Treponema pallidum distribution patterns in mucocutaneous lesions of primary and secondary syphilis: an immunohistochemical and ultrastructural study.

To study the different patterns of Treponema pallidum distribution in primary and secondary syphilis, 34 biopsy specimens of 8 patients with primary and 26 with secondary syphilis were assessed. Histopathological features, silver stain, and immunohistochemical T pallidum polyclonal antibody expression were investigated. The number and distribution of spirochetes were evaluated, and ultrastructural studies were performed. Spirochetes were identified with Warthin-Starry stain in 17 specimens (4/8 primary and 13/26 secondary syphilis), whereas immunohistochemical analysis disclosed spirochetes in 29 (8/8 primary and 21/26 secondary syphilis). In secondary syphilis, an epitheliotropic pattern characterized by abundant spirochetes in the lower mucosa/epidermis in an intercellular distribution was observed. In contrast, primary syphilis exhibited a mixed epitheliotropic and vasculotropic pattern further manifested by treponemes surrounding the vascular walls. These differences were statistically significant. Ultrastructural examination confirmed these results. Immunohistochemistry shows greater sensitivity when compared with Warthin-Starry staining. The immunohistochemical pattern of T pallidum distribution may permit the diagnostic differentiation of primary from secondary syphilis.

Assessment of the clinical significance of intestinal spirochaetosis.

BACKGROUND: Spirochaetes are well known causative agents of diarrhoea in veterinary medicine. However, there is no agreement as to whether or not they have any clinical significance in humans. AIMS: To assess the symptoms associated with intestinal spirochaetosis, their response to treatment and the natural history of untreated cases. METHODS: A retrospective review of all cases of intestinal spirochaetosis identified within an eight year period in a single university teaching hospital was performed. A chart review and follow up telephone interview was performed to assess the indications for colonoscopy that led to the diagnosis, treatment received, and duration and nature of symptoms. RESULTS: 18 cases were identified. The indications for colonoscopy were diarrhoea in 50% and rectal bleeding in 16.7%; also investigation of constipation, anaemia and abdominal pain, and in two cases reassessment of chronic proctitis. Two subjects were treated with metronidazole and two were treated with aminosalicylates. 69% had complete resolution of symptoms at follow-up, 15% had persistent symptoms and 15% had intermittent symptoms. Of the two patients treated with metronidazole, one had resolution of symptoms and one has persistent abdominal pain. CONCLUSION: Symptoms do not appear to parallel spirochaete persistence or eradication and therefore it seems appropriate to adopt a wait and see approach to treatment of patients in whom spirochaetes are identified, giving a trial of antimicrobial treatment only in those who have severe or persistent symptoms. Careful consideration of both host and pathogen should be undertaken.

Infective colitis associated with human intestinal spirochetosis.

AIM: Our study reports the detection and identification of intestinal spirochetosis in patients with colonic diseases in a tertiary-care hospital over a 12-year period, and includes a description of all cases we diagnosed. METHODS: Our patients (8323) underwent colonoscopy and histopathological examinations including transmission electron microscopy (TEM) and light microscopy. Specimens from patients suspected of intestinal spirochetosis at histopathology (17 patients) underwent microbiological investigation performed by culture and molecular methods (16S restriction fragment length polymorphism-polymerase chain reaction [RFLP-PCR], nox RFLP-PCR assays). RESULTS: Seventeen cases were diagnosed: seven patients were infected by B. aalborgi, one by B. pilosicoli, two by both species and four by Brachyspira spp. diagnosed both histopathology and microbiology (culture and molecular methods: 16S RFLP-PCR and nox RFLP-PCR assays). Three cases were referred to as Brachyspira spp. infections using only histopathology, including TEM. CONCLUSIONS: Our results demonstrated that intestinal spirochetosis, although rarely occurring, might play a role in chronic diarrhea and suggested a pathogenetic mechanism of intestinal spirochetosis based on the destruction of colonic microvilli and colitis histologically documented, providing additional clinical and pathological information on this entity. This study suggests that metronidazole seems to be the drug of choice for the eradication of intestinal spirochetosis.

Spirochetal cyst forms in neurodegenerative disorders,...hiding in plain sight.

