Inoculation effects of Nitrospirillum amazonense and biofertilizer in sugarcane / Efeitos da inoculação de Nitrospirillum amazonense e biofertilizante em cana-de-açúcar

In this study, we analyzed the hypothesis that the combination of Nitrospirillum amazonense strain BR11145 with biological fertilizer prepared using Microgeo® and native microbiome from location of the product application results in morphological and nutritional gains for the initial development of sugarcane plants and soil chemistry. For this purpose, pre-sprouted sugarcane seedlings were grown in a greenhouse mesocosm experiment using soil amended with nitrogen/phosphorus/potassium fertilizer. The experimental treatments consisted of: 1) biological fertilizer with the addition of N. amazonense (100 mL ha-1), 2) biological fertilizer without the addition of N. amazonense, 3) inoculation with N. amazonense at a dose of 100 mL ha-1 with 2x108 viable cell mL-1, 4) inoculation with N. amazonense at a dose of 200 mL ha-1 with 2x108 viable cell mL-1, and 5) control, without the addition of biological fertilizer and N. amazonense. The biological fertilizer was applied at dose of 300 L ha-1, which was split at planting (200 L ha-1) and in the post-emergence phase (100 L ha-1). After 164 days of planting, it was detected an increase in leaf length +3, number of green leaves, leaf area and sulfur content in the leaves of sugarcane plants that received the biological fertilizer with the addition of N. amazonense. In conclusion, the combination of N. amazonense with biological fertilizer revealed positive effects through morphological and nutritional characteristics in sugarcane plants during their early stages of development when compared to plants grown only with the inoculation of N. amazonense, biological fertilizer or mineral fertilizers, with few notable positive effects on soil chemistry.

Neste estudo avaliou-se a hipótese de que a combinação de Nitrospirillum amazonense estirpe BR11145 com adubo biológico preparado com base em microbioma autóctone da localidade de aplicação do produto, produzido com Microgeo®, resulta em benefícios morfológicos e nutricionais para o desenvolvimento inicial de plantas de cana-de-açúcar e para a química do solo. Para tanto, mudas pré-brotadas de cana-de-açúcar foram crescidas em mesocosmos num experimento conduzido em casa-de-vegetação com solo enriquecido com fertilizante à base de nitrogênio/fósforo/potássio. Os tratamentos avaliados foram: 1) adubo biológico acrescido de N. amazonense (100 mL ha-1), 2) adubo biológico sem o acréscimo de N. amazonense, 3) inoculação de N. amazonense na dose de 100 mL ha-1 (2x108 células viáveis por mL), 4) inoculação de N. amazonense na dose de 200 mL ha-1 (2x108 células viáveis por mL), e 5) testemunha, sem a adição de fertilizante biológico e N. amazonense. O adubo biológico foi aplicado na dose de 300 L ha-1 sendo esta parcelada no plantio (200 L ha-1) e na fase de pós-emergência (100 L ha-1). Após 164 dias do plantio constatou-se aumento no comprimento da folha +3, número de folhas verdes, área foliar e no teor de enxofre nas folhas das plantas de cana-de-açúcar que receberam o adubo biológico combinado com N. amazonense. Conclui-se que a combinação de N. amazonense com o adubo biológico produzido com Microgeo® revelou efeitos positivos por meio de caraterísticas morfológicas e nutricionais em plantas de cana-de-açúcar durante os seus estádios iniciais de desenvolvimento quando comparadas com plantas crescidas apenas com a inoculação de N. amazonense, fertilizante biológico ou fertilizantes minerais, com poucos efeitos positivos notáveis na química do solo.

A multi-attribute approach to evaluating the impact of biostimulants on crop performance.

