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Natural history of Schistosoma mansoni infection in mice: egg production, egg passage in the feces, and contribution of host and parasite death to changes in worm numbers.

Mice, C57Bl/6N (B6) and BALB/cAnN (BALB), infected with Schistosoma mansoni were examined 8-26 weeks postinfection (PI) to estimate the fecundity of the worms and the contribution of death of worms and the death of heavily infected mice to the decrease in worm numbers in chronic infections. Portal worms were recovered by perfusion and the lungs were examined for parasites shunted from the portal circulation. Animals that died were more heavily infected than those that survived. Between eight and 12 weeks PI, this loss of worms resulted in a net decrease of approximately 19% of worm pairs in surviving BALB mice, but of only 4% in B6 mice. Loss of portal worms to the lungs after the eighth week of infection was 9-13% of portal worms in BALB mice and 3-4% in B6 mice. The estimated rates of egg production by S. mansoni decreased slightly with time in both strains of mice. At 12 and 20 weeks PI, tissue eggs per worm pair and eggs passed in the feces per worm pair often decreased as the intensity of infection increased. We do not consider the loss of worms in the murine host relevant to most infections in humans because of the high intensity of infection relative to body size in mice and the high frequency of severe portal obstruction in murine infections.

Animals↗

Detection by polymerase chain reaction of Schistosoma mansoni DNA in human serum and feces.

A novel method for the detection of Schistosoma mansoni in human samples that is based on the amplification of a highly repeated DNA sequence has been developed. By use of simple DNA extraction techniques and a rapid 2-step polymerase chain reaction (PCR), it was possible to amplify S. mansoni DNA in human fecal and serum samples. The high sensitivity of the approach enabled the detection of the parasite DNA in fecal samples containing as few as 2.4 eggs per gram of feces, which makes it 10 times more sensitive than the Kato-Katz examination. A detection limit of I fg of Schistosoma sp. DNA was determined when pure DNA was used as PCR template. The amplification reaction showed to be specific giving no cross-reaction with DNA from other helminths. The PCR assay developed in this study may constitute a valuable alternative for the diagnosis of the Schistosoma sp. infection.

Animals↗

Comparison of autofluorescence and iodine staining for detection of Isospora belli in feces.

To evaluate the sensitivity of autofluorescence for detection of Isospora oocysts, wet preparations of 192 stool samples from patients with chronic diarrhea were examined by fluorescence microscopy and by light microscopy after iodine staining used for routine screening for ova and parasites. Silicon-chambered glass coverslips were used for fluorescence microscopy. Isospora oocysts were detected in 46 iodine-stained concentrated stool samples; 91 samples were positive by autofluorescence. According to the maximum likelihood estimates, examination by fluorescence (95.7%; 95% confidence interval [CI], 85.2-99.5) was significantly more sensitive than iodine staining (48.4%; 95% CI, 37.7-59.1). Examination for autofluorescence is a simple, highly sensitive, inexpensive, and easily applicable method to detect Isospora spp. oocysts in feces.

Animals↗

Thermal inactivation of Escherichia coli O157:H7 isolated from ground beef and bovine feces, and suitability of media for enumeration.

