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The effect of seasons on the soil microbiome in a Larix gmelinii forest of Mohe, China, where winter temperatures are generally below − 40 °C, was evaluated with metagenomics analysis. Taxonomic profiling using sequencing information revealed that Proteobacteria, Actinobacteria, Acidobacteria and Verrucomicrobia were the dominant phyla in spring, summer, and fall, as were Bradyrhizobium, Chthoniobacter, Streptomyces, Acid Candidatus Koribacter at the genus level. Some species that were abundant in spring and fall greatly diminished in abundance in summer. Clusters of orthologous groups (COG) of proteins, carbohydrate-active enzymes (CAZy), Kyoto Encyclopedia of Genes and Genomes (KEGG) and NCBI databases were used to elucidate the function of diverse proteins and metabolites of the microbial community of L. gmelinii forest. COG analysis showed that fewer genes were detected in spring than in fall and summer, indicating that many soil microbes in the L. gmelinii forest were not tolerant to cold. Based on KEGG analysis, some pathways in the soil microbes were activated in spring and autumn and deactivated in summer. CAZy analysis revealed that most CAZy were more active in summer than in spring or autumn and were severely inhibited in the spring. Many functional pathways, proteins, and CAZy involved in the community changes were concerned with cold or heat resistance. Therefore, the soil in the L. gmelinii forest can be a valuable resource for further research on heat and cold tolerance of soil microbes.

Some groups of fossil beetles are difficult to study because the characters needed for identification or classification are hidden and difficult to study. Ptinidae is one of these groups. We descri...

Significance Since Darwin, insect pollination was thought to be a key contributor to the Cretaceous radiation of angiosperms. Both insects and angiosperms were common during the mid-Cretaceous, but direct evidence for a Cretaceous insect-angiosperm pollination mode was until now absent. Here, we report a specialized beetle-angiosperm pollination mode preserved in Burmese amber where a tumbling flower beetle is carrying tricolpate pollen grains that belongs to the eudicots that comprise the majority of extant angiosperm species. Our study provides direct evidence of insect pollination of Cretaceous flowers, which is further supported by the flower-visiting body shape, specialized pollen-feeding mouthparts, and zoophilous pollen grains. These findings demonstrate that insect pollination of flowering plants was well established 99 million years ago., Insect pollination of flowering plants (angiosperms) is responsible for the majority of the world’s flowering plant diversity and is key to the Cretaceous radiation of angiosperms. Although both insects and angiosperms were common by the mid-Cretaceous, direct fossil evidence of insect pollination is lacking. Direct evidence of Cretaceous insect pollination is associated with insect-gymnosperm pollination. Here, we report a specialized beetle-angiosperm pollination mode from mid-Cretaceous Burmese amber (99 mega-annum [Ma]) in which a tumbling flower beetle (Mordellidae), Angimordella burmitina gen. et sp. nov., has many tricolpate pollen grains attached. A. burmitina exhibits several specialized body structures for flower-visiting behavior including its body shape and pollen-feeding mouthparts revealed by X-ray microcomputed tomography (micro-CT). The tricolpate pollen in the amber belongs to the eudicots that comprise the majority of extant angiosperm species. These pollen grains exhibit zoophilous pollination attributes including their ornamentation, size, and clumping characteristics. Tricolpate pollen grains attached to the beetle’s hairs are revealed by confocal laser scanning microscopy, which is a powerful tool for investigating pollen in amber. Our findings provide direct evidence of insect pollination of Cretaceous angiosperms, extending the range insect-angiosperm pollination association by at least 50 million years. Our results support the hypothesis that specialized insect pollination modes were present in eudicots 99 million years ago.

Cryptococcus neoformans is an encapsulated yeast pathogen that infects immunocompromised patients to cause fungal meningitis, resulting in hundreds of thousands of deaths each year. F-box protein Fbp1, the key component of the E3 ubiquitin ligase, plays a critical role in fungal development and virulence in fungal pathogens. In this study, we identified a potential substrate of Fbp1, the vacuolar morphogenesis protein Vam6-like protein Vlp1, and evaluated its role in virulence in C. neoformans. Deletion or overexpression of the VLP1 gene results in abnormal capsule formation and melanin production of C. neoformans. Stress tolerance assay showed that the vlp1Δ mutant was sensitive to SDS and NaCl but not to CFW or Congo red, indicating that Vlp1 might regulate the cell membrane integrity in C. neoformans. Fungal virulence assay showed that Vlp1 was essential for the pathogenicity of C. neoformans, as vlp1Δ mutants are avirulent in the mouse systematic infection model of cryptococcosis. The progression of fungal infection revealed that the vlp1Δ mutants were gradually eliminated from the lungs of the mice after infection. Moreover, the vlp1Δ mutants showed a proliferation defect inside macrophages and a viability defect in the host complement system, which likely contributes to the virulence attenuation of the vlp1Δ mutants. In summary, our results revealed that the vacuolar morphogenesis protein Vam6-like protein Vlp1 is essential for the pathogenicity of C. neoformans.

Within this study the subfamily Praemordellinae is reviewed and the holotype of Praemordella martynovi Scegoleva-Barovskaja, 1929 is re-described. The genera Cretanaspis Huang and Yang, 1999, Mirimordella Liu et al., 2007, Bellimordella Liu et al., 2008 and Wuhua Wang and Zhang, 2011 is attributed to Praemordellinae. One new species, Wuhua peregrina sp. nov., is erected based on two well-preserved specimens from the Middle Jurassic Daohugou Biota (Inner Mongolia, China). The morphological characters of Praemordellinae are given and a key to genera of Praemordellinae is presented. Praemordellinae is closely related to extant Mordellidae in having humpbacked body, strongly deflexed head, filiform antennae and pectinate tarsal claws, but it is differing in that the pygidium absent and hind femora not well developed. It is also similar to Ripiphoridae in having a convex body and simple tarsi, but differs Ripiphoridae in having filiform antennae. Praemordellinae is probably a stem group including the ancestor of Mordellidae and Ripiphoridae. Also, within this paper, the fossil record of Mordellidae is summarized.

