RESUMEN

Heat stress (HS) may result in economic losses to pig producers across the USA and worldwide. Despite significant advancements in management practices, HS continues to be a challenge. In this study, an in-feed antibiotic (carbadox, CBX) and antibiotic alternatives ( [XPC], and [SGX] fermentation products) were evaluated in a standard pig starter diet as mitigations against the negative effects of HS in pigs. A total of 100 gilts were obtained at weaning (6.87 ± 0.82 kg BW, 19.36 ± 0.72 d of age) and randomly assigned to dietary treatments (2 rooms/treatment, 2 pens/room, 6 to 7 pigs/pen). After 4 wk of dietary acclimation, half of the pigs in each dietary group (1 room/dietary treatment) were exposed to repeated heat stress conditions (RHS; daily cycles of 19 h at 25°C and 5 h at 40°C, repeated for 9 d), and the remaining pigs were housed at constant thermal neutral temperature (25°C, [NHS]). Pigs subjected to RHS had elevated skin surface temperature ( < 0.05; average 41.7°C) and respiration rate ( < 0.05; 199 breaths per minute (bpm) during HS, and overall reduced ( < 0.05) BW, ADG, ADFI, and G:F regardless of dietary treatment. Independent of diet, RHS pigs had significantly shorter ( < 0.05) jejunum villi on d 3 and d 9 compared to NHS pigs. Heat stress resulted in decreased villus height to crypt depth ratio (V:C) in pigs fed with control diet with no added feed additive (NON) and CBX diets at d 3, whereas the pigs fed diets containing XPC or SGX showed no decrease. Transcriptional expression of genes involved in cellular stress (, , , ), tight junction integrity (, , ), and immune response (, , and ) were measured in the ileum mucosa. Pigs in all dietary treatments subjected to RHS had significantly higher ( < 0.05) transcript levels of and , and an upward trend ( < 0.07) of mRNA expression. RHS pigs had higher ( < 0.05) transcript levels of and in NON diet, in XPC and CBX diets, and in SGX diet compared to the respective diet-matched pigs in the NHS conditions. Neither RHS nor diet affected peripheral natural killer () cell numbers or NK cell lytic activity. In conclusion, pigs subjected to RHS had decreased performance, and supplementation with fermentation products in the feed (XPC and SGX) protected pigs from injury to the jejunum mucosa.

Asunto(s)

Alimentación Animal/análisis , Fenómenos Fisiológicos Nutricionales de los Animales , Suplementos Dietéticos , Porcinos/fisiología , Animales , Antibacterianos/farmacología , Dieta/veterinaria , Femenino , Fermentación , Calor/efectos adversos , Íleon/efectos de los fármacos , Mucosa Intestinal/efectos de los fármacos , Distribución Aleatoria , Estrés Fisiológico , Porcinos/inmunología , Destete

RESUMEN

The qseBC-encoded quorum-sensing system regulates the motility of Escherichia coli O157:H7 in response to bacterial autoinducer 3 (AI-3) and the mammalian stress hormones epinephrine (E) and norepinephrine (NE). The qseC gene encodes a sensory kinase that autophosphorylates in response to AI-3, E, or NE and subsequently phosphorylates its cognate response regulator QseB. In the absence of QseC, QseB downregulates bacterial motility and virulence in animal models. In this study, we found that 8- to 10-month-old calves orally inoculated with a mixture of E. coli O157:H7 and its isogenic qseBC mutant showed significantly higher fecal shedding of the qseBC mutant. In vitro analysis revealed similar growth profiles and motilities of the qseBC mutant and the parental strain in the presence or absence of NE. The magnitudes of the response to NE and expression of flagellar genes flhD and fliC were also similar for the qseBC mutant and the parental strain. The expression of ler (a positive regulator of the locus of enterocyte effacement [LEE]), the ler-regulated espA gene, and the csgA gene (encoding curli fimbriae) was increased in the qseBC mutant compared to the parental strain. On the other hand, growth, motility, and transcription of flhD, fliC, ler, espA, and csgA were significantly reduced in the qseBC mutant complemented with a plasmid-cloned copy of the qseBC genes. Thus, in vitro motility and gene expression data indicate that the near-parental level of motility, ability to respond to NE, and enhanced expression of LEE and curli genes might in part be responsible for increased colonization and fecal shedding of the qseBC mutant in calves.

