The first comprehensive molecular detection of six honey bee viruses in Iran in 2015-2016.

At least 18 viruses have been reported in the honey bee (Apis mellifera L.). However, severe diseases in honey bees are mainly caused by six viruses, and these are the most important in beekeeping. These viruses include: deformed wing virus (DWV), acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV), sacbrood virus (SBV), kashmir bee virus (KBV), and black queen cell virus (BQCV). In this study, we evaluated 89 Iranian honey bee apiaries (during the period 2015-2016) suffering from symptoms of depopulation, sudden collapse, paralysis, or dark coloring, by employing reverse transcription-PCR. Samples were collected from four regions (Mazandaran, Hormozgan, Kurdistan, and Khorasan Razavi) of Iran. Of the 89 apiaries examined, 16 (17.97%), three (3.37%), and three (3.37%) were infected by DWV, ABPV, and CBPV, respectively. The study results for the other viruses (SBV, KBV, and BQCV) were negative. The present study evaluated the presence of the six most important honey bee viruses in bee colonies with suspected infections, and identified remarkable differences in the distribution patterns of the viruses in different geographic regions of Iran.

Characterization and Tissue Tropism of Newly Identified Iflavirus and Negeviruses in Glossina morsitans morsitans Tsetse Flies.

Tsetse flies cause major health and economic problems as they transmit trypanosomes causing sleeping sickness in humans (Human African Trypanosomosis, HAT) and nagana in animals (African Animal Trypanosomosis, AAT). A solution to control the spread of these flies and their associated diseases is the implementation of the Sterile Insect Technique (SIT). For successful application of SIT, it is important to establish and maintain healthy insect colonies and produce flies with competitive fitness. However, mass production of tsetse is threatened by covert virus infections, such as the Glossina pallidipes salivary gland hypertrophy virus (GpSGHV). This virus infection can switch from a covert asymptomatic to an overt symptomatic state and cause the collapse of an entire fly colony. Although the effects of GpSGHV infections can be mitigated, the presence of other covert viruses threaten tsetse mass production. Here we demonstrated the presence of two single-stranded RNA viruses isolated from Glossina morsitans morsitans originating from a colony at the Seibersdorf rearing facility. The genome organization and the phylogenetic analysis based on the RNA-dependent RNA polymerase (RdRp) revealed that the two viruses belong to the genera Iflavirus and Negevirus, respectively. The names proposed for the two viruses are Glossina morsitans morsitans iflavirus (GmmIV) and Glossina morsitans morsitans negevirus (GmmNegeV). The GmmIV genome is 9685 nucleotides long with a poly(A) tail and encodes a single polyprotein processed into structural and non-structural viral proteins. The GmmNegeV genome consists of 8140 nucleotides and contains two major overlapping open reading frames (ORF1 and ORF2). ORF1 encodes the largest protein which includes a methyltransferase domain, a ribosomal RNA methyltransferase domain, a helicase domain and a RdRp domain. In this study, a selective RT-qPCR assay to detect the presence of the negative RNA strand for both GmmIV and GmmNegeV viruses proved that both viruses replicate in G. m. morsitans. We analyzed the tissue tropism of these viruses in G. m. morsitans by RNA-FISH to decipher their mode of transmission. Our results demonstrate that both viruses can be found not only in the host's brain and fat bodies but also in their reproductive organs, and in milk and salivary glands. These findings suggest a potential horizontal viral transmission during feeding and/or a vertically viral transmission from parent to offspring. Although the impact of GmmIV and GmmNegeV in tsetse rearing facilities is still unknown, none of the currently infected tsetse species show any signs of disease from these viruses.

Temperature-dependent development and temperature thresholds of codling moth (Lepidoptera: Tortricidae) in Iran.

Developmental rate models and biological parameters estimated from them, especially lower and upper temperature thresholds and optimal temperature, can help to forecast phenological events of codling moth, Cydia pomonella L. (Lepidoptera: Tortricidae), in apple orchards. We studied the developmental time of immature stages of codling moth at eight constant temperatures ranging from 10 to 35 degrees C and modeled their developmental rate as a function of temperature using 13 published nonlinear and 2 linear models. Data were fitted to developmental rate models and temperature thresholds and the optimal temperatures were estimated. The models were evaluated based on adjusted coefficient of determination (R(2)(adj)) and Akaike information criterion (AIC), in addition to coefficient of determination (R(2)) and residual sum of squares (RSS). The thermal constants were 79.80, 312.60, 232.03, and 615.32 DD for egg, larva, pupa, and overall immature stages of codling moth, respectively, using the Ikemoto and Takai linear model. The Ikemoto and Takai linear model estimated lower temperature thresholds as 9.97, 8.94, 10.04, and 9.63 degrees C for egg, larva, pupa, and overall immature stages, respectively. Among the nonlinear models, the third-order polynomial fit the data well. This model estimates optimal temperature accurately. Brière-1 and Brière-2 accurately estimated the lower and upper temperature thresholds considering model evaluation criteria and accuracy of estimations.

Molecular identification and phylogenetic analysis of chronic bee paralysis virus in Iran.

Chronic bee paralysis virus (CBPV) is an unclassified polymorphic single-stranded RNA virus. Among the viruses infecting honeybees, CBPV is known to induce significant losses in honeybee colonies. In this study, a total number of eighty-nine suspected apiaries from four regions of Iran (including Mazandaran, Khorasan Razavi, Hormozgan, and Kurdistan) were sampled and submitted for molecular identification. Three positive samples were detected by RT-PCR. All positive samples were confirmed by sequencing. The phylogenetic tree which displays the molecular relationship between the viruses of different Iranian geographic regions and references isolates was constructed. The Iranian isolates formed two distinct phylogenetic groups (Group 1 and Group 2). The IR-CPV-GMG-1, IR-CPV-GMG-2, IR-CPV-GMG-4, and IR-CPV-GMG-6 formed Group 1 and IR-CPV-GMG-3, IR-CPV-GMG-5, and IR-CPV-GMG-7 were in Group 2 as a distinct group. Iranian isolates in group 1 were similar to European and East Asian CBPVs. This research was the first phylogenetic analysis of CBPV in Iran. Further researches are needed to study the other aspects of this virus-like genetic characteristics and pathogenesis in Iran.