Four Species of Under-reported Parasitic Arthropods in Mexico and their Potential Role as Vectors of Pathogens.

Numerous arthropod taxa are important in human and veterinary medicine. The salivary secretions and feces of arthropods can cause allergic reactions in host vertebrates or harbor pathogens. Also, bites can be a risk factor for secondary infections. Documenting the diversity of arthropods of medical and veterinary importance remains an important aspect of disease control and prevention. We provide new records of ectoparasitic arthropods from Mexico that are of potential medical or veterinary relevance. Scanning electron microscopy along with amplification and sequencing of a fragment of the mitochondrial gene (16S rRNA) was used to confirm some species identities. We report the cat louse Felicola subrostratus from cats and the chewing louse Heterodoxus spiniger from dogs, which are common ectoparasites but largely not reported in Mexico. The chigger Eutrombicula alfreddugesi is common on wild lizards (Squamata). For the first time, E. alfreddugesi is reported on Hemidactylus frenatus (common house gecko). This reptile has a close relationship with humans and its chiggers can cause dermatitis (i.e., trombiculiasis) or transmit pathogens. In addition, the common bed bug Cimex lectularius is reported for the first time in the state of Yucatan, an atypical area for its natural distribution. Phylogenetic analysis revealed that Cimex lectularius from Yucatan is closely related to genetic sequences of Cimex lectularius from China. Knowing the regional distribution of arthropods allows the design and implementation of prevention strategies for those that have potential roles as reservoirs or vectors.

[MORPHOLOGICAL DIVERSITY OF FLEAS' STRUCTURES (SIPHONAPTERA) PART 3: GENERAL CHARACTERISTICS AND FEATURES OF THORACIC SETATION].

The paper continues a series of publications (Medvedev, 2015a, b) devoted to the analysis of flea structural features. In the present publication, structural features of flea thoracic setation are analyzed for the first time. Six characters with 53 states in the structure of the chaetom of the pronotum, meso-, and metathoracic. segments are distinguished. The author analyzes the cases of the formation of similar states stipulated by processes of reduction or, by contrast, by strengthening of the chaetom in fleas of separate genera belonging to different superfamilies in relation to the type or peculiarities of parasitism in different flea species. In spite of the presence of strong variability in arrangement and number of thoracic setae, some examples of marking of groups in the range families and superfamilies by certain states of the chaetotaxy of the pronotum and metepimera are revealed.

Surface ultrastructure of the eggs of Malacopsylla grossiventris and Phthiropsylla agenoris (Siphonaptera: Malacopsyllidae).

The fleas of the Malacopsyllidae are known only by their adults. In this study, we describe the eggs of Malacopsylla grossiventris and Phthiropsylla agenoris and compare their surface ultrastructure using scanning electron microscopy. Eggs of both flea species are similar in their general appearance, i.e., the presence of disks and absence of lateral aeropyles, but they differ in size, texture of the surface, and number of micropyles and aeropyles. In addition, the eggs of M. grossiventris and P. agenoris are larger (719-800 µm) than most of the flea eggs known (<600 µm), and the length/width ratio is 2.0, while it varies between 1.6 and 1.8 in other species. As opposed to other large species, malacopsyllids have expandable (telescoping) abdomens that display growth. It is possible that a clutch of eggs has more than 2 eggs. Females of M. grossiventris were observed fixed with their mouthparts to the skin of their hosts. Indeed, some of these specimens were observed copulating on the venter of their hosts. These results contribute to the knowledge about the biology of malacopsyllids.

Jumping mechanisms and performance of snow fleas (Mecoptera, Boreidae).

