Characterization of novel <Emphasis Type="Italic">Trichoderma</Emphasis> spp. isolates as a search for effective biocontrollers of fungal diseases of economically important crops in Argentina

Monoconidial cultures of 33 isolates of Trichoderma from Buenos Aires Province, Argentina were characterized on the basis of twenty eight morphological, physiological and biochemical features. All of them were screened for proteinase, endochitinase and β-1,3 glucanase activity. Universally primed PCR (UP-PCR) and inter-simple sequence repeat (ISSR) techniques were used to examine the genetic variability among isolates, which resulted in 127 bands for the total number of isolates. These results were subjected to numerical analysis revealing 20 haplotypes grouped in five clusters. The ability of Trichoderma isolates to antogonize soil-borne fungal plant pathogens using a dual culture assay was done against five fungal species: Alternaria sp., Bipolaris sorokiniana, Fusarium graminearum, F. solani, and Pyricularia oryzae. The highest inhibition values (85% RI) were obtained against B. sorokiniana and P. oryzae. Three isolates of T. harzianum named as FCCT2, FCCT3 and FCCT9 were capable of causing a high growth inhibition on four of the fungal species assayed, which was in agreement with their higher extracellular hydrolytic activity. Our results suggest that these isolates have the potential to be effective agents for biocontrol of cereal and tomato fungal pathogens.     

Biological variation in Trichinella species and genotypes

At present, the genus Trichinella comprises seven species of which five have encapsulated muscle larvae (T. spiralis, T. nativa, T. britovi, T. nelsoni and T. murrelli) and two do not (T. pseudospiralis and T. papuae) plus three genotypes of non-specific status (T6, T8 and T9). The diagnostic characteristics of these species are based on biological, biochemical and genetic criteria. Of biological significance is variation observed among species and isolates in parameters such as infectivity and immunogenicity. Infectivity of Trichinella species or isolates is determined, among other considerations, by the immune status of the host in response to species- or isolate-specific antigens. Common and particular antigens determine the extent of protective responses against homologous or heterologous challenge. The kinetics of isotype, cytokine and inflammatory responses against T. spiralis infections are isolate-dependent. Trichinella spiralis and T. pseudospiralis induce different dose-dependent T-cell polarizations in the early host response, with T. spiralis initially preferentially promoting Th1-type responses before switching to Th2 and T. pseudospiralis driving Th2-type responses from the outset.     

Resistance to freezing temperatures in Aedes (Ochlerotatus) albifasciatus (Macquart) eggs (Diptera: Culicidae) from two different climatic regions of Argentina

Aedes (Ochlerotatus) albifasciatus (Macquart) has the capacity to proliferate in different kinds of climates within its distribution range in South America. With the aim of studying local thermal adaptations of eggs, we exposed egg stocks from two climatically different localities: temperate humid pampa (Buenos Aires) and cold arid Patagonian (Sarmiento), to freezing conditions and then evaluated the effect on some features at this level. First, we thermally described the substrate where this species lays its eggs in the arid region. A typical thermal condition during winter was 10 h at ?12° C. Second, we evaluated the effect of freezing on primary hatching (vs total hatching) and embryo survival. We also compared the proportion of embryonated eggs from both populations. The proportions of embryonated eggs were not different between localities, with averages of 78% and 83% in Sarmiento and Buenos Aires, respectively. Survival was equally successful after freezing in the two localities with an average range between 94–99%. Whether or not the eggs from Buenos Aires and Sarmiento were under freezing conditions, hatching was more than 98% after the first flooding. The results suggest that eggs of Ae. albifasciatus from Sarmiento and Buenos Aires have the same ability to survive at extreme temperatures (<0° C), showing a regional thermal adaptation rather than a local one.     

Conservation of diagnostic antigen epitopes among biologically diverse isolates of Trichinella spiralis

Crude and immunoaffinity-purified excretory-secretory antigens derived from a domestic pig isolate of Trichinella spiralis were used in an enzyme-linked immunosorbent assay to test serum from mice infected with 25 different pig and wild animal isolates of T. spiralis sspp. All of the sera were found positive by ELISA using either of the antigen preparations, indicating all isolates shared certain antigen epitopes. Excretory-secretory antigens were prepared from 3 distinct isolates of T. spiralis sspp.--Trichinella spiralis spiralis (pig isolate), Trichinella spiralis nativa (polar bear isolate), and Trichinella spiralis pseudospiralis--and compared by electrophoresis and monoclonal antibody binding. While protein profiles varied among the isolates, a monoclonal antibody recognizing a major immunodiagnostic antigen epitope bound all 3 antigen preparations. However, this antigen epitope occurred on different molecular weight excretory-secretory proteins from the different isolates.     

