Gut microflora of two saltmarsh detritivore thalassinid prawns,Upogebia africana andCallianassa kraussi

The presence and digestive capabilities of bacteria associated with the digestive systems and habitats of two saltmarsh-burrowing detritivore thalassinid prawns (Upogebia africana andCallianassa kraussi) was examined.U. africana is a filter-feeding prawn inhabiting muddy deposits, whereasC. kraussi, a deposit feeder, inhabits coarser more sandy deposits. Scanning electron microscopy was used to examine the gut lining and associated microflora and the nature of the ingested food of both prawns. The gut contents of both prawns included plant fragments, fragmented diatoms, partially degraded protozoa, and bacteria attached to organic matter. In bothU. africana andC. kraussi the midgut walls and gut contents were extensively coated by filamentous bacteria which were absent in the hindgut. The hindgut epithelium ofU. africana was coated by mats of rodshaped bacteria, not reported in marine invertebrates previously. The digestive glands of both species contained bacteria in the lumen. Isolation of gut and habitat bacteria suggests that bothU. africana andC. kraussi maintain a gut microflora distinct from the habitat microflora. Bacteria isolated from the guts of both species of prawn differed from those isolated from their respective habitats with regards to both the genera isolated and their digestive capabilities. The dominant genera isolated from the guts of bothU. africana andC. kraussi wereVibrio andPseudomonas, with an unidentified fermenter andPseudomonas, respectively dominating in the digestive glands. Bacteria of the genusAcinetobacter dominated the isolates from the habitats of both species of prawn. Resident gut bacteria isolated from the guts of both species of prawn exhibited lipase, protease, chitinase, and lysozyme, but not cellulase activity, and may contribute to nitrogen aquisition by the prawns. Isolates from the prawns' habitat exhibited alginase, gelatinase, and lipase activity, a few (3%) fromU. africana habitat having cellulases. In this study a distinction between resident gut bacteria and transient gut bacteria was made. Results suggest that some habitat bacteria remain viable in the guts ofU. africana, but not inC. kraussi.     

News & Notes: Isolation of Vibrio harveyi from Diseased Kuruma Prawns Penaeus japonicus

Outbreaks of high mortality among the cultured kuruma prawn Penaeus japonicus without overt gross signs occurred during August and December of 1994 in I-Lan, Taiwan. Eleven luminous bacterial strains were isolated from the hepatopancreas of moribund prawns from five different farms by use of tryptic soy agar (TSA, supplemented with 2% NaCl) and/or thiosulfate citrate bile salt sucrose (TCBS) agar. These strains, together with our two previously unpublished isolates, were characterized and identified to be Vibrio harveyi in comparison with two ATCC Type strains and one strain previously isolated from the tiger prawn, P. monodon. Received: 22 November 1995 / Accepted: 24 February 1996     

The relative abundance and seawater requirements of gram-positive bacteria in near-shore tropical marine samples

The relative abundance of gram-positive bacteria in a variety of near-shore marine samples was determined using the KOH method. Gram-positive bacteria accounted for 14%, 25%, 31 %, and 12%, respectively, of the colony-forming bacteria obtained from seawater, sediments, and the surfaces of algae and invertebrates. A total of 481 gram-positive strains were isolated representing a wide range of morphological groups including regular and irregular rods, cocci, and actinomycetes. Seventy-seven percent of the strains characterized did not form spores and were aerobic, catalase-positive rods with regular to irregular cell morphologies. Eighty-two percent of the strains tested showed an obligate requirement of seawater for growth. None of the cocci tested required seawater or sodium for growth. This is the first report documenting that gram-positive bacteria can compose a large percentage of the culturable, heterotrophic bacteria associated with the surfaces of tropical marine algae. Correspondence to: P.R. Jensen     

基于PCR-DGGE基因指纹的对虾体内优势细菌组成分析

采用不依赖分离培养的16S rDNA的PCR-DGGE基因指纹技术对刀额新对虾与中国对虾的鳃部与肠道优势细菌种群组成进行比较分析。研究发现:对虾鳃部与肠道存在着丰富多样的细菌;根据DGGE指纹图的聚类分析发现不同对虾及同一种对虾的鳃部与肠道内的细菌组成差异性非常大;同时也发现不同对虾体内有相同的细菌存在。首次尝试建立基于16S rDNA的PCR-DGGE基因指纹的对虾体内细菌组成揭示方法,对于今后建立对虾与养殖水体微生物和相关疾病的关系具有重要意义。     

Distribution and Physiology of Aerobic Bacteria Containing Bacteriochlorophyll a on the East and West Coasts of Australia

