Genetic Diversity of the Cryptococcus gattii Species Complex in Mato Grosso State,Brazil

Mycopathologia - Cryptococcosis is caused by fungi of the genus Cryptococcus. Owing to its importance, this study aimed to analyze the genetic diversity of C. gattii isolates from animals, humans,...     

Molecular Epidemiology Reveals Genetic Diversity amongst Isolates of the Cryptococcus neoformans/C. gattii Species Complex in Thailand

To gain a more detailed picture of cryptococcosis in Thailand, a retrospective study of 498 C. neoformans and C. gattii isolates has been conducted. Among these, 386, 83 and 29 strains were from clinical, environmental and veterinary sources, respectively. A total of 485 C. neoformans and 13 C. gattii strains were studied. The majority of the strains (68.9%) were isolated from males (mean age of 37.97 years), 88.5% of C. neoformans and only 37.5% of C. gattii strains were from HIV patients. URA5-RFLP and/or M13 PCR-fingerprinting analysis revealed that the majority of the isolates were C. neoformans molecular type VNI regardless of their sources (94.8%; 94.6% of the clinical, 98.8% of the environmental and 86.2% of the veterinary isolates). In addition, the molecular types VNII (2.4%; 66.7% of the clinical and 33.3% of the veterinary isolates), VNIV (0.2%; 100% environmental isolate), VGI (0.2%; 100% clinical isolate) and VGII (2.4%; 100% clinical isolates) were found less frequently. Multilocus Sequence Type (MLST) analysis using the ISHAM consensus MLST scheme for the C. neoformans/C. gattii species complex identified a total of 20 sequence types (ST) in Thailand combining current and previous data. The Thai isolates are an integrated part of the global cryptococcal population genetic structure, with ST30 for C. gattii and ST82, ST83, ST137, ST141, ST172 and ST173 for C. neoformans being unique to Thailand. Most of the C. gattii isolates were ST7 = VGIIb, which is identical to the less virulent minor Vancouver island outbreak genotype, indicating Thailand as a stepping stone in the global spread of this outbreak strain. The current study revealed a greater genetic diversity and a wider range of major molecular types being present amongst Thai cryptococcal isolates than previously reported.     

Cryptococcus neoformans and Cryptococcus gattii Isolated from the Excreta of Psittaciformes in a Southern Brazilian Zoological Garden

Cryptococcus neoformans, a major pathogen in immunocompromised patients, is a ubiquitous free-living fungus that can be isolated from soils, avian excreta and plant material. To further study potential saprophytic sources of this yeast in the Southern Brazilian State Rio Grande do Sul, we analyzed fecal samples from 59 species of captive birds kept in cages at a local Zoological Garden, belonging to 12 different orders. Thirty-eight environmental isolates of C. neoformans were obtained only from Psittaciformes (Psittacidae, Cacatuidae and Psittacula). Their variety and serotype were determined, and the genetic structure of the isolates was analyzed by use of the simple repetitive microsatellite specific primer M13 and the minisatellite specific primer (GACA)₄ as single primers in the PCR. The varieties were confirmed by pulsed-field gel electrophoresis (PFGE). Thirty-three isolates (87%) were from the var. grubii, serotype A, molecular type VNI and five (13%) were Cryptococcus gattii, serotype B, molecular type VGI. All the isolates were mating type α. Isolates were screened for some potential virulence factors. Quantitative urease production by the environmental isolates belonging to the C. gattii was similar to the values usually obtained for clinical ones.     

Genetic Diversity and Natural Population Structure of Cacao (Theobroma cacao L.) from the Brazilian Amazon Evaluated by Microsatellite Markers