Thermal response in pre‐imaginal biology of Ochlerotatus albifasciatus from two different climatic regions

The biological processes on mosquito could be variable in response to local climatic characteristics. The thermal effects on time and the rate larval development, immature survival and adult size in local populations of Ochlerotatus albifasciatus (Macquart) (Diptera: Culicidae) from cold (Sarmiento) and temperate (Buenos Aires) regions from Argentina were evaluated. This species affects livestock production and human health. Larvae of both regions were placed in breeding thermal baths (11–32 °C range). Development and survival were recorded daily until adult emergence. The development temperature threshold and thermal constant for Sarmiento (4.59 ± 3.08 °C, 204.08 ± 7.83 degree days) was lower and higher than Buenos Aires, respectively (8.06 ± 1.81 °C, 149.25 ± 2.6 degree days). At cold temperatures (11–16 °C), Sarmiento larvae demonstrated 5 days faster development and higher survival (56%) than Buenos Aires (15%), whereas at warm temperatures (20–32 °C) were up to 2 days slower and similar survival (16% vs. 18%). The size did not show differences between populations. An Ochlerotatus albifasciatus population seems to present local thermal responses. The favourable temperature for survival and rate of development would vary within a cold or warm range, and these differential responses would explain the wide geographical distribution in different climatic regions of southern South America.     

Ribo HRM--detection of inter- and intra-species polymorphisms within ribosomal DNA by high resolution melting analysis supported by application of artificial allelic standards

Ribo HRM, a single-tube PCR and high resolution melting (HRM) assay for detection of polymorphisms in the large subunit ribosomal DNA expansion segment V, was developed on a Trichinella model. Four Trichinella species: T. spiralis (isolates ISS3 and ISS160), T. nativa (isolates ISS10 and ISS70), T. britovi (isolates ISS2 and ISS392) and T. pseudospiralis (isolates ISS13 and ISS1348) were genotyped. Cloned allelic variants of the expansion segment V were used as standards to prepare reference HRM curves characteristic for single sequences and mixtures of several cloned sequences imitating allelic composition detected in Trichinella isolates. Using the primer pair Tsr1 and Trich1bi, it was possible to amplify a fragment of the ESV and detect PCR products obtained from the genomic DNA of pools of larvae belonging to the four investigated species: T. pseudospiralis, T. spiralis, T. britovi and T. nativa, in a single tube Real-Time PCR reaction. Differences in the shape of the HRM curves of Trichinella isolates suggested the presence of differences between examined isolates of T. nativa, T. britovi and T. pseudospiralis species. No differences were observed between T. spiralis isolates. The presence of polymorphisms within the amplified ESV sequence fragment of T. nativa T. britovi and T. pseudospiralis was confirmed by sequencing of the cloned PCR products. Novel sequences were discovered and deposited in GenBank (GenBank IDs: JN971020-JN971027, JN120902.1, JN120903.1, JN120904.1, JN120906.1, JN120905.1). Screening the ESV region of Trichinella for polymorphism is possible using the genotyping assay Ribo HRM at the current state of its development. The Ribo HRM assay could be useful in phylogenetic studies of the Trichinella genus.     

On the taxonomic status of a Dugesia species from Buenos Aires (Platyhelminthes: Turbellaria: Tricladida)

The taxonomic and distributional statuses of the freshwater planarians of South America are imperfectly understood, and a comprehensive review is sorely needed. The species known from the vicinity of Buenos Aires, Argentina, was first described by Böhmig (1902) under the name Planaria dubia Borelli, 1895. Subsequent authors (Kenk, 1930, 1974; Cazzaniga & Curino, 1987) have cast doubt on this assignation. Some have assigned it to Dugesia anceps Kenk; Hyman (1959) was inclined to regard the Buenos Aires planarian as a geographical variant of Dugesia sanchezi Hyman. A comparison of D. sanchezi from the mountainous region of central Chile with specimens from Buenos Aires confirms that the Argentinian material is a distinct species.The differences between the two species are subtle. The Argentinian specimens are distinctive in their possession of infranucleate epithelia of the atria, penis, and bursal canal, as well as in their penial structure. In both species, the epithelia of the atria and bursal canal are glandular. Those of D. sanchezi are nucleate and their secretions are cyanophilous. In the Argentinian specimens, these same epithelia are infranucleate; but while the secretion of the bursal canal is indeed cyanophilous, that of the atrium is erythrophilous. The presence, in these same specimens, of cyanophilous glands opening into the penial vesicles, and of erythrophilous glands whose secretions contribute to a viscous plug in the ejaculatory duct, suggests a difference in the physiology of copulation.It is apparent that the Argentinian specimens are distinct from D. sanchezi Hyman, and probably they represent a new species, one to be described in detail elsewhere. In any case, Hyman's (1959) suggestion that D. sanchezi occurs near Buenos Aires is mistaken; so also is Cazzaniga's & Curino's (1987) identification of specimens from Buenos Aires as D. anceps.     