Aerobic heterotrophic bacteria containing bacteriochlorophyll were isolated from specimens from a wide variety of marine environments on the west (Shark Bay, Lake Clifton, Lake Heyward, and Perth) and east (near Townsville and Brisbane) coasts of Australia. The bacteria were found in a high proportion (10 to 30%) of the total heterotrophic bacterial strains isolated from marine algae, seagrasses, stromatolites, the epiphytes on stromatolites, seawater, and sands; in some cases they constituted up to 49% of the total. This is much higher than the previous report of 6% from Japan. A high percentage, 13%, was also found in the seawater of Hamelin Pool, at Shark Bay, where the salinity was 66%. The number of these bacteria was generally low in seawater and sands, with a few exceptions. There were no aerobic bacteriochlorophyll-containing bacteria on sponges or corals. The isolated strains were orange or pink, and most had absorption maxima around 800 and 850 to 870 nm, the latter range being the absorption of bacteriochlorophyll a in vivo. The maximum bacteriochlorophyll content was 1 nmol/mg (dry weight) of bacterial cells. Most of the bacteria did not grow phototrophically under anaerobic conditions in a broth medium containing succinate. Cells and cell extracts grown under aerobic conditions had photochemical activities such as reversible photooxidations of the reaction center and cytochrome(s). Some strains showed denitrifying activity. The optimal salinity for bacterial growth varied between strains.     

In vivo humoral and cellular reactions, and fate of injected bacteria Aeromonas hydrophila in freshwater prawn Macrobrachium rosenbergii

Live non-opsonized and opsonized Aeromonas hydrophila were injected into juveniles of freshwater prawn Macrobrachium rosenbergii to study the cells involved in phagocytosis, distribution of bacteria, cellular reactions and clearance of both forms of bacteria from the system. The bacteria were rapidly distributed to various tissues viz., gills, heart, hepatopancreas within 1h, and the tissues revealed haemocytic nodule formation after 3 h of injection. There was rapid clearance of both the forms of bacteria from the circulation. However, clearance efficiency was significantly (P < 0.05) faster in the case of opsonized bacteria at 12 h after injection. Similarly, the nodule formation, that was prominent in cardiac musculature, was rapidly eliminated from the tissues of the group injected with opsonized bacteria as compared to non-opsonized bacteria injected group, thus confirming the existence of opsonic factors in haemolymph of this prawn. In another experiment, various dose levels of bacteria were injected intramuscularly into prawns and haemolymph was collected after 1, 6, 24, 72 h and 7 days of injection to study various immune parameters. Although, no major alterations in the total and differential haemocyte counts were observed in bacteria injected prawns compared to control, there was a significant decline in phenoloxidase activity in the highest dose bacteria injected group at the earlier phase and a rise in agglutinin levels at the later phase of the experimental period in the higher dose groups.     

Marine bacterioplankton at the Weddell Sea ice edge,distribution of psychrophilic and psychrotrophic populations

Summary In the eastern Weddell Sea on several transects from ice-covered, through ice melt, to open-ocean stations, total and heterotrophic bacteria were estimated to document an enhanced bacteriological biomass expected near the ice edge. The highest numbers of bacteria were found in melted ice cores, with 4.2·10³ CFUml^–1 and 1.1·10⁷ Cells ml^–1. Although brine from pore water samples average more than one order of magnitude less cells per ml, the highest bacterial production, 2.2·10⁷ cells l^–1 day^–1, was recorded in brine samples. All quantitatively studied bacterial parameters were lower under the ice than in the ice samples but there were no clear vertical gradients in the water column. In the studied spring situation, sea ice occurrence seems to play only a minor role in the general distribution of the seawater bacterioplankton. The bacterial community structure was investigated by carrying out 29 morphological and biochemical tests on 118 isolated strains. The bacterial communities inhabiting Antarctic pack ice differ from those found in underlying seawater. Although non fermentative Gram-negative rods were always dominant in seawater, Vibrio sp. represented more than 25% of the strains isolated from some ice samples. The results clearly indicated that a large majority of the bacteria isolated from seawater must be considered psychrotrophic but that truly psychrophilic strains occurred in melted ice and brine samples.Data presented here were collected during the European Polarstern Study (EPOS) sponsored by the European Science Foundation     

