<i>In Vitro</i>-Propagation of <i>Agave tequilana</i> Weber cv. azul in a Temporary Immersion System

In Mexico, there is a need to produce large quantities of plantlets for the establishment and replanting of blue (cv. azul) agave production areas. Most of these plots are within the origin denomination area (DOT, Spanish acronym) of the distilled product of this plant, known as tequila. The objective of this study was to develop an in vitro-propagation protocol for Agave tequilana Weber cv. azul using segmented stems in both: solid and liquid media. A disinfection and in vitro technique were developed to obtain shoots, through plantlets collected in commercial plots, which attained 100% surface-disinfection and budding rate. At the multiplication stage, the effects of 6-Benzylaminopurine (BA) (0.0, 4.4 and 13.2 μM) and kinetin (0.0, 9.4, 18.8 and 37.6 μM) were evaluated on lateral-shoot production of segmented sagittal stems. These were cultivated on Murashige & Skoog (MS) medium, with the addition of 3.0% sucrose and 8 g L⁻¹ agar. It was observed that BA and kinetin increased the number of shoots per explant, obtaining up to 18 and 26, respectively. Furthermore, it was found that just the sagittal segmentation of explants increased axillary budding. On the other hand, segmented-stem bases were grown in MS liquid medium with 3.0% sucrose, inside a RITA® system, programmed by a 5 min immersion step with a frequency of every 4 h. The effect of Indole−3-Acetic acid (IAA) (0.57, 2.9, 5.7 μM) was evaluated, while maintaining a concentration of BA (13.2 μM). It was observed that the greatest concentration of IAA led to the formation of more than 20 buds per explant. These results offer a new methodology to increase the efficiency of A. tequilana Weber cv. azul-in vitro multiplication by sagittal segmentation of stems and the addition of BA and/or IAA.     

Trypanoxyuris (Trypanoxyuris) minutus (Schneider, 1866) among Alouatta guariba clamitans (Cabrera, 1940) in the state of Rio de Janeiro,Brazil

This study reports on infection by Trypanoxyuris (Trypanoxyuris) minutus among Alouatta guariba clamitans through biometry on adults and eggs. A total of 58 adult helminths were recovered from the small segment of the animal's large intestine. This study is the first report of this parasite among A. guariba clamitans in Rio de Janeiro.     

Detection of Toxoplasma gondii DNA by polymerase chain reaction in experimentally desiccated tissues

Despite toxoplasmosis being a common infection among human and other warm-blooded animals worldwide, there are no findings about Toxoplasma gondii evolutionary forms in ancient populations. The molecular techniques used for amplification of genetic material have allowed recovery of ancient DNA (aDNA) from parasites contained in mummified tissues. The application of polymerase chain reaction (PCR) to paleoparasitological toxoplasmosis research becomes a promising option, since it might allow diagnosis, acquisition of paleoepidemiological data, access to toxoplasmosis information related origin, evolution, and distribution among the ancient populations. Furthermore, it makes possible the analysis of parasite aDNA aiming at phylogenetic studies. To standardize and evaluate PCR applicability to toxoplasmosis paleodiagnostic, an experimental mummification protocol was tested using desiccated tissues from mice infected with the ME49 strain cysts, the chronic infection group (CIG), or infected with tachyzoites (RH strain), the acute infection group (AIG). Tissues were subjected to DNA extraction followed by PCR amplification of T. gondii B1 gene. PCR recovered T. gondii DNA in thigh muscle, encephalon, heart, and lung samples. AIG presented PCR positivity in encephalon, lungs, hearts, and livers. Based on this results, we propose this molecular approach for toxoplasmosis research in past populations.     

Plant core DNA barcode performance at a local scale: identification of the conifers of the state of Hidalgo,Mexico

DNA barcoding constitutes a fundamental tool for species identification, especially for highly diverse geographic regions. Here, we characterize and evaluate the plant core barcoding regions matK and rbcL to identify the 25 conifer species from the state of Hidalgo, Mexico, including 10 species in various threat categories. Sequence quality, linguistic complexity, and the presence of the barcode gap were estimated. Two methods were compared for successful species identification: BRONX (Barcode Recognition Obtained with Nucleotide eXposés) and the least inclusive clade. We generated 77 sequences for matK and 88 for rbcL. The matK region had higher haplotype diversity and nucleotide diversity (Π), including six indels. The analysis of 77 specimens with complete sequences (matK + rbcL) resulted in 21 nonspecies-specific unique haplotypes for the 25 conifer species. Higher sequence quality and linguistic complexity were observed in rbcL than in matK. Every diagnosable species had a barcode gap. Ninety-seven specimens were assigned unambiguously to family and genus, regardless of the marker or method employed. The analysis of matK with BRONX produced the highest species level identification success (44%). Despite the low specimen identification success at the specific level, it will be possible to establish local management, conservation, and monitoring projects for at least half of the threatened species even when specimens do not exhibit diagnostic morphological characters. The low divergence between closely related species may result from the slow rate of molecular evolution of the core barcoding markers or from hybridization or incomplete lineage sorting. Similar identification success is expected for groups with comparable life history traits under similar conditions as this study. A reduction in the geographic area will not necessarily translate into higher identification success, especially for high-diversity regions and centres of diversification.