Mesocosm experiments for evaluating the biological efficacy of ozone treatment of marine ballast water

Ballast water is a major pathway for the transfer of non-indigenous species in aquatic environments. The objectives of this study were to determine the ability of ozone to reduce the numbers of a spectrum of marine organisms collected from Puget Sound, Washington in replicated mesocosm (280 l) experiments, and estimate the minimum ozone concentrations as measured by total residual oxidant (TRO) required to reduce organism densities. Ozone treatment was effective in removing bacteria, phytoplankton, and mesozooplankton with initial TRO concentrations of 2–5 mg l⁻¹ as Br₂. Persistence of TRO resulted in an extended period of toxicity and cumulative mortality. TRO decay allowed bacteria populations to multiply when TRO levels fell below 0.5–1.0 mg l⁻¹ as Br₂. Phytoplankton chlorophyll a concentrations were rapidly reduced by ozone treatment and did not increase in any treatments or controls because of lack of light. Overall mesozooplankton viability was rapidly reduced by 90–99% in treatment TRO levels above 1.85 mg l⁻¹ as Br₂. Our study outlines novel protocols that can be used for testing different potential ballast water treatment systems in replicated and controlled mesocosm experiments.     

Shipboard trials of an ozone-based ballast water treatment system

Legislation introduced by the United Nations International Maritime Organization (IMO) has focused primarily on standards defining successful treatments designed to remove invasive species entrained in ballast water. An earlier shipboard study found that ozone introduced into salt water ballast resulted in the formation of bromine compounds, measured as total residual oxidants (TRO) that were toxic to both bacteria and plankton. However, the diffuser system employed to deliver ozone to the ballast water tanks resulted in patchiness in TRO distribution and toxicity to entrained organisms. In this follow-up study, the shipboard diffuser system was replaced by a single Venturi-type injection system designed to deliver a more homogeneous biocide distribution. Within-tank variability in TRO levels and associated toxicity to zooplankton, phytoplankton and marine bacteria was measured via a matrix of tubes deployed to sample different locations in treated and untreated (control) tanks. Three trials were conducted aboard the oil tanker S/T Prince William Sound in the Strait of Juan de Fuca off Port Angeles, Washington State, USA, between June and December 2007. Mortalities of plankton and bacteria and oxidant concentrations were recorded for treated and untreated ballast water up to 3 days following treatment, and residual toxicity beyond this period was measured by bioassay of standard test organisms. Results indicated uniform compliance with current IMO standards, but only partial compliance with other existing and pending ballast water legislation.     

Setting a size-exclusion limit to remove toxic dinoflagellate cysts from ships' ballast water

Dinoflagellate cysts are well-recognized biological constituents of ships' ballast tanks. They are present in ballast water, sediments and residual water in drained tanks, and in biofilms formed on interior tank surfaces. Therefore, cysts have the potential to be released during ballast discharge. The International Maritime Organization's (IMO) Ballast Water Management Convention (promulgated February 2004) stipulates a performance standard (Annex, Regulation D2) requiring discharged ballast water contain <10 viable organisms between 10 and 50 microm per ml and <10 viable organisms 50 microm per m3. The proposed size limit has potential to exclude both the smallest toxic and the largest toxic and non-toxic dinoflagellate (and other microalgal) cysts from discharged ballast water. Despite the appropriateness of size cutoffs however, ballast water containing predominantly small cysts (<50 microm) could be deemed in compliance with the performance standard, even without treatment, while ballast water having the same concentration of larger cysts (>50 microm) could require a multiple-log reduction in abundance before its permissible discharge. Also of concern, it remains uncertain whether ballast-water treatment can remove sufficient organisms, including dinoflagellate cysts, to meet the performance standard.     

Effects of sonication and advanced chemical oxidants on the unicellular green alga Dunaliella tertiolecta and cysts, larvae and adults of the brine shrimp Artemia salina: a prospective treatment to eradicate invasive organisms from ballast water

