Comparison of fluorinated polymers against stainless steel, glass and polypropylene in microbial biofilm adherence and removal.

Biofilm formation is a long-standing problem in ultrapure water and bioprocess fluid transport lines. The standard materials used in these applications (316L stainless steel, polypropylene and glass) have long been known to be good surfaces for the attachment of bacteria and other biological materials. To compare the relative tenacity of biofilms grown on materials used in manufacturing processes, a model system for biofilm attachment was constructed that approximates the conditions in industrial process systems. New fluorinated polymers were compared to the above materials by evaluating the surface area coverage of bacterial populations on materials before and after mild chemical treatment. In addition, contact angle studies compared the relative hydrophobicity of surfaces to suspensions of bacteria in growth media, and scanning electron microscopy and atomic force microscopy studies were used to characterize surface smoothness and surface defects. Biofilm adherence to polymer-based substrata was determined to be a function of both surface finish and surface chemistry. Specifically, materials that are less chemically reactive, as indicated by higher contact angle, can have rougher surface finishes and still be amenable to biofilm removal.

Immobilization of bacteria and Saccharomyces cerevisiae in poly(tetrafluoroethylene) membranes.

A novel method for immobilization of bacteria and Saccharomyces cerevisiae cells is described. Microorganisms may be entrapped in a matrix of poly(tetrafluoroethylene) (PTFE) fibrils. Cells are blended with an aqueous emulsion of PTFE stabilized with Triton X-100 surfactant to form a thick paste. The paste is calendered biaxially in a standard rubber mill. This process causes fibrillation of the PTFE and formation of the fibril matrix, which serves only to impart physical integrity to the composite microporous membrane. The cells trapped in the membrane were shown to be viable by incubation of the membrane on solid media and in broth culture. This bioactive membrane represents a new means of immobilization of cells for bioprocessing.

Detection of antigens in urine of mice and humans infected with Borrelia burgdorferi, etiologic agent of Lyme disease.

Lyme disease is a seasonal tick-borne malady which has worldwide distribution. Early and accurate diagnosis of Lyme disease is essential for successful antibiotic therapy. Symptoms are too vague to make an early diagnosis based on conventional criteria. We report the detection of antigens of Borrelia burgdorferi, causative agent of Lyme disease, in the urine of infected mice and humans. This technique may eventually provide a rapid diagnostic test for the early and accurate detection of this illness.

Characterization of a circular plasmid from Borrelia burgdorferi, etiologic agent of Lyme disease.

Borrelia burgdorferi, the etiologic agent of Lyme disease, was recently shown to contain plasmid DNA. Two plasmid species have been described in strain CT1, a Wisconsin tick isolate: a 9.2-kilobase entity; and a larger, 70-kilobase entity. Characterization of the 9.2-kilobase entity by using DNase I and restriction endonucleases demonstrated that the plasmid is supercoiled and exists as a stable dimer in this strain. The role played by the plasmid in B. burgdorferi is unknown.

New infectious spirochete isolated from short-tailed shrews and white-footed mice.

A spirochete with two periplasmic flagella was isolated from the blood or tissues of spleens and kidneys from short-tailed shrews (Blarina brevicauda) and white-footed mice (Peromyscus leucopus) in Connecticut and Minnesota. After inoculation, the shrew-mouse spirochete infected Swiss mice and Syrian hamsters. This spirochete is morphologically and serologically distinct from the species of Treponema, Borrelia, Leptospira, and Spirochaeta examined.

Prevalence of Borrelia burgdorferi and Babesia microti in mice on islands inhabited by white-tailed deer.

Borrelia burgdorferi and Babesia microti were isolated from 35 of 51 white-footed mice (Peromyscus leucopus) and meadow voles (Microtus pennsylvanicus) captured on two Narragansett Bay, R.I., islands inhabited by deer, the principal host for the adult stages of the vector tick, Ixodes dammini. Immature ticks parasitized mice from both islands. From 105 mice captured on four other islands not inhabited by deer neither pathogen was isolated, nor were I. dammini found.

Genetic analysis of Borrelia.

DNA homology studies were performed on Borrelia hermsii, B. duttoni, B. crocidurae and the O. coriaceus spirochete. These organisms had a genus level relationship with each other. B. hermsii was 17 to 53% homologous with B. duttoni, B. crocidurae, and the O. coriaceus spirochetes; B. crocidurae was 17-32% related to B. hermsii, B. anserina and the O. coriaceus spirochete, and the O. coriaceus spirochete was 28-50% related to B. hermsii, B. crocidurae and B. anserina. The O. coriaceus spirochete was 10% homologous with B. burgdorferi, the etiologic agent of Lyme disease. Plasmid DNA was detected in these strains of Borrelia. Southern blot analysis of plasmid DNA from these strains suggested little or no homology with plasmid DNA from B. hermsii used as a probe. A plasmid of 9.2 kilobases was seen in B. burgdorferi strain CT1. This plasmid was shown to be a tandem repeat with unit length of 4.6 kilobases. Sequences corresponding to this plasmid were detected in several fresh tick isolates from North America, and were not seen in any animal, human or European isolates surveyed. The role in the biology of the spirochete played by this plasmid is unknown.

Culturing Borrelia burgdorferi from spleen and kidney tissues of wild-caught white-footed mice, Peromyscus leucopus.

Borrelia burgdorferi was isolated most frequently from tissue of spleen (n = 13) and kidney (n = 10) and less often from blood (n = 5) of wild-caught Peromyscus leucopus. Prevalence of infection tended to be highest at sites where Lyme disease was most common (e.g., 5 of 6 mice were positive in East Haddam, Connecticut). Spirochetes were not isolated in Danbury or New Hartford, areas where this malady is rare. However, in Fairfield, where the disease is also uncommon, 4 of 9 mice were infected. Larval and nymphal I. dammini, containing borreliae, parasitized P. leucopus at all sites where B. burgdorferi was cultured from mice. Borreliae were also detected in D. variabilis feeding on hosts at two of the sites. P. leucopus appears to be an excellent animal to identify focal areas of B. burgdorferi.

