Search

Bristle crabs of the genus Pilumnus (Brachyura: Heterotremata: Pilumnidae) are common inhabitants of European waters. They are easily identifiable as a genus, but with the exception of P. inermis, intrageneric classification turns out to be quite complex. There is no general agreement on the number and distinction of species. Therefore, this genus is well-suited for comparative molecular studies. Specimens of the Pilumnus hirtellus complex, here defined as including Pilumnus hirtellus, P. villosissimus, P. spinifer, P. aestuarii, and an undescribed species, were gathered from throughout the Mediterranean Sea and the eastern Atlantic Ocean. DNA sequence data were obtained from the barcoding region of the cytochrome oxidase 1 mitochondrial gene and used for reconstruction of a phylogenetic tree and a haplotype network. The morphology of the gastric ossicles was compared in the search of separating characters. Our results give evidence for five genetic clusters within the P. hirtellus complex. There is negligible geographic variation within these clusters. Unambiguous mtDNA sequences within morphologically variable local populations argue against possible hybridization. The here encountered evolutionary units are relatively young and possibly allow to study ongoing processes of morphological, genetic, and ecological differentiation, leading to speciation and radiations in the coastal marine environment.

The pathway and frequency of species' introductions can affect the extent, impact, and management of biological invasions. Here, we examine the pathway of introduction of the aquatic plant Cabomba caroliniana (fanwort) into Canada and the northern United States using plastid DNA sequence (intergenic spacers atpF-atpH, trnH-psbA, and trnL-trnF) and DNA content analyses. We test the hypothesis that the spread of fanwort is a result of commercial trade by comparing a Canadian population (Kasshabog Lake, ON) to native populations from southern U.S., introduced populations in northern U.S., and plants from commercial retailers. Thirteen plastid haplotypes were identified throughout North America, including one dominant haplotype, which was present in all C. caroliniana populations. Several rare haplotypes were used to infer shared colonization history. In particular, the Canadian population shared two rare alleles with a population from Massachusetts, suggesting range expansion of C. caroliniana from the northern U.S. However, the possibility of a commercial introduction cannot be excluded, as common alleles were shared between the Canadian population and both commercial and southern U.S. sources. Variation in C. caroliniana genome size was bimodal and populations were classified into "high" and "low" categories. The Canadian population had DNA contents similar to several northern U.S. populations (low DNA content). This may provide additional support for range expansion from these introduced populations rather than from commercial sources or populations in the southern U.S., which had high DNA content.

