High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire.

Each B-cell receptor consists of a pair of heavy and light chains. High-throughput sequencing can identify large numbers of heavy- and light-chain variable regions (VH and VL) in a given B-cell repertoire, but information about endogenous pairing of heavy and light chains is lost after bulk lysis of B-cell populations. Here we describe a way to retain this pairing information. In our approach, single B cells (>5 × 104 capacity per experiment) are deposited in a high-density microwell plate (125 pl/well) and lysed in situ. mRNA is then captured on magnetic beads, reverse transcribed and amplified by emulsion VH:VL linkage PCR. The linked transcripts are analyzed by Illumina high-throughput sequencing. We validated the fidelity of VH:VL pairs identified by this approach and used the method to sequence the repertoire of three human cell subsets-peripheral blood IgG+ B cells, peripheral plasmablasts isolated after tetanus toxoid immunization and memory B cells isolated after seasonal influenza vaccinatio

Capturing Single Cell Genomes of Active Polysaccharide Degraders: An Unexpected Contribution of Verrucomicrobia

Microbial hydrolysis of polysaccharides is critical to ecosystem functioning and is of great interest in diverse biotechnological applications, such as biofuel production and bioremediation. Here we demonstrate the use of a new, efficient approach to recover genomes of active polysaccharide degraders from natural, complex microbial assemblages, using a combination of fluorescently labeled substrates, fluorescence-activated cell sorting, and single cell genomics. We employed this approach to analyze freshwater and coastal bacterioplankton for degraders of laminarin and xylan, two of the most abundant storage and structural polysaccharides in nature. Our results suggest that a few phylotypes of Verrucomicrobia make a considerable contribution to polysaccharide degradation, although they constituted only a minor fraction of the total microbial community. Genomic sequencing of five cells, representing the most predominant, polysaccharide-active Verrucomicrobia phylotype, revealed significant enrichment in genes encoding a wide spectrum of glycoside hydrolases, sulfatases, peptidases, carbohydrate lyases and esterases, confirming that these organisms were well equipped for the hydrolysis of diverse polysaccharides. Remarkably, this enrichment was on average higher than in the sequenced representatives of Bacteroidetes, which are frequently regarded as highly efficient biopolymer degraders. These findings shed light on the ecological roles of uncultured Verrucomicrobia and suggest specific taxa as promising bioprospecting targets. The employed method offers a powerful tool to rapidly identify and recover discrete genomes of active players in polysaccharide degradation, without the need for cultivation

Proteases of haematophagous arthropod vectors are involved in blood-feeding, yolk formation and immunity : a review

Ticks, triatomines, mosquitoes and sand flies comprise a large number of haematophagous arthropods considered vectors of human infectious diseases. While consuming blood to obtain the nutrients necessary to carry on life functions, these insects can transmit pathogenic microorganisms to the vertebrate host. Among the molecules related to the blood-feeding habit, proteases play an essential role. In this review, we provide a panorama of proteases from arthropod vectors involved in haematophagy, in digestion, in egg development and in immunity. As these molecules act in central biological processes, proteases from haematophagous vectors of infectious diseases may influence vector competence to transmit pathogens to their prey, and thus could be valuable targets for vectorial control

Serious case reviews: The lived experience of Black children

Despite the many high‐profile Black child deaths in England, race as a factor remains a largely underexplored factor of serious case reviews (SCRs). Evidence from analysis of SCRs indicates that race receives limited attention, or is virtually absent. Given that the main function of SCRs is to provide opportunities for learning lessons to improve practice, the way in which issues of race and culture may influence child protection processes for Black children is therefore of critical importance. In this article, we employ content analysis to examine the extent that race and cultural factors are considered in SCRs involving Black children. It is argued that race is often an important factor influencing Black children's experiences of abuse and neglect, as well as their encounters in the child protection system. This article therefore poses two key questions: (a) What questions are asked about race, ethnicity, and culture in SCRs concerning Black children? (b) How did the SCRs extract lessons to be learnt for improving practice to safeguard Black children? By extending the analysis of race and ethnicity in SCRs, this article furthers our understandings of the needs of Black children in the child protection system