Involvement of birds in the epidemiology of the Lyme disease agent Borrelia burgdorferi.

Borrelia burgdorferi, the causative agent of Lyme disease, was isolated from the liver of a passerine bird, Catharus fuscescens (veery), and from larval Ixodes dammini (tick) feeding on Pheucticus ludovicianus (rose-breasted grosbeak) and Geothlypis trichas (common yellowthroat). In indirect immunofluorescence antibody tests, isolates reacted with polyclonal and monoclonal (H5332) antibodies. Studies on the DNA composition of the veery liver isolate and the strain cultured from an I. dammini larva indicated that both were B. burgdorferi and not Borrelia anserina or Borrelia hermsii. The veery liver isolate infected hamsters and a chick. In contrast, B. anserina infected chicks but not hamsters. B. burgdorferi is unique among Borrelia spp. in being infectious to both mammals and birds. We suggest that the cosmopolitan distribution of B. burgdorferi may be caused by long-distance dispersal of infected birds that serve as hosts for ticks.

Peromyscus leucopus and Microtus pennsylvanicus simultaneously infected with Borrelia burgdorferi and Babesia microti.

Borrelia burgdorferi, the etiologic agent of Lyme disease, and Babesia microti, the causative agent of human babesiosis, were isolated from 71 and 57%, respectively, of 14 specimens of Peromyscus leucopus and Microtus pennsylvanicus collected from Prudence and Patience Islands, R.I. Both pathogens were isolated from five individual rodents. The presence of these two infectious organisms in the same mammal suggests that individual larval Ixodes dammini may ingest both pathogens and subsequently transmit them in the nymphal stage.

Identification of endemic foci of Lyme disease: isolation of Borrelia burgdorferi from feral rodents and ticks (Dermacentor variabilis).

Borrelia burgdorferi, the etiological agent of Lyme disease, was isolated from the blood, kidneys, spleens, eyes, or livers of white-footed mice (Peromyscus leucopus) (n = 19 of 22) and from the blood, kidneys, or spleens of eastern chipmunks (Tamias striatus) (n = 2 of 2) captured at three foci for Lyme disease in eastern Connecticut. These bacteria were cultured most frequently from spleens (n = 19) and kidneys (n = 15). B. burgdorferi persisted in one mouse for at least 60 days. One spirochetemic mouse had infected Ixodes dammini and Dermacentor variabilis larvae attached, suggesting that these ticks may have acquired spirochetes from the host. Spirochetes isolated from P. leucopus, T. striatus, and D. variabilis larvae were serologically and genetically indistinguishable from reference B. burgdorferi isolates. We conclude that isolation of spirochetes from feral rodents is a method for identifying endemic areas of Lyme disease.

Genetic relationship of lyme disease spirochetes to Borrelia, Treponema, and Leptospira spp.

Genetic studies were performed on the following spirochetes: three Lyme disease spirochetes isolated from Ixodes ticks and from human spinal fluid; three species of North American borreliae; four species of Treponema; and two species of Leptospira. The mol% G+C values for Lyme disease spirochetes were 27.3 to 30.5%, similar to values of 28.0 to 30.5% for Borrelia species but different from the values of Leptospira or Treponema species which ranged from 35.3 to 53%. Lyme disease spirochetes represent a new species of Borrelia, with DNA homologies of 31 to 59% with the three North American strains of Borrelia studied. These studies also showed that Lyme disease spirochetes from three sources constituted a single species, with DNA homologies ranging from 76 to 100%. A high degree of relatedness was also seen between the three North American borreliae, with homology varying from 77 to 95%, indicating that these spirochetes represent a single species. Lyme disease spirochetes and Borrelia species exhibited almost no homology with Leptospira and Treponema species (0 to 2%). Plasmids were detected in the three Lyme disease spirochetes and in the three North American borreliae.

Taxonomy of the Lyme disease spirochetes.

Morphology, physiology, and DNA nucleotide composition of Lyme disease spirochetes, Borrelia, Treponema, and Leptospira were compared. Morphologically, Lyme disease spirochetes resemble Borrelia. They lack cytoplasmic tubules present in Treponema, and have more than one periplasmic flagellum per cell end and lack the tight coiling which are characteristic of Leptospira. Lyme disease spirochetes are also similar to Borrelia in being microaerophilic, catalase-negative bacteria. They utilize carbohydrates such as glucose as their major carbon and energy sources and produce lactic acid. Long-chain fatty acids are not degraded but are incorporated unaltered into cellular lipids. The diamino amino acid present in the peptidoglycan is ornithine. The mole % guanine plus cytosine values for Lyme disease spirochete DNA were 27.3-30.5 percent. These values are similar to the 28.0-30.5 percent for the Borrelia but differed from the values of 35.3-53 percent for Treponema and Leptospira. DNA reannealing studies demonstrated that Lyme disease spirochetes represent a new species of Borrelia, exhibiting a 31-59 percent DNA homology with the three species of North American borreliae. In addition, these studies showed that the three Lyme disease spirochetes comprise a single species with DNA homologies ranging from 76-100 percent. The three North American borreliae also constitute a single species, displaying DNA homologies of 75-95 percent. Lyme disease spirochetes and Borrelia exhibited little or no DNA homology (0-2 percent) with the Treponema or Leptospira. Plasmids were present in the three Lyme disease spirochetes and the three North American borreliae.