Cytochrome C Oxidase Research Articles

A new species of Janiralata Menzies, 1951 (Isopoda: Asellota, Janiridae) from Korea

A new isopod species, Janiralata kwangsooi sp. nov., from Dokdo Island located in the East Sea off the Korean Peninsula is described here. This species is distinguished from its congeners by the number of antennular articles, setation of mouthparts and pereopods, length and width ratio of appendages, and shape of male pleopods I and II. To aid species identification, a taxonomic key to species of Janiralata Menzies, 1951 in the Japanese and Korean waters is provided. A partial sequence of mitochondrial cytochrome-c-oxidase (COI) of this new species is also provided for DNA barcoding and compared with those of congeners publicly available in GenBank.

Microbiome of lovebug (Plecia longiforceps) in Seoul, South Korea.

Lovebugs appeared in large numbers across a wide area in Seoul, South Korea, in June 2023. The sudden appearance of exotic insects not only discomforts people but also fosters anxiety, as their potential for pathogen transmission would be unknown. In this study, targeted next-generation sequencing (NGS) of the 16S rRNA gene V4 region was performed using iSeq 100 to screen for bacteria in lovebugs. Forty-one lovebugs (20 females and 21 males) collected in Seoul, Korea, were identified as Plecia longiforceps based on mitochondrial cytochrome oxidase subunit 1 sequencing data using PCR. We analyzed the microbiome of the lovebugs and detected 453 species of bacteria. Among all bacteria screened based on NGS, Rickettsia was detected in all samples with an average relative abundance of 80.40%, followed by Pandoraea and Ewingella. Diversity (alpha and beta) between females and males did not differ; however, only Tumebacillus showed a higher relative abundance in females. Sequencing analysis of Rickettsia using a gltA gene-specific primer by PCR showed that it had higher sequence similarity to the Rickettsia symbiont of arthropods than to the spotted fever group rickettsiae. Eleven samples in which Pandoraea was detected by iSeq 100 were confirmed by PCR and exhibited 100% sequence identity to Pandoraea oxalativorans strain DSM 23570. Consequently, the likelihood of pathogen transmission to humans is low. The applied method may play a crucial role in swiftly identifying bacterial species in the event of future outbreaks of exotic insects that may be harmful to humans.IMPORTANCELovebugs have recently emerged in large numbers in Seoul, causing major concern regarding potential health risks. By performing the next-generation sequencing of the 16S rRNA gene V4 region, we comprehensively examined the microbiome of these insects. We identified the presence of numerous bacteria, including Rickettsia and Pandoraea. Reassuringly, subsequent tests confirmed that these detected bacteria were not pathogenic. The present study addresses health concerns related to lovebugs and shows the accuracy and efficiency of our detection technique. Such methods prove invaluable for rapidly identifying bacterial species during potential outbreaks of unfamiliar insects, thereby ensuring public safety.

Genetic diversity and haplotypes of Cysticercus tenuicollis isolates from slaughtered sheep and goats in Elazig and Bingol provinces of Turkey.

