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tempo and mode of allopatric divergence in the weakly electric fish Sternopygus dariensis in the Isthmus of Panama
Full text linkSpatial isolation is one of the main drivers of allopatric speciation, but the extent to which spatiallysegregated populations accumulate genetic differences relevant to speciation is not always clear. We used data from ultraconserved elements (UCEs) and whole mitochondrial genomes (i.e., mitogenomes) to explore genetic variation among allopatric populations of the weakly electric fish Sternopygus dariensis across the Isthmus of Panama. We found strong genetic divergence between eastern and western populations of S. dariensis. Over 77% of the UCE loci examined were differentially fixed between populations, and these loci appear to be distributed across the species’ genome. Population divergence occurred within the last 1.1 million years, perhaps due to global glaciation oscillations during the Pleistocene. Our results are consistent with a pattern of genetic differentiation under strict geographic isolation, and suggest the presence of incipient allopatric species within S. dariensis. Genetic divergence in S. dariensis likely occurred in situ, long after the closure of the Isthmus of Panama. Our study highlights the contribution of spatial isolation and vicariance to promoting rapid diversification in Neotropical freshwater fishes. The study of spatially-segregated populations within the Isthmus of Panama could reveal how genetic differences accumulate as allopatric speciation proceeds.Spatial isolation is one of the main drivers of allopatric speciation, but the extent to which spatiallysegregated populations accumulate genetic differences relevant to speciation is not always clear. We used data from ultraconserved elements (UCEs) and whole mitochondrial genomes (i.e., mitogenomes) to explore genetic variation among allopatric populations of the weakly electric fish Sternopygus dariensis across the Isthmus of Panama. We found strong genetic divergence between eastern and western populations of S. dariensis. Over 77% of the UCE loci examined were differentially fixed between populations, and these loci appear to be distributed across the species’ genome. Population divergence occurred within the last 1.1 million years, perhaps due to global glaciation oscillations during the Pleistocene. Our results are consistent with a pattern of genetic differentiation under strict geographic isolation, and suggest the presence of incipient allopatric species within S. dariensis. Genetic divergence in S. dariensis likely occurred in situ, long after the closure of the Isthmus of Panama. Our study highlights the contribution of spatial isolation and vicariance to promoting rapid diversification in Neotropical freshwater fishes. The study of spatially-segregated populations within the Isthmus of Panama could reveal how genetic differences accumulate as allopatric speciation proceeds
Environmental Conditions May Shape the Patterns of Genomic Variations in Leishmania panamensis
Full text linkDue to the absence of transcriptional regulation of gene expression in Leishmania parasites, it is now well accepted that several forms of genomic variations modulate the levels of critical proteins through changes in gene dosage. We previously observed many of these variations in our reference laboratory strain of L. panamensis (PSC-1 strain), including chromosomes with an increased somy and the presence of a putative linear minichromosome derived from chromosome 34. Here, we compared the previously described genomic variations with those occurring after exposure of this strain to increasing concentrations of trivalent antimony (SbIII), as well as those present in two geographically unrelated clinical isolates of L. panamensis. We observed changes in the somy of several chromosomes, amplifications of several chromosomal regions, and copy number variations in gene arrays after exposure to SbIII. Occurrence of amplifications potentially beneficial for the Sb-resistant phenotype appears to be associated with the loss of other forms of amplification, such as the linear minichromosome. In contrast, we found no evidence of changes in somy or amplification of relatively large chromosomal regions in the clinical isolates. In these isolates, the predominant amplifications appear to be those that generate genes arrays; however, in many cases, the amplified arrays have a notably higher number of copies than those from the untreated and Sb-treated laboratory samples.Due to the absence of transcriptional regulation of gene expression in Leishmania parasites, it is now well accepted that several forms of genomic variations modulate the levels of critical proteins through changes in gene dosage. We previously observed many of these variations in our reference laboratory strain of L. panamensis (PSC-1 strain), including chromosomes with an increased somy and the presence of a putative linear minichromosome derived from chromosome 34. Here, we compared the