Rates of thermal inactivation of five strains of Escherichia coli O157:H7 isolated from ground beef implicated in outbreaks of hemorrhagic colitis and five strains isolated from bovine feces were determined. Ground beef (22% fat, 10 g), inoculated with individual test strains at populations ranging from 6.85 to 7.40 log10 CFU g-1 of beef, was formed into patties (0.3 cm thick and 8.0 cm in diameter) and sealed in polyethylene bags. For each strain and treatment temperature (54.4, 58.9, 62.8, 65.6, or 68.3 degrees C), 6 bags were simultaneously immersed into a recirculating water bath. Viable cells in patties heated for various lengths of time were enumerated by plating diluted samples on sorbitol MacConkey agar supplemented with 4-methylumbelliferyl-beta-D-glucuronide (MSMA) and modified eosin methylene blue (MEMB) agar. Regardless of strain or treatment temperature, higher numbers of E. coli O157:H7 cells were generally recovered on MEMB agar than on MSMA, indicating the inferiority of MSMA as a recovery medium for quantitative determination of E. coli O157:H7 cells in heat-processed ground beef. Significantly (P < or = 0.05) higher D values when enumeration was done using MEMB agar compared with MSMA. Mean D values for combined strain data at 54.4, 58.9, 62.8, and 65.6 degrees C from cultures on MEMB agar were 123.90, 6.47, 0.62, and 0.20 min, respectively, whereas D values of 25.5, 5.21, and 0.18 min were obtained at the same temperatures from cultures on MSMA. Results suggest that cooking ground beef patties to an internal temperature of 68.3 degrees C for 40 s will inactivate at least 99.99% of E. coli O157:H7 cells; z values of 4.0 and 5.1 degrees C were calculated from mean D values obtained from MEMB agar and MSMA, respectively, as recovery media. Differences in D values existed among strains but rates of thermal inactivation do not appear to be correlated with the sources of the isolates.

Animals↗

Survey of Salmonella serotypes shed in feces of beef cows and their antimicrobial susceptibility patterns.

Salmonella prevalence on cow-calf operations was studied as a part of a national study of health and management of the U.S. beef cow-calf industry and was conducted as part of the National Animal Health Monitoring System. Within this study, the prevalence of Salmonella spp. shed in feces was determined. A total of 5,049 fecal samples were collected from 187 beef cow-calf operations each visited on a single occasion. The number of fecal samples collected per operation was predetermined based on herd size. Salmonellae were recovered from 1 or more fecal samples collected on 11.2% (21 of 187) of the operations. Overall 78 salmonellae representing 22 serotypes were recovered from 1.4% (70 of 5,049) of samples. Multiple serotypes were recovered from eight samples from a single operation. The five most common serotypes were Salmonella Oranienburg (21.8% of isolates), and Salmonella Cerro (21.8%), followed by Salmonella Anatum (10.3%), Salmonella Bredeney (9.0%), and Salmonella Mbandaka (5.1%). The most common serogroups identified were C1 (33.3%), K (21.8%), B (16.7%), and E (15.4%). Even though the recovery rate of salmonellae from fecal samples was very low, 43.6% (34 of 78) and 38.5% (30 of 78) of the isolates were among the 10 most common serotypes from cattle with clinical signs of disease or isolated from humans, respectively. The majority of the isolates (50 of 78; 64.1%) were recovered from fecal samples from two operations. All isolates were screened for resistance to a panel of 17 antimicrobics, and 87.2% (68 of 78) were susceptible to all of the antimicrobics. The resistant isolates were most commonly resistant to streptomycin (n = 9) and/or sulfamethoxazole (n = 9). Nine isolates showed multiple (> or =2 antimicrobics) resistance most commonly to streptomycin and sulfamethoxazole (n = 6).

Animals↗

Broiler carcass contamination with Campylobacter from feces during defeathering.