Uptake and release of ship-borne ballast water is a major factor contributing to introductions of aquatic phytoplankton and invasive macroinvertebrates. Some invasive unicellular algae can cause harmful algal blooms and produce toxins that build up in food chains. Moreover, to date, few studies have compared the efficacy of ballast water treatments against different life history phases of aquatic macroinvertebrates. In the present study, the unicellular green alga Dunaliella tertiolecta, and three discrete life history phases of the brine shrimp Artemia salina, were independently used as model organisms to study the efficacy of sonication as well as the advanced oxidants, hydrogen peroxide and ozone, as potential ballast water treatments. Algal cells and brine shrimp cysts, nauplii, and adults were subjected to individual and combined treatments of sonication and advanced oxidants. Combined rather than individual treatments consistently yielded the highest levels of mortality in algal cells (100% over a 2 min exposure) and in brine shrimp (100% and 95% for larvae and adults, respectively, over a 2 min exposure). In contrast, mortality levels in brine shrimp cysts (66% over 2 min; increased to 92% over a 20 min exposure) were moderately high but consistently lower than that detected for larval or adult shrimp. Our results indicate that a combination of sonication and advanced chemical oxidants may be a promising method to eradicate aquatic unicellular algae and macroinvertebrates in ballast water.     

Ship board testing of a deoxygenation ballast water treatment

A ship board trial of a deoxygenation method for treating ballast water was carried out during a voyage from Southampton (United Kingdom) to Manzanillo (Panama). A nutrient solution added to two ballast tanks encouraged bacterial growth, resulting in a gradual change to an anoxic environment. Samples were taken from two treated tanks and two untreated tanks to assess changes in the abundance and viability of zooplankton, phytoplankton and bacteria. The work was carried out before the International Maritime Organization (IMO) standard was agreed so only a broad indication of whether the results achieved the standard was given. For the zooplankton, the standard would have been achieved within 5 or 7 days but the phytoplankton results were inconclusive. The biological efficacy was the result of the combination of several factors, including the treatment, pump damage and an increase in the water temperature during the voyage.     

Results from the first ballast water sampling study in the Mediterranean Sea - the Port of Koper study

The ongoing transfer of harmful organisms by shipping, especially via ballast water transport, may result in a change of biodiversity, alteration of ecosystems, negative impacts on human health and economic loss. Species introductions which cause irreversible consequences to receiving environments and economies call for particular attention. One critical issue is a need to evaluate the quantities and processes of species introductions. Consequently ballast water was sampled on 15 ships calling at the Port of Koper, Slovenia. This was the first ballast water sampling study in the Mediterranean Sea. This paper summarises the sampling results. Samples were analysed for all types of aquatic organisms including bacteria. The results may be considered as background information for an initial risk assessment of future species introductions - an important tool for the implementation of ballast water management measures.     

Microbial distribution and abundance in response to physical and biological processes on the continental shelf of southeastern U.S.A.

The distribution and abundance of bacteria and phytoplankton on the continental shelf of the southeastern United States were observed in relation to physical processes. Phytoplankton production was influenced by inputs of water of reduced salinity from the estuaries and by inputs of high salinity, low-temperature water from the west front of the Gulf Stream. The distribution of chlorophyll suggests that in each case production is influenced both by inputs of nutrients and by the enhanced vertical stability associated with the stratification of waters of different densities. The standing stock of bacteria on the inner shelf, 10⁶ ml^?1, is little changed by the influx of water of reduced salinity. On the outer shelf, where the usual standing stock of bacteria is 10⁵ ml^?1, the numbers increase to 10⁶ ml^?1 in and above intrusions of Gulf Stream water in which phytoplankton blooms have developed, suggesting that the bacteria respond to products of both phytoplankton and zooplankton production. Adenylate energy charge values in the waters of the southeastern shelf are variable and volatile. At times values of 0.7 to 0.8 are widespread over most of the shelf, while at other times values <0.6 are common, with localized patches of high values. Both autotroph-dominated and heterotroph-dominated microbial communities show these variations.     

On board short-time high temperature heat treatment of ballast water: a field trial under operational conditions

A ballast water short-time high temperature heat treatment technique was applied on board a car-carrier during a voyage from Egypt to Belgium. Ballast water from three tanks was subjected for a few seconds to temperatures ranging from 55 degrees C to 80 degrees C. The water was heated using the vessel's heat exchanger steam and a second heat exchanger was used to pre-heat and cool down the water. The treatment was effective at causing mortality of bacteria, phytoplankton and zooplankton. The International Maritime Organization (IMO) standard was not agreed before this study was carried out, but comparing our results gives a broad indication that the IMO standard would have been met in some of the tests for the zooplankton, in all the tests for the phytoplankton; and probably on most occasions for the bacteria. Passing the water through the pump increased the kill rate but increasing the temperature above 55 degrees C did not improve the heat treatment's efficacy.     

