Urbanization,land tenure security and vector-borne Chagas disease

Modern cities represent one of the fastest growing ecosystems on the planet. Urbanization occurs in stages; each stage characterized by a distinct habitat that may be more or less susceptible to the establishment of disease vector populations and the transmission of vector-borne pathogens. We performed longitudinal entomological and epidemiological surveys in households along a 1900 × 125 m transect of Arequipa, Peru, a major city of nearly one million inhabitants, in which the transmission of Trypanosoma cruzi, the aetiological agent of Chagas disease, by the insect vector Triatoma infestans, is an ongoing problem. The transect spans a cline of urban development from established communities to land invasions. We find that the vector is tracking the development of the city, and the parasite, in turn, is tracking the dispersal of the vector. New urbanizations are free of vector infestation for decades. T. cruzi transmission is very recent and concentrated in more established communities. The increase in land tenure security during the course of urbanization, if not accompanied by reasonable and enforceable zoning codes, initiates an influx of construction materials, people and animals that creates fertile conditions for epidemics of some vector-borne diseases.     

Modelling the proportion of influenza infections within households during pandemic and non-pandemic years

Background

The key epidemiological difference between pandemic and seasonal influenza is that the population is largely susceptible during a pandemic, whereas, during non-pandemic seasons a level of immunity exists. The population-level efficacy of household-based mitigation strategies depends on the proportion of infections that occur within households. In general, mitigation measures such as isolation and quarantine are more effective at the population level if the proportion of household transmission is low.

Methods/Results

We calculated the proportion of infections within households during pandemic years compared with non-pandemic years using a deterministic model of household transmission in which all combinations of household size and individual infection states were enumerated explicitly. We found that the proportion of infections that occur within households was only partially influenced by the hazard h of infection within household relative to the hazard of infection outside the household, especially for small basic reproductive numbers. During pandemics, the number of within-household infections was lower than one might expect for a given because many of the susceptible individuals were infected from the community and the number of susceptible individuals within household was thus depleted rapidly. In addition, we found that for the value of at which 30% of infections occur within households during non-pandemic years, a similar 31% of infections occur within households during pandemic years.

Interpretation

We suggest that a trade off between the community force of infection and the number of susceptible individuals in a household explains an apparent invariance in the proportion of infections that occur in households in our model. During a pandemic, although there are more susceptible individuals in a household, the community force of infection is very high. However, during non-pandemic years, the force of infection is much lower but there are fewer susceptible individuals within the household.     

Aquatic polymers can drive pathogen transmission in coastal ecosystems

Gelatinous polymers including extracellular polymeric substances (EPSs) are fundamental to biophysical processes in aquatic habitats, including mediating aggregation processes and functioning as the matrix of biofilms. Yet insight into the impact of these sticky molecules on the environmental transmission of pathogens in the ocean is limited. We used the zoonotic parasite Toxoplasma gondii as a model to evaluate polymer-mediated mechanisms that promote transmission of terrestrially derived pathogens to marine fauna and humans. We show that transparent exopolymer particles, a particulate form of EPS, enhance T. gondii association with marine aggregates, material consumed by organisms otherwise unable to access micrometre-sized particles. Adhesion to EPS biofilms on macroalgae also captures T. gondii from the water, enabling uptake of pathogens by invertebrates that feed on kelp surfaces. We demonstrate the acquisition, concentration and retention of T. gondii by kelp-grazing snails, which can transmit T. gondii to threatened California sea otters. Results highlight novel mechanisms whereby aquatic polymers facilitate incorporation of pathogens into food webs via association with particle aggregates and biofilms. Identifying the critical role of invisible polymers in transmission of pathogens in the ocean represents a fundamental advance in understanding and mitigating the health impacts of coastal habitat pollution with contaminated runoff.     

