The occurrence of Campylobacter jejuni in raw cows' milk

Faeces and raw milk from individual cows were examined for the presence of Campylobacter jejuni . After drawing milk, the lactoperoxidase system was inactivated by raising the pH to 7.5. The organism was isolated from 22% of 904 faecal samples and from 4.5% of 904 milk samples. From laboratory experiments it could be concluded that inactivation of the lactoperoxidase system resulted in a better isolation of C. jejuni from raw cows' milk.     

The occurrence of Campylobacter jejuni in raw cows' milk

Faeces and raw milk from individual cows were examined for the presence of Campylobacter jejuni. After drawing milk, the lactoperoxidase system was inactivated by raising the pH to 7.5. The organism was isolated from 22% of 904 faecal samples and from 4.5% of 904 milk samples. From laboratory experiments it could be concluded that inactivation of the lactoperoxidase system resulted in a better isolation of C. jejuni from raw cows' milk.     

Isolation of Campylobacter jejuni from raw milk

Campylobacter jejuni was isolated from raw milk by a method that can routinely detect less than or equal to 1 organism per ml. This procedure was used in a survey of 195 separate farms and showed a 1.5% incidence of C. jejuni in milk from bulk tanks.     

Isolation of Campylobacter jejuni from raw milk.

Campylobacter jejuni was isolated from raw milk by a method that can routinely detect less than or equal to 1 organism per ml. This procedure was used in a survey of 195 separate farms and showed a 1.5% incidence of C. jejuni in milk from bulk tanks.     

Communicable disease associated with milk and dairy products in England and Wales 1951-80.

In England and Wales between 1951 and 1980 233 reported outbreaks of communicable disease attributed to milk or dairy products affected nearly 10 000 people, of whom four died. Tuberculosis and brucellosis have been controlled, but milk-borne outbreaks of salmonellosis and campylobacter enteritis due to raw or defectively pasteurised milk are common and may be increasing in number. Universal heat treatment of milk is an effective preventive measure, and it is regrettable that the continued sale of untreated milk is to be permitted in England and Wales.     

Q fever through consumption of unpasteurised milk and milk products – a risk profile and exposure assessment

Q fever is a zoonotic disease caused by the bacterium Coxiella burnetii which is endemic in cattle, sheep and goats in much of the world, including the United Kingdom (UK). There is some epidemiological evidence that a small proportion of cases in the developed world may arise from consumption of unpasteurised milk with less evidence for milk products such as cheese. Long maturation at low pH may give some inactivation in hard cheese, and viable C. burnetii are rarely detected in unpasteurised cheese compared to unpasteurised milk. Simulations presented here predict that the probability of exposure per person to one or more C. burnetii through the daily cumulative consumption of raw milk in the UK is 0·4203. For those positive exposures, the average level of exposure predicted is high at 1266 guinea pig intraperitoneal infectious dose 50% units (GP_IP_ID₅₀) per person per day. However, in the absence of human dose–response data, the case is made that the GP_IP_ID₅₀ unit represents a very low risk through the oral route. The available evidence suggests that the risks from C. burnetii through consumption of unpasteurised milk and milk products (including cheese) are not negligible but they are lower in comparison to transmission via inhalation of aerosols from parturient products and livestock contact.     

Clinical aspects of Campylobacter jejuni infections in adults.

Campylobacter jejuni is an almost ubiquitous, microaerophilic, gram-negative rod. Outbreaks have been associated with drinking raw milk or contaminated water and eating poultry. Campylobacter jejuni accounts for 3.2% to 6.1% of cases of diarrheal illness in the general population of the United States, and infected patients frequently present with abdominal pain and fever. Less frequently, C jejuni is responsible for bacteremia, septic arthritis, septic abortion, and other extraintestinal infections. Reactive arthritis, Reiter''s syndrome, the Guillain-Barré syndrome, and pancreatitis may accompany or follow C jejuni enterocolitis. Campylobacter jejuni is an important cause of diarrheal illness and is a more commonly identified stool organism than Salmonella or Shigella species. Recurrent and chronic infection is generally reported in immunocompromised hosts.     

Campylobacter enteritis associated with consumption of unpasteurised milk.

In October and November 1978 two outbreaks of enteritis occurred in the north of England. Symptoms lasted two to over eight days but in no case necessitated admission to hospital. Faecal specimens from most of the patients were found to contain thermophilic Campylobacter sp. Inquiry disclosed that all patients had consumed unpasteurised milk from local farms. Examination of rectal swabs from the cattle concerned and milk socks yielded strains of Campylobacter sp indistinguishable from those isolated from the patients. It was therefore concluded that, since campylobacters are not known to be excreted in milk, faecal contamination of the milk had probably occurred and had led to these outbreaks. Evidence suggests that thermophilic Campylobacter sp is an occasional contaminant of milk. So long as unpasteurised milk continues to be distributed further outbreaks will probably occur.     