A sample of 94 accessions of Theobroma cacao L. (cacao), representing four populations from the Brazilian Amazon (Acre, Rondônia, lower Amazon and upper Amazon) were analyzed using microsatellite markers to assess the genetic diversity and the natural population structure. From the 19 microsatellite loci tested, 11 amplified scorable products, revealing a total of 49 alleles, including two monomorphic loci. The Brazilian upper Amazon population contained the largest genetic diversity, with the most polymorphic loci, the highest observed heterozygosity; and the majority of rare alleles, thereby this region might be considered part of the center of diversity of the species. The observed heterozygosity for all the Brazilian populations (H _o = 0.347) was comparable with values reported for other similar upper Amazon Forastero cacao populations, with the Acre and Rondônia displaying the lowest values. The lower Amazon population, traditionally defined as highly homozygous, presented an unexpectedly high observed heterozygosity (H _o = 0.372), disclosing rare and distinct alleles, with large identity with the upper Amazon population. It was hypothesized that part of the lower Amazon population might derive from successive natural or intentional introduction of planting material from other provenances, mainly upper Amazon. Most of the loci exhibited a lower observed heterozygosity than expected, suggesting that self-pollination might be more common than usually assumed in cacao, but excess of homozygotes might also derive from sub-grouping (Wahlund effect) or from sampling related individuals. Most of the gene diversity was found to occur within groups, with small differentiation between the four Brazilian Amazon populations, typical of species with high gene flow.     

Successful treatment of Cryptococcus gattii neurocryptococcosis in a 5-year-old immunocompetent child from the French Guiana Amazon region

Compared to the incidence in adults, cryptococcosis is rare among children. We report a case of neurocryptococcosis due to Cryptococcus gattii in a five-year-old girl without identified risk factors living in French Guiana. Neurological surgery in combination with long-term antifungal treatment with amphotericin B and 5-flucytosine successfully resolved the cryptococcal infection. Subsequent molecular characterization of the Cryptococcus isolate revealed that the infection was caused by a C. gattii genotype AFLP6B/VGIIb strain.     

Genotypic Analysis of Klebsiella pneumoniae Isolates in a Beijing Hospital Reveals High Genetic Diversity and Clonal Population Structure of Drug-Resistant Isolates

Background

The genetic diversity and the clinical relevance of the drug-resistant Klebsiella pneumoniae isolates from hospital settings are largely unknown. We thus conducted this prospective study to analyze the molecular epidemiology of K. pneumoniae isolates from patients being treated in the 306 Hospital in Beijing, China for the period of November 1, 2010–October 31, 2011.

Methodology/Principal Findings

Antibiotic susceptibility testing, PCR amplification and sequencing of the drug resistance-associated genes, and multilocus sequence typing (MLST) were conducted. A total of 163 isolates were analyzed. The percentage of MDR, XDR and PDR isolates were 63.8% (104), 20.9 (34), and 1.8% (3), respectively. MLST results showed that 60 sequence types (STs) were identified, which were further separated by eBURST into 13 clonal complexes and 18 singletons. The most dominant ST was ST15 (10.4%). Seven new alleles and 24 new STs were first identified in this study. Multiple logistic regression analysis revealed that certain clinical characteristics were associated with those prevalent STs such as: from ICU, from medical ward, from community acquired infection, from patients without heart disease, from patients with treatment success, susceptible to extended spectrum cephalosporin, susceptible to cephamycins, susceptible to fluoroquinolones, and with MDR.

Conclusions/Significance

Our data indicate that certain drug-resistant K. pneumoniae clones are highly prevalent and are associated with certain clinical characteristics in hospital settings. Our study provides evidence demonstrating that intensive nosocomial infection control measures are urgently needed.     

Estimating the Population Size and Genetic Diversity of Amur Tigers in Northeast China

Over the past century, the endangered Amur tiger (Panthera tigris altaica) has experienced a severe contraction in demography and geographic range because of habitat loss, poaching, and prey depletion. In its historical home in Northeast China, there appears to be a single tiger population that includes tigers in Southwest Primorye and Northeast China; however, the current demographic status of this population is uncertain. Information on the abundance, distribution and genetic diversity of this population for assessing the efficacy of conservation interventions are scarce. We used noninvasive genetic detection data from scats, capture-recapture models and an accumulation curve method to estimate the abundance of Amur tigers in Northeast China. We identified 11 individual tigers (6 females and 5 males) using 10 microsatellite loci in three nature reserves between April 2013 and May 2015. These tigers are confined primarily to a Hunchun Nature Reserve along the border with Russia, with an estimated population abundance of 9–11 tigers during the winter of 2014–2015. They showed a low level of genetic diversity. The mean number of alleles per locus was 2.60 and expected and observed heterozygosity were 0.42 and 0.49, respectively. We also documented long-distance dispersal (~270 km) of a male Amur tiger to Huangnihe Nature Reserve from the border, suggesting that the expansion of neighboring Russian populations may eventually help sustain Chinese populations. However, the small and isolated population recorded by this study demonstrate that there is an urgent need for more intensive regional management to create a tiger-permeable landscape and increased genetic connectivity with other populations.     

