[Immunohistochemical diagnosis of colonic spirochetosis with anti-treponema antibody in patients consulting for chronic diarrhea. Results of a prospective study conducted in 137 patients]. / Diagnostic immunohistochimique de la spirochétose colique avec l'anticorps anti-tréponème chez les patients consultant pour une diarrhée chronique. Résultats d'une étude prospective menée chez 137 patients.

AIM: To assess the incidence of colonic spirochetosis, diagnosed by immunohistochemical stain with anti-Treponema pallidum antibody, in a prospective study of colonic biopsies of patients presenting with chronic diarrhea. MATERIAL AND METHODS: From March 2017 to March 2018 the colonic biopsies of patients presenting with chronic diarrhea were stained with Hematoxylin Eosin and anti-Treponema pallidum antibody. The positive cases were also stained with Steiner stain. RESULTS: A total of 137 colonic biopsies were assessed and 3 cases were positive for immunohistochemical stain with anti-Treponema pallidum antibody (2% of the patients). One case was easy to diagnose with HE stain but the 2 other cases were not. The bacteria were stained with Steiner stain, but less easily seen than with the immunohistochemical stain. No patient was treated with antibiotics. DISCUSSION AND CONCLUSION: The colonic spirochetosis can be easily diagnosed by pathologists with immunohistochemical stain with anti-Treponema pallidum antibody. The bacteria are more easily diagnosed with immunohistochemical stain than with HE stain or Steiner stain. However, colonic spirochetosis is rarely diagnosed on colonic biopsies of patients presenting with chronic diarrhea (2% of the patients in our study). Due to the rarity of the entity, and the cost of immunohistochemical stain and the weak benefit for the patient (no patient in our study was treated with antibiotics for colonic spirochetosis) we cannot advise to perform systematic immunohistochemical stain with anti-Treponema pallidum antibody in all the colonic biopsies of patients presenting with chronic diarrhea.

Human intestinal spirochetosis in Japanese patients aged less than 20 years: Histological analysis of colorectal biopsy and surgical specimens obtained from 479 patients.

Human intestinal spirochetosis (HIS) is a condition in which spirochetes attach to and colonize the colorectal epithelium. To our knowledge, no comprehensive studies of HIS in young patient have been published in a developed country. This study aimed to determine the incidence and clinicopathological manifestations of HIS in Japanese patients aged less than 20 years. We retrospectively reviewed 3605 biopsy and 92 surgical specimens obtained from 479 patients admitted to Shinshu University Hospital between 1997 and 2014. All slides were reviewed independently by two pathologists to confirm the histological presence of spirochetes. Among 387 patients who underwent biopsy, the most common pathologic diagnosis was ulcerative colitis (12.6%, n = 49). Additionally, about half of the biopsy specimens showed non-specific, mildly inflamed mucosa (50.6%, n = 196); only one of these cases was HIS. On the other hand, among the surgical specimens, we found no cases of HIS. We concluded that the incidence of HIS in Japanese young patients was 0.2% (1/479 cases). The incidence of HIS in Japanese young patients was very low, and one HIS case was associated with colitis with abdominal pain.

Occurrence of blood parasites in seabirds admitted for rehabilitation in the Western Cape, South Africa, 2001-2013.

Blood parasites are generally uncommon in seabirds, and knowledge on their epidemiology is further limited by the fact that they often inhabit remote locations that are logistically difficult or expensive to study. We present a long term data set of blood smear examinations of 1909 seabirds belonging to 27 species that were admitted to a rehabilitation centre in Cape Town (Western Cape, South Africa) between 2001 and 2013. Blood parasites were detected in 59% of species (16/27) and 29% of individuals examined (551/1909). The following blood parasites were recorded: Babesia ugwidiensis, Babesia peircei, Babesia sp., Plasmodium sp., Leucocytozoon ugwidi, Hepatozoon albatrossi, Haemoproteus skuae and Spirochaetales. Several of the records are novel host-parasite associations, demonstrating the potential of rehabilitation centres for parasite and disease surveillance, particularly for species infrequently sampled from which no host-specific parasites have been described.

