Arachidonic acid induction of Rho-mediated transendothelial migration in prostate cancer.

BACKGROUND: Bone metastases in prostate cancer (CaP) result in CaP-related morbidity/mortality. The omega-6 polyunsaturated fatty acid (PUFA) arachidonic acid (AA) and lipophilic statins affect metastasis-like behaviour in CaP cells, regulating the critical metastatic step of CaP migration to the bone marrow stroma. METHODS: Microscopic analysis and measurement of adhesion and invasion of CaP cells through bone marrow endothelial cells (BMEC) was undertaken with AA stimulation and/or simvastatin (SIM) treatment. Amoeboid characteristics of PC-3, PC3-GFP and DU-145 were analysed by western blotting and Rho assays. RESULTS: The CaP cell lines PC-3, PC3-GFP and DU-145 share the ability to migrate across a BMEC layer. Specific amoeboid inhibition decreased transendothelial migration (TEM). AA stimulates amoeboid characteristics, driven by Rho signalling. Selective knock-down of components of the Rho pathway (RhoA, RhoC, Rho-associated protein kinase 1 (ROCK1) and ROCK2) showed that Rho signalling is crucial to TEM. Functions of these components were analysed, regarding adhesion to BMEC, migration in 2D and the induction of the amoeboid phenotype by AA. TEM was reduced by SIM treatment of PC3-GFP and DU-145, which inhibited Rho pathway signalling. CONCLUSIONS: AA-induced TEM is mediated by the induction of a Rho-driven amoeboid phenotype. Inhibition of this cell migratory process may be an important therapeutic target in high-risk CaP.

Prevalence of Group A beta-haemolytic Streptococcus isolated from children with acute pharyngotonsillitis in Aden, Yemen.

OBJECTIVES: To estimate the prevalence of Group A beta-haemolytic streptococcus (GAS) and non-GAS infections among children with acute pharyngotonsillitis in Aden, Yemen, to evaluate the value of a rapid diagnostic test and the McIsaac score for patient management in this setting and to determine the occurrence of emm genotypes among a subset of GAS isolated from children with acute pharyngotonsillitis and a history of acute rheumatic fever (ARF) or rheumatic heart disease (RHD). METHODS: Group A beta-haemolytic streptococcus infections in school-aged children with acute pharyngotonsillitis in Aden, Yemen, were diagnosed by a rapid GAS antigen detection test (RADT) and/or GAS culture from a throat swab. The RADT value and the McIsaac screening score for patient management were evaluated. The emm genotype of a subset of GAS isolates was determined. RESULTS: Group A beta-haemolytic streptococcus pharyngotonsillitis was diagnosed in 287/691 (41.5%; 95% CI 37.8-45.3) children. Group B, Group C and Group G beta-haemolytic streptococci were isolated from 4.3% children. The RADT had a sensitivity of 238/258 (92.2%) and specificity of 404/423 (95.5%) against GAS culture. A McIsaac score of ≥4 had a sensitivity of 93% and a specificity of 82% for confirmed GAS infection. The emm genotypes in 21 GAS isolates from children with pharyngitis and a history of ARF and confirmed RHD were emm87 (11), emm12 (6), emm28 (3) and emm5 (1). CONCLUSION: This study demonstrates a very high prevalence of GAS infections in Yemeni children and the value of the RADT and the McIsaac score in this setting. More extensive emm genotyping is necessary to understand the local epidemiology of circulating strains.

Confusing medical terms: disease that may or may not exist.

Patients often quote diseases or illnesses that either do not exist per se or are hard to prove that they exist. Often symptoms are vague and, therefore, difficult for patients to qualify in a language clinicians can understand, interpret and act upon. Physicians often perpetuate this by giving 'diagnoses of exclusion', or using poor explanations, oversimplifications, conflicting diagnostic criteria or vague historical terms that have now evolved into something else. However, the history taker must be able to interpret the subtle language barrier that exists between doctor and patient. In this short review of the literature, some commonly quoted conditions are examined more closely to try and understand further the terminology used by both patients and clinicians alike.

Ligand-independent activation of EphA2 by arachidonic acid induces metastasis-like behaviour in prostate cancer cells.

