Nutritional Value of Moss for Arctic Ruminants: a Test with Muskoxen

Abstract: Although moss is commonly found in the feces of arctic herbivores, we do not know the digestible value of this forage for ruminants. We compared grass hay (Bromus sp.) with moss (Hylocomium splendens, Tomenthypnum nitens) from 2 locations in Alaska, USA: Cape Krusenstern National Monument and Fairbanks. We evaluated forages by digestion in ruminally fistulated muskoxen (Ovibos moschatus) by suspending forages in polyester bags before and after the rumen was acclimated with moss for 15 consecutive days. Ruminal degradation was not affected by acclimation to moss. Hay lost dry matter during 48 hours of ruminal incubation (-49%), whereas moss gained dry matter (+44-57%). Incubated moss gained nitrogen (+435-680%), as well as fiber (+18%), and one moss gained ash (+121%). Mass gained by moss in the rumen was probably due to the combined effect of microbial colonization and adsorption of fibrous particles onto the sponge-like matrix. We evaluated postruminal degradation of forages by incubation in acid-pepsin. Ruminally incubated mosses lost little nitrogen in acid-pepsin even though ruminally incubated hay lost 23% nitrogen on acid digestion. Consumption of moss during winter may be a net cost of selecting plants within moss communities when lichens and graminoids are scarce. Moss in feces may, therefore, indicate low availability of favored foods for muskoxen and other arctic ruminants that are confined to small winter ranges. Increasing concentrations of moss in the feces and, thus, the diet of muskoxen may alert wildlife managers to shifts in winter range quality or forage access due to changing snow conditions.     

Detection of Helicobacter pylori DNA in feces and saliva by polymerase chain reaction: a review

The polymerase chain reaction (PCR), known for its high sensitivity and specificity, has been used for the detection of Helicobacter pylori DNA in bodily materials such as feces and saliva. Since fecal specimens contain PCR inhibitors, DNA before PCR amplification has been purified using various biochemical, immunological and physical pre-PCR steps. Several PCR protocols, differing from each other in the selection of genomic targets and primers, have produced varying degrees of specificity and sensitivity in detecting H. pylori DNA. PCR identified antimicrobial resistance of H. pylori in feces. It also detected virulence factor genes such as the cytotoxin-associated gene (cagA) and vacuolating cytotoxin gene (vacA) in feces and saliva. While the cagA gene was detected in 50-60% of fecal specimens, it was found in 25% of salivary specimens from patients. There was considerable variation in the detection rate of H. pylori DNA in salivary samples. The detection rate in saliva with the most effective primer pair was lower than that observed in feces, making saliva a less suitable specimen for the diagnosis of H. pylori infection. There is controversy regarding the permanent presence of H. pylori in saliva. Whether the salivary and gastric specimens of an individual harbor identical or different strains has not been resolved. PCR cannot distinguish between living and dead organisms. However, it can offer quick results on fecal and salivary specimens, which may contain fastidious and slow-growing H. pylori in low numbers.     

Influence of season on the incidence of DNA hypodiploidy in urinary cytology

OBJECTIVE: This study was conducted to determine if an incidence of hypodiploidy in urinary specimens is related to seasonal temperature changes. MATERIALS AND METHODS: DNA ploidy was evaluated on 10,846 urinary specimens fixed in buffered alcohol (MOPSO/NaCl + ETOH) and received over a one year period from numerous sites throughout the United States. The percentage of hypodiploid (DNA index < 0.8) cases was evaluated in each month. As a control, DNA ploidy results from 3, 755 prostate biopsies, fixed in 10% neutral buffered formalin, received during the winter and summer months of the same year, were evaluated. RESULTS: The average percentage of hypodiploidy in cytologic specimens during the summer months was 19.6% compared to 5. 4% in the winter and early spring months (range: 20.6-4.8%). The average percentage of hypodiploid cells in histologic specimens was 0.8% for both the summer and winter months (range: 1.73-0.36%). CONCLUSIONS: The rate of hypodiploidy in urinary cytology seems to be temperature related. The hypodiploidy rate of histologic specimens fixed in formalin shows no fluctuation with the seasons.     

