An atlas of human vector-borne microbe interactions reveals pathogenicity mechanisms.

Vector-borne diseases are a leading cause of death worldwide and pose a substantial unmet medical need. Pathogens binding to host extracellular proteins (the "exoproteome") represents a crucial interface in the etiology of vector-borne disease. Here, we used bacterial selection to elucidate host-microbe interactions in high throughput (BASEHIT)-a technique enabling interrogation of microbial interactions with 3,324 human exoproteins-to profile the interactomes of 82 human-pathogen samples, including 30 strains of arthropod-borne pathogens and 8 strains of related non-vector-borne pathogens. The resulting atlas revealed 1,303 putative interactions, including hundreds of pairings with potential roles in pathogenesis, including cell invasion, tissue colonization, immune evasion, and host sensing. Subsequent functional investigations uncovered that Lyme disease spirochetes recognize epidermal growth factor as an environmental cue of transcriptional regulation and that conserved interactions between intracellular pathogens and thioredoxins facilitate cell invasion. In summary, this interactome atlas provides molecular-level insights into microbial pathogenesis and reveals potential host-directed targets for next-generation therapeutics.

Adiponectin in the mammalian host influences ticks' acquisition of the Lyme disease pathogen Borrelia.

Arthropod-borne pathogens cause some of the most important human and animal infectious diseases. Many vectors acquire or transmit pathogens through the process of blood feeding. Here, we report adiponectin, the most abundant adipocyte-derived hormone circulating in human blood, directly or indirectly inhibits acquisition of the Lyme disease agent, Borrelia burgdorferi, by Ixodes scapularis ticks. Rather than altering tick feeding or spirochete viability, adiponectin or its associated factors induces host histamine release when the tick feeds, which leads to vascular leakage, infiltration of neutrophils and macrophages, and inflammation at the bite site. Consistent with this, adiponectin-deficient mice have diminished pro-inflammatory responses, including interleukin (IL)-12 and IL-1ß, following a tick bite, compared with wild-type animals. All these factors mediated by adiponectin or associated factors influence B. burgdorferi survival at the tick bite site. These results suggest a host adipocyte-derived hormone modulates pathogen acquisition by a blood-feeding arthropod.

mosGILT controls innate immunity and germ cell development in Anopheles gambiae.

Gene-edited mosquitoes lacking a gamma-interferon-inducible lysosomal thiol reductase-like protein, namely (mosGILTnull) have lower Plasmodium infection, which is linked to impaired ovarian development and immune activation. The transcriptome of mosGILTnull Anopheles gambiae was therefore compared to wild type (WT) mosquitoes by RNA-sequencing to delineate mosGILT-dependent pathways. Compared to WT mosquitoes, mosGILTnull A. gambiae demonstrated altered expression of genes related to oogenesis, 20-hydroxyecdysone synthesis, as well as immune-related genes. Serendipitously, the zero population growth gene, zpg, an essential regulator of germ cell development was found to be one of the most downregulated genes in mosGILTnull mosquitoes. These results provide a crucial missing link between two previous studies on the role of zpg and mosGILT in ovarian development. This study further demonstrates that mosGILT has the potential to serve as a target for the biological control of mosquito vectors and to influence the Plasmodium life cycle within the vector.

Bulk and single-nucleus RNA sequencing highlight immune pathways induced in individuals during an Ixodes scapularis tick bite.

Ticks are hematophagous arthropods that use a complex mixture of salivary proteins to evade host defenses while taking a blood meal. Little is known about the immunological and physiological consequences of tick feeding on humans. Here, we performed the first bulk and single-nucleus RNA sequencing (snRNA-seq) of skin and blood of four persons presenting with naturally acquired, attached Ixodes scapularis ticks. Pathways and individual genes associated with innate and adaptive immunity were identified based on bulk RNA sequencing, including interleukin-17 signaling and platelet activation pathways at the site of tick attachment or in peripheral blood. snRNA-seq further revealed that the Hippo signaling, cell adhesion, and axon guidance pathways were involved in the response to an I. scapularis bite in humans. Features of the host response in these individuals also overlapped with that of laboratory guinea pigs exposed to I. scapularis and which acquired resistance to ticks. These findings offer novel insights for the development of new biomarkers for I. scapularis exposure and anti-tick vaccines for human use.

