Ploidy level stability of adventitious shoots of sour cherry ‘?a?anski Rubin’ and Gisela 5 cherry rootstock

The capacity of regeneration of adventitious shoots from leaf explants was studied in sour cherry ???a?anski Rubin?? (Prunus cerasus L.) and cherry rootstock Gisela 5 (P. cerasus?×?P. canescens). Regeneration assay included thirty different combinations of plant growth regulators. 6-benzyladenine (BA) and thidiazuron (TDZ) were applied either individually or each combined with different concentrations of indole-3-butyric acid, ??-naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D). ???a?anski Rubin?? showed higher regeneration capacity in comparison with Gisela 5 regarding the total number of treatments inducing regeneration as well as the highest frequency of regeneration achieved. In both genotypes, 8.9???M BA was more effective than both 4.5 and 9.0???M TDZ in inducing adventitious regeneration, but only when combined with auxins. The highest frequency of regeneration (20.8?%) in ???a?anski Rubin?? was achieved on medium supplemented with 8.9???M BA combined with 5.4???M NAA, while in Gisela 5 the highest value (8.3?%) was obtained when BA was combined with 4.5???M 2,4-D. Flow cytometry combined with 4??-6-diamidino-2-phenylindole staining was employed to estimate DNA ploidy levels and relative nuclear DNA content in adventitious regeneration-derived shoots, in vitro shoots of axillary origin and in vivo control plants from open field. No significant differences in nuclear DNA content were detected among plants of different origin. Chromosome counting in root tip meristems also showed normal tetraploid chromosome number (2n?=?4x?=?32) in ???a?anski Rubin?? shoots and normal triploid chromosome number (2n?=?3x?=?24) in Gisela 5 shoots regenerated in vitro. The results obtained suggest that no major genetic instability occurred during adventitious regeneration under the described experimental conditions.     

Ornithodoros peruvianus Kohls, Clifford & Jones (Ixodoidea: Argasidae) in Chile: a Tentative Diagnosis

Three argasid tick larvae were collected on April 2, 2010, from a common vampire bat, Desmodus rotundus, captured in the Parque Nacional Pan de Azúcar (26°09?? S, 70°41?? W), Region of Atacama, Chile. The larvae were diagnosed as Ornithodoros, and further comparative analysis showed them to be Ornithodoros peruvianus Kohls, Clifford & Jones or a species close to it. Phylogenetic analysis based on 16S mitochondrial rRNA gene sequences of Ornithodoros species plus four Argas species was carried out to clarify the taxonomic position of the larvae. This is the first finding of ticks parasitizing D. rotundus in Chile.     

The effect of iron-chelating agents on Magnetospirillum magneticum strain AMB-1: stimulated growth and magnetosome production and improved magnetosome heating properties

The introduction of various iron-chelating agents to the Magnetospirillum magneticum strain AMB-1 bacterial growth medium stimulated the growth of M. magneticum strain AMB-1 magnetotactic bacteria and enhanced the production of magnetosomes. After 7?days of growth, the number of bacteria and the production of magnetosomes were increased in the presence of iron-chelating agents by factors of up to ??2 and ??6, respectively. The presence of iron-chelating agents also produced an increase in magnetosome size and chain length and yielded improved magnetosome heating properties. The specific absorption rate of suspensions of magnetosome chains isolated from M. magneticum strain AMB-1 magnetotactic bacteria, measured under the application of an alternating magnetic field of average field strength ??20?mT and frequency 198?kHz, increased from ??222?W/g_Fe in the absence of iron-chelating agent up to ??444?W/g_Fe in the presence of 4???M rhodamine B and to ??723?W/g_Fe in the presence of 4???M EDTA. These observations were made at an iron concentration of 20???M and iron-chelating agent concentrations below 40???M.     

