Correction to: An outbreak of cutaneous leishmaniasis due to Leishmania major in an endemic focus in central Iran.

[This corrects the article DOI: 10.1007/s12639-022-01474-6.].

An outbreak of cutaneous leishmaniasis due to Leishmania major in an endemic focus in central Iran.

Zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania major is an important endemic disease and a major public health concern in Iran. Following an outbreak of leishmaniasis in 2013 in one of the important foci in Iran, the researchers were prompted to determine the underlying causes of the epidemic. Adult sand flies were collected using sticky traps and aspirating tubes and infection with Leishmania parasites was evaluated. Also, rodents were captured using Sherman live traps and stray dogs were hunted and were tested for the presence of leishmanial infection. Active case detection was also conducted and data related to each household were recorded using a researcher-designed form. Nested PCR and PCR-RLFP techniques were employed to determine Leishmania infection in the samples. Phlebotomus papatasi was the most dominant species among the 7 different species of sand flies collected in this study. Sergentomyia clydei, Sergentomyia theodori, and Sergentomyia grekovi were identified for the first time in the study area. 20% of collected Ph. papatasi species and one Meriones libycus were infected with Leishmania major. Stray dogs demonstrated no lesions on different parts of their body. The relative frequency of active lesions and scars on the dogs were respectively 5.49% and 1.23% in 2013 and 5.82% and 0.56% in 2014. The highest number of ZCL cases due to L. major was recorded in Ardestan city. Phlebotomus papatasi is the main vector and M. libycus seems to be the primary reservoir host of ZCL in the suburbs of this city. Furthermore, due to the low density of rodent reservoir hosts in the study area, we support the hypothesis that humans may have had a role in maintaining the parasite cycle in the population.

A Comparative Study on the Biodiversity and Species Richness of Phlebotomine Sand Flies (Diptera: Psychodidae) in Kermanshah and Khuzestan Provinces of Iran.

Background: Leishmaniasis is one of the most important vector-borne and neglected tropical diseases in many parts of the world. The main objective of this study was to determine the biodiversity of phlebotomine sand flies (Diptera: Psychodidae, Phlebotominae) in Khuzestan and Kermanshah Provinces of Iran. Methods: Sampling was conducted in Khuzestan and Kermanshah Provinces using sticky paper traps and CDC light traps. The samples were then stored in 96% alcohol-containing vials, mounted, and identified. Afterward, the alpha diversity (using Simpson's, Shannon-Weiner's diversity, evenness, Maghalef's, Menhinick's, Hill N 1 and N 2 indices) and beta diversity indices (using Sorensen's and Jaccard's coefficients) were calculated. Results: A total of 4302 sand flies were caught and identified which comprised mostly of Phlebotomus papatasi, Ph. alexandri and Ph. sergenti. It was found that among the four counties of Ahvaz, Shush, Shushtar and Dezful in Khuzestan Province, Shush had the lowest, and Shushtar, the highest amount of species diversity and evenness. In the four studied counties of Kermanshah Province, species diversity in Kermanshah County had the lowest amount and Sarpol-e-Zahab, the highest. The species richness in Kermanshah County was the lowest, while Qasr-e-Shirin County had the highest amount. Conclusion: An overview of the biodiversity of phlebotomine sand flies in Kermanshah County in Kermanshah Province, and Shush County in Khuzestan Province, showed less stability in community structure of these vectors, which can be counted as an alarm for emerging dominant vectors among the studied counties and have the potential to increase the prevalence of leishmaniasis.

Biodiversity and Spatial Distribution of Mosquitoes (Diptera: Culicidae) in Kurdistan Province, Western Iran.

