Development of MALDI-TOF mass spectrometry for the identification of lice isolated from farm animals.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is now routinely used for the rapid identification of microorganisms isolated from clinical samples and has been recently successfully applied to the identification of arthropods. In the present study, this proteomics tool was used to identify lice collected from livestock and poultry in Algeria. The MALDI-TOF MS spectra of 408 adult specimens were measured for 14 species, including Bovicola bovis, B. ovis, B. caprae, Haematopinus eurysternus, Linognathus africanus, L. vituli, Solenopotes capillatus, Menacanthus stramineus, Menopon gallinae, Chelopistes meleagridis, Goniocotes gallinae, Goniodes gigas, Lipeurus caponis and laboratory reared Pediculus humanus corporis. Good quality spectra were obtained for 305 samples. Spectral analysis revealed intra-species reproducibility and inter-species specificity that were consistent with the morphological classification. A blind test of 248 specimens was performed against the in-lab database upgraded with new spectra and validated using molecular tools. With identification percentages ranging from 76% to 100% alongside high identification scores (mean = 2.115), this study proposes MALDI-TOF MS as an effective tool for discriminating lice species.

TITLE: Développement de la spectrométrie de masse MALDI-TOF MS pour l'identification de poux isolés d'animaux de ferme. ABSTRACT: La Spectrométrie de Masse à Temps de Vol par Désorption/Ionisation Laser Assistée après Matrice est maintenant utilisée pour l'identification rapide des microorganismes isolés à partir d'échantillons cliniques et a récemment été appliquée avec succès pour l'identification des arthropodes. Dans cette étude, cet outil protéomique a été utilisé pour identifier les poux prélevés sur le bétail et la volaille en Algérie. Les spectres MALDI-TOF MS de 408 spécimens adultes ont été mesurés pour 14 espèces, dont Bovicola bovis, B. ovis, B. caprae, Haematopinus eurysternus, Linognathus africanus, L. vituli, Solenopotes capillatus, Menacanthus stramineus, Menopon gallinae, Chelopistes meleagridis, Goniocotes gallinae, Goniodes gigas, Lipeurus caponis et Pediculus humanus corporis élevé en laboratoire. Des spectres de bonne qualité ont été obtenus pour 305 échantillons. L'analyse spectrale a révélé une reproductibilité intra-espèce et une spécificité inter-espèces qui concordaient avec la classification morphologique. Un test à l'aveugle de 248 échantillons a été effectué par rapport à la base de données de notre laboratoire mise à niveau avec de nouveaux spectres et validée à l'aide d'outils moléculaires. Avec des pourcentages d'identification allant de 76 à 100 % et des scores d'identification élevés (moyenne : 2,115), cette étude propose MALDI-TOF MS comme un outil efficace pour distinguer les espèces de poux.

Lip b 1 is a novel allergenic protein isolated from the booklouse, Liposcelis bostrychophila.

BACKGROUND: Booklice, belonging to the order Psocoptera, are small household insect pests that are distributed worldwide. Liposcelis bostrychophila, a common home-inhabiting species of booklouse, infests old books, sheets of paper, and stored food. Recent entomological and serological studies demonstrated that L. bostrychophila accounted for the majority of detectable insects in house dust and could be a potent inducer of respiratory allergy. Our recent proteomic analysis identified a potent allergenic protein from L. bostrychophila, designated Lip b 1, and determined its partial amino acid sequences. METHODS: Cloning of cDNAs for Lip b 1 was performed by large-scale transcriptome analysis (RNA-seq) and subsequent reverse transcription polymerase chain reaction. The full-length amino acid sequences deduced from Lip b 1 cDNAs were bioinformatically analyzed. The recombinant proteins of glutathione S-transferase (GST)-fused Lip b 1 were analyzed by Western blot and enzyme-linked immunosorbent assay. RESULTS: Lip b 1 cDNAs encoding two types of 254-amino acid proteins were cloned. The clones shared 87% identity, and the deduced molecular weights and isoelectric points were consistent with those determined in our previous study. The two types of Lip b 1 proteins in the GST-fused form were similarly reactive with sera from allergic patients sensitized with L. bostrychophila. CONCLUSIONS: Lip b 1 is a novel protein possibly causing booklouse allergy.

The effects of insect extracts and some insect-derived compounds on the settling behavior of Liposcelis bostrychophila.

