Application of computer vision and deep learning models to automatically classify medically important mosquitoes in North Borneo, Malaysia.

Mosquito-borne diseases have emerged in North Borneo in Malaysia due to rapid changes in the forest landscape, and mosquito surveillance is key to understanding disease transmission. However, surveillance programmes involving sampling and taxonomic identification require well-trained personnel, are time-consuming and labour-intensive. In this study, we aim to use a deep leaning model (DL) to develop an application capable of automatically detecting mosquito vectors collected from urban and suburban areas in North Borneo, Malaysia. Specifically, a DL model called MobileNetV2 was developed using a total of 4880 images of Aedes aegypti, Aedes albopictus and Culex quinquefasciatus mosquitoes, which are widely distributed in Malaysia. More importantly, the model was deployed as an application that can be used in the field. The model was fine-tuned with hyperparameters of learning rate 0.0001, 0.0005, 0.001, 0.01 and the performance of the model was tested for accuracy, precision, recall and F1 score. Inference time was also considered during development to assess the feasibility of the model as an app in the real world. The model showed an accuracy of at least 97%, a precision of 96% and a recall of 97% on the test set. When used as an app in the field to detect mosquitoes with the elements of different background environments, the model was able to achieve an accuracy of 76% with an inference time of 47.33 ms. Our result demonstrates the practicality of computer vision and DL in the real world of vector and pest surveillance programmes. In the future, more image data and robust DL architecture can be explored to improve the prediction result.

Population genetic structure of tropical bed bug (Hemiptera: Cimicidae) populations and their breeding pattern in Iraq.

A study was conducted to investigate the population genetic structure and breeding pattern of 140 tropical bed bugs, Cimex hemipterus (F.) (Hemiptera: Cimicidae), collected from 14 infested sites in major cities in Iraq. The samples were genotyped using a set of 7 polymorphic microsatellite markers. High genetic variety was seen among populations, with an average of 2-9 alleles per locus. The number of alleles across 7 microsatellite loci was between 6 and 18. There was a notable disparity in the alleles per loci when comparing the overall population to those within it. The overall population exhibited an average observed heterozygosity of 0.175 and an average expected heterozygosity of 0.730. Among the population, the average observed heterozygosity was 0.173, while the average expected heterozygosity was 0.673. Analysis of molecular variance (AMOVA) revealed that 93% of the genetic variability was within the populations, and 7% was among them. The genetic differentiation coefficient (FST = 0.045), indicates a low degree of genetic differentiation and a high degree of inbreeding (FIS = 0.761), as indicated by notably significant positive inbreeding coefficients. Admixed individuals were revealed using STRUCTURE and neighbor-joining phylogenetic trees, demonstrating moderate gene flow between populations and a lack of genetic structure in the regional groups. Thus, both active dispersion and human-mediated dispersion possess the potential to influence the low population genetic structure of tropical bed bug C. hemipterus populations in Iraq, which can have implications toward tropical bed bug and management strategies.

Phylogenetic relationship of subterranean termite Coptotermes gestroi (Blattodea: Rhinotermitidae) inhabiting urban and natural habitats.

Coptotermes termites were collected from forestry habitats at University Sains Malaysia, Penang, while urban samples were collected from residentials from Penang and Kedah, Malaysia. Mitochondrial DNA markers, Cytochrome Oxidase 1 (CO1), and 16S ribosomal RNA (16S rRNA) genes were amplified and sequenced to confirm the species of the termite samples as Coptotermes gestroi. Through Blastn, all 25 CO1 and 16S rRNA sequences, respectively from urban and natural habitats were found to be 99.54-100.00 % similar to C. gestroi reference sequences from previous studies in Peninsular Malaysia. The phylogenetic trees constructed using Neighboring-joining (NJ) and Maximum Likelihood (ML) methods resulted in CO1 sequences clustering in two clades and 16S rRNA sequences clustering in a single clade. The overall mean distance was low for the C. gestroi populations from natural habitats and urban settings (FST = 0.004). Analysis of natural habitat populations using CO1 sequences revealed two haplotypes within the population, with a haplotype diversity (Hd) of 0.045 ± 0.005, while the urban population shared a common haplotype with the natural habitat populations and there was no haplotype diversity recorded between the populations. Urban and natural habitats included only one haplotype for 16S rRNA sequences, indicating a lack of nucleotide diversity. Based on the findings, a non-significant difference between the natural habitat and urban population suggests C. gestroi inhabiting both regions likely originated from a similar source and underwent population homogeneity in different settings facilitated by anthropogenic dispersal.