Campylobacter jejuni: molecular biology and pathogenesis

Campylobacter jejuni is a foodborne bacterial pathogen that is common in the developed world. However, we know less about its biology and pathogenicity than we do about other less prevalent pathogens. Interest in C. jejuni has increased in recent years as a result of the growing appreciation of its importance as a pathogen and the availability of new model systems and genetic and genomic technologies. C. jejuni establishes persistent, benign infections in chickens and is rapidly cleared by many strains of laboratory mouse, but causes significant inflammation and enteritis in humans. Comparing the different host responses to C. jejuni colonization should increase our understanding of this organism.     

Changes in peptic digestibility of bovine beta-lactoglobulin as a result of food processing studied by capillary electrophoresis and immunochemical methods

Digestion studies constitute a functional tool for allergen characterisation. This strategy for investigating allergenic proteins relates to the observation of increased proteolytic resistance of some proteins recognised to exhibit allergenic potential. beta-Lactoglobulin (betaLG) is one of the major whey proteins, a potent milk allergen and shows a high stability against peptic hydrolysis in its native form. In order to study the impact of milk fermentation process on its digestibility, two complementary analytical methods were applied: capillary zone electrophoresis (CZE) to quantitatively study proteolytic degradation of betaLG isolated from different fermented bovine milk products, and enzyme linked immunosorbent assay (ELISA) to assess differences in immunoreactivity. betaLG, isolated from either raw or pasteurised cow's milk (CM), as expected, showed only minimal digestibility (less than 10% in 2 h). However, when raw milk or pasteurised milk was fermented, the rate of peptic digestion of the protein significantly increased (up to 45% in 2 h). In accordance with changes in digestibility, the immunochemical response for all fermented samples was lower than that of non-fermented references. Raw and pasteurised milk "naturally" fermented in our laboratory only resulted in a slight reduction (betaLG detected is still in the range of milligrams per gram sample), whereas the industrially manufactured sour milk as well as the "Acidophilus milk" reflected a remarkably lower level of immunoreactivity (55-56 microg/g sample).     

Large outbreak of Salmonella enterica serotype paratyphi B infection caused by a goats' milk cheese,France, 1993: a case finding and epidemiological study.

OBJECTIVE--To assess the magnitude of a nationwide outbreak of infection with Salmonella enterica serotype paratyphi B and identify the vehicle and source of infection. DESIGN--A case finding study of S paratyphi B infection between 15 August and 30 November 1993; a pair matched case-control study; an environmental investigation at a processing plant that produced a raw goats'' milk cheese incriminated in the outbreak; phage typing and genotyping of food and human S paratyphi B isolates. SETTING--France, 15 August to 30 November 1993. SUBJECTS--273 patients with S paratyphi B infection; 59 pairs of cases and controls matched for age, sex, and city of residence. MAIN OUTCOME MEASURES--Numbers of cases and incidence rates by region of residence and age; matched odds ratios for dairy food preferences. RESULTS--Among the 273 cases there was one death; 203 (78%) strains belonged to phage type 1 var 3. The incidence of infection was greatest in the region where goats'' milk cheese is commonly produced. Comparison of cases and controls showed a 12-fold greater risk of illness (95% confidence interval 1.6 to 92.3) from eating brand A unpasteurised goats'' milk cheese. S paratyphi B isolates of phage type 1 var 3 were recovered from cheese A, goats'' milk at the plant processing cheese A, and goats'' milk supplied to the plant by a single farm. Genotypic IS 200 typing of food and human 1 var 3 phage type isolates showed a common IS 200 pattern. CONCLUSION--This outbreak emphasises the potential health hazards of widely distributed unpasteurised milk products in France and the need for their close bacterial monitoring.     

Application of real-time PCR for quantitative detection of Campylobacter jejuni in poultry, milk and environmental water

Campylobacter jejuni is a leading human food-borne pathogen. The rapid and sensitive detection of C. jejuni is necessary for the maintenance of a safe food/water supply. In this article, we present a real-time polymerase chain reaction (PCR) assay for quantitative detection of C. jejuni in naturally contaminated poultry, milk and environmental samples without an enrichment step. The whole assay can be completed in 60 min with a detection limit of approximately 1 CFU. The standard curve correlation coefficient for the threshold cycle versus the copy number of initial C. jejuni cells was 0.988. To test the PCR system, a set of 300 frozen chicken meat samples, 300 milk samples and 300 water samples were screened for the presence of C. jejuni. 30.6% (92/300) of chicken meat samples, 27.3% (82/300) of milk samples, and 13.6% (41/300) of water samples tested positive for C. jejuni. This result indicated that the real-time PCR assay provides a specific, sensitive and rapid method for quantitative detection of C. jejuni. Moreover, it is concluded that retail chicken meat, raw milk and environmental water are commonly contaminated with C. jejuni and could serve as a potential risk for consumers in eastern China, especially if proper hygienic and cooking conditions are not maintained.     