Development of Bacteroides 16S rRNA gene TaqMan-based real-time PCR assays for estimation of total, human, and bovine fecal pollution in water

Bacteroides species are promising indicators for differentiating livestock and human fecal contamination in water because of their high concentration in feces and potential host specificity. In this study, a real-time PCR assay was designed to target Bacteroides species (AllBac) present in human, cattle, and equine feces. Direct PCR amplification (without DNA extraction) using the AllBac assay was tested on feces diluted in water. Fecal concentrations and threshold cycle were linearly correlated, indicating that the AllBac assay can be used to estimate the total amount of fecal contamination in water. Real-time PCR assays were also designed for bovine-associated (BoBac) and human-associated (HuBac) Bacteroides 16S rRNA genes. Assay specificities were tested using human, bovine, swine, canine, and equine fecal samples. The BoBac assay was specific for bovine fecal samples (100% true-positive identification; 0% false-positive identification). The HuBac assay had a 100% true-positive identification, but it also had a 32% false-positive rate with potential for cross-amplification with swine feces. The assays were tested using creek water samples from three different watersheds. Creek water did not inhibit PCR, and results from the AllBac assay were correlated with those from Escherichia coli concentrations (r2= 0.85). The percentage of feces attributable to bovine and human sources was determined for each sample by comparing the values obtained from the BoBac and HuBac assays with that from the AllBac assay. These results suggest that real-time PCR assays without DNA extraction can be used to quantify fecal concentrations and provide preliminary fecal source identification in watersheds.     

黄喉貂日活动节律及食性的季节变化

2016年4月至2017年4月,采用红外相机技术和粪便分析法(频率法和剩余物干重法)对四川栗子坪国家级自然保护区内黄喉貂不同季节的日活动节律及食物组成变化进行了研究。结果显示:(1)黄喉貂主要在白天活动(昼间独立照片数占总独立照片数的85. 64%),不同季节黄喉貂日活动节律无显著差异(χ~2=126. 950,df=132,P=0. 608),但在不同季节其日活动高峰出现时间不同,春季的活动高峰在16:00~19:00(31. 65%),夏季活动高峰在15:00~18:00 (26. 32%),秋季活动高峰在13:00~16:00 (34. 31%),冬季活动高峰在11:00~14:00 (25. 00%),并且冬季夜间活动与其他季节相比明显增多;(2)黄喉貂取食食物有兽类、鸟类、昆虫类和植物类等,但兽类是黄喉貂最主要的食物来源,在一年中以兽类的出现频率最高,为95. 28%,兽类剩余物的总相对干重百分比达80. 99%,其次是植物、鸟类和昆虫;(3)黄喉貂对食物类别的利用表现出明显的季节差异,春、夏、秋三个季节黄喉貂粪便中兽类所占比重最多,春季鸟类出现频率较高,冬季黄喉貂粪便中植物所占比重明显增多。本研究表明,黄喉貂在不同季节其日活动节律和食性均表现出一定的差异,这可能与其繁殖特性和生理代谢需求有关。本研究揭示了黄喉貂的日活动节律及食性的季节性变化,充实了黄喉貂的生物、生态学资料,也为该物种的保护管理提供了参考资料。     

Development of Bacteroides 16S rRNA Gene TaqMan-Based Real-Time PCR Assays for Estimation of Total, Human, and Bovine Fecal Pollution in Water

Bacteroides species are promising indicators for differentiating livestock and human fecal contamination in water because of their high concentration in feces and potential host specificity. In this study, a real-time PCR assay was designed to target Bacteroides species (AllBac) present in human, cattle, and equine feces. Direct PCR amplification (without DNA extraction) using the AllBac assay was tested on feces diluted in water. Fecal concentrations and threshold cycle were linearly correlated, indicating that the AllBac assay can be used to estimate the total amount of fecal contamination in water. Real-time PCR assays were also designed for bovine-associated (BoBac) and human-associated (HuBac) Bacteroides 16S rRNA genes. Assay specificities were tested using human, bovine, swine, canine, and equine fecal samples. The BoBac assay was specific for bovine fecal samples (100% true-positive identification; 0% false-positive identification). The HuBac assay had a 100% true-positive identification, but it also had a 32% false-positive rate with potential for cross-amplification with swine feces. The assays were tested using creek water samples from three different watersheds. Creek water did not inhibit PCR, and results from the AllBac assay were correlated with those from Escherichia coli concentrations (r² = 0.85). The percentage of feces attributable to bovine and human sources was determined for each sample by comparing the values obtained from the BoBac and HuBac assays with that from the AllBac assay. These results suggest that real-time PCR assays without DNA extraction can be used to quantify fecal concentrations and provide preliminary fecal source identification in watersheds.     

