Fecal cup for collection of feces in male rats

A cup was designed which fits around the tail of male rats. The cup was used to collect feces and made it possible to separate urine and feces completely from each other. The cup was easy to apply, remove, empty and replace, and its use did not cause noticeable stress to the animal.     

血液微生态环境代表性指标检测结果影响因素分析

目的探讨采血容器、检测时间等因素对血液微生态环境代表性指标检测结果的影响。方法采用不同采血容器（采血管和采血杯）分别在5、15、30和60 min采集受检者静脉血1次,经全自动分析仪对血液微生态环境代表性指标白细胞（WBC）、红细胞（RBC）、血红蛋白（HGB）和血小板（PLT）进行检测分析。结果不论是采用采血管还是采血杯,在5、15、30和60 min检测时血液微生态环境代表性指标WBC、RBC、HGB及PLT的结果比较差异无统计学意义（P〉0.05）。结论不论是采用采血管还是采血杯采血,在1 h内检测,都不会对血液微生态环境代表性指标的检测结果造成较大影响。     

Evaluation of the association of parasitism with mortality of wild European rabbits Oryctolagus cuniculus (L.) in southwestern Australia

Abundances of the parasitic nematodes Trichostrongylus retortaeformis and Passalurus ambiguus, and 8 Eimeria species were estimated by fecal egg and oocyst output in 12 discrete free-ranging populations of wild rabbits (Oryctolagus cuniculus) in southwestern Australia. Comparisons of parasite egg and oocyst counts were made between those rabbits known to have survived at least 2 mo after fecal samples were collected and those rabbits that did not survive. There were significant negative relationships between parasite egg and oocyst counts and survival when all age groups and collection periods were pooled for several species of coccidia and for T. retortaeformis. However, when the same comparisons were made within rabbit age groups and within collection periods, there were very few significant differences even where sample sizes were quite large. The differences indicated by the pooled analysis for coccidia were most likely due to an uneven host age distribution with respect to survival, combined with an uneven distribution of the oocyst counts with rabbit age. The result for T. retortaeformis was similarly affected but by a seasonal pattern. Parasitism by nematodes and coccidia did not appear to be an important mortality factor in these rabbit populations, at least at the range of host densities we examined. This suggests that other factors must have been responsible for the observed pattern of density-dependent regulation in these rabbits.     

Predation on artificial nests in a forest dominated landscape – the effects of nest type, patch size and edge structure

We studied the effects of forest patch size and forest edge structure on nest predation in a boreal coniferous forest landscape. The following predictions were tested. Nest predation should be higher in small than in large stands, in edges than in interior areas of forest stands, and in barren forest/clear–cut edges created by forestry than in natural forest/open marsh edges. Four types of artificial above ground nests (total of 261) were used; open cup nests with reindeer Rangifer t. tarandus hair, open cup nests with domestic hen Gallus domesticus feathers, and unlined open cup and nest–box nests. Nests were baited with one Japanese quail Coturnix coturnix japonica egg. Nest–boxes were depredated significantly less than open cup nests of all types. No edge- or stand size–related nest predation was found. The predation rate, regardless of the nest type, did not differ relative to the edge type and vegetation characteristics. However, better horizontal visibility of open cup nests due to more open vegetation structure increased predation risk in man–made edges compared to inherent edges. The results suggest that edge–related nest predation is absent or weak in forest dominated landscapes. This may be due to predator types present in the landscape and/or predators habitat use in forest dominated areas. Therefore, it might be that findings documented in other areas, such as in agricultural dominated landscapes, cannot be directly applied to managed forest landscapes.     

Non-invasive genotyping of transgenic animals using fecal DNA

For genotyping of transgenic animals, many IACUC guidelines recommend the use of fecal DNA when possible because this approach is non-invasive. Existing methods for extracting fecal DNA may be costly or involve the use of toxic organic solvents. Furthermore, feces contain an abundance of PCR inhibitors that may hinder DNA amplification when they are co-purified with fecal DNA. Here the authors describe a cost-effective, non-toxic method for genotyping transgenic animals by using the reagent AquaStool to extract fecal DNA and remove PCR inhibitors. Genotyping results obtained from fecal DNA samples extracted using AquaStool were reliably accurate when compared with results obtained from tail DNA samples. Because it is non-invasive, the authors believe that use of this method for genotyping transgenic animals using fecal DNA samples may improve animal welfare.     

