Management of <Emphasis Type="Italic">Tetranychus urticae</Emphasis> (Acari: Tetranychidae) in strawberry fields with <Emphasis Type="Italic">Neoseiulus californicus</Emphasis> (Acari: Phytoseiidae) and acaricides

The purpose of this study was to evaluate the performance of Neoseiulus californicus (McGregor) for the control of Tetranychus urticae Koch in commercial strawberry fields, under greenhouse conditions, in association or not with the use of acaricides. The N. californicus strain used in this study was tolerant or resistant to several pesticides. Three experiments were carried out in the State of São Paulo, Brazil. For the first experiment, the initial infestation of T. urticae was 87.1 active stages per leaflet. Two applications of propargite were made on the first and the 14th day of the experiment. Approximately 2 h after each propargite application, N. californicus was released at a rate of 3.0 and 1.9 adult mites per plant, respectively, for each application. The population of T. urticae decreased from 87.1 to 2.8 mites per leaflet in the first three weeks. After this period, the population of T. urticae was maintained at low levels (≤1.5 mites/leaflet) until the end of the experiment (10th week). Propargite and dimethoate sprayed on the strawberry field did not affect significantly the population of this predaceous mite. For the second experiment, the infestation of T. urticae was 29.1 mites per leaflet, when the acaricide chorfenapyr was applied on the strawberry field. The release of N. californicus (2 mites per plant) was made 2 weeks after spraying the acaricide. The population of T. urticae was maintained at low levels (≤2.8 mites/leaflet) for 8 weeks (evaluation period). The T. urticae infestations in plots with N. californicus were significantly lower than in non-release plots, for the experiments 1 and 2. In the third experiment, the initial infestation of T. urticae was 40.5 mites per leaflet (55.5 active stages/leaflet on release plants; 25.5 active stages/leaflet on non-release plants). Three releases of N. californicus (average rate of 3.0 adult mites/plant), without any acaricide application, were not sufficient to reduce significantly the T. urticae population in release plots (release plants + non-release plants) in 6 weeks from the first release, however, the spider mite population decreased from 55.5 to 7.8 active stages per leaflet on release plants, during this period. Interplant dispersal of N. californicus was low in this strawberry field with high infestation of T. urticae. The studies indicate the viability of the use of this strain of N. californicus for the control of T. urticae in strawberry fields under greenhouse conditions, especially in association with selective acaricides.     

Species- and density-dependent induction of volatile organic compounds by three mite species in cassava and their role in the attraction of a natural enemy

Upon damage by herbivores, plants induce an array of volatile organic compounds (VOCs) that mediate ecological interactions involving communication with organisms of the second and third trophic levels. VOC-mediated tritrophic interactions have largely been studied in various systems, including cassava (Manihot esculenta), but little is known about the chemical nature of herbivore-induced VOCs in this crop and the response they evoke in natural enemies. Several tetranychid and predatory mites are associated with cassava. Here, VOC emissions from uninfested plants and plants infested with 200 or 400 Mononychellus tanajoa, a specialist herbivore on cassava, and the generalists Tetranychus urticae and T. gloveri were measured. Dual-choice experiments were also conducted to assess the preference of inexperienced (reared on prey-infested bean plants) and experienced (adapted on prey-infested cassava plants) predatory mites, Neoseiulus idaeus (Phytoseiidae), between odors of uninfested plants versus odors of plants infested with M. tanajoa, T. urticae or T. gloveri. Two hundred individuals significantly increased the emissions of (Z)-3-hexen-1-ol, (E)-β-ocimene, β-caryophyllene, alloaromadendrene and (E)-geranyl acetone in T. urticae-infested plants, and (E)-β-ocimene and methyl salicylate (MeSA) in T. gloveri-infested plants. Four hundred individuals significantly increased the emissions of (Z)-3-hexen-1-ol, MeSA, α-pinene and D-limonene in M. tanajoa-infested plants. In addition, T. urticae at this density induced (E)-β-ocimene, D-limonene, (E)-geranyl acetone and six compounds that were not detected in other treatments. Tetranychus gloveri-infested plants induced the emissions of (E)-2-hexenal and D-limonene. Regardless of the infesting species, inexperienced N. idaeus did not discriminate between uninfested or infested plants. Upon experience, they discriminated between the odors of uninfested and T. urticae-damaged plants. Our findings reveal that mite infestations in cassava result in density-dependent and species-specific emission of VOCs, and that N. idaeus relies on associative learning to forage for its prey.     

