Acquired resistance to Borrelia burgdorferi infection in the rabbit. Comparison between outer surface protein A vaccine- and infection-derived immunity.

Intradermal inoculation of the rabbit with Borrelia burgdorferi, sensu lato, results in the consistent development of erythema migrans (EM), dermal infection, and visceral dissemination of the spirochete. Within 5 mo, EM as well as dermal and visceral infection are cleared and the animals exhibit immunity to reinfection. This study compares infection-derived immunity with acquired resistance resulting from the administration of a lipidated recombinant outer surface protein A (OspA) vaccine presently undergoing human trial. 4 of 11 OspA vaccinated rabbits, challenged intradermally at each of 10 sites with 10(5) low passage B. burgdorferi, developed EM as well as dermal and disseminated infection. After identical challenge, 2 of the 11 infection-immune rabbits developed a dermal infection, but not EM or disseminated infection. Further, ELISA anti-OspA titers did not correlate with the status of immunity for either OspA vaccinated or infection-immune rabbits. Prechallenge ELISA anti-OspA titers were relatively low in the infection-immune group. This study demonstrates that a state of partial immunity to experimental Lyme disease may result that could potentially mask infection. Further, our data strongly suggest that immunogen(s) other than OspA is/are responsible for stimulating acquired resistance in the infection-immune rabbit.

Virulent strain associated outer membrane proteins of Borrelia burgdorferi.

We have isolated and purified outer membrane vesicles (OMV) from Borrelia burgdorferi strain B31 based on methods developed for isolation of Treponema pallidum OMV. Purified OMV exhibited distinct porin activities with conductances of 0.6 and 12.6 nano-Siemen and had no detectable beta-NADH oxidase activity indicating their outer membrane origin and their lack of inner membrane contamination, respectively. Hydrophobic proteins were identified by phase partitioning with Triton X-114. Most of these hydrophobic membrane proteins were not acylated, suggesting that they are outer membrane-spanning proteins. Identification of palmitate-labeled lipoproteins revealed that several were enriched in the OMV, several were enriched in the protoplasmic cylinder inner membrane fraction, and others were found exclusively associated with the inner membrane. The protein composition of OMV changed significantly with successive in vitro cultivation of strain B31. Using antiserum with specificity for virulent strain B31, we identified OMV antigens on the surface of the spirochete and identified proteins whose presence in OMV could be correlated with virulence and protective immunity in the rabbit Lyme disease model. These virulent strain associated outer membrane-spanning proteins may provide new insight into the pathogenesis of Lyme disease.

Rabbit model of Lyme borreliosis: erythema migrans, infection-derived immunity, and identification of Borrelia burgdorferi proteins associated with virulence and protective immunity.

Erythema migrans (EM), persistent skin infection, and visceral dissemination can be induced reproducibly in the adult male New Zealand White rabbit by intradermal injection of as few as 10(3) Borrelia burgdorferi. EM was found to persist for 7 +/- 3 d. Skin culture positivity (infection) cleared within a mean of 6.7 +/- 1.4 wk after infection and similarly visceral infection was not demonstrated after 8 wk; infection-derived immunity to intradermal challenge was evident 5 mo after initial infection. The extent of the protection against EM and dermal infection induced by untreated infection was directly related to the extent of prior in vitro passage of the B31 strain. Initial infection with as few as 4 x 10(3) B31 passage 4 induced complete protection against EM and skin infection upon subsequent challenge with 4 x 10(7) B31, passage 4. Initial infection with B31 passage 27 led to partial protection against EM along with complete protection against skin infection. Initial infection with passage 47 led to partial protection against EM, but conferred no protection against skin infection. Using serum from rabbits fully immune to reinfection, we defined a set of B. burgdorferi proteins present in virulent B31, but absent in the avirulent American Type Culture Collection B31 strain, termed "va" for virulent strain associated. The va proteins of B31 passages 1, 27, and 47 differed strikingly, thus raising the possibility that these changes may relate in a causal way to the differences in induction of protective immunity observed.

Acid-adaptation does not increase the resistance of Listeria monocytogenes to irradiation in a seafood salad.

