First Steps towards the Pharmaceutical Development of Ovules Containing Lactobacillus Strains: Viability and Antimicrobial Activity as Basic First Parameters in Vaginal Formulations.

In the majority of Latin-American countries, including Argentina, there is a limited availability of vaginal bioproducts containing probiotics in the market. In addition, the conventional treatments of genital tract infections in women represent a high cost to the public health systems. The future development of this type of bioproducts that employ specific lactobacilli strains would not only have a meaningful impact on women's health but would also represent a significant challenge to the pharmaceutical industry. The aims of the work described in this paper were (i) to study different pharmaceutical formulations of vaginal ovules containing Lactobacillus fermentum L23 and L. rhamnosus L60, to determine in which formulation lactobacilli viability was sustained for longer time and (ii) to evaluate if probiotic strains maintained both the antimicrobial activity and biofilm-producing ability after being recovered from the ovules. In this study, we developed and characterized three pharmaceutical formulations containing different glycerol amounts and specific lactobacilli strains. Three relevant parameters, cell viability, antimicrobial activity, and biofilm production, by lactobacilli recovered from the ovules were tested. Although the viability of L23 and L60 strains was mainly influenced by high ovule's glycerol proportion, they survived at 4 °C during the 180 days. Both lactobacilli's antimicrobial activity and biofilm-producing ability were maintained for all treatments. In conclusion, employing a much reduced number of components, we were able to select the most suitable pharmaceutical formulation which maintained not only lactobacilli viability for a long period of time but also their antimicrobial activity and biofilm-producing ability.

Resistência bacteriana e ação das bacteriocinas de Lactobacillus spp em Staphylococcus aureus isolados de mastite bovina / Bacterial resistance and bacteriocins action in Staphylococcus aureus isolated from bovine mastitis

Staphylococcus aureus is the major pathogen causing intramammary infections in dairy cattle worldwide. Among the factors that contribute to its spread and infectious potential is the ability to overcome the mechanisms of antimicrobials activity. The present work investigated the antimicrobial resistance pattern and sensibility to bacteriocins produced by strains of Lactobacillus spp of 30 isolates of S. aureus from mastitis. From this, 29 are beta-lactamase producers. Eight isolates (26.6 percent) showed resistance to at least four antibiotics being considered multiresistent. All of them were mecA-positive. Otherwise, all isolates tested showed sensibility to at least one of the four bacteriocin producer strains. Due to the significant depletion of the efficacy of antimicrobials, pathogen growth inhibition by bacteriocins seems an alternative of biological control in infectious processes.

Antimicrobial sensitivity and adherence study in strains of coagulase-negative Staphylococcus spp.

Antimicrobial sensitivity was evaluated from 120 strains of coagulase-negative Staphylococcus (CoNS), isolated from urinary infections. The isolated species were identified by conventional methods and sensitivity to vancomycin, tetracyclin, norfloxacin, cephalothin erythromycin, clindamycin, oxacillin, penicillin, cyprofloxacin, ampicillin and ampicillin/sulbactam was tested by the agar dilution technique and the diffusion in disc technique. The distribution of the species was: Staphylococcus epidermidis 26 (21.6%), S. haemolyticus 48 (40.0%), S. saprophyticus 40 (33.4%) and S. simulans 6 (5%). CoNS strains extracted from urinary infections showed a high percentage of vancomycin and tetracyclin sensitivity, and a low sensitivity to beta-lactamic antibiotics, except ampicillin/sulbactam and cephalotin. Isolation percentage resistance to oxacillin was less than 48%. Strains resistant to oxacillin were considered as resistant to both penicillin and ampicillin. From 42% to 80% of S. haemolyticus, S. saprophyticus, Staphylococcus epidermidis and S. simulans were positive to the adherence test. Results from this work showed that slime producing CoNS strains, isolated in Río Cuarto Hospital, had different percentages of resistance to the studied antimicrobial agents.

Epidemiology of sexually transmitted diseases in Río Cuarto, Argentina.

