Background Hydrosalpinx is considered one of the obstacles that could hinder the success of IVF techniques due to the toxic effect of the hydrosalpinx fluid pouring into the uterine cavity. Tubal disconnection by either hysteroscopic or laparoscopic approaches is considered the standard in the operative management of hydrosalpinx prior to IVF cycles. The aim of the current study was to compare the success rate of hysteroscopic tubal occlusion using electro-coagulation with laparoscopic tubal disconnection in cases of hydrosalpinx prior to in vitro fertilization (IVF). Methods A total of 108 women with unilateral or bilateral hydrosalpinx, who were candidates for tubal occlusion before IVF, were equally randomized into two groups. Group A ( n = 54) underwent hysteroscopic tubal occlusion using the roller-ball electro-coagulation of the interstitial part of the tube and the uterine cornual area, and group B ( n = 54) underwent laparoscopic tubal disconnection using bipolar coagulation and a proximal tubal cut. Operative time, complications, postoperative pain measured by the visual analogue score (VAS score), and postoperative hospital stay were recorded for both groups. The success rate of tubal occlusion was assessed 1 month later using a post-menstrual hysterosalpingogram (HSG). Results Laparoscopic tubal disconnection was more successful than the hysteroscopic approach regarding tubal occlusion rate (96.15% vs. 86.67% respectively, p = 0.044). The operative time and postoperative pain VAS scores in the hysteroscopy group (3.65 ± 1.03 min and 1.81 ± 1.35, respectively) were significantly lower than that in the laparoscopy group (17.48 ± 4.70 min and 4.06 ± 1.65, respectively) with p < 0.001. Conclusion Although laparoscopic tubal disconnection is more successful, the hysteroscopic approach is an alternative which has its own limitations that can be assessed by hysterosalpingogram, especially when laparoscopy is contraindicated, technically difficult, or refused by the patient. Trial registration It was first registered at ClinicalTrials.gov on 30/07/2019 with registration number NCT04037813 .

Abstract A severe isolate of Prunus necrotic ringspot virus (PNRSV) was isolated from apple orchards in the vicinity of Nubaria city, Beheira governorate, Egypt. Infected-apple trees showed chlorotic, necrotic ringspots, and shoot holes on leaves. Severely infected- trees withered, became useless, and were removed causing severe economic losses. Reverse transcriptase (RT) polymerase chain reaction (PCR), RT-PCR, using degenerate primer pair for the coat protein (CP) gene of Ilarvirus amplified products similar to those produced from peach and apricot isolates of PNRSV-infecting stone fruits). Dot blotting immuno-binding assay (DBIA showed a positive reaction between PNRSV-infected apple sap and an Egyptian antiserum for PNRSV. Purified preparation from infected leaves, using the electro-elution technique yielded nucleoprotein, which had Amax and Amin at 260 and 240 nm respectively. Electron microscopy examination showed spherical virions with ca. 26 nm in diameter.

Cotton leaf curl Gezira virus (CLCuGeV), in association with a DNA-beta satellite, is reported as a causal agent of cotton leaf curl disease. This study reports CLCuGeV detection in field samples of Capsicum annuum and Cucumis melo from Egypt and represents an expansion of the virus host range beyond the Malvaceae family to the two plant families, Solanaceae and Cucurbitaceae .