The cestode Taenia hydatigena uses canids, primarily dogs, as definitive hosts, whereas the metacestode larval stage cysticercus infects a range of intermediate hosts, including domestic animals such as goats, sheep and pigs. In this study, we aimed to determine the genetic differences and haplotypes of Cysticercus tenuicollis isolates obtained from sheep and goats slaughtered in the Bingol and Elazig provinces of Turkey. C. tenuicollis isolates were collected from 44 sheep and 26 goats slaughtered in slaughterhouses in Bingol and Elazig. After the isolation of total genomic DNA from C. tenuicollis isolates, the genetic characterization of the partial mitochondrial cytochrome oxidase 1 (CO1) gene region (866bp) was amplified using specific primers by polymerase chain reaction (PCR), the products were then sequenced, and haplotype and genetic diversity analyses were carried out. As a result of the haplotype network analyses, 34 different haplotypes were detected around the main haplotype (Hap02) arranged in a star-like configuration and separated from other haplotypes by 1-28 mutation steps and covering 22.85% (16/70) of all isolates. Twenty-seven polymorphic fields were detected, 77.77% (21/27) of which were parsimony-informative, and secondary haplotype and nucleotide diversity were observed. Additionally, we detected high intraspecies haplotype diversity (hd: 0.933) and high nucleotide diversity (π: 0.00383), with 27 different nucleotide variation positions among the haplotypes of the isolates. Tajima's D value was negative, indicating population expansion and/or selection purification. The significantly negative Fu's Fs values indicated recent population expansion or the presence of expected rare haplotypes. The results of this study confirmed that C. tenuicollis isolates clustered in one lineage and were closely related to the relevant reference sequences in different countries, confirming the circulation of C. tenuicollis in different geographical regions.

CoSFISH: a comprehensive reference database of COI and 18S rRNA barcodes for fish.

Fish, being a crucial component of aquatic ecosystems, holds significant importance from both economic and ecological perspectives. However, the identification of fish at the species level remains challenging, and there is a lack of a taxonomically complete and comprehensive reference sequence database for fish. Therefore, we developed CoSFISH, an online fish database. Currently, the database contains 21 535 cytochrome oxidase I sequences and 1074 18S rRNA sequences of 21 589 species, belonging to 8 classes and 90 orders. We additionally incorporate online analysis tools to aid users in comparing, aligning and analyzing sequences, as well as designing primers. Users can upload their own data for analysis, in addition to using the data stored in the database directly. CoSFISH offers an extensive fish database and incorporates online analysis tools, making it a valuable resource for the study of fish diversity, phylogenetics and biological evolution. Database URL: http://210.22.121.250:8888/CoSFISH/home/indexPage.

Biodiversity of Demersal Fish Communities in the Cosmonaut Sea Revealed by DNA Barcoding Analyses.

The Cosmonaut Sea is one of the least accessed regions in the Southern Ocean, and our knowledge about the fish biodiversity in the region is sparse. In this study, we provided a description of demersal fish diversity in the Cosmonaut Sea by analysing cytochrome oxidase I (COI) barcodes of 98 fish samples that were hauled by trawling during the 37th and 38th Chinese National Antarctic Research Expedition (CHINARE) cruises. Twenty-four species representing 19 genera and 11 families, namely, Artedidraconidae, Bathydraconidae, Bathylagidae, Channichthyidae, Liparidae, Macrouridae, Muraenolepididae, Myctophidae, Nototheniidae, Paralepididae and Zoarcidae, were discriminated and identified, which were largely identical to local fish occurrence records and the general pattern of demersal fish communities at high Antarctic shelf areas. The validity of a barcoding gap failed to be detected and confirmed across all species due to the indicative signals of two potential cryptic species. Nevertheless, DNA barcoding still demonstrated to be a very efficient and sound method for the discrimination and classification of Antarctic fishes. In the future, various sampling strategies that cover all geographic sections and depth strata of the Cosmonaut Sea are encouraged to enhance our understanding of local fish communities, within which DNA barcoding can play an important role in either molecular taxonomy or the establishment of a dedicated local reference database for eDNA metabarcoding analyses.

DNA barcoding for identification of forensically important synanthropic flesh flies (Diptera: Sarcophagidae) from Eastern India