Here is proposed a hypothesis that a completely unsuspected biology exists for pathogenic spirochetes, namely that the cystic spirochetal forms (long thought to be static and resting or just a dormant cohort) actually are capable of killing mammalian host cells. At least two "lethal" scenarios are proposed; first, the host cell destruction from the "inside out" by small caliber cystic forms invading the host cell cytoplasm, and second host cell destruction by engulfment of entire host cells by large caliber cystic spirochetal forms. Conventional thinking about spirochetal cyst forms is divided between two polar spheres of influence; one a majority community that completely denies the existence of spirochetal cyst forms, and a second group of academically persecuted individuals who accepts the precepts of such antebellum scientists as Schaudinn, Hoffman, Dutton, Levaditi, Balfour, Fantham, Noguchi, McDonough, Hindle, Steiner, Ingraham, Coutts, Hampp, Warthin, Ovcinnikov, and Delamater. Microscopic images of cystic spirochetes are difficult to ignore, but as has been the case in this century, academic "endowments" have nearly expunged all cystic spirochetal image data from the current textbook versions of what is the truth about the spirochetaceae. If the image database from the last century is obliterated; many opportunities to diagnose will be lost. Variously sized cystic spirochetal profiles within diseased nerve cells explain the following structures: Lewy body of Parkinson's disease, Pick body, ALS spherical body, Alzheimer plaque. Borrelia infection is therefore a unifying concept to explain diverse neurodegenerative diseases, based not entirely on a corkscrew shaped profile in diseased tissue, but based on small, medium and large caliber rounded cystic profiles derived from pathogenic spirochetes which are..."hiding in plain sight".

A life cycle for Borrelia spirochetes?

Subsequent to Schaudinn and Hoffman's visualization of Treponema pallidum in 1905, many distinguished syphilologists proposed that spirochetes have a life cycle. What is the "essence" of a life cycle? Simply put, life cycles are diverse arrays of life forms, which emerge in an ordered sequence; which are "connected" to one another across primary and secondary hosts, and constitute a cycle with "circular" relationship between hosts. Fecal-oral life cycles and blood-to-blood life cycles are exemplary of host parasite relationships in this realm. The "blood-to-blood" begins and ends with an insect taking a blood "meal". In this operatic scenario, a "blood-less" insect functions simultaneously as a hypodermic needle and as an incubator for some of the infectious components. The initial phase is inside the body fluid compartment of an insect. The second phase is in the blood or body fluid of a warm-blooded mammal. Third, is the phase inside the cell of a mammalian host. And a final portion of the "life" marked by "death" of the parasitized mammalian cells and the release of infectious parasites which return to the "warm" blood where the "cold blooded" vector again takes a blood meal. The cycle then begins again. In each phase of a blood to blood life cycle, the infectious agent changes its shape. Blood phase "profiles" look different from "tissue phase" profiles. Some of the tissue phase profiles may be "invisible". Borrelia spirochetes offer an excellent example of a life cycle, by virtue of the insect vector to mammalian "piece", the blood and intracellular residence "pieces" and the morphologic diversity "piece". Stereotypes of what a spirochete "should " look like, have actually produced a state of "perseveration" in spirochetal pathobiology. We have been "stuck" like a broken record, on the corkscrew form, and have failed to see the rest of the life cycle. Cystic, granular, and cell wall deficient spirochetal profiles, which were well known in the 19th and 20th centuries by such titans as Schaudinn, Hoffman, Noguchi, Delamater, Steiner, and Mattman, have been repudiated by professional microbiologists, and by pathologists who practice and who confer the status of 21st century truths in microbiology matters. Proper microscopic study, as is required by Dr. Robert Koch's second postulate, for establishing links between microbes and disease, presupposes that the microscopist be aware of the complete array of morphologic repertoires of the alleged pathogen. (Morphologies, which are herein introduced.).

Morphological properties of a human intestinal spirochete first isolated from a patient with diarrhea in Japan.

A human intestinal spirochete isolated from a rectal biopsy specimen was morphologically characterized. The isolate was comma-shaped, 3-6 microm in length, 0.2 micro m in diameter and had tapered ends. The surface layer, external to the outer envelope, was amorphous. Four string-like periplasmic flagella with a diameter of 20 nm were presented at each end of the SDS-treated cells. Thin sections of the bacterial cell revealed a high-density cytoplasmic membrane and flagella in the periplasmic space between the cytoplasmic membrane and the outer envelope. Three segments of equal length were observed in some of the cells, while other cells were bi-segmented and more frequently observed.

A TEM/SEM study of the microbial plaque overlying the necrotic gingival papillae of HIV-seropositive, necrotizing ulcerative periodontitis.