BACKGROUND: High intake of omega-6 polyunsaturated fatty acids (PUFA) has been associated with clinical progression in prostate cancer (CaP). This study investigates the signalling mechanism by which the omega-6 PUFA arachidonic acid (AA) induces prostatic cellular migration to bone marrow stroma. METHODS: Western blot analysis of the PC-3, PC3-GFP, DU 145 and LNCaP cells or their lipid raft (LR) components post AA stimulation was conducted in association with assays for adhesion and invasion through the bone marrow endothelial monolayers. RESULTS: Arachidonic acid increased transendothelial migration of PC3-GFP cells (adhesion 37%±0.08, P=0.0124; transmigration 270%±0.145, P=0.0008). Akt, Src and focal adhesion kinase (FAK) pathways were induced by AA and integrally involved in transendothelial migration. LR were critical in AA uptake and induced Akt activity. Ephrin receptor A2 (EphA2), localised in LR, is expressed in DU 145 and PC-3 cells. Arachidonic acid induced a rapid increase of EphA2 Akt-dependent/ligand-independent activation, while knockdown of the EphrinA1 ligand decreased AA induced transendothelial migration, with an associated decrease in Src and FAK activity. Arachidonic acid activated Akt in EphA2(-) LNCaP cells but failed to induce BMEC transendothelial invasion. CONCLUSION: Arachidonic acid induced stimulation of EphA2 in vitro is associated fundamentally with CaP epithelial migration across the endothelial barrier.

Multilocus sequence typing of human and canine C. upsaliensis isolates.

Risk of Campylobacter infection in humans has been associated with many sources, including dogs. C. upsaliensis is the most common species found in canines, and has been occasionally isolated from symptomatic humans. This study aimed to investigate the genetic diversity of 41 C. upsaliensis isolates carried by dogs and from nine isolates carried by humans using Multilocus sequence typing (MLST). We identified considerable genetic diversity amongst the C. upsaliensis isolates from both dogs and humans, identifying 45 different sequence types (STs). All STs were new, apart from that of the reference strain. Only three STs were found in more than one isolate: ST-72 (2 isolates), ST-98 (2 isolates) and ST-104 (3 isolates). ST-104 was the only ST to be encountered in both dogs and humans. Thirty-one of the 45 STs were assigned to one of 13 clonal complexes (CCs). Four of these CCs contained STs originating from both humans and dogs. None of the CCs contained exclusively human isolates, and two isolates from dogs within the same kennel belonged to the same CC. The large amount of diversity found in both dog and human isolates of C. upsaliensis, combined with the relatively small database, made it difficult to assign strains to sources of infection. This emphasizes the need to increase the size of the database. Dog and human isolates occasionally grouped together, however there were insufficient human-derived isolates to determine whether or not dogs are a common source of infection. Although C. upsaliensis infection is rare in humans, dogs still remain a potential source, and are therefore a possible zoonotic risk. Further work is needed to investigate the epidemiology of C. upsaliensis infection in humans.

Diagnostic efficacy of activated partial thromboplastin time waveform and procalcitonin analysis in pediatric meningococcal sepsis.

OBJECTIVE: A biphasic activated partial thromboplastin time waveform predicts sepsis and disseminated intravascular coagulation in adults. This has not been previously investigated in children. Our aim is to ascertain whether there are changes in the activated partial thromboplastin time waveform in children with meningococcal disease and to compare its diagnostic use with procalcitonin. SETTING: Alder Hey Children's National Health Service Foundation Trust, Liverpool, UK. PATIENTS: Thirty-six children admitted to the hospital for the treatment of suspected meningococcal disease had activated partial thromboplastin time waveform and procalcitonin analysis performed at admission. The light transmittance level at 18 secs was used to quantitate the waveform. Severity of disease was assessed using the Glasgow Meningococcal Septicaemia Prognostic Score, Pediatric Risk of Mortality III score, and the Pediatric Logistic Organ Dysfunction score. MEASUREMENTS AND MAIN RESULTS: Twenty-four children had proven meningococcal disease, 12 had a presumed viral illness, and 20 control subjects were recruited. Transmittance level at 18 secs was lower in children with meningococcal disease and those with a viral illness (p < .0001) and control subjects (p < .0005). Sensitivity and specificity was 0.91 and 0.96 for transmittance level at 18 secs and 0.92 and 1 for procalcitonin in identifying meningococcal disease. There was a significant difference in procalcitonin between children with meningococcal disease and those with a viral illness and control subjects (p < .0005). A negative correlation was found between transmittance level at 18 secs and length of hospital stay (p < .0001), C-reactive protein (p < .0001), procalcitonin (p < .0001), Glasgow Meningococcal Septicaemia Prognostic Score (p < .01), Pediatric Risk of Mortality III score (p < .0001), and Pediatric Logistic Organ Dysfunction score score (p < .0001). CONCLUSION: The activated partial thromboplastin time waveform is abnormal in children with meningococcal disease and may be a useful adjunct in the diagnosis and management of sepsis in children.

