Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B stra... more Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B strains isolated from various sources revealed two homogeneous clusters, clusters BI and BII. The type B strains differed from reference strain ATCC 3502 by 413 coding sequence (CDS) probes, sharing 88% of all the ATCC 3502 genes represented on the microarray. The two Nordic type B clusters differed from each other by their response to 145 CDS probes related mainly to transport and binding, adaptive mechanisms, fatty acid biosynthesis, the cell membranes, bacteriophages, and transposon-related elements. The most prominent differences between the two clusters were related to resistance to toxic compounds frequently found in the environment, such as arsenic and cadmium, reflecting different adaptive responses in the evolution of the two clusters. Other relatively variable CDS groups were related to surface structures and the gram-positive cell wall, suggesting that the two clusters possess different antigenic properties. All the type B strains carried CDSs putatively related to capsule formation, which may play a role in adaptation to different environmental and clinical niches. Sequencing showed that representative strains of the two type B clusters both carried subtype B2 neurotoxin genes. As many of the type B strains studied have been isolated from foods or associated with botulism, it is expected that the two group I C. botulinum type B clusters present a public health hazard in Nordic countries. Knowing the genetic and physiological markers of these clusters will assist in targeting control measures against these pathogens.
Good governance is considered as the application of the regulations and legal principles regulati... more Good governance is considered as the application of the regulations and legal principles regulationg the public administration. This study examined whether the principle of good governance is realised in the compulsory actions applied by Finnish authorities in the sector of environmental health and fooc control. The results indicated that there are marked shortcomings in implementing administrative procedures, protecting legal rights and applying the principle of good governance. The most significant deficiencies were noted with the hearing process, argumentations of the decisions and instructions for appeals. The practices for compulsory actions seemed to vary in different municipalities, suggesting that entrepreneurs may be placed in an unequal position depending on the geographical location of the company.
Food control research is a rather new field of research, directed towards evaluating the efficacy... more Food control research is a rather new field of research, directed towards evaluating the efficacy and influence of food control on food safety. Food control is financed by society, which regulates needs in order to estimate the return on that investment. Does food control truly impact food safety and promote human health? Through food control research we may lay the scientific basis for the development of new control measures and policies. The occurrence of food-borne illnesses and food-related malpractices highlight the importance of an effective food control strategy. According to a broad literature search, the number of scientific publications in this field of research is growing, which may in part, reflect growing public concern about food safety. The main areas covered in food control research today include evaluating the hazard analysis of critical control point (HACCP) system, assessing the economic impact of implementing food control measures and evaluating national food con...
Food control systems are sometimes challenged when requirements set by law are intentionally viol... more Food control systems are sometimes challenged when requirements set by law are intentionally violated by Food Business Operators (FBOs) deliberately putting food on the market with the intention of deceiving the consumer (food fraud/adulteration). There is also a growing concern that in some cases food frauds could be considered more risky than traditional food safety threats since their public health risks are often unconventional and difficult to detect. In this study, food frauds/adulterations published in the EU Rapid Alert System for Food and Feed (RASFF) (n = 376) in 2008–2012, notifications of recalls published by the Finnish Food Safety Authority (Evira) (n = 50) in 2008–2012 and local Finnish cases (n = 16) in 2003–2012 were analysed to determine the overall pattern of reported frauds/adulterations in 2008–2012. Products originating from outside the borders were most often reported by both RASFF (92%) and Evira (90%), whereas local Finnish cases mainly dealt with domestic p...