The purpose of this study was to examine by transmission (TEM) and scanning electron microscopy (SEM) the supragingival microbial plaque overlying the ulcerated gingival papillae of necrotizing ulcerative periodontitis (NUP) lesions in HIV-seropositive patients. The microbiota of NUP and HIV-seropositive patients with periodontitis has been reported to be similar to that of conventional periodontitis in non-infected subjects, although several investigators have also reported high recovery rates of microbes not generally associated with the indigenous oral microbial flora. Light and electron microscopic observations and microbial culture studies indicate a similar high prevalence of spirochetes in both necrotizing ulcerative gingivitis (NUG) and NUP. In addition, several studies have reported more frequent isolation of Candida albicans from diseased periodontal sites in HIV-seropositive patients than from non-diseased sites. Ten male and six female patients, each HIV-seropositive and exhibiting NUP, constituted the study population. Two biopsies of involved gingival papillae from between posterior teeth were obtained from each patient and processed for examination by both TEM and SEM. Microscopic examination revealed a surface biofilm comprised of a mixed microbial flora of various morphotypes in 81.3% of biopsy specimens. The subsurface flora featured dense aggregations of spirochetes in 87.5% of specimens. Zones of aggregated polymorphonuclear leukocytes and necrotic cells were also noted. Yeasts were observed in 65.6% of specimens and herpes-like viruses in 56.5% of the specimens. Collectively, except for the presence of yeast and viruses, the results suggest that the microbial flora and possibly the soft tissue lesions of NUP and necrotizing ulcerative gingivitis are very similar.

Brachyspira aalborgi infection in four Australian children.

AIM: The clinical presentation of four children and adolescents (two males and two females with a mean age of 12.4 years; range 9-16 years) with colorectal spirochetosis is discussed. RESULTS: Symptoms included persistent diarrhea (n = 2), rectal bleeding (n = 1) and abdominal pain (n = 2). In all patients, colorectal spirochetosis was an unanticipated finding on colonic histology, and the presence of spirochetes was confirmed by the use of electron microscopy. Spirochetes were identified as Brachyspira aalborgi by using PCR amplification of the bacterial 16S rRNA and nicotinamide adenine dinucleotide oxidase sequences in all four patients. No other enteric pathogens were found. CONCLUSIONS: Although all patients appeared to respond to antibiotic treatment, the clinical significance of B. aalborgi as a human pathogen requires further investigation.

Canaleparolina darwiniensis, gen. nov., sp. nov., and other pillotinaceous spirochetes from insects.

We describe two new pillotinaceous spirochetes (Canaleparolina darwiniensis, Diplocalyx cryptotermitidis) and identify for the first time Hollandina pterotermitidis from both the subterranean termite Cryptotermes cavifrons and the wood-eating cockroach Cryptocercus punctulatus based on morphometric analysis of transmission electron micrographic thin sections. C. darwiniensis, gen. nov., sp. nov., limited to near Darwin, Australia, invariably is present on the surface of the treponeme-studded trichomonad Mixotricha paradoxa, a consistent inhabitant of the hindgut of healthy termite Mastotermes darwiniensis. The spirochete both attached to the surface of protists and free-swimming in the paunch (hindgut) lumen of the insect has 16 periplasmic flagella (16:32:16) and imbricated wall structures that resemble flattened crenulations of Pillotina. The flagella surround half the protoplasmic cylinder. C. darwiniensis is the largest (0.5 microm diameter x 25 microm length) of the three epibiotic bacteria (two spirochetes, one rod) that comprise the complex cortex of its host Mixotricha paradoxa. Several criteria distinguish Diplocalyx cryptotermitidis sp. nov. isolated from Cryptotermes cavifrons intestine: smaller diameter, fewer flagella, absence of inner and outer coats of the outer membrane, wider angle subtended by its flagella and, most notably, cytoplasmic tubule-associated centers, which are periodic electron dense spheres within the protoplasmic cylinder from which emanate cytoplasmic tubules up to 24 nm in diameter. This is also the first report of abundant populations of Hollandina in Cryptotermes cavifrons (those populations belong to the species H. pterotermitidis). Morphometric analysis of the first thin sections of any spirochetes (published nearly 40 years ago by A.V. Grimstone) permits us to identify the large (0.9 microm diameter) free-swimming intestinal symbiont of Cryptocercus punctulatus also as Hollandina pterotermitidis.

In vitro conversion of Borrelia burgdorferi to cystic forms in spinal fluid, and transformation to mobile spirochetes by incubation in BSK-H medium.

The purpose of this study was to examine the structural alterations of Borrelia burgdorferi when exposed to spinal fluid. Normal, mobile spirochetes were inoculated into spinal fluid, and the spirochetes were converted to cysts (spheroplast L-forms) after 1-24 h. When these cystic forms were transferred to a rich BSK-H medium, the cysts were converted back to normal, mobile spirochetes after incubation for 9 to 17 days. The cultures were examined by dark field microscopy (DFM), interference contrast microscopy (ICM) and transmission electron microscopy (TEM). When neuroborreliosis is suspected, it is necessary to realize that B. burgdorferi can be present in a cystic form, and these cysts have to be recognized by microscopy. This study may also explain why cultivation of spinal fluid often is negative with respect to B. burgdorferi.

