Migratory flexibility in native Hawai'ian amphidromous fishes

We assessed the prevalence of life history variation across four of the five native amphidromous Hawai'ian gobioids to determine whether some or all exhibit evidence of partial migration. Analysis of otolith Sr.: Ca concentrations affirmed that all are amphidromous and revealed evidence of partial migration in three of the four species. We found that 25% of Lentipes concolor (n = 8), 40% of Eleotris sandwicensis (n = 20) and 29% of Stenogobius hawaiiensis (n = 24) did not exhibit a migratory life-history. In contrast, all individuals of Sicyopterus stimpsoni (n = 55) included in the study went to sea as larvae. Lentipes concolor exhibited the shortest mean larval duration (LD) at 87 days, successively followed by E. sandwicensis (mean LD = 102 days), S. hawaiiensis (mean LD = 114 days) and S. stimpsoni (mean LD = 120 days). These findings offer a fresh perspective on migratory life histories that can help improve efforts to conserve and protect all of these and other at-risk amphidromous species that are subject to escalating anthropogenic pressures in both freshwater and marine environments.     

Molecular identification of two species of myiasis-causing Cuterebra by multiplex PCR and RFLP

The myiasis-causing flies Cuterebra grisea (Coquillet) and Cuterebra fontinella (Clark) (Diptera: Oestridae) are normally parasites of mice, predominantly of the genus Peromyscus. The morphological similarities of these species and the existence of intermediate morphotypes bearing characters of both species make the identification of adults problematic; furthermore the identification of larvae is apparently not possible. This study presents two molecular approaches to discriminate between these species using specific band patterns: (i) species-specific primers designed in the cytochrome oxidase II (COII) region used in multiplex polymerase chain reaction (PCR) and (ii) restriction fragment length polymorphism (RFLP) on amplified segments of cytochrome oxidase I (COI) gene. Both methods were tested on Cuterebra larvae and on adult museum specimens. The two techniques showed a clear difference between C. grisea and C. fontinella, although species-specific primers were more successful than RFLP for degraded DNA. No intraspecific variation in RFLP and species-specific amplifications were detected for the two species of Cuterebra. The results exhibit discrepancies between molecular and morphological identification, suggesting that some of the adults were misidentified.     

PCR–RFLP analysis of mitochondrial DNA in tench Tinca tinca

Polymorphism was detected at ND1, ND6, D‐loop and cyt b segments of mtDNA in 105 tench (Tinca tinca L.), using the polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) technique and five composite haplotypes were identified. The diversity indices and the results of the population comparisons revealed that the identified markers provide a powerful tool for further studies on this species.     

Survival and behavioral characteristics of amphidromous goby larvae of Sicyopterus japonicus (Tanaka, 1909) during their downstream migration

To understand the ecology and environmental tolerances of newly hatched larvae of the amphidromous fish Sicyopterus japonicus during their downstream migration, the salinity tolerance of eggs, 0-15 day old larvae, and adults, and the temperature tolerance, specific gravity and phototaxis of hatched larvae were examined. Tolerances of adults were measured as survival after a 24 h challenge in freshwater (FW), brackish water (1/3 SW) and seawater (SW). The survival rate of adult S. japonicus was 100% in FW and 1/3 SW, while none survived in SW. Hatching success of eggs (30 eggs each) was significantly higher in FW (mean: 73%) and 1/3 SW (73%) than in SW (19%). Tolerance of newly hatched larvae to salinity and temperature was investigated in different combinations of salinities (FW, 1/3 SW and SW) and temperatures (18, 23 and 28 °C). Larval survival was significantly different in each salinity and temperature. Survival rate was significantly higher in 1/3 SW than in FW and higher in SW than in FW at 23 °C and 28 °C. At the latter part of the experiment, there was no survival in FW and at 28 °C. Survival was higher in lower temperatures, but larval development did not occur in FW. Specific gravity of newly hatched larvae was 1.036 at 28 °C and 1.034 at 23 °C. When exposed to a light source on one side of an aquarium, larval distribution was not affected. Our results indicated larval S. japonicus are more adapted to brackish water and seawater than freshwater, while the adults and eggs are more adapted to freshwater and brackish water than seawater. This is consistent with their amphidromous life history with growth and spawning occurring in freshwater and the larval stage utilizing marine habitats.     

