Phase 1: Recovery of Morgellons-Related Particles
from Water Samples
The descriptive information below is a phase I summary report presenting the findings of an investigative research study. This work is supported through a grant from the Morgellons Research Foundation (MRF). Phase I is the first of three phases proposed in this research project. Phase I is a “look-see” initial study to determine what, if any, organisms, pathogens and/or materials in household water samples may be associated with Morgellons Disease. Phase I is only a findings presentation and no conclusions as to association and/or cause of Morgellons Disease are drawn. This initial phase lays the ground work for phases II and III as well as future research projects.
It is hoped that this information will encourage other researchers and scholars with knowledge in this field, to contribute to the present information base by becoming involved in this and additional research studies related to Morgellons disease.
Phase I – Brief Summary
There are six particle types that are consistently recovered from the skin surface of those suffering from Morgellons disease, 1) ribbon-like fibers, 2) rounded fibers, 3) capsule-like particles, 4) black flakes/grains, 5) worm-like particles, and 6) stellate-shaped (“starfish-shaped”) particles. The fibers are often pigmented and may luminesce under ultraviolet light.
Current Morgellons research at a laboratory in Massachusetts shows that individuals affected by Morgellons disease have been in contact with soil and/or water containing cyanobacteria (blue-green algae), algae, aquatic fungi, water molds and lichen (algae and fungi). This assemblage of organisms, and associated bacterial populations, is common in soil and aquatic environments where cryptobiotic soils are present and/or in environments where nutrient rich conditions promote the development of algae blooms.
Dermal contact with a water source and/or inhalation of aerial dust containing cyanobacteria and algae may lead to the progressive colonization of organisms that are capable of feeding on or consuming these photosynthetic producers, thus contributing to the wide range of symptoms reported by Morgellons sufferers. Incidental growth of these opportunistic populations, such as actinomyces, aquatic fungi and true fungi, are known to promote disease in humans, as they consist of species capable of degrading either cellulose or keratin (skin/hair). Evidence for the presence of opportunistic micro- organisms in samples is indicated by the occurrence of capsule-like Morgellons particles that have been identified as parasitized pine pollen grains. Chytrid populations are obligate parasites of frogs, nematodes, algae, cyanobacteria, plants, and pine pollen (see photo below).
These findings serve as a focus for further understanding the ecological significance of the organisms identified in this study and the role they may play as causative agents of Morgellons disease.
Recovery of Morgellons-Related Particles
from Water Samples
TABLE OF CONTENTS
3.0 Recovery of Morgellons Particles In Hot Water Tank Samples
3.1 Ribbon-like fibers
3.2 Rounded fibers
3.3 Capsule-like particles
3.4 Stellate-shaped (starfish-shaped) particles
3.5 Worm-like particles
3.6 Black flakes
4.0 Ultraviolet Fiber Counts-Before Tank Removal and after Pipe Replacement
5.0 Microscopic Observations-Before Tank Removal and after Pipe Replacement
6.0 Culture Results- Before Tank Removal and after Pipe Replacement
6.1 Total coliform bacteria
6.2 Heterotrophic bacteria
6.4 Membrane Filter Growth
7.0 Fiber Culture Study
8.0 Sample Preservation and Archival
9.0 Ecological Significance
10.0 Summary and Conclusions
10.1 Ribbon-like fibers
10.2 Rounded fibers
10.3 Capsule-like particles
10.4 Stellate-shaped (starfish-shaped) particles
10.5 Worm-like particles
10.6 Black flakes
There are six particle types that are consistently recovered from the skin surface of those suffering from Morgellons disease, 1) ribbon-like fibers, 2) rounded fibers, 3) capsule-like particles, 4) black flakes/grains, 5) worm-like particles, and 6) stellate-shaped (“starfish-shaped”) particles. The fibers are often pigmented and may luminesce under ultraviolet light. These six particle types were recovered from a hot water tank that was suspected of harboring contaminants causing chronic skin irritation and other debilitating symptoms to the scalp, neck and shoulders of the homeowner. The hot water tank had been idle for more than 2 years and is thought to have been contaminated by soil/groundwater organisms. Replacement of the hot water system and water pipes eliminated these contaminants and resulted in a marked improvement in the homeowner’s health. Microscopic examination and microbial analysis of water samples collected before hot tank removal show the presence of numerous fiber-producing microorganisms. Tentative identification is based on morphological characteristics and include the following primary organisms of interest: 1) gliding bacteria, cyanobacteria or blue-green algae, and rhizosphere bacteria (slime-producing bacteria), 2) actinomyces (spore-forming filamentous bacteria), 2) plasmodial slime molds, oomycete water molds, chytrids and algae (protists), and 3) zygomycetes and ascomycetes (fungi/yeast). Of particular interest is the keratin degrading aquatic fungi or chytrids. Evidence for their presence in hot water tank samples is indicated by the occurrence of capsule-like Morgellons particles that have been identified as parasitized pine pollen grains. Chytrid populations are obligate parasites of frogs, nematodes, algae, cyanobacteria, and plants. Based on the findings of this Phase I study it is proposed that prolonged dermal exposure to algae-rich conditions may promote the incidental growth of opportunistic keratin degrading populations causing chronic symptoms associated with Morgellons disease. These findings serve as a focus for further understanding the ecological significance of the organisms identified in this study and the role they may play as causative agents for this disease. Membrane filters, culture slants, and water samples have been archived for further study.
Morgellons disease is often overlooked or dismissed because our current knowledge of the causative agent(s) is limited. However, symptoms associated with this condition have been reported by thousands of individuals in the United States alone, and examination of particles recovered from affected individuals show a remarkable similarity in morphology.