An Ixodes scapularis Protein Disulfide Isomerase Contributes to Borrelia burgdorferi Colonization of the Vector.

Borrelia burgdorferi causes Lyme disease, the most common tick-transmitted illness in North America. When Ixodes scapularis feed on an infected vertebrate host, spirochetes enter the tick gut along with the bloodmeal and colonize the vector. Here, we show that a secreted tick protein, I. scapularisprotein disulfide isomerase A3 (IsPDIA3), enhances B. burgdorferi colonization of the tick gut. I. scapularis ticks in which ispdiA3 has been knocked down using RNA interference have decreased spirochete colonization of the tick gut after engorging on B. burgdorferi-infected mice. Moreover, administration of IsPDIA3 antiserum to B. burgdorferi-infected mice reduced the ability of spirochetes to colonize the tick when feeding on these animals. We show that IsPDIA3 modulates inflammatory responses at the tick bite site, potentially facilitating spirochete survival at the vector-host interface as it exits the vertebrate host to enter the tick gut. These data provide functional insights into the complex interactions between B. burgdorferi and its arthropod vector and suggest additional targets to interfere with the spirochete life cycle.

Non-invasive diagnostics of Liberibacter disease on tomatoes using a hand-held Raman spectrometer.

MAIN CONCLUSION: Hand-held Raman spectroscopy can be used for confirmatory, non-invasive and non-destructive detection and identification of two haplotypes of Liberibacter disease on tomatoes. Using this spectroscopic approach, structural changes in carotenoids, xylan, cellulose and pectin that are associ-ated with this bacterial disease can be determined. 'Candidatus Liberibacter solanacearum' (Lso) is a phloem-limited Gram-negative bacterium that infects crops worldwide. In North America, two haplotypes of Lso (LsoA and LsoB) are transmitted by the potato psyllid, Bactericera cockerelli (Sulc), and infect many solanaceous crops such as potato and tomato. Infected plants exhibit chlorosis, severe stunting, leaf cupping, and scorching. Polymerase chain reaction (PCR) and potato tuber frying are commonly used methods for diagnostics of the plant disease caused by Lso. However, they are time-consuming, costly, destructive to the sample, and often not sensitive enough to detect the pathogen in the early infection stage. Raman spectroscopy (RS) is a noninvasive, nondestructive, analytical technique, which probes chemical composition of analyzed samples. In this study, we demonstrate that Lso infection can be diagnosed by non-invasive spectroscopic analysis of tomato leaves three weeks following infection, before the development of aerial symptoms. In combination with chemometric analyses, Raman spectroscopy allows for 80% accurate diagnostics of Liberibacter disease caused by each of the two different haplotypes. This diagnostics approach is portable and sample agnostic, suggesting that it could be utilized for other crops and could be conducted autonomously.

Lso-HPE1, an Effector of 'Candidatus Liberibacter solanacearum', Can Repress Plant Immune Response.

'Candidatus Liberibacter solanacearum' is a plant pathogen affecting the families Solanaceae and Apiaceae in different parts of the world. 'Ca. L. solanacearum' is a Gram-negative, fastidious α-proteobacterium that is vectored by different psyllid species. Plant-pathogenic bacteria are known for interfering with the host physiology or defense mechanisms, often by secreting bacterial effectors. Effector proteins are critical for virulence; therefore, the identification of effectors could help with disease management. In this study, we characterized the Sec-translocon-dependent 'Ca. L. solanacearum'-hypothetical protein effector 1 (Lso-HPE1). We compared this protein sequence in the different 'Ca. L. solanacearum' haplotypes. We predicted the signal peptide and validated its function using Escherichia coli's alkaline phosphatase fusion assay. Agrobacterium tumefaciens-mediated transient expression in Nicotiana benthamiana demonstrated that Lso-HPE1 from 'Ca. L. solanacearum' haplotypes A and B were able to inhibit the induction of cell death in plants. We also compared gene expression of the Lso-HPE1- transcripts in 'Ca. L. solanacearum' haplotypes A and B in tomato and in the vector Bactericera cockerelli. This work validates the identification of a Sec-translocon-dependent 'Ca. L. solanacearum' protein possibly involved in suppression of plant cell death.