Coloration and ontogenetic features of fluviatile species of <Emphasis Type="Italic">Rhinogobius</Emphasis> (Gobioidei: Gobiidae) in Amami-oshima Island,Ryukyu Islands,Japan

 The fluviatile Rhinogobius sp. 1 (hereinafter R. sp. 1) (Japanese name kibara-yoshinobori or English name yellow belly medium-egg type) is broadly distributed in the Ryukyu Islands ranging from Amami-oshima Island to Iriomote Island. However, based on the molecular evidence, this species has probably had multiple origins from amphidromous species. We compared the coloration of specimens at Amami-oshima I. with that of the other specimens (islands) in the Ryukyu Is. Amami-oshima I. specimens of R. sp. 1 were different from the specimens of other populations in having five to six cross lines on the caudal fin. We examined the early life history of fluviatile Rhinogobius at Amami-oshima I., based on rearing experiments, and estimated the age of wild specimens by counting otolith increments. Spawned egg capsules of “Rhinogobius sp. 1” at Amami-oshima I. were larger than those of amphidromous species. Their developmental stage in newly hatched larvae was similar to that of Okinawan R. sp. 1 in having a straight notochord; however, it had a flexed notochord earlier than that of Rhinogobius sp. 2 (hereinafter R. sp. 2) (aobara-yoshinobori or blue belly medium-egg type). The ontogenetic trajectory of “Rhinogobius sp. 1” in Amami-oshima I. was similar to Okinawan R. sp. 1 (30 days in larval duration) but different from R. sp. 2 (15 days). In amphidromous relatives of R. sp. 2, developmental timing during larval period was earlier than in the amphidromous relatives of R. sp. 1. It has been suggested that the phenomenon of fluvial landlocking has been accompanied somewhat by egg size increase (on the basis of a few examples). However, the ontogenetic trajectories of fluviatile Rhinogobius species may be a reflection of those of ancestral amphidromous species. Received: March 18, 2002 / Revised: November 7, 2002 / Accepted: December 10, 2002     

Spatiotemporal patterns of research on Southern Hemisphere amphidromous galaxiids: A semi–quantitative review

Amphidromy is a distinctive life-history strategy of some fish species that involves spawning in fresh or brackish water followed by dispersal to sea by newly hatched larvae, where they develop for a short period. Individuals then return to freshwater as juveniles, where they feed and grow, before maturing and spawning. Six amphidromous species from the Southern Hemisphere genus Galaxias (G. truttaceus, G. fasciatus, G. argenteus, G. postvectis, G. brevipinnis, G. maculatus) are recreationally, culturally, and economically important as the juveniles are harvested. Due to ongoing population declines and a lack of critical demographic information, there is growing concern about the management of the species. Here, we used semi-quantitative review, culturomics, and bibliometric tools to analyse peer-reviewed research conducted on the six amphidromous species of Galaxias to: (i) understand how spatiotemporal patterns of research have shifted over the last five decades, and (ii) identify critical research gaps. Forty percent of studies (n = 295) covered a spatial extent of 10 km or less and 87% of studies lasted less than 2 years – studies were largely small and short relative to the species' ranges and their longevity. Additionally, we found important research gaps; for example, studies on the effects of climate change and the associated effects of disturbance, and the marine phase are scarce in the peer-reviewed literature. Finally, we suggest that quantitative models have been underutilized as tools for studying amphidromous galaxiids and should be embraced to answer questions not readily addressed with field and laboratory-based techniques. If these species are to be effectively managed, their population dynamics across spatiotemporal scales must be understood and critical and long-standing gaps in research knowledge addressed.     

Riverine life history of the amphidromous goby Sicyopterus japonicus (Gobiidae: Sicydiinae) in the Ota River, Wakayama, Japan

To understand the riverine life history of the amphidromous goby, Sicyopterus japonicus, studies were conducted in the Ota River, Wakayama, Japan. They were distributed from 3 to 23 km upstream from the river mouth, and abundance was higher in the middle reaches than in the upper and lower reaches. Fish were not observed in rapids during winter, suggesting a seasonal change of habitat. Body length ranged from 24 to 120 mm SL. Both females and males ranged from 1 to 6 years old, and males had larger asymptotic length than females. Condition factor showed two peaks in July and November, which appeared to correspond to the spawning season and preparation for over wintering. Gonad somatic index increased in summer with a peak in August indicating a summertime spawning season that was confirmed by collections of the newly hatched larvae migrating downstream. Eggs of 0.5 mm diameter were attached to stones and newly hatched larvae made continuous vertical movements of sinking and upward swimming and the distance of each movement was longer in freshwater than in seawater. The newly hatched larvae were collected at nighttime (19:00–24:00) mainly in August. Those larvae were very small (mean: 1.4 mm TL) with a yolk sac and no eye pigmentation. Low wintertime temperatures are likely an important determinant of the spawning and recruitment seasons and seasonal changes of activity of this species that lives at much higher latitudes than all other species of the subfamily.     

