Mealybug salivary microbes inhibit induced plant defenses.

BACKGROUND: Phenacoccus solenopsis is a polyphagous invasive mealybug that caused serious damage to crops worldwide. Phloem-sucking hemipterans are known to carry symbiotic microbes in their saliva. However, the role of salivary bacteria of P. solenopsis in modulating plant defenses remains limited. Exploring the impact of salivary bacteria on plant defense responses will contribute to the development of new targets for efficient control of invasive mealybugs. RESULTS: Salivary bacteria of the invasive mealybug P. solenopsis can suppress herbivore-induced plant defenses and thus enhance mealybug fitness. Mealybugs treated with an antibiotic showed decreased weight gain, fecundity and survival. Untreated mealybugs suppressed jasmonic acid (JA)-regulated defenses but activated salicylic acid (SA)-regulated defenses in cotton plants. In contrast, antibiotic-treated mealybugs triggered JA-responsive gene expression and JA accumulation, and showed shortened phloem ingestion. Reinoculating antibiotic-treated mealybugs with Enterobacteriaceae or Stenotrophomonas cultivated from mealybug saliva promoted phloem ingestion and fecundity, and restored the ability of mealybugs to suppress plant defenses. Fluorescence in situ hybridization visualization revealed that Enterobacteriaceae and Stenotrophomonas colonize salivary glands and are secreted into the mesophyll cells and phloem vessels. Exogenous application of the bacterial isolates to plant leaves inhibited JA-responsive gene expression and activated SA-responsive gene expression. CONCLUSION: Our findings imply that symbiotic bacteria in the saliva of the mealybug play an important role in manipulating herbivore-induced plant defenses, enabling this important pest to evade induced plant defenses and promoting its performance and destructive effects on crops. © 2023 Society of Chemical Industry.

The role of ectopic P granules protein 5 homolog (EPG5) in DHPG-induced pain sensitization in mice.

Nociplastic pain is a severe health problem, while its mechanisms are still unclear. (R, S)-3,5-Dihydroxyphenylglycine (DHPG) is a group I metabotropic glutamate receptor (mGluR) agonist that can cause central sensitization, which plays a role in nociplastic pain. In this study, after intrathecal injection of 25 nmol DHPG for three consecutive days, whole proteins were extracted from the L4~6 lumbar spinal cord of mice 2 h after intrathecal administration on the third day for proteomics analysis. Based on the results, 15 down-regulated and 20 up-regulated proteins were identified in mice. Real-time quantitative PCR (RT-qPCR) and western blotting (WB) revealed that the expression of ectopic P granules protein 5 homolog (EPG5) mRNA and protein were significantly up-regulated compared with the control group, which was consistent with the proteomics results. Originally identified in the genetic screening of Caenorhabditis elegans, EPG5 is mainly involved in regulating autophagy in the body, and in our study, it was mainly expressed in spinal neurons, as revealed by immunohistochemistry staining. After the intrathecal injection of 8 µL adeno-associated virus (AAV)-EPG5 short hairpin RNA (shRNA) to knock down spinal EPG5, the hyperalgesia caused by DHPG was relieved. Altogether, these results suggest that EPG5 plays an important role in DHPG-induced pain sensitization in mice.

SDC1-dependent TGM2 determines radiosensitivity in glioblastoma by coordinating EPG5-mediated fusion of autophagosomes with lysosomes.

Glioblastoma multiforme (GBM) is the most common brain malignancy insensitive to radiotherapy (RT). Although macroautophagy/autophagy was reported to be a fundamental factor prolonging the survival of tumors under radiotherapeutic stress, the autophagic biomarkers coordinated to radioresistance of GBM are still lacking in clinical practice. Here we established radioresistant GBM cells and identified their protein profiles using tandem mass tag (TMT) quantitative proteomic analysis. It was found that SDC1 and TGM2 proteins were overexpressed in radioresistant GBM cells and tissues and they contributed to the poor prognosis of RT. Knocking down SDC1 and TGM2 inhibited the fusion of autophagosomes with lysosomes and thus enhanced the radiosensitivity of GBM cells. After irradiation, TGM2 bound with SDC1 and transported it from the cell membrane to lysosomes, and then bound to LC3 through its two LC3-interacting regions (LIRs), coordinating the encounter between autophagosomes and lysosomes, which should be a prerequisite for lysosomal EPG5 to recognize LC3 and subsequently stabilize the STX17-SNAP29-VAMP8 QabcR SNARE complex assembly. Moreover, when combined with RT, cystamine dihydrochloride (a TGM2 inhibitor) extended the lifespan of GBM-bearing mice. Overall, our findings demonstrated the EPG5 tethering mode with SDC1 and TGM2 during the fusion of autophagosomes with lysosomes, providing new insights into the molecular mechanism and therapeutic target underlying radioresistant GBM.Abbreviations: BafA1: bafilomycin A1; CQ: chloroquine; Cys-D: cystamine dihydrochloride; EPG5: ectopic P-granules 5 autophagy tethering factor; GBM: glioblastoma multiforme; GFP: green fluorescent protein; LAMP2: lysosomal associated membrane protein 2; LIRs: LC3-interacting regions; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; NC: negative control; RFP: red fluorescent protein; RT: radiotherapy; SDC1: syndecan 1; SNAP29: synaptosome associated protein 29; SQSTM1/p62: sequestosome 1; STX17: syntaxin 17; TGM2: transglutaminase 2; TMT: tandem mass tag; VAMP8: vesicle associated membrane protein 8; WT: wild type.