Prevalence of rheumatic heart disease among school-children in Aden, Yemen.

BACKGROUND: Rheumatic heart disease (RHD) is an important contributor to cardiovascular disease in children and adults in Yemen. This is the first report to determine the prevalence of RHD among school-children in the city of Aden. METHODS: A cross-sectional case-finding survey of RHD was conducted in 6000 school-children aged 5-16 years. Echocardiography was undertaken in those with clinical signs of organic heart disease. RESULTS: The prevalence of RHD was 36·5/1000 school-children, which is one of the highest reported among school echocardiography surveys in the world. RHD was more common in 10-16-year-old students. RHD was diagnosed in more than one member of the families of 53 (24·2%) of the children. Mitral regurgitation (MR) was detected in 49·8%, 26·6% had MR with mitral valve prolapse and 17·8% had combined MR and aortic regurgitation. Fifty-eight children were diagnosed with congenital heart disease (CHD), representing a prevalence of 9·7/1000. The main types of CHD were mitral valve prolapse, patent ductus arteriosus, atrial septal defect, pulmonary stenosis and aortic stenosis. Congenital mitral valve prolapse found in 36 children was three times more common in males than females. Children with RHD were more likely to be from low-income families with poor housing and greater overcrowding (49·3%, 39·3% and 64·8%) than children with CHD (44·8%, 32·8% and 48·3%, respectively). CONCLUSIONS: The high prevalence of RHD is a major public health problem in Yemen. Urgent screening surveys and an RHD prophylactic programme of appropriate management of group A ß-haemolytic streptococcal pharyngotonsilitis are required.

The prevalence of antimicrobial-resistant Escherichia coli in sympatric wild rodents varies by season and host.

AIMS: To investigate the prevalence and temporal patterns of antimicrobial resistance in wild rodents with no apparent exposure to antimicrobials. METHODS AND RESULTS: Two sympatric populations of bank voles and wood mice were trapped and individually monitored over a 2- year period for faecal carriage of antimicrobial-resistant Escherichia coli. High prevalences of ampicillin-, chloramphenicol-, tetracycline- and trimethoprim-resistant E. coli were observed. A markedly higher prevalence of antimicrobial-resistant E. coli was found in wood mice than in bank voles, with the prevalence in both increasing over time. Superimposed on this trend was a seasonal cycle with a peak prevalence of resistant E. coli in mice in early- to mid-summer and in voles in late summer and early autumn. CONCLUSIONS: These sympatric rodent species had no obvious contact with antimicrobials, and the difference in resistance profiles between rodent species and seasons suggests that factors present in their environment are unlikely to be drivers of such resistance. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings suggest that rodents may represent a reservoir of antimicrobial-resistant bacteria, transmissible to livestock and man. Furthermore, such findings have implications for human and veterinary medicine regarding antimicrobial usage and subsequent selection of antimicrobial-resistant organisms.

Comparison of Arcobacter isolation methods, and diversity of Arcobacter spp. in Cheshire, United Kingdom.

The aims of this study were, firstly, to compare five published methods for the isolation of Arcobacter spp. from animal feces in order to determine the most sensitive and specific method. Second, we analyzed the resulting isolates by multilocus sequence typing (MLST) in order to investigate the diversity of the isolates recovered. Third, we investigated the ability to recover Arcobacter spp. from frozen fecal samples. Seventy-seven fecal samples from cattle, sheep, and badgers were subjected to five isolation methods, based on published methods for the isolation of Arcobacter and Campylobacter spp. Thirty-nine Arcobacter butzleri isolates were analyzed using a multilocus sequence typing scheme. The survival of Arcobacter spp. in frozen samples was investigated by freezing the fecal samples at -80°C for 7 days and then applying the same five isolation methods. The most sensitive and specific method used an Arcobacter-specific broth in conjunction with modified charcoal cefoperazone deoxycholate agar (mCCDA) with added antibiotics. Freezing of fecal samples led to a reduction in the recovery of Arcobacter spp. by approximately 50%. The 39 allelic profiles obtained by MLST could be divided into 11 sequence types (STs). We have identified the most sensitive and specific method for the isolation of Arcobacter spp. from animal feces and demonstrated that the freezing of fecal samples prior to isolation reduces arcobacter recovery. MLST analysis of the isolates revealed a high level of diversity.