The first reported case of intestinal spirochaetosis in Japan.

A 65-year-old Japanese male consulted Ozuchi Prefectural Hospital (Iwate, Japan) on 19 January 1994 complaining of weight loss. Cecal mucosal biopsy material, which was stained with hematoxylin-eosin revealed a thick, basophilic fuzzy fringe covering the surface epithelium. Transmission and scanning electron microscopy observations demonstrated the presence of slightly wavy spirochaetes with tapered ends, which were attached to the surface epithelium of the colonic mucosa via one of these ends. The patient did not display any clinical symptoms of inflammatory bowel disease, and laboratory tests eliminated an immunodeficiency condition. Thus, in the present case, the intestinal spirochaetes appear to be harmless commensals. This paper presents the first reported case of intestinal spirochaetosis in Japan.

Intestinal spirochetosis: first cases reported in Brazil and the use of immunohistochemistry as an aid in histopathological diagnosis.

Colonization of the colon and rectum by intestinal spirochetes is detected for the first time in Brazil in 4 of 282 (1.41%) patients who had undergone sigmoidoscopy and/or colonoscopy with a histopathological diagnosis of chronic non specific-colitis. This frequency is probably underestimated, since surgically obtained specimens were not considered in the present study. Histopathological diagnosis was performed using routine stains like hematoxylin-eosin which showed the typical, of 3-microns thick hematoxyphilic fringe on the brush border of the surface epithelium, and by silver stains like the Warthin-Starry stain. Immunohistochemical procedures using two, polyclonal, primary antibodies, one against Treponema pallidum and the other against Leptospira interrogans serovar copenhageni serogroup Icterohaemorrhagiae cross-reacted with spirochetal antigen/s producing a marked contrast of the fringe over the colonic epithelium, preserving the spiral-shaped morphology of the parasite. In one case with marked diarrhea, immunohistochemistry detected spirochetal antigen/s within a cell in an intestinal crypt, thus demonstrating that the infection can be more widely disseminated than suspected using routine stains. Immunohistochemical procedures, thus, greatly facilitate the histological diagnosis of intestinal spirochetosis and may contribute to a better understanding of the pathogenesis of the disease. Transmission and scanning electron microscopy performed in one case showed that the spirochete closely resembled the species designated as Brachyspira aalborgi.

Spirochetes isolated from dairy cattle with papillomatous digital dermatitis and interdigital dermatitis.

Two groups of spirochetes were isolated from papillomatous digital dermatitis (PDD) lesions in dairy cattle. The two groups could be readily differentiated on the basis of morphologic and immunologic characteristics and enzymatic activity. A spirochete isolated from an interdigital dermatitis (IDD) lesion appeared morphologically and antigenically similar to spirochetes in one of the PDD groups and exhibited an identical enzyme activity pattern. The two groups of PDD spirochetes had characteristics most consistent with the genus Treponema. The PDD and IDD isolates differed morphologically from previously described bovine Treponema spp. Although spirochetes have been observed to be one of the predominant bacterial morphotypes in PDD and IDD and are found invading the stratum spinosum and dermal papillae in PDD lesions, the significance of these spirochetes in the etiopathogenesis of PDD and IDD is presently unknown.

Intestinal spirochetosis and acquired immunodeficiency syndrome: ultrastructural studies of two cases.

Two cases of intestinal spirochetosis (IS) with acquired immunodeficiency syndrome are reported. In case 1, a 48-year-old homosexual black man presented with a 1-month history of alternating watery diarrhea and constipation, which dissipated following the removal of two colonic hyperplastic polyps containing IS. In case 2, a 26-year-old homosexual black man presented with a 3-month history of persistent bloody diarrhea and was found to have chronic shigellosis and IS. Pathologic findings of IS were similar in both cases. Basophilic fringes typical of IS covered the surfacing colonic epithelium and consisted of dense growths of spirochetes adherent to and oriented perpendicular to the plasma membranes of the surfacing epithelium. The spirochetes measured 3 to 5 microns in length and 0.2 micron in width, contained four to eight axial fibrils, and closely resembled Brachyspira aalborgi ultrastructurally. These cases are notable because the histopathologic changes of IS were more extensive than generally described. There was involvement of both the right colon and rectum by IS in case 2, and in both cases there was extension of the IS down into the crypts of Lieberkühn, spirochetal invasion of the colonic mucosa, and a conspicuous inflammatory response by macrophages in the underlying lamina propria.

