The evolution of myiasis in humans and other animals in the Old and New Worlds (part I): phylogenetic analyses

Myiasis, the infestation of live vertebrates with dipterous larvae, seems to take two distinct forms that, it has been suggested, evolved from two distinct phylogenetic roots: saprophagous and sanguinivorous. However, the convergent evolution of morphological and life-history traits seems to have had a major role in simplifying this overall assessment of the evolutionary routes by which myiasis arose. Moreover, this somewhat simplistic division is further complicated by the existence of both ectoparasitic and endoparasitic species of myiasis-causing Diptera, the evolutionary affinities of which remain to be resolved. To understand how different forms of parasitism arose, the evolution of the various groups of myiasis-causing flies must be separated from the evolution of the myiasis habit per se. Until recently, evolutionary studies of myiasis-causing flies were little more than discussions of morphology-based taxonomy. Since the mid-1990s, however, several formal phylogenies - based on both morphological and, increasingly, molecular data - have been published, enabling reassessment of the hypotheses concerning myiasis evolution. In part I of this review, we focus on some recent landmark studies in this often-neglected branch of parasitology and draw together phylogenetic studies based on molecular and morphological data to provide a framework for the subsequent analysis of biochemical, immunological, behavioural, biogeographical and fossil evidence relating to the evolution of myiasis.     

Shift from morphological to recent advanced molecular approaches for the identification of nematodes

Nematodes are the most diverse but most minor studied microorganisms found in soil, water, animals, or plants. Either beneficial or pathogenic, they significantly affect human and animal health, plant production and ultimately affect the environmental equilibrium. Knowledge of their taxonomy and biology are the main issues to answer the different challenges associated with these microorganisms. The classical morphology-based nematode taxonomy and biodiversity studies have proved insufficient to identify closely related taxa and have challenged most biologists. Several molecular approaches have been used to supplement morphological methods and solve these problems with markable success. The molecular techniques range from enzyme analysis, protein-based information to DNA sequence analysis. For several decades, efforts have been made to integrate molecular approaches with digital 3D image-capturing technology to improve the identification accuracy of such a taxonomically challenging group and communicate morphological data. This review presents various molecular techniques and provides examples of recent advances in these methods to identify free-living and plant-parasitic nematodes.     

分子生物学技术在蚧虫分类鉴定中的应用

近年来利用分子生物学技术对蚧虫进行分类的研究越来越多。本文主要介绍蚧虫分子生物学研究基础以及RFLP、RAPD、SSR、ISSR和DNA Barcoding技术在蚧虫种类鉴定中的应用概况,并对其应用前景及可能存在的问题进行讨论,以期促进蚧虫类昆虫分子生物学研究。     

Molecular characterization of the mitochondrial cytochrome oxidase I gene of Oestridae species causing obligate myiasis

A 688-bp region of the mitochondrial cytochrome oxidase I gene was sequenced from larvae of 18 species of Oestridae causing obligate myiasis. Larvae belonged to the four Oestridae subfamilies (Cuterebrinae, Gasterophilinae, Hypodermatinae and Oestrinae), which are commonly found throughout the world. Analysis of both nucleotide and amino acid data was performed. Nucleotide sequences included 385 conserved sites and 303 variable sites; mean nucleotide variation between all species was 18.1% and variation within each subfamily ranged from 5.3% to 13.34%. Intraspecific pairwise divergences ranged from 0.14% to 1.59%, and interspecific variation ranged from 0.7% to 27%. Of the 229 amino acids, 76 were variable (60 of which were phylogenetically informative), with some highly conserved residues identified within each subfamily. Phylogenetic analysis showed a strong divergence among the four subfamilies, concordant with classical taxonomy based on morphological and biological features. This study provides the first molecular data set for myiasis-causing Oestridae species, providing an essential database for the molecular identification of these parasites and the assessment of phylogenetic relationships within family Oestridae.     

Algal Taxonomy in Limnology: An Example of the Declining Trend of Taxonomic Studies?

The recent decline in taxonomic studies is well recognized. Algae-related papers (390) published in five leading limnology journals (1971–2004) were consulted to assess similar trends in limnology by taking algae as a test aquatic group. The study showed a decrease of algae-only studies and an increase of multi-group studies (algae plus one or more aquatic groups). Identification of species decreased while mentioning of ecological groups (phytoplankton, epiphytes, etc.) increased while presenting results. Species identification, however, was not associated with number of aquatic groups or number of algal species included in a study. Problems probably lie with the old-fashioned image of taxonomy and it being threatened by the recent advancement in evolutionary and molecular biology. Issues like the changing research patterns in freshwater ecology, scope of limnological works, its workers and relevant journals are also shaping the status of traditional taxonomy in limnology. Practices such as giving of at least (algal) genera in community studies, and of the current names of studied species in physiological/molecular works and also in studies on particular supra-generic taxa (Class, Family, etc.), could help sustaining of taxonomy in limnology.     

