The Centriole Mystique.

Centrioles organize the microtubule network and mitotic spindle and, as basal bodies, nucleate cilia and flagella. They undergo a beguiling process in which one appears to give rise to another and at a baffling orthogonal geometry. Nucleic acid-based replication has been pondered during cycles of zeniths and nadirs of plausibility, the latter now the state. Centrioles can also arise de novo, and thus the longstanding focus on centriole 'replication' may have led us astray from ground truth. We are in an era in which the assembly pathways of most intracellular machines are becoming understood in considerable detail. But apart from knowing the structure and parts list, little in our extant knowledge conveys how centrioles arise. Here the matters at hand are summarized, and a siren call is sounded.

Assessing the Parasitic Burden in a Late Antique Florentine Emergency Burial Site.

Excavation (2008-2014) carried out under the Uffizi Gallery (Florence, Italy) led to the discovery of 75 individuals, mostly buried in multiple graves. Based on Roman minted coins, the graves were preliminarily dated between the second half of the 4th and the beginning of the 5th centuries CE. Taphonomy showed that this was an emergency burial site associated with a catastrophic event, possibly an epidemic of unknown etiology with high mortality rates. In this perspective, paleoparasitological investigations were performed on 18 individuals exhumed from 9 multiple graves to assess the burden of gastrointestinal parasitism. Five out of eighteen individuals (27.7%) tested positive for ascarid-type remains; these are considered as "decorticated" Ascaris eggs, which have lost their outer mammillated coat. Roundworms (genus Ascaris) commonly infest human populations under dire sanitary conditions. Archaeological and historical evidence indicates that Florentia suffered a period of economic crisis between the end of 4th and the beginning of the 5th centuries CE, and that the aqueduct was severely damaged at the beginning of the 4th century CE, possibly during the siege of the Goths (406 CE). It is more than plausible that the epidemic, possibly coupled with the disruption of the aqueduct, deeply affected the living conditions of these individuals. A 27.7% frequency suggests that ascariasis was widespread in this population. This investigation exemplifies how paleoparasitological information can be retrieved from the analysis of sediments sampled in cemeteries, thus allowing a better assessment of the varying frequency of parasitic infections among ancient populations.

Differential Change in the Prevalence of the Ascaris, Trichuris and Clonorchis infection Among Past East Asian Populations.

As we learn more about parasites in ancient civilizations, data becomes available that can be used to see how infection may change over time. The aim of this study is to assess how common certain intestinal parasites were in China and Korea in the past 2000 years, and make comparisons with prevalence data from the 20th century. This allows us to go on to investigate how and why changes in parasite prevalence may have occurred at different times. Here we show that Chinese liver fluke (Clonorchis sinensis) dropped markedly in prevalence in both Korea and China earlier than did roundworm (Ascaris lumbricoides) and whipworm (Trichuris trichiura). We use historical evidence to determine why this was the case, exploring the role of developing sanitation infrastructure, changing use of human feces as crop fertilizer, development of chemical fertilizers, snail control programs, changing dietary preferences, and governmental public health campaigns during the 20th century.

Historical roots of centrosome research: discovery of Boveri's microscope slides in Würzburg.

Boveri's visionary monograph 'Ueber die Natur der Centrosomen' (On the nature of centrosomes) in 1900 was founded primarily on microscopic observations of cleaving eggs of sea urchins and the roundworm parasite Ascaris. As Boveri wrote in the introductory paragraph, his interests were less about morphological aspects of centrosomes, but rather aimed at an understanding of their physiological role during cell division. The remarkable transition from observations of tiny dot-like structures in fixed and sectioned material to a unified theory of centrosome function (which in essence still holds true today) cannot be fully appreciated without examining Boveri's starting material, the histological specimens. It was generally assumed that the microscope slides were lost during the bombing of the Zoological Institute in Würzburg at the end of WWII. Here, I describe the discovery of a number of Boveri's original microscope slides with serial sections of early sea urchin and Ascaris embryos, stained by Heidenhain's iron haematoxylin method. Some slides bear handwritten notes and sketches by Boveri. Evidence is presented that the newly discovered slides are part of the original material used by Boveri for his seminal centrosome monograph.

Reconstitution of amoeboid motility in vitro identifies a motor-independent mechanism for cell body retraction.