Clinicopathologic study of intestinal spirochetosis in Japan with special reference to human immunodeficiency virus infection status and species types: analysis of 5265 consecutive colorectal biopsies.

BACKGROUND: Previous studies reported that the incidence of intestinal spirochetosis was high in homosexual men, especially those with Human Immunodeficiency Virus infection. The aim of the present study was to clarify the clinicopathological features of intestinal spirochetosis in Japan with special reference to Human Immunodeficiency Virus infection status and species types. METHODS: A pathology database search for intestinal spirochetosis was performed at Tokyo Metropolitan Cancer and Infectious Disease Center Komagome Hospital between January 2008 and October 2011, and included 5265 consecutive colorectal biopsies from 4254 patients. After patient identification, a retrospective review of endoscopic records and clinical information was performed. All pathology slides were reviewed by two pathologists. The length of the spirochetes was measured using a digital microscope. Causative species were identified by polymerase chain reaction. RESULTS: Intestinal spirochetosis was diagnosed in 3 out of 55 Human Immunodeficiency Virus-positive patients (5.5%). The mean length of intestinal spirochetes was 8.5 µm (range 7-11). Brachyspira pilosicoli was detected by polymerase chain reaction in all 3 patients. Intestinal spirochetosis was also diagnosed in 73 out of 4199 Human Immunodeficiency Virus-negative patients (1.7%). The mean length of intestinal spirochetes was 3.5 µm (range 2-8). The species of intestinal spirochetosis was identified by polymerase chain reaction in 31 Human Immunodeficiency Virus-negative patients. Brachyspira aalborgi was detected in 24 cases (78%) and Brachyspira pilosicoli in 6 cases (19%). Both Brachyspira aalborgi and Brachyspira pilosicoli were detected in only one Human Immunodeficiency Virus-negative patient (3%). The mean length of Brachyspira aalborgi was 3.8 µm, while that of Brachyspira pilosicoli was 5.5 µm. The length of Brachyspira pilosicoli was significantly longer than that of Brachyspira aalborgi (p < 0.01). The lengths of intestinal spirochetes were significantly longer in Human Immunodeficiency Virus-positive patients than in Human Immunodeficiency Virus-negative patients (p < 0.05). CONCLUSIONS: The incidence of intestinal spirochetosis was slightly higher in Human Immunodeficiency Virus-positive patients than in Human Immunodeficiency Virus-negative patients. However, no relationship was found between the Human Immunodeficiency Virus status and intestinal spirochetosis in Japan. Brachyspira pilosicoli infection may be more common in Human Immunodeficiency Virus-positive patients with intestinal spirochetosis than in Human Immunodeficiency Virus-negative patients with intestinal spirochetosis.

Human intestinal spirochetosis is significantly associated with sessile serrated adenomas/polyps.

It remains unclear whether or not human intestinal spirochetosis (HIS) has any associated symptoms or lesions. In this study, we assessed the prevalence of HIS in sessile serrated adenomas/polyps (SSA/Ps) and their possible association. Following identification of early cecal cancer with SSA/P accompanied by a colonization of HIS, we went on to conduct a retrospective case-control study using endoscopically resected SSA/P specimens to examine the frequency of HIS infection in SSA/Ps. Nineteen SSA/P cases and 172 controls were obtained. The rate of HIS infection was significantly higher at 52.6% (10/19) in the SSA/P cases compared to the controls at 8.1% (14/172). Our SSA/P series were associated with a remarkably higher rate of HIS than controls or than previously reported. This is the first report to provide evidence for potential association between HIS and SSA/Ps.

High prevalence of spirochetosis in cholera patients, Bangladesh.

The microbes that accompany the etiologic agent of cholera, Vibrio cholerae, are only now being defined. In this study, spirochetes from the genus Brachyspira were identified at high titers in more than one third of cholera patients in Bangladesh. Spirochetosis should now be tracked in the setting of cholera outbreaks.

