Caracterización de cepas clínicas y ambientales de Salmonella enterica subsp. enterica serovar Heidelberg aisladas en Chile / Characterization of Salmonella Heidelberg strains isolated in Chile

Background: Salmonella Heidelberg (S. Heidelberg) causes gastroenteritis and sometimes bacteremia and endocarditis. In other countries, this serovar has multidrug resistance including extended-spectrum β-lactamases (ESBLs) and AmpC (β-lactamases (AmpC), associated with the blaCMY-2 gene. In Chile, an outbreak by S. Heidelberg occurred in 2011, the phenotypic and genetic characteristics of Chilean strains are unknown. Aim: To determine the antimicrobial susceptibility, presence of plasmids and virulence factor genes in S. Heidelberg strains isolated in Chile over the period 2006-2011. Material and Methods: In sixty-one S. Heidelberg clinical and environmental strains collected by the Public Health Institute in Chile during 2006-2011, antimicrobial susceptibility, plasmids and virulence factor genes (invA, sifA, pefA, agfA, lpfA and, stkD) were studied. Results: S. Heidelberg had a high susceptibility to sulfamethoxazole-trimethoprim, gentamicin, ceftriaxone, ceftiofur, chloramphenicol, amoxicillin-clavulanic acid and ampicillin. However, 52% had decreased susceptibility to ciprofloxacin and 33% resistance to tetracycline. ESBLs were detected in three strains isolated from blood cultures, environment and human feces. The latter strain was positive for AmpC and blaCMY-2 gene. Fifty three of 61 strains showed one to seven plasmids of 0.8 to approximately 30 kb. Most plasmids were small with sizes between 0.8 and 2 kb. All isolates were positive for all genes except pefA. Conclusions: S. Heidelberg isolated from Chilean samples was susceptible to first-line antimicrobials, except tetracycline and ciprofloxacin. The emergence of strains with ESBLs and AmpC should be a warning. The strains were homogeneous for virulence genes, but heterogeneous in their plasmids.

Rapid, specific, and sensitive detection of the ureR_1 gene in Klebsiella pneumoniae by loop-mediated isothermal amplification method

Klebsiella pneumoniae is one of the main pathogenic bacteria that causes nosocomial infections, such as pneumonia, urinary tract infection, and sepsis. Therefore, the rapid and accurate detection of K. pneumoniae is important for the timely treatment of infectious patients. This study aimed to establish a loop-mediated isothermal amplification (LAMP) method for the rapid and sensitive detection of K. pneumoniae-specific gene ureR_1 (Gene ID: 11847803). The ureR_1 gene was obtained through local and online BLAST, and the specific primers were designed for its detection. Positive reactions were observed on all 140 K. pneumoniae clinical isolates while all the 82 non-K. pneumoniae clinical isolates were negative. Plasmids with the specific gene and the mouse blood with K. pneumoniae were used for sensitivity analysis. The detection limit of the LAMP was 1 bacterium/reaction. The results showed that the LAMP targeted to ureR_1 is a fast, specific, sensitive, inexpensive, and suitable method for the detection of K. pneumoniae.

Virulence factors, antimicrobial resistance, and plasmid content of Escherichia coli isolated in swine commercial farms / Fatores de virulência, resistência aos antimicrobianos, presença de plasmídeos em Escherichia coli isoladas de amostras clínicas e ambientais de suínos

