Valdir de Queiroz Balbino

Detection of Bovine Papillomavirus Types, Co-Infection and a Putative New BPV11 Subtype in Cattle

The aim of this study was to evaluate the presence of different types of Bovine papillomavirus (BPV) in cattle skin lesions and to identify new viral types in Brazil. A total of 72 skin lesions were analysed from 66 different bovines by PCR using degenerate and specific primers, and subsequent sequencing. Sequencing quality was determined using Staden package with Phred 30. Similarity analysis was performed with BioEdit and BLAST programs to verify the identity with known BPV types. Phylogenetic analysis was carried out using Maximum Likelihood method with TIM3 + G as nucleotide substitution model in PAUP*, and 1000 non-parametric bootstrap replicates. Analyses revealed the presence of ten different types of BPV in the samples, with the exception of BPV7. The presence of co-infections was very high as almost all samples (89%) were co-infected. A putative new BPV11 subtype was also found in lesions from different animals. These results add significant knowledge about the prevalence and diversity of BPV infection in Brazilian cattle, which could be used in future studies aiming at the development of more specific treatment and diagnostic methods.

Lutzomyia longipalpis s.l. in Brazil and the impact of the Sao Francisco River in the speciation of this sand fly vector

Lutzomyia longipalpis s.l. (Diptera: Psychodidae) is the principal vector of Leishmania infantum chagasi in the Americas, and constitutes a complex of species. Various studies have suggested an incipient speciation process based on behavioral isolation driven by the chemotype of male sexual pheromones. It is well known that natural barriers, such as mountains and rivers can directly influence population divergence in several organisms, including insects. In this work we investigated the potential role played by the Sao Francisco River in eastern Brazil in defining the current distribution of Lu. longipalpis s.l. Our studies were based on analyses of polymorphisms of the cytochrome b gene (cyt b) sequences from Lu. longipalpis s.l. available in public databases, and from additional field-caught individuals. Altogether, 9 distinct populations and 89 haplotypes were represented in the analyses. Lu. longipalpis s.l. populations were grouped according to their distribution in regards to the 10°S parallel: north of 10°S (<10°S); and south of 10°S (>10°S). Our results suggest that although no polymorphisms were fixed, moderate genetic divergences were observed between the groups analyzed (i.e., FST= 0.184; and Nm = 2.22), and were mostly driven by genetic drift. The population divergence time estimated between the sand fly groups was about 0.45 million years (MY), coinciding with the time of the change in the course of the Sao Francisco River, during the Mindel glaciation. Overall, the polymorphisms on the cyt b haplotypes and the current speciation process detected in Lu. longipalpis s.l. with regards to the distribution of male sexual pheromones suggest a role of the Sao Francisco River as a significant geographical barrier in this process.

Species of Lasiodiplodia associated with papaya stem-end rot in Brazil

This study aims to identify and characterize species of Lasiodiplodia associated with stem-end rot of papaya in six different populations in the Northeast of Brazil. Fungal identifications were made using a combination of morphology together with a phylogenetic analysis based on partial translation elongation factor 1-α sequence (EF-1α) and internal transcribed spacers (ITS). Five species of Lasiodiplodia were identified: Lasiodiplodia brasiliense sp. nov., L. hormozganensis, L. marypalme sp. nov., L. pseudotheobromae and L. theobromae. Only L. theobromae had previously been reported in papaya, while all the other species are reported for the first time in association with this host in Brazil and worldwide. Lasiodiplodia theobromae was the most prevalent species. All species of Lasiodiplodia were pathogenic on papaya fruit, with L. hormozganensis being the most virulent.

Meta-analysis of Brazilian genetic admixture and comparison with other Latin America countries.

This study aims at performing a systematic review and meta‐analysis with the studies of genetic admixture inference of Brazilian population and to compare these results with the genetic admixture levels in other Latin American countries.

Phlebotomine sandflies (Diptera: Psychodidae) of the Atlantic forest in Recife, Pernambuco state, Brazil: the species coming to human bait, and their seasonal and monthly variations over a 2-year period.

Abstract In a study of the phlebotomine sandflies (Diptera: Psychodidae) in a forest reserve in Recife, Pernambuco state, north–eastern Brazil, the sandflies landing on human bait between 1.00 and 1.42 h after sunset were collected weekly for 2 years. Although 10,287 sandflies of 10 Lutzomyia species were collected, almost all (96.5%) of the sandflies caught were Lu. umbratilis. This species and several others caught are potential vectors of some of the Leishmania parasites that cause human disease. The recorded landing rate for Lu. umbratilis peaked, at the high level of 333.3 flies/person-hour, during the collections made in May 2003. The relative rarity in the collections of males of some of the species caught probably indicates that these species do not lek on their bloodmeal sources. It is likely that the sizes of the local populations of species that are not very anthrophilic, such as Lu. flaviscutellata, are much larger than indicated by the collections made on human bait.

