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  1. Colloff MJ, Cameron SL
    Zootaxa, 2014;3780:263-81.
    PMID: 24871836 DOI: 10.11646/zootaxa.3780.2.3
    The genus Austronothrus was previously known from three species recorded only from New Zealand. Austronothrus kinabalu sp. nov. is described from Sabah, Borneo and A. rostralis sp. nov. from Norfolk Island, south-west Pacific. A key to Austronothrus is included. These new species extend the distribution of Austronothrus beyond New Zealand and confirms that the subfamily Crotoniinae is not confined to former Gondwanan landmasses. The distribution pattern of Austronothrus spp., combining Oriental and Gondwanan localities, is indicative of a curved, linear track; consistent with the accretion of island arcs and volcanic terranes around the plate margins of the Pacific Ocean, with older taxa persisting on younger island though localised dispersal within island arc metapopulations. Phylogenetic analysis and an area cladogram are consistent with a broad ancestral distribution of Austronothrus in the Oriental region and on Gondwanan terranes, with subsequent divergence and distribution southward from the Sunda region to New Zealand. This pattern is more complex than might be expected if the New Zealand oribatid fauna was derived from dispersal following re-emergence of land after inundation during the Oligocene (25 mya), as well as if the fauna emanated from endemic, relictual taxa following separation of New Zealand from Gondwana during the Cretaceous (80 mya).
  2. Dang K, Doggett SL, Lee CY
    J Econ Entomol, 2023 Feb 10;116(1):29-39.
    PMID: 35639556 DOI: 10.1093/jee/toac068
    The residual performance of two pyrethroid-neonicotinoid mixture formulations: Temprid SC (10.5% beta-cyfluthrin and 21% imidacloprid) and Tandem (3.5% lambda-cyhalothrin and 11.6% thiamethoxam) on two substrates (glass and filter paper) against eight pyrethroid-resistant strains (BM-MY, BP-MY, CH-MY, GL-MY, KL-MY, SAJ-MY, TT-MY, and QLD-AU) of the tropical bed bug, Cimex hemipterus (F.) (Hemiptera: Cimicidae) collected from Malaysia, and Australia were evaluated. The aging effect of treatment residues on glass was also investigated. A susceptible C. lectularius L. strain (Monheim) was used for comparison. Temprid SC showed varying levels of performance against all C. hemipterus strains: TT-MY (PR50 = 6.5-fold, high performance), BM-MY, GL-MY, SAJ-MY, and QLD-AU (12.8-21.6-fold, moderate performance), BP-MY, and KL-MY (48.2-49-fold, poor performance), CH-MY (128.2-fold, very poor performance). On the other hand, Tandem displayed high performance against all C. hemipterus strains (1.8-8.3-fold). Tandem caused faster mortality than Temprid SC for all strains. Temprid SC and Tandem residues killed C. hemipterus significantly faster on glass than filter paper. Compared with fresh residues, the efficacy of Temprid SC residues significantly declined after one week of aging, while the effectiveness of Tandem residues declined after two weeks of aging. Further investigations using the topical assay method with a diagnostic dose of imidacloprid found two strains (CH-MY and GL-MY) resistant to imidacloprid. The six other strains (BM-MY, BP-MY, KL-MY, SAJ-MY, TT-MY, and QLD-AU) were susceptible.
  3. Kim DY, Billen J, Doggett SL, Lee CY
    J Econ Entomol, 2017 Jun 01;110(3):1179-1186.