ABSTRACT Sarcophagidae is a significant Diptera family because several of its species are among the first to visit a corpse, and it helps in minPMI estimation. Sarcophagidae species identification for a layman is difficult due to identical morphological appearance and if the fly is a female. Also, fly specimens get damaged due to improper storage of forensic specimens collected from crime scenes. Thus, in this study, we aimed to check the efficiency of cytochrome oxidase 1 (COI) for identifying eleven sarcophagid flies species. Flesh flies were collected from various biogeographic regions of West Bengal. COI barcodes successfully differentiated between species with a very significant K2P interspecific genetic divergence, which ranged between 3.12% and 20.11%, and intraspecific genetic divergences ranged from 0% to 1%. The subgenera Uroxanthisca and Sinonipponia are used in the phylogenetic analysis for the first time in the oriental context. Phylogenetic analysis was performed using a Neighbour Joining (NJ) tree constructed using Kimura two-parameter (K2P) in MEGA X software. Thus, COI barcodes proved to be quite effective as an alternate method of identifying flesh fly species, and future studies should include additional markers for further phylogenetic investigations of the Sarcophagidae family.

DNA Barcoding Revealed a High Percentage of Mislabeling in Commercial Fish Products: The First Empirical Survey in South Texas

Seafood mislabeling is a widespread problem that have produced a growing distrust of seafood industry. In this study, we examined the prevalence of mislabeling in fish samples from regional markets in the greater Houston area and close-by coastal communities. A total of 63 fish fillet samples were purchased, labeled, and stored at -20˚C in individual packages until DNA extraction. DNA fragments of roughly 700 base pairs encoding cytochrome oxidase I (COI) were PCR-amplified from each DNA sample. With 99.6–100% nucleotide identity in the GenBank and BOLD databases, all samples were correctly identified at the species level. The scientific names identified by DNA barcoding were matched with legally acceptable market names using the US Food and Drug Administration (FDA) Seafood List. Out of the 63 samples examined, 13 samples (20.6%) were mislabeled. Moreover, 24 samples (38.1%) did not use the acceptable market names, indicating that the FDA policy is poorly implemented in seafood industry. The first DNA barcoding survey this area warranted the need of continuous monitoring and the dissemination of the regulation combined with taxonomic knowledge.

6-BA Delays the Senescence of Postharvest Cabbage Leaves by Inhibiting Respiratory Metabolism.

6-BA, a small molecule compound of cytokinins, has been proven to delay leaf senescence in different species, including Chinese flowering cabbage; however, its specific mechanism remains relatively unknown. In this study, the application of external 6-BA delayed leaf senescence in Chinese flowering cabbage, showing that 6-BA effectively prevented the decrease in the maximum quantum yield (Fv/Fm) and overall chlorophyll content and suppressed the expression of the senescence-associated gene BrSAG12 over a 7-day period of storage. Moreover, treatment with 6-BA decreased the respiratory rate, NAD(H) content, the activities of hexose phosphate isomerase (PHI), succinate dehydrogenase (SDH), cytochrome c oxidase (CCO), and ascorbic acid oxidase (AAO) using enzyme-linked immunosorbent assay, and the transcriptional abundance of related genes by real-time quantitative polymerase chain reaction. Furthermore, 6-BA also increased the activity and expression levels of glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphate gluconate dehydrogenase (6-PGDH). The group treated with 6-BA retained elevated levels of NADP (H), ATP, total ATPase, and nicotinamide adenine dinucleotide kinase (NADK) activity, as well as the expression of respiratory enzymes. Molecular docking indicated that 6-BA hinders the glycolysis pathway (EMP), tricarboxylic acid cycle (TCA), and cytochrome pathway (CCP), and sustains elevated levels of the pentose phosphate pathway (PPP) through interactions with the PHI, SDH, 6-PGDH, G6PDH, CCO, and AAO proteins, consequently delaying postharvest leaf senescence in Chinese flowering cabbage.