Virulence factors and antimicrobial resistance patterns of Escherichia coli isolates were evaluated. A total of 80 E. coli isolates were evaluated, being 64 from clinical samples (intestinal content and fragments of organs from diarrheic piglets), seven from feces of clinically healthy piglets and sows, and nine environmental samples (five from facilities, two from feed, one from insect, and one from waste). Molecular characterization was performed by PCR detection of fimbriae and toxin genes and plasmid content determination. The isolates were also characterized according to their resistance or sensitivity to the following drugs: ampicillin, trimethoprim:sulfamethoxazole, tetracycline, amikacine, colistin, norfloxacin, florfenicol, enrofloxacin, cefalexin, trimethoprim, neomycin, chloramphenicol, and gentamicin. From 80 E. coli isolates, 53.8 percent were classified as enterotoxigenic E. coli (ETEC), 2.5 percent were shiga toxin-producing E. coli (STEC), and 43.8 percent showed a non specific pattern and were unclassified. One fecal isolate from non-diarrheic piglet was classified as ETEC by PCR. Clinical isolates showed resistance mainly for tetracycline and trimethoprim:sulfamethoxazole. Plasmidial DNA was observed in 70 isolates, being 78.5 percent of clinical isolates, 8.57 percent of non-diarrheic feces, and 12.8 percent of environment.

Os fatores de virulência e a resistência aos antimicrobianos foram avaliados em Escherichia coli. Um total de 80 isolados de E. coli, sendo 64 de amostras clínicas (conteúdo intestinal e fragmentos de órgãos de leitões diarreicos), sete das fezes de porcas e leitões saudáveis e nove de amostras ambientais (cinco de instalações, dois de alimentos, um de inseto e um de esterqueira). A caracterização molecular feita pela PCR objetivou detectar fimbrias e toxinas, bem como a determinação do conteúdo de plasmídeos. Os isolados foram caracterizados quanto à resistência ou sensibilidade às seguintes drogas: ampicilina, sulfazotrim, tetraciclina, amikacina, colistina, norfloxacina, florfenicol, enrofloxacina, cefalexina, trimetoprim, neomicina, cloranfenicol e gentamicina. Dos 80 isolados, 53,8 por cento foram classificados como E. coli enterotoxigênica (ETEC), 2,5 por cento como E. coli produtora de shiga toxina (STEC) e 43,8 por cento, por não apresentarem padrão específico, não foram classificadas. Pela PCR, um isolado de fezes de suíno sem diarreia foi classificado como ETEC. Os isolados das amostras clínicas foram principalmente resistentes à tetraciclina e à sulfazotrim. Em 70 isolados, observaram-se DNA plasmidial, destes 78,5 por cento foram obtidos de amostras clínicas, 8,57 por cento de leitões sadios e 12,8 por cento de amostras ambientais.

Virulence factors and antimicrobial resistance of Escherichia coli isolated from urinary tract of swine in southern of Brazil / Fatores de virulência e resistência aos antimicrobianos de Escherichia coli isoladas do trato urinário de suínos do Sul do Brasil

The present study determined the molecular and resistance patterns of E. coli isolates from urinary tract of swine in Southern of Brazil. Molecular characterization of urinary vesicle samples was performed by PCR detection of virulence factors from ETEC, STEC and UPEC. From a total of 82 E. coli isolates, 34 (38.63 percent) harbored one or more virulence factors. The frequency of virulence factors genes detected by PCR were: pap (10.97 percent), hlyA (10.97 percent), iha (9.75 percent), lt (8.53 percent), sta (7.31 percent) sfa (6.09 percent), f4 (4.87 percent), f5 (4.87 percent), stb (4.87 percent), f6 (1.21 percent) and f41 (1.21 percent). Isolates were resistant to penicillin (95.12 percent), lincomycin (93.9 percent), erythromycin (92.68 percent), tetracycline (90.24 percent), amoxicillin (82.92 percent), ampicillin (74.39 percent), josamycin (79.26 percent), norfloxacin (58.53 percent), enrofloxacin (57.31 percent), gentamicin (39.02 percent), neomycin (37.8 percent), apramycin (30.48 percent), colistine (30.48 percent) and cefalexin (6.09 percent). A number of 32 (39.02 percent) E. coli isolates harbored plasmids.

