Humoral Responses against BQ.1.1 Elicited after Breakthrough Infection and SARS-CoV-2 mRNA Vaccination.

The Omicron BQ.1.1 variant is now the major SARS-CoV-2 circulating strain in many countries. Because of the many mutations present in its Spike glycoprotein, this variant is resistant to humoral responses elicited by monovalent mRNA vaccines. With the goal to improve immune responses against Omicron subvariants, bivalent mRNA vaccines have recently been approved in several countries. In this study, we measure the capacity of plasma from vaccinated individuals, before and after a fourth dose of mono- or bivalent mRNA vaccine, to recognize and neutralize the ancestral (D614G) and the BQ.1.1 Spikes. Before and after the fourth dose, we observe a significantly better recognition and neutralization of the ancestral Spike. We also observe that fourth-dose vaccinated individuals who have been recently infected better recognize and neutralize the BQ.1.1 Spike, independently of the mRNA vaccine used, than donors who have never been infected or have an older infection. Our study supports that hybrid immunity, generated by vaccination and a recent infection, induces higher humoral responses than vaccination alone, independently of the mRNA vaccine used

Intra-Host Evolution Analyses in an Immunosuppressed Patient Supports SARS-CoV-2 Viral Reservoir Hypothesis.

Throughout the SARS-CoV-2 pandemic, several variants of concern (VOCs) have been identified, many of which share recurrent mutations in the spike glycoprotein's receptor-binding domain (RBD). This region coincides with known epitopes and can therefore have an impact on immune escape. Protracted infections in immunosuppressed patients have been hypothesized to lead to an enrichment of such mutations and therefore drive evolution towards VOCs. Here, we present the case of an immunosuppressed patient that developed distinct populations with immune escape mutations throughout the course of their infection. Notably, by investigating the co-occurrence of substitutions on individual sequencing reads in the RBD, we found quasispecies harboring mutations that confer resistance to known monoclonal antibodies (mAbs) such as S:E484K and S:E484A. These mutations were acquired without the patient being treated with mAbs nor convalescent sera and without them developing a detectable immune response to the virus. We also provide additional evidence for a viral reservoir based on intra-host phylogenetics, which led to a viral substrain that evolved elsewhere in the patient's body, colonizing their upper respiratory tract (URT). The presence of SARS-CoV-2 viral reservoirs can shed light on protracted infections interspersed with periods where the virus is undetectable, and potential explanations for long-COVID cases

Disentangling molecular and clinical stratification patterns in beta-galactosidase deficiency

INTRODUCTION: This study aims to define the phenotypic and molecular spectrum of the two clinical forms of β-galactosidase (β-GAL) deficiency, GM1-gangliosidosis and mucopolysaccharidosis IVB (Morquio disease type B, MPSIVB). METHODS: Clinical and genetic data of 52 probands, 47 patients with GM1-gangliosidosis and 5 patients with MPSIVB were analysed. RESULTS: The clinical presentations in patients with GM1-gangliosidosis are consistent with a phenotypic continuum ranging from a severe antenatal form with hydrops fetalis to an adult form with an extrapyramidal syndrome. Molecular studies evidenced 47 variants located throughout the sequence of the GLB1 gene, in all exons except 7, 11 and 12. Eighteen novel variants (15 substitutions and 3 deletions) were identified. Several variants were linked specifically to early-onset GM1-gangliosidosis, late-onset GM1-gangliosidosis or MPSIVB phenotypes. This integrative molecular and clinical stratification suggests a variant-driven patient assignment to a given clinical and severity group. CONCLUSION: This study reports one of the largest series of b-GAL deficiency with an integrative patient stratification combining molecular and clinical features. This work contributes to expand the community knowledge regarding the molecular and clinical landscapes of b-GAL deficiency for a better patient management

Production of Multiple Brain-Like Ganglioside Species Is Dispensable for Fas-Induced Apoptosis of Lymphoid Cells

Activation of an acid sphingomyelinase (aSMase) leading to a biosynthesis of GD3 disialoganglioside has been associated with Fas-induced apoptosis of lymphoid cells. The present study was undertaken to clarify the role of this enzyme in the generation of gangliosides during apoptosis triggered by Fas ligation. The issue was addressed by using aSMase-deficient and aSMase-corrected cell lines derived from Niemann-Pick disease (NPD) patients. Fas cross-linking elicited a rapid production of large amounts of complex a- and b-series species of gangliosides with a pattern and a chromatographic behavior as single bands reminiscent of brain gangliosides. The gangliosides were synthesized within the first ten minutes and completely disappeared within thirty minutes after stimulation. Noteworthy is the observation that GD3 was not the only ganglioside produced. The production of gangliosides and the onset of apoptotic hallmarks occurred similarly in both aSMase-deficient and aSMase-corrected NPD lymphoid cells, indicating that aSMase activation is not accountable for ganglioside generation. Hampering ganglioside production by inhibiting the key enzyme glucosylceramide synthase did not abrogate the apoptotic process. In addition, GM3 synthase-deficient lymphoid cells underwent Fas-induced apoptosis, suggesting that gangliosides are unlikely to play an indispensable role in transducing Fas-induced apoptosis of lymphoid cells