Three sets of experiments were conducted to explore the increase in recovery of Campylobacter from broiler carcasses after defeathering. In the first set of experiments, live broilers obtained from a commercial processor were transported to a pilot plant, and breast skin was sampled by a sponge wipe method before and after defeathering. One of 120 broiler breast skin samples was positive for Campylobacter before defeathering, and 95 of 120 were positive after defeathering. In the second set of experiments, Campylobacter-free flocks were identified, subjected to feed withdrawal, and transported to the pilot plant. Carcasses were intracloacally inoculated with Campylobacter (10(7) CFU) just prior to entering the scald tank. Breast skin sponge samples were negative for Campylobacter before carcasses entered the picker (0 of 120 samples). After defeathering, 69 of 120 samples were positive for Campylobacter, with an average of log10 2.7 CFU per sample (approximately 30 cm2). The third set of experiments was conducted using Campylobacter-positive broilers obtained at a commercial processing plant and transported live to the pilot plant. Just prior to scalding, the cloacae were plugged with tampons and sutured shut on half of the carcasses. Plugged carcasses were scalded, and breast skin samples taken before and after defeathering were compared with those collected from control broilers from the same flock. Prior to defeathering, 1 of 120 breast skin sponge samples were positive for the control carcasses, and 0 of 120 were positive for the plugged carcasses. After passing through the picker, 120 of 120 control carcasses had positive breast skin sponge samples, with an average of log10 4.2 CFU per sample (approximately 30 cm2). Only 13 of 120 plugged carcasses had detectable numbers of Campylobacter on the breast skin sponge, with an average of log10 2.5 CFU per sample. These data indicate that an increase in the recovery of Campylobacter after defeathering can be related to the escape of contaminated feces from the cloaca during defeathering.

Animals↗

Prevalence of Listeria spp. in feces and carcasses at a lamb packing plant in Brazil.

The objective of this work was to study the occurrence of Listeria species in feces and on dressed and cooled carcasses of lambs at a packing plant in Brazil. Listeria spp. were recovered on Oxford and Palcam agars. The 35 fecal samples yielded Listeria welshimeri (20%) and Listeria innocua (8.6%). The 69 carcass samples yielded L. innocua (34.8%), Listeria monocytogenes (4.3%), and Listeria ivanovii (1.5%). More Listeria spp. were recovered with two selective agars than with either agar alone.

Agar↗

Characterization of O157:H7 and other Escherichia coli isolates recovered from cattle hides, feces, and carcasses.

In a previous study, the seasonal prevalence was reported for stx+ Escherichia coli O157:H7 in feces and on hides and carcasses of cattle at processing. Overall, 1,697 O157:H7 isolates have now been characterized for the incidence of (i) eae(O157), hlyA, stx1, and stx2 in the recovered isolates and (ii) presumptive rough and presumptive nonmotile isolates. Seven O157:H7 isolates (0.4%) lacked stx genes, although they carried eae and hlyA. All but one of the isolates carried both eae and hlyA. Approximately two-thirds of the isolates (64% when one isolate per sample was considered) carried both stx1 and stx2. E. coli O157:H7 cells that harbored both stx1 and stx2 were more often recovered from hides in the fall (79% of the fall hide isolates) and winter (84% of the winter hide isolates) than in the spring (53%) and summer (59%). Isolates recovered from preevisceration carcasses showed a similar but not statistically significant trend. Twenty-three of the 25 O157:H7 isolates carrying stx1 but not stx2 were recovered during summer. Fifteen presumptive rough and 117 presumptive nonmotile stx+ O157:H7 isolates were recovered. Ten (67%) of the presumptive rough isolates were recovered during summer. Ninety-five of the presumptive nonmotile isolates (81%) were recovered during fall. Forty-eight percent of the false-positive isolates (175 of 363) tentatively identified as O157:H7 were O157+ H7- and lacked eae(O157), hlyA, and stx. These data suggest that in beef processing samples (i) there are minor seasonal variations in the prevalence of stx genes among E. coli O157:H7 isolates, (ii) presumptive rough and presumptive nonmotile stx+ O157:H7 isolates are present, (iii) E. coli O157:H7 isolates lacking stx genes may be rare, and (iv) O157+ H7- isolates lacking stx genes can result in many false-positive results.

Animals↗

Survival of Escherichia coli O157:H7 in bovine feces over time under various temperature conditions.

