Placebo-resistant gut bacteria: Akkermansia muciniphila spp. and Familial Mediterranean fever disease.

Introduction: Despite numerous investigations into the impact of drugs/probiotics on the gut microbiota composition in Familial Mediterranean Fever (FMF) patients, the question as to whether there exists a significant bacterial diversity(ies) independent of the placebo effect that can be reliably considered in clinical and nutritional trials remains unresolved. Methods: This study represents the in augural analysis of the placebo's influence on the gut microbiota of both healthy individuals and FMF afflicted men, utilizing previously collected data from PhyloChip™ DNA microarray experiments. A total of 15 healthy and 15 FMF male volunteers, aged 18 to 50, participated in this partially randomized placebo trial, which is accessible through the GEO Series accession number GSE111835. Results and Discussion: Key findings from current investigations include i. the anticipated divergence in gut bacteria resistance to placebo between healthy and FMF individuals, ii. the minor impact of placebo on gut bacterial diversities in healthy individuals, with Enterobacteriaceae diversities identified as placebo-resistant among "healthy" gut bacteria, and iii. the comprehensive influence of placebo on all bacterial phyla in the gut microbiome of FMF patients, extending to nearly all bacterial genera, except for the resilience of gut Akkermansia muciniphila spp. to placebo in FMF patients. This study underscores the susceptibility of Faecalibacterium, Blautia, and Clostridium genera to placebo. Consequently, this investigation holds significance for the proper design of placebo-controlled trials and establishes a foundation for further exploration of the gut-brain axis. Furthermore, it contributes valuable insights to discussions regarding proposals for probiotic therapies, particularly focusing on Faecalibacterium spp., Blautia spp., and Clostridium spp.

Gut Microbiota between Environment and Genetic Background in Familial Mediterranean Fever (FMF).

The gastrointestinal tract hosts the natural reservoir of microbiota since birth. The microbiota includes various bacteria that establish a progressively mutual relationship with the host. Of note, the composition of gut microbiota is rather individual-specific and, normally, depends on both the host genotype and environmental factors. The study of the bacterial profile in the gut demonstrates that dominant and minor phyla are present in the gastrointestinal tract with bacterial density gradually increasing in oro-aboral direction. The cross-talk between bacteria and host within the gut strongly contributes to the host metabolism, to structural and protective functions. Dysbiosis can develop following aging, diseases, inflammatory status, and antibiotic therapy. Growing evidences show a possible link between the microbiota and Familial Mediterranean Fever (FMF), through a shift of the relative abundance in microbial species. To which extent such perturbations of the microbiota are relevant in driving the phenotypic manifestations of FMF with respect to genetic background, remains to be further investigated.

Specific changes in faecal microbiota are associated with familial Mediterranean fever.

OBJECTIVES: Familial Mediterranean fever (FMF) can be complicated by AA amyloidosis (AAA), though it remains unclear why only some patients develop amyloidosis. We examined the gut microbiota composition and inflammatory markers in patients with FMF complicated or not by AAA. METHODS: We analysed the gut microbiota of 34 patients with FMF without AAA, 7 patients with FMF with AAA, 19 patients with AAA of another origin, and 26 controls using 16S ribosomal RNA gene sequencing with the Illumina MiSeq platform. Associations between bacterial taxa and clinical phenotypes were evaluated using multivariate association with linear models statistical method. Blood levels of interleukin (IL)-1ß, IL-6, tumour necrosis factor-α and adipokines were assessed by ELISA; indoleamine 2,3-dioxygenase (IDO) activity was determined by high-performance liquid chromatography. RESULTS: Compared with healthy subjects, specific changes in faecal microbiota were observed in FMF and AAA groups. Several operational taxonomic units (OTUs) were associated with FMF. Moreover, two OTUs were over-represented in FMF-related AAA compared with FMF without AAA. Additionally, higher adiponectin levels and IDO activity were observed in FMF-related AAA compared with FMF without AAA (p<0.05). CONCLUSION: The presence of specific changes in faecal microbiota in FMF and in FMF-related AAA suggests that intestinal microorganisms may play a role in the pathogenesis of these diseases. These findings may offer an opportunity to use techniques for gut microbiota manipulation.

Lactobacillus acidophilus INMIA 9602 Er-2 strain 317/402 probiotic regulates growth of commensal Escherichia coli in gut microbiota of familial Mediterranean fever disease subjects.

