[Prevalence of Trypanosoma cruzi infection in opossum (Didelphis albiventris) in Santiago del Estero, Argentina]. / Estudio de la prevalencia de la infección por Trypanosoma cruzi en zarigüeyas (Didelphis albiventris) en Santiago del Estero, Argentina.

The opossum of the genus Didelphis is one of the principal wild reservoirs of Trypanosoma cruzi and is widely distributed in the Western Hemisphere. Didelphis albiventris is the most common marsupial in Amamá and Trinidad, two communities in the province of Santiago del Estero, Argentina. The D. albiventris population is replaced every year, and the opossum normally has two reproductive periods, one at the beginning of the spring and another at the beginning of the summer. The two litters are weaned, and they leave the mother's marsupial pouch to join the population, the first (G1) at the beginning of the summer and the second (G2) at the beginning of the fall. Between 1988 and 1991 409 D. albiventris opossums were studied, and xenodiagnoses showed that 35% of them were infected with T. cruzi. Annual cycles of renewed infection were observed, with prevalences that ranged between 22% and 43%. The acquisition of the parasite occurred over the entire year, from the summer through the spring. The prevalence of infection increased with age. The G1 individuals tended to present higher prevalences than the G2 individuals, probably from being exposed to transmission for a longer period of time. In the first two (younger) age categories for the opossums, G2 individuals showed higher prevalences than did the G1 individuals. This indicates a significant increase in transmission intensity during the fall. Opossums should be regarded as a potential source of T. cruzi entry to the domestic transmission cycle.

Comparative meiotic studies in Triatoma sordida (Stål) and T. guasayana Wygodzinsky & Abalos (Reduviidae, Heteroptera).

Triatoma sordida and T. guasayana are competent Trypanosoma cruzi vectors, with overlapping distribution areas in Argentina. Both species are morphologically similar, and their immature stages are hard to discriminate. Cytogenetic studies in the genus Triatoma reveal scarce karyotypic variations, being 2n = 20 + XY the most frequent diploid number in males. In the present work the meiotic behaviour of different Argentinian populations of T. sordida and T. guasayana has been analyzed; the meiotic karyotype of both species has also been compared. The species differ in total chromosome area and in the relative area of the sex chromosomes. These meiotic karyotypic differences constitute an additional tool for the taxonomic characterization of T. sordida and T. guasayana. The analysis of an interpopulation hybrid of T. sordida (Brazil x Argentina) reveals a regular meiotic behaviour; despite the presence of heteromorphic bivalents. Our observations support the hypothesis that karyotype variations through the gain or loss of heterochromatin can not be considered as a primary mechanism of reproductive isolation in Triatoma.

Specific detection of a probiotic Lactobacillus strain in faecal samples by using multiplex PCR.

Lactobacillus gasseri 4B2 is a human isolate characterised by a strong autoaggregating phenotype mediated by APF (aggregation-promoting factor), a secreted protein. Two primer pairs were developed for simultaneous amplification of a specific fragment of the APF gene and a highly conserved region of the 16S rRNA gene. The specificity of this protocol was checked in DNA samples isolated from single and mixed cultures of Lactobacillus. The same amplification protocol was successfully used directly adding whole bacterial cells to PCR reaction tubes. The suitability of this method for in vivo studies was investigated through feeding L. gasseri 4B2 to mice and analysing colony forming units obtained by plating faecal samples on selective medium. The methodology allows a fast and reliable identification of the target strain without any DNA extraction procedure.

Use of RAPD and 16S rDNA sequencing for the study of Lactobacillus population dynamics in natural whey culture.

The development of communities of the thermophilic microflora of natural whey culture for Parmigiano Reggiano cheese production was studied by means of molecular techniques. RAPD analysis facilitates the identification of the Lactobacillus strains involved in this microbial association and permitted the study of population dynamics during two cycles of whey fermentation. Analysis of RAPD fingerprints revealed the presence of four biotypes that dominate the whey fermentation process. Sequence analysis of 16S rDNA demonstrated that the strains isolated from whey belong to Lact. helveticus and Lact. delbrueckii ssp. lactis species.