Asunto(s)

Escherichia coli O157/crecimiento & desarrollo , Escherichia coli O157/fisiología , Proteínas de Escherichia coli/genética , Intestinos/microbiología , Percepción de Quorum , Animales , Derrame de Bacterias , Bovinos , Escherichia coli O157/efectos de los fármacos , Escherichia coli O157/genética , Heces/microbiología , Eliminación de Gen , Perfilación de la Expresión Génica , Locomoción , Norepinefrina/metabolismo

RESUMEN

The presence of E. coli O157:H7 in cattle illustrates the complex, interrelated nature of the environment, livestock production practices, food safety, and the science of microbiology, particularly microbial ecology. Enterohemorrhagic E. coli, including E. coli O157:H7, can cause severe human diseases that can be debilitating and life threatening. Cattle are currently considered to be the definitive source for E. coli O157:H7 in the food supply, but this view may be simplistic and incomplete. E. coli O157:H7, appears widespread among U.S. cattle herds, while individual animal prevalence is low and transient. Most individual animals appear to be a transient reservoir for E. coli O157:H7 although the issue of carrier animals still remains unresolved. Epidemiological studies of the cattle production system have not clearly identified risk factors or management practices that affect E. coli O157:H7 prevalence in cattle feces. The problem of E. coli O157:H7 increases during the summer and fall months, but the environmental factors that contribute to this increase are poorly understood. Possible environmental factors that may influence E. coli O157:H7 shedding in cattle include livestock feed and waste handling practices as well as insects and microbial interactions in soil and water. Studies of E. coli O157:H7 ecology in cattle and the environment have been limited, but they suggest that a consideration of other independent, environmental sources of this microbe seems appropriate. The natural ecology of cholera may serve as a useful environmental model for pursuing additional environmental research on the occurrence and transmission of E. coli O157:H7 in nature.

Asunto(s)

Enfermedades de los Bovinos/microbiología , Infecciones por Escherichia coli/veterinaria , Escherichia coli O157/patogenicidad , Animales , Bovinos , Reservorios de Enfermedades , Ecosistema , Microbiología Ambiental , Infecciones por Escherichia coli/microbiología , Eucariontes/microbiología , Microbiología de Alimentos , Humanos , Seguridad , Estados Unidos , Virulencia

RESUMEN

Methanogenic bacteria, which are common inhabitants of the animal digestive tract, contain the fluorescent compound F420 (coenzyme 420), a 7,8-didemethyl-8-hydroxy-5-deazariboflavin chromophore. F420 was characterized as an initial step in determining if this compound would be useful as a fluorescent marker for the detection of fecal and ingesta contamination. Using a single anion exchange chromatographic process, F420 was separated from other cell components of a Methanobrevibacter sp. cell culture. The extent of separation was determined spectroscopically. To aid in the development of possible techniques for the detection of fecal contamination using F420 as a marker, further spectroscopic investigation of F420 was conducted using steady-state and time-resolved fluorescence methods. The fluorescence lifetime of F420 in an elution buffer of pH 7.5 was found to be 4.2 ns. At higher pH values, the fluorescence decay, F(t), was best described by a sum of two exponentials: at pH 13, F(t) = 0.31 exp(-t/4.20 ns) + 0.69 exp(-t/1.79 ns). Further investigation using front-faced fluorescence techniques has shown that emission from F420 can be collected efficiently from samples of methanogen cell cultures as well as from fecal material.

Asunto(s)

Euryarchaeota/aislamiento & purificación , Heces , Contaminación de Alimentos/análisis , Microbiología de Alimentos , Riboflavina/análisis , Animales , Bovinos , Sistema Digestivo/microbiología , Euryarchaeota/química , Femenino , Riboflavina/análogos & derivados , Rumen/microbiología , Espectrometría de Fluorescencia/métodos , Espectrofotometría/métodos