Vibrio cholerae genomic diversity within and between patients

Cholera is a severe, water-borne diarrhoeal disease caused by toxin-producing strains of the bacterium Vibrio cholerae. Comparative genomics has revealed ‘waves’ of cholera transmission and evolution, in which clones are successively replaced over decades and centuries. However, the extent of V. cholerae genetic diversity within an epidemic or even within an individual patient is poorly understood. Here, we characterized V. cholerae genomic diversity at a micro-epidemiological level within and between individual patients from Bangladesh and Haiti. To capture within-patient diversity, we isolated multiple (8 to 20) V. cholerae colonies from each of eight patients, sequenced their genomes and identified point mutations and gene gain/loss events. We found limited but detectable diversity at the level of point mutations within hosts (zero to three single nucleotide variants within each patient), and comparatively higher gene content variation within hosts (at least one gain/loss event per patient, and up to 103 events in one patient). Much of the gene content variation appeared to be due to gain and loss of phage and plasmids within the V. cholerae population, with occasional exchanges between V. cholerae and other members of the gut microbiota. We also show that certain intra-host variants have phenotypic consequences. For example, the acquisition of a Bacteroides plasmid and non-synonymous mutations in a sensor histidine kinase gene both reduced biofilm formation, an important trait for environmental survival. Together, our results show that V. cholerae is measurably evolving within patients, with possible implications for disease outcomes and transmission dynamics

Host plasma low density lipoprotein particles as an essential source of lipids for the bloodstream forms of Trypanosoma brucei.

In contrast to mammalian cells, bloodstream forms of Trypanosoma brucei show no activity for fatty acid and sterol synthesis and critically depend on plasma low density lipoprotein (LDL) particles for their rapid growth. We report here that these parasites acquire such lipids by receptor-mediated endocytosis of LDL, subsequent lysosomal degradation of apoprotein B-LDL, and utilization of these lipids. Uptake of LDL-associated [3H]sphingomyelin and of LDL-associated [3H]cholesteryl oleate paralleled each other, and that of 125I-apoprotein B-LDL showed saturation and could be inhibited by unlabeled LDL or by anti-LDL receptor antibodies. Metabolism of lipids carried by LDL was abolished by chloroquine and by the thiol protease inhibitor, leupeptin. Sphingomyelin was cleaved by an acid sphingomyelinase to yield ceramide, which was itself split up into sphingosine and fatty acids. The latter were further incorporated into phosphatidylcholine, triacylglycerols, or cholesteryl esters. Similarly, cholesteryl oleate was hydrolyzed by an acid lipase to yield free cholesterol, which was reesterified with fatty acids, presumably in the cytosol. Like free cholesterol, LDL provided substrate for cholesterol esterification. In the culture-adapted procyclic form of T. brucei, which is capable of sterol synthesis, exogenous LDL-cholesterol rather than endogenously synthesized sterol was utilized for sterol esterification. Interference with exogenous supply of lipids via receptor-mediated endocytosis of LDL should be explored to fight against trypanosomiasis

Uneven X inactivation in a female monozygotic twin pair with Fabry disease and discordant expression of a novel mutation in the alpha-galactosidase A gene.

We describe two female monozygotic (MZ) twins heterozygous for Fabry disease, an X linked disorder resulting from the deficient activity of alpha-galactosidase A. While one of the twins was clinically affected, the other was asymptomatic. Enzymatic assay of alpha-galactosidase in blood leucocytes, skin fibroblasts, Epstein-Barr virus transformed lymphoid cell lines, and hair follicles of the twins and their parents confirmed the heterozygous status of the twins and indicated that Fabry disease had occurred as a result of a de novo mutation. The son of the unaffected twin sister was shown to be hemizygous. Molecular analysis of the alpha-galactosidase A gene permitted the identification of an as yet undescribed point mutation at position 10182 of exon 5 which causes an Asp to Asn substitution at codon 231. Single strand conformation polymorphism (SSCP) analysis again showed the heterozygous status of the twins and a normal pattern in their parents. The basis for the discordant expression of this d novo mutation in the twins was investigated by studying their X inactivation status. Analysis of the inactive X specific methylation at the androgen receptor gene showed unbalanced inactivation in the twins' fibroblasts and in opposite directions. While the maternally derived X chromosome was preferentially active in the asymptomatic twin, the paternal X chromosome was active in the other, affected twin and was found in her hemizygotic nephew. These data suggest that the paternal X chromosome carries the de novo alpha-galactosidase A mutation and that uneven X inactivation is the underlying mechanism for disease expression in this novel female MZ twin pair. This is the first documented case of female twins discordant for Fabry disease

Danon disease: Further clinical and molecular heterogeneity

Two families of Greek patients with subclinical to severe cardiomyopathy are presented. The diagnosis of Danon disease was supported by a total lack of LAMP2 immunostaining in cultured skin fibroblasts and muscle biopsies. The LAMP2 mutation carried by one patient (c.928G>A) has already been reported but with different symptoms. The second patient had a novel point deletion. This has not been described previously, but it could be detected easily by restriction analysis. This mutation was also found in the patient's brother, and it was associated with severe cardiomyopathy leading to heart failure. Surprisingly, the proband also had partial reduction of α-galactosidase A activity, despite the absence of characteristic clinical features of Fabry disease. A substitution in the GLA gene (c.937G>T) was found, and its involvement in the cardiac disease is discussed. © 2009 Wiley Periodicals, Inc