Cholera is an acute diarrheal illness burdening several developing countries caused by toxigenic Vibrio cholerae, where endemics maintain a seasonal pattern and occur more than once a year. Cholera is endemic in certain regions of Africa and South America, and southern Asia, where outbreaks are associated with poor hygiene and sanitary conditions. Every year, 3–5 million cases of cholera are diagnosed, and it causes more than 100,000 deaths worldwide. Cholera toxin is secreted by the V. cholerae bacteria and causes extreme secretory diarrhea, most commonly in poor hygiene environment. Watery diarrhea, vomiting, and abdominal cramps characterize the illness and approximately 5–10% of patients die of severe fluid loss if left untreated. The structure and function of the cholera toxin, its subunits, receptor, and impact on hyperactivation of cyclic adenosine monophosphate (cAMP) were sufficiently described in the 1970s. These findings fit with epidemiologic observations, which determined that the cholera toxin must first enter intestinal cells by binding to monosialoganglioside (GM1) on the host’s epithelial surface. The correlation between increase risk of V. cholerae infection and individuals with a particular ABO blood group type is unclear because of the scarce information and few studies conducted. Thus, this study reviews published research articles to better understanding the association between the blood group O and susceptibility to developing severe cholera symptoms. Several large studies have recorded an association between ABO blood groups and different infectious agents). Anthropological surveys suggest that the racial and geographic distribution of human blood types reflects tendencies towards specific erythrocyte types susceptible to infectious disease, such as cholera and malaria. Experimentally, the V. cholerae toxin has been extensively used as an experimental adjuvant, and its association with ABO groups is of practical importance for the development of an oral cholera vaccination. The results of previous studies provided strong evidence that individuals with blood type O are more vulnerable than other persons to severe cholera symptoms, even though the biologic basis for this association remains unknown.

No description available.

Les souches humaines de rotavirus du groupe A (RVA) reconnaissent des glycannes fucosylés de la famille des Histo-Blood Group Antigens (HBGAs) et des gangliosides via la protéine de capside VP8*. L’interaction avec les gangliosides est essentielle pour l’entrée cellulaire et l’absence de ligands fucosylés dû au polymorphisme génétique des HBGAs est associée à une résistance à la gastroentérite sévère. Nos objectifs étaient de délimiter la contribution des HBGAs et du ganglioside GM1a dans le processus d’infection et d’explorer les conséquences du polymorphisme des HBGAs sur la transmission du virus et l’efficacité des vaccins vivants disponibles.génoty Ces travaux ont permis de montrer la concordance entre la spécificité glycannique des VP8* P[8], génotype le plus fréquent en France, et la sensibilité HBGA-dépendante à la gastroentérite sévère. La reconnaissance des HBGAs par les souches humaines de RVA apparaît donc essentielle pour l’infection symptomatique. Néanmoins, nos résultats suggèrent que l’attachement aux HBGAs correspond à un événement précoce puisqu’il n’est pas nécessaire pour l’infection de cellules peu différenciées par les souches P[8] adaptées à la culture. La contribution du GM1a dans l’infection reste incertaine. Enfin, nous avons montré que la reconnaissance des HBGAs est conservée entre des souches P[8] récentes et anciennes, indiquant que le polymorphisme des HBGAs pourrait contribuer à expliquer le défaut d’efficacité des vaccins dans les régions où la fréquence d’individus n’exprimant pas les ligands fucosylés est élevée. / Human strains of rotavirus A (RVAs) recognize fucosylated glycans of the histo-blood group family (HBGAs) as well as gangliosides through the VP8* protein of their capsid. Interaction with gangliosides is essential for cell entry and lack of fucosylated ligands due to HBGAs genetic polymorphism is associated with resistance to RVA gastroenteritis. Our goals are to delineate the contribution of HBGAs and gangliosides in the infection process and to explore the consequences of HBGAs polymorphisms on the virus transmission and efficacy of the available live vaccines. This study highlighted the concordance between the glycan specificity of P[8] VP8*, the most common genotype in France, and the HBGA-dependant susceptibility to RVA gastroenteritis. The recognition of HBGAs by human RVA strains therefore appears essential to the infection. Yet, our results suggest that HBGA binding corresponds to an early event since it is not required for infection of poorly differentiated cells by cell culture-adapted P[8] strains. The contribution of GM1a on infection remains unclear. Finally, we showed that HBGA recognition is conserved between recent and older P[8] strains, suggesting that HBGAs polymorphism may contribute to explain the low efficacy of vaccines in areas where the frequency of individuals who do not express fucosylated ligands is high.

