Mark R. Hall

Influence of omega-3 fatty acids on the growth of human colon carcinoma in nude mice

The present study investigated the influence of dietary omega-3 fatty acid supplementation on the growth of human colon carcinoma xenograft in athymic nude mice. Four diets were fed to evaluate the effect of levels and types of fat on colon tumor growth. Animals were maintained on a standard diet modified by addition of fats containing omega-3 and omega-6 fatty acids to represent high and low fat intakes for 53 days. The final mean estimated tumor weight for the high fat corn oil (24%) fed group was 2,302 mg, whereas the low fat (8% corn oil) group was 1,681 mg. The final mean tumor weight of the high fat menhaden oil fed group was 782 mg representing a 66% decrease in growth compared to the high fat corn oil group and a decrease of 54% compared to the low corn oil fed group. The high fat golden algae oil fed group resulted in a mean final tumor weight of 223 mg representing a 90% inhibition of tumor growth relative to the high fat corn oil fed group and 87% inhibition of growth compared to the low fat corn oil fed group. These findings indicate that dietary omega-3 fatty acids possess significant tumor suppressing properties and that the primary tumor suppressing fatty acid is docosahexaenoic acid. Histopathologic examination of control and treated tumors and expression array analyses (human cytokine and apoptosis arrays) support the tumor growth inhibition data and provide evidence for discussion of possible mechanisms for the observed growth inhibition.

Investigations of the incidence of bovine trichomoniasis in Nevada and of the efficacy of immunizing cattle with vaccines containing Tritrichomonas foetus.

Trichomonas cultures taken from 2389 bulls showed that approximately 4.7% of them were infected. Correlation of these data with the ranches from which diagnostic samples were obtained indicated that in the period of 1984 through 1987 26.7 to 44.1% of ranches had at least one infected bull. Thirty-four 18-month-old Holstein heifers were assigned to one of three groups, controls n = 12 animals, soluble vaccine n = 11 animals, and whole vaccine n = 11 animals to determine the effect of Tritrichomonas foetus vaccines on the reproductive performance of T . foetus infected animals. Heifers were bred with T . foetus infected bulls beginning two weeks after the second T . foetus vaccination. All immunized animals developed antibody titers of at least 1:1000 following vaccination. In addition, all control and immunized animals became infected with T . foetus . However, the duration of infection was approximately two weeks shorter in immunized animals. Approximately 42% (5 of 12) of control heifers remained infected with T . foetus for the duration of the experiment, while only 18% (2 of 11) of each of the vaccine groups remained infected for the duration of the experiment. Finally, 27% (3 of 11) of heifers in each of the vaccine groups were pregnant at slaughter, while none of the control heifers were pregnant at slaughter. Therefore, both vaccine formulations appeared to protect heifers (P<0.05) from fetal loss due to trichomoniasis.

Confirmation of the association of human papillomavirus with human colon cancer

The human papillomavirus (HPV) has been shown to be associated with neoplasms of the human colon using immunohistochemistry and in situ hybridization. We now report our use of the polymerase chain reaction and Southern blotting to investigate that same association. We selected 38 carcinomas, 21 adenomas, and 24 normal mucosal samples for the current study. Tissue sections were prepared, and then DNA was extracted and subjected to 40 cycles of amplification using Thermus aquaticus DNA polymerase and a set of degenerate primers. Amplified products were analyzed by agarose gel electrophoresis and Southern blotting. The L1 region of the HPV genome was identified in 13 of 38 carcinomas (32%), 8 of 21 adenomas (38%), and 2 of 24 normal biopsy specimens (8%). These observations validate our previous results and confirm the presence of HPV in human colon mucosa and tumors of that mucosa.