Stress adaptation of microbial cells enables the cells to survive better when they are subsequently exposed to other types of stresses. In the food industry, pathogens are commonly stressed during food processing and this is a concern where pathogens such as Listeria monocytogenes are involved. Research was conducted to determine if acid adaptation of L. monocytogenes provides resistance to ionizing irradiation. Three different strains of L. monocytogenes were acid-adapted using three different acids (acetic, citric, lactic) in Tryptic Soy Broth, at a pH 5.5 for 1 h, 4 h, or continuous acid exposure. The acid-adapted L. monocytogenes were then exposed to a low level of gamma irradiation (0.59-0.72 kGy) along with a non-acid adapted L. monocytogenes control. In a test tube study, the 1-h acetic acid-adapted L. monocytogenes strains showed the greatest difference from the control, a reduced kill of 1.1 log CFU/g but this difference was not significant by ANOVA (p=0.054). The reduction achieved after 4 h and continuous acid exposure also did not significantly differ from the control. To determine whether acid adaptation affected radiation resistance within a food product, a refrigerated storage shelf-life study was completed. Acetic acid was used to acid adapt a three-strain cocktail of L. monocytogenes for a period of 1 h. The organisms were then inoculated into a seafood salad (pH 5.15) and subsequently exposed to low dose gamma irradiation (0.7 to 4.5 kGy). L. monocytogenes was reduced or eliminated by irradiation regardless of acid adaptation; no increased resistance was observed.

Effect of gamma irradiation on Listeria monocytogenes in frozen, artificially contaminated sandwiches.

Gamma irradiation has been shown to effectively control L monocytogenes in uncooked meats but has not been extensively studied in ready-to-eat foods. The presence of Listeria in ready-to-eat foods is often due to postprocess contamination by organisms in the food-manufacturing environment. Because gamma irradiation is applied after products are packaged, the treated foods are protected from environmental recontamination. Currently, a petition to allow gamma irradiation of ready-to-eat foods is under review by the Food and Drug Administration. This study was conducted to determine if gamma irradiation could be used to control L. monocytogenes in ready-to-eat sandwiches. Ham and cheese sandwiches were contaminated with L. monocytogenes, frozen at -40 degrees C, and exposed to gamma irradiation. Following irradiation, sandwiches were assayed for L. monocytogenes. A triangle test was performed to determine if irradiated and nonirradiated sandwiches differed in sensory quality. We found that the D10-values ranged from 0.71 to 0.81 kGy and that a 5-log reduction would require irradiation with 3.5 to 4.0 kGy. The results of a 39-day storage study of sandwiches inoculated with 10(7) CFU of L monocytogenes per g indicated that counts for nonirradiated sandwiches remained fairly constant. Counts for sandwiches treated with 3.9 kGy decreased by 5 log units initially and then decreased further during storage at 4 degrees C. Sensory panelists could distinguish between irradiated and nonirradiated sandwiches but were divided on whether irradiation adversely affected sandwich quality. Our results suggest that manufacturers of ready-to-eat foods could use gamma irradiation to control L. monocytogenes and improve the safety of their products.

The phenomena of care in Sicilian-Canadian culture: an ethnonursing case study.

This study examined the caring beliefs and behaviors of first- and second-generation Sicilians who immigrated to Canada and now reside in an Ontario metropolitan area. Madeleine Leininger's ethnonursing model of "Cultural Care Diversity and Universality" (1978, 1985, 1988) provided the framework; qualitative information was gained from the perceptions and cognitions of two Sicilian-Canadian families. Emphasis was placed on cultural and social structure dimensions that depicted caring, health beliefs, and related practices. The research questions guiding this inquiry were: a) What care values, care beliefs, and behavior patterns are deemed to promote/preserve well-being in this culture? b) Who provides care in this culture and under what circumstances? Implications for nursing that emerged included: provision of continuity of care, the importance of the involvement of the family, and the development of innovative means to enhance self-care practices to prevent illness and to reduce aversion to the professional healthcare system.

An exercise test to evaluate fitness for wheelchair activity.

The purpose of this study was to develop a wheelchair ergometer (WERG) test to evaluate fitness for manual wheelchair activity. Thirty able-bodied females participated in a progressive intensity, discontinuous test where exercise bouts were 4 min in duration interspersed with 5-min rest periods. Physiological responses of oxygen uptake (VO2), respiratory exchange ratio (R), net mechanical efficiency (ME), pulmonary ventilation (VO) and heart rate (HR) were determined during the final minute of exercise at power output (PO) levels of 30, 60, 90, 120 and 150 kpm/min. These responses were generally found to be linearly related to PO, however, net ME initially increased with PO and plateaued at approximately 11 per cent at 90 kpm/min. Criteria for fitness evaluation were based upon: (1) magnitude of physiological responses at each PO level; and (2) the maximal PO level completed.