Chromosome Instability in Pony of Esperia Breed Naturally Infected by Intestinal Strongylidae.

The Pony of Esperia is an Italian autochthonous horse breed reared in the wild on the Aurunci and Ausoni Mountains. Currently, it is considered an endangered breed, as its population consists of 1623 animals. It is therefore essential to identify all aspects that can improve the management and economy of its breeding, favoring its diffusion. In this paper, the effects of intestinal strongyle infection on the chromosome stability of peripheral blood lymphocytes (PBLs) was evaluated through aneuploidy and chromosome aberration (gap, chromatid and chromosome breaks, and the number of abnormal cells) test. Statistical difference in the mean values of aneuploidy, cells with chromosome abnormalities, and chromosome and chromatid breaks were observed between ponies with high fecal egg counts (eggs per gram > 930) and those with undetectable intestinal strongylosis. The causes of this phenomenon and possible repercussions on the management of Pony of Esperia are discussed in the paper.

Nicotiana tabacum as a dead-end trap for adult Diaphorina citri: A potential biological tactic for protecting citrus orchards.

The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), is a key vector of the causal agents of Huanglongbing (HLB), a devastating disease affecting citrus almost worldwide. Nicotiana tabacum L. is an important commercial crop in China. Field observations suggested that D. citri adults die on N. tabacum leaves when grown nearby citrus orchards. In this study, the preference for and survivorship of D. citri adults on N. tabacum and their feeding behavior were investigated. The results showed that D. citri adults were attracted to N. tabacum and to the green leaf volatiles (GLVs) (Z)-3-hexenol and (E)-2-hexenol. The survival of D. citri adults on N. tabacum was less than 30 h, which was shorter than that for adults without food (35 h) and on a suitable host Murraya exotica L. (29 days). Electrical penetration graph (EPG) recordings revealed that the pathway phase of D. citri on N. tabacum leaves consisted of four waveforms-the non-probing phase (NP), the pathway phase (PP, including intercellular probing of activity in the phloem (C) and phloem penetration (D)), phloem salivation (E1), and phloem ingestion (E2). Diaphorina citri only secreted saliva and ingested sap from phloem on N. tabacum leaves and spent the longest duration in phloem sap ingestion (E2). Moreover, L-nicotine, an important defense compound against insects in N. tabacum plants, was highly toxic to D. citri. These results suggested that N. tabacum plants could help to sustainably control the spread of D. citri and HLB when growing in and around citrus orchards.

Intestinal antiviral signaling is controlled by autophagy gene Epg5 independent of the microbiota.

Mutations in the macroautophagy/autophagy gene EPG5 are responsible for Vici syndrome, a human genetic disease characterized by combined immunodeficiency. Previously, we found that epg5-/- mice exhibit hyperinflammation in the lungs mediated by IL1B/IL-1ß and TNF/TNFα, resulting in resistance to influenza. Here, we find that disruption of Epg5 results in protection against multiple enteric viruses including norovirus and rotavirus. Gene expression analysis reveals IFNL/IFN-λ responsive genes as a key alteration. Further, mice lacking Epg5 exhibit substantial alterations of the intestinal microbiota. Surprisingly, germ-free mouse studies indicate Epg5-associated inflammation of both the intestine and lung is microbiota-independent. Genetic studies support IFNL signaling as the primary mediator of resistance to enteric viruses, but not of microbial dysbiosis, in epg5-/- mice. This study unveils an important role, unexpectedly independent of the microbiota, for autophagy gene Epg5 in host organism protection by modulating intestinal IFNL responses.Abbreviations: CTNNB1: catenin (cadherin associated protein), beta 1; DAPI: 4',6-diamidino-2-phenylindole; EPG5: ectopic P-granules autophagy protein 5 homolog (C. elegans); FT: fecal transplant; IFI44: interferon-induced protein 44; IFIT1: interferon-induced protein with tetratricopeptide repeats 1; IFNG/IFN-γ: interferon gamma; IFNL/IFN-λ: interferon lambda; IFNLR1: interferon lambda receptor 1; IL1B/IL-1ß: interleukin 1 beta; ISG: interferon stimulated gene; GF: germ-free; LEfSe: linear discriminant analysis effect size; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MNoV: murine norovirus; MX2: MX dynamin-like GTPase 2; OAS1A: 2'-5' oligoadenylate synthetase 1A; RV: rotavirus; SPF: specific-pathogen free; SQSTM1/p62: sequestosome 1; STAT1: signal transducer and activator of transcription 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK-binding kinase 1; TNF/TNFα: tumor necrosis factor.