Effects of crop type on persistence and control efficacy of <Emphasis Type="Italic">Beauveria bassiana</Emphasis> against the two spotted spider mite

The influence of five different crops (beans, cucumber, eggplant, maize, and tomato) on persistence and the control efficacy of Beauveria bassiana (Balsamo) Vuillemin to Tetranychus urticae Koch was investigated. Plants were artificially infested with T. urticae. B. bassiana conidia were suspended in Break-thru^® solution (0.01%) at three different concentrations (0.5, 1, 2 g l^?1) and sprayed once onto plants. Plants in the control treatment were sprayed with Break-thru^® solution alone. The number of conidia and levels of mite mortality were evaluated. Colony forming unit counts of conidia recovered from plant leaves declined over time regardless of the concentrations of B. bassiana applied. The rate of decline was different among crops and significantly higher on maize. However, more than 50% of the initial population of conidia were still viable on crops three weeks after application. The longer persistence of B. bassiana on crops did not result in better control of mites. Rather, mortality was positively correlated to the amount of inoculum deposited on leaves immediately after spraying. Mortality of mites was concentration-dependent and influenced by the host crop, with less control observed on maize and tomato than eggplant, beans and cucumber. Results were consistent between trials conducted and confirmed the hypothesis that the type of crop can influence the efficacy of B. bassiana against T. urticae.     

In Vitro Susceptibility of a Large Collection of <Emphasis Type="Italic">Candida</Emphasis> Strains Against Fluconazole and Voriconazole by Using the CLSI Disk Diffusion Assay

We evaluated all Candida spp. isolates obtained from patients admitted to two tertiary care hospitals between 1999 and 2003 in the city of São Paulo, Brazil. The in vitro activities of fluconazole (FCZ) and voriconazole were determined by the agar disk diffusion test using the Clinical and Laboratory Standards Institute M44-A guidelines. The inhibition zone diameters were read and interpreted automatically by the BIOMIC^® image-analysis plate reader system. We tested a total of 4,625 strains, including 2,393 strains of C. albicans (51.7%), 658 of C. tropicalis (14.2%), 503 of C. glabrata (10.9%), 495 of C. parapsilosis (10.7%), 292 of C. rugosa (6.3%), 195 of C. guilliermondii (4.2%) and 89 of other Candida species (2.0%). Only 2.0% of the strains tested were classified as dose-dependent susceptible (DDS), and 5.8% of them were resistant to FCZ. The resistance or DDS to fluconazole was verified mainly among C. glabrata (7.8%), C. krusei (67.9%) and C. rugosa (65.1%). Voriconazole exhibited better activity in vitro than fluconazole, even in isolates fluconazole resistant. The resistance of fluconazole and voriconazole did not increase in the isolates of Candida spp. during the evaluated period.     

Identification of spider-mite species and their endosymbionts using multiplex PCR

Spider mites of the genus Tetranychidae are severe crop pests. In the Mediterranean a few species coexist, but they are difficult to identify based on morphological characters. Additionally, spider mites often harbour several species of endosymbiotic bacteria, which may affect the biology of their hosts. Here, we propose novel, cost-effective, multiplex diagnostic methods allowing a quick identification of spider-mite species as well as of the endosymbionts they carry. First, we developed, and successfully multiplexed in a single PCR, primers to identify Tetranychus urticae, T. evansi and T. ludeni, some of the most common tetranychids found in southwest Europe. Moreover, we demonstrated that this method allows detecting multiple species in a single pool, even at low frequencies (up to 1/100), and can be used on entire mites without DNA extraction. Second, we developed another set of primers to detect spider-mite endosymbionts, namely Wolbachia, Cardinium and Rickettsia in a multiplex PCR, along with a generalist spider-mite primer to control for potential failure of DNA amplification in each PCR. Overall, our method represents a simple, cost-effective and reliable method to identify spider-mite species and their symbionts in natural field populations, as well as to detect contaminations in laboratory rearings. This method may easily be extended to other species.     