Sexually transmitted diseases (STD) are a group of transmittable diseases acquired fundamentally through sexual contact. STD are a social problem resulting from demographic explosion and changes in sexual conduct, which affects teenagers and adults of all socioeconomic strata. The goal of this work was to establish the actual state of the different STD within the studied population. Samples of vaginal fluids, endocervical materials and urethral exudates taken from 2,630 patients during five years were processed. 1,341 samples tested positive to one or more of the microorganisms, 1,099 corresponding to female patients and 242 to male patients. The microorganisms found in women were: Gardnerella vaginalis (39.1%), Candida albicans (21.3%), Trichomonas vaginalis (16.8%), Chlamydia trachomatis (11.5%); Neisseria gonorrhoeae (3.4%), Mycoplasma hominis (2.6%); Ureaplasma urealyticum (4.1%) and Treponema pallidum (1.6%). Associations were: Gardnerella vaginalis with Trichomonas vaginalis (6%), Gardnerella vaginalis with Candida albicans (5.1%); Trichomonas vaginalis with Neisseria gonorrhoeae (2.2%) and Gardnerella vaginalis with Chlamydia trachomatis (2.1%). In men, gonococcic urethritis represented 37.8%, non-gonococcic urethritis 55.4% and Treponema pallidum 6.8%. A decrease in syphilis, gonococcic urethritis and gonococcic cervicitis was observed, increasing the prevalence of non-gonococcic urethritis and cervicitis. This study showed that in our environment the actual tendency of STD is still high.

Prevalencia de Chlamydia trachomatis, micoplasmas urogenitales y sus asociaciones en un grupo de mujeres promiscuas / Prevalence of Chlamydia trachomatis, urogenital micoplasmas and their association to a group of promiscuous women

Chalamydia trachomatis, Ureaplasma urealyticum y Mycoplasma hominis son microorganismos responsables de infecciones urogenitales. Son aislados con considerable frecuencia del tracto genital femenino. En este trabajo se estudiaron 100 exudados vaginales de mujeres promiscuas que concurrieron a la división de Bacteriología del Hospital Central de Río Cuarto. En todas las muestras se investigó la presencia de C. trachomatis, U. urealyticum, M. hominis. La prevalencia hallada fue: C. trachomatis 17 por ciento; U. urealyticum 57 por ciento; M. hominis 21 por ciento y Neisseria gonorrhoeae 2 por ciento. Las asociaciones más frecuentes fueron: C. trachomatis-Trichomonas vaginalis, micoplasmas-T. vaginalis y Gardnerella vaginalis-Candida albicans con un 18 por ciento, 15 por ciento y 8 por ciento respectivamente

Prevalencia de Chlamydia trachomatis, micoplasmas urogenitales y sus asociaciones en un grupo de mujeres promiscuas / Prevalence of Chlamydia trachomatis, urogenital micoplasmas and their association to a group of promiscuous women

Chalamydia trachomatis, Ureaplasma urealyticum y Mycoplasma hominis son microorganismos responsables de infecciones urogenitales. Son aislados con considerable frecuencia del tracto genital femenino. En este trabajo se estudiaron 100 exudados vaginales de mujeres promiscuas que concurrieron a la división de Bacteriología del Hospital Central de Río Cuarto. En todas las muestras se investigó la presencia de C. trachomatis, U. urealyticum, M. hominis. La prevalencia hallada fue: C. trachomatis 17 por ciento; U. urealyticum 57 por ciento; M. hominis 21 por ciento y Neisseria gonorrhoeae 2 por ciento. Las asociaciones más frecuentes fueron: C. trachomatis-Trichomonas vaginalis, micoplasmas-T. vaginalis y Gardnerella vaginalis-Candida albicans con un 18 por ciento, 15 por ciento y 8 por ciento respectivamente (AU)

[A biotype study of Gardnerella vaginalis isolated from patients with and without symptoms of bacterial vaginosis]. / Estudio de los biotipos de Gardnerella vaginalis, aisladas de pacientes con y sin síntomas de vaginosis bacteriana.

METHODS: A simple and reproducible proposed for Benito et al. scheme for identification biotypes of Gardnerella vaginalis has been developed, based on reactions for lipase, hippurate, hydrolysis, and beta-galactosidase. RESULTS: 11 biotypes were found among 130 strains from women with and without bacterial vaginosis (non-specific vaginitis) 1A, 5B; 8B, 5C, 8C, 1E, 6G and 7G in women with vaginosis and the biotypes 1A, 5B, 8B, 1C, 8F and 6H in women without vaginosis. These biotypes 1A; 5B and 8B were found in two groups. CONCLUSIONS: These data suggest that some biotypes of G. vaginalis are associated with bacterial vaginosis.

