Introduction. El Alférez, a village in Los Montes de María (Bolívar, Colombia) and a macro-focus of leishmaniasis, recorded its first case in 2018, evidencing changes in the distribution and eco-epidemiology of the disease, although interactions between vectors and local fauna remain unknown. Objective. To evaluate the diversity of sandflies and their blood meal sources in the community of El Alférez in the municipality of El Carmen de Bolívar (Bolívar, Colombia). Materials and methods. In 2018, sandflies were collected using LED-based light traps in domestic, peridomestic, and sylvatic ecotopes and identified at the species level. Multiplex polymerase chain reaction targeting the mitochondrial cytochrome B gene was used to analyze blood from the digestive tract. Results. Lutzomyia evansi was the most abundant species (71.85%; n = 485/675), followed by Lu. panamensis, Lu. gomezi, Lu. trinidadensis, Lu. dubitans, Lu. abonnenci, and Lu.aclydifera. Twenty-five percent of the species had blood meals from Canis familiaris (36.00%; n = 9/25), Ovis aries (36.00%; n=9:/25), Bos taurus (24.00%; n = 6/25), Sus scrofa (20.00%; n = 5/25), and Homo sapiens (8.00%; n = 2/25). Lutzomyia evansi registered the highest feeding frequency (68.00%; n = 17/25), predominantly on a single (44.00%; n = 11/25) or multiple species (24.00%; n = 6/25). Conclusion. Results indicate a eclectic feeding behavior in Lu. evansi, implying potential reservoir hosts for Leishmania spp. and increasing transmission risk. This study is a first step towards understanding the diversity of mammalian blood sources used by sandflies, that may be crucial for vector identification and formulation of effective control measures.
Introducción. En 2018, en la vereda El Alférez de Los Montes de María (Bolívar, Colombia), un macrofoco de leishmaniasis, se reportó el primer caso y se evidenciaron cambios en la distribución y ecoepidemiología de la enfermedad. No obstante, las interacciones entre vectores y fauna local aún son desconocidas. Objetivo. Evaluar la diversidad de flebotomíneos y sus fuentes de alimentación sanguínea en la comunidad de El Alférez del municipio de El Carmen de Bolívar (Bolívar, Colombia). Materiales y métodos. En el 2018, se recolectaron flebotomíneos mediante trampas de luz led ubicadas en el domicilio, el peridomicilio y en el área silvestre, y se identificaron a nivel de especie. Se utilizó la reacción en cadena de la polimerasa múltiple dirigida al gen mitocondrial citocromo B para analizar la sangre del aparato digestivo. Resultados. Lutzomyia evansi fue la especie más abundante (71,85 %; n = 485/675), seguida por Lu. panamensis, Lu. gomezi, Lu. trinidadensis, Lu. dubitans, Lu. abonnenci y Lu. aclydifera. El 25 % (n = 25/100) de las especies analizadas tuvieron como fuentes de ingesta sanguínea a Canis familiaris (36 %; n = 9/25), Ovis aries (36 %; n = 9/25), Bos taurus (24 %; n = 6/25), Sus scrofa (20 %; n = 5/25) y Homo sapiens (8 %; n = 2/25). Lutzomyia evansi fue la especie con la mayor frecuencia de alimentación (68 %; n = 17/25), predominantemente de una sola especie (44 %; n = 11/25) o de varias (24 %; n = 6/25).

Using integrative tools can be effective for species identification, especially in complex groups like Astyanax. Astyanax bimaculatus group is composed of six valid species, including A. lacustris. \"A. altiparanae\", \"A. asuncionensis\", and \"A. jacuhiensis\" are considered as junior synonyms of A. lacustris. Seeking to test the operational taxonomic unit (OTU) status of the junior synonyms of A. lacustris (\"A. altiparanae\", \"A. asuncionensis\", and \"A. jacuhiensis\"), we used analyses through mitochondrial DNA (COI and Cytb), cytogenetic markers (classical and molecular), and morphometry (\"truss network\"). Analysis of mitochondrial DNA sequences separated A. lacustris from the other synonymized species. The cytogenetic and morphometric analyses did not corroborate the synonymization and suggest that besides A. lacustris, the OTUs A. altiparanae, A. asuncionensis, and A. jacuhiensis are valid species. The analysis of different characters proposed by the integrative taxonomy used on the same individuals could provide greater reliability and minimize the underestimation of biodiversity.

