Repeated praziquantel treatments remodel the genetic and spatial landscape of schistosomiasis risk and transmission.

Repeated treatments with praziquantel reduce schistosomiasis prevalence and morbidity, but transmission persists and populations often recover within a few years. To identify factors associated with persistence, we surveyed and treated all identified Schistosoma mansoni infections in two rural Brazilian communities (Jenipapo and Volta do Rio) in 2009, 2012 and 2013. Eggs were collected from all infected individuals and genotyped with 11 microsatellite markers to evaluate parasite differentiation and diversity. After successive rounds of community-wide treatment, prevalence decreased from 45% to 24% then 16%. Intensity of infection decreased by 57% over this period, and the number of eggs transmitted to the environment decreased by 92%. During all time periods the majority of eggs were excreted by those >15years of age. The incidence was 23% in 2012 and 15% in 2013, consistent with a decrease in transmission. There was little immigration or gene flow over a distance of 6km. On reinfection, infrapopulations were moderately differentiated indicating that pretreatment multilocus genotypes were not fully reacquired. The effective population size responded to census population decline more rapidly than differentiation. Reinfection was concentrated in the downstream portion of Jenipapo, consistent with the observed increased human fecal contamination. At this scale and in this area S. mansoni infections exist on a fragmented landscape with a highly focal pattern of transmission that may facilitate future elimination.

The relative contribution of immigration or local increase for persistence of urban schistosomiasis in Salvador, Bahia, Brazil.

Urbanization is increasing across the globe, and diseases once considered rural can now be found in urban areas due to the migration of populations from rural endemic areas, local transmission within the city, or a combination of factors. We investigated the epidemiologic characteristics of urban immigrants and natives living in a neighborhood of Salvador, Brazil where there is a focus of transmission of Schistosoma mansoni. In a cross-sectional study, all inhabitants from 3 sections of the community were interviewed and examined. In order to determine the degree of parasite differentiation between immigrants and the native born, S. mansoni eggs from stools were genotyped for 15 microsatellite markers. The area received migrants from all over the state, but most infected children had never been outside of the city, and infected snails were present at water contact sites. Other epidemiologic features suggested immigration contributed little to the presence of infection. The intensity and prevalence of infection were the same for immigrants and natives when adjusted for age, and length of immigrant residence in the community was positively associated with prevalence of infection. The population structure of the parasites also supported that the contribution from immigration was small, since the host-to-host differentiation was no greater in the urban parasite population than a rural population with little distant immigration, and there had been little differentiation in the urban population over the past 7 years. Public health efforts should focus on eliminating local transmission, and once eliminated, reintroduction from distant migration is unlikely.

IL-22 and IL-22 binding protein (IL-22BP) regulate fibrosis and cirrhosis in hepatitis C virus and schistosome infections.

UNLABELLED: Interleukin (IL)-22 acts on epithelia, hepatocytes, and pancreatic cells and stimulates innate immunity, tissue protection, and repair. IL-22 may also cause inflammation and abnormal cell proliferation. The binding of IL-22 to its receptor is competed by IL-22 binding protein (IL-22BP), which may limit the deleterious effects of IL-22. The role of IL-22 and IL-22BP in chronic liver diseases is unknown. We addressed this question in individuals chronically infected with schistosomes or hepatitis C virus (HCV). We first demonstrate that schistosome eggs stimulate production of IL-22 transcripts and inhibit accumulation of IL22-BP transcripts in schistosome-infected mice, and that schistosome eggs selectively stimulate production of IL-22 in cultures of blood leukocytes from individuals chronically infected with Schistosoma japonicum. High IL-22 levels in cultures correlated with protection against hepatic fibrosis and portal hypertension. To test further the implication of IL-22/IL-22BP in hepatic disease, we analyzed common genetic variants of IL22RA2, which encodes IL-22BP, and found that the genotypes, AA, GG of rs6570136 (P = 0.003; odds ratio [OR] = 2), and CC, TT of rs2064501 (P = 0.01; OR = 2), were associated with severe fibrosis in Chinese infected with S. japonicum. We confirmed this result in Sudanese (rs6570136 GG [P = 0.0007; OR = 8.2], rs2064501 TT [P = 0.02; OR = 3.1]), and Brazilians (rs6570136 GG [P = 0.003; OR = 26], rs2064501 TC, TT (P = 0.03; OR = 11]) infected with S. mansoni. The aggravating genotypes were associated with high IL22RA2 transcripts levels. Furthermore, these same variants were also associated with HCV-induced fibrosis and cirrhosis (rs6570136 GG, GA [P = 0.007; OR = 1.7], rs2064501 TT, TC (P = 0.004; OR = 2.4]). CONCLUSIONS: These results provide strong evidence that IL-22 protects against and IL-22BP aggravates liver fibrosis and cirrhosis in humans with chronic liver infections. Thus, pharmacological modulation of IL-22 BP may be an effective strategy to limit cirrhosis.