Allozyme analysis of Trichinella isolates from various host species and geographical regions.

Allozyme analysis was carried out on 152 Trichinella isolates from synanthropic and wild animals and from humans; the isolates were collected from 5 continents. The analysis, involving 27 enzymes, revealed the presence of 8 distinct gene pools, termed T1-T8. Four of the genetic groups represent the 4 previously proposed species: Trichinella spiralis sensu stricto (T1), Trichinella nativa (T2), Trichinella nelsoni (T7), and Trichinella pseudospiralis (T4). The other 4, T3, T5, T6, and T8 are distinct from previously described species. The absence of allozymic hybrid patterns among even sympatric groups indicates a lack of gene flow among the groups. Principal component analysis and the unweighted pair group method of analysis were used to assemble allozyme patterns of the 152 isolates into discrete groups and to show their relative relationships. Both analyses indicated the presence of 8 primary clusters that correlated with the gene pools revealed by direct allozyme profile analysis. The absence of evidence of gene flow among the gene pools and the high level of allozymic differentiation between the cluster groups support the concept that the genus Trichinella is composed of several sibling species.     

Identificación presuntiva de Candida spp. y de otras levaduras de importancia clínica: utilidad de Brilliance Candida Agar

Fungal infections caused by yeasts have increased during the last decades and invasive forms represent a serious problem for human health. Candida albicans is the species most frequently isolated from clinical samples. However, other emerging yeast pathogens are increasingly responsible for mycotic infections, and some of them are resistant to some antifungal drugs. Consequently, it is necessary to have methods that can provide a rapid presumptive identification at species level.Numerous chromogenic agar media have been shown to be of value as diagnostic tools. We have compared a chromogenic medium, Brilliance Candida Agar, with CHROMagar Candida, the chromogenic medium most used in our country. A multicentre study was conducted in 16 Hospitals belonging to the Mycology Net of Buenos Aires City Government. A total of 240 yeast isolates were included in this research.The new chromogenic agar showed results very similar to those obtained with CHROMagar Candida.     

Trichinella spiralis: genetic evidence for synanthropic subspecies in sylvatic hosts

Isolates of the nematode genus Trichinella from sylvatic hosts differ in their potential to reproduce in domestic swine. The structure of the genomic DNA from 13 sylvatic isolates from North America and 5 pig isolates, 4 from North America and 1 from Asia, was examined and correlated with the infectivity of the isolate for domestic pigs. DNA restriction fragment length differences, identified by ethidium bromide staining and by hybridization with 32P-labeled ribosomal RNA, served as molecular markers to classify each isolate. All 5 pig isolates and 8 of 13 sylvatic isolates had a high infectivity and reproductive capacity in pigs. All isolates that were highly infectious for pigs regardless of host origin had similar DNA characteristics and were classified operationally as T. spiralis spiralis (pig) and those of the second group as T. spiralis ssp. A DNA clone of repetitive DNA from T. s. spiralis, pBP2, was selected from a library of genomic DNA in plasmid pUC8. When used as a probe, pBP2 hybridized only to the DNA of T. s. spiralis isolates, thus making it a useful diagnostic reagent to predict whether new isolates are highly infectious for pigs (i.e., T. s. spiralis). These results show that T. s. spiralis occurs in wild mammals and this should be considered a serious obstacle to efforts to eradicate trichinellosis from domestic swine.     

Comparison of three subspecies of Trichinella spiralis by scanning electron microscopy

The surface morphology of three subspecies of Trichinella spiralis was examined by SEM in an attempt to find characteristics useful for distinguishing the subspecies. The subspecies studied were T. spiralis spiralis, which had been maintained in swine and laboratory animals for about 50 yr; T. spiralis nativa collected from Ursus maritimus at 58 degrees N latitude and 95 degrees W longitude in 1976; and, T. spiralis pseudospiralis, which was derived from the original isolation of this subspecies from Procyon lotor at 43 degrees N latitude and 47 degrees 30'E longitude in 1972. All three subspecies were passed in CFW mice and adult worms were collected from the small intestine at 4, 5, 6, 7, 8, and 11 days PI. Characteristics examined included labial and cephalic papillae, cuticular ridges and folds, hypodermal gland cell pores, pseudobursal lobes, genital papillae, cloacal aperture, copulatory bell and vulval morphology. Previous reports of subspecies differences within Trichinella spiralis in the number and distribution of hypodermal gland cell pores, position of genital papillae, shape of the cloacal aperture and shape of pseudobursal lobes were not confirmed and are believed to have been in error resulting from artifacts of fixation and a lack of knowledge of variations within the subspecies caused by low numbers of samples. Differences in surface morphology were not found among the three subspecies. The available names of the recognized biological populations of Trichinella were used at the subspecies level rather than species level because this more clearly represents the state of our knowledge of the nematodes. The question of whether the epidemiology of trichinosis is complicated by the presence of more than one species has not been answered, and it is important that our nomenclature reflect this.     