BACKGROUND AND OBJECTIVETomato Chlorosis Virus (ToCV) is a white fly-transmitted and phloem-limited crinivirus reported in this study for the first time in Egypt. ToCV caused drastic reduction in tomato yield since 2013. The aim of this study is to characterize the virus incidence using biological, serological and molecular tools.MATERIAL AND METHODSThe B. tabaci MEAM1 white fly was used for virus isolation and propagation. Identity of ToCV , its natural hosts were confirmed with RT-PCR using a specific primer pair for ToCV-heat shock protein 70 homologue (HSP70h) gene, sequencing and phylogenetic studies. ToCV was purified using the innovative electro-elution technique. The induced antiserum for the Egyptian isolate of the virus (ToCV-Giza) was used for DAS-ELISA and dot blotting immuno-assays to evaluate the virus presence in tomato and other natural hosts.RESULTSThe ToCV-Giza isolate was donated an accession number "MH667315.1" from the GenBank. Blastx sequence analysis of the HSP70h gene indicated 97-99% of amino acid similarities with many tested ToCV isolates. Phylogenetic studies showed the clustering of all ToCV isolates including ToCV-Giza in a separate group from the other tested criniviruses. The virus had a UV spectrum of a nucleoprotein with Amax and Amin at 260 and 240 nm, respectively and A260/280 ratio of 1.33. Out of 52 different tested plant species within 22 families, 44 were positive hosts for ToCV. Thirty seven out of these 44 plant species were considered as new hosts for ToCV in the present study. These included Ammi majus and Coriandrum sativum (Apiaceae), cabbage (Brassicaceae), sweet potato (Convolvulaceae), melon, cucumber, luffa (Cucurbitaceae), soybean, cowpea, faba bean (Fabaceae), Egyptian and American Cotton (Malvaceae). Several ornamentals either herbal type or woody trees belonging to Acanthaceae, Amaranthaceae, Euophorbiaceae, Moraceae and Rubiaceae were also recognized for the first time as hosts for ToCV.CONCLUSIONThe obtained results confirmed the wide distribution of ToCV in its natural hosts in Egypt. Hygienic measures including control of the virus vector and removing of natural hosts should be strictly implicated.

Background and purpose The functional outcomes of incontinence and high stool frequency resulting from restorative surgery are often criticised. The aim of this study was to assess the taeniectomy pouch in comparison with other pouches described in the literature. Material and methods This was a prospective cohort study. All patients who were candidate for low rectal resection presenting to the colorectal unit at Cairo University hospitals during the period February 2013 to February 2015 were included in the study (90 patients). Safety and feasibility of the new technique were assessed, including operative time, leakage, postoperative urgency, incontinence, number of daily motions and difficulty in evacuation. These parameters were assessed clinically, by means of defecography and anorectal manometry. Results The mean age of patients was 49.6 years. Percentages of postoperative mortality and leakage were 2.2% and 3.4%, respectively. Mean operative time was 117 minutes. Mean numbers of daily motions were 3.04 and 1.52 at 3 and 12 months, respectively. Mean Wexner score for continence at 3 and 12 months were 3.21 and 1.32, respectively. Mean resting pressure was 51.63 mmHg, squeeze pressure was 130.42 mmHg and mean threshold volume was 118.68 ml. Conclusions Taeniectomy is a novel technique for pouch formation after low rectal resection, which can be used as an alternative to other pouches, especially the widely used transverse coloplasty.

Polyclonal antibodies to recombinant (r) coat proteins (r CP) of Potato virus X (PVX) and Potato leaf roll virus (PLRV) were developed. The effectiveness of these antisera was determined by indirect ELISA (I-ELISA), immuno blotting, and double antibody sandwich (DAS) enzyme linked immunosorbent assay (ELISA), i.e. DAS-ELISA. The measured titers, with indirect-ELISA, expressed as reciprocal of the dilution end-points were 6000 for PLRV; whereas, PVX had a titer of 5000. The CP genes of PVX, and PLRV were amplified by reverse transcription-polymerase chain reaction (RT-PCR), cloned and expressed into pBAD-C terminal 6xHis Tag TOPO expression vector in Escherichia coli. For immunization, the CP fractions from bacterial lysates of each virus were purified, under native and denatured conditions, by nickel-nitrilotriacetic acid (Ni-NTA) batch chromatography and equally mixed. The fused proteins were 6.6 mg/20 ml for PVX and 9.0 mg/20 ml for PLRV bacterial culture. Antigenicity of the purified CP fraction was measured with Western blotting (WB) analysis. The r CP of each of PVX, and PLRV purified under naturing and denaturing conditions were injected into rabbits for antibody productions. The PVX- and PLRV-induced antisera reacted in I-ELISA, indirect plate trapped antigen enzyme-linked immunosorbent assay (IPTA- ELISA), and dot blotting immuno binding assay (DBIA). The data indicated that the produced r antisera were efficient and accurate in differentiation between negative and positive samples in DAS- ELISA upon testing leaves and sprouts of several potato varieties tests. Therefore, these r antisera are suitable for the certification program of potatoes due to their low cost, high specificity, feasibility, and their endless supply from r bacterial clones carrying the CP genes for these viruses.

