Solenopsis invicta virus-1A (SINV-1A): distinct species or genotype of SINV-1?

We have cloned and sequenced a 2845 bp cDNA representing the 3'-end of either a new picorna-like virus species or genotype of Solenopsis invicta virus-1 (SINV-1). Analysis of the nucleotide sequence revealed 1 large open reading frame. The amino acid sequence of the translated open reading frame was most identical to structural proteins of SINV-1 (97%), followed by the Kashmir bee virus (KBV, 30%), and acute bee paralysis virus (ABPV, 29%). A PCR-based survey for SINV-1 and the new species or genotype (tentatively named S. invicta virus-1A, SINV-1A) using RNA extracts of S. invicta collected around Gainesville, Florida, revealed a mean colony infestation rate of 25% by SINV-1 and 55% by SINV-1A. Both SINV-1 and SINV-1A were found to co-infect 17.5% of the nests surveyed. Although the data preclude definitive species or genotype assignment, there is no doubt that SINV-1A is distinct from SINV-1, identifiable, and infects S. invicta. We provide a simple RT-PCR technique capable of discerning SINV-1 and SINV-1A infection of S. invicta.     

Detection of Thelohania solenopsae (Microsporidia: Thelohaniidae) in Solenopsis invicta (Hymenoptera: Formicidae) by multiplex PCR

Oligonucleotide primer pairs were designed to unique areas of the small subunit (16S) rRNA gene of Thelohania solenopsae and a region of the Gp-9 gene of Solenopsis invicta. Multiplex PCR resulted in sensitive and specific detection of T. solenopsae infection of S. invicta. The T. solenopsae-specific primer pair only amplified DNA from T. solenopsae and T. solenopsae-infected S. invicta. This primer pair did not produce any amplification products from DNA preparations from uninfected S. invicta, seven additional species of microsporidia (including Vairimorpha invictae), or Mattesia spp. The Gp-9-specific primers recognized and amplified DNA from Solenopsis xyloni, Solenopsis richteri, Solenopsis geminata, the invicta/richteri hybrid, and monogyne and polygyne S. invicta, but not from T. solenopsae, and, as such, served as a positive control verifying successful DNA preparation. Multiplex PCR detected T. solenopsae in worker fire ants infected with as few as 5000 spores. Furthermore, multiplex PCR detected T. solenopsae in all developmental stages of S. invicta. However, detection could be made more sensitive by using only the T. solenopsae-specific primer pair; ants infected with as few as 10 spores were able to be discerned. Multiplex PCR detection of T. solenopsae offers the advantages of a positive control, a single PCR amplification, detection of all developmental stages, and increased sensitivity and specificity compared with microscopy.     

Colony genetic structure and queen mating frequency in fire ants of the subgenus Solenopsis (Hymenoptera: Formicidae)

Colony genetic structure was studied in natural populations of three fire ant taxa, Solenopsis richteri Forel, S. geminata (Fabr.), and hybrid S. invicta/richteri , using allozyme markers. All colonies studied exhibited arrays of female genotypes predicted under a model of monogyny (single functional queen) and monoandry (single insemination of queens). Males produced in the colonies appear to originate exclusively from the foundress queen, rather than from any virgin females present in the colonies. Thus these social insect colonies represent simple, albeit enormous, family groups. Single insemination and foundress parentage of males appear to be conserved reproductive traits in the subgenus Solenopsis , whereas another major determinant of colony genetic structure, the number of functional queens, is evolutionarily labile in this group.     

