Many bacterial pathogens produce extracellular proteases that are involved in the degradation of ... more Many bacterial pathogens produce extracellular proteases that are involved in the degradation of the host extracellular matrix. Dichelobacter nodosus, which causes ovine footrot, is one such pathogen, Mutagenesis and virulence studies revealed that AprV2, one of three secreted subtilisin-like D. nodosus proteases, is required for virulence. Our work challenges the previous hypothesis that the elastase activity of AprV2 is important for disease progression, since aprV2 mutants were virulent when complemented with a variant with impaired elastase activity. These data reveal that an unusual extended disulphide-tethered loop functions as an exosite that governs the ability of AprV2 to degrade insoluble extracellular matrix components. The disulphide bond and Tyr92, located at the exposed end of the loop, were functionally important. Bioinformatics suggests that other pathogens utilize a similar mechanism, providing a new paradigm for understanding the role of proteases in disease.<br>
Escherichia coli O157 is a human pathogen carried asymptomatically by cattle and shed in their fa... more Escherichia coli O157 is a human pathogen carried asymptomatically by cattle and shed in their faeces. Infection can occur from the consumption of contaminated beef or by direct contact. Large variations of E. coli O157 shedding in cattle exist and vary in the number of cattle positive for E. coli O157 and the amount of bacteria (c.f.u./g faeces) shed by positive animals. To investigate E. coli O157 shedding and super-shedding (>104 c.f.u./g) we used daily sampling over two 8-day periods; in January 2013 (n = 12) and February 2013 (n = 21). Samples were tested by direct faecal culture for enumeration and by immunomagnetic separation to detect lower levels of shedding. We identified three patterns of shedding, similar to previously observed descriptions: intermittent, transient and consistent. The most commonly observed pattern was intermittent shedding and variation in the level of shedding could be large. This extreme variation is demonstrated by a heifer from which E. coli O157...
To investigate public health implications of antibiotics to control post‐weaning scours, we surve... more To investigate public health implications of antibiotics to control post‐weaning scours, we surveyed 22 commercial pig herds in southeastern Australia. Fifty faecal samples per herd were collected from pre‐ and post‐weaned piglets. Presumptive Escherichia coli isolates were confirmed by MALDI‐TOF MS. Isolates (n = 325) were screened for susceptibility to 19 veterinary antibiotics using MIC broth microdilution. All 325 E. coli isolates underwent further testing against 27 antibiotics used in human medicine and were screened for ETEC adhesin and enterotoxin genes (F4 (K88), F5 (K99), F6 (987P), F18, F41, STa, STb, Stx2e and LT) by multiplex PCR. Isolates identified as phenotypically resistant to third‐generation cephalosporin (3GC) and aminoglycoside antibiotics were screened by multiplex PCR/reverse line blot to detect common β‐lactam and aminoglycosides resistance genes, confirmed by sequencing. Twenty (6.1%) of the E. coli isolates were resistant to 3GC antibiotics and 24 (7.4%) to the aminoglycoside antibiotic gentamicin. Genetic analysis revealed six different extended spectrum β‐lactamase (ESBL) genes (blaCTX‐M‐1, ‐14, ‐15, ‐27, blaSHV‐12 and blaCMY‐2‐like genes), four of which have not been previously reported in Australian pigs. Critically, the prevalence of 3GC resistance was higher in non‐pathogenic (non‐ETEC) isolates and those from clinically normal (non‐diarrhoeal) samples. This highlights the importance of non‐ETECE. coli as reservoirs of antimicrobial resistance genes in piglet pens. Antimicrobial resistance surveillance in pig production focused on diagnostic specimens from clinically‐affected animals might be potentially misleading. We recommend that surveillance for emerging antimicrobial resistance such as to 3GC antibiotics should include clinically healthy pigs.