The presence of Dahlia mosaic virus D10 (DMV-D10) was confirmed for the first time in dahlia (Dahlia variabilis) in Egypt. DMV-D10 was recently described as a caulimovirus that exists as an endogenous pararetroviral sequence (EPRS). DMV-D10 was confirmed by amplification of the ORF1 (encoding for the movement protein) using species specific primers (D10F1/R1). The expected size (900 bp) was amplified from 36 samples with no evidence of infection with either DMV or DCMV. The same dahlia plants were tested for the presence of CMV, INSV, TSV, and TSWV and they were all negative. Sequence comparisons of the Egyptian DMV-D10 ORF1, GenBank accession HM007162, amplified from these samples revealed that the amplicon had the highest sequence identity (96%) with that of DMV-D10 (isolated from US dahlia cultivars). Cluster dendogram based on the amino acid sequences of movement protein of all known caulimoviruses placed D10-US (isolated from US dahlia cultivars), D10-NZ (isolated from New Zealand dahlia cultivars), D10-DC (isolated from D. coccinea) and D10-Egypt (isolated from Egyptian dahlia cultivars) in one clade.

Two isolates of PNRSV were isolated from peach and apricot trees at the Experimental Station of the Faculty of Agriculture, Cairo Univer sity, Giza, Egypt. The peach isolate of PNRSV (PNRSV-PF) was differentiated from the apricot isol ate (PNRSV-AP) by ten differential host species. Both isolates were purified successfully w ith the electro-elution technique. Both isolates had A max and A min at 260 and 240 nm respectively. The A260/280 ratios were 1.55 and 1.6 0 for the AP and the PF isolates, respectively. Electron mic roscopy examination showed spherical virions with ca 27-29 nm in diameter. Both isolates had mo lecular weight of coat protein subunits of 29 kDa, determined by polyacrylamide gel electrophores is (SDS-PAGE). Antisera raised against the two isolates detected their counter antigens in pea ch and apricot trees. Both antisera cross reacted with their homologous and heterologous antigens in dot blot immunoassay (DBIA) and agar-double diffusion (ADD) tests. The two virus isolates appea red to belong to the same sero-group and represent two different pathotypes. PNRSV-AP varie d in sero-grouping of that of beet necrotic ringspot ilarvirus, a tentative isolate of PNRSV, w hen examined in ADD test. Indirect ELISA showed that 64.2 % of 210 tested apricot trees were infected, while 150-inspected peach trees showed 29.5 % infection. Four sets of primers were used to amplify both movement protein (MP) and coat protein (CP) genes of the two Egyptian iso lates of PNRSV isolated from apricot and peach trees. Amplicons of the correct size (~ 894 bp) fo r the MP gene and (~ 704 bp) for the CP gene were obtained from the two examined isolates of PNR SV. Nested PCR using specific primers for both the MP and the CP genes confirmed the authenti city of the PCR amplified products. RT-PCR detected successfully the presence of PNRSV in the pollen grains of infected apricot and peach trees. Nucleotide sequences of the MP genes of the two isolates were revised by the GenBank and given the accession # EU100388 for the peach isolate and EU106649 for apr icot isolate. Phylogenetic analysis of RNA 3-MP showed ca 65 % si milarity between PNRSV-AP and PNRSV- PF; indicating that the two isolates of PNRSV are d istantly related. The relatedness between the two isolates and other PNRSV isolates is discussed.