Analysis of Binary Multivariate Longitudinal Data via 2-Dimensional Orbits: An Application to the Agincourt Health and Socio-Demographic Surveillance System in South Africa

We analyse demographic longitudinal survey data of South African (SA) and Mozambican (MOZ) rural households from the Agincourt Health and Socio-Demographic Surveillance System in South Africa. In particular, we determine whether absolute poverty status (APS) is associated with selected household variables pertaining to socio-economic determination, namely household head age, household size, cumulative death, adults to minor ratio, and influx. For comparative purposes, households are classified according to household head nationality (SA or MOZ) and APS (rich or poor). The longitudinal data of each of the four subpopulations (SA rich, SA poor, MOZ rich, and MOZ poor) is a five-dimensional space defined by binary variables (questions), subjects, and time. We use the orbit method to represent binary multivariate longitudinal data (BMLD) of each household as a two-dimensional orbit and to visualise dynamics and behaviour of the population. At each time step, a point (x, y) from the orbit of a household corresponds to the observation of the household, where x is a binary sequence of responses and y is an ordering of variables. The ordering of variables is dynamically rearranged such that clusters and holes associated to least and frequently changing variables in the state space respectively, are exposed. Analysis of orbits reveals information of change at both individual- and population-level, change patterns in the data, capacity of states in the state space, and density of state transitions in the orbits. Analysis of household orbits of the four subpopulations show association between (i) households headed by older adults and rich households, (ii) large household size and poor households, and (iii) households with more minors than adults and poor households. Our results are compared to other methods of BMLD analysis.     

Computationally exact methods for stochastic periodic dynamics: Spatiotemporal dispersal and temporally forced transmission

The dynamics of many diseases and populations possess distinct recurring phases. For example, many species breed only during a subset of the year and the infection dynamics of many pathogens have transmission rates that vary with season. Here I investigate computational methods for studying transient and long-term behaviour of stochastic models which have periodic phases—several different potential techniques for studying long-term behaviour will be contrasted. I illustrate the results with two studies: The first is of a spatially realistic metapopulation model of malleefowl (Leipoa ocellata), a species which disperses only during a quarter of the year; this model is used to highlight the advantages and disadvantages of the particular methods presented. The second study is of a model for disease dynamics which incorporates seasonality in both the rate of within-population transmission and also in the rate of transmission effected via aerosol importation. This model has applications to studying disease invasion and persistence in captive-breeding populations. We demonstrate, via comparison to appropriately matched models with constant transmission rates and also no aerosol transmission, that seasonality and aerosol importation may alter control choices, with possibly an increase in the threshold population size for local control surveillance, transfer of importance to limiting aerosol transmission, and the use of temporally targetted surveillance. The methodology presented is the gold-standard for dealing with many phased processes in ecology and epidemiology, but its application is limited to systems of small size.     

Estimating and interpreting secondary attack risk: Binomial considered biased

The household secondary attack risk (SAR), often called the secondary attack rate or secondary infection risk, is the probability of infectious contact from an infectious household member A to a given household member B, where we define infectious contact to be a contact sufficient to infect B if he or she is susceptible. Estimation of the SAR is an important part of understanding and controlling the transmission of infectious diseases. In practice, it is most often estimated using binomial models such as logistic regression, which implicitly attribute all secondary infections in a household to the primary case. In the simplest case, the number of secondary infections in a household with m susceptibles and a single primary case is modeled as a binomial(m, p) random variable where p is the SAR. Although it has long been understood that transmission within households is not binomial, it is thought that multiple generations of transmission can be neglected safely when p is small. We use probability generating functions and simulations to show that this is a mistake. The proportion of susceptible household members infected can be substantially larger than the SAR even when p is small. As a result, binomial estimates of the SAR are biased upward and their confidence intervals have poor coverage probabilities even if adjusted for clustering. Accurate point and interval estimates of the SAR can be obtained using longitudinal chain binomial models or pairwise survival analysis, which account for multiple generations of transmission within households, the ongoing risk of infection from outside the household, and incomplete follow-up. We illustrate the practical implications of these results in an analysis of household surveillance data collected by the Los Angeles County Department of Public Health during the 2009 influenza A (H1N1) pandemic.     

Hrp?mutant bacteria as biocontrol agents: Toward a sustainable approach in the fight against plant pathogenic bacteria

Sustainable agriculture necessitates development of environmentally safe methods to protect plants against pathogens. Among these methods, application of biocontrol agents has been efficiently used to minimize disease development. Here we review current understanding of mechanisms involved in biocontrol of the main Gram-phytopathogenic bacteria-induced diseases by plant inoculation with strains mutated in hrp (hypersensitive response and pathogenicity) genes. These mutants are able to penetrate plant tissues and to stimulate basal resistance of plants. Novel protection mechanisms involving the phytohormone abscisic acid appear to play key roles in the biocontrol of wilt disease induced by Ralstonia solanacearum in Arabidopsis thaliana. Fully understanding these mechanisms and extending the studies to other pathosystems are still required to evaluate their importance in disease protection.     