Sublethal effects of abamectin on the biological performance of the predatory mite, <Emphasis Type="Italic">Phytoseius plumifer</Emphasis> (Acari: Phytoseiidae)

Traditionally estimating pesticide effects by measuring only lethal effect may underestimate the total negative effect on beneficial arthropods and sublethal effects should be assessed to estimate the total effect of their applications. In this study, sublethal effect of the acaricide abamectin (Vermectin^® 1.8% EC, Giah, Iran) on the predatory mite Phytoseius plumifer (Canestrini &; Fanzago) (Acari: Phytoseiidae) fed on Tetranychus urticae Koch was assessed in laboratory conditions. The adult predators were exposed to the residues of the acaricide on fig leaves and the LC₅₀ value was determined based on a concentration–response analysis. The results showed that sublethal concentrations (LC₁₀, LC₂₀ and LC₃₀) of abamectin severely affected the fecundity and longevity of the treated females of P. plumifer. Furthermore, reproductive and life table parameters of the subsequent generation were affected. The results indicated that adverse effects of abamectin on population growth of P. plumifer were significant, so the results from this study can be used to develop approximate guidelines for the use of abamectin in order to minimize their impact on P. plumifer and related natural enemies.     

Acaricidal property of the essential oil from <Emphasis Type="Italic">Lippia gracilis</Emphasis> against <Emphasis Type="Italic">Tetranychus urticae</Emphasis> and a natural enemy, <Emphasis Type="Italic">Neoseiulus californicus</Emphasis>, under greenhouse conditions

The essential oil from the leaves of Lippia gracilis was investigated for fumigant and residual activity against Tetranychus urticae (Acari: Tetranychidae) and Neoseiulus californicus (Acari: Phytoseiidae). The results were compared to eugenol, Ortus^® and Azamax^®, as positive controls. Gas chromatography (GC) and GC/mass spectrometry analysis enabled the identification of 28 compounds, accounting for 99.1?±?0.6% of the essential oil. The major constituents were carvacrol (61%), p-cymene (11%) and thymol (11%). Mites were more susceptible to the oil in fumigant tests than in residual tests. Among the components, thymol and β-caryophyllene had the greatest fumigant and residual toxicity against T. urticae, respectively. The role of selected constituents (carvacrol, p-cymene, thymol, limonene, β-pinene, 1,8-cineole, terpinolene and β-caryophyllene) in the acaricidal properties of the L. gracilis essential oil is also discussed. Fumigant and residual effects of Lippia oil were more selective than eugenol with regard to a natural enemy of T. urticae, Neoseiulus californicus. Experiments under greenhouse conditions demonstrated greater toxicity of the Lippia oil in comparison to the positive control at 24, 48 and 72 h after treatment. The results suggest that Lippia oil is a good candidate for the formulation of a botanical acaricide for the integrated management of T. urticae.     

Exploring virulence of new and less studied species of <Emphasis Type="Italic">Metarhizium</Emphasis> spp. from Brazil for two-spotted spider mite control

The two-spotted spider mite Tetranychus urticae is an important pest of strawberry crops in Brazil and many other countries. Focus for biocontrol studies involving entomopathogenic fungi has been on three species from the genus Metarhizium: M. anisopliae sensu stricto (s.s.), M. brunneum and M. robertsii. Also, the species Beauveria bassiana has been studied for spider mite control and one isolate (ESALQPL63) is commercially available in Brazil. New and undescribed Metarhizium species have been found recently in Brazil and provide a pool of isolates with potential for biocontrol in Brazil and probably also elsewhere. The mortality of adult females of T. urticae when exposed to four new Brazilian species of Metarhizium was compared to the mortality when exposed to M. anisopliae s.s., M. brunneum, M. pingshaense, M. robertsii and Beauveria bassiana ESALQPL63. Fungal suspensions were sprayed onto mites at 10⁷ conidia/mL with 0.05% Tween 80 in laboratory bio-assays. We measured total mortality and percentage sporulating cadavers 10 days after exposure and calculated median lethal time (LT₅₀). The lowest LT₅₀ (4.0 ± 0.17) was observed for mites treated with Metarhizium sp. Indet. 1 (ESALQ1638), which also performed well with respect to mortality after 10 days and capacity to sporulate from cadavers. Among the other little studied species tested, M. pingshaense (ESALQ3069 and ESALQ3222) and Metarhizium Indet. 2 (ESALQ1476) performed well and were comparable to B. bassiana (ESALQPL63). The new Metarhizium isolates and species thus showed potential for biological control.     

