SEARCH · Search PubMed
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Cryptosporidium oocysts were concentrated by an ether-phosphate-buffered saline sedimentation technique and then separated by density gradient centrifugation. This two-step method yielded highly concentrated oocysts largely free of bacteria and fecal debris.
The apparent causative organism from the only reported case of type E infant botulism was isolated and characterized. Except for its ability to produce type E botulinal toxin, this organism (strain 5262) would be unquestionably identified as Clostridium butyricum. This is the second time an organism resembling a defined Clostridium species other than a member of the C. botulinum group has been implicated in infant botulism.
A total of 2,693 fecal specimens, with 1,422 from healthy persons and 1,271 from patients suffering from enteric diseases, was investigated to isolate species of the Morganella-Proteus-Providencia group and to evaluate the role of these bacteria in intestinal disorders. Most strains were isolated from two media, i.e., blood agar and tryptophan agar. Two of the species were more frequently found in diarrheal cases than in healthy controls. These species were Morganella morganii and Proteus mirabilis. Two new species of Enterobacteriaceae, i.e., Proteus penneri and Providencia rustigianii, were found in 33 and 5 people, respectively. However, these two species were not found more frequently in the diarrheal cases.
A blood-free, charcoal-based selective medium (CSM) consisting of a Columbia agar base, activated charcoal (4 g/liter), hematin (0.032 g/liter), sodium pyruvate (0.1 g/liter), cefoperazone (32 mg/liter), vancomycin (20 mg/liter), and cycloheximide (100 mg/liter) supported the growth of Campylobacter jejuni and C. coli with colony counts equivalent to those obtained on antibiotic-free horse blood agar. CSM was compared to Skirrow medium (SKM) for the recovery of C. jejuni and C. coli from stools of patients with diarrhea, the media being incubated for 2 days under reduced oxygen tension at 43 degrees C. These campylobacters were isolated from 35 (2.9%) of 1,227 stools tested (29 on both media, 5 on CSM alone, and one on SKM alone). Whenever C. jejuni and C. coli were recovered, growth was pure on 29 CSM cultures (85%), but on only 11 SKM cultures (37%). Complete suppression of "contaminating" flora occurred in 704 CSM cultures (57%) compared with 426 SKM cultures (35%). CSM more effectively suppressed contaminating pseudomonads, gram-positive organisms, and yeasts than did SKM; both media failed to suppress members of the family Enterobacteriaceae in about a quarter of the samples. Studies on 20 representative Enterobacteriaceae contaminants showed that susceptibility to cefoperazone and growth on CSM were markedly dependent on inoculum size; 12 strains were inhibited by cefoperazone (32 mg/liter) at inoculum sizes of 5 X 10(2) and 5 X 10(4) but not 5 X 10(6) organisms, indicating that the frequency of contaminants on CSM could probably be reduced further by ensuring that stools were not inoculated too heavily on CSM. Our findings confirm that charcoal is an effective substitute for blood in media for growing campylobacters, and that CSM is a highly effective blood-free selective medium for isolating C. jejuni and C. coli from stools.
Cultures of Aeromonas species were tested for production of a toxin recently purified by Asao et al. (T. Asao, Y. Kinoshita, S. Kozaki, T. Uemura, and G. Sukaguchi, Infect. Immun. 46:122-127, 1984) and described as a hemolysin with enterotoxic and cytotoxic activity. The toxin was produced by only 63% of Aeromonas sobria strains and by 93% of Aeromonas hydrophila strains. Also, 54% of A. hydrophila strains produced another cytotoxic entity.
