Sources and survival of Listeria monocytogenes on fresh, leafy produce.

Listeria monocytogenes is an intracellular human pathogen which enters the body through contaminated food stuffs and is known to contaminate fresh leafy produce such as spinach, lettuce and rocket. Routinely, fresh leafy produce is grown and processed on a large scale before reaching the consumer through various products such as sandwiches and prepared salads. From farm to fork, the fresh leafy produce supply chain (FLPSC) is complex and contains a diverse range of environments where L. monocytogenes is sporadically detected during routine sampling of produce and processing areas. This review describes sources of the bacteria in the FLPSC and outlines the physiological and molecular mechanisms behind its survival in the different environments associated with growing and processing fresh produce. Finally, current methods of source tracking the bacteria in the context of the food supply chain are discussed with emphasis on how these methods can provide additional, valuable information on the risk that L. monocytogenes isolates pose to the consumer.

Fate of Escherichia coli O145 present naturally in bovine slurry applied to vegetables before harvest, after washing and simulated wholesale and retail distribution.

AIMS: To determine the fate of Escherichia coli on vegetables that were processed through commercial wash treatments and stored under simulated retail conditions at 4°C or wholesale at fluctuating ambient temperatures (0-25°C, dependent on season). METHODS AND RESULTS: Bovine slurry that was naturally contaminated with E. coli O145 was applied without dilution or diluted 1:10 using borehole water to growing potatoes, leeks or carrots. Manure was applied 1 week prior to harvest to simulate a near-harvest contamination event by manure deposition or an application of contaminated water to simulate a flooding event or irrigation from a contaminated water source. At harvest, crops were contaminated at up to 2 log cfu g-1 . Washing transferred E. coli into the water of a flotation tank used for potato washing and did not completely remove all traces of contamination from the crop. Manure-contaminated potatoes were observed to contain 0·72 cfu E. coli O145 g-1 after processing and retail storage. Manure-contaminated leeks harboured 0·73-1·55 cfu E. coli O145 g-1 after washing and storage. There was no cross-contamination when leeks were spray washed. Washing in an abrasive drum resulted in less than perfect decontamination for manure-contaminated carrots. There were five post-distribution isolations from carrots irrigated with contaminated water 24 h prior to harvest. CONCLUSIONS: Standard commercial washing and distribution conditions may be insufficient to reliably control human pathogenic E. coli on fresh produce. SIGNIFICANCE AND IMPACT: Previous speculation that the cause of a UK foodborne disease outbreak was soil from imperfectly cleaned vegetables is plausible.

Ineffective hand washing and the contamination of carrots after using a field latrine.

UNLABELLED: A study was undertaken to simulate the likely effects of a field worker with poor hygienic practices that had returned to work too soon after recovering from an infection by an enteric pathogen. The studies simulated a variety of hand-washing practices from no washing to washing with soap and water followed by an application of alcohol gel after using a field latrine. The numbers of generic Escherichia coli isolated from workers' hands declined with increasing thoroughness of hand-washing treatments with unwashed hands > water > water and soap > water, soap and alcohol gel. Where gloves were worn the counts obtained for the treatments were significantly reduced, but it was observed that unwashed hands contaminated gloves during the process of putting them on. Hand contamination following the use of a field latrine transferred contamination to carrots. These results suggest that if no gloves are worn it would be best practice to wash hands with water and soap and apply alcohol gel after using a field latrine. Wearing gloves reduced the risk of contaminating handled produce but workers should still wash their hands after using a field latrine before applying gloves. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that inadequate hand hygiene in the field following the use of a field latrine can transfer bacterial contamination to hand-harvested carrots. Where fresh produce crops are to be handled by workers, wearing gloves reduces the risk of contaminating produce but workers should still wash their hands after using a field latrine before applying gloves. If no gloves are worn it would be best practice to wash hands with water and soap and apply alcohol gel after using a field latrine.

Spot urinary protein analysis for excluding significant proteinuria in pregnancy.

