Integrated treatment of shrimp effluent by sedimentation, oyster filtration and macroalgal absorption: a laboratory scale study

Effluent water from shrimp ponds typically contains elevated concentrations of dissolved nutrients and suspended particulates compared to influent water. Attempts to improve effluent water quality using filter feeding bivalves and macroalgae to reduce nutrients have previously been hampered by the high concentration of clay particles typically found in untreated pond effluent. These particles inhibit feeding in bivalves and reduce photosynthesis in macroalgae by increasing effluent turbidity. In a small-scale laboratory study, the effectiveness of a three-stage effluent treatment system was investigated. In the first stage, reduction in particle concentration occurred through natural sedimentation. In the second stage, filtration by the Sydney rock oyster, Saccostrea commercialis (Iredale and Roughley), further reduced the concentration of suspended particulates, including inorganic particles, phytoplankton, bacteria, and their associated nutrients. In the final stage, the macroalga, Gracilaria edulis (Gmelin) Silva, absorbed dissolved nutrients. Pond effluent was collected from a commercial shrimp farm, taken to an indoor culture facility and was left to settle for 24 h. Subsamples of water were then transferred into laboratory tanks stocked with oysters and maintained for 24 h, and then transferred to tanks containing macroalgae for another 24 h. Total suspended solid (TSS), chlorophyll a, total nitrogen (N), total phosphorus (P), NH4+, NO3-, and PO43-, and bacterial numbers were compared before and after each treatment at: 0 h (initial); 24 h (after sedimentation); 48 h (after oyster filtration); 72 h (after macroalgal absorption). The combined effect of the sequential treatments resulted in significant reductions in the concentrations of all parameters measured. High rates of nutrient regeneration were observed in the control tanks, which did not contain oysters or macroalgae. Conversely, significant reductions in nutrients and suspended particulates after sedimentation and biological treatment were observed. Overall, improvements in water quality (final percentage of the initial concentration) were as follows: TSS (12%); total N (28%); total P (14%); NH4+ (76%); NO3- (30%); PO43-(35%); bacteria (30%); and chlorophyll a (0.7%). Despite the probability of considerable differences in sedimentation, filtration and nutrient uptake rates when scaled to farm size, these results demonstrate that integrated treatment has the potential to significantly improve water quality of shrimp farm effluent. (C) 2001 Elsevier Science B.V. All rights reserved.

Detection of dengue viral RNA in Aedes aegypti (Diptera : Culicidae) exposed to sticky lures using reverse-transcriptase polymerase chain reaction

Active surveillance for dengue (DEN) virus infected mosquitoes can be an effective way to predict the risk of dengue infection in a given area. However, doing so may pose logistical problems if mosquitoes must be kept alive or frozen fresh to detect DEN virus. In an attempt to simplify mosquito processing, we evaluated the usefulness of a sticky lure and a seminested reverse-transcriptase polymerase chain reaction assay (RT-PCR) for detecting DEN virus RNA under laboratory conditions using experimentally infected Aedes aegypti (L.) mosquitoes. In the first experiment, 40 male mosquitoes were inoculated with 0.13 mul of a 10(4) pfu/ml DEN-2 stock solution. After a 7-d incubation period, the mosquitoes were applied to the sticky lure and kept at room temperatures of 23-30 degreesC. Following 7,10,14, and 28 d application, 10 mosquitoes each were removed from the lure pooled and assayed for virus. DEN virus nucleic acid was clearly detectable in all pools up to 28 d after death. A second study evaluated sensitivity and specificity using one, two, and five DEN-infected mosquitoes removed after 7, 10, 14, 21 and 30 d application and tested by RT-PCR. All four DEN serotypes were individually inoculated in mosquitoes and evaluated using the same procedures as experiment 1. The four serotypes were detectable in as few as one mosquito 30 d after application to the lure with no evidence of cross-reactivity. The combination of sticky lures and RT-PCR show promise for mosquito and dengue virus surveillance and warrant further evaluation.