Extracts of whole booklice (Liposcelis bostrychophila)-sequentially extracted in hexane and aqueous 80% methanol (80%MeOH)-repel conspecifics. A methanol-soluble fraction (MFr) of the 80% methanol extract was more repellent than either its corresponding water fraction (WFr) or the hexane extract. The repellent effect of the MFr was repeatable across extracts prepared on different occasions over a 1 month period. Gas chromatography, mass-spectrometry (GC-MS) analyses showed that saturated (C(16); C(18)) monoenoic (C(16:1); C(18:1)) and a dienoic fatty acid (C(18:2)) and the corresponding methyl esters of all but C(16:1) and C(18) constituted approximately 95% and 30%, of the detected compounds in the methanol fractions and the hexane extract, respectively. Qualitative thin layer chromatography showed that cholesterol was present in methanol fractions and the hexane extract, and also enabled tentative identification of triacylglycerols and phospholipids in the methanol fractions. Extracts of wheatgerm, dried skimmed milk powder, active yeast, and wholemeal flour-L. bostrychophila dietary components-were analyzed by GC-MS, and C(16), C(18:1) and C(18:2) were detected, indicating that C(18) and the methyl esters were not directly extractable and/or that they were products of booklice metabolism. A fatty acid amide (stearamide) previously identified in cuticular extracts of L. bostrychophila was not detected, and therefore was not responsible for the observed biological activity. Pure fatty acids and fatty acid methyl esters repelled settling of L. bostrychophila at 10 mM, with the exception of palmitic and stearic acids, indicating, among other things, a difference between the efficacy of saturated and unsaturated fatty acids. The effect of concentrations <10 mM was less significant, although palmiteoleic acid appeared to be attractive to L. bostrychophila at 0.1 mM. Fatty acids and fatty acid methyl esters were at a much lower concentration than 10 mM in the repellent methanol fractions, indicating that an interaction between known and as yet unidentified compounds is likely. The significance of fatty acids in relation to the biology and behavior of L. bostrychophila and their potential for use in traps and monitoring are discussed.

The neglected saliva: medically important toxins in the saliva of human lice.

Although there has been a great deal of research effort within the last two decades on identifying the active components of the saliva of blood-sucking ticks, mosquitoes, biting flies, fleas and bugs, essentially neglected have been the human lice. Despite initial reports in the early part of this century suggestive of vasodilatory, anticoagulant and immunosuppressive properties of the saliva, for the next 50 years there were no biochemical studies on the active principles. Very recently, anatomical and biochemical studies have begun to characterize the bioactive molecules in lice saliva. The louse stocks a salivary vasodilator in excess over what is needed for a single bite, and injects similar amounts at each successive bite. The vasodilator in lice saliva appears to have different pharmacological properties than peroxidative, oxidative and maxidilan types of vasodilators reported from other blood-sucking insects. Possible anticoagulant activities have also been characterized. This belated, but welcome, interest comes at a time of resurgence of lice-born disease in certain parts of Africa, and of resistance to chemical control in Europe and North America.

Toxins produced by arthropod parasites: salivary gland proteins of human body lice and venom proteins of chelonine wasps.

A review is presented of our ongoing research projects on the protein components of the saliva of human body lice and of the non-paralyzing venom of wasps in the subfamily Cheloninae. Sodium dodecyl sulfate-polyacryamide gel electrophoretic analysis of lice salivary gland proteins showed a predominance of high and intermediate mol. wt proteins. Immunoblotting with a low titer polyclonal antiserum to lice salivary proteins indicated that some, but not all, of the predominant high mol. wt salivary gland proteins are injected into the host during feeding. The venom of a Chelonus sp. wasp contains a chitinase, and a 33,000 mol. wt protein with a primary structure composed mostly of a series of 12 tandem repeats of a 14-residue sequence. The N-terminus of this protein and its homologs in a related species of Ascogaster share a conserved adjacent pair of acidic residues. Epitope mapping/immunoprecipitation experiments now in progress will provide information on which linear motifs are on the surface of the protein, and will thereby provide information on the tertiary structure of the protein.

Analysis of the electromobility of different protein fractions of Bovicola bovis females (Mallophaga: Insecta).

The authors characterize in females of Bovicola bovis the total proteins of this species by means of SDS-page determining the R(mb), Rx, Rf and mW of each protein in particular and studying finally the number of protein bands in function with their mW.