Molecular identification of a novel deltaproteobacterium as the etiologic agent of epizootic bovine abortion (foothill abortion)

ABSTRACT Epizootic bovine abortion (EBA) is endemic in Californias coastal range and the foothill regions of the Sierra Nevada, where it has been the primary diagnosed cause of abortion in beef cattle for >50 years. Investigation of these losses has defined a specific fetal syndrome characterized by late-term abortion or birth of weak or dead calves. Although the unusual clinical presentation and unique fetal pathology associated with EBA have been recognized since the 1950s, the identity of the etiologic agent is unknown. In this study, suppression-hybridization PCR was used to identify a fragment of the 16S rRNA gene of a previously undescribed bacterium in thymus tissue derived from affected fetuses. Phylogenetic analysis revealed that this pathogen was a deltaproteobacterium closely related to members of the order Myxococcales. A specific PCR was subsequently developed to detect the presence of this bacterium in DNA extracted from fetal thymuses. Using histopathology as the definitive diagnosis for EBA, this PCR demonstrated 100% specificity and 88% sensitivity. The bacterium was also detected in the argasid tick Ornithodoros coriaceus, which is the recognized vector of EBA. These data imply a close association between this novel agent and the etiology of EBA.

Experimental transmission of epizootic bovine abortion (foothill abortion)

Advances in defining the biology of epizootic bovine abortion (EBA), including identification of the etiologic agent, have been hampered by the inability to reproduce the disease with confidence. Experimental reproduction of EBA, by feeding the tick vector Ornithodoros coriaceus on susceptible pregnant heifers, is not reliable. The primary objectives of this study were to identify specific tissue(s) obtained from EBA-infected fetuses that could transmit the disease, and then utilize such an infectious challenge system to better define the pathogen, host immunity and geographic distribution of the agent. Described here is the ability to routinely reproduce EBA following inoculation of cryopreserved suspensions of homogenized thymus into susceptible pregnant heifers. This challenge system permitted experiments demonstrating the agent was non-filterable, inactivated upon sonication and susceptible to antibiotics. These findings suggest a prokaryotic microbe and represent a major advance in EBA research. Additional experiments demonstrated that inoculation of the cryopreserved EBA-infectious tissue into heifers, prior to breeding, conferred immunity. Furthermore, such immunized heifers were resistant to challenge with heterologous sources of infectious tissue, suggesting monovalent vaccine development might be feasible. Lastly, challenge studies employing animals from Central Nevada, an area considered free of EBA, demonstrated partial immunity, suggesting the pathogen, and possibly the disease, enjoy a broader distribution than previously thought.

Growth and enrichment medium for detection and isolation of Shiga toxin-producing Escherichia coli in cattle feces.

Detection methods of Shiga toxin-producing Escherichia coli (STEC) in cattle feces varied in using enrichment media containing different antibiotic combinations. To examine efficacy of a new detection method for STEC, three O157:H7 (ATCC 43889, 43890, and 43895) and 41 non-O157:H7 (members of the O1, O15, O26, O86, O103, O111, O125, O127, O128, O136, O146, O153, O158, O165, O166, and O169 serogroups) isolates were tested. These isolates were grown in tryptic soy broth for 6 h, and their concentrations were determined before inoculation of tubes containing 1 g of cattle feces (sterile [experiment 1; evaluating growth] and fresh [experiment 2; evaluating enrichment]) to simulate the high and low levels of STEC shedding by cattle (10(5) versus 102 CFU/g feces, respectively). Eight STEC isolates (the three O157:H7 and five non-O157:H7 selected at random) were tested at a very low level (10 CFU/g feces). The feces were incubated in 50 ml of brain heart infusion broth containing potassium tellurite, novobiocin, and vancomycin (2.5, 20, and 40 mg/liter, respectively) and cefixime (50 microg/liter) at 37 degrees C for 12 h and tested for STEC (VTEC [verotoxin-producing E. coli]-Screen assay [agglutination immunoassay]). Potential STEC isolates were recovered, characterized biochemically, serotyped, and tested for toxin production using Vero (African green monkey kidney) cell toxicity assay and agglutination immunoassay. In both experiments, all the STEC isolates used for fecal inoculation were recovered at the concentrations tested. Our medium supported growth of and enrichment for a wide range of STEC isolates.

