Wax "Tails" Enable Planthopper Nymphs to Self-Right Midair and Land on Their Feet.

The striking appearance of wax 'tails' - posterior wax projections on planthopper nymphs - has captivated entomologists and naturalists alike. Despite their intriguing presence, the functional roles of these formations remain largely unexplored. This study leverages high-speed imaging to uncover the biomechanical implications of wax structures in the aerial dynamics of planthopper nymphs (Ricania sp.). We quantitatively demonstrate that removing wax tails significantly increases body rotations during jumps. Specifically, nymphs without wax undergo continuous rotations, averaging 4.2 ± 1.8 per jump, in contrast to wax-intact nymphs, who do not complete a full rotation, averaging only 0.7 ± 0.2 per jump. This along with significant reductions in angular and translational velocity from takeoff to landing suggest that aerodynamic drag forces on wax structures effectively counteract rotation. These stark differences in body rotation correlate with landing success: nymphs with wax intact achieve a near perfect landing rate of 98.5%, while those without wax manage only a 35.5% success rate. Jump trajectory analysis reveals that wax-intact jumps transition from parabolic to asymmetric shapes at higher take-off velocities and show a significantly greater reduction in velocity from takeoff to landing compared to wax-removed jumps, demonstrating how wax structures help nymphs achieve more stable, controlled descents. Our findings confirm the aerodynamic self-righting functionality of wax tails in stabilizing planthopper nymph landings, advancing our understanding of the complex relationship between wax morphology and aerial maneuverability, with broader implications for wingless insect aerial adaptations and bioinspired robotics.

Wavelength-specific negatively phototactic responses of the burrowing mayfly larvae Ephoron virgo.

Mayflies are typically negatively phototactic during larval development, whereas the adults possess positive phototaxis. However, no extensive research has been done into the wavelength dependence of phototaxis in any mayfly larvae. We measured the repellency rate of Ephoron virgo larvae to light as a function of wavelength in the 368-743 nm spectral range. We established that the magnitude of repellence increased with decreasing wavelength and the maximal responses were elicited by 400 nm violet light. This wavelength dependence of phototaxis is similar to the recently reported spectral sensitivity of positive phototaxis of the twilight-swarming E. virgo adults. Negative phototaxis not only facilitates predation evasion: avoidance of the blue-violet spectral range could also promote the larvae to withdraw towards the river midline in the case of a drop in the water level, when the underwater light becomes enriched with shorter wavelengths as a result of the decreasing depth of overhead river water.

Wax "tails" enable planthopper nymphs to self-right midair and land on their feet.

The striking appearance of wax 'tails' - posterior wax projections on planthopper nymphs - has captivated entomologists and naturalists alike. Despite their intriguing presence, the functional roles of these structures remain largely unexplored. This study leverages high-speed imaging to uncover the biomechanical implications of these wax formations in the aerial dynamics of planthopper nymphs (Ricania sp.). We quantitatively demonstrate that removing wax tails significantly increases body rotations during jumps. Specifically, nymphs without wax projections undergo continuous rotations, averaging 4.3 ± 1.9 per jump, in contrast to wax-intact nymphs, who narrowly complete a full rotation, averaging only 0.7 ± 0.2 per jump. This suggests that wax structures effectively counteract rotation through aerodynamic drag forces. These stark differences in body rotation correlate with landing success: nymphs with wax intact achieve a near perfect landing rate of 98.5%, while those without wax manage only a 35.5% success rate. Jump trajectory analysis reveals transitions from parabolic to Tartaglia shapes at higher take-off velocities for wax-intact nymphs, illustrating how wax structures assist nymphs in achieving stable, controlled descents. Our findings confirm the aerodynamic self-righting functionality of wax tails in stabilizing planthopper landings, advancing our understanding of the complex interplay between wax morphology and aerial maneuverability, with broader implications for the evolution of flight in wingless insects and bioinspired robotics.

Oxygen extraction efficiency of the tidally-ventilated rectal gills of dragonfly nymphs.

