Standardization of in-house polymerase chain reaction for the identification of Mycobacterium tuberculosis at the reference Tropical Disease Hospital in the State of Goiás, Brazil

This study compares smear, growth in Lowenstein-Jensen medium, and in-house polymerase chain reaction (PCR) techniques for the detection of Mycobacterium tuberculosis. A total of 72 specimens from 72 patients with clinical symptoms of tuberculosis, including 70 sputum and two bronchial aspirate samples, were tested in parallel by smear, culture, and in-house PCR techniques. From these, 48 (66.6%) were negative by the 3 methods, 2 (2.8%) were smear positive and negative by culture and in-house PCR, 11 (15.3%) were both smear and culture negative, and in-house PCR positive, 7 (9.7%) were positive by the 3 methods, 2 (2.8%) were positive by smear and culture, and negative by PCR, 2 (2.8%) were positive by culture and PCR, but smear negative. After the resolution of discrepancies in PCR results, the sensitivity and specificity for in-house PCR technique to M. tuberculosis relative to the culture, were 81.8% and 81.9%, respectively. These results confirm that this method, in-house PCR, may be a sensitive and specific technique for M. tuberculosis detection, occurring in both positive and negative smear and negative cultures.     

Detection of Leishmania braziliensis in human paraffin-embedded tissues from Tucumán, Argentina by polymerase chain reaction

American cutaneous leishmaniasis (ACL) is an endemic disease in Northern Argentina. We applied the polymerase chain reaction (PCR) followed by a hybridization labelled probe to 21 paraffin embedded human skin biopsies, already analyzed histologically, from leishmaniasis endemic areas in the province of Tucumán, Argentina. We used primers previously designed to detect a Leishmania-specific 120-base-pair fragment of kinetoplast DNA minicircle, other two primer pairs that amplify kDNA minicircles belonging to the L. braziliensis and L. mexicana complexes respectively, and specific oligonucleotide primers to detect L. (V.) braziliensis which amplify the sequence of the ribosomal protein L-14 of this species. The PCR-hybridization showed a sensitivity of 90.5% when compared to the histopathology test which was 61.9%. Five of the total samples analyzed were positive for the L. braziliensis complex whilst none was positive for the L. mexicana complex. The specific primers for L. (V.) braziliensis detected the parasite in four samples. These results are consistent with those reported for close endemic areas and demonstrate that the causative agent of human leishmaniasis in the analyzed cases was L. (V.) braziliensis. PCR should be used as a diagnostic tool for tegumentary leishmaniasis, especially in the mucosal form, and as a valuable technique for the identification of the Leishmania species that causes the disease in certain areas.     

Populations and production of fish in two small tributaries of the Paraná River,Paraná, Brazil

Mean biomass (153-1) and production (P) of fish in two small tributaries of the Paraná River (Paraná, Brazil) were 61 kg ha^–1 and 48 kg ha^–1 yr^–1 in the Caracu River and 29 kg ha^–1 and 26 kg ha^–1 yr^–1 in the Agua do Rancho River, respectively. Matrix correlation analysis revealed high positive correlations of both 153-2 and P to maximum depth and hiding places and, at a lower level of significance, to mean depth, pH and oxygen level. Lower 153-3 and P values were found in the Agua do Rancho River, whose valley has retained a more natural character, rich canopy and scarcity of macrophytes, but also lower conductivity and nitrogen and phosphate levels than those in the Caracu River.Address for correspondence     

Biological behaviour in mice of Trypanosoma cruzi isolates from Amazonas and Paraná, Brazil

The biological behaviour of 23 Trypanosoma cruzi isolates in Swiss mice was compared. Nineteen isolates were obtained from patients in the acute phase of Chagas disease (13), sylvatic reservoir hosts (Didelphis marsupialis) (3), and triatomine bugs (Rhodnius robustus) (3) from four regions of the State of Amazonas (AM). Four isolates were obtained from chronic chagasic patients in the State of Paraná (PR): three autochthones, and one allochthone from the State of Minas Gerais. Only one isolate was unable to infect the mice. The AM and PR isolates showed the largest number of significant differences from each other. The former had lower mean values in the pre-patent (5.4 days) and patent (4.6 days) periods (PP), with the parasitaemia (Pmax) reaching a peak of 9.9×10(4) blood trypomastigotes (BT)/mL of blood by the 7th day following inoculation. The AM isolates also had higher positivity to fresh-blood examination (FBE) (84.1%) compared to haemoculture (HC) (58.7%) and polymerase chain reaction (PCR) (33.3%), in addition to higher mortality (2.9%). The PR isolates had higher values for PP (18.5 days) and Pmax (99.9×10(4)BT/mL) as well as higher positivity to FBE (87.2%), HC (100%), and PCR (83.3%). The correlations between the biological behaviour of the T. cruzi isolates and the clinical and epidemiological characteristics of Chagas disease are discussed.     

