The development of linear regression models using environmental variables to explain the spatial distribution of Fasciola hepatica infection in dairy herds in England and Wales

Fasciolosis caused by Fasciola hepatica is a major cause of economic loss to the agricultural community worldwide as a result of morbidity and mortality in livestock. Spatial models developed with the aid of Geographic Information Systems (GIS) can be used to develop risk maps for fasciolosis for use in the formulation of disease control programmes. Here we investigate the spatial epidemiology of F. hepatica in dairy herds in England and Wales and develop linear regression models to explain observed patterns of exposure at a small spatial unit, the postcode area. Exposure data used for the analysis were taken from an earlier study of F. hepatica infection, performed in the winter of 2006/7. Climatic, environmental, soil, livestock and pasture variables were considered as potential predictors. The performance of models that used climate variables for 5 years average data, contemporary data and a combination of both for England and Wales, and for England only, was compared. All models explained over 70% of the variation in the prevalence of exposure. The best performing models were those built using 5 year average and contemporary weather data. However, the fit of these models was only slightly better than the fit of models using weather data from one time period only. Rainfall was a consistent predictor in all models. Other model covariates included temperature, the negative predictors of soil pH and slope and the positive predictors of poor quality land, as determined by the Agricultural Land Classification, and very fine sand content of soil. Choroplethic risk maps showed a good match between the observed F. hepatica exposure values and exposure values fitted by the models. The development of these detailed spatial models is the first step towards the development of a spatially specific, temporal forecasting system for liver fluke in the United Kingdom.     

Fasciola hepatica in Snails Collected from Water-Dropwort Fields using PCR

Fasciola hepatica is a trematode that causes zoonosis mainly in cattle and sheep and occasionally in humans. Fascioliasis has been reported in Korea; however, determining F. hepatica infection in snails has not been done recently. Thus, using PCR, we evaluated the prevalence of F. hepatica infection in snails at 4 large water-dropwort fields. Among 349 examined snails, F. hepatica-specific internal transcribed space 1 (ITS-1) and/or ITS-2 markers were detected in 12 snails and confirmed using sequence analysis. Morphologically, 213 of 349 collected snails were dextral shelled, which is the same aperture as the lymnaeid snail, the vectorial host for F. hepatica. Among the 12 F. hepatica-infected snails, 6 were known first intermediate hosts in Korea (Lymnaea viridis and L. ollula) and the remaining 6 (Lymnaea sp.) were potentially a new first intermediate host in Korea. It has been shown that the overall prevalence of the snails contaminated with F. hepatica in water-dropwort fields was 3.4%; however, the prevalence varied among the fields. This is the first study to estimate the prevalence of F. hepatica infection using the vectorial capacity of the snails in Korea.     

A Case of Fasciola hepatica Infection Mimicking Cholangiocarcinoma and ITS-1 Sequencing of the Worm

Fascioliasis is a zoonotic infection caused by Fasciola hepatica or Fasciola gigantica. We report an 87-year-old Korean male patient with postprandial abdominal pain and discomfort due to F. hepatica infection who was diagnosed and managed by endoscopic retrograde cholangiopancreatography (ERCP) with extraction of 2 worms. At his first visit to the hospital, a gallbladder stone was suspected. CT and magnetic retrograde cholangiopancreatography (MRCP) showed an intraductal mass in the common bile duct (CBD) without proximal duct dilatation. Based on radiological findings, the presumed diagnosis was intraductal cholangiocarcinoma. However, in ERCP which was performed for biliary decompression and tissue diagnosis, movable materials were detected in the CBD. Using a basket, 2 living leaf-like parasites were removed. The worms were morphologically compatible with F. hepatica. To rule out the possibility of the worms to be another morphologically close species, in particular F. gigantica, 1 specimen was processed for genetic analysis of its ITS-1 region. The results showed that the present worms were genetically identical (100%) with F. hepatica but different from F. gigantica.     