An ever-growing collection of commercial biostimulants is becoming available in a wide variety of forms and compositions to improve crop performance. Given the intricate nature of deciphering the underlying mechanisms of commercial products, which typically comprise various biological components, it is crucial for research in this area to have robust tools to demonstrate their effectiveness in field trials. Here, we took a multi-attribute approach to evaluating the impact of biostimulants on crop performance. First, we assessed the impact of a biostimulant on the soil and rhizosphere microbiomes associated to crops in eight reference farms, including corn (3 farms), soybean (2), cotton (2) and sugarcane (1), in different biomes and production contexts in Brazil and Paraguay. Second, we modeled a set of integrated indicators to measure crop responses to biostimulant application, including five analytical themes as follows: i) crop development and production (9 indicators), ii) soil chemistry (9), iii) soil physics (5), iv) soil biology (6) and v) plant health (10). Amplicon 16S rRNA and ITS sequencing revealed that the use of the biostimulant consistently changes the structure of bacterial and fungal communities associated with the production system for all evaluated crops. In the rhizosphere samples, the most responsive bacterial taxa to biostimulant application were Prevotella in cotton; Prauserella and Methylovirgula in corn; and Methylocapsa in sugar cane. The most responsive fungal taxa to biostimulant use were Arachnomyces in soybean and cotton; and Rhizophlyctis in corn. The proposed integrated indicators yielded highly favorable positive impact indices (averaging at 0.80), indicating that biostimulant-treated fields correlate with better plant development and crop performance. Prominent indices were observed for indicators in four themes: soil biology (average index 0.84), crop production (0.81), soil physics (compaction reduction 0.81), and chemical fertility (0.75). The multi-attribute approach employed in this study offers an effective strategy for assessing the efficacy of biostimulant products across a wide range of crops and production systems.

Co-infection patterns of vector-borne zoonotic pathogens in owned free-ranging dogs in central Chile.

We investigated the co-occurrence of the nine of the most relevant canine vector-borne pathogens (CVBP) using conventional and real-time PCR and evaluated risk factors and potential non-apparent haematological alterations associated with co-infection in 111 rural, owned, free-ranging dogs in the Metropolitan Region of Chile.At least one pathogen was detected in 75% of the dogs. DNA of Anaplasma platys (Ap; 36%), Candidatus Mycoplasma haematoparvum (CMhp; 31%), Mycoplasma haemocanis (Mhc; 28%), Trypanosoma cruzi (17%), Leishmania spp. (4.5%), and Acanthocheilonema reconditum (1%) was detected. All dogs were negative for Ehrlichia spp., Rickettsia spp., Bartonella spp., Piroplasmida, and Hepatozoon spp. Thirty-eight dogs (34%) were coinfected. CMhp was involved in 71%, Mhc in 58%, and Ap in 50% of the co-infections. The most common co-infection pattern was CMhp-Mhc (37% of the cases). The prevalence of Ap was higher in juvenile than in adult dogs, whereas the opposite was found for CMhp and Mhc. Adult dogs were four times more likely of being co-infected than juveniles. Co-infected animals showed higher white blood cell count, segmented neutrophil count, and GGT levels than non-co-infected dogs. Clinically healthy but infected dogs may act as reservoirs of CVBP, and their free-ranging behavior would facilitate the spread of these pathogens to other dogs as well as human beings or wild carnivores.

Large-scale survey for canine vector-borne parasites in free-ranging dogs and foxes from six diverse bioclimatic regions of Chile.

Chile is a large country with a marked range of climate conditions that make it an ideal scenario for the study of vector-borne parasites (VBPs); however, knowledge about their distribution is limited to a few confined areas of this country. The presence of Hepatozoon spp., piroplasmids, Leishmania spp. and filarioids was investigated through molecular and serological methods in blood and serum samples of 764 free-ranging rural dogs, 154 Andean foxes (Lycalopex culpaeus), and 91 South American grey foxes (Lycalopex griseus) from six bioclimatic regions across Chile. Hepatozoon spp. DNA was exclusively detected in foxes (43% prevalence), including sequences closely related to Hepatozoon felis (24.1%; only Andean foxes), Hepatozoon americanum (16.2%; only grey foxes), and Hepatozoon canis (1.25%; in one grey fox). Risk factor assessment identified a higher probability of Hepatozoon infection in juvenile foxes. DNA of piroplasmids was detected in 0.7% of dogs (Babesia vogeli) but in no fox, whilst antibodies against Babesia sp. were detected in 24% of the dogs and 25% of the foxes, suggesting a wider circulation of canine piroplasmids than previously believed. A positive association between the presence of antibodies against Babesia and high Rhipicephalus sanguineus sensu lato burden was observed in dogs. Leishmania spp. DNA and antibodies were detected in 0.8% and 4.4% of the dogs, respectively. Acanthocheilonema reconditum was the only blood nematode detected (1.5% of the dogs and no fox). Differences in prevalence among bioregions were observed for some of the VBPs. These results expand our knowledge about the occurrence of vector-borne parasites in Chile, some of which are firstly reported herein. This information will facilitate the diagnosis of vector-borne diseases in domestic dogs and improve the control measures for both domestic and wild canids.