Dragonfly nymphs breathe water using tidal ventilation, a highly unusual strategy in water-breathing animals owing to the high viscosity, density and low oxygen (O2) concentration of water. This study examines how well these insects extract O2 from the surrounding water during progressive hypoxia. Nymphs were attached to a custom-designed respiro-spirometer to simultaneously measure tidal volume, ventilation frequency and metabolic rate. Oxygen extraction efficiencies (OEE) were calculated across four partial pressure of oxygen (pO2) treatments, from normoxia to severe hypoxia. While there was no significant change in tidal volume, ventilation frequency increased significantly from 9.4 ± 1.2 breaths per minute (BPM) at 21.3 kPa to 35.6 ± 2.9 BPM at 5.3 kPa. Metabolic rate increased significantly from 1.4 ± 0.3 µl O2 min-1 at 21.3 kPa to 2.1 ± 0.4 µl O2 min-1 at 16.0 kPa, but then returned to normoxic levels as O2 levels declined further. OEE of nymphs was 40.1 ± 6.1% at 21.3 kPa, and did not change significantly during hypoxia. Comparison to literature shows that nymphs maintain their OEE during hypoxia unlike other aquatic tidal-breathers and some unidirectional breathers. This result, and numerical models simulating experimental conditions, indicate that nymphs maintain these extraction efficiencies by increasing gill conductance and/or lowering internal pO2 to maintain a sufficient diffusion gradient across their respiratory surface.

Ixodes ricinus tick bacteriome alterations based on a climatically representative survey in Hungary.

IMPORTANCE: Climate-sensitive disease vectors, such as ticks, respond to the environment with changes in their microbiome. These changes can affect the emergence or re-emergence of various vector-borne pathogens, such as the causative agent of Lyme borreliosis (LB) or tick-borne encephalitis. This aspect is particularly emphasized in light of climate change. The climatically representative assessment of microbiome differences in various developmental stages of the most common Central European tick species, Ixodes ricinus, deepens our understanding of the potential climatic factors behind microbial relative abundance and interaction changes. This knowledge can support the development of novel disease vector control strategies.

Acmella oleracea extracts as green pesticides against eight arthropods attacking stored products.

Developing sustainable control tools for managing noxious pests attacking stored foodstuffs is a timely research challenge. Acmella oleracea (L.) R. K. Jansen is a crop widely cultivated for its multiple usages on an industrial level. In this study, the extracts prepared with A. oleracea aerial parts were applied on wheat kernels for the management of eight important arthropod pests attacking stored products, i.e., Cryptolestes ferrugineus, Tenebrio molitor, Oryzaephilus surinamensis, Trogoderma granarium, Tribolium castaneum, Tribolium confusum, Alphitobius diaperinus (adults/larvae), and Acarus siro (adults/nymphs). Extraction of A. oleracea was optimized on the base of the yield and content of spilanthol and other N-alkylamides which were analysed by HPLC-DAD-MS. Two concentrations of n-hexane or methanol extracts (500 ppm and 1000 ppm), obtained through Soxhlet extraction, were tested to acquire mortality data on the above-mentioned pests after 4, 8, and 16 h and 1 to 7 days of exposure. Both extracts achieved complete mortality (100.0%) of C. ferrugineus adults. In the case of A. diaperinus adults, mortalities were very low at any concentrations of both extracts. In general, the n-hexane extract was more efficient than methanol extract against almost all species and stages. Considering both extracts, the susceptibility order, from most to least susceptible species/stage, was C. ferrugineus adults > A. diaperinus larvae > C. ferrugineus larvae > T. granarium adults > T. molitor larvae > O. surinamensis adults > O. surinamensis larvae > T. confusum larvae > T. castaneum larvae > A. siro adults > T. molitor adults > A. siro nymphs > T. granarium larvae > T. castaneum adults > T. confusum adults > A. diaperinus adults. Our research provides useful knowledge on the efficacy of N-alkylamides-rich A. oleracea extracts as grain protectants, pointing out the importance of targeting the most susceptible species/ developmental stages.

A remarkable new species of the flat bug genus Nesoproxius (Hemiptera, Aradidae), the first Oceanian representative with brachyptery.

A new flat bug species, Nesoproxiuskishimotoisp. nov., from the Oceanian region (Ogasawara Islands, Japan) is described. It is the first brachypterous representative in the genus Nesoproxius. The sexual dimorphism, nymph, and habitat are also described for the first time in this genus. A key to the species of Nesoproxius is also provided.

Lanternflies (Hemiptera: Fulgoridae) of Taiwan.