Our previously described (H. Goossens, M. De Boeck, and J. P. Butzler, Eur. J. Clin. Microbiol. 2:389-393, 1983) selective medium, consisting of cefoperazone (15 mg/liter), rifampin (10 mg/liter), colistin (10,000 IU/liter), and amphotericin B (2 mg/liter) (medium M1), for the isolation of Campylobacter jejuni and Campylobacter coli from stool specimens was modified as follows: cefoperazone (30 mg/liter), rifampin (10 mg/liter), and amphotericin B (2 mg/liter) (medium M2). A comparative study of the isolation of Campylobacter spp. from stool specimens was carried out with medium M1; medium M2; a selective blood-free medium consisting (per liter) of charcoal (4 g), ferrous sulfate (0.25 g), sodium pyruvate (0.25 g), casein hydrolysate (3 g), sodium deoxycholate (1 g), nutrient broth no. 2 (25 g), agar (12 g), and cefoperazone (32 mg) (medium M3); and Preston medium containing (per liter) trimethoprim (10 mg), rifampin (10 mg), polymyxin B (5,000 IU), and cycloheximide (100 mg) (medium M4). We also included a filtration system in which membrane filters were applied directly to the surface of the nonselective blood-free medium distributed in small petri dishes. A total of 5,276 stool specimens were tested: 2,788 stool specimens were tested on M1 and M3 in study 1; 2,488 stool specimens were inoculated on the four selective media in study 2, and the last 986 specimens of the 2,488 were tested in parallel with the filtration system. In study 2, 128 Campylobacter strains were isolated from 126 different patients; 85.0, 88.3, 82.5, and 66.6% of these strains were isolated on M1, M2, M3, and M4, respectively. No contaminating fecal flora was found on 65.4, 70.7, 62.4, and 40.3% of the M1, M2, M3, and M4 plates, respectively. Furthermore, C. coli was found to be more susceptible to antibiotics present in the selective media, particularly colistin and polymyxin B, than was C. jejuni. We therefore recommend M2 for the isolation of Campylobacter spp. Finally, the filtration method was found to be easy and cheap; although the sensitivity was low, this method allowed the isolation of new Campylobacter spp. which seem to be associated with diarrhea.
Five selective media were evaluated for their effectiveness in the primary isolation of Aeromonas spp. from human patients with acute diarrhea and from healthy domestic animals. Sheep blood agar with 30 mg of ampicillin per liter (ASBA30) yielded a significantly higher percentage of positive specimens as compared with the four other media. The effective combination of two selective media with which 98% of all isolates were detected and with which all of the three human-associated Aeromonas spp. could be isolated was ASBA30-DNase-toluidine blue agar. ASBA30 was the most sensitive medium since it permitted more growth of Aeromonas colonies and effectively suppressed competing microflora. We recommend the use of ASBA30-DNase-toluidine blue agar for investigations in which an attempt is made to assess the significance of Aeromonas spp. in the etiology of human diarrhea.
Since May 1983, our laboratory has, upon request, cultured stools for Yersinia spp. by using direct plating on cefsulodin-irgasan-novobiocin agar and a 3-week cold enrichment procedure. We isolated bacteria identified as Y. intermedia from six adult patients. All isolates were recovered only by the cold enrichment procedure and misidentified as Y. enterocolitica by the API 20E system (Analytab Products, Plainview, N.Y.). Final identification was made on the basis of results obtained with conventional tube biochemical tests. The isolates were tested for the following characteristics associated with virulence in Y. enterocolitica: lack of pyrazinamidase activity, autoagglutinability, presence of a 40- to 50-megadalton plasmid, production of heat-stable enterotoxin, and mouse lethality. All isolates tested had pyrazinamidase activity, and none were autoagglutinable. However, one isolate possessed a 40-megadalton plasmid. None produced enterotoxin or were lethal for mice. Review of the medical histories of the patients revealed that four of the six had diarrhea; however, none had disease typical of that caused by Y. enterocolitica. Our data confirmed the limited pathogenic potential of Y. intermedia and suggested that its isolation was without clinical significance in our patients. Conventional biochemical tests were required for reliable identification of Y. intermedia.
Between June 1982 and May 1987 Aeromonas species were isolated from 208 of 34,311 (0.61%) fecal samples submitted to a Regional Public Health Laboratory in The Netherlands. Aeromonas isolates were found most frequently in summer and rarely in winter. Of 169 Aermonas isolates that were available for further study, 19% were isolated from patients with a mixed infection, 5% from patients with underlying diseases, and 15% from patients who used medication that could predispose the intestinal tract to colonization with Aeromonas species. Aeromonas species that produced cytotoxins to Vero cells (cytotoxigenic) were found in hybridization groups 1 (11% of all isolates), 2 (1%), 3 (2%), and 8 (25%) and were identified phenotypically as A. hydrophila or A. sobria. Aeromonas species that did not produce cytotoxins to Vero cells (noncytotoxigenic) were found in hybridization groups 4 (57%) and 5A (4%) and were identified phenotypically as A. caviae. Distribution of Aeromonas species by age showed a predominance of noncytotoxigenic strains in children under the age of 5 years (46% of all noncytotoxigenic strains), while cytotoxigenic strains were mainly cultured from patients aged 50 years or older (54% of all cytotoxigenic strains). Significant correlations were found between cytotoxigenic strains and hospitalization, foreign travel, and contact with surface water. Cytotoxigenic strains were isolated significantly more often than noncytotoxigenic strains from patients with diarrhea, but in a multivariate analysis including age, previous medication, underlying disease, and foreign travel, this association was not significant.