Morgellons disease is a “fiber disease” that presents as a chronic skin condition accompanied by itching, burning, and crawling sensations. As the disease progresses, fibers of unknown origin are found to protrude from the pores of the skin surface and/or from skin lesions. Other debilitating symptoms may accompany these primary symptoms and include general weakness, fatigue, and mental impairment. The disease is progressive and life threatening, and has been shown to cause neurological problems in some individuals.
In January 2008, a laboratory in Massachusetts received samples of cold and hot water collected from a private residence where it was believed that the hot water tank might contain contaminants associated with a chronic skin condition. The property owner reported symptoms of itching, burning, stinging, and crawling sensations on the scalp, neck, and shoulder regions. Fatigue and weakness also reportedly compromised the overall health of the owner. The history of the property revealed that the hot water tank had not been used in over 2 years.
Microscopic examination of water samples collected from the hot water tank, the shower, and kitchen locations revealed the presence of conspicuously colored filaments and other particles. Sketches and descriptions of 6 distinct particle types were recorded:
- ribbon-like filaments (clear, blue, red, green, black)
- rounded, smooth filaments in single strands or in loosely interwoven balls
- capsule-like structures (gray)
- starfish-shaped particles
- worm-like tubes
- black, irregular-edged flakes
The hot water tank sample contains an abundance of all 6 particle types, whereas the hot shower sample contains only long, rod-shaped cells (red or blue) presumably related to the filaments found in the hot water tank samples. The cold water sample did not contain any of the 6 particle types. Hot water samples, especially the hot shower sample also contained a thick biofilm population associated with the shower plumbing.
While researching the possible origins of these particles, several reports were found that described particles remarkably similar, if not identical, to those described above. These 6 particle types are commonly recovered from the skin surface of those suffering from Morgellons disease and are shown in Figure 1.
Individuals suffering from Morgellons exhibit chronic skin infections ranging from mild (rashes and dryness) to severe infections (unidentified fibers that exude from skin pores and/or lesions). After reviewing reports that described fibers or filaments associated with Morgellons, which luminesce under UV light, a UV lamp (360 nm) was used to confirm that the fibers obtained from the hot water tank and hot water shower contained luminescent blue, green, and red filaments. Based on the presence of these fibers, and other particle types from the hot water tank, the property owner replaced the hot water tank with a passive hot water system.
A second round of water samples were collected and submitted for analysis after the new hot water system was installed. Samples from cold (kitchen) and hot (shower) water sources were analyzed for the presence of Morgellons-related particles to confirm that the hot water plumbing no longer contained luminescent fibers. UV fiber counts, however, were higher after tank removal, most likely resulted from pipe disturbance during tank removal. The well- developed, sticky biofilm coating on the inside of the hot water pipes (produced by slime-producing bacteria) is likely to have captured UV filaments over time which were then sloughed off during pipe disturbance when the hot water tank was removed. The higher counts reported for the hot water shower sample coincided with an increase in symptoms experienced by the property owner who reportedly used the shower after the tank had been removed. Based on these results, the owner replaced the cold and hot water plumping on the property.
A final round of water sample analysis was performed after the plumbing had been replaced. No indication of water contamination with Morgellons-related particles was found in these samples and the health of the property owner improved markedly. During completion of this study, investigation into the possible source of these particles was initiated. Water samples, membrane filter concentrates, microscopic slides and cultures deemed of value for further research were archived. A phased approach for further studies was submitted to the Morgellons Research Foundation.
The tasks associated with each research phase are detailed in a Grant Proposal provided to the Morgellons Research Foundation. The long-term goal is to provide screening services to individuals affected by Morgellons disease, and to collect database information and samples that would be made available for future research studies.
This report documents the recovery of Morgellons-related particles during the Phase I study of the water samples collected from the residence before, during, and after hot water tank and pipe removal. Microscopic observations, microbial culture results, and the ecological significance of organisms identified are reviewed with respect to identifying the role they may play as causative agents of Morgellons and the possible origin of the particle types associated with this disease. This study is considered preliminary, with tentative identification of organisms based on morphological and growth characteristics. Water samples, particles, and cultures were archived for future study.
A total of 15 samples (Table 1), were analyzed before hot water tank removal, during pipe replacement, and after pipe replacement over the course of 3 months (January through March 2008). This Phase I study is based on the following assumptions:
- Morgellons disease is a “fiber disease” that presents as a chronic skin condition and is accompanied by itching, burning, and crawling sensations. Skin symptoms can range from mild (rashes, burning, crawling sensations) to severe (fiber growth and open lesions).
- Individuals with severe symptoms have reported the presence of six common Morgellons particle-types.
- Burning and crawling sensations experienced by the homeowner occur on the scalp, neck and shoulder regions, and is believed to have resulted from exposure to “contaminants” during hot water shower use. Hot water tank samples were found to contain an abundance of Morgellons-related particles.
- Fibers are considered to be biological in origin.
Water sample results from this Phase I study are summarized and the characteristics of contaminated samples collected before tank and pipe removal are compared with contaminant-free samples collected after pipe replacement.
Water samples were submitted for analysis in clean, one-gallon containers and were processed upon receipt. Samples were processed for archival purposes, for microscopic examination, and for microbial culture isolation. For sample archiving, a 750 ml portion of each water sample was membrane filtered (0.45 um) to concentrate particulate matter onto sterile filters that were then placed into sterile containers containing 100 ml of the original water sample. Sample concentrates have been refrigerated at 2oC for future study.