ABSTRACT Clostridium botulinum is a diverse group of anaerobic spore-forming organisms that produ... more ABSTRACT Clostridium botulinum is a diverse group of anaerobic spore-forming organisms that produce lethal botulinum neurotoxins (BoNT) during their growth. BoNTs cause a paralytic condition, botulism, to man and animals. The most common forms of human botulism include the classical foodborne botulism due to ingestion of BoNT preformed in food, and infant botulism due to spore germination, growth, and toxin production in the infant’s intestine. Botulism is diagnosed by detecting BoNT and/or C. botulinum in the patient and in suspected food samples. There are several drawbacks related to the diagnostics of botulism; the standard bioassay for toxin detection employs the use of laboratory animals, making it laborious and expensive and possessing ethical concern. Selective media for culturing the organism are not available. Neurotoxin gene-specific PCR has facilitated the detection of C. botulinum. In this chapter a multiplex PCR for simultaneous detection of C. botulinum types A, B, E, and F in foods and feces is described. The method involves sample dilution and homogenization, and two-step enrichment followed by cell wash, cell lysis, and multiplex PCR. Quantification is obtained by the most-probable-number technique. Depending on the type of sample material, the detection limit of the assay varies from 10−2 to 103 spores per gram of sample material. Key WordsClostridium botulinum–botulism–botulinum neurotoxin–BoNT–bot–multiplex PCR
ABSTRACT We evaluated the consistency and efficacy of inspection processes of 17 Finnish municipa... more ABSTRACT We evaluated the consistency and efficacy of inspection processes of 17 Finnish municipal food control units by analyzing inspection reports of 83 restaurants over the 2007-2011 period and by interviewing food business operators. Fifty-six officials of the food control units responded to an electronic questionnaire about their inspection activities. We found several factors related to the food establishment, the food control official and the working unit of the official that can affect the inspection processes and the efficacy of the controls. The use of checklists and templates for inspection reports enhanced the consistency and efficacy of the controls. The templates also reduced the time used for, and increased the quality of, reporting. Time limits for correcting non-compliances had a significant effect on the efficacy of controls.
The main responsibility of the local food control lies in the local food and environmental health... more The main responsibility of the local food control lies in the local food and environmental health units. The producer is responsible for the safety of the food produced, and applies an in-house control system to achieve this goal. The local food safety authority evaluates the in-house control system and assures that it is targeted to the most critical steps in the production to efficiently minimize the food safety risks. The main challenges in implementing efficient food control system are the variability in the knowledge of the producer about food hygiene, inadequate resources in control units and harmonization of the control measures in national level.
The antibacterial properties of 13 essential oils, derived from spices grown in Finland, were exa... more The antibacterial properties of 13 essential oils, derived from spices grown in Finland, were examined with an agar diffusion method against 12 bacterial strains. The organisms tested included both spoilage and pathogenic bacteria. The gram-positive bacteria appeared to be more sensitive than the gram-negative organisms, Clostridium botulinum and Clostridium perfringens being the most sensitive. Oregano, savory, and thyme showed the broadest antibacterial activity by distinctly inhibiting the growth of all the organisms tested. By gas chromatography-mass spectrometry analysis, differences were noted in the composition of oregano and thyme oils in comparison to previous reports.
ABSTRACT Enforcement measures are an administrative tool used by food control authorities for mak... more ABSTRACT Enforcement measures are an administrative tool used by food control authorities for making food business operators (FBOs) comply with food safety regulations. This study evaluates the effectiveness and uniformity of the use of enforcement measures in local food control units in Finland. The study investigates whether and how rapidly the use enforcement measures leads to compliance with food safety regulations and whether the enforcement processes and their durations vary according to the seriousness of non-compliance. Enforcement decisions were requested from 29 (34%) local food control units, ten of which had used no enforcement measures during the study period. A total of 188 enforcement cases from 19 control units were analysed. The food safety violations were categorized into critical and non-critical, based on their likeliness to directly contribute to food contamination or foodborne outbreaks.
Applied and Environmental Microbiology - AEM, 2009
Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B stra... more Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B strains isolated from various sources revealed two homogeneous clusters, clusters BI and BII. The type B strains differed from reference strain ATCC 3502 by 413 coding sequence (CDS) probes, sharing 88% of all the ATCC 3502 genes represented on the microarray. The two Nordic type B clusters differed from each other by their response to 145 CDS probes related mainly to transport and binding, adaptive mechanisms, fatty acid biosynthesis, the cell membranes, bacteriophages, and transposon-related elements. The most prominent differences between the two clusters were related to resistance to toxic compounds frequently found in the environment, such as arsenic and cadmium, reflecting different adaptive responses in the evolution of the two clusters. Other relatively variable CDS groups were related to surface structures and the gram-positive cell wall, suggesting that the two clusters possess dif...