Previously, we reported a positive effect the probiotic formulation, Lactobacillus acidophilus INMIA 9602 Er-2 strain 317/402 (Narine strain), had on the blood characteristics of patients with familial Mediterranean fever disease (FMF). The aim of this investigation was to evaluate the effect of the Narine probiotic on growth characteristics in the predominant commensal Escherichia coli isolates from the gut microbiota in FMF-positive study participants. Bacterial growth of 192 prevalent commensal E. coli isolates found in the volunteer participants' guts was evaluated using Verhulst's logistic function. This study showed that the duration of the preparatory growth phase for the E. coli isolates collected from FMF-positive volunteers was significantly shorter, whereas the duration of the logarithmic growth phase was significantly longer (P < 0·03) than that of the isolates collected from healthy participants. The Narine probiotic formulation caused a significant extension (P < 0·001) of the preparatory growth phase in the commensal E. coli isolated from FMF subjects a month after the Narine probiotic administration was terminated. The data suggest that the mathematical model characterizes the growth of commensal E. coli isolates from FMF-positive participants and it can be useful in a decision-making process on the practical use of probiotics during FMF. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to demonstrate the effects of Narine, containing the probiotic Lactobacillus acidophilus, on the growth of gut commensal Escherichia coli from study participants with familial Mediterranean fever disease (FMF). Verhulst's logistic function was demonstrated to act as a possible tool for the evaluation and quantification of effects produced by the probiotic formulation in FMF participants.

Small Intestinal Bacterial Overgrowth Affects the Responsiveness to Colchicine in Familial Mediterranean Fever.

OBJECTIVE: Familial Mediterranean fever (FMF) is an autosomal recessive disease due to a MEFV gene mutation. Since Helicobacter pylori infection has been described to increase the severity and frequency of FMF attacks, we evaluate if overgrowth of small intestinal bacterial (SIBO), associated with a release of bacterial products, can affect the response to colchicine in FMF patients poorly responsive to colchicine. METHODS: We revised our Periodic Fever Centre database to detect FMF patients who were poorly responsive to colchicine, without a well-defined cause of drug resistance. They were evaluated for SIBO presence, then treated with decontamination therapy. RESULTS: Among 223 FMF patients, 49 subjects show colchicine resistance, and no other known causes of colchicine unresponsiveness has been found in 25 patients. All 25 patients underwent glucose breath test; 20 (80%) of them were positive, thus affected by SIBO. After a successful decontamination treatment, 11 patients (55%) did not show FMF attacks during the following three months (p < 0.01), while 9 of them revealed a significant reduction of the number of attacks compared to three months before (p < 0.01). CONCLUSION: The SIBO eradication improves laboratory and clinical features of FMF patients. Thus, patients with unresponsiveness to colchicine treatment should be investigated for SIBO.

[ANTIBIOTIC RESISTANCE OF ESCHERICHIA COLI OF THE INTESTINAL MICROBIOTA IN PATIENTS WITH FAMILIAL MEDITERRANEAN FEVER].

We used clinical bacteriological analysis and high-density DNA-microchips (PhyloChip) to study the quality and the quantity of commensal bacteria of the genus Escherichia in patients with familial mediterraneanfever (periodic disease). The intestinal microbiota of these patients contained a large number of operational taxonomic units of these bacteria. The study of antibiotic resistance of Escherichia coli from the intestinal microbiota in patients with familial mediterranean fever reveald a large number of resistant and multiresistant isolates. Therapy with commercial probiotic Narine (Vitamax-E, Armenia) reduced the number of operational taxonomic units of commensal bacteria and the frequency of multiresistant isiolates. The mechanism of action of Narine probiotic on intestinal bacteria and their resistance to antibiotics is discussed

Digoxigenin-labeled in situ hybridization for the detection of Streptococcus suis DNA in polyserositis and a comparison with biotinylated in situ hybridization.