U zemljama Zapadnog Balkana zabeleženo je intenzivno sušenje brojnih drvenastih i žbunastih vrsta tokom poslednje decenije. Simptomi bolesti bili su tipični za one koje izazivaju gljivični patogeni iz familije Botryosphaeriaceae. Ciljevi ovog istraživanja bili su: 1. izolacija gljiva iz familije Botryosphaeriaceae iz obolelih stabala u Srbiji, Crnoj Gori i Bosni i Hercegovini i njihova karakterizacija, 2. da se utvrde domaćini i distribucija gljiva iz familije Botryosphaeriaceae u Srbiji, Crnoj Gori i Bosni i Hercegovini, u kontinentalnom i mediteranskom području, 3. da se utvrde domaćini gljive Diplodia sapinea u Srbiji i Crnoj Gori, 4. da se utvrdi genetički diverzitet i razmena genotipova između različitih populacija gljiva Neofusicoccum parvum i D. sapinea u Srbiji i Crnoj Gori. Karakterizacija izolata najpre je izvršena uz pomoć DNA sekvenci internog transkribovanog spejsera (ITS) rDNA i identifikovano je šest glavnih klada. Reprezentativni izolati svake klade su zatim dodatno okarakterisani uz pomoć DNA sekvenci translacionog elongacionog faktora 1-alpha (TEF 1-α), β-tubulin-2 (BT2) dela gena i velike podjedinice (LSU) rRNA, kao i morfoloških karakteristika aseksualnog izgleda ovih gljiva. D. sapinea-slični izolati su dodatno okarakterisani uz pomoć TEF 1-α dela gena. Genetički diverzitet i protok genotipova gljiva N. parvum i D. sapinea utvrđeni su uz pomoć DNA sekvenci ITS regiona, TEF 1-α, BT2 dela gena i mikrosatelitskih markera. Izolati gljive D. sapinea poreklom iz različitih domaćina proučavani su na nivou populacija poreklom iz Cedrus spp. i Pinus spp. Izolati gljiva D. sapinea i N. parvum koji potiču iz regiona sa različitim uslovima spoljašnje sredine proučavani su na nivou populacija iz kontinentalnog (CR) i Mediteranskog (MR) regiona. Identifikovano je deset vrsta gljiva iz familije Botryosphaeriaceae od kojih su osam i to Dothiorella sarmentorum, Neofusicoccum parvum, Botryosphaeria dothidea, Phaeobotryon cupressi, Sphaeropsis visci, Diplodia seriata, D. sapinea i Diplodia mutila bili nauci poznati taksoni. Preostale dve vrste mogle su biti identifikovane samo do nivoa roda i to kao Dothiorella spp. Takođe, opisan je Dichomera sin-aseksualni izgled gljiva D. sapinea, Dothiorella sp. 2 i B. dothidea, kao i jedinstvene morfološke karakteristike osam nauci poznatih vrsta... Extensive die-back and mortality of various trees and shrubs has been observed in parts of the Western Balkans region during the past decade. The disease symptoms have been typical of those caused by fungal pathogens residing in the Botryosphaeriaceae. The aims of this study were to 1. isolate and characterize Botryosphaeriaceae species associated with diseased trees in Serbia, Montenegro, Bosnia and Herzegovina, 2. consider the host range and distribution of Botryosphaeriaceae species, spanning a Mediterranean and a Continental region, 3. consider the host range of Diplodia sapinea in Serbia and Montenegro 4. determine genetic diversity and genotype flow between different populations of Neofusicoccum parvum and D. sapinea from Serbia and Montenegro. Isolates were initially characterized based on the DNA sequence data for the internal transcribed spacer (ITS) rDNA and six major clades were identified. Representative isolates from each clade were further characterized using DNA sequence data for the translation elongation factor 1-alpha (TEF 1-α), β-tubulin-2 (BT2) and large subunit (LSU) rRNA gene regions, as well as the morphology of the asexual morphs. D. sapinea-like isolates were additionally characterized using TEF 1- α. Genetic diversity and genotype flow of N. parvum and D. sapinea were determined using DNA sequence data of the ITS region, TEF 1-α, BT2 and microsatellite markers. The relationship between isolates of D. sapinea from different hosts was considered using populations from Cedrus spp. and Pinus spp. The relationships between isolates from environmentally different regions included populations of each species from the Continental (CR) and Mediterranean (MR) regions. Ten species of the Botryosphaeriaceae were identified of which eight, i.e. Dothiorella sarmentorum, Neofusicoccum parvum, Botryosphaeria dothidea, Phaeobotryon cupressi, Sphaeropsis visci, Diplodia seriata, D. sapinea and Diplodia mutila were known taxa. The remaining two species could be identified only as Dothiorella spp. Dichomera syn-asexual morphs of D. sapinea, Dothiorella sp. 2 and B. dothidea, as well as unique morphological characters for a number of the known species are described...