Clostridium botulinum type B was detected by multiplex PCR in the intestinal contents of a sudden... more Clostridium botulinum type B was detected by multiplex PCR in the intestinal contents of a suddenly deceased 11-week-old infant and in vacuum cleaner dust from the patient's household. C. botulinum was also isolated from the deceased infant's intestinal contents and from the household dust. The genetic similarity of the two isolates was demonstrated by pulsed-field gel electrophoresis and randomly amplified polymorphic DNA analysis, thereby confirming that dust may act as a vehicle for infant botulism that results in sudden death.
The largest reported outbreak of type C botulism in fur production animals is described. Epidemio... more The largest reported outbreak of type C botulism in fur production animals is described. Epidemiological investigation of 117 out of 157 (response rate, 74.5%) farms revealed that 44,130 animals died or were euthanized, while 8,033 animals with milder symptoms recovered. The overall death rate in all animals at risk was 21.7%. The death rates were significantly higher in blue and shadow foxes (24.2 and 27.8%, respectively) than in silver and blue silver foxes and minks (below 4%). All minks had been immunized against botulinum toxin type C. Deaths were associated with feed manufactured by a local processor, 83 of whose customer farms (70.9%) reported dead or sick animals. Five feedlots out of 19 delivered to the farms on the day preceding the onset of the outbreak (day 2) were associated with a death rate higher than 40%. These feedlots consisted of fresh feed processed on day 2 and feed processed 1 day earlier (day 1). In laboratory analysis, the day 2 feed contained botulinum toxin type C (>600 minimum lethal doses/g), while the day 1 feed did not contain toxin. Toxin was not detected in feed raw-material samples. Clostridium botulinum type C was detected by PCR in some feed components and in feed. However, as the feed temperature was continuously 8°C or below and the pH was continuously 5.6 or below according to the manufacturer, it seems unlikely that spore germination and toxin formation occurred during overnight storage. Hence, the events leading to toxin formation were not determined.
A total of 294 honey samples produced in Denmark, Norway and Sweden were studied for the presence... more A total of 294 honey samples produced in Denmark, Norway and Sweden were studied for the presence of Clostridium botulinum types A, B, E and F by using a multiplex-PCR method. The samples consisted of honeycombs taken directly from beehives, and extracted honey representing several hives or apiaries. The prevalence of C. botulinum showed a significant variation between Denmark, Norway and Sweden, the proportions of positive samples being 26%, 10% and 2%, respectively. The major serotype detected was type B. When analysed with pulsed-field gel electrophoresis (PFGE) using restriction enzyme SacII, the 24 strains isolated produced eight different PFGE patterns. At a similarity level of 95%, four clusters were produced, three of which contained 20 of the 24 analysed strains. One of the clusters included isolates from both Denmark and Norway. D
To determine the prevalence of Clostridium botulinum types A, B, E, and/or F in pig intestinal sa... more To determine the prevalence of Clostridium botulinum types A, B, E, and/or F in pig intestinal samples, different PCR-based methods were compared using artificially contaminated faeces. The methods included a multiplex PCR targeted to botulinum neurotoxin genes A, B, E, and F or a nested PCR targeted to toxin genes B, E, and F, combined with different pre-enrichment protocols and forms of templates. A method including the two-step enrichment followed by DNA extraction and multiplex PCR yielded the highest number of positives. This assay protocol was employed to investigate 100 pig intestinal samples. The sample materials studied included colon wall, intestinal content, and mucus peeled from the colon wall. Three pigs (3%) were positive for C. botulinum type B, and no other toxinotypes were detected in any sample. The number of positive samples was higher when colon wall or peeled mucus was analyzed compared to the intestinal content. C. botulinum was isolated from two PCR-positive samples and confirmed to be type B by PCR. Both isolates were shown to be proteolytic and thus to represent C. botulinum group I.