Squash plants showing leaf curling, yellow mottling, and reduced fruit set were observed in fields in Giza, Egypt in spring 2005. These particular symptoms had not been observed previously in zucchini squash plants in Egypt, but were reminiscent of those caused by begomoviruses (Geminiviridae) that are known to occur in the region, including Watermelon chlorotic stunt virus. Squash plants were heavily infested with the whitefly Bemisia tabaci (Genn.), the only known vector of begomoviruses. Total nucleic acids were isolated from symptomatic squash leaves using the cetyltrimethylammoniumbromide method, and extracts were subjected to polymerase chain reaction (PCR) analysis using two sets of PCR primers. One primer set (prAV2644 and prAC1154) was designed to amplify a fragment that contains the entire viral coat protein (Cp), while the second primer set (prBV1855 and prBC656) was designed to amplify the common region (CR) of DNA-B of begomoviruses (1). The expected size fragments were cloned and the sequence was determined for five clones each. Unexpectedly, the Cp and the CR-B fragments shared their highest nucleotide sequence (nt) identity among well-characterized begomoviruses to the bipartite Squash leaf curl virus (SLCV) native to the western United States. A third primer set (prAC344 and prAV1134) (1) was subsequently used to amplify the remainder of the putative SLCV DNA-A. The fragment was cloned and the DNA sequence was determined. Assembly of the overlapping DNA-A fragments resulted in a complete DNA-A component sequence of 2,636 nt, which is identical to the expected size of the SLCV DNA-A component (GenBank Accession No. DQ285019). Comparison with the latter sequence indicated that the Egyptian squash isolate shared 98% nt identity with SLCV. The sequence for the DNA-B fragment (1,162 nt) shared 94% nt identity with SLCV and was deposited in GenBank as Accession No. DQ285020. The high-shared nt identity with SLCV (2) from the United States suggests that this isolate, herein SLCV-EG, has been introduced into Egypt. The relatively low DNA-B nt sequence identity was a not a surprise since this component is normally less conserved even between strains of a single begomoviral species. Introduction of SLCV is not only potentially significant to the domestic production of crop species in the Cucurbitaceae but also for legume crops. SLCV has a broad host range that also includes members of the Fabaceae, which includes species that contribute significant sources of protein for much of Egypt's population. The virus thus far is thought to be present only in Lower Egypt, however, it could feasibly threaten legume and cucurbit crops if it spreads to Upper Egypt. To our knowledge, this is the first begomovirus of New World origin to become established in the Old World. References: (1) A. M. Idris and J. K. Brown. Phytopathology 88:648, 1998. (2) S. G. Lazarowitz. Virology 180:70, 1991.

Post-transcriptional gene silencing (PTGS) is a nucleotide sequence specific RNA turnover mechanism produced by plant cell as a natural mechanism against virus infection. Double strands of short (21-25 nt) interference RNA (siRNA) mediate this natural defense mechanism. The PTGS mechanism is becoming powerful tools for reducing expression of a certain viral gene and controlling infection. In the purpose of controlling the tomato yellow leaf curl virus (TYLCV), three regions from the viral genome located in the overlap regions between C1/C2, C2/C3 and V1/V2 were used for studying their efficiency in working as siRNA to target viral DNA accumulation in plant cells. Constructs were prepared by cloning each of the three regions both in the sense or sense/antisense directions in the binary vector pFGC5941 and introduced into tomato and tobacco plant tissues via Agro-infiltration technique. Additional construct containing the three regions together in the sense/antisense directions was also used as a control to prove that either one of the selected regions may activate the siRNA mechanism. All plants were challenged with virus using the TYLCV-Eg infectious clone 10 days post infiltration. The obtained results revealed that only sense/antisense constructs inhibit or reduce accumulation of viral genome due to the induction of siRNA mechanism. However, infiltered plants with the construct containing the as C1/C2 region proved to be the best region for initiating PTGS as it inhibited the accumulation of the viral genome completely as well as it prevented the appearance of viral symptoms in all tested plants. This may be due to its role in silencing the expression of two very important viral genes which express the replication associated protein (Rep) and transactivator protein for the viral sense promoter. However, the other two regions showed a high, but not a complete, level of interference for genome accumulation and viral infection symptoms. The obtained results may also be explained based on the length of the cloned fragments, as it was only 1١٦ bp for C1/C2 region compared to the other regions (280 bp for C2/C3 and 260 bp for V1/V2) and / or the type of the genes they interfere. It can be concluded that this mechanism can result in higher viral resistance in tomato against TYLCV