No Evidence of Apoptotic Response of the Potato Psyllid Bactericera cockerelli to "Candidatus Liberibacter solanacearum" at the Gut Interface.

"Candidatus Liberibacter solanacearum" is a pathogen transmitted by the potato psyllid Bactericera cockerelli (Sulc) (Hemiptera: Triozidae) in a persistent manner. In this study, we investigated the molecular interaction between "Ca. Liberibacter solanacearum" and the potato psyllid at the gut interface. Specifically, we focused on the apoptotic response of potato psyllids to the infection by two "Ca. Liberibacter solanacearum" haplotypes, LsoA and LsoB. To this end, we first quantified and localized "Ca. Liberibacter solanacearum" in the gut of adult psyllids. We then evaluated the existence of an apoptotic response in the insect gut using microscopy analyses to visualize the nuclei and the actin cytoskeleton of the gut cells and DNA fragmentation analyses by agarose gel electrophoresis. We also performed annexin V cell death assays to detect apoptosis. Finally, we annotated apoptosis-related genes from the potato psyllid transcriptome and evaluated their expression in response to "Ca. Liberibacter solanacearum" infection. The results showed no cellular markers of apoptosis despite the large amount of "Ca. Liberibacter solanacearum" present in the psyllid gut. In addition, only three genes potentially involved in apoptosis were regulated in the psyllid gut in response to "Ca. Liberibacter solanacearum": the apoptosis-inducing factor AIF3 was downregulated in LsoA-infected psyllids, while the inhibitor of apoptosis IAPP5 was downregulated and IAP6 was upregulated in LsoB-infected psyllids. Overall, no evidence of apoptosis was observed in the gut of potato psyllid adults in response to either "Ca. Liberibacter solanacearum" haplotype. This study represents a first step toward understanding the interactions between "Ca. Liberibacter solanacearum" and the potato psyllid, which is crucial to developing approaches to disrupt their transmission.

NDUFA4L2 expression predicts poor prognosis in clear cell renal cell carcinoma patients.

BACKGROUND: NDUFA4L2 is overexpressed in VHL-deficient cell lines and neuroblastoma. The clinical significance of NDUFA4L2 in clear cell renal cell carcinoma (ccRCC) has not been well studied. Therefore, we evaluated the prognostic value of NDUFA4L2 in ccRCC patients. METHODS: In our study, NDUFA4L2 expression in 86 cases of ccRCC and adjacent normal tissues was monitored by immunohistochemistry, semi-quantitative RT-PCR, and Western blot analyses. The relationship between NDUFA4L2 expression and the clinical features of ccRCC was assessed. RESULTS: The results showed that NDUFA4L2 protein expression was found to be higher in ccRCC tissues 81.4% (70/86) than in normal tissues 26.7% (23/86) (p = 0.021). The average level of NDUFA4L2 mRNA expression was found to be 122.23 ± 6.018 and 21.34 ± 1.036 in ccRCC tissue and adjacent normal tissue (p < 0.001). NDUFA4L2 expression levels were correlated with some clinical features of ccRCC. Multivariate analysis showed NDUFA4L2 expression was an independent prognostic factor for ccRCC patients. CONCLUSIONS: Our study has provided the significant clinical relevance of NDUFA4L2 in ccRCC and suggested that ccRCC patients with NDUFA4L2 overexpression may be suitable as a potential therapeutic target for ccRCC patients.

Characterization and expression of genes encoding three small heat shock proteins in Sesamia inferens (Lepidoptera: Noctuidae).