The draft genome dataset of the American cockroach, Periplaneta americana (Linnaeus, 1758) (Blattidae: Blattinae).

Periplaneta americana is a cosmopolitan pest cockroach endemic to tropical and subtropical climates. It occurs frequently in urban sewer and wastewater system and transit in human proximities, spreading pathogens that causes serious public health concerns such as asthma, allergies, and others. By using the Next-generation Sequencing (NGS) known as Illumina NovaSeq 6000, this article documents for the draft genome data set of P. americana collected in Penang Island, Malaysia. This article displays the pair-end 150 bp genome dataset and results on the sequence quality. This genome dataset presents the information for further understanding of P. americana populations at molecular level and the opportunity to develop effective control and management strategies for the species. This dataset is available under Sequence Read Archive (SRA) databases with the SRR23867103.

Metagenomic 16S rRNA amplicon data of gut microbial diversity in three species of subterranean termites (Coptotermes gestroi, Globitermes sulphureus and Macrotermes gilvus).

In this paper, we present the metagenomic dataset of gut microbial DNA of the lower group of subterranean termites, i.e. Coptotermes gestroi, and the higher groups, i.e. Globitermes sulphureus and Macrotermes gilvus, in Penang, Malaysia. Two replicates of each species were sequenced using Next-Generation Sequencing (Illumina MiSeq) and analysed via QIIME2. The results returned with 210,248 sequences in C. gestroi, 224,972 in G. sulphureus, and 249,549 in M. gilvus. The sequence data were deposited in the NCBI Sequence Read Archive (SRA) under BioProject number of PRJNA896747. The community analysis showed that Bacteroidota is the most abundant phylum in C. gestroi and M. gilvus, while Spirochaetota is prevalent in G. sulphureus.

Metagenomic 16S rDNA amplicon data of microbial diversity of Cimex hemipterus (F.) (Hemiptera: Cimicidae) treated with insect growth regulators (IGR).

The metagenomics dataset presented here is based on bacterial 16S rDNA gene amplicons of DNA extracted from tropical bed bugs (Cimex hemipterus). Amplicon-based sequencing was performed using the Illumina MiSeq platform, and the raw sequence data were analyzed using QIIME (version 2022.8.3). The metagenome sequence comprised ten samples that include C1 (133 511bps), C2 (108 920bps), CH1 (106 562bps), CH2 (101 778bps), P1 (103 618bps), P2 (133 258bps), T1 (113 558bps), T2 (133 952bps), TM1 (125 335bps), and TM2 (118 345bps). The sequence data is readily accessible at the NCBI SRA under bio project PRJNA918835. The most abundant microbial community present in the C. hemipterus is the Proteobacteria, with more than 99% of the abundance.

Human profiling from STR and SNP analysis of tropical bed bug, Cimex hemipterus, for forensic science.

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.

Efficacy and toxicity of cockroach gel bait (imidacloprid 2.5% and fipronil 0.05%) against American cockroach infestation in sewer system.

Background: The American cockroach, Periplaneta americana, is the most prevalent domiciliary and invasive urban pest in the sewer and waste water system. It poses a substantial threat to human public health and home allergens. This study was conducted at Universiti Sains Malaysia main campus to highlight and provide information on the control and management of American cockroach infestations in sewage systems by comparing the efficiency of fipronil 0.05% gel bait with imidacloprid 2.5% gel bait. A total number of adult and nymph stages of trapped American cockroaches was recorded using glass jar traps at eight sampling sites. Gel baits were placed beneath the lids of each manhole shaft along the inner wall perimeter. Results: The use of fipronil and imidacloprid gel baits in the sewer system resulted in a significant difference (P = 0.013). Imidacloprid 2.5% gel bait, compared to fipronil 0.05% gel bait, is the most effective treatment technique for reducing American cockroaches' population in sewer systems, with a high reduction percentage mean for both adult (91.17%) and nymph (85.50%) stages. Conclusion: As a conclusion, imidacloprid gel bait can effectively control cockroaches in sewer systems up to eight weeks.