Can orange essential oil reduce the severity of parasitic infection in sheep?

In this study, a three-step experiment was conducted to evaluate the effectiveness of orange essential oil (OEO) to reduce the severity of parasitic infection in lambs naturally infected with gastrointestinal parasites. In the first and second stages of the experiment, the lambs were distributed in groups (n = 10): without treatment (control), treated with levamisole (7.5 mg/kg), and treated either with 200, 300 or 400 mg/kg OEO in the first stage and with two doses of 300 or 350 mg/kg each of OEO with a six-hour interval in the second stage. In the third stage of the experiment, 20 lambs were distributed into control and treated groups (two doses of 350 mg/kg OEO) and slaughtered for determining the parasite count in the abomasum and small intestine. Evaluations were performed on D0, D3, D7, D10, and D14 by determining the egg count per gram of feces (EPG), and the percentage of EPG reduction was calculated in comparison to D0. Coproculture was performed on days D0 and D14 of the first and second stages of the study. In the first stage, the EPG increased by 20.90 and 100.99% and decreased by 5.54% on D14 in the groups treated with 200, 300, or 400 mg/kg OEO, respectively. At the second stage, the EPG of the OEO groups decreased by more than 96% on D3, but on D14 was decreased by 0.82% and 77.07% in the 2 × 300 and 2 × 350 mg/kg groups, respectively. Haemonchus sp. was the highest percentage of collected larvae in all coproculture tests, ranging from 90 to 100%. In the third stage, there was no difference (P > 0.05) in the mean number of parasites between the OEO groups and the control. Although the decrease in EPG of more than 96% on D3 did not persist in the other evaluations, and immature forms and adult parasites were collected at slaughter, the EPG reduction of 77.07% on D14 with the dose of 700 mg/kg (OEO2 × 350) suggests that OEO can help to reduce the severity of parasitic infection and, consequently, to diminish pastures contamination, when used together with other methods to control gastrointestinal parasites in sheep.

The application of nanoparticles in cancer immunotherapy: Targeting tumor microenvironment.

The tumor development and metastasis are closely related to the structure and function of the tumor microenvironment (TME). Recently, TME modulation strategies have attracted much attention in cancer immunotherapy. Despite the preliminary success of immunotherapeutic agents, their therapeutic effects have been restricted by the limited retention time of drugs in TME. Compared with traditional delivery systems, nanoparticles with unique physical properties and elaborate design can efficiently penetrate TME and specifically deliver to the major components in TME. In this review, we briefly introduce the substitutes of TME including dendritic cells, macrophages, fibroblasts, tumor vasculature, tumor-draining lymph nodes and hypoxic state, then review various nanoparticles targeting these components and their applications in tumor therapy. In addition, nanoparticles could be combined with other therapies, including chemotherapy, radiotherapy, and photodynamic therapy, however, the nanoplatform delivery system may not be effective in all types of tumors due to the heterogeneity of different tumors and individuals. The changes of TME at various stages during tumor development are required to be further elucidated so that more individualized nanoplatforms could be designed.

Impact of supplementation with different protein sources on the parasitological profile of ovine matrices and development of lambs / Impacto da suplementação com diferentes fontes protéicas no perfil parasitológico de matrizes ovinas e no desenvolvimento de cordeiros

The aim was to evaluate the impact of supplementation with different protein sources from agro-industrial production on the parasitological profile of ovine matrices and the development of their offspring. The protein sources used were: soybean bran, cotton pie, babassu pie, and coconut pie. A total of 60 ovine matrices were used, with confirmed pregnancy and supplemented in the final third of gestation at 0.5% of live weight in pastures of deferred Marandu grass. The evaluations occurred from the final third of gestation and in the lactation phase until weaning. The data collected in the matrices were: counting of eggs per gram of feces (EPG) and identification of infective larvae (Faecal test), evaluation of the degree of anemia by the FAMACHA© method, body weight (kg), and body condition score (BCS). In the lambs, the biometric measurements were taken from the 30 days of life until weaning. In Faecal test, the prevalence was 98% of Haemonchus spp. There was no effect of the protein sources for EPG (3476.0) and FAMACHA©. However, in the mats supplemented with coconut pie were observed the lowest body weights (36.9 kg) and lower BCS (2.7). Babassu, coconut and cotton cakes, and soybean meal used as a protein source in the supplementation of ovine matrices kept in deferred pastures of Marandu grass at the 0.5% level of live weight maintain the production indexes consistent with the physiology stage of the matrices even with a high endo-parasitic load.