A test protocol for reliable detection of Clostridium botulinum types A and B spores in honey by ... more A test protocol for reliable detection of Clostridium botulinum types A and B spores in honey by polymerase chain reaction (PCR) was developed and used for a prevalence survey of C. botulinum spores in 190 honey samples. The inhibiting effects of honey on microbial growth and PCR analysis were overcome by using a method of supernatant filtration (SF) in the preparation of the samples before enrichment and PCR. By using this method, an inoculum of 0.1 spore of C. botulinum/g honey could be detected. In the prevalence survey, spores of C. botulinum were detected in 8 (7%) of the 114 Finnish and in 12 (16%) of the 76 imported honey samples. The quantity of spores in PCR-positive samples varied from less than 18 to 140 spores/kg. Neurotoxin gene sequences corresponding to C. botulinum type A were detected in 17 samples and proteolytic type B in 12 samples by PCR analysis. Both types A and B were detected in nine samples. Strains of C. botulinum type A were isolated from 14 and type B from 2 of the 20 PCR-positive samples. This is the first report of type A spores of C. botulinum being detected and isolated in Fennoscandia.
Whitefish eggs were confirmed by pulsed-field gel electrophoresis to cause type E foodborne botul... more Whitefish eggs were confirmed by pulsed-field gel electrophoresis to cause type E foodborne botulism in a 54-yearold patient in Finland. Botulinum neurotoxin and/or nonproteolytic Clostridium botulinum type E organisms were detected in fecal and gastric samples from the patient and in suspected whitefish eggs. Apart from C. botulinum type E, proteolytic type B organisms were detected in the patient's gastric content. This was considered to be insignificant with respect to the clinical disease, suggesting botulinal spores to be occasionally present in the human gastrointestinal tract without any apparent clinical significance. This is the first domestic case of foodborne botulism in Finland.
Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B stra... more Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B strains isolated from various sources revealed two homogeneous clusters, clusters BI and BII. The type B strains differed from reference strain ATCC 3502 by 413 coding sequence (CDS) probes, sharing 88% of all the ATCC 3502 genes represented on the microarray. The two Nordic type B clusters differed from each other by their response to 145 CDS probes related mainly to transport and binding, adaptive mechanisms, fatty acid biosynthesis, the cell membranes, bacteriophages, and transposon-related elements. The most prominent differences between the two clusters were related to resistance to toxic compounds frequently found in the environment, such as arsenic and cadmium, reflecting different adaptive responses in the evolution of the two clusters. Other relatively variable CDS groups were related to surface structures and the gram-positive cell wall, suggesting that the two clusters possess different antigenic properties. All the type B strains carried CDSs putatively related to capsule formation, which may play a role in adaptation to different environmental and clinical niches. Sequencing showed that representative strains of the two type B clusters both carried subtype B2 neurotoxin genes. As many of the type B strains studied have been isolated from foods or associated with botulism, it is expected that the two group I C. botulinum type B clusters present a public health hazard in Nordic countries. Knowing the genetic and physiological markers of these clusters will assist in targeting control measures against these pathogens.
Good governance is considered as the application of the regulations and legal principles regulati... more Good governance is considered as the application of the regulations and legal principles regulationg the public administration. This study examined whether the principle of good governance is realised in the compulsory actions applied by Finnish authorities in the sector of environmental health and fooc control. The results indicated that there are marked shortcomings in implementing administrative procedures, protecting legal rights and applying the principle of good governance. The most significant deficiencies were noted with the hearing process, argumentations of the decisions and instructions for appeals. The practices for compulsory actions seemed to vary in different municipalities, suggesting that entrepreneurs may be placed in an unequal position depending on the geographical location of the company.
