Experimental inoculation of wild turkeys (Meleagris gallopavo) with Mycobacterium bovis.

Although avian species are known to be susceptible to infection with Mycobacterium spp. organisms, much remains unknown about the susceptibility of birds to infection with M. bovis. The objective of this current study was to determine if wild turkeys (Meleagris gallopavo) can be infected with M. bovis when inoculated by the oral or intratracheal route. Six turkeys were orally inoculated and another six were inoculated via the trachea with a high dose of M. bovis, 1 x 10(5) CFU/ml. Six turkeys were sham-inoculated controls. Two turkeys from each treatment group were sacrificed on days 30, 60, and 90 postinoculation. There were no gross or microscopic lesions consistent with mycobacteriosis in the 23 inoculated turkeys over the 90-day duration of this study. Fecal cultures were also consistently negative for M. bovis when sampled before inoculation and on days 1, 30, and 60 postinoculation. Two intratracheally inoculated turkeys were positive for M. bovis in visceral tissues at 30 days postinoculation. However, this finding was only indicative of passive persistence of mycobacteria in the tissues and not of infection, as there were no attendant lesions or clinical compromise to support infection. Thus, it can be concluded that young wild turkeys are resistant to infection with M. bovis and, therefore, pose minimal threat as reservoir or spillover hosts for this organism.

Neocarzinostatin-induced DNA strand scission and subsequent cell cycle traverse in HeLa S3 cells.

HeLa cells were synchronized at the G1-S boundary by double thymidine block and treated for 1 hr with varying concentrations of the antibiotic anticancer protein neocarzinostatin (NCS). Cells were then released from the block and allowed to resume their cycle. Aliquots were removed at various times in order to monitor cell cycle progression and to assess repair of DNA strand breaks. Dose-dependent DNA strand breakage occurred at all concentrations of NCS tested down to 0.05 microgram/ml. Flow cytometry revealed that NCS-treated cells were delayed in entering S phase and that once in S phase their rate of progression through it was retarded significantly. At all concentrations of NCS tested, the majority of cells did not enter G2 by 12 hr. Untreated cells, on the other hand, completed mitosis by this time. NCS-treated cells had little ability to repair DNA breaks. There appeared to be a correlation between the initial number of NCS-induced DNA breaks and the delayed entry into the S phase but little correlation between the lack of strand scission repair and the retarded progress through S phase.

Retinoblastoma protein inhibits IFN-gamma induced apoptosis.

Regulation of apoptosis (programmed cell death) is critical for maintaining tissue homeostasis. Recent studies indicate a tight coupling between cellular proliferation and apoptosis as cell cycle regulators such as Cyclin D, E1A and E7 appear to influence both events. Each of these modulators is able to bind to and inhibit the function of the retinoblastoma tumor suppressor protein (RB). RB functions, in part, by binding to and inactivating E2F transcription factors, preventing expression of E2F-activated genes associated with G1/S cell-cycle progression. Loss of functional RB deregulates E2F activity and, depending on cell type and environmental factors, promotes tumorigenesis or apoptotic death. To determine the effect of RB on IFN-gamma induced apoptosis, we treated RB-defective carcinoma cell lines and their respective RB-constituted sister clones with IFN-gamma and examined the cells for alterations characteristic of apoptosis. We observed that RB-defective cells, but not the RB-reconstituted clones, decreased in size following IFN-gamma treatment. IFN-gamma treatment caused increased cell detachment in the RB-defective lines but did not affect adherence of the RB-reconstituted clones. Assays for DNA fragmentation revealed lower molecular weight DNA and the apoptosis-associated oligo-nucleosomal ladder following IFN-gamma treatment of the RB-defective cells while higher molecular weight DNA was present in the IFN-gamma treated, RB-reconstituted lines. IFN gamma-induced apoptosis in RB-defective cells was enhanced by serum stimulation, which is also characteristic of p53-dependent E2F-1-mediated apoptosis. However, IFN-gamma induced apoptosis in RB-defective lines does not require wild-type p53 suggesting that, upon IFN-gamma induction, deregulated E2F-mediated apoptosis can also proceed via p53-independent pathways.

