Keenan, T. A. | bioRxiv Science

bioRxiv2026

Freeze tolerance of a beneficial lady beetle, Hippodamia convergens

The convergent lady beetle Hippodamia convergens is widespread in the Americas and considered an important beneficial insect due to its use in pest control. Early work on this species characterized the beetles as freeze-avoidant (freeze-intolerant), suggesting they survive low winter temperatures by physiologically preventing ice formation to temperatures as low as -15{degrees}C. Here, we show that H. convergens can be freeze-tolerant if ice formation occurs at relatively high temperatures. There was 100% survival following inoculative freezing at -0.5{degrees}C and exposure to -3{degrees}C for 20 hours, as well as freezing that spontaneously occurred in fed beetles exposed to -4{degrees}C for 4 hours. Males exposed to 0{degrees}C or -4{degrees}C for 4 hours had similar mating behaviours (latency to first mating, copulation duration) as control beetles exposed to 4{degrees}C, although sample sizes were too small to determine whether freezing itself had an effect on these behaviours. Several putative cryoprotectants were detected in fat body tissue of H. convergens: glycerol, proline, trehalose, and myo-inositol - in order of abundance. Exposure to -3{degrees}C for 20 hours, whether frozen or unfrozen, did not statistically affect cryoprotectant accumulation, although there was a trend towards increased glycerol following freezing. This study is the first to describe inoculative freeze tolerance in a lady beetle and establishes a baseline for future studies that examine the mechanisms underlying this freeze tolerance and the effects of freezing on reproductive behaviour. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=69 SRC="FIGDIR/small/744689v1_ufig1.gif" ALT="Figure 1"> View larger version (16K): org.highwire.dtl.DTLVardef@1c0382org.highwire.dtl.DTLVardef@12a5932org.highwire.dtl.DTLVardef@145f4adorg.highwire.dtl.DTLVardef@1c2cb22_HPS_FORMAT_FIGEXP M_FIG C_FIG O_LIHippodamia convergens can tolerate freezing at high subzero temperatures C_LIO_LIMating behaviour of males is normal after chilling or freezing C_LIO_LICryoprotectant accumulation may support overwintering survival C_LI

zoology↗

bioRxiv2026

Overwintering under the ice: seasonal shifts in cold and hypoxia stress physiology of three winter-active pond insects

Winter poses harsh physiological challenges for insects living in temperate ponds due to the combination of low temperatures and reduced oxygen levels (hypoxia). Despite these stressors, water boatmen (Hesperocorixa sp.), backswimmers (Notonecta sp.) and diving beetles (Laccophilus maculosus) remain active year-round in eastern Nova Scotia, including in ice- covered ponds. However, the underlying mechanisms that allow pond insects to survive overwintering have not been widely studied. We hypothesized that cold and hypoxia tolerance would improve from September to March in these insects, with changes in whole-animal and biochemical correlates of tolerance to both stressors. We collected insects from the field and stocked them in outdoor freshwater mesocosms. Every two months, between September 2023 and April 2024, we characterized whole animal responses and biochemical changes. Whole insect cold tolerance assays indicated a trend of improved ability to sustain voluntary muscle control (CTmin) at lower temperatures in winter-collected insects and higher internal fluid freezing temperatures (SCP). There were no changes in whole animal correlates of hypoxia tolerance over time (surface respiration frequency, submersion and surface time). Potential cryoprotectants like proline, myo-inositol and trehalose increased in concentration in multiple insect taxa during winter but were relatively low compared to terrestrial insect studies. As lactate dehydrogenase activity did not change, there is little evidence that the insects experienced functional hypoxia sufficient to induce anaerobic metabolism. Overall, this research has established a baseline cold and hypoxia tolerance dataset in understudied pond insects. Summary StatementPond insects remain active during low temperature and low oxygen conditions associated with ice cover in winter, supported by physiological and biochemical changes.

zoology↗