The family Fulgoridae belongs to the order Hemiptera, superfamily Fulgoridea, with approximately 770 described species worldwide. Their extraordinary appearance attracts the attention of both entomologists and the public. In addition to the evolutionary issue of their special appearance, certain species are also considered notorious pests (e.g., Lycorma delicatula). Several issues widely exist in previous taxonomic studies of lanternflies: (1) application of uncertain morphological characters leads to synonymy or misidentification; (2) descriptions of male genitalia are incomplete; (3) information of nymphal morphology is insufficient. Therefore, this study aims to provide a comprehensive taxonomic study of Fulgoridae from Taiwan. In this study, eight species in six genera from Taiwan were reported, of which Limois westwoodii was recorded for the first time from Taiwan. Lycorma olivacea was proposed as a new junior synonym of L. meliae. The fifth-instar nymph of Saiva formosana was described for the first time. Detailed descriptions of these lanternflies and an identification key to adults of Fulgoridae from Taiwan were also included.

20E biosynthesis gene CYP306A1 confers resistance to imidacloprid in the nymph stage of Bemisia tabaci by detoxification metabolism.

BACKGROUND: Difference in physiology level between the immature and mature stages of insects likely contribute to different mechanisms of insecticide resistance. It is well acknowledged that insect 20-hydroxyecdysone (20E) plays an important role in many biological processes in the immature stage, whether 20E confers insecticide resistance at this specific stage is still poorly understood. By gene cloning, reverse transcription quantitative real-time PCR, RNA interference (RNAi) and in vitro metabolism experiments, this study aimed to investigate the potential role of 20E-related genes in conferring imidacloprid (IMD) resistance in the immature stage of the whitefly Bemisia tabaci Mediterranean. RESULTS: After identification of low to moderate IMD resistance in the whitefly, we found CYP306A1 of the six 20E-related genes was overexpressed in the nymph stage of the three resistant strains compared to a laboratory reference susceptible strain, but not in the adult stage. Further exposure to IMD resulted in an increase in CYP306A1 expression in the nymph stage. These results together imply that CYP306A1 may be implicated in IMD resistance in the nymph stage of the whitefly. RNAi knockdown of CYP306A1 increased the mortality of nymphs after treatment with IMD in bioassay, suggesting a pivotal role of CYP306A1 in conferring IMD resistance in the nymph stage. Additionally, our metabolism experiments in vivo showed that the content of IMD reduced by 20% along with cytochrome P450 reductase and heterologously expressed CYP306A1, which provides additional evidence for the important function of CYP306A1 in metabolizing IMD that leads to the resistance. CONCLUSION: This study uncovers a novel function of the 20E biosynthesis gene CYP306A1 in metabolizing imidacloprid, thus contributing to such resistance in the immature stage of the insect. These findings not only advance our understanding of 20E-mediated insecticide resistance, but also provide a new target for sustainable pest control of global insect pests such as whitefly. © 2023 Society of Chemical Industry.

Aphrophoridae as Potential Vectors of Xylella fastidiosa in Tunisia.

The present study is an update on the situation of potential vectors of Xylella fastidiosa in Tunisia. Investigations in nine Tunisian regions (Nabeul, Bizerte, Béja, Jendouba, Zaghouan, Kairouan, Ben Arous, Tunis and Manouba) from 2018 to 2021 allowed for the observation of 3758 Aphrophoridae among a total of 9702 Auchenorrhyncha individuals collected by sweep net. Four Aphrophoridae species were identified with Philaenus tesselatus as most abundant (62%), followed by Neophilaenus campestris (28%), Neophilaenus lineatus (5%) and Philaenus maghresignus (5%). Aphrophoridae individuals were found to be particularly abundant in both forests of Nabeul and Jendouba, secondarily in olive groves and dry grassland. Furthermore, their distribution on weed hosts was followed in these two regions where nymphs and adults are widely distributed. P. tesselatus appears to be the most abundant species as determined either by conventional sweep netting for adults or by plant sampling on Sonchus, Smyrnium, Cirsium, Rumex, Polygonum and Picris for nymphs. Limited numbers of adults of P. maghresignus were detected by sweep netting, while nymphs of this species were found on Asphodelus microcarpus only. N. campestris was found in high numbers on plants belonging to the Poaceae family in forests, dry grassland and olive groves whereas N. lineatus occurred on herbs under or near olive trees and in dry grasslands.

Multiple physiological cohorts comprise seasonal activity of wild Amblyomma americanum (Acari: Ixodidae) nymphs.

