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The worldwide resurgence of bed bugs [both Cimex lectularius L. and Cimex hemipterus (F.)] over the past two decades is believed in large part to be due to the development of insecticide resistance. The transcriptomic and genomic studies since 2010, as well as morphological, biochemical and behavioral studies, have helped insecticide resistance research on bed bugs. Multiple resistance mechanisms, including penetration resistance through thickening or remodelling of the cuticle, metabolic resistance by increased activities of detoxification enzymes (e.g. cytochrome P450 monooxygenases and esterases), and knockdown resistance by kdr mutations, have been experimentally identified as conferring insecticide resistance in bed bugs. Other candidate resistance mechanisms, including behavioral resistance, some types of physiological resistance (e.g. increasing activities of esterases by point mutations, glutathione S-transferase, target site insensitivity including altered AChEs, GABA receptor insensitivity and altered nAChRs), symbiont-mediated resistance and other potential, yet undiscovered mechanisms may exist. This article reviews recent studies of resistance mechanisms and the genes governing insecticide resistance, potential candidate resistance mechanisms, and methods of monitoring insecticide resistance in bed bugs. This article provides an insight into the knowledge essential for the development of both insecticide resistance management (IRM) and integrated pest management (IPM) strategies for successful bed bug management.

Bed bugs pose significant public health challenges in tropical regions like Indonesia. This study aimed to identify the species of bed bugs involved in bed bug infestations across Indonesia using morphological and molecular approaches. Specimens were collected from 10 locations, including Central Java, Papua, and Kalimantan. A total of 101 C. hemipterus specimens were collected and examined morphologically, of which 23 were further analyzed using COX1 gene sequencing. Morphological identification confirmed all samples as C. hemipterus , with pronotum width-to-length ratios consistent with established keys. Molecular analysis of COX1 gene sequences revealed 98.61% to 99.77% similarity to reference sequences from Malaysia and Iraq. Phylogenetic analysis revealed strong genetic similarity among Southeast Asian populations, with minor regional variations and a distinct sub-branch for Iranian samples. The findings highlight the adaptability of C. hemipterus to diverse environments and its widespread prevalence in Indonesia. This study provides insight into genetic diversity and a foundation for future epidemiological studies.

In recent decades, the common and the tropical bed bugs have experienced a resurgence in many parts of the world. The evolution of insecticide resistance in bed bug populations is considered a significant factor contributing to this resurgence. We analyzed samples of Cimex lectularius L. and Cimex hemipterus (F.) from Europe (Spain 41, Switzerland 2, the Czech Republic 1), Asia (Hong Kong 34), North America (USA 14, Mexico 3), and South America (Colombia 3) to assess the prevalence and mechanisms of insecticide resistance. We identified specimens morphologically and barcoded them by sequencing the mitochondrial Cytochrome c oxidase subunit I (COI) and the 16S ribosomal RNA (16S rRNA) genes. Additionally, we screened segments of the voltage-gated sodium channel (VGSC) and the nicotinic acetylcholine receptor (nAChR) genes for point mutations associated with insecticide resistance and measured the activity of detoxifying enzymes. All samples from North America and Europe were identified as C. lectularius, whereas specimens from Hong Kong were C. hemipterus. Out of 64 C. lectularius samples tested for knockdown resistance (kdr) mutations, 90.6% contained at least 1 known mutation. All 35 C. hemipterus samples exhibited kdr mutations. A new mutation was identified in the pyrethroid target site in both common (F1524C) and tropical (F1450C) bed bugs. No resistance-associated mutations in the nAChR gene were found. Several populations that exhibited kdr mutations also showed elevated activity of detoxifying enzymes. The high frequency of kdr-associated mutations in bed bug populations from Spain and Hong Kong limits the efficacy of pyrethroids for their control.

