We demonstrated that a temperature-phased anaerobic bioprocess can convert acid whey from Greek-yogurt production into valuable medium-chain carboxylic acids (MCCAs). Thermophilic and mildly acidic conditions in the first phase promoted a Lactobacillus spp.-dominated microbiome that converted sugars from acid whey into lactic acid. The lactic acid-rich effluent was then fed to a mesophilic second phase in which a more diverse microbiome performed chain elongation to produce MCCAs. The overall SCOD conversion efficiency for acid-whey conversion into MCCAs was 53.5%.

Discovering new ways to inactivate pathogens in human waste is critical for the improvement of worldwide access to sanitation and for the reduction of the environmental impact of conventional waste treatment processes. Here, we utilized the carboxylate platform and chain elongation to produce n-butyric acid and n-caproic acid via the anaerobic fermentation of human fecal material. Then, we inactivated Ascaris eggs through exposure to these carboxylic acids. Using batch experiments with human fecal material as substrate, we accumulated n-butyric acid and n-caproic acid at total concentrations (uncharged acid plus conjugate base) of 257 and 27.1 mM, respectively. We then showed that carboxylic acids at these concentrations inactivated Ascaris eggs when the pH was below the pK for the acids, causing them to exist primarily in the uncharged forms. We observed that uncharged carboxylic acids affected viability rather than the pH itself or conjugate bases. In addition, we modeled the viability of Ascaris eggs as a function of uncharged carboxylic acid concentration for n-butyric acid and n-caproic acid at exposure times of 2, 6, 12, and 20 days. The results presented here indicate that in situ biological production of carboxylic acids in HFM provides a promising method of pathogen inactivation and may lead to new developments in sanitation technology and treatment of fecal sludge.

In the U.S., several states have attempted to mitigate greenhouse-gas emissions by banning food wastes from landfills. As a result, U.S.-based companies are now providing decentralized food-waste management systems for supermarkets and restaurants, which include storage as a slurry. It is unclear, however, which storage conditions (factors) would affect the spontaneous microbial activity, resulting in a different fermentation product spectra, and how this would further affect post-treatment. Here, we performed two experiments to mimic: (1) storage and (2) subsequent anaerobic digestion. For the food-waste storage system, we designed a mixed-level fractional factorial analysis with 12 experimental combinations, including separating food waste into: carbohydrate-rich, lipid-rich, and protein-rich food waste. We found that all factors that we tested affected the fermentation outcome. We observed that relatively low pH levels of 3–4, which were achieved due to rapid lactate accumulation by microbial activity during storage, coincided with greater lactate production and a maximum chemical oxygen demand (COD) selectivity of 90%. Food-waste storage followed classical ensilage dynamics with homofermentation to lactate in combination with low pH preventing the subsequent breakdown of lacate into other carboxylic acids and hydrogen gas. The mechanistic understanding provides an opportunity to optimize lactate production, which is ideal for subsequent methane or chemical production.

Ascaris eggs are commonly used as indicators for pathogen inactivation during the treatment of fecal sludge and wastewater due to their highly resistant lipid membrane and ability to survive in the environment for long periods of time. Current guidelines suggest that thermal treatment alone cannot inactivate Ascaris eggs at temperatures below 45 °C, although some evidence in the literature suggests this to be incorrect. Here, we performed a controlled experiment to test the effect of mesophilic temperatures on Ascaris inactivation. We exposed Ascaris suum eggs to a temperature gradient between 34°C and 45 °C under anaerobic and aerobic conditions to observe the required exposure times for a 3-log reduction. Indeed, we found that temperatures lower than 45 °C did inactivate these eggs, and the required exposure times were up to two orders of magnitude shorter than suggested by current guidelines. Results from the anaerobic exposures were used to develop a time-temperature relationship that is appropriate for Ascaris inactivation at mesophilic temperatures. Data from the literature demonstrated that our relationship is conservative, with faster inactivation occurring under environmental conditions when Ascaris eggs were suspended in fecal sludge or manure. A specific aerobic relationship was not developed, but we demonstrated that aerobic conditions cause faster inactivation than anaerobic conditions. Therefore, the anaerobic relationship provides a conservative guideline for both conditions. We demonstrate that relatively low temperatures can considerably impact Ascaris viability and suggest that mesophilic temperatures can be used in waste treatment processes to inactivate pathogens. The development of safe, low-input, mesophilic treatment processes is particularly valuable for ensuring universal access to safe sanitation and excreta management.

Innovative, low-cost methods for inactivating pathogens in human fecal material (HFM) are needed, particularly in expanding urban areas where conventional sewer systems and centralized wastewater treatment plants are not feasible. To address this challenge, we have developed a bioprocess that utilizes open cultures of anaerobic bacteria (i.e., microbiomes) to produce carboxylic acids using HFM as substrate. When the pH is sufficiently low, the carboxylic acids exist in the undissociated form and inactivate pathogens. Here, we used real solid waste (SW) collected from container-based, urine-diverting dry toilets (UDDT) in Nairobi, Kenya to conduct lab-scale and field-scale trials. Through these trials, we investigated operating conditions required to use carboxylic acid fermentation in sanitation waste treatment processes. We tested three different inoculum treatments and determined that the microbiome in UDDT-SW is wellsuited to produce carboxylic acids without the need for an external inoculum. We also tested co-fermentation of UDDT-SW with carbohydrate-rich food waste as a means of reducing the pH. We found that when food waste was incorporated in a way that maintained the pH between 4.8 and 5.2, then the food waste was quickly converted to carboxylic acids, and the low pH created high concentrations of undissociated carboxylic acids. The resulting concentrations of undissociated carboxylic acids resulted in Ascaris inactivation within 15 days. However, we found that a temperature ≥30 • C is required for carboxylic acid production to occur.