Comparison of different drying and storage methods on quantifiable concentrations of fecal steroids in the cheetah

Fecal steroid analysis is a powerful tool that can provide important information on the health, physiology, and reproductive status of nondomestic species. However, studying free‐ranging animals requires that feces be stored and transported from the collection site to the laboratory in a manner that prevents degradation or alteration of steroid metabolites. To determine the effects of different handling and storage methods on fecal steroids, 30 fresh fecal samples from five captive cheetahs were collected, thoroughly mixed, separated into aliquots, and processed (stored or dried) under different conditions. Concentrations of gonadal and adrenal steroid hormones were analyzed in feces stored frozen at –20°C or at room temperature in 95% ethanol. Both frozen and ethanol‐stored aliquots were desiccated using a lyophilizer, solar oven, or conventional oven. The steroid values from aliquots stored and desiccated using the different methods were compared to those obtained using the optimal storage method of freezing at –20°C and desiccating in a lyophilizer (control). Concentrations of corticoid, estrogen, progestagen, and androgen metabolites in fecal extracts were quantified by radioimmunoassay. Androgen metabolite concentrations were not significantly affected (P > 0.05) by storage or drying methods. Fecal samples stored at room temperature in ethanol and lyophilized also had steroid concentrations that did not differ (P > 0.05) from controls. However, the concentrations of corticoid and estrogen metabolites were significantly lower (P < 0.05), and progestagen metabolites were significantly higher (P < 0.05) in samples desiccated in solar and conventional ovens without regard to storage method. These results suggest that storage of fecal samples at room temperature in ethanol is the best alternative to freezing for subsequent analysis of steroid hormone concentrations. Differences in measured concentrations of hormones in oven‐desiccated samples could be due to hormone degradation or shifts in the immunodominant metabolite. Therefore, validation of storage and processing techniques should be included in the development of any new fecal steroid analysis methodology. Zoo Biol 21:215–222, 2002. © 2002 Wiley‐Liss, Inc.     

Validation of a field-friendly extraction and storage method to monitor fecal steroid metabolites in wild orangutans

Measuring hormone metabolites from feces is the most often used method to assess hormonal status in wildlife. Although immediate freezing of fecal samples collected in the field is the best method to minimize the risk of degradation of hormones over time, this is often not possible in remote field sites. Therefore, alternative storage and preservation methods for fecal samples are required in these conditions. We conducted an experiment to investigate if fecal glucocorticoid (FGCM) and progesterone metabolite (pregnanediol-3-glucuronide; PdG) levels measured from samples that were extracted with a simple, field-friendly methodology correlate with those generated from frozen samples. We also evaluated whether storing fecal samples in alcohol is a suitable alternative to preserve FGCM and PdG concentrations long-term (i.e. over a 9-month period) at locations where fecal extraction is not feasible. Finally, we tested if the hormone concentrations in unpreserved fecal samples of orangutans change over 14 h when stored at ambient conditions, representing the maximum duration between sample collection and return to the camp. FGCM and PdG levels measured from samples that were extracted with the field-friendly method showed strong correlations with those generated from frozen samples, and mean levels did not differ significantly between these methods. FGCM concentrations showed no significant change compared to control samples when fecal samples were stored for up to 6 months in alcohol at ambient temperature and PdG concentrations even remained stable for up to 9 months of storage. FGCM concentrations of fecal samples kept at ambient temperature for up to 14 h post-defecation did not significantly differ compared to control samples frozen immediately after collection. These results provide the basis for the successful monitoring of the physiological status of orangutans living in remote natural settings, like those included in the Indonesian reintroduction programs.     

Blood collection from the tail of a rat

Techniques for blood collection from the rat include puncture of the heart, retro-orbital plexus, jugular vein, saphenous vein, tail blood vessels, carotid artery, abdominal aorta, and vena cava. Most techniques (except saphenous vein and tail blood vessel puncture) require anesthesia. The following discussion focuses on two methods of blood collection - ventral tail artery puncture and dorsal or lateral tail vein puncture.     