Effects of pathogen dependency in a multi-pathogen infectious disease system including population level heterogeneity – a simulation study

Background

Increased computational resources have made individual based models popular for modelling epidemics. They have the advantage of incorporating heterogeneous features, including realistic population structures (like e.g. households). Existing stochastic simulation studies of epidemics, however, have been developed mainly for incorporating single pathogen scenarios although the effect of different pathogens might directly or indirectly (e.g. via contact reductions) effect the spread of each pathogen. The goal of this work was to simulate a stochastic agent based system incorporating the effect of multiple pathogens, accounting for the household based transmission process and the dependency among pathogens.

Methods

With the help of simulations from such a system, we observed the behaviour of the epidemics in different scenarios. The scenarios included different household size distributions, dependency versus independency of pathogens, and also the degree of dependency expressed through household isolation during symptomatic phase of individuals. Generalized additive models were used to model the association between the epidemiological parameters of interest on the variation in the parameter values from the simulation data. All the simulations and statistical analyses were performed using R 3.4.0.

Results

We demonstrated the importance of considering pathogen dependency using two pathogens, and showing the difference when considered independent versus dependent. Additionally for the general scenario with more pathogens, the assumption of dependency among pathogens and the household size distribution in the population cohort was found to be effective in containing the epidemic process. Additionally, populations with larger household sizes reached the epidemic peak faster than societies with smaller household sizes but dependencies among pathogens did not affect this outcome significantly. Larger households had more infections in all population cohort examples considered in our simulations. Increase in household isolation coefficient for pathogen dependency also could control the epidemic process.

Conclusion

Presence of multiple pathogens and their interaction can impact the behaviour of an epidemic across cohorts with different household size distributions. Future household cohort studies identifying multiple pathogens will provide useful data to verify the interaction processes in such an infectious disease system.

     

Social contact patterns in Vietnam and implications for the control of infectious diseases

Background

The spread of infectious diseases from person to person is determined by the frequency and nature of contacts between infected and susceptible members of the population. Although there is a long history of using mathematical models to understand these transmission dynamics, there are still remarkably little empirical data on contact behaviors with which to parameterize these models. Even starker is the almost complete absence of data from developing countries. We sought to address this knowledge gap by conducting a household based social contact diary in rural Vietnam.

Methods and Findings

A diary based survey of social contact patterns was conducted in a household-structured community cohort in North Vietnam in 2007. We used generalized estimating equations to model the number of contacts while taking into account the household sampling design, and used weighting to balance the household size and age distribution towards the Vietnamese population. We recorded 6675 contacts from 865 participants in 264 different households and found that mixing patterns were assortative by age but were more homogenous than observed in a recent European study. We also observed that physical contacts were more concentrated in the home setting in Vietnam than in Europe but the overall level of physical contact was lower. A model of individual versus household vaccination strategies revealed no difference between strategies in the impact on R ₀.

Conclusions and Significance

This work is the first to estimate contact patterns relevant to the spread of infections transmitted from person to person by non-sexual routes in a developing country setting. The results show interesting similarities and differences from European data and demonstrate the importance of context specific data.     

Calculation of Disease Dynamics in a Population of Households

Early mathematical representations of infectious disease dynamics assumed a single, large, homogeneously mixing population. Over the past decade there has been growing interest in models consisting of multiple smaller subpopulations (households, workplaces, schools, communities), with the natural assumption of strong homogeneous mixing within each subpopulation, and weaker transmission between subpopulations. Here we consider a model of SIRS (susceptible-infectious-recovered-susceptible) infection dynamics in a very large (assumed infinite) population of households, with the simplifying assumption that each household is of the same size (although all methods may be extended to a population with a heterogeneous distribution of household sizes). For this households model we present efficient methods for studying several quantities of epidemiological interest: (i) the threshold for invasion; (ii) the early growth rate; (iii) the household offspring distribution; (iv) the endemic prevalence of infection; and (v) the transient dynamics of the process. We utilize these methods to explore a wide region of parameter space appropriate for human infectious diseases. We then extend these results to consider the effects of more realistic gamma-distributed infectious periods. We discuss how all these results differ from standard homogeneous-mixing models and assess the implications for the invasion, transmission and persistence of infection. The computational efficiency of the methodology presented here will hopefully aid in the parameterisation of structured models and in the evaluation of appropriate responses for future disease outbreaks.     