Cryptococcus neoformans is a ubiquitous yeast pathogen that often infects the human central nervous system (CNS) to cause meningitis in immunocompromised individuals. Although numerous signaling pathways and factors important for fungal sexual reproduction and virulence have been investigated, their precise mechanism of action remains to be further elucidated. In this study, we identified and characterized a novel zinc finger protein Zfp1 that regulates fungal sexual reproduction and virulence in C. neoformans. qRT-PCR and ZFP1 promoter regulatory activity assays revealed a ubiquitous expression pattern of ZFP1 in all stages during mating. Subcellular localization analysis indicates that Zfp1 is targeted to the cytoplasm of C. neoformans. In vitro assays of stress responses showed that zfp1Δ mutants and the ZFP1 overexpressed strains ZFP1OE are hypersensitive to SDS, but not Congo red, indicating that Zfp1 may regulate cell membrane integrity. Zfp1 is also essential for fungal sexual reproduction because basidiospore production was blocked in bilateral mating between zfp1Δ mutants or ZFP1 overexpressed strains. Fungal nuclei development assay showed that nuclei in the bilateral mating of zfp1Δ mutants or ZFP1 overexpressed strains failed to undergo meiosis after fusion, indicating Zfp1 is important for regulating meiosis during mating. Although zfp1Δ mutants showed normal growth and produced normal major virulence factors, virulence was attenuated in a murine model. Interestingly, we found that the ZFP1 overexpressed strains were avirulent in a murine systemic-infection model. Overall, our study showed that the zinc finger protein Zfp1 is essential for fungal sporulation and virulence in C. neoformans.

Primaevomordellida burmitina gen. et sp. nov. (Coleoptera: Mordellidae) is described on the base of two well-preserved specimens from Cenomanian Burmese amber. Due to the general body plan and the preservation of elongated pygidium, it is apparently the first true pintail beetle (subfamily Mordellinae). The simple hind leg structures placed the new species into the basal tribe Raynoldsiellini; however, the comparative morphology of the new species shows advanced characters similar to tribes Mordellini and Mordellistenini. The angiosperm plant records in Burmese amber forests were reviewed and the mordellids-angiosperm interactions are discussed.

A new species of tumbling flower beetle (Coleoptera: Mordellidae) Tomoxia succinea sp. nov. is described based on two Baltic amber specimens. The new species differs from the only extant European species of this genus, Tomoxia bucephala Costa, 1854, mainly in the shape of the scutellum and the length ratio of particular body parts. The fossil records of Mordellidae found in Baltic amber are summarized; their paleoecology in the Paleocene–Eocene of the Baltic region and the composition and distribution of the family Mordellidae are discussed.

Myrmecophilous and termitophilous interactions likely contributed to the competitive advantage and evolutionary success of eusocial insects, but how these commensal and parasitic relationships originated is unclear due to absence of fossil records. New extinct cockroaches of the still living family Blattidae are reported here from the Cretaceous Myanmar amber (99 Ma) and are the earliest known inhabitants of complex ant nests, demonstrating that this specialised myrmecophily originated shortly after ant eusociality and appeared in the fossil record. Cretaceous stem aposematic Blattidae are known from the amber of Myanmar and Lebanon and we report them here also from the Syrian amber. Concurrent evolution suggests that the collective internal defence of early ants was weak and allowed infiltrations by numerous unrelated organisms, At the same time, the contemporary presence of ant mimicking myrmecomorphs suggests a need for strong external protection against visually hunting predators. Myrmecophily is supported by morphological adaptations (lack of wide fat body and feeding of adult male; short, fossorial legs; shortened cerci; oligomerised antenna; hairy surface structures) and camouflage behaviour, documented by sediment and own feces covering. Moreover the same piece of amber contains ants, ant mimics and other undescribed ant nest-visiting insects as syninclusions. Another species preserved along with two termites is a putative termitophile. Abundant comparatively large parasitic cockroaches influenced Mesozoic tropical forest ecosystems by affecting the early evolution of complex nests of eusocial insects. Rainforest rudiments in South Yunnan yielded observation of analogical still living, formally undescribed species.

The new family Apotomouridae fam. nov. (Coleoptera: Polyphaga: Tenebrionoidea) is described based on Multispinus multispinosus gen. et sp. nov. and Apotomoura fortiscrura gen. et. sp. nov. from Burmese amber (lower Cenomanian, ca. 99 Ma). The retractile wedge-shaped body and tarsal formula 5-5-4 placed Apotomouridae fam. nov. in Tenebrionoidea. The new family shares similarities with the Mordellidae, whereas the absence of pygidium and the combination of other microstructures assigned it more closely to the Jurassic beetle genus Wuhua. The morphological characters suggest the similar flower feeding style as to Mordellidae and Ripiphoridae, which represent the flower diet and pollinating strategy in the Tenebrionoidea.

The capsule of the fungal pathogen Cryptococcus neoformans consists of glucuronoxylomannan (GXM), glucuronoxylomannogalactan (GXMGal), and mannoproteins (MPs). MPs are a kind of glycoproteins with low content but high immunogenicity, which can stimulate the immune protection of the host. However, there is not much information about the role of mannoproteins in virulence of the human fungal pathogen C. neoformans. In this study, we reported the identification and functional analysis of a predicted mannoprotein Cmp1 that regulates fungal virulence in C. neoformans. Gene expression pattern analysis indicates that the CMP1 gene was ubiquitously expressed at all stages of cryptococcal development. Subcellular localization analysis indicated that Cmp1 was localized in the cytoplasm of cryptococcal cells. Disruption or overexpression of CMP1 results in impairing capsule formation in Cryptococcus, but it does not affect the melanin production and sensitivity under various stress conditions, nor does it affect the sexual reproduction process of Cryptococcus. Survival assay showed that the pathogenicity of the cmp1&Delta, mutant or the CMP1 overexpression strain was significantly attenuated in a murine inhalation model of cryptococcosis. In conclusion, our findings implied that the mannoprotein Cmp1 is required for the virulence of C. neoformans.

Zinc is one of the essential trace elements in eukaryotes and it is a critical structural component of a large number of proteins. Zinc finger proteins (ZNFs) are zinc-finger domain-containing proteins stabilized by bound zinc ions and they form the most abundant proteins, serving extraordinarily diverse biological functions. In recent years, many ZNFs have been identified and characterized in the human fungal pathogen Cryptococcus neoformans, a fungal pathogen causing fatal meningitis mainly in immunocompromised individuals. It has been shown that ZNFs play important roles in the morphological development, differentiation, and virulence of C. neoformans. In this review, we, first, briefly introduce the ZNFs and their classification. Then, we explain the identification and classification of the ZNFs in C. neoformans. Next, we focus on the biological role of the ZNFs functionally characterized so far in the sexual reproduction, virulence factor production, ion homeostasis, pathogenesis, and stress resistance in C. neoformans. We also discuss the perspectives on future function studies of ZNFs in C. neoformans.

