Cuticular extracts induce aggregation in head lice.

The human head lice Pediculus humanus capitis (De Geer) (Phthiraptera: Pediculidae) are strict, obligate human ectoparasites that spends their entire life cycle in the host and cause skin irritation and derived infections. Despite the health-related importance, few studies have evaluated the chemical communication among these insects. Here, we evaluate the response of lice of both sexes to cuticular extracts using two solvents of different polarity (hexane and methanol). Cuticular extracts that elicited an attraction response towards head lice were analysed by gas chromatography-mass spectrometry (GC-MS) to determine the cuticular lipid profile. Both lice sexes were attracted to the hexane extracts but not the methanol extracts, suggesting the non-polarity of the compounds present in the cuticle. Chemical analyses of hexane extracts from males and females showed high similarity in major compounds. This study provides the first evidence that lice respond to cuticle extracts, which may be important to understand aggregation behaviour.

Biophysical properties of human body louse nit related proteins: LNSP1, Agp9 and Agp22.

The human parasitic head and body lice lay their eggs on either hair or clothing. Attachments of the eggs are possible because the female lice secret a glue substance from the accessory gland along with the egg, which hardens into a nit sheath that secures and protects the egg (The "nit" commonly refers to either the louse egg with an embryo or the empty hatched egg). Proteins called the louse nit sheath protein (LNSP) are suggested to be the major proteins of the nit sheath, but transcriptome profiling of the accessory glands indicated other proteins such as Agp9 and Agp22 are also expressed in the glands. In this study, human body louse LNSP1 (partial), Agp9, and Agp22 are recombinantly produced using the E. coli expression system, and the biophysical properties characterized. Circular dichroism analysis indicated that the secondary structure elements of LNSP1 N-terminal and middle-domains, Agp9, and Agp22 are prominently random coiled with up to 10-30% anti-parallel ß-sheet element present. Size-exclusion chromatography profiles of LNSP1 proteins further suggested that the ß-sheets made of the smaller N-terminal domain stacks onto the ß-sheets of the larger middle-domain.

Characterization of nit sheath protein functions and transglutaminase-mediated cross-linking in the human head louse, Pediculus humanus capitis.

BACKGROUND: Head louse females secrete liquid glue during oviposition, which is solidified to form the nit sheath over the egg. Recently, two homologous proteins, named louse nit sheath protein (LNSP) 1 and LNSP 2, were identified as adhesive proteins but the precise mechanism of nit sheath solidification is unknown. METHODS: We determined the temporal transcriptome profiles of the head louse accessory glands plus oviduct, from which putative major structural proteins and those with functional importance were deduced. A series of RNA interference (RNAi) experiments and treatment of an inhibitor were conducted to elucidate the function and action mechanism of each component. RESULTS: By transcriptome profiling of genes expressed in the louse accessory glands plus uterus, the LNSP1 and LNSP2 along with two hypothetical proteins were confirmed to be the major structural proteins. In addition, several proteins with functional importance, including transglutaminase (TG), defensin 1 and defensin 2, were identified. When LNSP1 was knocked down via RNA interference, most eggs became nonviable via desiccation, suggesting its role in desiccation resistance. Knockdown of LNSP2, however, resulted in oviposition failure, which suggests that LNSP2 may serve as the basic platform to form the nit sheath and may have an additional function of lubrication. Knockdown of TG also impaired egg hatching, demonstrating its role in the cross-linking of nit sheath proteins. The role of TG in cross-linking was further confirmed by injecting or hair coating of GGsTop, a TG inhibitor. CONCLUSIONS: Both LNSP1 and LNSP2 are essential for maintaining egg viability besides their function as glue. The TG-mediated cross-linking plays critical roles in water preservation that are essential for ensuring normal embryogenesis. TG-mediated cross-linking mechanism can be employed as a therapeutic target to control human louse eggs, and any topically applied TG inhibitors can be exploited as potential ovicidal agents.

Head Louse Feces: Chemical Analysis and Behavioral Activity.

Human head lice Pediculus humanus capitis (De Geer) (Phthiraptera: Pediculidae) are insect parasites closely associated with humans, feeding on the blood of their hosts and causing them skin irritation and probable secondary infections. Despite being a severe nuisance, very few studies have reported on intraspecific chemical communication in head lice. Here, we evaluated the attractive response of head lice to the volatile compounds and solvent extracts from their feces. We also chemically analyzed the main volatile components of these feces and those of the feces' extracts. Head lice were attracted to the methanol extract of their feces but not to the hexane or dichloromethane extracts, suggesting the polar nature of bioactive chemicals present in head louse feces. Follow-up chemical identifications, in fact, showed the presence of hypoxanthine, uric acid, and another purine tentatively identified as either guanine or iso-guanine. Additionally, head lice were significantly attracted by volatiles emitted from samples containing feces. The volatiles emanated from feces alone contained 19 identified substances: 2-pentanone, hexanal, heptanal, 3-methyl-3-buten-1-ol, octanal, sulcatone, nonanal, acetic acid, 2-ethyl-1-hexanol, decanal, 1-octanol, butyric acid, 1-nonanol, hexanoic acid, octanoic acid, 2,6-dimethyl-7-octen-2-ol, 2-undecanone, geranylacetone, and hexadecane. The major compounds found were decanal, nonanal, hexanal, and acetic acid, together representing approximately 60% of the identified compounds. This work represents the first chemical evidence of intraspecies communication among head lice. The results support the existence of active substances present in the feces of P. humanus capitis that may be involved in its aggregation behavior.

Human identification by lice: A Next Generation Sequencing challenge.