Fasciola hepatica in wild boar (Sus scrofa) from Italy.

Fasciola hepatica is a trematode infecting ruminants worldwide, occasionally reported in a wide range of animal species, including humans. According to the WHO, fasciolosis is recognized as a re-emerging neglected tropical disease, responsible for endemic and epidemic outbreaks in humans. Although the main hosts of the parasite are represented by cattle, sheep and goats, wildlife may be involved in its circulation. Here we firstly report F. hepatica in a wild boar from Italy (southern area) and characterize it both morphologically and molecularly. The nad1 gene analysis of specimens analyzed, revealed a high genetic similarity with those of humans from Iran and Peru, as well as a close phylogenetic relationship to those in ruminants from Brazil, Ecuador and Egypt. Considering the increase in the wild boar populations in urban and peri-urban areas, a potential role of this ungulate in the circulation of this zoonotic trematode is suggested.

Molecular detection of vector-borne agents in ectoparasites and reptiles from Brazil.

Trombidiformes and Mesostigmata mites, as well as Ixodida ticks, infest ectothermic tetrapods worldwide, potentially acting as vectors of bacteria, viruses and protozoa. The relationship among ectoparasites, transmitted pathogenic agents (e.g., Borrelia spp., Coxiella spp., Hepatozoon spp., and Rickettsia spp.) and ectothermic hosts has been scarcely investigated. This research focuses on a large collection of Brazilian herpetofauna screened for the presence of arthropod ectoparasites and vector-borne microbial agents. Reptiles (n = 121) and amphibians (n = 49) from various locations were infested by ectoparasites. Following genomic extraction, microbial agents were detected in 81 % of the Acari (i.e. n = 113 mites and n = 26 ticks). None of the mites, ticks and tissues from amphibians yielded positive results for any of the screened agents. Blood was collected from reptiles and processed through blood cytology and molecular analyses (n = 48). Of those, six snakes (12.5 %) showed intraerythrocytic alterations compatible with Hepatozoon spp. gamonts and Iridovirus inclusions. Hepatozoon spp. similar to Hepatozoon ayorgbor and Hepatozoon musa were molecularly identified from seven hosts, two mite and two tick species. Rickettsia spp. (e.g., Rickettsia amblyommatis, Rickettsia bellii-like, Rickettsia sp.) were detected molecularly from four mite species and Amblyomma rotundatum ticks. Phylogenetic analyses confirmed the molecular identification of the above-mentioned microbial agents of mites and ticks related to snakes and lizards. Overall, our findings highlighted that the Brazilian herpetofauna and its ectoparasites harbour potentially pathogenic agents, particularly from the northern and south-eastern regions. The detection of several species of spotted fever group Rickettsia pointed out the potential role of ectothermic hosts and related arthropod ectoparasites in the epidemiological cycle of these bacteria in Brazil.

Molecular detection of vector-borne agents in ectoparasites and reptiles from Brazil

Trombidiformes and Mesostigmata mites, as well as Ixodida ticks, infest ectothermic tetrapods worldwide, potentially acting as vectors of bacteria, viruses and protozoa. The relationship among ectoparasites, transmitted pathogenic agents (e.g., Borrelia spp., Coxiella spp., Hepatozoon spp., and Rickettsia spp.) and ectothermic hosts has been scarcely investigated. This research focuses on a large collection of Brazilian herpetofauna screened for the presence of arthropod ectoparasites and vector-borne microbial agents. Reptiles (n = 121) and amphibians (n = 49) from various locations were infested by ectoparasites. Following genomic extraction, microbial agents were detected in 81 % of the Acari (i.e. n = 113 mites and n = 26 ticks). None of the mites, ticks and tissues from amphibians yielded positive results for any of the screened agents. Blood was collected from reptiles and processed through blood cytology and molecular analyses (n = 48). Of those, six snakes (12.5 %) showed intraerythrocytic alterations compatible with Hepatozoon spp. gamonts and Iridovirus inclusions. Hepatozoon spp. similar to Hepatozoon ayorgbor and Hepatozoon musa were molecularly identified from seven hosts, two mite and two tick species. Rickettsia spp. (e.g., Rickettsia amblyommatis, Rickettsia bellii-like, Rickettsia sp.) were detected molecularly from four mite species and Amblyomma rotundatum ticks. Phylogenetic analyses confirmed the molecular identification of the above-mentioned microbial agents of mites and ticks related to snakes and lizards. Overall, our findings highlighted that the Brazilian herpetofauna and its ectoparasites harbour potentially pathogenic agents, particularly from the northern and south-eastern regions. The detection of several species of spotted fever group Rickettsia pointed out the potential role of ectothermic hosts and related arthropod ectoparasites in the epidemiological cycle of these bacteria in Brazil.