Three unique groups of spirochetes isolated from digital dermatitis lesions in UK cattle.

Bovine digital dermatitis (BDD) is a severe infectious cause of lameness which has spread through dairy cattle populations worldwide, causing serious welfare and agricultural problems. Spirochetes are the main organisms implicated and have previously proven difficult to isolate. This study aimed to isolate and characterise the range of spirochetes associated with BDD in the UK. Twenty-three spirochete isolates were obtained from 30 BDD lesions, which by 16S rRNA gene and flaB2 gene analysis clustered within the genus Treponema as three phylogroups; groups 1 (Treponema medium/Treponema vincentii-like), 2 (Treponema phagedenis-like) and 3 (Treponema denticola/Treponema putidum-like). The treponemes displayed large genotypic and phenotypic diversity between phylogroups and differed from named treponeme species. A previously isolated contagious ovine digital dermatitis spirochete was located within one of the three phylogroups, group 3, and could also be identified within this group on the basis of phenotype testing, suggesting BDD and contagious ovine digital dermatitis may share the same aetiological agent. A strain isolated from a bovine interdigital dermatitis lesion, could be identified as part of BDD isolate group 2, suggesting bovine interdigital dermatitis and BDD may have the same causative agent. Two common enzyme activities, C4 esterase and C8 esterase lipase, were identified in all BDD associated treponemes suggesting common metabolic pathways for sharing this novel niche or even common virulence traits. Further studies are required to determine whether the three groups of novel treponemes are representative of new treponeme taxa and to delineate how they interact with bovine tissues to cause disease.

Spirochetes in otology: are we testing for the right pathogens?

OBJECTIVE: To determine the prevalence rate of otosyphilis and Lyme-associated labyrinthine disease among patients presenting with idiopathic, progressive sensorineural hearing loss and, thus, to provide recommendations regarding the diagnostic workup. STUDY DESIGN: A prospective cohort study of patients presenting to a university hospital otology clinic with idiopathic, progressive sensorineural hearing loss between 1998 and 2006. SUBJECTS AND METHODS: Cases in which hearing loss was clearly attributable to other causes were excluded, leaving 181 patients (mean age 55.6 years). History, physical examination, autoimmune laboratory tests, syphilis and Lyme serology, audiometric assessment, and MRI were performed in all cases. RESULTS: Nine cases (4.9%) of otosyphilis and none of Lyme disease were diagnosed by serological tests. CONCLUSION: Routine serological testing for otosyphilis, but not Lyme disease, is recommended for all cases of idiopathic, progressive sensorineural hearing loss.

Human intestinal spirochetosis in Japan; its incidence, clinicopathologic features, and genotypic identification.

Human intestinal spirochetosis is a common condition in Western countries, but is not well recognized in Japan. To demonstrate the incidence and clinicopathologic findings of human intestinal spirochetosis in Japan, we retrospectively investigated biopsy, and endoscopically or surgically resected specimens of the large intestine. Among a series of 2556 samples, 11 cases of human intestinal spirochetosis were detected (0.4%). Together with additional nine cases sporadically found, 20 cases of human intestinal spirochetosis were subjected to molecular detection of two strains of spirochetes (Brachyspira aalborgi and Brachyspira pilosicoli) by amplifying species-specific portion of 16S ribosomal RNA and NADH oxydase gene by polymerase chain reaction. B. aalborgi was detected in all cases examined, three of which revealed dual infection of both species. Our results suggest that human intestinal spirochetosis infection is relatively rare, and B. aalborgi is the most prevalent species in Japan. Most of human intestinal spirochetosis were asymptomatic, although symptomatic in exceptional cases. In addition, we emphasize a usefulness of immunostaining with anti-Treponema pallidum and anti-Mycobacterium bovis polyclonal antibodies for detecting the spirochetes.

Human intestinal spirochetosis: analysis of the symptoms of 209 patients.