不同寡营养培养条件下南海水体细菌群落结构及其对碳源的利用特征

【背景】绝大多数海洋微生物不可培养,为挖掘海洋生态系统中可培养的微生物资源,研究者尝试寡营养培养等方法。【目的】比较不同寡营养培养条件下南海水体细菌数量、群落结构及其对碳源的利用特征差异。【方法】采用原2216E培养液(Y)、稀释10倍(Y-10)和稀释50倍(Y-50)的2216E培养液培养南海海水样品,用荧光定量PCR法和16S rRNA基因检测细菌数量和菌群结构;利用平板计数法计数异养细菌的数量,纯化鉴定可培养细菌;采用Biolog EcoPlateTM微板法分析不同培养基中细菌群落对碳源的利用特征。【结果】Y组细菌总数高于Y-10组和Y-50组,差异不显著(P>0.05),但异养细菌数量显著高于Y-10组和Y-50组(P<0.05)。16S rRNA基因测序结果表明,不同稀释倍数下的细菌群落结构差异明显,Y组检测出10门193属,优势类群为Proteobacteria(56.44%)和Bacteroides (37.27%);Y-10组检测出15门220属,优势类群为Proteobacteria (40.30%)、Bacteroides(36.91%)和Firmic...     

Distribution of Aerobic Bacteria Which Contain Bacteriochlorophyll a

Sixteen strains of aerobic bacteria which contain bacteriochlorophyll a were isolated from the samples collected in aerobic marine environments: thalli of Enteromorpha linza, Porphyra sp., Sargussum horneri; beach sand; and the surface seawater from Aburatsubo Inlet. When they occurred, their proportions among the aerobic heterotrophic populations ranged from 0.9 to 1.1% in the seaweed samples and from 1.2 to 6.3% in the beach sand samples and were 0.9% in the seawater sample. The results suggested that the aerobic photopigmented bacteria widely inhabit aerobic marine environments.     

Numerical Taxonomy of Aerobic,Gram-negative Bacteria associated with Oysters and Surrounding Seawater of the Mediterranean Coast

A numerical taxonomic study was performed on 245 strains of heterotrophic, aerobic, marine bacteria, plus 26 reference strains. The isolates were obtained from oysters and seawater sampled monthly over one year, by direct plating on Marine Agar. The strains were characterised by 93 morphological, biochemical, physiological and nutritional tests. Clustering yielded 46 phena at 0.60 S level (S_J coefficient). Some could be identified as species of Alteromonas, Shewanella, Deleya, Flavobacterium, Oceanospirillum, Pseudomonas and marine Agrobacterium-like organisms, others were unidentified groups. Several phena seem to correspond to as yet undescribed taxa.     

Passive immunization of the tiger prawn, Penaeus monodon, using rabbit antisera to Vibrio harveyi

Passive immunization, toxicity neutralization and the persistence of passive protection in the tiger prawn ( Penaeus monodon ) were investigated using rabbit antisera to the formalinized extracellular products (ECP) (RαECP) and/or formalinized bacterial cells (RαBC) of luminescent Vibrio harveyi strain 820514 originally isolated from diseased tiger prawns. Rabbit antiserum to bovine serum albumin (RαBSA) or phosphate-buffered saline (PBS, pH 7·2) both served as controls. The toxicity of ECP to prawns was neutralized by pre-incubation with RαECP. Passive immunization by pre-injection of RαBC or RαECP into prawns 3 d in advance protected against a lethal dose challenge of bacteria. To determine the persistence of passive protection by rabbit antiserum in tiger prawns, the RαBC, RαECP, RαBSA or PBS were injected into prawns. At 10, 17 or 24 d post-immunization, groups of prawns were given a lethal dose challenge of bacteria. The prawns in the two control groups were all killed within the first 2 d following challenge at all three challenge dates, Pre-injection with RαBC and RαECP provided total protection for 10 and 17 d, respectively, with all treated prawns surviving for at least 2 weeks post-challenge. This is the first study using mammalian antisera to investigate toxicity neutralization, passive immunization and persistence of passive protection by rabbit antisera in prawns. The results could be useful in future studies on virulence mechanisms and disease control of vibriosis in cultured prawns.     

Succession of sea-ice bacterial communities in the Baltic Sea fast ice

Coastal fast ice and underlying water of the northern Baltic Sea were sampled throughout the entire ice winter from January to late March in 2002 to study the succession of bacterial biomass, secondary production and community structure. Temperature gradient gel electrophoresis (TGGE) and sequencing of TGGE fragments were applied in the community structure analysis. Chlorophyll-a and composition of autotrophic and heterotrophic assemblages were also examined. Overall succession of ice organism assemblages consisted of a low-productive stage, the main algal bloom, and a heterotrophic post-bloom situation, as typical for the study area. The most important groups of organisms in ice in terms of biomass were dinoflagellates, plasticidic flagellates, rotifers and ciliates. Ice bacteria showed a specific succession not directly dependent on the overall succession events of ice organisms. Sequenced 16S rDNA fragments were mainly affiliated to α-, β-, and γ-proteobacterial phyla and Cytophaga–Flavobacterium–Bacteroides-group, and related to sequences from cold environments, also from the Baltic Sea. Temporal clustering of the TGGE fingerprints was stronger than spatial, although lower ice and underlying water communities always clustered together, pointing to the importance of ice maturity and ice–water interactions in shaping the bacterial communities.     

