Increased biogas production at wastewater treatment plants through co-digestion of sewage sludge with grease trap sludge from a meat processing plant

The feasibility of co-digesting grease trap sludge from a meat-processing plant and sewage sludge was studied in batch and reactor experiments at 35 degrees C. Grease trap sludge had high methane production potential (918 m(3)/tVS(added)), but methane production started slowly. When mixed with sewage sludge, methane production started immediately and the potential increased with increasing grease trap sludge content. Semi-continuous co-digestion of the two materials was found feasible up to grease trap sludge addition of 46% of feed volatile solids (hydraulic retention time 16d; maximum organic loading rate 3.46 kgVS/m(3)d). Methane production was significantly higher and no effect on the characteristics of the digested material was noticed as compared to digesting sewage sludge alone. At higher grease trap sludge additions (55% and 71% of feed volatile solids), degradation was not complete and methane production either remained the same or decreased.     

Efficiency of different enrichment and isolation procedures for the detection of Salmonella serotypes in edible offal

Rapid detection systems for Salmonella in foodstuffs are currently being developed. However, existing standards still call for application of traditional methods employing pre-enrichment followed by selective enrichment and isolation. The efficacy of various methods was tested using 264 chicken and lamb organ meats. Pre-enrichment was carried out in Tryptone Soy Broth (TSB) and enrichment in Tetrathionate Brilliant Green Broth (TTB) at 37°C, Selenite Broth with Brilliant Green and Sulphapyridine at 37°C and 43°C, and Rappaport-Vassiliadis Broth (RV 10) at 42°C. The isolation media were Brilliant Green Agar (BGA), Deoxycholate Citrate Agar, Hektoen Enteric Agar (HEA) and Salmonella-Shigella Agar.
Enrichment in RV/42°C followed by isolation on BGA as recommended by ISO standard no. 6579 and enrichment in TTB/37°C followed by isolation in HEA, no longer recommended by that standard, produced the best results. Low percentages of positive samples and difficulties in detecting Salmonella are the result of interference by competing organisms (Enterobacteriaceae) and the number of salmonellas present after enrichment.
A total of 528 samples (TSB, eggs, lamb liver and chicken liver) were inoculated with Salm. enteritidis, Salm. kapemba and Salm. virchow , and the preceding experiment was repeated. All the TSB and egg samples tested positive, but the percentage of positive samples from the lamb and chicken liver was only 81–92%. Recovery of the salmonellas did not depend upon the method employed or the serotype inoculated but instead on interference by competing flora and the numbers of Salmonella present in the samples.     

Evaluation of Culture Media for the Isolation of Salmonellas from Sewage Sludge

Ten samples of sewage sludge were examined by various methods for the isolation of salmonellas using three types of enrichment broth: Muller-Kauffmann Tetrathionate Broth (MKTB), Selenite F Broth (SFB), and Brilliant Green Broth (BGB), two temperatures (37°C and 43°C) and three selective media: Deoxycholate Citrate Agar incubated aerobically (DCA), and anaerobically (DCA(N)), Brilliant Green Agar (BGA), and Bismuth Sulphite Agar (BSA). The results suggest that a combination of pre-enriched MKTB incubated at 37°C and plated on to BGA at 24 and 48 h was the best method, but when examining contaminated material such as sewage sludge, it appears unwise to rely on one single method.     

Sanitation of faeces from source-separating dry toilets using urea

Aim:  To develop a reliable and simple method to produce safe fertilizers from human excreta using urea for sanitation of faeces.
Methods and Results:  Urea was added to faecal matter (17% dry matter) at concentrations of 0·5–2% (w/w) and inactivation of Salmonella enterica subspecies 1 serovar Typhimurium (Salm. Typhimurium), Enterococcus spp . and the Salm. Typhimurium bacteriophage 28B was monitored at 14, 24 and 34°C. Urea additions enhanced inactivation and inactivation rates were positively related to increasing NH₃ (aq) concentration and temperature. Salm. Typhimurium was the most sensitive of the organisms studied, while Enterococcus spp. showed more persistence, especially at lower temperatures. The bacteriophage was the most resistant organism studied.
Conclusions:  Salmonella reduction levels that meet requirements for safe reuse of faeces as fertilizer (i.e. 6 log₁₀ reduction) can be achieved for 1% urea within 2 months at 14°C or within 1 week at 24°C and 34°C.
Significance and Impact of the Study:  The relationships between organism inactivation rates and temperature, ammonia and pH were identified. Urea treatment proved to be a robust and efficient option for safe recycling of plant nutrients.     