Each water sample was prepared for microscopic study by filtering 250 ml and 500 ml portions through sterile 0.45 um membrane filters. During filtration, an aliquot of the filter concentrate was transferred to sterile glass microscope slides. Filters and microscope (wet and dry) slide mounts were examined under a phase contrast microscope (40 x to 4000 x) and under an UV (360 nm) light source. The number of UV fibers recovered on the filters was counted for each water sample as an initial means of evaluating the degree of water sample contamination. Selective media was used to culture heterotrophic bacteria (R2 agar), total coliform bacteria (mEndo agar), and fungi/yeast (Sabsouraud dextrose agar) by placing membrane filters (100 ml portions from each sample) onto the agar surface. Cultures were grown at 35o C for 24 hours (total coliform), 48 hours (heterotrophs), and for 5 days at 20o C (fungi/yeast). Standard sterile procedures were used to culture and to preserve colonies of interest on agar slants for future study (1).
Particles similar to those described by individuals suffering from Morgellons disease were recovered from hot water tank water samples. The tank is known to have remained idle for more than 2 years. The conspicuous presence of these particles indicates that the homeowner’s symptoms may be Morgellons-related. Selected photographs of each of the 6 particle-types recovered from the hot water tank sample are shown in Figure 1 and are briefly described below. A more detailed discussion of their probable/possible origin is summarized in Section 10.0
Abundant flat, ribbon-like filaments of varying size and color are present in the hot tank water sample, suggesting that there are multiple biological sources for these particles. Sizes range from widths of 1 um to 1 mm, and lengths of 10 um to 3 cm. These delicate, thread-like filaments occurred in red, blue, green, brown, gray, and white, and range from translucent to opaque. Translucent varieties display a linear core region.
Most filaments occur as single strands, although there are ribbon-like fibers also found in interwoven “fuzzballs” (see below). Clear or white filaments were observed to luminesce under UV light, emitting primarily blue, green, and less commonly red colors.
Ribbon-like fibers were occasionally found to grade from a rounded, tube-like morphology into a flattened ribbon-like form, indicating that upon aging the original morphology was disrupted (or degraded) over time.
Rounded fibers also abundant and occur in similar sizes and colors (including black) as those described for the ribbon-like fibers. This suggests that both fiber-types may originate from similar biological sources. Rounded fibers are thread-like, lack cross-walls (aseptate) and occur singly or in entangled masses. Some forms show marked rigidity with conspicuous textures ranging from smooth (hollow, sheath-like), to banded (with vacuoles), scaly, or ropy. These textured fibers are also longer than most of the other fiber-types (1 cm to 3 cm).
Interwoven “fuzzballs” (entangled masses) consist of numerous pigmented fibers intertwined into a conspicuous structure. These often consist of a variety of ribbon-like and rounded filaments, with translucent varieties exhibiting fluorescence under UV light. Although rounded fibers occur primarily as single strands, many are likely to have originated from these interwoven masses.
For both ribbon-like and rounded fibers it was noted that a single strand could vary in color, with one portion grading into another color. Aged rounded fibers were also observed to be “shedding” or “peeling” an outer sheath.
Oval, capsule-like particles do not vary appreciably in size (50 um x 100 um) or color (pale gray) which suggest a common origin for these particles. Upon close inspection, the region between the darker pole areas consists of a translucent, light gray membrane appearing as a deflated membranous envelope. These particles are often found among entangled filaments.
These structures vary in size and color, although all seem to consist of 6 to 9 “tentacles”. There are two distinct forms present; clear or translucent varieties 1-2 mm in diameter with tubular or flattened (ribbon-like) “arms”, and larger, light brown opaque varieties up to 3 mm in diameter also with ribbon-like “arms”. Both varieties appear to consist of hyphae-like extensions originating from a single source point (monocentric thallus).
Particles resembling jelly-like worm tubes (3 um x 2 mm) were observed to be slightly curved and translucent. These particles are not abundant and do not contain any structures consistent with invertebrates (mouth parts, intestines etc.). In some cases their proximity to partially intact stellate-shaped particles clearly suggest that these worm-like tubes may be “tentacle” fragments. None of these worm-like particles exhibit motility or were associated with egg larvae.
The ubiquitous distribution of black flakes in nature is problematic in the study of these particles in relation to human disease. Potential inorganic (iron, sulfur, manganese etc.) and organic detritus sources abound. The study of these particles is perhaps best performed from skin samples.
Well-documented cases of black-grained mycetomas caused by fungi and/or actinomycetes (filamentous bacteria) indicate that dematiaceous (dark colored) hyphae-producing microbes account for the dark grains associated with these skin conditions (2). There are however, other unexplored sources that may merit further consideration as the ecology of Morgellons-related particle associations are better understood. Of particular interest are the black flakes observed to occur along the hyphae of an aquatic fungal colony (chytrid) recovered from the hot water tank (Figure 4A). Note that a black coating occurs along the entire length of one of the branched hyphae.
These 6 particle types occur only in hot tank water samples and suggest that their source originated from town drinking water flowing through the water mains. The integrity and condition of these water mains is likely to have an impact on the residence’s water quality.
Phase I water samples were filtered and microscopically examined before and after tank and pipe replacement to ensure that particles had been removed from the residence.
Examination of membrane filters reveal an abundance of UV luminescent fibers in water samples before hot tank replacement and before pipe installation is complete, with hot water samples containing more fibers than cold water samples ((Table 2). Although UV luminescence is known to occur widely in nature and in itself is not an indicator of conditions adverse to human health, the relative ease of enumeration and reported association with Morgellons skin disease resulted in their use as a general marker for potential water contamination.
Translucent/clear ribbon-like and rounded fibers were found to emit blue or blue-green luminescence, and in a few cases, red. Interwoven “fuzzballs” and red UV luminescent fibers were recovered in hot water tank samples only. A typical membrane filter from the hot water tank containing abundant UV fibers is shown in Figure 2.
Microscope slide mounts were also prepared during membrane filtration and were examined to identify other characteristic structures present.