Laboratory evaluation of two native fishes from tropical North Queensland as biological control agents of subterranean Aedes aegypti

The ability of 2 freshwater fishes, eastern rainbow fish Melanotaenia splendida splendida and fly-specked hardyhead Craterocephalus stercusmuscarum stercusmuscarum. native to North Queensland to prey on immature Aedes aegypti was evaluated under laboratory conditions. The predation efficiency of the 2 species was compared to the exotic guppy, Poecilia reticulata, which is commonly used as a biological control agent of mosquito larvae. Of the 3 fish species tested, M. s. splendida was shown to be the most promising agent for the biological control of Ae. aegypti that breed in wells. Melanotaenia s. splendida consumed significantly greater numbers of immature Ae. aegypti than P. reticulata, irrespective of developmental stage or light conditions. Unlike C. s. stercusmuscarum, M, s. splendida could be handled, transported, and kept in captivity for extended periods with negligible mortality. However, M. s. splendida was also an efficient predator of Litoria caerulea tadpoles, a species of native frog found in wells during the dry season. This result may limit the usefulness of M. s. splendida as a biological control agent of well-breeding Ae. aegypti and suggests that predacious copepods, Mesocyclops spp., are more suitable. However, the use of M. s. splendida as a mosquito control agent in containers that are unlikely to support frog populations (e.g., aquaculture tanks and drinking troughs) should be given serious consideration.

Drug susceptibility testing of anaerobic protozoa

A simple technique for routine, reproducible global surveillance of the drug susceptibility status of the anaerobic protozoa Trichomonas, Entamoeba, and Giardia is described, Data collected using this technique can be readily compared among different laboratories and with previously reported data. The technique employs a commercially available sachet and bag system to generate a low-oxygen environment and log, drug dilutions in microtiter plates, which can be monitored without aerobic exposure, to assay drug-resistant laboratory lines and clinically resistant isolates. MICs (after 2 days) of 3.2 and 25 muM indicated metronidazole-sensitive and highly clinically resistant isolates of T. vaginalis in anaerobic assays, respectively. The aerobic MICs were 25 and > 200 muM. MICs (1 day) of 12.5 to 25 muM were found for axenic lines of E. histolytica, and MICs for G. duodenalis (3 days) ranged from 6.3 muM for metronidazole-sensitive isolates to 50 muM for laboratory metronidazole-resistant lines. This technique should encourage more extensive monitoring of drug resistance in these organisms.

Analysis of the microbial community structure and function of a laboratory scale enhanced biological phosphorus removal reactor

A laboratory scale sequencing batch reactor (SBR) operating for enhanced biological phosphorus removal (EBPR) and fed with a mixture of volatile fatty acids (VFAs) showed stable and efficient EBPR capacity over a four-year-period. Phosphorus (P), poly-beta-hydroxyalkanoate (PHA) and glycogen cycling consistent with classical anaerobic/aerobic EBPR were demonstrated with the order of anaerobic VFA uptake being propionate, acetate then butyrate. The SBR was operated without pH control and 63.67+/-13.86 mg P l(-1) was released anaerobically. The P% of the sludge fluctuated between 6% and 10% over the operating period (average of 8.04+/-1.31%). Four main morphological types of floc-forming bacteria were observed in the sludge during one year of in-tensive microscopic observation. Two of them were mainly responsible for anaerobic/aerobic P and PHA transformations. Fluorescence in situ hybridization (FISH) and post-FISH chemical staining for intracellular polyphosphate and PHA were used to determine that 'Candidatus Accumulibacter phosphatis' was the most abundant polyphosphate accumulating organism (PAO), forming large clusters of coccobacilli (1.0-1.5 mum) and comprising 53% of the sludge bacteria. Also by these methods, large coccobacillus-shaped gammaproteobacteria (2.5-3.5 mum) from a recently described novel cluster were glycogen-accumulating organisms (GAOs) comprising 13% of the bacteria. Tetrad-forming organisms (TFOs) consistent with the 'G bacterium' morphotype were alphaproteobacteria , but not Amaricoccus spp., and comprised 25% of all bacteria. According to chemical staining, TFOs were occasionally able to store PHA anaerobically and utilize it aerobically.

The modelling of froth zone recovery in batch and continuously operated laboratory flotation cells

This paper proposes an integrated methodology for modelling froth zone performance in batch and continuously operated laboratory flotation cells. The methodology is based on a semi-empirical approach which relates the overall flotation rate constant to the froth depth (FD) in the flotation cell; from this relationship, a froth zone recovery (R,) can be extracted. Froth zone recovery, in turn, may be related to the froth retention time (FRT), defined as the ratio of froth volume to the volumetric flow rate of concentrate from the cell. An expansion of this relationship to account for particles recovered both by true flotation and entrainment provides a simple model that may be used to predict the froth performance in continuous tests from the results of laboratory batch experiments. Crown Copyright (C) 2002 Published by Elsevier Science B.V. All rights reserved.