The objective of this study was to develop digoxigenin-labeled in situ hybridization (ISH) for the detection of Streptococcus suis in naturally infected pigs with polyserositis and to compare it with biotinylated ISH. Digoxigenin-labeled hybridization signals for S. suis were observed in cells that had infiltrated the fibrous polyserositis and microcolonies in the blood vessels. Mock hybridization showed no hybridization signals for endogenous digoxigenin. Biotinylated hybridization signals for S. suis were observed in cells that had infiltrated the fibrous polyserositis. However, similar hybridization signals were also observed in the fibrous inflammatory area using mock hybridization for endogenous biotin. The present study demonstrated that digoxigenin-labeled ISH is a valuable diagnostic tool for specific detection of S. suis in polyserositic tissues without nonspecific reactions compared with biotinylated ISH.

Program of vaccination and antibiotic treatment to control polyserositis caused by Haemophilus parasuis under field conditions.

The present study investigated the effects of vaccinating sows and piglets or piglets alone against Haemophilus parasuis on the prevalence of H. parasuis in nasal swabs, on the humoral and cellular immune responses, and on the production parameters of piglets at 3 Korean farms with a clinical history of polyserositis caused by H. parasuis. Piglets born to vaccinated or non-vaccinated sows were subdivided into 3 groups: vaccinated sows and vaccinated pigs (VS-VP), non-vaccinated sows and vaccinated pigs (NVS-VP), and non-vaccinated sows and non-vaccinated pigs (NVS-NVP). The proportion of piglets with positive nasal swabs was significantly lower (P < 0.05) in the vaccinated animals (VS-VP and NVS-VP groups) than in the non-vaccinated animals (NVS-NVP group) at 35 and 60 d of age at the 3 farms. The overall growth performance (from 7 to 60 d of age) of the vaccinated piglets was significantly better (P < 0.05) than that of the non-vaccinated piglets at the 3 farms. Piglets in the VS-VP group had significantly higher levels (P < 0.05) of H. parasuis-specific IgG antibodies, lymphocyte proliferation, and interferon-γ-secreting cells than piglets in the NVS-VP and NVS-NVP groups on days 1, 7, 21, 35, and 60 after birth at the 3 farms.

Ce projet visait à étudier les effets de la vaccination contre Haemophilus parasuis des truies et des porcelets ou des porcelets uniquement sur la prévalence d'H. parasuis dans des écouvillons nasaux, sur les réponses immunitaires humorale et cellulaire, et sur les paramètres de production des porcelets dans trois fermes coréennes avec une histoire de cas cliniques de polysérosites causés par H. parasuis. Les porcelets nés de truies vaccinées et non-vaccinées ont été répartis en trois groupes : truies vaccinées et porcelets vaccinés (VS-VP), truies non-vaccinées et porcelets vaccinés (NVS-VP), et truies non-vaccinées et porcelets non-vaccinés (NVS-NVP). La proportion de porcelets positifs pour H. parasuis à partir de l'écouvillon nasal était significativement plus faible (P < 0,05) chez les animaux vaccinés (groupes VS-VP et NVS-VP) que chez les animaux non-vaccinés (groupe NVS-NVP) à 35 et 60 jours d'âge sur les trois fermes. Sur les 3 fermes, les performances de croissance globales (de 7 à 60 jours d'âge) des porcelets vaccinés étaient significativement meilleures (P < 0,05) que celles des porcelets non-vaccinés. Sur les trois fermes, les porcelets du groupe VS-VP avaient des niveaux significativement plus élevés (P < 0,05) d'anticorps IgG spécifiques contre H. parasuis, de prolifération lymphocytaire, et de cellules secrétant de l'interféron-γ que les porcelets dans les groupes NVS-VP et NVS-NVP aux jours 1, 7, 21, 35, et 60 après la naissance.(Traduit par Docteur Serge Messier).

TLR2 and TLR4 polymorphisms in familial Mediterranean fever.

It has been suggested that MEVF mutations offer advantage against infections, including tuberculosis. Bearing in mind the central role of TLR-2 and TLR-4 in the recognition of pathogens, we conducted this study to examine whether the TLR2-R753Q, TLR4-D299G, TLR4-T399I common polymorphisms are associated with susceptibility to familial Mediterranean fever (FMF) or affect the course of the disease. A cohort of 169 FMF patients and 245 healthy bone marrow donors were enrolled in the study. FMF patients appeared with a significantly lower frequency of the TLR4-D299G mutated allele (3.2% vs 6.9%, p = 0.032). No association was observed with the other analyzed polymorphisms. Moreover, we found no association between polymorphisms and the frequency of attacks or the development of amyloidosis. Our results may reinforce the hypothesis that FMF patients display a better defense against pathogens, providing an additional mechanism and suggesting a positive selection advantage in the area of the Mediterranean basin.