Recent advances in genomic and post-genomic technologies have facilitated a genome-wide analysis of the insecticide resistance-associated genes in insects. Through bed bug, Cimex lectularius transcriptome analysis, we identified 14 molecular markers associated with pyrethroid resistance. Our studies revealed that most of the resistance-associated genes functioning in diverse mechanisms are expressed in the epidermal layer of the integument, which could prevent or slow down the toxin from reaching the target sites on nerve cells, where an additional layer of resistance ( kdr ) is possible. This strategy evolved in bed bugs is based on their unique morphological, physiological and behavioral characteristics and has not been reported in any other insect species. RNA interference-aided knockdown of resistance associated genes showed the relative contribution of each mechanism towards overall resistance development. Understanding the complexity of adaptive strategies employed by bed bugs will help in designing the most effective and sustainable bed bug control methods.

Recent resurgence of the bedbug Cimex lectularius is a global problem on the public health. On account of the worldwide rise of insecticide-resistant bedbug populations, exploration of new approaches to the bedbug control and management is anticipated. In this context, gene silencing by RNA interference (RNAi) has been considered for its potential application to pest control and management, because RNAi enables specific suppression of target genes and thus flexible selection of target traits to be disrupted. In this study, in an attempt to develop a control strategy targeting reproduction of the bedbug, we investigated RNAi-mediated gene silencing of vitellogenin (Vg), a major yolk protein precursor essential for oogenesis. From the bedbug transcriptomes, we identified a typical Vg gene and a truncated Vg gene, which were designated as ClVg and ClVg-like, respectively. ClVg gene was highly expressed mainly in the fat body of adult females, which was more than 100 times higher than the expression level of ClVg-like gene, indicating that ClVg gene is the primary functional Vg gene in the bedbug. RNAi-mediated suppression of ClVg gene expression in adult females resulted in drastically reduced egg production, atrophied ovaries, and inflated abdomen due to hypertrophied fat bodies. These phenotypic consequences are expected not only to suppress the bedbug reproduction directly but also to deteriorate its feeding and survival indirectly via behavioral modifications. These results suggest the potential of ClVg gene as a promising target for RNAi-based population management of the bedbug.

The bed bug, Cimex lectularius , has re-established itself as a ubiquitous human ectoparasite throughout much of the world during the past two decades. This global resurgence is likely linked to increased international travel and commerce in addition to widespread insecticide resistance. Analyses of the C. lectularius sequenced genome (650 Mb) and 14,220 predicted protein-coding genes provide a comprehensive representation of genes that are linked to traumatic insemination, a reduced chemosensory repertoire of genes related to obligate hematophagy, host–symbiont interactions, and several mechanisms of insecticide resistance. In addition, we document the presence of multiple putative lateral gene transfer events. Genome sequencing and annotation establish a solid foundation for future research on mechanisms of insecticide resistance, human–bed bug and symbiont–bed bug associations, and unique features of bed bug biology that contribute to the unprecedented success of C. lectularius as a human ectoparasite. The bed bug, Cimex lectularius , is a ubiquitous human ectoparasite with global distribution. Here, the authors sequence the genome of the bed bug and identify reductions in chemosensory genes, expansion of genes associated with blood digestion and genes linked to pesticide resistance.

The IMD pathway is an innate immune signalling pathway regulating antibacterial humoral defence responses, and is highly conserved among diverse insects and other organisms. However, genomic studies among hemipterans suggested that certain insects from this order may lack the Imd gene, a key component of the IMD immune pathway. Our previous work identified Imd in the brown-winged green stinkbug Plautia stali, which showed low sequence homology to other insect Imd genes but exhibited functionality in the IMD pathway. Using the P. stali Imd sequence as a query, we surveyed genomic data of the common bedbug Cimex lectularius and the brown marmorated stinkbug Halyomorpha halys, both of which were thought to lack an Imd gene, and identified Imd-like gene sequences in each. RNA interference experiments demonstrated that, in both species, these Imd-like genes function similarly to the canonical Imd gene. 3D protein structure predictions confirmed that, despite extensive sequence divergence, the Imd-like translation products were structurally similar to Imd proteins of other organisms. High levels of sequence diversity and positive selection were observed with some Imd-like genes of bedbugs and stinkbugs, suggesting an ongoing evolutionary arms race between insect hosts and their microbial symbionts.