In addition to the six Morgellons-related particles present in hot water tanks samples (Figure 3), additional structures of interest were also present and are shown in Figure 4. Microscope slide mounts show a diversity of structures including aquatic fungal (chytrid) colonies (note black coating along hyphae), bacterial colonies, biofilm from the tank walls, and structures indicative of the presence of actinomyces, blue-green algae (cyanobacteria), and algae. Circular cell arrangements (L) have been found to occur in both tank and shower samples before new pipes were installed, and are likely associated with populations indicative of poor water quality.
In contrast to the hot water tank samples that contain all 6 particle types, hot shower water samples contained only the ribbon-like and rounded fibers. Microscope slide mounts of water samples before tank and pipe replacement contain numerous microscopic features of note (Figure 4). In addition to ribbon-like and rounded fibers, the following conspicuous structures are also present:
- Long, hyphae-like filaments, possibly fiber fragments or immature stages of rounded fibers found in hot water tank samples. Rod-shaped cells display red, or less commonly blue, pigmentation and are 4-20 um in length (A).
- Triangular arrangements of rod-shaped cells characteristic of aquatic fungal spores (chytrids), 5 um in length (B).
- Spores/hyphae with red or blue pigmentation indicative of actinomyces, 0.5-1.0 um (C).
- Clear “crystals” consisting of 6-sided polyhedral shapes (3-8 um) often containing green, rod-shaped cells characteristic of slime-encapsulated algae/cyanobacteria cells (D).
- Various rounded structures, often colonized by bacterial cells, characteristic of unicellular algae (E).
- Circular/oval structures resembling sporangia or oogonium characteristic of water molds (F)
- Round to oval spore-bearing sporangia characteristic of var. hyphae-producing organisms (G).
- Abundant germ tube structures (bacteria, protists, or yeasts, not shown).
Despite the abundance of Morgellons-related particles in hot water tank samples, it is apparent that the shower head acts as a “sieve” filtering out all but the smallest particles. It is likely that the long, pigmented hyphae-like fragments found in shower water are related to the larger fibers or filaments present in hot tank water. Unless the larger filaments in the hot tank remain viable under thermophilic conditions, it is likely that temperatures above 60oC combined with agitated water tank conditions results in the fragmentation of the original fibers and the dispersal of resting or resistant stages of filament-producing organisms. Upon release of these structures into a more conducive growth environment (through the shower head), filaments may then be able to resume normal growth. The other structures, such as the “clear crystals”, slime-encapsulated algal cells, are present due to their ecological association with these filamentous microorganisms, and/or may be juvenile forms of the larger filament. The microscopic structures described above may play important roles in the origin of the particle types associated with Morgellons and should not be overlooked.
None of these microscopic structures are found in cold water samples or in samples collected after the hot water tank and pipes had been replaced. These samples are considered to be free of “contaminants”. The structures described above are considered to represent the primary structures indicative of “contaminated” water conditions for kitchen, tub, and shower water. Organisms tentatively identified with these structures are considered common soil/freshwater inhabitants and are considered to have originated from town water, at the source, or during transport through the water mains where soils come in contact with flowing water.
Phase I samples were cultured for bacterial and fungi/yeast populations using selective media to further elucidate the types of organisms present in the water samples before and after pipe replacement.
Culture results for total coliform bacteria, heterotrophic plate counts, and fungi/yeast counts are presented in (Table 3), for all Phase I water samples. Total coliform populations are commonly used to assess the water quality of drinking water. Their presence is used as an indicator to the potential presence of other pathogenic microorganisms known to be detrimental to human health if ingested (1). Results show that total coliform bacteria occur only in hot water shower samples before tank and pipe replacement, indicating that water from this source could have posed a health concern if consumed. Hot water tank samples do not contain total coliform populations, most likely because elevated temperatures prevented growth of these non-spore-forming microbes. Their presence in hot shower samples, however, indicates that biofilm communities coating the pipes protect the bacterial population from excessive heat. Shower pipes were reportedly the oldest original plumping remaining on the property. Upon pipe replacement, hot shower samples no longer contain coliform bacteria.
Heterotrophic bacterial cultures confirm the presence of heavy biofilm growth in the plumbing, particularly before tank and pipe replacement ((Table 3). Cultures containing confluent or spreading bacterial growth and/or TNTC populations indicate that slime-producing biofilms have colonized tank and pipe surfaces. It was noted that the membrane filters containing confluent growth are stained blue, and may be attributed to the production of a soluble blue pigment, indochrome, that is produced by streptomycete populations (2).
Although normal biofilm microflora are present in all water systems, where a complex community of surface-associated microbes are enclosed within a polysaccaride slime matrix, incidental “contamination” or colonization of biofilms by pathogenic organisms is recognized as an important concern in human health (4). Biofilms have been increasingly recognized as sources of opportunistic pathogens, particularly as causative agents of nosocomial (hospital-related) infections. Biofilms have been found to be involved in a wide variety of chronic opportunistic infections of mucosal and systemic sites (sinusitis, urinary tract infections, middle-ear infections, dental plaque, gingivitis, endocarditis, infections of cystic fibrosis). The persistence of many infections has been shown to be related to enhanced antibiotic production by populations to ensure survival in a highly competitive environment. This in turn has lead to an alarming increase in population resistance to antimicrobial agents that make infections difficult to treat.
As shown in Figure 5, antibiotic-producing actinomyces populations are found to occur in hot water tank populations. A clearing (inhibition) zone surrounding the antibiotic-producing population is present and confirms that the hot water tank is the source of potentially harmful biofilm organisms. Microscope slide mounts of the antibiotic-producing population show a growth morphology consistent with actinomyces, whereas the surrounding population is consistent with gliding bacteria.