O objetivo foi avaliar o impacto da suplementação com diferentes fontes proteicas da produção agroindustrial no perfil parasitológico das matrizes ovinas e no desenvolvimento de seus filhotes. As fontes de proteína utilizadas foram: farelo de soja, torta de algodão, torta de babaçu e torta de coco. Foram utilizadas 60 matrizes ovinas, com prenhez confirmada e suplementadas no terço final da gestação com 0,5% do peso vivo em pastagens de capim Marandu diferido. As avaliações ocorreram desde o terço final da gestação e na fase de lactação até o desmame. Os dados coletados nas matrizes foram: contagem de ovos por grama de fezes (OPG) e identificação de larvas infectantes (teste fecal), avaliação do grau de anemia pelo método FAMACHA ©, peso corporal (kg) e escore de condição corporal (ECC). Nos cordeiros, as medidas biométricas foram realizadas a partir dos 30 dias de vida até o desmame. No teste fecal, a prevalência foi de 98% de Haemonchus spp. Não houve efeito das fontes de proteína para OPG (3476,0) e FAMACHA ©. No entanto, nas matrizes suplementadas com torta de coco foram observados os menores pesos corporais (36,9 kg) e menorECC (2,7). Torta de Babaçu, coco, algodão e farelo de soja utilizados como fonte protéica na suplementação de matrizes ovinas mantidas em pastos diferidos de capim Marandu no nível de 0,5% de peso vivo mantêm os índices de produção consistentes com o estágio fisiológico das matrizes, mesmo com uma carga endo-parasitária alta.

Implication of the fecal egg count reduction test (FECRT) in sheep for better use of available drugs / Implicação do teste de redução da contagem de ovos nas fezes (TRCOF) em ovinos para melhor uso dos anti-helmínticos disponíveis

Abstract The aim here is to present data on the efficacy of anthelmintics in sheep flocks in Rio de Janeiro, Brazil, and to discuss the interpretation of the fecal egg count reduction test (FECRT) for each nematode genus. Fecal eggs counts and pre- and post-treatment coprocultures were performed, the former to evaluate the efficacy of and the latter to determine the overall parasite prevalence. An additional efficacy test was performed at Farm # 1 a year after the initial test. Severe anthelmintic resistance was found for the flocks, with no FECRT sensitivity at any of the 22 farms evaluated. However, an analysis of the infective larvae showed that some drugs were effective against certain parasitic genera; i.e., levamisole was more effective against Haemonchus spp. and moxidectin against Trichostrongylus spp. In the additional FECRT performed at Farm # 1, moxidectin and nitroxynil were ineffective separately, but when applied in combination they were highly effective due to their efficacy against Haemonchus (nitroxynil) and Trichostrongylus (moxidectin), respectively. The use of the FECRT targeting the parasitic nematode species prevalent on farms may make it possible to choose more effective anthelmintics.

Resumo O objetivo deste trabalho foi apresentar dados sobre a eficácia de anti-helmínticos em rebanhos ovinos no Rio de Janeiro, Brasil, e discutir a interpretação do teste de redução da contagem de ovos nas fezes (TRCOF) para cada gênero de nematoide. A contagem de ovos fecais (OPG) e coprocultura pré e pós-tratamento foram realizadas para avaliar a eficácia e a prevalência geral do parasito, respectivamente. Um teste de eficácia adicional foi realizado na Fazenda # 1 após um ano do teste inicial. Resistência anti-helmíntica grave foi encontrada, não havendo sensibilidade no TRCOF em nenhuma das 22 fazendas avaliadas. No entanto, na análise das larvas infectantes observou-se que algumas drogas foram eficazes contra certos gêneros parasitários; por exemplo, o levamisol foi mais eficaz contra Haemonchus spp. e a moxidectina contra Trichostrongylus spp. No TRCOF adicional realizado na Fazenda 1, a moxidectina e o nitroxinil foram ineficazes separadamente, mas quando aplicados em combinação, foram altamente eficazes devido à sua eficácia contra Haemonchus spp. (nitroxinil) e Trichostrongylus spp. (moxidectina), respectivamente. O TRCOF visando às espécies de nematoides parasitas prevalentes nas fazendas pode possibilitar a escolha de anti-helmínticos mais eficazes.

C-myc/miR-150/EPG5 axis mediated dysfunction of autophagy promotes development of non-small cell lung cancer.