    PMID: 28334370 DOI: 10.1093/jee/tox039
    The climbing abilities of two bed bug species, Cimex lectularius L. and Cimex hemipterus (F.), were determined by evaluating their escape rates from smooth surface pitfall traps using four commercial bed bug monitors (Verifi Bed Bug Detector, ClimbUp Insect Interceptor, BlackOut Bed Bug Detector, and SenSci Volcano Bed Bug Detector). All detectors were used in the absence of lures or attractants. Unlike C. lectularius, adult C. hemipterus were able to escape from all traps. On the other hand, no or a low number nymphs of both species escaped, depending on the evaluated traps. Examination of the vertical friction force of adults of both species revealed a higher vertical friction force in C. hemipterus than in C. lectularius. Scanning electron microscope micrograph observation on the tibial pad of adult bed bugs of C. hemipterus showed the presence of a greater number of tenent hairs on the tibial pad than on that of adult C. lectularius. No tibial pad was found on the fourth and fifth instars of both species. Near the base of the hollow tenent hairs is a glandular epithelium that is better developed in adult C. hemipterus than in adult C. lectularius. This study highlights significant morphological differences between C. lectularius and C. hemipterus, which may have implications in the monitoring and management of bed bug infestations.
  4. Dang K, Doggett SL, Leong XY, Veera Singham G, Lee CY
    J Econ Entomol, 2021 12 06;114(6):2473-2484.
    PMID: 34693975 DOI: 10.1093/jee/toab205
    The modern resurgence of the common (Cimex lectularius L.) and tropical bed bugs (C. hemipterus [F.]) is thought to be primarily due to insecticide resistance. While there are many reports on insecticide resistance mechanisms in C. lectularius, such information in C. hemipterus is limited. We examined dichloro-diphenyl-trichloroethane (DDT), malathion, deltamethrin, permethrin, lambda-cyhalothrin resistance, and the underlying mechanisms in several C. hemipterus strains (Australia: Queensland [QLD-AU]; Malaysia: Kuala Lumpur [KL-MY], Tanjung Tokong [TT-MY], Christian [CH-MY], and Green Lane [GL-MY]). We used a surface contact method, synergism studies (utilizing piperonyl butoxide [PBO], S,S,S-tributyl phosphorotrithioate [DEF], and diethyl maleate [DEM]), and molecular detection of kdr mutations. Results demonstrated that all C. hemipterus strains possessed high resistance to DDT and the pyrethroids and moderate to high resistance to malathion. Synergism studies showed that deltamethrin resistance in all strains was significantly (P < 0.05) inhibited by PBO. In contrast, deltamethrin resistance was not affected in DEF or DEM. Similar findings were found with lambda-cyhalothrin resistance. Malathion resistance was significantly (P < 0.05) reduced by DEF in all strains. Resistance to DDT was not affected by DEM in all strains. Multiple kdr mutations (M918I, D953G, and L1014F) were detected by molecular analyses. TT-MY strain was found with individuals possessing three kdr mutation combinations; D953G + L1014F (homozygous susceptible: M918), M918I + D953G + L1014F (heterozygous resistant: I918), and M918I + D953G + L1014F (homozygous resistant: I918). Individuals with M918I + D953G + L1014F (homozygous resistant: I918) survived longer on deltamethrin (>12 h) than those (≤1 h) with other combinations. M918I + L1014F mutations most likely conferred super-kdr characteristic toward pyrethroids and DDT in C. hemipterus.
  5. Khalid MF, Lee CY, Doggett SL, Veera Singham G
    PLoS One, 2019;14(6):e0218343.
    PMID: 31206537 DOI: 10.1371/journal.pone.0218343
    Many insect species display daily variation of sensitivity to insecticides when they are exposed to the same concentration at different times during the day. To date, this has not been investigated in bed bugs. To address this, we explored circadian rhythms in insecticide susceptibility, xenobiotic metabolizing (XM) gene expressions, and metabolic detoxification in the common bed bug, Cimex lectularius. An insecticide susceptible Monheim strain of C. lectularius was most tolerant of deltamethrin during the late photophase at ZT9 (i.e. nine hours after light is present in the light-dark cycle (LD) cycle) and similarly repeated at CT9 (i.e. nine hours into the subjective day in constant darkness (DD)) suggesting endogenous circadian involvement in susceptibility to deltamethrin. No diel rhythm was observed against imidacloprid insecticide despite significant daily susceptibility in both LD and DD conditions. Rhythmic expressions of metabolic detoxification genes, GSTs1 and CYP397A1 displayed similar expression patterns with total GST and P450 enzyme activities in LD and DD conditions, respectively. The oscillation of mRNA levels of GSTs1 and CYP397A1 was found consistent with peak phases of deltamethrin susceptibility in C. lectularius. This study demonstrates that circadian patterns of metabolic detoxification gene expression occur within C. lectularius. As a consequence, insecticide efficacy can vary dramatically throughout a 24 hour period.