Population Structure and Genetic Diversity in Sweet Cassava Cultivars from Paraná, Brazil

F _is (0.170) and F _it (0.259) values indicated a number of heterozygotes present in the population under study lower than that necessary to reach the Hardy-Weinberg equilibrium. The genetic variability found among the traditional sweet cassava cultivars assessed was considered wide, and the groups that were most distant were mostly cultivars from Toledo and Maringá.     

Prion protein gene polymorphisms in sheep in the state of Paraná, Brazil

To determine the polymorphisms of the prion protein gene in sheep from the state of Paraná, Brazil, 323 animals of meat breeds (Suffolk, Hampshire Down, Texel, Ile de France, Dorper, Dorset, Santa Inês and crossbreds) were genotyped by restriction fragment length polymorphism (RFLP) analysis. The most frequent allele was ARQ, with a frequency of 0.61, followed by ARR (0.30). VRQ and AHQ alleles were present at very low frequencies (0.13 and 0.05 respectively), and the ARH allele was not found. Seven genotypes were identified (ARR/ARR, ARR/ARQ, ARQ/ARQ, ARR/VRQ, ARR/AHQ, ARQ/VRQ and ARQ/AHQ), of which ARQ/ARQ was the most frequent (0.41). The Santa Inês breed and crossbred animals showed the highest genotypic variability.     

Prevalence and virulence properties of Vibrio cholerae non-O1, Aeromonas spp. and Plesiomonas shigelloides isolated from Cambé Stream (State of Paraná, Brazil)

The incidence of Vibrio cholerae, Aeromonas spp. and Plesiomonas shigelloides was determined in water samples from Cambé Stream. The samples were collected from seven different sites. The serogroups, virulence markers and drug resistance profiles were also evaluated. Twelve Aer. hydrophila, 12Aer. caviae, eight Aer. sobria, seven Ple. shigelloides and two V. cholerae non-O1 were isolated. They belonged to different serogroups and all produced haemolysis in different assays. Five of the Aeromonas strains and one of V cholerae non-O1 were positive for enterotoxin activity. Haemagglutination and its inhibition, using erythrocytes of different origins, was variable for Aeromonas spp. and V. cholerae, while none of the Ple. shigelloides haemagglutinated in association with any type of erythrocyte. All isolates exhibited multiple drug resistance. These results indicate that the occurrence of V. cholerae non-O1, Aeromonas spp. and Ple. shigelloides, in water used for vegetable irrigation, human recreation and animal consumption, among others, represents a potential risk for humans.     

Environmental Influence on Planktonic Cladocerans and Copepods in the Floodplain of the Upper River Paraná, Brazil

The abundance of microcrustacean species recorded in plankton samples from the littoral and pelagic zones of the Upper Paran River, State of Mato Grosso do Sul, Brazil, was evaluated in relation to abiotic and biotic environmental factors. The cladocerans Bosminopsis deitersi and Bosmina hagmann i were most abundant. The presence or absence of aquatic macrophytes also influenced abundance and species composition. The true zooplankton species were most abundant at the pelagic stations, while species of Chydoridae and Macrothricidae contributed markedly to microcrustacean abundance and species richness at littoral stations. Species abundance was also strongly affected by water level variation. Microcrustaceans, mainly bosminids, were most abundant during low water. During high water, bosminids decreased in abundance while most other species increased. The higher variation of the environmental factors during high water probably explains the shift in abundance patterns. Changes in water level increased mixing of littoral and pelagic microcrustaceans between sampling stations.     

Molecular differentiation of species of the genus Zungaro (Siluriformes, Pimelodidae) from the Amazon and Paraná-Paraguay River basins in Brazil