A review of the varunid crab genus Parapyxidognathus Ward, 1941 (Decapoda: Brachyura: Varunidae), with descriptions of two new species

Abstract Species of Parapyxidognathus Ward, 1941 are distributed across tropical Indo-West Pacific to temperate downstream rivers and in estuaries. The genus previously included only one species, Parapyxidognathus deianira (De Man, 1888). We examined specimens from Parapyxidognathus and its closely related genus, Pyxidognathus A. Milne-Edwards, 1879 from the eastern Indian Ocean and western Pacific, focusing on their morphological characters. The results also support the reclassification of Pyxidognathus fluviatilis Alcock, 1900 into Parapyxidognathus, as well as the establishment of two new species from the western Pacific, Pa. tshiansuesp. nov. and Pa. ongiasp. nov. The identity of the species is supported by evidence from mitochondrial 16S rDNA and cytochrome oxidase subunit 1 (COI) genes. There are now four species in Parapyxidognathus distinguishable by the characters of the anterolateral teeth of the carapace, the pleon, and the first gonopods of the male, and the female vulvae.

Field monitoring and genetic diversity of the large copper butterfly <i>Lycaena dispar</i> (Lepidoptera: Lycaenidae)

The large copper butterfly Lycaena dispar (Haworth, 1803) (Lepidoptera: Lycaenidae), which is found across Europe and Asia, has been categorized as a near-threatened species (NT) in South Korea from 2012 mainly due to limited distribution. The species has been infrequently observed outside its traditional range in the northwestern region, but no nationwide survey has yet been conducted. In addition, the population genetic data of L. dispar, which is informative to evaluate its vulnerability, remains unknown. In this study, we visited 36 sites spread across all South Korean provinces to verify the distributional range of the species. Mitochondrial cytochrome oxidase subunit I (COI) was also partially sequenced for 53 individuals from nine sites. We observed L. dispar at 15 sites in six provinces, including the two previously known provinces, indicating a southward range expansion. The in-field monitoring and genetic data collectively suggested that L. dispar does not have a limited distribution nor is it isolated, indicating that it should be reclassified as less vulnerable. Our study demonstrates that the combination of field and genetic data can provide a more reliable assessment of the stability of a species.

Genetic variability of Aedes aegypti (Diptera: Culicidae) in El Salvador and Honduras: presence of a widespread haplotype and implications for mosquito control

BackgroundThis study examined population genetics of Aedes aegypti in El Salvador and Honduras, two adjacent countries in Central America. Aedes aegypti is associated with yellow fever, dengue, chikungunya, and Zika. Each year, thousands of cases of dengue are typically reported in El Salvador and Honduras.MethodsIn El Salvador, collections were obtained from five Departments. In Honduras, samples were obtained from six municipalities in four Departments. Mitochondrial DNA cytochrome oxidase I (COI) was sequenced, and consensus sequences were combined with available sequences from El Salvador to determine haplotype number, haplotype diversity, nucleotide diversity, and Tajima’s D. A haplotype network was produced to examine the relationship between genotypes.ResultsIn El Salvador, there were 17 haplotypes, while in Honduras there were 4 haplotypes. In both El Salvador and Honduras, Haplotype 1 is most abundant and widespread. In El Salvador, haplotype H2 was also widespread in 10 of 11 sampled municipalities, but it was not present in Honduras. The capital of El Salvador (San Salvador) and the eastern region of ES had the highest haplotype diversity of regions sampled.ConclusionsHaplotype 1 and H2 each belong to different phylogenetic lineages of Ae. aegypti. The most geographically widespread haplotype (H1) may have been present the longest and could be a remnant from previous eradication programs. These data may contribute to future control programs for Ae. aegypti in the two countries.Graphical

The fungicide cymoxanil impairs respiration in Saccharomyces cerevisiae via cytochrome c oxidase inhibition.

Cymoxanil (CYM) is a widely used synthetic acetamide fungicide, but its biochemical mode of action remains elusive. Since CYM inhibits cell growth, biomass production, and respiration in Saccharomyces cerevisiae, we used this model to characterize the effect of CYM on mitochondria. We found it inhibits oxygen consumption in both whole cells and isolated mitochondria, specifically inhibiting cytochrome c oxidase (CcO) activity during oxidative phosphorylation. Based on molecular docking, we propose that CYM blocks the interaction of cytochrome c with CcO, hampering electron transfer and inhibiting CcO catalytic activity. Although other targets cannot be excluded, our data offer valuable insights into the mode of action of CYM that will be instrumental in driving informed management of the use of this fungicide.

