Diversity of Culturable Bacteria Isolated From the Feces of Wild Anopheles darlingi (Diptera: Culicidae) Mosquitoes From the Brazilian Amazon.

Microorganisms living in the midgut of Anopheles mosquitoes have been studied to fight vector-borne diseases, such as malaria. Studies on the microbiota of the Neotropical Anopheles darlingi, the most important Brazilian vector for malaria, have been reported for the same purpose. Our aims were to isolate and identify culturable bacteria from An. darlingi mosquito guts through their feces and to estimate the species richness and the frequency distribution of the sampled bacteria. Sixty wild females of An. darlingi mosquitoes were captured at two rural locations, near Porto Velho, Rondônia, Brazil. Bacteria were isolated from mosquito feces, which were collected using cages which permit the collection of feces on LB nutrient agar plates. Sixty bacterial colonies were isolated and stored in glycerol at -80°C. Bacteria were identified by sequencing their 16S rRNA gene obtained using PCR and Sanger sequencing. To aid in species identification, MALDI-TOF, VITEK2, and BBL Crystal were used as complementary protocols. The sequences obtained from the 60 bacterial isolates were compared to sequences deposited in GenBank (NCBI) using BLAST. Homology greater than 97% between the query and the subject was used as the criteria for assigning the identity of each isolate. Fourteen species from eight different genera were identified among the 60 isolates. The most frequent species were Serratia liquefaciens (20%) and Serratia marcescens (15%). Due to their established apathogenicity and according to previous studies, we suggest Serratia and Pantoea species as suitable for paratransgenesis development to fight malaria in Brazilian Amazon.

Multilocus analysis of divergence and introgression in sympatric and allopatric sibling species of the Lutzomyia longipalpis complex in Brazil.

BACKGROUND: Lutzomyia longipalpis, the main vector of visceral leishmaniasis in Latin America, is a complex of sibling species. In Brazil, a number of very closely related sibling species have been revealed by the analyses of copulation songs, sex pheromones and molecular markers. However, the level of divergence and gene flow between the sibling species remains unclear. Brazilian populations of this vector can be divided in two main groups: one producing Burst-type songs and the Cembrene-1 pheromone and a second more diverse group producing various Pulse song subtypes and different pheromones. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed 21 nuclear loci in two pairs of Brazilian populations: two sympatric populations from the Sobral locality (1S and 2S) in northeastern Brazil and two allopatric populations from the Lapinha and Pancas localities in southeastern Brazil. Pancas and Sobral 2S are populations of the Burst/Cembrene-1 species while Lapinha and Sobral 1S are two putative incipient species producing the same pheromone and similar Pulse song subtypes. The multilocus analysis strongly suggests the occurrence of gene flow during the divergence between the sibling species, with different levels of introgression between loci. Moreover, this differential introgression is asymmetrical, with estimated gene flow being higher in the direction of the Burst/Cembrene-1 species. CONCLUSIONS/SIGNIFICANCE: The results indicate that introgressive hybridization has been a crucial phenomenon in shaping the genome of the L. longipalpis complex. This has possible epidemiological implications and is particularly interesting considering the potential for increased introgression caused by man-made environmental changes and the current trend of leishmaniasis urbanization in Brazil.

Investigation into the prevalence, persistence and antibiotic resistance profiles of staphylococci isolated from euro currency.