Clonal Diversity and Fine‐scale Genetic Structure in a High Andean Treeline Population

Clonal propagation becomes more abundant with increasing altitudes as environmental conditions worsen. To date, little attention has been paid to the way in which clonal propagation affects genetic diversity and the fine‐scale spatial genetic structure (FSGS) of clonal alpine trees. An AFLP study was undertaken to quantify the clonal and genetic diversity and FSGS of the vulnerable treeline species Polylepis reticulata in Ecuador. We successfully genotyped 32 and 75 ramets within 4 m × 100 m (coarse scale) and 4 m × 4 m (fine scale) transects of one population, respectively. Higher genotypic diversity was detected at the coarse scale than at the fine scale, while lower genetic diversity was detected for P. reticulata than other Polylepis spp. at both scales. Significantly stronger FSGS was detected at the ramet level than the genet level for P. reticulata within a spatial distance of 3 m. The studied P. reticulata population showed pronounced FSGS (Sp = 0.012 at the genet level, a statistic reflecting declining pairwise kinship with distance) revealed restricted gene dispersal, which implies restricted seed dispersal for this population, assuming pollen flow is as extensive as that described for other wind‐pollinated tree species. Our results revealed that clonal diversity is a function of both sample size and the spatial scale of the sampling area. The findings highlights that clonal propagation has affected FSGS within a spatial distance of 3 m for this species.     

A Genome-Wide Analysis of Populations from European Russia Reveals a New Pole of Genetic Diversity in Northern Europe

Several studies examined the fine-scale structure of human genetic variation in Europe. However, the European sets analyzed represent mainly northern, western, central, and southern Europe. Here, we report an analysis of approximately 166,000 single nucleotide polymorphisms in populations from eastern (northeastern) Europe: four Russian populations from European Russia, and three populations from the northernmost Finno-Ugric ethnicities (Veps and two contrast groups of Komi people). These were compared with several reference European samples, including Finns, Estonians, Latvians, Poles, Czechs, Germans, and Italians. The results obtained demonstrated genetic heterogeneity of populations living in the region studied. Russians from the central part of European Russia (Tver, Murom, and Kursk) exhibited similarities with populations from central–eastern Europe, and were distant from Russian sample from the northern Russia (Mezen district, Archangelsk region). Komi samples, especially Izhemski Komi, were significantly different from all other populations studied. These can be considered as a second pole of genetic diversity in northern Europe (in addition to the pole, occupied by Finns), as they had a distinct ancestry component. Russians from Mezen and the Finnic-speaking Veps were positioned between the two poles, but differed from each other in the proportions of Komi and Finnic ancestries. In general, our data provides a more complete genetic map of Europe accounting for the diversity in its most eastern (northeastern) populations.     

长江流域棉花综合群体品系的遗传多样性分析

以15个长江流域棉花杂交种为基础材料,花粉混合互交构建了综合群体,从中选育出29个棉花株系。通过田间试验对12个主要性状进行考察,衍生株系间霜前花率的变异最大,子棉产量及构成因素次之,纤维品质性状的变异最小。主成分分析表明,纤维品质、产量及构成因素、霜前花率、衣分和株高等前5个主成分,对变异方差的贡献率分别为24.312%、19.662%、13.287%、10.812%、9.085%。基于SSR的分子标记差异,绝大多数衍生株系聚在一类,遗传差异较小,明显区别于黄河流域棉花品种。     

High Genetic Diversity of Newcastle Disease Virus in Wild and Domestic Birds in Northeastern China from 2013 to 2015 Reveals Potential Epidemic Trends