Effect of Sodium Metabisulphite and Storage Temperature on the Survival of Vibrio cholerae on Prawns (Penaeus monodon)

Summary Sodium metabisulphite is traditionally used to control a non-microbiological spoilage symptom of prawns known as blackspot. This study was undertaken to evaluate the effect of sodium metabisulphite, at levels currently used to control blackspot, on the survival of V. cholerae on prawns during simulated commercial storage. Fresh prawns (Penaeus monodon) were divided into two batches, one of which was exposed to seawater while the other was treated with sodium metabisulphite (1%) in seawater. Two V. cholerae O1 strains were inoculated onto the prawns in separate experiments. Each of the batches were further sub-divided and stored either under chilled conditions (1.5 °C) for 14 days or under frozen conditions (−25 °C) for 49 days. At appropriate time intervals whole prawns were sampled and dilutions plated onto TCBS Cholera medium to determine numbers of V. cholerae. Experiments were performed in triplicate and uninoculated controls were included. Results indicated a significant (P < 0.05) decrease in numbers of V. cholerae on both chilled (~4 log units) and frozen (~2 log units) prawns over the storage period. A significantly (P < 0.05) greater decrease in numbers of the pathogen (up to 1.8 log units) on chilled sodium-metabisulphite-treated, as compared to chilled untreated, prawns was apparent. Sodium metabisulphite had a negligible effect on numbers of V. cholerae on frozen prawns. The use of sodium metabisulphite could be optimized and used to enhance the safety of chilled seafood in cholera-endemic countries.     

Influence of temperature on the growth potential of Southern polar marine bacteria

Regular surveys of heterotrophic microflora from seawater were conducted in the subantarctic (Kerguelen archipelago) and Antarctic (Terre Adélie area). Although a predominance of psychrophilic bacteria could be expected for such polar marine environments, there were no significant differences between results obtained after incubation at two different temperatures (4°C for 21 days or 18°C for 6 days). To investigate this further, four sets of bacterial strains were isolated from the subantarctic area (early fall, late fall, spring, and summer) and one set of Antarctic bacteria was isolated in summer. The growth rates of the 143 strains collected were determined at four different temperatures (4, 7, 20, and 30°C). The results clearly indicated that a large majority of the isolated bacteria must be considered psychrotrophic and not truly psychrophilic strains.     

Cultivation of particle-associated heterotrophic bacteria during a spring phytoplankton bloom in the North Sea

Seawater contains free-living and particle-attached bacteria. Only a small fraction is cultivable on plates. As free-living and particle-associated bacteria differ in their physiological traits, their cultivability on plates may coincide with particle association. Using filtration and Imhoff sedimentation cones, particles were collected during a spring phytoplankton bloom off Helgoland (North Sea) in order to obtain particle-associated bacteria as inocula. Direct dilution plating resulted in 526 strains from 3 µm filtration retentates and 597 strains from settled particles. Motile Gammaproteobacteria from the genera Pseudoalteromonas, Shewanella, Psychrobacter, Vibrio and Colwellia, as well as particle-attached Flavobacteriia affiliating with the genera Tenacibaculum and Gramella, were frequently isolated. As a result, a diverse collection comprised of 266 strains was deposited. Two strains were most likely to represent novel genera and 78 strains were probably novel species. Recently, a high-throughput cultivation study from the same site using seawater as an inoculum had retrieved 271 operational phylogenetic units (OPUs) that represented 88% of the 4136 characterized strains at the species level. A comparison of 16S rRNA gene sequences revealed that the collection obtained matched 104 of the 271 seawater OPUs at the species level and an additional 113 at the genus level. This large overlap indicated a significant contribution of particle-associated bacteria to the cultivable microbiome from seawater. The presence of 49 genera not identified in the larger seawater study suggested that sample fractionation was an efficient strategy to cultivate rare members of the planktonic microbiome. The diverse collection of heterotrophic bacteria retrieved in this study will be a rich source for future studies on the biology of particle-associated bacteria.     