入侵中国大陆的红火蚁的鉴定及发生为害调查

根据形态特征鉴定结果首次证实红火蚁SolenopsisinvictaBuren入侵中国大陆 ,并调查了广东省吴川市红火蚁发生为害情况。调查结果表明 ,发生区内部分地点红火蚁发生密度较高 ,主要发生于较稳定的生态环境中 ,如荒坡、草地、长满杂草的田埂等。红火蚁已对当地农业生产、人们的身体健康、日常生活等造成了不利影响。此外 ,Cytb基因序列分析结果表明 ,采自美国佛罗里达和广东吴川的红火蚁与采自广东 4个地方的热带火蚁S .geminata (Fabr.)两物种间有 61个变异位点 ,种内变异为 0。限制性酶切多态实验 (RFLP)的结果显示 ,BamHI酶在红火蚁的特异扩增片段中有一个识别位点 ,而在热带火蚁中无酶切位点 ;MspI酶在热带火蚁的扩增片段中也有一个识别位点 ,而在红火蚁中无识别位点。有酶切位点的扩增产物在酶切后分别获得近 2 0 0bp与 2 5 0bp的片段各一带。因此 ,用PCR RFLP的方法可以简单快速地鉴别红火蚁和热带火蚁。     

Solenopsis invicta virus-1 tissue tropism and intra-colony infection rate in the red imported fire ant: a quantitative PCR-based study

Quantitative real-time PCR was employed to measure the Solenopsis invicta virus 1 (SINV-1) load in tissues, individuals, and among colonies of the red imported fire ant, S. invicta Buren. Among tissues examined from SINV-1-infected adults and larvae, the alimentary canal (specifically the midgut) consistently had the highest number of SINV-1 genome copies (91.1 and 99.9%, respectively). Negative staining of a supernatant of the gut homogenate demonstrated the presence of spherical virus particles with a diameter of 30-35 nm, consistent with SINV-1. The number of SINV-1 genome copies in infected larvae and workers from the same queenright colonies were similar to each other. In other words, the infection rate was consistent among both developmental stages. No significant differences were observed in SINV-1 genome copy number among infected colonies sampled during the winter and summer. Although the SINV-1 infection rate of summer-collected mounds was previously shown to be six-times higher than winter-collected mounds, the intra-colony infection rate appears to be unaffected by season. Perhaps less inter-mound interaction during the winter months among S. invicta restricts spread of the virus. A positive correlation between intra-colony infection rate and mean SINV-1 genome copy number per ant was also observed. Based on these results, it is likely that SINV-1 replicates in gut epithelia of S. invicta and virus is shed into the gut lumen where it may be transmitted to nestmates by trophallaxis.     

Use of ribosomal DNA sequence data to characterize and detect a neogregarine pathogen of Solenopsis invicta (Hymenoptera: Formicidae)

A neogregarine parasite of the red imported fire ant, Solenopsis invicta, was discovered recently in Florida and tentatively placed in the Mattesia genus based on morphological characterization. S. invicta infected with this Mattesia species exhibited a characteristic yellowing of the cuticle which was designated Mattesia "yellow-head disease" (YHD). The 18S rRNA gene sequence from Mattesia YHD was elucidated and compared with the neogregarine pathogens, Mattesia geminata and Ophriocystis elektroscirrha. The sequence data support the previous conclusion that Mattesia YHD is a new species that infects S. invicta. Furthermore, high sequence identity between Mattesia YHD, M. geminata (95.7%), and O. elektroscirrha (86.2%) correctly place the YHD organism in the Mattesia genus and Neogregarinorida order. Oligonucleotide primer pairs were designed to unique areas of the 18S rRNA genes of Mattesia YHD and S. invicta. Multiplex PCR resulted in sensitive and specific detection of Mattesia YHD infection of S. invicta.     

Interspecific competition between Solenopsis invicta and two native ant species, Pheidole fervens and Monomorium chinense

This study was designed to understand the effects of the interspecific competition between red imported fire ant, Solenopsis invicta Buren and two native ant species, Pheidole fervens Smith and Monomorium chinense Santschi, by conducting colony interference and individual confrontation tests under laboratory conditions. The colony interference test showed that both native ant species, owing to their numerical advantage, killed the Solenopsis invicta virus-1 (SINV-1)-infected or healthy queens of S. invicta. Significantly less time was required for M. chinense to kill all SINV-1-infected S. invicta compared with the time required to kill the healthy S. invicta. Compared with healthy S. invicta, SINV-1-infected S. invicta spent a longer time eliminating the P. fervens colonies. In confrontation tests, M. chinense killed a significantly higher number of infected S. invicta minors than they did healthy minors, but the number of S. invicta majors (either infected or healthy) killed was substantially less. This study found that the viral infection weakened the competitive ability of S. invicta and made them prone to be eliminated by M. chinense but not by P. fervens.     