OBJECTIVE: Colonization of the colorectal mucosa with spirochetes is very rare. Owing to the small number of cases, it is not clear from the currently available publications whether spirochetes colonizing the colorectal mucosa are harmless commensals or pathogenic organisms. Furthermore, the reported complaints of these patients cannot be pooled to identify a characteristic complex of symptoms. The aim of the present work was to describe the symptoms associated with intestinal spirochetosis in a population of 209 patients, and to elucidate the effect of antibiotic treatment on these symptoms. MATERIAL AND METHODS: A total of 209 carefully processed questionnaires providing information on the symptoms, treatment and post-treatment symptoms of patients with spirochetosis were evaluated statistically and descriptively with the aid of the SPSS program, and the results were compared with those reported in the currently available literature. RESULTS: Of the 209 patients 168 (80.4%) were males, and the average age of the entire population at establishment of the diagnosis was 50.75 years. The most common symptoms reported were abdominal pain (46%), diarrhoea (51%) and alternating diarrhoea and constipation (13%). In this population, homosexuality and HIV infection played only a small role (6.5% homosexual patients, 3.8% HIV infected). In 72 of the 84 patients who received treatment (86%), the antibiotic employed was metronidazole, and the symptoms improved in 44 of the 84 patients (5%). Twenty-six of the 84 patients (30.9%) were investigated by colonoscopy/biopsy after receiving medical treatment. Biopsies in 20 of these patients no longer revealed infection with spirochetes, and symptoms were found to have improved in 11 of the 20 patients (55%). CONCLUSIONS: If intestinal spirochetosis is diagnosed to be the sole intestinal pathology in symptomatic patients, the bacteria should be eradicated with metronidazole and a colonoscopy/biopsy follow-up performed, where indicated, in patients with persisting symptoms. Significant results regarding symptoms and treatment of intestinal spirochetosis can be achieved only in a prospective, placebo-controlled, randomized, crossover study. In view of the low prevalence of this condition, such a study is difficult to implement.

Human intestinal spirochaetosis in Parma: a focus on a selected population during 2002-2005.

BACKGROUND AND AIM OF THE WORK: Human intestinal spirochaetosis (HIS) is a large bowel infection characterised by the colonization of the intestinal mucosa by spirochaetes belonging to the genus Brachyspira. The causative agents of HIS are Brachyspira aalborgi and Brachyspirapilosicoli. Symptoms of the infection, even if not specific, are long standing diarrhoea, abdominal pain, meteorism and rectal bleeding and sometimes they can suggest the clinical suspect of inflammatory bowel diseases or rectal carcinoma. Since poor data were available on the prevalence of this infection, the aim of our study was to describe the occurrence of this infection in our area in the period 2002-2005. METHODS: During a period of 4 years we analysed 297 faecal samples from 99 patients selected by potential risk factors and symptomatology suspected for HIS. The diagnosis of HIS was performed by isolation and a molecular assay based on 16S rDNA restriction fragment length polymorphism (RFLP)-polymerase chain reaction (PCR). RESULTS: From 2002 to 2005 we detected 12 cases of intestinal spirochaetosis, 7 caused by Brachyspira aalborgi, 4 by Brachyspirapilosicoli and one by both spirochaetes, which represented the first case of a mixed infection by 2 intestinal spirochaetes in our area. CONCLUSIONS: Despite the fact that HIS seems to be a low prevalence infection in our area, in a strongly selected population we found 12 cases of this infection (12.12%). These results stimulate us to extend the research of intestinal spirochaetosis in the general population, when long standing gastrointestinal disorders and potential risk factors are present.

Risk factors associated with Brachyspira hyodysenteriae PCR-positivity in East-European pig production units.