Parentage Assignment Using Microsatellites Reveals Multiple Mating in Aedes aegypti (Diptera: Culicidae): Implications for Mating Dynamics.

The mosquito Aedes aegypti is the primary vector of the dengue, yellow fever, and chikungunya viruses. Evidence shows that Ae. aegypti males are polyandrous whereas Ae. aegypti females are monandrous in mating. However, the degree to which Ae. aegypti males and females can mate with different partners has not been rigorously tested. Therefore, this study examined the rates of polyandry via parentage assignment in three sets of competitive mating experiments using wild-type male and female Ae. aegypti. Parentage assignment was monitored using nine microsatellite DNA markers. All Ae. aegypti offspring were successfully assigned to parents with 80% or 95% confidence using CERVUS software. The results showed that both male and female Ae. aegypti mated with up to 3-4 different partners. Adults contributed differentially to the emergent offspring, with reproductive outputs ranging from 1 to 25 viable progeny. This study demonstrates a new perspective on the capabilities of male and female Ae. aegypti in mating. These findings are significant because successful deployment of reproductive control methods using genetic modification or sterile Ae. aegypti must consider the following criteria regarding their mating fitness: 1) choosing Ae. aegypti males that can mate with many different females; 2) testing how transformed Ae. aegypti male perform with polyandrous females; and 3) prioritizing the selection of polyandrous males and/or females Ae. aegypti that have the most offspring.

Analysis of Transcriptome Difference between Blood-Fed and Starved Tropical Bed Bug, Cimex hemipterus (F.) (Hemiptera: Cimicidae).

The reference transcriptome for Cimex hemipterus (tropical bed bug) was assembled de novo in this study, and differential expression analysis was conducted between blood-fed and starved tropical bed bug. A total of 24,609 transcripts were assembled, with around 79% of them being annotated against the Eukaryotic Orthologous Groups (KOG) database. The transcriptomic comparison revealed several differentially expressed genes between blood-fed and starved bed bugs, with 38 of them being identifiable. There were 20 and 18 genes significantly upregulated in blood-fed and starved bed bugs, respectively. Differentially expressed genes (DEGs) were revealed to be associated with regulation, metabolism, transport, motility, immune, and stress response; endocytosis; and signal transduction. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed an enrichment of genes encoding steroid biosynthesis, glycosaminoglycan biosynthesis, butanoate metabolism, and autophagy in both blood-fed and starved bed bugs. However, in blood-fed bed bugs, genes involved in histidine metabolism, caffeine metabolism, ubiquinone/terpenoid-quinone biosynthesis, and sulfur relay system were enriched. On the other hand, starvation activates genes related to nicotinate and nicotinamide metabolism, fatty acid elongation, terpenoid backbone biosynthesis, metabolism of xenobiotics by cytochrome P450, riboflavin metabolism, apoptosis, and protein export. The present study is the first to report a de novo transcriptomic analysis in C. hemipterus and demonstrated differential responses of bed bugs in facing blood-feeding and starvation.

Laboratory efficacy and toxicology of two commercial insecticides (deltamethrin and fenitrothion) against two German cockroach field strains.

The German cockroach, Blattella germanica (L.), a prominent pest that requires management due to its detrimental economic and medical consequences. Several research discovered that German cockroaches were insecticide resistant, mainly commercial pesticides. One-week-old nymphs from two strains in Penang, Malaysia (Georgetown strain: EL and Greenlane strain: IC strain) were tested in the laboratory against two commercial insecticides, Cislin® 25 (deltamethrin) and Sumithion 50 (fenitrothion). The concentration of solutions used in the residual test based on the manufacturer labeling. Cislin® was tested at 1.90 ppm, 1.60 ppm, 1.30 ppm, and 1.0 ppm; Sumithion 50 was tested at 25 and 27.00 ppm, 23.00 ppm, 18.00 ppm, 14.00 ppm using the residual method. Probit analysis and one-way analysis of variance (ANOVA) were used to compare the data. Cislin® 25 and Sumithion 50 were more effective and high toxicity against the IC strain compared with the EL strain. Sumithion 50 demonstrated a fast knockdown time on cockroaches, but Cislin® 25 showed no knockdown time. Sumithion 50 showed a significant mortality rate in cockroaches within a short period of time compared to Cislin® 25. Both insecticides were found to be effective against both strains, but Sumithion 50 is more effective at controlling cockroaches than Cislin® 25.