Autophagy (macroautophagy) is an evolutionarily conserved degradation pathway involved in bulk degradation of cytoplasmic organelles, old protein, and other macromolecules and nutrient recycling during starvation. Extensive studies on functions of autophagy-related genes have revealed that autophagy plays a role in cell differentiation and pathogenesis of pathogenic fungi. In this study, we identified and characterized 14 core autophagy machinery genes (ATGs) in C. neoformans. To understand the function of autophagy in virulence and fungal development in C. neoformans, we knocked out the 14 ATGs in both α and a mating type strain backgrounds in C. neoformans, respectively, by using biolistic transformation and in vivo homologous recombination. Fungal virulence assay showed that virulence of each atgΔ mutants was attenuated in a murine inhalation systemic-infection model, although virulence factor production was not dramatically impaired in vitro. Fungal mating assays showed that all the 14 ATGs are essential for fungal sexual reproduction as basidiospore production was blocked in bilateral mating between each atgΔ mutants. Fungal nuclei development assay showed that nuclei in the bilateral mating of each atgΔ mutants failed to undergo meiosis after fusion, indicating autophagy is essential for regulating meiosis during mating. Overall, our study showed that autophagy is essential for fungal virulence and sexual reproduction in C. neoformans, which likely represents a conserved novel virulence and sexual reproduction control mechanism that involves the autophagy-mediated proteolysis pathway.

Cryptococcus neoformans is the most common cause of deadly fungal meningitis. This fungus has a complex inositol acquisition and utilization system, and our previous studies have shown the importance of inositol utilization in cryptococcal development and virulence. However, how inositol utilization is regulated in this fungus remains unknown. In this study, we found that inositol, irrespective of the presence of glucose in the media, represses the expression of C. neoformans genes involved in inositol pyrophosphate biosynthesis, including the gene encoding inositol hexakisphosphate kinase Kcs1. Kcs1 was recently reported to regulate inositol metabolism in Saccharomyces cerevisiae and to impact virulence in C. neoformans. To examine the potential role of Kcs1 in inositol regulation in C. neoformans, we generated the kcs1Δ mutant and compared its phenotype with the wild type strain. We found that Kcs1 negatively regulates inositol uptake and catabolism in C. neoformans, but, in contrast to Kcs1 function in S. cerevisiae, does not appear to regulate inositol biosynthesis. Together, these results show that Kcs1 functions to fine-tune inositol acquisition to maintain inositol homeostasis in C. neoformans.

The Pb-accumulator Sedum alfredii is a good phytoremediation material, and widely used in the phytoremediation research of soils contaminated with Pb. The root exudates from it may be playing a significant role in the process of phytoremediation. In this study, the metabonomics method which based on gas chromatography–mass spectrometry (GC–MS) and pattern recognition analysis was used to identify the remarkable root exudates from S. alfredii under different Pb stresses, including exposure concentrations (0, 10, 50, 200 and 1000 µmol/L) and times (4 and 8 days). And batch extraction experiments were used to verify the roles of these remarkable root exudates. According to the results, 11 metabolites were considered as the remarkable metabolites. Oxalic acid, galactonic acid and glyceric acid can remove Pb in soil, and the removal effect was: oxalic acid > galactonic acid > glyceric acid. Xylose, glucose and maltose have no removal effect for Pb in soil.

Mesozoic tenebrionid and leiodid beetles are poorly known. Here we describe Praezolodinus pilosus gen. et sp. nov. (Tenebrionidae: Zolodininae) and Cretoptomaphagus microsoma gen. et sp. nov. (Leiodidae: Cholevinae), both from mid-Cretaceous Burmese amber (ca. 99 Ma). Praezolodinus pilosus is placed in the basal group Zolodininae due to (1) galea preserved with fine and long pubescence, (2) aedeagus inverted, pimeloid type. Cretoptomaphagus microsoma is assigned to Leiodidae and included in Cholevinae based on its (1) characteristic interrupted five-segmented antennal club, and (2) cephalic configuration, with the posterodorsal contour of the exposed part of the head abutting with the anterior pronotal edge. Cretoptomaphagus microsoma is the second Cholevinae described from the Mesozoic. Its strong morphological similarity with extant members of Cholevinae makes this fossil an astonishing example of long-term morphological stasis in Coleoptera.

Cryptococcus neoformans is a basidiomycete human fungal pathogen causing lethal meningoencephalitis, mainly in immunocompromised patients. Oxidoreductases are a class of enzymes that catalyze redox, playing a crucial role in biochemical reactions. In this study, we identified one Cryptococcus oxidoreductase-like protein-encoding gene OLP1 and investigated its role in the sexual reproduction and virulence of C. neoformans. Gene expression patterns analysis showed that the OLP1 gene was expressed in each developmental stage of Cryptococcus, and the Olp1 protein was located in the cytoplasm of Cryptococcus cells. Although it produced normal major virulence factors such as melanin and capsule, the olp1&Delta, mutants showed growth defects on the yeast extract peptone dextrose (YPD) medium supplemented with lithium chloride (LiCl) and 5-fluorocytosine (5-FC). The fungal mating analysis showed that Olp1 is also essential for fungal sexual reproduction, as olp1&Delta, mutants show significant defects in hyphae growth and basidiospores production during bisexual reproduction. The fungal nuclei imaging showed that during the bilateral mating of olp1&Delta, mutants, the nuclei failed to undergo meiosis after fusion in the basidia, indicating that Olp1 is crucial for regulating meiosis during mating. Moreover, Olp1 was also found to be required for fungal virulence in C. neoformans, as the olp1&Delta, mutants showed significant virulence attenuation in a murine inhalation model. In conclusion, our results showed that the oxidoreductase-like protein Olp1 is required for both fungal sexual reproduction and virulence in C. neoformans.

The dual roles of baculovirus for the control of natural insect populations as an insecticide, and for foreign gene expression and delivery, have called for a comprehensive understanding of the molecular mechanisms governing viral infection. Here, we demonstrate that theBombyx moriNiemann-Pick C1 (BmNPC1) is essential for baculovirus infection in insect cells. Both pretreatment ofBombyx moriembryonic cells (BmE) with NPC1 antagonists (imipramine or U18666A) and down-regulation of NPC1 expression resulted in a significant reduction in baculovirus BmNPV (Bombyx morinuclear polyhedrosis virus) infectivity. Furthermore, we show that the major glycoprotein gp64 of BmNPV, responsible for both receptor binding and fusion, is able to interact predominantly with the BmNPC1 C domain, with an enhanced binding capacity at low pH conditions, indicating that NPC1 most likely plays a role during viral fusion in endosomal compartments. Our results, combined with previous studies identifying an essential role of hNPC1 in filovirus infection, suggest that the glycoprotein of several enveloped viruses possess a shared strategy of exploiting host NPC1 proteins during virus intracellular entry events.IMPORTANCEBmNPV is one of the most important members of theBaculoviridae; many viruses in this family have been frequently employed as viral vectors for foreign gene delivery or expression and as biopesticides, but their host receptors still remain unclear. Here, we describe that the intracellular cholesterol transporter BmNPC1 is indispensable for BmNPV infection in insect cells, and it interacts with the major viral glycoprotein gp64. Our study on the role of BmNPC1 in baculovirus infection has further expanded the list of the enveloped viruses that require host NPC1 proteins for entry, and will ultimately help us to uncover the molecular mechanism of the involvement of NPC1 proteins in the entry process of many enveloped viruses.