Molecular identification of two species of myiasis-causing Cuterebra by multiplex PCR and RFLP

The myiasis-causing flies Cuterebra grisea (Coquillet) and Cuterebra fontinella (Clark) (Diptera: Oestridae) are normally parasites of mice, predominantly of the genus Peromyscus. The morphological similarities of these species and the existence of intermediate morphotypes bearing characters of both species make the identification of adults problematic; furthermore the identification of larvae is apparently not possible. This study presents two molecular approaches to discriminate between these species using specific band patterns: (i) species-specific primers designed in the cytochrome oxidase II (COII) region used in multiplex polymerase chain reaction (PCR) and (ii) restriction fragment length polymorphism (RFLP) on amplified segments of cytochrome oxidase I (COI) gene. Both methods were tested on Cuterebra larvae and on adult museum specimens. The two techniques showed a clear difference between C. grisea and C. fontinella, although species-specific primers were more successful than RFLP for degraded DNA. No intraspecific variation in RFLP and species-specific amplifications were detected for the two species of Cuterebra. The results exhibit discrepancies between molecular and morphological identification, suggesting that some of the adults were misidentified.     

龙胆族(龙胆科)分类与进化研究进展

龙胆是著名的高山植物,广布世界温带地区,不仅可作药用,也可用于园艺观赏。龙胆科分为6个族,其中龙胆族占龙胆科物种数的一半以上;龙胆族包括龙胆亚族和獐牙菜亚族,分类与进化方面的相关研究众多。随着分子生物学在龙胆族研究中的应用,近些年对多个类群进行了研究并发表了多个新属和新种。该文从属间和属内两个层次梳理了龙胆族近年来在分类和进化研究中的最新进展,总结了新发表的类群。并分析了现存的问题,重点强调在今后的研究中用特别注意以下几点:(1)形态性状的准确掌握对龙胆族的研究至关重要;(2)发表新类群应当谨慎;(3)单拷贝核基因有助于解析龙胆族复杂的系统发育关系;(4)关注杂交和多倍化等在龙胆族进化中的作用。     

木霉菌及其系统分类学研究回顾

从木霉菌(Trichodermaspp.)的分类历史、主要分类系统、分子生物学在木霉菌分类研究中的应用、我国木霉菌分类研究状况等几个方面,对木霉菌的分类研究进展进行了简要回顾。近些年来国际上对木霉菌的分类研究取得了显著进展,陆续发表了许多新种,目前木霉属已报道有3组60种和2个变型。另外,对木霉分类研究中存在的问题也进行了讨论。     

Nematode parasites of fishes: recent advances and problems of their research

Although nematodes (Nematoda) belong to the most frequent and the most important parasites of fishes in the freshwater, brackish-water and marine environments throughout the world, the present knowledge of these parasites remains still incomplete, especially as to their biology and ecology, but also taxonomy, phylogeny, zoogeography, and the like. However, a certain progress in the research of fish nematodes has been achieved during recent years. An overview of some of the most important discoveries and results obtained is presented. As an example, existing problems in the taxonomy of these nematodes are shown in the dracunculoid family Philometridae (presently including 109 species in 9 genera), where they are associated mainly with some biological peculiarities of these mostly important tissue parasites. Nematodes of the Dracunculoidea as a whole remain poorly known; for example, of 139 valid species parasitizing fishes, 81 (58%) are known by females only and the males have not yet been described for members of 8 (27%) of genera. A taxonomic revision of this nematode group, based on detailed morphological, life history and molecular studies of individual species, is quite necessary; for the time being, Moravec (2006) has proposed a new classification system of dracunculoids, where, based on previous molecular studies, the Anguillicolidae is no longer listed in Dracunculoidea, but in an independent superfamily Anguillicoloidea. Important results have recently been achieved also in the taxonomy of fish nematodes belonging to other superfamilies, as well as in studies of their geographical distribution and diversity in different parts of the world and those of their biology. Opportunities for more detailed studies of fish nematodes have recently greatly improved with the use of some new methods, in particular SEM and DNA studies. There is a need to create a new classification system of these parasites reflecting phylogenetic relationships; a prerequisite for this is taxonomic revisions of different groups based on detailed studies of individual species, including mainly their morphology, biology and genetics. Further progress should concern studies on various aspects of biology, ecology and host-parasite relationships, because these data may have practical implications.     