Nymphs of the hard tick Amblyomma americanum (L.) are an important life stage in the maintenance and transmission of tick-borne pathogens. As pathogen composition can vary across developmental cohorts, it is essential to understand the demographic structure of the questing population. Amblyomma americanum nymphs often display a second peak in activity during late summer, but it is unknown whether this peak represents older overwintered ticks or younger newly molted ticks. The objective of this study was to examine the heme concentration in field-captured A. americanum nymphs to determine if the questing population consists of one physiological cohort or multiple cohorts in a season. Ticks were collected from March to August in an old field of primarily non-native grasses, and heme concentration was used to assess physiological age. LOESS modeling depicted that heme concentration in the population declined from March to early July but increased in later sampling sessions. As ticks cannot replenish declining heme stores without a blood meal, a late-summer spike in heme concentration demonstrates that newly molted nymphs are entering the active population. The vector potential of these newly emerging nymphs may differ from those collected earlier in the year as pathogen diversity depends on reservoir host dynamics and timing of larval feeding.

The Molecular Detection of Bacterial Infections of Public Health Importance in Hard Tick (Ixodidae) Nymphs Collected from the Forest Fringes of Western Ghats in the Goa, Karnataka and Maharashtra States of India.

A survey was conducted to determine the human tick-borne bacterial infections in the nymphs which were collected from Western Ghats' fringe forest areas. Tick nymphs were collected using the flagging method from the villages where cases Kyasanur Forest Disease (KFD) were previously reported in the states of Goa, Karnataka and Maharashtra. A total of 200 tick pools consisting of 4587 nymphs were tested by PCR for the detection of bacteria of public health importance, such as Coxiella burnetii and Rickettsia spp. Of these, four pools (4.8%) in Karnataka and three pools (4.4%) in Maharashtra were positive for Coxiella burnetii, while none of the samples from Goa state were positive. Rickettsia spp. were positively obtained from Maharashtra (51.5%), Goa (35.42%) and Karnataka (26.19%). The sequence results of Rickettsia spp. showed similarity to the spotted fever group Candidatus Rickettsia shennongii, Rickettsia conorii subsp. heilongjiangensis and Rickettsia spp. strain koreansis. Individuals are entering into the forest areas for various reasons are more likely to infect with Coxiella burnetii. and Rickettsia spp.

Morphometrics of the Tropical Bed Bug (Hemiptera: Cimicidae) From Cape Coast, Ghana.

Bed bugs, Cimex lectularius (L.) (Hemiptera: Cimicidae) and Cimex hemipterus (F.), have become established worldwide in recent years largely due to the development of insecticide resistance. However, limited attention has been given to ongoing morphological and macroevolutionary changes within the species and their populations, which could have implications for their control. Here, we evaluated whether bed bugs of the species C. hemipterus inhabiting different communities in Cape Coast, Ghana are undergoing segregation, which could lead to possible speciation. We also aimed to provide a morphometric description of all nymphal stages. Nine-bed bug populations of C. hemipterus were field-collected in Cape Coast and were subjected to geometric morphometric analysis. The multivariate parameters applied distinguished various populations from each of the locations, indicating the presence of morphologically distinct subpopulations of C. hemipterus. Shape-based segregation and shape changes associated with the insect pronotum (which is an important taxonomic character in the Cimicidae) were evident across the populations. Through this comparative study of C. hemipterus, we showed that possible subpopulations of this bed bug are being spread from Ghana. The nymphal stages (first-fifth) of C. hemipterus were distinguished by the length of the last three antennal segment and pronota width; such information contributes to the taxonomic knowledge of the species.

Predicting the habitat suitability of Schistosoma intermediate host Bulinus truncatus, its predatory aquatic insect Odonata nymph, and the associated aquatic plant Ceratophyllum demersum using MaxEnt.

Schistosomiasis is one of the most important parasitic diseases in tropical and subtropical areas. Its prevalence is associated with the distribution of freshwater snails, which are their intermediate hosts. Thus, control of freshwater snails is the solution to reduce the transmission of this disease. This will be achieved by understanding the relationship between the snails and their habitats including natural enemies and associated aquatic plants as well as the factors affecting their distribution. In this study, Maximum Entropy model (MaxEnt) was used for mapping and predicting the possible geographic distribution of Bulinus truncatus snail (the intermediate host of Schistosoma haematobium), Odonata nymph (predatory aquatic insect), and Ceratophyllum demersum (the associated aquatic plant) in Egypt based on topographic and climatic factors. The models of the investigated species were evaluated using the area under receiver operating characteristic curve. The results showed that the potential risk areas were along the banks of the Nile River and its irrigation canals. In addition, the MaxEnt models revealed some similarities in the distribution pattern of the vector, the predator, and the aquatic plant. It is obvious that the predictive distribution range of B. truncatus was affected by altitude, precipitation seasonality, isothermality, and mean temperature of warmest quarter. The presence of B. truncatus decreases with the increase of altitude and precipitation seasonality values. It could be concluded that the MaxEnt model could help introducing a predictive risk map for Schistosoma haematobium prevalence and performing better management strategies for schistosomiasis.