previously described genomic variations with those occurring after exposure of this strain to increasing concentrations of trivalent antimony (SbIII), as well as those present in two geographically unrelated clinical isolates of L. panamensis. We observed changes in the somy of several chromosomes, amplifications of several chromosomal regions, and copy number variations in gene arrays after exposure to SbIII. Occurrence of amplifications potentially beneficial for the Sb-resistant phenotype appears to be associated with the loss of other forms of amplification, such as the linear minichromosome. In contrast, we found no evidence of changes in somy or amplification of relatively large chromosomal regions in the clinical isolates. In these isolates, the predominant amplifications appear to be those that generate genes arrays; however, in many cases, the amplified arrays have a notably higher number of copies than those from the untreated and Sb-treated laboratory samples
Environmental Conditions May Shape the Patterns of Genomic Variations in Leishmania panamensis
Full text linkDue to the absence of transcriptional regulation of gene expression in Leishmania parasites, it is now well accepted that several forms of genomic variations modulate the levels of critical proteins through changes in gene dosage. We previously observed many of these variations in our reference laboratory strain of L. panamensis (PSC-1 strain), including chromosomes with an increased somy and the presence of a putative linear minichromosome derived from chromosome 34. Here, we compared the previously described genomic variations with those occurring after exposure of this strain to increasing concentrations of trivalent antimony (SbIII), as well as those present in two geographically unrelated clinical isolates of L. panamensis. We observed changes in the somy of several chromosomes, amplifications of several chromosomal regions, and copy number variations in gene arrays after exposure to SbIII. Occurrence of amplifications potentially beneficial for the Sb-resistant phenotype appears to be associated with the loss of other forms of amplification, such as the linear minichromosome. In contrast, we found no evidence of changes in somy or amplification of relatively large chromosomal regions in the clinical isolates. In these isolates, the predominant amplifications appear to be those that generate genes arrays; however, in many cases, the amplified arrays have a notably higher number of copies than those from the untreated and Sb-treated laboratory samples.Due to the absence of transcriptional regulation of gene expression in Leishmania parasites, it is now well accepted that several forms of genomic variations modulate the levels of critical proteins through changes in gene dosage. We previously observed many of these variations in our reference laboratory strain of L. panamensis (PSC-1 strain), including chromosomes with an increased somy and the presence of a putative linear minichromosome derived from chromosome 34. Here, we compared the previously described genomic variations with those occurring after exposure of this strain to increasing concentrations of trivalent antimony (SbIII), as well as those present in two geographically unrelated clinical isolates of L. panamensis. We observed changes in the somy of several chromosomes, amplifications of several chromosomal regions, and copy number variations in gene arrays after exposure to SbIII. Occurrence of amplifications potentially beneficial for the Sb-resistant phenotype appears to be associated with the loss of other forms of amplification, such as the linear minichromosome. In contrast, we found no evidence of changes in somy or amplification of relatively large chromosomal regions in the clinical isolates. In these isolates, the predominant amplifications appear to be those that generate genes arrays; however, in many cases, the amplified arrays have a notably higher number of copies than those from the untreated and Sb-treated laboratory samples
Forest disturbance and vector transmitted diseases in thelowland tropical rainforest of central Panama
Full text linkobjective To explore possible changes in the community attributes of haematophagous insects as afunction of forest disturbance. We compare the patterns of diversity and abundance, plus thebehavioural responses of three epidemiologically distinct vector assemblages across sites depictingvarious levels of forest cover.methods Over a 3-year period, we sampled mosquitoes, sandflies and biting-midges in forestedhabitats of central Panama. We placed CDC light traps in the forest canopy and in the understorey togather blood-seeking females.results We collected 168 405 adult haematophagous dipterans in total, including 26 genera and 86species. Pristine forest settings were always more taxonomically diverse than the disturbed forest sites,confirming that disturbance has a negative impact on species richness. Species of Phlebotominae andCulicoides were mainly classified as climax (i.e. forest specialist) or