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Borrelia burgdorferi outer surface protein (Osp) A has been used as a Lyme disease vaccine that blocks transmission: OspA antibodies of immune hosts enter ticks during blood feeding and destroy spirochetes before transmission to the host can occur. B. burgdorferi produce OspA in the gut of unfed Ixodes scapularis ticks, and many spirochetes repress OspA production during the feeding process. This preferential expression suggests that OspA may have an important function in the vector. Here we show that OspA mediates spirochete attachment to the tick gut by binding to an I. scapularis protein. The binding domains reside in the central region and COOH-terminus of OspA. OspA also binds to itself, suggesting that spirochete-spirochete interactions may further facilitate adherence in the gut. OspA-mediated attachment in the tick provides a possible mechanism for how stage-specific protein expression can contribute to pathogenesis during the B. burgdorferi natural cycle.

The agent of human granulocytic ehrlichiosis (HGE) is an emerging tick-borne pathogen that resides in neutrophils and can be cultured in a promyelocytic (HL-60) cell line. In response to microbes, polymorphonuclear leukocytes normally activate the NADPH oxidase enzyme complex and generate superoxide anion (O2-). However, HL-60 cells infected with HGE bacteria did not produce O2- upon activation with PMA. RT-PCR demonstrated that HGE organisms inhibited mRNA expression of a single component of NADPH oxidase, gp91phox, and FACS analysis showed that plasma membrane-associated gp91phox protein was reduced on the infected cells. Infection with HGE organisms also decreased gp91phox mRNA levels in splenic neutrophils in a murine model of HGE, demonstrating this phenomenon in vivo. Therefore, HGE bacteria repress the respiratory burst by down-regulating gp91phox, the first direct inhibition of NADPH oxidase by a pathogen.

Thrombotic thrombocytopenic purpura (TTP) is an unusual complication of systemic lupus erythematosus (SLE). There are no randomized, prospective studies of its treatment. The association of plasma infusions and (or) plasmapheresis with steroids improves survival when compared with steroid treatment alone, but these patients still have a higher mortality than those with "classic" TTP. The role of immunosuppressive drugs in the management of this disorder remains uncertain. We report two cases of TTP in SLE which presumably benefited from the addition of cyclophosphamide to the treatment with plasmapheresis and steroids.

The role of interleukin (IL)-11, a cytokine with potent anti-inflammatory properties, in murine Lyme disease was investigated. Borrelia burgdorferi-infected mice treated with IL-11 developed less arthritis than did control animals. In contrast, IL-11 blocking antibodies increased Lyme arthritis. Murine Lyme carditis was not affected by either IL-11 or IL-11 antibodies. Administration of IL-11 was associated with increased production of mRNA for IL-12 and inducible nitric oxide synthase but not interferon-gamma or IL-4 in B. burgdorferi-infected mice, suggesting a predominant effect of IL-11 on the innate immune response. These data show that IL-11 selectively reduced joint but not cardiac inflammation caused by B. burgdorferi in mice.

The expression of a Borrelia burgdorferi gene, erpT, was investigated throughout the spirochaete life cycle in the arthropod vector and the murine host. Three phage clones from a B. burgdorferi DNA expression library synthesized a 30 kDa antigen that was recognized by antibodies in the sera of B. burgdorferi-infected mice but not mice hyperimmunized with B. burgdorferi lysates. Differential antibody binding suggested that this protein was preferentially expressed in vivo. This antigen was designated ErpT, based upon 99.6% homology with the BBF01 sequence in the B. burgdorferi genome. ErpT was not detected on spirochaetes cultured in BSK II medium by indirect immunofluorescence or in B. burgdorferi lysates by immunoblotting, implying that ErpT is not readily produced in vitro. erpT mRNA was not discernible by Northern blot but was identified by RNA polymerase chain reaction in vitro, indicating that erpT is expressed at low levels by cultured spirochaetes. erpT expression was then investigated in the vector and mice because B. burgdorferi do not normally reside in culture medium. RNA polymerase chain reaction and immunofluorescence studies demonstrated that erpT was expressed by a small minority of B. burgdorferi (11/500, 2.2%) within unfed ticks and then repressed during engorgement. erpT mRNA or ErpT antibodies were first detected in B. burgdorferi-infected mice at 4 weeks, suggesting that erpT was not expressed in the early stages of murine infection. Then, during persistent infection, RNA polymerase chain reaction showed that erpT was expressed by B. burgdorferi within the joints, heart and spleen, but not by spirochaetes in the skin. Immunization of mice with ErpT was antigenic but was not protective. These studies demonstrate that B. burgdorferi erpT is differentially expressed throughout the B. burgdorferi life cycle, in both the vector and the mammalian host, and is primarily expressed in extracutaneous sites during murine infection.