Newcastle disease (ND), caused by the virulent Newcastle disease virus (NDV), is one of the most important viral diseases of birds globally, but little is currently known regarding enzootic trends of NDV in northeastern China, especially for class I viruses. Thus, we performed a surveillance study for NDV in northeastern China from 2013 to 2015. A total 755 samples from wild and domestic birds in wetlands and live bird markets (LBMs) were collected, and 10 isolates of NDV were identified. Genetic and phylogenetic analyses showed that five isolates from LBMs belong to class I subgenotype 1b, two (one from wild birds and one from LBMs) belong to the vaccine-like class II genotype II, and three (all from wild birds) belong to class II subgenotype Ib. Interestingly, the five class I isolates had epidemiological connections with viruses from southern, eastern, and southeastern China. Our findings, together with recent prevalence trends of class I and virulent class II NDV in China, suggest possible virus transmission between wild and domestic birds and the potential for an NDV epidemic in the future.     

Several Different Lactase Persistence Associated Alleles and High Diversity of the Lactase Gene in the Admixed Brazilian Population

Adult-type hypolactasia is a common phenotype caused by the lactase enzyme deficiency. The −13910 C>T polymorphism, located 14 Kb upstream of the lactase gene (LCT) in the MCM6 gene was associated with lactase persistence (LP) in Europeans. This polymorphism is rare in Africa but several other variants associated with lactase persistence were observed in Africans. The aims of this study were to identify polymorphisms in the MCM6 region associated with the lactase persistence phenotype and to determine the distribution of LCT gene haplotypes in 981 individuals from North, Northeast and South Brazil. These polymorphisms were genotyped by PCR based methods and sequencing. The −13779*C,−13910*T, −13937*A, −14010*C, −14011*T LP alleles previously described in the MCM6 gene region that acts as an enhancer for the LCT gene were identified in Brazilians. The most common LP allele was −13910*T. Its frequency was highly correlated with European ancestry in the Brazilian populations investigated. The −13910*T was higher (0.295) in southern Brazilians of European ancestry and lower (0.175) in the Northern admixed population. LCT haplotypes were derived from the 10 LCT SNPs genotyped. Overall twenty six haplotypes previously described were identified in the four Brazilian populations studied. The Multidimensional Scaling analysis showed that Belém, in the north, was closer to Amerindians. Northeastern and southern Afro-descendants were more related with Bantu-speaking South Africans whereas the Southern population with European ancestry grouped with Southern and Northern Europeans. This study shows a high variability considering the number of LCT haplotypes observed. Due to the highly admixed nature of the Brazilian populations, the diagnosis of hypolactasia in Brazil, based only in the investigation of the −13910*T allele is an oversimplification.     

Population Genetic Analysis of Streptomyces albidoflavus Reveals Habitat Barriers to Homologous Recombination in the Diversification of Streptomycetes

Examining the population structure and the influence of recombination and ecology on microbial populations makes great sense for understanding microbial evolution and speciation. Streptomycetes are a diverse group of bacteria that are widely distributed in nature and a rich source of useful bioactive compounds; however, they are rarely subjected to population genetic investigations. In this study, we applied a five-gene-based multilocus sequence analysis (MLSA) scheme to 41 strains of Streptomyces albidoflavus derived from diverse sources, mainly insects, sea, and soil. Frequent recombination was detected in S. albidoflavus, supported by multiple lines of evidence from the pairwise homoplasy index (Φ_w) test, phylogenetic discordance, the Shimodaira-Hasegawa (SH) test, and network analysis, underpinning the predominance of homologous recombination within Streptomyces species. A strong habitat signal was also observed in both phylogenetic and Structure 2.3.3 analyses, indicating the importance of ecological difference in shaping the population structure. Moreover, all three habitat-associated groups, particularly the entomic group, demonstrated significantly reduced levels of gene flow with one another, generally revealing habitat barriers to recombination. Therefore, a combined effect of homologous recombination and ecology is inferred for S. albidoflavus, where dynamic evolution is at least partly balanced by the extent that differential distributions of strains among habitats limit genetic exchange. Our study stresses the significance of ecology in microbial speciation and reveals the coexistence of homologous recombination and ecological divergence in the evolution of streptomycetes.     

