Evolution of methodology for the study of adaptability and stability in cultivated species.

The GxE interaction only became widely discussed from evolutionary studies and evaluations of the causes of behavioral changes of species cultivated in environments. In the last 60 years, several methodologies for the study of adaptability and stability of genotypes in multiple environments trials were developed in order to assist the breeder's choice regarding which genotypes are more stable and which are the most suitable for the crops in the most diverse environments. The methods that use linear regression analysis were the first to be used in a general way by breeders, followed by multivariate analysis methods and mixed models. The need to identify the genetic and environmental causes that are behind the GxE interaction led to the development of new models that include the use of covariates and which can also include both multivariate methods and mixed modeling. However, further studies are needed to identify the causes of GxE interaction as well as for the more accurate measurement of its effects on phenotypic expression of varieties in competition trials carried out in genetic breeding programs.

Diallelic analysis to obtain cowpea (Vigna unguiculata L. Walp.) populations tolerant to water deficit.

The purpose of this study was to identify parents and obtain segregating populations of cowpea (Vigna unguiculata L. Walp.) with the potential for tolerance to water deficit. A full diallel was performed with six cowpea genotypes, and two experiments were conducted in Teresina, PI, Brazil in 2011 to evaluate 30 F2 populations and their parents, one under water deficit and the other under full irrigation.

Epidemiological and molecular characteristics of HIV-1 infection among female commercial sex workers, men who have sex with men and people living with AIDS in Paraguay

An HIV seroprevalence and molecular study was conducted among 935 subjects: 723 female commercial sex workers, 92 men who have sex with men and 120 HIV-positive volunteers. The reported injection drug use rates were 0.7% in female commercial sex workers and 3% in men who have sex with men. Sexually transmitted infections were reported in 265 (37%) of the female commercial sex workers and 38 (41%) of the men who have sex with men. A total of 20 (2.8%) female commercial sex workers and 12 (13%) men who have sex with men became HIV infected during the study period. A history of sexually transmitted infection increased the risk of subsequent HIV infection twofold (adjusted odds ratio of 2.5) among the female commercial sex workers, while cocaine use had an adjusted odds ratios of 6.61 among men who have sex with men. From 130 samples, and based on heteroduplex mobility assaying for the env gene, with sequencing of part of pol and/or full genomes, subtype B was the predominant subtype identified (66%); followed by subtype F (22%) and subtype C (4%). Recombinant CRF12-BF strains were identified in 6% and CRF17_BF was identified in 2%.

Study of salt stress tolerance in spelt

World population is increasing at an alarming rate while food productivity is decreasing due to the effect of various abiotic stresses. Soil salinity is one of the most important abiotic stress and a limiting factor for worldwide plant production. In addition to its important effects on yield, salt stress affects numerous cellular activities, including cell wall composition, photosynthesis, protein synthesis, ions and organic solutes. Up to 20% of the irrigated arable land in arid and semiarid regions is already salt affected and is still expanding. Improving salt tolerant varieties is of major importance, and efforts should be focused on finding adaptive mechanisms which are involved in salinity tolerance. In this study, several spelt wheat (Triticum aestivum var. Spelta) genotypes and one cultivar of modern bread wheat were used to screen them for salt tolerance. Spelt is an old-European cereal crop currently attracting renewed interest as a food grain because it is said to be harder than wheat and requires less fertilizer. Spelt wheat is also becoming very attractive genetic source by plant breeders due to its wide adaptation ability to various stressful conditions such as soil salinity. In this study morphological parameters (e.g., leaf appearance; shoot elongation), dry matter production, mineral nutrients (especially Na and K), and activity of antioxidative enzymes were measured to select superior genotypes of spelt for salt tolerance. The results showed that Spelt genotype Sp41 is a salt sensitive genotype and genotypes Sp69, Sp96 and Sp912 are good candidates for salt tolerant genotypes.