Mestrado em Ecologia Aplicada Species persist over time, due to exchange of individuals between subpopulations. In the marine environment, most benthic organisms have complex life cycles including pelagic larvae that are transported by ocean currents promoting species dispersal. Larval dispersal connects geographically distant populations and determines population structure. The knowledge about this biologic process provides relevant information for conservation of marine populations. This study investigates the genetic structure and connectivity of deep-sea mussel populations between fragmented habitats in the NE Atlantic and Mediterranean. The mitochondrial Cytochrome Oxidase I (mtCOI) gene was used to analyze site-specific genetic diversity and the population structure of two mussel species, Idas modiolaeformis and “Idas” simpsoni. Populations of each species are not geographically isolated. The presence of one dominant haplotype for each species suggests shared ancestral polymorphisms between Mediterranean and NE Atlantic populations. The overall high genetic differentiation observed in I. modiolaeformis indicates that the metapopulation is structured. Distant populations, located in Atlantic and E Mediterranean, revealed low genetic distances, suggesting gene flow between the two regions. Genetic and geographical distances support an island model of I. modiolaeformis population structure. A major drawback of this study is concerned with the discrepant number of individuals among populations. Further research will be needed, using more specimens and other gene markers, to investigate connectivity patterns at different spatial scales. As espécies persistem ao longo do tempo devido à troca de indivíduos entre subpopulações. No ambiente marinho, a maioria dos organismos bentónicos têm ciclos de vida complexos, envolvendo larvas pelágicas que são transportadas por correntes oceânicas contribuindo para dispersão das espécies. A dispersão larvar estabelece conectividade entre populações geograficamente separadas e afeta a estrutura da população. O conhecimento deste processo biológico promove informações importantes para a conservação de populações marinhas. Este estudo investiga a estrutura genética e conectividade de populações de mexilhão de profundidade entre habitats fragmentados no NE Atlântico e Mediterrânico. O gene mitocondrial, Citocromo Oxidase I (mtCOI), foi utilizado para analisar diversidade genética por local e a estrutura populacional de duas espécies de mexilhão, Idas modiolaeformis e "Idas" simpsoni. As populações de cada uma das espécies não se encontram geograficamente isoladas. A presença de um haplótipo dominante para cada espécie sugere a partilha de polimorfismos ancestrais entre populações do Mediterrâneo e do NE Atlântico. As populações de I. modiolaeformis demonstraram uma elevada diferenciação genética, indicando estruturação da metapopulação. Populações distantes umas das outras, localizadas no Atlântico e E Mediterrâneo, revelaram baixas distâncias genéticas, sugerindo fluxo genético entre as duas regiões. Distâncias genéticas e geográficas suportam o modelo de ilha como o modelo para a estrutura populacional de I. modiolaeformis. Uma grande desvantagem deste estudo está relacionada com o número discrepante de indivíduos entre populações. Para investigar os padrões de conectividade em diferentes escalas espaciais serão necessários mais estudos, utilizando mais espécimes e outros marcadores genéticos.

Mestrado em Ecologia Aplicada Submitted by Carla Jesus (cjesus@ua.pt) on 2017-05-17T09:44:25Z No. of bitstreams: 1 MScThesis_AnaRitaAndrilMonteiro_60522.pdf: 3534346 bytes, checksum: fd367e8c643b5a1a5ae759f5272a0ae1 (MD5) Made available in DSpace on 2017-05-17T09:44:25Z (GMT). No. of bitstreams: 1 MScThesis_AnaRitaAndrilMonteiro_60522.pdf: 3534346 bytes, checksum: fd367e8c643b5a1a5ae759f5272a0ae1 (MD5) Previous issue date: 2017-01-05

Reported malaria cases in the Americas had been reduced to about one-half million by 2012. To advance towards elimination of this disease, it is necessary to gain insights into how the malaria parasite is evolving, including the emergence, spread and persistence of new haplotypes in affected regions. In here, the genetic diversity of the three major P. vivax merozoite genes was analyzed. From P. vivax-infected blood samples obtained in southern Mexico (SMX) during 2006–2007, nucleotide sequences were achieved for: the 42 kDa carboxyl fragment of the merozoite surface protein-1 (msp1 42 ), domains I-II of the apical membrane antigen-1 (ama1 I-II ), and domain II of the Duffy binding protein (dbp II ). Gene polymorphism was examined and haplotype networks were developed to depict parasite relationships in SMX. Then genetic diversity, recombination and natural selection were analyzed and the degree of differentiation was determined as FST values. The diversity of P. vivax merozoite genes in SMX was less than that of parasites from other geographic origins, with dbp II