Food control research is a rather new field of research, directed towards evaluating the efficacy... more Food control research is a rather new field of research, directed towards evaluating the efficacy and influence of food control on food safety. Food control is financed by society, which regulates needs in order to estimate the return on that investment. Does food control truly impact food safety and promote human health? Through food control research we may lay the scientific basis for the development of new control measures and policies. The occurrence of food-borne illnesses and food-related malpractices highlight the importance of an effective food control strategy. According to a broad literature search, the number of scientific publications in this field of research is growing, which may in part, reflect growing public concern about food safety. The main areas covered in food control research today include evaluating the hazard analysis of critical control point (HACCP) system, assessing the economic impact of implementing food control measures and evaluating national food con...
Food control systems are sometimes challenged when requirements set by law are intentionally viol... more Food control systems are sometimes challenged when requirements set by law are intentionally violated by Food Business Operators (FBOs) deliberately putting food on the market with the intention of deceiving the consumer (food fraud/adulteration). There is also a growing concern that in some cases food frauds could be considered more risky than traditional food safety threats since their public health risks are often unconventional and difficult to detect. In this study, food frauds/adulterations published in the EU Rapid Alert System for Food and Feed (RASFF) (n = 376) in 2008–2012, notifications of recalls published by the Finnish Food Safety Authority (Evira) (n = 50) in 2008–2012 and local Finnish cases (n = 16) in 2003–2012 were analysed to determine the overall pattern of reported frauds/adulterations in 2008–2012. Products originating from outside the borders were most often reported by both RASFF (92%) and Evira (90%), whereas local Finnish cases mainly dealt with domestic p...
ABSTRACT Clostridium botulinum is a diverse group of anaerobic spore-forming organisms that produ... more ABSTRACT Clostridium botulinum is a diverse group of anaerobic spore-forming organisms that produce lethal botulinum neurotoxins (BoNT) during their growth. BoNTs cause a paralytic condition, botulism, to man and animals. The most common forms of human botulism include the classical foodborne botulism due to ingestion of BoNT preformed in food, and infant botulism due to spore germination, growth, and toxin production in the infant’s intestine. Botulism is diagnosed by detecting BoNT and/or C. botulinum in the patient and in suspected food samples. There are several drawbacks related to the diagnostics of botulism; the standard bioassay for toxin detection employs the use of laboratory animals, making it laborious and expensive and possessing ethical concern. Selective media for culturing the organism are not available. Neurotoxin gene-specific PCR has facilitated the detection of C. botulinum. In this chapter a multiplex PCR for simultaneous detection of C. botulinum types A, B, E, and F in foods and feces is described. The method involves sample dilution and homogenization, and two-step enrichment followed by cell wash, cell lysis, and multiplex PCR. Quantification is obtained by the most-probable-number technique. Depending on the type of sample material, the detection limit of the assay varies from 10−2 to 103 spores per gram of sample material. Key WordsClostridium botulinum–botulism–botulinum neurotoxin–BoNT–bot–multiplex PCR
ABSTRACT We evaluated the consistency and efficacy of inspection processes of 17 Finnish municipa... more ABSTRACT We evaluated the consistency and efficacy of inspection processes of 17 Finnish municipal food control units by analyzing inspection reports of 83 restaurants over the 2007-2011 period and by interviewing food business operators. Fifty-six officials of the food control units responded to an electronic questionnaire about their inspection activities. We found several factors related to the food establishment, the food control official and the working unit of the official that can affect the inspection processes and the efficacy of the controls. The use of checklists and templates for inspection reports enhanced the consistency and efficacy of the controls. The templates also reduced the time used for, and increased the quality of, reporting. Time limits for correcting non-compliances had a significant effect on the efficacy of controls.
The main responsibility of the local food control lies in the local food and environmental health... more The main responsibility of the local food control lies in the local food and environmental health units. The producer is responsible for the safety of the food produced, and applies an in-house control system to achieve this goal. The local food safety authority evaluates the in-house control system and assures that it is targeted to the most critical steps in the production to efficiently minimize the food safety risks. The main challenges in implementing efficient food control system are the variability in the knowledge of the producer about food hygiene, inadequate resources in control units and harmonization of the control measures in national level.