Morphological and phylogenetic analyses of Lutzomyia migonei from three Brazilian states.

Lutzomyia migonei is incriminated as a vector of Leishmania braziliensis, the main causative agent of cutaneous leishmaniasis in Brazil. Recently, this phlebotomine sand fly species has been suggested as a vector for Leishmania infantum, which causes zoonotic visceral leishmaniasis. Considering the widespread distribution of Lu. migonei in South America, the existence of isolated populations has been hypothesized. Three Lu. migonei populations, two from north-eastern Brazil (Machados, Pernambuco State, and Baturité, Ceará State) and other from the south-eastern region (Niterói, Rio de Janeiro State) were analysed both morphologically and genetically. Though no significant morphological differences were found amongst the sand fly specimens analysed, discriminant analysis based on specific morphometric characters (i.e., length of wing, antennal segment 3 and coxite for males, and length of wing and antennal segment 3 for females), showed that specimens from Machados were closer to Baturité than to Niterói. The molecular analysis of cytochrome c oxidase subunit I gene sequences also supported this observation by the distinct separation of two monophyletic clades, grouping specimens from Machados and Baturité separately from those of Niterói. Our results suggest the existence of different populations within the distribution range of Lu. migonei. Whether these populations are reproductively isolated and/or present differences in terms of vector competence/capacity for L. braziliensis and L. infantum needs to be further investigated.

Biological compatibility between two temperate lineages of brown dog ticks, Rhipicephalus sanguineus (sensu lato).

BACKGROUND: The brown dog tick Rhipicephalus sanguineus (sensu stricto) is reputed to be the most widespread tick of domestic dogs worldwide and has also been implicated in the transmission of many pathogens to dogs and humans. For more than two centuries, Rh. sanguineus (s.s.) was regarded as a single taxon, even considering its poor original description and the inexistence of a type specimen. However, genetic and crossbreeding experiments have indicated the existence of at least two distinct taxa within this name: the so-called "temperate" and "tropical" lineages of Rh. sanguineus (sensu lato). Recent genetic studies have also demonstrated the existence of additional lineages of Rh. sanguineus (s.l.) in Europe and Asia. Herein, we assessed the biological compatibility between two lineages of Rh. sanguineus (s.l.) found in southern Europe, namely Rhipicephalus sp. I (from Italy) and Rhipicephalus sp. II (from Portugal). METHODS: Ticks morphologically identified as Rh. sanguineus (s.l.) were collected in southern Portugal and southern Italy. Tick colonies were established and crossbreeding experiments conducted. Morphological, biological and genetic analyses were conducted. RESULTS: Crossbreeding experiments confirmed that ticks from the two studied lineages were able to mate and generate fertile hybrids. Hybrid adult ticks always presented the same genotype of the mother, confirming maternal inheritance of mtDNA. However, larvae and nymphs originated from Rhipicephalus sp. I females presented mtDNA genotype of either Rhipicephalus sp. I or Rhipicephalus sp. II, suggesting the occurrence of paternal inheritance or mitochondrial heteroplasmy. While biologically compatible, these lineages are distinct genetically and phenotypically. CONCLUSIONS: The temperate lineages of Rh. sanguineus (s.l.) studied herein are biologically compatible and genetic data obtained from both pure and hybrid lines indicate the occurrence of paternal inheritance or mitochondrial heteroplasmy. This study opens new research avenues and raises question regarding the usefulness of genetic data and crossbreeding experiments as criteria for the definition of cryptic species in ticks.