Pinus nigra J.F. Arnold, European black pine, is a typical component of Mediterranean and sub-Mediterranean coniferous forests with highly fragmentary distribution. Western Mediterranean populations of this species have been studied genetically to date, while eastern populations from the central Balkans, which are larger and more abundant, are still genetically understudied. We analyzed seven populations of P. nigra representing all infraspecific taxa recognized within the central Balkans (subspecies nigra with varieties nigra and gocensis Dordevic; and subspecies pallasiana (Lamb.) Holmboe with varieties pallasiana and banatica (Endl.) Georgescu et Ionescu), with three chloroplast microsatellites (cpDNA SSRs) and one mitochondrial (mtDNA) locus. Although our molecular data failed to support circumscription of studied infraspecific taxa, we found that genetic patterns at both genomes are in accordance with those found previously in westward populations of this species, that is - exceptionally high levels of genetic diversity (H-T = 0.949) and low genetic differentiation (G(ST) = 0.024) at the cpDNA level, and moderate levels of genetic diversity (H-T = 0.357) and genetic differentiation (G(ST) = 0.358) at the mtDNA level. Based on genealogical relations of mtDNA types currently present in Balkans' and Iberian/African populations, we inferred that the ancestral gene pool of P. nigra already harbored polymorphism at position 328 prior to the divergence to two lineages currently present in westward and eastward parts of the species range distribution. Subsequent occurrence of three mutations, which distinguish these two lineages, suggests their long-term isolation. Ministry of Education, Science, and Technology of the Republic of Serbia {[}173029, 173030]

Brevipalpus phoenicis es considerado un vector importante del Citrus leprosis virus C (CiLV-C) agente causal de la leprosis tipo citoplásmico, enfermedad que afecta cítricos de variedades dulces principalmente. En este estudio se obtuvieron muestras de Brevipalpus phoenicis en huertos de naranja dulce (Citrus sinensis) de ocho y 12 localidades de Brasil y México, respectivamente. La estructura poblacional, la variación genética, así como el análisis filogenético de las muestras recolectadas fueron estudiados a partir de secuencias de nucleótidos del gen citocromo oxidasa subunidad I (COI). Se observó la existencia de dos grupos morfológicos (“B” y “C”) en muestras de ambos países, siendo éste el primer reporte del grupo “C” en el territorio mexicano. La diversidad genética en las poblaciones de Brasil fue mayor con la presencia de 17 haplotipos mientras que las poblaciones mexicanas provenientes de Tabasco, Veracruz y Chiapas sólo presentaron cuatro. La mayor variación (44.92%) se observó entre las localidades de muestreo de cada país, seguida por la variación entre muestras de cada localidad (29.89%). Aunque el nivel de variación entre países fue bajo (25.18%), este fue significativo, sugiriendo que las poblaciones de ambos países están geográficamente estructuradas. _______________ GENETIC VARIATION OF POPULATIONS OF Brevipalpus phoenicis (ACARI: TENUIPALPIDAE) VECTOR OF Citrus leprosis virus CYTOPLASMIC TYPE. ABSTRACT: Brevipalpus phoenicis is considered an important vector of Citrus leprosis virus C (CiLV-C) causal agent of the cytoplasmic-type leprosis, a disease affecting sweet citrus orchards. Samples of Brevipalpus phoenicis were obtained from sweet orange (Citrus sinensis) orchards in eight and twelve localities in Brazil and Mexico respectively. Population genetic structure and genetic variation was assessed in the samples collected by analyzing nucleotide sequences of cytochrome oxidase subunit I gene (COI). We found the existence of two morphological groups (“B” and “C”) in samples of both countries; this represents the first report of group “C” for Mexico. The genetic diversity among Brazilian populations was higher with seventeen haplotypes while Mexican populations from Tabasco, Veracruz and Chiapas only presented four. The biggest variation was observed among localities of each country (44.92%), followed by variation among sampling sites of each locality (29.89%). The variation between countries was low (25.18%), however, still significant suggesting a geographically structured populations between Brazil and Mexico.

