Enumeration of heterotrophs, fecal coliforms and Escherichia coli in water: comparison of 3M Petrifilm plates with standard plating procedures

A total of 177 naturally contaminated water samples were analyzed by membrane filtration according to the Standard Methods for the Examination of Water and Wastewater published by the American Public Health Association. Filters were incubated in parallel on mHPC-agar and 3M™ Petrifilm™ Aerobic Count Plates (Petrifilm™ AC plates) for heterotrophic counts. Fecal coliforms and Escherichia coli were enumerated on mFC-agar and 3M™ Petrifilm™ E. coli/Coliform Count Plates (Petrifilm™ EC plates). Typical colonies on each media type were confirmed following standard procedures. Heterotrophic counts were between 10³ and 10⁴ CFU/mL and the average log₁₀ counts obtained on Petrifilm™ AC plates were about two-fold lower than on mHPC-agar. Counts for fecal coliforms and E. coli were between 10² and 10³ CFU/mL. Average log₁₀ counts for confirmed fecal coliforms obtained on Petrifilm™ EC plates were slightly lower than on mFC agar with a correlation coefficient of 0.949. The average log₁₀ counts for confirmed E. coli on Petrifilm™ EC plates and on mFC agar were statistically not different (P=0.126) with a correlation coefficient of 0.879. Specificity of Petrifilm™ EC plates and mFC agar was evaluated by comparing typical colony counts with confirmed counts. On mFC agar, counts for typical colonies were by 2 log₁₀ CFU higher than the actual confirmed counts. In contrast, on Petrifilm™ EC plates typical colony counts were almost identical to confirmed colony counts for both fecal coliforms and E. coli. This comparison illustrates the high specificity of Petrifilm™ EC plates for enumeration of both fecal coliforms and E. coli in water.     

Treatment of sewage water from tourist areas by anaerobic fixed-bed reactor

A laboratory-scale anaerobic fixed-bed reactor, operating at ambient temperature (30 to 35°C), was used to treat sewage water from tourist areas in Cuba at hydraulic retention times (HRT) ranging from 4 to 72 h. The total chemical oxygen demand (T-COD), total biological oxygen demand (T-BOD) and total suspended solids removal varied between 30 and 80%, 40 and 95% and 25 and 80%, respectively. Total and faecal coliforms were reduced by 98.1 to 99.9% and by 99.0% to 99.9% respectively, despite the marked decrease in HRT from 72 to 4 h.     

Faecal pellets in streams: their binding, breakdown and utilization

1. Faecal pellets of Gammarus (shredders) and Simulium larvae (suspension feeders) are bound by exopolymers. Immediately after egestion, Gammarus pellets are covered by a peritrophic membrane that breaks up within hours, although pellets remain intact because of internal binding materials. 2. Although they expand soon after egestion, the faecal pellets of Gammarus and Simulium remain intact for more than 30 days. Their internal structure is altered and the main agents of this change are bacteria that have survived passage through the gut (and become bound within pellets). 3. When disrupted physically, freshly egested (1‐ to 2‐day old) Simulium faecal pellets break up into relatively large pieces whereas freshly egested Gammarus faecal pellets break apart into much smaller pieces. Disruption of 30‐day old Simulium faecal pellets results in similar sized pieces to those from freshly egested pellets, but disruption of 30‐day old Gammarus pellets produces pieces that are two orders of magnitude larger than those resulting from disruption of freshly egested pellets. 4. Faecal pellets of Gammarus and Simulium are eaten by stream invertebrates and are sites of microbial breakdown. Faecal pellets are a source of organic matter for benthic invertebrates, bacteria and, indirectly, for plants.     

Review: Markers and proxies to monitor ruminal function and feed efficiency in young ruminants

Developing the rumen’s capacity to utilise recalcitrant and low-value feed resources is important for ruminant production systems. Early-life nutrition and management practices have been shown to influence development of the rumen in young animals with long-term consequences on their performance. Therefore, there has been increasing interest to understand ruminal development and function in young ruminants to improve feed efficiency, health, welfare, and performance of both young and adult ruminants. However, due to the small size, rapid morphological changes and low initial microbial populations of the rumen, it is difficult to study ruminal function in young ruminants without major invasive approaches or slaughter studies. In this review, we discuss the usefulness of a range of proxies and markers to monitor ruminal function and nitrogen use efficiency (a major part of feed efficiency) in young ruminants. Breath sulphide and methane emissions showed the greatest potential as simple markers of a developing microbiota in young ruminants. However, there is only limited evidence for robust indicators of feed efficiency at this stage. The use of nitrogen isotopic discrimination based on plasma samples appeared to be the most promising proxy for feed efficiency in young ruminants. More research is needed to explore and refine potential proxies and markers to indicate ruminal function and feed efficiency in young ruminants, particularly for neonatal ruminants.     