Crawling movement in eukaryotic cells requires coordination of leading-edge protrusion with cell body retraction [1-3]. Protrusion is driven by actin polymerization along the leading edge [4]. The mechanism of retraction is less clear; myosin contractility may be involved in some cells [5] but is not essential in others [6-9]. In Ascaris sperm, protrusion and retraction are powered by the major sperm protein (MSP) motility system instead of the conventional actin apparatus [10, 11]. These cells lack motor proteins [12] and so are well suited to explore motor-independent mechanisms of retraction. We reconstituted protrusion and retraction simultaneously in MSP filament meshworks, called fibers, that assemble behind plasma membrane-derived vesicles. Retraction is triggered by depolymerization of complete filaments in the rear of the fiber [13]. The surviving filaments reorganize to maintain their packing density. By packing fewer filaments into a smaller volume, the depolymerizing network shrinks and thereby generates sufficient force to move an attached load. Our work provides direct evidence for motor-independent retraction in the reconstituted MSP motility system of nematode sperm. This mechanism could also apply to actin-based cells and may explain reports of cells that crawl even when their myosin activity is compromised.

Dephosphorylation of major sperm protein (MSP) fiber protein 3 by protein phosphatase 2A during cell body retraction in the MSP-based amoeboid motility of Ascaris sperm.

The crawling movement of nematode sperm requires coordination of leading edge protrusion with cell body retraction, both of which are powered by modulation of a cytoskeleton based on major sperm protein (MSP) filaments. We used a cell-free in vitro motility system in which both protrusion and retraction can be reconstituted, to identify two proteins involved in cell body retraction. Pharmacological and depletion-add back assays showed that retraction was triggered by a putative protein phosphatase 2A (PP2A, a Ser/Thr phosphatase activated by tyrosine dephosphorylation). Immunofluorescence showed that PP2A was present in the cell body and was concentrated at the base of the lamellipod where the force for retraction is generated. PP2A targeted MSP fiber protein 3 (MFP3), a protein unique to nematode sperm that binds to the MSP filaments in the motility apparatus. Dephosphorylation of MFP3 caused its release from the cytoskeleton and generated filament disassembly. Our results suggest that interaction between PP2A and MFP3 leads to local disassembly of the MSP cytoskeleton at the base of the lamellipod in sperm that in turn pulls the trailing cell body forward.

Inactivation of Ascaris suum in a biodrying compost system.

Pathogen contamination of waterways is a serious concern in dairy farming areas where livestock waste is applied to agricultural fields. As an alternative, a biodrying composting system dries collected livestock waste, reduces the strong odors, and has been proposed as a means of reducing, and even eliminating pathogens present in the waste. Therefore, the survival of pathogens in a biodrying composting system was investigated. Dairy farm livestock waste was piled in a biodrying storage shed where forced aeration and natural decomposition processes heated a major portion of the waste pile to temperatures exceeding 55 degrees C. Ascaris suum eggs were used as the surrogate species and inoculated into special chambers and placed at three different elevations at different intervals along the length of the pile. Control chambers were stored in water at 4 degrees C in the laboratory. Both compost and control chambers were removed at Day 4, 8, 12, 16, and 20. The eggs were extracted from the chamber medium and analyzed for viability. No viable eggs were recovered from any of the chambers removed from the compost pile, while >or=90% viability was observed in the control chambers. High temperatures and continued drying were the major contributing factors to the inactivation of the helminth eggs. The complete inactivation of A. suum eggs by the biodrying process encourages the storage and treatment of manure to high temperatures and reduced moisture conditions before field spreading to reduce the risk of harmful pathogens contaminating waterways and potential drinking water supplies.

Structure of MFP2 and its function in enhancing MSP polymerization in Ascaris sperm amoeboid motility.

The simplicity and specialization of the cell motility machinery of Ascaris sperm provides a powerful system in which to probe the basic molecular mechanism of amoeboid cell motility. Although Ascaris sperm locomotion closely resembles that seen in many other types of crawling cell, movement is generated by modulation of a cytoskeleton based on the major sperm protein (MSP) rather than the actin present in other cell types. The Ascaris motility machinery can be studied conveniently in a cell-free in vitro system based on the movement of plasma membrane vesicles by fibres constructed from bundles of MSP filaments. In addition to ATP, MSP and a plasma membrane protein, reconstitution of MSP motility in this cell-free extract requires cytosolic proteins to orchestrate the site-specific assembly and bundling of MSP filaments that generates locomotion. One of these proteins, MFP2, accelerates the rate of movement in this assay. Here, we describe crystal structures of two isoforms of MFP2 and show that both are constructed from two domains that have the same fold based on a novel, compact beta sheet arrangement. Patterns of conservation observed in a structure-based analysis of MFP2 sequences from different nematode species identified regions that may be putative functional interfaces involved both in interactions between MFP2 domains and also with other components of the sperm motility machinery. Analysis of the growth of fibres in vitro in the presence of added MFP2 indicated that MFP2 increases the rate of locomotion by enhancing the effective rate of MSP filament polymerization. This observation, together with the structural data, suggests that MFP2 may function in a manner analogous to formins in actin-based motility.

Electron miscroscopy and histochemical studies on four Egyptian helminthes eggs of medical importance.

