DNA Reference Reagents for Genotyping RH Variants.

Patients who carry Rhesus (RH) blood group variants may develop Rh alloantibodies requiring matched red blood cell transfusions. Serologic reagents for Rh variants often fail to specifically identify variant Rh antigens and are in limited supply. Therefore, red blood cell genotyping assays are essential for managing transfusions in patients with clinically relevant Rh variants. Well-characterized DNA reference reagents are needed to ensure quality and accuracy of the molecular tests. Eight lyophilized DNA reference reagents, representing 21 polymorphisms in RHD and RHCE, were produced from an existing repository of immortalized B-lymphoblastoid cell lines at the Center for Biologics Evaluation and Research/US Food and Drug Administration. The material was validated through an international collaborative study involving 17 laboratories that evaluated each DNA candidate using molecular assays to characterize RHD and RHCE alleles, including commercial platforms and laboratory-developed testing, such as Sanger sequencing, next-generation sequencing, and third-generation sequencing. The genotyping results showed 99.4% agreement with the expected results for the target RH polymorphisms and 87.9% for RH allele agreement. Most of the discordant RH alleles results were explained by a limited polymorphism coverage in some genotyping methods. Results of stability and accelerated degradation studies support the suitability of these reagents for use as reference standards. The collaborative study results demonstrate the qualification of these eight DNA reagents for use as reference standards for RH blood group genotyping assay development and analytical validation.

Viral Infections, Are They a Trigger and Risk Factor of Alzheimer's Disease?

Alzheimer's Disease (AD), a progressive and debilitating condition, is reported to be the most common type of dementia, with at least 55 million people believed to be currently affected. Many causation hypotheses of AD exist, yet the intriguing link between viral infection and its possible contribution to the known etiology of AD has become an attractive focal point of research for the field and a challenging study task. In this review, we will explore the historical perspective and milestones that led the field to investigate the viral connection to AD. Specifically, several viruses such as Herpes Simplex Virus 1 (HSV-1), Zika virus (ZIKV), and severe cute respiratory syndrome coronavirus 2 (SARS-CoV-2), along with several others mentioned, include the various viruses presently considered within the field. We delve into the strong evidence implicating these viruses in the development of AD such as the lytic replication and axonal transport of HSV-1, the various mechanisms of ZIKV neurotropism through the human protein Musashi-1 (MSI1), and the spread of SARS-CoV-2 through the transfer of the virus through the BBB endothelial cells to glial cells and then to neurons via transsynaptic transfer. We will also explore beyond these mere associations by carefully analyzing the potential mechanisms by which these viruses may contribute to AD pathology. This includes but is not limited to direct neuronal infections, the dysregulation of immune responses, and the impact on protein processing (Aß42 and hyperphosphorylated tau). Controversies and challenges of the virus-AD relationship emerge as we tease out these potential mechanisms. Looking forward, we emphasize future directions, such as distinct questions and proposed experimentations to explore, that the field should take to tackle the remaining unanswered questions and the glaring research gaps that persist. Overall, this review aims to provide a comprehensive survey of the past, present, and future of the potential link between viral infections and their association with AD development while encouraging further discussion.

Dengue Virus-2 Infection Affects Fecundity and Elicits Specific Transcriptional Changes in the Ovaries of Aedes aegypti Mosquitoes.

Dengue fever (DF), caused by the dengue virus (DENV), is the most burdensome arboviral disease in the world, with an estimated 400 million infections each year. The Aedes aegypti mosquito is the main vector of DENV and transmits several other human pathogens, including Zika, yellow fever, and chikungunya viruses. Previous studies have shown that the pathogen infection of mosquitoes can alter reproductive fitness, revealing specific vector-pathogen interactions that are key determinants of vector competence. However, only a handful of studies have examined the effect of DENV infection in A. aegypti, showing a reduction in lifespan and fecundity over multiple blood meals. To provide a more comprehensive analysis of the impact of DENV infection on egg laying and fecundity, we assessed egg laying timing in DENV-2 blood-fed mosquitoes (infected group) compared to mock blood-fed mosquitoes (control group). We confirmed a significant decrease in fecundity during the first gonadotrophic cycle. To further investigate this phenotype and the underlying DENV-2 infection-dependent changes in gene expression, we conducted a transcriptomic analysis for differentially expressed genes in the ovaries of A. aegypti infected with DENV-2 vs. mock-infected mosquitoes. This analysis reveals several DENV-2-regulated genes; among them, we identified a group of 12 metabolic genes that we validated using reverse transcription-quantitative PCR (RT-qPCR). Interestingly, two genes found to be upregulated in DENV-infected mosquito ovaries exhibited an antiviral role for DENV-2 in an Aedes cell line. Altogether, this study offers useful insights into the virus-vector interface, highlighting the importance of gene expression changes in the mosquito's ovary during DENV-2 infection in the first gonadotrophic  cycle,  triggering  antiviral  responses  that  may  possibly  interfere  with mosquito reproduction. This information is extremely relevant for further investigation of A. aegypti's ability to tolerate viruses since virally infected mosquitoes in nature constitute a powerful source of supporting viruses during intra-epidemic periods, causing a huge burden on the public health system.