RESUMEN

OBJECTIVE: To determine whether numbers of coliform bacteria in feces of dairy cattle changed during the periparturient period and whether fluctuations were associated with changes in dry-matter intake. ANIMALS: 12 healthy Holstein cows. PROCEDURE: Fecal samples were collected on a semi-regular basis (i.e., 3 to 7 times/wk) beginning 4 to 6 weeks before the anticipated parturition date and continuing through the third day (5 cows) or second week (7 cows) after parturition, and total numbers of fecal coliform bacteria were determined. Daily feed intake of 7 cows was monitored. RESULTS: For 11 cows, fecal coliform bacterial counts between 34 and 25 days prior to parturition were low and relatively constant (< 102 change in number of bacteria). Coliform bacteria were not detected in 4 to 8% of fecal samples from 10 cows. All cows had a 10(4) to 10(7) increase in number of colony forming units/g of feces near the time of parturition. Number of fecal coliform bacteria peaked within 7 days of parturition in 9 cows and within 12 days of parturition in 3. Number of fecal coliform bacteria was not correlated with feed intake. CONCLUSIONS AND CLINICAL RELEVANCE: Cows may have large increases in fecal coliform bacteria count during the periparturient period; however, periparturient cows do not continually shed high numbers of coliform bacteria, and coliform bacteria may not always be detectable by conventional culture methods. Changes in fecal coliform bacteria count did not correlate with changes in dry-matter intake.

Asunto(s)

Enterobacteriaceae/aislamiento & purificación , Heces/microbiología , Trabajo de Parto , Periodo Posparto , Preñez/fisiología , Animales , Bovinos , Industria Lechera , Femenino , Embarazo , Análisis de Regresión

RESUMEN

Shiga toxin-producing Escherichia coli (STEC) is an important cause of food-borne illness in humans. Ruminants appear to be more frequently colonized by STEC than are other animals, but the reason(s) for this is unknown. We compared the frequency, magnitude, duration, and transmissibility of colonization of sheep by E. coli O157:H7 to that by other pathotypes of E. coli. Young adult sheep were simultaneously inoculated with a cocktail consisting of two strains of E. coli O157:H7, two strains of enterotoxigenic E. coli (ETEC), and one strain of enteropathogenic E. coli. Both STEC strains and ETEC 2041 were given at either 10(7) or 10(10) CFU/strain/animal. The other strains were given only at 10(10) CFU/strain. We found no consistent differences among pathotypes in the frequency, magnitude, and transmissibility of colonization. However, the STEC strains tended to persist to 2 weeks and 2 months postinoculation more frequently than did the other pathotypes. The tendency for persistence of the STEC strains was apparent following an inoculation dose of either 10(7) or 10(10) CFU. One of the ETEC strains also persisted when inoculated at 10(10) CFU. However, in contrast to the STEC strains, it did not persist when inoculated at 10(7) CFU. These results support the hypothesis that STEC is better adapted to persist in the alimentary tracts of sheep than are other pathotypes of E. coli.

Asunto(s)

Infecciones por Escherichia coli/veterinaria , Escherichia coli O157/crecimiento & desarrollo , Escherichia coli O157/patogenicidad , Escherichia coli/crecimiento & desarrollo , Escherichia coli/patogenicidad , Ovinos/microbiología , Animales , Anticuerpos Antibacterianos/sangre , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/transmisión , Enfermedades de las Ovejas/microbiología , Enfermedades de las Ovejas/transmisión , Toxina Shiga I/inmunología , Toxina Shiga II/inmunología , Virulencia

RESUMEN

Semi-automated detection of Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 and non-O157:H7 Shiga toxin-producing E. coli (STEC) was achieved using fluorogenic polymerase chain reaction (PCR). These PCR assays were designed to amplify 80, 120 and 150 bp regions of virulence genes stx1, stx2 and eaeA, respectively, using specific primers. The fluorogenic probes were used for specific detection of amplified products of the stx1 and stx2 genes of STEC, and the eaeA gene of EHEC O157:H7. For multiplex PCR assay, the three sets of primers and fluorogenic probes were included in one reaction to simultaneously amplify and detect any of the three targeted virulence genes. In non-multiplex PCR assay, each of the three virulence genes was amplified and detected in independent reactions. The specificity of these assays was evaluated using suspensions of STEC and other bacterial species lacking stx1, stx2 and eaeA. The multiplex assay detected all STEC harbouring any combination of three virulence genes. Three non-multiplex PCR reactions identified types of Shiga toxin genes carried by a STEC and identified STEC as either EHEC O157:H7 or non-O157:H7 STEC. Sensitivity limits of these assays in beef and faeces inoculated with EHEC O157:H7 were 5.8 to 580 cfu and 1.2 to 1200 cfu, respectively. These assays can be completed within 8-10 h when performed simultaneously or within 13 h if the multiplex assay is used as an initial screen for detecting STEC and the non-multiplex assay is used for subsequent detection of stx1 and stx2 of STEC and eaeA of EHEC O157:H7

