Pre- and post-synaptic cortical cholinergic deficits are proportional to amyloid plaque presence and density at preclinical stages of Alzheimer's disease.

Amyloid imaging has identified cognitively normal older people with plaques as a group possibly at increased risk for developing Alzheimer's disease-related dementia. It is important to begin to thoroughly characterize this group so that preventative therapies might be tested. Existing cholinotropic agents are a logical choice for preventative therapy as experimental evidence suggests that they are anti-amyloidogenic and clinical trials have shown that they delay progression of mild cognitive impairment to dementia. A detailed understanding of the status of the cortical cholinergic system in preclinical AD is still lacking, however. For more than 30 years, depletion of the cortical cholinergic system has been known to be one of the characteristic features of AD. Reports to date have suggested that some cholinergic markers are altered prior to cognitive impairment while others may show changes only at later stages of dementia. These studies have generally been limited by relatively small sample sizes, long postmortem intervals and insufficient definition of control and AD subjects by the defining histopathology. We, therefore, examined pre- and post-synaptic elements of the cortical cholinergic system in frontal and parietal cortex in 87 deceased subjects, including non-demented elderly with and without amyloid plaques as well as demented persons with neuropathologically confirmed AD. Choline acetyltransferase (ChAT) activity was used as a presynaptic marker while displacement of (3)H-pirenzepine binding by oxotremorine-M in the presence and absence of GppNHp was used to assess postsynaptic M1 receptor coupling. The results indicate that cortical ChAT activity as well as M1 receptor coupling are both significantly decreased in non-demented elderly subjects with amyloid plaques and are more pronounced in subjects with AD and dementia. These findings confirm that cortical cholinergic dysfunction in AD begins at the preclinical stage of disease and suggest that cholinotropic agents currently used for AD treatment are a logical choice for preventative therapy.

Circle of Willis atherosclerosis: association with Alzheimer's disease, neuritic plaques and neurofibrillary tangles.

The role of intracranial atherosclerosis in Alzheimer's disease (AD) has been a subject of debate since the first decade of the last century. The initial "vascular hypothesis" of AD was rejected after a series of mid-twentieth century gross anatomical postmortem studies that showed an inconstant relationship between intracranial atherosclerosis and senile dementia. These early studies did not utilize statistical methods, however, and the investigators did not appear to consider the possibility that intracranial atherosclerosis might have a probabilistic, rather than an absolute, effect on AD risk. Recent studies by three independent groups have found a significant statistical association between postmortem measures of circle of Willis atherosclerosis and AD. The present study was undertaken to further address the validity of this association in a large autopsy series, including cases diagnosed neuropathologically with vascular dementia (VaD) and non-AD dementias. Postmortem gross anatomical grading of circle of Willis atherosclerosis was performed in 397 subjects classified by neuropathological diagnosis, including 92 non-demented elderly controls, 215 with AD, 30 with VaD and 60 with non-AD dementias. Circle of Willis atherosclerosis was more severe in subjects with AD and VaD than in control subjects, while it was equivalent between control subjects and subjects with non-AD dementias. Increasing atherosclerotic grade increased the odds ratios (OR) for the diagnoses of both AD and VaD and also increased the ORs for both increased neuritic plaque density and higher Braak neurofibrillary tangle stage. The significance of these associations was retained after consideration of the effects of age, gender and the apolipoprotein E-epsilon4 allele. The results suggest that the statistical association between intracranial atherosclerosis and AD is not an artifact of diagnostic misclassification or of unequal distribution of the apolipoprotein E-epsilon4 allele.

16S rRNA-based analysis of microbiota from the cecum of broiler chickens.

The microbiota of the intestinal tract of chickens plays an important role in inhibiting the establishment of intestinal pathogens. Earlier culturing and microscopic examinations indicated that only a fraction of the bacteria in the cecum of chickens could be grown in the laboratory. Therefore, a survey of cecal bacteria was done by retrieval of 16S rRNA gene sequences from DNA isolated from the cecal content and the cecal mucosa. The ribosomal gene sequences were amplified with universal primers and cloned or subjected to temporal temperature gradient gel electrophoresis (TTGE). Partial 16S rRNA gene sequences were determined from the clones and from the major bands in TTGE gels. A total of 1,656 partial 16S rRNA gene sequences were obtained and compared to sequences in the GenBank. The comparison indicated that 243 different sequences were present in the samples. Overall, sequences representing 50 phylogenetic groups or subgroups of bacteria were found, but approximately 89% of the sequences represented just four phylogenetic groups (Clostridium leptum, Sporomusa sp., Clostridium coccoides, and enterics). Sequences of members of the Bacteroides group, the Bifidobacterium infantis subgroup, and of Pseudomonas sp. each accounted for less than 2% of the total. Sequences related to those from the Escherichia sp. subgroup and from Lactobacillus, Pseudomonas, and Bifidobacterium spp. were generally between 98 and 100% identical to sequences already deposited in the GenBank. Sequences most closely related to those of the other bacteria were generally 97% or less identical to those in the databases and therefore might be from currently unknown species. TTGE and random cloning indicated that certain phylogenetic subgroups were common to all birds analyzed, but sequence data from random cloning also provided evidence for qualitative and quantitative differences among the cecal microbiota of individual birds reared under very similar conditions.