Use of PCR for direct detection of Campylobacter species in bovine feces

This study reports on the use of PCR to directly detect and distinguish Campylobacter species in bovine feces without enrichment. Inhibitors present in feces are a major obstacle to using PCR to detect microorganisms. The QIAamp DNA stool minikit was found to be an efficacious extraction method, as determined by the positive amplification of internal control DNA added to bovine feces before extraction. With nested or seminested multiplex PCR, Campylobacter coli, C. fetus, C. hyointestinalis, and C. jejuni were detected in all fecal samples inoculated at approximately 10(4) CFU g(-1), and 50 to 83% of the samples inoculated at approximately 10(3) CFU g(-1) were positive. At approximately 10(2) CFU g(-1), C. fetus, C. hyointestinalis, and C. jejuni (17 to 50% of the samples) but not C. coli were detected by PCR. From uninoculated bovine feces, a total of 198 arbitrarily selected isolates of Campylobacter were recovered on four commonly used isolation media incubated at three temperatures. The most frequently isolated taxa were C. jejuni (152 isolates) and C. lanienae (42 isolates), but isolates of C. fetus subsp. fetus, Arcobacter butzleri, and A. skirrowii also were recovered (相似文献   

Feather deuterium measurements reveal origins of migratory western loggerhead shrikes (Lanius ludovicianus excubitorides) wintering in Mexico

Understanding the winter distributions of migrant birds is important because productivity and recruitment are influenced by conditions at several locations and periods in the life cycle of individuals. The western loggerhead shrike, Lanius ludovicianus excubitorides , is a threatened species in Canada, and its decline is attributed to potential limitations on the wintering grounds. We examined patterns of stable-hydrogen isotope (δD) distributions in feathers of loggerhead shrikes, primarily of L. l. excubitorides , during winter at three regions in north and central Mexico, to establish relative abundance and origins of migrants. We also investigated potential movements of Mexican winter resident individuals. Using shrike museum specimens of known summer provenance, a shrike deuterium base map for Mexico was developed from isotopic measurement of feathers of resident shrikes and use of a recently established feather base map for raptors in North America. Stable hydrogen isotope analyses of inner secondary feather (s9) of all loggerhead shrikes examined in Mexico during winter indicated that north-central (Region A), north-eastern (Region B) and south-central (Region C) sites in Mexico consisted of 28.1%, 73.7% and 63.8% of migrant individuals from northern breeding grounds, respectively. Isotopic evidence suggested movements of a few local residents birds (7.9%) into the Chihuahuan desert from south-western USA and north-eastern Mexico to winter.     

Genotyping of Escherichia coli from environmental and animal samples

The triplex PCR of Clermont et al. [Clermont, O., Bonacorsi, S., Bingen, E., 2000. Rapid and simple determination of the Escherichia coli phylogenetic groups. Appl. Environ. Microbiol. 66, 4555-4558.] was used to genotype E. coli isolates from the Mid-Atlantic region of the USA, obtained from freshwater, animal internal organs, and feces. Of 445 isolates subjected to genotyping, 118 isolates (26%) were genotype A, 111 (25%) genotype D, 140 (31%) genotype B1, and 76 (17%) genotype B2. All four genotypes were present in three sets of freshwater stream samples. When isolates from chicken cecal ingesta, cecal mucosa, and tracheal mucosa were screened, there was selective distribution of genotypes in these organs. Genotype D was rarely encountered in feces, milk, and intestinal tissues of dairy cows, while all four genotypes were represented in goose feces. Isolates from the feces of zoo animals reared in the US demonstrated a predominance of genotype B1. Thirty-six of the A isolates in our overall collection were subgenotype A(0), in which none of the three amplicons are observed; confirmation that these isolates were E. coli was done using an ancillary lacZ PCR assay. We conclude that the genotyping triplex PCR assay, used in combination with traditional culture methods, can be useful in categorizing E. coli from environmental and veterinary sources in the Mid-Atlantic region of the USA.     

