Characterization of 32 microsatellite loci for the Pacific red snapper, <Emphasis Type="Italic">Lutjanus peru</Emphasis>, through next generation sequencing

We developed a set of hypervariable microsatellite markers for the Pacific red snapper (Lutjanus peru), an economically important marine fish for small-scale fisheries in the west coast of Mexico. We performed shotgun genome sequencing with the 454 XL titanium chemistry and used bioinformatic tools to search for perfect microsatellite loci. We selected 66 primer pairs that were synthesized and genotyped in an ABI PRISM 3730XL DNA sequencer in 32 individuals from the Gulf of California. We estimated levels of genetic diversity, deviations from linkage and Hardy–Weinberg equilibrium, estimated the frequency of null alleles and the probability of individual identity for the new markers. We reanalyzed 16 loci in 16 individuals to estimate genotyping error rates. Eighteen loci failed to amplify, 16 loci were discarded due to unspecific amplifications and 32 loci (14 tetranucleotide and 18 dinucleotide) were successfully scored. The average number of alleles per locus was 21 (±6.87, SD) and ranged from 8 to 34. The average observed and expected heterozygosities were 0.787 (±0.144 SD, range 0.250–0.935) and 0.909 (±0.122 SD, range 0.381–0.965), respectively. No significant linkage was detected. Eight loci showed deviations from Hardy–Weinberg equilibrium, and from these, four loci showed moderate null allele frequencies (0.104–0.220). The probability of individual identity for the new loci was 1.46^?62. Genotyping error rates averaged 9.58%. The new markers will be useful to investigate patterns of larval dispersal, metapopulation dynamics, fine-scale genetic structure and diversity aimed to inform the implementation of spatially explicit fisheries management strategies in the Gulf of California.     

Development and characterization of microsatellite markers for Echinopsis rhodotricha and cross-amplification in other species of Cactaceae

Located in central South America, the Chaco is a large subtropical dry forest characterized by plants with xeromorphic features, including numerous cacti. In the Brazilian Chaco at the northeasternmost part of the Chaquenian region, Echinopsis rhodotricha (Cactaceae, Trichocereeae) is one of the species threatened by intense deforestation caused by expanding pastureland. This study characterizes the microsatellite loci isolated from E. rhodotricha and cross-amplification in thirteen other cactus species. Twelve microsatellite loci were developed from an enriched genomic library. Eight of these were polymorphic and characterized in 48 individuals from three E. rhodotricha populations. The loci showed a mean of 3.2 alleles per locus and overall levels of expected and observed heterozygosities ranging from 0.00 to 0.83 and 0.00 to 0.66, respectively. Five loci showed significant departures from the Hardy–Weinberg equilibrium and also exhibited signs of null alleles. Cross-amplification in other Cactaceae species was successful, ranging from one (Ferocactus latispinus, Cacteae and Harrisia adscendens, Trichocereeae) to twelve loci (Echinopsis calochlora, Trichocereeae). The development of these microsatellite markers will contribute to investigations of population structure, genetic diversity, and gene flow in E. rhodotricha populations, as well as in other cactus species, providing information useful for the creation and delimitation of conservation areas in the Brazilian Chaco region.     

Characterization of six microsatellite DNA loci for Sorex arizonae

Sorex arizonae is a rare species that occupies a narrow range of habitat types in several mountain ranges of New Mexico, Arizona and Northern Mexico. Here we identify and characterize six microsatellite loci for this species. We screened 63 individuals from four different localities from New Mexico and Arizona to analyse genetic variability. Alleles ranged from three to 16. Heterozygosity ranged from 40% to 78%. Most polymorphic loci were in Hardy–Weinberg equilibrium with the exception of one locus. Primers appear to have reasonable cross‐species applicability as five loci amplified in another shrew species (Sorex monticolus).     

Isolation and characterization of microsatellite DNA markers in the Pacific abalone,Haliotis discus hannai

We developed 17 new microsatellite markers in Haliotis discus hannai. All loci were found to be polymorphic with an average of 13.1 alleles per locus (range 3–28). The mean observed and expected heterozygosities were 0.77 (range 0.17–1.00) and 0.79 (range 0.42–0.96), respectively. Six loci deviated significantly from Hardy–Weinberg proportions, and thus should be used with caution. The high variabilities revealed in this study suggest that these microsatellite loci should provide useful markers for studies of trait mapping, kinship and population genetics.     