AIMS: The study set out to sample € 10 banknotes for the presence of coagulase-positive staphylococci (CPS) such as Staphylococcus aureus (S. aureus) and coagulase-negative staphylococci (CoNS) in Southern Ireland, to assess the levels of antibiotic resistance among those isolated, and determine the persistence of S. aureus on € 10 banknotes and € 2 coins. METHODS AND RESULTS: We report that 97% of € 10 banknotes screened (n = 155) harboured multiple species of staphylococci. From the generated bank of strains, a total of 150 representative staphylococci isolates were used for further study, 71 were CPS and 79 were CoNS. Of these, we found that 62% of the staphylococci demonstrated resistance to at least one of the first-line antibiotics (52.11% of CPS isolates and 76.71% of the CoNS isolates). Resistance to multiple antibiotics was seen in 31.18% of the resistant isolates. In relation to persistence studies, S. aureus was shown to remain viable on euro banknotes and coins for significant periods (on average, 19.33 days on € 10 banknotes and 16.67 days on € 2 coins) as determined using bioluminescence. CONCLUSIONS: We advocate the expansion of antibiotic surveillance programs, with a view to tracking/monitoring antibiotic resistance dissemination among environmental contaminants. Additionally, we propose that 'cashless transactions' should be encouraged in high-risk environments such as hospitals and healthcare settings, as well as stricter infection controls. SIGNIFICANCE AND IMPACT OF THE STUDY: Although it is accepted that circulating currency has the potential to harbour disease-causing pathogens, studies investigating prevalence and persistence of such pathogens on euro currency are virtually nonexistent. In an attempt to rectify this, we examined the prevalence of staphylococci on € 10 banknotes in Ireland and reported relatively high levels of antibiotic resistance among the isolates. Furthermore, we have established the persistence of S. aureus on euro currency for the first time.

Molecular and behavioral differentiation among Brazilian populations of Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae).

BACKGROUND: Lutzomyia longipalpis is the primary vector of American visceral leishmaniasis. There is strong evidence that L. longipalpis is a species complex, but until recently the existence of sibling species among Brazilian populations was considered a controversial issue. In addition, there is still no consensus regarding the number of species occurring in this complex. METHODOLOGY/PRINCIPAL FINDINGS: Using period, a gene that controls circadian rhythms and affects interpulse interval periodicity of the male courtship songs in Drosophila melanogaster and close relatives, we analyzed the molecular polymorphism in a number of L. longipalpis samples from different regions in Brazil and compared the results with our previously published data using the same marker. We also studied the male copulation songs and pheromones from some of these populations. The results obtained so far suggest the existence of two main groups of populations in Brazil, one group representing a single species with males producing Burst-type copulation songs and cembrene-1 pheromones; and a second group that is more heterogeneous and probably represents a number of incipient species producing different combinations of Pulse-type songs and pheromones. CONCLUSIONS/SIGNIFICANCE: Our results reveal a high level of complexity in the divergence and gene-flow among Brazilian populations of the L. longipalpis species complex. This raises important questions concerning the epidemiological consequences of this incipient speciation process.

Multilocus analysis of introgression between two sand fly vectors of leishmaniasis.

BACKGROUND: The phlebotomine sand flies (Diptera:Psychodidae) Lutzomyia (Nyssomyia) intermedia Lutz & Neiva 1912 and Lutzomyia (Nyssomyia) whitmani Antunes & Coutinho 1932 are two very closely related species and important vectors of American cutaneous leishmaniasis. Two single-locus studies have revealed evidence for introgression between the two species in both mitochondrial and nuclear genomes. These findings have prompted the development of a multilocus approach to investigate in more detail the genetic exchanges between the two species. RESULTS: We analyzed ten nuclear loci using the "isolation with migration" model implemented in the IM program, finding evidence for introgression from L. intermedia towards L. whitmani in three loci. These results confirm that introgression is occurring between the two species and suggest variation in the effects of gene flow among the different regions of the genome. CONCLUSION: The demonstration that these two vectors are not fully reproductively isolated might have important epidemiological consequences as these species could be exchanging genes controlling aspects of their vectorial capacity.

Germline polymorphism of codon 727 of human thyroid-stimulating hormone receptor is associated with toxic multinodular goiter.