Survival of dehydrated cells of Salmonella typhimurium LT2 at high temperatures

Cells of Salmonella typhimurium LT2 were dehydrated on hydrophobic membranes (Millipore FGLP2500) placed in a controlled atmosphere chamber held at 57% equilibrium relative humidity (ERH) and 37°C. Dehydration for 48 h under the above conditions increased the heat resistance of Salm. typhimurium LT2 when measured as the surviving fraction after a heat challenge of 135°C for 30 min. Results also showed that little or no death occurred during heat challenges of 1 h at temperatures of up to 100°C. The survival of Salm. typhimurium LT2 was measured as the ability to form colonies on solid media tryptone soy broth plus 1.2% agar (TSBA) after 24 h at 37°C. Incorporation of sodium pyruvate, at a concentration of 0.2% into the recovery medium, did not enhance the recovery of heated Salm. typhimurium LT2. Dehydrated cells of S. typhimurium LT2 showed a triphasic death curve. Increasing the period of dehydration from 48 h to 34 d, reduced initial numbers due to die off but did not alter the shape of the subsequent survival curve. and accepted 22 June 1989     

Factors affecting the survival of Salmonella and Escherichia coli in anaerobically fermented pig waste

The survival of Salmonella typhimurium was investigated in acidogenic, anaerobically fermented pig wastes and in synthetic media, each containing volatile fatty acids (VFA). Salm. typhimurium survived at pH 6·8, but not at pH 4·0, when incubated at 37°C for 24 h in either fermented or synthetic medium containing VFA. The minimum inhibiting concentration of VFA for Salm. typhimurium after 48 h incubation at 30°C at pH 4·0 was 0·03 mol/l and for Escherichia coli it was 0·09 mol/l. Fermented pig wastes in a digester, maintained at pH 5·9, were inoculated with Salm. typhimurium and then incubated at 37°C for 24 h. The pH was adjusted to either 4·0 or 5·0 and after a further 48 h at 30°C, Salm. typhimurium survived at pH 5·0 but not at pH 4·0. It was concluded that pH is critical in determining the survival of this organism in acidogenic anaerobically fermented pig waste.     

Salmonella in animal slurry can be destroyed by aeration at low temperatures

Cattle and other animals infected by Salmonella can emit high numbers of these bacteria. To determine an effective means for reducing this bacterial group in animal slurry, samples were subjected to aeration in laboratory experiments and in farm-scale slurry tanks. A clear reduction in Salmonella levels was found in laboratory experiments at temperatures from 4 to 40 °C. Aeration in farm-scale slurry tanks increased the temperature above the ambient temperatures (often less than 0 °C) to maxima ranging between 19 and 40 °C. Farm-scale aeration resulted in similar reductions in Salmonella as those achieved in laboratory experiments. Thus, reductions, ranging from greater than 99% of the initial number to no detectable Salmonella , could be reached after 2–5 weeks using aeration processes with cattle slurries contaminated by Salm. infantis or pig slurry contaminated by Salm. typhimurium . These results suggest that farmers can control the spread of Salmonella from slurry to agricultural fields. The reduction mechanisms remain unknown, though the increase in pH (to 7·6–9·0) found in slurries after aeration might exert a decreasing effect on these bacteria.     

Discovery of an agent in wastewater sludge that reduces the heat required to inactivate reovirus.

An agent that causes heat inactivation of reovirus to occur at reduced temperatures has been found in both raw and anaerobically digested sludge. This agent is originally associated with sludge solids but can be washed from these solids by blending with water. The activity of the agent was considerably greater in alkaline than in acid solutions, probably because it is insoluble at low pH. The agent was shown to be nonvolatile and heat stable up to 300 degrees C but was inactivated within 30 min at 400 degrees C. The rate of heat inactivation of reovirus by the agent was found to occur in a bimodal fashion and to be relatively rapid, even at 35 degrees C. Finally, it was found that this agent did not accelerate heat inactivation of poliovirus, but instead may be the component of sludge previously found to protect poliovirus against inactivation by heat.     

The removal of Salmonella enteritidis in activated sludge

Salmonella destruction efficiencies of 99% were obtained after 10 h aeration at 15°C in a laboratory model of the activated sludge process. This study demonstrated that, in a batch process, the removal of salmonellas occurred in three phases. (i) By 4 h, 90% of the original inoculum had disappeared from the activated sludge, probably due mainly to predation by ciliated protozoa. The remaining 10% was distributed between the liquid phase (approximately 90%) and the sludge floc (approximately 10%). (ii) During the next 2 h this situation was inverted so that, by 6 h, more than 80% of the remaining salmonellas were then adsorbed to floc, leaving less than 20% in liquid suspension. (iii) From 6 h onwards there was a much slower decline of the remaining salmonellas attached to floc. The addition of dioctyl sodium sulphosuccinate after 4 h, inactivated the ciliated protozoa populations and completely eliminated the continued reduction of salmonellas from activated sludge observed previously.     