Background Plasmodium vivax is a protozoan parasite with an extensive worldwide distribution, being highly prevalent in Asia as well as in Mesoamerica and South America. In southern Mexico, P. vivax transmission has been endemic and recent studies suggest that these parasites have unique biological and genetic features. The msp1 gene has shown high rate of nucleotide substitutions, deletions, insertions, and its mosaic structure reveals frequent events of recombination, maybe between highly divergent parasite isolates. Methods The nucleotide sequence variation in the polymorphic icb5-6 fragment of the msp1 gene of Mexican and worldwide isolates was analysed. To understand how genotype diversity arises, disperses and persists in Mexico, the genetic structure and genealogical relationships of local isolates were examined. To identify new sequence hybrids and their evolutionary relationships with other P. vivax isolates circulating worldwide two haplotype networks were constructed questioning that two portions of the icb5-6 have different evolutionary history. Results Twelve new msp1 icb5-6 haplotypes of P. vivax from Mexico were identified. These nucleotide sequences show mosaic structure comprising three partially conserved and two variable subfragments and resulted into five different sequence types. The variable subfragment sV1 has undergone recombination events and resulted in hybrid sequences and the haplotype network allocated the Mexican haplotypes to three lineages, corresponding to the Sal I and Belem types, and other more divergent group. In contrast, the network from icb5-6 fragment but not sV1 revealed that the Mexican haplotypes belong to two separate lineages, none of which are closely related to Sal I or Belem sequences. Conclusions These results suggest that the new hybrid haplotypes from southern Mexico were the result of at least three different recombination events. These rearrangements likely resulted from the recombination between haplotypes of highly divergent lineages that are frequently distributed in South America and Asia and diversified rapidly.

The Holarctic genus Reticulitermes shows seven species within the Mediterranean Basin. While phylogeny and systematics at continental level has been deeply investigated, a few studies concentrated on local ranges. To gain a clearer picture of the diversity and evolution of the Italian species Reticulitermes lucifugus, we analyzed the mitochondrial cytochrome oxidase II (COII) gene marker in newly collected colonies across the Peninsula. Data were gathered with all R. lucifugus sequences available from previous studies; COII sequences of the closely related Iberian taxa were also added to the data set. Maximum-likelihood, median-joining and statistical parsimony network elaborations on the resulting 119 colonies all agreed in indicating that: (i) the Sardo-Corsican subspecies R. lucifugus corsicus, strictly related to Southern Italian populations (including the Sicilian ones), is phylogenetically closer to the Iberian Reticulitermes grassei; and (ii) R. lucifugus lucifugus peninsular populations are structured into three clusters. The phylogenetic relationships and the biogeography of extant taxa suggest a scenario in which R. lucifugus ancestors colonized the Italian region through the Sardo-Corsican microplate during its Oligocene-Miocene anticlockwise rotation. Moreover, well after the colonization took place, northward range expansion might have produced the presently observed genetic diversity, as inferred from haplotype and nucleotide diversity estimates. On the whole, this study highlights the evolution of Italian Reticulitermes taxa and supports the importance of a wide taxon sampling especially when dealing with organisms easily dispersed by human activities.

Background The soil fungus Rhizoctonia solani anastomosis group 3 (AG-3) is an important pathogen of cultivated plants in the family Solanaceae. Isolates of R. solani AG-3 are taxonomically related based on the composition of cellular fatty acids, phylogenetic analysis of nuclear ribosomal DNA (rDNA) and beta-tubulin gene sequences, and somatic hyphal interactions. Despite the close genetic relationship among isolates of R. solani AG-3, field populations from potato and tobacco exhibit comparative differences in their disease biology, dispersal ecology, host specialization, genetic diversity and population structure. However, little information is available on how field populations of R. solani AG-3 on potato and tobacco are shaped by population genetic processes. In this study, two field populations of R. solani AG-3 from potato in North Carolina (NC) and the Northern USA; and two field populations from tobacco in NC and Southern Brazil were examined using sequence analysis of two cloned regions of nuclear DNA (pP42F and pP89). Results Populations of R. solani AG-3 from potato were genetically diverse with a high frequency of heterozygosity, while limited or no genetic diversity was observed within the highly homozygous tobacco populations from NC and Brazil. Except for one isolate (TBR24), all NC and Brazilian isolates from tobacco shared the same alleles. No alleles were shared between potato and tobacco populations of R. solani AG-3, indicating no gene flow between them. To infer historical events that influenced current geographical patterns observed for populations of R. solani AG-3 from potato, we performed an analysis of molecular variance (AMOVA) and a nested clade analysis (NCA). Population differentiation was detected for locus pP89 (Φ ST = 0.257, significant at P < 0.05) but not for locus pP42F (Φ ST = 0.034, not significant). Results based on NCA of the pP89 locus suggest that historical restricted gene flow is a plausible explanation for the geographical association of clades. Coalescent-based simulations of genealogical relationships between populations of R. solani AG-3 from potato and tobacco were used to estimate the amount and directionality of historical migration patterns in time, and the ages of mutations of populations. Low rates of historical movement of genes were observed between the potato and tobacco populations of R. solani AG-3. Conclusion The two sisters populations of the basidiomycete fungus R. solani AG-3 from potato and tobacco represent two genetically distinct and historically divergent lineages that have probably evolved within the range of their particular related Solanaceae hosts as sympatric species., BMC Evolutionary Biology, 7, ISSN:1471-2148

