Mitogenomic Characterization of Cameroonian Endemic Coptodon camerunensis (Cichliformes: Cichlidae) and Matrilineal Phylogeny of Old-World Cichlids.

The mitogenomic evolution of old-world cichlids is still largely incomplete in Western Africa. In this present study, the complete mitogenome of the Cameroon endemic cichlid, Coptodon camerunensis, was determined by next-generation sequencing. The mitogenome was 16,557 bp long and encoded with 37 genes (13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNA genes, and a control region). The C. camerunensis mitogenome is AT-biased (52.63%), as exhibited in its congener, Coptodon zillii (52.76% and 53.04%). The majority of PCGs start with an ATG initiation codon, except COI, which starts with a GTG codon and five PCGs and ends with the TAA termination codon and except seven PCGs with an incomplete termination codon. In C. camerunensis mitogenome, most tRNAs showed classical cloverleaf secondary structures, except tRNA-serine with a lack of DHU stem. Comparative analyses of the conserved blocks of two Coptodonini species control regions revealed that the CSB-II block was longer than other blocks and contained highly variable sites. Using 13 concatenated PCGs, the mitogenome-based Bayesian phylogeny easily distinguished all the examined old-world cichlids. Except for Oreochromini and Coptodinini tribe members, the majority of the taxa exhibited monophyletic clustering within their respective lineages. C. camerunensis clustered closely with Heterotilapia buttikoferi (tribe Heterotilapiini) and had paraphyletic clustering with its congener, C. zillii. The Oreochromini species also displayed paraphyletic grouping, and the genus Oreochromis showed a close relationship with Coptodinini and Heterotilapiini species. In addition, illustrating the known distribution patterns of old-world cichlids, the present study is congruent with the previous hypothesis and proclaims that prehistoric geological evolution plays a key role in the hydroclimate of the African continent during Mesozoic, which simultaneously disperses and/or colonizes cichlids in different ichthyological provinces and Rift Lake systems in Africa. The present study suggests that further mitogenomes of cichlid species are required, especially from western Africa, to understand their unique evolution and adaptation.

Development of PCR-RFLP Technique for Identify Several Members of Fusarium incarnatum-equiseti Species Complex and Fusarium fujikuroi Species Complex.

Fusarium incarnatum-equiseti species complex (FIESC) contain over 40 members. The primer pair Smibo1FM/Semi1RM on the RPB2 partial gene has been reported to be able to identify Fusarium semitectum. The F. fujikuroi species complex (FFSC) contains more than 50 members. The F. verticillioides as a member of this complex can be identified by using VER1/VER2 primer pair on the CaM partial gene. In this research, the Smibo1FM/Semi1RM can amplify F. sulawesiense, F. hainanense, F. bubalinum, and F. tanahbumbuense, members of FIESC at 424 bp. The VER1/VER2 can amplify F. verticillioides, F. andiyazi, and F. pseudocircinatum, members of FFSC at 578 bp. Polymerase chain reaction-restriction fragment length polymorphism by using the combination of three restriction enzymes EcoRV, MspI, and HpyAV can differentiate each species of FIESC used. The two restriction enzymes HpaII and NspI can distinguish each species of FFSC used. The proper identification process is required for pathogen control in the field in order to reduce crop yield loss.

Phytophthora palmivora from Sulawesi and Java Islands, Indonesia, reveals high genotypic diversity and lack of population structure.

Phytophthora palmivora is the causal agent of cocoa black pod disease, one of the primary diseases of cocoa in Indonesia. A better understanding of P. palmivora population genetics is needed to aid the development of relevant disease management strategies. This study is the first population genetic study of P. palmivora in Indonesia using microsatellite markers based on the alleles genotyping method. The microsatellite markers were used to determine the genotype of 44 P. palmivora isolates from Sulawesi (24) and Java (20) islands. The total number of observed multilocus genotypes (MLG) from both populations was 34. The genotypic diversity of P. palmivora from Sulawesi (2.90; 16.0; 0.938) and Java (2.76; 14.3; 0.930) islands was high as seen from Shannon's diversity index (H), Stoddart and Taylor's Index (G), and Simpson's Index (λ) respectively. Evenness and Nei's unbiased gene diversity exhibited similarly high levels from both populations. The linkage disequilibrium test indicated that sexual recombination occurred in the Java population (P = 0.312). Analysis of molecular variance (AMOVA) and Bayesian clustering revealed five genetic clusters, and isolates from both islands were evenly distributed across the five gene clusters. All genetic diversity was from within individuals. P. palmivora from Sulawesi and Java showed a high genotypic diversity but a lack of genetic differentiation among populations (Fst = 0.006). Both populations formed one highly diverse group. Minimum spanning network analysis showed no particular grouping of MLGs, and shared MLGs from both populations indicated long-distance migration of P. palmivora facilitated by human activities.