Influences of different space allowance on reproductive performances in buffalo

The aim of this study was to evaluate the effect of two different conditions of space allowance on reproductive performance and oxidative parameters, biochemical and hormonal profiles in buffalo. The trial was carried out on one hundred pluriparous buffaloes divided into two different groups. Buffaloes in group HDG (high density group - n = 50) were maintained in open yards that allowed 10 m2/head while those in group LDG (low density group -n = 50) were maintained in 22 m2/head. After 60 days, 45 buffaloes in each group underwent synchronization of ovulation by Ovsynch and were artificially inseminated to assess the reproductive efficiency. On the day of AI blood samples were collected to evaluate oxidative stress, hormonal and metabolic profile. Furthermore, on the same day, blood, saliva and hair samples were collected to assess cortisol levels. Simultaneously, Five buffaloes/group, were synchronized but not inseminated and on the day of the hypothetical timed artificial insemination (TAI), follicular fluid was recovered by OPU and blood samples were collected from each animal to evaluate the redox status on both plasma and follicular fluid. Conception rate on day 70 post-AI was similar between the two groups (57.5 vs. 62.5%, in LDG and HDG, respectively). No significant differences were found on redox status, metabolic and hormonal profile and cortisol levels between the groups. In conclusion, on the conditions of this experiment it was observed that the space allowance of 10 m 2/head did not affect reproductive efficiency in buffalo cows.

Influences of different space allowance on reproductive performances in buffalo

The aim of this study was to evaluate the effect of two different conditions of space allowance on reproductive performance and oxidative parameters, biochemical and hormonal profiles in buffalo. The trial was carried out on one hundred pluriparous buffaloes divided into two different groups. Buffaloes in group HDG (high density group - n = 50) were maintained in open yards that allowed 10 m2/head while those in group LDG (low density group -n = 50) were maintained in 22 m2/head. After 60 days, 45 buffaloes in each group underwent synchronization of ovulation by Ovsynch and were artificially inseminated to assess the reproductive efficiency. On the day of AI blood samples were collected to evaluate oxidative stress, hormonal and metabolic profile. Furthermore, on the same day, blood, saliva and hair samples were collected to assess cortisol levels. Simultaneously, Five buffaloes/group, were synchronized but not inseminated and on the day of the hypothetical timed artificial insemination (TAI), follicular fluid was recovered by OPU and blood samples were collected from each animal to evaluate the redox status on both plasma and follicular fluid. Conception rate on day 70 post-AI was similar between the two groups (57.5 vs. 62.5%, in LDG and HDG, respectively). No significant differences were found on redox status, metabolic and hormonal profile and cortisol levels between the groups. In conclusion, on the conditions of this experiment it was observed that the space allowance of 10 m 2/head did not affect reproductive efficiency in buffalo cows.(AU)

Genetic characterization of Rhipicephalus sanguineus (sensu lato) ticks from dogs in Portugal.

BACKGROUND: The taxonomic status of the brown dog tick Rhipicephalus sanguineus (sensu stricto) is a subject of on-going debate; there is a consensus that populations of this tick species should be referred to as R. sanguineus (sensu lato) until its taxonomic status is resolved. Recent genetic studies revealed the existence of more than one lineage of R. sanguineus (s.l.) in temperate countries. In this study, we assessed the genetic identity of ticks collected from rural dogs living in several areas located in all major geographical regions of Portugal. METHODS: A total of 347 ticks were collected from rural dogs living in different regions of Portugal. These ticks were morphologically identified and partial mitochondrial 16S rRNA gene sequences (~300 bp) were obtained from representative specimens. RESULTS: The ticks were morphologically identified as Ixodes ricinus (seven males and 27 females), Rhipicephalus bursa (one male), Rhipicephalus pusillus (one female) and R. sanguineus (s.l.) (two larvae, 101 nymphs, 108 males and 100 females). Partial mitochondrial 16S rRNA gene sequences were obtained from 58 R. sanguineus (s.l.) specimens, and all of them were genetically identified as belonging to the so-called temperate lineage of R. sanguineus (s.l.) CONCLUSIONS: These results strongly suggest that the temperate species of R. sanguineus (s.l.) is the only representative of this tick group found on dogs in Portugal. It also adds weight to the hypothesis that Rhipicephalus turanicus is not present in this country, although further investigations are necessary to confirm this.