Advances in the sequencing of the genome of the adenophorean nematode Trichinella spiralis

The adenophorean nematodes are evolutionarily distant from other species in the phylum Nematoda. Interspecific comparisons of predicted proteins have supported such an ancient divergence. Accordingly, Trichinella spiralis represents a basal nematode representative for genome sequencing focused on gaining a deeper insight into the evolutionary biology of nematodes. In addition, molecular characteristics that are conserved across the phylum could be of great value for control strategies with broad application. In this review, we describe and summarize progress that has been made on the sequencing and analysis of the T. spiralis genome. The genome sequence was used in preliminary analyses for the investigation of specific questions relating to the biology of T. spiralis and, more generally, to parasitic nematodes. For instance, we evaluated an unusually large DNase II-like protein family, predicted proteins of prospective interest in the parasite-host muscle cell interaction, anthelmintic targets and prospective intestinal genes, the encoded proteins (potentially) linked to immunological control against other nematodes. The results are discussed in relation to characteristics that are broadly conserved among evolutionary distant nematodes. The results lead to expectations that this genome sequence will contribute to advances in research on T. spiralis and other parasitic nematodes.     

Genetic variability of Phytophthora sojae isolates from Argentina

Phytophthora sojae causes root and stem rot, one of the most important diseases of soybean worldwide. Genetic diversity of 32 Phytophthora sojae isolates of different geographic origin from Argentina was evaluated with RAPD markers. The isolates were collected from diseased soybean plants and soil samples from Santa Fe, Buenos Aires, C6rdoba and Entre Rios provinces, in the Pampeana Region. DNA was amplified with 20 decanucleotides primers. Seven primers amplified 49 fragments, of which 35 were polymorphic, indicating high variability. RAPD analysis detected intraspecific variability even among isolates of the same geographic origin.     

Environmental characteristics of the cemeteries of Buenos Aires City (Argentina) and infestation levels of Aedes aegypti (Diptera: Culicidae)

Cemeteries with many water-filled containers, flowers, sources of human blood, and shade are favorable urban habitats for the proliferation of Aedes aegypti, a vector of yellow fever and dengue. A total of 22,956 containers was examined in the five cemeteries of the city of Buenos Aires, Argentina. The vector was found in four cemeteries that showed an average infestation level of 5.5% (617 positive out of 11,196 water-filled containers). The four cemeteries positive for Ae. aegypti showed significantly different (p<0.01) infestation levels. Vegetation cover and percentage of infestation were significantly correlated (p<0.01), but neither cemetery area nor number of available containers were significantly related to the proportion of positive vases. Our results suggest that the cemeteries of Buenos Aires represent a gradient of habitat favorableness for this vector species, some of which may act as foci for its proliferation and dispersal.     

On the habitat and isolation of Gymnocharacinus bergi (Osteichthyes: Characidae)

Synopsis Gymnocharacinus bergi is a rare Paranensean fish which is the only characiform almost lacking scales in the adult. It is endemic and the only species in a peculiar spot — a tributary of the Valcheta creek — in the Somuncurá plateau in northern Patagonia, Argentina, over 300 km from the nearest place with a paranensean fish fauna. Besides its geographical isolation, G. bergi occurs within an area with climatic features drastically different from those currently associated with fishes from Neotropical temperate zones. We tested the assumption that water temperature in the naked characin habitat do not agree with the northern Patagonia climate. We also considered the isolation of G. bergi within the framework of an increasing inpoverishment of the paranensean ichthyofauna along a NE-SW axis in the Buenos Aires province. For this we applied a decrement equation used in island biogeography. Our findings demonstrate that the existence of G. bergi in its isolated habitat is possible because of the thermal traits of the water at the sources of the creeks, its temperature being independent of the climate of the area. The chemical composition of water was found to be within the range of common environments in the Buenos Aires pampas inhabited by several species of Paranensean fishes. Geographically, G. bergi lives in the last of a series of habitats which show a decreasing number of species correlated with the increasing distance from the La Plata River. Conservation status of the species is briefly discussed.     