Fish species of the Zungaro genus (Siluriformes, Pimelodidae) are amongst the largest migratory fish in Latin America and have considerable economic importance for commercial fishing in Brazil. However, natural populations of this large catfish are experiencing a severe decline. There are significant taxonomical inconsistencies for this fish. Two geographically separated species of the fish were initially described, one endemic in the Amazon and another in the Paraná-Paraguay River basins. A taxonomic review had recently proposed that there is only one Zungaro species in Brazil, based on morphological data. We made a molecular study of Zungaro populations in an attempt to solve taxonomical inconsistencies and to analyze genetic diversity in natural populations of this genus. We analyzed two regions of the mitochondrial DNA (the control region and the ATPase 6 gene region) of individuals sampled from the Paraná-Paraguay River and Amazon River basins. Analyses based on p-distances and maximum likelihood phylogenetic models showed a genetic difference between populations corresponding to different species. Genetic differentiation between Zungaro populations was at the same level as that observed between other Siluriformes species, using the same DNA sequences. We conclude that Zungaro species of the Paraná-Paraguay River basin do not belong to the same species found in the Amazon basin. This finding has a significant implication for conservation of this fish, given that populations are disappearing at a high rate in the Paraná-Paraguay River basin, mainly due to impoundments.     

Lepidophylloides corumbataensis sp. nov. from the Guadalupian in the Paraná Basin,southern Brazil

A new species of the isoetalean leaf genus Lepidophylloides is described. This Guadalupian species from the Paraná Basin, Lepidophylloides corumbataensis, found as silicified microphylls in the Corumbataí Formation in the municipality of Piracicaba, São Paulo state, in southern Brazil, is the first species of the genus to be formally described for Brazil, and possibly the first for Gondwanaland. This is also the first time that surviving palisade tissue has been found in a species of Lepidophylloides. The new species is compared to known species and shown to be distinct from all previously described from Euramerican and Cathaysian floras. The organization of the xylem in arch-like or crescent-shaped bundles suggests a relation to Cathaysian species. The close association with Lycopodiopsis derbyi stems (which have true leaf scars and must have suffered leaf abscission) suggests that they are different organs from the same species.     

Length-Weight Relationships for freshwater fishes from streams of the Atlantic Rainforest,State of Paraná, Brazil

This study reports the Length-Weight Relationships (LWR) for 10 freshwater fish species sampled in the Atlantic Rainforest, Southern Brazil. Sampling was conducted between 2016 and 2017, in nine streams, using electrofishing. Collected fishes were fixed in formalin, and thus some shrinking was unavoidable. The values of parameter b of the LWR (W = aL^b) ranged between 2.5 and 3.5. We report for the first time the LWR for seven species: Geophagus iporanguensis, Isbrueckerichthys duseni, Hypostomus interruptus, Kronichthys lacerta, Chasmocranu lopezi, Astyanax lacustris e Neoplecostomus ribeirensis. We highlight the importance of the LWR data in supporting further studies on fish stocks and environment in the Atlantic Rainforest, thereby contributing to the database needed to develop fisheries management and maintenance of ecosystem services.     

Neotropical Monogenoidea. 32. Cacatuocotyle paranaensis n. g., n. sp. (Dactylogyridae,Ancyrocephalinae) from Characidium spp. (Teleostei,Characidae) from the State of Paraná, Brazil

Cacatuocotyle paranaensis n. sp. (Dactylogyridae, Ancyrocephalinae) is described from the gills of the characid fishes Characidium lanei Travassos and C. pterostictum Gomes collected from two streams on the coast of the State of Paraná, Brazil. Cacatuocotyle n. g. is proposed for species possessing a single cephalic lobe (terminal), one pair of head organs, a convex haptor with thickened muscular anterior margins, one anchor-bar complex (ventral), seven pairs of ventral hooks (one pair associated with the anchor shafts; one central pair anterior to the bar; five submarginal bilateral pairs) and a sinistral vaginal aperture.     

HLA polymorphism and evaluation of European, African, and Amerindian contribution to the white and mulatto populations from Paraná, Brazil

Polymorphism of classical HLA-A, HLA-B, HLA-C, HLA-DR, and HLA-DQ genes differs greatly among populations, both in frequencies and in the presence of alleles and haplotypes particular to population groups, making these genes powerful tools for the study of origins of populations and their degree of admixture. Antigen, allele, and haplotype frequencies, together with linkage disequilibrium patterns, are reported for 2 populations in the southern Brazilian state of Paraná, one of predominantly European ancestry (white), the other of predominantly African and European ancestry (mulatto). Genetic distance estimates between the 2 groups and other populations studied previously, and of degree of admixture, were performed. In accordance with phenotypic classification, the white population is of predominantly European origin (80.6%), with a smaller contribution of African (12.5%) and Amerindian (7.0%) genes. The mulatto population consists of African (49.5%) and European (41.8%) ancestry, with a smaller but significant contribution of Amerindian (8.7%) ancestry. On the basis of history and population genetics, there is controversy regarding the Amerindian contribution to Paraná's gene pool. These results provide a better picture of Paraná's ethnic constitution and on the Amerindian contribution to the white and mulatto populations.     