High Genetic Diversity and Structured Populations of the Oriental Fruit Moth in Its Range of Origin

The oriental fruit moth Grapholita ( = Cydia) molesta is a key fruit pest globally. Despite its economic importance, little is known about its population genetics in its putative native range that includes China. We used five polymorphic microsatellite loci and two mitochondrial gene sequences to characterize the population genetic diversity and genetic structure of G. molesta from nine sublocations in three regions of a major fruit growing area of China. Larval samples were collected throughout the season from peach, and in late season, after host switch by the moth to pome fruit, also from apple and pear. We found high numbers of microsatellite alleles and mitochondrial DNA haplotypes in all regions, together with a high number of private alleles and of haplotypes at all sublocations, providing strong evidence that the sampled area belongs to the origin of this species. Samples collected from peach at all sublocations were geographically structured, and a significant albeit weak pattern of isolation-by-distance was found among populations, likely reflecting the low flight capacity of this moth. Interestingly, populations sampled from apple and pear in the late season showed a structure differing from that of populations sampled from peach throughout the season, indicating a selective host switch of a certain part of the population only. The recently detected various olfactory genotypes in G. molesta may underly this selective host switch. These genetic data yield, for the first time, an understanding of population dynamics of G. molesta in its native range, and of a selective host switch from peach to pome fruit, which may have a broad applicability to other global fruit production areas for designing suitable pest management strategies.     

Genetic Diversity of Giardia duodenalis: Multilocus Genotyping Reveals Zoonotic Potential between Clinical and Environmental Sources in a Metropolitan Region of Brazil

Background

Giardia duodenalis is a flagellate protozoan that parasitizes humans and several other mammals. Protozoan contamination has been regularly documented at important environmental sites, although most of these studies were performed at the species level. There is a lack of studies that correlate environmental contamination and clinical infections in the same region. The aim of this study is to evaluate the genetic diversity of a set of clinical and environmental samples and to use the obtained data to characterize the genetic profile of the distribution of G. duodenalis and the potential for zoonotic transmission in a metropolitan region of Brazil.

Methodology/Principal Findings

The genetic assemblages and subtypes of G. duodenalis isolates obtained from hospitals, a veterinary clinic, a day-care center and important environmental sites were determined via multilocus sequence-based genotyping using three unlinked gene loci. Cysts of Giardia were detected at all of the environmental sites. Mixed assemblages were detected in 25% of the total samples, and an elevated number of haplotypes was identified. The main haplotypes were shared among the groups, and new subtypes were identified at all loci. Ten multilocus genotypes were identified: 7 for assemblage A and 3 for assemblage B.

Conclusions/Significance

There is persistent G. duodenalis contamination at important environmental sites in the city. The identified mixed assemblages likely represent mixed infections, suggesting high endemicity of Giardia in these hosts. Most Giardia isolates obtained in this study displayed zoonotic potential. The high degree of genetic diversity in the isolates obtained from both clinical and environmental samples suggests that multiple sources of infection are likely responsible for the detected contamination events. The finding that many multilocus genotypes (MLGs) and haplotypes are shared by different groups suggests that these sources of infection may be related and indicates that there is a notable risk of human infection caused by Giardia in this region.     

High prevalence and mortality due to Histoplasma capsulatum in the Brazilian Amazon: An autopsy study

BackgroundHistoplasmosis is acquired by inhalation of spores of the dimorphic fungus Histoplasma spp. Although this pathogen is distributed worldwide, it is more prevalent in the Americas. However, the real burden of histoplasmosis remains undefined in many endemic regions.MethodologyWe conducted a series of 61 autopsies to individuals who died in a hospital in the Brazilian Amazon focused on infectious diseases. We performed a detailed histological and microbiological evaluation with genetic characterization of Histoplasma strains with the aim to evaluate the contribution of histoplasmosis to morbidity and mortality. Additionally, we assessed the clinicopathological correlation.Principal findingsEvidence of Histoplasma infection was detected in 21 patients (34%). Eight cases were disseminated infections, all of them occurred in HIV-positive patients. Six cases were localized histoplasmosis, limited to the lungs. In seven patients Histoplasma DNA was detected by PCR in patients with no histological lesions. Histoplasma infection was detected in 38% of HIV-positive patients and was a major contributor to death in 22% of them. Lungs, liver and spleen were affected in all cases of disseminated histoplasmosis. Phylogenetic analysis of the strains suggested a high diversity of Histoplasma species circulating in the Brazilian Amazon. Histoplasmosis was clinically missed in 75% of the disseminated infections.ConclusionsThe high incidence of histoplasmosis, the low index of clinical suspicion, and the severity of the disseminated disease highlight the need of proactively implementing sensitive routine screening methods for this pathogen in endemic areas. Antifungal prophylaxis against Histoplasma should be encouraged in the severely immunocompromised HIV patients in these areas. In conclusion, substantial mortality is associated with disseminated histoplasmosis among HIV-positive patients in the Brazilian Amazon.     