Rapid and progressive advances in molecular biology techniques and the advent of Next Generation Sequencing (NGS) have opened new possibilities for analyses also in the identification of entomological matrixes. Insects and other arthropods are widespread in nature and those found at a crime scene can provide a useful contribution to forensic investigations. Entomological evidence is used by experts to define the postmortem interval (PMI), which is essentially based on morphological recognition of the insect and an estimation of its insect life cycle stage. However, molecular genotyping methods can also provide an important support for forensic entomological investigations when the identification of species or human genetic material is required. This case study concerns a collection of insects found in the house of a woman who died from unknown causes. Initially the insects were identified morphologically as belonging to the Pediculidae family, and then, human DNA was extracted and analyzed from their gastrointestinal tract. The application of the latest generation forensic DNA assays, such as the Quantifiler(®) Trio DNA Quantification Kit and the HID-Ion AmpliSeq™ Identity Panel (Applied Biosystems(®)), individuated the presence of human DNA in the samples and determined the genetic profile.

Mechanism of phospho-ubiquitin-induced PARKIN activation.

The E3 ubiquitin ligase PARKIN (encoded by PARK2) and the protein kinase PINK1 (encoded by PARK6) are mutated in autosomal-recessive juvenile Parkinsonism (AR-JP) and work together in the disposal of damaged mitochondria by mitophagy. PINK1 is stabilized on the outside of depolarized mitochondria and phosphorylates polyubiquitin as well as the PARKIN ubiquitin-like (Ubl) domain. These phosphorylation events lead to PARKIN recruitment to mitochondria, and activation by an unknown allosteric mechanism. Here we present the crystal structure of Pediculus humanus PARKIN in complex with Ser65-phosphorylated ubiquitin (phosphoUb), revealing the molecular basis for PARKIN recruitment and activation. The phosphoUb binding site on PARKIN comprises a conserved phosphate pocket and harbours residues mutated in patients with AR-JP. PhosphoUb binding leads to straightening of a helix in the RING1 domain, and the resulting conformational changes release the Ubl domain from the PARKIN core; this activates PARKIN. Moreover, phosphoUb-mediated Ubl release enhances Ubl phosphorylation by PINK1, leading to conformational changes within the Ubl domain and stabilization of an open, active conformation of PARKIN. We redefine the role of the Ubl domain not only as an inhibitory but also as an activating element that is restrained in inactive PARKIN and released by phosphoUb. Our work opens up new avenues to identify small-molecule PARKIN activators.

Lipid and protein composition at different developmental stages of Pediculus capitis (Arthropoda, Phthiraptera).

Protein and lipid compositions were studied at different developmental stages of Pediculus capitis De Geer 1778. Phosphatidylcholine was found to be the predominant lipid at all stages and in both sexes. Palmitic and oleic acids were the main fatty acids throughout the 3 stages studied. A marked decline was observed in the total lipid content and triacylglyceride concentration during development, suggesting that their consumption is an energy source. The electrophoretic mobility revealed the predominance of a 320-kDa protein in eggs and adult females, whereas 2 major proteins of 514 and 439 kDa were found in nymphs, as well as in male and female adults. Two very high density lipoprotein fractions were isolated by ultracentrifugation of egg cytosol in a density gradient of NaBr. Both reserve lipoproteins contained phospholipids and triacylglycerols as the predominant lipids and a protein band of around 320 kDa. The structure of this band is likely to be similar to that found in females in a vitellogenic state.

Histochemical analysis of the nit of Pediculus humanus capitis (Anoplura: Pediculidae).

Histochemical stainings of head louse nits were performed and observations indicated that the nit sheath was nonchitinous. One discernable membrane composed of hyaluronic acid was established that encircled the larvae within the egg. The structural framework of the egg included a definite lattice around each aeropyle of the operculum, through which oxygen exchange occurred.

Molecular composition of the louse sheath.

Flash pyrolysis-gas chromatography/mass spectrometry was used to assess the chemical composition of the head louse's nit sheath. The pyrolyzate of the female insect's secretions, which form a cement-like cylinder holding the egg onto the hair, is dominated by amino acid derivatives and fatty acids. No chitin-specific compounds were detected in the sheath. These results, contrary to previous reports, show that the polymeric complex of the sheath is composed of proteinaceous moieties, possibly cross-linked to aliphatic components. This study constitutes the first chemical characterization of the pyrolysis products of insect (louse) glue and unequivocally confirms that louse sheaths are not chitinous, as suggested by earlier histochemical studies. Development of agents that might loosen nits from the hair shaft is dependent on research that addresses the chemical composition of the nit sheath.

Characterization of body louse midgut proteins recognized by resistant hosts.

The human body louse, Pediculus humanus, showed eighteen midgut proteins ranging between 12 and 117 kDa, when analysed by SDS-PAGE electrophoresis. Seven of them (12 kDa, 17 kDa, 29 kDa, 35 kDa, 40 kDa, 55 kDa and 97 kDa) were major bands based on their intensity of staining. The immunization of rabbits with a midgut extract elicited the production of protective polyclonal antibodies. These antibodies reacted strongly with all major midgut proteins as well as with 63 kDa and 117 kDa proteins when tested by the Western blot technique. The analysis of the proteins revealed that the 12 kDa, 25 kDa, 29 kDa, 35 kDa, 45 kDa, 87 kDa and 97 kDa proteins are glycosylated and none of them contained a lipid moiety. By electroelution, the proteins of 35 kDa and 63 kDa were purified. On trypsinization, the proteins of 35 kDa and 63 kDa produced four major fragments (F1, F2, F3, and F4) when resolved on a 18% SDS-PAGE. The F1 fragment of the 35 kDa protein reacted with the polyclonal antibodies by the immunoblot technique.