Next generation sequencing improves the resolution of detecting mixed host blood meal sources in field collected arboviral mosquito vectors.

Accurate knowledge of blood meal hosts of different mosquito species is critical for identifying potential vectors and establishing the risk of pathogen transmission. We compared the performance of Miseq next generation sequencing approach relative to conventional Sanger sequencing approach in identification of mosquito blood meals using genetic markers targeting the 12S rRNA and cytochrome oxidase I (COI) genes. We analysed the blood meals of three mosquito vector species (Aedes aegypti, Aedes simpsoni s.l. and Culex pipiens s.l.) collected outdoors, and compared the frequency of single- versus multiple-blood feeding. Single host blood meals were mostly recovered for Sanger-based sequencing of the mitochondrial 12S rRNA gene, whereas Miseq sequencing employing this marker and the COI marker detected both single and multiple blood meal hosts in individual mosquitoes. Multiple blood meals (two or more hosts) which mostly included humans were detected in 19%-22.7% of Ae. aegypti samples. Most single host blood meals for this mosquito species were from humans (47.7%-57.1%) and dogs (9.1%-19.0%), with livestock, reptile and rodent hosts collectively accounting for 4.7%-28.9% of single host blood meals. The frequency of two or more host blood meals in Ae. simpsoni s.l. was 26.3%-45.5% mostly including humans, while single host blood meals were predominantly from humans (31.8%-47.4%) with representation of rodent, reptile and livestock blood meals (18.2%-68.2%). Single host blood meals from Cx. pipiens s.l. were mostly from humans (27.0%-39.4%) and cows (11.5%-27.36%). Multiple blood meal hosts that mostly included humans occurred in 21.2%-24.4% of Cx. pipiens s.l. samples. Estimated human blood indices ranged from 53%-76% for Ae. aegypti, 32%-82% for Ae. simpsoni s.l. and 26%-61% for Cx. pipiens s.l. and were consistently lower for Sanger-based sequencing approach compared to Miseq-based sequencing approach. These findings demonstrate that Miseq sequencing approach is superior to Sanger sequencing approach as it can reliably identify mixed host blood meals in a single mosquito, improving our ability to understand the transmission dynamics of mosquito-borne pathogens.

Prevalence and molecular characterization of Hysterothylacium species infecting Pandora (Pagellus erythrinus) in the Mediterranean Sea of Egypt

Species of the genus Hysterothylacium are aquatic roundworms (nematodes) belonging to the family Raphidascarididae. Some species in this family are known to be associated with zoonotic diseases in humans after they consume their parasitic larvae in raw or undercooked fish. The aim of this research was to report the prevalence, morphology, and molecular characteristics of Hysterothylacium species in Pagellus erythrinus. A total of Two hundred fish were purchased from the fish market in Damanhour, Beheira Province, between December 2021 and November 2022 and subjected to examination. For molecular characterization, the internal transcribed spacer (ITS) region of nuclear ribosomal DNA and the mitochondrial cytochrome oxidase subunit 2 (COX-2) gene were used. Hysterothylacium species were morphologically described and identified from the intestine of Pagellus erythrinus in Beheira Province, Egypt. The PCR amplified 1087 bp and 629 bp of the target sequences of the ITS region and COX-2 gene, respectively. Sequence analysis revealed the Hysterothylacium thalassini species. The identified species provided novel biological data for the Hysterothylacium nematode in Pagellus erythrinus. The prevalence of Hysterothylacium species recovered from the intestine was 55%. The highest prevalence of 72% has been reported in summer compared to the lowest prevalence of 38% in the winter. Females had a higher prevalence of 61.8% than males, with 44.2%. The first detection, prevalence, and molecular characterization of H. thalassini in Pagellus erythrinus from Beheira Province, Egypt, was presented in this study.