Environmental Contamination as a Risk Factor for Intra-Household Staphylococcus aureus Transmission

Background

The household is a recognized community reservoir for Staphylococcus aureus. This study investigated potential risk factors for intra-household S. aureus transmission, including the contribution of environmental contamination.

Methods

We investigated intra-household S. aureus transmission using a sample of multiple member households from a community-based case-control study examining risk factors for CA-MRSA infection conducted in Northern Manhattan. During a home visit, index subjects completed a questionnaire. All consenting household members were swabbed, as were standardized environmental household items. Swabs were cultured for S. aureus. Positive isolates underwent further molecular characterization. Intra-household transmission was defined as having identical strains among two or more household members. Multiple logistic regression was used to identify independent risk factors for transmission.

Results

We enrolled 291 households: 146 index cases, 145 index controls and 687 of their household contacts. The majority of indexes were Hispanic (85%), low income (74%), and female (67%), with a mean age of 31 (range 1–79). The average size of case and control households was 4 people. S. aureus colonized individuals in 62% of households and contaminated the environment in 54% of households. USA300 was the predominant clinical infection, colonizing and environmental strain. Eighty-one households had evidence of intra-household transmission: 55 (38%) case and 26 (18%) control households (P<.01). Environmental contamination with a colonizing or clinical infection strain (aOR: 5.4 [2.9–10.3] P<.01) and the presence of a child under 5 (aOR: 2.3 [1.2–4.5] P = .02) were independently associated with transmission. In separate multivariable models, environmental contamination was associated with transmission among case (aOR 3.3, p<.01) and control households (aOR 27.2, p<.01).

Conclusions

Environmental contamination with a colonizing or clinical infection strain was significantly and independently associated with transmission in a large community-based sample. Environmental contamination should be considered when treating S. aureus infections, particularly among households with multiple infected members.     

An additional step in the transmission of Yersinia pestis?

Plague, caused by the bacterium Yersinia pestis, is a mammalian vector-borne disease, transmitted by fleas that serve as the vector between rodent hosts. For many pathogens, including Y. pestis, there are strong evolutionary pressures that lead to a reduction in ‘useless genes'', with only those retained that reflect function in the specific environment inhabited by the pathogen. Genetic traits critical for survival and transmission between two environments, the rodent and the flea, are conserved in epizootic/epidemic plague strains. However, there are genes that remain conserved for which no function in the flea–rodent cycle has yet been observed, indicating an additional environment may exist in the transmission cycle of plague. Here, we present evidence for highly conserved genes that suggests a role in the persistence of Y. pestis after death of its host. Furthermore, maintenance of these genes points to Y. pestis traversing a post-mortem path between, and possibly within, epizootic periods and offering insight into mechanisms that may allow Y. pestis an alternative route of transmission in the natural environment.     

Enteric Pathogens in Stored Drinking Water and on Caregiver’s Hands in Tanzanian Households with and without Reported Cases of Child Diarrhea

Background

Diarrhea is one of the leading causes of mortality in young children. Diarrheal pathogens are transmitted via the fecal-oral route, and for children the majority of this transmission is thought to occur within the home. However, very few studies have documented enteric pathogens within households of low-income countries.

Methods and Findings

The presence of molecular markers for three enteric viruses (enterovirus, adenovirus, and rotavirus), seven Escherichia coli virulence genes (ECVG), and human-specific Bacteroidales was assessed in hand rinses and household stored drinking water in Bagamoyo, Tanzania. Using a matched case-control study design, we examined the relationship between contamination of hands and water with these markers and child diarrhea. We found that the presence of ECVG in household stored water was associated with a significant decrease in the odds of a child within the home having diarrhea (OR = 0.51; 95% confidence interval 0.27–0.93). We also evaluated water management and hygiene behaviors. Recent hand contact with water or food was positively associated with detection of enteric pathogen markers on hands, as was relatively lower volumes of water reportedly used for daily hand washing. Enteropathogen markers in stored drinking water were more likely found among households in which the markers were also detected on hands, as well as in households with unimproved water supply and sanitation infrastructure.