Asunto(s)

Adhesinas Bacterianas , Proteínas de la Membrana Bacteriana Externa/genética , Toxinas Bacterianas/genética , Proteínas Portadoras , Escherichia coli O157/aislamiento & purificación , Proteínas de Escherichia coli , Escherichia coli/aislamiento & purificación , Reacción en Cadena de la Polimerasa/métodos , Animales , Automatización , Bovinos , Cartilla de ADN , ADN Bacteriano/análisis , Escherichia coli/genética , Escherichia coli O157/genética , Heces/microbiología , Fluorescencia , Humanos , Carne/microbiología , Sensibilidad y Especificidad , Toxinas Shiga , Factores de Tiempo

RESUMEN

Toxin-producing Escherichia coli expressing F18 fimbriae colonizes the small intestines of weaned pigs and causes diarrhea, edema disease, or both. The F18 family is composed of two antigenic variants, F18ab and F18ac. Because many strains do not express F18 fimbriae in vitro, identification and differentiation of these two variants are difficult. Single-strand conformational polymorphism (SSCP) analysis is a rapid method for identifying genetic mutations and polymorphisms. The F18 major fimbrial subunit genes (fedA) of 138 strains were amplified by PCR, and genetic differences were detected by SSCP analysis. The SSCP analysis of the fedA gene differentiated F18ab+ strains from F18ac+ strains. Most strains classified as F18ab+ by SSCP analysis contained Shiga toxin 2e and enterotoxin genes. Most strains classified as F18ac+ by SSCP analysis contained only enterotoxin genes. The SSCP analysis was a useful method for predicting the antigenicity of F18+ E. coli and could also be used for analysis of other virulence genes in E. coli and other pathogenic bacteria.

Asunto(s)

Proteínas Bacterianas/metabolismo , Proteínas de Escherichia coli , Escherichia coli/clasificación , Escherichia coli/genética , Proteínas Fimbrias , Fimbrias Bacterianas/genética , Polimorfismo Conformacional Retorcido-Simple , Secuencia de Aminoácidos , Animales , Antígenos Bacterianos/química , Antígenos Bacterianos/genética , Antígenos Bacterianos/inmunología , Proteínas Bacterianas/química , Proteínas Bacterianas/inmunología , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/veterinaria , Genes Bacterianos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Especificidad de la Especie , Porcinos , Enfermedades de los Porcinos/microbiología

RESUMEN

Two groups of calves were subjected to dietary stress by withholding of food beginning 1 or 14 days after inoculation with 10(10) CFU of Escherichia coli O157:H7. Following treatment, neither group had a significant increase in fecal shedding of E. coli O157:H7. A third group of calves had food withheld for 48 h prior to inoculation with 10(7) CFU of E. coli O157:H7. These calves were more susceptible to infection and shed significantly more E. coli O157:H7 organisms than calves maintained on a normal diet.

Asunto(s)

Bovinos/microbiología , Escherichia coli O157/aislamiento & purificación , Ayuno , Heces/microbiología , Animales

RESUMEN

The role of Escherichia coli heat-stable enterotoxin B (STb) in neonatal porcine diarrhea caused by enterotoxigenic E. coli was examined by comparing adherent isogenic strains with or without STb. The cloned STb gene (in the plasmid pRAS1) was electroporated into a nonenterotoxigenic strain (226M) which expresses the F41 adhesin. Strain 226M pRAS1 adhered and expressed STb in vivo, causing fluid secretion in ligated ileal loops in neonatal pigs. Although strain 226M pRAS1 caused very mild diarrhea in some orally inoculated neonatal pigs, the weight loss in these pigs was similar to that caused by the parental strain without STb. We conclude that STb does not significantly contribute to diarrhea caused by enterotoxigenic E. coli in neonatal pigs.