Although Escherichia coli O157:H7 prevalence estimates in cattle have increased over time due to improvements in detection methods, fecal sample transport conditions from farm to microbiological laboratories for further analysis may be a factor for prevalence underestimation. The objective of this study was to compare and determine the survival characteristics of E. coli O157:H7 in bovine feces under various storage conditions that could be encountered during transport. Fecal pats were inoculated with a four-strain cocktail of antibiotic-resistant E. coli O157:H7 to contain approximately 1 x 10(5) CFU/g. Inoculated and control samples were taken after 0, 24, 48, 120, and 168 h at each storage temperature and examined for presence and numbers of E. coli O157:H7. Each sample was subdivided and placed at each of the four following temperatures: 37, 23, 4.4 degrees C, and in a plastic cooler with refrigerant packs (0, 4, 4, 21, and 23 degrees C at five sampling times, respectively) to simulate transportation conditions. A statistically significant decrease in the population of the pathogen was observed after 48 h in samples held at 37 degrees C (P < 0.01) and after 168 h at 4.4 degrees C (P = 0.02). At 37 degrees C, E. coli O157:H7 was not detected after 48 h, either by direct plating (P < 0.01) or by immunomagnetic separation. Overall, the results of this study showed that E. coli O157:H7 survived without significant detriment in bovine fecal material inside the cooler for up to 168 h. These results indicate that shipment and storage under these conditions before microbiological analysis would be acceptable and should not affect pathogen detection.

Animals↗

Isolation and characterization of Yersinia enterocolitica from swine feces recovered during the National Animal Health Monitoring System Swine 2000 study.

A national study was conducted for the isolation of pathogenic Yersinia enterocolitica in pig feces in the United States as part of the National Animal Health Monitoring System's Swine 2000 study. Fecal samples collected from swine operations from September 2000 to March 2001 from 77 production sites in 15 of the top 17 swine-producing states were tested for the presence of pathogenic Y. enterocolitica. After enrichment of swine fecal samples in irgasan-ticarcillin-potassium chlorate broth, the enriched cultures were plated on cefsulodin-irgasan-novobiocin agar for isolation of presumptive Y. enterocolitica. The isolates were confirmed as pathogenic Y. enterocolitica by the fluorogenic 5' nuclease PCR assay targeting the chromosomal attachment invasion ail gene. Of 2793 fecal samples tested, 106 (3.80%) ail-positive strains of Y. enterocolitica were isolated. These 106 ail-positive isolates originated from 7 of the 15 participating states. The predominant serotype O:3 (n = 79 of 106) was distributed in five states (n = 5 of 7). Serotype O:5 (n = 27 of 106) was also found in five states (n = 5 of 7). All isolates contained the virulence plasmid and expressed virulence-associated phenotypic characteristics. These results indicate that swine in the United Stares harbor Y. enterocolitica that can potentially cause human illness.

Animals↗

Seasonal changes in the prevalence of ova of Diphyllobothrium ursi and Baylisascaris transfuga in the feces of the black bear (Ursus americanus).

An analysis of 114 samples of feces from black bears (Ursus americanus) from La Verendrye Park, Quebec revealed that the prevalence of eggs of Diphyllobothrium ursi and Baylisascaris transfuga varied seasonally. D. ursi ova were most prevalent during the summer and fall and at their lowest in spring while the converse was true for B. transfuga. This phenomenon possibly is associated with the cessation of feeding during the winter denning of black bears.

Animals↗

Detection of parvoviruses in wolf feces by electron microscopy.

One hundred fifteen wolf (Canis lupus) feces were collected between 1980 and 1984 from northeastern Minnesota and were examined for canine parvovirus by negative contrast electron microscopy. Of these, seven (6%) samples revealed the presence of parvovirus. Some of these viruses were able to grow in cell cultures forming intranuclear inclusion bodies and giant cells.

Animals↗

Radiometric culture of Mycobacterium avium paratuberculosis from the feces of tule elk.