Slime-producing, filamentous forms that have been tentatively identified in hot water tank and shower samples include: myxobacteria, cytophagales, cyanobacteria, and root-associated nitrogen fixing bacteria (3, 5,6). An important non-filament producing microorganism that is widely known to occur in biofilms is the human pathogen Pseudomonas aeruginosa and should not be overlooked in considering possible causative agents involved in Morgellons disease.
As pipes are replaced (Table 3), fast growing, confluent biofilm populations are eliminated, allowing slower-growing populations to develop. Clear, punctiform (pin head-sized) colonies (i.e. rhizosphere bacteria) and brightly-colored yellow and orange colonies (i.e. myxobacteria) can now be discerned, and are considered to represent the normal composition of microflora associated with water entering the property.
Prior to tank and pipe replacement fungi/yeast water sample populations consisted of an abundance (TNTC) of fast growing, pasty, tan-colored yeast colonies that coated the membrane filters. It is likely that mucoidal colonies of actinomyces bacteria also occured in these cultures as they are able to degrade the complex substrates that make the Sabs media fungi/yeast-selective. Cultures have been archived for further study, particularly with regard to the presence of pathogenic Candidia species, also a known component of biofilms (7). Filamentous fungal colonies were isolated only from hot water tank samples and included 1 type of dematiaceous (dark pigmentation), and 3 light tan to olive-colored types. These have also been archived to assess the potential presence of fungal dermatophytes.
Following pipe replacement, water sample cultures, with the exception of the cold kitchen sample, show that fungi/yeast populations have been eliminated. It is likely that frequent food-related contaminants, specifically yeast, were present in this post- replacement kitchen sample.
In addition to cultures grown on selective agar media, populations requiring minimal nutrient conditions were encouraged to grow on moistened membrane filters (100 ml to 500 ml) from hot water tank samples. Slow-growing protists (slime molds, water molds) and fungi (zygomycetes) were observed at 8 weeks (Figure 6). Diagnostic features used for tentative identification include: slime mold structures (plasmodia, coiled spore-bearing capillitia, stalked sporangia), water mold structures (cobweb-like aseptate hyphae, oogonia, antheridium, fertilization tubes, sporangia), and zygomycete fungi/yeast structures (germ tubes, zygospores, sporangiophores, sporangia, somatic hyphae, and rhizoids).
Microscope slides were also re-examined after 8 weeks without prior moistening. Of particular note was the colonization of rounded fibers by filamanetous, slime-producing gliding bacteria (Figure 7). As a result of the low nutrient and dry conditions of the glass slide, gliding motility was facilitated for these organisms and explains why growth was not previously observed in freshly prepared wet mounts. The lack of fruiting structures indicates that the colonies are most likely members of the genus Cytophagales, a common soil/water inhabitant capable of degrading chitin and cellulose (3).
Unpublished fiber data suggest that the composition of Morgellons-related body fibers consist of cellulose. However, further studies are needed to more fully characterize the composition(s) of the various Morgellons-related fibers present. The delicate thread-like morphology of these fibers often makes it difficult to isolate individual strands without damage. During Phase I study it was noted that special care is needed to prevent contamination during recovery of Morgellons-related fibers. Airborne fibers were found to be problematic, particularly with UV luminescent fibers from clothing and plastic sources. In addition to contaminant concerns, fiber studies need to carefully distinguish between dermal sources, including skin depth sampled, and environmental sources (water or air born fibers).
A preliminary investigation to determine whether fibers recovered from the hot water tank were viable and could grow under heterotrophic, aerobic conditions was conducted. Black, green, and red fibers from hot water tank samples were placed on the surface of a pour plate containing tryptic soy agar and incubated at 35oC for 6 weeks. The plate was examined daily to observe progressive changes (Figure 8).
Initial growth consisted of the formation of clear “crystals”, encapsulated algal cells, studded throughout the agar (some 6-sided) which coincided with the formation of greenish pigmentation throughout the agar substrate. Following algal growth, the agar surface was rapidly covered with confluent gray/brown bacterial growth. Conspicuous raised circular ridge formation was followed by the development of brown/black clusters of filaments inside the circles (Figure 8A and Figure 8C). These features are consistent with actinomycete morphology and development (8). Actinomyces are known to feed on living algae (and other organisms) and are often found in association with algae mats.
Translucent, green filaments up to 2 mm in length were also noted after 2 weeks. A wet slide mount revealed the presence of a blue-green UV luminescent fiber, characteristic of blue-green algae or cyanobacteria that produce phycocyanin, a blue, soluble luminescent pigment. None of the original fibers displayed longitudinal growth suggesting that fibers are sterile or that growth conditions were not adequate to facilitate further growth. Colonization of the agar by algae and bacteria is likely to have been initiated by microbes that had adhered to the surfaces of the introduced fibers.
Of particular note was a clearing zone observed at 10 days surrounding an introduced red fiber (Figure 8B). This inhibition zone confirms that antibiotic and/or toxin producing microbes are present as shown in previous hot water tank biofilm cultures (Figure 5).
At 4 weeks, actinomycete cyst formation occurred on the agar surface (with white, felty colony growth surrounding the cyst. Numerous translucent fibers were also present (Figure 8C and Figure 8E). Final observations at 6 weeks include the formation of abundant clear, rectangular “crystals” consisting of sheathed filaments of cyanobacteria containing typical elongated akinete cells or spores (Figure 8D). It was noted that the black fiber introduced onto the agar surface had turned red, perhaps as a result of surface bacteria dispersing into the agar over time, revealing the fiber’s “true” color (Figure 8F).
The presence of cyanobacteria, algae, and actinomyces in this fiber culture is consistent with observations of microscope slide mounts for hot water tank and shower samples.
Membrane filters containing fibers and other Morgellons-related particles have been archived under sterile conditions. Water sample concentrates and selected cultures (slants) have been refrigerated for further study. All microscope slide mounts have been secured under sterile conditions. (Table 4), lists all archived samples from the Phase I study.