Discovery of an agent in wastewater sludge that reduces the heat required to inactivate reovirus.

An agent that causes heat inactivation of reovirus to occur at reduced temperatures has been found in both raw and anaerobically digested sludge. This agent is originally associated with sludge solids but can be washed from these solids by blending with water. The activity of the agent was considerably greater in alkaline than in acid solutions, probably because it is insoluble at low pH. The agent was shown to be nonvolatile and heat stable up to 300 degrees C but was inactivated within 30 min at 400 degrees C. The rate of heat inactivation of reovirus by the agent was found to occur in a bimodal fashion and to be relatively rapid, even at 35 degrees C. Finally, it was found that this agent did not accelerate heat inactivation of poliovirus, but instead may be the component of sludge previously found to protect poliovirus against inactivation by heat.     

Microbial populations and sludge characteristics in thermophilic aerobic sewage sludge digestion

Summary Estimates of bacterial numbers from raw sewage sludge and sludge treated by thermophilic aerobic digestion were compared with simple indicators of sludge quality and concentrations of potential substrates. Significant differences were found between sludge types for all but one of the variables examined (frequency of dividing cells). During a stable period of digestor operation, the average number of viable obligate thermophiles present in digested sludge (1.63 × 10⁶ ml^–1) was approximately 10²-fold greater than in feed sludge (1.10 × 10⁴ ml^–1). Total numbers of bacteria were slightly greater in digested sludge (3.24 × 10¹⁰ ml^–1) than in feed sludge (2.39 × 10 ml^–10), as were viable counts of bacteria at incubation temperatures of 37°C and 55°C. Significant correlation was found between viable counts of bacteria at 37°C and 55°C for digested sludge, and 65°C and 55°C for feed sludge. The numbers of obligate thermophiles present and the total of bacteria present were related to the temperature and pH of the digested sludge and inversely related to the numbers ofEscherichia coli and coliforms present, which were not detected at temperatures greater than 50°C.     

The Effects of pH and Levels of Organic Matter on the Death Rates of Salmonellas in Chicken Scald-tank Water

The immersion scalding of chicken carcases at 52°C for 2.5 min was found to result in the accumulation of bacteria and organic matter in the water. Disassociation of ammonium urate present in chicken faeces caused the pH value of the water to fall rapidly from 8.4 to 6.0. This pH was maintained for most of the time the tank was in operation. The change in pH value greatly increased the heat resistance of some common salmonella serotypes with a strain of Salmonella typhimurium being the most resistant, having a D52°C value of 34.5 min. The pH value of the water was found to be close to the optimum for the heat resistance of salmonellas. In scald water at pH 5.9–6.0 the death rate of Salm. typhimurium was shown to be independent of organic matter levels indicating that pH was the major influence on heat resistance.     

The growth of salmonellas in hydrated animal feed supplemented with antibiotics

The growth of antibiotic-sensitive salmonellas in hydrated animal feed incubated at 37°C was relatively unaffected by high concentrations (> 800 ppm) of bacteriostatic agents such as chlortetracycline. On the other hand, they were killed by low concentrations (< 50 ppm) of bactericidal agents such as ampicillin or furazolidone. The aminoglycoside antibiotics apramycin and neomycin killed the salmonellas when present at concentrations in excess of those likely to be recommended for therapeutic purposes (> 300 ppm).     

Dual anaerobic co-digestion of sewage sludge and confectionery waste

Three configurations for a dual digestion system were examined. The units were based on three 5 l completely stirred tank reactors (CSTR). A first-stage thermophilic digester was used to provide the feed to each of the two second-stage mesophilic (35°C) digesters. Using a mixture of sewage sludge and strong confectionery waste, the thermophilic digester was operated at 55°C with a hydraulic retention time of 4 h. The mesophilic digesters were operated at hydraulic retention times of 8, 12 and 15 days. In terms of the reduction of volatile solids (VS), the three dual digestion configurations were similar but were more effective than the single-stage reactor which was used as a control. However, based on the specific methane yield (m³ CH₄/kg VS removed), the configuration with a first stage operating at 55°C and a secondary digester at 35°C with a hydraulic retention time of 12 days was the most effective. This configuration also maintained a more stable pH, irrespective of the quality of the feed sludge.     

Inactivation of poliovirus in digested sludge.