The pink stem borer, Sesamia inferens (Walker), is a major pest of rice and is endemic in China and other parts of Asia. Small heat shock proteins (sHSPs) encompass a diverse, widespread class of stress proteins that have not been characterized in S. inferens. In the present study, we isolated and characterized three S. inferens genes that encode members of the α-crystallin/sHSP family, namely, Sihsp21.4, Sihsp20.6, and Sihsp19.6. The three cDNAs encoded proteins of 187, 183 and 174 amino acids with calculated molecular weights of 21.4, 20.6 and 19.6 kDa, respectively. The deduced amino acid sequences of the three genes showed strong similarity to sHSPs identified in other lepidopteran insects. Sihsp21.4 contained an intron, but Sihsp20.6 and Sihsp19.6 lacked introns. Real-time quantitative PCR analyses revealed that Sihsp21.4 was most strongly expressed in S. inferens heads; Whereas expression of Sihsp20.6 and Sihsp19.6 was highest in eggs. The three S. inferens sHSP genes were up-regulated during low temperature stress. In summary, our results show that S. inferens sHSP genes have distinct regulatory roles in the physiology of S. inferens.

Transplantation of en bloc kidneys from cardiac deceased small pediatric donors: 2 case reports and literature review.

OBJECTIVE: To gain an insight into the transplantation with donor kidneys from extended criterion donation after cardiac death (DCD) and to improve the management during and after renal transplantation METHODS: Renal transplantation in 2 patients who used organs from small pediatric donors (<3 years) was performed. The graft kidneys were procured from 1 donor aged 11 months and the other 1 year and 7 months. The 2 donors were diagnosed as brain death caused by serious infantile hepatitis syndrome and severe craniocerebral injury, respectively. After the cardiac death, en bloc organ resection was performed. En bloc kidneys were transplanted to 2 adult recipients who were 37 and 41 years old, respectively. RESULTS: The recipients were followed-up for 6 months. Both of them developed large volume of bloody drainage in the early post-operational period and relieved after relevant treatment. The kidney grafts functioned well and no other surgical complications or acute rejections happened during the follow-up. CONCLUSION: Based on modified peri-operative techniques, it is safe to perform renal transplantation with kidneys procured from cardiac death donors who are younger than 3 years old, an important source to increase the number of organs available for transplantation, yet the vascular complications require attention.

Signaling between mammalian adiponectin and a mosquito adiponectin receptor reduces Plasmodium transmission.

IMPORTANCE: When a female mosquito takes a blood meal from a mammalian host, components of the blood meal can affect mosquito fitness and indirectly influence pathogen infectivity. We identified a pathway involving an Anopheles gambiae adiponectin receptor, which, triggered by adiponectin from an incoming blood meal, decreases Plasmodium infection in the mosquito. Activation of this pathway negatively regulates lipophorin expression, an important lipid transporter that both enhances egg development and Plasmodium infection. This is an unrecognized cross-phyla interaction between a mosquito and its vertebrate host. These processes are critical to understanding the complex life cycle of mosquitoes and Plasmodium following a blood meal and may be applicable to other hematophagous arthropods and vector-borne infectious agents.

Metabolomic changes associated with acquired resistance to Ixodes scapularis.

Guinea pigs repeatedly exposed to Ixodes scapularis develop acquired resistance to the ticks (ATR). The molecular mechanisms of ATR have not been fully elucidated, and partially involves immune responses to proteins in tick saliva. In this study, we examined the metabolome of sera of guinea pigs during the development of ATR. Induction of components of the tyrosine metabolic pathway, including hydroxyphenyllactic acid (HPLA), were associated with ATR. We therefore administered HPLA to mice, an animal that does not develop ATR, and exposed the animals to I. scapularis. We also administered nitisinone, a known inhibitor of tyrosine degradation, to another group of mice. The mortality of I. scapularis that fed on mice given HPLA or nitisinone was 26 % and 72 % respectively, compared with 2 % mortality among ticks that fed on control animals. These data indicate that tick bites alter the guinea pig metabolome, and that the tyrosine metabolism pathway can potentially be targeted for I. scapularis control.

[Renal transplantation with kidneys procured from cardiac deceased post-liver transplantation donor: 2 cases report and literature review].