Prevalence and shedding patterns of Campylobacter spp. in longitudinal studies of kennelled dogs.

Dog ownership is considered a risk factor for campylobacteriosis in humans. This study investigated the prevalence and shedding of Campylobacter spp. in kennelled dogs. Faecal samples (n=399) were collected in longitudinal studies from 52 dogs in two kennels. Campylobacter spp. were isolated using charcoal-based selective agars and direct PCR. The prevalence of Campylobacter spp. in dogs in boarding kennels ranged from 46% (95% CI 22, 72) on entry, to 50% (95% CI 30, 70) overall, and in dogs in 'rescue' kennels from 68% (95% CI 49, 84) on entry to 73% (95%, CI 56, 87) overall. C. upsaliensis was isolated from 62% (95% CI 48, 73) of the dogs, whilst C. jejuni was isolated from 15% (95% CI 7, 26) of animals. The majority of infected dogs entered the kennels already carrying Campylobacter spp., and remained infected throughout their stay. However, in some cases, shedding appeared to commence after kennelling. Given that the prevalence of C. upsaliensis and C. jejuni was relatively high in dogs from both boarding and rescue kennels, such animals may pose a zoonotic risk.

Molecular characterization of rotavirus strains circulating among children with acute gastroenteritis in Madagascar during 2004-2005.

A survey was undertaken of the etiology of acute gastroenteritis in children <16 years of age in Antananarivo, Madagascar, from May 2004 through May 2005. With use of electron microscopy of fecal specimens, 104 (36%) of 285 children were found to be infected with rotavirus. Rotavirus strain characterization was undertaken using enzyme-linked immunosorbent assay, electropherotyping, reverse-transcription polymerase chain reaction genotyping, and nucleotide sequencing. The predominant group A rotavirus strain types identified were P[4]G2 (62%) and P[8]G9 (23%). Nucleotide sequence analysis of the VP7 genes of selected Malagasy G2 and G9 strains demonstrated similarity with those of other recently identified African rotavirus strains belonging to the same genotype.

An investigation of the RWPE prostate derived family of cell lines using FTIR spectroscopy.

Interest in developing robust, quicker and easier diagnostic tests for cancer has lead to an increased use of Fourier transform infrared (FTIR) spectroscopy to meet that need. In this study we present the use of different experimental modes of infrared spectroscopy to investigate the RWPE human prostate epithelial cell line family which are derived from the same source but differ in their mode of transformation and their mode of invasive phenotype. Importantly, analysis of the infrared spectra obtained using different experimental modes of infrared spectroscopy produces similar results. The RWPE family of cell lines can be separated into groups based upon the method of cell transformation rather than the resulting invasiveness/aggressiveness of the cell line. The study also demonstrates the possibility of using a genetic algorithm as a possible standardised pre-processing step and raises the important question of the usefulness of cell lines to create a biochemical model of prostate cancer progression.

Spatio-temporal epidemiology of Campylobacter jejuni enteritis, in an area of Northwest England, 2000-2002.

A total of 969 isolates of Campylobacter jejuni originating in the Preston, Lancashire postcode district over a 3-year period were characterized using multi-locus sequence typing. Recently developed statistical methods and a genetic model were used to investigate temporal, spatial, spatio-temporal and genetic variation in human C. jejuni infections. The analysis of the data showed statistically significant seasonal variation, spatial clustering, small-scale spatio-temporal clustering and spatio-temporal interaction in the overall pattern of incidence, and spatial segregation in cases classified according to their most likely species-of-origin.

Detection of coronaviruses in children with acute gastroenteritis in Maddina, Saudi Arabia.

BACKGROUND: The role of coronaviruses in paediatric gastro-enteritis is not well defined. We investigated the detection rate and epidemiological features of infection with coronavirus in children receiving hospital care for acute gastro-enteritis in Maddina, Saudi Arabia. METHODS: Stool specimens were collected from children less than 5 years of age who were either hospitalised in Maddina or given oral rehydration therapy as outpatients between April 2004 and April 2005. Coronaviruses were detected by electron microscopy. RESULTS: Coronaviruses were detected in 63 (6%) of 984 children with acute gastro-enteritis and were more commonly detected in outpatients (47/423, 11%) than in inpatients (16/561, 3%). The median age (range) of children with coronavirus infection was 42 months (10-60). Coronaviruses were detected throughout the year with the highest detection rate at the end of the winter season. CONCLUSIONS: Coronaviruses were commonly identified in children with diarrhoea in Saudi Arabia. Their role in paediatric gastro-enteritis warrants further evaluation.