The aim of this research was to compare the accuracy of urinary protein/creatinine ratio (PCR) and albumin/creatinine ratio (ACR) in defining optimal cut-off points to rule-out significant proteinuria (>300 mg/24 h) in pregnancy. The secondary outcome measure was to determine the investigation of choice to evaluate proteinuria used by maternity units in the UK. PCR and ACR were calculated on first (PCR1, ACR1) void urine samples of the 24-hour urinary protein collection (24UP). Sensitivity and specificity was calculated for different cut-off points for PCR1 and ACR1 to rule-out significant proteinuria. An online survey was sent to RCOG members questioning them on their investigation of choice to evaluate proteinuria. We concluded from our results that both PCR and ACR are good rule-out tests for significant proteinuria in pregnancy using cut-off points of <20 mg/mmol and <2.5 mg/mmol. PCR is the investigation of choice in 56% of UK units studied.

Distribution and decline of human pathogenic bacteria in soil after application in irrigation water and the potential for soil-splash-mediated dispersal onto fresh produce.

AIMS: To improve our understanding of the survival and splash-mediated transfer of zoonotic agents and faecal indicator bacteria introduced into soils used for crop cultivation via contaminated irrigation waters. METHODS AND RESULTS: Zoonotic agents and an Escherichia coli marker bacterium were inoculated into borehole water, which was applied to two different soil types in early-, mid- and late summer. Decline of the zoonotic agents was influenced by soil type. Marker bacteria applied to columns of two soil types in irrigation water did not concentrate at the surface of the soils. Decline of zoonotic agents at the surface was influenced by soil type and environmental conditions. Typically, declines were rapid and bacteria were not detectable after 5 weeks. Selective agar strips were used to determine that the impact of water drops 24-87 µl could splash marker bacteria from soil surfaces horizontal distances of at least 25 cm and heights of 20 cm. CONCLUSIONS: Soil splash created by rain-sized water droplets can transfer enteric bacteria from soil to ready-to-eat crops. Persistence of zoonotic agents was reduced at the hottest part of the growing season when irrigation is most likely. SIGNIFICANCE AND IMPACT OF THE STUDY: Soil splash can cause crop contamination. We report the penetration depths and seasonally influenced declines of bacteria applied in irrigation water into two soil types.

The air-borne distribution of zoonotic agents from livestock waste spreading and microbiological risk to fresh produce from contaminated irrigation sources.

AIMS: To assess the risks of zoonotic agents in dissemination of livestock wastes into the environment by airborne distribution. To subsequently assess the survival time of zoonotic agents, introduced in irrigation water, on the phylloplane of produce. METHODS AND RESULTS: An Escherichia coli marker was introduced into pig slurry which was spread using a rain gun sprayer. Air sampling was undertaken to determine the distance reached by the marker. No recoveries were observed at a distance of 250 m. Borehole water, contaminated with zoonotic agents, was used to irrigate field plots sown with lettuce and spinach. Decline in bacterial numbers on the phylloplane was observed with time. After initial rapid decreases, we were unable to detect any pathogen from the phylloplane, 1 month after contamination. CONCLUSIONS: These preliminary results suggest that the risks to public health from the aerosolized spread of bacteria during slurry spreading by rain gun are low. Although, zoonotic agents on crop phylloplanes perish quickly, the risks of overhead irrigation of fresh produce 3 weeks before harvest should still be considered. SIGNIFICANCE AND IMPACT OF THE STUDY: These preliminary results improve our understanding on the fate of zoonotic agents in the environment. Spreading liquid livestock wastes by an airborne mechanism may not pose a significant public health risk. Detection of zoonotic agents 3 weeks after contamination of lettuce and spinach means that consideration should be given by the farmers until the time of harvest, when irrigating fresh produce with water that may have been directly or indirectly contaminated by livestock wastes.

Risk Assessment or Assessment of Risk? Developing an Evidence-Based Approach for Primary Producers of Leafy Vegetables To Assess and Manage Microbial Risks.

Over the last 10 years, some high-profile foodborne illness outbreaks have been linked to the consumption of leafy greens. Growers are required to complete microbiological risk assessments (RAs) for the production of leafy crops supplied either to retail or for further processing. These RAs are based primarily on qualitative judgements of hazard and risks at various stages in the production process but lack many of the steps defined for quantitative microbiological RAs by the Codex Alimentarius Commission. This article is based on the discussions of an industry expert group and proposes a grower RA approach based on a structured qualitative assessment, which requires all decisions to be based on evidence and a framework for describing the decision process that can be challenged and defended within the supply chain. In addition, this article highlights the need for evidence to be more easily available and accessible to primary producers and identifies the need to develop hygiene criteria to aid validation of proposed interventions.