Cryptococcus neoformans is the main etiologic agent of cryptococcal meningitis and causes a significant number of deadly infections per year. Although it is well appreciated that host immune responses are crucial for defense against cryptococcosis, our understanding of factors that control the development of effective immunity to this fungus remains incomplete. In previous studies, we identified the F-box protein Fbp1 as a novel determinant of C. neoformans virulence. In this study, we found that the hypovirulence of the fbp1Δ mutant is linked to the development of a robust host immune response. Infection with the fbp1Δ mutant induces a rapid influx of CCR2+ monocytes and their differentiation into monocyte-derived dendritic cells (mo-DCs). Depletion of CCR2+ monocytes and their derivative mo-DCs resulted in impaired activation of a protective inflammatory response and the rapid death of mice infected with the fbp1Δ mutant. Mice lacking B and T cells also developed fungal meningitis and succumbed to infection with the fbp1Δ mutant, demonstrating that adaptive immune responses to the fbp1Δ mutant help to maintain the long-term survival of the host. Adaptive immune responses to the fbp1Δ mutant were characterized by enhanced differentiation of Th1 and Th17 CD4+ T cells together with diminished Th2 responses compared to the H99 parental strain. Importantly, we found that the enhanced immunogenicity of fbp1Δ mutant yeast cells can be harnessed to confer protection against a subsequent infection with the virulent H99 parental strain. Altogether, our findings suggest that Fbp1 functions as a novel virulence factor that shapes the immunogenicity of C. neoformans., IMPORTANCE Cryptococcus neoformans is the most common cause of deadly fungal meningitis, with over 270,000 infections per year. Immune responses are critically required for the prevention of cryptococcosis, and patients with impaired immunity and low CD4+ T cell numbers are at high risk of developing these deadly infections. Although it is well appreciated that the development of protective immunity is shaped by the interactions of the host immune system with fungal cells, our understanding of fungal products that influence this process remains poor. In this study, we found that the activity of F-box protein 1 (Fbp1) in highly virulent C. neoformans clinical strain H99 shapes its immunogenicity and thus affects the development of protective immune responses in the host. The identification of this new mechanism of virulence may facilitate the future development of therapeutic interventions aimed at boosting antifungal host immunity.

Corn (Zea Mays L.) seedlings were in four cadmium (Cd) levels of 0-1mg/L in a hydroponic system to analyze the antioxidant enzyme system, Cd content in the shoots and roots, growth responces in the leaves of corn and phytosiderphore release. There was a significant decrease in superoxide dismutase (SOD) activities, chlorophyll concentration (Chl), shoot and root biomass and phytosiderphore release from Cd 0mg/L to 1mg/L. This indicated that Cd stress induced an oxidative stress response in corn seedlings, characterized by a decrease in activities of SOD and chl concentration. Root and shoot Cd contents of corn increased with their exposure Cd level, and the highest Cd concentration occurred in roots, followed by leaves.

This work evaluated the effect of cadmium (Cd) on the physiological responses of wheat (Triticum aestivum L.). For this purpose, seedlings of wheat were cultivated under controlled conditions. The plant chlorophyll content of young leaves, biomass production, superoxide dismutase (SOD) activities and phytosiderophores (PS) release by roots were assessed. Results showed that Cd decreased the chlorophyll content of young leaves, accompanied by a significant shoot and root biomass reduction across all Cd treatments. Cadmium also inhibited PS release in wheat.

Nosema bombycis is an obligate intracellular parasite, which can cause pebrine disease. To investigate the effects of N. bombycis infection, 5th-instar silkworms were challenged with N. bombycis isolate CQ1, and two-dimensional gel electrophoresis analysis was performed to analyze the differentially expressed proteins in infected and uninfected silkworm fat bodies 1, 2, 4, 6 and 8days post-infection (dpi). 46 differentially expressed proteins were identified at the 5 time points using MALDI-TOF/TOF MS. The changed proteins mainly involved in immune response, energy metabolism, and molecular synthesis. Overall, the identified proteins may provide important insights into the mechanisms of the silkworm response to N. bombycis infection.

Soil bacteria play a key role in the ecological and evolutionary responses of agricultural ecosystems. Domestic herbivore grazing is known to influence soil bacterial community. However, the effects of grazing and its major driving factors on soil bacterial community remain unknown for different plant community compositions under increasing grazing intensity. Thus, to investigate soil bacterial community diversity under five plant community compositions (Grass; Leymus chinensis; Forb; L. chinensis & Forb; and Legume), we performed a four-year field experiment with different grazing intensity treatments (no grazing; light grazing, 4 sheep·ha-1; and heavy grazing, 6 sheep·ha-1) in a grassland in China. Total DNA was obtained from soil samples collected from the plots in August, and polymerase chain reaction (PCR) analysis and denaturing gradient gel electrophoresis (DGGE) fingerprinting were used to investigate soil bacterial community. The results showed that light grazing significantly increased indices of soil bacterial community diversity for the Forb and Legume groups but not the Grass and L. chinensis groups. Heavy grazing significantly reduced these soil bacterial diversity indices, except for the Pielou evenness index in the Legume group. Further analyses revealed that the soil N/P ratio, electrical conductivity (EC), total nitrogen (TN) and pH were the major environmental factors affecting the soil bacterial community. Our study suggests that the soil bacterial community diversity was influenced by grazing intensity and plant community composition in a meadow steppe. The present study provides a baseline assessment of the soil bacterial community diversity in a temperate meadow steppe.

Fungal meningitis is a serious disease caused by a fungal infection of the central nervous system (CNS) mostly in individuals with immune system deficiencies. Fungal meningitis is often fatal without proper treatment, and the mortality rate remains unacceptably high even with antifungal drug interventions. Currently, cryptococcal meningitis is the most common fungal meningitis in HIV-1/AIDS, and its disease mechanism has been extensively studied. The key steps for fungi to infect brain and cause meningitis after establishment of local infection are the dissemination of fungal cells to the bloodstream and invasion through the blood brain barrier to reach the CNS. In this review, we use cryptococcal CNS infection as an example to describe the current molecular understanding of fungal meningitis, including the establishment of the infection, dissemination, and brain invasion. Host and microbial factors that contribute to these infection steps are also discussed.

The ubiquitin-proteasome system is one of the major protein turnover mechanisms that plays important roles in the regulation of a variety of cellular functions. It is composed of E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3 ubiquitin ligases that transfer ubiquitin to the substrates that are subjected to degradation in the 26S proteasome. The Skp1, Cullin, F-box protein (SCF) E3 ligases are the largest E3 gene family, in which the F-box protein is the key component to determine substrate specificity. Although the SCF E3 ligase and its F-box proteins have been extensively studied in the model yeast Saccharomyces cerevisiae, only limited studies have been reported on the role of F-box proteins in other fungi. Recently, a number of studies revealed that F-box proteins are required for fungal pathogenicity. In this communication, we review the current understanding of F-box proteins in pathogenic fungi.

