In vitro cultivation and morphological characterization of phloemic trypanosomatids isolated from coconut trees.

Plant trypanosomatids cause lethal vascular wilting in palms of the Arecaceae family. Infections, affecting plants in South and Central America, can result in significant economic loss. The study of trypanosomatids that cause these diseases has been complicated due to the inability to culture these organisms for in vitro analyses. To develop a protocol that would facilitate studies of trypanosomatids, continuous in vitro cultures of phloemic trypanosomatids were established from apical stems of diseased coconut trees collected in endemic and non-endemic regions of Brazil (the states of Bahia and Rio de Janeiro, respectively). Although attempts at establishing axenic cultures were unsuccessful, it was found that trypanosomatid co-cultures could be successfully established and maintained. The procedure was to preculture media with 10(4)Aedes albopictus cells in Grace's medium supplemented with 10% heat-inactivated fetal bovine serum (without antibiotics or fungicides) for 3 d before adding 10(6) trypanosomatids/ml harvested from either fresh apical stem extracts or with 2 mm(3) fragments of coconut apical stems. By day 7 under these conditions the parasites grew exponentially. Using this strategy, two isolates were identified and have been maintained in our laboratory for over 400 passages, demonstrating the efficacy of this culturing procedure. In situ the organisms were observed in vascular bundles and inside sieve elements of the phloem of diseased palms. In vitro parasites retained their mobility. Morphometric analysis revealed differences between Bahia and Rio de Janeiro isolates.

A new intracellular pathway of haem detoxification in the midgut of the cattle tick Boophilus microplus: aggregation inside a specialized organelle, the hemosome.

The hard tick Boophilus microplus ingests large volumes of cattle blood, as much as 100 times its own mass before feeding. Huge amounts of haem are produced during haemoglobin digestion, which takes place inside acidic lysosomal-type vacuoles of the digest cells of the midgut. Haem is a promoter of free radical formation, so haemoglobin digestion poses an intense oxidative challenge to this animal. In the present study we followed the fate of the haem derived from haemoglobin hydrolysis in the digest cells of the midgut of fully engorged tick females. The tick does not synthesize haem, so during the initial phase of blood digestion, absorption is the major route taken by the haem, which is transferred from the digest cells to the tick haemocoel. After this absorptive period of a few days, most of the haem produced upon haemoglobin degradation is accumulated in the interior of a specialized, membrane-delimited, organelle of the digest cell, herein called hemosome. Haem accounts for 90% of the hemosome mass and is concentrated in the core of this structure, appearing as a compact, non-crystalline aggregate of iron protoporphyrin IX without covalent modifications. The unusual FTIR spectrum of this aggregate suggests that lateral propionate chains are involved in the association of haem molecules with other components of the hemosome, which it is proposed is a major haem detoxification mechanism in this blood-sucking arthropod.

Report of Balanorchis anastrophus in Pará state with surface topography by scanning electron microscopy

Balanorchis anastrophus Fischoeder, 1901 from the reticulum from Bos taurus is reported for the first time in State of Pará, Brazil. The surface topography as revealed by scanning electron microscopy is presented