Maximization of cytochrome C oxidase enzyme activity by optimizing color conversion from red organic light-emitting diodes

Photobiomodulation (PBM) therapy using red and near-infrared (NIR) light can effectively activate cytochrome C oxidase (CCO) enzyme to promote wound healing in living organisms. To maximize this therapeutic effect, the use of a light source with a spectrum absorbed by the CCO enzyme is necessary. A red/NIR light source optimized for CCO enzyme was obtained by partially color-converting a red organic light-emitting diode with copper indium sulfide/zinc sulfide quantum dots (QDs). To increase color conversion efficiency, the dispersion of QDs was improved by using a fibrous color conversion layer (FCCL). By optimizing the QD content in the FCCL, a red/NIR light source similar to the absorption spectrum of CCO enzyme was obtained with a spectral matching ratio of 81.7 %. When this light source was used in PBM therapy, it was experimentally verified that it showed a much faster wound closure effect compared to general red light, and it was proven that this was due to increased activity of the CCO enzyme. This study is a method of obtaining a red/NIR light source for PBM suitable for a specific enzyme performing a specific function, and is particularly useful due to its excellent form factor of being large, thin, and flexible.

Molecular taxonomy of fish larvae in the Northwestern Arabian gulf: A baseline study from Kuwait's first marine protected area

Sulaibikhat Bay, a shallow embayment dominated by tides and extensive mud flats along the northern coast of Kuwait Bay, serves as a crucial nursery ground for various commercially important species of finfish and shellfish in the Northwestern Arabian Gulf (NWAG). As the location of Kuwait's inaugural marine protected area (MPA), Sulaibikhat Bay plays a vital role in fostering numerous fish and marine invertebrate species of commercial importance in the Arabian/Persian Gulf. Accurate identification of larval specimens from this habitat is essential for understanding their spawning dynamics and recruitment process. However, limitations in taxonomic identification hinder scientific understanding of fish larval recruitment. To address this, our baseline study employed an integrated taxonomic approach, combining conventional morphology with molecular techniques. Fish larvae collected from the MPA and surrounding waters in Spring (2022) were identified up to the genus and species levels using partial gene sequencing of 16 S rRNA and mitochondrial cytochrome oxidase (COI) genes. Morphological analysis indicated the prevalence of Sparidae, Sillaginidae, and Dorosomatidae families. Molecular analysis confirmed the presence of these families and identified two species, Crenidens crenidens (Sparidae) and Nematalosa nasus (Dorosomatidae), along with two genera, Diplodus sp. (Sparidae) and Sillago sp. (Sillaginidae), which exhibited maximum similarity to available COI sequences of Diplodus sargus, and Sillago maculata, respectively. This represents the first baseline molecular study on fish larvae from NWAG, laying the groundwork for future in-depth analyses with more larval specimens and spatiotemporal samplings from the entire exclusive economic zone (EEZ) of Kuwait in the Arabian/Persian Gulf.

Pterygodermatites (Mesopectines) whartoni (Nematoda: Rictulariidae) encysted larvae in invasive Cuban treefrogs (Osteopilus septentrionalis) from Florida, United States.

Species of Pterygodermatites are spirurid nematodes that have expanded their geographic distribution worldwide. They infect a variety of mammalian definitive hosts with few reports of potential paratenic infections in amphibian and reptile hosts. In this study, we report Pterygodermatites sp. larvae identified in free-ranging, invasive Cuban treefrogs (Osteopilus septentrionalis), from central Florida, United States. Encysted larvae were recovered from the skeletal muscle and/or the coelomic cavity of three frogs; molecular characterization of the small subunit (18S) ribosomal RNA and cytochrome oxidase I genes of the parasites matched reported sequences of Pterygodermatites (Mesopectines) whartoni (Tubangui, 1931). This is a parasite native to Southeastern Asia and to the best of the authors' knowledge, it is the first report of the species in the New World. The recovery of invasive Pterygodermatites from invasive Cuban treefrogs in North America highlights the growing concern regarding the potential impact non-native parasites and invasive species may have on native wildlife populations.