Exercise program for wheelchair activity.

Poor physical fitness of wheelchair-dependent individuals may result in excessive cardiorespiratory responses during locomotion. The purpose of this study was to develop and implement an interval training program (ITP) incorporating wheelchair ergometer (WERG) exercise to improve fitness for wheelchair activity. Of thirteen able-bodied female volunteers, seven were selected to participate in a 5-week ITP, while the remainder served as sedentary controls. Both the exercise training (ET) group and the sedentary control (SC) group completed a standardized fitness test on the WERG before (pre-test) and after (post-test) the 5-week period. Following the ITP, submaximal heart rate, pulmonary ventilation and oxygen uptake responses of the ET group were generally found to be significantly lower during the post-test. These improvements in WERG exercise performance were not observed in the SC group. The ITP may have contributed to adaptations of upper body muscles, improved cardiorespiratory function, and/or a higher level of skill for wheelchair propulsion. We conclude that applying the concepts of interval training to wheelchair exercise may substantially improve the performance and fitness characteristics of wheelchair users. This could reduce the relative stresses of wheelchair locomotion and lead to a higher level of rehabilitation.

Decadal and shorter period variability of surf zone water quality at Huntington Beach, California.

The concentration of fecal indicator bacteria in the surf zone at Huntington Beach, CA, varies over time scales that span at least 7 orders of magnitude, from minutes to decades. Sources of this variability include historical changes in the treatment and disposal of wastewater and dry weather runoff, El Niño events, seasonal variations in rainfall, spring-neap tidal cycles, sunlight-induced mortality of bacteria, and nearshore mixing. On average, total coliform concentrations have decreased over the past 43 years, although point sources of shoreline contamination (storm drains, river outlets, and submarine outfalls) continue to cause transiently poor water quality. These transient point sources typically persist for 5-8 yr and are modulated by the phase of the moon, reflecting the influence of tides on the sourcing and transport of pollutants in the coastal ocean. Indicator bacteria are very sensitive to sunlight therefore, the time of day when samples are collected can influence the outcome of water quality testing. These results demonstrate that coastal water quality is forced by a complex combination of local and external processes and raise questions about the efficacy of existing marine bathing water monitoring and reporting programs.

Cloning and molecular characterization of plasmid-encoded antigens of Borrelia burgdorferi.

Thirteen independent clones that encode Borrelia burgdorferi antigens utilizing antiserum from infection-immune rabbits were identified. The serum was adsorbed against noninfectious B. burgdorferi B31 to enrich for antibodies directed against either infection-associated antigens of B. burgdorferi B31 or proteins preferentially expressed during mammalian infection. The adsorption efficiency of the immune rabbit serum (IRS) was assessed by Western immunoblot analysis with protein lysates derived from infectious and noninfectious B. burgdorferi B31. The adsorbed IRS was used to screen a B. burgdorferi expression library to identify immunoreactive phage clones. Clones were then expressed in Escherichia coli and subsequently analyzed by Western blotting to determine the molecular mass of the recombinant B. burgdorferi antigens. Southern blot analysis of the 13 clones indicated that 10 contained sequences unique to infectious B. burgdorferi. Nucleotide sequence analysis indicated that the 13 clones were composed of 9 distinct genetic loci and that all of the genes identified were plasmid encoded. Five of the clones carried B. burgdorferi genes previously identified, including those encoding decorin binding proteins A and B (dbpAB), a rev homologue present on the 9-kb circular plasmid (cp9), a rev homologue from the 32-kb circular plasmid (cp32-6), erpM, and erpX. Additionally, four previously uncharacterized loci with no known homologues were identified. One of these unique clones encoded a 451-amino-acid lipoprotein with 21 consecutive, invariant 9-amino-acid repeats near the amino terminus that we have designated VraA (for "virulent strain-associated repetitive antigen A"). Since all the antigens identified are recognized by serum from infection immune rabbits, these antigens represent potential vaccine candidates and, based on the identification of dbpAB in this screen, may also be involved in pathogenic processes operative in Lyme borreliosis.