Acid‐insoluble ash as a measure of dry matter digestibility in captive African elephants (Loxodonta africana)

There are limited data on the diet dry matter digestibility (DMD) of captive African elephants. Although the total fecal collection method is the standard for determining DMD, it is labor‐intensive, time‐consuming, and expensive. The acid‐insoluble ash (AIA) marker technique has been used successfully to determine DMD in ruminants and monogastrics. The objective of this study was to assess how accurately the AIA marker technique could estimate the DMD of captive African elephants (Loxodonta africana). Three mature male African elephants at Disney's Animal Kingdom in Florida were used in this study. The animals were offered a Bermuda grass hay‐based ration, and the total dry matter intake (DMI) and total fecal output were measured daily over a 7‐day period to determine the total collection DMD. The feed ingredients and fecal samples were also analyzed for AIA. Although there were differences (P<0.05) in total DMI and total fecal outputs, the DMD values did not (P>0.05) differ (35.1±0.72 vs. 37.1±0.72 for total collection and AIA, respectively). There was a linear (y=0.9461x; R²=0.74) relationship between the total collection and AIA marker technique DMD values. These results suggest that AIA can be used to accurately estimate the DMD of captive African elephants. Zoo Biol 0:1–5, 2005. © 2005 Wiley‐Liss, Inc.     

A biological validation procedure for the measurements of fecal outputs and fecal cortisol metabolites in male Syrian hamsters

Monitoring fecal outputs and fecal cortisol metabolites (FCM), a noninvasive technique, has been used to investigate physiological responses to stress and relationships between hormones and behavior in an increasing number of species. The aim of this study was to investigate whether measurements of fecal outputs and FCM can be used as indexes to repeatedly and precisely monitor stress levels in male Syrian hamsters using a social defeat as a biological validation method. The feces voided by each animal were collected every 3 h for at least 1 day before and after experiencing a single fighting interaction, and the extracted FCM during the pre- and post-fight phases was quantified by enzyme immunoassays. During the pre-fight baseline phase, both the number of fecal pellets and the FCM levels fluctuated throughout the whole day. Although the number of fecal pellets did not differ between the dark and light cycles, the levels of FCM were significantly higher during the dark cycle than during the light cycle. During the post-fight phase, the experience of fighting did not result in a significant difference in the number of fecal pellets per hour between the winner and loser groups, but did considerably increase the total amount of fecal outputs in both groups. The level of FCM was significantly higher in the loser group than in the winner group during the 1st and 7th 3-h collection periods after the fight, which indicated that the experience of defect affected the behavioral and physiological responses of the losers. Our findings suggest that measurement of FCM is sensitive enough to distinguish the stress levels between winners and losers after experiencing a fight. The measurements of fecal outputs and FCM levels provide new opportunities to longitudinally and frequently monitor behavioral and hormonal responses to stress in hamsters and other small laboratory animals.     

Evaluation of porcelain cup soil water samplers for bacteriological sampling

The validity of obtaining soil water for fecal coliform analyses by porcelain cup soil water samplers was examined. Numbers from samples of manure slurry drawn through porcelain cups were reduced 100- to 10,000,000-fold compared to numbers obtained from the external manure slurry, and 65% of the cups yielded coliform-free samples. Fecal coliforms adsorbed to cups apparently were released, thus influencing the counts of subsequent samples. Fecal coliforms persisted in soil water samplers buried in soil and thus could significantly influence the coliform counts of water samples obtained a month later. These studies indicate that porcelain cup soil water samplers do not yield valid water samples for fecal coliform analyses.     

Cup blocks the precocious activation of the orb autoregulatory loop

Translational regulation of localized mRNAs is essential for patterning and axes determination in many organisms. In the Drosophila ovary, the germline-specific Orb protein mediates the translational activation of a variety of mRNAs localized in the oocyte. One of the Orb target mRNAs is orb itself, and this autoregulatory activity ensures that Orb proteins specifically accumulate in the developing oocyte. Orb is an RNA-binding protein and is a member of the cytoplasmic polyadenylation element binding (CPEB) protein family. We report here that Cup forms a complex in vivo with Orb. We also show that cup negatively regulates orb and is required to block the precocious activation of the orb positive autoregulatory loop. In cup mutant ovaries, high levels of Orb accumulate in the nurse cells, leading to what appears to be a failure in oocyte specification as a number of oocyte markers inappropriately accumulate in nurse cells. In addition, while orb mRNA is mislocalized and destabilized, a longer poly(A) tail is maintained than in wild type ovaries. Analysis of Orb phosphoisoforms reveals that loss of cup leads to the accumulation of hyperphosphorylated Orb, suggesting that an important function of cup in orb-dependent mRNA localization pathways is to impede Orb activation.     