Toxic multinodular goiter (TMNG) represents a frequent cause of endogenous hyperthyroidism, affecting 5-15% of such patients (with higher frequencies reported in iodine-deficient areas of the world). Although mutations of human TSH receptor (hTSHR) have been described in autonomously functioning thyroid nodules (AFTN), the role of such mutations in the pathogenesis of TMNG remains unclear. To search for alterations of hTSHR in AFTN and TMNG, we performed bidirectional, dye primer automated fluorescent DNA sequencing of the entire transmembrane domain and cytoplasmic tail of hTSHR (TMD+CT-hTSHR) using DNA extracted from nodular regions of 24 patients with TMNG and 7 patients with AFTN. Eight of the 24 patients (33.3%) showed heterozygote polymorphism of codon 727 on the cytoplasmic tail of hTSHR with an amino acid substitution of aspartic acid to glutamic acid. Three of 24 (12.5%) patients with TMNG were found to carry a heterozygote mutation of codon 703, resulting in substitution of alanine with glycine. One patient had multiple heterozygote mutations including I606M (Ile to Met), A703G (Ala to Gly), Q720E (Gln to Glu), and D727E (Asp to Glu). Two patients exhibited silent polymorphism of codons 460 and 618. We found no mutation of the TMD+CT-hTSHR in 7 patients with AFTN, except for a silent polymorphism of codon 460 in 1. DNA fingerprinting of codon 727 using restriction enzyme NlaIII and genomic DNA confirmed the sequencing results in all cases, indicating that the sequence alterations were not somatic in nature. This technique was also used to examine peripheral blood genomic DNA from 52 normal individuals and 49 patients with Graves' disease; 33.3% of TMNG (P = 0.019 vs. normal subjects), 16.3% of Graves' disease patients (P = 0.10 vs. normal subjects), and 9.6% of normal individuals were heterozygous for the D727E polymorphism. Expression of the D727E hTSHR variant in eukaryotic cells (COS-7) resulted in an exaggerated cAMP response to TSH stimulation compared to that of the wild-type hTSHR. These findings indicate that a germline polymorphism of codon D727E of hTSHR is associated with TMNG, suggesting that its presence is an important predisposing genetic factor in the pathogenesis of TMNG.

Ischemic injury and extracellular amino acid accumulation in hippocampal area CA1 are not dependent upon an intact septo-hippocampal pathway.

The septo-hippocampal pathway contains a major gamma-aminobutyric acid (GABA) projection to dendritic fields within the hippocampus. To determine the importance of the septo-hippocampal pathway in ischemia-induced accumulation of GABA and subsequent cell death in area CA1 of hippocampus, septo-hippocampal deafferentation of adult gerbils was performed. Electrolytic lesions were produced in the medial or medial plus lateral septal regions in gerbils 7 days prior to being subjected to 5 min forebrain ischemia. The extent of deafferentation of the dorsal hippocampus was determined histochemically by acetylcholinesterase staining. Both the medial and medial plus lateral septal lesions produced nearly complete loss of acetylcholinesterase staining in the dorsal hippocampus indicating relatively complete deafferentation. During and following ischemia, in vivo microdialysis was used to measure extracellular GABA accumulation, which reached concentrations up to 1060 +/- 143% of basal. Septo-hippocampal deafferentation in both groups of lesioned animals failed to prevent the accumulation of GABA (and glutamate) induced by ischemia, indicating that ischemia-induced GABA accumulation in area CA1 arises principally from intrinsic GABAergic interneurons. Ischemic animals with medial septal lesions did not demonstrate neuroprotection or increased damage in the stratum pyramidale 7 days after reperfusion. Since the septo-hippocampal pathway provides the source of GABAergic disinhibition within the hippocampus, neither disinhibition nor the septo-hippocampal input appear to play an important role in the development of ischemia-induced neuronal death in the hippocampus.

Evolution of neuroablative surgery for involuntary movement disorders: an historical review.

Surgical therapy of involuntary movement disorders has evolved during the past century from gross destructive ablations of the central nervous system to refined, accurate, discrete lesioning of sites deep within the brain. The understanding of neuroanatomic and physiological systems improved tremendously through experimentation in animals and empirical observations of surgery in humans. A continuum of accumulated knowledge has been achieved through ablation or lesioning of virtually all aspects of the central and peripheral nervous system predicated on previous successes or failures. This compilation of surgical history of involuntary movement disorders has provided present neurosurgeons with the foundations on which they base their therapeutic measures and will direct future endeavors within this field.