disturbance-generalist, which tendto decrease in abundance along with rising levels of disturbance. In contrast, a significant portion ofmosquito species, including primary and secondary disease vectors, was classified as colonists (i.e.disturbed-areas specialists), which tend to increase in numbers towards more disturbed forest habitats.At pristine forest, the most prevalent species of Phlebotominae and Culicoides partitioned the verticalniche by being active at the forest canopy or in the understorey; yet this pattern was less clear indisturbed habitats. Most mosquito species were not vertically stratified in their habitat preference.conclusion We posit that entomological risk and related pathogen exposure to humans is higher inpristine forest scenarios for Culicoides and Phlebotominae transmitted diseases, whereas forestdisturbance poses a higher entomological risk for mosquito-borne infections. This suggests that theDilution Effect Hypothesis (DEH) does not apply in tropical rainforests where highly abundant, yetunrecognised insect vectors and neglected zoonotic diseases occur. Comprehensive, community levelentomological surveillance is, therefore, the key for predicting potential disease spill over in scenariosof pristine forest intermixed with anthropogenic habitats. We suggest that changes in forest qualityshould also be considered when assessing arthropod-borne disease transmission risk.objective To explore possible changes in the community attributes of haematophagous insects as afunction of forest disturbance. We compare the patterns of diversity and abundance, plus thebehavioural responses of three epidemiologically distinct vector assemblages across sites depictingvarious levels of forest cover.methods Over a 3-year period, we sampled mosquitoes, sandflies and biting-midges in forestedhabitats of central Panama. We placed CDC light traps in the forest canopy and in the understorey togather blood-seeking females.results We collected 168 405 adult haematophagous dipterans in total, including 26 genera and 86species. Pristine forest settings were always more taxonomically diverse than the disturbed forest sites,confirming that disturbance has a negative impact on species richness. Species of Phlebotominae andCulicoides were mainly classified as climax (i.e. forest specialist) or disturbance-generalist, which tendto decrease in abundance along with rising levels of disturbance. In contrast, a significant portion ofmosquito species, including primary and secondary disease vectors, was classified as colonists (i.e.disturbed-areas specialists), which tend to increase in numbers towards more disturbed forest habitats.At pristine forest, the most prevalent species of Phlebotominae and Culicoides partitioned the verticalniche by being active at the forest canopy or in the understorey; yet this pattern was less clear indisturbed habitats. Most mosquito species were not vertically stratified in their habitat preference.conclusion We posit that entomological risk and related pathogen exposure to humans is higher inpristine forest scenarios for Culicoides and Phlebotominae transmitted diseases, whereas forestdisturbance poses a higher entomological risk for mosquito-borne infections. This suggests that theDilution Effect Hypothesis (DEH) does not apply in tropical rainforests where highly abundant, yetunrecognised insect vectors and neglected zoonotic diseases occur. Comprehensive, community levelentomological surveillance is, therefore, the key for predicting potential disease spill over in scenariosof pristine forest intermixed with anthropogenic habitats. We suggest that changes in forest qualityshould also be considered when assessing arthropod-borne disease transmission risk
Diverse novel phleboviruses in sandflies from the Panama Canal area, Central Panama
Full text linkThe genus Phlebovirus (order Bunyavirales, family Phenuiviridae) comprises 57 viruses that are grouped into nine speciescomplexes. Sandfly-transmitted phleboviruses are found in Europe, Africa and the Americas and are responsible for febrile illness and infections of the nervous system in humans. The aim of this study was to assess the genetic diversity of sandflytransmitted phleboviruses in connected and isolated forest habitats throughout the Panama Canal area in Central Panama. In total, we collected 13 807 sandflies comprising eight phlebotomine species. We detected several strains pertaining to five previously unknown viruses showing maximum pairwise identities of 45–78 % to the RNA-dependent RNA polymerase genes of phleboviruses. Entire coding regions were directly sequenced from infected sandflies as virus isolation in cell culture was not successful. The viruses were tentatively named La Gloria virus (LAGV), Mona Grita virus (MOGV), Peña Blanca virus (PEBV), Tico virus (TICV) and Tres Almendras virus (TRAV). Inferred phylogenies and p-distance-based analyses revealed that PEBV groups with the Bujaru phlebovirus species-complex, TRAV with the Candiru phlebovirus speciescomplex and MOGV belongs to the proposed Icoarci