Characterization of mannose-binding lectin plasma levels and genetic polymorphisms in HIV-1-infected individuals

INTRODUCTION: The present study investigated the association between mannose-binding lectin (MBL) gene polymorphism and serum levels with infection by HIV-1. METHODS: Blood samples (5mL) were collected from 97 HIV-1-infected individuals resident in Belém, State of Pará, Brazil, who attended the Special Outpatient Unit for Infections and Parasitic Diseases (URE-DIPE). CD4+ T-lymphocyte count and plasma viral load were quantified. A 349bp fragment of exon 1 of the MBL was amplified via PCR, using genomic DNA extracted from controls and HIV-1-infected individuals, following established protocols. MBL plasma levels of the patients were quantified using an enzyme immunoassay kit. RESULTS: Two alleles were observed: MBL*O, with a frequency of 26.3% in HIV-1-infected individuals; and the wild allele MBL*A (73.7%). Similar frequencies were observed in the control group (p > 0.05). Genotype frequencies were distributed according to the Hardy-Weinberg equilibrium in both groups. Mean MBL plasma levels varied by genotype, with statistically significant differences between the AA and AO (p < 0.0001), and AA and OO (p < 0.001) genotypes, but not AO and OO (p = 0.17). Additionally, CD4+ T-lymphocytes and plasma viral load levels did not differ significantly by genotype (p > 0.05). CONCLUSIONS: The results of this study do not support the hypothesis that MBL gene polymorphism or low plasma MBL concentrations might have a direct influence on HIV-1 infection, although a broader study involving a large number of patients is needed.

Restriction enzyme analysis of the human cytomegalovirus genome in specimens collected from immunodeficient patients in Belém, State of Pará, Brazil

INTRODUCTION: Human cytomegalovirus is an opportunistic betaherpesvirus that causes persistent and serious infections in immunodeficient patients. Recurrent infections occur due to the presence of the virus in a latent state in some cell types. It is possible to examine the virus using molecular methods to aid in the immunological diagnosis and to generate a molecular viral profile in immunodeficient patients. The objective of this study was to characterize cytomegalovirus genotypes and to generate the epidemiological and molecular viral profile in immunodeficient patients. METHODS: A total of 105 samples were collected from immunodeficient patients from the City of Belém, including newborns, hemodialysis patients, transplant recipients and HIV+ patients. An IgG and IgM antibody study was completed using ELISA, and enzymatic analysis by restriction fragment length polymorphism (RFLP) was performed to characterize viral genotypes. RESULTS: It was observed that 100% of the patients had IgG antibodies, 87% of which were IgG+/IgM-, consistent with a prior infection profile, 13% were IgG+/IgM+, suggestive of recent infection. The newborn group had the highest frequency (27%) of the IgG+/IgM+ profile. By RFLP analysis, only one genotype was observed, gB2, which corresponded to the standard AD169 strain. CONCLUSIONS: The presence of IgM antibodies in new borns indicates that HCMV continues to be an important cause of congenital infection. The low observed genotypic diversity could be attributed to the small sample size because newborns were excluded from the RFLP analysis. This study will be continued including samples from newborns to extend the knowledge of the general and molecular epidemiology of HCMV in immunodeficient patients.

Molecular analysis and dimorphism of azole-susceptible and resistant Candida albicans isolates

INTRODUCTION: Candida albicans is responsible for superficial or systemic infections known as candidiasis, which may be found in infected tissue as unicellular budding yeasts, hyphae, or pseudohyphae. In this study, the effects of both fluconazole and itraconazole antifungal agents on the hyphal formation and genotypic characterization of C. albicans isolates classified as either susceptible or resistant were investigated. METHODS: The hyphal production of five C. albicans isolates under the action of antifungal agents was investigated by culturing yeast on growth medium and on hyphal induction medium. The genotypic characterization was carried out for 13 isolates of C. albicans using the random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) method. RESULTS: The dimorphism analysis showed that the hyphal formation was higher in resistant than in the susceptible isolates to both azoles. The RAPD-PCR method identified the formation of two different groups. In group A, four resistant and two susceptible isolates were clustered, and in group B, one resistant and six susceptible isolates were clustered. CONCLUSIONS: Considering that hyphal formation was higher in resistant isolates in the presence of azole drugs, we confirmed that the hyphal production is closely related to susceptibility to azoles. These drugs may affect the morphogenesis of C. albicans depending on their susceptibility to these drugs. In relation to RAPD-PCR, most resistant isolates classified in group A and susceptible isolates in group B demonstrated that this method presented a similar standard between the two groups, suggesting that by this technique, a strong correlation between genotypes and fluconazole-resistant samples may be found.