OBJECTIVE: To better understand the pre-operation evaluation of donor kidneys from extended criteria donation after cardiac death and to improve the management during and after renal transplantation. METHODS: Both of the donor kidneys were from the donor who underwent liver transplantation 5 years ago in the Center of Organ Transplantation of Central South University. The donor was admitted because of liver function deterioration which led to hepatic coma, brain death, hepatorenal syndrome and cardiac death sequentially. Deceased donor score (DDS) and "zero point" kidney biopsy were applied to evaluate the donor kidney. After thorough examination of the donor and the renal function, renal transplantation was performed on 2 recipients. RESULTS: The recipients were followed up by 6 months, both of whom developed pulmonary infection and relieved after treatments. The kidney grafts functioned well and no surgical complication and no acute rejection occurred during the follow-up. CONCLUSION: Proper evaluation of the donor organs ensures the safety of renal transplantation with kidneys from cardiac death donors who underwent liver transplantation, which is an important way to increase the number of organs for transplantation, yet the long-term effects need further observation.

Potato psyllids mount distinct gut responses against two different 'Candidatus Liberibacter solanacearum' haplotypes.

'Candidatus Liberibacter solanacearum' (Lso) is a bacterial pathogen infecting several crops and causing damaging diseases. Several Lso haplotypes have been identified. Among the seven haplotypes present in North America, LsoA and LsoB are transmitted by the potato psyllid, Bactericera cockerelli (Sulc), in a circulative and persistent manner. The gut, which is the first organ pathogen encounters, could be a barrier for Lso transmission. However, the molecular interactions between Lso and the psyllid vector at the gut interface remain largely unknown. In this study, we investigated the global transcriptional responses of the adult psyllid gut upon infection with two Lso haplotypes (LsoA and LsoB) using Illumina sequencing. The results showed that each haplotype triggers a unique transcriptional response, with most of the distinct genes elicited by the highly virulent LsoB. The differentially expressed genes were mainly associated with digestion and metabolism, stress response, immunity, detoxification as well as cell proliferation and epithelium renewal. Importantly, distinct immune pathways were triggered by LsoA and LsoB in the gut of the potato psyllid. The information in this study will provide an understanding of the molecular basis of the interactions between the potato psyllid gut and Lso, which may lead to the discovery of novel molecular targets for the control of these pathogens.

Metabolomic changes associated with acquired resistance to Ixodes scapularis.

Guinea pigs repeatedly exposed to Ixodes scapularis develop acquired resistance to the ticks (ATR). The molecular mechanisms of ATR have not been fully elucidated, and partially involve immune responses to proteins in tick saliva. In this study, we examined the metabolome of sera of guinea pigs during the development of ATR. Induction of components of the tyrosine metabolic pathway, including hydroxyphenyllactic acid (HPLA), were associated with ATR. We therefore administered HPLA to mice, an animal that does not develop ATR, and exposed the animals to I. scapularis . We also administered nitisinone, a known inhibitor of tyrosine degradation, to another group of mice. The mortality of I. scapularis that fed on mice given HPLA or nitisinone was 26% and 72% respectively, compared with 2% mortality among ticks that fed on control animals. These data indicate that metabolic changes that occur after tick bites contribute to ATR.

Anopheles gambiae mosGILT regulates innate immune genes and zpg expression.

Gene-edited mosquitoes lacking a g amma-interferon-inducible lysosomal thiol reductase-like protein, namely ( mosGILT null ) have lower Plasmodium infection, which is linked to impaired ovarian development and immune activation. The transcriptome of mosGILT null A. gambiae was therefore compared to wild type (WT) by RNA-sequencing to delineate mosGILT-dependent pathways. Compared to WT mosquitoes, mosGILT null A. gambiae demonstrated altered expression of genes related to oogenesis, 20-hydroxyecdysone synthesis, as well as immune-related genes. Serendipitously, the zero population growth gene, zpg , an essential regulator of germ cell development was found to be one of the most downregulated genes in mosGILT null mosquitoes. These results provide the crucial missing link between two previous studies on the role of zpg and mosGILT in ovarian development. This study further demonstrates that mosGILT has the potential to serve as a target for the biological control of mosquito vectors and to influence the Plasmodium life cycle within the vector.

Specific mRNA lipid nanoparticles and acquired resistance to ticks.