Verotoxin-Producing Escherichia coli in Culled Beef Cows Grazing Rangeland Forages

The objective of this study was to assess prevalence of verotoxin-producing Escherichia coli (VTEC) in culled beef cows at the time of shipping to slaughter. Feces were collected from 82 cows on eight Nevada ranches during fall and winter (from September to January) after grazing rangeland forages. A random sample (n = 154) of potential VTEC isolates were tested for verotoxicity and were screened for the presence (polymerase chain reaction) and expression (VTEC-reversed passive latex agglutination assay) of the toxin genes (i.e., VT1 and VT2). Seventeen isolates from four ranches were VTEC. Of these, four had the VT1 gene, five had the VT2 gene, seven had both genes, and one did not have either gene despite its toxicity to Vero cells. Except for one isolate (i.e., untypeable that reacted with VT1-latex beads without having VT1 gene), the genotype and phenotype data of the VTEC isolates matched. Another isolate (08:H– [nonmotile]) was verotoxic, but neither had nor expressed the toxin genes. Of the 17 isolates, four (from one cow) were O157:H7, 11 (from five cows on three ranches) were non-O157:H7 (two O8:H–, three O105:H–, three O116:H–, and three O141:H–), and two were untypeable. Because some of these VTEC serotypes (i.e., O8:H–, O141:H–, and O157:H7) are known to cause human illnesses, it is beneficial to identify VTEC-positive cows before slaughter. This is a critical step in any pre-or post-harvest strategy to minimize the risk of beef contamination with such pathogens.

Diagnosis of epizootic bovine abortion in Nevada and identification of the vector

In the 43 years since the first description in California, epizootic bovine abortion (EBA) has been considered but not definitively diagnosed as a cause of late-term abortions on Nevada ranches. Examination of aborted full-term bovine fetuses obtained from Nevada ranches revealed gross abnormalities consistent with EBA (enlarged lymph nodes, petechial hemorrhages of the oral mucosa and conjunctiva, ascites, and splenohepatomegaly), and EBA was confirmed by histologic examination of fetal tissues. The histologic thymic changes were characteristic of EBA and included severe histocytic thymusitis with depletion of thymocytes, interlobular hemorrhage, and fibrinocellular exudation. The gross enlargement of lymph nodes was the result of cortical follicular hyperplasia and histiocytic lymphadenitis. In addition, widespread, predominately nonsuppurative histologic lesions typical of EBA were observed in most organs, including the brain, lung, heart, liver, and spleen. Furthermore, the presence of Ornithodorus coriaceus, the argasid tick vector of EBA, was established by tick collection using CO2 traps. The tick was identified on ranches and in geographic areas (northern and northwestern counties of Nevada) coincident with diagnosis of multiple cases of EBA. This study establishes the presence of EBA as a cause of late-term abortion in Nevada. Additionally, identification of the EBA tick vector, O. coriaceus, in the same areas as the abortions provides strong evidence that the disease is endemic.

From Ill-defined Problems to Informed Decisions

Decision makers such as military leaders and security analysts are increasingly being asked to make decisions on ill-defined problems. These problems may contain uncertain or incomplete data, and are often complex to piece together. Consequently, decision makers rely heavily on intuition, knowledge and experience. We argue for rich narratives that encapsulate both explicit data and implicit knowledge, supported by three levels of provenance: data, analytical and reasoning. Our hypotheses is that visual analytics tools and methods can help to provide a valuable means to make sense of these complex data, and to help make this tacit knowledge explicit, to support the construction and presentation of the decision.

Occurrence of verotoxin-producing Escherichia coli in dairy heifers grazing an irrigated pasture

Verotoxin-producing Escherichia coli (VTEC) produce one or two toxins known as VT1 and VT2. These toxins have been associated with several human illnesses. Dairy cattle harboring VTEC represent a potential health hazard because they enter the food chain as ground beef. The objective of this study was to assess the occurrence of VTEC in dairy heifers. A total of 91 fecal samples were rectally collected during four periods (spring, summer, fall, and winter of 1999) from 23 heifers. A random sample (n=530) of potential VTEC isolates were tested for verotoxicity and were screened by the polymerase chain reaction (PCR) for presence or absence of VT1 and/or VT2 genes. Thirteen isolates from two heifers (from the winter collection) were verotoxic and were confirmed as E. coli. VTEC were only detected during winter with an occurrence rate of 9.5%. Using PCR, five isolates had the VT1 gene while the remaining eight had the VT2 gene. The sequence and expression of VT1 and VT2 genes were confirmed. No E. coli O157:H7 was detected, but serotyping revealed that the five VT1-positive isolates were O26:NM (a non-motile strain of O26). The remaining eight isolates were untypeable. Identification of VTEC-positive cattle before slaughter is a critical step in any on-farm strategy to minimize the risk of beef contamination with such pathogens.