  6. Dang K, Doggett SL, Veera Singham G, Lee CY
    Parasit Vectors, 2017 Jun 29;10(1):318.
    PMID: 28662724 DOI: 10.1186/s13071-017-2232-3
    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.
  7. Yong HH, Hamann SL, Borland R, Fong GT, Omar M, ITC-SEA project team
    Soc Sci Med, 2009 Oct;69(7):1025-31.
    PMID: 19695758 DOI: 10.1016/j.socscimed.2009.07.042
    In recent years, attempts have been made to incorporate religion into tobacco control efforts, especially in countries like Malaysia and Thailand where religion is central to the lives of people. This paper is a prospective examination of the perceived relevance and role of religion and religious authorities in influencing smoking behaviour among Muslims in Malaysia and Buddhists in Thailand. Data were collected from 1482 Muslim Malaysian and 1971 Buddhist Thai adult smokers who completed wave 1 (early 2005) of the International Tobacco Control Southeast Asia Survey (ITC-SEA). Respondents were asked about the role of religion and religious leadership on smoking at Wave 1 and among those recontacted, quitting activity at Wave 2. Results revealed that over 90% of both religious groups reported that their religion guides their day-to-day behaviour at least sometimes, but Malaysian Muslims were more likely to report that this was always the case. The majority (79% Muslims and 88% Buddhists) of both groups believed that their religion discourages smoking. About 61% of the Muslims and 58% of the Buddhists reported that their religious leaders had encouraged them to quit before and a minority (30% and 26%, respectively) said they would be an influential source to motivate them to quit. Logistic regression models suggest that these religious factors had a clear independent association with making quitting attempts in both countries and this translated to success for Malaysian Muslims but not for the Thai Buddhists. Taken together, results from this study indicate that religion and religious authorities are both relevant and important drivers of quitting, but whether this is always enough to guarantee success is less clear. Religion can be a culturally relevant vehicle to complement other tobacco control efforts.
  8. Schwartz TM, Hillis SL, Sridharan R, Lukyanchenko O, Geiser W, Whitman GJ, et al.
    J Med Imaging (Bellingham), 2020 Mar;7(2):022408.
    PMID: 32042859 DOI: 10.1117/1.JMI.7.2.022408
    Purpose: Computer-aided detection (CAD) alerts radiologists to findings potentially associated with breast cancer but is notorious for creating false-positive marks. Although a previous paper found that radiologists took more time to interpret mammograms with more CAD marks, our impression was that this was not true in actual interpretation. We hypothesized that radiologists would selectively disregard these marks when present in larger numbers. Approach: We performed a retrospective review of bilateral digital screening mammograms. We use a mixed linear regression model to assess the relationship between number of CAD marks and ln (interpretation time) after adjustment for covariates. Both readers and mammograms were treated as random sampling units. Results: Ten radiologists, with median experience after residency of 12.5 years (range 6 to 24) interpreted 1832 mammograms. After accounting for number of images, Breast Imaging Reporting and Data System category, and breast density, the number of CAD marks was positively associated with longer interpretation time, with each additional CAD mark proportionally increasing median interpretation time by 4.35% for a typical reader. Conclusions: We found no support for our hypothesis that radiologists will selectively disregard CAD marks when they are present in larger numbers.
  9. Li M, Mathai A, Lau SLH, Yam JW, Xu X, Wang X
    Sensors (Basel), 2021 Jan 05;21(1).