Molecular characterization of the hepatitis B virus in autochthonous and endogenous populations in the Western Brazilian Amazon

INTRODUCTION: Hepatitis B virus (HBV) infection is a serious public health issue worldwide. Hepatitis B virus is classified into eight genotypes, varying from A to H, with distinct geographical distributions. In Brazil, the most frequent genotypes are A, D, and F. METHODS: This study aimed to characterize the HBV genotypes in cases of hepatitis B virus and hepatitis D virus (HDV) co-infections in an endemic area in the Western Brazilian Amazon. We analyzed 86 serum samples reactive for HBsAg from indigenous and non-indigenous populations obtained from previous serological surveys. RESULTS: Of the 86 reactive serum samples, 39 were found to be HBV-DNA-positive by semi-nested PCR. The genotypes were established by sequencing the amplified S gene region. We obtained 20 sequences classified into three genotypes: A, D, and F. Genotype A was the most frequent (60%), followed by D (35%) and F (5%). CONCLUSIONS: The distribution of the HBV genotypes reflected the pattern of historical occupation of the region.

Biological behavior of Trypanosoma cruzi stocks obtained from the state of Amazonas, Western Brazilian Amazon, in mice

INTRODUCTION: The biological diversity of circulating Trypanosoma cruzi stocks in the Amazon region most likely plays an important role in the peculiar clinic-epidemiological features of Chagas disease in this area. METHODS: Seven stocks of T. cruzi were recently isolated in the State of Amazonas, Brazil, from humans, wild mammals, and triatomines. They belonged to the TcI and Z3 genotypes and were biologically characterized in Swiss mice. Parasitological and histopathological parameters were determined. RESULTS: Four stocks did not promote patent parasitemia in mice. Three stocks produced low parasitemia, long pre-patent periods, and a patent period of 1 day or oscillating parasitemia. Maximum parasitemia ranged from 1,400 to 2,800 trypomastigotes/0.1mL blood. Mice inoculated with the T. cruzi stocks studied showed low positivity during fresh blood examinations, ranging from 0% to 28.6%. In hemoculture, positivity ranged from 0% to 100%. Heart tissue parasitism was observed in mice inoculated with stocks AM49 and AM61. Stock AM49 triggered a moderate inflammatory process in heart tissue. A mild inflammatory process was observed in heart tissue for stocks AM28, AM38, AM61, and AM69. An inflammatory process was frequently observed in skeletal muscle. Examinations of brain tissue revealed inflammatory foci and gliosis in mice inoculated with stock AM49. CONCLUSIONS: Biological and histopathological characterization allowed us to demonstrate the low infectivity and virulence of T. cruzi stocks isolated from the State of Amazonas.

Genetic diversity of dengue virus serotypes 1 and 2 in the State of Paraná, Brazil, based on a fragment of the capsid/premembrane junction region

INTRODUCTION: The precise identification of the genetic variants of the dengue virus is important to understand its dispersion and virulence patterns and to identify the strains responsible for epidemic outbreaks. This study investigated the genetic variants of the capsid-premembrane junction region fragment in the dengue virus serotypes 1 and 2 (DENV1-2). METHODS: Samples from 11 municipalities in the State of Paraná, Brazil, were provided by the Central Laboratory of Paraná. They were isolated from the cell culture line C6/36 (Aedes albopictus) and were positive for indirect immunofluorescence. Ribonucleic acid (RNA) extracted from these samples was submitted to the reverse transcription polymerase chain reaction (RT-PCR) and nested PCR. RESULTS: RT-PCR revealed that 4 of the samples were co-infected with both serotypes. The isolated DENV-1 sequences were 95-100% similar to the sequences of other serotype 1 strains deposited in GenBank. Similarly, the isolated DENV-2 sequences were 98-100% similar to other serotype 2 sequences in GenBank. According to our neighbor-joining tree, all strains obtained in this study belonged to genotype V of DENV-1. The DENV-2 strains, by contrast, belonged to the American/Asian genotypes. CONCLUSIONS: The monitoring of circulating strains is an important tool to detect the migration of virus subtypes involved in dengue epidemics.