Anatomy and genetic diversity of two populations of Schinus terebinthifolius (Anacardiaceae) from the Tibagi River basin in Paraná, Brazil

Knowledge of the effects of flooding on plant survival is relevant for the efficiency of management and conservation programs. Schinus terebinthifolius is a tree of economic and ecological importance that is common in northeast Brazil. Flooding tolerance and genetic variation were investigated in two riparian populations of S. terebinthifolius distributed along two different ecological regions of the Tibagi River basin. Flooding tolerance was evaluated through the investigation of young plants, submitted to different flooding intensities to examine the morphological and anatomical responses to this stress. The growth rate of S. terebinthifolius was not affected by flooding, but total submersion proved to be lethal for 100% of the plants. Morphological alterations such as hypertrophied lenticels were observed in both populations and lenticel openings were significantly higher in plants from one population. Genetic analysis using DNA samples obtained from both populations showed a moderate degree of genetic variation between populations (13.7%); most of the variation was found within populations (86.3%). These results show that for conservation purposes and management of degraded areas, both populations should be preserved and could be used in programs that intend to recompose riparian forests.     

Spatial and temporal variability of saphrophytic bacteria in the surface waters of Paranaguá and Antonina Bays,Paraná, Brazil

Spatial and temporal variability in the concentration of saprophytic bacteria was investigated in surface waters of Paranaguá and Antonina bays.Seven points along a profile from the entrance of Paranaguá Bay to the innermost part of Antonina Bay, were sampled monthly from November 1985 to November 1986. Temperature, dissolved oxygen, pH and precipitation data were also measured.ZoBell 2216E culture medium was utilized with freshwater and 32% salinity water.Principal component analysis of biotic and abiotic data showed a gradient from the innermost bay to the baymouth bar with an increase in bacteria concentration at Antonina Bay. Significant variation was not observed among sampling stations at Paranaguá Bay, except for February 1986 when there was a homogeneous increase of bacteria throughout both bays in association with an increase in temperature and precipitation. There was also no strong variation of saprophyte numbers at sampling locations near Paranaguá city, despite domestic and harbour waste input.We conclude that the outer region of the estuary is highly influenced by the adjacent ocean and that the inner part has typical estuarine characteristics. The region may be considered little polluted.     

Survey of microcystins in water between 1995 and 1996 in Paraná, Brazil using ELISA

An enzyme-linked immunosorbent assay (ELISA) based on a monoclonal antibody was used to determine microcystin (MC) concentrations in water supplies and water plant samples collected between November 1995 and October 1996, from five regions of Paraná, Brazil. In addition, the presence of Microcystis sp. was monitored. Of the 50 samples obtained, 12 were from an urban lake, 8 from human water supplies, 10 from recreational lakes, 13 from farm waters used for animal pasture and 7 from aquaculture facilities. M. aeruginosa was positive in all locations. MCs were positive (>50 pg ml(-1)) in 9 samples (2 samples from human water supplies, 5 from recreational lakes and 2 from animal pasture). Heavy contamination with MCs was observed in water samples collected in May 1996 from 2 recreation (swimming-fishing sites at Itaipu dam, 6380 and 10,000 pg ml(-1)) and human supplies (6627 pg ml(-1)) samples. At these sites, a large bloom of Microcystis sp. was detected. Treatment with 1 ppm Cl- reduced MCs levels, although 267 pg ml(-1) remained in the water plant samples. Our data showed frequent occurrence of Microcystis sp., which may be a hazard to humans and animals in the state of Paraná. More detailed investigations are required to evaluate the risk of natural MC contamination in the water supplied in this region.     

Correlation of Candida species and symptoms among patients with vulvovaginal candidiasis in Maringá, Paraná, Brazil

Vulvovaginal candidiasis (VVC) is an infection caused by abnormal yeast growth in the mucosa of the female genital tract which is commonly diagnosed in gynecology. The aim of this study was to correlate the frequency of yeasts and their respective species in asymptomatic women with different clinical manifestation of VVC; evaluate possible relationships between number of fungus colonies and symptoms in this pathology. All patients who visited the laboratory within a period of five months, for routine examinations of vaginal secretion, independent of the presence or absence of symptoms of VVC were included in this study. Of these, women with immunodeficiency or with an infection of the genital tract by another agent were excluded. Candida albicans was the most frequently yeast isolated (60%). Among non-C. albicans yeasts, 61.5% were isolated of the asymptomatic women, 38.7% from patients with VVC and 11.1% of those from patients with RVVC. C. albicans was associated with symptoms of VVC and while, the presence of non-C. albicans yeasts with asymptomatic women. However, there was no association between the number of fungal colonies and symptoms.     