Hepatitis C virus quantification in serum and saliva of HCV-infected patients.

The hepatitis C virus (HCV) can be detected in blood and other bodily fluids, such as saliva, semen and gastric juices. The aim of this study was to compare the HCV viral loads in the serum and saliva of infected patients. Twenty-nine patients with detectable HCV RNA in their serum and saliva were included in this study. The HCV viral loads were determined through quantitative real-time polymerase chain reactions. The median viral RNA levels were 5.78 log10 copies in the serum and 3.32 log10 copies in the saliva. We observed that the salivary HCV viral load was significantly lower than the viral load in the serum. Further studies are required to understand the role of saliva in the diagnosis, management and potential transmission of HCV.

Hepatitis C virus quantification in serum and saliva of HCV-infected patients

The hepatitis C virus (HCV) can be detected in blood and other bodily fluids, such as saliva, semen and gastric juices. The aim of this study was to compare the HCV viral loads in the serum and saliva of infected patients. Twenty-nine patients with detectable HCV RNA in their serum and saliva were included in this study. The HCV viral loads were determined through quantitative real-time polymerase chain reactions. The median viral RNA levels were 5.78 log10 copies in the serum and 3.32 log10 copies in the saliva. We observed that the salivary HCV viral load was significantly lower than the viral load in the serum. Further studies are required to understand the role of saliva in the diagnosis, management and potential transmission of HCV.

First insights into the genetic diversity of Mycobacterium tuberculosis strains in Salvador, Bahia State, Brazil.

This study constitutes a first attempt to describe the genetic population structure of Mycobacterium tuberculosis circulating in Salvador, Bahia State, Brazil. A total of 56 confirmed cases of pulmonary tuberculosis, identified between March and June 2008, were analyzed using restriction fragment length polymorphism (IS6110-RFLP). The study population was characterized by a predominance of males (71.43%) over 30 years of age (68.75%). Forty-one isolates were found to belong to a single pattern (73.2%), while 15 (26.7%) were found in group patterns, forming six clusters. The higher level of diversity observed is much more suggestive of endogenous reactivation than recent transmission.

First insights into the genetic diversity of Mycobacterium tuberculosis strains in Salvador, Bahia State, Brazil / Primeiro ensaio sobre diversidade genética das cepas de Mycobacterium tuberculosis em Salvador, Bahia, Brasil

This study constitutes a first attempt to describe the genetic population structure of Mycobacterium tuberculosis circulating in Salvador, Bahia State, Brazil. A total of 56 confirmed cases of pulmonary tuberculosis, identified between March and June 2008, were analyzed using restriction fragment length polymorphism (IS6110-RFLP). The study population was characterized by a predominance of males (71.43 percent) over 30 years of age (68.75 percent). Forty-one isolates were found to belong to a single pattern (73.2 percent), while 15 (26.7 percent) were found in group patterns, forming six clusters. The higher level of diversity observed is much more suggestive of endogenous reactivation than recent transmission.

Este é o primeiro estudo realizado na Bahia, Brasil, visando à descrição da estrutura da população genética circulante do Mycobacterium tuberculosis na cidade de Salvador. Um total de 56 casos confirmados de tuberculose pulmonar, identificados entre março e junho de 2008, foi analisado pelo método Restriction Fragment Lenght Polymorphism (IS6110-RFLP). A população de estudo foi caracterizada como a maioria do sexo masculino (71,43 por cento), idade acima de 30 anos (68,75 por cento). Quarenta e um isolados (73,21 por cento) com padrão único, enquanto 15 (26,75 por cento) apresentaram padrões agrupáveis, formando seis clusters. A alta taxa de diversidade das cepas de M. tuberculosis observada é mais sugestiva de reativação endógena do que transmissão recente.