[Survey of sexually transmitted diseases in the region of Rio Cuarto]. / Relevamiento de las enfermedades de transmisión sexual en la región de Río Cuarto.

Sexually transmitted diseases (STD) are acquired mainly through sexual intercourse, being one of the most frequent groups of infectious diseases worldwide and consequently an important public health problem. The aim of this paper was to determine the current state of STD and to compare different diagnostic methods in the population studied. A total of 1060 samples from vaginal flows, endocervical material and urethral discharge were studied during 3 years. Of the total samples, 583 were positive, 493 in women and 90 in men. Microorganisms found in women were: Gardnerella vaginalis (39.3%), Candida albicans (21.1%), Trichomonas vaginalis (17.3%), Candida trachomatis (11.3%), Neisseria gonorrhoeae (3.2%): Mycoplasma hominis and Ureaplasma urelyticum (6.5%) and Treponema pallidum (1.4%), the associations found were, Gardnerella vaginalis with Trichomonas vaginalis 5.5%; Gardnerella vaginalis with Candida albicans 4.9%; Trichomonas vaginalis with Neisseria gonorrhoeae (2.2%) and Gardnerella vaginalis with Chlamydia trachomatis (1.9%). In men, gonococcal urethritis (UG) represented 37.7% non UG 55.6% and Treponema pallidum 6.7%. These results indicate a decrease in sifilis and in UG when compared to previous studies showing that gonococcal cervicitis had also decreased. We found an important increase in the prevalence of urethritis and non gonococcal cervicitis in agreement with world statistics which consider these diseases as the most common venereal ones. It is necessary to increase the search for Chlamydia trachomatis in pregnant women due to vertical transmission. It should be noted that, in spite of certain fluctuations, the incidence of the STD in our area is still unacceptably high.

[In vitro inhibition of the growth of Gardnerella vaginalis by bacteriocins produced by strains of Pseudomonas aeruginosa]. / Inhibición in vitro del crecimiento de Gardnerella vaginalis por bacteriocinas producidas por cepas de Pseudomonas aeruginosa.

BACKGROUND: Pseudomonas aeruginosa with inhibitory capacity in vitro was studied on Gardnerella vaginalis strains. METHODS: Antimicrobial activity was demonstrated by inhibitory halos of bacterial growth on solid media by two methods: crossed streak and agar well diffusion. The inhibitory activity of this substance produced by P. aeruginosa was characterized as bacteriocin by: activity spectrum sensitivity proteolytic enzyme, chloroform, heat, pH, ultraviolet, irradiation effect and molecular weight. RESULTS: Four strains of P. aeruginosa producers of bacteriocins were chosen for this study and contacted with 40 strains of G. vaginalis. The producing strain D inhibited 70% of these G. vaginalis strains. The strains B and C inhibited 55% and 52.5%, respectively. The 3 strains presented a wide rank of action but the strain A had effect on a few strains of G. vaginalis. CONCLUSIONS: This work showed the inhibitory in vitro effect of bacteriocins of P. aeruginosa on strains of G. vaginalis. The results obtained suggest the probable topic application of bacteriocins as an alternative of conventional therapeutic on this infection biological control.

[Clinical and microbiological evaluation of bacterial vaginosis in women in a hospital from Rio Cuarto, Argentina]. / Evaluación clínica y microbiológica de vaginosis bacteriana en mujeres de un hospital de Río Cuarto, Argentina.

One hundred and seventy nine women of discharge vaginal were studied. Endocervical and exocervical samples were obtained. The vaginal fluids samples were processed by the following tests: pH, fishy odor, clue cells observation was done by Gram reaction. Gardnerella vaginalis was identified through biochemical tests. G. vaginalis (23.4%), C. albicans (21.8%), T. vaginalis (14.5%), N. gonorrhoeae (5.5%) and C. trachomatis (11.7%), were found in 42 women. A 7.1% of forms compatible with Mobiluncus spp. were observed by direct Gram. The relation between isolation of G. vaginalis and the presence of clue cells was 95.2%. There was a sensitivity of 95.2%, 100% specificity and a predictive positive value of 100%.