Development and Characterization of Novel Polymorphic Microsatellite Markers for Tapinoma indicum (Hymenoptera: Formicidae).

Tapinoma indicum (Forel) (Hymenoptera: Formicidae) is a nuisance pest in Asia countries. However, studies on T. indicum are limited, especially in the field of molecular biology, to investigate the species characteristic at the molecular level. This paper aims to provide valuable genetic markers as tools with which to study the T. indicum population. In this study, a total of 143,998 microsatellite markers were developed based on the 2.61 × 106 microsatellites isolated from T. indicum genomic DNA sequences. Fifty selected microsatellite markers were amplified with varying numbers of alleles ranging from 0 to 19. Seven out of fifty microsatellite markers were characterized for polymorphism with the Hardy-Weinberg equilibrium (HWE) and linkage disequilibrium (LD) analysis. All seven microsatellite markers demonstrated a high polymorphic information content (PIC) value ranging from 0.87 to 0.93, with a mean value of 0.90. There is no evidence of scoring errors caused by stutter peaks, no large allele dropout, and no linkage disequilibrium among the seven loci; although loci Ti-Tr04, Ti-Tr09, Ti-Te04, Ti-Te13, and Ti-Pe5 showed signs of null alleles and deviation from the HWE due to excessive homozygosity. In conclusion, a significant amount of microsatellite markers was developed from the data set of next-generation sequencing, and seven of microsatellite markers were validated as informative genetic markers that can be utilized to study the T. indicum population.

Characterization of bacterial communities associated with blood-fed and starved tropical bed bugs, Cimex hemipterus (F.) (Hemiptera): a high throughput metabarcoding analysis.

With the development of new metagenomic techniques, the microbial community structure of common bed bugs, Cimex lectularius, is well-studied, while information regarding the constituents of the bacterial communities associated with tropical bed bugs, Cimex hemipterus, is lacking. In this study, the bacteria communities in the blood-fed and starved tropical bed bugs were analysed and characterized by amplifying the v3-v4 hypervariable region of the 16S rRNA gene region, followed by MiSeq Illumina sequencing. Across all samples, Proteobacteria made up more than 99% of the microbial community. An alpha-proteobacterium Wolbachia and gamma-proteobacterium, including Dickeya chrysanthemi and Pseudomonas, were the dominant OTUs at the genus level. Although the dominant OTUs of bacterial communities of blood-fed and starved bed bugs were the same, bacterial genera present in lower numbers were varied. The bacteria load in starved bed bugs was also higher than blood-fed bed bugs.

Isolation and characterization of human DNA recovered from Cimex hemipterus (F.) (Hemiptera:Cimicidae).

The ability to isolate and generate a DNA profile from human DNA recovered from tropical bed bugs (Cimex hemipterus) for identifying individuals can be useful for public health, forensic, and medical entomology. In this study, genomic DNA was recovered from both male and female bed bugs at every time interval tested (0, 1, 3, 5, 7, 14, 30, and 45 days post blood meal). The total DNA concentrations recovered from male bed bugs ranged from 12.93 to 65.97 ng/µL, while the total DNA concentrations from female bed bugs ranged from 8.93 to 44.53 ng/µL. However, based on the results from the BLAST search and PCR products, human DNA could be detected from female bed bugs at 0, 3, 5, 14, and 30 days post blood meal using the D18S51 marker. Concentrations of PCR products of the D18S51 locus from male bed bugs ranged from 4.20 to 35.50 ng/µL, whereas, for female bed bugs, concentrations ranged from 4.31 to 22.47 ng/µL. These were generally higher compared to the PCR products of the first hypervariable part (HVR1) marker. The results indicate the HVR1 locus was less sensitive than the D18S51 locus.

Biting rhythm and demographic attributes of Aedes albopictus (Skuse) females from different urbanized settings in Penang Island, Malaysia under uncontrolled laboratory conditions.