Schistosoma mansoni population structure and persistence after praziquantel treatment in two villages of Bahia, Brazil.

Praziquantel has been used to treat schistosome infections since 1979 and currently is the only chemotherapeutic agent in production for this purpose, raising concerns about the potential for the emergence of drug resistance. In practice, 10-20% of infected patients will continue to excrete eggs after treatment. It is not understood to what degree this represents selection of a resistant population or incomplete elimination due to the presence of immature worms at the time of treatment. We used a population genetics approach to test whether or not persistent Schistosomamansoni parasites were drawn from the same population as susceptible parasites. In this study, stool samples were collected from 96% of individuals in two small Brazilian communities (populations 482 and 367) and examined for S.mansoni eggs. The combined prevalence of S.mansoni infections in the villages was 41%. Total egg DNA was extracted from each sample and was genotyped at 15 microsatellite markers. Day-to-day variation of the infrapopulation from an individual human host was low (median differentiation using Jost's D=0.010), so that a single stool was representative of the genotypes present in stool eggs, at least in the short term. Average pairwise analysis of D among all pre-treatment infrapopulations suggested moderate differentiation (mean D=0.082 and 0.122 for the two villages), whereas the pre-treatment component population differentiation between the two communities was 0.047. The differentiation of the component population remaining after treatment from the fully susceptible component population was low (mean D=0.007 and 0.020 for the two villages), suggesting that the persistent parasites were not selected by praziquantel treatment. We will continue to follow these communities for evidence of selection or changes in population structure.

IL-8 is associated with non-viremic state and IFN-γ with biochemical activity in HCV-seropositive blood donors.

OBJECTIVE: The aim of this study was to explore a possible association between the pattern of serum cytokines with the virological and biochemical status of hepatitis C virus (HCV)-seropositive blood donors. METHODS: 23 non-viremic and 33 viremic HCV-seropositive blood donors based on HCV-RNA tests, and 29 healthy individuals were included. Cytometric bead array assays were performed to detect cytokines. RESULTS: The subjects were classified as low, medium or high cytokine producers based on the tertile distribution. The absence of detectable viremia was associated with high IL-1ß and IL-8 producers. Conversely, elevated levels of IL-6, IL-10 and IL-12 were associated with detectable viremia. An increased frequency of high IL-1ß producers was observed frequently in the non-viremic recombinant immunoblot assay (RIBA)-indeterminate subjects, while the high IL-4, IL-6, IL-8, IL-10 and IL-12 producers were more frequent in the non-viremic RIBA-positive subjects. Furthermore, the levels of IL-1ß and IL-8 were higher in viremic subjects with a low level of alanine-aminotransferase (ALT), whereas the level of IFN-γ was increased among viremic subjects with a high ALT level. CONCLUSION: IL-1ß and IL-8 were more likely to be associated with a non-viremic or less severe HCV infection, whereas IL-2 and IFN-γ levels correlated with a high ALT level.

Prevalence and genotypes of hepatitis C virus among injecting drug users from Salvador-BA, Brazil.

Hepatitis C virus (HCV) is the major infectious disease agent among injecting drug users (IDUs), with seroprevalence ranging from 50-90%. In this paper, serological and virological parameters were investigated among 194 IDUs, 94 ex-IDUs and 95 non-IDUs that were sampled by the "snowball" technique in three localities renowned for both intense drug use and trafficking activities in Salvador, Brazil. The majority of the participants were male, but sex and mean age differed significantly between IDUs/ex-IDUs and non-IDUs (p < 0.05). Anti-HCV screening revealed that 35.6%, 29.8% and 5.3% of samples from IDUs, ex-IDUs and non-IDUs, respectively, were seropositive. HCV-RNA detection confirmed that the prevalence of infection was 29.4%, 21.3% and 5.3% for IDUs, ex-IDUs and non-IDUs, respectively. Genotyping analysis among IDUs/ex-IDUs determined that 76.9% were infected with genotype 1, 18.5% with genotype 3 and 4.6% with a mixed genotype; this result differed significantly from non-IDUs, where genotype 3 was the most frequent (60%), followed by genotype 1 (20%) and a mixed genotype (20%). We report a significantly higher prevalence of HCV infection in IDUs/ex-IDUs compared to the control group (p < 0.001). Although the sample size of our study was small, the differences in HCV genotype distribution reported herein for IDUs/ex-IDUs and non-IDUs warrant further investigation.