Casein kinases regulate a wide range of cellular functions in eukaryotes, including phosphorylation of proteins that are substrates for degradation via the ubiquitin-proteasome system (UPS). Our previous study demonstrated that Fbp1, a component of the SCF FBP1 E3 ligase complex, was essential for Cryptococcus virulence. Because the Saccharomyces cerevisiae homolog of Fbp1, Grr1, requires casein kinase I (Yck1 and Yck2) to phosphorylate its substrates, we investigated the function of casein kinase I in Cryptococcus neoformans . In this report, we identified a C. neoformans casein kinase I protein homolog, Cck1. Similar to Fbp1, the expression of Cck1 is negatively regulated by glucose and during mating. cck1 null mutants showed significant virulence attenuation in a murine systemic infection model, but Cck1 was dispensable for the development of classical virulence factors (capsule, melanin, and growth at 37°C). cck1 mutants were hypersensitive to SDS treatment, indicating that Cck1 is required for cell integrity. The functional overlap between Cck1 and Fbp1 suggests that Cck1 may be required for the phosphorylation of Fbp1 substrates. Interestingly, the cck1 mutant also showed increased sensitivity to osmotic stress and oxidative stress, suggesting that Cck1 regulates both cell integrity and the cellular stress response. Our results show that Cck1 regulates the phosphorylation of both Mpk1 and Hog1 mitogen-activated protein kinases (MAPKs), demonstrating that Cck1 regulates cell integrity via the Mpk1 pathway and regulates cell adaptation to stresses via the Hog1 pathway. Overall, our study revealed that Cck1 plays important roles in regulating multiple signaling pathways and is required for fungal pathogenicity.

Cryptococcus neoformans is the leading cause of fungal meningitis in immunocomprised populations. Although extensive studies have been conducted on signal transduction pathways important for fungal sexual reproduction and virulence, how fungal virulence is regulated during infection is still not understood. In this study, we identified the F-box protein Fbp1, which contains a putative F-box domain and 12 leucine-rich repeats (LRR). Although fbp1 mutants showed normal growth and produced normal major virulence factors, such as melanin and capsule, Fbp1 was found to be essential for fungal virulence, as fbp1 mutants were avirulent in a murine systemic-infection model. Fbp1 is also important for fungal sexual reproduction. Basidiospore production was blocked in bilateral mating between fbp1 mutants, even though normal dikaryotic hyphae were observed during mating. In vitro assays of stress responses revealed that fbp1 mutants are hypersensitive to SDS, but not calcofluor white (CFW) or Congo red, indicating that Fbp1 may regulate cell membrane integrity. Fbp1 physically interacts with Skp1 homologues in both Saccharomyces cerevisiae and C. neoformans via its F-box domain, suggesting it may function as part of an SCF (Skp1, Cullins, F-box proteins) E3 ligase. Overall, our study revealed that the F-box protein Fbp1 is essential for fungal sporulation and virulence in C. neoformans , which likely represents a conserved novel virulence control mechanism that involves the SCF E3 ubiquitin ligase-mediated proteolysis pathway.

Corn (Zea Mays L.) seedlings were in four cadmium (Cd) levels of 0-1mg/L in a hydroponic system to analyze the antioxidant enzyme system, Cd content in the shoots and roots and growth responces in the leaves of corn. There was a significant increase in malondialdehyde (MDA) concentration from Cd 0mg/L to 1mg/L, and peroxidase (POD) and catalase (CAT) activities in the leaves of corn subjected to 0-1mg/L Cd. However there was a significant decrease in superoxide dismutase (SOD) activities subjected to 0-1mg/L Cd. This indicated that Cd stress induced an oxidative stress response in corn seedlings, characterized by an accumulation of MDA, decrease in activities of SOD and increase in activities of POD and CAT. Root and leaf Cd contents of corn increased with their exposure Cd level, and the highest Cd concentration occurred in roots, followed by leaves.

The objective of this research was to determine the effect of iron (Fe) deficiency on the uptake of cadmium (Cd) by three plants: tomato (Lycopersicon esculentum Mill.), morel (Solanum nigrum L.), and leaf red beet (Beta vulgaris var.cicla L.) in hydroponic culture. We also investigated the influence of Fe deficiency on rhizosphere pH, bioconcentration factor (BCF), and translocation factor (TF) in three plants. The plants were grown in Hoagland's solution for 28 days, then some plants were still in full strength Hoagland's solution (+Fe) for seven days, the others were generated in full strength Hoagland's solutions without Fe (−Fe) for seven days. After the seedlings had grown for five weeks, the seedlings were exposed for four daysat concentrations of Cd 0 mg L−1 (control), 0.1 mg L−1, and 1 mg L−1. The results showed that Fe deficiency induced Cd uptake, biomass decrease and solution pH decrease in hydroponic culture by three plants. Values of BCF and TF indicated that three plants had dissimilar abil...

Atg4 is a unique cysteine protease responsible for the cleavage of the carboxyl terminus of Atg8 during the formation of autophagosomes in yeast. Here we report that MoAtg4, an Atg4 homologue in Magnaporthe oryzae, controls cell differentiation and pathogenicity by interacting with MoAtg8, an autophagic protein essential for autophagic cell death and pathogenicity. Yeast complementation assay revealed that MoATG4 can functionally complement the defects of the yeast ATG4 deletion mutant. The direct interaction between MoAtg4 and MoAtg8 was detected in both yeast two hybrid and bimolecular fluorescence complementation (BiFC) assays. We also specify a cysteine residue, Cys206, as the active residue within MoAtg4 for the cleavage of MoAtg8 in vitro. Expression pattern analysis revealed that MoATG4 gene is expressed throughout growth and development by M. oryzae and can be induced by starvation and MoAtg4 protein localized in the cytoplasm of M. oryzae. Deletion of MoATG4 in M. oryzae caused significant reduction of aerial hyphae, conidiation, perithecia formation and delay of conidial germination and appressorium formation. Furthermore, as a result of lower turgor pressure of the appressorium, the DeltaMoatg4 mutant lost its ability to penetrate rice and barley. The developmental and pathogenic phenotypes were recovered by reintroduction of an intact copy of MoATG4 into the mutant, suggesting that MoATG4 is indispensable in the development of M. oryzae and essential to pathogenicity of this fungus.