A summary of the tentatively-identified microorganisms that were observed during this study is presented in (Table 5). These are considered to be the primary organisms of interest for further Phase II Morgellons-related studies. There are three groups of filament-producing organisms that occur in Phase I water samples: bacteria, protists, and fungi/yeasts. Selected characteristics for each group were reviewed to provide an overview of their potential ecological significance in relation to each other and to their origin. Characteristics selected for review are based on the following Morgellons-related considerations:
- terrestrial vs. aquatic predominance (motility and dispersal)
- reported symptoms of itching, crawling, burning (motility, toxins)
- filamentous structures and morphology (growth habit)
- capsule-like particles (resting stages, reproductive structures, parasitology)
- black-flake particles (resting stages, coatings or films from secondary metabolites)
- UV and non-UV pigmentation
- stellate-structures (hyphae morphology, germination morphology)
- cell wall composition (cellulose vs.chitin)
- nutrient requirements (ability to degrade various substrates)
- ecology (where they are found, i.e. stagnant water conditions, and what role they play)
- human health (known reported diseases)
Based on these characteristics it is evident that most of the organisms of interest would flourish under stagnant or calm hot water tank conditions. Aquatic organisms would reside in the water column and include the water molds, algae, and chytrids. Biofilm-related populations associated with the tank walls would include the slime-producing gliding bacteria, cyanobacteria, and nitrogen-fixing bacteria. Actinomyces, present in both aquatic and terrestrial (tank wall) habitats, would be abundant, feeding on algae and other detritus. These are known not only for their unique keratin degrading abilities (skin and hair), but are also for their ability to degrade many other substrates that other microorganisms find difficult or are unable to degrade. Antibiotic production also gives them an added advantage as successful competitors.
Based on the limited abundance of filamentous varieties of “true” fungi (cell walls composed of chitin) isolated from Phase I samples, and that Morgellons-related fibers consist of aseptate rather than septate varieties, it is probable that fungi are limited to the more primitive zygomycetes (pin molds). The occurrence of yeast in Phase I cultures are of importance as certain species are known to cause serious human diseases, particularly the dark colored, dimorphic varieties that alternate between hyphae-producing and budding life-cycles.
The organisms listed in (Table 5), constitute an interrelated ecosystem, where some play roles as symbiants (cyanobacteria, protists, zygomycetes) and others as predators (gliding bacteria, actinomycetes, chytrids, protists). Most of these organisms are primarily soil inhabitants that were transported from their natural soil or terrestrial habitat into an aquatic water tank environment. The incidental transport of aquatic species from wet soils into an “artificial” environment where “natural” components of a fresh water system are lacking (and, similarly, where terrestrial components are missing for the soil organisms) means that organisms adapted to their new environment. The blue-green algae (cyanobacteria) and green/brown algae occupy the lowest tier in the food web as “producers” (photosynthetic plant matter), followed by the “consumers” that graze on algae and living microorganism, and lastly by the “decomposers” that breakdown the decaying organic matter. The lack of sunlight stimulated the formation of resting stages for photosynthetic algae, those species able to grow under darkness became dominant. Other organisms would also resort to resting stages (spores, cysts) as nutrients were consumed.
A notable aquatic component that is lacking in this “artificial” closed (tank) system is other freshwater grazers such as zooplankton that are known to feed on chytrids. An abundance of chytrids may have occurred under stagnant tank conditions given the lack of this important consumer. Chytrid populations are of particular interest because they produce aseptate filaments similar to those described for Morgellons-related fibers. In addition to cellulose, they are also able to degrade keratin (skin and hair), an unusual trait shared by only one other aseptate hyphae-producing organism of interest, the actinomyces. Both are parasitic to another common tank inhabitant, notably the blue-green bacteria and the algae. It is proposed that the ecological relationships between these three groups of organisms of interest play a dominant role in the progression of Morgellons disease:
- cyanobacteria and algae (producers)
- actinomyces and chytrids (cellulose and keratin consumers)
- slime-producing bacteria and slime molds (algae decomosers)
Initial human exposure to hot water flowing from an environment that contains elevated populations of algae, actinomyces, and chytrids, and where surfaces are heavily coated with biofilm microflora could result in a range of dermal contact symptoms. The severity and longevity of these symptoms would depend on the relative proportions of cellulose- and keratin- degrading organisms, as well as the duration of dermal exposure. Upon release from the shower head, algal cells would adhere to an individual’s skin surface, followed by colonization of the algae by cellulose degrading organisms (algae plant pathogens), including the incidental colonization of the skin surface by species capable of degrading keratin (notably actinomyces and chyrtids). Plant pathogens are known to cause human diseases precisely in this way, where substrate colonization is incidental, and they establish themselves as an opportunistic pathogen in humans (9, 10). Dead algae are in turn colonized by alga decomposers, thus completing the food cycle.
The progression of infection proposed above (aquatic) could also be envisioned for aerial dust rich in algae and filamentous cyanobacteria. Airborne algae in dust is well known, especially in association with soil algal “blooms”. Actinomyces and chytrids (pine pollen parasites) are also reported to occur in aerial dust. It is interesting to note that although aerial dust samples can contain particles that are to be composed of a particular composition, such as cellulose, the composition of the actual disease-causing skin fibers may not coincide. Studies that focus on cell wall composition of Morgellons-related fibers should use fibers specifically collected from skin samples, and should not be confused with fibers from water or aerial dust sources.