A new species of Neohagenulus Traver, 1938 from Hispaniola (Ephemeroptera, Leptophlebiidae, Hagenulinae, Hagenulini).

Here, I report a new species of the genus Neohagenulus Traver, 1938 from the Dominican Republic. The genus was believed to be endemic to Puerto Rico until now. Neohagenulushodeceki sp. nov. is described at the nymphal stage. Some discussion on the tribe Hagenulini is also provided.

A Pilot Study on the Microbiome of Amblyomma hebraeum Tick Stages Infected and Non-Infected with Rickettsia africae.

Variation in tick microbiota may affect pathogen acquisition and transmission but for many vector species, including Amblyomma hebraeum, components and determinants of the microbiome are unidentified. This pilot study aimed to determine baseline microbial community within A. hebraeum nymphs infected- and non-infected with Rickettsia africae from the environment, and within adult ticks infected- and non-infected with R. africae collected from cattle sampled from two locations in the Eastern Cape province of South Africa. Adult A. hebraeum ticks (N = 13) and A. hebraeum nymph (N = 15) preliminary screened for R. africae were randomly selected and subjected to Illumina sequencing targeting the v3-v4 hypervariable regions of the 16S rRNA gene. No significant difference in microbial community composition, as well as rarefied OTU richness and diversity were detected between adults and nymphs. Nymphs showed a higher richness of bacterial taxa indicating blood-feeding could have resulted in loss of microbial diversity during the moulting stage from nymph to adult. Core OTUs that were in at least 50% of nymphs and adults negative and positive for Rickettsia at 1% minimum relative abundance were Rickettsia, Coxiella and Ruminococcaceae UCG-005 with a single genus Arsenophonus occurring only in nymphs negative for Rickettsia. Ehrlichia spp. was present in only four nymphal ticks positive for Rickettsia. Interestingly, Rickettsia aeschlimannii was found in one nymph and one adult, indicating the first ever detection of the species in A. hebraeum. Furthermore, A. hebraeum harboured a Coxiella-like endosymbiont, which should be investigated further as Coxiella may affect the viability and transmission of other organisms.

Transcriptomic Analysis of Salivary Glands of Ornithodoros brasiliensis Aragão, 1923, the Agent of a Neotropical Tick-Toxicosis Syndrome in Humans.

Tick salivary glands produce and secrete a variety of compounds that modulate host responses and ensure a successful blood meal. Despite great progress made in the identification of ticks salivary compounds in recent years, there is still a paucity of information concerning salivary molecules of Neotropical argasid ticks. Among this group of ticks, considering the number of human cases of parasitism, including severe syndromes and hospitalization, Ornithodoros brasiliensis can be considered one of the major Neotropical argasid species with impact in public health. Here, we describe the transcriptome analysis of O. brasiliensis salivary glands (ObSG). The transcriptome yielded ~14,957 putative contigs. A total of 368 contigs were attributed to secreted proteins (SP), which represent approximately 2.5% of transcripts but ~53% expression coverage transcripts per million. Lipocalins are the major protein family among the most expressed SP, accounting for ~16% of the secretory transcripts and 51% of secretory protein abundance. The most expressed transcript is an ortholog of TSGP4 (tick salivary gland protein 4), a lipocalin first identified in Ornithodoros kalahariensis that functions as a leukotriene C4 scavenger. A total of 55 lipocalin transcripts were identified in ObSG. Other transcripts potentially involved in tick-host interaction included as: basic/acid tail secretory proteins (second most abundant expressed group), serine protease inhibitors (including Kunitz inhibitors), 5' nucleotidases (tick apyrases), phospholipase A2, 7 disulfide bond domain, cystatins, and tick antimicrobial peptides. Another abundant group of proteins in ObSG is metalloproteases. Analysis of these major protein groups suggests that several duplication events after speciation were responsible for the abundance of redundant compounds in tick salivary glands. A full mitochondrial genome could be assembled from the transcriptome data and confirmed the close genetic identity of the tick strain sampled in the current study, to a tick strain previously implicated in tick toxicoses. This study provides novel information on the molecular composition of ObSG, a Brazilian endemic tick associated with several human cases of parasitism. These results could be helpful in the understanding of clinical findings observed in bitten patients, and also, could provide more information on the evolution of Neotropical argasids.

Effects of seasonal variation on the biology and morphology of the dusky cotton bug, Oxcarenus laetus (Kirby).