    PMID: 33466530 DOI: 10.3390/s21010313
    Due to medium scattering, absorption, and complex light interactions, capturing objects from the underwater environment has always been a difficult task. Single-pixel imaging (SPI) is an efficient imaging approach that can obtain spatial object information under low-light conditions. In this paper, we propose a single-pixel object inspection system for the underwater environment based on compressive sensing super-resolution convolutional neural network (CS-SRCNN). With the CS-SRCNN algorithm, image reconstruction can be achieved with 30% of the total pixels in the image. We also investigate the impact of compression ratios on underwater object SPI reconstruction performance. In addition, we analyzed the effect of peak signal to noise ratio (PSNR) and structural similarity index (SSIM) to determine the image quality of the reconstructed image. Our work is compared to the SPI system and SRCNN method to demonstrate its efficiency in capturing object results from an underwater environment. The PSNR and SSIM of the proposed method have increased to 35.44% and 73.07%, respectively. This work provides new insight into SPI applications and creates a better alternative for underwater optical object imaging to achieve good quality.
  10. Dang K, Singham GV, Doggett SL, Lilly DG, Lee CY
    J Econ Entomol, 2017 04 01;110(2):558-566.
    PMID: 28115498 DOI: 10.1093/jee/tow296
    The performance of five insecticides (bendiocarb, deltamethrin, DDT, malathion, and imidacloprid) using three application methods (oil-based insecticide films on filter paper, and acetone-based insecticide deposits on two substrates: filter paper and glass) was assessed against a susceptible strain of Cimex lectularius (L.) and two resistant strains of Cimex hemipterus (F.). Substrate type significantly affected (P 
  11. Leong XY, Kim DY, Dang K, Singham GV, Doggett SL, Lee CY
    J Econ Entomol, 2020 02 08;113(1):353-366.
    PMID: 31586445 DOI: 10.1093/jee/toz266
    This study examined the presence of insecticide resistance in different developmental stages (adults, first instars, and eggs) of the tropical bed bug, Cimex hemipterus (F.) using several insecticide formulations. Adults and first instars of five strains (Queensland, Kuala Lumpur, Bukit Mertajam, Saujana, and Krystal Point) were evaluated using the surface contact method and compared with a susceptible strain (Monheim) of the common bed bug Cimex lectularius L. The insecticide formulations were used at their label rates in this study: Tandem (thiamethoxam [11.6%], lambda-cyhalothrin [3.5%]) at 183.96 mg/m2; Temprid SC (imidacloprid [21%], beta-cyfluthrin [10.5%]) at 106.13 mg/m2; Sumithion 20CS (fenitrothion [20%]) at 250 mg/m2; Pesguard FG161 (d-tetramethrin [4.4%], cyphenothrin [13.2%]) at 110 mg/m2; and Sumithrin 10SEC (d-phenothrin [10%]) at 100 mg/m2. Results showed a very high level of resistance to Pesguard FG161 (388.3 to >605.0 times) and Sumithrin (302.9 to >365.5 times) in all adults of the strains tested, whereas low to high levels of resistance were registered for Tandem (1.4-4.7 times), Temprid (7.3-16.7 times), and Sumithion (1.2-14.6 times) for adults of all bed bug strains. For first instars, resistance to the former two formulations were high to very high (31.4-118.1 times). In contrast, they showed lower resistance to Tandem, Temprid, and Sumithion (1.0-10.2 times). An immersion method used to test on bed bug eggs found high to very high resistance toward all tested formulations. Results demonstrate that the resistance level varies between bed bug developmental stages.
  12. Shekhawat KS, Chauhan A, Sakthidevi S, Nimbeni B, Golai S, Stephen L
    Indian J Dent Res, 2020 8 10;31(3):354-357.
    PMID: 32769266 DOI: 10.4103/ijdr.IJDR_352_18
    Background: Work-related musculoskeletal pain (MSPs) is not uncommon among dentist and often limits their work efficiency impacting their quality of life.

    Aim: The present research was conducted to identify site-specific pain resulting from musculoskeletal disorders (MSDs) among practicing dentists and determine its impact on their quality of life.

    Setting and Design: A cross-sectional questionnaire study conducted among practicing dentists of Puducherry Taluk, Puducherry, India.