During a 3-year period, "Campylobacter upsaliensis" was isolated from 99 patients. Phenotypic characterization and numerical analysis of protein electrophoregrams showed evidence that "C. upsaliensis" is a distinct Campylobacter species with unique characteristics. The MBCs of 13 antibiotics were determined. In general, these organisms were highly susceptible to drugs that were present in the selective isolation media, making none of the available selective media suitable for the isolation of "C. upsaliensis." Ten strains were found to be resistant to erythromycin (MBCs, greater than or equal to 12.50 mg/liter). Plasmid DNA was detectable in 89 of the 99 strains; 16 plasmid profiles could be identified. Plasmid pattern 16, containing four plasmids of 52, 32, 5.5, and 2.6 megadaltons, represented 60.7% of the plasmid-containing strains. None of the "C. upsaliensis" strains could be agglutinated with antisera against heat-labile antigens from C. jejuni, C. coli, or C. laridis. "C. upsaliensis" was found to be susceptible to serum killing and was readily phagocytized by human polymorphonuclear cells.
A new solid-phase immune electron microscopy double-antibody colloidal-gold technique (SPIEMDAGT) was developed and compared with direct electron microscopy, direct immune electron microscopy, and enzyme immunoassay for detecting rotavirus. Guinea pig and rabbit antirotavirus antisera were used as capture and detector antibodies, respectively, and goat anti-rabbit immunoglobulin G-gold complexes were employed as a label. Animal rotavirus in cell culture media and human virus in stool specimens were detected by this method. On average, SPIEMDAGT detected 800 times more virus particles than direct electron microscopy and 45 times more particles than direct immune electron microscopy and yielded 20% more positives than enzyme immunoassay. SPIEMDAGT could detect not only viral antigen associated with morphologically recognizable particles but also antigen present when whole virus particles were not visible.
On the basis of antigenic variability in the VP7 outer capsid glycoprotein, at least 14 G serotypes exist for group A rotaviruses. Serotypic diversity exists among bovine rotaviruses (BRV), with serotypes G1, G6, G8, and G10 reported for cattle. Although G1 and G8 rotaviruses were originally described for humans, the recent isolation of G6 and G10 rotaviruses from humans further emphasizes the serotypic similarity between human and bovine rotaviruses and the possible zoonotic potential of rotaviruses. Results of our previous studies have indicated that more than 24% of BRV-positive field samples from diarrheic calves were nonreactive with cDNA probes or monoclonal antibodies to serotypes G6, G8, and G10. In this study, cDNA probes were prepared by polymerase chain reaction amplification of the hyperdivergent regions of the VP7 genes (nucleotides 51 to 392) from human (G1, G2, and G3) and porcine (G4, G5, and G11) rotaviruses. These probes were used in a dot blot hybridization assay to further characterize the G types of 59 BRV strains (fecal samples from diarrheic calves in Ohio, Nebraska, Washington, and South Dakota) that were nonreactive with cDNA probes to G6, G8, and G10. Rotaviruses belonging to serotypes G1 (n = 7), G2 (n = 1), G3 (n = 2), and G11 (n = 3) were identified among the BRV field samples. The BRV associated with these G types accounted for 22% of the samples tested; the other 78% of these samples remained untypeable with these probes. To our knowledge, this is the first report in the United States of the identification among BRV isolates of rotavirus serotypes G1, G2, G3, and G11.