Banana bunchy top virus is a multicomponent virus, comprising of at least six integral components, with a circular single-stranded DNA genome. In this study, specific rabbit polyclonal antibodies against bacterially expressed coat protein of Banana Bunchy Top virus (BBTV, genus Nanovirus) were produced using a recombinant DNA approach. The BBTV capsid protein (CP) gene located on component 3 was cloned in an expression vector pQE-30 (Qiagen). Expression of the CP with an N-terminal hexahistidine tag in Escherichia coli M15 cells was induced by adding isopropyl-3-D-1-thiogalactoside (IPTG) to a final concentration of 1 mM. About 13 mg of bacterially expressed CP was purified from 1 litre of bacterial liquid culture using a Ni-NTA resin column (Qiagen). The expressed CP which migrated as a protein of approximately 21 kDa in sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis (PAGE) was identified by its strong reaction with polyclonal antibodies produced against BBTV purified particles (BBTV-AS) and Ni-NTA-AP conjugate in Western blots. Expressed and purified CP (SDS-PAGE 21 kDa band) was injected into a white rabbit, using two subcutaneous and three intramuscular injections at weekly intervals. The antiserum produced was evaluated for BBTV and FBNYV detection in Western blot and dot blot immunoassays (DBIA). The antiserum raised against the expressed CP (BBTV-AS6xHis) gave strong BBTV-specific DBIA reactions and very weak background reactions with non-infected tissues, similar to those produced by polyclonal antibodies raised against BBTV purified virion (BBTV-AS). Furthermore, (BBTV-AS6xHis) polyclonal antibody reacted specifically with both the denatured recombinant protein and the disrupted BBTV virus particle as well as FBNYV purified particles in DBIA. These results showed that the (BBTV-AS6xHis) polyclonal antibody is useful for the detection of BBTV in infected tissues by dot blot tests.

Two isolates of faba bean necrotic yellows nanovirus (FBNYV) and banana bunchy top babuvirus (BBTV) were purified. Cellulase added to the extraction buffers (EB) enhanced extraction of both viruses. An acidic pH was used for FBNYV extraction. Several trials for purification of BBTV indicated that alkaline condition surpassed acidic condition in virus extraction. Polyethylene glycol and NaCl were used to concentrate both viruses. Electroelution coupled with freezing and thawing were employed in further purification steps. Purified FBNYV yielded 677 µg/ g tissue and with A 260/280 ratio of 1.43. Purified BBTV yielded 12.9 µg /g tissue and with A 260/280 ratio of 1.3. Both viruses had a single-capsid protein of 20 kDa. An antiserum raised for the intact virions of FBNYV detected the purified virus and was able to cross react with BBTV in dot blotting immunobinding assay (DBIA) and in western blotting (WB). Antiserum for the whole virions of BBTV detected the virus in its vector Pentalonia nigronervosa as well as in infected banana tissues. This antiserum also cross-reacted with FBNYV in DBIA and in WB but failed to react with its own antigen of BBTV. On the otherhand, an antiserum raised for the coat protein of BBTV was able to detect both FBNYV and BBTV in WB. Such results suggest the presence of continuous epitopes in FBNYV virions and discontinuous epitopes in BBTV virions. Polymerase chain reaction (PCR) experiments indicated that primers for the coat protein and replicase genes of BBTV amplified products in FBNYV genome similar to those produced in BBTV genome. Similar results were obtained upon using immunocapture (IC) PCR (IC-PCR) technique; indicating strong serologic relationships between the two viruses. The use of specific BBTV primers for nanovirus detection is of a great value in sensitivity and suitable for large scale testing.