Direct somatic embryogenesis from leaf and petiole explants of Spathiphyllum ‘Supreme’ and analysis of regenerants using flow cytometry

This study established a method of regenerating Spathiphyllum ??Supreme?? through direct somatic embryogenesis. Somatic embryos occurred in leaf and petiole explants cultured in the dark on a Murashige and Skoog basal medium supplemented with 2.27, 4.54, or 9.08???M N-phenyl-N??-1,2,3-thiadiazol-5-ylurea (TDZ) in combination with 1.08???M ??-naphthalene acetic acid or 2.26???M 2,4-dichlorophenoxyacetic acid (2,4-D). Explants with somatic embryos were transferred to fresh medium containing the same concentrations of growth regulators under lighted conditions for embryo conversion. The highest frequencies of leaf explants with somatic embryos and embryo conversion were both 84.4?%, which were induced by 9.08???M TDZ with 2.26???M 2,4-D. The frequencies for somatic embryo induction and embryo conversion were both 100?% when petiole explants were induced by 4.54???M TDZ with 2.26???M 2,4-D. The number of plantlets produced per leaf explant and petiole explant were as high as 67.4 and 74.4, respectively. Plantlets after transplanting to a soilless substrate grew vigorously in a shaded greenhouse. Liners were stable without phenotypic variation. Flow cytometry analysis of randomly selected plants showed that they all had a single identical peak. The mean nuclear DNA index for ??Supreme?? was 1.568, and the nuclear DNA content was 14.222?pg 2C^?1. The estimated genome size for ??Supreme?? was 6,954.5?Mbp 1C^?1 with a CV at 4.008?%. The results suggest that the regenerated plants have a stable ploidy level and this established regeneration method can be used for highly effective propagation of uniform Spathiphyllum ??Supreme??.     

A genetic linkage map of an apple rootstock progeny anchored to the Malus genome sequence

An apple rootstock progeny raised from the cross between the very dwarfing ??M.27?? and the more vigorous ??M.116?? (??M.M.106???×???M.27??) was used for the construction of a linkage map comprising a total of 324 loci: 252 previously mapped SSRs, 71 newly characterised or previously unmapped SSR loci (including 36 amplified by 33 out of the 35 novel markers reported here), and the self-incompatibility locus. The map spanned the 17 linkage groups (LG) expected for apple covering a genetic distance of 1,229.5?cM, an estimated 91% of the Malus genome. Linkage groups were well populated and, although marker density ranged from 2.3 to 6.2?cM/SSR, just 15 gaps of more than 15?cM were observed. Moreover, only 17.5% of markers displayed segregation distortion and, unsurprisingly in a semi-compatible backcross, distortion was particularly pronounced surrounding the self-incompatibility locus (S) at the bottom of LG17. DNA sequences of 273 SSR markers and the S locus, representing a total of 314 loci in this investigation, were used to anchor to the ??Golden Delicious?? genome sequence. More than 260 of these loci were located on the expected pseudo-chromosome on the ??Golden Delicious?? genome or on its homeologous pseudo-chromosome. In total, 282.4?Mbp of sequence from 142 genome sequence scaffolds of the Malus genome were anchored to the ??M.27???×???M.116?? map, providing an interface between the marker data and the underlying genome sequence. This will be exploited for the identification of genes responsible for traits of agronomic importance such as dwarfing and water use efficiency.     

Encapsulation technology for short-term storage and conservation of a woody climber, Decalepis hamiltonii Wight and Arn.

An efficient protocol was developed for short-term storage and conservation of a woody medicinal climber, Decalepis hamiltonii, using encapsulated nodal segments. The encapsulation of nodal segments was significantly affected by the concentrations of sodium alginate (Na-alginate) and calcium chloride (CaCl₂·2H₂O). A gelling matrix of 4?% Na-alginate and 100?mM CaCl₂·2H₂O was found most suitable for the production of ideal Ca-alginate beads. Maximum shoot re-growth (77.00?±?2.09?%) was recorded on Murashige and Skoog (MS) basal medium supplemented with 5.0???M 6-benzyladenine (BA), 0.5???M indole-3-acetic acid (IAA) and 30.0???M adenine-sulphate (ADS). Microshoots, recovered from encapsulated nodal segments (capsule) were best rooted on half-strength MS medium containing 2.5???M ??-naphthalene acetic acid (NAA). Complete plantlets (with shoot and root) were successfully acclimatized and established in field where they grew well without any detectable variation.     