Alloimmunisation becomes a problem when serological discrepancies occur in matching antigens between donors and patients for blood transfusion. The rate of alloimmunisation has been increased especially in multiply transfused patients. Blood group genotyping (BGG) is a DNA-based assay that aids in reducing this situation. Currently, many platforms of BGG have become available, in which every technique has its own advantages and disadvantages. All these platforms lack the ability to identify novel alleles that might have an unknown clinical significance. The advent of next-generation sequencing (NGS) offers identification of the unprecedented alleles due to its basis of sequence-based typing. Moreover, it provides an extreme high-throughput which may be able to screen many donors and patients in a single run. In this project, two approaches have been developed in generating sequencing libraries followed by sequencing on the Ion Torrent Personal Genome MachineTM platform (Ion PGMTM). The first approach was amplicon-based target selection using Ion AmpliseqTM Custom Panel, designated as Human Erythrocyte Antigens and Human Platelet Antigens Panel (HEA and HPA Panel). This panel assay screens 11 blood group systems, as well as 16 human platelet antigens. The outcome was extraordinary, in particular four novel alleles had been identified out of 28 samples, one in the RHCE gene 208C > T (Arg70Trp) in exon 2 and three in the KEL gene. The first SNP was 331G > A (Ala111Thr) in exon 4. The second SNP was 1907C > T (Ala636Val) in exon 17 and the third SNP was 2165T > C (Leu722Pro) in exon 19. However, some issues occurred regarding co-amplification of homologous genes. The second approach was a long-range polymerase chain reaction (LR-PCR) based approach. This method provided a high resolution assay by amplification of entire genes, including the non-coding area, of the Kell and Rh blood group systems. The Kell blood group was initially utilised as a model in order to apply the same approach on the Rh system. Most alleles encoding the antigens of the Kell blood group, especially the high prevalence ones, were identified. The Rh LR-PCR approach was carried out by amplification of the RHD and RHCE genes with seven amplicons. For five RhD-positive samples no mutations were observed within the coding areas. On the other hand, five serotyped weak D samples were genotyped as; two weak D type 1, two weak D type 2 and one DAR3.1 weak partial D 4.0 (RHD*DAR3.01). Regarding the RHCE, the following antigens (C, c, E, c) were predicted properly from the sequencing data. Moreover, the RHCE*ceVS.02 was identified. 64 and 39 intronic SNPs were identified in RHD and RHCE genes, respectively. The intronic SNPs assisted the genotyping process by identifying the haplotype of interest. Interestingly, the novel allele identified in the RHCE gene by the HEA and HPA Panel was confirmed to belong to the RHCE gene by the LR-PCR approach, indicating the panel misaligned it to the RHD gene. In conclusion, NGS paves the way to be an alternative substitution to the previous molecular techniques. It would supplant the conventional serology for typing blood for transfusion.

Serological discrepancies in matching blood group antigens between donors and patients for blood transfusion may lead to alloimmunisation, especially in multiply transfused patients. Blood group genotyping (BGG) has contributed in reducing this issue. ABO, Fy and Jk antigens are among those to be causative for alloimmunisation through transfusion or pregnancy. The number of alleles of these clinically significant blood groups is ever increasing. Currently, all commercially available high-throughput BGG platforms are only based on pre-defined polymorphisms. Consequently, novel or rare alleles that might have clinical significance are not identified. Next generation sequencing (NGS) circumvents this issue by providing high-throughput comprehensive genotyping of blood group genes in discovery mode to find all existing and novel mutations. Accordingly, a large number of individuals can be genotyped in a single run. Here, we describe an NGS-based method coupled with long-range polymerase chain reaction (LR-PCR) for high-throughput, rapid and extensive genotyping of FY, JK and ABO blood group genes. The Ion Torrent Personal Genome Machine (PGMTM) was used for sequencing the entire FY, JK and ABO blood group genes including flanking regions. Accordingly, high resolution genotyping was obtained. 53 genomic DNA samples were sequenced and genotyped for FY, 67 for JK and 47 for ABO. Sequencing data were aligned to the gene reference sequence derived from the human genome (hg19) to analyse variants. Analysis was accomplished by software packages, such as Ion Torrent SuiteTM plugins. Sanger sequencing of cDNA and cDNA clones was used to confirm findings in the JK gene. The sequencing data had a coverage depth of more than 5000x for FY, 700x for JK and 600x for ABO. NGS data matched with the serological phenotypes of FY alleles FY*A, FY*B and FY*02 Null main polymorphisms, such as FY*A/FY*B (125G > A) in exon 2 and (-67 T > C) in the promotor region. JK variant analysis revealed that the JK*01W.01 allele (130G > A) is common (10/67 samples) with normal antigenicity. The previously described silencing polymorphism (810G > A), leading to a purported JK*B null allele, restores a splice site and does not correlate with loss of Jkb antigenicity (10/67 samples). JK intron analysis revealed several new JK alleles described in this thesis. All 7 exons, introns and the flanking regions of the ABO gene were covered by only four amplicons. Several rare O alleles were found, such as O73 and O75, while one suggested novel O allele was characterised by a missense SNP 482G > A (Arg161His) in exon 7. The ABO reference sequence from hg19 appeared to resemble (O01 and O02) alleles. The intronic SNPs might be used to distinguish between alleles more accurately as a correlation of the intronic SNPs with the alleles was noted for the homozygous O alleles. It is predicted that NGS-based genotyping will replace not only microarray-based genotyping but also serology in the blood group typing of individuals, with great advancements in technology and molecular knowledge being expected in the near future.

Diss. (sammanfattning) Umeå : Univ., 2004. / Härtill 4 uppsatser. I publ. felaktig serietitel: Umeå University medical dissertation.

Thesis (Doctoral)--Ludwig-Maximilians-Universität zu München, 1934.

Thesis (doctoral)--Lund University, 1997. / Added t.p. with thesis statement inserted.

Thesis--Copenhagen. / "Oversigt": p. [166]-170. "References": p. [171]-172.

Thesis--Copenhagen. / English summary: p. [138]-139. "Litteraturfortegnelse": p. [140]-148.