An outbreak of virulent footrot was investigated in a flock of 605 Merino cross-bred sheep in Bhu... more An outbreak of virulent footrot was investigated in a flock of 605 Merino cross-bred sheep in Bhutan. Conventional control methods in the preceding eight years had reduced its prevalence from 36-79% in different components of the flock to about 15% overall. Only one serogroup (B) of Dichelobacter nodosus was identified among 40 isolates cultured from affected sheep. A vaccine prepared from this strain was used in a pilot trial to compare the response of 14 treated and 14 untreated sheep. All affected, vaccinated animals in this trial healed quickly and were protected against re-infection while additional cases developed among untreated sheep during a period favourable for the spread of footrot. The serogroup B vaccine was administered to the whole flock for two successive years. No other footrot treatment was given during these or subsequent years. The whole flock was examined three times, foot by foot, for two years and twice yearly for another two years. When vaccination began the...
The fimbrial subunit (pili) gene that dominates the main protective immunogen was amplified and c... more The fimbrial subunit (pili) gene that dominates the main protective immunogen was amplified and cloned from D. nodosus serotype E by PCR. An expression plasmid was constructed by cloning the pili gene into pME290. The plasmid harbouring the pili sequence was designated pME290-Pili. The pME290-Pili was transformed into the host competent cell PAK/2pfs and the recombinant Pili was expressed and the supernatant of the cultures of the transformant cell PAK/2pfs. The recombinant pili was purified by MgCl_(2) from the supernatant of the culture of the transformed PAK/2pfs. The specific reaction of the recombinant pili and antiserum of D. nodosus serotype E pili was demonstrated by cross electrophoresis. The recombinant pili was expressed at high level in PAK/2pfs and amounted to 11.8% of the total protein of the transformed host cell PAK/2pfs.
Virulent footrot is an economically significant disease in most sheep-rearing countries. The dise... more Virulent footrot is an economically significant disease in most sheep-rearing countries. The disease can be controlled with vaccine targeting the fimbriae of virulent strains of the essential causative agent,However, the bacterium is immunologically heterogeneous, and ten distinct fimbrial serogroups have been identified. Ideally, in each outbreak the infecting strains must be cultured and serogrouped so that the appropriate serogroup-specific mono- or bi-valent vaccine can be administered, because multivalent vaccines lack efficacy due to antigenic competition. If clinical disease expression is suspected to be incomplete, culture-based virulence tests are required to confirm the diagnosis, because control of benign footrot is economically unjustifiable. Both diagnosis and vaccination are conducted at flock-level. The aims of this study were to develop a PCR-based procedure for detecting and serogroupingdirectly from foot swabs, and to determine whether this could be done accurately...
Diarrhoeal diseases in piglets caused by Escherichia coli are responsible for substantial losses ... more Diarrhoeal diseases in piglets caused by Escherichia coli are responsible for substantial losses each year in the Australian pig industry. A cross-sectional survey was conducted (September 2013-May 2014) across 22 commercial pig herds located in southeastern Australia: NSW (n = 9); VIC (n = 10); and SA (n = 3), to estimate the prevalence of E. coli associated diarrhoea in pre- and post-weaned piglets and to identify key risk factors associated with E. coli disease. A questionnaire on management and husbandry practices was included. Faecal samples (n = 50 from each herd) were tested for the presence of β-haemolytic E. coli. Species level identification was confirmed by matrix-assisted laser desorption / ionisation time-of-flight mass spectrometry (MALDI-TOF/MS). ETEC virulence and enterotoxin genes (F4, F5, F6, F18, F41, STa, STb and LT) were screened for by multiplex PCR. This study assessed 60 potential risk factors for E. coli disease in post-weaned piglets, with 2 key factors-rec...
Dichelobacter nodosus is a fastidious, strictly anaerobic bacterium, an obligate parasite of the ... more Dichelobacter nodosus is a fastidious, strictly anaerobic bacterium, an obligate parasite of the ruminant hoof, and the essential causative agent of virulent ovine footrot. The clinical disease results from a complex interplay between the pathogen, the environment, and the host. Sheep flocks diagnosed with virulent but not benign footrot in Australia may be quarantined and required to undergo a compulsory eradication program, with costs met by the farmer. Virulence of D. nodosus at least partially depends on the elaboration of a protease encoded by aprV2 and manifests as elastase activity. Laboratory virulence tests are used to assist diagnosis because clinical differentiation of virulent and benign footrot can be challenging during the early stages of disease or when the disease is not fully expressed due to unfavorable pasture conditions. Using samples collected from foot lesions from 960 sheep from 40 flocks in four different geographic regions, we evaluated the analytical charac...