The effect of docking length on the risk of tail biting,tail-directed behaviour,aggression and activity level of growing pigs kept under commercial conditions

Tail biting in domestic pigs relates to a range of risk factors, primarily in the pigs’ environment. Preventive tail docking is widely used, and various experimental approaches suggest that docking reduces the risk of tail biting. However, whether the docking length affects the prevalence of tail biting outbreaks is less studied, as is how a shortened tail will affect pigs’ social behaviour. The aim of this study was to investigate how three different tail docking lengths, measured at docking, as well as retained intact tails (Short: 2.9 cm; Medium: 5.7 cm; Long: 7.5 cm; and Undocked) affected tail biting risk and behaviour directed at other finisher pigs with the same docking length treatment. Tail lesions were scored weekly, as was behaviour at pen level after introduction to finisher pens and until a potential outbreak of tail biting or slaughter. Pigs from four commercial herds (258 litters) entered the study. Before the pigs entered the finisher section and data collection started, some pigs were excluded, mainly due to tail biting outbreaks in the weaner section. The risk of a tail biting outbreak differed significantly between treatments (P=0.001), with a lowered risk of a tail biting outbreak in Short pens compared with Undocked (P<0.001) and Medium (P<0.05), and was affected by herd as well (P<0.001). Pens in the Long and Undocked treatments were pooled for the behavioural analysis due to low representation, especially in the Undocked treatment. The probability of tail contacts, where a pig interacted with a pen mate’s tail, differed between docking length treatments and was highest in the Long/Undocked compared with the Short treatment (P<0.01), but docking length did not affect aggressive behaviour. Docking length affected the risk of a tail biting outbreak and the frequency of tail-directed behaviour in our participating herds, of which three reported a high prevalence of tail biting problems. Only the shortest docking length treatment (Short) reduced the tail biting risk, but did not completely prevent tail biting outbreaks.     

The stimulatory effect of calpactin (annexin II) on calcium-dependent exocytosis in chromaffin cells: requirement for both the N-terminal and core domains of p36 and ATP

Calpactin, or calpactin heavy chain (p36), reconstitute secretion in digitonin-permeabilized adrenal chromaffin cells after a reduction in their secretory potential resulting from the loss of cytosolic components. We have characterized the stimulatory effect of p36, which resulted in an increase in both the extent and the rate of exocytosis. A mixture of other annexins (p70 and p32) did not have any effect on secretion at similar or greater concentrations than p36. Controlled proteolysis of p36 using chymotrypsin was carried out, and the 33,000 molecular weight core and 3000 molecular weight tail peptide isolated. In contrast to p36, p33 had no effect on exocytosis, even at high calcium concentrations. The N-terminal tail peptide and a synthetic peptide based on the tail of p36 [Ac-calpactin-(1-15)-NH2] had no effect on endogenous secretion, or secretion stimulated by exogenous p36. The results show that both the tail and core domains are required for p36 to stimulate exocytosis. The tail domain is unlikely to be required for interaction with cellular components but probably has a regulatory effect on the core domain. Endogenous secretion and the stimulatory effect of p36 were markedly inhibited by depletion of ATP. The ATP requirement for p36 action was not due to a requirement for phosphorylation by protein kinase C (PKC), since the PKC inhibitor staurosporine partially inhibited endogenous secretion but did not affect the stimulation of exocytosis due to exogenous p36.     