By SEM the Fasciola gigantica egg is ovoid with a small knob like operculum, while the egg of Heterophyes heterophytes is broad oval with the operculum more tapering. The egg shell of fertilized Ascaris lumbricoides has interconnected ridges and peak-like projections, while the egg of Enterobius vermicularis is flattened with a thicker margin at the curved side. By TEM, Fasciola egg shell consists of fine reticulum fibrils of three layers. The outer lipoprotein of perivitelline membrane beneath which 2 membranes separated by inclusions, middle of protein globules and inner lipoprotein layer with minute electron-dense granules of melanin or polymer origin, in some parts of the shell giving the egg its brown coloration. The Heterophyes egg shell is more or less similar to that of Fasciola but lacking the minute electron-dense granules. The egg shell of Ascaris has outer ulterine layer with three consecutive layers, basal lipoprotein layer and the inner lipid or ascaroside layer which is the most resistant layer. The Enterobius egg shell consists of five layers, external uterine, internal uterine, vitelline, chitinous and lipid layer. Histochemically, Fasciola egg shell consists of nine amino-acids, and that of Heterophyes consists of ten amino acids. In Ascaris, the lipid layer characteristically consists 25% protein and 75% lipid. The histochemical examination of Enterobius as a detailed example, showed different degrees of reactions with mercuric bromophenol blue, diazotization coupling, Sakaguchi reaction, Sudan black and Mallory's triple stain. Temperature showed marked effect on eggs survival. Eggs of Fasciola and Heterophyes withstand more low temperatures but those of Ascaris and Enterobius withstand more high ones. There are marked correlations between the egg shell constitution, histochemical compositions on one hand and water permeability and egg dryness on the other hand. The results were photographed and discussed.

Modelling and estimation of physical parameters in a sludge drying system.

In this paper is presented the study of a Sludge Drying System used to kill pathogenic organisms living in sludge. The system is modeled and the physical parameters thermal capacity, thermal resistance and thermal time constant are estimated using conventional estimation methods.

Posterior end mark, a novel maternal gene encoding a localized factor in the ascidian embryo.

Ascidian embryogenesis is regarded as a typical 'mosaic' type. Recent studies have provided convincing evidence that components of the posterior-vegetal cytoplasm of fertilized eggs are responsible for establishment of the anteroposterior axis of the embryo. We report here isolation and characterization of a novel maternal gene, posterior end mark (pem). After fertilization, the pem transcript is concentrated in the posterior-vegetal cytoplasm of the egg and later marks the posterior end of developing ascidian embryos. Despite its conspicuous localization pattern, the predicted PEM protein shows no significant homology to known proteins. Overexpression of this gene by microinjection of synthesized pem mRNA into fertilized eggs results in development of tadpole larvae with deficiency of the anteriormost adhesive organ, dorsal brain and sensory pigment-cells. Lineage tracing analysis revealed that the anterior epidermis and dorsal neuronal cells were translocated posteriorly into the tail region, suggesting that this gene plays a role in establishment of anterior and dorsal patterning of the embryo. The ascidian tadpole is regarded as a prototype of vertebrates, implying a similar function of pem in vertebrate embryogenesis.

Reconstitution in vitro of the motile apparatus from the amoeboid sperm of Ascaris shows that filament assembly and bundling move membranes.

We have developed an in vitro motility system from Ascaris sperm, unique amoeboid cells that use filament arrays composed of major sperm protein (MSP) instead of an actin-based apparatus for locomotion. Addition of ATP to sperm extracts induces formation of fibers approximately 2 microns in diameter. These fibers display the key features of the MSP cytoskeleton in vivo. Each fiber consists of a meshwork of MSP filaments and has at one end a vesicle derived from the plasma membrane at the leading edge of the cell. Fiber growth is due to filament assembly at the vesicle; thus, fiber elongation results in vesicle translocation. This in vitro system demonstrates directly that localized polymerization and bundling of filaments can move membranes and provides a powerful assay for evaluating the molecular mechanism of amoeboid cell motility.

Ascariasis en primates no humanos del Perú / Ascariasis in non human primates of Peru

Se señala el segundo caso en el Perú de infección por Ascaris lumbricoides en un primate no humano, Pithecia monachus "guapo negro"

GABA-immunoreactive neurons in the nematode Ascaris.