Isolation, identification and characterization of phytoplankton-lytic bacterium CH-22 against Microcystis aeruginosa

A bacterial strain named CH-22 showing phytoplankton-lytic activity against bloom-forming cyanobacterium Microcystis aeruginosa was isolated from Lake Chaohu of Anhui Province, China. The isolated strain was identified as Pseudomonas putida by morphology and homology research based on 16S rDNA. The phytoplankton-lytic activity was confirmed by reduction in cell number and chlorophyll a concentration of M. aeruginosa in an infected culture for a defined period. The initial bacterial and M. aeruginosa densities affected the phytoplankton-lytic activity significantly. When the 15% (150 μL/mL) concentration of bacterial cultures was infected, the highest phytoplankton-lytic activity reached to 98.8% after 7 days. When the initial M. aeruginosa density was less than 3×10⁶ cells/mL, about 90.0% of chlorophyll a was removed. Obvious reduction in phycocyanin concentration in the treated M. aeruginosa suggests that isolated strain may possibly inhibit the synthesis of photosynthetic apparatus. Supernatants of bacterial cultures showed higher phytoplankton-lytic activity, suggesting that phytoplankton-lytic bacterium P. putida indirectly attacked M. aeruginosa cells by secretion of extracellular antialgal substances, which is characterized as anti-heat shock. The isolated P. putida also showed effective phytoplankton-lytic activity against a wide range of phytoplankton. These results suggest that indigenous bacteria isolated from eutrophic lake may be employed to regulate the ecological balance between the phytoplankton and bacteria, and consequently, to reduce the occurrence of cyanobacterial blooms in freshwaters.     

A low-energy intensive electrochemical system for the eradication of Escherichia coli from ballast water: process development, disinfection chemistry, and kinetics modeling

The invasion of biological organisms via ballast water has created threats to the environment and human health. In this study, a cost-effective electrochemical disinfection reactor was developed to inactivate Escherichia coli, one of the IMO-regulated indicator microbes, in simulated ballast water. The complete inactivation of E. coli could be achieved within a very short time (150, 120, or 60 s) with an energy consumption as low as 0.0090, 0.0074 or 0.0035 kWh/m(3) for ballast water containing E. coli at concentrations of 10(8), 10(7) and 10(6) CFU/100 mL, respectively. Electrochemical chlorination was the major disinfection mechanism in chloride-abundant electrolytes, whereas oxidants such as ozone and free radicals contributed to 20% of the disinfection efficiency in chloride-free electrolytes. Moreover, a disinfection kinetics model was successfully developed to describe the inactivation of E. coli.     

Ballast water as a vector of coral pathogens in the Gulf of Mexico: the case of the Cayo Arcas coral reef

The discharge of nutrients, phytoplankton and pathogenic bacteria through ballast water may threaten the Cayo Arcas reef system. To assess this threat, the quality of ballast water and presence of coral reef pathogenic bacteria in 30 oil tankers loaded at the PEMEX Cayo Arcas crude oil terminal were determined. The water transported in the ships originated from coastal, oceanic or riverine regions. Statistical associations among quality parameters and bacteria were tested using redundancy analysis (RDA). In contrast with coastal or oceanic water, the riverine water had high concentrations of coliforms, including Vibrio cholerae 01 and, Serratia marcescens and Sphingomona spp., which are frequently associated with “white pox” and “white plague type II” coral diseases. There were also high nutrient concentrations and low water quality index values (WQI and TRIX). The presence of V. cholerae 01 highlights the need for testing ballast water coming from endemic regions into Mexican ports.     

Importance of ciliated protozoa in relation to the bacterial and phytoplanktonic biomass in an oligo-mesotrophic lake,during the spring diatom bloom

The abundance and the biomass of bacterial, phytoplanktonic, and ciliate communities were estimated at different depths during the spring planktonic development in an oligo-mesotrophic lake (the Pavin lake).The bacterial population, which consists mainly of free bacteria (94% of the total bacterial abundance), displays only low cell densities (0.6 to 7 × 10⁵ cells · ml^–1) and represents low biomass values (0.9 to 11.5 µgC·l^–1) The bacteria represent from 0.9 to 23.8% (M = 9.7%) of the microplanktonic biomass (with the exclusion of heterotrophic nanoflagellates, i.e. bacteria + phytoplankton + ciliates, size range 0.2–160 µm). The abundance of the phytoplankton varies between 0.5 and 1.8 × 10⁶ cells·l^–1, and the biomass values between 12 and 118 µC·l^–1. The phytoplankton population constitutes the largest part of the microplanktonic biomass (51.9 to 96.6%, M = 80.6%), and the diatomMelosira italica subsp.subarctica is the largely dominant species of this community. The population of ciliates, essentiallyOligotrichida andScuticociliatida, displays densities between 1.3 and 38.3 × 10³ cells·l^–1 (M = 6.7 × 10³ cells·l^–1), and biomass values vary from 0.10 to 16.30 µgC·l^–1 (M = 6.01 µgC·l^–1). The ciliates constitute thus from 0.1 to 26.4% (M = 9.8%) of the microplanktonic biomass. Whereas the oligotrichs are best represented in the euphotic zone, the small-sized scuticociliates dominate in the hypolimnion. Besides, species having symbionts and considered to be mixotrophic (Strobilidium gyrans, Strombidium viride, Stokesia vernalis) develop preferentially in the epilimnion and constitute more than 50% of the total ciliate biomass.     