... Ellis, Dr. Bill Blackwell, Colin Cavanagh, Suresh Khatkar, Jiri Tasler, Juha Virolainen, Quyn... more ... Ellis, Dr. Bill Blackwell, Colin Cavanagh, Suresh Khatkar, Jiri Tasler, Juha Virolainen, Quynh Vu, Helen McGregor, Sandra Saville, Eileen Risby, Ron Henderson, David Palmer, Mathew VanDjick, Marie Wildrige for their friendship and ...
As part of an outbreak-specific footrot vaccination field trial a total of 1282 footrot lesion sa... more As part of an outbreak-specific footrot vaccination field trial a total of 1282 footrot lesion samples were collected from 2 sheep flocks on King Island, Tasmania. Breeding rams were shared between the two flocks, suggesting a common source of infection. All samples were tested for Dichelobacter nodosus. A total of 1047 D. nodosus isolates were obtained in pure culture (490 from 670 lesion samples from flock 1, and 557 from 612 lesion samples from flock 2) were tested by agglutination and PCR tests for the 9 common Australian serogroups A to I. After the first rounds of a specific vaccination program, a significant proportion of the isolates of D. nodosus from these flocks were found to be negative in the serogrouping tests and the prevalence of the disease remained high in both. Those isolates were tested retrospectively against New Zealand and Nepal serogroup M antisera and found to be positive. Fimbrial gene (fimA) sequences of three isolates collected over three years were ident...
SUMMARY We compared the use of recto-anal mucosal swab (RAMS) culture and faecal culture for the ... more SUMMARY We compared the use of recto-anal mucosal swab (RAMS) culture and faecal culture for the detection of E. coli O157 in a mob of Merino sheep. Fifty Merino wethers and maiden ewes housed in indoor pens were sampled on five occasions. We detected E coli O157 in 32% (16/50) of sheep, with weekly prevalence ranging from 4% (2/50) to 16% (8/50). Overall, 12·5% (2/16) were detected by RAMS culture only, and 37·5% (6/16) were detected by faecal culture only. The level of agreement between the two sampling methods was moderate [kappa statistic = 0·583, 95% confidence interval (CI) 0·460-0·707]. The relative sensitivities of RAMS and faecal culture were 67% (95% CI 41-86) and 57% (95% CI 34-77), respectively. We identified four super-shedding sheep using direct faecal culture. Although the majority of culture-positive sheep were detected at one sampling point only, 3/4 super-shedding sheep were culture-positive at two sampling points, and 1/4 was culture-positive at four sampling poin...
Ovine footrot is a major disease affecting sheep welfare and production. The anaerobic Gram-negat... more Ovine footrot is a major disease affecting sheep welfare and production. The anaerobic Gram-negative bacterium Dichelobacter nodosus is the essential transmitting agent. Monovalent or bivalent vaccines induce high levels of D. nodosus antibodies and are the basis of several successful footrot treatment, control and eradication programs. Due to the rapid rate of disease transmission within a flock, the presence of therapeutic vaccination non-responders has major implications for a control program. The aim of this study was to assess the immunological basis of a therapeutic vaccination non-response. Sheep (n=120) were infected with D. nodosus in an artificial pen challenge. Once disease had established, animals were vaccinated with a serogroup specific D. nodosus fimbrial vaccine. Based on the response to therapeutic vaccination, animals were allocated into one of three groups: (i) TVNR where disease persisted despite vaccination (ii) non-diseased, where disease never established and ...