Fasciclin family proteins have been identified as cell adhesion molecules in various organisms. In this study, a novel Magnaporthe oryzae fasciclin-like protein encoding gene, named MoFLP1, was isolated from a subtractive suppressive cDNA library and functionally analyzed. Sequence analysis showed that the MoFLP1 gene contains an open reading frame (ORF) of 1050 nucleotides encoding 349 amino acids with a calculated molecular weight of 35.85 kDa and a pI of 7.76. The deduced MoFLP1 protein contains a 17-amino acid secretion signal sequence and an 18-amino acid sequence with the characteristics of a glycosylphosphotidylinositol (GPI) anchor additional signal at its N- and C-terminuses, respectively. Potential N-glycosylation sites and domains involving cell adhesion were also identified in MoFLP1. Sequence analysis and subcellular localization by the expression of MoFLP1-GFP fusion construct in M. oryzae indicated that the MoFLP1 protein is probably localized on the vacuole membrane. Two MoFLP1 null mutants generated by targeted gene disruption exhibited marked reduction of conidiation, conidial adhesion, appressorium turgor, and pathogenicity. Our results indicate that fasciclin proteins play important roles in fungal development and pathogenicity in M. oryzae.

Phytoremediation is a promising tool in removing pollutants from the environment or in rendering them harmless. The objective of this research was to determine the effect of iron-deficiency on the uptake of cadmium by Solanum nigrum L. Results showed that iron-deficiency induced cadmium uptake, biomass decrease and changes in pH and Eh in hydroponic culture. Under iron-deficiency status, the decrease in pH and the increase in Eh might result in higher cadmium availability. Bioconcentration and translocation factors indicated that iron-deficiency status affected cadmium accumulation and translocation in Solanum nigrum L.

Cryptococcus neoformans is a human fungal pathogen that often causes lung and brain infections in immunocompromised patients, with a high fatality rate. Our previous results showed that an F-box protein, Fbp1, is essential for Cryptococcus virulence independent of the classical virulence factors, suggesting a novel virulence control mechanism. In this study, we show that Fbp1 is part of the ubiquitin-proteasome system, and we further investigated the mechanism of Fbp1 function during infection. Time course studies revealed that the fbp1 Δ mutant causes little damage in the infected lung and that the fungal burden in the lung remains at a low but persistent level throughout infection. The fbp1 Δ mutant cannot disseminate to other organs following pulmonary infection in the murine inhalation model of cryptococcosis but still causes brain infection in a murine intravenous injection model, suggesting that the block of dissemination of the fbp1 Δ mutant is due to its inability to leave the lung. The fbp1 Δ mutant showed a defect in intracellular proliferation after phagocytosis in a Cryptococcus -macrophage interaction assay, which likely contributes to its virulence attenuation. To elucidate the molecular basis of the SCF(Fbp1) E3 ligase function, we analyzed potential Fbp1 substrates based on proteomic approaches combined with phenotypic analysis. One substrate, the inositol phosphosphingolipid-phospholipase C1 (Isc1), is required for fungal survival inside macrophage cells, which is consistent with the role of Fbp1 in regulating Cryptococcus -macrophage interaction and fungal virulence. Our results thus reveal a new determinant of fungal virulence that involves the posttranslational regulation of inositol sphingolipid biosynthesis.

Silverleaf whitefly (SLW), Bemisia argentifolii Bellows and Perring, is one of the most noxious pests of numerous field and vegetable crops, causing billions of dollars worth of damage throughout the world. SLW is a phloem feeder whose feeding is likely to interfere with phloem transport. The aim of this study was to test the hypothesis that SLW infestation impairs carbohydrate export from source leaves, and consequently increases their carbohydrate content. The youngest fully expanded leaves of cotton (Gossypium hirsutum L., cv. Siv'on), grown under SLW-infested and noninfested conditions, were characterized for their diurnal changes in carbohydrate content and photoassimilate export. SLW infestation induced a considerable reduction in net photosynthetic rate (P n ), coupled with increased sucrose, glucose and fructose and decreased starch concentrations. Export rate was determined after 14 CO 2 pulse-labeling both by in situ monitoring of leaf radioactivity and by analyzing the content and radioactivity of the major carbon metabolites. Radioactive counting indicated a lower rate of 14 C efflux for the infested plants. A similar trend was found for the specific activities of sucrose and the three soluble sugars combined (sucrose, glucose and fructose). A single exponential decay function with asymptote was fitted to the above efflux curves. All the calculated exponential coefficients demonstrated lower export rates after SLW injury. These results indicate that SLW impairs photoassimilate export, suggesting possible down-regulation of P n due to increased foliar soluble sugar contents.

The silverleaf whitefly (SLW), Bemisia argentifolii (Bellows and Perring), has been a severe pest of numerous field and vegetable crops for many years. Nevertheless, little has been reported on the complex interactions between SLW and its host plants. This study was aimed at investigating the effects of SLW on cotton (Gossypium hirsutum L.) photosynthesis and productivity under various irrigation and N fertilization regimes. Cotton plants (cv. Siv'on) were examined during an entire growing season in two factorial-designed screenhouse experiments consisting of two irrigation regimes (I) × two N application rates × two SLW levels (noninfested vs. infested). Buildup of the SLW population was markedly higher on N-fertilized plants than on N-deficient plants. Seed cotton yield was reduced by SLW infestation under all combinations of I × N treatments by about 40%. Yield reduction (noninfested minus infested) was significantly correlated with the number of SLW adults on the maximally infested leaf at peak flowering (r 2 = 0.94, P < 0.05 in 1995; r 2 = 0.91, P < 0.05 in 1996). Photosynthetic rate decreased 50% at about 60 d after SLW introduction. The relative photosynthetic rate (infested/noninfested) was significantly correlated with SLW adult density on the maximally infested leaf (r 2 = 0.72, P < 0.001 in 1995; r 2 = 0.53, P < 0.01 in 1996). The reduced photosynthetic rate was associated with reductions in chlorophyll variable fluorescence (Fv/Fm) and fluorescence yield. No substantial changes were found in stomatal conductance, intercellular CO 2 concentration, and leaf chlorophyll content. These findings indicate that SLW infestation impairs, either directly or indirectly, the photochemical reaction of the photosynthetic system in cotton plants.

Cryptococcus is a major fungal pathogen that frequently causes systemic infection in patients with compromised immunity. Glucose, an important signal molecule and the preferred carbon source for Cryptococcus, plays a critical role in fungal development and virulence. Cryptococcus contains more than 50 genes sharing high sequence homology with hexose transporters in Saccharomyces cerevisiae. However, there is no report on their function in glucose sensing or transport. In this study, we investigated two hexose transporter-like proteins (Hxs1 and Hxs2) in Cryptococcus that share the highest sequence identity with the glucose sensors Snf3 and Rgt2 in S. cerevisiae. The expression of HXS1 is repressed by high glucose, while the HXS2 expression is not regulated by glucose. Functional studies showed that Hxs1 is required for fungal resistance to oxidative stress and fungal virulence. The hxs1Δ mutant exhibited a significant reduction in glucose uptake activity, indicating that Hxs1 is required for glucose uptake. Heterologous expression of Cryptococcus HXS1 rendered the S. cerevisiae mutant lacking all 20 hexose transporters a high glucose uptake activity, demonstrating that Hxs1 functions as a glucose transporter. Heterologous expression of HXS1 in the snf3Δ rgt2Δ double mutant did not complement its growth in YPD medium containing the respiration inhibitor antimycin A, suggesting that Hxs1 may not function as a glucose sensor. Taken together, our results demonstrate that Hxs1 is a high-affinity glucose transporter and required for fungal virulence.