Succession of Carrion Fauna in the Arid Region of San Juan Province, Argentina and Its Forensic Relevance

The succession of carrion fauna and the decomposition stages were studied in the arid environment of San Juan Province, Argentina (31°32??34.7?? S; 68°34??39.4?? W). Two pig carcasses (Sus scrofa) were placed in wire mesh cages, 100?m apart from each other. Each carcass was surrounded by pitfall traps, and a modified Malaise trap was placed above. Daily samplings were carried out to collect the insects present in the carcasses and the traps, and body and environmental temperature were measured. The main colonizer species was Chrysomya albiceps (Wiedemann) and its larvae were dominant over other Diptera. The first adult blowflies emerged at 8.9?days. The time to reach the remains stage was 8?days shorter than for other South American sites. We recorded the early arrival of adult Dermestes maculates De Geer and Dermestes ater De Geer 2?days after the beginning of the assay, and larvae of these species were recorded 4?days after. We determined a 1.5-day error in the postmortem interval estimation using the temperatures measured in the assay and those recorded by the nearest meteorological station.     

Development of Geocoris varius and G. proteus (Hemiptera: Geocoridae) provided with Ephestia kuehniella (Lepidoptera: Pyralidae) eggs

We studied the development of Geocoris varius (Uhler) and Geocoris proteus Distant reared on Ephestia kuehniella Zeller eggs at 20, 24, 26, 30, 33, or 36?°C. The lower developmental thresholds (T ₀) and the thermal constants (K) of eggs and nymphs of G. varius were 13.3?°C, 151.1 degree-days and 13.4?°C, 433.0 degree-days, respectively; those of G. proteus were 16.1?°C, 98.3 degree-days and 16.9?°C, 226.9 degree-days, respectively. The hatch rate of G. varius eggs was significantly lower at 33?°C than at ??30?°C, and no eggs hatched at 36?°C. That of G. proteus was lowest at 20?°C and did not decline significantly at 36?°C. The survival rate throughout the nymphal period increased with temperature up to 30?°C in G. varius, and it was lowest at 20?°C in G. proteus. Thus, the optimal rearing temperatures for immature stages appear to be about 24?C30?°C for G. varius and 26?C33?°C for G. proteus. It might be possible to improve the efficiency of their mass production by controlling the rearing temperature in the above ranges. This would also make the developmental stages of nymphs more uniform and so prevent cannibalism in mass rearing.     

Biotic and Abiotic Factors Affecting Brevicoryne brassicae (L.) (Hemiptera: Aphididae) and the Associated Hyperparasitoid Alloxysta fuscicornis Hartig (Hymenoptera: Figitidae) Morphologies

This study investigates the influence of biotic and abiotic factors associated with the morphological development of Brevicoryne brassicae (Linnaeus) and the associated hyperparasitoid Alloxysta fuscicornis (Hartig). The experiment involved the examination of whether aphid size was influenced by their vertical distribution and density on the host plant, as well as whether variations in hyperparasitoid size and symmetry were correlated with those of their aphid hosts. An aphid multivariate size index was obtained using principal component analysis, while symmetry was evaluated in terms of fluctuating asymmetry (FA). Samples were collected in 2007 on cabbage plants cultivated at an experimental farm located in Uberlandia, Minas Gerais, Brazil (18°56??54???S; 48°12??46???W). The results demonstrated that the size of B. brassicae was negatively associated with temperature, but not with its vertical distribution on the host plant. Temperature was also negatively correlated with hyperparasitoid size. During warmer periods, females produced large quantities of small-sized offspring, whereas an opposite pattern, i.e. the production of fewer offspring of larger size took place during colder periods. This type of adjustment involving trade-offs between physiological and morphological mechanisms, as well as individual interaction with abiotic environmental factors, such as temperature, can be considered an adaptive plastic response in order to increase the chances of survival at a given locality. The encountered relationship between aphid and hyperparasitoid sizes may be an after effect of their indirectly biotic interaction. Hyperparasitoid FA was dependent on the width of the mummified aphids. However, the hypothesis that temperature and vertical distribution on the host plant might influence FA was not confirmed.     

