Transformation of a virulence associated gene of Haemophilus somnus into a strain lacking the gene.

The role of a 76 kDa surface antigen (p76) of Haemophilus somnus in virulence was investigated. The p76 gene from a virulent isolate of H. somnus (strain 2336) was introduced into an asymptomatic carrier strain (129Pt) lacking this gene.

Developmentally regulated transcripts and evidence of differential mRNA processing in Giardia lamblia.

Although encystation and excystation are crucial to transmission of Giardia lamblia, little is known about the regulation of these very distinct differentiation processes. Fingerprinting of giardial mRNA populations throughout the time course of differentiation demonstrated complex patterns in mRNA differential display. Certain transcripts appeared or increased, while others decreased or disappeared at specific times, in response to physiologic stimuli that mimic key stages in parasite descent through the host gastrointestinal tract.

Cell biology of the primitive eukaryote Giardia lamblia.

Giardia lamblia is an extremely primitive or early-diverging eukaryote that has been considered to have no typical ER or Golgi apparatus, although it is a complex and highly developed cell. Both the trophozoite and cyst have unusual surface proteins that enable these stages to survive in very different and hostile environments. We found that G.

Specialized surface adaptations of Giardia lamblia.

Although Giardia lamblia trophozoites were first described by Von Leeuwenhoek in his own diarrheic stool, relatively little is known of the basic biology of this common parasite or the pathophysiology of giardiasis. In particular, there is little specific information about trophozoite properties that cause diarrhea, as neither toxins nor conventional virulence factors have been identified. Therefore, parasite adaptations that promote cyst survival in the external environment and infection and trophozoite persistence in the small intestine, may be viewed as key virulence properties.

A lipoprotein-cholesterol-albumin serum substitute stimulates Giardia lamblia encystation vesicle formation.

We found previously that the A6 clone of Giardia lamblia strain WB that did not encyst in vitro was blocked at an early stage in differentiation, as it did not form encystation secretory vesicles (ESV) efficiently or express cyst antigens, in comparison with the related clone C6. We now report that A6 formed ESV normally in the suckling mouse model. Therefore, we asked whether our serum-containing encystation media might lack a stimulus or component or contain an inhibitor of ESV formation to which A6 was especially sensitive.

Bovine vaginal antibody responses to immunoaffinity-purified surface antigen of Tritrichomonas foetus.

Bovine trichomoniasis is a prevalent sexually transmitted disease of cattle caused by the protozoan Tritrichomonas foetus. Currently, diagnosis is most often made by culture. In order to provide a faster immunodiagnostic approach, a specific enzyme-linked immunosorbent assay (ELISA) was investigated.

Antigenic switching of TSA 417, a trophozoite variable surface protein, following completion of the life cycle of Giardia lamblia.

Expression of TSA 417, the predominant cysteine-rich variable surface protein of Giardia lamblia WB clone C6 trophozoites, did not change during encystation in vitro. However, in vitro excystation of cysts derived in vitro or in vivo consistently produced TSA 417 nonexpressing trophozoite populations, suggesting that completion of the life cycle leads to antigenic switching.

Conservation of a protective surface antigen of Tritrichomonas foetus.

Bovine trichomoniasis is a sexually transmitted disease caused by the flagellated protozoan Tritrichomonas foetus. A protective surface antigen was previously identified and immunoaffinity purified from T. foetus isolate D1 with cross-reactive monoclonal antibodies (MAbs) TF1.

Giardia lamblia: absence of cyst antigens and reduced secretory vesicle formation and bile salt uptake in an encystation-deficient subline.

Encystation of Giardia lamblia entails the appearance of a number of new antigens, as well as formation of a novel class of large encystation-specific secretory vesicles (ESV) that transport stage-specific proteins to the nascent cyst wall. The monoclonal antibody GCSA-1, which was raised against purified cyst walls, recognizes protein species of approximately 26-46 kDa that are regulated by exposure to bile (plus lactic acid) and alkaline pH, the factors that induce encystation. The GCSA-1 epitope is maximally expressed after approximately 14 hr of encystation and localizes to the interior, but not the membrane of the ESV as shown by frozen section immunoelectron microscopy.

Giardia lamblia: post-translational processing and status of exposed cysteine residues in TSA 417, a variable surface antigen.

Giardia lamblia develops and thrives within the harsh and variable environment of the human small intestine. To survive this environment, Giardia has evolved a unique family of antigenically variable, extremely cysteine-rich surface proteins. We have characterized the expression of one of these antigens, TSA 417, at the gene and protein levels.

Molecular analysis of a gene encoding a serum-resistance-associated 76 kDa surface antigen of Haemophilus somnus.

Haemophilus somnus is a Gram-negative bacterial bovine pathogen which can cause disease or be carried asymptomatically. We previously showed that four serum-sensitive isolates from asymptomatic carriers lacked a 13.4 kb sequence of chromosomal DNA that was present in two virulent serum-resistant strains.

Site-directed mutations in the relaxase operon of RP4.

Mutations were constructed by site-directed mutagenesis in the relaxase operon of the broad-host-range plasmid RP4. The mutations were constructed in smaller plasmids, recombined into the 60-kb RP4 plasmid, and tested for their ability to transfer. The relaxase operon contains the transfer genes traJ, traH, and traI, which are involved in nicking at the transfer origin to generate the single strand destined to be transferred to the recipient cell.

Mutational analysis of essential IncP alpha plasmid transfer genes traF and traG and involvement of traF in phage sensitivity.

Although the broad-host-range IncP plasmids can vegetatively replicate in diverse gram-negative bacteria, the development of shuttle vector systems has established that the host range for IncP plasmid conjugative transfer is greater than the range of bacteria that sustain IncP replicons. Towards understanding IncP plasmid conjugation and the connection between IncP conjugation and Agrobacterium tumefaciens T-DNA transfer to plants, two sets of mutants were generated in the larger transfer region (Tra1) of the IncP alpha plasmid RK2. Mutagenesis strategies were chosen to minimize transcriptional polar effects.

Human secretory and serum antibodies recognize environmentally induced antigens of Giardia lamblia.

The variability in duration and severity of infection with Giardia lamblia is likely to be due to trophozoite interactions with immune and nonimmune components of the small intestinal milieu. Despite its potential importance, nothing is known of the isotype or the specificity of the secretory antibody response to G. lamblia.