Virulent footrot is a significant disease of sheep in most sheep farming countries; a strain/sero... more Virulent footrot is a significant disease of sheep in most sheep farming countries; a strain/serogroup of the anaerobic bacterium Dichelobacter nodosus is the essential transmitting agent. Commercial multivalent footrot vaccines containing nine fimbrial serogroups (A through I) of D. nodosus produce relatively low and short term antibody responses due to antigenic competition, in contrast to higher and longer responses provided by monovalent or bivalent vaccines. The latter were important components of successful eradication programs for endemic footrot caused by either one or two serogroups of D. nodosus in Nepal, Bhutan, and several flocks in Australia. However, the presence of up to six serogroups in some Australian flocks and the use of an annual bivalent vaccination regime to progressively eradicate serogroups would require a long term program. In this study we report the results of a sequential vaccination trial testing different time intervals between different bivalent vaccinations. Intervals of 12, 9, 6, 3 and 0 months were tested. The 1st vaccination was with recombinant fimbrial antigens for serogroups A and B while the 2nd vaccination was with D and E. There were no significant differences between the antibody responses for time intervals of 3, 6, 9 and 12 months whereas there was a reduced response when sheep were vaccinated with two bivalent vaccines (four antigens) concurrently, indicating antigenic competition. Therefore an inter-vaccination interval of 3 months can be applied between two different bivalent vaccines without detrimental impact on the humoral immune responses to the various fimbrial antigens of D. nodosus. These results could have wider applications in vaccination against diseases caused by multivalent or multistrain microbes.
Dichelobacter nodosus is the essential causative agent of footrot in sheep. The type IV fimbriae ... more Dichelobacter nodosus is the essential causative agent of footrot in sheep. The type IV fimbriae of D. nodosus are required for virulence, are highly immunogenic and immunoprotective, and can be divided into 10 major serogroups. Fimbrial variation has been postulated to have arisen because of genetic recombination within the fimbrial gene region perhaps as a means of evading the immune response invoked by infection. To show that antigenic variation in these fimbriae could occur after natural transformation and subsequent homologous recombination, a suicide plasmid containing the fimbrial subunit gene, fimA, of a serogroup G strain was used to convert a serogroup I strain to serogroup G. The resultant mutants were shown by Western blotting and slide agglutination to produce serogroup G fimbriae, but by two independent methods to still have the genotype of the parent type I strain. These data have significant implications for the use of fimbrial vaccines for the control of ovine footrot and suggest that benign strains of D. nodosus could play an important role as a reservoir of alternative fimbrial antigens.
Many bacterial pathogens produce extracellular proteases that are involved in the degradation of ... more Many bacterial pathogens produce extracellular proteases that are involved in the degradation of the host extracellular matrix. Dichelobacter nodosus, which causes ovine footrot, is one such pathogen, Mutagenesis and virulence studies revealed that AprV2, one of three secreted subtilisin-like D. nodosus proteases, is required for virulence. Our work challenges the previous hypothesis that the elastase activity of AprV2 is important for disease progression, since aprV2 mutants were virulent when complemented with a variant with impaired elastase activity. These data reveal that an unusual extended disulphide-tethered loop functions as an exosite that governs the ability of AprV2 to degrade insoluble extracellular matrix components. The disulphide bond and Tyr92, located at the exposed end of the loop, were functionally important. Bioinformatics suggests that other pathogens utilize a similar mechanism, providing a new paradigm for understanding the role of proteases in disease.<br>
Escherichia coli O157 is a human pathogen carried asymptomatically by cattle and shed in their fa... more Escherichia coli O157 is a human pathogen carried asymptomatically by cattle and shed in their faeces. Infection can occur from the consumption of contaminated beef or by direct contact. Large variations of E. coli O157 shedding in cattle exist and vary in the number of cattle positive for E. coli O157 and the amount of bacteria (c.f.u./g faeces) shed by positive animals. To investigate E. coli O157 shedding and super-shedding (>104 c.f.u./g) we used daily sampling over two 8-day periods; in January 2013 (n = 12) and February 2013 (n = 21). Samples were tested by direct faecal culture for enumeration and by immunomagnetic separation to detect lower levels of shedding. We identified three patterns of shedding, similar to previously observed descriptions: intermittent, transient and consistent. The most commonly observed pattern was intermittent shedding and variation in the level of shedding could be large. This extreme variation is demonstrated by a heifer from which E. coli O157...