The immunology of myiasis: parasite survival and host defense strategies

Infestations by dipterous larvae that feed on dead or living vertebrate tissues for a variable period are known as myiases; these infestations reduce host physiological functions, destroy host tissues and cause significant economic losses to livestock worldwide. Recent advances in understanding the specific and nonspecific immune responses of hosts to infestation by myiasis-causing larvae and the immunological strategies evolved by larvae against the host are reviewed here. The practical implications of immunological knowledge for diagnostic and vaccination strategies are also discussed, with a view to developing environmentally sustainable control methods to be used as an alternative to chemical treatments.     

Presidential address: rediscovering parasites using molecular tools--towards revising the taxonomy of Echinococcus, Giardia and Cryptosporidium

Molecular biology has provided parasitologists with a fantastic variety of techniques that have had a major impact on research into parasites and parasitism. Molecular tools have revealed the extent and nature of genetic diversity in parasites and this information has made a significant contribution to studies on the population genetics and evolutionary biology of parasites. Similarly, epidemiology has benefited enormously from the application of molecular tools in terms of studying parasite life cycles and transmission, and in the development of specific and sensitive methods for diagnosis and surveillance. However, the theme I wish to develop in this paper is concerned with the contribution molecular tools have made to parasite taxonomy and systematics, and in particular, the fact that in many cases molecular tools are validating the proposals made many years ago by taxonomists and biologists which were discounted or not fully accepted at the time. To do this I have chosen four examples (Echinococcus, Entamoeba, Giardia, Cryptosporidium) where recent research involving molecular characterisation has confirmed observations made many years ago and has resulted in a need to revise the taxonomy of different groups of parasites.     

A review of recent taxonomic concepts and developments in the Gracilariaceae (Rhodophyta)

Despite much effort in the past decade to resolve the taxonomy of algae in the Gracilariaceae, the family remains problematic. Genetic concepts have become equivocal as their diagnostic reproductive features have been reported as mixed in single species. Species concepts are often also uncertain since the full phenotypic range of a species is seldom known or appreciated. This paper reviews some of the recent taxonomic controversy from the viewpoints of morphology and molecular biology. As more observations are made, features like spermatangial configuration and mode of gonimoblast nutrition, once considered characteristic of genera and subgenera, are emerging as a continuum of types that may vary within species. Culture studies to establish the reproductive anatomy of a life history and to test crossabilities are invaluable to species concepts but also may yield false results owing to clonal variability and intraspecific sterility. Characteristics of DNA have been effective in clarifying some aspects of gracilariacean taxonomy and phylogeny, and may be able to resolve the major inconsistencies if more widely applied. NRCC 38089     

Relationship between digestive enzymes and food habit of Lutzomyia longipalpis (Diptera: Psychodidae) larvae: Characterization of carbohydrases and digestion of microorganisms

The sandfly Lutzomyia longipalpis (Lutz and Neiva, 1912) is the main vector of American Visceral Leishmaniasis. In spite of its medical importance and several studies concerning adult digestive physiology, biochemistry and molecular biology, very few studies have been carried out to elucidate the digestion in sandfly larvae. Even the breeding sites and food sources of these animals in the field are largely uncharacterized. In this paper, we describe and characterize several carbohydrases from the gut of L. longipalpis larvae, and show that they are probably not acquired from food. The enzyme profile of this insect is consistent with the digestion of fungal and bacterial cells, which were proved to be ingested by larvae under laboratory conditions. In this respect, sandfly larvae might have a detritivore habit in nature, being able to exploit microorganisms usually encountered in the detritus as a food source.     

Genetic Approaches for Studying Myiasis-causing Flies: Molecular Markers and Mitochondrial Genomics

“Myiasis-causing flies” is a generic term that includes species from numerous dipteran families, mainly Calliphoridae and Oestridae, of which blowflies, screwworm flies and botflies are among the most important. This group of flies is characterized by the ability of their larvae to develop in animal flesh. When the host is a live vertebrate, such parasitism by dipterous larvae is known as primary myiasis. Myiasis-causing flies can be classified as saprophagous (free-living species), facultative or obligate parasites. Many of these flies are of great medical and veterinary importance in Brazil because of their role as key livestock insect-pests and vectors of pathogens, in addition to being considered important legal evidence in forensic entomology. The characterization of myiasis-causing flies using molecular markers to study mtDNA (by RFLP) and nuclear DNA (by RAPD and microsatellite) has been used to identify the evolutionary mechanisms responsible for specific patterns of genetic variability. These approaches have been successfully used to analyze the population structures of the New World screwworm fly Cochliomyia hominivorax and the botfly Dermatobia hominis. In this review, various aspects of the organization, evolution and potential applications of the mitochondrial genome of myiasis-causing flies in Brazil, and the analysis of nuclear markers in genetic studies of populations, are discussed.     