The larva of Libellula foliata (Kirby, 1889) (Odonata: Libellulidae)

The larva of Libellula foliata (Kirby) is described based upon mature larvae from the Biosphere Reserve of ??El Triunfo?? in the state of Chiapas. It belongs to the small group of species without dorsal protuberances, L. composita (Hagen), L. comanche Calvert and L. saturata Uhler. The following combination of characters permits the separation of L. foliata larva from the other aforementioned larvae: tergites 6?C10 uniformly colored, no lateral spines on segments 8?C9, 5?C6 palpal setae and 3 long premental setae. After this finding, only the larvae of Libellula gaigei Gloyd and L. nodisticta Hagen remain undiscovered for the Mexican species of Libellula.     

Effect of Low-Temperature Storage on Diaeretiella rapae (McIntosh) (Hymenoptera: Braconidae)

We evaluated the effects of constant low-temperature storage on Diaeretiella rapae (McIntosh) (Braconidae, Aphidiinae). Diaeretiella rapae mummies were stored at 5?±?1°C for 0–36 days. The percentage of D. rapae emergence varied (100-83%) after 0–32 days of storage. After 32 days, emergence reduced to 55%. According to our Probit analysis, 50% mortality (LT50) of the population of D. rapae was reached after 40 days of storage at 5°C. Storage for up to 32 days did not negatively affect emergence and survival. Cold exposure of D. rapae for 36 days did not influence morphological malformations, sex ratio, and emergence of the F1 generation. After 4–36 days of storage, D. rapae showed a gradual decrease in emergence, longevity, reproductive capacity, and F1 sex ratio. Diaeretiella rapae can be stored for up to 24 days at 5°C, at which time the percentage of parasitism and the F1 sex ratio remain above 38% and at 0.50, respectively.     

The effects of water temperature and hormone treatments on circulating LH and ovulation in tench (Tinca tinca)

The effectiveness of three hormone treatments commonly used for artificial reproduction of tench was evaluated under three thermal regimes, with focus on the need for dopamine antagonist. Mature females were divided into nine groups (n?=?8) and gradually exposed, over a 24?h period, to three temperature regimes: cold (18.1?±?0.02?°C), optimal (22.05?±?0.03?°C), and warm (26.3?±?0.01?°C). Each temperature regime comprised three experimental groups injected with one of three hormone treatments: carp pituitary extract (CPE; 3?mg?kg^?1); [D-Arg⁶, Pro⁹, NEt]-sGnRH (10???g?kg^?1); and [D-Arg⁶, Pro⁹, NEt]-sGnRH (10???g?kg^?1)?+?metoclopramide (20?mg?kg^?1) (combined treatment). No differences were found between ovulation induction (ovulation rate????75?%) with sGnRHa alone and with the combined treatment; whereas CPE at cold and warm water temperatures was significantly less effective (P?<?0.05) than above mentioned treatments. Administration of sGnRHa alone induced a gradual increase in luteinizing hormone (LH) levels with LH peaks close to ovulation, in contrast to the immediate LH surge with high LH levels throughout the entire study observed with the combined treatment under all thermal regimes. LH levels induced by GnRHa alone were significantly lower (P?<?0.05) at all temperatures compared to the combined treatment, with the exception of the final sample at 26?°C, when no difference was recorded. Based on these results we recommend the application of sGnRHa without the addition of dopamine antagonist as a reliable method for inducing ovulation in tench under suboptimal temperature conditions.     