Background Genetic variation in the ribosomal DNA (rDNA) internal transcribed spacer (ITS) region has been studied among fungi. However, the numbers of ITS sequence polymorphisms in the various Candida species and their associations with sources of invasive fungal infections remain poorly investigated. Here, we characterized the intraspecific and interspecific ITS diversity of Candida spp. strains collected from patients with bloodstream or oroesophageal candidiasis. Methods We selected cultures of representative medically important species of Candida as well as some rare and emerging pathogens. Identification was performed by micromorphology and by biochemical testing using an ID32C® system, as well as by the sequencing of rDNA ITS. The presence of intraspecific ITS polymorphisms was characterized based on haplotype networks, and interspecific diversity was characterized based on Bayesian phylogenetic analysis. Results Among 300 Candida strains, we identified 76 C. albicans, 14 C. dubliniensis, 40 C. tropicalis, 47 C. glabrata, 34 C. parapsilosis (sensu stricto), 31 C. orthopsilosis, 3 C. metapsilosis, 21 Meyerozyma guilliermondii (C. guilliermondii), 12 Pichia kudriavzevii (C. krusei), 6 Clavispora lusitaniae (C. lusitaniae), 3 C. intermedia, 6 Wickerhamomyces anomalus (C. pelliculosa), and 2 C. haemulonii strains, and 1 C. duobushaemulonii, 1 Kluyveromyces marxianus (C. kefyr), 1 Meyerozyma caribbica (C. fermentati), 1 Pichia norvegensis (C. norvegensis), and 1 Lodderomyces elongisporus strain. Out of a total of seven isolates with inconsistent ID32C® profiles, ITS sequencing identified one C. lusitaniae strain, three C. intermedia strains, two C. haemulonii strains and one C. duobushaemulonii strain. Analysis of ITS variability revealed a greater number of haplotypes among C. albicans, C. tropicalis, C. glabrata and C. lusitaniae, which are predominantly related to endogenous sources of acquisition. Bayesian analysis confirmed the major phylogenetic relationships among the isolates and the molecular identification of the different Candida spp. Conclusions Molecular studies based on ITS sequencing are necessary to identify closely related and emerging species. Polymorphism analysis of the ITS rDNA region demonstrated its utility as a genetic marker for species identification and phylogenetic relationships as well as for drawing inferences concerning the natural history of hematogenous infections caused by medically important and emerging Candida species. Electronic supplementary material The online version of this article (doi:10.1186/s12879-015-0793-3) contains supplementary material, which is available to authorized users.