UNLABELLED: The objective of the present study was to determine the risk factors for swine dysentery in East-European middle-to-large sized, farrow-to-finish units, with separate breeding and grower-finisher facilities. Samples of faeces from 10 breeding animals (3-10% of the female inventory) and 10 grower finisher pigs (80-140 days of age) were collected for polymerase chain reaction testing (PCR) for Brachyspira hyodysenteriae (B. hyo). Of 139 farrow-to-finish units, 51 (36.7%) were positive, 49 (35.3%) were negative, and 39 (28.1%) were inconclusive for B. hyo by PCR. In breeding subunits, twelve variables passed the screening criterion for risk factors (P < .2) for B. hyo PCR positivity. The odds of the breeding subunits being B. hyo PCR positive were 3.5 times greater when the grower-finisher subunit was positive and the fibre content of the diet was > 6%. Use of 'all-in-all-out' farrowing policy and having >60% multiparous sows each reduced the odds of being B. hyo PCR-positive by 4-fold. In grower-finisher subunits, fourteen variables passed the screening criterion for risk factors (P < .2) for B. hyo PCR positivity. B. hyo PCR-positive status of the breeding subunits and higher fibre content of the diet were the most influential variables, with the odds of the grower-finisher subunits being B. hyo PCR positive being almost 8 times greater when the breeding subunit was also B. hyo PCR positive. Grower-finisher B. hyo PCR positivity was also associated with the percentage of pigs housed on concrete slats, with the odds of being positive being 7.5-times higher for subunits where more that 70% of the animals were kept on concrete slats compared with all other floor types. There was a strong association between grower-finisher status and whether the animals were on outdoor lots, with the odds of being B. hyo PCR positive being substantially lower for pigs on outdoor lots compared with all other surfaces. IN CONCLUSION: All-in-all-out management in the breeding units, B. hyo negativity of adjacent grower-finisher units, high fibre content of the diet, and older parity structure in a sow herd may reduce the risk of swine dysentery. In grower-finisher units, slatted flooring is associated with a higher risk, while B. hyo negativity of the breeding units, the fibre content of the diet, and outdoor production are associated with lower risk of swine dysentery.

The prevalences of Brachyspira spp. and Lawsonia intracellularis in Swedish piglet producing herds and wild boar population.

The aim of the present study was to survey the prevalences of the enteric pathogens Brachyspira hyodysenteriae, Brachyspira pilosicoli and Lawsonia intracellularis in Swedish growing pigs and in the Swedish wild boar population and to relate these findings to clinical signs. The study included 105 randomly selected herds, constituting approximately one third of Swedish herds with a herd size of >100 sows. The herds were located all over the country. In these herds, growth promoters were not used and pigs sampled were not subjected to any medication. From each herd, samples were taken from 10 growing pigs aged 8-12 weeks, corresponding to approximately 2.5% of all growing pigs present in the herd at the sampling occasion. If possible, the samples were taken from pigs with diarrhoea. Forty-eight faecal samples and 71 rectal swabs were also taken from free-living wild boars (31 piglets, 19 growers and 21 adult animals) at shooting. The samples were analysed by culture and biochemical tests for the presence of Brachyspira spp. and by nested PCR for the presence of L. intracellularis. Brachyspira hyodysenteriae was not demonstrated in any sample. Brachyspira intermedia was detected in 22 samples originating from 15 herds, Brachyspira innocens/Brachyspira murdochii was detected in 370 samples from 82 herds and B. pilosicoli was detected in 134 samples originating from 34 herds. In 21 herds and in 534 samples, no Brachyspira spp. were detected. Lawsonia intracellularis was demonstrated in 285 samples from 50 herds. Further, 418 samples from conventional herds were negative with respect to L. intracellularis and in 345 samples the PCR had been inhibited. All samples from the wild boars were negative for Brachyspira spp., 12 of 48 samples were negative for L. intracellularis, and in 36 wild boar samples, the PCR was inhibited.

Prevalence of Lawsonia intracellularis, Brachyspira hyodysenteriae and Salmonella in swine herds.