Quinone outside inhibitor (QoI) has been applied to manage taro leaf blight caused by Phytophthora colocasiae in southeastern of China for many years. The risk of P. colocasiae to QoI and the potential resistant mechanism remain unknown. In this study, the 74 P. colocasiae strains were sampled from southeastern of China. Sequence analysis of the QoI target Cytb showed one nucleotide variant in the fragment of this gene in this population, producing two haplotypes. The nucleotide variant leads to codon change at 142 (GGT to GCT) producing A142 (alanine) and G142 (glycine) in Hap_1 and Hap_2 strains, respectively. The sensitivity differentiation to azoxystrobin of two haplotypes were observed in vitro. The Hap_1 and Hap_2 strains were confirmed resistant and sensitive by control efficacy of label rate fungicide application, which was 3.0% and 88.8% treated with 500 μg/mL azoxystrobin, respectively. In addition, 10.0 μg/mL azoxystrobin plus 50 μg/mL salicylhydroxamic acid (SHAM) supplemented in PDA medium was identified as a discriminatory dose for differentiation of these two phenotype strains. The azoxystrobin resistant frequency reached 86.5%, indicating prevalence of QoI resistance in the field. Further fitness related features showed that no significant difference in temperature sensitivity, mycelial growth rate, sporangia production, zoospore release and aggressiveness between azoxystrobin-resistant and sensitive strains indicating no potential fitness cost for azoxystrobin resistance. Taken together, azoxystrobin resistance need to be taken into consideration to manage taro leaf blight in southeastern of China.

Avian haemosporidian parasites are spread worldwide and pose a threat to their hosts occasionally. A complete life cycle of these parasites requires two hosts: vertebrate and invertebrate (a blood-sucking insect that acts as a vector). In this study, we tested wild-caught mosquitoes for haemosporidian infections. Mosquitoes were collected (2021-2023) in several localities in Lithuania using a sweeping net and a CDC trap baited with CO2, morphologically identified, and preparations of salivary glands were prepared (from females collected in 2022-2023). 2093 DNA samples from either individual after dissection (1675) or pools (418 pools/1145 individuals) of female mosquito\'s abdomens were screened using PCR for the detection of haemosporidian parasite DNA. Salivary gland preparations were analyzed microscopically from each PCR-positive mosquito caught in 2022 and 2023. The average prevalence of haemosporidian parasites for all analyzed samples was 2.0 % and varied between 0.6 % (2021) and 3.5 % (2022). DNA of Plasmodium ashfordi (cytochrome b genetic lineage pGRW02), P. circumflexum (pTURDUS1), P. homonucleophilum (pSW2), P. matutinum (pLINN1), P. vaughani (pSYAT05), Haemoproteus brachiatus (hLK03), H. majoris (hWW2), and H. minutus (hTUPHI01) were detected in mosquitoes. Coquilletidia richiardii (3.5 %) and Culex pipiens (2.9 %) were mosquito species with the highest prevalence of haemosporidian parasite DNA detected. Mixed infections were detected in 16 mosquitoes. In one of the samples, sporozoites of P. matutinum (pLINN1) were found in the salivary gland preparation of Culex pipiens, confirming this mosquito species as a competent vector of Plasmodium matutinum and adding it to the list of the natural vectors of this avian parasite.

Wild birds are common hosts to numerous intracellular parasites such as single-celled eukaryotes of the family Eimeriidae (order Eucoccidiorida, phylum Apicomplexa). We investigated the infection rates, phylogeny, and pathogenicity of Isospora and Lankesterella parasites in wild and captive passerine birds. Blood and tissue samples of 815 wild and 15 deceased captive birds from Europe were tested using polymerase chain reaction and partial sequencing of the mitochondrial cytochrome b and cytochrome c oxidase I and the nuclear 18S rRNA gene. The infection rate for Lankesterella in wild birds was 10.7% compared to 5.8% for Isospora. Chromogenic in situ hybridization with probes targeting the parasites\' 18S rRNA was employed to identify the parasites\' presence in multiple organs, and hematoxylin-eosin staining was performed to visualize the parasite stages and assess associated lesions. Isospora parasites were mainly identified in the intestine, spleen, and liver. Extraintestinal tissue stages of Isospora were accompanied by predominantly lymphohistiocytic inflammation of varying severity. Lankesterella was most frequently detected in the spleen, lung, and brain; however, infected birds presented only a low parasite burden without associated pathological changes. These findings contribute to our understanding of Isospora and Lankesterella parasites in wild birds.

Membrane proteins carrying redox cofactors are key subunits of respiratory chain complexes, yet the exact path of their folding and maturation remains poorly understood. Here, using cryo-EM and structure prediction via Alphafold2, we generated models of early assembly intermediates of cytochrome b (Cytb), a central subunit of complex III. The predicted structure of the first assembly intermediate suggests how the binding of Cytb to the assembly factor Cbp3-Cbp6 imposes an open configuration to facilitate the acquisition of its heme cofactors. Moreover, structure predictions of the second intermediate indicate how hemes get stabilized by binding of the assembly factor Cbp4, with a concomitant weakening of the contact between Cbp3-Cbp6 and Cytb, preparing for the release of the fully hemylated protein from the assembly factors.