Untersuchungen zum Einfluß limnischer Makrophyten auf die Absterbegeschwindigkeit von Escherichia coli im Wasser

In bioassays in aquaria, the dying of Escherichia coli was accelerated by Glyceria maxima, Schoenoplectus lacustris, Alisma plantago-aquatica as well as Mentha aquatia as compared with check tanks without any plants (initial concentration 15 × 10³ colony-forming units per ml). As expected, by this the antibacterial effect of the limnic macrophytes attains its maximum effectivity within the first days of the period of investigation. The decimal time of reduction is shortened by one third to half, the centimal reduction time is shortened by one fourth to one third. Mean contact times of 7… 11 h are required for the decimation of bacteria by 90 %, and mean contact times of 16… 19 h are necessary for a 99 % reduction of the population. Possibly, the antibacterial effect in mixed stands of different plant species is increased additively or superadditively.     

Electrochemical disinfection for ballast water management: technology development and risk assessment

Ballast water is essential in maintaining the balance and structural integrity of ships during voyage. However, it has created biological invasion threats to the ocean environment. An innovative electrochemical technology was developed in this study. The microorganisms regulated by the International Maritime Organization (D2) were used as the target organisms. It was found that the required energy to meet the D2 was below 0.006 kWh/m3. The size of disinfector (m3) was about 0.5% of treatment flow rate (m3/h). The complete disappearance of chlorine in seawater was achieved after three days. The ballast tank corrosion was not worsened due to the application of technology. The ecotoxicity studies showed no toxic effect on fish, invertebrate, and algae. Finally, the environmental risk assessment showed the treated water did not pose threats to the environment. It can therefore be concluded that the technology provides a cost-effective and environmental friendly solution to ballast water management.     

Evolution of phytoplankton cultures after ultraviolet light treatment

Introducing invasive species in new environments through ballast water is a specific problem of contamination and has recently become one of the main concerns of Maritime Organizations. Ultraviolet-C radiation (UV-C) is a technological alternative to prevent this maritime pollution. This study addresses the effect of UV-C on different phytoplankton cultures and also the ability to recover following exposure to damage. A UV-C low-pressure lamp irradiates the cultures. The distance from the source and the thickness of the layer prevent part of the energy from reaching the culture and the disinfective process is diminished. Some cultures such as Chlorella autotrophica and Chaetoceros calcitrans can easily recover from UV-C damage. However, Phaeocystis globosa does not have this ability. C. calcitrans forms cysts and exhibits two different behaviours depending on the dose applied.     

Real-time PCR and NASBA for rapid and sensitive detection of Vibrio cholerae in ballast water

Transport of ballast water is one major factor in the transmission of aquatic organisms, including pathogenic bacteria. The IMO-guidelines of the Convention for the Control and Management of Ships' Ballast Water and Sediments, states that ships are to discharge <1 CFU per 100 ml ballast water of toxigenic Vibrio cholerae, emphasizing the need to establish test methods. To our knowledge, there are no methods sensitive and rapid enough available for cholera surveillance of ballast water. In this study real-time PCR and NASBA methods have been evaluated to specifically detect 1 CFU/100ml of V. cholerae in ballast water. Ballast water samples spiked with V. cholerae cells were filtered and enriched in alkaline peptone water before PCR or NASBA detection. The entire method, including sample preparation and analysis was performed within 7 h, and has the potential to be used for analysis of ballast water for inspection and enforcement control.     