Background: Mosquitoes (Diptera: Culicidae) have always been considered as the vector/s of viral and parasitic diseases. This study aimed to conduct a comprehensive survey on the species composition, spatial distribution, and biodiversity indices of mosquitoes in Kurdistan Province, western Iran. Methods: This study was carried out in 10 counties of Kurdistan Province. The immature stages of mosquitoes were collected monthly from June to September. ArcGIS software was used to spatial analysis and create maps. Alpha diversity indices were calculated using the related formula. Results: Totally, 5831 larvae belonging to the family Culicidae were collected. Twelve species were identified including: Anopheles claviger, An. maculipennis s.l, An. superpictus s.l, Culiseta. longiareolata, Cs. subochrea, Culex hortensis, Cx. mimeticus, Cx. perexiguus, Cx. pipiens, Cx. theileri, Cx. modestus and Cx. territans. Based on this analysis, the high-risk areas of the province are determined as Anopheles in the west, Culex in the north, and the Culiseta in the south of the province. Analyzing the Alpha biodiversity indices showed Baneh and Sarabad had the maximum and Bijar had the minimum mosquito biodiversity. Conclusion: The western counties of the province are regarded as the hotspots for anopheline mosquitos. Moreover, reporting of malaria cases in the past, bordering with Iraq and the high traffic of travelers have made these areas as potential foci for malaria transmission. So that, routine entomological inspections are proposed to detect any suspicious vector or case entrance.

Persistence assessment of SARS-CoV-2-specific IgG antibody in recovered COVID-19 individuals and its association with clinical symptoms and disease severity: A prospective longitudinal cohort study.

BACKGROUND: Antibodies play an important role in neutralizing invading pathogens and protecting the host against re-infection. Thus, the accurate assessment of antibodies during a pandemic can provide important evidence for monitoring pathogen exposure, understanding the role of antibodies in protective immunity, and helping vaccine development. METHODS: In this study, 96 west Iranian recovered COVID-19 subjects were recruited and, based on clinical symptoms and disease severity, categorized into three different groups: mild, moderate, and severe. In addition, the presence and dynamic change of SARS-CoV-2-specific IgG antibody three, four-, and six months post symptom onset (PSO) were measured. Also, the association between IgG antibody titer with clinical symptoms and disease severity was examined. RESULTS: Although in real-time RT-PCR-positive samples negative IgG antibody results were found, most subjects mount humoral immune responses that could raise a robust SARS-CoV-2-specific IgG antibody. Furthermore, this antibody persisted in the serum of most recovered COVID-19 subjects at least six months PSO and demonstrated little to no decrease. Also, specific IgG antibody titer was strongly correlated with clinical symptoms and disease severity. CONCLUSIONS: These results provide an insight into the presence and persistence of the SARS-CoV-2-specific IgG antibody. Although serological tests could not be used as the primary diagnostic test, they may support real-time RT-PCR results. Also, they could be used for diagnosing COVID-19 subjects tested later outside of the optimal period. Thus, the SARS-CoV-2-specific IgG antibody is an excellent marker of COVID-19 infection or vaccination and provides an additional diagnostic tool for verifying results and helps monitor and control COVID-19 spread.

An Eco-Epidemiological Study on Zoonotic Cutaneous Leishmaniasis in Central Iran.

BACKGROUND: Leishmaniasis is an expanding neglected tropical disease in the world reporting from 98 countries including Iran. This study focused on eco-epidemiological determinants of the disease following a rapid and unexpected increase of leishmaniasis incidence in a strategic residential district in North-East of Isfahan County, Iran. METHODS: This study was accomplished from Apr 2012 to Jan 2014 in a strategic residential zone in North-East of Isfahan County, Esfahan, Iran. Monthly activity, parity, Leishmania infection and susceptibility tests, were determined on sand flies. Some portion of inhabitants and school children were surveyed to find active or passive cases of leishmaniasis and also wild rodents were collected to determine reservoir host. RESULTS: Totally 5223 sand flies belonging to Phlebotomus and Sergentomyia genus were collected and identified; Ph. papa-tasi was the dominant species and started to appear in May and disappeared in Oct. The majority of living dissected sand flies were unfed and parous. Ph. papatasi showed 4.6% Leishmania infection through direct examination and 39.54% by nested-PCR respectively. Phlebotomus papatasi was susceptible against deltametrin 0.05%. Totally 2149 people were surveyed and incidence and prevalence of zoonotic cutaneous leishmaniasis estimated as 45.39 and 314.40 per 1000 population. Rodents showed 73.91% and 80% Leishmania infection by direct examination and nested-PCR respectively. CONCLUSION: Cutaneous leishmaniasis due to L. major has been established in this area. Rodent control operation and personal protection are highly recommended to control the disease in this focus.