The dusky cotton bug (Oxycarenus laetus, KIRBY) a pest of several crops. The effects of winter and summer on the biology and morphology was investigated. The sampled eggs of dusky cotton bug (DCB) were kept under controlled environment for biological and morphological investigations. In winter, the mating duration of DCB was observed longer significantly (74.2 days), egg development period (3.93 days), an egg laying period (3.6 days) and hatching period (6.66 days) noted longer in winter season. Interestingly, average number of hatched egg (16.8 days) observed significantly higher in summer and the percentage of the hatching of eggs (81.95%) were also observed higher in summer as compare to winter. Whereas, the longevity of all nymph stages in winter longer days as compare to summer nymph stages. Moreover, differences were also observed between male and female development days between winter and summer. In the winter, female DCB development was suggestively higher as compare to summer (24 days). Whereas, the developmental days were noted considerably more in winter for males as compare to summer (14.93 days). On other hand, for morphological parameters, no differences were observed between winter and summer population of DCB.

Identification and characterization of microRNAs in the immature stage of the beneficial predatory bug Arma chinensis Fallou (Hemiptera: Pentatomidae).

MicroRNAs (miRNAs) are a type of small noncoding RNAs that regulate gene expression at the posttranscriptional level and can influence significant biological processes. Arma chinensis (Hemiptera: Pentatomidae) is a predaceous insect species that preys upon a wide variety of insect pests. It is important to explore and understand the molecular mechanisms involving miRNAs in regulating developmental and other gene expression for beneficial insects. However, examination of miRNAs associated with Hemiptera, especially predatory bugs, has been absent or scarce. This study represents the first comprehensive analysis of predatory bug A. chinensis encoded miRNAs through high throughput sequencing and predicts genes and biological processes regulated by the newly identified miRNAs through analyzing their differential expression in and across five nymphal instars. A total of 64 A. chinensis miRNAs, including 46 conserved miRNAs and 18 novel miRNAs, were identified by analysis of high throughput sequence reads mapped to the genome. A total of 2913 potential gene targets for these 64 miRNAs were predicted by comprehensive analyses utilizing miRanda, PITA, and RNAhybrid. Gene Ontology annotation of predicted target genes of A. chinensis suggested the key processes regulated by miRNAs involved biological processes, regulation of cellular processes, and transporter activity. Kyoto Encyclopedia of Genes and Genomes pathway predictions included the Toll and Imd signaling pathway, Valine, leucine and isoleucine degradation, Steroid biosynthesis, the AGE-RAGE signaling pathway in diabetic complications, and Alanine, aspartate and glutamate metabolism. This newly identified miRNAs through analyzing their differential expression, assessment of their predicted functions forms a foundation for further investigation of specific miRNAs.

A Comparison of Questing Substrates and Environmental Factors That Influence Nymphal Ixodes pacificus (Acari: Ixodidae) Abundance and Seasonality in the Sierra Nevada Foothills of California.

In California, the western blacklegged tick, Ixodes pacificus Cooley and Kohls, is the principal vector of the Borrelia burgdorferi sensu lato (sl) complex (Spirochaetales: Spirochaetaceae, Johnson et al.), which includes the causative agent of Lyme disease (B. burgdorferi sensu stricto). Ixodes pacificus nymphs were sampled from 2015 to 2017 at one Sierra Nevada foothill site to evaluate our efficiency in collecting this life stage, characterize nymphal seasonality, and identify environmental factors affecting their abundance and infection with B. burgdorferi sl. To assess sampling success, we compared the density and prevalence of I. pacificus nymphs flagged from four questing substrates (logs, rocks, tree trunks, leaf litter). Habitat characteristics (e.g., canopy cover, tree species) were recorded for each sample, and temperature and relative humidity were measured hourly at one location. Generalized linear mixed models were used to assess environmental factors associated with I. pacificus abundance and B. burgdorferi sl infection. In total, 2,033 substrates were sampled, resulting in the collection of 742 I. pacificus nymphs. Seasonal abundance of nymphs was bimodal with peak activity occurring from late March through April and a secondary peak in June. Substrate type, collection year, month, and canopy cover were all significant predictors of nymphal density and prevalence. Logs, rocks, and tree trunks had significantly greater nymphal densities and prevalences than leaf litter. Cumulative annual vapor pressure deficit was the only significant climatic predictor of overall nymphal I. pacificus density and prevalence. No associations were observed between the presence of B. burgdorferi sl in nymphs and environmental variables.