Ignacio Méndez

Nuclear genome size analysis of Agave tequilana Weber

Abstract Flow cytometric analysis of DNA content was performed using nuclei isolated from internal basal parts of young leaves in 8 varieties of Agave tequilana. The distributions of nuclear DNA content showed that more than 85% of leaf nuclei were in G0/G1 phase of the cell cycle. Nuclei with DNA content higher than 4C were not detected, indicating the absence of endopolyploidy. A. tequilana varieties lineño, azul listado, azul, moraleño, xiguin, and pata de mula were found to be diploid (2n =2x=60). These diploid varieties displayed a 2.5% variation in 2C DNA content, which ranged from 8.304 pg for var. lineño to 8.517 pg for var. pata de mula. No statistically significant differences were detected among all diploid varieties considered. This observation demonstrates that Agave belongs to plant taxa with a stable genome size. The mean 1Cx value (DNA amount of monoploid chromosome set .) calculated for diploid cultivars was equal to 4.202 pg DNA. Triploid var. bermejo (2n=3x=90) had 2C=12.513 pg DNA, i. e. approximately three times the 1Cx value. 2C DNA amounts of tetraploid (4x) and pentaploid (5x) var. chato were equal to 16.01 pg and 20.11 pg DNA, respectively. These values are also multiples of the 1Cx value, indicating a relationship between ploidy level and 2C DNA content. Nevertheless, the 1Cx values calculated for tetraploids and pentaploids were lower than the 1Cx value determined for diploids. This observation indicates a loss of certain DNA sequences after polyploidization. The results obtained form a basis for the application of flow cytometry in taxonomy, breeding and biotechnology of Agave tequilana.

Aflatoxin M1 in pasteurized and ultrapasteurized milk with different fat content in Mexico.

High per capita milk consumption in Mexico indicated a strong need for documentation of aflatoxin M1 (AFM1) levels in milk. A survey of 580, 2-liter samples (n = 290), was conducted to quantify AFM1 using high-performance liquid chromatography, considering two maximum tolerance levels (0.05 and 0.5 microg/liter). We relate aflatoxin levels in the seven most consumed brands from different regions, with two processes (pasteurized and ultrapasteurized), different expiration dates, and different fat content: whole fat (28, 30, and 33 g), half-skimmed (10, 16, and 20 g), light (1, 2, and 4 g), and with vegetable oil. Pasteurization and ultrapasteurization did not diminish AFM1 contamination present at levels of 0 to 8.35 microg/liter in 40% of the milk samples at concentrations > or = 0.05 microg/liter and in 10% of the samples at > or = 0.5 microg/liter. Statistically significant relationships were AFM1 contamination with brand (P = 0.002 at the > or = 0.05 microg/liter level and P = 0.034 at the > or = 0.5 microg/ liter level) and higher AFM1 levels with mild or warm seasons of the year (P = 0.0003). Samples with greater fat content had slightly more probability (P = 0.067) of being contaminated by AFM1 at the > or = 0.5 microg/liter level. The milk with the lowest contamination of AFM1 was a brand imported as powder and rehydrated in Mexico.

Aflatoxin B1 and its interconverting metabolite aflatoxicol in milk: the situation in Mexico

Between 1996 and 1998, 580 litres of milk in Mexico were surveyed for aflatoxin B1 (AFB1) and its metabolite aflatoxicol (AFL), which are mutagenic and carcinogenic mycotoxins that interconvert AFB1–AFL–AFB1. The seven most consumed brands from different regions of Mexico included pasteurized and ultrapasteurized milk with four different fat levels: whole fat (28–33 g l−1), half-skimmed (10–20 g l−1), light (1–4 g l−1) and with vegetable oil (33 g l−1). Aflatoxins in each sample were concentrated with total aflatoxin immunoaffinity columns and quantitated by high-performance liquid chromatography. A milk sample was considered contaminated if it contained ≥0.05 μg l−1 AFL. Pasteurization and ultrapasteurization of milk did not control contamination with AFL, which was present in 13% of samples at ≥0.05 μg l−1 and in 8% at ≥0.5 μg l−1, with a range of AFL from 0 to 12.4 μg l−1. AFB1 was present mainly in traces (0–0.4 μg l−1). The safest milk in relation to AFL contamination was imported milk powder with vegetable oil. There was a significant correlation between contamination of milk with AFL and the autumn (p<0.0002); the fat content was not significant.