The house shrew (Suncus murinus-S. montanus species complex) colonized regions across southern Asia and the Indian Ocean following human activity. The house shrew is distributed on islands of the Ryukyu Archipelago, the southernmost part of Japan, but the evolutionary history of the shrew on those islands and possible associations between these populations and humans remain unknown. In this study, we conducted phylogenetic and population genetic analyses based on both nuclear and mitochondrial genome sequences of house shrews. Phylogenetic analyses based on mitochondrial cytochrome b (cytb) sequences revealed that shrews from the Ryukyu Archipelago showed strong genetic affinity to Vietnamese and southern Chinese shrews. Demographic analyses of cytb sequences indicated a rapid population expansion event affecting the haplotype group in Vietnam, southern China, and the Ryukyu Archipelago 3300-7900 years ago. Furthermore, gene flow between Ryukyu (Yonaguni Island) and Taiwan and between Ryukyu and Vietnam inferred from f4 statistics of the nuclear genomes suggested repeated immigration to Ryukyu in recent years. The present study demonstrates that the Nagasaki population has a different origin from the Ryukyu population. These findings elucidate the complex pattern of genetic admixture in house shrews and provide insights into their evolutionary history.

Haemoproteus parasites are the most diverse among Haemosporida. However, their natural vectors (Culicoides) are still poorly investigated and were identified for only a few parasite species and lineages. The application of an integrative approach (insect dissection, microscopic analysis, and molecular-based methods) is necessary in these studies, which have been carried out by a few research groups, mainly in Europe. The aim of this study was (i) to determine the Culicoides species that are naturally infected by Haemoproteus parasites, and which can support its complete sporogonic development, and (ii) to investigate the prevalence of Culicoides species and Haemoproteus parasite lineages in different study sites. In total, 1953 parous Culicoides females, from 11 species, were collected in four different localities in Lithuania and were dissected and analyzed using an integrative approach. The most abundant was C. pictipennis (30.3%). Parasite DNA was found in 7.9% of all investigated Culicoides, of which ~30% had sporozoites in their salivary glands, confirming their vector competence for these parasites. The Botanical Garden presented the highest number of Culicoides parous females, Culicoides species, and parasite lineages, as well as the highest positivity for sporozoites. Culicoides reconditus was confirmed as a natural vector of Haemoproteus parasites, sporozoites of six Haemoproteus lineages were reported for the first time, and 12 new interactions between Haemoproteus parasite lineages and Culicoides species were identified. Haemoproteus parasites seem to be transmitted by a high number of Culicoides species, with C. kibunensis, C. pictipennis, and C. segnis being the most important vectors.

BACKGROUND: The two-spotted spider mite Tetranychus urticae causes significant damage to ornamental, cotton, sugarcane and horticultural crops in Australia. It has a long history of developing resistance to many acaricides including bifenazate. A mutation in the conserved cd1- and ef-helices of the Qo pocket of cytochrome b is recognized as the primary mechanism of bifenazate resistance. To investigate the resistance mechanisms against bifenazate in Australian two-spotted spider mite, we sequenced the complete mitochondrion genome of five mite strains including a susceptible and bifenazate-resistant strain.
RESULTS: We identified a novel mutation D252N in the G126S background at cytochrome b being the cause of bifenazate resistance in a bifenazate-resistant strain, Bram. We validated the role of this mutation combination by reciprocal crosses between a bifenazate resistant and susceptible strain. By doing these crosses we confirmed the pattern of inheritance was maternal. Additionally, mitochondrial heteroplasmy was not observed by single mite genotyping of the mutations in cytb in a known bifenazate-resistant strain Bram. The phylogenetic analysis with the complete mitochondrion genome sequences revealed that Australian two-spotted spider mite strains are closely related to the green form of T. urticae found in China.
CONCLUSIONS: The novel mutation D252N found in the cytochrome b in the G126S background was revealed to be the main cause of bifenazate resistance in the Australian T. urticae strain Bram. © 2024 Society of Chemical Industry.

Tomato wilt disease caused by Fusarium oxysporum f. sp. lycopersici (Fol) results in a decrease in tomato yield and quality. Pyraclostrobin, a typical quinone outside inhibitor (QoI), inhibits the cytochrome bc1 complex to block energy transfer. However, there is currently limited research on the effectiveness of pyraclostrobin against Fol. In this study, we determined the activity of pyraclostrobin against Fol and found the EC50 values for pyraclostrobin against 100 Fol strains (which have never been exposed to QoIs before). The average EC50 value is 0.3739 ± 0.2413 μg/mL, indicating a strong antifungal activity of pyraclostrobin against Fol, as shown by unimodal curves of the EC50 values. Furthermore, we generated five resistant mutants through chemical taming and identified four mutants with high-level resistance due to the Cytb-G143S mutation and one mutant with medium-level resistance due to the Cytb-G137R mutation. The molecular docking results indicate that the Cytb-G143S or Cytb-G137R mutations of Fol lead to a change in the binding mode of Cytb to pyraclostrobin, resulting in a decrease in affinity. The resistant mutants exhibit reduced fitness in terms of mycelial growth (25 and 30 °C), virulence, and sporulation. Moreover, the mutants carrying the Cytb-G143S mutation suffer a more severe fitness penalty compared to those carrying the Cytb-G137R mutation. There is a positive correlation observed among azoxystrobin, picoxystrobin, fluoxastrobin, and pyraclostrobin for resistant mutants; however, no cross-resistance was detected between pyraclostrobin and pydiflumetofen, prochloraz, or cyazofamid. Thus, we conclude that the potential risk of resistance development in Fol toward pyraclostrobin can be categorized as ranging from low to moderate.