Rationale: Lung cancer is the leading cause of cancer death worldwide, and treatment options are limited to mainly cytotoxic agents. Here we reveal a novel role of miR-150 in non-small cell lung cancer (NSCLC) development and seek potential new therapeutic targets. Methods: The miR-150-mediated autophagy dysfunction in NSCLC cells were examined using molecular methods in vitro and in vivo. The upstream regulatory element and downstream target of miR-150 were identified in vitro and validated in vivo. Potential therapeutic methods (anti-c-myc or anti-miR-150) were tested in vitro and in vivo. Clinical relevance of the c-myc/miR-150/EPG5 axis in NSCLC was validated in human clinical samples and large genomics database. Results: miR-150 blocked the fusion of autophagosomes and lysosomes through directly repressing EPG5. The miR-150-mediated autophagy defect induced ER stress and increased cellular ROS levels and DNA damage response, and promoted NSCLC cell proliferation and tumor growth. Knockdown of EPG5 promoted NSCLC cell proliferation, and attenuated the effects of miR-150. c-myc gene was identified as a miR-150 transcriptional factor which increased miR-150 accumulation, therefore pharmacologically or genetically inhibiting c-myc/miR-150 expression significantly inhibited NSCLC cell growth in vitro and in vivo. Both c-myc and miR-150 were significantly over-expressed in NSCLC, while EPG5 was down-regulated in NSCLC. Expression levels of these molecules were well correlated, and also well correlated with patient survival. Conclusions: Our findings suggest that c-myc/miR-150/EPG5 mediated dysfunction of autophagy contributes to NSCLC development, which may provide a potential new diagnostic and therapeutic target in NSCLC.

Body position of the stink bug Dichelops melacanthus (Dallas) during feeding from stems of maize seedlings / Posição do corpo do percevejo Dichelops melacanthus (Dallas) durante a alimentação em hastes de plântulas de milho

Abstract The Neotropical green-belly stink bug, Dichelops melacanthus (Dallas) is a major pest of maize, Zea mays L. in the main production areas of Brazil. It usually feeds on the stems of young plants (seedlings) causing heavy damage by affecting the plant growth and mitigating seed yield. Laboratory studies were conducted to determine body position (upward or downward) of the bug on plant (seedling) stem during feeding and not feeding activities. Ten visual records were taken per day, each spaced one hour during 30 days of 10 adult bugs of similar age exposed to maize seedlings inside cages (plastic tubes). At each observation, it was recorded if the bug was feeding on the stem (i.e., stylets inserted into the plant tissue) or not, and its body position. During feeding, waveforms were recorded using the EPG (electropenetrography) technique, which were correlated with histological studies to reveal the feeding sites. Results indicated that when they were feeding, the majority of the bugs were in the downward position. In contrast, when the bugs were on the plants, and not feeding, they were mostly in the upward position. Waveforms generated using the EPG coupled with histological studies demonstrated that during ingestion bugs fed from the xylem vessels and from the parenchyma tissue using cell rupture strategy in the latter. No clear explanation was found to explain the preferred downward body position during ingestion, but some hypothesis are speculated.

Resumo O percevejo barriga-verde, Dichelops melacanthus (Dallas) é uma praga importante do milho, Zea mays L. nas principais áreas produtoras do Brasil. Usualmente alimenta-se nas hastes de plantas jovens (plântulas) causando danos severos no seu crescimento e reduzindo o rendimento de grãos. Estudos foram conduzidos em laboratório para determinar a posição do corpo (voltado para cima ou para baixo) dos percevejos nas hastes das plântulas de milho durante as atividades de alimentação e não-alimentação. Foram feitas dez observações por dia, espaçadas por uma hora, durante 30 dias em 10 percevejos adultos com idade semelhante sobre plântulas de milho colocadas em gaiolas (tubos de plástico). Em cada observação, anotou-se se o percevejo estava se alimentando (i.e., estiletes bucais inseridos no tecido vegetal) ou não, e a posição do corpo. Durante a alimentação, ondas eletromagnéticas foram registradas utilizando-se o EPG (técnica da eletropenetrografia), as quais foram correlacionadas com estudos histológicos para revelar os locais de alimentação. Os resultados indicaram que quando os percevejos estavam se alimentando, a maioria estava voltado para baixo. Em contraste, os percevejos sem se alimentar nas plântulas estavam a maioria voltados para cima. As ondas geradas pelo EPG junto com os estudos histológicos revelaram que os percevejos se alimentaram dos vasos do xilema e do tecido parenquimatoso usando a estratégia de ruptura celular no último. Não foi encontrada uma explicação clara para a preferência dos percevejos em se alimentarem na posição voltados para baixo, mas algumas hipóteses são especuladas.

Cytokine profiles of Necator americanus and Plasmodium falciparum co-infected patients in rural Ghana.