Assembly of secondary intermediates during deoxyribonucleic acid replication in transformed human fibroblasts.

The elongation of replicative DNA was studied in transformed WI-38 cells (designated 2RA). Shear effects were avoided by use of an alkaline sucrose gradient sedimentation method whereby cells were lysed directly on top of gradients, at 4 degrees C in the dark. The earliest detected intermediate is a short (2 S) piece of DNA which is converted first to a 25 S piece and then to a 100 S piece, within 10 min. The 100 S piece is next converted to a 212 S, and a 370 S, and finally to a chromosomal DNA of about 450 S. This pattern is quite different from that previously reported by us for normal WI-38 cells, where there was a 50 S intermediate which was not quickly converted into a much larger size, but which gradually elongated, by addition of smaller pieces, to a larger size, of 100 S.; another difference was the time required for formation of the 100 S piece, i.e., 75 min (Rawles, J.W., Jr. and Collins, J.M. (1977) J. Biol. Chem. 252, 4762-4766).

Resistance of Mallard ducks (Anas platyrhynchos) to experimental inoculation with Mycobacterium bovis.

The purpose of this study was to investigate whether mallard ducks (Anas platyrhynchos) are susceptible to infection with Mycobacterium bovis by either oral or intratracheal inoculation and to assess their potential role in the spread of bovine tuberculosis. Six ducks were orally inoculated with 1.0 x 10(5) colony-forming units of M. bovis, six ducks were intratracheally inoculated with the same dose, and six ducks served as sham-inoculated controls. The study length was 90 days postinoculation, with samples of two birds from each group necropsied at 30-day intervals. Both fecal and tissue samples were collected for mycobacterial culture. None of the inoculated ducks shed M. bovis in their feces at any culture point (days 1, 30, and 60) during the study. No evidence of illness or weight loss was present during the course of the study, and only one duck had M. bovis isolated from any tissue, although there were no associated microscopic lesions. Mallard ducks were highly resistant to infection with M. bovis following high-dose inoculation and did not shed the organism in their feces. This study was conducted using high-dose inoculation; therefore, it appears that ducks are unlikely to play any significant role in the transmission of M. bovis between infected and uninfected mammalian hosts.

Demonstration of Ac-Arg-Gly-Asp-Ser-NH2 as an antiaggregatory agent in the dog by intracoronary administration.

This study compared the anti-platelet effect of Ac-RGDS-NH2 which is a peptide fragment from fibrinogen to Ac-RGES-NH2 in which the aspartic acid (D) of Ac-RGDS-NH2 has been replaced by glutamic acid (E). When Ac-RGDS-NH2 was infused intracoronary at concentrations of 100-400 mM, acute platelet-dependent thrombus formation in the dog coronary artery was inhibited. However, infusion of Ac-RGES-NH2 intracoronary at similar concentrations to Ac-RGDS-NH2 failed to inhibit platelet-dependent thrombus formation in the dog. Ac-RGDS-NH2 and Ac-RGES-NH2 were also tested for their ability to inhibit collagen-induced platelet aggregation in vitro. Ac-RGDS-NH2 elicited concentration-dependent inhibition of collagen-induced aggregation with no effect of Ac-RGES-NH2 on collagen-induced platelet aggregation. Thus, Ac-RGDS-NH2 is an effective antiplatelet agent after intracoronary administration in the dog and also inhibits collagen-induced platelet aggregation in vitro. Ac-RGDS-NH2 is a specific inhibitor of platelet aggregation as replacement of the aspartic acid in Ac-RGDS-NH2 with glutamic acid results in complete loss of biological activity.

Vitamin D3 reduces the apoptotic effect of IFN-gamma but does not facilitate HLA class II inducibility in RB-defective cells.