Flightless snow fleas (snow scorpion flies, Mecoptera, Boreidae) live as adults during northern hemisphere winters, often jumping and walking on the surface of snow. Their jumping mechanisms and performance were analysed with high speed imaging. Jumps were propelled by simultaneous movements of both the middle and hind pairs of legs, as judged by the 0.2 ms resolution afforded by image rates of 5000 frames s(-1). The middle legs of males represent 140% and the hindlegs 187% of the body length (3.4 mm), and the ratio of leg lengths is 1:1.3:1.7 (front:middle:hind). In preparation for a jump the middle legs and hindlegs were rotated forwards at their coxal joints with the fused mesothorax and metathorax. The first propulsive movement of a jump was the rotation of the trochantera about the coxae, powered by large depressor muscles within the thorax. The acceleration time was 6.6 ms. The fastest jump by a male had a take-off velocity of 1 m s(-1), which required 1.1 µJ of energy and a power output of 0.18 mW, and exerted a force about 16 times its body weight. Jump distances of about 100 mm were unaffected by temperature. This, and the power per mass of muscle requirement of 740 W kg(-1), suggests that a catapult mechanism is used. The elastic protein resilin was revealed in four pads at the articulation of the wing hinge with the dorsal head of the pleural ridge of each middle leg and hindleg. By contrast, fleas, which use just their hindlegs for jumping, have only two pads of resilin. This, therefore, provides a functional reference point for considerations about the phylogenetic relationships between snow fleas and true fleas.

[Phenomenon of Yersinia pestis biofilm formation in flea organism].

For the first time substantiated was the role of the phenomenon of Yersinia pestis biofilm--extracellular matrix envelope (EME)--formation as the basis that determines the nature of plague agent interaction with flea organism. Implication of vector's proventriculus in the process of biofilm formation was demonstrated. Ultrastucture of plague microbe conglomerates in flea proventriculus and midgut was analysed and uniform mechanism of their formation was elucidated. The role of Yersinia pestis biofilm in plague agent preservation in the gut of ectoparasites and soil of rodents burrows was discussed. PCR analysis confirmed the presence of plague microbe in plague infected corpses and fleas excrements stored at +8-10 degreesC for 7 years 9 months.

Sand flea (Tunga spp.) infections in humans and domestic animals: state of the art.

Tungiasis is a parasitic disease of humans and animals caused by fleas (Siphonaptera) belonging to the genus Tunga. Two species, Tunga penetrans (L.) and Tunga trimamillata, out of 10 described to date, are known to affect man or domestic animals; the other eight are exclusive to a few species of wild mammals. Tunga penetrans and T. trimamillata originated from Latin America, although the first species is also found in sub-Saharan Africa (between 20 degrees N and 25 degrees S). Hundreds of millions of people are at risk of infection in more than 70 nations, mostly in developing countries. The second species has been reported only in Ecuador and Peru. Males and non-fertilized females of Tunga are haematophagous ectoparasites; pregnant females penetrate the skin where, following dilatation of the abdomen, they increase enormously in size (neosomy) and cause inflammatory and ulcerative processes of varying severity. The importance of Tunga infection in humans concerns its frequent localization in the foot, which sometimes causes very serious difficulty in walking, thereby reducing the subject's ability to work and necessitating medical and surgical intervention. Tungiasis in domestic animals can be responsible for economic losses resulting from flea-induced lesions and secondary infections. Because tungiasis represents a serious problem for tropical public health and because of the recent description of a new species (Tunga trimamillata), it seems appropriate to review current knowledge of the morphology, molecular taxonomy, epidemiology, pathology, treatment and control of sand fleas of the genus Tunga.

Uniquely insidious: Yersinia pestis biofilms.

Bubonic plague, one of history's deadliest infections, is transmitted by fleas infected with Yersinia pestis. The bacteria can starve fleas by blocking their digestive tracts, which stimulates the insects to bite repeatedly and thereby infect new hosts. Direct examination of infected fleas, aided by in vitro studies and experiments with the nematode Caenorhabditis elegans, have established that Y. pestis forms a biofilm in the insect. The extracellular matrix of the biofilm seems to contain a homopolymer of N-acetyl-d-glucosamine, which is a constituent of many bacterial biofilms. A regulatory mechanism involved in Y. pestis biofilm formation, cyclic-di-GMP signaling, is also widespread in bacteria; yet only Y. pestis forms biofilms in fleas. Here, the historical background of bubonic plague is briefly described and recent studies investigating the mechanisms by which these unique and deadly biofilms are formed are discussed.

Investigations on the life cycle and morphology of Tunga penetrans in Brazil.