Mechanisms of interspecific competition among an invasive and two native fire ants

The mechanisms of interspecific competition among an invasive and two native Solenopsis fire ant forms were investigated in a series of laboratory experiments. In separate trials each with a different food resource, the native S. geminata × xyloni retrieved the greatest amount of a protein- and lipid-rich artificial food resource and a high protein natural food resource, and the native S. geminata retrieved the greatest amount of a high carbohydrate food resource. In trials investigating aspects of interference competition at the colony level, the invasive S. invicta proved to be initially more aggressive than S. geminata , but less aggressive than S. geminata × xyloni . Solenopsis invicta eventually controlled more of the foraging arenas against both native forms when colonies were equivalent by worker biomass, but not when colonies were equivalent by worker number. When paired with S. invicta , S. geminata suffered a significantly greater proportional reduction in both workers and entire colonies when colonies were initially standardized by worker biomass, but not when colonies were standardized by worker number. When paired with S. invicta , a significantly greater proportional reduction of workers occurred in S. geminata × xyloni , regardless of how colonies were standardized. In pairwise trials at the individual level, majors always exhibited significantly less mortality than minors, regardless of the Solenopsis form. The majors of both native forms suffered significantly less mortality than those of S. invicta . Superiority in colony-level interference ability appears to be an important mechanism allowing S. invicta to displace native Solenopsis forms. The ability of S. invicta to reach high population densities, because of intrinsic biological characteristics or an escape from natural enemies, plays an important contributory role. Similar mechanisms may underlie the success of other invasive ant species.     

Similarity in venom alkaloid chemistry of alate queens of imported fire ants: implication for hybridization between Solenopsis richteri and S. invicta in the Southern United States

Both cis- and trans-2-methyl-6-undecylpiperidines, MC11P, have been previously reported as the major components of the venom of alate queens of the imported fire ants, Solenopsis richteri (black) and S. invicta (red). To identify the minor components of venom alkaloids from alate queens and compare the venom alkaloid chemistry of alate queen of their hybrid (S. richteri×S. invicta) with that of the two parental fire ant species (S. richteri and S. invicta), silica-gel short-column chromatography was utilized for separating cis-stereoisomers of venom alkaloids from trans-stereoisomers. GC/MS Analyses of venom-alkaloid chemistry of alate queens demonstrated that fewer alkaloid peaks were detected in the chromatograms of the alate queens compared to those of workers. Three new compounds, 7, 12, and 13, were detected as minor components in the venom of alate queens of all three fire ant species. Alate queens of hybrid fire ants showed cis- and trans-alkaloid patterns similar to those of the parental species. Similarity in venom-alkaloid chemistry of alate queens of S. richteri and S. invicta, and their hybrid may indicate their reproductive compatibility in the hybrid zone in southern United States, where all three species occur sympatrically.     

红火蚁及其他两种蚂蚁毒腺细菌鉴定与多样性分析(英文)