The leishmanicidal effect of Lucilia sericata larval saliva and hemolymph on in vitro Leishmania tropica.

BACKGROUND: Leishmaniasis is a major parasitic disease worldwide, except in Australia and Antarctica, and it poses a significant public health problem. Due to the absence of safe and effective vaccines and drugs, researchers have begun an extensive search for new drugs. The aim of the current study was to investigate the in vitro leishmanicidal activity of larval saliva and hemolymph of Lucilia sericata on Leishmania tropica. METHODS: The effects of different concentrations of larval products on promastigotes and intracellular amastigotes of L. tropica were investigated using the mouse cell line J774A.1 and peritoneal macrophages as host cells. The 3-(4.5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and direct observation and counting method were used to assess the inhibitory effects and cell cytotoxicity of the larval products. The effects of larval products on the amastigote form of L. tropica were quantitatively estimated by calculating the rate of macrophage infection, number of amastigotes per infected macrophage cell, parasite load and survival index. RESULTS: The 50% cytotoxicity concentration (CC50) value of both larval saliva and hemolymph was 750 µg/ml, and the 50% inhibitory concentration (IC50) values were 134 µg/ml and 60 µg/ml for larval saliva and larval hemolymph, respectively. The IC50 for Glucantime, used a positive control, was (11.65 µg/ml). Statistically significant differences in viability percentages of promastigotes were observed for different doses of both larval saliva and hemolymph when compared with the negative control (p ≤ 0.0001). Microscopic evaluation of the amastigote forms revealed that treatment with 150 µg/ml larval hemolymph and 450 µg/ml larval saliva significantly decreased the rate of macrophage infection and the number of amastigotes per infected macrophage cell. CONCLUSION: Larval saliva and hemolymph of L. sericata have acceptable leishmanicidal properties against L. tropica.

Emerging of Cutaneous Leishmaniais Due to Leishmania major in a New Focus in Esfahan Province, Central Iran.

BACKGROUND: Esfahan Province is considered as one of the main focus of zoonotic cutaneous leishmaniasis (ZCL) in Iran. Recently, ZCL distribution is expanding through this province leading to report of new cases in non-endemic areas. In the current study epidemiological aspects of ZCL has been investigated in Naein County in Esfahan Province. METHODS: Adult sand flies were collected from beginning to the end of their seasonal activity. Rodents were caught by Sherman live traps once a month for one year. To active case detection, a hundred households in each selected village were visited in November and December 2016. Nested-PCR was employed to detect Leishmania parasite in the vector, reservoir and human. RESULTS: Totally 1562 sand flies including Phlebotomus sergenti, Phlebotomus papatasi, Sergentomyia sintoni and Sergentomyia mervinae were collected and identified. No Leishmania infection was detected in the collected sand flies. All of the 30 collected rodents were identified as Rhombomys opimus, and of these 3.3% and 26.7% were infected by Leishmania major using microscopic and molecular technique respectively. Totally, 914 individuals were investigated and the ulcer and scar rates of ZCL calculated to be at 1.1 and 15.3 per 1000 population, respectively. Molecular results confirmed L. major infection in human and reservoir samples. CONCLUSION: It is concluded that ZCL is established in the area in low endemicity, and it is extrapolated the disease will not be a serious increasing health problem in the near future in this region.

Molecular Identification of Leishmania Species in Phlebotomus alexandri (Diptera: Psychodidae) in Western Iran.

BACKGROUND: Visceral and cutaneous leishmaniasis are common in some areas of Iran and consider as health problems. Phlebotomus alexandri has been incriminated as a suspected vector for the both form of leishmaniasis. METHODS: This study was carried out in 4 western provinces of Iran. Sand flies were collected using sticky traps and light traps from indoor and outdoor resting places. Nested PCR was employed to detect Leishmania parasites among collected sand flies. RESULTS: Seven hundred and twenty two P. alexandri females were collected and pooled in 179 batches. Results of nested PCR showed, out of 9 samples from East Azerbaijan Province, only one sample was infected by Leishmania infantum. Of 34 individual and pooled samples from Kermanshah Province, only one pooled sample was infected with Leishmania major and among 30 individual and pooled samples in Fars Province, five specimens were infected by L. major, L. infantum, Leishmania donovani and Leishmania tropica. Furthermore, out of 108 individual and pooled samples from Khuzestan Province, 10 samples showed infection with L. major and L. infantum. CONCLUSION: The results of this study showed that P. alexandri is more active in hot zones than in moderate zones and this species may be considered as a permissive species.