Blood Meals With Active and Heat-Inactivated Serum Modifies the Gene Expression and Microbiome of Aedes albopictus.

The Asian "tiger mosquito" Aedes albopictus is currently the most widely distributed disease-transmitting mosquito in the world. Its geographical expansion has also allowed the expansion of multiple arboviruses like dengue, Zika, and chikungunya, to higher latitudes. Due to the enormous risk to global public health caused by mosquitoes species vectors of human disease, and the challenges in slowing their expansion, it is necessary to develop new and environmentally friendly vector control strategies. Among these, host-associated microbiome-based strategies have emerged as promising options. In this study, we performed an RNA-seq analysis on dissected abdomens of Ae. albopictus females from Manhattan, KS, United States fed with sugar and human blood containing either normal or heat-inactivated serum, to evaluate the effect of heat inactivation on gene expression, the bacteriome transcripts and the RNA virome of this mosquito species. Our results showed at least 600 genes with modified expression profile when mosquitoes were fed with normal vs. heat-inactivated-containing blood. These genes were mainly involved in immunity, oxidative stress, lipid metabolism, and oogenesis. Also, we observed bacteriome changes with an increase in transcripts of Actinobacteria, Rhodospirillaceae, and Anaplasmataceae at 6 h post-feeding. We also found that feeding with normal blood seems to particularly influence Wolbachia metabolism, demonstrated by a significant increase in transcripts of this bacteria in mosquitoes fed with blood containing normal serum. However, no differences were observed in the virome core of this mosquito population. These results suggest that heat and further inactivation of complement proteins in human serum may have profound effect on mosquito and microbiome metabolism, which could influence interpretation of the pathogen-host interaction findings when using this type of reagents specially when measuring the effect of Wolbachia in vector competence.

Vertebrate Responses against Arthropod Salivary Proteins and Their Therapeutic Potential.

The saliva of hematophagous arthropods contains a group of active proteins to counteract host responses against injury and to facilitate the success of a bloodmeal. These salivary proteins have significant impacts on modulating pathogen transmission, immunogenicity expression, the establishment of infection, and even disease severity. Recent studies have shown that several salivary proteins are immunogenic and antibodies against them may block infection, thereby suggesting potential vaccine candidates. Here, we discuss the most relevant salivary proteins currently studied for their therapeutic potential as vaccine candidates or to control the transmission of human vector-borne pathogens and immune responses against different arthropod salivary proteins.

What came first, the virus or the egg: Innate immunity during viral coinfections.

Infections with any pathogen can be severe and present with numerous complications caused by the pathogen or the host immune response to the invading microbe. However, coinfections, also called polymicrobial infections or secondary infections, can further exacerbate disease. Coinfections are more common than is often appreciated. In this review, we focus specifically on coinfections between viruses and other viruses, bacteria, parasites, or fungi. Importantly, innate immune signaling and innate immune cells that facilitate clearance of the initial viral infection can affect host susceptibility to coinfections. Understanding these immune imbalances may facilitate better diagnosis, prevention, and treatment of such coinfections.

Common Differences: The Ability of Inflammasomes to Distinguish Between Self and Pathogen Nucleic Acids During Infection.

The innate immune system detects the presence of pathogens based on detection of non-self. In other words, most pathogens possess intrinsic differences that can distinguish them from host cells. For example, bacteria and fungi have cell walls comprised of peptidoglycan and carbohydrates (like mannans), respectively. Germline encoded pattern recognition receptors (PRRs) of the Toll-like receptor (TLR) and C-type lectin receptor (CLR) family have the ability to detect such unique pathogen associated features. However, some TLRs and members of the RIG-I-like receptor (RLR), NOD-like receptor (NLR), or AIM2-like receptor (ALR) family can sense pathogen invasion based on pathogen nucleic acids. Nucleic acids are not unique to pathogens, thus raising the question of how such PRRs evolved to detect pathogens but not self. In this chapter, we will examine the PRRs that sense pathogen nucleic acids and subsequently activate the inflammasome signaling pathway. We will examine the selective mechanisms by which these receptors distinguish pathogens from "self" and discuss the importance of such pathways in disease development in animal models and human patients.