Neuropsychiatric complications after temporal lobe limbic system surgery.

Although control of seizure activity is the main goal of temporal lobectomies for complex partial epilepsy, the results of surgery must be viewed within the larger context of possible ramifications of limbic system disruption. The neuropsychiatric complications may be significant and influence the patient's perceived qualify of life, thus detracting from the benefits of seizure control. It is apparent from the previous studies that the modalities affected are influenced by the laterality of the resection, with left-sided lobectomies more often affecting verbal memory and learning, whereas right-sided resections may cause visual/spatial memory and cognitive declines. Preoperative neuropsychologic testing may help to predict these possible complications. General psychosocial function, memory, and cognition all seem to be less affected when complete seizure control is attained, with nearly 80% experiencing no significant change in neuropsychologic testing and the gain and losses being equal in magnitude. Also, younger patients and those with a shorter history of seizure activity may fare better after surgery because of a greater capacity for neuronal plasticity to subserve these functions. These findings support the contention that early diagnosis and surgical treatment may help to minimize potential neuropsychiatric complications in conjunction with improved postoperative seizure control.

History of spinal cord stereotaxy.

Stereotactic and functional neurosurgery has experienced a remarkable degree of development during the last 50 years, from the plaster of Paris frame of Spiegel and Wycis to the technology of frameless stereotaxis. Although predominantly used for intracranial procedures, stereotaxy has its roots in experimental studies of the spinal cord. The field of spinal cord stereotaxy has not received the same amount of attention as supratentorial surgery, but there have been significant contributions to the field that have helped to further our understanding of spinal cord anatomy and physiology. Now that frameless stereotaxis has reached clinical practice, there may be further developments in the field of spinal surgery: this technique may prove useful for spinal fusion operations and, possibly, intramedullary operations as well.

Recurrence of a cerebral arteriovenous malformation after surgical excision. Case report.

Complete excision of a cerebral arteriovenous malformation (AVM) should eliminate the future risk of an associated intracranial hemorrhage. Because total removal of an AVM may be difficult to assess at the time of surgery, postoperative angiography has become the accepted standard for documenting that the removal has been accomplished. However, even angiography confirmed excision of an AVM does not completely ensure against rebleeding. Regrowth of an AVM with subsequent hemorrhage can occur. This has been documented in children and is attributed to forces acting on the immature vasculature of these younger patients. The authors report the case of an older patient whose AVM recurred when he was 28 years of age, despite an angiography proven complete excision, and emphasize that, even in adults, angiography documentation of total removal does not always eliminate the risk of reformation of an AVM.

Glomus jugulare tumor metastatic to the sacrum after high-dose radiation therapy: case report.

A 47-year-old woman with left ear pain and hearing loss was diagnosed with a glomus jugulare tumor for which she received radiation therapy as the primary treatment. Over a period of 20 years, she developed temporal bone necrosis, brain stem calcifications, local tumor recurrence, and eventually metastases to her lungs and sacrum. This case underscores the often indolent nature of glomus jugulare tumors, the late sequelae of radiation therapy for benign intracranial tumors, and the potential of these tumors to metastasize. This patient's history suggests that aggressive surgical resection should be considered early for such tumors, particularly because radiation treatment does not ablate the tumor. This is only the second reported case of a glomus jugulare tumor metastatic to the sacrum.

Failure to detect conventional enterotoxins in classical enteropathogenic (serotyped) Escherichia coli strains of proven pathogenicity.

Ammonium sulfate-precipitated supernatants of classical enteropathogenic Escherichia coli strains were negative when investigated for enterotoxin production in rabbit ligated ileal loops, rabbit skin vascular permeability factor tests, suckling mice, and Y-1 adrenal cells. They also failed to stimulate guanylate cyclase activity in homogenates of rabbit, rat, and infant mouse intestines. Furthermore, DNA from enteropathogenic E. coli lacked sequences that encode heat-labile and heat-stable enterotoxins. These studies fail to show conventional enterotoxin synthesis by classical enteropathogenic E. coli.