The role of ifosfamide in cervical cancer.

A series of phase II studies using ifosfamide as a single agent and in combination with cisplatin and bleomycin (BIP) in advanced and recurrent cervical cancer were coordinated at the West Midlands Cancer Research Campaign Clinical Trials Unit, Birmingham, UK. The aim of these studies was to identify single agents and combination regimens that might be of value for palliation and have potential for neoadjuvant and adjuvant therapy in primary treatment. Ninety-eight patients were studied. Seventy-nine patients with disease not amenable to radical local therapy were treated with single-agent ifosfamide or the BIP combination. In 30 patients treated with single-agent ifosfamide, ten objective responses (33%) were seen, with one complete response. In 49 patients treated with BIP, 34 objective responses (69%) were seen, with ten complete responses (20%). Eleven (79%) of 14 patients with primary inoperable disease had at least a 50% reduction in tumor bulk before radical local radiotherapy. Toxicity resulted in alopecia, nausea and vomiting, myelosuppression, infection, reduction in renal function, and disturbance of consciousness. There was no evidence that neoadjuvant chemotherapy enhanced the acute toxic effects of pelvic radiotherapy. These data indicate that ifosfamide is highly active in cervical cancer and that in combination with bleomycin and cisplatin, it can be used for effective palliation and cytoreduction in around 70% of patients. Ifosfamide-containing regimens have potential for use as neoadjuvant and adjuvant therapy in patients at high risk of recurrence with conventional treatment. These hypotheses are currently being tested in prospective randomized trials.

Effects of carbidopa and of intravenous saline infusion into normal and hypertensive subjects on urinary free and conjugated dopamine.

OBJECTIVE: To investigate the possible role played by endogenous dopamine as a modulator of renal sodium (Na+) reabsorption after a combined Na+ and volume load. DESIGN: A randomized placebo-controlled study. METHODS: Ten healthy volunteers and four hypertensive patients were subjected to intravenous infusions of 21 0.9% saline (308 mmol Na+) administered from 1000 to 1300 h after oral administration of placebo or of carbidopa, a dopamine decarboxylase inhibitor. RESULTS: Studies on control subjects after placebo showed that natriuresis occurred during the 6 h after commencement of the saline infusion, with falls in plasma albumin concentration, plasma renin activity and plasma aldosterone concentration; in comparison with results of mock infusion (6 mmol Na+) there was no change in the urinary excretion of dopamine and noradrenaline (In their free or conjugated forms). There was, however, a marked surge in excretion of urinary conjugated dopamine and in the dopamine: noradrenaline ratio from 1300 to 1600 h, after either type of infusion. Administration of carbidopa before the saline infusion resulted in a marked decrease in excretion of urinary free dopamine, but had no effect on the surge in excretion of urinary conjugated dopamine. Saline infusion decreased proximal fractional Na+ reabsorption. Administration of carbidopa delayed but did not prevent this decrease. The effects of saline infusion and of carbidopa on the urinary excretion of dopamine and noradrenaline from hypertensive patients were similar to those observed with the healthy volunteers. CONCLUSIONS: These findings indicate that volume expansion by intravenous saline infusion has no appreciable effect on the urinary free dopamine excretion from normal or hypertensive humans; with any apparent increase, it is important to exclude the possibility of conversion of conjugates to free dopamine in vitro. Furthermore, that carbidopa administration did not inhibit the afternoon surge of conjugated dopamine suggests that administration of carbidopa is deficient as a tool to investigate the functional role of the renal dopamine system.

Efficacy of an angiotensin II receptor antagonist in managing hyperaldosteronism.