Simultaneous purification of fucoxanthin isomers from brown seaweeds by open-column and high-performance liquid chromatography.

Simultaneous purification of fucoxanthin isomers from brown seaweeds by two steps of open-column chromatography (OCC) and reversed-phase (RP)-high-performance liquid chromatography (HPLC) is described. Analysis and identification of fucoxanthin isomers were performed by chromatographic and spectrophotometric properties such as retention time (tR), spectral shape, maximal absorption wavelength (λmax), Q-ratio, and mass spectrometry (MS) data including the ratio of fragment ions. The optimal conditions for a simultaneous separation and purification were examined by changing several parameters of HPLC, i.e., mobile phase composition, equilibration time, and column oven temperature. The purification procedure consisted of the following two steps: first, highly purified fucoxanthin fraction was obtained by a silica-gel OCC. Then, four major fucoxanthin isomers, all-trans, 13'-cis, 13-cis, and 9'-cis, were simultaneously separated and purified by RP-HPLC with an analytical C30 column and gradient elution in a mixture of water, methanol, and methyl tert-butyl ether. The purity of fucoxanthin isomers purified was >95% for all-trans and 9'-cis, 85% for 13'-cis, and >80% for 13-cis. A large-scale purification by RP-HPLC using a preparative C18 column was effective for the purification of all-trans and 9'-cis with a yield of 95%. This developed technique was fully applicable to analyze the enhanced production of fucoxanthin isomers by iodine-catalyzed stereomutation which composed of 9 isomer species including 9-cis.

Variability of Lecanicillium spp. Mycoparasite of Coffee Leaf Rust Pathogen (Hemileia vastatrix) in Indonesia.

<b>Background and Objective:</b> Coffee leaf rust disease caused by <i>Hemileia vastatrix</i> resulted in high yield loss and difficult to control. Several chemical fungicides have been used to control this disease. However, the effectiveness of chemical control is low, so it is necessary to find other methods such as biological control. <i>Lecanicillium</i> spp. is well-known as mycoparasite on <i>H. vastatrix</i> uredospores but the study in Indonesia is still limited. This study aimed to collect and investigated the genetic variability of <i>Lecanicillium</i> spp. at various coffee plantations in Indonesia. <b>Materials and Methods:</b> Samples of <i>Lecanicillium </i>spp. were collected from 20 districts in 7 provinces throughout Indonesia. Morphology of colony and conidia were identified by visual examination and by viewed under the light microscope. Genetic variability was conducted using Rep-PCR and clustered with UPGMA. <b>Results:</b> Morphological observation in this study revealed all isolates collected from uredospores of <i>H. vastatrix</i> were similar with <i>Lecanicillium </i>spp. Genetic variability analysis clustered the 80 isolates into eight clusters with their specific characters. <b>Conclusion:</b> Morphological identification in this study showed that 80 isolates of mycoparasite on <i>H. vastatrix</i> belong to <i>Lecanicillium</i> spp. Further study using the molecular technique is needed to identity the species of <i>Lecanicillium</i>.

Multi-genetic Analysis of Colletotrichum spp. Associated with Postharvest Disease of Fruits Anthracnose in Special Region of Yogyakarta, Indonesia.

BACKGROUND AND OBJECTIVE: Postharvest disease caused by Colletotrichum spp. caused major losses. The species of Colletotrichum are difficult to distinguish if only seen from their morphology. This study investigated Colletotrichum isolates associated with tropical fruits anthracnose using multi-genetic analysis and the cross-infection potency of each isolate among tropical fruits. MATERIALS AND METHODS: The fruit samples were collected from markets in the Special Region of Yogyakarta, Indonesia and its surrounding area. The fruits affected by anthracnose subjected to isolation, resulting in 15 isolates. Morphology of colony and conidia then characterized and clustered with UPGMA. The seven representative isolates were selected for molecular identification. The multi-genetic analysis was used by combining ITS, Glyceraldehyde 3-phosphate dehydrogenase (gapdh) and tub2 sequence genes. A cross-infection test was conducted by using selected species from the multi-genetic analysis. RESULTS: Multi-genetic analysis clustered the selected isolates into four species. Isolates from banana, avocado, papaya and citrus belonged to gloeosporioides species complex, including C. siamense, C. asianum and C. gloeosporioides. Isolates from apple, guava, mango and citrus belonged to acutatum species complex, including C. sloanei. The cross-infection test in this study showed that C. siamense could cause anthracnose on banana, apple, citrus and avocado, C. asianum on avocado, papaya, apple and citrus, C. gloeosporioides on citrus and apple, C. sloanei on apple, guava, citrus and papaya. CONCLUSION: The C. siamense, C. asianum, C. gloeosporioides and C. sloanei found associated with tropical fruits anthracnose. The potency of the cross-infection test revealed the board range in the pathogenicity of the Colletotrichum isolates.