phlebovirus species-complex, whereas LAGV and TICV seem to be distant members of the Bujaru phlebovirus species-complex. No specific vector or habitat association was found for any of the five viruses. Relative abundance of sandflies was similar over habitat types. Our study shows that blood-feeding insects originating from remote and biodiverse habitats harbour multiple previously unknown phleboviruses. These viruses should be included in future surveillance studies to assess their geographic distribution and to elucidate if these viruses cause symptoms of disease in animals or humans.The genus Phlebovirus (order Bunyavirales, family Phenuiviridae) comprises 57 viruses that are grouped into nine speciescomplexes. Sandfly-transmitted phleboviruses are found in Europe, Africa and the Americas and are responsible for febrile illness and infections of the nervous system in humans. The aim of this study was to assess the genetic diversity of sandflytransmitted phleboviruses in connected and isolated forest habitats throughout the Panama Canal area in Central Panama. In total, we collected 13 807 sandflies comprising eight phlebotomine species. We detected several strains pertaining to five previously unknown viruses showing maximum pairwise identities of 45–78 % to the RNA-dependent RNA polymerase genes of phleboviruses. Entire coding regions were directly sequenced from infected sandflies as virus isolation in cell culture was not successful. The viruses were tentatively named La Gloria virus (LAGV), Mona Grita virus (MOGV), Peña Blanca virus (PEBV), Tico virus (TICV) and Tres Almendras virus (TRAV). Inferred phylogenies and p-distance-based analyses revealed that PEBV groups with the Bujaru phlebovirus species-complex, TRAV with the Candiru phlebovirus speciescomplex and MOGV belongs to the proposed Icoarci phlebovirus species-complex, whereas LAGV and TICV seem to be distant members of the Bujaru phlebovirus species-complex. No specific vector or habitat association was found for any of the five viruses. Relative abundance of sandflies was similar over habitat types. Our study shows that blood-feeding insects originating from remote and biodiverse habitats harbour multiple previously unknown phleboviruses. These viruses should be included in future surveillance studies to assess their geographic distribution and to elucidate if these viruses cause symptoms of disease in animals or humans
α-Glucosidase inhibitors from a mangrove associated fungus, Zasmidium sp. strain EM5-10
Full text linkMangroves plants and their endophytes represent a natural source of novel and bioactive compounds. In our ongoing research on mangrove endophytes from the Panamanian Pacifc Coast, we have identifed several bio‑ active endophytic fungi. From these organisms, an isolate belonging to the genus Zasmidium (Mycosphaerellaceae) showed 91.3% of inhibition against α-glucosidase enzyme in vitroMangroves plants and their endophytes represent a natural source of novel and bioactive compounds. In our ongoing research on mangrove endophytes from the Panamanian Pacifc Coast, we have identifed several bio‑ active endophytic fungi. From these organisms, an isolate belonging to the genus Zasmidium (Mycosphaerellaceae) showed 91.3% of inhibition against α-glucosidase enzyme in vitr
Development of Monoclonal Antibodies Against Cry1Ac/Ab Protein for Designing of Sandwich ELISA to Detect BT Toxin from Cotton Seeds and Leaves
Full text linkoai:repositorio-indicasat.org.pa:123456789/77The design of the study is to develop monoclonal antibodies against Cry1Ac/Ab protein for designing os sandwich ELISA(hybridoma technology). Hybridoma technology was invented by Cesar Milstein and Georges J.F Kohler in the year 1975 and is an unique method used to produce identical antibodies in maximum quantities. Monoclonal antibodies were developed by immunization of Balb/C mice with Cry1Ac/Ab Protein. Titer values of mice tail bleeds were checked and the best mice with higher titer value was used for fusion. Immunized mice spleen cells were fused with Myeloma cells (SP2-O), using polyethylene glycol (PEG) and the fused cells were incubated with HAT medium for 12 days and initially 400 positive hybridoma clones were obtained, of which 13 potential clones were selected using indirect ELISA against Cry1Ac/Ab recombinant antigen. Cross reactivity was ruled out using indriet ELSA against cry proteins such as Cry2A, Cry1F and CP4EPSPS using. Cloning was carried out twice for all 13 clones by limiting dilution factor and pure single clones were selected. The class IgG/IgM/IgA and sub classes IgG1, IgG2, IgG3 antibodies are determined by isotyping. Determination of class and subclass of an antibody is very important for selecting proper purification methods. Commercially available rapid isotyping kits were used for isotyping which provides the information of 1) IgG, IgM, IgA, IgG2a, IgG2b or IgG3 2) Light chain identification as either kappa or lambda. All pure clones were preserved in Liquid Nitrogen for future use to develop immunological kits for detection