The retinoblastoma protein (RB) regulates the cell cycle by binding and inactivating the E2F transcription factors, which prevents transcription of genes required for DNA synthesis. RB has been shown to inhibit IFN-gamma-mediated apoptosis, possibly by regulating premature entry into S phase. RB is also required for high level IFN-gamma induction of HLA class II genes, which encode antigen presenting molecules, but not for other IFN-gamma inducible genes as demonstrated in previous reports describing the analysis of RB-transformants of the RB-defective cell lines, MDA-468-S4 (S4) and H2009. The IFN-gamma response of the HLA class II genes takes much longer than does the response of the other IFN-gamma inducible genes, raising the question of whether RB facilitates HLA class II inducibility by maintaining cell viability over the long time course required for HLA class II induction. Thus, we sought to learn whether IFN-gamma induced apoptosis in an RB-defective cell line could be prevented independently of RB and whether doing so would facilitate HLA class II inducibility in the RB-defective line. Our results indicated that cotreating the RB-defective S4 cells with IFN-gamma and Vitamin D3 decreased the number of cells containing subdiploid DNA compared to cells treated with IFN-gamma alone, suggesting that Vitamin D3 reduced IFN-gamma-mediated apoptosis. S4 cells cotreated with Vitamin D3 and IFN-gamma also had decreased cell detachment, further indicating that Vitamin D3 decreased IFN-gamma induced apoptosis. However, Vitamin D3 cotreatment resulted in no detectable increase in HLA-DR, the most prominent HLA class II molecule, indicating that the effect of RB on HLA class II induction is not exclusively due to its ability to inhibit IFN-gamma induced apoptosis.

Acute lethality of D- and L-ethionine in Swiss mice.

The acute, 7-day LD50 in Swiss mice of D-ethionine administered intraperitoneally, was determined as 185 mg/kg with 95% confidence limits of 163 and 210 mg/kg. L-ethionine, the stereoisomer, which is more potent in inhibiting liver RNA synthesis, was not lethal at 2500 mg/kg. This acute toxicity of D-ethionine was suppressed by L-methionine and adenine. Blood accumulation in the thoracic cavity and pericardium was the probable cause of death.

Comparison of postmortem techniques for the detection of Mycobacterium bovis in white-tailed deer (Odocoileus virginianus).

A retrospective study of various diagnostic postmortem techniques used in a 4-year surveillance program for detection of Mycobacterium bovis infection in wild white-tailed deer (Odocoileus virginianus) was conducted. The tests evaluated were routine histopathology, acid-fast staining, detection of acid-fast bacilli in culture, and an M. tuberculosis group-specific genetic probe applied to pure cultures. Each of these techniques were compared with a reference or "gold standard" of mycobacterial culture and identification. Histopathology, the most rapid form of testing for M. bovis infection in white-tailed deer samples, had a sensitivity of 98% and a specificity of 87%, resulting in a positive predictive value of 94%. The detection of acid-fast bacilli by staining was less sensitive than histopathology (90%), but its higher specificity (97%) resulted in a positive predictive value of 99%. The detection of acid-fast bacilli on culture was both highly specific (93%) and sensitive (100%). The group-specific genetic probe had the highest sensitivity and specificity and produced results in complete agreement with those of mycobacterial culture, suggesting that this technique could be used as the new "gold standard" for this particular wildlife tuberculosis surveillance program.

Experimental inoculation of pigeons (Columba livia) with Mycobacterium bovis.