Elevated systemic antibodies towards commensal gut microbiota in autoinflammatory condition.

BACKGROUND: Familial Mediterranean fever (FMF) is an autoinflammatory condition, which is characterized by acute, self-limiting episodes of fever and serositis and chronic subclinical inflammation in remission. Here we investigated the consequence of this condition on the level of systemic antibodies directed towards common intestinal bacteria. METHODOLOGY/PRINCIPAL FINDINGS: The level of systemic antibodies towards the antigens of Bacteroides, Parabacteroides, Escherichia, Enteroccocus and Lactobaccilus was measured by ELISA in FMF patients at various stages of the disease and in healthy controls. The difference between remission and attack was not significant. IgG antibodies against the antigens of Bacteroides, Parabacteroides, Escherichia and Enteroccocus were significantly increased in FMF compared to control while IgA levels were not significantly affected. Western blot analyses demonstrated the IgG reactivity against multiple antigens of commensal bacteria in FMF. Serological expression cloning was performed to identify these antigens. No single dominant antigen was identified; the response was generalized and directed against a variety of proteins from Bacteroides, Parabacteroides, Escherichia, and other gut commensals. CONCLUSIONS/SIGNIFICANCE: This autoinflammatory syndrome is characterized by the increased systemic reactivity against commensal gut microbiota. This is probably the consequence of hypersensitivity of the inflammasome in FMF that triggers the inflammation and contributes to the excessive translocation of bacteria and bacterial antigens through the gut barrier.

Predominant role of host genetics in controlling the composition of gut microbiota.

BACKGROUND: The human gastrointestinal tract is inhabited by a very diverse symbiotic microbiota, the composition of which depends on host genetics and the environment. Several studies suggested that the host genetics may influence the composition of gut microbiota but no genes involved in host control were proposed. We investigated the effects of the wild type and mutated alleles of the gene, which encodes the protein called pyrin, one of the regulators of innate immunity, on the composition of gut commensal bacteria. Mutations in MEFV lead to the autoinflammatory disorder, familial Mediterranean fever (FMF, MIM249100), which is characterized by recurrent self-resolving attacks of fever and polyserositis, with no clinical signs of disease in remission. METHODOLOGY/PRINCIPAL FINDINGS: A total of 19 FMF patients and eight healthy individuals were genotyped for mutations in the MEFV gene and gut bacterial diversity was assessed by sequencing 16S rRNA gene libraries and FISH analysis. These analyses demonstrated significant changes in bacterial community structure in FMF characterized by depletion of total numbers of bacteria, loss of diversity, and major shifts in bacterial populations within the Bacteroidetes, Firmicutes and Proteobacteria phyla in attack. In remission with no clinical signs of disease, bacterial diversity values were comparable with control but still, the bacterial composition was substantially deviant from the norm. Discriminant function analyses of gut bacterial diversity revealed highly specific, well-separated and distinct grouping, which depended on the allele carrier status of the host. CONCLUSIONS/SIGNIFICANCE: This is the first report that clearly establishes the link between the host genotype and the corresponding shifts in the gut microbiota (the latter confirmed by two independent techniques). It suggests that the host genetics is a key factor in host-microbe interaction determining a specific profile of commensal microbiota in the human gut.

Autoinflammatory syndromes and infections: pathogenetic and clinical implications.

The autoinflammatory syndromes are a group of disorders characterized by recurrent episodes of seemingly unprovoked inflammation without significant levels of autoantobodies and antigen specific T cells. Although a direct association between defective innate immune responses to bacterial components and these diseases has not been formally established, much ongoing research is aimed towards confirmation of that hypothesis. This article will review recent advances in the study of a subset of NOD-like receptors (NLRs), which control the activation of caspase-1 through the assembly of a large protein complex called inflammasome. Moreover, we will review recent progresses in understanding of a range of autoinflammatory conditions in humans.

Effect of Helicobacter pylori infection and eradication therapy on interleukin-6 levels in patients with Familial Mediterranean Fever.