Effect of dispersed oil on heterotrophic bacterial communities in cold marine waters

Mesocosm studies were conducted to evaluate the effect of dispersed oil on total and heterotrophic bacterial communities of under-ice seawater from the St. Lawrence Estuary. A regular survey of bacterial changes in the oil-contaminated seawater was performed during a two week period. The bacterial community structure was investigated by carrying out 27 morphological and biochemical tests on 168 isolated strains. The results show a detectable but transient response of the bacterial community to crude oil addition. While total bacterial counts were approximately constant during the experiment, dispersed oil induced an increase in heterotrophic bacterial microflora (from 10⁴ to 10⁵ bacteria ml^-1 after two weeks of contamination). The dispersed oil appeared to have an inhibitory effect on some components of the bacterial community. A decrease of most probable number values was observed just after addition of crude oil in the most polluted tanks and one day later in the less polluted tank. However, except for the most polluted tank, this adverse effect disappeared rapidly. While the dispersed oil induced a total disappearance of some components of the bacterial community in the most polluted tank, the structure of the bacterial community in the less polluted tank appeared relatively unchanged after 14 days of contamination.Offprint requests to: Daniel Delille     

News & Notes: Virulence of Vibrio alginolyticus Isolated from Diseased Tiger Prawn,Penaeus monodon

Outbreaks of mass mortality among cultured tiger prawns (Penaeus monodon) with white spotted syndrome (WSS) in the carapace occurred in the summer of 1994 in I-Lan, Taiwan. A swarming strain Val was isolated from hemolymph of the moribund prawns with tryptic soy agar (TSA, supplemented with 1% NaCl, Oxoid) and/or thiosulfate citrate bile salt sucrose (TCBS, Difco) agar. This strain was characterized and identified to be Vibrio alginolyticus. The strain was susceptible to antibiotics such as chloramphenicol, ciprofloxacin, doxycycline hydrochloride, nalidixic acid, oxolinic acid, and oxytetracycline while resistant to ampicillin, novobiocin, penicillin G, sulfisoxazole, and sulfonamide. The bacteria and their extracellular products (ECP) were lethal to both tiger prawns (P. monodon) and kuruma prawns (P. japonicus) with LD₅₀ values of 1.13 × 10⁵, 2.46 × 10⁵ CFU/g, and 0.23, 0.63 μg protein/g prawn body weight, respectively.     

Effects of eyestalk ablation on growth and food conversion efficiency of the freshwater prawn Macrobrachium lanchesteri (de Man)

Bilateral eyestalk ablation in the freshwater prawn Macrobrachium lanchesteri results in high mortality, while unilateral eyestalk ablated prawns exhibited a high survival rate. There was marked increase in the growth of bilateral eyestalk-ablated prawns (47.70 mg/prawn) as compared to those that were unilaterally ablated (19.19 mg/prawn).     

Characterisation of the culturable heterotrophic bacterial community in a small eutrophic lake (Priest Pot)

The community composition and structure of planktonic heterotrophic bacteria (903 isolates) sampled from a small eutrophic lake in northern England (Priest Pot) was studied with respect to season (four samples) and depth (to 3.1 m). Bacteria (887) were isolated on tryptic soy broth agar and identified to 48 genera using fatty acid methyl ester analysis. The two most abundant genera isolated were Aeromonas and Pseudomonas which, respectively, dominated the middle to bottom depths in August and all depths in February. The structure of the sampled community was described using: species richness, Simpson's index and the Shannon-Wiener index. All three indices detected a number of significant differences with depth demonstrating stratification. The greatest stratification of the bacterial community was observed in August when bacterial counts correlated strongly and negatively with diversity. Using structural measures was found to be preferable to the use of species frequencies in the analysis of perturbation and succession in community structure. Insensitivity to one or more of eight antibiotics was observed in 71% (61/86) of the isolates tested particularly in Gram-negative genera. Bacteriocinogeny and lysogeny was observed in 36% (32/90) of isolates. Using sensitive indicator strains, two of 10 producing strains produced virus, while the others produced bacteriocins.     

Nile perch,Lates niloticus,predation on the freshwater prawn,Caridina nilotica,in the Nyanza Gulf,Lake Victoria,East Africa

Synopsis The percentage of Nile perch, Lates niloticus, containing the prawn, Caridina nilotica, declined from 60–85%, for fish between 5 and 40 cm total length, to less than 10% for fish larger than 80 cm in length. The maximum number of prawns eaten by a Nile perch increased steadily, from 10 prawns for a fish in the 5–9.9 cm length group, to 913 prawns in a fish in the 60–69.9 cm length group, and then fell sharply for larger fish. The mean number of Caridina eaten by fish that were foraging on this animal followed a similar trend. Gill raker spacing increased in direct proportion to fish length and this may prevent fish larger than 70 cm in length from being effective prawn predators.