Acquired resistance to ticks can develop when animals are repeatedly exposed to ticks. Recently, acquired resistance to Ixodes scapularis was induced in guinea pigs immunized with an mRNA-lipid nanoparticle vaccine (19ISP) encoding 19 I. scapularis proteins. Here, we evaluated specific mRNAs present in 19ISP to identify critical components associated with resistance to ticks. A lipid nanoparticle containing 12 mRNAs which included all the targets within 19ISP that elicited strong humoral responses in guinea pigs, was sufficient to induce robust resistance to ticks. Lipid nanoparticles containing fewer mRNAs or a single mRNA were not able to generate strong resistance to ticks. All lipid nanoparticles containing salp14 mRNA, however, were associated with increased redness at the tick bite site - which is the first manifestation of acquired resistance to ticks. This study demonstrates that more than one I. scapularis target within 19ISP is required for resistance to ticks, and that additional targets may also play a role in this process.

A tick C1q protein alters infectivity of the Lyme disease agent by modulating interferon γ.

In North America, the Lyme disease agent, Borrelia burgdorferi, is commonly transmitted by the black-legged tick, Ixodes scapularis. Tick saliva facilitates blood feeding and enhances pathogen survival and transmission. Here, we demonstrate that I. scapularis complement C1q-like protein 3 (IsC1ql3), a tick salivary protein, directly interacts with B. burgdorferi and is important during the initial stage of spirochetal infection of mice. Mice fed upon by B. burgdorferi-infected IsC1ql3-silenced ticks, or IsC1ql3-immunized mice fed upon by B. burgdorferi-infected ticks, have a lower spirochete burden during the early phase of infection compared with control animals. Mechanically, IsC1ql3 interacts with the globular C1q receptor present on the surface of CD4+ and CD8+ T cells, resulting in decreased production of interferon γ. IsC1ql3 is a C1q-domain-containing protein identified in arthropod vectors and has an important role in B. burgdorferi infectivity as the spirochete transitions from the tick to vertebrate host.

CD55 Facilitates Immune Evasion by Borrelia crocidurae, an Agent of Relapsing Fever.

Relapsing fever, caused by diverse Borrelia spirochetes, is prevalent in many parts of the world and causes significant morbidity and mortality. To investigate the pathoetiology of relapsing fever, we performed a high-throughput screen of Borrelia-binding host factors using a library of human extracellular and secretory proteins and identified CD55 as a novel host binding partner of Borrelia crocidurae and Borrelia persica, two agents of relapsing fever in Africa and Eurasia. CD55 is present on the surface of erythrocytes, carries the Cromer blood group antigens, and protects cells from complement-mediated lysis. Using flow cytometry, we confirmed that both human and murine CD55 bound to B. crocidurae and B. persica. Given the expression of CD55 on erythrocytes, we investigated the role of CD55 in pathological B. crocidurae-induced erythrocyte aggregation (rosettes), which enables spirochete immune evasion. We showed that rosette formation was partially dependent on host cell CD55 expression. Pharmacologically, soluble recombinant CD55 inhibited erythrocyte rosette formation. Finally, CD55-deficient mice infected with B. crocidurae had a lower pathogen load and elevated proinflammatory cytokine and complement factor C5a levels. In summary, our results indicate that CD55 is a host factor that is manipulated by the causative agents of relapsing fever for immune evasion. IMPORTANCE Borrelia species are causative agents of Lyme disease and relapsing fever infections in humans. B. crocidurae causes one of the most prevalent relapsing fever infections in parts of West Africa. In the endemic regions, B. crocidurae is present in ~17% of the ticks and ~11% of the rodents that serve as reservoirs. In Senegal, ~7% of patients with acute febrile illness were found to be infected with B. crocidurae. There is little information on host-pathogen interactions and how B. crocidurae manipulates host immunity. In this study, we used a high-throughput screen to identify host proteins that interact with relapsing fever-causing Borrelia species. We identified CD55 as one of the host proteins that bind to B. crocidurae and B. persica, the two causes of relapsing fever in Africa and Eurasia. We show that the interaction of B. crocidurae with CD55, present on the surface of erythrocytes, is key to immune evasion and successful infection in vivo. Our study further shows the role of CD55 in complement regulation, regulation of inflammatory cytokine levels, and innate immunity during relapsing fever infection. Overall, this study sheds light on host-pathogen interactions during relapsing fever infection in vivo.