Stimulation of resistance to Taenia taeniaeformis in the rat by infection with Fasciola hepatica

Homologous resistance to F. hepatica and T. taeniaeformis and cross resistance between these two parasites was investigated in the rat. Rats given a primary infection with F. hepatica were challenged with either F. hepatica or T. taeniaeformis. Conversely rats given a primary infection with T. taeniaeformis were challenged with either F. hepatica or T. taeniaeformis.Infection with F. hepatica generated significant resistance against challenge with F. hepatica given 9 weeks later. Similarly, infection with T. taeniaeformis protected against challenge with T. taeniaeformis given 6 weeks later. Infection with F. hepatica also generated significant resistance against challenge with T. taeniaeformis given 4, 8 or 9 weeks later. Primary infection with T. taeniaeformis did not protect against challenge with F. hepatica.     

Higher non-return rate associated with Mycobacterium avium subspecies paratuberculosis infection at early stage in Holstein dairy cows

The effects of infection by Mycobacterium avium paratuberculosis (Map) on dairy cows are poorly documented and quite controversial. This retrospective study aimed at quantifying the variation in non-return to service of Holstein dairy cows according to their Map-infection status. Three different statuses were defined based on both individual and herd tests results: ELISA positive cow, all tests negative cow in a negative herd and all tests negative cow in a positive herd. Whatever the age at Map testing, the status was attributed to a cow from its first lactation onwards. Non-return to service was determined at 200 days after first and second services. The study was performed from 1999 to 2007 on 185,950 AI from 48,914 cows in early stage of the infection in 1069 herds by logistic regression controlling for known factors influencing non-return rate. Non-return rate was higher for infected cows compared to negative cows from negative herds (RR of 1.10 or +3.9 points of % of non-return rate). The effect was significant for parities 1 and 2 (RR of 1.11 and 1.12, respectively) but not for higher ones. This effect was lower when comparing positive cows to negative cows in the same herds but relative risks were still above 1. The hypothesis that the effect of Map on non-return depends upon the stage of infection is formulated.     

Schistosoma mansoni and Fasciola hepatica: Cross-resistance in mice

Cross-resistance in Schistosoma mansoni and Fasciola hepatica infections were studied in mice. A primary infection of S. mansoni, 7 to 28 days old, did not stimulate a significant level of resistance to heterologous challenge with F. hepatica. In contrast, in older S. mansoni infections (54–65 days old) there was a significant level of resistance to a challenge with F. hepatica. The F. hepatica worm burden was reduced by 34.0 to 72.5% in separate experiments. Challenge infection with F. hepatica did not influence the number of S. mansoni in primary infections. No heterologous resistance to S. mansoni was found in mice with 7- and 23-day-old F. hepatica infections. However, primary infections with F. hepatica, 28, 32, 42, and 50 days old, conferred significant resistance to a heterologous challenge with S. mansoni. The established schistosome worm burden was reduced by 41.5 to 50.4%. In no case was the primary F. hepatica burden reciprocally influenced by challenge infection with S. mansoni.     

Monitoring of Fasciola Species Contamination in Water Dropwort by cox1 Mitochondrial and ITS-2 rDNA Sequencing Analysis

Fascioliasis, a food-borne trematode zoonosis, is a disease primarily in cattle and sheep and occasionally in humans. Water dropwort (Oenanthe javanica), an aquatic perennial herb, is a common second intermediate host of Fasciola, and the fresh stems and leaves are widely used as a seasoning in the Korean diet. However, no information regarding Fasciola species contamination in water dropwort is available. Here, we collected 500 samples of water dropwort in 3 areas in Korea during February and March 2015, and the water dropwort contamination of Fasciola species was monitored by DNA sequencing analysis of the Fasciola hepatica and Fasciola gigantica specific mitochondrial cytochrome c oxidase subunit 1 (cox1) and nuclear ribosomal internal transcribed spacer 2 (ITS-2). Among the 500 samples assessed, the presence of F. hepatica cox1 and 1TS-2 markers were detected in 2 samples, and F. hepatica contamination was confirmed by sequencing analysis. The nucleotide sequences of cox1 PCR products from the 2 F. hepatica-contaminated samples were 96.5% identical to the F. hepatica cox1 sequences in GenBank, whereas F. gigantica cox1 sequences were 46.8% similar with the sequence detected from the cox1 positive samples. However, F. gigantica cox1 and ITS-2 markers were not detected by PCR in the 500 samples of water dropwort. Collectively, in this survey of the water dropwort contamination with Fasciola species, very low prevalence of F. hepatica contamination was detected in the samples.     

