An outbreak of Cyclospora infection on a cruise ship.

In 2010, an outbreak of cyclosporiasis affected passengers and crew on two successive voyages of a cruise ship that departed from and returned to Fremantle, Australia. There were 73 laboratory-confirmed and 241 suspected cases of Cyclospora infection reported in passengers and crew from the combined cruises. A case-control study performed in crew members found that illness was associated with eating items of fresh produce served onboard the ship, but the study was unable conclusively to identify the responsible food(s). It is likely that one or more of the fresh produce items taken onboard at a south-east Asian port during the first cruise was contaminated. If fresh produce supplied to cruise ships is sourced from countries or regions where Cyclospora is endemic, robust standards of food production and hygiene should be applied to the supply chain.

Population genetic analysis of Serpulina pilosicoli and its molecular epidemiology in villages in the eastern Highlands of Papua New Guinea.

The population genetics of Serpulina pilosicoli and its molecular epidemiology in villages in the Eastern Highlands province of Papua New Guinea were investigated. Multilocus enzyme electrophoresis (MLEE) was used to analyse 164 isolates from humans and animals. These were divided into 33 electrophoretic types (ETs), four of which contained 65% of the isolates. The mean genetic diversity (n = number of ETs) for 145 human isolates was 0.18, and the mean number of alleles at five polymorphic loci was 2.6. The species appeared to be recombinant, as there was a lack of linkage disequilibrium, and 25% of all the possible combinations of alleles was present in the population. PFGE analysis using the enzymes M/ul and Sa/l divided 157 of the isolates into 99 PFGE types, demonstrating the existence of considerable strain diversity in a geographically restricted area. The two techniques were in excellent agreement; however, PFGE was more discriminatory for strain typing than was MLEE. Nine out of 19 (47.4%) culture-positive individuals were colonized by the same PFGE type of S. pilosicoli when retested after 6 weeks. For three individuals, the PFGE profiles of the second isolate differed from the first in only one or two DNA bands, while the other seven individuals were colonized with distinct PFGE types on each occasion. In two cases, strains with the same PFGE pattern were isolated from humans and dogs, suggesting that cross-species transmission of S. pilosicoli may occur naturally and that the infection can be zoonotic.

Development and evaluation of polymerase chain reaction tests as an aid to diagnosis of swine dysentery and intestinal spirochaetosis.

Polymerase chain reaction (PCR) tests were established for detection of Serpulina hyodysenteriae, the agent of swine dysentery, and S. pilosicoli, the agent of intestinal spirochaetosis. Both reactions were specific when tested with DNA from 107 strains of various intestinal spirochaetes. For diagnostic use, faeces were plated to selective medium, and diatomaceous earth extraction used to obtain DNA prior to PCR. This procedure detected 10(3)-10(4) cells of either organism seeded into 0.2 g of faeces. When applied to 63 samples from pigs of eight piggeries naturally infected with either S. hyodysenteriae or S. pilosicoli, both PCRs were specific, more rapid, and detected more positive samples than did routine faecal culture and isolation.

The prevalence of Serpulina pilosicoli in humans and domestic animals in the Eastern Highlands of Papua New Guinea.

In a survey of five villages in the Eastern Highlands of Papua New Guinea, Serpulina pilosicoli was isolated from rectal swabs from 113 of 496 individuals (22.8%). Colonization rates ranged from 22.6-30.1% in four of the villages but was only 8.6% in the other village. In comparison colonization was demonstrated in only 5 of 54 indigenous people (9.3%) and none of 76 non-indigenous people living in an urban environment in the same region. Colonization did not relate to reported occurrence of diarrhoea, age, sex, or length of time resident in a village. A second set of 94 faecal specimens was collected from 1 village 6 weeks after the first set. S. pilosicoli was isolated from 27 of 29 individuals (93.1%) who were positive on the first sampling and from 7 of 65 individuals (10.8%) who previously were negative. In this case, isolates were significantly more common in watery stools than in normal stools. The annual incidence of infection in the village was calculated as 93.6%, with an average duration of infection of 117 days. S. pilosicoli could not be isolated from any village pig (n = 126) despite its confirmed presence in 17 of 50 commercial pigs (34.0%) sampled at a local piggery. Four of 76 village dogs (5.3%) and 1 of 2 village ducks were colonized with S. pilosicoli, suggesting the possibility of cross transmission between humans and animals.

Molecular analysis of isolates of Salmonella typhi obtained from patients with fatal and nonfatal typhoid fever.

Molecular characterization of a total of 52 human isolates of Salmonella typhi from Papua New Guinea was performed by using pulsed-field gel electrophoresis (PFGE) after digestion of chromosomal DNA with three restriction endonucleases, XbaI (5'-TCTAGA-3'), AvrII (5'-CCTAGG-3'), and SpeI (5'-ACTAGT-3'). Of the 52 isolates tested, 11 were obtained from patients with fatal typhoid fever and 41 were obtained from patients with nonfatal disease. The 52 isolates showed limited genetic diversity as evidenced by only three different PFGE patterns detected following digestion with XbaI (patterns X1 to X3; F [coefficient of similarity] = 0.86 to 1.0), four patterns detected following digestion with AvrII (patterns A1 to A4; F =0.78 to 1.0), and two patterns detected following digestion with SpeI (patterns S1 and S2; F = 0.97 to 1.0). Of the 52 isolates, 37 were phage typed, and all belonged to phage type D2. All 11 isolates obtained from patients with fatal typhoid fever were identical (F = 1.0) and possessed the PFGE pattern combination X1S1A1, whereas the 41 isolates from patients with nonfatal typhoid fever had various PFGE pattern combinations, the most common being X2S1A2 (39%), X1S1A1 (24%), and X1S1A2 (15%). Thus, all the isolates from patients with the fatal disease had the X1 and A1 patterns, whereas the majority of the isolates from patients with nonfatal typhoid fever possessed the X2 and A2 patterns. The data suggest that there is an association among strains of S. typhi between genotype, as assessed by PFGE patterns, and the capability to cause fatal illness. Analysis of blood and fecal isolates of S. typhi from the same patient also indicated that some genetic changes occur in vivo during the course of infection.