BACKGROUND: Necator americanus (hookworm) and Plasmodium falciparum co-infections are common in endemic communities in rural Ghana. Human immune responses to P. falciparum and hookworm are complex, and the dynamics of cytokine levels and effector mediators are poorly understood. This study aimed to determine the effect of hookworm and P. falciparum co-infection on parasite intensities and cytokine profiles in individuals before and after deworming drug treatment. METHODS: In this cross-sectional study conducted in the Kintampo North Municipality of Ghana blood and stool samples were analyzed from 984 participants (aged 4-88 years). Stool samples were collected at baseline from all participants and examined for the presence of hookworm using the Kato-Katz method. Blood and stool samples were analysed again two weeks after albendazole treatment of hookworm infected individuals. Malaria parasitaemia was estimated by light microscopy and P. falciparum-specific 18S rRNA gene PCR method used for species identification. Serum levels of circulating cytokines interleukins -5, -10 (IL-5, IL-10), tumor necrosis factor [TNF]-α, and eotaxin [CCL11] were determined using ELISA based methods. RESULTS: Malaria parasitaemia was significantly reduced in hookworm and P. falciparum co-infected individuals (p = 0.0018) while hookworm intensity was similar between groups. IL-10 level was significantly higher in the co-infected individuals (39.9 ±â€¯12.2 pg/ml) compared to the single infected or the uninfected group (10.7 ±â€¯7.6 mg/ml). IL-5 level was higher in the hookworm only infected individual. TNF-α levels were higher in all infected groups compared to the uninfected controls. CCL11 levels were significantly higher in subjects infected with hookworm only or co-infected with hookworm and P. falciparum. There was a significantly negative correlation (rs = -0.39, p = 0.021) between hookworm eggs per gram of stool and CCL11 levels in the group mono-infected with hookworm which was not affected by treatment. Treatment with albendazole led to a significant reduction of TNF-α (p = 0.041), IL-5 (p = 0.01) and IL-10 (p = 0.001) levels. CONCLUSION: This study shows that in the absence of other helminths, co-infection of hookworm with P. falciparum may modulate blood parasitemia levels and cytokine responses. Data also show that deworming drug treatment alters these cytokine profiles in hookworm infected subjects. Future studies to elucidate the potential mechanisms underlying these observations should include an assessment of parasite specific cellular responses.

A non-canonical autophagy-dependent role of the ATG16L1T300A variant in urothelial vesicular trafficking and uropathogenic Escherichia coli persistence.

50% of Caucasians carry a Thr300Ala variant (T300A) in the protein encoded by the macroautophagy/autophagy gene ATG16L1. Here, we show that the T300A variant confers protection against urinary tract infections (UTIs), the most common infectious disease in women. Using knockin mice carrying the human T300A variant, we show that the variant limits the UTI-causing bacteria, uropathogenic Escherichia coli (UPEC), from establishing persistent intracellular reservoirs, which can seed UTI recurrence. This phenotype is recapitulated in mice lacking Atg16l1 or Atg7 exclusively in the urothelium. We further show that mice with the T300A variant exhibit urothelial cellular abnormalities, including vesicular congestion and aberrant accumulation of UPK (uroplakin) proteins. Importantly, presence of the T300A variant in humans is associated with similar urothelial architectural abnormalities, indicating an evolutionarily conserved impact. Mechanistically, we show that the reduced bacterial persistence is independent of basal autophagic flux or proinflammatory cytokine responses and does not involve Atg14 or Epg5. However, the T300A variant is associated with increased expression of the small GTPase Rab33b; RAB33B interacts with ATG16L1, as well as other secretory RABs, RAB27B and RAB11A, important for UPEC exocytosis from the urothelium. Finally, inhibition of secretory RABs in bladder epithelial cells increases intracellular UPEC load. Together, our results reveal that UPEC selectively utilize genes important for autophagosome formation to persist in the urothelium, and that the presence of the T300A variant in ATG16L1 is associated with changes in urothelial vesicle trafficking, which disrupts the ability of UPEC to persist, thereby limiting the risk of recurrent UTIs. Abbreviations: 3-PEHPC: 3-pyridinyl ethylidene hydroxyl phosphonocarboxylate; ATG: autophagy; ATG16L1: autophagy related 16 like 1; BECs: bladder epithelial cells; dpi: days post infection; hpi: hours post infection; IF: immunofluorescence; IL1B: interleukin 1 beta; IL6: interleukin 6; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MVB: multivesicular bodies; T300A: Thr300Ala; TNF: tumor necrosis factor; QIR(s): quiescent intracellular reservoir(s); siRNA: short interfering RNA; UPEC: uropathogenic Escherichia coli; UTI(s): urinary tract infection(s); TEM: transmission electron microscopy; WT: wild type.