In the present study, the life cycle of Tunga penetrans was established in Wistar rats in the laboratory, and the morphology of the resulting developmental stages was studied by means of light and scanning electron microscopy. It was seen that the females enter at a nonfertilized stage through the skin of their hosts. Only there the copulation occurs, while females and males brought together in a Petri dish showed no interest in each other. In any way -- fertilized or not -- the females start about 6 days after penetration and hypertrophy with the ejection of eggs. While fertilized eggs proceed to development, the unfertilized ones remain arrested. The eggs are ovoid and measure about 600 x 320 mum. The larvae hatch from the eggs 1-6 days (mean 3-4) after ejection. Formation of larvae 2 took at least another day, while 4 up to 10 days more were needed until this larva starts pupation (mean 5-7 days). The formation of the adult fleas inside the puparium occurred within 9-15 days (with a maximum hatch at day 12). Adult female fleas having reached the skin of a host start blood sucking within 5 min and prepare to enter the skin. After 24 h, the flea stacked already with two thirds of its body inside the skin. After 40 h, the penetration was completed, and feeding and hypertrophical enlargement started, which was completed on day 6, when eggs became ejected. When studying the morphology of the fleas obtained from different hosts, slight variations were seen, which, however, are not significant for a species separation but may be an indication of the presence of different strains/races or the beginning of such a formation.

Simultaneous false-colour imaging of birefringence, extinction and transmittance at camera speed.

A polarized light imaging technique is introduced that simultaneously captures and unfolds transmittance, fast/slow axes directions and birefringence or linear dichroism from single camera exposures. The technique, based on the rotating polarizer method, is capable of sampling an object through polarizers at four different angles in a single image with the use of an image multiplexer. The range of possible applications is demonstrated with examples from the fields of biology, medicine, forensics and material sciences.

Investigations on the biology, epidemiology, pathology and control of Tunga penetrans in Brazil. VI. Natural history of the infestation in laboratory-raised Wistar rats.

Tungiasis is endemic in many countries in Latin America, the Caribbean and sub-Saharan Africa, and it is associated with severe morbidity. The pathophysiological and immunological characteristics of the ectoparasitosis are not well understood, and no effective therapy is currently available. The aim of this study was to describe the natural history of tungiasis in laboratory-raised Wistar rats. The rats were exposed in the laboratory to the parasite or were kept in a natural environment with an intense transmission of Tunga penetrans. The time course of the infestation was determined, and lesions were photographed, described clinically in detail and biopsied. Biopsies were examined histopathologically and by light and scanning electron microscopy. Based on these findings, the natural history of tungiasis in Wistar rats was described and divided in five stages. Our data show that the natural history of tungiasis in Wistar rats and humans is almost identical, except that in the animals, the basement membrane disrupts 5 days after penetration and provokes an intense infiltration of the dermis, while in humans, the basement membrane remains intact. The study indicates that the Wistar rat is an appropriate model for the study of clinical and pathological aspects of tungiasis. Using this model should enable a better understanding of the pathophysiology and immunology of the ectoparasitosis.

Scanning electron microscopy studies of sensilla and other structures of the head of Polygenis (Polygenis) tripus (Siphonapera: Rhopalopsyllidae).

The external morphology of the head and appendages of adult Polygenis (Polygenis) tripus [Jordan, K., 1933. Descriptions of Siphonaptera. Nov. Zool. 39, 66-69] (Siphonaptera: Rhopalopsyllidae) was described based on scanning electron microscopy. Chaetal, trichoid and basiconic sensilla were encountered and their positions noted, comparisons being made between male and female specimens. Those of the basiconic type were particularly abundant on the frontal and occipital regions of the head and were surrounded by several pores, indicating the considerable sensory importance of these areas. Frontal tubercle was observed as a projection emerging from a depression in the frontal head region. The antennae of males were more developed, presenting greater number of sensilla on the scape and about 370 spatulate setae on the internal surface of the clava. Three large chemoreceptors, digitate sensilla ca. 11-13 microm were observed on the lateral surface of the antennal clava of both sexes between antennomeres 4-7. Females present a characteristic distribution of basiconic sensilla on the dorsal margin and median external surface of the clava. Trichoid sensilla were observed on all segments of the maxilary palps while basiconic sensilla were only recognized on the distal three segments.

Field trial of the efficacy of a combination of imidacloprid and permethrin against Tunga penetrans (sand flea, jigger flea) in dogs in Brazil.