Further observations on the nature and characteristics of cross protection against Fasciola hepatica produced in sheep by infection with Cysticercus tenuicollis

Dineen J. K., Kelly J. D. and Campbell N. J. 1978. Further observations on the nature and characteristics of cross protection against Fasciola hepatica produced in sheep by infection with Cysticercus tenuicollis. International Journal for Parasitology8: 173–176. Previous studies showed that sheep infected with Cysticercus tenuicollis were protected against a subsequent infection with Fasciola hepatica given at 12 weeks (Campbell, Kelly. Townsend & Dineen, 1977). The present studies showed that these animals were again protected against re-challenge with F. hepatica at 9 months. Resistance was detected about 10 weeks after re-challenge with metacercariae.Sheep in which the initial C. tenuicollis infections were terminated by anthelmintic at 12 weeks, were resistant to the primary infection with F. hepatica but became fully susceptible to the re-challenge at 9 months.These results suggest that maintenance of resistance depends upon persistence of the C. tenuicollis infections. They also indicate that resistance is maintained by cysts in the peritoneum which is remote from the reactive site (liver).Infection with F. hepatica at 3 weeks after infection with C. tenuicollis prevented cross protection against both the primary infection with F. hepatica and re-challenge at 9 months.     

Genotypic characterization and species identification of Fasciola spp. with implications regarding the isolates infecting goats in Vietnam

Ribosomal RNA sequences (361 or 362 bp) of the second internal transcribed spacer 2 (ITS-2) and a portion of mitochondrial cox1 (423 bp) for Fasciola spp. obtained from specimens collected in indigenous and hybrid goats and sheep in Vietnam were characterized for genotypic status and hybridization/introgression. Alignment of 48 ITS-2 sequences (also those from goats and sheep in this study) indicates that F. gigantica and F. hepatica differ typically from each other at seven sites whereas one of these is a distinguishing deletion (T) at the 327th position in F. gigantica relative to F. hepatica. The isolates from the mountainous goats in the North of Vietnam (Yen Bai province) showed the ITS-2 composition relatively identical to that of F. hepatica. The ITS-2 sequences from populations of Fasciola isolates in goats had probably experienced introgression/hybridization as reported previously in other ruminants and humans. All Vietnamese goat-of-origin specimens had high pairwise percentage of mitochondrial cox1 sequences to F. gigantica (97-100%), and very low identity to F. hepatica (91-93%), suggesting their maternal linkage to be traced to F. gigantica. The presence of hybrid and/or introgressed populations of liver flukes bearing genetic material from both F. hepatica and F. gigantica in the goats/sheep in Vietnam, regardless of indigenous or imported hosts, appears to be the first demonstration from a tropical country.     

Isolation of Fasciola hepatica genus-specific antigens

Santiago de Weil N., Hillyer G. V. and Pacheco E. 1984. Isolation of Fasciola hepatica genus-specific antigens. International Journal for Parasitology14: 197–206. The Fasciola hepatica antigens which induce antibody formation in acute fascioliasis were isolated by acid elution after reacting an F. hepatica tegument antigen extract with a CNBr-Sepharose 4B column coupled with IgG obtained from the serum of rabbits infected with fascioliasis for 6–10 weeks. These isolated antigens were further separated by gel filtration using a column packed with Sephacryl S-200. In this manner three major peaks were obtained. The best serologic antigens were found in peak 2 which had a mol. wt range of 14,000–43,000. This peak contains genus-specific F. hepatica antigens which are highly reactive with fascioliasis serum. These antigens do not cross-react with either Schistosoma mansoni or with bovine serum albumin by gel diffusion. Monitoring by ELISA and gel diffusion with heterologous and homologous antisera showed that as purification by antibody affinity chromatography proceeded, cross reactivity with S. mansoni was eliminated. The rabbit antiserum obtained against peak 2, when tested by immunoelectrophoresis with a crude F. hepatica extract shows one main band identical to the main band observed with serum from acutely infected rabbits. Up to two other minor bands can be detected using concentrated homologous antisera. Fractions obtained from preparative iso-electric focusing of the F. hepatica tegument extract were reacted with sera from rabbits with acute fascioliasis. Two main bands were observed in immunodiffusion with antigens eluting in a pH range of 7.4–8.7. When these fractions were monitored with anti peak 2 antisera, two precipitin bands appeared with antigens eluting in a pH range of 7.4–7.9. The F. hepatica genus-specific antigen pool was applied to ELISA to evaluate its ability to detect antibody in a primary F. hepatica infection in rabbits. A rise in absorbance values could be detected by 2 weeks of infection, reached high levels by 6 weeks and remained high through 28 weeks of infection.     