Ingested foreign (phage M13) DNA survives transiently in the gastrointestinal tract and enters the bloodstream of mice

Is the epithelial lining of the mammalian gastrointestinal (GI) tract a tight barrier against the uptake of ingested foreign DNA or can such foreign DNA penetrate into the organism? We approached this question by pipette-feeding circular or linearized double-stranded phage M13 DNA to mice or by adding M13 DNA to the food of mice whose fecal excretions had previously been shown to be devoid of this DNA. At various post-prandial times, the feces of the animals was tested for M 13 DNA sequences by Southern or dot blot hybridization or by the polymerase chain reaction (PCR). On Southern blot hybridization, the majority of M13 DNA fragments were found in the size range between < 200 and 400 by (base pairs). For the PCR analysis, synthetic oligodeoxyribonucleotide primers were spaced on the M13 DNA molecule such that the sizes of the persisting M13 DNA fragments could be determined. We also extracted DNA from whole blood or from sedimented blood cells of the animals at different times after feeding M t3 DNA and examined these DNA preparations for the presence of M13 DNA by dot blot hybridization or by PCR. M13 DNA fragments were found between 1 and 7 h postprandially in the feces of mice. By PCR analysis, fragments of 712, 976, and 1692 by in length were detected. In DNA from blood, M13 DNA fragments of up to 472 by were found by PCR between 2 and 6 h after feeding. Dot blot or Southern blot hybridization revealed M13 DNA at 2 and 4 h, but not at 1, 8 or 24 h after feeding. This DNA was shown to be DNase sensitive. M13 DNA was found both in blood cells and in the serum. A segment of about 400 by of the DNA amplified by PCR from feces or blood was analyzed for its nucleotide sequence which was found to be identical to that of authentic M13 DNA, except for a few deviations. M13 DNA could not be detected in the feces or in the blood of the animals prior to feeding or prior to 1 h and later than 7 h after feeding. These controls attest to the validity of the results and also argue against the possibility that the murine GI tract had been colonized by phage M13. Moreover, M13 DNA-positive bacterial colonies were never isolated from the feces of animals that had ingested M13 DNA. The results of reconstitution experiments suggested that 2 to 4% of the orally administered M13 DNA could be detected in the GI tract of mice. A proportion of about 0.01% to 0.1% of the M13 DNA fed could be retrieved from the blood.     

Botanical and chemical composition of rumen contents of Sika deer on Mt Goyo,northern Japan

Botanical and chemical compositions of the rumen contents of 58 Sika deer on Mt Goyo, northern Japan, collected from summer of 1988 to spring of 1989, were analyzed.Sasa nipponica, a dwarf bamboo, was important in summer (35.0%) and winter (61.4%), but it decreased to 5.6% and was replaced by browse leaves in fall, and to 28.0% and was replaced by dead leaves, twigs and bark of woody plants in winter. Crude protein was 20–25% lower in the washed fraction than in the gross fraction. It was highest (16.2%) in summer and lowest (8.6%) in winter. High protein content in summer and fall foods was attributed to forb and browse leaves. Seasonal fluctuation in protein content in the foods of these Sika deer was greater than red deer on Rhum, Scotland and smaller than wapiti in Yellowstone National Park, Wyoming, USA. Climatically, winter severity of Mt Goyo is intermediate between the two localities, which seems to explain the seasonal fluctuation of the protein level. Crude fiber wasca 33% in the ‘washed’ fraction, and did not change seasonally.     

Molecular analyses of novel methanotrophic communities in forest soil that oxidize atmospheric methane