Eight polymorphic microsatellite markers for kelp bass,Paralabax clathratus,amplified in three multiplex polymerase chain reaction sets

Kelp bass, Paralabrax clathratus, is an important sport fishery of California, USA and Baja, Mexico. Here we describe eight microsatellite loci developed for this species. Two loci were derived from known primers for other species in the family, Serranidae and six were developed anew using a clone enrichment protocol. Loci were arranged into three multiplex PCR (polymerase chain reaction) sets for fast throughput; 564 individuals from nine populations across the species’ range were genotyped. Polymorphism ranged from four to 47 alleles and all populations at all loci displayed Hardy–Weinberg and linkage equilibrium.     

Twelve microsatellites for two deep sea polychaete tubeworm species,Lamellibrachia luymesi and Seepiophila jonesi,from the Gulf of Mexico

The vestimentiferan tubeworms Lamellibrachia luymesi and Seepiophila jonesi are found at hydrocarbon seeps in the Gulf of Mexico. Primers for polymorphic microsatellite loci were developed from genomic libraries of L. luymesi (five loci) and from S. jonesi tissue (eight loci) and were used to screen individuals collected from nine northern Gulf of Mexico hydrocarbon seep sites. Loci had from four to more than 50 alleles with high expected levels of heterozygosity. Cross‐species amplification, tested on seven vestimentiferan species including both hydrothermal vent and cold seep species, was generally strong in similar species but weak in more genetically distant species.     

Isolation and characterization of microsatellite loci in the flat‐headed bat (Tylonycteris pachypus)

Microsatellite loci were developed in the flat‐headed bat (Tylonycteris pachypus) from genomic DNA using an enriched library method. Nine loci were tested on 48 individuals sampled from Guangxi Province, China. The mean number of observed alleles per locus was 6.4 (range 4–12). Observed and expected heterozygosity values ranged from 0.24 to 0.83 and from 0.30 to 0.89, respectively. One locus revealed significant departure from Hardy–Weinberg equilibrium and no significant linkage disequilibrium was detected between loci pairs. These markers will be used to examine genetic structure and parentage analysis in this species.     

Eleven new highly polymorphic microsatellite loci in the yellow croaker,Pseudosciaena crocea

We developed 11 new microsatellite markers in Pseudosciaena crocea by screening an enriched genomic library using nonradioactive polymerase chain reaction (PCR) techniques. All loci were found to be polymorphic with an average of 14.9 alleles per locus (range four to 30). The mean observed and expected heterozygosities were 0.86 (range 0.57–1.00) and 0.90 (range 0.62–0.98), respectively. Four loci showed significant Hardy–Weinberg disequilibrium. The high variabilities revealed in this study suggest that these microsatellite loci should provide useful markers for population genetic studies of P. crocea.     

Polymorphic microsatellites isolated from the cave fish Astyanax fasciatus

Six polymorphic microsatellite loci from a library of the tetra Astyanax fasciatus from Mexico were isolated. Amplification and heterozygosity were tested in four cave and four surface populations. These loci were developed for population genetic study to detect gene flow between cave and surface populations.     

Isolation and characterization of microsatellite loci from a threatened rattlesnake (New Mexico Ridge‐nosed Rattlesnake,Crotalus willardi obscurus)

Six novel microsatellite loci are identified from genomic DNA of the threatened New Mexico Ridge‐nosed Rattlesnake (Crotalus willardi obscurus). Data from the Animas Mountains (New Mexico) population demonstrate these loci: (i) are highly variable with 5–24 alleles per locus, expected heterozygosities between 0.35 and 0.92, and observed heterozygosities between 0.32 and 0.91; (ii) are sufficiently variable for assigning parentage with total exclusionary power for the first parent of 0.96, and 0.99 for the second parent; and (iii) amplify similar size fragments in other rattlesnakes (C. atrox, C. lutosus, C. scutulatus, and C. tigris).     