The population densities of the common bed bug, Cimex lectularius, have recently exploded worldwide. This demographic boom is mostly due to the evolution of insecticide resistance, which appears to be mainly driven by one autosomal locus in this species, identified by a Quantitative Trait Loci analysis. However, the exact gene content of this locus is still unclear, in particular regarding the inclusion of the voltage-gated sodium channel gene, due to uncertainty in previous assemblies available. To resolve this ambiguity, and more generally to provide useful resources to fight this hematophagous human parasite, we combined short, long, and Hi-C reads to produce a chromosome-scale assembly for this species. Three competing assembly strategies were used, all of which resulted in 13 autosomes plus two X chromosomes, consistent with previous cytological studies and a very recent chromosome-scale assembly. The best assembly had a total length of 507 Mb, an N50 of 35 Mb, encoded 98% of complete BUSCO genes, and covered 99% of the previous reference genome. This chromosome-scale assembly revealed that the main insecticide-resistance locus does indeed contain the voltage-gated sodium channel gene, as well as other genes possibly involved in insecticide resistance. Additionally, a population genomics analysis showed that this 7.65 Mb locus is highly differentiated between insecticide-resistant and susceptible strains, confirming previous results. We hope this high-quality, complete, and annotated genome of C. lectularius will serve as a useful resource to understand the mechanisms of insecticide resistance evolution and, more generally, better control bed bug populations.

Cimex lectularius L. populations have been documented worldwide to be resistant to pyrethroids and neonicotinoids, insecticides that have been widely used to control bed bugs. There is an urgent need to discover new active ingredients with different modes of action to control bed bug populations. Fipronil, a phenylpyrazole that targets the GABA receptor, has been shown to be highly effective on bed bugs. However, because fipronil shares the same target site with dieldrin, we investigated the potential of fipronil resistance in bed bugs. Resistance ratios in eight North American populations and one European population ranged from 1.4- to >985-fold, with highly resistant populations on both continents. We evaluated metabolic resistance mechanisms mediated by cytochrome P450s, esterases, carboxylesterases, and glutathione S-transferases using synergists and a combination of synergists. All four detoxification enzyme classes play significant but variable roles in bed bug resistance to fipronil. Suppression of P450s and esterases with synergists eliminated resistance to fipronil in highly resistant bed bugs. Target-site insensitivity was evaluated by sequencing a fragment of the Rdl gene to detect the A302S mutation, known to confer resistance to dieldrin and fipronil in other species. All nine populations were homozygous for the wild-type genotype (susceptible phenotype). Highly resistant populations were also highly resistant to deltamethrin, suggesting that metabolic enzymes that are responsible for pyrethroid detoxification might also metabolize fipronil. It is imperative to understand the origins of fipronil resistance in the development or adoption of new active ingredients and implementation of integrated pest management programs.

Tropical bed bugs, Cimex hemipterus , which commonly feeds on human blood, may be useful in forensic applications. However, unlike the common bed bug, Cimex lectularius , there is no information regarding tropical bed bug, C. hemipterus , being studied for its applications in forensics. Thus, in this study, lab-reared post-feeding tropical bed bugs were subjected to Short Tandem Repeat (STR) and Single Nucleotide Polymorphism (SNP) analyses to establish the usage of tropical bed bugs in forensics. Several post-feeding times (0, 5, 14, 30, and 45 days) were tested to determine when a complete human DNA profile could still be obtained after the bugs had taken the blood meal. The results showed that complete STR and SNP profiles could only be obtained from the D0 sample. The profile completeness decreased over time, and partial STR and SNP profiles could be obtained up to 45 days post-blood meal. The generated SNP profiles, complete or partial, were also viable for HIrisPlex-S phenotype prediction. In addition, field-collected bed bugs were also used to examine the viability of the tested STR markers, and the STR markers detected mixed profiles. The findings of this study established that the post-blood meal of tropical bed bugs is a suitable source of human DNA for forensic STR and SNP profiling. Human DNA recovered from bed bugs can be used to identify spatial and temporal relations of events.