The effect of anaerobically digested sludge on poliovirus during incubation at temperatures between 28 and 4 C was studied. Although virus was fully recoverable from sludge, its infectivity decreased in proportion to the time and temperature of incubation. The rate ranged from greater than 1 log per day at 28 C to about 1 log every 5 days at 4 C. The mechanism of inactivation was studied at the lower temperature where the sedimentation coefficients of most inactivated particles were not detectably modified. The ribonucleic acid (RNA) of these particles appeared to have been nicked and had an average sedimentation value about 70% that of RNA from infectious virus. Since the specific infectivity of RNA from particles recovered from sludge was directly proportional to that of the particles from which it was extracted, loss of infectivity was probably due to inactivation of RNA. Some breakdown was also found in the two largest viral proteins of inactivated particles. Thus, the mechanism of inactivation may be cleavage of viral proteins followed by nicking of encapsulated RNA. Because no virucidal activity was found in raw sludge, this component of digested sludge appears to be a product of the digestion process.     

Inactivation of poliovirus in digested sludge.

The effect of anaerobically digested sludge on poliovirus during incubation at temperatures between 28 and 4 C was studied. Although virus was fully recoverable from sludge, its infectivity decreased in proportion to the time and temperature of incubation. The rate ranged from greater than 1 log per day at 28 C to about 1 log every 5 days at 4 C. The mechanism of inactivation was studied at the lower temperature where the sedimentation coefficients of most inactivated particles were not detectably modified. The ribonucleic acid (RNA) of these particles appeared to have been nicked and had an average sedimentation value about 70% that of RNA from infectious virus. Since the specific infectivity of RNA from particles recovered from sludge was directly proportional to that of the particles from which it was extracted, loss of infectivity was probably due to inactivation of RNA. Some breakdown was also found in the two largest viral proteins of inactivated particles. Thus, the mechanism of inactivation may be cleavage of viral proteins followed by nicking of encapsulated RNA. Because no virucidal activity was found in raw sludge, this component of digested sludge appears to be a product of the digestion process.     

Formation of viable but nonculturable Salmonella during starvation in chemically defined solutions

Salmonella enteritidis enters a viable-but-nonculturable state when exposed to starvation in aquatic environments. This study determined starvation survival of this pathogen in chemically defined solutions and tested the ability of nonselective enrichment to detect viable-but-nonculturable cells. Starvation of Salm. enteritidis at 7°C in 7.35 mmol 1^-1 potassium phosphate buffer resulted in complete loss of culturability after 5 weeks with maintenance of a substrate-responsive population of over 10000 cell ml^-1. Starvation at 21°C and starvation in saline solutions or lower concentrations of phosphate buffer resulted in prolonged survival of a culturable population although this population was lower than the total viable population. Enrichment using lactose broth did not allow resuscitation of viable-but-nonculturable cells even after 5 d of incubation at 35°C.     

Comparison of Muller-Kauffmann tetrathionate broth with Rappaport-Vassiliadis (RV) medium for the isolation of salmonellas from sewage sludge

The Rappaport-Vassiliadis (RV, modification of Rappaport's) medium was superior to Muller-Kauffmann tetrathionate broth (MKT) for the enrichment and isolation of salmonellas from sewage sludge samples, either naturally contaminated or artificially inoculated with salmonella. Isolation rates for RV (42°C) were higher even when MKT (43°C) performance was improved by the use of brilliant green sulphamandelate agar. Kinetic results show that the suppression of the competing flora was poorer in MKT than in RV. The use of high inoculum to medium volume ratios (e.g. 1:5000) increased the isolation rates from RV and kinetic data explain why this was so.     

The effect of slurry storage and anaerobic digestion on survival of pathogenic bacteria

The decline in viable numbers of Salmonella typhimurium, Yersinia enterocolitica and Listeria monocytogenes in beef cattle slurry is temperature-dependent; they decline more rapidly at 17°C than at 4°C. Mesophilic anaerobic digestion caused an initial rapid decline in the viable numbers of Escherichia coli, Salm. typhimurium, Y. enterocolitica and L. monocytogenes. This was followed by a period in which the viable numbers were not reduced by 90%. The T₉₀ values of E. coli, Salm. typhimurium and Y. enterocolitica ranged from 0.7 to 0.9 d during batch digestion and 1.1 to 2.5 d during semi-continuous digestion. Listeria monocytogenes had a significantly higher mean T₉₀ value during semi-continuous digestion (35.7 d) than batch digestion (12.3 d). Anaerobic digestion had little effect in reducing the viable numbers of Campylobacter jejuni.