Prevalence of Campylobacter spp. in a cross-sectional study of dogs attending veterinary practices in the UK and risk indicators associated with shedding.

Campylobacteriosis is a major cause of gastroenteritis in humans and some studies have suggested that dog ownership is a risk factor for the condition. To determine the prevalence, species distribution, and risk indicators for Campylobacter spp. infecting dogs attending veterinary practices in UK, faecal samples were collected in a cross-sectional study from 249 dogs with and without clinical signs. The prevalence of Campylobacter spp. was 38% (95% CI 32, 44), with Campylobacter upsaliensis accounting for 94 (98%) of the isolates and Campylobacter jejuni for the remainder. Multivariable analysis indicated that younger dogs were more likely to carry C. upsaliensis and the high prevalence of this pathogen supports the hypothesis that dogs, particularly younger animals, may be an important source of C. upsaliensis infection for humans. However the prevalence of C. jejuni, the most common Campylobacter spp. associated with disease in humans, was low (1.2%, 95% CI 0.3, 3).

Risk factors for the carriage of Campylobacter upsaliensis by dogs in a community in Cheshire.

Samples of faeces were taken from 183 healthy pet dogs in a census-based, cross-sectional study in Cheshire; culture methods were used to detect any Campylobacter species and a direct PCR was used to detect Campylobacter upsaliensis. Forty-six of the dogs were positive for C upsaliensis by either culture or direct PCR, giving a prevalence of 25.1 per cent (95 per cent confidence interval [CI] 19.0 to 32.1 per cent). One sample was positive by culture for Campylobacter jejuni (95 per cent CI 0.0 to 3.0 per cent) and one for Campylobacter lari. Multivariable logistic regression identified risk factors for the carriage of C upsaliensis by a dog as: living with another dog that also carried C upsaliensis; being small rather than medium-sized; being less than three years old; living in a household that kept fish; being fed commercial dog treats; and being fed human food titbits, particularly in the dog's bowl.

Typing of Campylobacter jejuni isolates from dogs by use of multilocus sequence typing and pulsed-field gel electrophoresis.

Campylobacter is a major cause of human gastroenteritis worldwide. Risk of Campylobacter infection in humans has been associated with many sources, including dogs. This study aimed to investigate whether C. jejuni carried by dogs could potentially be a zoonotic risk for humans and if there were common sources of C. jejuni infection for both humans and dogs. Multilocus sequence typing (MLST) together with macrorestriction analysis of genomic DNA using SmaI and pulsed-field gel electrophoresis (PFGE) were both used to analyze 33 C. jejuni isolates obtained from various dog populations, including those visiting veterinary practices and from different types of kennels. MLST data suggested that there was a large amount of genetic diversity between dog isolates and that the majority of sequence types found in isolates from these dogs were the same as those found in isolates from humans. The main exception was ST-2772, which was isolated from four samples and could not be assigned to a clonal complex. The most commonly identified clonal complex was ST-45 (11 isolates), followed by ST-21 (4 isolates), ST-508 (4 isolates), and ST-403 (3 isolates). The profiles obtained by macrorestriction PFGE were largely in concordance with the MLST results, with a similar amount of genetic diversity found. The diversity of sequence types found within dogs suggests they are exposed to various sources of C. jejuni infection. The similarity of these sequence types to C. jejuni isolates from humans suggests there may be common sources of infection for both dogs and humans. Although only a small number of household dogs may carry C. jejuni, infected dogs should still be considered a potential zoonotic risk to humans, particularly if the dogs originate from kennelled or hunt kennel dog populations, where the prevalence may be higher.

Neutrophil TLR4 expression is reduced in the airways of infants with severe bronchiolitis.