Comparison of arboreal beetle catches in wet and dry collection cups with Lindgren multiple funnel traps

We compared the effectiveness of a dry collection cup (with an insecticide killing strip) to a wet collection cup (containing antifreeze) for use with Lindgren multiple-funnel traps in catching several common species of bark and wood-boring beetles, and their associates in southern pine forests. All traps were baited with either the binary combination of ethanol and (-)-alpha-pinene or the quaternary combination of (+/-)-ipsenol, (+/-)-ipsdienol, ethanol, and (-)-alpha-pinene. We found that cup treatment had little, if any, effect on catches of Ips avulsus (Eichhoff) and I. grandicollis (Eichhoff) (Coleoptera: Scolytidae), Alaus myops (F.) (Elateridae), Chalcophora Solier species (Buprestidae), Temnochila virescens (F.) (Trogositidae), and Lasconotus Erichson species (Colydiidae). In contrast, catches of the following species were significantly less (by 40-97%) in traps with dry cups than in traps with wet cups: Hylobius pales Herbst and Pachylobius picivorus LeConte (Curculionidae); Buprestis lineata F. (Buprestidae); Acanthocinus obsoletus (Olivier), Arhopalus rusticus nubilus (LeConte), Monochamus titillator (F.) and Xylotrechus sagittatus sagittatus (Cerambycidae); Hylastes porculus Erichson and Xyleborinus saxeseni (Ratzeburg) (Scolytidae); and Thanasimus dubius (F.) (Cleridae). The same was true in at least one experiment for the following species: Dendroctonus terebrans (Olivier), Hylastes salebrosus Eichhoff, Hylastes tenuis Eichhoff, and Xylosandrus crassiusculus (Motschulsky) (Scolytidae). We conclude that cup treatment can have a significant impact on catches of some arboreal beetles in baited multiple-funnel traps. Anyone using multiple-funnel traps to capture arboreal beetles should evaluate the potential impacts arising from their choice of collection cup treatment to their trapping objectives and expectations. The issue of cup treatment may be particular important at low population levels when maximum trap efficiency is required such as in the detection of exotic insects at ports-of-entry and within quarantine and containment zones.     

Feasibility and Recommendations for Swift Fox Fecal DNA Profiling

Abstract: Genetic profiling using fecal samples collected noninvasively in the wild can provide managers with an alternative to live-trapping. However, before embarking on a large-scale survey, feasibility of this methodology should be assessed for the focal species. Costs associated with fecal genotyping can be high because of the need for multiple amplifications to prevent and detect errors. Assessing the relative amount of target DNA before genotyping means samples can be eliminated where error rates will be high or amplification success will be low, thereby reducing costs. We collected fecal samples from an endangered population of swift fox (Vulpes velox) and employed target-DNA quantification and a screening protocol to assess sample quality before genetic profiling. Quantification was critical in identifying samples of low quality (68%, <0.2 ng/μL). Comparison of the amplification, from a subset of loci in 25 samples that did not meet the screening criteria, confirmed the effectiveness of this method. The protocol, however, used a considerable amount of DNA, and an assessment of the locus and sample variability allowed us to refine it for future population surveys. Although we did not use <50% of the samples we collected, the remaining samples provided 36 unique genotypes, which corresponded to approximately 70% of animals estimated to be present in the study area. Although obtaining fecal DNA from small carnivores is challenging, our protocol, including the quantification and qualification of DNA, the selection of markers, and the use of postgenotyping analyses, such as DROPOUT, CAPWIRE, and geographic information, provides a more cost-effective way to produce reliable results. The method we have developed is an informative approach that wildlife managers can use to conduct population surveys where the collection of feces is possible without the need for live-trapping.     

Persistence and distribution of pollution indicator bacteria on land used for disposal of piggery effluent.

Numbers of pollution indicator bacteria (fecal coliforms and fecal streptococci) were assessed on land to which effluent from intensively housed pigs had been applied. Topsoil (to a 30-mm depth) was found to provide a more favorable environment for fecal coliform persistence than was pasture or subsoil. Times required for a 90% reduction in number (T90) in topsoil (calculated by linear regression of log counts obtained in a 6-week period after effluent application) ranged from 7 to 20 days (mean T90, 11 days). T90 values for fecal coliforms fell within this range irrespective of the season of application and for a number of soil types and climatic conditions. The range in die-off times was encountered irrespective of the fecal coliform count in the applied effluent or the application regimen (125 to 1,000 kg of elemental nitrogen in the form of effluent per ha; return periods, 3 to 12 months). Autumn and winter conditions were conducive to the persistence of a survivor tail of these bacteria at 10(1) to 10(3) cells per g of topsoil. Fecal streptococci survived similarly on soil and pasture (T90, ca. 14 days) and appeared slightly more suited to survival in the environment than did fecal coliforms. Contamination of subsoils after effluent applications occurred at a rate well in excess of the infiltration capacity of the soil, presumably by percolation of the effluent through soil cracks. Contamination levels of subsoils in the experimental area generally remained low.     

