A MORPHOLOGICAL AND ULTRASTRUCTURAL-STUDY OF THE PARACLOACAL (SCENT) GLANDS OF THE MARSUPIAL METACHIRUS-NUDICAUDATUS GEOFFROY, 1803

Morphological and ultrastructural features of the paracloacal glands of Metachirus us nudicaudatus are described. Two pairs of glands, one on the right and the other on the left of the anal canal, are formed, each consisting of a major and a minor portion. Their wall is made up of three layers: a mucosal, a muscular and an external capsule. The inner one is a mucosa the epithelium of which contains holocrine cells characterized by lipid droplets and intermediate filaments. The surrounding vascular lamina propria contains flattened tubular apocrine glands whose epithelial cells contain abundant endoplasmic reticulum, prominent Golgi complexes and numerous secretory granules. The middle layer is formed by skeletal striated muscle and the outer (third layer) consists of dense connective tissue. Each gland originates from a single duct. Transverse sections show that each duct, except in the female major gland, is in fact formed by a duct system. One of these ducts comes from the central cavity, lined by holocrine epithelium, and the others result from the branched tubular glands of the lamina propria.UNIV FED SAO PAULO,CTR MICROSCOPIA ELETR,SAO PAULO,BRAZILUNIV FED SAO PAULO,CTR MICROSCOPIA ELETR,SAO PAULO,BRAZILWeb of Scienc

Changes in Cell Wall Synthesis and Ultrastructure during Paradoxical Growth Effect of Caspofungin on Four Different Candida Species ▿

Paradoxical growth (PG) has been described for echinocandins and is characterized by cell growth at drug concentrations above the MIC. In this study, two isolates each of Candida albicans, C. tropicalis, C. orthopsilosis, and C. parapsilosis, all of which displaying PG in response to caspofungin, were subjected to MIC, minimal fungicidal concentration (MFC), and time-kill curve assays to evaluate the levels of PG. Cell wall components and ultrastructural modifications of the PG cells were also investigated. The results showed that when cell growth and survival were evaluated by MFC or time-kill curve assays, high concentrations of caspofungin did not show fungicidal activity against PG cells. Furthermore, for C. parapsilosis and C. orthopsilosis, time-kill curves were more discriminatory than MFCs in detecting the PG effect. The four different Candida species studied demonstrated similar alterations in cell wall components and ultrastructure associated with PG. In PG cells, β-1,3-glucan content decreased from 2.7- to 7.8-fold, whereas chitin content increased from 4.0- to 6.6-fold. An electron microscopy study of the PG cells revealed morphological alterations, clumping of cells, enlarged cells, the absence of filamentation, abnormal septa, and accumulation of chitin in the cell wall. Also, PG cells basically exhibited a single dark high-density layer in the cell wall, indicating the loss of the β-1,3-glucan layer. Our results present novel details about the ultrastructural alterations that occur in C. albicans, C. parapsilosis, C. orthopsilosis, and C. tropicalis during PG and show that chitin is the major component of the cell walls of PG cells. Stimulation of chitin synthesis may represent a rescue mechanism against caspofungin activity

Light and Transmission Electronic Microscopy Evaluation of Lyophilized Corneas

Purpose: Cornea storage for longer periods is still a challenge for corneal Surgeons. The purpose of this study was to find a method to lyophilize corneas for anterior lamellar transplant and to evaluate. them by light and transmission electronic microscopy.Methods: Corneal flaps were created by using a microkeratome. Corneas were lyophilized with a cryoprotectant (2.3 mol sacarousis for 40 minutes) and without a cryoprotectant in a lyophilization machine (Modulyon D). The corneas were rehydrated with distilled water, balanced saline solution (BSS), and phosphate-buffered saline, after which they were evaluated by microscopy. A cornea that did not undergo lyophilization served as a control.Results: Lyophilization without a cryoprotectant did not preserve the corneal structure. This finding was also observed when lyophilizing and rehydrating the corneas with distilled water or phosphate-buffered saline. We found that lyophilizing corneas and rehydrating them with 11 mL of BSS for 30 minutes preserved the general corneal structure, the parallelism of the collagen fibers, the Bowman layer, and the epithelial basement membrane for 15 and 30 days and for as long as 1 year or more.Conclusions: Lyophilization with sacarousis and rehydration with BSS may be a good method for anterior lamellar transplantation.Univ Fed Sao Paulo, Sao Paulo, BrazilSorocaba Eye Bank, Sorocaba, BrazilUniv Fed Sao Paulo, Sao Paulo, BrazilWeb of Scienc

Primers and probes used in this study.

<p>¹The nucleotide numbering refers to the numbering from the Tohama I genome [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132623#pone.0132623.ref019" target="_blank">19</a>]. Segment accession numbers: BX640414 (<i>prn</i>), BX640422 (<i>ptxP</i>, <i>ptxA</i>, <i>ptxB</i>), BX640416 (<i>fimD</i> and <i>bvgS</i>) and BX640414 (<i>tcfA</i>).</p><p>²The “colder”primer forming only two hydrogen bonds at the specific base has a one base extension at its 5´-end to compensate the reduced T_m.</p><p><b>A,</b> Primers prnAF (top) and prnAR (bottom) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132623#pone.0132623.ref014" target="_blank">14</a>] were used for <i>prn</i> sequencing. <b>B,</b><b>C-1,</b> ARMS-qPCR primers for the determination of the <i>ptxA</i>, <i>ptxB</i>, <i>fimD</i>, <i>bvgS</i> and <i>tcfA</i> alleles <b>(B)</b> as well as for the discrimination between the <i>ptxP1</i>-like and <i>ptxP3</i> alleles <b>(C-1)</b>. For each gene, the forward primer is shown on top, then the reverse primer corresponding to the Tohama I sequence and the reverse primer corresponding to the alternative allele, with the base at the SNP site underlined, and finally the labelled hydrolysis probe. Lower case letters, artificial base replacement with both sequences at the SNP site; FAM, 6-carboxyfluorescein; TET, 4,5,6,7-tetrachlorofluorescein; BHQ1, Black Hole Quencher-1; Cy5, red sulfoindocyanine dye. <b>C-2,</b> oligonucleotides for the <i>fimD</i> non-discriminatory control assay.</p

Genomic variations in <i>prn</i> variants of Austrian <i>B</i>. <i>pertussis</i> samples (n = 77) collected from 2002–2008. Frequencies are given as numbers and as percentage of successfully typed samples (in brackets).

<p>¹To distinguish the prn variants Prn1 and Prn7, the sequence of the second polymorphic region is needed as well.</p><p>Genomic variations in <i>prn</i> variants of Austrian <i>B</i>. <i>pertussis</i> samples (n = 77) collected from 2002–2008. Frequencies are given as numbers and as percentage of successfully typed samples (in brackets).</p