To investigate public health implications of antibiotics to control post‐weaning scours, we surve... more To investigate public health implications of antibiotics to control post‐weaning scours, we surveyed 22 commercial pig herds in southeastern Australia. Fifty faecal samples per herd were collected from pre‐ and post‐weaned piglets. Presumptive Escherichia coli isolates were confirmed by MALDI‐TOF MS. Isolates (n = 325) were screened for susceptibility to 19 veterinary antibiotics using MIC broth microdilution. All 325 E. coli isolates underwent further testing against 27 antibiotics used in human medicine and were screened for ETEC adhesin and enterotoxin genes (F4 (K88), F5 (K99), F6 (987P), F18, F41, STa, STb, Stx2e and LT) by multiplex PCR. Isolates identified as phenotypically resistant to third‐generation cephalosporin (3GC) and aminoglycoside antibiotics were screened by multiplex PCR/reverse line blot to detect common β‐lactam and aminoglycosides resistance genes, confirmed by sequencing. Twenty (6.1%) of the E. coli isolates were resistant to 3GC antibiotics and 24 (7.4%) to the aminoglycoside antibiotic gentamicin. Genetic analysis revealed six different extended spectrum β‐lactamase (ESBL) genes (blaCTX‐M‐1, ‐14, ‐15, ‐27, blaSHV‐12 and blaCMY‐2‐like genes), four of which have not been previously reported in Australian pigs. Critically, the prevalence of 3GC resistance was higher in non‐pathogenic (non‐ETEC) isolates and those from clinically normal (non‐diarrhoeal) samples. This highlights the importance of non‐ETECE. coli as reservoirs of antimicrobial resistance genes in piglet pens. Antimicrobial resistance surveillance in pig production focused on diagnostic specimens from clinically‐affected animals might be potentially misleading. We recommend that surveillance for emerging antimicrobial resistance such as to 3GC antibiotics should include clinically healthy pigs.
An outbreak of virulent footrot was investigated in a flock of 605 Merino cross-bred sheep in Bhu... more An outbreak of virulent footrot was investigated in a flock of 605 Merino cross-bred sheep in Bhutan. Conventional control methods in the preceding eight years had reduced its prevalence from 36-79% in different components of the flock to about 15% overall. Only one serogroup (B) of Dichelobacter nodosus was identified among 40 isolates cultured from affected sheep. A vaccine prepared from this strain was used in a pilot trial to compare the response of 14 treated and 14 untreated sheep. All affected, vaccinated animals in this trial healed quickly and were protected against re-infection while additional cases developed among untreated sheep during a period favourable for the spread of footrot. The serogroup B vaccine was administered to the whole flock for two successive years. No other footrot treatment was given during these or subsequent years. The whole flock was examined three times, foot by foot, for two years and twice yearly for another two years. When vaccination began the...
The fimbrial subunit (pili) gene that dominates the main protective immunogen was amplified and c... more The fimbrial subunit (pili) gene that dominates the main protective immunogen was amplified and cloned from D. nodosus serotype E by PCR. An expression plasmid was constructed by cloning the pili gene into pME290. The plasmid harbouring the pili sequence was designated pME290-Pili. The pME290-Pili was transformed into the host competent cell PAK/2pfs and the recombinant Pili was expressed and the supernatant of the cultures of the transformant cell PAK/2pfs. The recombinant pili was purified by MgCl_(2) from the supernatant of the culture of the transformed PAK/2pfs. The specific reaction of the recombinant pili and antiserum of D. nodosus serotype E pili was demonstrated by cross electrophoresis. The recombinant pili was expressed at high level in PAK/2pfs and amounted to 11.8% of the total protein of the transformed host cell PAK/2pfs.
Virulent footrot is an economically significant disease in most sheep-rearing countries. The dise... more Virulent footrot is an economically significant disease in most sheep-rearing countries. The disease can be controlled with vaccine targeting the fimbriae of virulent strains of the essential causative agent,However, the bacterium is immunologically heterogeneous, and ten distinct fimbrial serogroups have been identified. Ideally, in each outbreak the infecting strains must be cultured and serogrouped so that the appropriate serogroup-specific mono- or bi-valent vaccine can be administered, because multivalent vaccines lack efficacy due to antigenic competition. If clinical disease expression is suspected to be incomplete, culture-based virulence tests are required to confirm the diagnosis, because control of benign footrot is economically unjustifiable. Both diagnosis and vaccination are conducted at flock-level. The aims of this study were to develop a PCR-based procedure for detecting and serogroupingdirectly from foot swabs, and to determine whether this could be done accurately...