The prevalence of Lawsonia intracellularis, Brachyspira hyodysenteriae and Salmonella spp. were investigated by multiplex PCR using fecal samples of pigs with diarrhea or a history of diarrhea. The overall herd prevalence of L. intracellularis, B. hyodysenteriae and Salmonella spp. were 46.5%, 37.2% and 51.1%, respectively. Also, the prevalence of L. intracellularis, B. hyodysenteriae and Salmonella spp. among all sampled pigs were 19.9%, 10.8% and 17.7%, respectively. Seventeen of 43 herds were positive with 2 enteric organisms, and 2 herds were positive with L. intracellularis, B. hyodysenteriae and Salmonella spp. simultaneously. It was notable that 11 of 12 herds with more than 2,000 pigs were affected with Salmonella spp., and that only 2 of 12 the herds were affected with B. hyodysenteriae. This study suggested that herds positive for L. intracellularis, B. hyodysenteriae and Salmonella spp. were distributed throughout Korea, although the relationship among other pathogens such as viral or parasitic ones and/or with metabolic disorders was not determined.

Amplification and sequencing of Brachyspira spp. specific portions of nox using paraffin-embedded tissue samples from clinical colitis in Austrian pigs shows frequent solitary presence of Brachyspiramurdochii.

Brachyspira infections are significant causes of enterocolitis in pigs. In order to differentiate pathogenic species (Brachyspira (Br.) hyodysenteriae, Brachyspira pilosicoli) from less pathogenic or non-pathogenic species (Brachyspira intermedia, Brachyspira innocens, Brachyspira murdochii) in paraffin-embedded tissue samples a polymerase chain reaction (PCR) protocol allowing identification of Brachyspira at species level in archival material was developed. This approach was complemented by sequencing of the PCR amplification products. All seven cases presented with clinical and morphological Brachyspira-associated enterocolitis. Br. hyodysenteriae was not identified in any of the cases, while Br. pilosicoli was identified in a single case in conjunction with Br. murdochii. One case each was found positive for Br. innocens and Br. intermedia. Interestingly, the majority of cases presented as single or double infections with Br. murdochii. In some of the pigs other pathogens, like porcine circovirus-2 or Lawsonia intracellularis were present. These observations point at the possibility that under certain conditions even Brachyspira species of low pathogenicity can multiplicate extensively and lead to Brachyspira-associated enterocolitis.

A cross-sectional study to investigate the occurrence and distribution of intestinal spirochaetes (Brachyspira spp.) in three flocks of laying hens.

A cross-sectional study was conducted on a commercial egg-producing farm with a history of wet litter. A total of 600 fresh caecal faecal samples were obtained from under cages of laying hens in three sheds each containing flocks of approximately 5400 hens. Samples were cultured for intestinal spirochaetes, and growth on the primary isolation plate was observed under a phase contrast microscope and subjected to PCRs specific for the intestinal spirochaetes Brachyspira intermedia and Brachyspira pilosicoli. Spirochaete isolates obtained in pure culture were assessed for their ability to cause haemolysis on blood agar and to produce indole, and were typed using pulsed field gel electrophoresis (PFGE). A 1250 base pair portion of the 16S rRNA gene of three B. intermedia and five unidentified isolates was sequenced, and the sequences compared with those of other Brachyspira species. Overall, 121 (20.2%) of the faecal samples contained spirochaetes as determined by growth on the plate and microscopy. Using PCR on the primary growth from these positive samples, 43 (7.2% overall) were shown to contain B. intermedia, 8 (1.3%) to contain B. pilosicoli, and 70 (11.7%) were PCR negative. Only 24 isolates of B. intermedia and five isolates of unknown species were obtained in pure culture. Comparative analysis of the 16S rRNA gene sequence identified the non-B. intermedia isolates as belonging to the proposed species "Brachyspira pulli". PFGE analysis of the B. intermedia strains identified them as having four major banding patterns. Individual patterns were found in hens from different flocks, suggesting cross-transmission of strains between flocks. No environmental sources of infection were identified. The youngest flock had a significantly lower level of colonisation with B. intermedia than the flock of intermediate age (P = 0.004), suggesting that following initial infection of individual young hens on this farm there was amplification and transmission of infection amongst members of the flock.