【目的】本研究旨在分析红火蚁Solenopsis invicta毒腺细菌群落多样性,并与热带火蚁Solenopsis geminata和聚纹双刺猛蚁Diacamma rugosum比较毒腺细菌群落差异。【方法】采用Illumine Hiseq 2500测序平台对红火蚁(工蚁、有翅蚁和蚁后)、热带火蚁(工蚁)及聚纹双刺猛蚁(工蚁)毒腺细菌群落16S rRNA基因V3-V4区测序,基于测序数据进行生物信息学分析。【结果】变形菌门(Proteobacteria)在红火蚁工蚁、有翅蚁和蚁后以及热带火蚁工蚁毒腺中占优势,而厚壁菌门(Firmicutes)在聚纹双刺猛蚁工蚁毒腺中占优势。与红火蚁工蚁和有翅蚁相比,柔壁菌门(Tenericutes)在红火蚁蚁后毒腺中更丰富。广州红火蚁蚁后毒腺中假单胞杆菌Pseudomonas相对丰度显著高于其在有翅蚁及工蚁中的。螺原体Spiroplasma相对丰度在热带火蚁毒腺中显著高于在聚纹双刺猛蚁工蚁毒腺中的。毒腺中细菌多样性分析发现,芽孢杆菌属Bacillus和乳酸杆菌属Lactobacillus在广西红火蚁工蚁中的相对丰度显著高于在广州红火蚁工蚁中的。然而,乳酸杆菌属细菌在聚纹双刺猛蚁工蚁毒腺中的相对丰度显著高于广西的热带火蚁工蚁毒腺中的。【结论】毒腺细菌组成和多样性在3种不同种类蚂蚁工蚁和红火蚁不同品级中存在差异。     

Alternative genetic foundations for a key social polymorphism in fire ants

Little is known about the genetic foundations of colony social organization. One rare example in which a single major gene is implicated in the expression of alternative social organizations involves the presumed odorant-binding protein gene Gp-9 in fire ants. Specific amino acid substitutions in this gene invariably are associated with the expression of monogyny (single queen per colony) or polygyny (multiple queens per colony) in fire ant species of the Solenopsis richteri clade. These substitutions are hypothesized to alter the abilities of workers to recognize queens and thereby regulate their numbers in a colony. We examined whether these same substitutions underlie the monogyny/polygyny social polymorphism in the distantly related fire ant S. geminata. We found that Gp-9 coding region sequences are identical in the polygyne and monogyne forms of this species, disproving our hypothesis that one or a few specific amino acid replacements in the protein are necessary to induce transitions in social organization in fire ants. On the other hand, polygyne S. geminata differs genetically from the monogyne form in ways not mirrored in the two forms of S. invicta, a well-studied member of the S. richteri clade, supporting the conclusion that polygyny did not evolve via analogous routes in the two lineages. Specifically, polygyne S. geminata has lower genetic diversity and different gene frequencies than the monogyne form, suggesting that the polygyne form originated via a founder event from a local monogyne population. These comparative data suggest an alternative route to polygyny in S. geminata in which loss of allelic variation at genes encoding recognition cues has led to a breakdown in discrimination abilities and the consequent acceptance of multiple queens in colonies.     

Examination of host genome for the presence of integrated fragments of Solenopsis invicta virus 1

A series of oligonucleotide primer pairs covering the entire genome of Solenopsis invicta virus 1 (SINV-1) were used to probe the genome of its host, S. invicta, for integrated fragments of the viral genome. All of the oligonucleotide primer sets yielded amplicons of anticipated size from cDNA created from an RNA template from SINV-1. However, no corresponding amplification was observed when genomic DNA (from 32 colonies of S. invicta) was used as template for the PCR amplifications. Host DNA integrity was verified by amplification of an ant-specific gene, SiGSTS1. The representation of fire ant colonies included both social forms, monogyne and polygyne, and those infected and uninfected with SINV-1. Furthermore, no amplification was observed from genomic DNA from ant samples collected from Argentina or the US. Thus, it appears that SINV-1 genome integration, or a portion therein, has not likely occurred within the S. invicta host genome.     

Infection characteristics of Solenopsis invicta virus 2 in the red imported fire ant, Solenopsis invicta