    Method and Materials: A closed-ended, self-administered questionnaire was distributed to 95 practicing dentists to identify site-specific MSP from the study subjects. Data on pain due to MSDs, frequency of pain, its impact on quality of life, relieving factors, patients attended per day, working hours per day, and awareness on ergonomics were also recorded.

    Statistical Analyses: The data were analyzed for descriptive statistics, and Chi-square tests was used for proportions.

    Results: Almost all respondents experienced pain due to MSDs. Approximately, 11.1% "always" experienced elbow pain; 5.6% "always" experienced pain in neck and back. Approximately, 83% "sometimes" experienced pain in the back. Pain in elbow was significantly associated with gender (P = 0.036), qualification (P = 0.029), and years of practice (P = 0.032). Approximately, 36% reported having an impact on their life.

    Conclusion: The magnitude of the problem is slowly shifting from "sometimes" to "always." Although small in proportion, pain due to MSDs has an impact on dental practitioners' quality of life, and elbow pain was reportedly higher in the study setting. Measures need to be implemented before MSD becomes a career limiting occupational hazard.

  13. Schutze MK, Krosch MN, Armstrong KF, Chapman TA, Englezou A, Chomič A, et al.
    BMC Evol. Biol., 2012;12:130.
    PMID: 22846393
    Bactrocera dorsalis s.s. is a pestiferous tephritid fruit fly distributed from Pakistan to the Pacific, with the Thai/Malay peninsula its southern limit. Sister pest taxa, B. papayae and B. philippinensis, occur in the southeast Asian archipelago and the Philippines, respectively. The relationship among these species is unclear due to their high molecular and morphological similarity. This study analysed population structure of these three species within a southeast Asian biogeographical context to assess potential dispersal patterns and the validity of their current taxonomic status.
  14. Takeda K, Yamauchi J, Miki A, Kim D, Leong XY, Doggett SL, et al.
    Sci Rep, 2019 04 24;9(1):6500.
    PMID: 31019205 DOI: 10.1038/s41598-019-42844-0
    The insect male accessory gland (MAG) is an internal reproductive organ responsible for the synthesis and secretion of seminal fluid components, which play a pivotal role in the male reproductive strategy. In many species of insects, the effective ejaculation of the MAG products is essential for male reproduction. For this purpose, the fruit fly Drosophila has evolved binucleation in the MAG cells, which causes high plasticity of the glandular epithelium, leading to an increase in the volume of seminal fluid that is ejaculated. However, such a binucleation strategy has only been sporadically observed in Dipteran insects, including fruit flies. Here, we report the discovery of binucleation in the MAG of the common bed bug, Cimex lectularius, which belongs to hemimetabolous Hemiptera phylogenetically distant from holometabolous Diptera. In Cimex, the cell morphology and timing of synchrony during binucleation are quite different from those of Drosophila. Additionally, in Drosophila, the position of the two nuclei in the adult stage changes as a result of the mating history or the nutrient conditions; however, it remains stable in Cimex. These differences suggest that binucleation in the Cimex MAG plays a unique role in the male reproductive system that is distinct from that of Drosophila.
  15. Dang K, Toi CS, Lilly DG, Lee CY, Naylor R, Tawatsin A, et al.
    Pest Manag Sci, 2015 Jul;71(7):1015-20.
    PMID: 25132449 DOI: 10.1002/ps.3880
    Bed bugs [both Cimex hemipterus (F.) and Cimex lectularius L.] are highly resistant to pyrethroids worldwide. An important resistance mechanism known as 'knockdown resistance' (kdr) is caused by genetic point mutations on the voltage-gated sodium channel (VGSC) gene. Previous studies have identified two point mutations (V419L and L925I) on the VGSC gene in C. lectularius that are responsible for kdr-type resistance. However, the kdr mutations in C. hemipterus have not been investigated.
  16. Xing J, Watkins WS, Witherspoon DJ, Zhang Y, Guthery SL, Thara R, et al.
    Genome Res, 2009 May;19(5):815-25.