Prevalence and genotypes of hepatitis C virus among injecting drug users from Salvador-BA, Brazil

Hepatitis C virus (HCV) is the major infectious disease agent among injecting drug users (IDUs), with seroprevalence ranging from 50-90 percent. In this paper, serological and virological parameters were investigated among 194 IDUs, 94 ex-IDUs and 95 non-IDUs that were sampled by the "snowball" technique in three localities renowned for both intense drug use and trafficking activities in Salvador, Brazil. The majority of the participants were male, but sex and mean age differed significantly between IDUs/ex-IDUs and non-IDUs (p < 0.05). Anti-HCV screening revealed that 35.6 percent, 29.8 percent and 5.3 percent of samples from IDUs, ex-IDUs and non-IDUs, respectively, were seropositive. HCV-RNA detection confirmed that the prevalence of infection was 29.4 percent, 21.3 percent and 5.3 percent for IDUs, ex-IDUs and non-IDUs, respectively. Genotyping analysis among IDUs/ex-IDUs determined that 76.9 percent were infected with genotype 1, 18.5 percent with genotype 3 and 4.6 percent with a mixed genotype; this result differed significantly from non-IDUs, where genotype 3 was the most frequent (60 percent), followed by genotype 1 (20 percent) and a mixed genotype (20 percent). We report a significantly higher prevalence of HCV infection in IDUs/ex-IDUs compared to the control group (p < 0.001). Although the sample size of our study was small, the differences in HCV genotype distribution reported herein for IDUs/ex-IDUs and non-IDUs warrant further investigation.

Limited evidence of HCV transmission in stable heterosexual couples from Bahia, Brazil

HCV infected patients frequently ask their physician about the risk of transmission to their partners. Although it is easy to answer that the risk does exist, it is difficult to quantify. We studied the transmission of HCV infection in stable heterosexual couples: anti-HCV positive patients in hemodialytic therapy and their partners. Thirty-four couples were tested by third generation ELISA and RIBA. Blood samples of anti-HCV positive patients were evaluated by RT-PCR and detected sequences were genotyped by restriction fragment length polymorphism. Concordance of infection was observed in only one couple in which both subjects were in dialytic therapy. One other partner had two positive ELISA tests and an indeterminate RIBA, with negative RT-PCR, which may suggest a false positive or a previous resolved infection. Either sexual relations, sharing of personal items and history of parenteral exposure (hemodialysis, blood transfusion) could explain transmission in the only couple with concordant infection. We observed, in accordance with previous reports, that this risk is minimal or negligible in stable heterosexual couples.

Analysis of Schistosoma mansoni population structure using total fecal egg sampling.

Many parasite populations are difficult to sample because they are not uniformly distributed between several host species and are often not easily collected from the living host, thereby limiting sample size and possibly distorting the representation of the population. For the parasite Schistosoma mansoni, we investigated the use of eggs, in aggregate, from the stools of infected individuals as a simple and representative sample. Previously, we demonstrated that microsatellite allele frequencies can be accurately estimated from pooled DNA of cloned S. mansoni adults. Here, we show that genotyping of parasite populations from reproductively isolated laboratory strains can be used to identify these specific populations based on characteristic patterns of allele frequencies, as observed by polyacrylamide gel electrophoresis and automated sequencer analysis of fluorescently labeled PCR products. Microsatellites used to genotype aggregates of eggs collected from stools of infected individuals produced results consistent with the geographic distribution of the samples. Preferential amplification of smaller alleles, and stutter PCR products, had negligible effect on measurement of genetic differentiation. Direct analysis of total stool eggs can be an important approach to questions of population genetics for this parasite by increasing the sample size to thousands per infected individual and by reducing bias.