Human immune response to Phlebotomus sergenti salivary gland antigens in a leishmaniasis-endemic focus in Iran.

Salivary proteins specific antibodies have been shown to be useful biomarkers of exposure to sand fly bites. This study aimed to investigate the level, duration, and dynamics of the human immune response against the SGL of Phlebotomus sergenti Parrot, 1917 (Diptera: Psychodidae), and to assess the immunoreactivity of human sera with SGL components in an endemic area of anthroponotic cutaneous leishmaniasis (ACL) in Iran. The study was carried out in 2-phase; longitudinal and cross-sectional. Sand flies were collected monthly from indoors and outdoors. In the longitudinal study, sera from healthy volunteers were collected monthly, and in the cross-sectional study, sera from healthy volunteers and patients with ACL lesion/s, were collected for immunoassay studies. The level of anti-P. sergenti saliva IgG was detected using the ELISA. Immunoreactivity of individual human sera with saliva components was also assessed by western blotting. Phlebotomus sergenti was the predominant sand fly species in the study area. The maximum and minimum percentages of IgG responses were seen in October (66%) and March (29%), respectively. Additionally, the cross-sectional study showed that 59.3% of the healthy volunteers and 80% of the patients were IgG positive. The antibody response against P. sergenti salivary gland was high during the sand fly active season and declined by the end of the activity of the vectors.  Antibody response against the SGL components of P. sergenti was transient and individual-specific. Some individuals shared a strong reaction against certain individual antigens, which could be considered as vector exposure markers for further investigation. LIST OF ABBREVIATIONS: ELISA: Enzyme-Linked Immunosorbent Assay; SDS PAGE: Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis; SGL: Salivary Gland Lysate; ACL: Anthroponotic Cutaneous Leishmaniasis; PBS: Phosphate Buffered Saline; BCA: Bicinchoninic Acid; PBS-T: Phosphate Buffered Saline Tween; FBS: Fetal Bovine Serum; HRP: Horseradish Peroxidase; TMB: 3,3',5,5'-Tetramethylbenzidine; PVDF: Polyvinylidene Difluoride; SGA: Salivary Gland Antigens; OD: Optical Density; KDa: Kilodalton; VL: Visceral Leishmaniasis; CL: Cutaneous Leishmaniasis; SGs: Salivary glands.

Epidemiological survey on Cutaneous Leishmaniasis in southwestern Iran.

BACKGROUND & OBJECTIVES: Cutaneous leishmaniasis (CL) is still a public health threat in many rural areas of Iran. It causes considerable morbidity among a huge number of people in the endemic areas. Khuzestan is considered as a CL focus in Iran and recently several cases have been reported from Shush County. The aim of the current study was to determine different aspects of CL epidemiology in this County. METHODS: The study was conducted in four villages in the rural district of Shush County. In order to survey the scar and acute rate of the disease, 100 inhabitants were questioned in each village. In addition, to determine the reservoir host and vector of the disease, rodents were captured using Sherman traps, and sandflies were collected from around the rodent burrows using sticky paper traps. RESULTS: The scar and acute lesion rates were reported as 5.26 and 0.57% among inhabitants, respectively. Fifty-four rodents including Tatera indica (55.5%), Nesokia indica (33.3%), Mus musculus (9.3%) and Rattus norvegicus (1.9%) were captured and identified. Nested PCR showed 3 out of 54 rodents infected with Leishmania major. In total, 1122 sandflies were collected, and Leishmania major detected in Phlebotomus papatasi (4.35%), Sergentomyia dentata (23%), and Sergentomyia clydei (10%). INTERPRETATION & CONCLUSION: Zoonotic cutaneous leishmaniasis due to L. major prevailed in this area. The common and the main vector was Ph. papatasi and, T. indica seems to be the main while N. indica as a secondary reservoir host of the disease in this area.