Research Note: Simple molecular discrimination of cultivated Porphyra species (Porphyra yezoensis and Porphyra tenera) and related wild species (Bangiales,Rhodophyta)

To discriminate between cultivated Porphyra species (Porphyra yezoensis and Porphyra tenera) and closely related wild Porphyra species, we developed a polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) analysis of the rbcL gene using five restriction enzymes. Although our previous PCR‐RFLP analyses of internal transcribed spacer (ITS) rDNA and plastid RuBisCO spacer regions could not always discriminate wild P. yezoensis, wild P. tenera, and closely related wild species, the PCR‐RFLP profiles of the rbcL gene were useful in discriminating samples collected from natural habitats. Therefore, PCR‐RFLP analysis of the rbcL gene will help in the simple identification of a large number of samples, not only for the establishment of reliable cultures as breeding material, but also for the taxonomic investigations of species that are closely related to cultivated Porphyra.     

Simultaneous maturation of two discrete oocyte batches in ayu

Ayu Plecoglossus altivelis including amphidromous and landlocked populations, are known for batch spawning. Unusual clutches at ovulation were observed from rearing stocks that were naturally induced to sexual maturation. Two peaks occurred simultaneously in the frequency distribution of egg size, compared to normal fish, which have clutches with unimodal size distribution. The eggs in the small size class could be fertilized, and small-sized larvae hatched from these eggs. To explain the alteration in egg size through divergence from amphidromous to landlocked populations, the role of batch allocation among oocytes is discussed.     

Simultaneous identification of Grapholita molesta Busck and Grapholita dimorpha Komai by PCR‐RFLP

Although several molecular diagnostic techniques are available for the identification of the apple‐feeding pests Grapholita molesta Busck and Grapholita dimorpha Komai, these pests are severely affecting apple orchards in Korea. These two pests may be misidentified or the available molecular diagnostic techniques may not facilitate the simultaneous identification of the morphological features of both species. In this study, we developed a multiplex assay for these two species using the polymerase chain reaction – restriction fragment length polymorphism (PCR‐RFLP) method. Sixty‐two specimens were collected from apples presumed infested with moth larvae and from pheromone traps from 2013 to 2014. Both species were identified morphologically, and a partial region of the cytochrome b gene was sequenced to design primers for PCR‐RFLP. Digestion profiles of G. molesta and G. dimorpha, using the Sau3A1 restriction enzyme, were characterized using three DNA fragments each for G. molesta (363 bp, 91 bp and 31 bp) and G. dimorpha (220 bp, 234 bp and 31 bp). The RFLP assay developed for both species in this study was more efficient and accurate than other currently used diagnostic assays and would be helpful to identify field‐collected specimens for pest control research.     

Molecular Species Identification of Spiny Lobster Phyllosoma Larvae of the Genus Panulirus from the Northwestern Pacific

To identify lobster phyllosoma larvae of the genus Panulirus occurring in waters adjacent to Japan, genetic variation within and between 10 Indo-Pacific lobster species was investigated using restriction fragment length polymorphism (RFLP) analysis for the 1300-base pair mitochondrial cytochrome oxidase I (COI) gene. RFLP analysis using two endonucleases (AluI and TaqI) enabled discrimination of all species, including the P. longipes complex. The diagnostic DNA markers, supplemented with nucleotide sequence analysis, were applied to 44 mid- to late-stage phyllosoma larvae (7.4 to 27.7 mm in body length) collected in the northwestern Pacific. These samples were unexpectedly variable in species composition, comprising P. japonicus (n = 16), P. longipes bispinosus (21), P. longipes longipes (1), P. “aka” (1), and P. penicillatus (5). Comparison of larval size at similar stages revealed that P. l. bispinosus larvae were significantly larger than P. japonicus.     

Why be amphidromous: expatrial dispersal and the place of source and sink population dynamics?