Cereal cyst nematode resistance gene <Emphasis Type="Italic">CreV</Emphasis> effective against <Emphasis Type="Italic">Heterodera filipjevi</Emphasis> transferred from chromosome 6VL of <Emphasis Type="Italic">Dasypyrum villosum</Emphasis> to bread wheat

Cereal cyst nematodes (CCN) are a global economic problem for cereal production. Heterodera filipjevi is one of the most commonly identified and widespread CCN species found in many wheat production regions of the world. Transferring novel genes for resistance to H. filipjevi from wild relatives of wheat is a promising strategy for protection of wheat crops. A set of wheat–Dasypyrum villosum chromosome addition lines, T6V#4S·6AL translocation lines and their donor parental lines were tested for their response to the nematode. D. villosum and wheat–D. villosum disomic addition line DA6V#4 were resistant. As T6V#4S·6AL translocation lines were susceptible, resistance was presumed to be located on chromosome 6V#4L. The objective of this study was to produce and characterize wheat–6V#4L translocations and confirm the chromosome location of the resistance. Introgression lines T6V#4L·6AS, T6V#4L-4BL·4BS and DT6V#4L were developed and subjected to molecular cytogenetic analysis. These and four additional wheat–6V#4 introgression lines were tested for response to H. filipjevi in the greenhouse. The results indicated that introgression lines DA6V#4, T6V#4L·6AS, T6V#4L-4BL·4BS, T6V#4L·6V#4S-7BS and DT6VL#4 had higher levels of H. filipjevi resistance than their recurrent parent. However, Del6V#4L-1 and translocation line T6V#4S·6AL were equally susceptible to wheat cv. Chinese Spring. The CCN resistance gene, temporarily named CreV, was therefore physically mapped to chromosome arm 6V#4L FL 0.80–1.00. Translocation chromosomes T6V#4L·6AS transferred to a modern wheat cv. Aikang 58 with its co-dominant molecular markers could be utilized as a novel germplasm for CCN resistance breeding in wheat.     

Larval aquatic and terrestrial mites infesting parthenogenetic <Emphasis Type="Italic">Ischnura hastata</Emphasis> (Odonata: Coenagrionidae) from the Azores islands

We report here the prevalence of parasitism by water mites (Arrenurus sp.) and terrestrial mites (Leptus killingtoni) on parthenogenetic Ischnura hastata (Odonata: Coenagrionidae) from the Azores islands. Leptus killingtoni was only found on the island of Pico, and the prevalence of infestation was highly variable among the different ponds studied, ranging from 0 to 41%. Leptus killingtoni was observed on three of the four odonate species from the archipelago: I. hastata, I. pumilio, and Sympetrum fonscolombii, all of them new hosts for this species. Aquatic mites have been found parasitizing I. hastata females on the island of São Miguel. The prevalence of mite parasitism by Arrenurus sp. on I. hastata was very low, ranging from 12% (2003) to 1% (2008), and in most of the studied ponds, no mites were found attached to females. Although I. hastata coexists with a sexual congener species in the Azores (I. pumilio), they are syntopic in only a small fraction of ponds. Therefore, a comparison between I. hastata and I. pumilio was insufficient to test the predictions of the Red Queen Hypothesis, and further research on parasitism rates in both species needs to be done. In any case, the low prevalence of mite parasitism found in the Azores, coupled with the fact that most of the populations in the archipelago are almost free from competitors and predators, could explain the persistence of these I. hastata parthenogenetic populations, despite their low levels of genetic variation.     