Pollination system of the Pilosocereus leucocephalus columnar cactus (tribe Cereeae) in eastern Mexico

It has been suggested that there is a geographic dichotomy in the pollination systems of chiropterophilous columnar cacti: in intra‐tropical areas they are pollinated almost exclusively by bats, whereas in extratropical areas they are pollinated by bats, birds and bees. However, currently the studies are clumped both taxonomically (mainly Pachycereeae species) and geographically (mainly in the Tehuacan Valley and the Sonoran Desert). This clumping limits the possibility of generalising the pattern to other regions or cactus tribes. Only four of the 36 chiropterophilous cacti in Pilosocereus have been studied. Despite the tropical distribution of two Pilosocereus species, bees account for 40–100% of their fruit set. We examined how specialised is the pollination system of P. leucocephalus in eastern Mexico. As we studied tropical populations, we expected a bat‐specialised pollination system. However, previous studies of Pilosocereus suggest that a generalised pollination system is also possible. We found that this cactus is mainly bat‐pollinated (bats account for 33–65% of fruit set); although to a lesser degree, diurnal visitors also caused some fruit set (7–15%). Diurnal visitors were more effective in populations containing honeybee hives. P. leucocephalus is partially self‐compatible (14–18% of fructification) but unable to set fruit without visitors. Despite the variation in pollination system, P. leucocephalus shows more affinity with other columnar cacti from tropical regions than with those from extratropical regions. Although we report here that a new species of tropical Pilosocereus is relatively bat‐specialised, this Cereeae genus is more flexible in its pollination system than the Pachycereeae genera.     

Columnar cacti as sources of energy and protein for frugivorous bats in a semi‐arid ecosystem

Columnar cacti constitute the dominant elements in the vegetation structure of arid and semi‐arid New World ecosystems representing a plethora of food resources for vertebrate consumers. Previous stable isotope analysis in Central Mexico showed that columnar cacti are of low importance to build tissue for frugivorous bats. We used carbon stable isotope analysis of whole blood and breath samples collected from four species of frugivorous bats (Sturnira parvidens, Sturnira ludovici, Artibeus jamaicensis, and Artibeus intermedius) to reconstruct the importance of cactus plants in their diet. Breath samples were collected within 10 min (B₁₀) of bat capture and ~12 h after capture (B₇₂₀), representing the oxidation of recently ingested food and of body reserves, respectively. We expected that bats relied primarily on non‐cactus food to construct tissues and fuel oxidative metabolism. Non‐cactus food strongly predominated for tissue building, whereas oxidative metabolism was supported by a moderate preponderance of non‐cactus food for B₁₀ samples, and a moderate preponderance of cactus food or an equal contribution of both sources for B₇₂₀ samples. Artibeus and Surnira species appear to cover a narrow part of the diet with cactus food, confirming that the incorporation of nutrients derived from these plants is not generalized among vertebrate consumers.     

Candy Barrel Cactus (<Emphasis Type="Italic">Echinocactus platyacanthus</Emphasis> Link &; Otto: A Traditional Plant Resource in Mexico Subject to Uncontrolled Extraction and Browsing

Candy Barrel Cactus ( Echinocactus platyacanthus Link & Otto [C Mex.]): A Traditional Plant Resource in Mexico Subject to Uncontrolled Extraction and Browsing. The candy barrel cactus (Echinocactus platyacanthus) is an endemic of the Chihuahuan Desert in México that is used as an ornamental plant, for fodder, and for human consumption. The uncontrolled use of this species has resulted in its being put under special legal protection. Although now illegal to harvest, many candy barrel cacti continue being destroyed in situ by both gathering and livestock grazing. This study describes the uses of this species and the characteristics of its natural population in central Mexico. In order to estimate density, percentage of damaged cactus, and height of candy barrel cactus, censuses were carried in stands of 2,500 m². The anthropogenic disturbance in each site was estimated by an environmental disturbance index (DI). Results show that the state of Hidalgo has the best conserved population (mean density = 1,111 ind/ha, stratus height = 137 cm, and only 2% of the total cacti injured). Populations in Puebla represent an intermediate state of conservation, while Querétaro has populations with the lowest density (435 ind/ha), the shorter height (62 cm), the highest foraging (18%), and the greatest DI (71.30). We recommend that conservation strategies for this species be implemented immediately, and that areas be established that exclude livestock and prevent extraction by humans.     