OBJECTIVE: To determine whether an angiotensin II receptor antagonist decreases blood pressure in patients with hyperaldosteronism and hypertension who are taking other antihypertensive agents. DESIGN: A double-blind randomized placebo-controlled crossover study. PATIENTS AND METHODS: Blood pressure and hormonal responses to 2-week courses of placebo/irbesartan (150 mg/day given by mouth at 08.05 h) were assessed in 10 patients with hyperaldosteronism. Clinic blood pressure was measured by sphygmomanometer, and plasma concentrations of aldosterone, cortisol, angiotensin II, electrolytes and renin activity (PRA) were determined weekly. Automated 24 h ambulatory blood pressure recordings were made at the end of the active and placebo phases. RESULTS: Irbesartan caused a post-dose decrease in ambulatory blood pressure (systolic, P = 0.02; diastolic, P = 0.05) in the period from 10.00 h to 20.00 h. Clinic blood pressure, measured at trough, was not significantly decreased. Plasma aldosterone decreased (P < 0.03) and PRA increased (P < 0.04) in the first week of active treatment with irbesartan, but differences between the placebo and active-treatment groups were not significant in the second week. There were no significant changes in plasma concentrations of angiotensin II, cortisol or potassium in either week. In the second week of irbesartan treatment, there were associations between change in plasma aldosterone and maximal change in ambulatory blood pressure (systolic and diastolic). CONCLUSION: Irbesartan has a role in combination antihypertensive treatment of patients with hyperaldosteronism.

Erythrocyte cation fluxes in the normotensive and hypertensive clients of a health screening clinic.

Erythrocyte cation transport was measured in vitro using 22Na+ and 86Rb+ uptake techniques in Caucasian men with newly-detected hypertension and in male control groups. The Na+, K+ cotransport [determined by ouabain-resistant frusemide-sensitive (ORFS) components of Na+ or Rb+ influx], sodium pump activity (determined by ouabain-sensitive Rb+ influx) and erythrocyte Na+ and K+ concentrations were not significantly altered in hypertensive men. The total Na+ influx in hypertensives (n = 59) was significantly greater (P less than 0.001) than in controls. The difference was mainly attributable to an increase in the ouabain-resistant frusemide-resistant component of this flux. The total Rb+ influx in hypertensives (n = 39) was also greater (P less than 0.005) than in controls. Overall, both total Na+ influx and total Rb+ influx were positively correlated (P less than 0.01) with diastolic blood pressure and with habitual dietary intake of alcohol. Multivariate analyses after controlling for the effect of blood pressure showed that mean corpuscular volume (MCV) and alcohol intake were statistically significant predictor variables for total Rb+ influx, although not for total Na+ influx. The results are compatible with increased diffusion of cations across the erythrocyte membrane in hypertension, but raise the question of a possible role of alcohol intake in mediating this effect.

Effects of dietary sodium deprivation on erythrocyte sodium concentration and cation transport in normotensive and untreated hypertensive subjects.

There is controversy about the effects of dietary sodium deprivation on cellular cation transport. Using washed erythrocytes for in vitro 22Na and 86Rb uptake studies, we studied the effects of a strict low-salt diet (20 mmol/day) for 4 days in 14 normotensive and 13 hypertensive subjects. Urinary sodium excretion fell from 147 +/- 13 to 18 +/- 3 mmol/24 h in the normotensive group and from 155 +/- 16 to 20 +/- 2 mmol/24 h in the hypertensive group. In both groups, there was a fall in plasma sodium concentration and activation of the renin-aldosterone axis. Both systolic and diastolic blood pressures fell in the hypertensive, but not the normotensive group. There were small but significant (P less than 0.025) decreases in cell cation concentrations and passive cation transport in the normotensive, but not the hypertensive group. No significant change in sodium pump activity or in Na+K+ cotransport was seen in either group. These observations provide no support for the concept that a decrease in dietary sodium intake can induce changes in cell cation transport, detectable in vitro, to which reduction in blood pressure may be attributed.

An examination of the renal transplant evaluation process focusing on cost and the reasons for patient exclusion.