New insights into diet breadth of polyphagous and oligophagous aphids on two Arabidopsis ecotypes.

We investigated whether plant ecotype might affect aphid performance and behavior. The probing behaviors of the polyphagous aphid Myzus persicae and the oligophagous aphid Brevicoryne brassicae on two ecotypes of Arabidopsis thaliana, WS and Col-0 were recorded using the direct current electrical penetration graph method (DC-EPG). Myzus persicae displayed a significant preference for the WS ecotype but was not greatly disturbed on Col-0, while B. brassicae discriminated between the two A. thaliana ecotypes, feeding less on WS than on Col-0. A Principal Component Analysis of aphid probing behavior data recorded on Col-0 and WS ecotypes showed that the one of M. persicae was positively correlated with the phloem ingestion phases while the one of B. brassicae was more related to nonfeeding phase. The survival of the aphid species was followed during early larval stages on the two ecotypes and a significantly higher mortality was observed of B. brassicae neonates compared to M. persicae, both reared on WS. Moreover, transcriptomic analysis of noninfested plant leaves from both ecotypes was monitored and underlined constitutive differences between Col-0 and WS gene expression that might explain the different aphid behaviors. Among a unigene set comprising 39 042 sequences for A. thaliana, 6% were differently expressed affecting, for example, the secondary metabolites and cell wall pathways: two third upregulated in WS and one third upregulated in Col-0. Thus, the "ecotype" variable should be taken into account when setting up a plant-insect experimental research.

Low-level expression of EPG5 leads to an attenuated Vici syndrome phenotype.

Vici syndrome is a multisystem disorder characterized by agenesis of the corpus callosum, oculocutaneous hypopigmentation, cataracts, cardiomyopathy, combined immunodeficiency, failure to thrive, profound developmental delay, and acquired microcephaly. Most individuals are severely affected and have a markedly reduced life span. Here we describe an 8-year-old boy with a history of developmental delay, agenesis of the corpus callosum, failure to thrive, myopathy, and well-controlled epilepsy. He was initially diagnosed with a mitochondrial disorder, based in part upon nonspecific muscle biopsy findings, but mitochondrial DNA mutation analysis revealed no mutations. Whole exome sequencing revealed compound heterozygosity for two EPG5 variants, inherited in trans. One was a known pathogenic mutation in exon 13 (c.2461C > T, p.Arg821X). The second was reported as a variant of unknown significance found within intron 16, six nucleotides before the exon 17 splice acceptor site (c.3099-6C > G). Reverse transcription-polymerase chain reaction of the EPG5 mRNA showed skipping of exon 17-which maintains an open reading frame-in 77% of the transcript, along with 23% expression of wild-type mRNA suggesting that intronic mutations may affect splicing of the EPG5 gene and result in symptoms. However, the expression of 23% wild-type mRNA may result in a significantly attenuated Vici syndrome phenotype.

Arabidopsis-Green Peach Aphid Interaction: Rearing the Insect, No-choice and Fecundity Assays, and Electrical Penetration Graph Technique to Study Insect Feeding Behavior.

Aphids constitute a large group of Hemipterans that use their slender stylets to tap into the sieve elements of plants from which they consume copious amounts of phloem sap, thus depriving the plant of photoassimilates. Some aphids also transmit viral diseases of plants. Myzus persicae Sülzer, commonly known as the green peach aphid (GPA), which is a polyphagous insect with a host range that covers 50 plant families, is considered amongst the top 3 insect pest of plants. The interaction between Arabidopsis thaliana and the GPA is utilized as a model pathosystem to study plant-aphid interaction. Here we describe the protocol used in our laboratories for rearing the GPA, and no-choice and fecundity bioassays to study GPA performance on Arabidopsis. In addition, we describe the procedure for the electrical penetration graph (EPG) technique to monitor feeding behavior of the GPA on Arabidopsis.

Muscle pathology in Vici syndrome-A case study with a novel mutation in EPG5 and a summary of the literature.

Vici syndrome is a disorder characterized by myopathy, cardiomyopathy, agenesis of the corpus callosum, immunodeficiency, cataracts, hypopigmentation, microcephaly, gross developmental delay and failure to thrive. It is caused by mutations in EPG5, which encodes a protein involved in the autophagy pathway. Although myopathy is part of the syndrome, few publications have described the muscle pathology. We present a detailed morphological analysis in a boy with Vici syndrome due to a novel homozygous one-base deletion in EPG5 (c.784delA), and we review the histopathological findings from previous reports. Muscle biopsy was performed at three months of age and demonstrated small vacuolated fibers, frequently with internal nuclei, and expressing developmental and fast myosin isoforms. There was an increase in acid phosphatase activity in the small fibers, which also showed LAMP-2 upregulation, glycogen accumulation and contained numerous p62-positive inclusions and some lipid droplets. Electron microscopy demonstrated hypoplastic fibers with massive glycogen accumulation and extensive disorganization of the myofibrils. This study expands the muscle pathological features of Vici syndrome and demonstrates a pattern of vacuolar myopathy with glycogen storage and immature, hypoplastic and atrophic muscle fibers. Increased lysosomes and accumulation of p62 are in line with a disturbance of the autophagic pathway as an essential part of the pathogenesis.