Uptake and persistence of human associated Enterococcus in the mussel Mytilus edulis: relevance for faecal pollution source tracking

Aims:  Micro-organisms and molecular markers for microbial source tracking (MST) in coastal waters are often present at low numbers, and often exhibit significant variability in time and space. In this study, we investigated the uptake, accumulation, and persistence of human associated Enterococcus in the mussel Mytilus edulis .
Methods and Results:  The human associated molecular markers esp in Enterococcus faecium , and M66 in Enterococcus faecalis were targetted by PCR in seawater and mussel samples from coastal sites affected by sewage contamination. Both native mussels and mussels transplanted from pristine to polluted sites were included. The results showed that the esp and M66 markers were often not detectable in seawater whereas mussels were enriched in the markers. Human associated E. faecalis accumulated rapidly in M. edulis , and reached maximum levels after 4–6 h with concentration 30–300 times greater than in the surrounding seawater. Enterococcus faecalis retained in M. edulis showed a survival comparable to planktonic E. faecalis in seawater with half lives of 30 and 22 h, respectively. Human associated markers remained detectable for 120 h in M. edulis after faecal contamination.
Conclusions:  The study demonstrated that native and transplanted M. edulis can accumulate and retain human associated molecular markers relevant for MST.
Significance and Impact of the Study:  Mussels should be considered as additional targets in MST studies in coastal waters.     

Phylogenetic analysis of Bacteroidales 16S rRNA gene sequences from human and animal effluents and assessment of ruminant faecal pollution by real‐time PCR

Aims: The aims of this study were to evaluate the host‐specific distribution of Bacteroidales 16S rRNA gene sequences from human‐ and animal‐related effluents and faeces, and to define a ruminant‐specific marker. Methods and Results: Bacteroidales 16S rRNA gene clone libraries were constructed from samples of effluent (sewage, bovine manure and pig slurry) and faeces (human, bovine, pig and wild bird), using PCR primers targeting order Bacteroidales. The phylogenetic analysis revealed six main distinct human‐, bovine‐, pig‐ and wild bird‐specific clusters. From the bovine‐specific cluster II, we designed a ruminant‐specific marker, Rum‐2‐Bac, and this showed 97% sensitivity (n = 30) and 100% specificity (n = 40) when tested by TaqMan^® real‐time PCR. Average concentrations of this marker in bovine and sheep faeces and in bovine manure were 8·2 ± 0·5, 8·4 ± 1·3 and 7 ± 0·5 log₁₀ copies per gram, respectively. It was also quantified in samples of runoff water impacted by bovine manure, with average concentrations of 5·1 ± 0·3 log₁₀ copies per millilitre water. Conclusions: Our results confirmed that some members of Bacteroidales isolated from effluents and faeces had host‐specific distributions. Identification of a bovine‐specific cluster made it possible to design a reliable ruminant‐specific marker. Significance and Impact of the Study: The host‐specific distribution of Bacteroidales sequences from effluents mirrored the host‐specific distribution of sequences observed in individual faeces. This efficient new ruminant‐specific Bacteroidales 16S rRNA marker represents a useful addition to the microbial source tracking toolbox.     

Evaluation of Colilert-18 for the detection of coliforms and Escherichia coli in tropical fresh water

AIMS: To evaluate the suitability of Colilert-18 in detecting Escherichia coli and total coliforms in tropical freshwater samples. METHODS AND RESULTS: Target organisms were isolated from yellow-fluorescent and yellow wells of Colilert-18/Quanti-Tray using m-TEC agar and m-ENDO LES agar respectively. All the selected isolates were first identified based on their fatty acid methyl ester profile. Isolates showing contradictory results to that of the Colilert-18 procedure were re-identified using API 20E strips. A total of 357 isolates, 177 from yellow-fluorescent wells and 180 from yellow wells, were identified. CONCLUSIONS: The false-positive and -negative rates for E. coli detection using Colilert-18 were 36.4% and 11%, respectively, while for coliform detection the false-positive rate was 10.3%. SIGNIFICANCE AND IMPACT OF THE STUDY: The high false-positive rate of Colilert-18, tempers its value for E. coli detection when used for tropical freshwater samples.