Asunto(s)

Toxinas Bacterianas/toxicidad , Diarrea/etiología , Enterotoxinas/toxicidad , Escherichia coli/patogenicidad , Animales , Animales Recién Nacidos , Adhesión Bacteriana , Proteínas de Escherichia coli , Íleon/microbiología , Porcinos

RESUMEN

AIM: To test the efficacy and safety of recombinant human epidermal growth factor (hEGF) on corneal re-epithelialisation following penetrating keratoplasty. METHODS: A prospective, randomised, placebo controlled study was carried out in which patients were matched for diagnosis and received either hEGF ophthalmic solution (30 micrograms/ml or 100 micrograms/ml) or placebo in a double masked fashion. Matched pairs of patients received donor corneas from the same donor and were operated by the same surgeon on the same day. At the end of surgery all donor epithelium was removed mechanically. Patients were examined twice daily and fluorescein stained photographs were taken until the epithelium had closed. The area of the defect was measured by planimetry of the fluorescein stained defect on the photographs. RESULTS: There were no significant differences in re-epithelialisation of the donor cornea between the placebo group and the group treated with 30 micrograms/ml hEGF. Time until complete closure was slightly longer with 100 micrograms/ml hEGF compared with 30 micrograms/ml hEGF and with placebo. Mean healing rate of the epithelial defect with 100 micrograms/ml hEGF was significantly slower than in the other groups. CONCLUSION: No significant acceleration of corneal re-epithelialisation was demonstrated with the use of recombinant hEGF after penetrating keratoplasty in humans.

Asunto(s)

Endotelio Corneal/efectos de los fármacos , Factor de Crecimiento Epidérmico/administración & dosificación , Queratoplastia Penetrante , Cicatrización de Heridas/efectos de los fármacos , Administración Tópica , Adulto , Método Doble Ciego , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Complicaciones Posoperatorias , Estudios Prospectivos , Factores de Tiempo

RESUMEN

Comparative fimbrial expression and adhesion studies were made on enterotoxigenic and verotoxigenic E. coli (ETEC and VTEC) strains isolated from cases of porcine postweaning diarrhoea or oedema disease. F107(F18ab) fimbriae--monitored by polyclonal and monoclonal antibodies and by electron microscopy--were poorly expressed on most VTEC strains. In contrast, 2134P(F18ac) fimbriae were more readily detected on most ETEC strains. The F18ac strains adhered in vivo to ligated intestinal loops in weaned pigs while the F18ab strains did not adhere or adhered weakly. Similarly, the F18ac strains adhered to isolated intestinal brush borders in weaned pigs but the F18ab strains (except for the F107 reference E. coli) did not adhere or adhered weakly in vitro. Neither the F18ab nor F18ac strains adhered to brush borders from newborn pigs. In vitro adhesion of F18ab and F18ac strains was mannose resistant and receptors for F18 seemed to differ from receptors for K88(F4). It is concluded that the antigenic variants of F18 fimbriae (F18ab and F18ac) are biologically distinct. F18ab fimbriae are expressed poorly both in vitro and in vivo and are frequently linked with the production of SLT-IIv and serogroup O139, while F18ac are more efficiently expressed in vitro and in vivo and most often are linked with enterotoxin (STa, STb) production, and serogroups O141, O157.

Asunto(s)

Diarrea/microbiología , Diarrea/veterinaria , Edematosis Porcina/microbiología , Infecciones por Escherichia coli/inmunología , Infecciones por Escherichia coli/microbiología , Escherichia coli/inmunología , Fimbrias Bacterianas/metabolismo , Animales , Anticuerpos Monoclonales/inmunología , Antígenos Bacterianos/análisis , Adhesión Bacteriana , Toxinas Bacterianas/química , Bioensayo , Células Cultivadas , Chlorocebus aethiops , Sondas de ADN/genética , Escherichia coli/genética , Escherichia coli/ultraestructura , Fimbrias Bacterianas/inmunología , Fimbrias Bacterianas/ultraestructura , Técnica del Anticuerpo Fluorescente Directa , Células HeLa , Humanos , Íleon/microbiología , Íleon/ultraestructura , Manosa/farmacología , Microscopía Electrónica , Microscopía Inmunoelectrónica , Hibridación de Ácido Nucleico , Reacción en Cadena de la Polimerasa , Receptores de Superficie Celular/inmunología , Porcinos , Células Vero