Survival of Campylobacter jejuni in raw, pasteurized and ultra-heat-treated goat's milk stored at different temperatures

The survival of a human strain of Campylobacter jejuni in raw, pasteurized and ultra-heat-treated goat's milk stored at 5°, 10°, 15° and 20°C was studied. No viable units were detected in raw milk after 24 h at 20°C and 48 h at 15°C. None were detected in pasteurized milk after 48 h at 20°C. In all other samples, there was a decline in viable units in the first 24 h but very little decline in the next 24 h period. The organism survived best at 5° and 10° C.     

Characterization of mono- and mixed-culture Campylobacter jejuni biofilms

Campylobacter jejuni, one of the most common causes of human gastroenteritis, is a thermophilic and microaerophilic bacterium. These characteristics make it a fastidious organism, which limits its ability to survive outside animal hosts. Nevertheless, C. jejuni can be transmitted to both humans and animals via environmental pathways, especially through contaminated water. Biofilms may play a crucial role in the survival of the bacterium under unfavorable environmental conditions. The goal of this study was to investigate survival strategies of C. jejuni in mono- and mixed-culture biofilms. We grew monoculture biofilms of C. jejuni and mixed-culture biofilms of C. jejuni with Pseudomonas aeruginosa. We found that mono- and mixed-culture biofilms had significantly different structures and activities. Monoculture C. jejuni biofilms did not consume a measurable quantity of oxygen. Using a confocal laser scanning microscope (CLSM), we found that cells from monoculture biofilms were alive according to live/dead staining but that these cells were not culturable. In contrast, in mixed-culture biofilms, C. jejuni remained in a culturable physiological state. Monoculture C. jejuni biofilms could persist under lower flow rates (0.75 ml/min) but were unable to persist at higher flow rates (1 to 2.5 ml/min). In sharp contrast, mixed-culture biofilms were more robust and were unaffected by higher flow rates (2.5 ml/min). Our results indicate that biofilms provide an environmental refuge that is conducive to the survival of C. jejuni.     

5'-methylthioadenosine nucleosidase is implicated in playing a key role in a modified futalosine pathway for menaquinone biosynthesis in Campylobacter jejuni

Menaquinone (vitamin K(2)) serves as an electron carrier in the electron transport chain required for respiration in many pathogenic bacteria. Most bacteria utilize a common menaquinone biosynthetic pathway as exemplified by Escherichia coli. Recently, a novel biosynthetic pathway, the futalosine pathway, was discovered in Streptomyces. Bioinformatic analysis strongly suggests that this pathway is also operative in the human pathogens Campylobacter jejuni and Helicobacter pylori. Here, we provide compelling evidence that a modified futalosine pathway is operative in C. jejuni and that it utilizes 6-amino-6-deoxyfutalosine instead of futalosine. A key step in the Streptomyces pathway involves a nucleosidase called futalosine hydrolase. The closest homolog in C. jejuni has been annotated as a 5'-methylthioadenosine nucleosidase (MTAN). We have shown that this C. jejuni enzyme has MTAN activity but negligible futalosine hydrolase activity. However, the C. jejuni MTAN is able to hydrolyze 6-amino-6-deoxyfutalosine at a rate comparable with that of its known substrates. This suggests that the adenine-containing version of futalosine is the true biosynthetic intermediate in this organism. To demonstrate this in vivo, we constructed a C. jejuni mutant strain deleted for mqnA2, which is predicted to encode for the enzyme required to synthesize 6-amino-6-deoxyfutalosine. Growth of this mutant was readily rescued by the addition of 6-amino-6-deoxyfutalosine, but not futalosine. This provides the first direct evidence that a modified futalosine pathway is operative in C. jejuni. It also highlights the tremendous versatility of the C. jejuni MTAN, which plays key roles in S-adenosylmethionine recycling, the biosynthesis of autoinducer molecules, and the biosynthesis of menaquinone.     

Procedure for increased recovery of Campylobacter jejuni from inoculated unpasteurized milk.