Population Structure and Genetic Diversity in Popular Rice Varieties of India as Evidenced from SSR Analysis

We report here on the phylogenetic analysis, population substructure, and identification of molecular tags of 25 popular rice varieties and four landraces from different ecological belts of India employing a set of 52 simple sequence repeat (SSR) markers. Genetic analysis using the SSR markers categorized the genotypes into two major clusters, distributed according to their pedigree. Population structure analysis suggested that the optimum number of subpopulations was three (K?=?3) in the popular varieties and landraces. At K?=?5 the allelic distribution was much more similar to the phylogenetic dendrogram. The molecular diversity and population structure analysis indicated that there is not much variation among the popular rice cultivars of India. The study has identified SSR markers producing unique alleles, which should aid in the precise identification, maintenance, and genetic purity analysis of rice varieties.     

Seed dispersal and population structure in Vriesea gigantea,a bromeliad from the Brazilian Atlantic Rainforest

Seed dispersal, population structure and the mating system of plant species can have great consequences on the genetic structure of populations. Vriesea gigantea is a bromeliad from southeastern Brazil which is self‐compatible and pollinator dependent for fruit set. Its populations are fertile in terms of the production of flowers, pollen, fruits and seeds. To assess the importance of seed supply for gene flow, colonization and distribution of adult individuals, the seed dispersal and population structure of V. gigantea were studied. Seeds are dispersed over short distances; most seeds land close to the mother plant. This pattern coincides with the reported aggregate distribution of bromeliad seedlings. Population structure results showed high seedling recruitment, because 51.3% developed into adults, although few juveniles reached this stage. This result is different from that for other bromeliad species from different habitat conditions. Seed dispersal and population structural patterns are consistent with previous molecular studies, revealing that V. gigantea populations are genetically structured, with low gene flow and a moderate outcrossing rate. © 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 164 , 317–325.     

Genetic Diversity and Antibiotic Resistance Patterns in a Campylobacter Population Isolated from Poultry Farms in Switzerland

The diversity and genetic interrelation of Campylobacter jejuni and C. coli isolated from Swiss poultry were assessed by three independent typing methods. Samples were derived prior to slaughter from 100 randomly selected flocks (five birds per flock) raised on three different farm types. The observed flock prevalence was 54% in total, with 50% for conventional and 69% for free-range farms. Birds held on farms with a confined roaming area had the lowest prevalence of 37%. Campylobacter isolates were characterized by amplified fragment length polymorphism (AFLP), restriction fragment length polymorphism of flaA PCR fragments (flaA-RFLP), and disk diffusion testing for eight antimicrobial agents that are commonly used in veterinary or human medicine in Switzerland. Analysis of the genotypic results indicates that the Campylobacter population in Swiss poultry is genetically highly diverse. Nevertheless, occasionally, isolates with identical or nearly identical characteristics were isolated from different farms or farm types in different locations. Genetic typing by AFLP and flaA-RFLP was found to be complementary. The majority of isolates (67%) were susceptible to all tested antibiotics; however, single, double, and triple resistances were observed in 7%, 23%, and 2% of the strains, respectively. There was no correlation between genotype and antibiotic resistance. Surprisingly, sulfonamide resistance was frequently found together with streptomycin resistance. Our findings illustrate the results of common genetic exchange in the studied bacterial population.