RESUMEN

We have previously identified two K88ac adhesion receptors (210 and 240 kDa) which are present in membrane preparations from adhesive but not nonadhesive porcine intestinal brush border cells; these adhesin receptors are postulated to be important determinants of the susceptibility of pigs to K88ac+ enterotoxigenic Escherichia coli infections (A.K. Erickson, J.A. Willgohs, S.Y. McFarland, D.A. Benfield, and D.F. Francis, Infect. Immun. 60:983-988, 1992). We now describe a procedure for the purification of these two receptors. Receptors were solubilized from adhesive intestinal brush border vesicles using deoxycholate and were purified by gel filtration chromatography on Sepharose CL-4B and then by hydroxyapatite chromatography. Amino acid compositional analyses indicated that the two receptors have similar amino acid compositions. The most distinguishing characteristic of both receptors is a high percentage of threonine and proline residues. Neuraminidase treatment caused the K88ac adhesin receptors to migrate with a slower mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels, indicating that these receptors are sialoglycoproteins. Results from lectin-binding studies indicated that the receptors contain O-linked oligosaccharides composed of galactosyl (beta-1,3)N-acetylgalactosamine, alpha-linked fucose, galactosyl(beta-1,4)N-acetylglucosamine, sialic acid, galactose, and N-acetylgalactosamine. Collectively, these characteristics indicate that the K88ac adhesin receptors are mucin-type sialoglycoproteins.

Asunto(s)

Antígenos Bacterianos , Proteínas de Escherichia coli , Escherichia coli/inmunología , Proteínas Fimbrias , Intestinos/química , Mucinas/química , Receptores Inmunológicos/química , Sialoglicoproteínas/química , Aminoácidos/análisis , Animales , Antígenos de Superficie/inmunología , Adhesión Bacteriana/fisiología , Secuencia de Carbohidratos , Carbohidratos/química , Fimbrias Bacterianas/inmunología , Lectinas/metabolismo , Microvellosidades/química , Datos de Secuencia Molecular , Receptores Inmunológicos/aislamiento & purificación , Sialoglicoproteínas/clasificación , Porcinos

RESUMEN

Mycobacteria are unusual causes of keratitis and other ocular infections but the outcome of infection is often serious. We report a case of keratitis due to Mycobacterium chelonae, a rapidly growing environmental mycobacterium, in a soft contact-lens wearer, and discuss the difficulty and delay in identifying the organism, twice erroneously identified as Nocardia asteroides on morphological grounds. Despite in vitro susceptibility, the response to anti-bacterial agents was negligible and a second keratoplasty was required after a recurrence of disease at the donor-host junction. We review the role of mycobacteria as the cause of keratitis and other forms of ocular disease.

Asunto(s)

Infecciones Bacterianas del Ojo/diagnóstico , Infecciones Bacterianas del Ojo/microbiología , Queratitis/microbiología , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Mycobacterium chelonae , Enfermedades Orbitales/microbiología , Adulto , Lentes de Contacto de Uso Prolongado/efectos adversos , Córnea/microbiología , Infecciones Bacterianas del Ojo/terapia , Femenino , Humanos , Queratitis/terapia , Infecciones por Mycobacterium no Tuberculosas/terapia , Mycobacterium chelonae/aislamiento & purificación

RESUMEN

Pasteurella haemolytica is the leading cause of economic loss to the beef cattle industry in the United States and an important etiologic agent worldwide. Study of P. haemolytica is hindered by researchers' inability to genetically manipulate the organism. A new restriction endonuclease, PhaI, an isoschizomer of SfaNI (R. J. Roberts, Methods Enzymol. 65:19-36, 1980), was isolated from P. haemolytica serotype 1, strain NADC-D60, obtained from pneumonic bovine lung. PhaI recognizes the 5-base nonpalindromic sequences 5'-GCATC-3' and 5'-GATGC-3'. Cleavage occurs 5 bases 3' from the former recognition site and 9 bases 5' from the latter recognition site. A gene encoding a methyltransferase which protects against PhaI cleavage was cloned from P. haemolytica NADC-D60 into Escherichia coli. Whereas unmethylated plasmid DNA containing a P. haemolytica origin of replication was unable to transform P. haemolytica when introduced by electroporation, the same plasmid DNA obtained from E. coli which contained a cloned PhaI methyltransferase gene could do so. The data indicate that PhaI is an effective barrier to the introduction and establishment of exogenous DNA in P. haemolytica.

