specific modification of the mandibular motor neurons in termites

Social insects exhibit a variety of caste-specific behavioral tendencies that constitute the basis of division of labor within the colony. In termites, the soldier caste display distinctive defense behaviors, such as aggressively attacking enemies with well-developed mandibles, while the other castes retreat into the colony without exhibiting any aggressive response. It is thus likely that some form of soldier-specific neuronal modification exists in termites. In this study, the authors compared the brain (cerebral ganglion) and the suboesophageal ganglion (SOG) of soldiers and pseudergates (workers) in the damp-wood termite, Hodotermopsis sjostedti. The size of the SOG was significantly larger in soldiers than in pseudergates, but no difference in brain size was apparent between castes. Furthermore, mandibular nerves were thicker in soldiers than in pseudergates. Retrograde staining revealed that the somata sizes of the mandibular motor neurons (MdMNs) in soldiers were more than twice as large as those of pseudergates. The enlargement of MdMNs was also observed in individuals treated with a juvenile hormone analogue (JHA), indicating that MdMNs become enlarged in response to juvenile hormone (JH) action during soldier differentiation. This enlargement is likely to have two functions: a behavioral function in which soldier termites will be able to defend more effectively through relatively faster and stronger mandibular movements, and a developmental function that associates with the development of soldier-specific mandibular muscle morphogenesis in termite head. The soldier-specific enlargement of mandibular motor neurons was observed in all examined species in five termite families that have different mechanisms of defense, suggesting that such neuronal modification was already present in the common ancestor of termites and is significant for soldier function.     

A single fluorescence-based LAMP reaction for identifying multiple parasites in mosquitoes

Vector-borne diseases, such as malaria and lymphatic filariasis, are co-endemic in large parts of the world. To develop a multiplex amplification method for the simultaneous detection of multiple insect-borne infectious diseases, we used LAMP with fluorescently labeled primers to identify the SPECT2 gene of Plasmodium berghei and the cytochrome oxidase subunit I gene of Dirofilaria immitis in mosquitoes. This technique could detect as few as 100 P. berghei-infected red blood cell-equivalents or one D. immitis microfilaria. Moreover, individual species of parasites in mosquitoes could be identified when a mixture of fluorescently labeled primer sets was used. These findings suggest that the multiplex LAMP assay is sensitive and specific enough to identify parasite-bearing mosquitoes in areas where several diseases occur simultaneously. This procedure could increase the efficiency and effectiveness of arthropod-borne disease elimination programs.