Disentangling the taxonomy of the subfamily Rasborinae (Cypriniformes, Danionidae) in Sundaland using DNA barcodes.

Sundaland constitutes one of the largest and most threatened biodiversity hotspots; however, our understanding of its biodiversity is afflicted by knowledge gaps in taxonomy and distribution patterns. The subfamily Rasborinae is the most diversified group of freshwater fishes in Sundaland. Uncertainties in their taxonomy and systematics have constrained its use as a model in evolutionary studies. Here, we established a DNA barcode reference library of the Rasborinae in Sundaland to examine species boundaries and range distributions through DNA-based species delimitation methods. A checklist of the Rasborinae of Sundaland was compiled based on online catalogs and used to estimate the taxonomic coverage of the present study. We generated a total of 991 DNA barcodes from 189 sampling sites in Sundaland. Together with 106 previously published sequences, we subsequently assembled a reference library of 1097 sequences that covers 65 taxa, including 61 of the 79 known Rasborinae species of Sundaland. Our library indicates that Rasborinae species are defined by distinct molecular lineages that are captured by species delimitation methods. A large overlap between intraspecific and interspecific genetic distance is observed that can be explained by the large amounts of cryptic diversity as evidenced by the 166 Operational Taxonomic Units detected. Implications for the evolutionary dynamics of species diversification are discussed.

The complete mitochondrial DNA sequence of Pectenocypris sp. (Actinopterygii: Cyprinidae) from Serkap River, Sumatra, Indonesia.

The whole mitochondrial genome of a small cyprinid freshwater fish Pectenocypris sp. collected from Serkap River, Central Sumatra, Indonesia was sequenced. This mitochondrial genome consisted of 16,589 bp and included 37 genes in the same order as in many other vertebrates including the human. Phylogenetic analysis suggested that this taxon clusters with Boraras maculatus among several Rasbora species.

Real-Time in Vivo Detection of H2O2 Using Hyperpolarized 13C-Thiourea.

Reactive oxygen species (ROS) are essential cellular metabolites widely implicated in many diseases including cancer, inflammation, and cardiovascular and neurodegenerative disorders. Yet, ROS signaling remains poorly understood, and their measurements are a challenge due to high reactivity and instability. Here, we report the development of 13C-thiourea as a probe to detect and measure H2O2 dynamics with high sensitivity and spatiotemporal resolution using hyperpolarized 13C magnetic resonance spectroscopic imaging. In particular, we show 13C-thiourea to be highly polarizable and to possess a long spin-lattice relaxation time (T1), which enables real-time monitoring of ROS-mediated transformation. We also demonstrate that 13C-thiourea reacts readily with H2O2 to give chemically distinguishable products in vitro and validate their detection in vivo in a mouse liver. This study suggests that 13C-thiourea is a promising agent for noninvasive detection of H2O2 in vivo. More broadly, our findings outline a viable clinical application for H2O2 detection in patients with a range of diseases.

Photoactivatable glycopolymers for the proteome-wide identification of fucose-α(1-2)-galactose binding proteins.

Although fucose-α(1-2)-galactose (Fucα(1-2)Gal)-containing glycans have been implicated in cognitive processes such as learning and memory, their precise molecular mechanisms are poorly understood. Here we employed the use of multivalent glycopolymers to enable the first proteome-wide identification of weak affinity, low abundance Fucα(1-2)Gal glycan-binding proteins (GBPs). Biotin-terminated glycopolymers containing photoactivatable cross-linking groups were designed to capture and enrich GBPs from rat brain lysates. Candidate proteins were tested for their ability to bind Fucα(1-2)Gal, and the functional significance of the interaction was investigated for the synaptic vesicle protein SV2a using a knockout mouse system. The results suggest a role for SV2a-Fucα(1-2)Gal interactions in SV2a trafficking and synaptic vesicle recycling. More broadly, our studies outline a general chemical approach for the systems-level discovery of novel GBPs.