O presente estudo teve por objetivo determinar os padrões moleculares e de resistência aos antimicrobianos de isolados de E. coli provenientes do trato urinário de suínos no Sul do Brasil. Os fatores estudados dividiram os patotipos ETEC, STEC e UPEC. Trinta e quatro (38,63 por cento) isolados avaliados apresentavam um ou mais dos fatores de virulência pesquisados. A freqüência dos genes de virulência detectados foram: pap (10,97 por cento), hlyA (10,97 por cento), iha (9,75 por cento), lt (8,53 por cento), sta (7,31 por cento) sfa (6,09 por cento), f4 (4,87 por cento), f5 (4,87 por cento), stb (4,87 por cento), f6 (1,21 por cento) e f41 (1,21 por cento). Os isolados foram resistentes à penicilina (95,12 por cento), lincomicina (93,9 por cento), eritromicina (92,68 por cento), tetraciclina (90,24 por cento), amoxacilina (82,92 por cento), ampicilina (74,39 por cento), josamicina (79,26 por cento), norfloxacina (58,53 por cento), enrofloxacina (57,31 por cento), gentamicina (39,02 por cento), neomicina (37,8 por cento), apramicina (30,48 por cento), colistina (30,48 por cento) e cefalexina (6,09 por cento). Trinta e dois (39,02 por cento) isolados de E. coli continham plasmídeos.

Aislamiento de plásmidos conjugativos en cepas de enterobacterias productoras de ß-lactamasas de espectro expandido / Isolation of conjugative plasmids in enterobacteriaceae strains producing beta-lactamases expanded sprectrum

Las ß-lactamasas de espectro expandido son enzimas capaces de hidrolizar el enlace amida de los oximino ß-lactámicos, (cefotaxime, ceflazidime y aztreonam). Las mismas son codificadas en plásmidos y por lo tanto pueden ser transferidas mediante conjugación a diversos géneros bacterianos. Diseminándose ampliamente en el ambiente hospitalario. En esta investigación se persigue, en cepas de enterobacterias aislar plásmidos que codifican para ß-lactamasas de espectro expandido y que sean capaces de transferirse mediante conjugación. Se estudio una población de 51 enterobacterias productoras de ß-lactamasas de espectro expandido aisladas de diferentes centros hospitalarios del área Metropolitana de Caracas. A las mismas se le determinó el perfil de resistencia a múltiples antibióticos mediante la metodología de Kirby-Bauer. Se detectaron las BLEE mediante dos ensayos fenotípicos basados en el efecto sinergístico con el ácido clavulánico y se tipificaron molecularmente por PCRIRFLP, seguidamente se transfirieron los plásmidos conjugativos en ensayos de conjugación en medio sólido y se aislaron los plásmidos de cepas donantes y transconjugautes, por el método de lisis alcalina. Los resultados fenotípicos indican una mayor proporción de BLEE con actividad ceflazidimasa y en menor grado actividad cefotaximasa. La tipificación molecular indicó que 60,8 por ciento de las cepas portan genes tipo SHV y 15,6 por ciento codifican ß-lactamasas de espectro expandido de la familia CTX-M. De 36 cepas conjugadas un 81 por ciento transfirió material plasmídico. El análisis de los aislamientos plasmídicos mostró la presencia en la totalidad de las transcojugantes de una banda de 25000 pb y en un 80 por ciento se evidenció una banda plasmídica mayor a 50000 pb. Se pudo constatar la cotransferencia de resistencia a otras familias de antibióticos.

Fragmentos de tropomiosina - estudos da estabilidade conformacional e da interação cabeça-cauda / Fragments of tropomyosin - studies of the conformational stability and of the head-tail interactions

A Tropomiosina (TM) está diretamente envolvida no processo de regulação da contração muscular, que é controlado por um mecanismo alostérico que envolve `Ca²+', troponina (Tn), actina (Ac) e miosina. A Tm é uma proteína flexível, de estrutura ®coled-coil¼, constituída de duas `alfaï- hélices com 284 aminoácidos cada uma. A molécula de Tm faz interações do tipo ®cabeça-cauda¼ com outra molécula de Tm através da sobreposição de aproximadamente 8 a 15 resíduos da extremidade N- terminal de uma molécula com 8 a 15 resíduos da extremidade C-terminal da outra molécula de Tm. Desta maneira, em baixas forças iônicas, formam-se filamentos lineares através de um processo de polimerização...