The interaction of two-spotted spider mites, <Emphasis Type="Italic">Tetranychus urticae</Emphasis> Koch,with Cry protein production and predation by <Emphasis Type="Italic">Amblyseius andersoni</Emphasis> (Chant) in Cry1Ac/Cry2Ab cotton and Cry1F maize

Crops producing insecticidal crystal (Cry) proteins from the bacterium, Bacillus thuringiensis (Bt), are an important tool for managing lepidopteran pests on cotton and maize. However, the effects of these Bt crops on non-target organisms, especially natural enemies that provide biological control services, are required to be addressed in an environmental risk assessment. Amblyseius andersoni (Acari: Phytoseiidae) is a cosmopolitan predator of the two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae), a significant pest of cotton and maize. Tri-trophic studies were conducted to assess the potential effects of Cry1Ac/Cry2Ab cotton and Cry1F maize on life history parameters (survival rate, development time, fecundity and egg hatching rate) of A. andersoni. We confirmed that these Bt crops have no effects on the biology of T. urticae and, in turn, that there were no differences in any of the life history parameters of A. andersoni when it fed on T. urticae feeding on Cry1Ac/Cry2Ab or non-Bt cotton and Cry1F or non-Bt maize. Use of a susceptible insect assay demonstrated that T. urticae contained biologically active Cry proteins. Cry proteins concentrations declined greatly as they moved from plants to herbivores to predators and protein concentration did not appear to be related to mite density. Free-choice experiments revealed that A. andersoni had no preference for Cry1Ac/Cry2Ab cotton or Cry1F maize-reared T. urticae compared with those reared on non-Bt cotton or maize. Collectively these results provide strong evidence that these crops can complement other integrated pest management tactics including biological control.     

Antixenosis and antibiosis response of common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis>) to two-spotted spider mite (<Emphasis Type="Italic">Tetranychus urticae</Emphasis>)

The two-spotted spider mite, Tetranychus uticae Koch (Acari: Tetranychidae), is globally one of the most devastating pests that feed on numerous crops, including common bean (Phaseolus vulgaris L.). This study was aimed to evaluate the effects of genotype and morphological attributes of common bean on T. uticae. Forty common bean accessions were used to investigate antixenosis and antibiosis through assessing mite feeding preference and reproduction under laboratory conditions. Three resistant (i.e., 56, 63, 238) and two susceptible (i.e., 182, 236) accessions, along with cultivars Naz (resistant) and Akhtar (susceptible), were used in a life-table study. Both antixenosis and antibiosis mechanism were observed in all of the accessions, albeit a negative correlation occurred. Significant differences were observed for all traits of T. urticae: developmental time of immature stages, reproduction, adult longevity and life-table parameters. Based on the intrinsic rate of increase, the accessions 56, 63, 182, 238, and cv. Naz impose high antibiotic effects on T. urticae. Although significant variation existed among accessions for morphological factors, only glandular trichomes correlated with mite fecundity and feeding preference.     

Species composition and arthropod pest feeding of phytoseiid mites in a Japanese pear greenhouse

A population survey of spider mites and phytoseiid mites was conducted on Japanese pear leaves in a greenhouse. For the survey, the method to estimate phytoseiid mite species composition using quantitative sequencing was modified to be applicable for phytoseiid mite species inhabiting in the greenhouse. Results show the dominant appearance of Neoseiulus californicus (McGregor), Neoseiulus womersleyi (Schicha), and Neoseiulus makuwa (Ehara) from the end of June to late September and their contribution in spider mite control. PCR-based method to detect the ribosomal internal transcribed spacer (ITS) sequences of spider mites from phytoseiid mites was developed. The method shows sensitivity to detect the ITS sequences of Tetranychus urticae Koch from single N. californicus adult at 168 h after ingestion of the spider mite. PCR-based method to detect the cytochrome c oxidase subunit I sequences of several arthropod pests belonging to Hemiptera, Thysanoptera, and Acari from phytoseiid mites was also developed. Results show that phytoseiid mites prey on Eriophyes chibaensis (Kadono) and Aphis gossypii (Glover), in addition to spider mites.     