gamma-Aminobutyric acid (GABA) immunoreactive neurons in the cephalic, somatic, and caudal regions of the Ascaris nervous system were visualized with serial section and whole-mount GABA immunocytochemistry. In the ventral and dorsal nerve cords, GABA-like immunoreactivity (GLIR) is localized to the neurites and cell bodies of identified inhibitory motor neurons and to two fibers, one in each cord, that arise from neurons in the nerve ring. GLIR is absent from identified excitatory motor neurons and from ventral cord interneurons. In neurons containing GLIR, immunoreactivity was present throughout the cell, which argues against an exclusive localization of GABA at conventional synapses. In whole mounts, ten GABA-immunoreactive neurons were present in the cephalic region. These include four nerve ring-associated cells (the RME-like cells), two bilaterally symmetrical pairs of lateral ganglia neurons (the amphid-GABA and deirid-GABA cells) and one bilaterally symmetrical pair of ventral ganglion cells (the VG-GABA cells). In sections, the RME-like cells and the VG-GABA cells were consistently stained through the cephalic region. However, anti-GABA staining of the lateral ganglia cells in sections was light, thus suggesting that they contain less GLIR than the other more intensely stained GABA-immunoreactive neurons. In the caudal region, a single GABA-immunoreactive neuron was present in the dorsal rectal ganglion. Our data suggest that these ten cephalic neurons, and a single dorsal rectal ganglion neuron, use GABA as a neurotransmitter.

Biophysical transport properties of the cuticle of Ascaris suum.

The transport properties of isolated cuticle from Ascaris suum were studied using standard two-chamber diffusion cells and a number of radiolabeled permeants which varied in molecular size, lipophilicity and electrical charge. The permeability coefficient of the collagen matrix (lipid-extracted cuticle) vs. molecular radius relationship showed the interdependence of molecular size and electrical charge of the permeants with respect to the aqueous pores of the negatively charged matrix. The permeability of neutral solutes decreased monotonically with size. Protonated amines permeated the aqueous pores faster than neutral solutes of comparable size, while the permeation of anions was slower. The average pore size was estimated to be 1.5 nm in radius. A biophysical model which accounted for diffusion of molecules within a fixed electrostatic field of force and for molecular sieving by the pore channels was used in the mechanistic interpretation of the data. The effective permeability coefficient of the non-lipid-extracted cuticle was delineated into the permeability coefficients of the water-filled collagen matrix and the lipoidal component of the cuticle to determine which layer was the rate-controlling barrier. While each solute was capable of penetrating the water-filled collagen matrix, the rate-determining step for the majority of compounds was passive diffusion across the lipid component, which controlled 75-99% of transport. The exception was water, for which transport kinetics was 75% matrix-controlled. In general, permeation across the lipid-filled tissue was more favorable for small lipophilic compounds because of molecular restriction not only in the aqueous pores, but also in the lipid-filled pores.

Immunocytochemical demonstration of a neuropeptide in Ascaris suum (Nematoda) using an antiserum to FMRFamide.

A FMRFamide-like peptide has been detected in the nematode Ascaris suum, using the peroxidase-anti-peroxidase (PAP) immunocytochemical technique. Positive reactions were obtained in both the central nervous system and the peripheral nervous system of the worm, the strongest reactions being in the anterior nerve ring, the cephalic papillary ganglia, the lateral ganglia and the dorso-rectal ganglion. Immunoreactivity was observed along the length of the main nerve cords of the worm and, to a lesser extent, in the pharyngeal nerve cords. The possible role of this neuropeptide in the physiology of the nematode is discussed.

Ascaris suum: immunoperoxidase and fluorescent probe analysis of host proteases and parasite proteinase inhibitors in developing eggs and second stage larvae.

Live Ascaris suum females were incubated in medium containing chymotrypsin liganded to fluorescein-5-isothiocyanate, and eggs in the parasite's genital tract took up the probe and fluoresced. Eggs passed by these worms into the medium containing fluorescent probe retained their fluorescence in formaldehyde-saline and by 65 days had developed into second stage infective larvae. Eggs passed naturally by untreated worms were incubated in media containing fluorescent probes and all of the eggs exposed to chymotrypsin liganded to fluorescein-5-isothiocyanate were extensively labeled. Control eggs were labeled sporadically and less intensely, indicating specificity in the uptake of environmental proteins. Chymotrypsin from the parasite's environment can bind to A. suum eggs, and this occurs both inside the worm's genital tract and outside of the parasite. Immunoperoxidase studies showed that IgG developed against chymotrypsin or against A. suum chymotrypsin/elastase isoinhibitors A or C, binds to antigens in cross sections of second stage larvae and their egg shell coats. This suggests that host chymotrypsin is retained during development and may be complexed to A. suum isoinhibitors A and C.

The survival of parasite eggs throughout the soil profile.

An experiment using lysimeters suggested that the eggs of Taenia saginata and Ascaris lumbricoides survive for only a short time when applied to pasture in sewage sludge. However, a subsequent experiment which followed the survival of eggs throughout the soil profile demonstrated that some T. saginata eggs could still be found at 200 days on the soil surface, and that survival increased down the profile. Rainfall is shown to be able to wash eggs into the soil where they may be afforded protection from radiation and desiccation; this may have little epidemiological significance.