Expressão de fatores peptídicos de crescimento e regulação do ciclo celular em células adrenocorticais / Expression of peptide factors of growth and regulation of cellular cycle in adrenocortical cells

ACTH (hormônio adrenocorticotrópico) e fatores peptídicos de crescimento regulam a proliferação celular em células adrenocorticais. Resulatdos preliminares de ensaios biológicos, cromatografia de heparina-sefarose e ®Western blotting¼ indicaram a produção de fatores peptídicos de crescimento das famílias de PDGF e de FGF na linhagem celular adrenocortical Y-1. No entanto a relação entre o estímulo promovido por ACTH e a expressão de fatores peptídicos de crescimento e sua contribuição para o controle do ciclo celular em células adrenocorticais ainda não estão definidas. Os principais objetivos deste projeto consistiram em 1. Detectar e caracterizar os fatores peptídicos de crescimento e seus correspondentes receptores expressos em células adrenorticais; 2. Investigar a regulação da expressão de fatores peptídicos de crescimento sob o estímulo de ACTH; 3. Definir os mecanismos de ação para os fatores peptídicos de crescimento localmente expressos e sua contribuição na regulação do ciclo celular...

Transposon [Tn5] and UV Irradiation as mutagenic agents for improving photorhabdus sp. Sinai 1 [S1], an insect - Pathogenic bacterium

A local isolate of symbiotic bacterium, Photorhabdus sp. Sinai 1 [S1] was isolated from Heterorhabditis sp. Sl insect-pathogenic nematodes and its character was detected. Antibacterial/antifungal antibiotic[s] and proteinase productions of Photorhabdus Sl play an important role in killing insects. Photorhabdus sp. [Sl] was treated with UV for different times. Survival percentage of the original strain dropped from 100% at zero time to 11.13%,0.23 and 0.022% after UV treatment for 1, 3 and 5 minutes, respectively. However, 32 mutants were isolated, 18 after 1 minute treatment and 14 after 3 minutes treatment. Wide ranges of proteinase production capacities were detected. Eight of the mutants isolated after UV treatment for 1 minute had lower production than Sl, while 4 mutants had the same productivity as Sl. Six mutants of l minute UV treatment, were higher producers than S1. Fourteen mutants were isolated after UV treatment for 3 minutes. Three of them had the same proteinase production at S1, 3 had lower production, and about 8 proved to be highly producer mutants. Mutant number 20 from UV 3 minutes treatment group proved to be the best producer. It produced 25% over than S1. Suicidal plasmid pSUP2021 was mobilized from Escherichia coli to photorhabdus bytransconjugation and 15 transconjugants were isolated. All of them proved electrophoritically to have the plasmid. Fourteen transconjugants were tested for their proteinase productivity, only two transconjugants [14%] were higher proteinase producers, one of them reached 133.3% in comparison with S1. All transconjugants, except one were higher antibacterial antibiotic producers thanS1. Transconjugant number 7 proved to be the best producer one. It produced about 150% antibacterial and 160% antifungal antibiotics in comparison with S1parental strain. Results also indicated that, Photorhabdus S1 mutations due to transposon Tn5 mutagenesis were more powerful than those induced by UV in proteinase production. Transposon Tn5 mutagenesis is one of the most important tool for antibiotic improvement in Photorhabdus

Construção e análise de mutantes fluorescentes da troponina I / Construction and analysis of Troponin I fluorescence mutants