It is being questioned if Helicobacter pylori infection, which causes a chronic inflammatory response, can increase the frequency and severity of attacks in patients with Familial Mediterranean Fever (FMF) and if the impact of inflammatory response can be diminished by eradication of the infection. To evaluate if there is difference in interleukin (IL)-6 levels of H. pylori-positive and -negative patients both before and during FMF attacks; if there is a change in IL-6 levels following successful eradication treatment; and if MEFV gene mutations have an effect on IL-6 levels. IL-6 levels were evaluated in 47 FMF patients before and during FMF attacks. Genetic testing to determine M694V, M694I, E148Q, V726V, M680I mutations was also performed in all patients. IL-6 levels were also determined after successful eradication of the infection in H. pylori-positive patients. IL-6 levels were compared in H. pylori-positive and -negative patients, and before and after eradication treatment in patients who cleared the infection. Number of patients in tested mutation groups was not enough to compare IL-6 levels in these groups. Thirty-four patients (73.9%) were H. pylori-positive. Before FMF attack there was no statistically significant difference in IL-6 levels of H. pylori-positive and -negative groups. IL-6 levels were significantly higher in both groups during the attacks than before the attacks (p < 0.05). There was a statistically significant decline in IL-6 levels both before and during FMF attacks, following eradication therapy in patients who cleared the infection (p < 0.05). In patients with homozygous M694V mutation, IL-6 levels before and during the FMF attacks were not significantly different in H. pylori-positive and -negative groups, despite a much lower level found in H. pylori-negative group (p > 0.05). Comparisons were not performed in other mutation groups because of small number of patients in each group. C-reactive protein (CRP) and fibrinogen levels were not significantly different between the groups (p > 0.05). We believe that the observation of IL-6 levels are lower both before and during FMF attacks both in H. pylori-negative patients and in patients who cleared the infection after eradication therapy is very important in the determination of the role of eradication of H. pylori on decreasing the frequency and severity of FMF attacks. As for today, the correlation between H. pylori infection and FMF seems unlikely; however, studies evaluating the interaction of cytokines in both diseases and their relations and roles will be needed to reach better conclusions.

[Study of volatile fatty acids in the blood of patients with periodic disease].

Microecological "failures" are an important pathogenetic factor of different diseases and, in the authors' opinion, periodic disease (PD) is one of them. PD is a recessive disease characterized by fever attacks and neutrophil-mediated serous inflammation. A genetic factor has been established to be responsible for half the cases of PD, the influence of non-hereditary factors, particularly a role of the host automicroflora in the genesis of an inflammatory process, has been little studied. The authors' early studies indicate that there are changes in the qualitative and quantitative composition of microbial molecules in the blood of patients with PD. The anaerobic bacterial metabolites that are volatile fatty acids (VFAs) represent biologically active substances that affect the growth of the microflora, on the one hand, and the host's immunological responsiveness, on the other. Out of VFAs, only is acetic acid detectable in small quantities in the blood of healthy individuals. The other VFAs, namely propionic, valeric, butyric, and caproic acids and their isomers, are absent. Gas chromatography was used for qualiitative and quantitative determination of the metabolites of anaerobic microorganisms in the blood of patients with PD (n = 13) during an attack and remission and in that of healthy volunteers (Armenians) (n = 5) of a control group from one Yerevan region. The blood samples from all the patients with PD displayed a significantly higher concentration of caproic acid while the latter was absent in the blood of the controls. This finding suggests that there is a specific shift in the structure of the microbiocenosis in patients with PD. It is conceivable that caproic acid plays a certain role in the pathogenesis of the disease under study. Further studies will deal with the association of some microbial molecules with the manifestation of an attack of PD, which may provide the key to the goal-oriented regulation of detected homeostatic disorders and to the management of the frequency of its attacks.

Is there any relationship between Chlamydophila pneumoniae infection and juvenile idiopathic arthritis?