DETERMINATION OF ENTOMOPATHOGENIC NEMATODE FAUNA IN ŞIRNAK, TURKIYE

This study focuses on the isolation, identification, and characterization of entomopathogenic nematodes (EPNs) in Şırnak Province, Turkey. Between July and September 2023, we collected 256 soil samples, from which 11 EPN isolates were successfully recovered,representing a 4.3% of the samples. These isolates were identified using morphological examinations and molecular diagnostics, including mitochondrial COI and 18S rDNA gene regions. Our findings revealed a diverse population of EPNs, predominantly Heterorhabditis bacteriophora with five isolates, alongside four Steinernema species (one Steinernema carpocapsae, one Steinernema affine, and two Steinernema sp.). Phylogenetic analysis, employing the mitochondrial cytochrome oxidase subunit I (COXI) and the 18S rDNA gene regions, indicated low genetic variability within H. bacteriophora and greater diversity among Steinernema species. The study contributes significant insights into the EPN biodiversity in Şırnak Province and underscores the potential of these nematodes in biological pest control, offering a sustainable alternative to chemical pesticides. The results highlight the importance of local EPN isolates in the development of effective biological control strategies and the conservation of agricultural biodiversity.

Range extension of Blackstripe Lizardfish, Synodus nigrotaeniatus Allen, Erdmann & Peristiwady, 2017 (Actinopterygii, Aulopiformes, Synodontidae), from Jakarta Bay, Indonesia

Three specimens (194.4–203.8 mm standard lengths) of the lizardfish Synodus nigrotaeniatus Allen, Erdmann & Peristiwady, 2017, collected from Jakarta Bay, western Indonesia, represent a significant range extension for the species, which was previously known from only six specimens collected in Lembeh Strait, North Sulawesi, eastern Indonesia. Morphological comparisons with available data from the type specimens and molecular data from the mitochondrial cytochrome oxidase subunit I (COI) gene are presented. The COI sequence data show a 4% genetic difference from the related Synodus sageneus Waite, 1905.

Genome divergence and reproductive incompatibility among populations of Ganaspis near brasiliensis.

During the last decade, the spotted wing drosophila, Drosophila suzukii, has spread from eastern Asia to the Americas, Europe, and Africa. This fly attacks many species of cultivated and wild fruits with soft, thin skins, where its serrated ovipositor allows it to lay eggs in undamaged fruit. Parasitoids from the native range of D. suzukii may provide sustainable management of this polyphagous pest. Among these parasitoids, host-specificity testing has revealed a lineage of Ganaspis near brasiliensis, referred to in this paper as G1, that appears to be a cryptic species more host-specific to D. suzukii than other parasitoids. Differentiation among cryptic species is critical for introduction and subsequent evaluation of their impact on D. suzukii. Here, we present results on divergence in genomic sequences and architecture and reproductive isolation between lineages of Ganaspis near brasiliensis that appear to be cryptic species. We studied five populations, two from China, two from Japan, and one from Canada, identified as the G1 vs G3 lineages based on differences in cytochrome oxidase l sequences. We assembled and annotated the genomes of these populations and analyzed divergences in sequence and genome architecture between them. We also report results from crosses to test reproductive compatibility between the G3 lineage from China and the G1 lineage from Japan. The combined results on sequence divergence, differences in genome architectures, ortholog divergence, reproductive incompatibility, differences in host ranges and microhabitat preferences, and differences in morphology show that these lineages are different species. Thus, the decision to evaluate the lineages separately and only import and introduce the more host-specific lineage to North America and Europe was appropriate.