The purpose of this pilot study was to determine if pigeons (Columba livia) are susceptible to infection with Mycobacterium bovis by either oral or intratracheal inoculation and to assess their possible role in the lateral transmission of bovine tuberculosis. Six pigeons were orally inoculated with 1.3 x 10(5) colony-forming units of M. bovis, six pigeons were intratracheally inoculated with the same dose, and six pigeons served as noninoculated controls. The study continued for 90 days postinoculation (PI), with groups of birds necropsied at 30-day intervals, and fecal samples and tissues were collected for mycobacterial culture. Two pigeons, one intratracheally inoculated and one orally inoculated, shed M. bovis in their feces at 1 day PI, and one intratracheally inoculated bird shed M. bovis in its feces 60 days PI. Whereas no illness or weight loss was present during the course of the study, 2 of 12 inoculated birds exhibited microscopic lesions of mycobacteriosis, and the organism was isolated from tissues of three inoculated birds. Pigeons are susceptible to infection with M. bovis after high dose inoculation and can shed the organism in their feces for up to 60 days PI; intratracheally inoculated birds appear more likely to become active fecal shedders of M. bovis. Although these were high dose inoculations under experimental conditions, pigeons may potentially play a role in the lateral transmission of bovine tuberculosis between infected and uninfected mammalian hosts.

Experimental inoculation of European starlings (Sturnus vulgaris) and American crows (Corvus brachyrhynchos) with Mycobacterium bovis.

The purpose of this series of pilot studies was to determine whether the passerine species studied are susceptible to infection with Mycobacterium bovis. Separate experiments were conducted on wild-caught starlings (Sturnus vulgaris) and American crows (Corvus brachyrhynchos). In each experiment, four birds were challenged intraperitoneally and four were challenged orally with microorganisms. Challenge dose was 1 x 10(5) colony-forming units of M. bovis cultured from a white-tailed deer (Odocoileus virginianus) case in Michigan. Birds were euthanatized at 1 and 2 mo postinoculation. Histologic lesions suggestive of mycobacteriosis, without the presence of acid-fast bacilli, were noted in all experimental groups. Mycobacterial cultures performed on pooled tissue samples were positive for M. bovis in only some of the intraperitoneal inoculates of each species.

The emerging epidemiology of rural AIDS.

The incidence of AIDS in rural areas is increasing rapidly. However, historically it has been overshadowed by AIDS in the epicenters. From 1991-1992 the increased percentage of cases was higher in nonmetropolitan areas than in any other areas of residence. The rate per 100,000 also increased at almost the same rate in rural areas as in the largest metropolitan areas, defined by the Centers for Disease Control (CDC) as having a population of more than 500,000, and in other metropolitan areas of 50,000 to 500,000 population, as designated by CDC. To date, the epidemiology of AIDS in rural areas has not been defined. This information is necessary to developing effective policies and programs. The research presented here reviews the literature on AIDS in specific areas and populations at risk as a basis for generating hypotheses for further study. The first wave of the epidemic, primarily affecting homosexual or bisexual men, is strongly evident in many rural locations. The second wave of the epidemic is strongly evident in the South and can be seen among high-risk groups such as black women, adolescents, migrant and seasonal farm workers, people who abuse alcohol, intravenous drug users, and users of crack cocaine, including those who trade sex for drugs. Poverty is a common characteristic of the second-wave population. Proximity to interstate highways as well as metropolitan areas may also be associated. A national study of the epidemiology of AIDS in rural America is imperative.

Environmental responses in small rural hospitals.

Major environmental forces are placing increased pressure on community small and rural hospitals to provide access to medical care in rural communities and to survive as free standing non-profit community institutions. This survey of chief executive officers of small and rural hospitals in two states examines their perceptions of their institution's environment and the degree to which multi-institutional arrangements are seen as an organizational response to a changing environment. Major differences among small and rural hospitals are explored.

HIV issues for rural hospitals in U.S. frontier areas.