First report of Lutzomyia (Nyssomyia) umbratilis Ward & Frahia, 1977 outside of Amazonian Region, in Recife, State of Pernambuco, Brazil (Diptera: Psychodidae: Phlebotominae)

Lutzomyia umbratilis, a known vector of Leishmania guyanensis in the north of Amazon basin, has been exclusively found in the Amazonian region. Here we report for the first time the occurrence of this species in northeastern Brazil. The epidemiological importance of the occurrence of this species in the Atlantic Forest is commented.

Molecular epidemiology of bovine papillomatosis and the identification of a putative new virus type in Brazilian cattle

Bovine papillomaviruses (BPVs) are a diverse group of double-stranded DNA viruses, of which 12 viral types have been detected and characterized so far. However, there is still a limited understanding of the diversity of BPV. Several putative new BPVs have been detected and some of these have been recently characterized as new viral types. However, only a very limited amount of information is available on the pathology associated with these novel viral types yet this information could be of significant value in improving our understanding of the biology of BPV. The objective of this study was to examine some of the epidemiological features of cutaneous bovine papillomatosis in Brazilian cattle, in particular to establish the relationship between BPV types isolated from beef and dairy cattle herds and the lesions they cause. Seventy-two cutaneous lesions were collected from 60 animals. Histopathological, PCR and sequencing assays were conducted to characterize the lesions and detect the BPV types responsible. Phylogenetic analysis was carried out using the maximum likelihood method. BPV types 1-6 and 8-10 were found, as well as a putative new BPV type that belongs to the Deltapapillomavirus genus. The tumors were all classified as fibropapillomas. This is believed to be the first record of BPV types 3 and 10 associated with fibropapillomas. These results confirm that there is a wide range of BPV types that infect cattle, and that an understanding of this diversity is necessary for improved methods of therapeutic treatment.

Staphylococcal toxin genes in strains isolated from cows with subclinical mastitis

The present study was carried out in 11 dairy herds in four municipal districts of the rural area of the State of Pernambuco, Brazil. Out of 984 quarter milk (246 cows), 10 (1.0%) were positive for clinical mastitis, 562 (57.1%) for subclinical mastitis and 412 (41.9%) were negative. A total of 81 Staphylococcus spp. isolates were obtained from milk samples from the cows diagnosed with subclinical mastitis. From these, 53 (65.0%) were S. aureus, 16 (20.0%) coagulase-positive staphylococci (CPS) and 12 (15.0%) coagulase-negative staphylococci (CNS). The isolates were further investigated for the presence of toxin genes by multiplex and uniplex PCR. The main gene observed was seg followed by seh, sei and sej. The distribution of these observed genes among the isolates obtained from different areas showed a regional pattern for the SEs. The presence of toxin genes in the strains isolated from bovine milk demonstrates a potential problem for public health.

Evaluation of the tannic acid inhibitory effect against the NorA efflux pump of Staphylococcus aureus

During the early periods of antibiotic usage, bacterial infections were considered tamed. However, widespread antibiotic use has promoted the emergence of antibiotic-resistant pathogens, including multidrug resistant strains. Active efflux is a mechanism for bacterial resistance to inhibitory substances, known simply as drug efflux pumps. The bacterium Staphylococcus aureus is an important pathogenic bacterium responsible for an array of infections. The NorA efflux pump has been shown to be responsible for moderate fluoroquinolone resistance of S. aureus. The inhibition of the efflux pump was assayed using a sub-inhibitory concentration of standard efflux pump inhibitors and tannic acid (MIC/8), where its capacity to decrease the MIC of Ethidium bromide (EtBr) and antibiotics due to the possible inhibitory effect of these substances was observed. The MICs of EtBr and antibiotics were significantly reduced in the presence of tannic acid, indicating the inhibitory effect of this agent against the efflux pumps of both strains causing a three-fold reduction of the MIC when compared with the control. These results indicate the possible usage of tannic acid as an adjuvant in antibiotic therapy against multidrug resistant bacteria (MDR).

Status of pyrethroid resistance and mechanisms in Brazilian populations of Tuta absoluta.

The tomato leafminer, Tuta absoluta, is a major pest of tomato crops worldwide. This study surveyed the resistance of T. absoluta populations from four regions in Brazil to pyrethroid insecticides, the frequencies of L1014F, T929I and M918T Na channel mutations, and the role of detoxification metabolism in the resistance. Resistance ratios varied from 1- to 11-times among populations and insecticides, but control failure likelihood assays showed that all pyrethroids assessed exhibited no efficacy at all (and thus, 98-100% control failure likelihood) against all T. absoluta populations. The activity of glutathione S-transferase and cytochrome P450-mediated N-demethylation in biochemical assays was significantly correlated with the level of resistance to deltamethrin and permethrin suggesting that these enzymes may play a role in resistance. TaqMan assays were used to screen for the presence of knockdown resistance (kdr) mutations and revealed that the L1014F kdr mutation was fixed in all populations and associated with two super-kdr mutations, M918T and particularly T929I, at high frequency. Altogether, results suggest that control failures are because of mutations in the domain II of the sodium channel, as a prevailing mechanism of resistance to pyrethroids in populations of T. absoluta in Brazil. But, enhanced cytochrome P450-dependent monooxygenases and GST activities also play an important role in the resistance of some populations, which reinforce that pyrethroids must not be used overall to control T. absoluta.