[Pseudomonas fluorescens: production of pyoverdine in human blood at 4 degrees C and cytotoxic effect of the pigment]. / Pseudomonas fluorescens: producción de pioverdina en sangre humana a 4 grados C y efecto citotóxico del pigmento.

Pseudomonas fluorescens PAB strain produced pyoverdine in a synthetic medium, this pigment was purified by solvent extraction and ion exchange, then sterilized by filtration. Where cytotoxic effect on human leukocytes was assayed, death and lysis was observed. Sublytic doses decreased leukocytes phagocytosis and chemotaxis. Bacteria grew and produced pigment in blood stored a 4 degrees C, with a pyoverdine production of 0.13 mg/ml of serum after 5 days of incubation.

[Pseudomonas microcins: characterization and effect of iron concentration]. / Microcinas en Pseudomonas: caracterización y efecto de la concentración de fierro.

Antibacterial substances were found in Pseudomonas, they were heat and trypsin resistant, molecular weight less than 10,000 daltons, produced in minimal media, optimum pH 7.0. Methionine and glycine inhibited them; other amino acids produced less antagonistic effect. An inverted relationship between susceptibility to microcin and Fe3+ concentration was observed in experiments with iron-depleted strains. Pseudomonas was sensitive to its own microcin in iron-free medium; this is an important finding in the study of the immunity of microcin-producing strains.

[Pyoverdin from Pseudomonas fluorescens: binding to the globulin fraction of serum and its effect on serological reaction titers]. / Pioverdina de Pseudomonas fluorescens: unión a la fracción globulina del suero y efecto sobre los títulos de reacciones serólogicas.

Pseudomonas fluorescens grows and produces pigment in refrigerated human blood at 4 degrees C. Saline precipitation of plasma showed that globulin and albumin fractions retained pyoverdin at different concentrations. By dialysis it was possible to determine that the pigment attached or aggregated to the protein in total plasma as well as in the fraction obtained by saline precipitation. A greater binding was observed at the globulin fraction. Gamma-globulin immunological activity was reduced due to the interaction with pyoverdin, as much in agglutination reaction (VDRL) as in neutralization (Streptolysin O).

Infección intrahospitalaria a Pseudomonas diminuta susceptible a piocinas / Intrahospitalary infection to pseudomanas diminuta susceptible to piocins

La incidencia de las infecciones por Pseudomonas ha incrementado especialmente en pacientes hospitalizados. En nuestro medio se identifican con cierta frecuencia Pseudomonas de diferentes materiales biológicos (orina, secreción óptica, lesiones fistulares, septicemias, catéteres y otros). Se aisló un total de 31 cepas de Pseudomonas de las cuales el 90% correspondió a P. aeruginosa y en dos casos se identificó P. diminuta, proveniente de dos pacientes internados en el hospital durante un tiempo prolongado. En uno de ellos se aisló de hemocultivo seriado y en el otro de lesiones fistulares. Ambas cepas presentaron el mismo biotipo, espectro de sensibilidad a antimicrobianos e igual susceptibilidad en la investigación de efecto inhibitorio sobre cepas del género. Este ensayo de detección de actividad bacteriocínica permitió confirmar la sospecha de una infección adquirida en el hospital. (AU)

Infección intrahospitalaria a Pseudomonas diminuta susceptible a piocinas / Intrahospitalary infection to pseudomanas diminuta susceptible to piocins

La incidencia de las infecciones por Pseudomonas ha incrementado especialmente en pacientes hospitalizados. En nuestro medio se identifican con cierta frecuencia Pseudomonas de diferentes materiales biológicos (orina, secreción óptica, lesiones fistulares, septicemias, catéteres y otros). Se aisló un total de 31 cepas de Pseudomonas de las cuales el 90% correspondió a P. aeruginosa y en dos casos se identificó P. diminuta, proveniente de dos pacientes internados en el hospital durante un tiempo prolongado. En uno de ellos se aisló de hemocultivo seriado y en el otro de lesiones fistulares. Ambas cepas presentaron el mismo biotipo, espectro de sensibilidad a antimicrobianos e igual susceptibilidad en la investigación de efecto inhibitorio sobre cepas del género. Este ensayo de detección de actividad bacteriocínica permitió confirmar la sospecha de una infección adquirida en el hospital.