Necrotizing typhlocolitis associated with a spirochete in rheas (Rhea americana).

Necrotizing typhlocolitis was diagnosed in 13 juvenile common rheas (Rhea americana) from three separate of geographically isolated Ohio flocks, with mortality ranging from 25% to 80%. At postmortem examination, a diphtheritic membrane covered ulcerated cecal mucosa. Histologically, cecal sections showed necrosis and granulomatous-to-suppurative inflammation that extended into the submucosa and often surrounded large eosinophilic colonies of bacteria. Warthin-Starry staining showed these colonies to be composed of entangled spirochetes that invaded the submucosa and frequently were present transmurally. Similar organisms were identified by Warthin-Starry staining in the cecum of a juvenile rhea from a fourth flock that histologically had mild lymphocytic typhlitis. Scanning and transmission electron microscopy demonstrated the presence of a spirochete in the ceca. Anaerobic culture yielded a gram-negative, beta-hemolytic spirochete. Coccidia, histomonads, and Salmonella spp. were consistently absent.

Unusual in vitro formation of cyst-like structures associated with human intestinal spirochaetosis.

Spirochaetes were isolated from rectal biopsies of three patients and successfully cultured. Enzymatic reactions and electron microscopy revealed spirochaetes resembling Brachyspira aalborgi. Examination of ultrathin sections of centrifugates of cultured spirochaetes yielded unusual cyst-like structures with an outer double membrane containing spirochaetes in different developmental stages. The protoplasmic cylinders seemed to originate from a large electron-dense focus in the cyst-like structures. The axial fibrils occurred as loosely distributed solitary structures in the cyst-like structures. Cysts were not found in the biopsy tissue. The encystment of the spirochaetes could be related to their protection, multiplication, spread and transmission.

Morphology as a basis for taxonomy of large spirochetes symbiotic in wood-eating cockroaches and termites: Pillotina gen. nov., nom. rev.; Pillotina calotermitidis sp. nov., nom. rev.; Diplocalyx gen. nov., nom. rev.; Diplocalyx calotermitidis sp. nov., nom. rev.; Hollandina gen. nov., nom.[TRUNCATED].

The purposes of this paper are (i) to present a framework for the morphometric analysis of large uncultivable spirochetes that are symbiotic in wood-eating cockroaches and termites; (ii) to revive, in accordance with the rules of the International Code of Nomenclature of Bacteria, the names of three genera (Pillotina, Diplocalyx, and Hollandina) and three species (Pillotina calotermitidis, Diplocalyx calotermitidis, and Hollandina pterotermitidis) for the same organisms to which the names were originally applied, because these names were not included on the 1980 Approved Lists of Bacterial Names; and (iii) to formally propose the name Clevelandina reticulitermitidis for a new genus and species of spirochetes from the termite Reticulitermes tibialis. None of these genera and species has been cultivated either axenically or in mixed culture; hence, all are based on type-descriptive material.

Rectal spirochaetosis in homosexual men: characterisation of the organism and pathophysiology.

Microbiological and electron microscopy studies were carried out on rectal biopsy specimens and faecal samples from eight practising male homosexuals and five heterosexual controls. Rectal spirochaetosis was present in five of the eight homosexual men. The organism was cultured and morphologically identified as a large anaerobic host associated treponeme. The degrees of infestation and depletion of microvilli were also measured. The results are discussed in relation to the possible clinical importance of rectal spirochaetosis.

Intestinal spirochetosis of the vermiform appendix.

A series of 681 surgically removed appendices were examined for spirochetes. Five hundred seventy-four appendices were removed because of suspected acute appendicitis; the rest were removed per occasionem. One hundred six of the former were histologically normal, whereas 421 showed acute appendicitis. The remaining 47 specimens showed a variety of other pathological conditions, for example, tumors and diverticula. Spirochetes were found in 13 (12.3%) of the appendices removed from patients clinically suspected to suffer from acute appendicitis, but whose appendices were otherwise histologically normal (pseudoappendicitis). Only 3 patients (0.7%) with histologically confirmed acute appendicitis (p less than 0.05) did show spirochetes in their appendices. Of the 107 patients who had their appendices removed per occasionem, 2 patients (1.9%) had spirochetosis (p less than 0.05). The ultrastructure of the spirochetes obtained from appendices with spirochetosis was studied by means of negative staining and ultrathin sectioning. The morphology of these spirochetes was very similar to that of Brachyspira aalborgi, a spirochete recently isolated from rectal biopsy specimens obtained from patients with colorectal spirochetosis.