The antibacterial properties of 13 essential oils, derived from spices grown in Finland, were exa... more The antibacterial properties of 13 essential oils, derived from spices grown in Finland, were examined with an agar diffusion method against 12 bacterial strains. The organisms tested included both spoilage and pathogenic bacteria. The gram-positive bacteria appeared to be more sensitive than the gram-negative organisms, Clostridium botulinum and Clostridium perfringens being the most sensitive. Oregano, savory, and thyme showed the broadest antibacterial activity by distinctly inhibiting the growth of all the organisms tested. By gas chromatography-mass spectrometry analysis, differences were noted in the composition of oregano and thyme oils in comparison to previous reports.
ABSTRACT Enforcement measures are an administrative tool used by food control authorities for mak... more ABSTRACT Enforcement measures are an administrative tool used by food control authorities for making food business operators (FBOs) comply with food safety regulations. This study evaluates the effectiveness and uniformity of the use of enforcement measures in local food control units in Finland. The study investigates whether and how rapidly the use enforcement measures leads to compliance with food safety regulations and whether the enforcement processes and their durations vary according to the seriousness of non-compliance. Enforcement decisions were requested from 29 (34%) local food control units, ten of which had used no enforcement measures during the study period. A total of 188 enforcement cases from 19 control units were analysed. The food safety violations were categorized into critical and non-critical, based on their likeliness to directly contribute to food contamination or foodborne outbreaks.
Applied and Environmental Microbiology - AEM, 2009
Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B stra... more Comparative genomic hybridization analysis of 32 Nordic group I Clostridium botulinum type B strains isolated from various sources revealed two homogeneous clusters, clusters BI and BII. The type B strains differed from reference strain ATCC 3502 by 413 coding sequence (CDS) probes, sharing 88% of all the ATCC 3502 genes represented on the microarray. The two Nordic type B clusters differed from each other by their response to 145 CDS probes related mainly to transport and binding, adaptive mechanisms, fatty acid biosynthesis, the cell membranes, bacteriophages, and transposon-related elements. The most prominent differences between the two clusters were related to resistance to toxic compounds frequently found in the environment, such as arsenic and cadmium, reflecting different adaptive responses in the evolution of the two clusters. Other relatively variable CDS groups were related to surface structures and the gram-positive cell wall, suggesting that the two clusters possess dif...
Clostridium botulinum type B was detected by multiplex PCR in the intestinal contents of a sudden... more Clostridium botulinum type B was detected by multiplex PCR in the intestinal contents of a suddenly deceased 11-week-old infant and in vacuum cleaner dust from the patient's household. C. botulinum was also isolated from the deceased infant's intestinal contents and from the household dust. The genetic similarity of the two isolates was demonstrated by pulsed-field gel electrophoresis and randomly amplified polymorphic DNA analysis, thereby confirming that dust may act as a vehicle for infant botulism that results in sudden death.
The largest reported outbreak of type C botulism in fur production animals is described. Epidemio... more The largest reported outbreak of type C botulism in fur production animals is described. Epidemiological investigation of 117 out of 157 (response rate, 74.5%) farms revealed that 44,130 animals died or were euthanized, while 8,033 animals with milder symptoms recovered. The overall death rate in all animals at risk was 21.7%. The death rates were significantly higher in blue and shadow foxes (24.2 and 27.8%, respectively) than in silver and blue silver foxes and minks (below 4%). All minks had been immunized against botulinum toxin type C. Deaths were associated with feed manufactured by a local processor, 83 of whose customer farms (70.9%) reported dead or sick animals. Five feedlots out of 19 delivered to the farms on the day preceding the onset of the outbreak (day 2) were associated with a death rate higher than 40%. These feedlots consisted of fresh feed processed on day 2 and feed processed 1 day earlier (day 1). In laboratory analysis, the day 2 feed contained botulinum toxin type C (>600 minimum lethal doses/g), while the day 1 feed did not contain toxin. Toxin was not detected in feed raw-material samples. Clostridium botulinum type C was detected by PCR in some feed components and in feed. However, as the feed temperature was continuously 8°C or below and the pH was continuously 5.6 or below according to the manufacturer, it seems unlikely that spore germination and toxin formation occurred during overnight storage. Hence, the events leading to toxin formation were not determined.