Forest and other upland soils are important sinks for atmospheric CH(4), consuming 20 to 60 Tg of CH(4) per year. Consumption of atmospheric CH(4) by soil is a microbiological process. However, little is known about the methanotrophic bacterial community in forest soils. We measured vertical profiles of atmospheric CH(4) oxidation rates in a German forest soil and characterized the methanotrophic populations by PCR and denaturing gradient gel electrophoresis (DGGE) with primer sets targeting the pmoA gene, coding for the alpha subunit of the particulate methane monooxygenase, and the small-subunit rRNA gene (SSU rDNA) of all life. The forest soil was a sink for atmospheric CH(4) in situ and in vitro at all times. In winter, atmospheric CH(4) was oxidized in a well-defined subsurface soil layer (6 to 14 cm deep), whereas in summer, the complete soil core was active (0 cm to 26 cm deep). The content of total extractable DNA was about 10-fold higher in summer than in winter. It decreased with soil depth (0 to 28 cm deep) from about 40 to 1 microg DNA per g (dry weight) of soil. The PCR product concentration of SSU rDNA of all life was constant both in winter and in summer. However, the PCR product concentration of pmoA changed with depth and season. pmoA was detected only in soil layers with active CH(4) oxidation, i.e., 6 to 16 cm deep in winter and throughout the soil core in summer. The same methanotrophic populations were present in winter and summer. Layers with high CH(4) consumption rates also exhibited more bands of pmoA in DGGE, indicating that high CH(4) oxidation activity was positively correlated with the number of methanotrophic populations present. The pmoA sequences derived from excised DGGE bands were only distantly related to those of known methanotrophs, indicating the existence of unknown methanotrophs involved in atmospheric CH(4) consumption.     

Survey of an otter<Emphasis Type="Italic">Lutra lutra</Emphasis> population in Southern Italy: site occupancy and influence of sampling season on species detection

In order to assess the current status of the otterLutra lutra Linnaeus, 1758 in Southern Campania (Italy), we surveyed 141 sites for spraints (faeces) in 2001. Fifty-eight sites were surveyed during the winter and in the following summer in order to test, through an estimation-based approach, the influence of sampling season on species detection. Site occupation in the study area was high (69.5%) and possibly underestimated, because the survey was affected by non-detection errors. Our analyses showed that winter surveying markedly underestimated true otter occupancy at the 58 sites (51.7% vs 97.1%), whereas summer surveying was very reliable (91.4%). Rains and floods may have removed spraints during winter, thereby reducing the detection probability to 0.534. These results suggest that otter standard surveys in areas with Mediterranean-type climates should be conducted during summer or periods without prolonged precipitation. Comparing our results with those of the 1985 National survey, we found an occupancy increase from 65.8 to 100%. We could not establish whether this change reflected a population increase or was due to possible non-detection errors that occurred in the 1985 survey. However, the present occupancy substantiates the strategic relevance of the study area for planning the conservation of Italian otters.     

Foraging dynamics of muskoxen in Peary Land, northern Greenland

Muskoxen Ovibos moschatus in northern Greenland (79-83°N) are at the northern limit of their distribution and exist under seasonal extremes dominated by nearly 10 months of winter, much of which is without sunlight. The period of summer vegetative growth is less than two months. In the Kap København area (82°30'N), diversity of plant species is low (76 species of vascular plants) and forage biomass in major vegetation types in summer varies from over 40 g m^-2 in sedge-dominated fens to ≤5 g m^-2 in polar barrens. Nonetheless, 90-95% of the ice-free area consists of barren ground or sparcely vegetated polar desert. During summer, muskoxen apparently foraged opportunistically to maximize intake, with sedges the major food item in fens while willows were the major dietary component when on willow-dominated slopes. Quality of summer forage was high during its early phenological stages, with 21-28% crude protein and 60-75% in vitro digestibility. Microhistological analysis of winter feces indicated dominance by graminoids. Muskoxen spent > 50% of their daily activity feeding, which fits a cline of increasing feeding time with increasing latitude in summer. Increased feeding times at high latitudes appears to be a function of both reduced forage biomass and need to maximize forage intake during the brief summer period when forage quality is high. Movement rates in summer while foraging were inversely related to available forage biomass. Seasonal activity of muskoxen peaks during the rutting period (July-September) and then declines gradually through early winter to a low in late winter (March-April).     

Comparison of PCR versus Culture for Detection of Mycobacterium bovis after Experimental Inoculation of Various Matrices Held under Environmental Conditions for Extended Periods