A troponina (Tn) regula a contração do músculo estriado esquelético de vertebrados. Ela é composta de três subunidades: troponina I (TnI), troponina C (TnC) e troponina T (TnT). A TnI tem a função inibitória que é neutralizada pela ligação de Ca²+ nos sítios regulatórios do N-domínio da TnC, e a TnT posiciona o complexo no filamento fino. Para monitorar o sinal do Ca²+ sendo transmitido da TnC para a TnI as propriedades espectrais únicas do 5-hidroxitriptofano (5HW) foram utilizadas. O 5HW foi incorporado em mutantes pontuais da TnI com um único códon para triptofano. Foram identificadas duas sondas espectrais intrínsicas na TnI capazes de detectar a ligação de Ca²+ na Tn: as TnIs com 5HW nas posições 100 e 121...

Marcadores de virulência em Aeromonas spp isoladas de materiais clínicos humanos e de água doce / Virulence markers in Aeromonas spp isolated from human clinical materials and fresh water

Foram estudadas 88 cepas de Aeromonas isoladas de materiais clínicos de humanos e de água doce. Dessas, 31 foram isoladas de fezes diarréicas, 10 de fezes näo diarréicas, 39 de material humano extra-intestinal e oito de água. Essas cepas foram estudadas quanto ao perfil de virulência. Foram submetidas aos seguintes testes: adesäo e invasäo em células Hela, produçäo de citotoxina, enterotoxina, hemolisinas alfa e beta, auto-aglutinaçäo a 37ºC, resistência ao soro humano normal, absorçäo do Vermelho Congo, isolamento e extraçäo de plasmídeo. Os marcadores de virulência foram caracterizados em proporçöes variadas: 68,2 por cento aderiram e 45,5 por cento invadiram células HeLa, 36,4 por cento produziram citotoxina, 31,8 por cento enterotoxina, 100 por cento produziram hemolisina ß e 34,1 por cento produziram hemolisina alfa quando semeadas em profundidade nas placas de ágar sangue, 73,9 por cento produziram hemolisina ß e 47,7 por cento hemolisina alfa em microplacas, 17 por cento auto-aglutinaram a 37ºC, 31,8 por cento foram resistentes ao soro humano e 100 por cento absorveram o Vermelho Congo. Das cepas estudadas, 8 por cento apresentaram um ou mais plasmídeos de tamanho variado. Conclui-se pela pouca correlaçäo entre fatores de virulência e tipo de material clínico com exceçäo da citotoxina que foi encontrada com maior freqüência em cepas isoladas de infecçöes extra-intestinais e a enterotoxina mais comum em fezes diarréicas.

Homology among extra-cryptic DNA bands and the typical plasmids in brazilian Yersina pestis strains

Yersina pestis, the etiologic agent of plague, harbors three well-characterized plasmids: pFra (90-110kb), pYV (70kb) and pPst (9.5kb). Furthermore, some extra-cryptic DNA bands have been observed in a number of wild strains from several foci of the world. Additional bands have also been reported in Brazilian strains. Looking for any relationship among these cryptic DNA bands and the three-prototypical plasmids, we analyzed twelve strains displaying different plasmid content. The DNA bands were hybridized by southern blot with probes directed at the genes cafl, lcrV and 'pla' located respectively on the plasmids pFra, pYV and pPst. The probes were constructed by PCR amplification and labeled with digoxigenin. The Pla probe hybridized with its target (pPst) and with bands of about 35 kb suggesting some homology among them. The Cafl probe hybridized with the target (pFra) as well as with higher bands. The LcrV also hybridized with the target (pYV) and both with the bands higher than pFra and the bands between pFra and pYV. These results suggest that the large-cryptic bands could represent some rearrangement, open circular or linearized forms of the pFras and pYV plasmids.