When is a family a family? Evidence from survey data and implications for family policy

This article examines the correspondence between common assumptions about the American family and actual patterns. The assessment is based on national data on individuals, households, and families. Findings indicate that the coresident nuclear model should be considered a model rather than the model of family. Past as well as current marital ties need to be considered in defining “family,” and divorce rather than death should be the expected cause of losing the main breadwinner in the family, except among elderly women. Parent-child ties to either young or adult children often span separate households. Coresidents can include individuals other than nuclear family members, and change rather than stability is the modal pattern in living arrangements. Rather than shaping concepts of the family from a single mold, policy makers and researchers are better advised to recognize the diversity and fluidity in family and household structures.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44653/1/10834_2006_Article_BF02353666.pd

"My Death Will Not [Be] in Vain": Testimonials from Last Gift Rapid Research Autopsy Study Participants Living with HIV at the End of Life.

End-of-life (EOL) HIV cure-related research provides a novel approach to studying HIV reservoirs. The Last Gift is a rapid autopsy research study at the University of California San Diego that enrolls terminally ill people living with HIV (PLWHIV) with a desire to contribute to HIV cure-related research. We conducted in-depth baseline and follow-up interviews with Last Gift study participants. We analyzed interview data applying conventional content analysis. Since summer 2017, 13 participants have been enrolled (n = 11 males and 2 females; aged 45-89 years) and 8 participants interviewed. Terminal illnesses included cancers, heart diseases, and neurodegenerative illnesses. Our analysis revealed five key themes: (1) The Last Gift study has tremendous meaning for participants at the end of their life. (2) HIV-specific altruism was a primary motivator to join the Last Gift study, nested within the context of community, scientific advancement, and moral obligation. (3) Participants did not expect physical benefits yet they perceived emotional/psychological, financial, and societal/scientific benefits. (4) There were minimal participant-perceived risks and concerns. (5) Last Gift participants expressed immense gratitude toward study staff. The Last Gift study provides a framework for ethical HIV cure-related research at EOL and highlighted participants' perspectives, motivations, and experiences. Knowing how PLWHIV understand and experience such studies will remain critical to designing ethical, fully informed HIV cure research protocols that are acceptable to PLWHIV

"My Death Will Not [Be] in Vain": Testimonials from Last Gift Rapid Research Autopsy Study Participants Living with HIV at the End of Life.

End-of-life (EOL) HIV cure-related research provides a novel approach to studying HIV reservoirs. The Last Gift is a rapid autopsy research study at the University of California San Diego that enrolls terminally ill people living with HIV (PLWHIV) with a desire to contribute to HIV cure-related research. We conducted in-depth baseline and follow-up interviews with Last Gift study participants. We analyzed interview data applying conventional content analysis. Since summer 2017, 13 participants have been enrolled (n = 11 males and 2 females; aged 45-89 years) and 8 participants interviewed. Terminal illnesses included cancers, heart diseases, and neurodegenerative illnesses. Our analysis revealed five key themes: (1) The Last Gift study has tremendous meaning for participants at the end of their life. (2) HIV-specific altruism was a primary motivator to join the Last Gift study, nested within the context of community, scientific advancement, and moral obligation. (3) Participants did not expect physical benefits yet they perceived emotional/psychological, financial, and societal/scientific benefits. (4) There were minimal participant-perceived risks and concerns. (5) Last Gift participants expressed immense gratitude toward study staff. The Last Gift study provides a framework for ethical HIV cure-related research at EOL and highlighted participants' perspectives, motivations, and experiences. Knowing how PLWHIV understand and experience such studies will remain critical to designing ethical, fully informed HIV cure research protocols that are acceptable to PLWHIV