Allozyme Genotype-Environment Relationships. I. Variation in Natural Populations of DROSOPHILA PACHEA

Remarkable geographic and seasonal uniformities in gene and genotype frequencies were found for four enzyme loci. Principal component analysis revealed two patterns of allele frequency changes in three of the allozyme loci (Est-b, Est-c and Acph) and one pattern in the Mdh-d locus. These patterns accounted for 90% to 100% of the variability at these loci. Significant positive correlations were obtained between several of the components and the variability in the non-saponifiable and alkaloid fractions of the senita cactus host plant. The genetic variance within a single locality (Guaymas, Mexico) was as great as the variance between all localities sampled. This is interpreted as a local founder effect. The temporary nature of the rotting cactus breeding site, the delayed maturation and the differential maturation rate of females and males are three factors proposed to cause forced outbreeding and counter the local founder effect. A combination of selection resulting from a direct interaction of the genotype and the environment, in addition to migration, is proposed to maintain the uniform gene frequencies.     

Isolation and characterization of microsatellite markers for funnel-eared bats Natalus mexicanus (Chiroptera: Natalidae) and cross-amplification using next-generation sequencing

This study presents the characterization of a series of 10 microsatellite loci developed for the funnel-eared bat Natalus mexicanus. A total of 35 pairs of primers, in 32 exemplars of N. mexicanus collected at two localities in Mexico, were selected by pyrosequencing for testing via cross-amplification in other bat species (Corynorhinus townsendii, Natalus major, Natalus tumidirostris, Nyctinomops laticaudatus and Tadarida brasiliensis). All 10 loci of this species amplified positively with medium and high resolution of genetic variation (2–15 alleles per locus, mean H_E 0.7217, mean H_O 0.6074). We consider that these polymorphic loci will be useful in genetic studies of this species as well as that of other bat families.     

Microsatellite loci for behavioural studies of rainbow bee‐eaters (Merops ornatus: Aves)

A set of polymorphic microsatellite loci was identified for population and behavioural studies of the rainbow bee‐eater (Merops ornatus). Eight loci were newly isolated from a rainbow bee‐eater genomic library enriched for GA‐containing repeats, and six loci from those previously reported for the European bee‐eater (Merops apiaster). From these two sources, the seven most variable loci that amplified reliably were unlinked and in Hardy–Weinberg equilibrium, had five to 13 alleles (mean 7.7) and an expected heterozygosity of 0.60–0.86 (mean 0.73).     

Development of microsatellite markers for the white mangrove (Laguncularia racemosa C.F. Gaertn., Combretaceae)

We developed nine new microsatellite markers for the white mangrove (Laguncularia racemosa C.F. Gaertn.), a mangrove species widely distributed in the Western Hemisphere. The loci were developed from a sample collected in Baja California Sur, Mexico, and were screened for polymorphism in 108 individuals from the Pacific Coast of Mexico and Costa Rica. The level of polymorphism was relatively low, from two to four alleles. These are the first set of polymorphic markers described for L. racemosa and should be of value in studying regional genetic differentiation among natural populations of this species.     

Endophytic bacteria in cacti seeds can improve the development of cactus seedlings

A plant–bacterium association between the giant cardon cactus Pachycereus pringlei and endophytic bacteria help seedlings establish and grow on barren rock. This cactus, together with other desert plants, is responsible for weathering ancient lava flows in the Baja California Peninsula of Mexico. When cardon seeds are inoculated with endophytic bacteria, the seedlings grow in pulverized rock for at least a year without fertilization and without showing distress. The bacteria–plant association released significant amounts of necessary nutrients from the substrate. When endophytic bacteria were eliminated from the seeds by antibiotics, development of seedlings stopped. In complementary experiments of sterile seeds inoculated with the same endophytic bacteria, plant growth was restored. This study and the previous one show that, under extreme environmental conditions, a symbiotic relationship is present between endophytic bacteria and their cactus host.     

PERMANENT GENETIC RESOURCES: Identification and characterization of 14 polymorphic microsatellite loci in the argasid tick Ornithodoros coriaceus

Fourteen polymorphic microsatellite loci (di, tetra and di‐tetra complexes) were developed for the argasid tick Ornithodoros coriaceus. Polymorphism was assessed for 56 individuals from two populations separated by ~95 km. All loci were polymorphic (X = 7, range 3–17 alleles). All loci were in Hardy–Weinberg equilibrium except for one locus (OrC 8) in a single population (P < 0.00119, after Bonferroni correction for multiple tests).