Asunto(s)

Enzimas de Restricción del ADN/metabolismo , Genes Bacterianos , Mannheimia haemolytica/enzimología , Metiltransferasas/genética , Transformación Bacteriana/genética , Secuencia de Bases , Clonación Molecular , Enzimas de Restricción del ADN/genética , Enzimas de Restricción del ADN/aislamiento & purificación , ADN Bacteriano/genética , Escherichia coli/genética , Metiltransferasas/biosíntesis , Datos de Secuencia Molecular

RESUMEN

We investigated the role of the rumen fermentation as a barrier to the foodborne pathogen, Escherichia coli O157:H7. Strains of E. coli, including several isolates of O157:H7, grew poorly in media which simulated the ruminal environment of a well-fed animal. Strains of E. coli O157:H7 did not display a superior tolerance to ruminal conditions which may facilitate their colonization of the bovine digestive tract. Unrestricted growth of E. coli was observed in rumen fluid collected from fasted cattle. Growth was inhibited by rumen fluid collected from well-fed animals. Well-fed animals appear less likely to become reservoirs for pathogenic E. coli. These results have implications for cattle slaughter practices and epidemiological studies of E. coli O157:H7.

Asunto(s)

Bovinos/microbiología , Escherichia coli/crecimiento & desarrollo , Contenido Digestivo/microbiología , Rumen/microbiología , Anaerobiosis , Animales , Medios de Cultivo , Escherichia coli/efectos de los fármacos , Escherichia coli/patogenicidad , Ácidos Grasos Volátiles/metabolismo , Ácidos Grasos Volátiles/farmacología , Privación de Alimentos , Concentración de Iones de Hidrógeno , Virulencia

RESUMEN

Fimbriae (pili) of enterotoxigenic Escherichia coli (ETEC), including K88, K99, 987P, and F41, are adhesins that facilitate intestinal colonization in neonatal pigs. K88 is also associated with some ETEC isolated from weaned pigs. Many ETEC isolates from weaned pigs do not express known adhesins and are termed 4P-. A novel bacterial adhesin, 2134P, was recently identified on two 4P- ETEC isolates from weaned pigs. In this study, we identified a 2134P-specific monoclonal antibody, mAb 6C7/C1, that blocked the binding of 2134P+ bacteria to intestinal epithelial cells. Indirect immunofluorescent antibody and immunoperoxidase assays using mAb 6C7/C1 confirmed that the 2134P adhesin is expressed in vivo by adherent bacteria in pigs challenge-exposed with 2134P+ ETEC. 2134P was detected on 31% of 189 postweaning diarrhea 4P- ETEC isolates from the National Animal Disease Center's culture collection by dot blot immunoperoxidase assays using mAb 6C7/C1. We conclude that 2134P is a bacterial adhesin and is an important virulence attribute of some ETEC that cause diarrhea in weaned pigs.

Asunto(s)

Antígenos Bacterianos/análisis , Adhesión Bacteriana/fisiología , Proteínas de la Membrana Bacteriana Externa/análisis , Diarrea/veterinaria , Infecciones por Escherichia coli/veterinaria , Fimbrias Bacterianas/fisiología , Enfermedades de los Porcinos/microbiología , Adhesinas de Escherichia coli , Animales , Anticuerpos Monoclonales , Adhesión Bacteriana/inmunología , Técnicas de Cultivo , Diarrea/microbiología , Escherichia coli/inmunología , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/epidemiología , Infecciones por Escherichia coli/microbiología , Fimbrias Bacterianas/inmunología , Microvellosidades/microbiología , Porcinos , Destete

RESUMEN

A DNA probe derived from the subunit gene of a cloned 987P determinant was used to characterize the locations of the 987P genes in several Escherichia coli strains. We examined E. coli 987, a nonpiliated mutant of strain 987 (I36) that does not express 987P in vitro, and a variant of I36 that expressed 987P following growth in vivo. We found that plasmid and chromosomal copies of the 987P subunit gene could be differentiated in strain 987 by restriction endonuclease analysis and Southern blot hybridization. The nonpiliated mutant I36 had lost the plasmid copy but retained the chromosomal copy of the 987P gene. A 987P-piliated variant of I36, which did not contain the 987P plasmid, colonized and caused diarrhea in neonatal pigs similarly to wild-type 987. The plasmid that hybridized with the 987P probe was transferred from strain 987 to an E. coli K-12 strain by conjugation. We were unable to demonstrate expression of 987P by these transconjugants. The data suggest that the chromosomal and plasmid copies of the 987P genes may interact to influence 987P expression.