The genus Oxytropis DC. is one of the largest genera in the Fabaceae family. The most plant species belonging to the Oxytropis genus have an important medicinal value. Currently the botanical taxonomy of the genus is complicated due to existence of many subgenera and sections that developed based on morphological traits. Also, in the literature there is luck of knowledge on phylogeny of Oxytropis species from Central Asian region. Therefore, the purpose of the present study was the clarification of taxonomic relationship of two Oxytropis species from SouthEast of Kazakhstan (O. almaatensis Bajt. and O. glabra DC.). The study was based on using phylogenetic analysis and haplotype network assessment based on sequences ITS (internal transcribed spacers), which is DNA marker of nuclear genome. Plant materials of O. almaatensis were collected from 2 populations in two neighboring Gorges in Trans Ili Alatau Mountains, O. glabra plant material was obtained from Herbarium of the Department of Biodiversity and Bioresources, al-Farabi Kazakh National University. Based on DNA sequences of ITS the phylogenetic and network relationships were investigated by using Neighbor Joining and Median Joining methods, respectively. The nucleotide sequences of ITS of O. almaatensis and O. glabra were aligned with sequences of 29 Oxytropis references found in the NCBI database. Out of the 601 aligned positions of ITS 33 (5.6 %) sites were found to be polymorphic nucleotides and used in evaluation of the genetic relationship of species. Constructed MJ haplotype network showed a very high congruence with the NJ phylogenetic tree. MJ network provided valuable additional hints in clarification of the taxonomic relationship among species involved in the analysis. In this study phylogenetic NJ tree and MJ network based on the variation of ITS sequences confirmed the monophyletic origin of the genus. The ITS haplotype network suggested that O. glabra is very diverse species and possibly played important role in the evolutionary processes of the genus in Central Asian region. The study is additional contribution in the molecular taxonomy of complex Oxytropis genus.

Vanda tricolor Lindl. is widespread in cultivation in South East Asia, being relatively easily cultivated in the garden, and is often seen growing floriferously on garden trees, fence posts and verandas. Whilst the species is widespread in cultivation in its native regions of Java and Bali, wild populations are small and highly fragmented. The author’s recent field trip to Java and Bali found few wild examples of the species, and the few plants found were in the most inaccessible trees, due to massive over-collection, often with no other V. tricolor plants to be found in the area. Vanda tricolor Lindl. is widespread in cultivation in South East Asia, being relatively easily cultivated in the garden, and is often seen growing floriferously on garden trees, fence posts and verandas. Whilst the species is widespread in cultivation in its native regions of Java and Bali, wild populations are small and highly fragmented. The author’s recent field trip to Java and Bali found few wild examples of the species, and the few plants found were in the most inaccessible trees, due to massive over-collection, often with no other V. tricolor plants to be found in the area.

Background. In Plasmodium falciparum, resistance to chloroquine (CQ) is conferred by a K to T mutation at amino acid position 76 (K76T) in the P. falciparum CQ transporter (PfCRT). To date, at least 15 pfcrt genotypes, which are represented by combinations of five amino acids at positions 72-76, have been described in field isolates from various endemic regions. To identify novel mutant pfcrt genotypes and to reveal the genetic relatedness of pfcrt genotypes, a large-scale survey over a wide geographic area was performed. Methods. Sequences for exon 2 in pfcrt, including known polymorphic sites at amino acid positions 72, 74, 75 and 76, were obtained from 256 P. falciparum isolates collected from eight endemic countries in Asia (Bangladesh, Cambodia, Lao P.D.R., the Philippines and Thailand), Melanesia (Papua New Guinea and Vanuatu) and Africa (Ghana). A haplotype network was constructed based on six microsatellite markers located -29 kb to 24 kb from pfcrt in order to examine the genetic relatedness among mutant pfcrt genotypes. Results. In addition to wild type (CVMNK at positions 72-76), four mutant pfcrt were identified; CVIET, CVIDT, SVMNT and CVMNT (mutated amino acids underlined). Haplotype network revealed that there were only three mutant pfcrt lineages, originating in Indochina, Philippines and Melanesia. Importantly, the Indochina lineage contained two mutant pfcrt genotypes, CVIET (n = 95) and CVIDT (n = 14), indicating that CVIDT shares a common origin with CVIET. Similarly, one major haplotype in the Melanesian lineage contained two pfcrt genotypes; SVMNT (n = 71) and CVMNT (n = 3). In Africa, all mutant pfcrt genotypes were the CVIET of the Indochina lineage, probably resulting from the intercontinental migration of CQ resistance from Southeast Asia. Conclusions. The number of CQ-mutant lineages observed in this study was identical to that found in previous studies. This supports the hypothesis that the emergence of novel CQ resistance is rare. However, in the mutant pfcrt genotypes, amino acid changes at positions 72, 74 and 75 appear to have recently been generated at least several times, producing distinct pfcrt mutant genotypes. The occurrence of new mutations flanking K76T may yield stronger resistance to CQ and/or a higher fitness than the original pfcrt mutant., Malaria Journal, 11, no92; 2012