A medical evaluation of prospective renal transplant recipients is performed to identify conditions that may exclude patients from transplantation because of unacceptable risks. Protocols for evaluating potential transplant candidates are available, but there is little information about reasons for excluding patients from transplantation. To assess the effectiveness and cost of our renal transplant-recipient evaluation process, we retrospectively reviewed patients excluded from renal transplantation between January 1993 and December 1995 to categorize the reasons for exclusion. We also examined the costs of the evaluation. The study group included all adults referred for kidney-only transplantation during the study period who were excluded from transplantation (n=125). Demographics of the 160 patients with end-stage renal disease (ESRD) who underwent renal transplantation during the study period were also examined. Compared with the patients who underwent transplantation, the excluded patients were older (48+/-14 v 43+/-12 years; P=0.006) and more likely to be women (66 of 125 patients; 53% v 57 of 160 patients; 36%; P=0.005) and diabetic (59 of 125 patients; 47% v 30 of 160 patients; 19%; P=0.005). The most common reason for excluding patients was medical contraindication (46%), followed by patient declined (25%), obesity (10%, defined as a body mass index [BMI] > or = 35), patient death (6%), and insurance/financial (5%). The medical reasons for exclusion were heart disease (38%), noncompliance (28%), miscellaneous (22%), and cancer (12%). Tests performed after the initial evaluation included cardiac testing (stress thallium or echocardiography and coronary angiography) in 50 patients, Doppler studies of the lower extremities in 28 patients, and hepatitis C polymerase chain reaction (PCR) or recombinant immunoblot assay (RIBA) assays in 8 patients. The cost of standard pretransplantation blood work for selected tests (ABO blood group typing, HLA, hepatitis B and C, and cytomegalovirus) was $709. Deferring such routine pretransplantation blood work until after the patient education session and history and physical examinations by nephrology and surgery in the 31 patients (25%) who declined transplantation at the initial visit would have resulted in considerable savings. Our evaluation process now includes prereferral information on a prospective recipient's medical problems, height and weight, and basic screening laboratory tests. This protocol has resulted in a more efficient and cost-effective evaluation process. Further examination of the cost-effectiveness of the transplant evaluation process is warranted.

Cytokine-mediated nitric oxide release--a common cytotoxic pathway in host-versus-graft and graft-versus-host reactions?

Serial cytokine and nitrate (as a measure of nitric oxide production) levels were assayed in nine consecutive patients undergoing allogeneic haemopoietic stem cell transplants. They were compared to those in 13 patients undergoing autologous transplants (transplant controls), 15 neutropenic patients with infective complications (patient controls) and 27 blood donors (normal controls). Peak nitrate, interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNF-alpha) levels were significantly higher in four allogeneic transplant patients with major non-infective complications compared to those without such complications, and control groups. Cytokine and nitrate levels peaked during conditioning therapy in the patients with veno-occlusive disease (one patient) and fulminant cholestatic liver failure (one patient), indicating that tissue damage may have been initiated during chemoradiotherapy in these patients, whereas peak levels occurred 2-3 days before graft rejection (one patient) and severe graft-versus-host disease (one patient), indicating a role for cytokine-induced nitric oxide release in the pathophysiology of these immune-mediated complications. Based on the data presented, it can be tentatively postulated that nitric oxide is a common proximate regulator of the immune response in host-versus-graft and graft-versus-host reactions.

The hormonal receptor status of uterine carcinosarcomas (mixed müllerian tumours): an immunohistochemical study.

AIM: To investigate the role of oestrogen and progesterone receptor status in uterine carcinosarcomas (mixed Müllerian tumours) to see whether the receptors were identifiable, and if so whether they were of significance clinically. METHODS: 11 cases of uterine carcinosarcoma were identified from clinical and pathology records. An immunohistochemical method was used to demonstrate oestrogen and progesterone hormone receptors on paraffin embedded material, with suitable tissue controls, staining being recorded. RESULTS: 10 of 11 cases showed staining for one or both hormone receptors in normal tissue adjacent to tumour. In four carcinosarcoma cases, staining for one or both receptors was shown within the epithelial component (appearing to correlate with the degree of epithelial differentiation); two of these cases had staining within sarcomatous areas. Two of the three patients still alive had epithelial hormone receptor positivity. CONCLUSIONS: Receptors for oestrogen and progesterone were found in four of 11 cases of uterine carcinosarcoma, using paraffin embedded material. There may be an association between hormone receptor positivity and clinical outcome.

Urine cyclic nucleotide concentrations in cancer and other conditions; cyclic GMP: a potential marker for cancer treatment.

