Antigenic and 3D structural characterization of soluble X4 and hybrid X4-R5 HIV-1 Env trimers.

BACKGROUND: HIV-1 is decorated with trimeric glycoprotein spikes that enable infection by engaging CD4 and a chemokine coreceptor, either CCR5 or CXCR4. The variable loop 3 (V3) of the HIV-1 envelope protein (Env) is the main determinant for coreceptor usage. The predominant CCR5 using (R5) HIV-1 Env has been intensively studied in function and structure, whereas the trimeric architecture of the less frequent, but more cytopathic CXCR4 using (X4) HIV-1 Env is largely unknown, as are the consequences of sequence changes in and near V3 on antigenicity and trimeric Env structure. RESULTS: Soluble trimeric gp140 Env constructs were used as immunogenic mimics of the native spikes to analyze their antigenic properties in the context of their overall 3D structure. We generated soluble, uncleaved, gp140 trimers from a prototypic T-cell line-adapted (TCLA) X4 HIV-1 strain (NL4-3) and a hybrid (NL4-3/ADA), in which the V3 spanning region was substituted with that from the primary R5 isolate ADA. Compared to an ADA (R5) gp140, the NL4-3 (X4) construct revealed an overall higher antibody accessibility, which was most pronounced for the CD4 binding site (CD4bs), but also observed for mAbs against CD4 induced (CD4i) epitopes and gp41 mAbs. V3 mAbs showed significant binding differences to the three constructs, which were refined by SPR analysis. Of interest, the NL4-3/ADA construct with the hybrid NL4-3/ADA CD4bs showed impaired CD4 and CD4bs mAb reactivity despite the presence of the essential elements of the CD4bs epitope. We obtained 3D reconstructions of the NL4-3 and the NL4-3/ADA gp140 trimers via electron microscopy and single particle analysis, which indicates that both constructs inherit a propeller-like architecture. The first 3D reconstruction of an Env construct from an X4 TCLA HIV-1 strain reveals an open conformation, in contrast to recently published more closed structures from R5 Env. Exchanging the X4 V3 spanning region for that of R5 ADA did not alter the open Env architecture as deduced from its very similar 3D reconstruction. CONCLUSIONS: 3D EM analysis showed an apparent open trimer configuration of X4 NL4-3 gp140 that is not modified by exchanging the V3 spanning region for R5 ADA.

Closely related Campylobacter jejuni strains from different sources reveal a generalist rather than a specialist lifestyle.

BACKGROUND: Campylobacter jejuni and Campylobacter coli are human intestinal pathogens of global importance. Zoonotic transmission from livestock animals or animal-derived food is the likely cause for most of these infections. However, little is known about their general and host-specific mechanisms of colonization, or virulence and pathogenicity factors. In certain hosts, Campylobacter species colonize persistently and do not cause disease, while they cause acute intestinal disease in humans. RESULTS: Here, we investigate putative host-specificity using phenotypic characterization and genome-wide analysis of genetically closely related C. jejuni strains from different sources. A collection of 473 fresh Campylobacter isolates from Germany was assembled between 2006 and 2010 and characterized using MLST. A subset of closely related C. jejuni strains of the highly prevalent sequence type ST-21 was selected from different hosts and isolation sources. PCR typing of strain-variable genes provided evidence that some genes differed between these strains. Furthermore, phenotypic variation of these strains was tested using the following criteria: metabolic variation, protein expression patterns, and eukaryotic cell interaction. The results demonstrated remarkable phenotypic diversity within the ST-21 group, which however did not correlate with isolation source. Whole genome sequencing was performed for five ST-21 strains from chicken, human, bovine, and food sources, in order to gain insight into ST-21 genome diversity. The comparisons showed extensive genomic diversity, primarily due to recombination and gain of phage-related genes. By contrast, no genomic features associated with isolation source or host were identified. CONCLUSIONS: The genome information and phenotypic data obtained in vitro and in a chicken infection model provided little evidence of fixed adaptation to a specific host. Instead, the dominant C. jejuni ST-21 appeared to be characterized by phenotypic flexibility and high genetic microdiversity, revealing properties of a generalist. High genetic flexibility might allow generalist variants of C. jejuni to reversibly express diverse fitness factors in changing environments.