Effect of Sex and Age on the Supercooling Point of the Winter-Active Diamesa mendotae Muttkowski (Diptera: Chironomidae)

Supercooling points (SCP) of adult Diamesa mendotae Muttkowski, 1915 (Diptera: Chironomidae) were determined at 3, 5, 9, 12 and 17 days post-collection. Supercooling points were recorded using surface contact thermometry and a cooling rate of ca. 1°C min^?1. Female SCPs averaged ?22.81, ?23.76, ?23.85, ?23.65, and ?20.87°C on each date post-collection and did not differ significantly. Male SCPs were statistically similar and averaged ?21.75, ?23.53, ?23.68, ?23.66, and ?22.92°C on each date post-collection. Paired comparisons of female/male SCPs on each date post-collection did not show significant differences over time. The overall mean SCP of D. mendotae (?23.05°C) is substantially lower than values of ?5.3°C and ?5.7°C published for adults and larvae of Belgica antarctica Jacobs 1900 collected at Palmer Station (Antarctica) and ?14.2°C for larvae of Paraclunio alaskensis Coquillett 1900 collected at Vancouver Island, British Columbia. In addition, the SCP of this species appears to be lower than that of other winter-active insect species reported in the literature. Although no adults survived after the SCP was recorded, further studies are necessary to determine if D. mendotae is a freeze-intolerant insect. Nevertheless, our results suggest that a low SCP could be used as a mechanism to facilitate emergence and adult activity of this species during winter conditions.     

Hair Mercury Concentrations of Lactating Mothers and Breastfed Infants in Iran (Fish Consumption and Mercury Exposure)

Coastal populations with high seafood consumption in the South Caspian Sea (Iran) have a significant exposure to dietary mercury. This study assesses the biomonitoring of mercury in mothers and breastfed infants in the South Caspian Sea. The mean of mercury concentration in the hair of 93 pairs of mothers and infants was obtained and was 3.55 and 1.89???g?g^?1, respectively. A statistically significant correlation (R?=?0.850, P?=?0.000) was seen between mercury concentration in the hair of mothers and infants. The results of this study indicate that hair mercury concentrations exceeded the USEPA reference dose of 1???g?g^?1 in 82.7?% of mothers and 61.2?% of infants. Also, 31?% of the mothers and 10.7?% of the infants had mercury concentrations more than the WHO ??threshold?? level (5???g?g^?1). The age and fish consumption of mothers were the factors that significantly affected the hair mercury concentration of mothers and infants. Number of dental amalgam fillings of mothers was the factor that only affected mercury in the hair of mothers. According to the results, we can conclude that the main determinant of mercury exposure was the intake of mercury through fish consumption of mothers.     

A forecasting model for the adult emergence of overwintered Monochamus alternatus (Coleoptera: Cerambycidae) larvae based on degree-days in Korea

Emergence data from 1999 to 2011 were analyzed to develop a forecasting model of the adult emergence of overwintered Monochamus alternatus Hope (Coleoptera: Cerambycidae) larvae. The cumulative emergence curves of M. alternatus adults showed high variation year to year and were fitted by the Weibull function to obtain an all-combined preliminary model that was then used to separate fractional models. The emergence curves were grouped into three fractional modes—advance, regular, and delay—based on the number of discrepant days between actual and predicted date by the preliminary model at 50 % emergence date (predicted ? observed), resulting in the mean number of discrepant days of 8.7, ?0.2, and ?8.0, respectively. Three assumptions were made to support the grouping: delay mode is selected when the third instars prevails in overwintering stages; advance mode is selected when the fourth instars prevail, since fourth instars immediately can go into pupal stage; regular mode is located between advance and delay mode. These three modes produced a 340, 420, and 500 degree-days (DDs) at 50 % emergence date, respectively, when each was fitted by the Weibull function, with DDs based on 11.9 °C from 1 January. The emergence models developed in this study considerably described the actual population phenology of M. alternatus, with the selection of advance, regular, and delay modes. Consequently, the models should be useful in establishing management strategies against M. alternatus in Korean forests.     