Bovine fecal samples (1 g) negative for Cryptosporidium sp. oocysts were seeded with 7 x 10(4) Cryptosporidium parvum oocysts and purified by either water-ether concentration, sucrose density flotation, or zinc sulfate flotation to evaluate oocyst recovery. The effect of these purification techniques on the viability of recovered oocysts was also evaluated. Significantly higher numbers of seeded oocysts were recovered by water-ether concentration (recovery rate, 46 to 75%) than by sucrose density (24 to 65%) or zinc sulfate (22 to 41%) flotation methods. In addition, water-ether concentration did not exert a significant effect on the viability of the population of oocysts recovered, whereas sucrose density flotation and zinc sulfate flotation selectively concentrated viable oocysts. The water-ether concentration procedure is recommended for use in epidemiological studies in which both oocyst enumeration and viability assessment are required.
The sensitivities of several plating and broth enrichment methods for the detection of Escherichia coli O157:H7 in (i) bovine fecal samples directly inoculated with E. coli O157:H7, (ii) fecal samples from cattle in herds previously positive for E. coli O157:H7, and (iii) fecal samples from calves shedding E. coli O157:H7 after experimental oral inoculation were compared. Three enrichment protocols and three plating protocols were evaluated with directly inoculated fecal samples. All broth enrichment methods were superior to direct plating when they were combined with subsequent plating on sorbitol-MacConkey with cefixime and tellurite (SMACct). SMACct was the most sensitive plating medium, and the three alternative broth enrichment methods gave similar improvements in sensitivity. Of 351 fecal samples from known positive herds, 24 samples (6.8%) were positive by one or more methods. By the most sensitive plating method, cultures of 10-g samples were slightly more sensitive (19 of 351 [5.4%]) than cotton-tipped swab fecal samples (14 of 351 [4.0%]); however, this difference was not significant. For samples from calves orally inoculated with E. coli O157:H7, separation by immunomagnetic beads was slightly more sensitive (79%) than broth enrichment followed by plating at two dilutions (10(-3) and 10(-4)) (71%); however, this difference was not significant. The combination of overnight enrichment of swab fecal samples (0.1 g) and plating on SMACct at two dilutions (10(-3) and 10(-4)) appears to be a sensitive method for detection in large-scale studies involving hundreds of samples per week.
The identification of a new murine pathogen, Helicobacter hepaticus, and its association with chronic active hepatitis and liver tumors prompted an evaluation of the prevalence of H. hepaticus in commercially available mice. Of the 28 different strains or stocks, totaling 160 mice from four major commercial vendors, cultured for H. hepaticus, 100% of mice from two outbred strains from one vendor were infected with H. hepaticus, whereas 9 of 13 inbred mouse strains from another vendor were infected. This high prevalence of H. hepaticus established a need for a rapid and reproducible, noninvasive assay for the screening of colony-maintained mice being used for biomedical research. The culturing of fecal material by using 0.45-microns-pore- size filtration for H. hepaticus consistently yielded reproducible results but required extended periods of time. (1 to 3 weeks) to obtain a definitive answer. Although it is rapid, the use of a direct PCR-based detection assay with fecal specimens is restricted by inhibitory agents. to circumvent these inhibitory agents and to augment our H. hepaticus culture technique, we have developed a novel PCR system in which the bacteria are isolated from fecal material in the presence of polyvinylpyropyrollidone and lysed by treatment with Chelex 100. The PCR is performed with Tth polymerase supplemented with a polymerase enhancer. By this PCR method, 24 H. hepaticus culture-positive and 30 H. hepaticus culture-negative fecal samples were correctly identified. Moreover, two samples which were PCR positive and culture negative initially were positive by both methods upon retesting of fresh material. Southern blot hybridizations and sequencing of PCR products showed them to be H. hepaticus specific. A comparison of results obtained under identical conditions indicated a 100-fold increase in sensitivity with Tth polymerase over Taq polymerase. This PCR method can be used as a noninvasive means of rapidly screening large numbers of colony mice for H. hepaticus.
Explore the source record for details and available documents.
The use of lacto-phenol cotton blue (LPCB) stain for wet mount preparation of stools to demonstrate intestinal parasites by routine microscopy was evaluated in this study. LPCB-stained trophozoites and cysts and helminthic ova could easily be detected and identified in LPCB wet mounts of stools. The stain is recommended for routine use in the wet mount preparation of stools in a parasitology laboratory.
Explore the source record for details and available documents.