To determine if Mycobacterium avium paratuberculosis has persisted in tule elk (Cervus elaphus nannodes) at Point Reyes National Seashore (California, USA), 100 fresh fecal samples were collected. Feces were cultured on a modified BACTEC 12B radiometric medium for detection of M. avium paratuberculosis. Four samples, coming from two separate groups of elk tested positive for M. avium paratuberculosis. Thus, a noninvasive technique was used to document the continued presence of M. avium paratuberculosis in elk at Point Reyes National Seashore. These findings document persistence of this organism for a period of at least 13 yr in a free ranging herd of elk, with a 6 yr absence of observed clinical signs.

Animals↗

Prevalence and serovars of Salmonella in the feces of free-ranging white-tailed deer (Odocoileus virginianus) in Nebraska.

To determine the prevalence and serovars of Salmonella in free-ranging deer, we cultured feces from white-tailed deer (Odocoileus virginianus) harvested by hunters during a regular firearm season in southeastern Nebraska (USA). We recovered Salmonella from 5 (1%; 95% confidence interval: 0.37-2.20%) of 500 samples and identified four different Salmonella enterica serovars [Litchfield (1), Dessau (1), Infantis (2), and Enteritidis (1)]. Although the prevalence of Salmonella in free-ranging deer appears to be low, the serovars recovered are known to be pathogenic to humans and animals.

Animals↗

[Milk substitutes with high ash and macro mineral contents--of importance in loose feces of young calves?].

In 3 feeding experiments with 23 calves (age: 5-18 days at trials' start) the effects of high ash and mineral contents in the diet on feces composition (dry matter, pH, concentrations of lactic and volatile fatty acids, mineral contents) and digestibility rates of organic matter, nutrients and minerals as well as on some parenteral parameters (electrolytes in serum, renal excretion) were tested. In all feeding experiments a change-over-design was used, during 10 days each of all calves was fed the control and experimental diet, respectively. In the first trial (8 calves) high ash and mineral contents in the milk replacer were caused by use of a special whey product (25% in the diet), in the second experiment (n = 7) comparable ash and macromineral concentrations were achieved by adding minerals to the basic diet, in the third trial (n = 8) the whey product was used in a lower proportion (15% of the diet). Ash concentrations in diet (g/kg dry matter): 68 versus 138 (trial 1), 75 versus 135 (trial 2), 75 versus 93 (trial 3). The differences in the ash contents were based mainly on differences in the sodium and potassium concentrations.

Animal Feed↗

Effects of sample storage and delayed secondary enrichment on detection of Salmonella spp in swine feces.

OBJECTIVE: To determine effects of fecal sample storage and delayed secondary enrichment (DSE) on detection of Salmonella spp in swine feces. Sample Population-Fecal samples obtained from 84 pigs in a commercial herd. PROCEDURE: Each fecal sample underwent 3 storage treatments: no storage (ie, processed on the day of collection), storage at 4 C for 6 days, and storage at -15 C for 14 days. After assigned storage treatments, all samples were enriched in Rappaport-Vassiladias (RV) broth (single enrichment) and plated on XLT4 agar. Delayed secondary enrichment was performed, using single enrichment broths that were stored for 4 days at room temperature. RESULTS: Of 504 cultures, 186 (36.9%) were Salmonella positive. A difference in proportions of samples with positive results was not found between same-day processing and storage at 4 C for 6 days. Compared with use of single enrichment for 24 hours (34% positive), use of DSE resulted in a greater proportion (40%; P < 0.001) of samples with positive results. Estimated relative sensitivities for the storage methods were 0.90, 0.85, and 0.71 for same-day processing, storage at 4 C for 6 days, and storage at -15 C for 14 days, respectively. CONCLUSIONS: Where practical, processing of fecal samples on the day of collection is recommended, although storage at 4 C for several days does not result in marked loss of sensitivity. Improved detection associated with DSE warrants further investigation and optimization.

Animals↗

Presence of Yersinia enterocolitica in tissues of orally-inoculated pigs and the tonsils and feces of pigs at slaughter.