Hepatitis B virus infection in children, adolescents, and their relatives: genotype distribution and precore and core gene mutations

INTRODUCTION:The objectives of this study were evaluate hepatitis B virus (HBV) serological markers in children and adolescents followed up at the Child Institute of the Hospital das Clínicas, Faculdade de Medicina de São Paulo, Universidade de São Paulo; identify chronic HBV carriers and susceptible individuals in the intrafamilial environment; characterize HBV genotypes; and identify mutations in the patients and household contacts. METHODS: Ninety-five hepatitis B surface antigen-positive children aged <19 years and 118 household contacts were enrolled in this study. Commercial kits were used for the detection of serological markers, and PCR was used for genotyping. RESULTS: Hepatitis B e antigen (HBeAg) was detected in 66.3% (63/95) of cases. Three of the 30 HBeAg-negative and anti-HBeAg-positive patients presented with precore mutations and 11 presented with mutations in the basal core promoter (BCP). Genotype A was identified in 39 (43.8%) patients, genotype D in 45 (50.6%), and genotype C in 5 (5.6%). Of the 118 relatives, 40 were chronic HBV carriers, 52 presented with the anti-HBc marker, 19 were vaccinated, and 7 were susceptible. Among the relatives, genotypes A, D, and C were the most frequent. One parent presented with a precore mutation and 4 presented with BCP mutations. CONCLUSIONS: Genotypes A and D were the most frequent among children, adolescents, and their relatives. The high prevalence of HBV in the families showed the possibility of its intrafamilial transmission.

Detection and molecular analysis of Toxoplasma gondii and Neospora caninum from dogs with neurological disorders

INTRODUCTION: Toxoplasma gondii and Neospora caninum are related Apicomplexa parasites responsible for systemic diseases in many species of animals, including dogs. METHODS: This study aimed to determine the occurrence of T. gondii and N. caninum infections in 50 dogs with neurological signs that were admitted to the Veterinary Hospital of Universidade Estadual Paulista, City of Botucatu, Brazil. All animals were screened for antibodies using an immunofluorescent antibody test for both parasites. Tissues of positive animals were bioassayed in mice (T. gondii) and gerbils (N. caninum), and DNA was analyzed using the polymerase chain reaction (PCR). Positive samples for T. gondii by PCR were typed using restriction fragment length polymorphism-PCR for 11 markers: SAG1, SAG2 (5′-3′-SAG2 and alt.SAG2), SAG3, Btub, GRA6, L358, c22-8, c29-6, PK1 and Apico, and CS3 marker for virulence analysis. RESULTS: Specific antibodies were detected in 11/50 (22%; 95% confidence interval (CI95%), 12.8-35.3%) animals for T. gondii and 7/50 (14%; CI95%, 7.02-26.3%) for N. caninum. In the bioassay and PCR, 7/11 (63.6%; CI95%, 34.9-84.8%) samples were positive for T. gondii and 3/7 (42.9%; CI95%I, 15.7-75.5%) samples were positive for N. caninum. Three different genotypes were identified, but only 1 was unique. CONCLUSIONS: These data confirm the presence of T. gondii and N. caninum in dogs from Brazil, indicating the importance of this host as a sentinel of T. gondii for human beings, and the genotypic variation of this parasite in Brazil.