Cyclic guanosine 3',5' monophosphate (cyclic GMP) and cyclic adenosine 3',5' monophosphate (cyclic AMP) have been determined in random urine specimens from 95 healthy individuals, 60 patients with non-cancerous conditions, 52 patients with benign tumours, and 74 patients with malignant tumours. Concentrations of cyclic GMP have also been determined in a number of other groups, including some undergoing cancer treatment. Ninety-three per cent of cancer patients had raised urinary cyclic GMP concentrations compared to the reference range for healthy subjects. For the non-cancerous and benign groups, 33% and 42% respectively had raised concentrations. The urine cyclic AMP concentrations were similar in all groups. Urine cyclic GMP appeared to rise early in the onset of malignant growth. Successful cancer treatment was accompanied by a dramatic fall in the urine cyclic GMP concentrations, whereas if the treatment was unsuccessful the level did not change. It is concluded that urine cyclic GMP may have important applications in the monitoring of cancer treatment.

A critical evaluation of three methods of studying cell proliferation in human cervical epithelium.

Three experiments describing applications of cell kinetic techniques to the human cervix are presented. The mitotic index is shown not to be associated with histologic differentiation, node metastasis, or survival rates in cervical squamous carcinoma, although inherent sources of inaccuracy may be a contributory factor. The second experiment, a new application of the metaphase arrest technique to cervical epithelium, allows an estimation of cell production rates in normal, dysplastic, wart-affected cervical epithelium, and in invasive carcinoma. Confidence limits are wide, rendering individual values imprecise in isolation. The third project describes a modification of in vitro labeling of cervical biopsy specimens using tritiated thymidine followed by autoradiography. Labeling indexes are produced for normal and pathological cervical epithelium. Although successful labeling was not achieved in every case, the labeling index for cervical intraepithelial neoplasia (CIN stage III) is significantly greater than for normal epithelium. The metaphase arrest technique appears to be the most applicable of the three to the study of the human cervix, yielding a useful parameter of cell proliferative activity. The labeling index gives an indication of activity that is useful for comparative studies, but that does not provide data regarding actual proliferative rates. More sophisticated experiments using radioactive materials are ethically inadmissable.

Adenocarcinoma of the uterine cervix: a study of 73 cases.

Seventy-three patients with adenocarcinoma of the cervix were seen between 1969 and 1983. This represented 8.1% of all carcinoma of cervix seen during that period. Survival rates in stage Ib were significantly worse for those with poorly differentiated lesions and for those with involved pelvic lymph nodes. Age at presentation appeared to decline over the period of the study. When the patients with stage Ib lesions were compared with a group of stage Ib squamous carcinomas treated in the same unit, there was no difference in age at presentation, node metastasis rates, or survival. Four patients had cervical intraepithelial neoplasia (CIN) in addition to adenocarcinoma.

Ultrasound placental grading and amniotic fluid quantitation in prolonged pregnancy.

Serial ultrasound examinations were performed every three to five days to monitor 200 singleton pregnancies with gestations of 42 weeks or longer. Scanning was used to measure the deepest amniotic fluid pool and to grade placental echogenic changes. Oligohydramnios, defined as an amniotic fluid pool less than 30 mm, was detected in 33 patients (17%) and was an indication for intervention. Grade III placenta was found in 53 patients (27%), and no grade could be assigned in 13 (7%). Intrauterine growth retardation and meconium-stained or absent amniotic fluid at amniotomy occurred significantly more often in association with oligohydramnios than with grade III placenta, which was found before spontaneous labor in only 24% of cases. Ultrasound placental grading appears to offer little useful additional information to amniotic fluid quantitation in satisfactory monitoring of prolonged pregnancies.

Loop diathermy cone biopsy.

A technique for cone biopsy using loop diathermy is presented. The diathermy loop consists of an insulated shaft attached to an insulated transverse or U-shaped arm, to which the loop wire is attached. The diathermy power is supplied by an electrosurgical unit. Among the first 50 cases, the mean age was 41.6 years; 72% of the cases were reported as cervical intraepithelial neoplasia III and 6% as microinvasive carcinoma. The mean length of the cone was 20.2 mm and the mean weight 3.9 g. Only 10% of the cones were reported as incompletely excised. No major complications were observed, and minor complications were reported in four patients (8%). We found this technique to be quick, simple, and economical. It provides a good histologic specimen with a low incidence of short-term morbidity.