A total of 294 honey samples produced in Denmark, Norway and Sweden were studied for the presence... more A total of 294 honey samples produced in Denmark, Norway and Sweden were studied for the presence of Clostridium botulinum types A, B, E and F by using a multiplex-PCR method. The samples consisted of honeycombs taken directly from beehives, and extracted honey representing several hives or apiaries. The prevalence of C. botulinum showed a significant variation between Denmark, Norway and Sweden, the proportions of positive samples being 26%, 10% and 2%, respectively. The major serotype detected was type B. When analysed with pulsed-field gel electrophoresis (PFGE) using restriction enzyme SacII, the 24 strains isolated produced eight different PFGE patterns. At a similarity level of 95%, four clusters were produced, three of which contained 20 of the 24 analysed strains. One of the clusters included isolates from both Denmark and Norway. D
To determine the prevalence of Clostridium botulinum types A, B, E, and/or F in pig intestinal sa... more To determine the prevalence of Clostridium botulinum types A, B, E, and/or F in pig intestinal samples, different PCR-based methods were compared using artificially contaminated faeces. The methods included a multiplex PCR targeted to botulinum neurotoxin genes A, B, E, and F or a nested PCR targeted to toxin genes B, E, and F, combined with different pre-enrichment protocols and forms of templates. A method including the two-step enrichment followed by DNA extraction and multiplex PCR yielded the highest number of positives. This assay protocol was employed to investigate 100 pig intestinal samples. The sample materials studied included colon wall, intestinal content, and mucus peeled from the colon wall. Three pigs (3%) were positive for C. botulinum type B, and no other toxinotypes were detected in any sample. The number of positive samples was higher when colon wall or peeled mucus was analyzed compared to the intestinal content. C. botulinum was isolated from two PCR-positive samples and confirmed to be type B by PCR. Both isolates were shown to be proteolytic and thus to represent C. botulinum group I.
A test protocol for reliable detection of Clostridium botulinum types A and B spores in honey by ... more A test protocol for reliable detection of Clostridium botulinum types A and B spores in honey by polymerase chain reaction (PCR) was developed and used for a prevalence survey of C. botulinum spores in 190 honey samples. The inhibiting effects of honey on microbial growth and PCR analysis were overcome by using a method of supernatant filtration (SF) in the preparation of the samples before enrichment and PCR. By using this method, an inoculum of 0.1 spore of C. botulinum/g honey could be detected. In the prevalence survey, spores of C. botulinum were detected in 8 (7%) of the 114 Finnish and in 12 (16%) of the 76 imported honey samples. The quantity of spores in PCR-positive samples varied from less than 18 to 140 spores/kg. Neurotoxin gene sequences corresponding to C. botulinum type A were detected in 17 samples and proteolytic type B in 12 samples by PCR analysis. Both types A and B were detected in nine samples. Strains of C. botulinum type A were isolated from 14 and type B from 2 of the 20 PCR-positive samples. This is the first report of type A spores of C. botulinum being detected and isolated in Fennoscandia.
Whitefish eggs were confirmed by pulsed-field gel electrophoresis to cause type E foodborne botul... more Whitefish eggs were confirmed by pulsed-field gel electrophoresis to cause type E foodborne botulism in a 54-yearold patient in Finland. Botulinum neurotoxin and/or nonproteolytic Clostridium botulinum type E organisms were detected in fecal and gastric samples from the patient and in suspected whitefish eggs. Apart from C. botulinum type E, proteolytic type B organisms were detected in the patient's gastric content. This was considered to be insignificant with respect to the clinical disease, suggesting botulinal spores to be occasionally present in the human gastrointestinal tract without any apparent clinical significance. This is the first domestic case of foodborne botulism in Finland.
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