Modeling and Predicting Seasonal Influenza Transmission in Warm Regions Using Climatological Parameters

Background

Influenza transmission is often associated with climatic factors. As the epidemic pattern varies geographically, the roles of climatic factors may not be unique. Previous in vivo studies revealed the direct effect of winter-like humidity on air-borne influenza transmission that dominates in regions with temperate climate, while influenza in the tropics is more effectively transmitted through direct contact.

Methodology/Principal Findings

Using time series model, we analyzed the role of climatic factors on the epidemiology of influenza transmission in two regions characterized by warm climate: Hong Kong (China) and Maricopa County (Arizona, USA). These two regions have comparable temperature but distinctly different rainfall. Specifically we employed Autoregressive Integrated Moving Average (ARIMA) model along with climatic parameters as measured from ground stations and NASA satellites. Our studies showed that including the climatic variables as input series result in models with better performance than the univariate model where the influenza cases depend only on its past values and error signal. The best model for Hong Kong influenza was obtained when Land Surface Temperature (LST), rainfall and relative humidity were included as input series. Meanwhile for Maricopa County we found that including either maximum atmospheric pressure or mean air temperature gave the most improvement in the model performances.

Conclusions/Significance

Our results showed that including the environmental variables generally increases the prediction capability. Therefore, for countries without advanced influenza surveillance systems, environmental variables can be used for estimating influenza transmission at present and in the near future.     

The First Case of Capillaria hepatica Infection in a Nutria (Myocastor coypus) in Korea

This study reports the first case of Capillaria hepatica infection in a nutria in Korea. Ten nutrias, captured near the Nakdong River, were submitted to our laboratory for necropsy. White-yellowish nodules were found in the liver of 1 of the nutrias at necropsy. Histologically, the lesions were granulomatous, and infiltrations of lipid-laden macrophages, eosinophils, and several multinucleated giant cells were observed. The lesions consisted of numerous eggs and necrotic hepatocytes. The eggs were lemon-shaped and had polar plugs at the ends of both long sides. The eggs were morphologically identified as those of C. hepatica. Worldwide, C. hepatica infection in nutrias is very rare. Nutrias are a kind of livestock, as well as wildlife; therefore, an epidemiological study for parasitic infections needs to be conducted.     

Assessing the potential impact of Salmonella vaccines in an endemically infected dairy herd

Salmonella spp. in cattle contribute to bacterial foodborne disease for humans. Reduction of Salmonella prevalence in herds is important to prevent human Salmonella infections. Typical control measures are culling of infectious animals, vaccination, and improved hygiene management. Vaccines have been developed for controlling Salmonella transmission in dairy herds; however, these vaccines are imperfect and a variety of vaccine effects on susceptibility, infectiousness, Salmonella shedding level, and duration of infectious period were reported. To assess the potential impact of imperfect Salmonella vaccines on prevalence over time and the eradication criterion, we developed a deterministic compartmental model with both replacement (cohort) and lifetime (continuous) vaccination strategies, and applied it to a Salmonella Cerro infection in a dairy farm. To understand the uncertainty of prevalence and identify key model parameters, global parameter uncertainty and sensitivity analyses were performed. The results show that imperfect Salmonella vaccines reduce the prevalence of Salmonella Cerro. Among three vaccine effects that were being considered, decreasing the length of the infectious period is most effective in reducing the endemic prevalence. Analyses of contour lines of prevalence or the critical reproduction ratio illustrate that, reducing prevalence to a certain level or zero can be achieved by choosing vaccines that have either a single vaccine effect at relatively high effectiveness, or two or more vaccine effects at relatively low effectiveness. Parameter sensitivity analysis suggests that effective control measures through applying Salmonella vaccines should be adjusted at different stages of infection. In addition, lifetime (continuous) vaccination is more effective than replacement (cohort) vaccination. The potential application of the developed vaccination model to other Salmonella serotypes related to foodborne diseases was also discussed. The presented study may be used as a tool for guiding the development of Salmonella vaccines.     