Different treatments were applied to Campylobacter jejuni-inoculated unpasteurized milk to identify means of enhancing the survival of the organism in refrigerated (4 degrees C) samples. The greatest survival occurred in milk supplemented with 0.01% sodium bisulfite and held under an atmosphere of 100% nitrogen (bisulfite-nitrogen), in most instances allowing isolation of C. jejuni from highly contaminated milk 15 or more days longer than from unsupplemented milk held in air (21% oxygen). Although a larger amount of Campylobacter was consistently recovered from milk treated with bisulfite-nitrogen, similar isolation rates (qualitative) resulted from milk stored in air and supplemented with 0.01% sodium bisulfite and 0.15% sodium thioglycolate when analyzed within 12 days after sampling. Milk samples to be transported and assayed at a later date would best be held refrigerated (4 degrees C) and supplemented with 0.01% sodium bisulfite and either 0.15% sodium thioglycolate or an atmosphere of 100% nitrogen.     

Procedure for increased recovery of Campylobacter jejuni from inoculated unpasteurized milk

Different treatments were applied to Campylobacter jejuni-inoculated unpasteurized milk to identify means of enhancing the survival of the organism in refrigerated (4 degrees C) samples. The greatest survival occurred in milk supplemented with 0.01% sodium bisulfite and held under an atmosphere of 100% nitrogen (bisulfite-nitrogen), in most instances allowing isolation of C. jejuni from highly contaminated milk 15 or more days longer than from unsupplemented milk held in air (21% oxygen). Although a larger amount of Campylobacter was consistently recovered from milk treated with bisulfite-nitrogen, similar isolation rates (qualitative) resulted from milk stored in air and supplemented with 0.01% sodium bisulfite and 0.15% sodium thioglycolate when analyzed within 12 days after sampling. Milk samples to be transported and assayed at a later date would best be held refrigerated (4 degrees C) and supplemented with 0.01% sodium bisulfite and either 0.15% sodium thioglycolate or an atmosphere of 100% nitrogen.     

Cross-contamination in the kitchen: effect of hygiene measures

Aims:  To determine the effect of hygiene measures on cross-contamination of Campylobacter jejuni at home and to select a safe tracer organism for C. jejuni.
Methods and Results:  Comparative tests were conducted with nonpathogenic Escherichia coli and Lactobacillus casei and L. casei was chosen as the safe tracer organism. Salads containing chicken breast fillet contaminated with a known number of C. jejuni and L. casei were prepared according to different cross-contamination scenarios and contamination levels of salads were determined. Cross-contamination could be strongly reduced when cleaning cutting board and cutlery with hot water (68°C), but generally was not prevented using consumer-style cleaning methods for hands and cutting board.
Conclusions:  Dish-washing does not sufficiently prevent cross-contamination, thus different cutting boards for raw meat and other ingredients should be used and meat–hand contact should be avoided or hands should be thoroughly cleaned with soap. Lactobacillus casei can be used as a safe tracer organism for C. jejuni in consumer observational studies.
Significance and Impact of the Study:  Cross-contamination plays an important role in the transmission of food-borne illness, especially for C. jejuni . This study delivers suitable data to quantitatively assess the risk of campylobacteriosis caused by cross-contamination and it shows the effect of different preventive hygiene measures.     

Cellular prion protein in ovine milk

Cellular prion protein, PrP(C), is essential for the development of prion diseases where it is considered to be a substrate for the formation of the disease-associated conformer, PrP(Sc). In sheep, PrP(C) is abundant in neuronal tissue and is also found at lower concentrations in a range of non-neuronal tissues, including mammary gland. Here, we demonstrate the presence of soluble PrP(C) in the non-cellular, non-lipid fraction of clarified ovine milk. Compared with brain-derived PrP(C), ovine milk PrP(C) displays an increased electrophoretic mobility. Ovine milk PrP(C) is mainly present as three species that differ in the extent of their N-linked glycosylation, with glycoform profiles varying among animals. Similar PrP(C) species are also present in fresh and commercial homogenised/pasteurised bovine milk, with additional N-terminal PrP(C) fragments detectable in ruminant milk and commercial milk products.     

Association of Campylobacter jejuni with laying hens and eggs

Laying hens were individually caged at 20 weeks of age and tested for fecal excretion of Campylobacter jejuni (minimum level of detection was 100 cfu/g) during a 42-week period. Peak rates of C. jejuni isolation (approximately 25% of hens positive) occurred at two different times, in October and in late April to early May. Before being segregated in late September, birds were allowed to consume fecal matter, litter, and communal drinking water, all likely sources of C. jejuni. The increased excretion rate in late April may have been due to a climatic change. A small portion (8.1%) of the hens chronically excreted (positive less than 30% of the sampling times) the organism, whereas C. jejuni was not detected in 33% of the hens, even though birds were likely exposed to the organism before being segregated. No correlation could be made between rates of C. jejuni excretion and egg production. Of 266 eggs from hens fecally excreting C. jejuni, the organism was isolated from two shell surfaces but no egg contents. Egg penetration studies revealed that the organism would not penetrate into the contents of the eggs but could be isolated occasionally from the inner shell and membranes of refrigerated eggs.