The role of Chlamydophila pneumoniae in the development and exacerbation of juvenile idiopathic arthritis (JIA) was investigated. Blood samples were taken from 60 JIA patients during an active disease period and for 4 weeks after. Synovial fluid samples were obtained from 20 of the 60 patients. In addition, 22 patients with familial Mediterranean fever (FMF) during the active period and 35 healthy children were included in the study as control groups. Synovial fluid samples were also obtained from three children with FMF. IgG, IgM and IgA levels against C. pneumoniae in serum samples were studied by immunofluorescence and IgG antibody and PCR studies were performed for C. pneumoniae DNA in synovial fluid samples. Twenty-nine (48.3 %) patients with JIA, 18 (81.8 %) patients with FMF and 22 (62.8 %) healthy children were found to be pre-infected with C. pneumoniae. Pre-infection with C. pneumoniae among FMF patients was found to be significantly higher than among those with JIA. We did not find a significant difference between JIA patients and healthy children. Chronic C. pneumoniae infection was observed only in six JIA patients, one FMF patient and two healthy children. Synovial fluid antibodies were found at higher than 1/512-fold dilution in one JIA patient and four times higher than normal serum in three JIA patients. C. pneumoniae DNA was not detected in any synovial fluid sample from FMF or JIA patients by PCR. In conclusion, C. pneumoniae infection does not have a triggering or a progressive effect on the clinical situation in JIA aetiopathogenesis, as a result of a multifactorial aetiology. New, extensive and serial studies (especially PCR studies of synovial tissue) are needed in order to confirm the indirect results.

[The spectrum and level of low-weight molecular compounds of microbial origin in familial Mediterranean fever]. / Spektr i uroven' soderzhaniia nizkomolekuliarnykh soedinenii mikrobnogo proiskhozhdeniia pri periodicheskoi bolezni.

Familial Mediterranean fever (FMF) is a disease whose etiopathogenesis is not clarified yet. The infectious theory of FMF has not been confirmed either. Nevertheless, the involvement of microbes in the trigger of an inflammatory process cannot be excluded since today's well-known pathogenetic processes in FMF are to be directly related to the key cells of an inflammatory response. According to the existing concept, homeostasis of small molecules originating from microbes (SMOM) in healthy individuals is achieved due to adequate immune system function with the preserved biocenosis of a macroorganism whose disturbance with resultant immune shifts triggers a cascade of inflammatory reactions in the body. An attempt was taken to reveal the participation of microorganisms at the onset of an inflammation in case of FMF by using chromatographic mass spectrometry to detect chemical components of microorganisms and their vital activity products. The method allows one to screen a large number of microbial markers in a clinical sample. Pronounced impairments in the homeostasis of non-protein SMOM were found in the blood of examined patients with FMF (n = 16). There was a uniformity of deviations from the normal values in all the examinees in the episodes and episode-free periods. These qualitative and quantitative deviations basically differ from the direction of changes with other diseases (n 59) or healthy individuals (n = 18). All significant deviations affect non-traditional participants of an inflammatory process in the host. The similar microecological breakages in the human body and their consequences have not been earlier detected and investigated. The findings show it necessary to continue studies to receive an answer to the question as to whether the detected homeostatic features of SMOM in patients with FMF are primary or what is their role in the etiopathogenesis.

Familial Mediterranean fever. No role of Mycobacterium tuberculosis in ten patients.

BACKGROUND: Tuberculosis (TB) and Familial mediterranean fever (FMF) are two common diseases in our region, Turkey. Both share some properties in common: Both cause AA type amyloidosis and have association with some immunological abnormalities. Upon incidentally observing Mycobacterium tuberculosis in bone marrow biopsies of three patients with FMF in a previous study, we intended to elucidate this association prospectively. MATERIAL AND METHODS: In this study, we examined prospectively 10 FMF patients, 5 male and 5 female, with a median duration of 31 years disease activity. All were under colchicine therapy. They had no sign of renal involvement. The bone marrow biopsies of these patients were examined for the presence of M. tuberculosis by Polymerase chain reaction (PCR), BACTEC culture and pathological stains. Pathological examination was performed for the existence of granuloma and amyloid deposition by hematoxylin-eosin, Crystal Violet and Congo red stains. RESULTS: The examination of all bone marrow specimens by the mentioned methods suggest that Mycobacterium tuberculosis has no role in the ethiopathogenesis of FMF. Although the patients had a positive family history of 60% for tuberculosis and in 80% of them with positive tuberculin skin test. CONCLUSIONS: We concluded that although there seemed to be a kind of association between both diseases, this relationship is not via the direct existence of bacteria itself. Considering high family history and skin test positivity, one should look for the presence of autoimmune mechanisms under this suspicious relationship between tuberculosis and FMF. Also, this is the first study examined the state of amyloidosis in the bone marrow at an earlier stage of FMF without overt renal findings.