DNA 1 components are satellite-like, single-stranded DNA molecules associated with begomoviruses (family Geminiviridae) that require the satellite molecule DNA beta to induce authentic disease symptoms in some hosts. They have been shown to be present in the begomovirus-DNA beta complexes causing cotton leaf curl disease (CLCuD) and okra leaf curl disease (OLCD) in Pakistan as well as Ageratum yellow vein disease (AYVD) in Singapore. We have cloned and sequenced a further 17 DNA 1 molecules from a diverse range of plant species and geographical origins. The analysis shows that DNA 1 components are associated with the majority of begomovirus-DNA beta complexes, being absent from only two of the complexes examined, both of which have their origins in Far East Asia. The sequences showed a high level of conservation as well as a common organization consisting of a single open reading frame (ORF) in the virion sense, a region of sequence rich in adenine and a predicted hairpin structure. In phylogenetic analyses, there was some evidence of grouping of DNA 1 molecules according to geographic origin, but less evidence for grouping according to host plant origin. The possible origin and function of DNA 1 components are discussed in light of these findings.

DNA β molecules are symptom-modulating, single-stranded DNA satellites associated with monopartite begomoviruses (family Geminiviridae). Such molecules have thus far been shown to be associated with Ageratum yellow vein virus from Singapore and Cotton leaf curl Multan virus from Pakistan. Here, 26 additional DNA β molecules, associated with diverse plant species obtained from different geographical locations, were cloned and sequenced. These molecules were shown to be widespread in the Old World, where monopartite begomoviruses are known to occur. Analysis of the sequences revealed a highly conserved organization for DNA β molecules consisting of a single conserved open reading frame, an adenine-rich region, and a region of high sequence conservation [the satellite conserved region (SCR)]. The SCR contains a potential hairpin structure with the loop sequence TAA/GTATTAC; similar to the origins of replication of geminiviruses and nanoviruses. Two major groups of DNA β satellites were resolved by phylogenetic analyses. One group originated from hosts within the Malvaceae and the second from a more diverse group of plants within the Solanaceae and Compositae. Within the two clusters, DNA β molecules showed relatedness based both on host and geographic origin. These findings strongly support coadaptation of DNA β molecules with their respective helper begomoviruses.

An isolate of tobacco ringspot virus (TRSV) was isolated from potato (Solanum tuberosum L.) plants showing calico symptoms and growing in commercial fields in El-Dakahlia govenorate. The isolated virus was designated as TRSV-DAK. The virus was transmitted mechanically to several hosts including Capsicum annuum L., Physalis floridana Thdb., Cucumis sativus L., and some cultivars of Nicotiana tabacum L. Symptoms ranged between chlorosis, ringspot, and necrosis. TRSV-DAK induced local lesions upon its inoculation on Vigna unguiculata Walp. The isolated virus was purified using an electro- elution technique. The purified virus has Amax at 260, Amin, at 245 and A260/280 of 1.34. Virus yield was 0.5 mg/ml. TRSV-DAK reacted scrologically with two authentic antisera for TRSV from the USA and France. A third locally induced antiserum for TRSV- DAK detected its presence in potato leaves, stems and tubers grown in both field and greenhouse upon using different immuno-sorbent techniques. Distribution of the virus was uneven in infected- potato plants. Ten potato cultivars were tested for pathogenecity to TRSV-DAK. Generally, plants infected under field conditions were more susceptible to TRSV-DAK infection when compared to those under greenhouse conditions. Cydon and Gigent potato cvs. were the least infected under greenhouse conditions showing 10 and 20% infection, respectively. The same two cvs. exhibited 60, 70% infection respectively in the field. EIectron microscopy examination of purified virus showed the presence of spherical particles with 27 nm in diameter. In tissue culture experiments, four tested antiviral compounds, namely salicylic acid (SA), benzoic acid (BA), jasmonic acid (JA) and linolcnic acid (LA), exhibited variable abilities to remove TRSV- DAK from infected potato explants. Increased concentrations of all used substances were positively correlated with the percentages of TRSV-free potato plantlets. JA induced the highest percentages of virus-free plantlets (being 96.7% at 0.3 mg/l) compared with other tested chemicals.