In a field trial in Brazil 17 dogs penetrated by females of the jigger flea, Tunga penetrans, were topically treated with a combination of 10% imidacloprid and 50% permethrin (Advantix), while 17 dogs remained untreated. The follow-up controls on days 7, 14, 21 and 28 post-treatment clearly showed that, beginning from day 7, the flea load in treated dogs decreased, so that most of the dogs became free of tungiasis lesions, while in the untreated group the flea load remained high. Since the dogs distribute the flea eggs throughout the village, leading to a high incidence of tungiasis in humans, treatment of dogs probably also decreases the number of cases of tungiasis in the latter.

Transmission of Yersinia pestis from an infectious biofilm in the flea vector.

Transmission of plague by fleas depends on infection of the proventricular valve in the insect's foregut by a dense aggregate of Yersinia pestis. Proventricular infection requires the Y. pestis hemin storage (hms) genes; here, we show that the hms genes are also required to produce an extracellular matrix and a biofilm in vitro, supporting the hypothesis that a transmissible infection in the flea depends on the development of a biofilm on the hydrophobic, acellular surface of spines that line the interior of the proventriculus. The development of biofilm and proventricular infection did not depend on the 3 Y. pestis quorum-sensing systems. The extracellular matrix enveloping the Y. pestis biofilm in the flea appeared to incorporate components from the flea's blood meal, and bacteria released from the biofilm were more resistant to human polymorphonuclear leukocytes than were in vitro-grown Y. pestis. Enabling arthropod-borne transmission represents a novel function of a bacterial biofilm.

Techniques of DNA-studies on prehispanic ectoparasites (Pulex sp., Pulicidae, Siphonaptera) from Animal mummies of the Chiribaya culture, southern Peru.

During a paleoparasitological survey of several animal mummies (Cavia aperea f. porcellus and Canis familiaris) from Chiribaya Baja, an archaeological site in Southern Peru, an unexpected find was made. In the well preserved fur, large numbers of mummified fleas (Pulex simulans/irritans) that parasitized the animals during life were encountered. Due to the relative recent event of the host mummification and the outstanding preservation of the fleas, an attempt for the retrieval of DNA was made. A DNA extraction and sequencing protocol for archaeological ectoparasitic remains has been established, taking additional studies for tissue and protein preservation into account. Tissue preservation was assessed with transmission electron microscopy and the protein preservation was tested through the racemisation ratios of aspartic acid. Regions of the 28S rDNA gene were successfully amplified and sequenced. Further research perspectives are outlined.

Tunga penetrans: molecular identification of Wolbachia endobacteria and their recognition by antibodies against proteins of endobacteria from filarial parasites.

In search of Wolbachia in human parasites, Wolbachia were identified in the sand flea Tunga penetrans. PCR and DNA sequencing of the bacterial 16S rDNA, the ftsZ cell division protein, the Wolbachia surface protein (wsp) and the Wolbachia aspartate aminotransferase genes revealed a high similarity to the respective sequences of endosymbionts of filarial nematodes. Using these sequences a phylogenetic tree was generated, that indicates a close relationship between Wolbachia from T. penetrans and from filarial parasites, but possibly as a member of a new supergroup. Ultrastructural studies showed that Wolbachia are abundant in the ovaries of neosomic fleas, whereas other, smaller and morphologically distinct, bacteria were observed in the lumen of the intestine. Wolbachia were labeled by immunohistology and immunogold electron microscopy using polyclonal antibodies against wsp of Drosophila, of the filarial parasite Dirofilaria immitis, or against hsp 60 from Yersinia enterocolitica. These results show that as in filariasis, humans with tungiasis are exposed to Wolbachia. Furthermore, antisera raised against proteins of Wolbachia from arthropods or from filarial parasites can be immunologically cross-reactive.

[Peculiarities of thoracic and abdominal combs of fleas (Siphonaptera)]. / Osobennosti stroeniia grudnykh i briushnykh ktenidiev blokh (Siphonaptera).