Glycogen-accumulating organisms in laboratory-scale and full scale wastewater treatment processes

Laboratory-scale sequencing batch reactors (SBRs) as models for wastewater treatment processes were used to identify glycogen-accumulating organisms (GAOs), which are thought to be responsible for the deterioration of enhanced biological phosphorus removal (EBPR). The SBRs (called Q and T), operated under alternating anaerobic-aerobic conditions typical for EBPR, generated mixed microbial communities (sludges) demonstrating the GAO phenotype. Intracellular glycogen and poly-beta-hydroxyalkanoate (PHA) transformations typical of efficient EBPR occurred but polyphosphate was not bioaccumulated and the sludges contained 1.8% P (sludge Q) and 1.5% P (sludge T). 16S rDNA clone libraries were prepared from DNA extracted from the Q and T sludges. Clone inserts were grouped into operational taxonomic units (OTUs) by restriction fragment length polymorphism banding profiles. OTU representatives were sequenced and phylogenetically analysed. The Q sludge library comprised four OTUs and all six determined sequences were 99.7% identical, forming a cluster in the gamma-Proteobacteria radiation. The T sludge library comprised eight OTUs and the majority of clones were Acidobacteria subphylum 4 (49% of the library) and candidate phylum OPU (39% of the library). One OTU (two clones, of which one was sequenced) was in the gamma-Proteobacteria radiation with 95% sequence identity to the Q sludge clones. Oligonucleotide probes (called GAOQ431 and GAOQ989) were designed from the gamma-Proteobacteria clone sequences for use in fluorescence in situ hybridization (FISH); 92 % of the Q sludge bacteria and 28 % of the T sludge bacteria bound these probes in FISH. FISH and post-FISH chemical staining for PHA were used to determine that bacteria from a novel gamma-Proteobacteria cluster were phenotypically GAOs in one laboratory-scale SBR and two fullscale wastewater treatment plants. It is suggested that the GAOs from the novel cluster in the gamma-Proteobacteria radiation be named 'Candidatus Competibacter phosphatis'.

Detection and stability of Japanese encephalitis virus RNA and virus viability in dead infected mosquitoes under different storage conditions

A semi-nested polymerase chain reaction (PCR) was evaluated for detection of Japanese encephalitis (JE) virus in infected mosquitoes stored under simulated northern Australian summer conditions. The effect of silica gel, thymol, and a combination of the two on RNA stability and virus viability in dead mosquitoes were also examined. While JE virus RNA was relatively stable in mosquitoes held for up to 14 days after death, viable virus was not detected after day 1. Thymol vapor inhibited fungal contamination. Detection of single mosquitoes infected with JE virus in large pools of mosquitoes was also investigated. Single laboratory-infected mosquitoes were detected in pools of less than or equal to200 mosquitoes and in pools diluted to 0.2/100 and 0.1/100 mosquitoes, using the semi-nested PCR. However, the ability to detect live virus decreased as pool size increased. The semi-nested PCR proved more expensive than virus isolation for pools of 100 mosquitoes. However, the semi-nested PCR was faster and more economical using larger pools. Results indicate that surveillance of JE virus in mosquitoes using the semi-nested PCR is an alternative to monitoring seroconversions in sentinel pigs.

Detection of West Nile virus antigen in mosquitoes and avian tissues by a monoclonal antibody-based capture enzyme immunoassay

