Biomedical subjects

Alan G Barbour

Publications and source records attributed to Alan G Barbour.

24 records · Page 2Linked to original sources

PubMed2004

Transgenic expression of RecA of the spirochetes Borrelia burgdorferi and Borrelia hermsii in Escherichia coli revealed differences in DNA repair and recombination phenotypes.

After unsuccessful attempts to recover a viable RecA-deficient mutant of the Lyme borreliosis agent Borrelia burgdorferi, we characterized the functional activities of RecA of B. burgdorferi, as well as RecA of the relapsing fever spirochete Borrelia hermsii and the free-living spirochete Leptospira biflexa, in a recA mutant of Escherichia coli. As a control, E. coli RecA was expressed from the same plasmid vector. DNA damage repair activity was assessed after exposure of the transgenic cells to UV light or the radiomimetic chemicals methyl methanesulfonate and mitomycin C. Recombination activity in the cells was assessed by using an assay for homologous recombination between repeats in the chromosome and by measuring the ability of the cells to foster lytic growth by red gam mutant bacteriophage lambda. Overall, we found that transgenic cells with recA genes of B. burgdorferi, B. hermsii, and L. biflexa had approximately equivalent activities in promoting homologous recombination in the lacZ duplication assay, but cells with B. burgdorferi recA and, most notably, B. hermsii recA were significantly less capable than cells with L. biflexa recA or E. coli recA in responding to DNA damage or in facilitating plaque formation in the phage assay. The comparatively poor function of Borrelia recA in the latter set of assays may be the consequence of impaired coordination in the loading of the transgenic RecA by RecBCD and/or RecFOR in E. coli.

Amino Acid Sequence↗

PubMed2004

Typing of Borrelia relapsing fever group strains.

Partial sequencing of the 16S-23S rDNA intergenic spacer showed two to four genotypes each for Borrelia hermsii and B. turicatae, both relapsing fever agents transmitted by argasid ticks, and for B. miyamotoi and B. lonestari, transmitted by ixodid ticks. Field surveys of Ixodes ticks in Connecticut and Sweden showed limited local diversity for B. miyamotoi.

Animals↗

PubMed2003

Immunization with the Chlamydia trachomatis major outer membrane protein, using the outer surface protein A of Borrelia burgdorferi as an adjuvant, can induce protection against a chlamydial genital challenge.

Two strains of mice C3H/HeN (H-2(k)) and BALB/c (H-2(d)) were immunized with the Chlamydia trachomatis mouse pneumonitis (MoPn) major outer membrane protein (MOMP) using the Borrelia burgdorferi outer surface protein A (OspA) as an adjuvant. As a control, groups of mice were inoculated with ovalbumin instead of MOMP. Female mice were immunized using three different routes: intramuscular (i.m.) plus subcutaneous (s.c.), intranasal (i.n.) and perivaginal and perisacral (p.vag.+p.sac.). Significant humoral and cell mediated immune responses developed particularly in mice inoculated by the i.m.+s.c. routes as determined by the levels of chlamydial specific antibody in the serum and genital secretions and a T-cell proliferative assay. Following immunization the animals were challenged in the genital tract with C. trachomatis MoPn and the course of the infection followed by vaginal cultures. Significant protection against infection was achieved in the C3H/HeN mice inoculated i.m.+s.c. with MOMP+OspA, as shown by the intensity and duration of vaginal cultures, and by the number of mice with positive cultures. On the other hand in BALB/c mice there was only a decrease in the number of animals with positive vaginal cultures. Six weeks after the challenge the mice were mated and the outcome of the pregnancy evaluated. In both the C3H/HeN and the BALB/c mice immunized i.m.+s.c. with MOMP+OspA there was significant protection against infertility as shown by the number of animals with bilateral fertility and number of embryos per uterine horn. In conclusion, immunization using C. trachomatis MOMP, and B. burgdorferi OspA as an adjuvant, can induce significant protection against a chlamydial genital challenge.

Adjuvants, Immunologic↗

PubMed2002

Laboratory testing for suspected Lyme disease.