Rotavirus genotyping in gastroenteritis cases of an infantile population from Western Brazilian Amazonia

INTRODUCTION: During the period from 2000 to 2002, 79 rotavirus-positive stool samples were collected from children presenting diarrhea in the Western Brazilian Amazon. METHODS: Molecular characterization of the G and P genotypes was performed using RT-PCR and electropherotyping analysis by polyacrylamide gel electrophoresis. RESULTS: A total of 59 samples were confirmed as group A rotavirus. A long electrophoretic profile was exhibited by the G1P[8], G3P[8], and G4P[8] genotypes. The G1P[8] genotype was found in greater proportion. The short electropherotype was exhibited only by G2 genotype strains. CONCLUSIONS: The proportion of the rotavirus genotypes observed was not different from that in other areas of Brazil. This study is the first genotyping of rotavirus in the Western Brazilian Amazon.

Occult hepatitis B virus infection in hemodialysis patients in Recife, State of Pernambuco, Brazil

INTRODUCTION: Persistence of the hepatitis B virus (HBV) genome in individuals negative for the HBV surface antigen (HBsAg) reflects occult infection. The aim of this study was to identify occult HBV infection among hemodialysis patients at 5 clinics in Recife, State of Pernambuco, Brazil, between August 2006 and August 2007. METHODS: Serum samples underwent enzyme-linked immunosorbent assay to investigate total antibodies against HBcAg (anti-HBc), HBsAg, and antibodies against HBsAg (anti-HBs). Samples that were HBsAg-negative were tested for total anti-HBc, and those that were positive for total anti-HBc were tested for anti-HBs. HBV DNA was investigated with an in-house PCR technique to identify samples positive for total anti-HBc. Subsequently, the samples positive for HBV DNA were sequenced to identify the genotype and mutations. RESULTS: The study population (n = 752) had a mean age of 50 15.1 years and included both sexes. All samples analyzed were negative for HBsAg. The seroprevalence of total anti-HBc was 26.7% (201/752), while that of anti-HBs was 67.2% (135/201). Total anti-HBc alone was detected in 5.7% of the patients. Occult infection was found in 1.5%, comprising genotypes A (33.3%, 1/3) and D (66.7%, 2/3). No mutations were found. CONCLUSIONS: The study detected occult hepatitis B virus infection in hemodialysis patients. Molecular studies on HBV are of fundamental importance because they identify patients that had been considered virus-negative but who, in reality, host the virus and have the ability to transmit it to other patients and staff.

Evaluation of the therapeutic response of hepatitis C in coinfected patients (HIV/HCV): a study of cases from a hospital for chronic liver diseases in the Eastern Brazilian Amazon

INTRODUCTION: The aim of this study was to evaluate the therapeutic response of hepatitis C in patients coinfected with human immunodeficiency virus (HIV-1). METHODS: A retrospective study of 20 patients coinfected with HIV-1/HCV who were treated in the outpatient liver clinic at the Sacred House of Mercy Foundation Hospital of Pará (Fundação Santa Casa de Misericórdia do Pará - FSCMPA) from April 2004 to June 2009. Patients were treated with 180µg PEG interferon-α2a in combination with ribavirin (1,000 to 1,250mg/day) for 48 weeks. The end point was the sustained virological response (SVR) rate (HCV RNA negative 24 weeks after completing treatment). RESULTS: The mean age of the patients was 40±9.5 years, of which 89% (n=17) were male, and the HCV genotypes were genotype 1 (55%, n=11/20), genotype 2 (10%, n=2/20) and genotype 3 (35%, n=7/20). The mean CD4+ lymphocyte count was 507.8, and the liver fibrosis stages were (METAVIR) F1 (25%), F2 (55%), F3 (10%) and F4 (10%). The early virological response (EVR) was 60%, the end-of-treatment virological response (EOTVR) was 45% and the SVR was 45%. CONCLUSIONS: The median HCV viral load was high, and in 85% of cases in which highly active antiretroviral therapy (HAART) was used, none of the patients with F3-F4 fibrosis responded to treatment. Of the twenty patients treated, 45% achieved SVR and 45% achieved EOTVR. Studies that include cases from a wider region are needed to better evaluate these findings.