Mild to moderate chronic skin conditions, such as dryness and/or rashes, could occur if exposure to water (and/or air born dust) rich in cellulose-containing organisms (algae and/or molds) was temporary or a single event. Skin colonization by opportunistic pathogens (actinomyces and chytrids) would therefore be minimal. More severe and chronic symptoms, however, could predominate if dermal contact were prolonged. Long-term colonization of the skin surface would occur as long as a cellulose-based food source (algae) was available. The importance of minimizing dermal contact by identifying and eliminating the source of “contaminating” particles is critical in promoting the successful recovery from this disease. The age of infected individuals, as well as many other variables (types and proportions of colonizing organisms), is also likely to affect the types of, and severity of, symptoms experienced by individuals.
Although the actinomyces and chytids in the tank water are the primary degraders of cellulose and keratin, slime-producing (gliding and root-associated) bacteria and slime molds may also play a significant role as algae decomposers, and may be important in the skin ecology of Morgellons disease. The gliding bacteria, particularly Herpetosiphon giganteus (Flexibacter), is known to occur in soils and fresh water, and it produces “occasional” hyphae-like protruberances consisting of interwoven filaments (more than 5 mm long) that rise from substrate surfaces (3, 11). Crawling motility, a common trait of slime molds and chytrids, is also a common feature of gliding bacteria, and it may be related to the crawling sensations reported by Morgellons sufferers (12). Musty odors are often reported by individuals with Morgellons disease, a common characteristic of both the actimomyces and gliding bacteria.
Other possible human health problems associated with dermal and/or aerial contact with the primary organisms of concern listed in (Table 5), are related to toxin-producing cyanobacteria and algae (cytotoxins, endotoxins, neurotoxins and hepatoxins), pathogenic microbes associated with biofilms (i.e. Pseudomonas aerigonosa), and yeast popoulations (Candidia albicans).
Based on the characteristics described for each organism of interest in Table 5, the following conclusions can be made with regard to the possible role each Morgellons-related particle may play.
10.1 Ribbon-like fibers
Microscopic observations show that ribbon-like fibers occur in various sizes and colors (green, greenish yellow, greenish blue, blue, black, clear/gray, and red), with the clear fibers often displaying a blue luminescence under UV light. Individual fibers consist of both rounded and ribbon-like morphologies, suggesting that ribbon-like morphologies may be indicative of fibers that have been degraded over time. Shedding or peeling of the outer fiber sheaths (ribbon-like remnants) of rounded varieties confirm this suspicion. The fiber’s color is likely related to the organisms internal, protoplasm, or cell wall pigmentation and/or pigmentation from organisms that are able to degrade or graze on filamentous organism’s surface. Many bacterial populations, especially the gliding bacteria and actinomyces, are well known for their brightly colored spores and vegetative cells, some of which luminesce under UV light (3).
Of the organisms listed in (Table 5), the only reference found for hyphae that consist of a ribbon-like morphology, are the aerial hyphae of the zygomycete fungi. Although not a common hot water tank inhabitant, it is possible that they constitute a small proportion of the total ribbon-like varieties present.
Rounded filaments are produced, to some extent, by all of the organisms of interest in (Table 5). Gliding bacterial cells can form long, filamentous colonies surrounded by an outer sheath of slime. Their gliding motility is due to their ability to produce this mucilage substance. As a result, many filament sheaths have a jelly-like appearance, especially the cyanobacteria. Colony growth of the myxobacteria and cytophagales bacteria in agar is characterized by spreading (confluent) growth and a pasty, wet appearance. These sheaths occur as distinct filaments (or fibers) that can remain intact for extended periods of time. Cyanobacteria produce greenish blue, unbranched filaments known as trichomes that have a distinct banded or bead-like texture with air pockets called vacuoles. Clear to grayish-colored filaments recovered from hot water tank samples consist of typical textures indicative of these organisms (Figure 1, particle # 2, type-B) although they are no longer pigmented due to fragmentation. Living specimens showing this banded texture were microscopically examined from fresh soils, and these exhibit a greenish yellow UV luminescence. It is likely that dead specimens from the hot water tank no longer contained the soluble UV pigment (phycocyanin and phycoerythrin) inside the filament’s protoplasm, confirming that UV luminescence for these fiber types can be “temporary” due to death and damage. Many other pigments reside in the cell wall rather than internally, such as the chlorophylls, and may also account for a fiber’s color, especially for the ribbon-like fibers, where the protoplasm may no longer be present.
Actinomyces have a remarkable fungus-like ability to produce spore-bearing hyphae that consist of long filaments composed of individual cells. Both spores and hyphae can be brightly pigmented and can occur in many colors that can be used to differentiate between species (3). Species displaying blue and red pigmentation occur in abundance in both hot water tank and shower samples before pipe replacement.
Nitrogen-fixing bacteria occur near the root zones or rhizosphere of plants and are important soil inhabitants, most commonly Rhizobium, Azotobacter, and Agrobacter species. Most rhizosphere organisms are not widely dispersed in soils apart from root zones and do not form extensive fibrous colonies (3). They are of special note, however, due to the brown to black pigments produced upon aging that may be of interest with regard to Morgellons-related black flakes. They can form conspicuous pigmented (red, pink) rod-like cells up to 5 um in length and may be one of the commonly rod-shaped structures noted in the microscope slide mounts (Figure 4). Cultures also show evidence of their presence, as they prefer anaerobic conditions and often form water-clear colonies on the bottom rather than the top surface of the agar. Some species have been reported to produce long spiral microfibrils (Rhizobium sp.). UV pigment production has also been reported for these organisms.
Protists produce many types of filaments that consist of cellulose. Plasmodial slime molds form fiber-like stalks that contain spore-bearing fruiting bodies. Spores inside this fruiting body are held loosely by capillitia and elator filaments that spring open to release the spores (13). Capillitia filaments have a “twisted”, ropy, or scaly texture and are frequently observed in water water tank samples (Figure 1, type-C fibers). These, as well as stalk filaments, occur in many colors, often with a single filament grading from one color into another. Oomycete water molds form a cobweb-like thallus composed of colorless, aseptate hyphae. Chytridomycetes or chytrids also form colorless hyphae, and some species form inside their host and then release zoospores through a long exit tube.