Spatial distribution of Triatoma guasayana (Hemiptera:Reduviidae) in hardwood forest biotopes in Santiago del Estero, Argentina.

In the study area Triatoma guasayana Wygodzinsky & Abalos is the only wild triatomine found sympatric with Triatoma infestans (Klug) in peridomestic premises. The Trypanosoma cruzi Chagas wild cycle is centered around the same biotopes occupied by T. guasayana, which are also visited mainly by opossums with annual prevalences of 29-50%. Twelve hectares were sampled for 4 consecutive years during all seasons. During that time, 420 T. Guasayana individuals were collected in 11.3% of 1,188 biotopes of 4 types, which included quimiles (the cactus Opuntia quimilo), chaguares (several species of bromeliads), trees, and logs. Quimiles had the highest percentage of positive biotopes (31.5%) followed by chaguares (22.3%), whereas 5% of the logs were found infested. During all seasons, 9.6-15.2% of biotopes were found infested. Distance to artificial biotopes was not statistically significant when comparing the frequency of triatomine numbers per biotope in all biotope-season combinations. With the exception of quimiles in the fall, the mean number of triatomines was higher in chaguares during all seasons. Triatomine abundance by biotope and season strata showed a clumped distribution, except for the quimiles biotope during the summer. When pooling by seasons, the mean number of triatomines in chaguares and quimiles biotope was higher than in logs and trees, with all biotopes showing a strong clumped distribution. When pooling by biotopes, the mean number of T. guasayana was relatively similar for all seasons, with a strong clumped distribution. The strong contagious distribution of T. guasayana in the hardwood forest biotopes may explain the maintenance of the wild cycle of T. cruzi, despite the low number and the low prevalences of the insect vector.

Aggregation-promoting factor in pig intestinal Lactobacillus strains.

Autoaggregation was frequently encountered among intestinal lactobacilli isolated from weaned pigs. The aggregation mechanism was shown to be mediated by the production of a proteinaceous aggregation-promoting factor in two strains of Lactobacillus reuteri. A 32 kDa aggregation-promoting protein was detected in these strains by cross-reaction with rabbit polyclonal antibodies for Aggregation-Promoting Factor produced by the human isolate Lact. plantarum 4B2. Coaggregation reactions of Lact. reuteri strains with pathogenic and non-pathogenic Escherichia coli were detected.

Interaction between Didelphis albiventris and Triatoma infestans in relation to Trypanosoma cruzi transmission.

This paper attempts to prove if a high Trypanosoma cruzi prevalence of opossums might be reached with few potential infective contacts. One non-infected Didelphis albiventris to T. cruzi and 10 infected nymphs of Triatoma infestans were left together during 23 hr in a device that simulated a natural opossum burrow. Twenty-six replicates were performed using marsupials and triatomines only once. Potentially infective contacts occurred in all the trials. From the 26 opossums used in trials, 54% did not eat any bug. Of the 260 bugs used, 21% were predated. In the 25 trials involving 205 surviving bugs, 36% of them did not feed. In 15/25 cases, > or = 60% of the triatomines were able to feed. The parasitological follow-up of 24 opossums showed that among 10 that had eaten bugs, 4 turned out infected and among the 14 that had not predate, 3 (21%) became positive. In sum, 7/24 (29%) of the marsupials acquired the infection after the experiment. This infection rate was similar to the prevalences found for the opossum population of Santiago del Estero, Argentina, suggesting that the prevalences observed in the field might be reached if each marsupial would encounter infected bugs just once in its lifetime.

Laboratory comparison of feeding success among Triatoma infestans, T. guasayana, and T. sordida (Hemiptera: Reduviidae).