A Survey of 108 hospital administrators in the eight states of the Mountain Census Region was conducted to identify frontier rural hospitals' experiences (fewer than 50 beds) in the provision of care and services to patients with HIV infection; to assess the availability of HIV care and services in these small, remote rural hospitals; and to assess the status of education and policy development related to HIV infection. Of the 62 hospitals that responded, 16 (26%) had provided care and services to HIV-infected patients. Acute inpatient and emergency room care were the services most commonly utilized. An additional 11 hospitals reported the presence of HIV-positive individuals in their medical service areas. Thus, nearly 44 percent of the hospitals were aware of the importance of addressing HIV infection as a local concern. Employees in the hospitals that had experienced caring for HIV-positive persons expressed more concern about acquiring HIV infection than those in hospitals that had not. Four nursing assistants, two registered nurses, and one dietary worker had refused to provide care. HIV education consisted primarily of video programs, presentations by in-house staff, and sending employees away to workshops. Despite this HIV education, most staff remain fearful of caring for HIV infected patients. Major concerns expressed by the hospital administrators were related to enforcing universal precautions, confidentiality, staff response, community acceptance, and cost of care. Only 30 hospitals (48%) had AIDS policies in effect, and these focused primarily on infection control and universal precautions.(ABSTRACT TRUNCATED AT 250 WORDS)

A typological approach to the study of rural HIV service delivery networks.

Despite the rapid growth of AIDS cases in nonmetropolitan areas, little is known about the characteristics and needs of HIV-positive rural residents or how rural areas are responding to the epidemic. This paper proposes a typology for distinguishing among rural environments and examining variations in HIV service networks. The typology identifies three dimensions that have a major effect on the development of rural HIV service networks: degree of rurality, the prevalence of AIDS, and the epidemiological and demographic characteristics of the infected populations. Data from four case studies are used to illustrate how variations in rural environments can affect the organization and delivery of HIV/AIDS care. The typology contributes to public policy discussions by identifying key attributes of rural environments that influence program planning and implementation and the transferability of service delivery models.

Economic impact of hospital closure on small rural counties, 1984 to 1988: demonstration of a comparative analysis approach.

Hospital closure in a rural community may affect the locale's economic prospects as well as the health of its residents. Studies of economic effects have primarily relied on modeling techniques rather than observation of actual change. This study demonstrates the use of a comparative analysis approach for estimating the economic effects of hospital closure on small rural counties. The experiences of 103 small rural counties at which a hospital closed between 1984 and 1988 was compared with a matched group of counties at which no closure took place. "Comparable" counties were selected based on seven scales measuring the similarity between a closure county and potential comparisons. Three scales examined population and economic characteristics in the year before closure; two scales measured change throughout a three-year period preceding closure; and two scales measured change throughout a five-year period preceding closure. Closure effects were measured through a multivariate analysis of the post-closure economic history of closure and comparison counties. The key assumption is that similar counties should have similar experiences over time. If an event occurs within some of these counties but not others, this event should have visible effects. Comparative analysis suggested that earned income in closure counties (excluding farming and mining income) was lower than in comparison counties subsequent to closure and that labor force growth was similarly affected. A comparative analysis approach produces results that parallel those obtained from economic modeling and should be considered for further research.

HIV infection in traditional rural communities.

The challenge to rural nurses to deliver knowledgeable and skilled nursing and health care to individuals with HIV infection and AIDS is indeed tremendous. Isolation of rural communities and health care facilities coupled with limited resources, financial concerns, conservative values of many traditional rural communities, and the tendency to exclude those who do not conform to community norms make it difficult to integrate the individual with HIV disease into the rural health care delivery system fully. Issues of particular concern to the rural nurse include maintenance of client confidentiality, obtaining and maintaining current knowledge and skills necessary to the provision of quality HIV nursing care, management of complex client health care problems, and provision of appropriate support services. Rural nurses must be innovative and creative in developing mechanisms to deal with these concerns. In addition, because rural nurses are well respected by the community and viewed as possessing a great deal of expertise in the delivery of health care, they are well positioned to provide leadership to the community in developing educational and care strategies to more effectively provide HIV care. Indeed, the delivery of high-quality HIV care in rural areas across the United States will likely depend on the expertise and leadership provided by rural nurses.