Concurrent Effects of High Hydrostatic Pressure, Acidity and Heat on the Destruction and Injury of Yeasts.

Saccharomyces cerevisiae ATCC 2373 and Zygosaccharomyces bailii ATCC 36947 were exposed to hydrostatic pressures ranging from 1,500 to 3,000 atmospheres for 10, 20 and 30 min in 0.1 M citrate buffer at pH 3.0, 4.0 and 5.0 at 25 and 45°C. Inactivation of inoculated yeast cultures was achieved in spaghetti sauce with meat at 25°C with 3,000 atmospheres for 10 min and also at 45°C and 2,500 atmospheres for 10 min. Viable counts were determined on potato dextrose agar (PDA) incubated at 30°C for 48 h. Pressure-induced injury was demonstrated by plate count differential between PDA and PDA supplemented with glucose (PDAG). A reduction of 7-log10 cycles colony forming units (CFU)/ml was seen for both strains at 3,000 atmospheres for 10 min at 25°C at all pH levels and at 2,250 atmospheres, pH 5.0 for 20 min at 45°C. At 2,000 atmospheres, pH 3.0 for 30 min, the increase in temperature from 25 to 45°C increased the inactivation of yeast by 6-log10 cycles. Lowering the pH from 5.0 to 3.0 enhanced lethality up to 2-log10 cycles at 2,250 atmospheres, 25°C for 30 min. Injury was most apparent at exposure parameters that produced 3- to 5-log10 cycle reductions on PDA. This was achieved (99% injury) at 2,250 atmospheres, 25°C for 30 min. These data indicate that mild heat and acidity contribute to the effectiveness of the inactivation and injury of yeast by high hydrostatic pressure (HHP).

Bacterial Contamination of Broiler Chickens before Scalding.

The bacterial load on the feathers (breast, thigh, and drum), skin (breast, thigh, and drum), and feet of a total of 40 broiler chickens from four different production units was characterized after the chickens were exsanguinated but before the carcasses were scalded. Each broiler carcass contained a mean of aerobic bacteria at or near 8log10/g, a level at which the carcasses would be considered spoiled. Escherichia coli counts ranged from 6.3 to 8.0 log10/g, with the highest presence on the feathers and breast skin and lower levels on the thigh and drum skin, and feet. Contamination of the carcasses followed a similar pattern for Salmonella spp., which ranged between 5.8 and 7.2 log10/g, and Campylobacter jejuni/coli which ranged between 6.1 and 7.5 log10/g. The incidence of potential pathogens was higher on the feathers, breast skin, and feet than on the thigh and drum skin. The incidence of E. coli ranged from 42.5 to 100%, Salmonella spp. ranged from 27.5 to 75%, and C. jejuni/coli ranged from 45 to 82.5%. The mean counts of microorganisms on broilers from the four different grow-out farms were significantly different (P > 0.05) from one another immediately after killing, although the magnitude of the differences was small in most cases. The means ranged from 7.3 to 8.0 log10/g for total colony-forming units (CFU), 6.7 to 7.6 log10/g for E. coli , 5.4 to 6.9 for Salmonella spp., and 5.7 to 7.9 for C. jejuni/coli . All birds were contaminated with E. coli , 60 to 100% with Salmonella spp., and 80 to 100% with C. jejuni/coli , depending on grow-out farm. C. jejuni/coli counts and incidences were higher than those of Salmonella spp., and usually lower than those of E. coli . This research demonstrates that broilers entering the processing plant are highly contaminated. Although processing seems to decrease the number and incidence of microorganisms on the carcasses, additional modifications in production, transportation, and processing are warranted to reduce the microbial population on the birds before they are slaughtered.