Conclusions

The prevalence of enteric pathogen genes and the human-specific Bacteroidales fecal marker in stored water and on hands suggests extensive environmental contamination within homes both with and without reported child diarrhea. Better stored water quality among households with diarrhea indicates caregivers with sick children may be more likely to ensure safe drinking water in the home. Interventions to increase the quantity of water available for hand washing, and to improve food hygiene, may reduce exposure to enteric pathogens in the domestic environment.     

Reverse Line Blot Macroarray for Simultaneous Detection and Characterization of Four Biological Warfare Agents

The need for a rapid detection and characterization of biowarfare (BW) agents cannot be over emphasized. With diverse array of potential BW pathogen available presently, rapid identification of the pathogen is crucial, so that specific therapy and control measures can be initiated. We have developed a multiplex polymerase chain reaction based reverse line blot macroarray to simultaneously detect four pathogens of BW importance viz. Bacillus anthracis, Yersinia pestis, Brucella melitensis and Burkholderia pseudomallei. The multiplex PCR utilizes 14 pairs of primers targeting 18 specific markers. These markers include genes which are genus specific, species-specific chromosomal sequences and virulence markers of plasmid origin. The assay was evaluated on various human, environment and animal isolates. The assay w successful in simultaneous detection and characterization of isolates of the four pathogens on as a single platform with sensitivity ranging from 0.3 pg to 0.3 ng of genomic DNA. The assay was able to detect 5 × 10² cfu/ml for B. anthracis, 8 × 10² cfu/ml for Yersinia sp., 1.4 × 10² cfu/ml for B. melitensis and 4 × 10² cfu/ml for B. pseudomallei.     

Molecular Analysis of Shower Curtain Biofilm Microbes

Households provide environments that encourage the formation of microbial communities, often as biofilms. Such biofilms constitute potential reservoirs for pathogens, particularly for immune-compromised individuals. One household environment that potentially accumulates microbial biofilms is that provided by vinyl shower curtains. Over time, vinyl shower curtains accumulate films, commonly referred to as “soap scum,” which microscopy reveals are constituted of lush microbial biofilms. To determine the kinds of microbes that constitute shower curtain biofilms and thereby to identify potential opportunistic pathogens, we conducted an analysis of rRNA genes obtained by PCR from four vinyl shower curtains from different households. Each of the shower curtain communities was highly complex. No sequence was identical to one in the databases, and no identical sequences were encountered in the different communities. However, the sequences generally represented similar phylogenetic kinds of organisms. Particularly abundant sequences represented members of the α-group of proteobacteria, mainly Sphingomonas spp. and Methylobacterium spp. Both of these genera are known to include opportunistic pathogens, and several of the sequences obtained from the environmental DNA samples were closely related to known pathogens. Such organisms have also been linked to biofilm formation associated with water reservoirs and conduits. In addition, the study detected many other kinds of organisms at lower abundances. These results show that shower curtains are a potential source of opportunistic pathogens associated with biofilms. Frequent cleaning or disposal of shower curtains is indicated, particularly in households with immune-compromised individuals.     

Historical Overview of Taenia asiatica in Taiwan

An overview of the epidemiological, biological, and clinical studies of Taenia and taeniasis in Taiwan for the past century is presented. The phenomenal observations that led to the discovery of Taenia asiatica as a new species, which differ from Taenia solium and Taenia saginata, are described. Parasitological surveys of the aborigines in Taiwan revealed a high prevalence of taeniasis, which might be due to the culture of eating raw liver of hunted wild boars. Chemotherapeutic deworming trials involving many patients with taeniasis were discussed. Praziquantel was found to be very effective, but sometimes complete worms could not be recovered from the feces after treatment, probably due to the dissolution of the proglottids. Atabrine, despite some side effects, can still be used, in properly controlled dosages, as the drug of choice for human T. asiatica infection if we need to recover the expelled worms for morphological examinations. Research results on the infection of T. asiatica eggs from Taiwan aborigines in experimental animals were also noted. Since the pig serve as the natural intermediate host of T. asiatica and the predilection site is the liver, a differential comparison of other parasitic pathogens that might cause apparently similar lesions is also presented.     

