Adriana Morales

Haplotype characterization of the COI mitochondrial gene in Chrysoperla externa (Neuroptera: Chrysopidae) from different environments in Jaboticabal, state of São Paulo, southeastern Brazil

The green lacewings (Chrysopidae) belong to the Order Neuroptera and are described as voracious predators in the larval stage and sometimes also in their adulthood. They are an important group used in integrated biological control in field and horticultural crops. Individuals of Chrysoperla externa were collected during 2007 until March 2008 in five different locations in Jaboticabal, SP, with all the seasons sampled. Thirty six sequences with 805 pairs of bases for the gene mitochondrial Citochrome Oxidase I (COI) were analysed. The genetic parameters revealed 24 haplotypes for this population, a total of 36 mutations and haplotype diversity of 0.956. The data of genetic distance and population structure calculated for this population considering the different areas and seasons, revealed a great genetic similarity and high degree of genetic sharing between individuals sampled. It showed that the species Chrysoperla externa from Jaboticabal, SP, is a single population, without genetic structure neither due to the area of origin nor to the seasons of the year.

Effects of antiproteolytic agents on corneal epithelial viability and matrix metalloproteinase-2 and metalloproteinase-9 activity in alkali-burned corneas of rats

OBJECTIVE To evaluate the effects of agents on corneal re-epithelization and metalloproteinase-2 and metalloproteinase-9 (MMP-2 and MMP-9) activities in corneas of rats submitted to ulceration. ANIMALS STUDIED Ninety eight healthy rats. PROCEDURES Corneal ulcers were created using 1N NaOH in their left eye. Eyes were treated every 6 h with 1% ethylenediaminetetraacetic acid (EDTA), 3% chondroitin sulfate (CS), 10% N-acetylcysteine NAc and saline (S) at 6-h intervals. Corneas were stained with fluorescein and photographed at the same time points. Following 20 h and 40-42 h of corneal injury, corneas were processed for scanning electron microscopy (SEM) to quantify microvilli density, and MMPs activities were analyzed using zymography. RESULTS The percentage of wound area and the time in hours for corneal re-epithelization did not differ significantly among treatment groups (P > 0.05). In first and the second moments, latent MMP-2 was significantly elevated in the eyes treated with NAC and CS (P < 0.001). Active MMP-2 did not change significantly among treatment groups in the first moment (P > 0.05); significantly higher activity was observed in the second moment in the eyes treated with CS (P <0.001). In the second moment, latent MMP-9 decreased significantly in eyes treated with EDTA and S (P < 0.01). Microvilli corneal density did not change significantly between healthy subjects and treatment groups (P > 0.05). CONCLUSION Any of the studied substances did not accelerate corneal re-epithelization and did not add protection to the corneal microvilli. Significant higher levels of active form of MMP-2 in 3% chondroitin sulfate-treated group may indicate that the agent acts as substrate for such enzyme. At the end of the experiment, 1% EDTA was the most efficient agent to inhibit significantly the latent form of MMP-9. However, any of the substances add benefit over saline on reducing the proteolytic activity in the cornea of rats after alkali injury.

Distinct genetic structure in populations of Chrysoperla externa (Hagen) (Neuroptera, Chrysopidae) shown by genetic markers ISSR and COI gene

Distinct genetic structure in populations of Chrysoperla externa (Hagen) (Neuroptera, Chrysopidae) shown by genetic markers ISSR and COI gene. Green lacewings are generalist predators, and the species Chrysoperla externa presents a great potential for use in biological control of agricultural pests due to its high predation and reproduction capacities, as well as its easy mass rearing in the laboratory. The adaptive success of a species is related to genetic variability, so that population genetic studies are extremely important in order to maximize success of the biological control. Thus, the present study used nuclear (Inter Simple Sequence Repeat - ISSR) and mitochondrial (Cytochrome Oxidase I - COI) molecular markers to estimate the genetic variability of 12 populations in the Sao Paulo State, Brazil, as well as the genetic relationships between populations. High levels of genetic diversity were observed for both markers, and the highest values of genetic diversity appear associated with municipalities that have the greatest areas of native vegetation. There was high haplotype sharing, and there was no correlation between the markers and the geographic distribution of the populations. The AMOVA indicated absence of genetic structure for the COI gene, suggesting that the sampled areas formed a single population unit. However, the great genetic differentiation among populations showed by ISSR demonstrates that these have been under differentiation after their expansion or may also reflect distinct dispersal behavior between males and females.