Temperature-dependent development of diapausing larvae of Chilo partellus (Swinhoe) (Lepidoptera: Crambidae)

Temperature-dependent development rate, percent diapause induction (hibernation at low temperature and aestivation at high temperature), and survival of diapausing larvae of Chilo partellus (Swinhoe, 1885) were examined on 13 constant temperatures ranging from 8 to 40 °C. Development of hibernating and aestivating larvae occurred from 10 to 25 °C and 27–38 °C, respectively. However, no development occurred at 8 °C and 40 °C. To determine actual thermal conditions that affect development and trigger both kind of diapause (hibernation and aestivation), various thermal parameters were estimated by fitting the development rate data to two linear (Ordinary equation and Ikemoto & Takai) models and thirteen non-linear models. The lower thermal thresholds (T_min) for development of diapausing larvae of C. partellus were calculated as 9.60 °C and 10.29 °C using the ordinary linear model and Ikemoto & Takai model, respectively. Similarly, the thermal constants (K) estimated using the ordinary linear model was 333.33 degree-days and that estimated with Ikemoto & Takai model was 338.92 degree-days. Among the non-linear models, Lactin-2 followed by Lactin-1 were found to be the best as these models estimated the critical temperatures (T_min, T_max and T_opt) similar to those of observed values. Conclusively, the Ikemoto & Takai linear model and Lactin-2 followed by Lactin-1 non-linear models are useful and efficient for describing temperature-dependent development and estimating the temperature thresholds of diapausing larvae of C. partellus. Our findings provided fundamental information for estimation of thermal requirement and temperature based development models for diapausing larvae of C. partellus. This information will be highly useful for predicting the occurrence, seasonal emergence, number of generations and population dynamics of C. partellus.     

Environmental induction of larval diapause and life-history consequences of post-diapause development in the Large Copper butterfly,Lycaena dispar (Lepidoptera: Lycaenidae)

Many insects in temperate zones withstand the adverse conditions of winter through entering diapause and the two most important environmental stimuli that induce diapause are photoperiod and ambient temperature. The Large Copper butterfly, Lycaena dispar Haworth (Lepidoptera: Lycaenidae), is a Palearctic butterfly that hibernates as larvae. Since this butterfly is a near threatened species in some regions, there has been a growing need for a standardized protocol for mass rearing of this butterfly based on the adequate knowledge of its ecology. In the present study, we first identified that L. dispar larvae were sensitive to the photoperiodic induction of diapause during their first larval instar. We then investigated to what extent the diapause-inducing effects of photoperiod could be modified by ambient temperatures in L. dispar larvae by exposing them to the range of day-lengths (L:D 14:10, 12:12, 10:14 and 8:16) at three different temperatures (15, 20 and 25 °C). All larvae were induced to enter diapause at low ambient temperature (15 °C) regardless of photoperiod, whereas most of them (86 %) exhibited direct development when temperature was high (25 °C). The photoperiodic induction of diapause was evident when day-length was shorter than 14 h at intermediate temperature (20 °C). Pre-diapause development was prolonged at low temperatures. Finally, we found that post-diapause development of L. dispar larvae was determined by both the chilling temperature experienced by diapausing larvae and the duration of larval diapause. Adult emergence was enhanced when larvae were chilled at 8 °C and when they had been under the state of diapause for 20 days before they were treated to terminate diapause.     