The disease cryptococcosis, caused by the fungus Cryptococcus neoformans, is acquired directly from environmental exposure rather than transmitted person-to-person. One explanation for the pathogenicity of this species is that interactions with environmental predators select for virulence. However, co-incubation of C. neoformans with amoeba can cause a “switch” from the normal yeast morphology to a pseudohyphal form, enabling fungi to survive exposure to amoeba, yet conversely reducing virulence in mammalian models of cryptococcosis. Like other human pathogenic fungi, C. neoformans is capable of microevolutionary changes that influence the biology of the organism and outcome of the host-pathogen interaction. A yeast-pseudohyphal phenotypic switch also happens under in vitro conditions. Here, we demonstrate that this morphological switch, rather than being under epigenetic control, is controlled by DNA mutation since all pseudohyphal strains bear mutations within genes encoding components of the RAM pathway. High rates of isolation of pseudohyphal strains can be explained by the physical size of RAM pathway genes and a hypermutator phenotype of the strain used in phenotypic switching studies. Reversion to wild type yeast morphology in vitro or within a mammalian host can occur through different mechanisms, with one being counter-acting mutations. Infection of mice with RAM mutants reveals several outcomes: clearance of the infection, asymptomatic maintenance of the strains, or reversion to wild type forms and progression of disease. These findings demonstrate a key role of mutation events in microevolution to modulate the ability of a fungal pathogen to cause disease., Author Summary Many diseases are contracted from the environment, rather than from sick people. It is unclear why those species are able to cause disease, since the selective pressures in the environment are presumed to be very different from those found within the host. Cryptococcus neoformans is a fungus that causes life-threatening lung and central nervous system disease in approximately one million people each year. The fungus is inhaled from environmental sources. One hypothesis to account for C. neoformans virulence is that amoeba are predators for this fungus, and surviving strains are pre-selected to be virulent in the human host. On the other hand, experiments have found that amoeba eat C. neoformans. A pseudohyphal cell type can survive, and while protecting against amoeba these cells are unable to cause disease in mouse models. We predicted that the pseudohyphal morphology reflected a change in function of a pathway of genes, and found that all pseudohyphal isolates contain mutations within genes for this pathway. The pseudohyphal trait is unstable, with reversion to normal yeast growth by counter-acting mutations. These mutations can occur during the course of mammalian infection. Our results show that mutation events account for a microevolution system currently described as phenotypic switching, and that mutations, at least under experimental conditions, can regulate pathogen adaptation and influence its host range.

Changes in the roots exudation of low molecular weight organic acids (LMWOA), cadmium (Cd) accumulation and translocation and soil solution pH in Solanum nigrum L., the Cd-hyperaccumulator were examined and compared with a non-hyperaccumulator Solanum lycopersicum L. The differences were significant (P < 0.05 or < 0.01) between S. nigrum and S. lycopersicum for all the five LMWOA at all soil levels. The total amount of LMWOA exuded by S. nigrum was significantly higher than that of S. lycopersicum (P < 0.05 or < 0.01). The S. nigrum accumulated and translocated more Cd than S. lycopersicum . The soil solution pH of S. nigrum was significantly lower than S. lycopersicum . The S. nigrum with higher concentrations of LMWOA in roots exudation accumulated more Cd in the plants. The results indicated tha LMWOA secretion by S. nigrum root, especially in Cd-contaminated soils, was likely to be an important role in Cd hyperaccumulation. Key words : Cadmium, Solanum nigrum L., low molecular weight organic acids, root exudates.

The objective of this research was to determine the effect of Fe status on the uptake of cadmium (Cd) by leaf red beet (Beta vulgaris var.cicla L.) in hydroponic culture. We also investigated the influence of Fe deficiency on rhizosphere pH, bioconcentration factor (BCF) and translocation factor (TF) in plants. After the seedlings had grown for five weeks, the seedlings were exposed for 4 days at at the concentrations of Cd 0 mg/L (control), 0.1 mg/L and 1 mg/L. The results showed that Fe deficiency induced Cd uptake, biomass decrease and solution pH decrease in hydroponic culture by plants. Fe-deficient plants BCF and TF were higher compared to Fe sufficient ones.

Cryptococcus neoformans is an AIDS-associated human fungal pathogen and the most common cause of fungal meningitis, with a mortality rate over 40% in AIDS patients. Significant advances have been achieved in understanding its disease mechanisms. Yet the underlying mechanism of a high frequency of cryptococcal meningitis remains unclear. The existence of high inositol concentrations in brain and our earlier discovery of a large inositol transporter ( ITR ) gene family in C. neoformans led us to investigate the potential role of inositol in Cryptococcus -host interactions. In this study, we focus on functional analyses of two major ITR genes to understand their role in virulence of C. neoformans . Our results show that ITR1A and ITR3C are the only two ITR genes among 10 candidates that can complement the growth defect of a Saccharomyces cerevisiae strain lacking inositol transporters. Both S. cerevisiae strains heterologously expressing ITR1A or ITR3C showed high inositol uptake activity, an indication that they are major inositol transporters. Significantly, itr1a itr3c double mutants showed significant virulence attenuation in murine infection models. Mutating both ITR1A and ITR3C in an ino1 mutant background activates the expression of several remaining ITR candidates and does not show more severe virulence attenuation, suggesting that both inositol uptake and biosynthetic pathways are important for inositol acquisition. Overall, our study provides evidence that host inositol and fungal inositol transporters are important for Cryptococcus pathogenicity.

The pathogenic fungus, Magnaporthe oryzae, has emerged as a model system for dissecting fungal-plant interactions. Considerable progress has been made in identifying genes necessary for the regulation of pre- and post-penetration events in the development and pathogenicity of the fungus, such as the appressorium (a dome-shaped infection structure) mediated penetration and invasive growth of the pathogen. However, the detailed molecular mechanisms of these developmental stages, especially appressorium morphogenesis, are still relatively poorly understood. Currently, we isolated the MNH6 gene for a nonhistone chromosomal protein and 24 autophagic genes from the genome of this fungus. These genes are highly conserved among other eukaryotes, including humans and plants, and they control the capability of surviving starvation, conidiation, conidial germination, lipid turnover, and appressorium turgor generation. Clarification of the functions of these autophagy genes and their interaction networks will allow us to understand the role of autophagic genes in fungal pathogenesis.