Solenopsis invicta virus 2 (SINV-2) is the second virus identified from the fire ant, S. invicta Buren. SINV-2 is unique among positive-strand RNA viruses from insects by possessing four cistrons in a monopartite genome. Fire ant colonies testing positive for SINV-2 by RT-PCR did not exhibit any discernable symptoms. RT-PCR-based surveys for SINV-2 among 688 fire ant mounds in Alachua County, Florida, sampled during the period January, 2006 through December, 2007 showed that the prevalence of SINV-2 among nests ranged from 1.6% to 16.4%. Unlike S. invicta virus 1, no seasonal-associated prevalence was observed with regard to SINV-2 infection among fire ant colonies. No social form specificity was evident; SINV-2 was found in both monogyne and polygyne S. invicta ants. Real-time quantitative PCR experiments showed that SINV-2 genome equivalents per individual ant ranged from 1.9 × 10⁷ in. pupae to 4.3 × 10¹¹ in. inseminated queens. The SINV-2 infection was detected in all ant stages examined (eggs, larvae, pupae, workers, and queens). Tissue tropism studies indicated that the alimentary canal (specifically the midgut) is most likely the susceptible tissue. SINV-2 was successfully transmitted to uninfected S. invicta ants by feeding a partially purified homogenate of SINV-2-infected ants. The SINV-2 transmission rate ranged from 30% to 80%, and both positive (genomic) and negative (replicative) SINV-2 RNA strands accumulated in recipient ants over the course of the experiment. These results indicated that SINV-2 replicates within S. invicta.     

中国和美国红火蚁毒腺生物碱组分的比较分析

【目的】比较两个入侵地红火蚁Solenopsis invicta毒腺生物碱成分及相对含量的地区间差异。【方法】以在中国广东省惠州、 广州、 深圳、 东莞、 珠海和美国阿拉巴马州采集的红火蚁工蚁为材料, 用正己烷溶剂浸提48 h获得浸提液, 将浸提液进行硅胶柱层析分离, 用气相色谱-质谱联用仪（GC-MS）分析其所含的生物碱成分及其相对含量。【结果】红火蚁毒腺生物碱含顺式生物碱组分和反式生物碱组分。通过总离子图对比, 得到了7种顺式生物碱、 8种四氢吡啶和7种反式生物碱, 并计算得到这些组分的相对百分含量。【结论】中国广东省5个地区红火蚁生物碱组成无差别, 相对含量略有差异, 惠州种群和深圳种群差别最大, 其他地区间没有差别。中美两国入侵地的红火蚁种群生物碱成分种类无差别, 其各个组分之间相对含量略有差异, 美国阿拉巴马州种群与中国深圳种群各组分相对含量差别最大, 与中国广州种群差别最小。另外, 在检测的中国广东样品中, 只确认了红火蚁的存在, 提示杂合蚁与黑火蚁还未传入中国。     

Prevalence of Thelohania solenopsae infected Solenopsis invicta newly mated queens within areas of differing social form distributions

Newly mated queens (NMQs) originating from monogyne red imported fire ant (Solenopsis invicta) colonies and following a mating flight, initiate new colonies by sealing themselves in a nuptial chamber and relying solely on their own fat and crop reserves, as well as no longer needed wing muscles to rear their first workers (claustral colony foundation). This method of colony-founding is rarely successful for polygyne-derived NMQs, whose low weight critically limits the number of first workers they are able to produce. However, this observation may be confounded by the parasitic microsporidium, Thelohania solenopsae, thus far found to persist only in association with polygyne colonies. Infections of this microsporidium reduce the weight of female alates and may explain why polygyne NMQs are unlikely to successfully found colonies claustrally. NMQs collected following mating flights in Gainesville and Ocala, Florida were sorted by weight, checked for insemination and T. solenopsae infection. Insemination levels were greater than 90% for all weight classes at both collection sites and were not related to infection. Infection levels were lower in Gainesville than Ocala, averaging 1.67% and 14.14%, respectively. Polygyne-derived NMQs collected in Ocala, defined here as weighing 12mg (social form correctly assigned in 85% of samples examined by PCR), had the highest infection levels, 25.37% (17/67) in 2003 and 21.43% (6/28) in 2004. We conclude that infection by T. solenopsae cannot be completely responsible for the inability of polygyne NMQs to claustrally establish colonies.     