The Potential Role of Humans in the Transmission Cycle of Leishmania major (Kinetoplastida: Trypanosomatidae), the Causative Agent of the Old World Zoonotic Cutaneous Leishmaniasis.

Zoonotic cutaneous leishmaniasis (ZCL) due to Leishmania major (Yakimoff and Schokhor, 1914) is known as a neglected tropical disease, transmitted by Phlebotomus papatasi (Scopoli, 1786) in North Africa and the Middle East. The main reservoirs of ZCL are desert gerbils and the role of humans as a reservoir host of the disease is not clearly defined and is therefore investigated in this study. In order to achieve this objective, Ph. papatasi sand flies were allowed to take blood through indirect blood sources (human, Rhombomys opimus (Lichtenstein, 1823) (great gerbil), and BALB/c mice) using artificial feeding or direct xenodiagnoses from ZCL patients. To detect Leishmania promastigotes inside digestive canal, blood-fed sand flies were dissected and examined under light microscope and then confirmed by nested-PCR. In indirect xenodiagnoses, promastigote forms of Leishmania parasite were observed in 7.1%, 12%, and 13.6% of sand flies which fed on Rh. opimus, BALB/c mice, or human blood sources, respectively. In direct xenodiagnoses, among 93 female lab-reared sand flies which fed on active L. major lesion(s), 2.1% Leishmania infection was subsequently observed in the sand flies. No infection was detected in those sand flies that fed on nonlesioned skin of CL patients. Humans can serve as a reservoir of ZCL since the data indicate that Ph. papatasi is able to acquire L. major parasites from an active lesion of ZCL patients and the parasites can complete metacyclogenesis in the sand fly.

wsp-based analysis of Wolbachia strains associated with Phlebotomus papatasi and P. sergenti (Diptera: Psychodidae) main cutaneous leishmaniasis vectors, introduction of a new subgroup wSerg.

Sand flies of Phlebotomus papatasi and P. sergenti are the main vectors of cutaneous leishmanisis (CL) in the old world. We aimed to screen Iranian P. papatasi and P. sergenti for their natural infections with Wolbachia and to determine their phylogenetic association with other species. Wolbachia surface protein (wsp) gene was PCR amplified from DNA extracted from Phlebotomus species, sequenced, and were analysed in combination with wsp sequences related to Phelebtominae and other insects. All Wolbachia-infecting Iranian sand flies of P. papatasi and P. sergenti were classified in the Supergroup A., Wolbachia isolated from P. sergenti were clustered in a new subgroup within Supergroup A so-called wSreg. The Wolbachia strains identified from the P. papatasi clustered mainly in the subgroup wPap and partly in wSerg. Multiple Wholbachia infection within a single population of P.papatasi warrants investigation on existence and intensity of cytoplasmic incompatibility between the wPap and wSerg subgroups.

Assessing the Ovarian Accessory Glands to Determine the Parity of Phlebotomus papatasi, Vector of Zoonotic Cutaneous Leishmaniasis, under Laboratory Condition.

BACKGROUND: Zoonotic cutaneous leishmaniasis (ZCL) is a neglected tropical disease prevailed in many rural areas of 17 out of 31 provinces in Iran. The main vector of the disease is Phlebotomus papatasi and the causative agent is Leishmania major in ZCL foci of Iran. In the current study we investigated the validity of accessory glands secretions as an indicator to recognize parous from nulliparous Ph. papatasi females under laboratory conditions. METHODS: Over 235 laboratory-reared females of Ph. papatasi were dissected in 6 groups including: newly emerged, one hour, one day, two days, and three days after blood feeding and also after oviposition under stereo microscope for their parity in 2014-2015. RESULTS: Transparent glands were compatible with nulliparous only in newly emerged sand flies. In sand flies dissected after oviposition, accessory glands were rather large as a result of oviposition though they were transparent. CONCLUSION: The accessory glands secretions could not be as an indicator for distinguishing parous from nulliparous of Ph. papatasi females.

Rearing and Biology of Phlebotomus sergenti, the Main Vector of Anthroponotic Cutaneous Leishmaniasis in Iran.