Genetic relationships between isolates of Serpulina (Treponema) hyodysenteriae, and comparison of methods for their subspecific differentiation.

Multilocus enzyme electrophoresis (MEE) was used to examine the extent of genetic diversity amongst 98 isolates of Serpulina (Treponema) hyodysenteriae. The species contained four major genetic divisions (A, B, C and D) and 29 electrophoretic types (ETs). Division D was relatively distinct, being separated from the other three divisions by fixed allelic differences at an average of 6.6 of 15 enzyme loci. Electrophoretic differences were compared with results of DNA restriction endonuclease analysis (REA) and serological typing of the isolates. Most isolates with the same or similar REA banding patterns shared the same or similar ETs. This demonstrated that both techniques could be used as sensitive and specific methods of identifying closely related isolates. However, using MEE analysis, some isolates that had quite different REA patterns were found to be genetically closely related. Therefore ET designations had an advantage over REA patterns in that they were readily quantifiable as a means of estimating genetic relatedness between isolates. Most isolates that were genetically similar to each other were of the same serological group, but some antigenic types were widely distributed across the genetic divisions.

Typing of Australian isolates of Treponema hyodysenteriae by serology and by DNA restriction endonuclease analysis.

A total of 91 isolates of Treponema hyodysenteriae which were obtained from 62 piggeries located around Australia were typed by serology and by DNA restriction endonuclease analysis (REA). The isolates fell into eight serogroups, of which groups B and D were the most common. Isolates with different REA patterns were recognised within serogroups, whilst a few isolates with the same REA pattern were placed into different serogroups. Some of the latter isolates were either from the same piggery or from farms with epidemiological links, thus indicating the bacteria may have altered their antigenic properties. A total of 31 different REA patterns were recognised amongst the Australian isolates. These comprised eight major patterns, with four of these being subdivided on the basis of minor differences in banding. Where a number of isolates were obtained from individual piggeries these all had the same REA pattern, and in one piggery isolates with the same pattern were recovered over a five year period. Plasmid bands were observed in 70 of the Australian isolates (77%), and in six of the seven overseas isolates included in the study for comparison. These plasmids did not affect the REA pattern. Of the States from which substantial numbers of isolates were examined, the greatest number of different strains (12 amongst 19 piggeries) were found from Victoria, and there were 10 REA patterns in strains from 24 piggeries in Queensland. Despite the large total number of different strains of T. hyodysenteriae in Australia, only three were found in more than one State.

Use of a whole chromosomal probe for identification of Treponema hyodysenteriae.

A whole chromosomal DNA probe labelled with photobiotin was used in a dot blot hybridisation to identify DNA from isolates of Treponema hyodysenteriae, the aetiological agent of swine dysentery. The probe was evaluated using DNA from 13 isolates of T hyodysenteriae and 13 isolates of non-T hyodysenteriae spirochaetes recovered from pigs. The initial test had both a sensitivity and specificity of 92.3 per cent, although when it was repeated the specificity fell to 84.6 per cent. The test was helpful in distinguishing between T hyodysenteriae and other morphologically similar treponemes that are part of the normal flora in the large intestine of pigs. The probe could also be used to detect as little as 10 ng of purified DNA from T hyodysenteriae, or DNA from 2 x 10(6) bacterial cells lysed directly onto nitrocellulose.

Serological grouping of Treponema hyodysenteriae.

Two Australian isolates of Treponema hyodysenteriae which did not fit within the current serological grouping system for these bacteria were examined by agarose gel double immunodiffusion tests (AGDP). Isolate Vic1 was serologically unique, and we propose that it becomes the type organism for a new sixth serological group of T. hyodysenteriae (Group F). Isolate Q1 was unusual in that lipopolysaccharide (LPS) extracted from it reacted strongly in AGDP with serum raised against the type organism for serogroup D (A1), and also weakly with serum raised against the type organism for serogroup B (WA1). The nature of this cross-reactivity was examined by using cross-absorbed antisera in AGDP, and by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis. The pattern of serological cross-reactivity between Q1, A1 and WA1 was complex and was not fully defined, but the isolate Q1 apparently shared low molecular weight 'serogroup' LPS antigens with A1, and shared higher molecular weight LPS antigens with WA1. On this basis Q1 was designated as belonging to serogroup D, although it was recommended that this be qualified as D (B) to indicate the presence of weak cross-reactivity with serogroup B. Such serological cross-reactivity may have significance in relation to the development of immunity to T.hyodysenteriae. Isolate Q1 may be a potentially useful organism for vaccine development because of its ability to induce a good serological response to LPS of treponemes from both serogroups D and B.