Multiple virus infections occur in individual polygyne and monogyne Solenopsis invicta ants

Concurrent infections of Solenopsis invicta colonies with S. invicta virus 1 (SINV-1), SINV-2, and SINV-3 has been reported. However, whether individual ants were capable of supporting multiple virus infections simultaneously was not known, nor whether the social form of the colony (polygyne or monogyne) had an influence on the occurrence of multiple infection rates in individual ants. S. invicta field populations were sampled sequentially to establish whether multiple virus infections co-occurred in individual worker ants. In addition, the intra-colony virus infection rates were compared in monogyne and polygyne field colonies to determine whether social form played a role in the viral infection prevalence. All combinations of virus infection (SINV-1, SINV-2, or SINV-3 alone, SINV-1 & SINV-2, SINV-1 & SINV-3, SINV-2 & SINV-3, and SINV-1, SINV-2 & SINV-3) were detected in individual worker ants as well as queens in the field. Thus, individual S. invicta ants can be infected simultaneously with all combinations of the S. invicta viruses. Colony social form did have an influence on the intra-colony prevalence of multiple S. invicta virus infections. Polygyne colonies exhibited significantly greater intra- and inter-colony single and multiple virus infections compared with monogyne colonies.     

红火蚁与热带火蚁间的干扰竞争

【目的】为了探讨入侵火蚁在我国成功定殖及其之间的竞争机制。【方法】运用行为学方法研究红火蚁Solenopsis invicta(Buren)和热带火蚁Solenopsis geminata(Fabricius)在个体水平和群体水平上的攻击性、攻击手段及合作能力。【结果】一对一攻击试验中,红火蚁和热带火蚁之间攻击级别多集中在3级,两种入侵蚂蚁间以相互威胁为主;红火蚁大型工蚁与热带火蚁兵、工蚁间最为好斗,其攻击级别达到4级的比例最高,分别为33.04%、37.92%。热带火蚁兵蚁与各型红火蚁间攻击强度差异不显著;热带火蚁工蚁与红火蚁小型工蚁之间的攻击性最强,其攻击性(3.49)显著高于热带火蚁工蚁与红火蚁大、中型工蚁的攻击性(3.32和2.97)。在攻击手段上,3级打斗时各型红火蚁更倾向以物理攻击主动威胁热带火蚁,而热带火蚁兵、工蚁会采取多种方式主动攻击红火蚁,双方皆以躲避应对为主;4级打斗时两种火蚁主要以混合攻击为主动或应对手段。群体攻击试验显示,红火蚁群体间攻击强度和合作性会随着群体数量的增加而显著增加,热带火蚁合作性较差,其群体对抗红火蚁的优势仅仅是由于个体数量的增加。【结论】红火蚁比热带火蚁具有更强的竞争优势。研究结果为入侵蚂蚁间不对称竞争机制和长期群落替代的内在原因提供理论基础。     

Predaceous ants, beach replenishment, and nest placement by sea turtles

Ants known for attacking and killing hatchling birds and reptiles include the red imported fire ant (Solenopsis invicta Buren), tropical fire ant [Solenopsis geminata (Fabr.)], and little fire ant [Wasmannia auropunctata (Roger)]. We tested whether sea turtle nest placement influenced exposure to predaceous ants. In 2000 and 2001, we surveyed ants along a Florida beach where green turtles (Chelonia mydas L.), leatherbacks (Dermochelys coriacea Vandelli), and loggerheads (Caretta caretta L.) nest. Part of the beach was artificially replenished between our two surveys. As a result, mean beach width experienced by nesting turtles differed greatly between the two nesting seasons. We surveyed 1,548 sea turtle nests (2000: 909 nests; 2001: 639 nests) and found 22 ant species. S. invicta was by far the most common species (on 431 nests); S. geminata and W. auropunctata were uncommon (on 3 and 16 nests, respectively). In 2000, 62.5% of nests had ants present (35.9% with S. invicta), but in 2001, only 30.5% of the nests had ants present (16.4% with S. invicta). Turtle nests closer to dune vegetation had significantly greater exposure to ants. Differences in ant presence on turtle nests between years and among turtle species were closely related to differences in nest placement relative to dune vegetation. Beach replenishment significantly lowered exposure of nests to ants because on the wider beaches turtles nested farther from the dune vegetation. Selective pressures on nesting sea turtles are altered both by the presence of predaceous ants and the practice of beach replenishment.