Quantitative Use of Fluorescent In Situ Hybridization To Examine Relationships between Mycolic Acid-Containing Actinomycetes and Foaming in Activated Sludge Plants

Abstract

The formation of viscous foams on aeration basins and secondary clarifiers of activated sludge plants is a common and widespread problem. Foam formation is often attributed to the presence of mycolic acidcontaining actinomycetes (mycolata). In order to examine the relationship between the number of mycolata and foam, we developed a group-specific probe targeting the 16S rRNA of the mycolata, a protocol to permeabilize mycolata, and a statistically robust quantification method. Statistical analyses showed that a lipase-based permeabilization method was quantitatively superior to previously described methods (P << 0.05). When mixed liquor and foam samples were examined, most of the mycolata present were rods or cocci, although filamentous mycolata were also observed. A nested analysis of variance showed that virtually all of the measured variance occurred between fields of view and not between samples. On this basis we determined that as few as five fields of view could be used to give a statistically meaningful sample. Quantitative fluorescent in situ hybridization (FISH) was used to examine the relationship between foaming and the concentration of mycolata in a 20-m 3 completely mixed activated sludge plant. Foaming occurred when the number of mycolata exceeded a certain threshold value. Baffling of the plant affected foaming without affecting the number of mycolata. We tentatively estimated that the threshold foaming concentration of mycolata was about 2 ؋ 10 6 cells ml ؊1 or 4 ؋ 10 12 cells m ؊2. We concluded that quantitative use of FISH is feasible and that quantification is a prerequisite for rational investigation of foaming in activated sludge.Microbe-based treatment problems occur in the plants of even the most sophisticated and experienced water utilities. One of the most intractable and widespread problems is the formation of viscous foams in activated sludge plants. Such foams appear as bacterial biomass floating on the surfaces of aeration basins and secondary sedimentation tanks. The presence of foam may lead to severe operational problems and may cause plants to fail effluent standards. The occurrence of foam is unpredictable, and there is uncertainty about the cause and mechanism of foaming; the control mechanisms used currently are empirical and frequently ineffective (45). It is thought that foams may form when gas bubbles are stabilized by the presence of hydrophobic particles and surfactants; the bubbles then rise to the surface and accumulate (45). The hydrophobic particles are assumed to be bacteria.Microscopic examination of foams usually reveals the presence of large numbers of one or two filamentous bacterial morphotypes. It is assumed that the morphotypes that are branched are mycolic acid-containing actinomycetes (mycolata) (13, 48) and that unbranched filaments are Microthrix parvicella (45). Putative mycolata morphotypes are the most widely reported morphotypes (39,43) and are often referred to as "nocardia." However, a wide range of mycolata are now associated with foaming. Plate cou...

Influence of Operating Parameters on the Biodegradation of Steroid Estrogens and Nonylphenolic Compounds during Biological Wastewater Treatment Processes

“…The removal efficiencies in this 199 study support these findings and were between 78 -80% in both the N/DN and N/DN-P 200 works for E1+E2+E3 which both had equivalent SRTs of up to 13 days (Table 1). 201 It has been postulated that increasing the sludge age increases the diversity of the 263 consortia of bacteria present in a treatment plant allowing the growth of EDC degrading 264 organisms (40) and that the ability to remove EDCs is assumed to be a property of some 265 of the slower growing organisms that can only colonise the treatment plant at long sludge 266 ages. It is unlikely that the presence of EDCs are specifically selecting for these 267 organisms, as the low concentrations of EDC found in wastewaters could only support a 268 small number of cells.…”

Evaluating the Measurement of Activated Sludge Foam Potential

“…and the filamentous species Microthrix parvicella have been associated with activated sludge foaming [22]. Such work has lead to the principle of threshold numbers of mycolata being required for the formation of stable activated sludge foams [7,40]. A subjective approach to assessing activated sludge foaming has been to simply classify foams using the degree of aeration basin foam coverage [41], although it has not always been possible to directly correlate foam coverage with the abundance of suspected foam-causing bacteria [2].…”

“…It is the stability and persistence of these foams which differentiates them from other bubble entities seen on the surface of aeration tanks [1]. In large volumes foam can result in a multitude of operational problems including: reduction in plant performance, physical hazard to operators from exposure to pathogens and walkway obstruction [2][3][4][5], blockage of pipes [1], reduction in oxygen transfer [6], interference with essential plant monitoring equipment [2], and risk to consent failure thorough solids loss from the final settlement stage of the treatment process [7][8][9]. Recently, the presence of foam has also been recognised in systems which employ advanced wastewater treatment such as membrane bioreactors (MBR) [10].…”

“…Studies have shown that sites suffering from activated sludge foaming are often complemented by the presence of large quantities of the filamentous species Microthrix parvicella and/or nocardioform actinomycetes which are found in large amounts within the foam and mixed liquor [2]. These filamentous species are strongly hydrophobic due to having cell walls composed of mycolic acid [7,[16][17][18]. In the absence of such species, non-stable foams are created [19].…”

Detección y caracterización por métodos fenotípicos y moleculares de mycolata formadores de espumas en estaciones depuradoras de aguas residuales domésticas con sistemas de fangos activos.

“…La sonda S-*-Myc-0657-a-A-16 diseñada por Davenport et al (2000), está dirigida a una región del rRNA 16S específica para todos los mycolata.…”