The purpose of this study was to compare the performance of a molecular detection technique (nested PCR) with that of mycobacterial culture in the detection of Mycobacterium bovis DNA in a set of 687 samples of experimentally inoculated environmental substrates (hay, soil, corn, water) exposed to natural weather conditions in Michigan. Four replicates of each substrate were used; half were autoclaved for sterilization, all were inoculated with 50,000 CFU of M. bovis isolated from Michigan livestock, and all were placed in outdoor enclosures, with half under shade and the other half exposed to direct sunlight. Samples were tested for the presence of M. bovis during one 12-month period, with monthly sample testing and during three 12-week periods (winter, spring, summer) with weekly sample testing. Samples were subjected to mycobacterial culture for isolation of M. bovis and a nested PCR with two primer sets targeting IS6110 to detect M. bovis DNA. In 128 samples tested during the 12-month period, M. bovis was not detectable by culture after 2 months but M. bovis DNA was detectable by PCR for at least 7 months. Of the 559 samples tested during the 12-week periods, PCR detected M. bovis DNA for up to 88 days in all of the sample types. There were no significant differences in the detection of M. bovis between shade and sun samples or between sterile and unsterilized samples, regardless of the detection method (PCR or culture). For use in epidemiologic investigations, the PCR assay was more rapid than mycobacterial culture, was not hindered by contaminating organisms, and detected M. bovis DNA in environment samples much longer after initial contamination than mycobacterial culture did.     

Diverse Wild Bird Host Range of Mycoplasma gallisepticum in Eastern North America

Emerging infectious diseases often result from pathogens jumping to novel hosts. Identifying possibilities and constraints on host transfer is therefore an important facet of research in disease ecology. Host transfers can be studied for the bacterium Mycoplasma gallisepticum, predominantly a pathogen of poultry until its 1994 appearance and subsequent epidemic spread in a wild songbird, the house finch Haemorhous mexicanus and some other wild birds. We screened a broad range of potential host species for evidence of infection by M. gallisepticum in order to answer 3 questions: (1) is there a host phylogenetic constraint on the likelihood of host infection (house finches compared to other bird species); (2) does opportunity for close proximity (visiting bird feeders) increase the likelihood of a potential host being infected; and (3) is there seasonal variation in opportunity for host jumping (winter resident versus summer resident species). We tested for pathogen exposure both by using PCR to test for the presence of M. gallisepticum DNA and by rapid plate agglutination to test for the presence of antibodies. We examined 1,941 individual birds of 53 species from 19 avian families. In 27 species (15 families) there was evidence for exposure with M. gallisepticum although conjunctivitis was very rare in non-finches. There was no difference in detection rate between summer and winter residents, nor between feeder birds and species that do not come to feeders. Evidence of M. gallisepticum infection was found in all species for which at least 20 individuals had been sampled. Combining the present results with those of previous studies shows that a diverse range of wild bird species may carry or have been exposed to M. gallisepticum in the USA as well as in Europe and Asia.     

Biological activities of organic compounds adsorbed onto ambient air particles: comparison between the cities of Teplice and Prague during the summer and winter seasons 2000-2001

The capital of the Czech Republic, Prague, appears today to be one of the most polluted residential areas in the country, whereas air pollution in the Northern Bohemia region (the former "Black Triangle Region") has substantially decreased during the last decade, especially with respect to the gaseous pollutant SO(2). This study evaluated the biological activities of complex mixtures of organic compounds adsorbed onto ambient air particles (PM10) collected during the summer and winter seasons of 2000-2001 at three monitoring sites--Teplice (TP), Prague-Smíchov (PRG-SM) (city centre) and Prague-Libus (PRG-LB) (suburban area). The following short-term in vitro assays with strikingly different endpoints were used: a bacterial mutagenicity test using the Salmonella typhimurium tester strain TA98 and YG1041, an acellular assay (CT DNA) combined with 32P-postlabelling to evaluate DNA adduct-forming potency and the chick embryotoxicity screening test (CHEST). The results of the mutagenicity test with the YG1041 strain, the acellular genotoxicity (DNA adducts) and the embryotoxicity tests responded to the amount of eight carcinogenic polycyclic aromatic hydrocarbons (PAHs) analysed in the EOM (dichloromethane extractable organic matter) samples tested. Nevertheless, the biological effects of the EOM did not differ between locations. The highest biological activity of the ambient air in terms of organic compounds associated with particles (per unit volume of air) was seen in the Prague city centre during both summer and winter seasons. At this location, B[a]P concentration ranged from 0.1 to 8.9 ng/m(3) (mean 0.3 and 3.6 ng/m(3) for summer and winter seasons, respectively), 13 PAHs ranged from 11 to 343 ng/m(3) (mean 52 and 160 ng/m(3) for summer and winter seasons, respectively). Generally, using in vitro tests, higher ambient air activity was found in the winter season as compared with the summer season at all three monitoring sites (TA98 +S9, approximately 4-fold; YG1041 -S9, approximately 5-fold; YG1041 +S9, approximately 8-fold; CT DNA +S9, approximately 10-fold; CHEST, approximately 10-fold; B[a]P, carcinogenic PAHs and total PAHs analysed, more than 10-fold). The different proportions of individual PAHs found in the summer and winter samples suggested traffic as a major emission source in the summer and, additionally, residential heating in the winter season at all three monitoring sites. The DNA adduct patterns resulting from the in vitro acellular assay also demonstrated similar major emission sources at all three locations. The study shows that particle-bound carcinogenic-PAH concentrations may be taken as an index for the biologically active (mutagenic, genotoxic, embryotoxic) components in air particulate samples. Therefore, high-quality monitoring data of carcinogenic PAHs may be useful for epidemiological studies of the impact of air pollution on the health of the population and for helping decision makers to improve our environment.     