Phylogenetic and metabolic diversity of bacteria associated with cystic fibrosis

In patients afflicted with cystic fibrosis (CF), morbidity and mortality are primarily associated with the adverse consequences of chronic microbial bronchial infections, which are thought to be caused by a few opportunistic pathogens. However, recent evidence suggests the presence of other microorganisms, which may significantly affect the course and outcome of the infection. Using a combination of 16S rRNA gene clone libraries, bacterial culturing and pyrosequencing of barcoded 16S rRNA amplicons, the microbial communities present in CF patient sputum samples were examined. In addition to previously recognized CF pathogens such as Pseudomonas aeruginosa and Staphylococcus aureus, >60 phylogenetically diverse bacterial genera that are not typically associated with CF pathogenesis were also detected. A surprisingly large number of fermenting facultative and obligate anaerobes from multiple bacterial phyla was present in each sample. Many of the bacteria and sequences found were normal residents of the oropharyngeal microflora and with many containing opportunistic pathogens. Our data suggest that these undersampled organisms within the CF lung are part of a much more complex microbial ecosystem than is normally presumed. Characterization of these communities is the first step in elucidating potential roles of diverse bacteria in disease progression and to ultimately facilitate advances in CF therapy.     

Estimating a Markovian Epidemic Model Using Household Serial Interval Data from the Early Phase of an Epidemic

The clinical serial interval of an infectious disease is the time between date of symptom onset in an index case and the date of symptom onset in one of its secondary cases. It is a quantity which is commonly collected during a pandemic and is of fundamental importance to public health policy and mathematical modelling. In this paper we present a novel method for calculating the serial interval distribution for a Markovian model of household transmission dynamics. This allows the use of Bayesian MCMC methods, with explicit evaluation of the likelihood, to fit to serial interval data and infer parameters of the underlying model. We use simulated and real data to verify the accuracy of our methodology and illustrate the importance of accounting for household size. The output of our approach can be used to produce posterior distributions of population level epidemic characteristics.     

Transmitting Plant Viruses Using Whiteflies

Whiteflies, Hemiptera: Aleyrodidae, Bemisia tabaci, a complex of morphologically indistinquishable species⁵, are vectors of many plant viruses. Several genera of these whitefly-transmitted plant viruses (Begomovirus, Carlavirus, Crinivirus, Ipomovirus, Torradovirus) include several hundred species of emerging and economically significant pathogens of important food and fiber crops (reviewed by^9,10,16). These viruses do not replicate in their vector but nevertheless are moved readily from plant to plant by the adult whitefly by various means (reviewed by^{2,6,7,9,10,11,17}). For most of these viruses whitefly feeding is required for acquisition and inoculation, while for others only probing is required. Many of these viruses are unable or cannot be easily transmitted by other means. Therefore maintenance of virus cultures, biological and molecular characterization (identification of host range and symptoms)^3,13, ecology^2,12, require that the viruses be transmitted to experimental hosts using the whitefly vector. In addition the development of new approaches to management, such as evaluation of new chemicals¹⁴ or compounds¹⁵, new cultural approaches^1,4,19, or the selection and development of resistant cultivars^7,8,18, requires the use of whiteflies for virus transmission. The use of whitefly transmission of plant viruses for the selection and development of resistant cultivars in breeding programs is particularly challenging⁷. Effective selection and screening for resistance employs large numbers of plants and there is a need for 100% of the plants to be inoculated in order to find the few genotypes which possess resistance genes. These studies use very large numbers of viruliferous whiteflies, often several times per year.Whitefly maintenance described here can generate hundreds or thousands of adult whiteflies on plants each week, year round, without the contamination of other plant viruses. Plants free of both whiteflies and virus must be produced to introduce into the whitefly colony each week. Whitefly cultures must be kept free of whitefly pathogens, parasites, and parasitoids that can reduce whitefly populations and/or reduce the transmission efficiency of the virus. Colonies produced in the manner described can be quickly scaled to increase or decrease population numbers as needed, and can be adjusted to accommodate the feeding preferences of the whitefly based on the plant host of the virus.There are two basic types of whitefly colonies that can be maintained: a nonviruliferous and a viruliferous whitefly colony. The nonviruliferous colony is composed of whiteflies reared on virus-free plants and allows the weekly availability of whiteflies which can be used to transmit viruses from different cultures. The viruliferous whitefly colony, composed of whiteflies reared on virus-infected plants, allows weekly availability of whiteflies which have acquired the virus thus omitting one step in the virus transmission process.