Increased frequency of CD56Bright NK-cells, CD3-CD16+CD56- NK-cells and activated CD4+T-cells or B-cells in parallel with CD4+CDC25High T-cells control potentially viremia in blood donors with HCV.

A detailed phenotypic analysis of major and minor circulating lymphocyte subsets is described in potential blood donors with markers of hepatitis C virus (HCV), including non-viremic and viremic groups. Although there were no changes in the hematological profile of either group, increased the levels of pre-NK cells (CD3-CD16+CD56-) and a lower frequency of mature NK cells (CD3-CD16+CD56+) characterized innate immunity in the non-viremic group. Both non-viremic and viremic groups displayed significantly increased levels of CD56(Bright) NK cells. Furthermore, this subset was significantly elevated in the viremic subgroup with a low viral load. In addition, an increase in the NKT2 subset was observed only in this subgroup. An enhanced frequency of activated CD4+ T-cells (CD4+HLA-DR+) was a characteristic feature of the non-viremic group, whereas elevated CD19+ B-cells and CD19+CD86+ cell populations were the major phenotypic features of the viremic group, particularly in individuals with a low viral load. Although CD4+CD25High T-cells were significantly elevated in both the viremic and non-viremic groups, it was particularly evident in the viremic low viral load subgroup. A parallel increase in CD4+CD25High T-cells, pre-NK, and activated CD4+ T-cells was observed in the non-viremic group, whereas a parallel increase in CD4+CD25High T-cells and CD19+ B-cells was characteristic of the low viral load subgroup. These findings suggest that CD56Bright NK cells, together with pre-NK cells and activated CD4+ T-cells in combination with CD4+CD25High T-cells, might play an important role in controlling viremia. Elevated CD56(Bright) NK cells, B-cell responses and a T-regulated immunological profile appeared to be associated with a low viral load.

Limited evidence of HCV transmission in stable heterosexual couples from Bahia, Brazil.

HCV infected patients frequently ask their physician about the risk of transmission to their partners. Although it is easy to answer that the risk does exist, it is difficult to quantify. We studied the transmission of HCV infection in stable heterosexual couples: anti-HCV positive patients in hemodialytic therapy and their partners. Thirty-four couples were tested by third generation ELISA and RIBA. Blood samples of anti-HCV positive patients were evaluated by RT-PCR and detected sequences were genotyped by restriction fragment length polymorphism. Concordance of infection was observed in only one couple in which both subjects were in dialytic therapy. One other partner had two positive ELISA tests and an indeterminate RIBA, with negative RT-PCR, which may suggest a false positive or a previous resolved infection. Either sexual relations, sharing of personal items and history of parenteral exposure (hemodialysis, blood transfusion) could explain transmission in the only couple with concordant infection. We observed, in accordance with previous reports, that this risk is minimal or negligible in stable heterosexual couples.

Potential immunomodulatory effects of plant lectins in Schistosoma mansoni infection.

Lectins are sugar-binding glycoproteins that can stimulate, in a non-antigen-specific fashion, lymphocytes, leading to proliferation and cytokine production. Some lectins are utilized as in vitro mitogenic lymphocyte stimulators and their use as immunomodulators against infectious diseases has been evaluated experimentally. In the experimental murine model, the immune response to schistosomiasis is Th1-like during the initial stage of infection, with a shift towards a Th2-like response after oviposition. We report the response of schistosomiasis patients' (n=37) peripheral blood mononuclear cells (PBMC) to stimulation by lectins, including newly isolated lectins from Brazilian flora, and by Schistosomamansoni soluble egg antigens (SEA). Cytokine production upon lectin stimulation ex vivo was assessed in PBMC supernatants, collected at 24 and 72 h, by sandwich ELISA to IL-5, IL-10, TNF-alpha and IFN-gamma. In PBMC from infected patients all but one of the lectins induced a Th2-like cytokine response, characterized by elevated IL-5 production that was higher than that induced by SEA stimulation alone. Our results show that the Th2 environment present during schistosomiasis is not affected and that it may be further stimulated by the presence of lectins.

Association of TGF-beta1 codon 25 (G915C) polymorphism with hepatitis C virus infection.