Toxoplasma gondii infections in cats from Paraná, Brazil: seroprevalence, tissue distribution, and biologic and genetic characterization of isolates

Cats are important in the epidemiology of Toxoplasma gondii because they are the only hosts that can excrete environmentally resistant oocysts. The prevalence of T. gondii was determined in 58 domestic cats from 51 homes from Santa Isabel do Ivai, Parana State, Brazil where a water-associated outbreak of acute toxoplasmosis had occurred in humans. Antibodies to T. gondii were found with the modified agglutination test in 49 of 58 (84.4%) cats at a serum dilution of 1:20. Tissues (brain, heart, and skeletal muscle) of 54 of these cats were bioassayed in T. gondii-free, laboratory-reared cats; T. gondii oocysts were excreted by 33 cats that were fed feline tissues. Brains from these 54 cats were bioassayed in mice; T. gondii was isolated from 7. Skeletal muscles and hearts of 15 cats were also bioassayed in mice; T. gondii was isolated from skeletal muscles of 9 and hearts of 13. The results indicate that T. gondii localizes in muscle tissue more than the brains of cats. In total there were 37 T. gondii isolates from 54 cats. Most isolates of T. gondii were virulent for mice. Genotyping of the 37 isolates of T. gondii, using the SAG2 locus, revealed that 15 isolates were type I and 22 were type III. The absence of type II genotype in cats in this study is consistent with the previous studies on T. gondii isolates from Brazil and is noteworthy because most T. gondii isolates from the United States are type II. These findings support the view that Brazilian and North American T. gondii isolates are genetically distinct. This is the first report of genotyping of T. gondii isolates from the domestic cat.     

Comparison of the Kato-Katz and Helmintex methods for the diagnosis of schistosomiasis in a low-intensity transmission focus in Bandeirantes, Paraná, southern Brazil

The diagnosis of schistosomiasis is problematic in low-intensity transmission areas because parasitological methods lack sensitivity and molecular methods are neither widely available nor extensively validated. Helmintex is a method for isolating eggs from large faecal samples. We report preliminary results of a comparative evaluation of the Helmintex and Kato-Katz (KK) methods for the diagnosis of schistosomiasis in a low-intensity transmission area in Bandeirantes, Paraná, southern Brazil. Eggs were detected by both methods in seven patients, whereas only Helmintex yielded positive results in four individuals. The results confirm the previously demonstrated higher sensitivity of the Helmintex method compared with the KK method.     

Effect of climatic conditions on natural mycoflora and fumonisins in freshly harvested corn of the State of Paraná, Brazil

Natural mycoflora associated with fumonisins were analyzed in 150 samples of freshly harvested corn from Central-Southern, Central-Western and Northern regions of the State of Paraná, Brazil and correlated to climatic conditions. The corn samples were frequently contaminated with Fusarium sp.(98.7 to 100%) and Penicillium sp. (93 to 100%), when compared to Aspergillus sp. (not detected to 27.7%). The highest contamination with potentially mycotoxigenic fungi occurred in corn harvested in the Central-Western region, where total mould and yeast counts ranged from 5.5 × 10³ to 5.2 × 10⁶ CFU/g, with 98.7% contaminated byFusarium sp. and 93% by Penicillium sp. In this region F. moniliforme (F. verticillioides) was the predominant Fusariumsp., and was isolated in 85.9% of the samples. Aspergillus sp. was isolated from 27.7% samples. FB₁ was detected in 100% of the samples (mean of 2.39 g/g) and FB₂ in 97.7% (mean of 1.09 g/g). Fumonisins were also detected in all samples from Northern region, with mean of 4.56 g/g (FB₁) and 2.20 g/g (FB₂).Considering 1.0 g/g as the threshold, 72% of the corn samples from the Central-West and 92% from the North were contaminated with concentrations above this value, in contrast to a 18.5% contamination rate from Central-Southern samples. Between corn planting to harvesting season, the average maximum temperature and relative humidity were 26 °C and 77.1%(Central-Southern), 27 °C and 69% (Northern)and 29.9 °C and 89.1% (Central-Western).Therefore, the higher fumonisins contamination of corn from Northern region when compared to the Central-South were due to the differences in rainfall levels (92.8 mm in Central-Southern, 202 mm in Northern) during the month preceding harvest.This revised version was published online in October 2005 with corrections to the Cover Date.