Development of a combined selection and enrichment PCR procedure for Clostridium botulinum Types B, E, and F and its use to determine prevalence in fecal samples from slaughtered pigs

A specific and sensitive combined selection and enrichment PCR procedure was developed for the detection of Clostridium botulinum types B, E, and F in fecal samples from slaughtered pigs. Two enrichment PCR assays, using the DNA polymerase rTth, were constructed. One assay was specific for the type B neurotoxin gene, and the other assay was specific for the type E and F neurotoxin genes. Based on examination of 29 strains of C. botulinum, 16 strains of other Clostridium spp., and 48 non-Clostridium strains, it was concluded that the two PCR assays detect C. botulinum types B, E, and F specifically. Sample preparation prior to the PCR was based on heat treatment of feces homogenate at 70 degrees C for 10 min, enrichment in tryptone-peptone-glucose-yeast extract broth at 30 degrees C for 18 h, and DNA extraction. The detection limits after sample preparation were established as being 10 spores per g of fecal sample for nonproteolytic type B, and 3.0 x 10(3) spores per g of fecal sample for type E and nonproteolytic type F with a detection probability of 95%. Seventy-eight pig fecal samples collected from slaughter houses were analyzed according to the combined selection and enrichment PCR procedure, and 62% were found to be PCR positive with respect to the type B neurotoxin gene. No samples were positive regarding the type E and F neurotoxin genes, indicating a prevalence of less than 1.3%. Thirty-four (71%) of the positive fecal samples had a spore load of less than 4 spores per g. Statistical analysis showed that both rearing conditions (outdoors and indoors) and seasonal variation (summer and winter) had significant effects on the prevalence of C. botulinum type B, whereas the effects of geographical location (southern and central Sweden) were less significant.     

Balancing sample accumulation and DNA degradation rates to optimize noninvasive genetic sampling of sympatric carnivores

Noninvasive genetic sampling, or noninvasive DNA sampling (NDS), can be an effective monitoring approach for elusive, wide‐ranging species at low densities. However, few studies have attempted to maximize sampling efficiency. We present a model for combining sample accumulation and DNA degradation to identify the most efficient (i.e. minimal cost per successful sample) NDS temporal design for capture–recapture analyses. We use scat accumulation and faecal DNA degradation rates for two sympatric carnivores, kit fox (Vulpes macrotis) and coyote (Canis latrans) across two seasons (summer and winter) in Utah, USA, to demonstrate implementation of this approach. We estimated scat accumulation rates by clearing and surveying transects for scats. We evaluated mitochondrial (mtDNA) and nuclear (nDNA) DNA amplification success for faecal DNA samples under natural field conditions for 20 fresh scats/species/season from <1–112 days. Mean accumulation rates were nearly three times greater for coyotes (0.076 scats/km/day) than foxes (0.029 scats/km/day) across seasons. Across species and seasons, mtDNA amplification success was ≥95% through day 21. Fox nDNA amplification success was ≥70% through day 21 across seasons. Coyote nDNA success was ≥70% through day 21 in winter, but declined to <50% by day 7 in summer. We identified a common temporal sampling frame of approximately 14 days that allowed species to be monitored simultaneously, further reducing time, survey effort and costs. Our results suggest that when conducting repeated surveys for capture–recapture analyses, overall cost‐efficiency for NDS may be improved with a temporal design that balances field and laboratory costs along with deposition and degradation rates.