Serological and biochemical characteristics of virulence plasmid of Yersinia enterocolitica isolates from chicken in the Islamic Republic of Iran

Pathogenic Yersinia enterocolitica harbour plasmid that is essential for virulence. We studied the characteristics of virulence plasmid using serological, biochemical and bioassay tests in Y. enterocolitica isolates of chicken using plasmid curing. Plasmid-cured isogenic derivatives [2029c and 2150c] were obtained from two isolates of Y. enterocolitica [RTCC 2029 and RTCC 2150]. The results demonstrated that plasmid-bearing isolates [2029 and 2150] were human-serum-resistant when grown at 37 °C, but were sensitive when grown at 25 °C, whereas plasmid-cured isolates [2029c and 2150c] were sensitive when grown at both temperatures. Also autoagglutination, calcium-dependency tests and experimental infection in mice demonstrated that these phenotypes were associated with the virulence plasmid

Construction of a cassette for cloning and analysis of replicons / Construccion de un cassette para el clonamiento y analisis de replicones

The aim of this work was the construction of a cassette, i.e., a non-replicative molecule formed by linkage of an antibiotic resistance gene and a multiple cloning site. This cassette would allow the cloning and analysis of a wide range of replicons. The aac(6')-lc amikacin gene was isolated and ligated to the multiple clining site of the pUC18 vector. This construction was HindIII digested and cloned in the HindIII site of the vector. The resulting pHJ13 clone conferred to the recipient cells the ability to grow in presence of amikacin (cassette marker) and ampicillin (vector gene). By restriction analysis, the cassette orientation was established. Cassette versatility is provided by the presence of the unaltered multiple cloning site segment, and also because it allows sequencing of any replication origin inserted. Cassette funcionality was demonstrated by ligation to a replicative region of H plasmid pHH1457. Presence of the ori region from pHH1457 and the aac(6')-lc gene was confirmed in E. coli transformed clones. The incompatibility properties of the pHH1457 and its capability to replicate in a Poll defective strain were preserved in the pHJII14 construct. Currently, the amikacin cassette is being used in the characterization of H Complex plasmids.

El objetivo de este trabajo es la construcción de un cassette ­ molécula no replicativa ­ formada por un gen de resistencia a un antibiótico y una región de múltiple sitios de clonamiento. Este cassette permitirá el clonamiento y análisis de una amplia variedad de replicones. El gen que determina resistencia a amikacina (aac (6')-Ic) fue aislado y ligado a la región de múltiple sitios de clonamiento del vector pUC18. La construcción resultante fue digerida con Hind III y clonada en el sitio Hind III del vector. El clon pHJ13 resultante confirió a las células receptoras la capacidad de crecer en presencia de amikacina (marcador del cassette) y ampicilina (marcador del vector). Mediante análisis con enzimas de restricción se determinó la orientación del cassette. La versatilidad del cassette se sustenta en la presencia, sin modificaciones, de la región de múltiple sitios de clonamiento, que permitirá obtener la secuencia de nucleótidos de cualquier origen de replicación insertado. La funcionalidad del cassette fue demostrada mediante el ligamiento a una región de replicación del plásmido pHH1457 (Complejo H). La presencia de la región ori de pHH1457 y del gen aac (6')-Ic fue confirmada en clones de E. coli. Las propiedades de incompatibilidad del plásmido H y su capacidad para replicarse en una cepa defectiva en PolI se conservaron en el plásmido pHJII14 construido. El cassette de amikacina está siendo utilizado en la caracterización de plásmidos del Complejo H. P

Plásmido de virulencia en yersinia enterocolítica 0-3 de origen murino / Virulence plasmid in yersinia enterocolitica 0-3 of murine origin