Success in obtaining a blood meal and rapid access to hiding places after feeding are the principal requirements triatomines have as they colonize artificial ecotopes. Feeding success and postfeeding location of 3rd and 5th instars of Triatoma sordida (Stål), of T. guasayana Wygodzinsky & Abalos, and T. infestans (Klug) were studied in an experimental box in which triatomines and a chicken were left to interact overnight. The bird was enclosed in a glass cube, slightly raised to allow triatomines to get in and out, turning the space outside into an extensive refuge area. Four bunches of dry corn husks and a wooden panel were also offered as shelters. The number of live and dead insects and their locations at the end of the experiment were recorded. Predation--as the percentage of missing insects--and success--as the percentage of insects alive and fed--were calculated. Interactions with the host were different among species and sometimes also between individuals of different life stages of the same species. Predation was always > 20%. T. sordida was the best exploiter of the blood source, because 3rd instars were more successful and 5th instars were as successful as the corresponding T. infestans stages. Performance of T. guasayana was equivocal because survival and feeding success were different for both instars. T. infestans showed a pronounced tendency to climb, and 3rd instars of T. guasayana were equally distributed in the upper half of the box and in the corn husks, whereas the majority of T. sordida and 5th instars of T. guasayana sheltered in the husks.

The interaction between poultry and Triatoma infestans Klug, 1834 (Hemiptera: Reduviidae) in an experimental model.

This paper compares the predation pressure that ducks and chickens exert on triatomines. For the tests, these birds were placed in individual boxes together with a known number of Triatoma infestans and left to interact from 6 p.m. till the next morning, involving a long lasting period of complete darkness limited by two short-term periods of semi-darkness. There was a shelter which could prevent the bugs from being predated. The number of live and dead triatomines was recorded, considering missing bugs as predated by the birds. Ducks exhibited a greater predatory activity than chickens, that could be due to a long term active period at night while chickens sleep motionless from sunset to dawn. Surviving triatomines that had fed on chickens outnumbered those fed on ducks suggesting that these were less accessible to the triatomine biting. If ducks are better than chickens to detect and eat bugs and to interfere with their feeding in the field, an increase in duck number might help to diminish triatomine density. Further research is needed to determine the feasibility of application of these experimental results.

Antimicrobial compounds from Lactobacillus casei and Lactobacillus helveticus.

Three strains of Lactobacillus casei and one of Lactobacillus helveticus were examined for antagonistic activity toward twenty five indicator strains of different species. Under conditions eliminating the effects of organic acid and hydrogen peroxide, culture supernatants of all the Lactobacillus strains exhibited a wide spectrum of inhibitory activity toward microorganisms of different genera. The inhibitory compound secreted by one strain of L. casei was active against Clostridium tyrobutyricum. The active components were insensitive to proteolytic enzyme and heat treatment.

Serum cholesterol levels in axenic mice colonized with Enterococcus faecium and Lactobacillus acidophilus.

Hypocholesterolemic effect was shown in axenic, mono, bicolonized and conventional mice: the effect was different depending on probiotic properties of intestinal microorganisms. Contamination by Enterococcus faecium CX determined the highest effect: haematic cholesterol level decrease was 16.9% in females and 7.8% in males. In mice contaminated by Lactobacillus acidophilus N5 the decrease of haematic cholesterol levels was less and not relevant in mice contaminated by conventional microflora. Enterococcus faecium CX and Lactobacillus acidophilus N5 strains were able to grow in presence of bile salts, to colonize intestinal tract, to survive at gastric conditions and to assimilate cholesterol (E. faecium more than L. acidophilus). The authors consider the possibility to associate probiotic strains with these characteristics for the health of consumers.

Plasmid transformation of Ruminococcus albus by means of high-voltage electroporation.

To apply recombinant DNA techniques to the genetic manipulation of cellulolytic ruminal bacteria, a plasmid vector transformation system must be available. The objective of this work was to develop a system for plasmid transformation of Ruminococcus albus. Using high voltage electrotransformation, pSC22 and pCK17 plasmid vectors, derived from lactic acid bacteria plasmids and replicating via single-stranded DNA intermediate, were successfully introduced into three freshly isolated R. albus strains and into R. albus type strain ATCC 27210. The optimization of the electrotransformation condition raised the electroporation efficiency up to 3 x 10(5) transformants per microgram of pSC22 plasmid.