Laboratory testing for B. burgdorferi infection is intended to substantiate a physician's clinical judgment of whether a patient has Lyme disease or not. Cultivation of B. burgdorferi from a patient's skin or blood is the gold standard for demonstration of active infection, but it is expensive and lacks clinical sensitivity. Detection of spirochetal DNA in clinical samples by PCR has better sensitivity, but PCR for B. burgdorferi has not yet been standardized for more routine diagnostic testing. Detection of antibodies to B. burgdorferi is the most practical and common approach for laboratory work-up of a case of suspected Lyme disease. Serologic assays fall short of 100% sensitivity and specificity, however, and examination of a single specimen in time does not discriminate between previous and ongoing infection. Because of a background false positivity even among healthy populations of nonendemic regions, serologic testing is recommended only when there is at least a one in five chance, in the physician's estimation, that the patient has active Lyme disease. The pretest likelihood of the disease is determined by the physician in the context of epidemiologic and clinical facts of the case. This estimate can serve to reassure patients who are at low risk of B. burgdorferi infection but are seeking a Lyme test for complaints of a more nonspecific nature. Although new subunit serologic assays based on recombinant proteins are becoming available commercially, the longstanding two-test approach, in which a positive or indeterminate result with a standardized, sensitive ELISA test is followed by verification with a more specific Western blot assay, still provides the physician with a reasonably accurate and reliable assessment of the presence of antibodies to B. burgdorferi. More recent challenges for serologic testing are seropositivity in the population as the result of immunization with the Lyme disease vaccine and the emergence of new Borrelia species that cause Lyme disease-like illnesses.

Antibodies, Bacterial↗

PubMed2002

Three major Lyme Borrelia genospecies (Borrelia burgdorferi sensu stricto, B. afzelii and B. garinii) identified by PCR in cerebrospinal fluid from patients with neuroborreliosis in Sweden.

The Lyme Borrelia genospecies Borrelia afzelii and B. garinii have previously been isolated using a culture method in Swedish patients with Lyme borreliosis (LB). There are reports suggesting that the genospecies distribution in human tissue specimens as determined by molecular methods is different from that obtained by culture. In the present study, we developed a nested PCR for detection of Lyme Borrelia-specific DNA in cerebrospinal fluid from Swedish patients with LB. The genospecies were subsequently identified by sequence analysis in a total of 7 PCR-positive patients. Two sequences were identified as B. burgdorferi sensu stricto (s. s.), 1 as B. afzelii and 4 as B. garinii. These are the first reported cases in which B. burgdorferi s. s. has been shown to be the causative agent of human LB in Sweden. The results of our study confirm that the use of direct molecular analytical methods for Borrelia genospecies identification in clinical specimens can provide epidemiological information additional to that obtained by culture.

Adolescent↗

PubMed2002

Fatal spirochetosis due to a relapsing fever-like Borrelia sp. in a northern spotted owl.

Acute septicemic spirochetosis was diagnosed in an adult male northern spotted owl (Strix occidentalis caurina) found dead in Kittitas County, Washington, USA. Gross necropsy findings included marked enlargement of the liver and spleen and serofibrinous deposits on the serous membranes lining the body cavities and the pericardial and perihepatic sacs. Microscopic observations included macrophage infiltration in the liver and spleen with mild thrombosis and multifocal necrosis, as well as hemorrhage and acute inflammation in the choroid plexus of the brain. No viruses or pathogenic bacteria were isolated from brain, liver, or spleen, and no parasites were found in blood smears or impression smears of the liver. Chlamydial culture attempts were unsuccessful and no chlamydial antibodies were detected in serum. In silver-stained microscopic sections and by transmission electron microscopy of liver, numerous long, thin, spiral-shaped bacteria were seen in the liver, spleen, cerebral ventricles, and within blood vessels in many organs. The organism was identified as a member of the Borrelia genus by sequence analysis of the PCR-amplified 16S rRNA gene. The most closely related species is B. hermsii, an agent of relapsing fever in humans in the western United States. This is the first report of a relapsing fever-related Borrelia in a wild bird.

Animals↗