Amphidromous fishes are found predominantly on the tropical and subtropical islands of the globe and there are few amphidromous species on continents. I suggest that this idiosyncratic distribution relates in part to problems in self-recruitment on islands that are often young or volcanic, and which may have streams with ephemeral flows across relatively short times scales. Amphidromy provides the ability to invade new habitats as these become available either on newly emergent (often volcanic) islands, or following perturbation after stream dewatering or the impacts of volcanism on older islands as a consequence of expatrial dispersal. Source/sink population dynamics may also be involved with islands ‘downstream’ in oceanic current systems behaving as sinks, with little or no self-recruitment. Streams in steep topography seem to be favoured by amphidromous species, perhaps because they provide more rapid transport to sea of the tiny, newly hatched larvae.     

Cytochrome-<Emphasis Type="Italic">b</Emphasis> variation in <Emphasis Type="Italic">Apis mellifera</Emphasis> samples and its association with COI–COII patterns

Five Mbo I (Mbo-A, Mbo-M, Mbo-C¹, Mbo-C² and Mbo-C³) and Hinf I (Hinf-1 to Hinf-5) patterns were observed in Apis mellifera samples after restriction of a 485 bp fragment of the mitochondrial cytochrome-b (cyt-b) gene. Associating the cyt-b Restriction fragment length polymorphism (RFLP) pattern of each sample to its respective previously established COI–COII (Dra I sites) pattern, five restriction patterns (Mbo-C¹, Mbo-C², Mbo-C³, Hinf-1 and Hinf-4) were observed in samples of maternal origin associated to the evolutionary branch C. No deletions or insertions were observed and the nucleotide substitution rate was estimated at 5.4%. Higher nucleotide diversity was observed among the branch C-haplotypes when compared with A and M lineages. Further studies are needed to confirm if the cyt-b + COI–COII haplotypes help to assign certain phylogeographic patterns to the branch C and to clarify phylogenetic relationships among A. mellifera subspecies.     

PCR–RFLP assays for discerning three weevil stem feeders (Ceutorhynchus spp.) (Col.: Curculionidae) on garlic mustard (Alliaria petiolata)

Polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) assays were developed to rapidly and cost-effectively differentiate the morphologically indistinguishable larval stages of three species, Ceutorhynchus alliariae, C. roberti and C. scrobicollis, proposed for the biological control of Alliaria petiolata. A PCR–RFLP using TaqαI restriction enzyme can differentiate the three target species only, while a PCR–RFLP double digestion using the restriction enzymes AluI and TaqαI should enable, based on virtual digestion, distinguishing them from 12 other Ceutorhynchus species associated with A. petiolata.     

Chloroplast DNA inheritance and variation in <Emphasis Type="Italic">Leucadendron</Emphasis> species (Proteaceae) as revealed by PCR-RFLP

The inheritance of chloroplast DNA (cpDNA) in Leucadendron species was studied by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) analysis. A total of 100 progeny from five interspecific crosses involving seven parental species were tested, and all progeny exhibited the cpDNA restriction fragment pattern of the female parent, indicating that cpDNA in Leucadendron is maternally inherited. PCR-RFLP was also employed to study cpDNA variation among 21 Leucadendron species. Parsimony analysis using a heuristic search resulted in a phylogenetic tree that showed limited agreement to the taxonomic classification of Leucadendron species, based on morphological characteristics. The incongruence between cpDNA phylogenetic and taxonomic groupings in Leucadendron may be due to reticulate evolution involving a combination of hybridization and introgression, convergent evolution and/or lineage sorting at the interspecific, intersubsectional and intersectional levels.     

Occurrence of additional Zoea-VI larvae in the mud crab, Scylla paramamosain (Estampador), reared in the laboratory

Mud crabs, Scylla spp. , are commercially important in many Indo-Pacific countries. The larval development of mud crabs has been reported previously as five zoeal and one megalopal stages. This paper reports larval rearing experiments that revealed variability in larval developmental stages in the mud crab Scylla paramamosain, one of four mud crab species. In addition to normal five zoeal stages, an alternative pathway of developing through six zoeal stages was observed for the crab. There were evidences suggested that the appearance of the additional Zoea-VI larvae was associated with unfavourable dietary conditions, including poor quality of diet, inadequate quantity of dietary supply and a period of starvation for newly hatched larvae. Based on exuviae and larval specimens, the morphology of the additional Zoea-VI larvae was described.     