Cytokines play a key role in the regulation of immune responses. In hepatitis C virus infection (HCV), the production of abnormal cytokine levels appears to contribute to the progression of the disease, viral persistence, and affects response to therapy. Cytokine genes are polymorphic at specific sites, and certain polymorphisms located within coding/regulatory regions have been shown to affect the overall expression and secretion of cytokines. The aim of the present study was to identify potential markers of cytokines genes associated with the susceptibility to HCV infection. The cohort was composed of 128 individuals infected by HCV and 94 healthy controls. Genotyping was carried out by PCR-SSP. The distributions of the following polymorphisms were compared in these groups: TNF-alpha (-308G/A [rs1800629]), TGF-beta1 (codon 10 T/C [rs1982073], codon 25 G/C [rs1800471]), IL-10 (-1082 A/G [rs 1800896]; -819T/C [rs1800871]; -592A/C [rs 1800872]), IL-6 (-174G/C [rs1800795]), and IFN-gamma (+874T/A [rs2430561]). This study demonstrated a statistically significant difference in the frequency of TGF-beta1 codon 25 polymorphism between healthy subjects and those infected with HCV. No associations were observed between polymorphisms of TNF-alpha, IFN-gamma, IL-10, TGF-beta1 codon 10, and IL-6 and HCV infection. These findings suggest that TGF-beta1 codon 25 polymorphism could be a host genetic factor associated with susceptibility to HCV infection.

Pseudotumoral form of neuroschistosomiasis: report of three cases.

Central nervous system (CSN) involvement in schistosomiasis is an ectopic manifestation with a large variety of clinical forms, including pseudotumoral, which occurs in isolated cases and is rare. Three patients with epidemiological indications of this pathology were examined; the clinical picture included lower-back pain irradiating to lower limbs, associated with progressive flaccid paraparesis and sphincterial disturbances in cases in which the spinal chord was involved; while in cases with encephalitic impairment, headache, dizziness and cerebellar syndrome, characterized by dysarthria and right-side dysgraphia, were present. Magnetic resonance imaging (MRI) showed a growing process in all cases; cerebrospinal fluid (CSF) characteristics and biological markers were compatible with neuroschistosomiasis (NS). Biopsy of the lesions confirmed this diagnosis in one case. After specific treatment with schistosomicides and corticosteroids, clinical, radiological and laboratorial improvement was observed.

Pseudotumoral form of neuroschistosomiasis: report of three cases

Central nervous system (CSN) involvement in schistosomiasis is an ectopic manifestation with a large variety of clinical forms, including pseudotumoral, which occurs in isolated cases and is rare. Three patients with epidemiological indications of this pathology were examined; the clinical picture included lower-back pain irradiating to lower limbs, associated with progressive flaccid paraparesis and sphincterial disturbances in cases in which the spinal chord was involved; while in cases with encephalitic impairment, headache, dizziness and cerebellar syndrome, characterized by dysarthria and right-side dysgraphia, were present. Magnetic resonance imaging (MRI) showed a growing process in all cases; cerebrospinal fluid (CSF) characteristics and biological markers were compatible with neuroschistosomiasis (NS). Biopsy of the lesions confirmed this diagnosis in one case. After specific treatment with schistosomicides and corticosteroids, clinical, radiological and laboratorial improvement was observed.

Biochemical and immunologic predictors of efficacy of treatment or reinfection risk for Schistosoma mansoni.

Most Schistosoma mansoni infections are egg-negative after a single dose of oxamniquine. A cohort of 661 infected children was treated at 6-month intervals and assessed for nutritional and parasitological status. Initial biochemical and immunologic markers were measured in a subset of 84 children. All were treated at the start of therapy and at 6 months. Immunoglobulins only served as markers for active infection. No markers were predictive of cure or reinfection, except initial infection intensity and serum low-density lipoprotein. Ten percent were persistently infected and had no change in infection intensity at any time-point. Several factors suggest that this group was biologically different. In addition to failing to reduce their worm burden, they had significantly higher initial intensity of infection (100 versus 65 eggs/g, P = 0.001) and significantly lower initial serum low-density lipoprotein (72 versus 104 mg/dL, P = 0.045). The biologic plausibility of this observation is discussed.