Background: Pathogenic strains of Yersinia enterocolitica harbor a virulence plasmid of 45-48 megadalton that can be detected using different techniques. Rodents are important reservoirs of Y enterocolitica. Aim: To investigate the carrier status of Y enterocolitica in murine rodents. Material and methods: Two hundred sixty one mice and rats were captured in rural and urban areas of Valdivia. Y enterocolitica was cultured from viscera and fecal homogenates. Virulence plasmids were detected using crystal violet binding. Results: Thirteen Y enterocolitica strains were isolated from 11 rodents. Ten strains belonged to the biotype 1 and three to the biotype 4, serotype O:3. The most frequently infected rodent species were R norvergicus (20 per cent), followed by A longipilis (11 per cent), A olivaceus (2 per cent) and O longicaudatus (2 per cent). Conclusions: Rodents, and specially the domestic rat, can be an important source of Y enterocolitica infection for human and susceptible animal species

A genetic study of a Staphylococus aureus plasmid involving cure and transference

High frequency transfer and elimination of drug resistance may indicate an extrachromosomal inheritance of genetic determinants. This study shows the cure and transfer of a small plasmid and tetracycline resistance in Staphylococcus aureus 1030 (55) Tetr strains. Several methods are available for plasmid elimination. We used ethidium bromide, an agent that binds to DNA, and thus inhibits DNA polymerase. This caused a high frequency of loss of the small plasmid and resistance to tetracycline. Transfer of tetracycline resistance was done in a mixed culture at a frequency of 10(-6). This type of study is very important to physicians and epidemiology investigators and provides better knowledge on antibiotic-resistance mechanisms that may occur in vivo in a hospital environment.

Isolation and curing of plasmid DNA from bacillus thuringiensis israelensis strains showing variable insecticidal activities

Bacillus thuringiensis israelensis 4Q1-WT [B. t. i.] have 9 plasmids. 14 mutates of this wild type were prepared with cured plasmids. The wild bacterial strain, the mutants and 14 industrial strains were compared according to the number, size of plasmids and their potential to induce mortality to Aedes caspius larvae. The 108 Kb plasmid is essential for B. t. israelensis to induce insect killing

Presencia de plasmidos de DNA de doble hebra en phaffia rhodozyma / Presence of double stranded plasmid DNA in phaffia rhodozyma

El análisis mediante electroforesis en gel de agarosa, de los ácidos nucleicos de tres cepas silvestres de phaffia rhodozyma, permitió determinar la presencia de elementos genéticos extracromosómicos de DNA de doble hebra en una de ellas, la cepa UCD 67-210. Esta cepa es portadora de al menos 6 bandas de DNA cromosómico y cuyos tamaños moleculares corresponden a 6.5, 5.9, 5.0, 4.4, 3,2 y 2.5 kb. Con el objetivo de determinar el tipo de ácido nucleico que constituye estos elementos, se estudió su comportamiento frente a diferentes nucleasas. El tratamiento con ribonucleasa A, ya sea en alta o baja fuerza iónica, no tiene efecto sobre las bandas electroforéticas, así como el tratamiento con nucleasa SI. Por el contrario, el tratamiento con desoxiribonucleasa pancreática conduce a una degradación completa de las bandas y del DNA cromosómico. Estos resultados sugieren que la naturaleza química de los plásmidos corresponde a DNA de doble hebra. Por otra parte, la visualización de los plásmidos en gel de agarosa, depende de la utilización de proteinasa K y SDS en el procedimiento de purificación de los plásmidos y en las condiciones de corrida electroférica, sugiriendo la presencia de un complejo DNA plásmidial-proteína en cada uno de estos elementos. Finalmente, el análisis mediante enzimas de restricción de dos de estos plásmidos, sugiere que estos elementos no están relacionados

Vibrio fluvialis attachs to but does not enter Hela cell monolayers

Considering the possibility that invasiveness could be a neglected factor of virulence in Vibrio fluvialis-linked enteritis, since a dysenteric form of the disease was seen in Bangladesh, we studied 12 Brazilian strains of the organism, six clinical and six environmental, to determine whether they might be able to enter into HeLa cell monolayers or would carry plasmids incidentally involved in invasiveness. Four human and two environmental isolates attached to but did not enter into the cells. Though five strains harbored plasmids,no relationship was found between the carriage of these genetic elements and adhesiveness