Diarrhoeal diseases in piglets caused by Escherichia coli are responsible for substantial losses ... more Diarrhoeal diseases in piglets caused by Escherichia coli are responsible for substantial losses each year in the Australian pig industry. A cross-sectional survey was conducted (September 2013-May 2014) across 22 commercial pig herds located in southeastern Australia: NSW (n = 9); VIC (n = 10); and SA (n = 3), to estimate the prevalence of E. coli associated diarrhoea in pre- and post-weaned piglets and to identify key risk factors associated with E. coli disease. A questionnaire on management and husbandry practices was included. Faecal samples (n = 50 from each herd) were tested for the presence of β-haemolytic E. coli. Species level identification was confirmed by matrix-assisted laser desorption / ionisation time-of-flight mass spectrometry (MALDI-TOF/MS). ETEC virulence and enterotoxin genes (F4, F5, F6, F18, F41, STa, STb and LT) were screened for by multiplex PCR. This study assessed 60 potential risk factors for E. coli disease in post-weaned piglets, with 2 key factors-rec...
Dichelobacter nodosus is a fastidious, strictly anaerobic bacterium, an obligate parasite of the ... more Dichelobacter nodosus is a fastidious, strictly anaerobic bacterium, an obligate parasite of the ruminant hoof, and the essential causative agent of virulent ovine footrot. The clinical disease results from a complex interplay between the pathogen, the environment, and the host. Sheep flocks diagnosed with virulent but not benign footrot in Australia may be quarantined and required to undergo a compulsory eradication program, with costs met by the farmer. Virulence of D. nodosus at least partially depends on the elaboration of a protease encoded by aprV2 and manifests as elastase activity. Laboratory virulence tests are used to assist diagnosis because clinical differentiation of virulent and benign footrot can be challenging during the early stages of disease or when the disease is not fully expressed due to unfavorable pasture conditions. Using samples collected from foot lesions from 960 sheep from 40 flocks in four different geographic regions, we evaluated the analytical charac...
... Ellis, Dr. Bill Blackwell, Colin Cavanagh, Suresh Khatkar, Jiri Tasler, Juha Virolainen, Quyn... more ... Ellis, Dr. Bill Blackwell, Colin Cavanagh, Suresh Khatkar, Jiri Tasler, Juha Virolainen, Quynh Vu, Helen McGregor, Sandra Saville, Eileen Risby, Ron Henderson, David Palmer, Mathew VanDjick, Marie Wildrige for their friendship and ...
As part of an outbreak-specific footrot vaccination field trial a total of 1282 footrot lesion sa... more As part of an outbreak-specific footrot vaccination field trial a total of 1282 footrot lesion samples were collected from 2 sheep flocks on King Island, Tasmania. Breeding rams were shared between the two flocks, suggesting a common source of infection. All samples were tested for Dichelobacter nodosus. A total of 1047 D. nodosus isolates were obtained in pure culture (490 from 670 lesion samples from flock 1, and 557 from 612 lesion samples from flock 2) were tested by agglutination and PCR tests for the 9 common Australian serogroups A to I. After the first rounds of a specific vaccination program, a significant proportion of the isolates of D. nodosus from these flocks were found to be negative in the serogrouping tests and the prevalence of the disease remained high in both. Those isolates were tested retrospectively against New Zealand and Nepal serogroup M antisera and found to be positive. Fimbrial gene (fimA) sequences of three isolates collected over three years were ident...