Susceptibility of the cigarette beetle Lasioderma serricorne (Coleoptera: Anobiidae) to hypoxia

The susceptibility of the cigarette beetle Lasioderma serricorne (F.) to hypoxia was examined at three different oxygen concentrations (0.5?C0.8, 1.0?C1.3, and 2.0?C2.3?%) and four different temperature/humidity (RH) conditions: 30?°C/75?% RH, 25?°C/75?% RH, 20?°C/43?% RH, and 15?°C/43?% RH. The influence of humidity on mortality was also examined at three humidity levels (21, 43, and 75?% RH) at 1.0?C1.3?% oxygen (O₂) and 25?°C. Our results revealed that adult beetles were the most tolerant at 2.0?C2.3?% O₂ and that the larvae were the most tolerant at O₂ levels <1.0?C1.3?%. Mortality increased with increasing temperatures and decreasing O₂ concentrations. At 30?°C, 75?% RH, and 0.5?C0.8?% O₂, the 99?% lethality (LT₉₉) of larvae was 6.9?days; however, it increased to 20?days when the temperature was decreased to 25?°C or when O₂ levels were increased to 1.0?C1.3?%. Humidity also influenced mortality of both larval and adult beetles. LT₉₉ values for larvae at 25?°C and 1.0?C1.3?% O₂ were 24.0, 44.6, and 50.2?days at 21, 43, and 75?% RH, respectively. Results of this study indicate that a controlled atmosphere (CA) with reduced oxygen levels (<0.5?C0.8?% O₂) represents an effective measure for disinfesting stored tobacco as an alternative to conventional phosphine fumigation at temperatures >30?°C.     

Occurrence of Stiretrus decastigmus (Herrich-Schaeffer) (Hemiptera: Pentatomidae) Preying on Microtheca semilaevis Stal (Coleoptera: Chrysomelidae)

Microtheca semilaevis Stal is a phythophagous insect associated to Brassicaceae, and frequently occurs with other species of beetles, such as Microtheca ochroloma Stal. Despite its importance to organic farmers, there is no information in the Brazilian literature about its natural enemies. Stiretrus decastigmus (Herrich-Schaeffer) was first found and here reported preying an adult of M. semilaevis in Chinese cabbage in Santa Maria, RS, Brazil (29°43??28??S, 53°43??19??W).     

Occurrence and distribution of plankton-associated and free-living toxigenic Vibrio cholerae in a tropical estuary of a cholera endemic zone

Cholera epidemics are thought to be influenced by changes in populations of estuarine Vibrio cholerae. We investigated the abundance and distribution of this bacterium, as ??free-living?? (<20???m fraction) and associated with microphytoplankton (>20???m) or zooplankton (>60???m), in the Karnaphuli estuary of Bangladesh during pre- and post-monsoon seasons. Cultivable Vibrio populations were ~10²?C10⁴ colony forming units (CFU) ml^?1 in the high saline zone (19?C23 practical salinity unit, PSU) and declined in freshwater (<10¹?CFU?ml^?1). Culture independent detection of toxigenic V. cholerae O1 and O139 serogroups revealed a higher abundance of ??free-living?? (10⁴?C10⁵ cells?l^?1) than those attached to plankton (10¹?C10³ cells?l^?1). However, ??free-living?? O1 and O139 cells were sometimes absent in the medium saline and freshwater areas (0.0?C11 practical salinity unit [PSU]). In contrast, plankton samples always harbored these serogroups despite changes in salinity and other physico-chemical properties. Microphytoplankton and zooplankton were dominated by diatoms and blue-green algae, and copepods and rotifers, respectively. Toxigenic V. cholerae abundance did not correlate with plankton abundance or species but had a positive correlation with chitin in the <20???m fraction, where suspended particulate matter (SPM), V. cholerae and chitin concentrations were highest. C:N ratios indicated that organic matter in SPM originated predominantly from plankton. The differential occurrence of ??free-living?? and attached V. cholerae suggests a pivotal function of plankton in V. cholerae spreading into freshwater areas. The probable association of this pathogen with organisms and particles in the nanoplankton (<20???m) fraction requires validation of the concept of the ??free living?? state of V. cholerae in aquatic habitats.