BACKGROUND: In respiratory syncytial virus (RSV) bronchiolitis, neutrophils account for >80% of cells recovered from the airways in bronchoalveolar lavage (BAL) fluid. This study investigated neutrophil activation and Toll-like receptor (TLR) expression in the blood and lungs of infants with severe RSV bronchiolitis. METHODS: BAL fluid and (blood) samples were collected from 24 (16) preterm and 23 (15) term infants ventilated with RSV bronchiolitis, and 12 (8) control infants. Protein levels and mRNA expression of CD11b, myeloperoxidase (MPO) and TLRs 2, 4, 7, 8 and 9 were measured in neutrophils. RESULTS: Blood neutrophils had more CD11b in preterm and term infants with RSV bronchiolitis than control infants (p<0.025) but similar amounts of MPO. BAL fluid neutrophils from infants with RSV bronchiolitis had greater amounts of CD11b and MPO than blood neutrophils and BAL fluid neutrophils from controls (p<0.01). Blood neutrophils from term infants with RSV bronchiolitis had less total TLR4 protein than preterm infants with RSV bronchiolitis (p = 0.005), and both had less than controls (p<0.04). Total TLR4 for each group was greater in BAL fluid neutrophils than in blood neutrophils. Blood neutrophils from preterm infants with RSV bronchiolitis had greater TLR4 mRNA expression than term infants with RSV bronchiolitis (p = 0.005) who had similar expression to controls (p = 0.625). CONCLUSIONS: In infants with severe RSV bronchiolitis, neutrophil activation starts in the blood and progresses as they are recruited into the airways. Total neutrophil TLR4 remains low in both compartments. TLR4 mRNA expression is unimpaired. This suggests that neutrophil TLR4 expression is deficient in these infants, which may explain why they develop severe RSV bronchiolitis.

Micronutrient concentrations in respiratory syncytial virus and human metapneumovirus in Yemeni children.

BACKGROUND: Acute respiratory infections (ARI) cause significant childhood mortality. Nutritional homeostasis, particularly micronutrient levels, is important in modulating response to infection. More information is required regarding micronutrient levels in ARI viral infections, especially newly identified viruses such as human metapneumovirus (HMPV). AIM: To describe zinc, copper, selenium and vitamins A and E concentrations in children with respiratory syncytial virus (RSV) and/or HMPV in relation to levels of C-reactive protein (CRP). METHODS: The presence of RSV/HMPV in nasopharyngeal aspirates (NPA) was identified in 246 children using RTPCR. Zinc, copper, selenium and vitamins A and E concentrations were measured using inductive coupled plasma mass spectrometry and high performance liquid chromatography. RESULTS: 183 children had RSV, 39 had HMPV and 24 were co-infected. Zinc concentrations were lower in children with HMPV than in children with RSV or RSV/HMPV co-infection. Copper concentrations were lower in children with RSV than in children with RSV/HMPV or HMPV and zinc/copper ratios were lower in children with HMPV/RSV or RSV than in children with HMPV alone. Retinol and a alpha-tocopherol were lower in children with RSV than in children with HMPV. Most children had low selenium concentrations. Children with RSV and raised CRP (>5 mg/L) had higher copper and lower zinc/copper ratios than those with low CRP (< or =5 mg/L). Children with HMPV and raised CRP had higher copper and lower zinc concentrations than children with low CRP. Children with RSV/HMPV and raised CRP had higher copper concentrations. Children with RSV/HMPV and raised CRP had higher a alpha-tocopherol concentrations. CONCLUSION: The profiles of micronutrients differ in children with RSV and HMPV and are confounded by CRP. These results may guide strategies for micronutrient supplementation in ARI.

Quantifying within- and between-animal variation and uncertainty associated with counts of Escherichia coli O157 occurring in naturally infected cattle faeces.

Cattle faeces are considered the most important reservoir for human infection with Escherichia coli O157. We have previously described shedding of E. coli O157 in the faeces of naturally infected cattle cohorts. However, the data require further investigation to quantify the uncertainty and variability in the estimates previously presented. This paper proposes a method for analysing both the presence and the quantity of E. coli O157 in cattle faecal samples, using two isolation procedures, one of which enumerates E. coli O157. The combination of these two measurements, which are fundamentally different in nature and yet measuring a common outcome, has necessitated the development of a novel statistical model for ascertaining the contribution of the various components of variation (both natural and observation induced) and for judging the influence of explanatory variables. Most of the variation within the sampling hierarchy was attributable to multiple samples from the same animal. The contribution of laboratory-level variation was found to be low. After adjusting for fixed and random effects, short periods of increased intensity of shedding were identified in individual animals. We conclude that within-animal variation is greater than between animals over time, and studies aiming to elucidate the dynamics of shedding should focus resources, sampling more within than between animals. These findings have implications for the identification of persistent high shedders and for assessing their role in the epidemiology of E. coli O157 in cattle populations. The development of this non-standard statistical model may have many applications to other microbial count data.