Significance of low-temperature growth associated with the fecal coliform response, indole production, and pectin liquefaction in Klebsiella

In the genus Klebsiella, the growth respnse in nutient broth at 10 degrees C correlates inversely with the operational definition of a fecal coliform and not merely with the ability to grow at 44.5 degrees C. Of the fecal coliform-positive Klebsiella, 97% did not grow at 10 degrees C after 72 h of incubation. Conversely, 97% of the fecal coliform-negative isolates grew at 10 degrees C. The amount of growth at 10 degrees C varied among the fecal coliform-negative isolates and was found to correlate with indole production and pectin liquefaction. Low-temperature growth associated with specific biochemical tests can be used to differentiate several groups in the genus Klebsiella. Three main groups were discerned. Group I consists of indole-negative, pectin-nonliquefying, fecal coliform-positive isolates that do not grow at 10 degrees C. Group II isolates are differentiated from group I by a fecal-coliform-negative response and growth at 10 degrees C. Group III are indole-positive, pectin-liquefying, fecal coliform-negative isolates that grow at 10 degrees C. In our culture collection, isolates of group I are most frequently of human/animal clinical origins, whereas isolates of groups II and III are predominantly derived from the environment.     

Significance of low-temperature growth associated with the fecal coliform response, indole production, and pectin liquefaction in Klebsiella.

In the genus Klebsiella, the growth respnse in nutient broth at 10 degrees C correlates inversely with the operational definition of a fecal coliform and not merely with the ability to grow at 44.5 degrees C. Of the fecal coliform-positive Klebsiella, 97% did not grow at 10 degrees C after 72 h of incubation. Conversely, 97% of the fecal coliform-negative isolates grew at 10 degrees C. The amount of growth at 10 degrees C varied among the fecal coliform-negative isolates and was found to correlate with indole production and pectin liquefaction. Low-temperature growth associated with specific biochemical tests can be used to differentiate several groups in the genus Klebsiella. Three main groups were discerned. Group I consists of indole-negative, pectin-nonliquefying, fecal coliform-positive isolates that do not grow at 10 degrees C. Group II isolates are differentiated from group I by a fecal-coliform-negative response and growth at 10 degrees C. Group III are indole-positive, pectin-liquefying, fecal coliform-negative isolates that grow at 10 degrees C. In our culture collection, isolates of group I are most frequently of human/animal clinical origins, whereas isolates of groups II and III are predominantly derived from the environment.     

Suction sampling technique for obtaining standardized cytologic specimens from the oral mucosa

To enhance the value of exfoliative cytology for the study of the oral mucosa, a simple apparatus was developed to permit adequate sampling of a specific site so that samples collected on different occasions could be compared. The device essentially consists of a collecting cup connected to a blood collection evacuation system. The collecting cup is a modified female half of a stainless-steel filter holder supporting a 13-mm-diameter cellulose filter of 0.05-microns pore size. Suction pressure is applied by means of a 10-mL glass tube of premeasured vacuum. After positioning the collecting cup on the selected site on the buccal mucosa, the vacuum (440 mm Hg) is applied for five seconds. The mucosa is drawn in against the filter, producing a monolayered imprint of cells. This sample may be disengaged from the filter by agitation into a solution; this allows quantitative cytologic studies, such as the measurement of cell numbers by an electronic counter or the estimation of the areas of cells and nuclei by computer-aided image analysis of Cytospin preparations. Five separate samplings from each of three test subjects produced a harvest of 3,000 to 7,000 epithelial cells per sample; the cellular areas ranged from 784 to 1,052 sq microns while the nuclear areas ranged from 18.4 to 21.8 sq microns.