of Cry1Ac/Ab present in the plant tissue.The design of the study is to develop monoclonal antibodies against Cry1Ac/Ab protein for designing os sandwich ELISA(hybridoma technology). Hybridoma technology was invented by Cesar Milstein and Georges J.F Kohler in the year 1975 and is an unique method used to produce identical antibodies in maximum quantities. Monoclonal antibodies were developed by immunization of Balb/C mice with Cry1Ac/Ab Protein. Titer values of mice tail bleeds were checked and the best mice with higher titer value was used for fusion. Immunized mice spleen cells were fused with Myeloma cells (SP2-O), using polyethylene glycol (PEG) and the fused cells were incubated with HAT medium for 12 days and initially 400 positive hybridoma clones were obtained, of which 13 potential clones were selected using indirect ELISA against Cry1Ac/Ab recombinant antigen. Cross reactivity was ruled out using indriet ELSA against cry proteins such as Cry2A, Cry1F and CP4EPSPS using. Cloning was carried out twice for all 13 clones by limiting dilution factor and pure single clones were selected. The class IgG/IgM/IgA and sub classes IgG1, IgG2, IgG3 antibodies are determined by isotyping. Determination of class and subclass of an antibody is very important for selecting proper purification methods. Commercially available rapid isotyping kits were used for isotyping which provides the information of 1) IgG, IgM, IgA, IgG2a, IgG2b or IgG3 2) Light chain identification as either kappa or lambda. All pure clones were preserved in Liquid Nitrogen for future use to develop immunological kits for detection of Cry1Ac/Ab present in the plant tissue
Molecular validation of anthropophilic Phlebotominae sandflies (Diptera: Psychodidae) in Central Panama
Full text linkSix Phlebotominae sand fly species are incriminated as biological vectors of human pathogens in Panama, but molecular corroboration is still needed. We aim at confirming the identity of Phlebotominae species documented as anthropophilic in Panama. Adult sandflies were collected from August 2010 to February 2012 in Central Panama using CDC light traps. Species confirmation was accomplished through molecular barcodes and allied sequences from GenBank. A total of 53,366 sand fly specimens representing 18 species were collected. Five species were validated molecularly as single phylogenetic clusters, but Psychodopygus thula depicted two genetically divergent lineages, which may be indicative of cryptic speciation.Six Phlebotominae sand fly species are incriminated as biological vectors of human pathogens in Panama, but molecular corroboration is still needed. We aim at confirming the identity of Phlebotominae species documented as anthropophilic in Panama. Adult sandflies were collected from August 2010 to February 2012 in Central Panama using CDC light traps. Species confirmation was accomplished through molecular barcodes and allied sequences from GenBank. A total of 53,366 sand fly specimens representing 18 species were collected. Five species were validated molecularly as single phylogenetic clusters, but Psychodopygus thula depicted two genetically divergent lineages, which may be indicative of cryptic speciation
Host affinity of endophytic fungi and the potential for reciprocal interactions involving host secondary chemistry
Full text linkPREMISE: Interactions between fungal endophytes and their host plants present useful systems for identifying important factors affecting assembly of host-associated microbiomes. Here we investigated the role of secondary chemistry in mediating host affinity of asymptomatic foliar endophytic fungi using Psychotria spp. and Theobroma cacao (cacao) as hosts. METHODS: First, we surveyed endophytic communities in Psychotria species in a natural common garden using culture-based methods. Then we compared differences in endophytic community composition with differences in foliar secondary chemistry in the same host species, determined by liquid chromatography–tandem mass spectrometry. Finally, we tested how inoculation with live and heat-killed endophytes affected the cacao chemical profile. RESULTS: Despite sharing a common environment and source pool for endophyte spores, different Psychotria host species harbored strikingly different endophytic communities that reflected intrinsic differences in their leaf chemical profiles. In T. cacao, inoculation with live and heat-killed endophytes produced distinct cacao chemical profiles not found in uninoculated plants or pure fungal cultures, suggesting that endophytes, like pathogens, induce changes in secondary chemical profiles of their host plant. CONCLUSIONS: Collectively our results suggest at least two potential processes: (1) Plant secondary chemistry influences assembly and composition of fungal endophytic communities, and (2) host colonization by endophytes subsequently induces changes in the host chemical landscape. We propose a series of testable predictions based on the possibility that reciprocal chemical interactions are a general property of plant–endophyte