Evaluations of the Removal of <Emphasis Type="Italic">Varroa destructor</Emphasis> in Russian Honey Bee Colonies that Display Different Levels of <Emphasis Type="Italic">Varroa</Emphasis> Sensitive Hygienic Activities

The removal of Varroa destructor was assessed in Russian honey bee (RHB) colonies with known levels of Varroa Sensitive Hygienic (VSH) and brood removal activities. The expression of grooming behaviour using individual bees was also measured using three groups of RHB displaying different VSH levels: low hygiene (RHB-LH, < 35% VSH), medium hygiene (RHB-MH, 35–70%) and high hygiene (RHB-HH, > 70%). Italian colonies (5.43–71.62% VSH) served as control. Our results demonstrated, for the first time, significant relationships between two hygienic responses (VSH activity measured as percent change in infestation and the actual brood removal of Varroa-infested donor comb) and two measurements of mite fall (trapped old mites/trapped mites or O/T and trapped young mites/trapped mites or Y/T). However, these relationships were only observed in RHB colonies. In addition, the RHB colonies that displayed the highest levels of hygiene (RHB-HH) also groomed longer in response to the presence of a V. destructor mite based on individual bee assays. The positive regressions between the two hygienic measurements and O/T and their negative regressions with Y/T suggest that the removal of infested brood prevented successful mite reproduction, ultimately suppressing V. destructor infestations in the RHB colonies. In addition, it is demonstrated that RHB resistance to V. destructor rests on both an increased hygienic response and the removal of phoretic mites, released by hygienic behaviour, through grooming. Both resistance traits are reflected in the O/T and Y/T ratios found in trapped mites from RHB colonies. None of the measurements involving mite injuries were associated with any measurements of hygiene and colony infestations.     

Expression Patterns of ABC Transporter Genes in Fluconazole-Resistant <Emphasis Type="Italic">Candida glabrata</Emphasis>

Clinical management of fungal diseases is compromised by the emergence of antifungal drug resistance in fungi, which leads to elimination of available drug classes as treatment options. An understanding of antifungal resistance at molecular level is, therefore, essential for the development of strategies to combat the resistance. This study presents the assessment of molecular mechanisms associated with fluconazole resistance in clinical Candida glabrata isolates originated from Iran. Taking seven distinct fluconazole-resistant C. glabrata isolates, real-time PCRs were performed to evaluate the alternations in the regulation of the genes involved in drug efflux including CgCDR1, CgCDR2, CgSNQ2, and CgERG11. Gain-of-function (GOF) mutations in CgPDR1 alleles were determined by DNA sequencing. Cross-resistance to fluconazole, itraconazole, and voriconazole was observed in 2.5 % of the isolates. In the present study, six amino acid substitutions were identified in CgPdr1, among which W297R, T588A, and F575L were previously reported, whereas D243N, H576Y, and P915R are novel. CgCDR1 overexpression was observed in 57.1 % of resistant isolates. However, CgCDR2 was not co-expressed with CgCDR1. CgSNQ2 was upregulated in 71.4 % of the cases. CgERG11 overexpression does not seem to be associated with azole resistance, except for isolates that exhibited azole cross-resistance. The pattern of efflux pump gene upregulation was associated with GOF mutations observed in CgPDR1. These results showed that drug efflux mediated by adenosine-5-triphosphate (ATP)-binding cassette transporters, especially CgSNQ2 and CgCDR1, is the predominant mechanism of fluconazole resistance in Iranian isolates of C. glabrata. Since some novel GOF mutations were found here, this study also calls for research aimed at investigating other new GOF mutations to reveal the comprehensive understanding about efflux-mediated resistance to azole antifungal agents.     

Chromosomal Sil system contributes to silver resistance in <Emphasis Type="Italic">E. coli</Emphasis> ATCC 8739

The rise of antibiotic resistance in pathogenic bacteria is endangering the efficacy of antibiotics, which consequently results in greater use of silver as a biocide. Chromosomal mapping of the Cus system or plasmid encoded Sil system and their relationship with silver resistance was studied for several gram-negative bacteria. However, only few reports investigated silver detoxification mediated by the Sil system integrated in Escherichia coli chromosome. Accordingly, this work aimed to study the Sil system in E. coli ATCC 8739 and to produce evidence for its role in silver resistance development. Silver resistance was induced in E. coli ATCC 8739 by stepwise passage in culture media containing increasing concentrations of AgNO₃. The published genome of E. coli ATCC 8739 contains a region showing strong homology to the Sil system genes. The role of this region in E. coli ATCC 8739 was assessed by monitoring the expression of silC upon silver stress, which resulted in a 350-fold increased expression. De novo sequencing of the whole genome of a silver resistant strain derived from E. coli ATCC 8739 revealed mutations in ORFs putative for SilR and CusR. The silver resistant strain (E. coli AgNO₃R) showed constitutive expression of silC which posed a cost of fitness resulting in retarded growth. Furthermore, E. coli AgNO₃R exhibited cross-resistance to ciprofloxacin and a slightly increased tolerance to ampicillin. This study demonstrates that E. coli is able to develop resistance to silver, which may pose a threat towards an effective use of silver compounds as antiseptics.     