An antigen capture immunoassay to detect West Nile (WN) virus antigen in infected mosquitoes and avian tissues has been developed. With this assay purified WN virus was detected at a concentration of 32 pg/0.1 ml, and antigen in infected suckling mouse brain and laboratory-infected mosquito pools could be detected when the WN virus titer was 10(2.1) to 10(3.7) PFU/0.1 ml. In a blindly coded set of field-collected mosquito pools (n = 100), this assay detected WN virus antigen in 12 of 18 (66.7%) TaqMan-positive pools, whereas traditional reverse transcriptase PCR detected 10 of 18 (55.5%) positive pools. A sample set of 73 organ homogenates from naturally infected American crows was also examined by WN virus antigen capture immunoassay and TaqMan for the presence of WN virus. The antigen capture assay detected antigen in 30 of 34 (88.2%) TaqMan-positive tissues. Based upon a TaqMan-generated standard curve of infectious WN virus, the limit of detection in the antigen capture assay for avian tissue homogenates was approximately 10(3) PFU/0.1 ml. The recommended WN virus antigen capture protocol, which includes a capture assay followed by a confirmatory inhibition assay used to retest presumptive positive samples, could distinguish between the closely related WN and St. Louis encephalitis viruses in virus-infected mosquito pools and avian tissues. Therefore, this immunoassay demonstrates adequate sensitivity and specificity for surveillance of WN virus activity in mosquito vectors and avian hosts, and, in addition, it is easy to perform and relatively inexpensive compared with the TaqMan assay.

Post-discharge surveillance: can patients reliably diagnose surgical wound infections?

Post-discharge surgical wound infection surveillance is an important part of many infection control programs. It is frequently undertaken by patient self-assessment, prompted either by a telephone or postal questionnaire. To assess the reliability of this method, 290 patients were followed for six weeks post-operatively. Their wounds were photographed and also covertly assessed for signs of infection by two experienced infection control nurses (ICNs). Patients also responded to a postal questionnaire seeking evidence of infection at both week four and week six post-surgery. Correlation between the patient's assessment of their wound and the ICNs diagnosis was poor (r = 0. 37) with a low positive predictive value (28.7%), although negative predictive value was high (98.2%). Assessment of photos for signs of infection by two experienced clinicians also correlated poorly with the ICNs diagnosis of infection (r = 0.54). The patient's recall of prescription of an antibiotic by their general practitioner (GP) for wound infection during the postoperative period correlated best with the ICNs diagnosis (r = 0.76). This latter measure, particularly when confirmed by the GP in those patients reporting an infection, appears to provide the most valid and resource efficient marker of post-discharge surgical wound infection.

Control of Aedes vectors of dengue in three provinces of Vietnam by use of Mesocyclops (Copepoda) and community-based methods validated by entomologic, clinical, and serological surveillance

We describe remarkable success in controlling dengue vectors, Aedes aegypti (L.) and Aedes albopictus (Skuse), in 6 communes with 11,675 households and 49,647 people in the northern provinces of Haiphong, Hung Yen, and Nam Dinh in Vietnam. The communes were selected for high-frequency use of large outdoor concrete tanks and wells. These were found to be the source of 49.6-98.4% of Ae. aegypti larvae, which were amenable to treatment with local Mesocyclops, mainly M. woutersi Van der Velde, M. aspericornis (Daday) and M. thermocyclopoides Harada. Knowledge, attitude, and practice surveys were performed to determine whether the communities viewed dengue and dengue hemorrhagic fever as a serious health threat; to determine their knowledge of the etiology, attitudes, and practices regarding control methods including Mesocyclops; and to determine their receptivity to various information methods. On the basis of the knowledge, attitude, and practice data, the community-based dengue control program comprised a system of local leaders, health volunteer teachers, and schoolchildren, supported by health professionals. Recycling of discards for economic gain was enhanced, where appropriate, and this, plus 37 clean-up campaigns, removed small containers unsuitable for Mesocyclops treatment. A previously successful eradication at Phan Boi village (Hung Yen province) was extended to 7 other villages forming Di Su commune (1,750 households) in the current study. Complete control was also achieved in Nghia Hiep (Hung Yen province) and in Xuan Phong (Nam Dinh province); control efficacy was greater than or equal to 99.7% in the other 3 communes (Lac Vien in Haiphong, Nghia Dong, and Xuan Kien in Nam Dinh). Although tanks and wells were the key container types of Ae. aegypti productivity, discarded materials were the source of 51% of the standing crop of Ae. albopictus. Aedes albopictus larvae were eliminated from the 3 Nam Dinh communes, and 86-98% control was achieved in the other 3 communes. Variable dengue attack rates made the clinical and serological comparison of control and untreated communes problematic, but these data indicate that clinical surveillance by itself is inadequate to monitor dengue transmission.