interactionsPREMISE: Interactions between fungal endophytes and their host plants present useful systems for identifying important factors affecting assembly of host-associated microbiomes. Here we investigated the role of secondary chemistry in mediating host affinity of asymptomatic foliar endophytic fungi using Psychotria spp. and Theobroma cacao (cacao) as hosts. METHODS: First, we surveyed endophytic communities in Psychotria species in a natural common garden using culture-based methods. Then we compared differences in endophytic community composition with differences in foliar secondary chemistry in the same host species, determined by liquid chromatography–tandem mass spectrometry. Finally, we tested how inoculation with live and heat-killed endophytes affected the cacao chemical profile. RESULTS: Despite sharing a common environment and source pool for endophyte spores, different Psychotria host species harbored strikingly different endophytic communities that reflected intrinsic differences in their leaf chemical profiles. In T. cacao, inoculation with live and heat-killed endophytes produced distinct cacao chemical profiles not found in uninoculated plants or pure fungal cultures, suggesting that endophytes, like pathogens, induce changes in secondary chemical profiles of their host plant. CONCLUSIONS: Collectively our results suggest at least two potential processes: (1) Plant secondary chemistry influences assembly and composition of fungal endophytic communities, and (2) host colonization by endophytes subsequently induces changes in the host chemical landscape. We propose a series of testable predictions based on the possibility that reciprocal chemical interactions are a general property of plant–endophyte interaction
Thermo-Energetic Study in Blood Infected with Plasmodium falciparum radiated at 2.45GHz
No full textMalaria is a disease that affects the world, caused by the parasite Plasmodium which resistance is emerging to medications that usually control it. Therefore, it is essential to seek alternative treatments like this one, our multidisciplinary work pursues, a variable radiofrequency energy at 2.45GHz to decrease the growth of parasitemia in blood. Experiments are carried on in vitro samples of blood infected with Plasmodium falciparum radiated in a microwave cavity controlling the rise of the temperature. The thermal properties of samples infected at different stages of the parasite are analyzed separately without using microwave radiation for thermal characterization. The energy absorbed by the infected sample is estimated using the specific heat of blood with Plasmodium falciparum. For thermo energetic control, radiation is applied in different duty cycles for a determined amount of energy observing an alteration which it is accentuated in energy radiation with more duty cycles and less power in watts. Our preliminary results show that thermo-energetic control system produce alterations in the growth of parasitemia levels. Furthermore, we confirm that using microwave radiation has a negative effect in the growth of Plasmodium falciparum. There is also, an increment of the specific heat as the parasitemia percentage increase in late stage parasites because in schizonts the Plasmodium falciparum changes the biophysical properties of the red blood cell. This thermo-energetic characterization could be transferred to in vivo samples, with the necessary adjustments for future applications.Malaria is a disease that affects the world, caused by the parasite Plasmodium which resistance is emerging to medications that usually control it. Therefore, it is essential to seek alternative treatments like this one, our multidisciplinary work pursues, a variable radiofrequency energy at 2.45GHz to decrease the growth of parasitemia in blood. Experiments are carried on in vitro samples of blood infected with Plasmodium falciparum radiated in a microwave cavity controlling the rise of the temperature. The thermal properties of samples infected at different stages of the parasite are analyzed separately without using microwave radiation for thermal characterization. The energy absorbed by the infected sample is estimated using the specific heat of blood with Plasmodium falciparum. For thermo energetic control, radiation is applied in different duty cycles for a determined amount of energy observing an alteration which it is accentuated in energy radiation with more duty cycles and less power in watts. Our preliminary results show that thermo-energetic control system produce alterations in the growth of parasitemia levels. Furthermore, we confirm that using microwave radiation has a negative effect in the growth of Plasmodium falciparum. There is also, an increment of the specific heat as the parasitemia percentage increase in late stage parasites because in schizonts the Plasmodium falciparum changes the biophysical properties of the red blood cell. This thermo-energetic characterization could be transferred to in vivo samples, with the necessary adjustments for future applications