Ecological insights from assessments of phenotypic plasticity in a Neotropical species of Drosophila.

Several authors have called attention to the evolutionary importance of phenotypic plasticity and niche construction, because such phenomena require a new status and a new perspective. Drosophila species are traditionally used as models in investigations of phenotypic plasticity, although the majority of such research has been conducted with species of the subgenus Sophophora, primarily Drosophila melanogaster. In this study, we investigated the phenotypic plasticity of Drosophila cardini, a Neotropical species of the subgenus Drosophila, and focused on the wing size, wing shape, thorax length and wing: thorax ratio of lines that were collected in the Brazilian savanna and exposed to different temperatures during growth. All of the analyzed traits presented plasticity to temperature, and the reaction norms were similar to those previously found in other drosophilid species; in addition, the maximum values were consistent with the temperature variations at the collection sites. The specimens that emerged at low temperatures were larger and had more rounded wings compared with those that emerged at high temperatures, which were smaller and had narrower wings. We hypothesized that the differences observed in the shape of the wings might be associated with flight performance. Nevertheless, further investigation of the relationships among wing shape, wing loading and flight performance is required. Investigations on phenotypic plasticity using species with diverse ecologies should help us to better understand how this phenomenon operates in nature, and studies of this type must be encouraged.

The white-nosed coati (Nasua narica) is a naturally susceptible definitive host for the zoonotic nematode Angiostrongylus costaricensis in Costa Rica.

Angiostrongylus costaricensis (Strongylida, Angiostrongylidae) is a roundworm of rodents, which may cause a severe or fatal zoonosis in several countries of the Americas. A single report indicated that the white-nosed coati (Nasua narica), acts as a potential free-ranging wildlife reservoir. Here we investigated the prevalence and features of A. costaricensis infection in two procyonid species, the white-nosed coati and the raccoon (Procyon lotor) from Costa Rica to better understand their possible role in the epidemiology of this zoonotic infection. Eighteen of 32 (56.2%) white-nosed coatis collected between July 2010 and March 2016 were infected with A. costaricensis but none of 97 raccoons from the same localities were diagnosed with this infection. First-stage larvae of A. costaricensis were obtained from feces of 17 fresh white-nosed coati carcasses by Baermann technique. Parasite identity was confirmed by morphology, histology and molecular characterization of target genes. These data demonstrate that the white-nosed coati is a naturally susceptible definitive host for A. costaricensis in Costa Rica contrary to findings in the raccoon.

Morphological and genetic diversity of Rhipicephalus sanguineus sensu lato from the New and Old Worlds.

BACKGROUND: The taxonomic status of the brown dog tick (Rhipicephalus sanguineus sensu stricto), which has long been regarded as the most widespread tick worldwide and a vector of many pathogens to dogs and humans, is currently under dispute. METHODS: We conducted a comprehensive morphological and genetic study of 278 representative specimens, which belonged to different species (i.e., Rhipicephalus bursa, R. guilhoni, R. microplus, R. muhsamae, R. pusillus, R. sanguineus sensu lato, and R. turanicus) collected from Europe, Asia, Americas, and Oceania. After detailed morphological examination, ticks were molecularly processed for the analysis of partial mitochondrial (16S rDNA, 12S rDNA, and cox1) gene sequences. RESULTS: In addition to R. sanguineus s.l. and R. turanicus, three different operational taxonomic units (namely, R. sp. I, R. sp. II, and R. sp. III) were found on dogs. These operational taxonomical units were morphologically and genetically different from R. sanguineus s.l. and R. turanicus. Ticks identified as R. sanguineus s.l., which corresponds to the so-called "tropical species" (= northern lineage), were found in all continents and genetically it represents a sister group of R. guilhoni. R. turanicus was found on a wide range of hosts in Italy and also on dogs in Greece. CONCLUSIONS: The tropical species and the temperate species (= southern lineage) are paraphyletic groups. The occurrence of R. turanicus in the Mediterranean region is confirmed. A consensual re-description of R. sanguineus s.s. and R. turanicus will be necessary to solve the taxonomic problems within the so-called R. sanguineus group.