Phytoremediation is a promising tool in removing pollutants from the environment or in rendering them harmless. Some previous reports showed that Fe deficiency in dicots induced the stimulated accumulation of a number of micronutrient metals other than Fe. The objective of this research was to determine the effect of Fe deficiency on the uptake of cadmium (Cd) by tomato (Lycopersicon. Esculentum Mill.) in hydroponic culture. Results showed that Fe deficiency induced cadmium uptake, biomass decrease and changes of pH and Eh in hydroponic culture. Under Fe deficiency status, the decrease in pH and the increase in Eh might result in higher cadmium availability. Values of bioconcentration factor (BCF) and translocation factor (TF) indicated that Fe-deficiency status affected cadmium accumulation and translocation in tomato. The results of this research would significantly improve the understanding of metal removal from the contaminated environment.

Autophagy is a ubiquitous degradative pathway for the bulk degradation of eukaryotic macromolecules and organelles in eukaryotic cells (Klionsky, 2005; Levine and Klionsky, 2004). Previously, the role of autophagy in turgor generation in plant pathogenic fungi was unknown. Currently, autophagy is confirmed as an important pathway for turgor accumulation in the appressorium (the tips of the invasive hyphae; Liu et al ., 2007b) using a technique of targeted gene replacement, deleting the genes that code for Magnaporthe oryzae homologs of yeast autophagy‐related ( ATG ) genes ATG2, ATG4, ATG5, ATG8, ATG9 , and ATG18 (Liu et al ., 2007a). All of these null mutants fail to breach the cuticle of the host. This chapter will first look at some methodologies to analyze the functions of autophagy‐related gene products at the biological, cellular, and molecular level in this model plant pathogenic fungi, and then provide some research evidence of the role of autophagy in the promotion of the formation of the infection structure and pathogenicity to point out some significant areas for further research in this field.

The promoter of NAR gene in Magnaporthe grisea was isolated and sequenced. The promoter sequences contained the “TATA” box, the “CAAT” box, and binding sites for fungal regulatory proteins. Programs that predict promoter sequences indicated that promoter sequence lies between locations 430 and 857 of the NAR promoter fragment. GFP expression under the NAR promoter and NAR transcript analysis revealed that this promoter is activated primarily at the mycelial stage in the rice blast fungus and could be used to express native or extrinsic genes in the mycelia of the rice blast fungus.

We have constructed a fungal subtractive suppressive library enriched in genes expressed during appressorium maturation in Magnaporthe grisea. Sequencing of 250 clones from the subtracted appressorium cDNA library revealed 142 unique genes, represented by 155 non-redundant ESTs (expressed sequence tags). Of these ESTs, 72 have not been previously isolated in M. grisea. RT-PCR analysis of 105 of the genes discovered found transcripts corresponding to 71 of the ESTs only in mature appressoria while transcripts corresponding to a further 34 of the isolated ESTs were expressed both in appressoria and conidia/mycelia. Genes specifically expressed in appressorium identified by SSH included a number that have been previously implicated in appressorium formation or function including GAS1, GAS3, and PTH11.

Cryptococcus neoformans is the most common cause of fungal meningitis, with high mortality and morbidity. The reason for the frequent occurrence of Cryptococcus infection in the central nervous system (CNS) is poorly understood. The facts that human and animal brains contain abundant inositol and that Cryptococcus has a sophisticated system for the acquisition of inositol from the environment suggests that host inositol utilization may contribute to the development of cryptococcal meningitis. In this study, we found that inositol plays an important role in Cryptococcus traversal across the blood-brain barrier (BBB) both in an in vitro human BBB model and in in vivo animal models. The capacity of inositol to stimulate BBB crossing was dependent upon fungal inositol transporters, indicated by a 70% reduction in transmigration efficiency in mutant strains lacking two major inositol transporters, Itr1a and Itr3c. Upregulation of genes involved in the inositol catabolic pathway was evident in a microarray analysis following inositol treatment. In addition, inositol increased the production of hyaluronic acid in Cryptococcus cells, which is a ligand known to binding host CD44 receptor for their invasion. These studies suggest an inositol-dependent Cryptococcus traversal of the BBB, and support our hypothesis that utilization of host-derived inositol by Cryptococcus contributes to CNS infection., Author Summary Cryptococcus neoformans is an AIDS-associated human fungal pathogen that annually causes over 1 million cases of meningitis world-wide, and more than 600,000 attributable deaths. Cryptococcus often causes lung and brain infection and is the leading cause of fungal meningitis in immunosuppressed patients. Why Cryptococcus frequently infects the central nervous system to cause fatal meningitis is an unanswered critical question. Our previous studies revealed a sophisticated inositol acquisition system in Cryptococcus that plays a central role in utilizing environmental inositol to complete its sexual cycle. Here we further demonstrate that inositol acquisition is also important for fungal infection in the brain, where abundant inositol is available. We found that inositol promotes the traversal of Cryptococcus across the blood-brain barrier (BBB), and such stimulation is fungal inositol transporter dependent. We also identified the effects of host inositol on fungal cellular functions that contribute to the stimulation of fungal penetration of the BBB. We propose that inositol utilization is a novel virulence factor for CNS cryptococcosis. Our work lays an important foundation for understanding how fungi respond to available host inositol and indicates the impact of host inositol acquisition on the development of cryptococcal meningitis.

Reduced plant productivity induced by silverleaf whitefly (SLW), Bemisia argentifolii Bellows and Perring, is often associated with reduced net photosynthetic rate (P n ). This study was aimed at characterizing the effects of SLW on photosynthetic activity in cotton and the underlying mechanisms. Cotton (Gossypium hirsutum L. cv. Siv'on) plants were examined under SLW-infested and non-infested conditions and subjected to measurements of gas exchange, chlorophyll content and chlorophyll fluorescence. SLW infestation significantly reduced the P n of cotton by 22-54% throughout the day as well as throughout most of the examined ranges of photosynthetic photon flux density (PPFD) and CO 2 concentrations. Stomatal conductance was also reduced by SLW injury, while intercellular CO 2 concentration (C i ) was not affected, suggesting that stomatal limitation is not the major factor limiting P n . This conclusion was further supported by estimates of stomatal limitation, based on P n /C i curves, which were not affected by SLW infestation. On the other hand, non-stomatal limitation induced by SLW was over 2-fold higher than stomatal limitation. Chlorophyll a and b contents were not modified by SLW infestation. Fluorescence ratio (F v /F m ) and the effective quantum yield of PSII were reduced by less than 10%; thus, impaired photochemical reaction is probably not the only factor limiting P n under SLW infestation. In conclusion, SLW-induced P n reduction in cotton was due to non-stomatal factors, possibly induced by impaired carbon fixation and/or carbohydrate export.