Karyotype studies in cultivars of Agave tequilana Weber

Abstract Agave tequilana cultivars “Lineño”, “Azul Listado”, “Moraleño”, “Azul”, “Sigüin”and “Pata de Mula” were diploids with 2n=2x=60; cult. “Bermejo” were triploid with 2n=3x=90, and cult. “Chato” tetraploid with 2n=4x=120 (x= 30). A. tequilana cultivars show a bimodal karyotype consisting of 10 large + 50 small chromosomes in diploids; 15 large + 75 small chromosomes in triploids and 20 large + 100 small chromosomes in tetraploids. In diploids they had secondary constriction in one pair of large chromosome and in the third or fourth largest homologous chromosome groups in triploids and tetraploids, respectively. The variation among cultivars was clearly evident in the arm ratio, proportion of different types of large and small chromosomes, mean of genome length and asymmetry index of karyotypes. The variation observed among Agave tequilana cultivars is possible due to rearrangements in the large and small chromosomes of the complement. These results support the separation of diploid cultivars “Azul”, “Azul Listado”, “Moraleño” and “Sigüin” from diploid “Pata de Mula” and polyploids “Bermejo” (3x) and “Chato” (4x).

Survey of aflatoxins in maize tortillas from Mexico City

In Mexico, maize tortillas are consumed on a daily basis, leading to possible aflatoxin exposure. In a survey of 396 2-kg samples, taken over four sampling days in 2006 and 2007 from tortilla shops and supermarkets in Mexico City, aflatoxin levels were quantified by HPLC. In Mexico, the regulatory limit is 12 µg kg−1 total aflatoxins for maize tortillas. In this survey, 17% of tortillas contained aflatoxins at levels of 3–385 µg kg−1 or values below the limit of quantification (12 µg kg−1 and 87% were below the regulatory limit. Average aflatoxin concentrations in 56 contaminated samples were: AFB1 (12.1 µg kg−1); AFB2 (2.7 µg kg−1); AFG1 (64.1 µg kg−1) and AFG2 (3.7 µg kg−1), and total AF (20.3 µg kg−1).

Vegetation patches improve the establishment of Salvia mexicana seedlings by modifying microclimatic conditions.

Human disturbance has disrupted the dynamics of plant communities. To restore these dynamics, we could take advantage of the microclimatic conditions generated by remaining patches of vegetation and plastic mulch. These microclimatic conditions might have great importance in restoring disturbed lava fields located south of Mexico City, where the rock is exposed and the soil is shallow. We evaluated the effects of both the shade projected by vegetation patches and plastic mulch on the mean monthly soil surface temperature (Tss) and photosynthetic photon flux density (PPFD) and on the survival and growth of Salvia mexicana throughout the year. This species was used as a phytometer of microsite quality. Shade reduced the Tss to a greater extent than mulch did. Both survival and growth were enhanced by shade and mulch, and the PPFD was related with seedling growth. During the dry season, plant biomass was lost, and there was a negative effect of PPFD on plant growth. At micro-meteorological scales, the use of shade projected by patches of vegetation and mulch significantly reduced the mortality of S. mexicana and enhanced its growth. Survival and growth of this plant depended on the environmental quality of microsites on a small scale, which was determined by the environmental heterogeneity of the patches and the landscape. For plant restoration, microsite quality must be evaluated on small scales, but on a large scale it may be enough to take advantage of landscape shade dynamics and the use of mulch to increase plant survival and growth.

Nuclear genome size and karyotype analysis in Mammillaria species (Cactaceae)

Abstract Seven species of Mammillaria were studied, all diploid: 2n=2x=22, x=11. Genome size was determined by flow cytometry, varied from 2C DNA =3.20 pg, 1570 Mbp (1 Cx) in M. crucigera, to 2C DNA = 3.04 pg, 1490 Mbp in M. flavicentra. The variation of these species in the nuclear content of DNA was 5% and was not significant (P = 0.3469). This indicates that they are small, very stable genomes, in which changes in DNA content are not very evident. The variation among species, however, was clearly evident in the relative length (L%) and arm ratio (r), the proportion of metacentrics and submetacentrics, and the position of satellites. The karyotype for M. albilanata, M. dixanthocentron and M. flavicentra was 11m, and in M. huitzilopochtli, 10m + 1 sm. Only one pair of chromosomes was observed with satellite in the four species. M. dixanthocentron and M. flavicentra, species considered synonymous, exhibited the satellite on different chromosomes. The interspecific variation observed among the species of Mammillaria is possibly due to spontaneous structural changes in their chromosomes. These mechanisms of restructuring in the genome of these species have not involved significant changes in nuclear DNA content. The Mammillaria species exhibited an endopolyploidy pattern with 2–16 C DNA content in the stem parenchyma, which may give them alternative strategies for adaptation in arid environments.