Unicellular green and brown algae cells are present in abundance in both hot water tank and shower samples before the pipes were replaced, and they are considered to play an important role in Morgellons-related symptoms. Filamentous forms may account for the red UV luminescent fibers present in hot water tank samples, however more information is needed to assess their significance in water samples.
All of the filaments or fibers described in this section are likely sources for the numerous types of fibers recovered during this study. Body fibers from individuals with Morgellons likely originated from both algae dermal contact (filamentous cyanobacteria), as well as from organisms colonizing the alga (cellulose degraders) and/or the skin surface (keratin degraders).
These particles are identified as parasitized pine pollen grains. Reports describe chytrid parasites that feed solely on pine pollen. Freshly collected pine pollen grains were microscopically examined and moistened with local lake water. Colonization of pollen grains by parasitic aquatic chyrids revealed that the central pollen regions were consumed or “deflated” leaving behind a capsule-like morphology identical to those recovered in hot water tank samples (Figure 9). Further ecological evidence confirming the likely presence of chyrids in the hot water tank samples is the abundance of algae. Chytrids are parasites of algae and consume them during growth in the hot water tank.
A simple pine pollen test is envisioned where archived water samples are inoculated with fresh pine pollen to determine if chytrids are present in samples before tank and pipe replacement (Phase II study).
Two distinct stellate morphologies are noted in hot water tank samples, a ribbon-like form and a rounded form (Figure 1). Note that similar forms are described for Morgellons-related fibers (above). These stellate particles, however, are not brightly pigmented. The rounded stellate forms are usually clear, with some showing a faint green interior color, whereas the ribbon forms are light brown in color. The rounded forms are remarkably similar to plant leaf hairs, and may account for the green (chlorophyll) pigmentation.
Another possible origin, due to their common association with pine pollen (also found to occur with freshly collected pollen), is from an organism that produces short hyphae that emanate from a single point, known as a monocentric thallus. Some species of parasitic chytrids produce a radiating thallus from a bulbous “holdfast”. Another monocentric thallus- producing organism includes the oomycete water molds (Rhipidiales sp.), and these would be worth investigating as a possible explanation for why there is multiple stellate forms present in samples (6). Ribbon forms may be degenerative forms of these organisms, or may simply be fungal spores in the process of germinating.
Worm-like particles are noted with either tapered or square ends. Microscopic observations show that the square-ended worm forms originate from fragmented stellate-shaped particles. Other worm forms do not appear fragmented. None have structures suggestive of invertebrate morphology. All have a jelly-like appearance, which suggest a possible algal origin.
There are several references to invertebrate parasitic associations by the organisms of interest listed in (Table 5) and include the water molds (nematodes, insect larvae), and the chytrids (nematodes, aquatic larvae). It is likely that open lesions may harbor secondary colonization by invertebrates. Further studies of lesion tissue samples are required to address the occurrence of possible invertebrates.
Black specks are widely reported to be associated with Morgellons disease. Microscopic observations show that they are often colonization by microorganisms, which is to be expected since microbes mainly occur on surfaces that can provide nutrients rather than as free floating cells.
Black-grained mycetoma is a well-known skin disease that is accompanied by lesions that discharge grains composed of short fungal hyphae. White or red grains consist of fine filaments indicative of actinomyces.
Considering that aseptate hyphae are the dominant fiber morphology in hot water tank samples, black grains or specks, assuming that they are biological in origin, are likely to originate from dark pigmented exudates or secondary metabolites produced by bacteria or protists (Table 5). Another possibility that warrants further investigation are the hard, black crusts (resting stages) produced by slime molds called sclerotia. Sclerotia are formed as the plasmodium slime mass becomes desiccated, providing a resting stage for these organisms to survive periods of dryness. Some Morgellons sufferers have reported the formation of black flecks 6-7 hours after bathing, and may be related to sclerotia formation as the skin surface dries. Further studies of their microscopic morphology (filamentous or grainy) are needed.
In conclusion, it is apparent from this study that the variability in Morgellons-related symptoms is likely to the reflect variability of the causative agents or organisms that promote skin and/or systemic symptoms. Appreciation of the ecological complexity and diversity of the causative agents is needed to further differentiate Morgellons-related symptoms into definitive and distinct (sub) categories. Symptoms could be regarded as distinct types, for example, toxin-generated systemic/neurological symptoms (algae), and fibrous growth-related symptoms (filamentous keratin degrading or associated organisms). The presence of antibiotic and/or toxin producing organisms may account for chronic, long-term symptoms associated with this disease.
Based on the abundance of algae and cyanobacteria (blue-gree algae) populations in contaminated water samples, the relationship of either water or airborne environments rich in these populations suggest that conditions promoting the development of algal “blooms” are important geographical markers associated with this disease. Aquatic and soil habitats where nitrogen and phosphorus concentrations become elevated due to waste discharge or wetland disturbance by draining or drying, are known to cause algae “blooms” as excess nutrients normally used by wetland plants instead are consumed by these organisms. Future studies should include a component related to the geographic distribution of affected individuals in relation to climate.
Based on the findings of this study, Phase II investigations will include, 1) further investigation into the origin of the remaining 5 Morgellons-related particles, 2) the development of water tests to readily determine if Morgellons-related contaminants (or organisms) are present (stains and selective media), and 3) further identification, at least to the genus level, of organisms of interest recovered to-date. Further ecological investigation into the possible role that algae, blue-green algae, and keratin degrading organisms (chytrids and actinomyces) may play as causative agents in this disease are also planned.
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