Indicadores morfométricos em cabeças de espécies brasileiras de Chrysoperla (Neuroptera, Chrysopidae)

The species C. externa, C. defreitasi, C. raimundoi and C. genanigra (Neuroptera, Chrysopidae) have great differences in the genitalia. These species were studied morphometrically by the distances between points of the head (eight linear measurements) to better identify them. The results, analyzed by multivariate statistics, showed that it is possible to identify measures that distinguish the individual, although some species are more distinguishable by morphometry. The canonical analysis showed that three of the four species, C. raimundoi, C. genanigra and C. externa, showed no major overlap between them, and their possible discrimination within the group, only C. defreitasi presented a low resolution according to this analysis, being overlapped with C. externa and C. genanigra, however, it is totally broken in C. raimundoi. The measures that contributed most to differentiation are related to the size of the head and especially the length of wrasse on the edge of the head, until the level of the eyes and antenna.

Population genetics of Chrysoperla externa (Neuroptera: Chrysopidae) and implications for biological control

Green lacewings are insects with great potential to be use in the biological control of agricultural pests, but relatively few studies have attempted to understand the genetic structure of these agents, especially those of predatory insects. The purpose of this study was to characterize genetically populations of C. externa using sequences of subunit I of the cytochrome oxidase, a mitochondrial gene, and examine the population structure of this species in sampled areas in São Paulo state. The results indicate high genetic diversity but no genetic structure, detected by AMOVA analysis, and high levels of haplotype sharing in the network. These genetic patterns could be a consequence of environmental homogeneity provided by agroecosystem (citrus orchard), allowing gene flow among populations. Probably there is a unique population in the area sampled that could be used as a population (genetic) source for mass-reared and posterior release in these farms.

Non-contact specular microscopy in aphakic and pseudophakic dogs

Cataract is one of the most common ocular diseases in dogs, and phacoemulsification is considered its treatment of choice. Posterior capsular opacity (PCO) is a frequent complication and may occur weeks or months after the surgery. It is known that intraocular lenses (IOL) used for the correction of post-operative hypermetropia are an adjunctive aid in the prevention of PCOs, mainly the foldable acrylics designs with square and truncated edges. However, IOL placement may result in endothelial cell loss. This study evaluated the morphology and cellularity of the endothelium by non-contact specular microscopy in aphakic and pseudophakic dogs. Endothelial cell density, endothelial cell area, coefficient of variation and hexagonal cells percent were analyzed for each group and between groups. During all evaluation periods, endothelial cell density did not alter in any group. From days 7 to 30, endothelial cell density was higher in pseudophakic dogs. However, values returned to normal and, by the end of the study, no significant difference was observed. Cell area and variation coefficient did not differ in each group or between groups. Hexagonal cells percent significantly reduced after phacoemulsification in both groups. With time, this condition returned to initial values. When both groups were compared, hexagonal cells percent were decreased in the pseudophakic group, immediately after the surgeries.

Keratitis due to microfilariae in dogs: a newly recognized disease

Parasitic agents have been associated with keratitis, but a diagnosis of parasitic keratitis has not been commonly made in domestic animals. The purpose of this study was to describe the clinical and histopathological findings in seven dogs with chronic keratitis caused by microfilariae diagnosed in Brazil. All dogs presented with superficial corneal opacities of varying degrees affecting the perilimbal and central regions of the cornea, with other opaque areas appearing as crystalline deposits and corneal vascularization. The lesions were bilateral and were associated with mild-to-moderate conjunctival hyperemia. There was no history of blepharospasm or pruritus, and no subjects presented with epithelial erosions. Corneal biopsy revealed free microfilariae in the corneal stroma, with varying degrees of inflammation and collagen fiber destruction. The microfilariae were also found in skin lesions by skin snip technique. No adult worms were found in these dogs, and no dogs were on heartworm preventative before diagnosis. Monthly doses of oral ivermectin improved ocular and dermal lesions. One dog showed complete remission with the treatment. The species of the microfilariae was not identified.