Nuclear genome size and cytotype analysis in Agave parviflora Torr. subsp. flexiflora Gentry (Asparagales, Asparagaceae)

This study presents the cytogenetic characterization by karyotyping and a determination of the DNA content by flow cytometry of wild adult plants, plants grown from seeds of wild plants and bulbils of Agave parviflora subsp. flexiflora from Bacadehuachi to Nacori Town, Sonora, Mexico. The analyzed plants were diploids (2n = 2x = 60) and had three different structural cytotypes. The cytotype observed in wild adult plants was 44m + 4sm + 10st + 2t, the cytotype found in plants grown from seeds was 48m + 8st + 4t and the cytotype displayed by bulbils was 44m + 2sm + 14st. Agave parviflora subsp. flexiflora showed a bimodal karyotype of 10 large + 50 small chromosomes. All diploid plants had a secondary constriction in one pair of the large chromosomes. The arm ratio, the proportion of different types of large and small chromosomes, the mean of genome length and the asymmetry index of karyotypes clearly varied among all three diploid cytotypes. The pattern of variation among all the types of plants is probably due to rearrangements in the large and small chromosomes of the complement. Differences in the amount of nuclear 2C DNA = 8.20 pg in diploid wild adult plants and 8.21 pg in bulbils were significantly different when compared to 2C DNA = 8.07 pg of diploid plants grown from seeds. The different types of plants displayed 1.7% variation in the 2C DNA content and the mean 2C DNA content was 8.16 pg; 1Cx value = 4.08 pg. The results here reported consist of basic and useful information to set conservation strategies and breeding approaches for Agave parviflora subsp. flexiflora.

Cytological and genical differentiation between cytotypes ofEcheandia nana(Anthericaceae)

Abstract The analysis of 9 populations of Echeandia nana showed all to be diploid,with 2n = 16, n −. 8 (x = 8). The analyzed populations displayed two cytotypes.Cytotype A − 10m+6sm, having two pairs of chromosomes with a satellite, was observed in four populations from the eastern flanks of the Pachuca mountain range. The five remaining populations from the western flank of the Pachuca and Sierra Nevada mountain ranges (Mexico) showed cytotype B + 6m+8sm+2st, having one pair of chromosomes with a satellite. The analysis of meiosis revealed heterozygotic exchanges. Analysis of MI showed three heteromorphic bivalents in cytotypes A and B. Analysis of AI showed sub-chromatid aberrations, (side arm bridges = SAB), which were more frequent in cytotype B (39.88+44.84%) than in cytotype A (2.86+31+53%). Cells with two bridges (SAB aberrations) were observed in cytotype B.The intraspecific cytological and genical differentiation of cytotypes A and B is probably the result of geographical isolation between populations of E. nana. This suggests that this species is undergoing through a major process of genomic differentiation involving heterozygotic chromosomal rearrangements; which favors a process of speciation between both cytotypes without the occurrence of significant morphological changes. This cytological and genical differentiation between cytotypes A and B was evident in the significant differences of the low number of fruits and viable seeds produced after cross-pollination among cytotypes (AxB: 0–16 fruits; 0–448 abortive seeds), relative to the larger values recorded after crosspollination within cytotypes (AxA: 4–48; 152–1824) (BxB: 3–89; 14–3382).

Brain-stem auditory evoked potentials in children with perinatal encephalopathies

OBJECTIVE We sought to describe if neurological damage, in terms of brain lesions, syndrome and syndrome severity led to abnormalities in the brain-stem auditory evoked potentials (BAEPs) in order to provide a profile of children that could be used as an indicator of subsequent neurological sequelae. We analyzed the BAEPs from a group of children having prior evidence of neurological damage and determined the presence of neurological sequelae when the subjects were 3 years old. METHODS Brain-stem auditory evoked potentials (BAEPs) were carried out in a group of 154 children with perinatal neurological damage. The children were classified with neurofunctional (clinical and EEG alterations) or organic and neurofunctional brain disease (clinical, EEG and image alteration) and were all followed from the first month of life and serially for 3 years. We used principal component analysis (PCA), clustered analysis and linear correlation to determine association between BAEPs, risk factors and future sequelae. RESULTS Latencies of BAEPs decreased significantly with age, and the time of conduction was modified by the presence of neurological damage. All statistical analyses suggested positive and significant associations between risk factors (trophism and condition at birth), and the latencies of waves I, III and V as well as with IPL III-V (interpeak latency) and I-V. PCA showed that IPL I-III was also positively associated with condition at birth, severity of the neurological syndrome and encephalopathy. In addition, we found that the presence and type of sequela reflected changes in the latencies of the waves, as well as IPLs, primarily those of IPL I-III. CONCLUSIONS Our results suggest that statistical methods are often needed to analyze neurological damage. The relation between BAEPs, risk factors and neurological sequelae allowed us to obtain a profile of children, which can be then used as an aid in the prognosis of children having a risk of developing neurological sequelae.