Recently, interleukin (IL)-6 was shown to be one of the earliest factors that trigger the differentiation of naive T cells into effector Th2 cells in vitro. Lyme arthritis was studied in IL-6-deficient mice, since joint inflammation is influenced by the T helper cell response against Borrelia burgdorferi. Arthritis incidence increased in B. burgdorferi-infected IL-6-deficient mice compared with that in controls. Furthermore, splenocytes of B. burgdorferi-infected IL-6-deficient mice produced significantly less IL-4 in response to Borrelia antigens than did C57BL/6 (B6) mice, and B. burgdorferi-specific IgG2b levels were significantly reduced in IL-6-deficient mice at 60 days of infection. These results extend previous in vitro observations by demonstrating an in vivo role for IL-6 in the differentiation of CD4 T cells toward a Th2 phenotype and further show that CD4 T cell responses influence murine Lyme arthritis.

Interleukin (IL)-4 is the most potent factor that causes naive CD4+ T cells to differentiate to the T helper cell (Th) 2 phenotype, while IL-12 and interferon gamma trigger the differentiation of Th1 cells. However, the source of the initial polarizing IL-4 remains unclear. Here, we show that IL-6, probably secreted by antigen-presenting cells, is able to polarize naive CD4+ T cells to effector Th2 cells by inducing the initial production of IL-4 in CD4+ T cells. These results show that the nature of the cytokine (IL-12 or IL-6), which is produced by antigen-presenting cells in response to a particular pathogen, is a key factor in determining the nature of the immune response.

Effects on bone remodeling have been attributed to epidermal growth factor (EGF). Sialoadenectomy (SX) removes the major source of EGF in rodents and decreases both salivary and serum EGF levels. EGF effects on rat alveolar bone remodeling manifested by molar drift (MD) and orthodontic tooth movement (OTM) were examined using the following two approaches: 1) EGF depletion by SX and replacement by orally administered EGF (50 micrograms.animal-1.day-1); 2) sham rats supplemented with matching amounts of EGF. MD and OTM were measured using cephalometric radiographs; bone formation was measured histomorphometrically using tetracycline labeling. Normal MD was not detected after SX, and alveolar bone formation was significantly reduced both around the tooth and in nondental sites. Replacement EGF given to SX rats and supplemental EGF administered to sham rats changed the direction and enhanced the rate of MD. A mesially directed orthodontic force applied to the molars of SX animals increased bone formation on the distal aspect of the tooth roots. Supplemental EGF did not significantly affect OTM. EGF affects alveolar bone remodeling, as manifested clinically by alterations in normal maxillary MD.

A DNA probe specific for genes coding for the heat-labile toxin type II (LT-II), heat-stable toxin type Ib (ST-Ib) and Shiga-like toxin type II (SLT-II), were used to examine 118 fecal Escherichia coli strains isolated from healthy pigs. Fifty-six (47.4%) of the isolated were hybridized with the LT-II probe. Thirty-nine strains (33%) possessed ST-Ib genes and five of the 118 isolates (4.2%) showed homology with the SLT-II DNA probe. E. coli that possessed unusual toxin genes for pigs were isolated with a high frequency, which indicates the importance of toxigenic clones found in nature. Uncommon virulence factors should be examined in order to improve the efficiency of diagnosis and control procedures.

Seventy-four strains of Aeromonas hydrophila isolated from water and sediments of the River Porma (León, N.W. Spain) were characterized biochemically and biologically. Fifty-seven strains (77.02%) were virulent for rainbow trout (Salmo gairdneri) by intramuscular challenge but showed differing degree of pathogenicity which could not be associated with the source. A lack of correlation between caseinase, haemolytic and cytotoxic activities of the strains and their isolation source was also observed. Only two surface characters, acriflavine 0.2% agglutination and non-agglutinating SP-/PAB-phenotypes, were significantly associated with water and sediment strains, respectively.

A genomic library of Aeromonas hydrophila F9 was constructed by using pBR322 as a vector. From that, two DNA fragments (5.8 and 11.6 kb) were isolated containing genetic information to complement trpA and trpB defects (5.8-kb fragment) and to complement trpA, trpB, trpC, trpD, and trpE defects (11.6-kb fragment) in Escherichia coli mutants. Evidence of the existence of a secondary promoter is given.

Aeromonas virulence is thought to depend on multigenic functions. The gene for an extracellular protease from Aeromonas hydrophila SO2/2 was cloned in Escherichia coli C600-1 by using pIJ860, bifunctional plasmid, as a vector. The gene encodes for a temperature-labile serine protease (P2) with a molecular mass of approx. 68 kDa which is highly inhibited by PMSF. The gene was expressed in Streptomyces lividans 1326 by transforming protoplasts with the original clone pPA2. We were also able to transfer and express the prt P2 gene in Pseudomonas putida by mating experiments. The protein P2 was secreted into the periplasms of both P. putida and E. coli C600-1 being identical in properties to one of the proteases secreted into the culture supernatant by A. hydrophila SO2/2.