Molecular characterization of Fasciola hepatica from Sardinia based on sequence analysis of genomic and mitochondrial gene markers

The aim of the present study is to investigate for the first time the genetic diversity of samples identified morphologically as Fasciola hepatica (Platyhelminthes: Trematoda: Digenea) (n = 66) from sheep and cattle from two localities of Sardinia and to compare them with available data from other localities by partial sequences of the first (ITS-1), the 5.8S, and second (ITS-2) Internal Transcribed Spacers (ITS) of nuclear ribosomal DNA (rDNA) genes, the mitochondrial cytochrome c oxidase subunit I (COI), and nicotinamide adenine dinucleotide dehydrogenase subunit I (ND1) genes. Comparison of the sequences from Sardinia with sequences of Fasciola spp. from GenBank confirmed that all samples belong to the species F. hepatica. The nucleotide sequencing of ITS rDNA showed no nucleotide variation in the ITS-1, 5.8S and ITS-2 rDNA sequences among all Sardinian samples, comparing with two ITS-2 haplotypes in standard F. hepatica, showing a substitution C/T in 20 position 859, reported previously from Tunisia, Algeria, Australia, Uruguay and Spain. The present study shows that in Sardinian sheep and cattle there is the most frequent haplotype (FhITS-H1) of F. hepatica species from South Europe. Considering NDI sequences, the phylogenetic trees showed reliable grouping among the haplotypes of F. hepatica from Sardinia and the mitochondrial lineage I, including the main N1 haplotype, observed previously from Europe (Russia, Belarus, Ukraine and Bulgaria), Armenia, West Africa (Nigeria), America (Uruguay and USA), Asia (Turkey, Japan, and China), Georgia, Turkmenistan, Azerbaijan and Australia. Furthermore, common haplotypes FhCOI-H1 and FhCOI-H2 of F. hepatica from Sardinia also corresponded mostly to the first lineage including the main C1 haplotype reported previously from Eastern European and Western Asian populations, they belonged just to a phylogenically distinguishable clade, as F. hepatica from Australia, France, Turkey, Uruguay, Russia, Armenia, Ukraine, Belarus, Turkmenistan, USA, Tunisia and Algeria, indicating that this is the main haplotype involved in the spread of F. hepatica throughout all continents.     

The impact of climate change on the geographical distribution of two vectors of Chagas disease: implications for the force of infection

Chagas disease, caused by the parasite Trypanosoma cruzi, is the most important vector-borne disease in Latin America. The vectors are insects belonging to the Triatominae (Hemiptera, Reduviidae), and are widely distributed in the Americas. Here, we assess the implications of climatic projections for 2050 on the geographical footprint of two of the main Chagas disease vectors: Rhodnius prolixus (tropical species) and Triatoma infestans (temperate species). We estimated the epidemiological implications of current to future transitions in the climatic niche in terms of changes in the force of infection (FOI) on the rural population of two countries: Venezuela (tropical) and Argentina (temperate). The climatic projections for 2050 showed heterogeneous impact on the climatic niches of both vector species, with a decreasing trend of suitability of areas that are currently at high-to-moderate transmission risk. Consequently, climatic projections affected differently the FOI for Chagas disease in Venezuela and Argentina. Despite the heterogeneous results, our main conclusions point out a decreasing trend in the number of new cases of Tr. cruzi human infections per year between current and future conditions using a climatic niche approach.     

Evaluation of losses in carcasses of cattle naturally infected with Fasciola hepatica: effects on weight by age range and on carcass quality parameters

Although fasciolosis is a relatively common disease, the productive and economic losses resulting from cattle with chronic fasciolosis are unclear. This paper aims to investigate the effect of fasciolosis on the parameters of carcass quality and discuss the hypothesis that the effects on weight differ among age ranges of cattle. For this, we analysed abattoir data of 30,151 bovines, from 928 farms, slaughtered in Uruguay in 2016, of which 33.9% (95% confidence interval (CI): 27.3–41.1%) had Fasciola hepatica (liver fluke). A mixed model was built to assess whether the effect of fasciolosis on weight differs depending on the age range, using the interaction term ‘age*F. hepatica’. The effect on the carcass parameters was tested using a proportional logistic regression. The interaction of age and F. hepatica was statistically significant (P < 0.001). Differences in carcass weights between infected and non-infected animals were observed mostly at younger ages (up to 30 months), with the highest difference observed in the 23–30 months age range (estimated marginal mean difference of 6.34 kg). Overall, the presence of F. hepatica was positively associated with poor conformations and lower fat scores of carcasses (P < 0.001). The carcasses of cattle infected with F. hepatica had 0.16 times greater odds of having worse conformation scores than carcasses of cattle without F. hepatica (proportional odds ratio (POR) = 1.16; 95% CI: 1.07–1.26). Similarly, carcasses of cattle with F. hepatica had 0.30 times (POR = 1.30, 95% CI: 1.23–1.39) greater odds of having poorer fat scores than carcasses of cattle without F. hepatica. Therefore, infection with F. hepatica is associated with poorer carcass quality parameters and lower weights, and the effect on weight differs across age ranges.     