Muroid rodents: Phylogeny and evolution

The Muroidea, a group of rodents that includes mice, rats, gerbils, hamsters and others, encompasses a tremendous diversity of fairly recent geological origin. The taxonomy, systematics, phylogeny and paleontology of the muroid rodents have progressed enormously during the last two decades, and many hypotheses on their evolutionary biology have been formalized. Nevertheless, there still remain important unanswered questions - regarding, for example, local conflicts between molecular and paleontological data, or the origin of the fast rate of DNA change in rats and mice - that need more investigation.     

Noninvasive molecular methods to identify live scarab larvae: an example of sympatric pest and nonpest species in New Zealand

Despite the negative impact that many scarab larvae have on agro-ecosystems, very little attention has been paid to their taxonomy. Their often extremely similar morphological characteristics have probably contributed to this impediment, which has also meant that they are very difficult to identify in the field. Molecular methods can overcome this challenge and are particularly useful for the identification of larvae to enable management of pest species occurring sympatrically with nonpest species. However, the invasive collection of DNA samples for such molecular methods is not compatible with subsequent behavioural, developmental or fitness studies. Two noninvasive DNA sampling and DNA analysis methods suitable for the identification of larvae from closely related scarab species were developed here. Using the frass and larval exuviae as sources of DNA, field-collected larvae of Costelytra zealandica (White) and Costelytra brunneum (Broun) (Scarabaeidae: Melolonthinae) were identified by multiplex PCR based on the difference in size of the resulting PCR products. This study also showed that small quantities of frass can be used reliably even 7 days after excretion. This stability of the DNA is of major importance in ecological studies where timeframes rarely allow daily monitoring. The approach developed here is readily transferable to the study of any holometabolous insect species for which morphological identification of larval stages is difficult.     

DNA序列在苔藓分子系统学研究中的应用

DNA是遗传信息的载体,直接对DNA核苷酸排列顺序的分析和比较是研究苔藓分子系统学的最彻底和最理想的方法。本文综述了DNA序列(叶绿体基因组、核基因组和线粒体基因组)在苔藓分子系统学中的应用,探讨了基因片段的选择策略。并提出只有将分子数据和传统分类学取得的研究成果结合起来,构建最合理的系统树,才能更好地推动苔藓分子系统学的发展。     

Techniques of molecular biology in morphological diagnosis of DNA and RNA viruses

Recognition of virus structure and biology as well as the increasingly more complete understanding of pathogenesis in infectious diseases have been possible due to the rapid development of the molecular biology techniques. In the recent few years, most of the studies employing those techniques in diagnosis of infectious diseases concerned the detection of novel viruses, clarification of the virus role in diseases of unknown aetiology and determination of the effect of virus mutants on the course of the infection. The pathogenetic mechanisms of chronic infections, oncogenesis and fibrogenesis are continued to be studied. This paper presents the advantages of using in situ hybridisation in the microscopical diagnosis of viruses. Moreover, principal techniques of amplifying the level of virus detection (in situ PCR and its variants, Immunomax) have been described. Direct application of the Immunomax technique in combination with the in situ hybridisation and with immunocytochemistry have been illustrated with our own studies on tissue expression of selected DNA viruses (HBV and HCMV).     

分子生物学技术在昆虫系统学研究中的应用

分子生物学技术应用于昆虫系统学研究,是８０年代末新兴起来的,近几年来发展相当迅速。为了把握这个研究方向,并促进这个研究领域的发展,作者从研究方法、研究内容、研究对象等方面着手,对近１０年来分子生物学技术应用于昆虫系统学中的研究进展进行了概括和总结。介绍了ＤＮＡ序列测定、ＲＦＬＰ,分子杂交技术、ＲＦＰＬ、分子杂交技术、ＲＡＰＤ、ＳＳＣＰ及ＤＳＣＰ等几种主要方法及其应用情况,并从种及种下阶元的分类鉴定