The structure of pseudosetae, spinelets, and spines of combs (ctenidia) was studied by means of light and SE microscopy in 80% of genera and subgenera of the World fauna. It is found out that peculiarities of ctenidiae in the prothorax and in tergites of the abdomen are characteristics of families and infraorders of fleas. Some characters of ctenidiae found in certain flea genera are reductions and apparently caused by habitation in some extremal conditions. An absence of ctenidiae in the unfraorder Pulicomorpha is compensated by more developed posterior margin of prothorax and general abbreviation of all thoracal segments. Reasons of ctenidiae absence, which is observed in certain genera of the infraorders Ceratophyllomorpha, Pygiopsyllomorpha and Hystricopsillomorpha associated with the same hosts, is not clear. It is confirmed, that distance between ctenidiae in different flea species associated with the same species host species, however it is recovered, that this distance correlates with the diameter of most thin hair of host. In some flea species the distance between ctenidia spices in females is larger, than in males. It is found, that sexual dimorphism by this character may not be expressed in certain species of closely related species group of fleas. It is suggested that ctenidiae were present even in the common ancestor of fleas. The hypothesis on origin of spines and pseudosetae from setae of the posterior walls of toracal and abdominal segments in the common ancestor of fleas is proposed.

Comparative study on the effects of three insecticides (fipronil, imidacloprid, selamectin) on developmental stages of the cat flea (Ctenocephalides felis Bouché 1835): a light and electron microscopic analysis of in vivo and in vitro experiments.

The effects of three insecticides (fipronil, imidacloprid and selamectin) on developmental stages of cat fleas (Ctenocephalides felis) were studied in vivo, in vitro and by means of light and electron microscopy. The results were documented by video. Adult fleas were attached to the skin of dogs that had been treated 7 days before with one of the three compounds. Furthermore, adult fleas were exposed exclusively to the hair and skin debris of such treated dogs or were placed on filter papers that had been impregnated with one of these three compounds or with the blood of treated dogs. Larval fleas were exposed to hair of treated dogs, to debris obtained by combing treated dogs, to dried blood samples of treated dogs or were placed onto filter papers impregnated with one of the three compounds. In these experiments with adult and larval fleas, it was noted that none of the three insecticides had a repellent effect on adult or larval fleas. Imidacloprid was the only compound that acted exclusively by body contact, and was apparently taken up by adult and larval fleas via the thin, non-sclerotized intersegmental membranes of the flea's body, shown when flea stages were exposed to hairs taken from dogs treated with one of the compounds or placed onto drug-impregnated filter papers. Imidacloprid killed larvae and adult fleas within 1 h, while it took at least 24 h until all adult fleas had died on fipronil- or selamectin-treated dogs, thus allowing longer feeding periods, increasing the risk of transmission of flea-derived diseases. Flea larvae covered with debris from dogs topically treated 7 days before with fipronil, imidacloprid or selamectin died, like the untreated control, within 16-28 h after exposure. This was, however, probably mainly due to a drying effect. Adult and larval fleas exposed to filter papers impregnated with the blood of treated dogs survived longer than 7 days, as did the untreated controls. All three drugs apparently acted on nerves and muscles and thus stopped motility.

Pulícidos: aspectos adaptativos y biológicos / Pulicides: adaptative and biological aspects

Las pulgas son insectos hematófagos que se caracterizan por una baja especificidad parasitaria. Son ectoparásitos de distintos animales y del hombre. El cuerpo de estos insectos está comprimido lateralmente y las antenas pueden replegarse dentro de un surco a los costados de la cabeza, características óptimas para un mejor desplazamiento entre los pelos y las plumas de los hospedadores. Los tres pares de patas son fuertes y poderosas, armadas con robustas espinas y en sus extremos con un par de grandes uñas en forma de garfios. El tercer par es el que está adaptado para saltar. Las piezas bucales son sucsopicadoras y modificadas en forma de estiletes. Algunas especies tienen unas hileras de cerdas (ctenidios) en la zona genal y/o en el dorso del primer segmento toráxico que evitan los movimientos de retroceso y la caída del cuerpo del hospedador. Esta característica también dificulta la extracción de los parásitos. La importancia de los pulícidos radica entre otras, por ser transmisores de la peste y el tifus. Se muestran bajo SEM y con microfotografías las adaptaciones al parasitismo, la biología y las afecciones cutáneas que se producen