Effect of saline water on early success of amphidromous fish

Organisms that migrate between rivers and the sea inevitably pass through estuarine habitats. Despite the potential importance of salinity and temperature fluctuations for metabolic adaptation, little is known about the impact of environmental changes in estuaries on the survival of residents. Ayu (Plecoglossus altivelis) is a migratory fish that inhabits estuarine brackish water in its early life stages. The recent decline in the abundance of populations ascending into rivers is of concern for local biodiversity. The present study aims to elucidate the ecological processes that determine the early success of Ayu larvae under variable environmental conditions. The effects of salinity and water temperature on the endogenous growth of newly hatched larvae from the same brood were examined experimentally based on morphological and metabolic characteristics. High salinity and high water temperature together appeared to require more energy for larval osmoregulation, resulting in the acceleration of yolk depletion and reduced growth of the notochord. Increasing the osmoregulative cost during the yolk-sac stage resulted in the faster induction of a state of starvation. Seawater is considered to have an adverse effect on the survival of newly hatched larvae, as it lowers the efficiency of foraging and predator avoidance due to an energetic tradeoff. More attention should be paid to the significance of estuarine environments involving brackish waters to ensure the early survival of amphidromous fish such as Ayu.     

Stable Isotope Analysis of Amphidromous Hawaiian Gobies Suggests Their Larvae Spend a Substantial Period of Time in Freshwater River Plumes

Synopsis We employed stable isotope analysis (δ¹³C, δ¹⁵N) to evaluate the sources of nutrients used by amphidromous gobiid fishes (Lentipes concolor, Sicyopterus stimpsoni, Awaous guamensis) caught migrating into and living in Hakalau Stream, Hawaii. Although considerable variation amongst the stable isotope values of stream items was noted across all 4 years of our study, the relationships between the fishes were relatively constant. Stable isotope values of recruiting gobies were consistently closer to those of both inshore plankton and freshwater adults than those of offshore plankton, suggesting that the larvae of these species derive much of their nutrition from inshore environments influenced by fresh water. Small differences between the stable values of these species further suggested that their larvae come from different inshore locations. After entering fresh water all species appear to swim rapidly upstream without feeding. Finally, once well upstream, adult L. concolor and A. guamensis appear to assume an omnivorous diet while adult S. stimpsoni rely upon autochthonous production within streams. We propose that freshwater food webs play an integral yet complex role in the lives of both larval and adult amphidromous Hawaiian fishes.     

Presence of Anopheles culicifacies B in Cambodia established by the PCR-RFLP assay developed for the identification of Anopheles minimus species A and C and four related species

Abstract A polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) assay developed for identification of five species of the Anopheles minimus Theobald group and a related mosquito species of the Myzomyia Series (Diptera: Culicidae) was applied to morphologically identified adult female specimens collected in Ratanakiri Province, north‐eastern Cambodia. In addition to finding An. aconitus Dönitz, An. minimus species A and An. pampanai Büttiker & Beales, some specimens showed a new restriction banding pattern. Siblings of specimens that exhibited this new PCR‐RFLP pattern were morphologically identified as An. culicifacies James sensu lato. Based on nucleotide sequences of the ribonuclear DNA internal transcribed spacer 2 region (ITS2) and the mitochondrial cytochrome oxidase I gene (COI), these specimens were recognized as An. culicifacies species B (sensu Green & Miles, 1980 ), the first confirmed record of the An. culicifacies complex from Cambodia. This study shows that the PCR‐RFLP assay can detect species not included in the initial set‐up and is capable of identifying at least seven species of the Myzomyia Series, allowing better definition of those malaria vector and non‐vector anophelines in South‐east Asia.     

Assessment of the degree of restriction fragment length polymorphism in Brassica

Summary The feasibility of creating a restriction fragment length polymorphism (RFLP) linkage map in Brassica species was assessed by screening EcoRI-, HindIII-, or EcoRV-digested total genomic DNA from several accessions of B. campestris, B. oleracea, and B. napus using random genomic DNA clones from three Brassica libraries as hybridization probes. Differences in restriction fragment hybridization patterns occurred at frequencies of 95% for comparisons of accessions among species, 79% for comparisons of accessions among subspecies within species, and 70% for comparisons among accessions within subspecies. In addition, species differences in the level of hybridization were noted for some clones. The high degree of polymorphism found even among closely related Brassica accessions indicates that RFLP analysis will be a very useful tool in genetic, taxonomic, and evolutionary studies of the Brassica genus. Development of RFLP linkage maps is now in progress.