In order to study the early events associated with infection of swine by Yersinia enterocolitica, 42 five-week-old crossbred piglets were inoculated per os with approximately 10(8) Y. enterocolitica O:3. Groups of 5 animals (and one negative control) were euthanized 30 min, 3, 6, 12, 24, 48 and 72 h following the infection. Palatine tonsils, retropharyngeal and mesenteric lymph nodes, esophagus, duodenum, jejunum, ileum (and Peyer's patches), stomach, liver, spleen and feces (from colon) were collected and analyzed for the presence of Y. enterocolitica by standard bacteriological procedures. Natural infections were also analyzed, as a complementary study, by taking one-gram samples of fecal material and tonsils from 291 pig carcasses less than 3 h after slaughter and culturing them for Y. enterocolitica using a cold enrichment technique. Within 30 min, Yersinia enterocolitica O:3 was already present at most sites. The presence of Y. enterocolitica in the liver of 3 out of 10 animals and also in the spleen of 3 out of 10 piglets, within the first 3 h postinfection, but not at later times (with one exception), probably indicated a transient bacteremia accompanying the initial stages of infection. The tonsils were colonized in most animals (13/20) as the bacteria remained present from 12 to 72 h postinfection, while only 4 out of 20 fecal samples were found to be positive over the same period. Up to 10(4) colony-forming units of Y. enterocolitica per gram of tonsil and fecal material were recovered. Finally, among the 291 animals sampled at the abattoir, a total of 79 were found positive, 70 of the tonsils sampled were positive, and bacteria were recovered in 17 fecal samples. It is therefore suggested that palatine tonsils are the most reliable tissue for the indication of an infection/colonization by Y. enterocolitica O:3 in swine and that the removal of this tissue during the slaughter process should be considered in order to minimize the possibility of contamination of meat products.

Abattoirs↗

Detection of Cryptosporidium parvum in human feces by PCR.

C. parvum has a high pathologic potential also for man, especially for immununosuppressed patients. The microscopic detection of cysts in feces is neither easy nor always reliable. During the recent years, considerable progress has been achieved in establishing PCR-based approaches for i) sensitive detection of C. parvum in a variety of specimen types [3, 9-16, 18, 21, 24, 25, 27, 29-32, 34], ii) identification of individual genotypes of C. parvum [2, 4-7, 17, 19, 20, 22, 23, 26, 28, 33] and iii) viability testing of C. parvum organisms [8, 9, 13, 29, 30]. The protocols published so far include nested PCR [3, 8, 18, 34], RT-PCR [13, 29], and use of the UNG carryover prevention system [10]. The aim of this work was to establish a PCR system for the detection of C. parvum oocysts in stool samples, applying the same specimen preparation procedure as applied for immunofluorescence. In addition, we combined the UNG carryover prevention system with the use of long, PCR-generated digoxigenin-labelled probes, thus achieving a sensitivity comparable to nested PCR und circumventing the contamination risks associated with nested PCR protocols. We developed a simplified sucrose-cushion-based protocol for preparation of clinical specimens (adopted from [1]), satisfying both the needs of immunofluorescence and PCR. When tested with stool samples spiked with C. parvum oocysts, the analytical sensitivity of PCR was 3,500 oocysts/ml stool (immunofluorescence: 3,000 oocysts/ml stool), demonstrating that both methods were equivalent with respect to analytical sensitivity. However, when PCR and immnunofluorescence were applied to clinical samples (n=5) with known positivity for C. parvum, only the specimen with the shortest duration of storage (5 weeks) could be correctly identified by PCR (clinical sensitivity: 20%). Our results demonstrate, that the PCR approach presented in this work is not suited for highly sensitive detection of C. parvum in faeces. This was mainly due to the fact that sucrose-gradient purified material was used, which relies on the presence of morphologically intact oocysts in the specimens. Desintegration of oocysts by excystation and/or storage may lower the parasite yield of the protocol drastically. As a consequence, a protocol extracting the entire DNA from faeces should be used for PCR for detection of C. parvum [34].

Animals↗