    PMID: 19411602 DOI: 10.1101/gr.085589.108
    We report an analysis of more than 240,000 loci genotyped using the Affymetrix SNP microarray in 554 individuals from 27 worldwide populations in Africa, Asia, and Europe. To provide a more extensive and complete sampling of human genetic variation, we have included caste and tribal samples from two states in South India, Daghestanis from eastern Europe, and the Iban from Malaysia. Consistent with observations made by Charles Darwin, our results highlight shared variation among human populations and demonstrate that much genetic variation is geographically continuous. At the same time, principal components analyses reveal discernible genetic differentiation among almost all identified populations in our sample, and in most cases, individuals can be clearly assigned to defined populations on the basis of SNP genotypes. All individuals are accurately classified into continental groups using a model-based clustering algorithm, but between closely related populations, genetic and self-classifications conflict for some individuals. The 250K data permitted high-level resolution of genetic variation among Indian caste and tribal populations and between highland and lowland Daghestani populations. In particular, upper-caste individuals from Tamil Nadu and Andhra Pradesh form one defined group, lower-caste individuals from these two states form another, and the tribal Irula samples form a third. Our results emphasize the correlation of genetic and geographic distances and highlight other elements, including social factors that have contributed to population structure.
  17. Leong XY, Lee CY, Veera Singham G, Chong Shu-Chien A, Naylor R, Naylor A, et al.
    J Econ Entomol, 2023 Feb 10;116(1):19-28.
    PMID: 35640206 DOI: 10.1093/jee/toac067
    Modern bed bugs are resistant to multiple insecticide classes, particularly the pyrethroids. The efficacy of pyrethroid-impregnated mattress liners marketed for bed bug management has been variable. This study evaluated the efficacy of a permethrin-impregnated mattress liner, ActiveGuard, against 24 bed bug strains, consisting of both Cimex hemipterus (F.) and Cimex lectularius L. A 'mat assay', employing an allethrin-impregnated mat, was used to establish the pyrethroid resistance profile of all strains. Three experiments were conducted to evaluate the effect of ActiveGuard exposure on bed bug knockdown: 1) exposing the bed bugs continuously on the liner for up to 24 d, 2) holding the bed bugs on the liner for either 4 or 6 h, and 3) placing a noninsecticide treated fabric above the liner with the bed bugs held continuously on top. Our results indicated that all modern strains (collected within the last 15 years during the current resurgence) were pyrethroid-resistant, although the magnitude of resistance was highly variable between strains. In the continuous exposure study, an incomplete knockdown was recorded for most modern bed bug strains, with some having no knockdown even up to 7 d of constant exposure. In the 4 or 6 h exposure study, the level of knockdown was reduced even further, and very few bed bugs were knocked down in the double fabric study. The results of this study indicate that pyrethroid-impregnated mattress liners are not likely to be effective in the management of most modern bed bug infestations involving either C. hemipterus or C. lectularius.
  18. Burton AC, Beirne C, Gaynor KM, Sun C, Granados A, Allen ML, et al.
    Nat Ecol Evol, 2024 May;8(5):924-935.
    PMID: 38499871 DOI: 10.1038/s41559-024-02363-2
    Wildlife must adapt to human presence to survive in the Anthropocene, so it is critical to understand species responses to humans in different contexts. We used camera trapping as a lens to view mammal responses to changes in human activity during the COVID-19 pandemic. Across 163 species sampled in 102 projects around the world, changes in the amount and timing of animal activity varied widely. Under higher human activity, mammals were less active in undeveloped areas but unexpectedly more active in developed areas while exhibiting greater nocturnality. Carnivores were most sensitive, showing the strongest decreases in activity and greatest increases in nocturnality. Wildlife managers must consider how habituation and uneven sensitivity across species may cause fundamental differences in human-wildlife interactions along gradients of human influence.
  19. Klionsky DJ, Abdel-Aziz AK, Abdelfatah S, Abdellatif M, Abdoli A, Abel S, et al.
    Autophagy, 2021 Jan;17(1):1-382.
    PMID: 33634751 DOI: 10.1080/15548627.2020.1797280
    In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
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