BACKGROUND: Establishment of sand flies laboratory colonies is essential to understand various biological aspects of Phlebotominae sand flies. The aims of the current study were to establish the colony of Phlebotomus sergenti Parrot (1917), the main vector of anthroponotic cutaneous leishmaniasis in old world, and to study biological parameters of this species. METHODS: The sand flies were reared at 26-28 °C temperature, 14:10 (light: dark) photoperiod and 70-80% relative humidity. Larval diet was a composted mixture of rabbit faces and rabbit pellets which is prepared through a special process. First to fifth generations of P. sergenti were used to define biological parameters. RESULTS: Results showed that, P. sergenti blood feeding percentage were 42% on chicken, 21% on BALB/c and 37% on golden hamster. Average time of blood digestion, egg incubation, 1st instar larva, pupa and adult emerging was recorded at 3.4, 8.7, 15, 33.3 and 41.2 days after blood feeding, respectively. Mean number of laid eggs was 55.1 and retained eggs were 35 per a female. Fecundity and production rate were 61.6%, and 42.2% respectively. Average longevity recorded at 15.2 days for females and 14.8 days for males. CONCLUSION: Colony of P. sergenti has been established for the first time in Iran. Average interval time from egg to adult of this species was 32.5 days. Chicken and golden hamster were recommended as a blood source for colony initiation and routine blood feeding, respectively.

Rodenticide Comparative Effect of Klerat® and Zinc Phosphide for Controlling Zoonotic Cutaneous Leishmaniasis in Central Iran.

BACKGROUND: Zoonotic cutaneous leishmaniasis (ZCL) is a neglected disease with public health importance that is common in many rural areas of Iran. In recent years, behavioral resistance and/or bait shyness against the common rodenticide among reservoir hosts of ZCL have been reported. The aim of this study was to evaluate the effectiveness of Klerat® and zinc phosphide against natural reservoir of ZCL. METHODS: This survey was carried out in four villages located 45 to 95 km far from Esfahan City Esfahan province, central Iran from April to November 2011. The rodent burrows were counted destroyed and reopened holes baited around all villages. Effect of rodent control operation on the main vector density and incidence of ZCL were evaluated. RESULTS: The reduction rate of rodent burrows after intervention calculated to be at 62.8% in Klerat® and 58.15% in zinc phosphide treated areas. Statistical analysis showed no difference between the densities of the vector in indoors and outdoors in intervention and control areas. The incidence of the disease between treated and control areas after intervention was statistically different (P< 0.05). CONCLUSION: Klerat® could be a suitable alternative for zinc phosphide in a specific condition such as behavior resistance or occurrence of bait shyness.

Assessing the Fauna of Aquatic Insects for Possible Use for Malaria Vector Control in Large River, Central Iran.

Insects with over 30,000 aquatic species are known as very successful arthropods in freshwater habitats. Some of them are applied as biological indicators for water quality control, as well as the main food supply for fishes and amphibians. The faunistic studies are the basic step in entomological researches; the current study was carried out emphasizing on the fauna of aquatic insects in Karaj River, northern Iran. A field study was carried out in six various sampling site of Karaj River during spring 2013. The aquatic insects were collected using several methods such as D-frame nets, dipping and direct search on river floor stones. Specimens were collected and preserved in Ethanol and identified by standard identification keys. Totally, 211 samples were collected belonging to three orders; Plecoptera, Trichoptera and Ephemeroptera. Seven genuses (Perla, Isoperla, Hydropsyche, Cheumatopsyche, Baetis, Heptagenia and Maccafferium) from five families (Perlidae, Perlodidae, Hydropsychidae, Batidae, Heptagenidae) were identified. The most predominant order was Plecoptera followed by Trichoptera. Karaj River is a main and important river, which provides almost all of water of Karaj dam. So, identification of aquatic species which exist in this river is vital and further studies about systematic and ecological investigations should be performed. Also, monitoring of aquatic biota by trained health personnel can be a critical step to describe water quality in this river. Understanding the fauna of aquatic insects will provide a clue for possible biological control of medically important aquatic insects such as Anopheles as the malaria vectors.