Use of DNA probes in the study of silage colonization by Lactobacillus and Pediococcus strains.

A technique to monitor lactic acid bacteria inoculants in silage, based on specific DNA probes, was developed and used to evaluate the colonization properties of two strains of Lactobacillus plantarum and one strain of Pediococcus pentosaceus which were used as maize silage inoculants in farm conditions. The results indicated that these three strains were able to dominate the natural microflora of the silage, representing more than the 95% of the bacterial biomass of the maize silage. These studies indicate that the colony hybridization with specific DNA probes may be an effective method for monitoring bacteria and evaluating the colonization properties of inoculants in maize silage.

Single-stranded DNA plasmid, vector construction and cloning of Bacillus stearothermophilus alpha-amylase in Lactobacillus.

Vector plasmids were constructed by ligating chloramphenicol and erythromycin resistance genes to TaqI-digested DNA of a cryptic plasmid from Lactobacillus plantarum. The minimal region of Lactobacillus plasmid DNA that was required for DNA replication was defined and a single-stranded DNA intermediate replication system was observed. Homologies with other origins of replication of plasmids from Gram-positive bacteria, replicating via rolling circle mechanism, were found. It was shown that the constructed vectors, named pPSC20 and pPSC22, were transformable into L. plantarum, Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus fermentum, Lactobacillus helveticus, Lactococcus lactis subsp. lactis, Bacillus subtilis, and Escherichia coli. Using plasmid pPSC22, the alpha-amylase gene of Bacillus stearothermophilus was cloned and expressed in several Lactobacillus species.

Antagonism and adhesion among isogenic strains of Lactobacillus reuteri in the caecum of gnotobiotic mice.

Interactions between a wild adhesive Lactobacillus reuteri strain isolated from chicken crop and its isogenic plasmid-free derivative were studied. Antagonism between these strains was tested in vitro or associated with gnotobiotic mice. The cured strains repressed the establishment of the wild, plasmid bearing strain in the gastrointestinal tract of gnotobiotic mice. Interactions between isogenic strains were independent of the amount of cells administered to mice. The cured derivative strain maintains its specific ability to adhere to epithelial cells of chicken crop.

In vivo transfer of pAM beta 1 from Lactobacillus reuteri to Enterococcus faecalis.

Trials were conducted to determine the in vivo transferability of plasmid-mediated antibiotic resistance between two strains of enteric Gram-positive bacteria. Germ-free mice were associated with the donor Lactobacillus reuteri DSM 20016 strain, carrying the broad host range pAM beta 1 plasmid, and with the Enterococcus faecalis JH2SS recipient strain. Analysis of faecal content of associated mice demonstrated that the in vivo transfer of this plasmid did occur and that frequencies of conjugation were affected by the presence of subtherapeutic levels of antibiotic in the diet.

An introduction to rod-shaped lactic-acid bacteria.

The genus Lactobacillus is mainly found in the intestinal tract of healthy humans and animals as well as in fermenting vegetables or plant materials, such as silage. It has a moderate diffusion in meat products and is rarely found in wines and beers. On the other hand, rod-shaped lactic-acid bacteria are largely used in the preparation of a variety of foods and feed products. As starter cultures, they are omnipresent in cheeses and dairy manufacturing. Specific fermentation processes have been developed in order to encourage the growth of the desired species, some of which are fastidious organisms such as L. delbrueckii ssp. bulgaricus, L. helveticus and L. sanfrancisco. In addition, a promising and interesting perspective is the use of rod-shaped lactic-acid bacteria, primarily L. acidophilus, L. reuteri and L. salivarius as probiotic starters to preserve the natural biological equilibrium of the intestinal tract.