Banding and FISH in three species of <Emphasis Type="Italic">Vernonia</Emphasis>, subsection <Emphasis Type="Italic">Macrocephalae</Emphasis> (Asteraceae,Vernonieae)

To complement our knowledge about the karyotypes of the genus Vernonia Schreb., different techniques of chromosome banding, including AgNOR, triple staining with fluorochromes CMA/DA/DAPI (CDD), and fluorescence in situ hybridization (FISH) for the 45S rDNA probe, were applied to three species of subsection Macrocephalae. Vernonia bardanoides was collected from an area of cerrado (savanna) vegetation in Itirapina, São Paulo State, Brazil, and V. linearifolia and V. tomentella were collected from areas of rocky, open altitudinal vegetation in Joaquim Felicio and Diamantina, respectively, in Minas Gerais State. All species showed two terminal CMA⁺ and NOR bands. FISH indicated two terminal 45S rDNA sites in V. linearifolia and V. tomentella, and six in V. bardanoides.     

A new species of <Emphasis Type="Italic">Senecio</Emphasis> (Asteraceae: Senecionae) from southeastern Brazil

Senecio altimontanus, a new species from the states of Minas Gerais and São Paulo, Brazil, is described and illustrated. Senecio altimontanus appears to be restricted to high-elevation grasslands (campos de altitude). This new species is characterized by its usually simple stems that are densely leafy apically, coriaceous leaves, and 1–5 corymbiform capitulescences. It is closely related to Senecio apensis from which it differs by the size, shape, and margins of the leaves, the number of phyllaries, and pubescence of the cypselae.     

Characterization of the genetic diversity of <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis> in São Paulo city,Brazil

Background

Tuberculosis is a major health problem in São Paulo, Brazil, which is the most populous and one of the most cosmopolitan cities in South America. To characterize the genetic diversity of Mycobacterium tuberculosis in the population of this city, the genotyping techniques of spoligotyping and MIRU were applied to 93 isolates collected in two consecutive years from 93 different tuberculosis patients residing in São Paulo city and attending the Clemente Ferreira Institute (the reference clinic for the treatment of tuberculosis).

Findings

Spoligotyping generated 53 different spoligotype patterns. Fifty-one isolates (54.8%) were grouped into 13 spoligotyping clusters. Seventy- two strains (77.4%) showed spoligotypes described in the international databases (SpolDB4, SITVIT), and 21 (22.6%) showed unidentified patterns. The most frequent spoligotype families were Latin American Mediterranean (LAM) (26 isolates), followed by the T family (24 isolates) and Haarlem (H) (11 isolates), which together accounted for 65.4% of all the isolates. These three families represent the major genotypes found in Africa, Central America, South America and Europe. Six Spoligo-International-types (designated SITs by the database) comprised 51.8% (37/72) of all the identified spoligotypes (SIT53, SIT50, SIT42, SIT60, SIT17 and SIT1). Other SITs found in this study indicated the great genetic diversity of M. tuberculosis, reflecting the remarkable ethnic diversity of São Paulo city inhabitants. The MIRU technique was more discriminatory and did not identify any genetic clusters with 100% similarity among the 93 isolates. The allelic analysis showed that MIRU loci 26, 40, 23 and 10 were the most discriminatory. When MIRU and spoligotyping techniques were combined, all isolates grouped in the 13 spoligotyping clusters were separated.

Conclusions

Our data indicated the genomic stability of over 50% of spoligotypes identified in São Paulo and the great genetic diversity of M. tuberculosis isolates in the remaining SITs, reflecting the large ethnic mix of the São Paulo city inhabitants. The results also indicated that in this city, M. tuberculosis isolates acquired drug resistance independently of genotype and that resistance was more dependent on the selective pressure of treatment failure and the environmental circumstances of patients.