The nature and extent of organisms in vessel sea-chests: A protected mechanism for marine bioinvasions

A total of 150 different organisms, including one plant species and 12 animal phyla were identified from sea-chests of 42 vessels visiting or operating in New Zealand between May 2000 and November 2004. Forty-nine percent of organisms were sessile, 42% mobile adults and the remaining 9% sedentary. Decapods were the most represented group with 19 species present among 79% of vessels. Forty percent of organisms were indigenous to New Zealand, 15% introduced, 10% non-indigenous, and 35% of unknown origin. Sea-chests have the potential to (1) transfer non-indigenous organisms between countries across oceanic boundaries; and (2) disperse both indigenous and introduced organisms domestically. The occurrence of adult mobile organisms is particularly significant and indicates that sea-chests may be of greater importance than ballast water or hull fouling for dispersing certain marine species. These findings emphasise the need to assess and manage biosecurity risks for entire vessels rather than different mechanisms (i.e., ballast water, hull fouling, sea-chests, etc.) in isolation.     

Food Type Effects on the Population Growth Patterns of Littoral Rotifers and Cladocerans

Littoral rotifers and cladocerans feed on bacteria, phytoplankton, and detritus. In comparison to planktonic organisms their species diversity is high, yet little is known about their biology. In this study we present data on the functional response of the cladocerans Macrothrix triserialis and Alona rectangula, and the rotifer Brachionus macracanthus to different levels of the alga Chlorella vulgaris. Based on the incipient limiting level, we chose the food concentration and tested the population growth on four types of diets for three cladoceran species — Macrothrix triserialis, Alona rectangula, and Chydorus sphaericus — and three rotifer species — Brachionus macracanthus, Lecane quadridentata, and Platyias quadricornis. We studied growth rates of each of these species on diets of live Chlorella vulgaris, heat‐killed Chlorella vulgaris, baker's yeast (Saccharomyces cerevisiae), and field collected detritus. The incipient limiting level was around 0.9 · 10⁶ cells mL^?1 of Chlorella for the tested zooplankton species. The population growth patterns showed that, in general, the cladocerans took 25…50 d to reach their peak population densities, which ranged from 10…75 ind. mL^?11. Among the rotifers L. quadridentata and B. macracanthus had equally high growth rates on live and heat‐killed forms of C. vulgaris while P. quadricornis grew best on detritus. The growth rates ranged from 0.03…0.21 d^?1. The similarities and differences between littoral and planktonic organisms in relation to food type and availability have been discussed.     

Electrochemical disinfection for ballast water management: Technology development and risk assessment

Ballast water is essential in maintaining the balance and structural integrity of ships during voyage. However, it has created biological invasion threats to the ocean environment. An innovative electrochemical technology was developed in this study. The microorganisms regulated by the International Maritime Organization (D2) were used as the target organisms. It was found that the required energy to meet the D2 was below 0.006 kWh/m³. The size of disinfector (m³) was about 0.5% of treatment flow rate (m³/h). The complete disappearance of chlorine in seawater was achieved after three days. The ballast tank corrosion was not worsened due to the application of technology. The ecotoxicity studies showed no toxic effect on fish, invertebrate, and algae. Finally, the environmental risk assessment showed the treated water did not pose threats to the environment. It can therefore be concluded that the technology provides a cost-effective and environmental friendly solution to ballast water management.     

Potential microbial bioinvasions via ships' ballast water, sediment, and biofilm

A prominent vector of aquatic invasive species to coastal regions is the discharge of water, sediments, and biofilm from ships' ballast-water tanks. During eight years of studying ships arriving to the lower Chesapeake Bay, we developed an understanding of the mechanisms by which invasive microorganisms might arrive to the region via ships. Within a given ship, habitats included ballast water, unpumpable water and sediment (collectively known as residuals), and biofilms formed on internal surfaces of ballast-water tanks. We sampled 69 vessels arriving from foreign and domestic ports, largely from Western Europe, the Mediterranean region, and the US East and Gulf coasts. All habitats contained bacteria and viruses. By extrapolating the measured concentration of a microbial metric to the estimated volume of ballast water, biofilm, or residual sediment and water within an average vessel, we calculated the potential total number of microorganisms contained by each habitat, thus creating a hierarchy of risk of delivery. The estimated concentration of microorganisms was greatest in ballast water>sediment and water residuals>biofilms. From these results, it is clear microorganisms may be transported within ships in a variety of ways. Using temperature tolerance as a measure of survivability and the temperature difference between ballast-water samples and the water into which the ballast water was discharged, we estimated 56% of microorganisms could survive in the lower Bay. Extrapolated delivery and survival of microorganisms to the Port of Hampton Roads in lower Chesapeake Bay shows on the order of 10(20) microorganisms (6.8 x 10(19) viruses and 3.9 x 10(18) bacteria cells) are discharged annually to the region.