SUMMARY We compared the use of recto-anal mucosal swab (RAMS) culture and faecal culture for the ... more SUMMARY We compared the use of recto-anal mucosal swab (RAMS) culture and faecal culture for the detection of E. coli O157 in a mob of Merino sheep. Fifty Merino wethers and maiden ewes housed in indoor pens were sampled on five occasions. We detected E coli O157 in 32% (16/50) of sheep, with weekly prevalence ranging from 4% (2/50) to 16% (8/50). Overall, 12·5% (2/16) were detected by RAMS culture only, and 37·5% (6/16) were detected by faecal culture only. The level of agreement between the two sampling methods was moderate [kappa statistic = 0·583, 95% confidence interval (CI) 0·460-0·707]. The relative sensitivities of RAMS and faecal culture were 67% (95% CI 41-86) and 57% (95% CI 34-77), respectively. We identified four super-shedding sheep using direct faecal culture. Although the majority of culture-positive sheep were detected at one sampling point only, 3/4 super-shedding sheep were culture-positive at two sampling points, and 1/4 was culture-positive at four sampling poin...
Ovine footrot is a major disease affecting sheep welfare and production. The anaerobic Gram-negat... more Ovine footrot is a major disease affecting sheep welfare and production. The anaerobic Gram-negative bacterium Dichelobacter nodosus is the essential transmitting agent. Monovalent or bivalent vaccines induce high levels of D. nodosus antibodies and are the basis of several successful footrot treatment, control and eradication programs. Due to the rapid rate of disease transmission within a flock, the presence of therapeutic vaccination non-responders has major implications for a control program. The aim of this study was to assess the immunological basis of a therapeutic vaccination non-response. Sheep (n=120) were infected with D. nodosus in an artificial pen challenge. Once disease had established, animals were vaccinated with a serogroup specific D. nodosus fimbrial vaccine. Based on the response to therapeutic vaccination, animals were allocated into one of three groups: (i) TVNR where disease persisted despite vaccination (ii) non-diseased, where disease never established and ...
Virulent footrot is a significant disease of sheep in most sheep farming countries; a strain/sero... more Virulent footrot is a significant disease of sheep in most sheep farming countries; a strain/serogroup of the anaerobic bacterium Dichelobacter nodosus is the essential transmitting agent. Commercial multivalent footrot vaccines containing nine fimbrial serogroups (A through I) of D. nodosus produce relatively low and short term antibody responses due to antigenic competition, in contrast to higher and longer responses provided by monovalent or bivalent vaccines. The latter were important components of successful eradication programs for endemic footrot caused by either one or two serogroups of D. nodosus in Nepal, Bhutan, and several flocks in Australia. However, the presence of up to six serogroups in some Australian flocks and the use of an annual bivalent vaccination regime to progressively eradicate serogroups would require a long term program. In this study we report the results of a sequential vaccination trial testing different time intervals between different bivalent vaccinations. Intervals of 12, 9, 6, 3 and 0 months were tested. The 1st vaccination was with recombinant fimbrial antigens for serogroups A and B while the 2nd vaccination was with D and E. There were no significant differences between the antibody responses for time intervals of 3, 6, 9 and 12 months whereas there was a reduced response when sheep were vaccinated with two bivalent vaccines (four antigens) concurrently, indicating antigenic competition. Therefore an inter-vaccination interval of 3 months can be applied between two different bivalent vaccines without detrimental impact on the humoral immune responses to the various fimbrial antigens of D. nodosus. These results could have wider applications in vaccination against diseases caused by multivalent or multistrain microbes.
Dichelobacter nodosus is the essential causative agent of footrot in sheep. The type IV fimbriae ... more Dichelobacter nodosus is the essential causative agent of footrot in sheep. The type IV fimbriae of D. nodosus are required for virulence, are highly immunogenic and immunoprotective, and can be divided into 10 major serogroups. Fimbrial variation has been postulated to have arisen because of genetic recombination within the fimbrial gene region perhaps as a means of evading the immune response invoked by infection. To show that antigenic variation in these fimbriae could occur after natural transformation and subsequent homologous recombination, a suicide plasmid containing the fimbrial subunit gene, fimA, of a serogroup G strain was used to convert a serogroup I strain to serogroup G. The resultant mutants were shown by Western blotting and slide agglutination to produce serogroup G fimbriae, but by two independent methods to still have the genotype of the parent type I strain. These data have significant implications for the use of fimbrial vaccines for the control of ovine footrot and suggest that benign strains of D. nodosus could play an important role as a reservoir of alternative fimbrial antigens.
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