Asunto(s)

Proteínas de la Membrana Bacteriana Externa/genética , Escherichia coli/genética , Fimbrias Bacterianas , Adhesinas de Escherichia coli , Animales , Animales Recién Nacidos , Mapeo Cromosómico , Diarrea/microbiología , Escherichia coli/patogenicidad , Regulación Bacteriana de la Expresión Génica , Genes Bacterianos , Plásmidos , Mapeo Restrictivo , Porcinos

RESUMEN

We report a retrospective analysis of the clinical indications for 3555 penetrating keratoplasties performed at our department between 1971 and 1990. The cases were distributed among 12 diagnostic categories. Regrafting was the most common indication overall, accounting for 1452 cases (40.8%). Other major indications were, in order of decreasing frequency, keratoconus (17%), scarring secondary to herpes simplex keratitis (11.7%), aphakic bullous keratopathy (5.9%) and interstitial keratitis (5%). Further analysis of the relative percentages in each category within each 5-year interval of the study period was carried out to identify any changes in incidence. Viral disease as an indication for penetrating keratoplasty has shown a gradual decrease in frequency, accounting for only 6.4% of the cases during the last 5-year period (1986-90) compared with 19.6% during the first 5 years (1971-75). This finding is consistent with the marked improvement in the recognition and medical treatment of herpes simplex keratitis. The increase in incidence of grafting for pseudophakic bullous keratopathy in 1986-90 (6.7%) compared with 1981-85 (1.1%) correlates well with the dramatic increase in the number of cataract extractions with intraocular lens implantations performed during that period.

Asunto(s)

Queratoplastia Penetrante/estadística & datos numéricos , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Preescolar , Femenino , Rechazo de Injerto/cirugía , Humanos , Lactante , Queratitis/cirugía , Queratitis Herpética/cirugía , Queratocono/cirugía , Masculino , Persona de Mediana Edad , Reoperación , Estudios Retrospectivos , Factores de Tiempo

RESUMEN

Three-week-old weaned and colostrum-deprived neonatal (less than 1 day old) pigs were inoculated to determine the pathogenicity of 2 enterotoxigenic Escherichia coli isolates that do not express K88, K99, F41, or 987P adhesins (strains 2134 and 2171). Strains 2134 and 2171 were isolated from pigs that had diarrhea after weaning attributable to enterotoxigenic E coli infection. We found that both strains of E coli adhered in the ileum and caused diarrhea in pigs of both age groups. In control experiments, adherent bacteria were not seen in the ileum of pigs less than 1 day old or 3 weeks old that were noninoculated or inoculated with a nonpathogenic strain of E coli. These control pigs did not develop diarrhea. Antisera raised against strains 2134 and 2171 and absorbed with the autologous strain, grown at 18 C, were used for bacterial-agglutination and colony-immunoblot assays. Both absorbed antisera reacted with strains 2134 and 2171, but not with strains that express K99, F41, or 987P adhesins. A cross-reaction was observed with 2 wild-type K88 strains, but not with a K12 strain that expresses K88 pili. Indirect immunofluorescence with these absorbed antisera revealed adherent bacteria in frozen sections of ileum from pigs infected with either strain. We concluded that these strains are pathogenic and express a common surface antigen that may be a novel adhesin in E coli strains that cause diarrhea in weaned pigs.

Asunto(s)

Diarrea/veterinaria , Enterotoxinas/biosíntesis , Infecciones por Escherichia coli/veterinaria , Escherichia coli/patogenicidad , Enfermedades de los Porcinos/microbiología , Adhesinas de Escherichia coli , Animales , Animales Recién Nacidos , Antígenos Bacterianos/biosíntesis , Antígenos Bacterianos/genética , Antígenos de Superficie/biosíntesis , Antígenos de Superficie/genética , Adhesión Bacteriana , Proteínas de la Membrana Bacteriana Externa/biosíntesis , Proteínas de la Membrana Bacteriana Externa/genética , Diarrea/microbiología , Enterotoxinas/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Infecciones por Escherichia coli/microbiología , Técnica del Anticuerpo Fluorescente , Regulación Bacteriana de la Expresión Génica , Íleon/microbiología , Íleon/ultraestructura , Microscopía Electrónica , Porcinos , Destete