A Semipersistent Plant Virus Differentially Manipulates Feeding Behaviors of Different Sexes and Biotypes of Its Whitefly Vector.

It is known that plant viruses can change the performance of their vectors. However, there have been no reports on whether or how a semipersistent plant virus manipulates the feeding behaviors of its whitefly vectors. Cucurbit chlorotic yellows virus (CCYV) (genus Crinivirus, family Closteroviridae) is an emergent plant virus in many Asian countries and is transmitted specifically by B and Q biotypes of tobacco whitefly, Bemisia tabaci (Gennadius), in a semipersistent manner. In the present study, we used electrical penetration graph (EPG) technique to investigate the effect of CCYV on the feeding behaviors of B. tabaci. The results showed that CCYV altered feeding behaviors of both biotypes and sexes of B. tabaci with different degrees. CCYV had stronger effects on feeding behaviors of Q biotype than those of B biotype, by increasing duration of phloem salivation and sap ingestion, and could differentially manipulate feeding behaviors of males and females in both biotype whiteflies, with more phloem ingestion in Q biotype males and more non-phloem probing in B biotype males than their respective females. With regard to feeding behaviors related to virus transmission, these results indicated that, when carrying CCYV, B. tabaci Q biotype plays more roles than B biotype, and males make greater contribution than females.

Body position of the stink bug Dichelops melacanthus (Dallas) during feeding from stems of maize seedlings

Abstract The Neotropical green-belly stink bug, Dichelops melacanthus (Dallas) is a major pest of maize, Zea mays L. in the main production areas of Brazil. It usually feeds on the stems of young plants (seedlings) causing heavy damage by affecting the plant growth and mitigating seed yield. Laboratory studies were conducted to determine body position (upward or downward) of the bug on plant (seedling) stem during feeding and not feeding activities. Ten visual records were taken per day, each spaced one hour during 30 days of 10 adult bugs of similar age exposed to maize seedlings inside cages (plastic tubes). At each observation, it was recorded if the bug was feeding on the stem (i.e., stylets inserted into the plant tissue) or not, and its body position. During feeding, waveforms were recorded using the EPG (electropenetrography) technique, which were correlated with histological studies to reveal the feeding sites. Results indicated that when they were feeding, the majority of the bugs were in the downward position. In contrast, when the bugs were on the plants, and not feeding, they were mostly in the upward position. Waveforms generated using the EPG coupled with histological studies demonstrated that during ingestion bugs fed from the xylem vessels and from the parenchyma tissue using cell rupture strategy in the latter. No clear explanation was found to explain the preferred downward body position during ingestion, but some hypothesis are speculated.

Resumo O percevejo barriga-verde, Dichelops melacanthus (Dallas) é uma praga importante do milho, Zea mays L. nas principais áreas produtoras do Brasil. Usualmente alimenta-se nas hastes de plantas jovens (plântulas) causando danos severos no seu crescimento e reduzindo o rendimento de grãos. Estudos foram conduzidos em laboratório para determinar a posição do corpo (voltado para cima ou para baixo) dos percevejos nas hastes das plântulas de milho durante as atividades de alimentação e não-alimentação. Foram feitas dez observações por dia, espaçadas por uma hora, durante 30 dias em 10 percevejos adultos com idade semelhante sobre plântulas de milho colocadas em gaiolas (tubos de plástico). Em cada observação, anotou-se se o percevejo estava se alimentando (i.e., estiletes bucais inseridos no tecido vegetal) ou não, e a posição do corpo. Durante a alimentação, ondas eletromagnéticas foram registradas utilizando-se o EPG (técnica da eletropenetrografia), as quais foram correlacionadas com estudos histológicos para revelar os locais de alimentação. Os resultados indicaram que quando os percevejos estavam se alimentando, a maioria estava voltado para baixo. Em contraste, os percevejos sem se alimentar nas plântulas estavam a maioria voltados para cima. As ondas geradas pelo EPG junto com os estudos histológicos revelaram que os percevejos se alimentaram dos vasos do xilema e do tecido parenquimatoso usando a estratégia de ruptura celular no último. Não foi encontrada uma explicação clara para a preferência dos percevejos em se alimentarem na posição voltados para baixo, mas algumas hipóteses são especuladas.