SHOREBIRDS OF COASTAL BUENOS AIRES PROVINCE, ARGENTINA

The Pampas shorebird community is comprised of species from diverse taxonomic and faunal origin. Four major groups can be recognized: North American migrants residing in coastal Buenos Aires Province during their non-breeding season, Patagonian species wintering locally but breeding further south, Pampean species breeding locally, and Brazil–Paraguayan species present during their breeding season. Because of differences among these groups in the timing and migration patterns, community composition shows marked seasonal variation. Both nesting and non-breeding birds use a diversity of habitat types ranging from littoral sandy beaches to upland grasslands. Throughout the year, however, and in all habitat types, non-breeders predominate in species number and abundance. Intra- and inter-specific patterns of aggression suggest both that resources are limiting, and that non-breeding migrants may play an important role in determining breeding community composition.     

COEVOLUTION AND AVIAN BROOD PARASITISM: COWBIRD EGGS SHOW EVOLUTIONARY RESPONSE TO HOST DISCRIMINATION

The Rufous Hornero (Furnarius rufus) is an important host of the brood parasitic Shiny Cowbird (Molothrus bonariensis) in Uruguay, but not in nearby Buenos Aires Province, Argentina. Eggs of the Shiny Cowbird are extremely variable in size, and horneros eject cowbird eggs with widths less than about 88% of the widths of their own eggs. Uruguayan cowbird eggs are, on average, 12% larger than those from Buenos Aires, a geographic pattern in egg size that corresponds to the pattern of successful host use. Uruguayan cowbird eggs are also wider per unit volume than eggs from Buenos Aires. Allometric analyses of egg width and volume indicate that this shape change is apparently an evolutionary response to selection exerted by the hornero. Such a response is absent in Buenos Aires because interactions between the cowbird and the hornero are probably of recent origin there.     

The systematic position of some nippostrongyline nematodes (Trichostrongylina: Heligmosomoidea) parasitic in Argentinean sigmodontine rodents

The systematic position of two nippostrongyline nematodes described from Argentinean sigmodontine rodents is clarified. The first species, Hassalstrongylus multiovatus Suriano & Navone, 1992, parasitic in Akodon simulator Thomas from the province of Tucumán, was studied on the basis of type and voucher material. H. multiovatus is proposed as a junior synonym of Trichofreitasia lenti Sutton & Durette-Desset, 1991, a parasite described from Oligoryzomys flavescens (Waterhouse) in the province of Buenos Aires. The holotype and three of seven paratypes deposited as H. multiovatus were identified as T. lenti. One male paratype was identified as Guerrerostrongylus uruguayensis Sutton & Durette-Desset, 1991, a parasite described from O. flavescens in Uruguay. Three female paratypes were identified as Guerrerostrongylus sp. The second species, Stilestrongylus scapteromys Suriano & Navone, 1996, parasitic in Scapteromys aquaticus Thomas from the province of Buenos Aires, was studied on voucher material. Stilestrongylus scapteromys and Malvinema frederici Digiani, Sutton & Durette-Desset, 2003, the type-species of Malvinema Digiani, Sutton & Durette-Desset, 2003, were described from the same host and geographical region. As they are considered to refer to one and the same taxon, the new combination Malvinema scapteromys n. comb. is proposed for this species.     

African ancestry of the population of Buenos Aires

The population of Argentina today does not have a “visible” black African component. However, censuses conducted during most of the 19th century registered up to 30% of individuals of African origin living in Buenos Aires city. What has happened to this African influence? Have all individuals of African origin died, as lay people believe? Or is it possible that admixture with the European immigrants made the African influence “invisible?” We investigated the African contribution to the genetic pool of the population of Buenos Aires, Argentina, typing 12 unlinked autosomal DNA markers in a sample of 90 individuals. The results of this analysis suggest that 2.2% (SEM = 0.9%) of the genetic ancestry of the Buenos Aires population is derived from Africa. Our analysis of individual admixture shows that those alleles that have a high frequency in populations of African origin tend to concentrate among 8 individuals in our sample. Therefore, although the admixture estimate is relatively low, the actual proportion of individuals with at least some African influence is approximately 10%. The evidence we are presenting of African ancestry is consistent with the known historical events that led to the drastic reduction of the Afro‐Argentine population during the second half of the 19th century. However, as our results suggest, this reduction did not mean a total disappearance of African genes from the genetic pool of the Buenos Aires population. Am J Phys Anthropol 128:164‐170, 2005. © 2005 Wiley‐Liss, Inc.