Schistosoma mansoni and Fasciola hepatica: Cross-resistance in mice with single-sex schistosome infections

Primary Schistosoma mansoni single-sex infections in mice, i.e., either male only or female only, did not stimulate any detectable level of heterologous resistance to challenge with Fasciola hepatica after 22 to 76 days, while statistically significant resistance to a challenge with F. hepatica was demonstrated in the presence of patent mixed-sex S. mansoni infections. Simultaneous infections with S. mansoni and F. hepatica induced a statistically significant reduction in the number of schisto-some worms established, i.e., the burden being reduced by 40.1 and 43.9%, respectively. There was no reduction of the F. hepatica worm burden. Similar features could be observed with a time interval of 48 hr between the S. mansoni infection and the F. hepatica challenge, i.e., the schistosome burden being reduced by 34.2 and 45.6%, respectively. Furthermore, simultaneous infections with S. mansoni and F. hepatica induced a statistically significant reduction of the egg production capacity per paired female schistosome worm as compared with that of the S. mansoni control group. Tissue egg counts of the various intestinal sections were reduced by 92.8–99.6%.     

Fasciola hepatica: Comparative studies on fascioliasis in rats and mice

Chapman C. B. and Mitchell G. F. 1982. Fasciola hepatica: comparative studies on fascioliasis in rats and mice. International Journal for Parasitology12: 81–91. Certain characteristics of infection differ between rats and mice exposed to metacercariae of the trematode parasite, Fasciola hepatica. Rats develop a degree of age-related resistance (and infected older females contain fewer parasites than older males), resistance to reinfection in infected rats is demonstrated readily though is partial, and a comparable degree of resistance can be obtained in recipients of infected rat serum provided the serum is given at about the time of challenge. None of these features of F. hepatica infection is seen in mice. Rats also differ from mice in that they can be vaccinated against infection (although again, resistance is incomplete) using larval antigen mixtures in adjuvants. Mice do respond to infection by production of antilarval antibodies and a slight IgG₁ hypergammaglobulinaemia and larvae will sensitize mice for delayed hypersensitivity. The results of this study indicate that sera from infected rats versus infected mice will be useful in pinpointing antigens of F. hepatica larvae which are involved in expression of partial host protection.     

Identification and differentiation of Fasciola hepatica and Fasciola gigantica using a simple PCR-restriction enzyme method

Accurate morphological differentiation between the liver fluke species Fasciola hepatica and Fasciola gigantica is difficult. We evaluated PCR-restriction enzyme profiles of internal transcribed spacer 1 (ITS1) that could aid in their identification. Fifty F. hepatica and 30 F. gigantica specimens were collected from different hosts in three provinces of Iran. For DNA extraction, we crushed fragments of the worms between two glass slides as a new method to break down the cells. DNA from the crushed materials was then extracted with a conventional phenol-chloroform method and with the newly developed technique, commercial FTA cards. A primer pair was selected to amplify a 463-bp region of the ITS1 sequence. After sequencing 14 samples and in silico analysis, cutting sites of all known enzymes were predicted and TasI was selected as the enzyme that yielded the most informative profile. Crushing produced enough DNA for PCR amplification with both the phenol-chloroform and commercial FTA card method. The DNA extracted from all samples was successfully amplified and yielded a single sharp band of the expected size. Digestion of PCR products with TasI allowed us to distinguish the two species. In all samples, molecular identification was consistent with morphological identification. Our PCR-restriction enzyme profile is a simple, rapid and reliable method for differentiating F. hepatica and F. gigantica, and can be used for diagnostic and epidemiological purposes.