Urbanization could potentially modify Aedes albopictus' ecology by changing the dynamics of the species, and affecting their breeding sites due to environmental changes, and thus contribute to dengue outbreaks. Thus, this study was conducted to evaluate the biting rhythm, fecundity and longevity of adult female Ae. albopictus in relation to urbanization strata; urban, suburban and rural areas in Penang Island, Malaysia. The experiments were done in comparison to a laboratory strain. Twenty-four hours biting activity of all the mosquito strains showed a clear bimodal biting activity, with morning and evening twilight peaks. The interaction effect between biting time and mosquito strains was not significant. Meanwhile, differences in fecundity among mosquito strains were statistically significant (F(3,442) = 10.559, P < 0.05) with urban areas having higher mean number of eggs (mean = 107.69, standard error = 3.98) than suburban (mean = 94.48, standard error = 5.18), and rural areas (mean = 72.52, standard error = 3.87). Longevity of adult females were significantly higher (F(3,441) = 31.259, P < 0.05) for mosquito strains from urban areas compared to the other strains. These findings would provide crucial information for the planning of control programs in Malaysia, particularly Penang.

Efficacy of Minimum Application of Chlorfluazuron Baiting to Control Urban Subterranean Termite Populations of Coptotermes gestroi (Wasmann) (Blattodea: Rhinotermitidae).

Termite infestations in urban areas are a serious problem because they cause negative economic effects, reduce the esthetic value of buildings, damage crops, and require household repairs. Chemical controls are the most common method used against subterranean termites, and baiting has emerged as one of the prominent control methods. The goal of this research was to determine the efficacy of termite baiting by treating one of six active termite stations (selective baiting) with chlorfluazuron baits to eradicate six populations of subterranean termites. This work shows that the placement of chlorfluazuron baits in one of the active stations was sufficient to destroy a colony that was interconnected with multiple chlorfluazuron-free stations. In general, it requires an average of 4-8 weeks for a quantity of less than 300 g of chlorfluazuron bait to remove a termite infestation at the study site.

Draft genome dataset of Tapinoma indicum (Forel) (Hymenoptera: Formicidae) in Penang Island, Malaysia.

Tapinoma indicum is a household pest that is widely distributed in Asian countries. It is known as nuisance pest that causes annoyance and disturbance by constructing nests and foraging in building for food and water. This article documents the draft genome dataset of T. indicum collected in Penang Island, Malaysia using the next-generation sequencing known as the Illumina platform. This article presents the pair-end 150 bp genome dataset and the quality of the sequencing result. This dataset provides the information for further understanding of T. indicum in the molecular aspect and the opportunity to develop a novel method for pest control and regulation. The dataset is available under Sequence Read Archive (SRA) databases with the accession number SRR10848807.

16S rRNA metagenomic data of microbial diversity of Pheidole decarinata Santschi (Hymenoptera: Formicidae) workers.

Metagenomic datasets of the microbial DNA of workers of a Pheidole decarinata Santschi (Hymenoptera: Formicidae) around houses with three replicates were presented. Next-generation sequencing of the microbial DNA was performed on an Illumina Miseq platform. QIIME (version 1.9.1) was used to analyze the raw fastq files. Metagenome of the three (3) samples consist of 333,708 sequences representing 137,359,149 bps with an average length of 413.67 bps. The sequence data is available at the NCBI SRA with the bioproject number PRJNA632430. Community analysis revealed Proteobacteria was the predominant (84.77%) microbial community present in the microbial DNA of workers of the P. decarinata.

Metagenomic 16S rDNA amplicon data of microbial diversity of guts of fully fed tropical bed bugs, Cimex hemipterus (F.) (Hemiptera: Cimicidae).

The metagenomic datasets of the microbial DNA from tropical bed bugs (Cimex hemipterus) after feeding on human blood were presented. Next-generation sequencing of the community DNA was carried out on an Illumina Miseq platform and the raw fastq files were analyzed using QIIME (version 1.9.1). The metagenome of three samples comprised of 108,198 sequences representing 44,646,263 bps with a mean length of 412.63 bps. The sequence data is accessible at the NCBI SRA under the bioproject number PRJNA600667. Community analysis showed Proteobacteria was the most abundance (more than 99%) microbial community that present in the guts of fully fed tropical bed bugs.